BIOMARKERS

Description

FIELD OF THE INVENTION

The present invention relates to the identification of biomarkers associated with cancer, in particular colorectal cancer and precursors thereof, i.e. high risk adenomas, and their use in screening methods, arrays for use in colorectal cancer screening methods, methods of treating colorectal cancer, and compounds for use in treating colorectal cancer, and kits for use in colorectal cancer screening methods.

BACKGROUND OF THE INVENTION

Colorectal cancer (CRC) is a significant health problem. It is the 3rd most common cancer worldwide, with over 1.200.000 new cases each year, with a fatal outcome for almost half the patients (Karsa L V, et al. Best Pract Res Clin Gastroenterol 2010; 24:381-96). Because CRC develops over many years, there is an excellent window of opportunity to detect the disease in an early, curable, or even premalignant stage. This can be achieved by screening of asymptomatic individuals. However, the currently available screening tests are either cumbersome or carry risk of complications and over treatment like colonoscopy.

The immunochemical fecal occult blood test (iFOBT), further referred to as fecal immunochemical test (FIT), is a widely used test that detects small traces of blood in stool, derived from lesions such as tumors that bleed in the colon. The FIT is a non-invasive method that makes use of an antibody directed against hemoglobin. However, not all colon tumors bleed and therefore the FIT has a sensitivity that leaves room for improvement (Duffy M J, et al. Int J Cancer 2011; 128:3-11; van Veen W, Mali W P. [Colorectal cancer screening: advice from the Health Council of the Netherlands]. Ned Tijdschr Geneeskd 2009; 153:A1441). Additional markers that detect other tumor characteristics besides bleeding in the stool could increase this sensitivity and the chance of identifying a colon tumor. Molecular changes resulting from the neoplastic process include changes in protein expression. The proteins for which expression is increased have the potential to serve as informative biomarkers with high diagnostic performance.

Several studies have been performed to identify proteins in stool and blood that can be used for the early detection of CRC and high risk colorectal adenomas, but most of these proteins have not been validated in a screening setting or failed to improve current tests for early detection, such as Carcinoembryonic antigen (CEA) or Calprotectin (Bosch L J, et al. Molecular tests for colorectal cancer screening. Clin Colorectal Cancer 2011; 10:8-23). There therefore remains a need for further tumor-specific protein markers to improve the current available non-invasive screening possibilities for CRC and high risk adenomas.

Recent technological advances in mass spectrometry can boost the discovery of novel protein markers (de Wit M, et al. Gut 2011; Jimenez C R, et al. J Proteomics 2010; 73: 1873-95). Tandem mass spectrometry-based approaches now have the power to analyze complex protein samples and to detect proteins in low concentrations (Cox J, Mann M. Annu Rev Biochem 2011; 80: 273-99). Although most biomarker discovery studies have been performed using tissue and/or cell line material followed by validation in stool or blood, the chemical composition of the biological sample used for screening may significantly affect the nature of biomarkers that can be identified. This holds especially for stool samples, in which the low pH, the protease- and glycosidase activities of bacteria, enzymes and other substances can disturb the specific detection of these markers (Young G P, Bosch L J W. Curr Colorectal Cancer Rep 2011; 7: 62-70). Measuring molecules directly in the biological sample that may be used for screening, i.e. stool, could therefore be a valuable approach to provide us with reliable biomarkers that are stable in the fecal environment. A recent study by Ang et al. has shown the feasibility of protein biomarker discovery in human stool samples using mass spectrometry (Ang C S, Nice E C. J Proteome Res 2010; 9: 4346-55).

The present invention aims to overcome some or all of the problems associated with the prior art.

SUMMARY OF THE INVENTION

According to a first aspect of the invention there is provided a method for screening for colorectal cancer, the method comprising:

- screening a biological sample obtained from an individual for one or more biomarkers selected from the group defined in Table 1 and/or Table 6, wherein the presence of or increased expression of the one or more biomarkers relative to a control sample is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer.

In a further aspect of the invention, there is provided a method for screening for colorectal cancer, the method comprising: screening a biological sample obtained from an individual for one or more biomarkers selected from the group defined in Table 1 and/or Table 6, wherein the presence of or increased expression of the one or more biomarkers relative to a control sample is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer, and wherein the one or more biomarkers were selected from secretomes significantly more abundant in colon tumor secretomes compared to normal colon secretomes.

In yet a further aspect of the invention, there is provided a method for determining the significance of one or more biomarkers for the determination whether an individual is at risk of suffering from or is suffering from colorectal cancer; comprising:

- comparing the presence of or increased expression of one or more biomarkers significantly more abundant in colon tumor secretomes compared to normal colon secretomes with the presence of one or more biomarkers in a biological control sample of an individual suffering from colorectal cancer, and selecting one or more biomarkers indicative of the presence of colorectal cancer.

In one embodiment, the one or more biomarkers is a protein or proteins.

Thus, the present invention provides protein signatures for diagnosing or predicting colorectal cancer in a subject. The present invention advantageously allows for detection of advanced and high-risk colonic adenomas and adenocarcinomas.

In one embodiment, the presence of the one or more biomarkers is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer.

In an alternative embodiment, the increased expression of the one or more biomarkers relative to a control sample is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer. The one or more biomarkers may be selected from the group defined in Table 2. The biomarkers in Table 2 represent a preferred subset of the gene products of Table 1 and/or Table 6, for which the levels of differential expression in CRC or high risk colorectal adenoma samples relative to control samples are considered statistically significant.

The one or more biomarkers may be selected from the group defined in Table 3. The biomarkers of Table 3 are differentially expressed in FIT-negative CRC samples relative to a control sample.

This particular subset of the markers of Table 1 and/or Table 6 are of particular importance as these markers have the potential to be used for the detection of CRC or susceptibility to CRC (i.e. detection of high risk colorectal adenomas) in those patients for whom the fecal immunochemical test, or any other test that aims to detect hemoglobin (such as the guaiac-based Fecal Occult Blood Test), gives a comes a negative result. The same applies to the markers of Tables 5 and 7, wherein the latter show the overlap between markers found in stool samples, and those found in secretomes of cancerous tumor tissue.

In this respect, Table 6 lists proteins that were found to be significantly more abundant in colon tumor secretomes compared to normal colon secretomes.

The one or more biomarkers may have a higher discriminative power than hemoglobin. By “higher discriminative power” it is meant that a biomarker has a higher sensitivity and specificity than hemoglobin. Thus, in one embodiment, the one or more biomarkers may be selected from the group consisting of: S100 calcium binding protein A8 (S100A8), complement component C4B (Chido blood group) 2 (C4A/C4B), transferrin (TF), alpha-2-macroglobulin (A2M), S100 calcium binding protein A9 (S100A9), proteinase 3 (PRTN3), Azurocidin (AZU1), lactotransferrin (LTF), hemopexin (HPX) and defensin, alpha 1 (DEFA1).

The one or more biomarkers may be selected from the group defined in Table 4. The group of biomarkers in Table 4 represent a subset of Table 1 and/or Table 6, and are markers which have been found to be expressed only in CRC samples and not in control samples.

In one embodiment, the biological sample is a stool sample.

The biological sample preferably may be screened for the one or more biomarkers using (targeted) mass spectrometry.

The biological sample may be screened for the one or more biomarkers using a binding agent capable of binding to the one or more biomarkers.

The binding agent may be an antibody or fragment thereof. The antibody or fragment thereof may be a recombinant antibody or fragment thereof. The antibody or fragment thereof may be selected from the group consisting of: scFv; Fab; a binding domain of an immunoglobulin molecule.

In one embodiment, the binding agent may be an aptamer.

The screening may be performed using an array. Any suitable array may advantageously be employed. The array may be a bead-based array. The array may be a surface-based array.

The control sample may comprise a biological sample, e.g. a stool sample from an individual known to be free from colorectal cancer or high risk colorectal adenomas.

The biological sample may also be analysed by a fecal immunochemical test. According to a second aspect of the present invention there is provided the use of one or more biomarkers selected from the group defined in Table 1 and/or Table 6 for diagnosing or predicting colorectal cancer in an individual.

The one or more biomarkers of the second aspect may be selected from the group defined in Table 2.

The one or more biomarkers of the second aspect may be selected from the group defined in Table 3.

The one or more biomarkers of the second aspect may have a higher discriminative power than hemoglobin.

Thus, the one or more biomarkers of the second aspect may be selected from the group consisting of: S100 calcium binding protein A8 (S100A8), complement component C4B (Chido blood group) 2 (C4A/C4B), transferrin (TF), alpha-2-macroglobulin (A2M), S100 calcium binding protein A9 (S100A9), proteinase 3 (PRTN3), Azurocidin (AZU1), lactotransferrin (LTF), hemopexin (HPX) and defensin, alpha 1 (DEFA1).

The one or more biomarkers of the second aspect may be selected from the group defined in Table 4.

The one or more biomarkers of the second aspect may be selected from the group defined in Table 5.

The one or more biomarkers of the second aspect may be selected from the group defined in Table 6.

The one or more biomarkers of the second aspect may be selected from the group defined in Table 7.

According to a third aspect of the invention there is provided an array for determining whether an individual is at risk of suffering from or is suffering from colorectal cancer, the array comprising one or more binding agent as defined according to certain embodiments of the first aspect of the invention.

The array may be for use in a method according to the first aspect of the invention. According to a fourth aspect of the invention there is provided a method for treating colorectal cancer, the method comprising:

- screening a biological sample, preferably a stool sample, obtained from an individual for one or more biomarkers selected from the group defined in Table 1 and/or Table 6, as applicable, wherein the presence of or increased expression of the one or more biomarkers relative is indicative that the individual is suffering from colorectal cancer; and administering a therapeutically active amount of a cancer therapeutic agent.

The one or more biomarkers of the fourth aspect may be selected from the group defined in Table 2.

The one or more biomarkers of the fourth aspect may be selected from the group defined in Table 3.

The one or more biomarkers of the fourth aspect may have a higher discriminative power than hemoglobin.

Thus, the one or more biomarkers of the fourth aspect may be selected from the group consisting of: S100 calcium binding protein A8 (S100A8), complement component C4B (Chido blood group) 2 (C4A/C4B), transferrin (TF), alpha-2-macroglobulin (A2M), S100 calcium binding protein A9 (S100A9), proteinase 3 (PRTN3), Azurocidin (AZU1), lactotransferrin (LTF), hemopexin (HPX) and defensin, alpha 1 (DEFA1).

The one or more biomarkers of the fourth aspect may be selected from the group defined in Table 4.

According to a fifth aspect of the invention there is provided a cancer therapeutic agent for use in a method for treating colorectal cancer, the method comprising:

- screening a stool sample obtained from an individual for one or more biomarkers selected from the group defined in Table 1, wherein the presence of or increased expression of the one or more biomarkers relative is indicative that the individual is suffering from colorectal cancer; and
- administering a therapeutically active amount of the cancer therapeutic agent.

The cancer therapeutic agent of the fourth or fifth aspect may comprise one or more therapeutic monoclonal antibody, one or more small molecule inhibitor or one or more chemotherapeutic agent or any combination thereof.

The one or more therapeutic monoclonal antibody may comprise one or more of bevacizumab, cetuximab or panitumumab or any combination thereof.

The one or more small molecule inhibitor may comprise one or more of erlotinib, sorafenib or alisertib or any combination thereof.

The one or more chemotherapeutic agent may comprise one or more of 5-FU, capecitabine, irinotecan oxaliplatin, or leucovorin or any combination thereof.

The one or more biomarkers of the fifth aspect may be selected from the group defined in Table 2.

The one or more biomarkers of the fifth aspect may be selected from the group defined in Table 3.

The one or more biomarkers of the fifth aspect may have a higher discriminative power than hemoglobin.

Thus, the one or more biomarkers of the fifth aspect may be selected from the group consisting of: S100 calcium binding protein A8 (S100A8), complement component C4B (Chido blood group) 2 (C4A/C4B), transferrin (TF), alpha-2-macroglobulin (A2M), S100 calcium binding protein A9 (S100A9), proteinase 3 (PRTN3), Azurocidin (AZU1), lactotransferrin (LTF), hemopexin (HPX) and defensin, alpha 1 (DEFA1).

The one or more biomarkers of the fifth aspect may be selected from the group defined in Table 4. The one or more biomarkers of the fifth aspect may be selected from the group defined in Table 5 and/or 7.

According to a sixth aspect of the invention there is provided a kit for screening for colorectal cancer in an individual, the kit comprising:

- (a) One or more binding agent which selectively bind to one or more biomarkers as defined in Table 1, or an array according to the third aspect;
- (b) Instructions for performing the method as defined in the first aspect.

The kit may, for example, be an ELISA kit. The one or more binding agent may, for example, comprise an antibody. The one or more binding agent may comprise an aptamer.

Any one or more features described for any aspect of the present invention or preferred embodiments or examples thereof, described herein, may be used in conjunction with any one or more other features described for any other aspect of the present invention or preferred embodiments or examples thereof described herein. The fact that a feature may only be described in relation to one aspect or embodiment or example does not limit its relevance to only that aspect or embodiment or example if it is technically relevant to one or more other aspect or embodiment or example.

DETAILED DESCRIPTION OF THE INVENTION

Colorectal Cancer

The most common colon cancer cell type is adenocarcinoma which accounts for 95% of cases. Other, rarer types include lymphoma and squamous cell carcinoma. Colorectal adenocarcinoma arises from precursor lesions called adenomas, of which only a minority progress to cancer. Adenomas that progress to cancer are referred to as high risk adenomas.

Protein markers have great potential to be applied for stool-based CRC screening, because they can be measured in small sample volumes with simple and relatively cheap assays, of which the widely used FIT is an excellent example (Bosch L J, et al. Molecular tests for colorectal cancer screening. Clin Colorectal Cancer 2011; 10; 8-23; Young G P, Bosch L J W. Curr Colorectal Cancer Rep 2011; 7: 62-70; Oort F A, et al. Aliment Pharmacol Ther 2010; 31: 432-9).

The present study has identified novel protein biomarkers by applying in-depth proteomics to stool samples. From a total of 830 detected human proteins, 134 were significantly enriched in stool samples from CRC patients compared to control stool samples, of which several showed higher discriminative power than hemoglobin and/or complementarity to hemoglobin.

The approach of measuring molecules directly in stool is of significance to reveal biomarkers that are stable in the fecal environment and detectable in the background of bacterial- and food-related molecules.

The present invention is advantageously used for screening for colorectal cancer, that is adenocarcinoma found in the colon. However, the methods of the invention should not be considered as being limited solely to the detection of colonic adenocarcinomas. Rather, the methods of the invention are also useful in the detection of advanced or high-risk colonic adenomas, thus enabling the identification of an individual at risk of developing colorectal cancer due to the presence of an advanced or a high-risk adenoma.

References herein to screening for colorectal cancer thus may include screening for advanced colonic adenomas and high-risk adenomas as well as colonic adenocarcinoma.

It is also expected that the biomarkers identified by the present invention may also find application for the diagnosis of adenocarcinomas present higher up the gastrointestinal tract.

Thus, the present invention may also provide a method for screening for gastrointestinal disease or gastrointestinal cancer, the method comprising: screening a biological sample, for example a stool sample, from an individual for one or more biomarkers selected from the group defined in Table 1, wherein the presence of or increased expression of the one or more biomarkers relative to a control sample is indicative that the patient is at risk of suffering from or is suffering from gastrointestinal disease or gastrointestinal cancer.

Sample for Screening

The sample for screening may include cell lines, biopsies, whole blood, blood serum, sputum, stool, urine, synovial fluid, wound fluid, cerebral-spinal fluid, tissue from eyes, intestine, kidney, brain, skin, heart, prostate, lung, breast, liver, muscle or connective tissue, the said tissue being optionally embedded in paraffin, histologic object slides, and all possible combinations thereof.

The preferred biological sample is stool.

The sample may be prepared by any conventional method for extracting proteins from a biological sample. One exemplary method can be found in Ang C S, Nice E C. J Proteome Res. 2010; 9:4346-55, the contents of which are incorporated herein by reference.

Biomarkers

The present invention provides a set of biomarkers which may be detected directly from a stool sample and which have been shown to be reliable indicators for the presence of advanced colonic adenomas or adenocarcinomas in an individual.

The biomarkers identified are listed in Table 1 and/or Table 6. In one embodiment, the methods of the invention screen for more than one biomarker from the group defined in Table 1, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers of the group defined in Table 1 and/or Table 6. In an alternative embodiment, the methods of the invention screen for more than ten of the biomarkers of the group defined in Table 1, for example, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, thirty, forty, fifty, sixty, seventy, eighty, ninety, one hundred of the biomarkers of the group defined in Table 1 and/or Table 6.

Thus, the methods of the invention screen for the presence of or increased expression of one or more of: complement component C4B (Chido blood group) 2 (C4A/C4B); glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2); glucose-6-phosphate isomerase (GPI); transketolase (TKT); N-acylaminoacyl-peptide hydrolase (APEH); histone cluster 1, H4c (HIST4H4 (includes others)); Fatty acid-binding protein 5 (psoriasis-associated) (FABP5); hexosaminidase B (beta polypeptide) (HEXB); epithelial cell adhesion molecule (EPCAM); NME1-NME2 walkthrough (NME1-NME2); Superoxide dismutase 2, mitochondrial (SOD2); Tu translation elongation factor, mitochondrial (TUFM); Glutathione synthetase (GSS); annexin A2 (ANXA2); ATP synthase, H+ transporting, mitochondrial F1 complex, beta polypeptide (ATP5B); 10 kDa heat shock protein (chaperonin 10) (HSPE1); glyoxalase I (GLO1); histone cluster 2, H2be (HIST2H2BE (includes others)); S100 calcium binding protein A4 (S100A4); S100 calcium binding protein A11 (S100A11); latexin (LXN); dehydrogenase/reductase (SDR family) member 11 (DHRS11); N-acetylglucosaminidase, alpha (NAGLU); Translin (TSN); Proteasome (prosome, macropain) subunit alpha type-4 (PSMA4); Proteasome (prosome, macropain) subunit alpha type-6 (PSMA6); ras-related C3 botulinum toxin substrate 1 (rho family, small GTP binding protein Rac1) (RAC1); Adenosylhomocysteinase (AHOY); fucosidase, alpha-L-1, tissue (FUCA1); S100 calcium binding protein P (S100P); Proteasome (prosome, macropain) subunit beta type-2 (PSMB2); X-prolyl aminopeptidase (aminopeptidase P) 1 (XPNPEP1); Keratin 18 (KRT18); Nuclear cap-binding protein subunit 1 80 kDa (NCBP1); mannosidase, alpha, class 2B, member 1 (MAN2B1); S100 calcium binding protein A6 (S100A6); valosin containing protein (VCP); quinolinate phosphoribosyltransferase (QPRT); major histocompatibility complex, class I, B (HLA-B); phosphoglycerate mutase 1 (brain) (PGAM1); ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3); serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10); myeloperoxidase (MPO); creatine kinase, mitochondrial 1B (CKMT1A/CKMT1B); proteinase 3 (PRTN3); elastase, neutrophil expressed (ELANE); MORC family CW-type zinc finger 1 (MORC1); ubiquitin B (UBB); phospholipase A2, group IIA (platelets, synovial fluid) (PLA2G2A); carbonic anhydrase IV (CA4); G elongation factor, mitochondrial 2 (GFM2); S100 calcium binding protein A7 (S100A7); Bactericidal permeability-increasing protein (BPI); collagen, type VI, alpha 5 (COL6A5); LIM homeobox 8 (LHX8); cysteine-rich secretory protein 3 (CRISP3); Azurocidin (AZU1); hemicentin 1 (HMCN1); Transglutaminase 3 (E polypeptide, protein-glutamine gamma-glutamyltransferase) (TGM3); CDC42 binding protein kinase alpha (DMPK-like) (CDC42BPA); Cathepsin G (CTSG); Resistin (RETN); methylmalonyl CoA mutase (MUT); armadillo repeat containing, X-linked 4 (ARMCX4); Integrin alpha-M (complement component 3 receptor 3 subunit) (ITGAM); Calcium channel, voltage dependent, R-type alpha-1E subunit (CACNA1E); T-cell lymphoma invasion and metastasis 2 (TIAM2); HIR histone cell cycle regulation defective homolog A (S. cerevisiae) (HIRA); dopey family member 2 (DOPEY2); integrin beta 1 binding protein 3 (ITGB1BP3); Sodium channel, voltage-gated, type VII, alpha (SCN7A); Rab3C, member RAS oncogene family (RAB3C); chromosome 9 open reading frame 79 (C9orf79); nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 4 (NFATC4); UDP-glucose glycoprotein glucosyltransferase 2 (UGGT2); Cornulin (CRNN); kielin/chordin-like protein (KCP); CD1E molecule (CD1E); coiled-coil domain-containing 18 (CCDC18); leukotriene A-4 hydrolase (LTA4H); albumin (ALB); alpha-2-macroglobulin (A2M); complement component 3 (C3); hemoglobin, beta (HBB); transferrin (TF); hemoglobin, alpha 1 (HBA1/HBA2); lactotransferrin (LTF); ceruloplasmin (ferroxidase) (CP); catalase (CAT); group-specific component (vitamin D-binding protein) (GC); serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1); fibrinogen gamma chain (FGG); S100 calcium binding protein A8 (S100A8); ferritin, light polypeptide (FTL); actin, beta (ACTB); fibronectin 1 (FN1); defensin, alpha 1 (DEFA1 (includes others)); serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1); retinol binding protein 4, plasma (RBP4); peroxiredoxin 2 (PRDX2); fibrinogen alpha chain (FGA); serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2); carbonic anhydrase II (CA2); orosomucoid 1 (ORM1/ORM2); lactate dehydrogenase A (LDHA); vitronectin (VTN); kininogen-1 (KNG1); actin, alpha, cardiac muscle 1 (ACTC1); leucine-rich alpha-2-glycoprotein 1 (LRG1); gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH); enolase 1, (alpha) (ENO1); profilin 1 (PFN1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 7 (SERPINA7); alpha-1-microglobulin/bikunin precursor (AMBP); lamin A/C (LMNA); apolipoprotein D (APOD); thyroid hormone receptor interactor 11 (TRIP11); complement component 4 binding protein, alpha (C4BPA); tropomyosin 4 (TPM4); filamin A, alpha (FLNA); haptoglobin (HP); hemopexin (HPX); hemoglobin, delta (HBD); fibrinogen beta chain (FGB); S100 calcium binding protein A9 (S100A9); complement component 5 (C5); solute carrier family 26, member 3 (SLC26A3); complement component 9 (C9); amyloid P component, serum (APCS); alpha-1-B glycoprotein (A1BG); complement C3-like (LOC100133511); inter-alpha (globulin) inhibitor H4 (plasma Kallikrein-sensitive glycoprotein) (ITIH4); complement component C8, alpha polypeptide (C8A); inter-alpha (globulin) inhibitor H1 (ITIH1); acyl-CoA dehydrogenase, very long chain (ACADVL); cDNA FLJ60317, highly similar to Aminoacylase-1 (ACY1); Ankyrin repeat domain-containing protein 35 (ANKRD35); baculoviral IAP repeat-containing 6 (BIRC6); Bleomycin hydrolase (BLMH); bone marrow stromal cell antigen 12 (BST1); hypothetical protein LOC643677 (C13orf40); Cytidine deaminase (CDA); chitinase 1 (chitotriosidase) (CHIT1); cathepsin C (CTSC); Cathepsin S (CTSS); Isoform 2 of Dedicator of cytokinesis protein 4 (DOCK4); Glutathione reductase (GSR); hect (homologous to the E6-AP (UBE3A) carboxyl terminus) domain and RCC1 (CHC1)-like domain (RLD) 1 (HERC1); hect domain and RLD 2 (HERC2); major histocompatibility complex, class II, DR beta 5 (HLA-DRB5); isocitrate dehydrogenase 1 (NADP+), soluble (IDH1); inter-alpha (globulin) inhibitor H2 (ITIH2); Uncharacterized protein KIAA1797 (KIAA1797); Lysozyme C (LYZ); Nebulin (NEB); NIMA (never in mitosis gene a)-related kinase 10 (NEK10); peptidase D (PEPD); quiescin Q6 sulfhydryl oxidase 1 (QSOX1); ribonuclease T2 (RNASET2); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 (SERPINA1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 (SERPINA3); serpin peptidase inhibitor, clade B (ovalbumin), member 3 (SERPINB3); SET domain containing 2 (SETD2); Shugoshin-like 2 (SGOL2); sialic acid acetylesterase (SIAE); spectrin repeat containing, nuclear envelope 1 (SYNE1); Transaldolase 1 (TALDO1); Taste receptor type 2 member 42 (TAS2R42); triosephosphate isomerase 1 (TPI1); Vinculin (VCL); Zymogen granule membrane protein 16 (ZG16); hypothetical protein LOC79887 (PLBD1); Isoform 1 of Serine/threonine-protein phosphatase 6 regulatory ankyrin repeat subunit A (ANKRD28); Cystatin-C(CST3); D-dopachrome decarboxylase (DDT); Synapse-associated protein 1 (SYAP1); Proteasome subunit alpha type-2 (PSMA2); SUB1 homolog (S. cerevisiae) (SUB1); Microfibril-associated glycoprotein 3 (MFAP3); Cathepsin D (CTSD); proteasome (prosome, macropain) subunit, beta type, 1 (PSMB1); proteasome (prosome, macropain) subunit, beta type, 5 (PSMB5); cDNA FLJ61112, highly similar to BTB/POZ domain-containing protein KCTD15 (KCTD15); prolyl 4-hydroxylase, beta polypeptide (P4HB); glutathione peroxidase 1 (GPX1); serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5); Isoform 1 of collagen, type IV, alpha 3 (Goodpasture antigen) binding protein (COL4A3BP); proteasome (prosome, macropain) subunit, beta type, 6 (PSMB6); Keratin 20 (KRT20); Calpain small subunit 1 (CAPNS1); peroxiredoxin 3 (PRDX3); NACC family member 2, BEN and BTB (POZ) domain containing (NACC2); Rho GDP-dissociation inhibitor 2 (ARHGDIB); Macrophage migration inhibitory factor (MIF); Ran-binding protein 6 (RANBP6); spinster homolog 3 (Drosophila) (SPNS3); minichromosome maintenance complex component 2 (MCM2); Fumarylacetoacetase (FAH); heat shock 70 kDa protein 8 (HSPA8); brain abundant, membrane attached signal protein 1 (BASP1); Branched-chain-amino-acid aminotransferase (BCAT2); Moesin (MSN); serpin peptidase inhibitor, clade B (ovalbumin), member 8 (SERPINB8); glucose-6-phosphate dehydrogenase (G6PD); Isoform 1 of UPF0557 protein C10orf119 (C10orf119); Prosaposin (PSAP); eukaryotic translation elongation factor 1 gamma (EEF1G); four and a half LIM domains 1 (FHL1); carboxypeptidase, vitellogenic-like (CPVL); tubulin tyrosine ligase-like family, member 3 (TTLL3); IPI:IPI00942608.1|ENSEMBLENSP0000 (unmapped; 26 kDa protein); proline-rich protein BstNI subfamily 2 (PRB1/PRB2); Protocadherin-8 (PCDH8); Alpha-2-macroglobulin-like protein 1 (A2ML1); Guanine deaminase (GDA); Lipocalin-1 (LCN1); Histone H1.4 (HIST1H1E); IPI:IPI00937064.1IREFSEQ:XP_—002342720 (ZAN); heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1);); Endoplasmic reticulum aminopeptidase 2 (ERAP2); 14-3-3 protein zeta/delta (YWHAZ); G-protein coupled receptor 39 (GPR39); similar to KIAA1783 protein (KIAA1783 protein); apolipoprotein A-I binding protein (APOA1BP); pleckstrin and Sec7 domain containing 2 (PSD2); prolylcarboxypeptidase (angiotensinase C) (PROP); Tubulin alpha-1C chain (TUBA1C); Calmodulin-like protein 5 (CALML5); ARP3 actin-related protein 3 homolog (yeast) (ACTR3); myosin, light chain 6, alkali, smooth muscle and non-muscle (MYL6); Vasodilator-stimulated phosphoprotein (VASP); ARP2 actin-related protein 2 homolog (yeast) (ACTR2); Rheumatoid factor (RF-IP18); Phosphoglycerate kinase 1 (PGK1); Solute carrier family 35 member F1 (SLC35F1); Solute carrier family 35 member F1 (SLC35F1); alkaline phosphatase, liver/bone/kidney (ALPL); I tropomyosin 3 (TPM3); Hexokinase-3 (HK3); Vimentin (VIM); Annexin A1 (ANXA1); IPI:IPI100930073.1|TREMBL:B2R853 (KRT6C); Keratin, type II cytoskeletal 6C (KRT6C); myosin, heavy chain 13, skeletal muscle (MYH13); cell cycle progression 1 (CCPG1); Hypothetical protein (H-INV); calcium channel, voltage-dependent, L type, alpha 1D subunit (CACNA1D); LY6/PLAUR domain containing 5 (LYPD5); aarF domain containing kinase 2 (ADCK2); Myosin-lc (MYO1C); amyloid beta precursor protein (cytoplasmic tail) binding protein 2 (APPBP2); integrin, alpha 2b (platelet glycoprotein IIb of IIb/IIIa complex, antigen CD41) (ITGA2B); tubulin, beta 6 (TUBB6); synaptotagmin-like 4 (SYTL4); aquaporin 4 (AQP4); cell division cycle 42 (GTP binding protein, 25 kDa) (CDC42); myosin, light chain 12B, regulatory (MYL12B); protein L-Myc-2-like (LOC100293553); RAP1B, member of RAS oncogene family (RAP1B); glycoprotein IX (platelet) (GP9); Destrin (DSTN); complement component 1, q subcomponent, C chain (C1QC); epidermal growth factor receptor pathway substrate 8 (EPS8); dual specificity phosphatase 3 (DUSP3); ras homolog gene family, member A (RHOA); myosin, light chain 9, regulatory (MYL9); peptidylprolyl isomerase A (cyclophilin A) (PPIA); Cofilin-1 (CFL1); and/or lactotransferrin (LTF), collagen, type XII, alpha 1 (COL12A1), agrin (AGRN), +MYB binding protein (P160) Ia (MYBBP1A), transformation/transcription domain-associated protein (TRRAP), annexin A6 (ANXA6), cytoskeleton associated protein 5 (CKAP5), minichromosome maintenance complex component 5 (MOMS), importin 4 (IPO4), neurobeachin-like 2 (NBEAL2), minichromosome maintenance complex component 4 (MCM4), 2′-5′-oligoadenylate synthetase 3, 100 kDa (OAS3), minichromosome maintenance complex component 3 (MCM3), NEDD8 activating enzyme E1 subunit 1 (NAE1), tripartite motif containing 28 (TRIM28), fused in sarcoma (FUS), phenylalanyl-tRNA synthetase, alpha subunit (FARSA), myeloid cell nuclear differentiation antigen (MNDA), suppressor of Ty 16 homolog (S. cerevisiae) (SUPT16H), DEAD (Asp-Glu-Ala-Asp) box polypeptide 5 (DDX5), tenascin C (TNC), nuclear import 7 homolog (S. cerevisiae) (NIP7), chromodomain helicase DNA binding protein 4 (CHD4), regulator of chromosome condensation 2 (RCC2), DNA (cytosine-5-)-methyltransferase 1 (DNMT1), exportin 4 (XPO4), chaperonin containing TCP1, subunit 5 (epsilon) (COTS), serine/arginine-rich splicing factor 9 (SRSF9), spectrin, beta, non-erythrocytic 2 (SPTBN2), TIMP metallopeptidase inhibitor 1 (TIMP1), nidogen 1 (NID1), ribonucleotide reductase M1 (RRM1), eukaryotic translation initiation factor 4 gamma, 1 (EIF4G1), component of oligomeric golgi complex 4 (COG4), polymerase (DNA directed), delta 1, catalytic subunit 125 kDa (POLD1), splicing factor 3b, subunit 2, 145 kDa (SF3B2), exosome component 2 (EXOSC2), minichromosome maintenance complex component 6 (MCM6), plastin 3 (PLS3), aldolase B, fructose-bisphosphate (ALDOB), SMG1 homolog, phosphatidylinositol 3-kinase-related kinase (C. elegans) (SMG1), G1 to S phase transition 1 (GSPT1), KH-type splicing regulatory protein (KHSRP), DEAD (Asp-Glu-Ala-Asp) box polypeptide 21 (DDX21), phosphatidylinositol transfer protein, beta (PITPNB), aquarius homolog (mouse) (AQR), heterogeneous nuclear ribonucleoprotein D-like (HNRPDL), annexin A3 (ANXA3), processing of precursor 1, ribonuclease P/MRP subunit (S. cerevisiae) (POP1), structural maintenance of chromosomes 2 (SMC2), dynein, cytoplasmic 1, light intermediate chain 2 (DYNC1LI2), peptidylprolyl isomerase D (PPID), vacuolar protein sorting 37 homolog B (S. cerevisiae) (VPS37B), adrenergic, beta, receptor kinase 1 (ADRBK1), DIS3 mitotic control homolog (S. cerevisiae) (DIS3), polymerase (RNA) I polypeptide A, 194 kDa (POLR1A), t-complex 1 (TCP1), plakophilin 3 (PKP3), La ribonucleoprotein domain family, member 1B (LARP1B), poly (ADP-ribose) polymerase 1 (PARP1), CD46 molecule, complement regulatory protein (CD46),

p21 protein (Cdc42/Rac)-activated kinase 2 (PAK2), ATP-binding cassette, sub-family E (OABP), member 1 (ABCE1), ubiquitin specific peptidase 14 (tRNA-guanine transglycosylase) (USP14), chaperonin containing TCP1, subunit 3 (gamma) (CCT3), Ran GTPase activating protein 1 (RANGAP1), deoxythymidylate kinase (thymidylate kinase) (DTYMK), N-myristoyltransferase 1 (NMT1), dynamin 1-like (DNM1L), interferon induced transmembrane protein 2 (1-8D) (IFITM2), fermitin family member 1 (FERMT1), tubulin folding cofactor D (TBCD), serine/arginine-rich splicing factor 10 (LOC100505793/SRSF10), STE20-like kinase (SLK), mucin 5AC, oligomeric mucus/gel-forming (MUCSAC/MUCSB), methionyl-tRNA synthetase (MARS), SMEK homolog 1, suppressor of mek1 (Dictyostelium) (SMEK1), high mobility group box 2 (HMGB2), non-POU domain containing, octamer-binding (NONO), transforming growth factor, beta-induced, 68 kDa (TGFBI), fibulin 2 (FBLN2), high density lipoprotein binding protein (HDLBP), collagen, type IV, alpha 2 (COL4A2), copine I (CPNE1), N(alpha)-acetyltransferase 50, NatE catalytic subunit (NAA50), LSM7 homolog, U6 small nuclear RNA associated (S. cerevisiae) (LSM7), structure specific recognition protein 1 (SSRP1), importin 8 (IP08), yippee-like 5 (Drosophila) (YPEL5), phosphoglucomutase 3 (PGM3), ring finger protein 40 (RNF40), structural maintenance of chromosomes 3 (SMC3), regenerating islet-derived family, member 4 (REG4), splicing factor 3a, subunit 3, 60 kDa (SF3A3), thrombospondin 1 (THBS1), chaperonin containing TCP1, subunit 6A (zeta 1) (CCT6A), PRP8 pre-mRNA processing factor 8 homolog (S. cerevisiae) (PRPF8), symplekin (SYMPK), far upstream element (FUSE) binding protein 1 (FUBP1), U2 small nuclear RNA auxiliary factor 1 (U2AF1), huntingtin (HTT), eukaryotic translation initiation factor 5B (EIFSB), nuclear autoantigenic sperm protein (histone-binding) (NASP), heterogeneous nuclear ribonucleoprotein K (HNRNPK), Y box binding protein 1 (YBX1), annexin A11 (ANXA11), RecQ protein-like (DNA helicase Q1-like) (RECQL), cortactin (CTTN), tubulin, beta 3 (TUBB3), pyrroline-5-carboxylate reductase-like (PYCRL), periplakin (PPL), phosphoglucomutase 2-like 1 (PGM2L1), chromosome 17 open reading frame 49 (C17orf49), mRNA turnover 4 homolog (S. cerevisiae) (MRTO4), methyltransferase like 1 (METTL1), squamous cell carcinoma antigen recognized by T cells 3 (SART3), S100 calcium binding protein A13 (S100A13), aminopeptidase-like 1 (NPEPL1), cyclin-dependent kinase 1 (CDK1), ubiquitin protein ligase E3 component n-recognin 1 (UBR1), Rho GTPase activating protein 18 (ARHGAP18), signal recognition particle 14 kDa (homologous Alu RNA binding protein) (SRP14), cathelicidin antimicrobial peptide (CAMP), splicing factor proline/glutamine-rich (SFPQ), RAS p21 protein activator (GTPase activating protein) 1 (RASA1), Ral GTPase activating protein, beta subunit (non-catalytic) (RALGAPB), laminin, beta 1 (LAMB1), RAB3 GTPase activating protein subunit 2 (non-catalytic) (RAB3GAP2), chaperonin containing TCP1, subunit 8 (theta) (CCT8), heterogeneous nuclear ribonucleoprotein L-like (HNRPLL), RAN binding protein 1 (RANBP1), kinetochore associated 1 (KNTC1), dyskeratosis congenita 1, dyskerin (DKC1), casein kinase 2, alpha 1 polypeptide (CSNK2A1), CAP-GLY domain containing linker protein 1 (CLIP1), chaperonin containing TCP1, subunit 2 (beta) (CCT2), tubulin tyrosine ligase-like family, member 12 (TTLL12), ataxia telangiectasia mutated (ATM), splicing factor 3a, subunit 1, 120 kDa (SF3A1), ribosomal protein S20 (RPS20), ubiquitin-conjugating enzyme E20 (UBE2O), translocated promoter region (to activated MET oncogene) (TPR), BRCA2 and CDKN1A interacting protein (BCCIP), gem (nuclear organelle) associated protein 5 (GEMIN5), ribonuclease P/MRP 30 kDa subunit (RPP30), loss of heterozygosity, 12, chromosomal region 1 (LOH12CR1), syntaxin binding protein 2 (STXBP2), ubiquitin-conjugating enzyme E2H (UBE2H), DIP2 disco-interacting protein 2 homolog B (Drosophila) (DIP2B), RAP1, GTP-GDP dissociation stimulator 1 (RAP1GDS1), heterogeneous nuclear ribonucleoprotein M (HNRNPM), LIM domain 7 (LMO7), RNA binding motif protein 25 (RBM25), aldehyde dehydrogenase 7 family, member A1 (ALDH7A1), cleavage and polyadenylation specific factor 1, 160 kDa (CPSF1), calponin 2 (CNN2), chaperonin containing TCP1, subunit 7 (eta) (CCT7), lysyl-tRNA synthetase (KARS), UDP-N-acteylglucosamine pyrophosphorylase 1 (UAP1), heat shock 70 kDa protein 4-like (HSPA4L), 138 kDa protein (138 kDa protein), thimet oligopeptidase 1 (THOP1), glutaredoxin 3 (GLRX3), phosphoglycerate dehydrogenase (PHGDH), CDV3 homolog (mouse) (CDV3), structural maintenance of chromosomes 4 (SMC4), RNA binding motif (RNP1, RRM) protein 3 (RBM3), hepatoma-derived growth factor (HDGF), heterogeneous nuclear ribonucleoprotein U (scaffold attachment factor A) (HNRNPU), nuclear receptor binding protein 1 (NRBP1), polymerase (RNA) I polypeptide B, 128 kDa (POLR1B), protein phosphatase 5, catalytic subunit (PPP5C), glucose-6-phosphate dehydrogenase (G6PD), arginase, liver (ARG1), 3-hydroxy-3-methylglutaryl-CoA synthase 1 (soluble) (HMGCS1), ubiquitin-like modifier activating enzyme 2 (UBA2), KIAA1033 (KIAA1033), annexin A4 (ANXA4), DEAD (Asp-Glu-Ala-Asp) box polypeptide 17 (DDX17), acidic (leucine-rich) nuclear phosphoprotein 32 family, member E (ANP32E), glucosamine (UDP-N-acetyl)-2-epimerase/N-acetylmannosamine kinase (GNE), KIAA0368 (KIAA0368), vacuolar protein sorting 4 homolog B (S. cerevisiae) (VPS4B), replication protein A1, 70 kDa (RPA1), eukaryotic translation initiation factor 2, subunit 1 alpha, 35 kDa (EIF2S1), eukaryotic translation initiation factor 3, subunit J (EIF3J), suppressor of Ty 6 homolog (S. cerevisiae) (SUPT6H), heat shock 105 kDa/110 kDa protein 1 (HSPH1), exportin 5 (XPO5), transcription elongation factor A (SII), 1 (TCEA1), Sjogren syndrome antigen B (autoantigen La) (SSB), AE binding protein 1 (AEBP1), LIM and cysteine-rich domains 1 (LMCD1), interleukin enhancer binding factor 3, 90 kDa (ILF3), WD repeat domain 61 (WDR61), N(alpha)-acetyltransferase 15, NatA auxiliary subunit (NAA15), serine/arginine-rich splicing factor 4 (SRSF4), ring finger protein 20 (RNF20), lactamase, beta 2 (LACTB2), NHP2 ribonucleoprotein homolog (yeast) (NHP2), chromosome 17 open reading frame 28 (C17orf28), CTP synthase II (CTPS2), fascin homolog 1, actin-bundling protein (Strongylocentrotus purpuratus) (FSCN1), tRNA nucleotidyl transferase, CCA-adding, 1 (TRNT1), splicing regulatory glutamine/lysine-rich protein 1 (SREK1),stromal antigen 1 (STAG1), oxysterol binding protein (OSBP), deoxyuridine triphosphatase (DUT), coiled-coil domain containing 25 (CCDC25), DEK oncogene (DEK), coiled-coil domain containing 72 (CCDC72), polymerase (RNA) II (DNA directed) polypeptide E, 25 kDa (POLR2E), phosphoserine phosphatase (PSPH), structural maintenance of chromosomes 1A (SMC1A), DEAD (Asp-Glu-Ala-Asp) box polypeptide 23 (DDX23), tRNA methyltransferase 11-2 homolog (S. cerevisiae) (TRMT112), COP9 constitutive photomorphogenic homolog subunit 2 (Arabidopsis) (COPS2), programmed cell death 5 (PDCD5), cyclin-dependent kinase 2 (CDK2), proteasome (prosome, macropain) 26S subunit, non-ATPase, 3 (PSMD3), RAN binding protein 2 (RANBP2), SERPINE1 mRNA binding protein 1 (SERBP1), O-linked N-acetylglucosamine (GlcNAc) transferase (UDP-N-acetylglucosamine:polypeptide-N-acetylglucosaminyl transferase) (OGT), non-SMC condensin I complex, subunit D2 (NCAPD2), SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily c, member 2 (SMARCC2), NOP10 ribonucleoprotein homolog (yeast) (NOP10), LPS-responsive vesicle trafficking, beach and anchor containing (LRBA), apoptosis inhibitor 5 (API5), signal recognition particle receptor (docking protein) (SRPR), transferrin receptor (p90, CD71) (TFRC), basic leucine zipper and W2 domains 2 (BZW2), ribonuclease, RNase A family, 3 (RNASE3), diazepam binding inhibitor (GABA receptor modulator, acyl-CoA binding protein) (DBI), FK506 binding protein 4, 59 kDa (FKBP4), chromosome 6 open reading frame 130 (C6orf130), cofactor of BRCA1 (COBRA1), flap structure-specific endonuclease 1 (FEN1), glucan (1,4-alpha-), branching enzyme 1 (GBE1), small nuclear ribonucleoprotein polypeptide B (SNRPB2), NSFL1 (p97) cofactor (p47) (NSFL1C), acyl-CoA thioesterase 7 (ACOT7), NOP2/Sun domain family, member 2 (NSUN2), chaperonin containing TCP1, subunit 4 (delta) (CCT4), kallikrein-related peptidase 6 (KLK6), glutaminyl-peptide cyclotransferase (QPCT), BCL2-associated athanogene 6 (BAG6), eukaryotic translation initiation factor 3, subunit C (EIF3C/EIF3CL), ATPase, H+ transporting, lysosomal 56/58 kDa, V1 subunit B2 (ATP6V1B2), matrix metallopeptidase 8 (neutrophil collagenase) (MMP8), proteasome (prosome, macropain) 26S subunit, ATPase, 5 (PSMC5), GTP cyclohydrolase I feedback regulator (GCHFR), poly(A) polymerase alpha (PAPOLA), hippocalcin-like 1 (HPCAL1), GTPase activating protein (SH3 domain) binding protein 1 (G3BP1), polymerase (RNA) Ill (DNA directed) polypeptide A, 155 kDa (POLR3A), superkiller viralicidic activity 2-like 2 (S. cerevisiae) (SKIV2L2), polymerase (RNA) II (DNA directed) polypeptide A, 220 kDa (POLR2A), collagen, type I, alpha 2 (COL1A2), fibrillarin (FBL), glutamyl-prolyl-tRNA synthetase (EPRS), ELAV (embryonic lethal, abnormal vision, Drosophila)-like 1 (Hu antigen R) (ELAVL1), nuclear cap binding protein subunit 2, 20 kDa (NCBP2), GCN1 general control of amino-acid synthesis 1-like 1 (yeast) (GCN1L1), histone acetyltransferase 1 (HAT1), stromal antigen 2 (STAG2), sorbitol dehydrogenase (SORD), REX2, RNA exonuclease 2 homolog (S. cerevisiae) (REXO2), heterogeneous nuclear ribonucleoprotein F (HNRNPF), thymopoietin (TMPO), ubiquitin specific peptidase 24 (USP24), KIAA1967 (KIAA1967), complement component 1, r subcomponent (C1R), annexin A7 (ANXA7), RuvB-like 2 (E. coli) (RUVBL2), acireductone dioxygenase 1 (ADI1), eukaryotic translation initiation factor 4A3 (EIF4A3), heterogeneous nuclear ribonucleoprotein U-like 2 (HNRNPUL2), ubiquitin protein ligase E3 component n-recognin 4 (UBR4), SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 2 (SMARCA2), cytochrome b5 reductase 2 (CYB5R2), splicing factor 3b, subunit 3, 130 kDa (SF3B3), G protein pathway suppressor 1 (GPS1), MAD2 mitotic arrest deficient-like 1 (yeast) (MAD2L1), phospholipase C, gamma 1 (PLCG1), eukaryotic translation initiation factor 4H (EIF4H), U2 small nuclear RNA auxiliary factor 2 (U2AF2), tankyrase 1 binding protein 1, 182 kDa (TNKS1BP1), transglutaminase 2 (C polypeptide, protein-glutamine-gamma-glutamyltransferase) (TGM2), heterogeneous nuclear ribonucleoprotein L (HNRNPL), inositol polyphosphate-5-phosphatase, 145 kDa (INPP5D), annexin A10 (ANXA10), BUD31 homolog (S. cerevisiae) (BUD31), phosphatidylinositol transfer protein, alpha (PITPNA), leucyl-tRNA synthetase (LARS), nicotinamide N-methyltransferase (NNMT), proteasome (prosome, macropain) 26S subunit, non-ATPase, 12 (PSMD12), v-crk sarcoma virus CT10 oncogene homolog (avian) (CRK), proteoglycan 2, bone marrow (natural killer cell activator, eosinophil granule major basic protein) (PRG2), versican (VCAN), exportin, tRNA (nuclear export receptor for tRNAs) (XPOT), EMG1 nucleolar protein homolog (S. cerevisiae) (EMG1), chromosome 11 open reading frame 73 (C11orf73), transportin 1 (TNPO1), latent transforming growth factor beta binding protein 2 (LTBP2), cold shock domain containing E1, RNA-binding (CSDE1), sulfiredoxin 1 (SRXN1), paraspeckle component 1 (PSPC1), ribosomal protein 53A (RPS3A), ISG15 ubiquitin-like modifier (ISG15), polymerase (RNA) II (DNA directed) polypeptide B, 140 kDa (POLR2B), general transcription factor IIi (GTF2I), NHP2 non-histone chromosome protein 2-like 1 (S. cerevisiae) (NHP2L1), proteasome (prosome, macropain) 26S subunit, non-ATPase, 10 (PSMD10), signal transducer and activator of transcription 1, 91 kDa (STAT1), elongation factor Tu GTP binding domain containing 1 (EFTUD1), mediator complex subunit 23 (MED23), eukaryotic translation initiation factor 2C, 2 (EIF2C2), RNA binding motif protein 4B (RBM4B), KIAA0664 (KIAA0664), core-binding factor, beta subunit (CBFB),

poly(A) binding protein, cytoplasmic 1 (PABPC1), nicotinamide phosphoribosyltransferase (NAMPT), cellular retinoic acid binding protein 2 (CRABP2), thyroid hormone receptor interactor 12 (TRIP12), DnaJ (Hsp40) homolog, subfamily C, member 9 (DNAJC9), StAR-related lipid transfer (START) domain containing 10 (STARD10), ring finger protein 213 (RNF213), eukaryotic translation initiation factor 2B, subunit 5 epsilon, 82 kDa (EIF2B5), bolA homolog 2 (E. coli) (BOLA2/BOLA2B), meningioma expressed antigen 5 (hyaluronidase) (MGEA5), Rab geranylgeranyltransferase, alpha subunit (RABGGTA), pyridoxal-dependent decarboxylase domain containing 1 (PDXDC1), exosome component 8 (EXOSC8), phosphoserine aminotransferase 1 (PSAT1), eukaryotic translation initiation factor 6 (EIF6), chromosome 16 open reading frame 13 (C16orf13), signal transducer and activator of transcription 3 (acute-phase response factor) (STAT3), EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1), defensin, alpha 6, Paneth cell-specific (DEFA6), pyrophosphatase (inorganic) 1 (PPA1), glutathione peroxidase 2 (gastrointestinal) (GPX2), unc-13 homolog D (C. elegans) (UNC13D), protein tyrosine phosphatase, non-receptor type 6 (PTPN6), myosin XVIIIA (MYO18A), fibulin 1 (FBLN1), ribosomal protein L19 (RPL19), diaphanous homolog 1 (Drosophila) (DIAPH1), MMS19 nucleotide excision repair homolog (S. cerevisiae) (MMS19), nudix (nucleoside diphosphate linked moiety X)-type motif 21 (NUDT21), splicing factor 3b, subunit 5, 10 kDa (SF3B5), SAP domain containing ribonucleoprotein (SARNP), ADP-ribosylation factor guanine nucleotide-exchange factor 2 (brefeldin A-inhibited) (ARFGEF2), guanylate binding protein 1, interferon-inducible (GBP1), HECT, UBA and WWE domain containing 1 (HUWE1), processing of precursor 7, ribonuclease P/MRP subunit (S. cerevisiae) (POP7), eukaryotic translation initiation factor 2B, subunit 3 gamma, 58 kDa (EIF2B3), N(alpha)-acetyltransferase 10, NatA catalytic subunit (NAA10), DnaJ (Hsp40) homolog, subfamily B, member 1 (DNAJB1), eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2), elongation factor Tu GTP binding domain containing 2 (EFTUD2), grancalcin, EF-hand calcium binding protein (GCA), neuronal cell adhesion molecule (NRCAM), DEAH (Asp-Glu-Ala-His) box polypeptide 16 (DHX16), small glutamine-rich tetratricopeptide repeat (TPR)-containing, alpha (SGTA), serine/threonine kinase 10 (STK10), smu-1 suppressor of mec-8 and unc-52 homolog (C. elegans) (SMU1), PRP4 pre-mRNA processing factor 4 homolog (yeast) (PRPF4), heterogeneous nuclear ribonucleoprotein D (AU-rich element RNA binding protein 1, 37 kDa) (HNRNPD), CTP synthase (CTPS), cleavage stimulation factor, 3′ pre-RNA, subunit 3, 77 kDa (CSTF3), cleavage stimulation factor, 3′ pre-RNA, subunit 1, 50 kDa (CSTF1), heterogeneous nuclear ribonucleoprotein U-like 1 (HNRNPUL1), ribosomal protein S5 (RPS5), protein tyrosine phosphatase, non-receptor type 11 (PTPN11), ladinin 1 (LAD1), component of oligomeric golgi complex 2 (COG2), cullin 2 (CUL2), ribosomal protein S17 (RPS17/RPS17L), proteasome (prosome, macropain) 26S subunit, non-ATPase, 4 (PSMD4), annexin A1 (ANXA1), annexin A2 (ANXA2), 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), azurocidin 1 (AZU1), baculoviral IAP repeat containing 6 (BIRC6), chitinase 3-like 1 (cartilage glycoprotein-39) (CHI3L1), carbamoyl-phosphate synthase 1, mitochondrial (CPS1), cathepsin G (CTSG), defensin, alpha 1 (DEFA1 (includes others)), deleted in malignant brain tumors 1 (DMBT1), elastase, neutrophil expressed (ELANE), integrin, alpha M (complement component 3 receptor 3 subunit) (ITGAM), lipocalin 2 (LCN2), lectin, galactoside-binding, soluble, 3 binding protein (LGALS3BP), minichromosome maintenance complex component 2 (MCM2), matrix metallopeptidase 9 (gelatinase B, 92 kDa gelatinase, 92 kDa type IV collagenase) (MMP9), myeloperoxidase (MPO), mucin 5AC, oligomeric mucus/gel-forming (MUC5AC/MUC5B), nuclear cap binding protein subunit 1, 80 kDa (NCBP1), neurofibromin 1 (NF1), olfactomedin 4 (OLFM4), PDS5, regulator of cohesion maintenance, homolog A (S. cerevisiae) (PDS5A), peptidoglycan recognition protein 1 (PGLYRP1), proteinase 3 (PRTN3), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), regenerating islet-derived 1 alpha (REG1A), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5), unc-45 homolog A (C. elegans) (UNC45A).

In one embodiment, the methods of the invention screen for one or more biomarkers, the presence of which in a sample is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer.

In an alternative embodiment, the methods of the invention screen for one or more biomarkers, the increased expression of which in a sample relative to a control sample is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer.

In a preferred embodiment, the methods of the invention screen for one or more biomarkers from the group defined in Table 2. The biomarkers provided in Table 2 represent a preferred subset of those defined in Table 1, the differential expression for which relative to control samples is considered statistically significant. Therefore, this group of biomarkers represents a panel of biomarkers from which any number of biomarkers may be selected for screening.

Thus, in one embodiment, the methods of the invention may screen for more than one biomarker from the group defined in Table 2, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers of the group defined in Table 2. In an alternative embodiment, the methods of the invention screen for more than ten of the biomarkers of the group defined in Table 2, for example, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, thirty, forty, fifty, sixty, seventy, eighty, ninety, one hundred of the biomarkers of the group defined in Table 2. Thus, the methods of the invention screen for the presence of or increased expression of one or more of: complement component C4B (Chido blood group) 2 (C4A/C4B); glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2); glucose-6-phosphate isomerase (GPI); transketolase (TKT); N-acylaminoacyl-peptide hydrolase (APEH); histone cluster 1, H4c (HIST4H4 (includes others)); Fatty acid-binding protein 5 (psoriasis-associated) (FABP5); hexosaminidase B (beta polypeptide) (HEXB); epithelial cell adhesion molecule (EPCAM); Nucleoside diphosphate kinase (NME1-NME2); Superoxide dismutase 2, mitochondrial (SOD2); Tu translation elongation factor, mitochondrial (TUFM); Glutathione synthetase (GSS); annexin A2 (ANXA2); ATP synthase, H+ transporting, mitochondrial F1 complex, beta polypeptide (ATP5B); 10 kDa heat shock protein (chaperonin 10) (HSPE1); glyoxalase I (GLO1); histone cluster 2, H2be (HIST2H2BE (includes others)); S100 calcium binding protein A4 (S100A4); S100 calcium binding protein A11 (S100A11); latexin (LXN); dehydrogenase/reductase (SDR family) member 11 (DHRS11); N-acetylglucosaminidase, alpha (NAGLU); Translin (TSN); Proteasome (prosome, macropain) subunit alpha type-4 (PSMA4); Proteasome (prosome, macropain) subunit alpha type-6 (PSMA6); ras-related C3 botulinum toxin substrate 1 (rho family, small GTP binding protein Rac1) (RAC1); Adenosylhomocysteinase (AHOY); fucosidase, alpha-L-1, tissue (FUCA1); S100 calcium binding protein P (S100P); Proteasome (prosome, macropain) subunit beta type-2 (PSMB2); X-prolyl aminopeptidase (aminopeptidase P) 1 (XPNPEP1); Keratin 18 (KRT18); Nuclear cap-binding protein subunit 1 80 kDa (NCBP1); mannosidase, alpha, class 2B, member 1 (MAN2B1); S100 calcium binding protein A6 (S100A6); valosin containing protein (VCP); quinolinate phosphoribosyltransferase (QPRT); major histocompatibility complex, class I, B (HLA-B); phosphoglycerate mutase 1 (brain) (PGAM1); ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3); serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10); myeloperoxidase (MPO); creatine kinase, mitochondrial 1B (CKMT1A/CKMT1B); proteinase 3 (PRTN3); elastase, neutrophil expressed (ELANE); MORC family CW-type zinc finger 1 (MORC1); ubiquitin B (UBB); phospholipase A2, group IIA (platelets, synovial fluid) (PLA2G2A); carbonic anhydrase IV (CA4); G elongation factor, mitochondrial 2 (GFM2); S100 calcium binding protein A7 (S100A7); Bactericidal permeability-increasing protein (BPI); collagen, type VI, alpha 5 (COL6A5); LIM homeobox 8 (LHX8); cysteine-rich secretory protein 3 (CRISP3); Azurocidin (AZU1); hemicentin 1 (HMCN1); Transglutaminase 3 (E polypeptide, protein-glutamine gamma-glutamyltransferase) (TGM3); CDC42 binding protein kinase alpha (DMPK-like) (CDC42BPA); Cathepsin G (CTSG); Resistin (RETN); methylmalonyl CoA mutase (MUT); armadillo repeat containing, X-linked 4 (ARMCX4); Integrin alpha-M (ITGAM); Calcium channel, voltage dependent, R-type alpha-1E subunit (CACNA1E); T-cell lymphoma invasion and metastasis 2 (TIAM2); HIR histone cell cycle regulation defective homolog A (S. cerevisiae) (HIRA); dopey family member 2 (DOPEY2); integrin beta 1 binding protein 3 (ITGB1BP3); Sodium channel, voltage-gated, type VII, alpha (SCN7A); Rab3C, member RAS oncogene family (RAB3C); chromosome 9 open reading frame 79 (C9orf79); nuclear factor of activated T-cells, calcineurin-dependent 4 (NFATC4); UDP-glucose glycoprotein glucosyltransferase 2 (UGGT2); Cornulin (CRNN); kielin/chordin-like protein (KCP); CD1E molecule (CD1E); coiled-coil domain-containing 18 (CCDC18); leukotriene A-4 hydrolase (LTA4H); albumin (ALB); alpha-2-macroglobulin (A2M); complement component 3 (C3); hemoglobin, beta (HBB); transferrin (TF); hemoglobin, alpha 1 (HBA1/HBA2); lactotransferrin (LTF); ceruloplasmin (ferroxidase) (CP); catalase (CAT); group-specific component (vitamin D-binding protein) (GC); serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1); fibrinogen gamma chain (FGG); S100 calcium binding protein A8 (S100A8); ferritin, light polypeptide (FTL); actin, beta (ACTB); fibronectin 1 (FN1); defensin, alpha 1 (DEFA1 (includes others)); serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1); retinol binding protein 4, plasma (RBP4); peroxiredoxin 2 (PRDX2); fibrinogen alpha chain (FGA); serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2); carbonic anhydrase II (CA2); orosomucoid 1 (ORM1/ORM2); lactate dehydrogenase A (LDHA); vitronectin (VTN); kininogen-1 (KNG1); actin, alpha, cardiac muscle 1 (ACTC1); leucine-rich alpha-2-glycoprotein 1 (LRG1); gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH); enolase 1, (alpha) (ENO1); profilin 1 (PFN1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 7 (SERPINA7); alpha-1-microglobulin/bikunin precursor (AMBP); lamin A/C (LMNA); apolipoprotein D (APOD); thyroid hormone receptor interactor 11 (TRIP11); complement component 4 binding protein, alpha (C4BPA); tropomyosin 4 (TPM4); filamin A, alpha (FLNA); haptoglobin (HP); hemopexin (HPX); hemoglobin, delta (HBD); fibrinogen beta chain (FGB); S100 calcium binding protein A9 (S100A9); complement component 5 (C5); solute carrier family 26, member 3 (SLC26A3); complement component 9 (C9); amyloid P component, serum (APCS); alpha-1-B glycoprotein (A1BG); complement C3-like (LOC100133511); inter-alpha (globulin) inhibitor H4 (plasma Kallikrein-sensitive glycoprotein) (ITIH4); complement component C8, alpha polypeptide (C8A); inter-alpha (globulin) inhibitor H1 (ITIH1).

In one embodiment, the method of the invention screens for one or more biomarkers capable of diagnosing or predicting CRC in an individual who tested negative in the fecal immunochemical test.

Thus, the methods of the invention may screen for one or more biomarkers selected from the group defined in Table 3. The biomarkers provided in Table 3 represent a preferred subset of those defined in Table 1, which have been found to be present in significantly higher levels in CRC samples which came back negative from the fecal immunochemical test. Thus, the group of Table 3 represents a panel of biomarkers from which any number of biomarkers may be selected for screening.

Thus, in one embodiment, the methods of the invention may screen for more than one biomarker from the group defined in Table 3, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers of the group defined in Table 3. In an alternative embodiment, the methods of the invention screen for more than ten of the biomarkers of the group defined in Table 3, for example, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, thirty, forty, fifty, sixty, seventy, eighty, ninety, one hundred of the biomarkers of the group defined in Table 3. Thus, the methods of the invention may screen for the presence of or increased expression of one or more of: complement component C4B (Chido blood group) 2 (C4A/C4B); glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2); glucose-6-phosphate isomerase (GPI); transketolase (TKT); N-acylaminoacyl-peptide hydrolase (APEH); hexosaminidase B (beta polypeptide) (HEXB); epithelial cell adhesion molecule (EPCAM); NME1-NME2 readthrough (NME1-NME2); Tu translation elongation factor, mitochondrial (TUFM); Glutathione synthetase (GSS); glyoxalase I (GLO1); latexin (LXN); Proteasome (prosome, macropain) subunit alpha type-4 (PSMA4); fucosidase, alpha-L-1, tissue (FUCA1); Keratin 18 (KRT18); Nuclear cap-binding protein subunit 1 80 kDa (NCBP1); mannosidase, alpha, class 2B, member 1 (MAN2B1); S100 calcium binding protein A6 (S100A6); major histocompatibility complex, class I, B (HLA-B); ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3); serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10); myeloperoxidase (MPO); proteinase 3 (PRTN3); phospholipase A2, group IIA (platelets, synovial fluid) (PLA2G2A); carbonic anhydrase IV (CA4); G elongation factor, mitochondrial 2 (GFM2); S100 calcium binding protein A7 (S100A7); Bactericidal permeability-increasing protein (BPI); collagen, type VI, alpha 5 (COL6A5); LIM homeobox 8 (LHX8); Azurocidin (AZU1); hemicentin 1 (HMCN1); methylmalonyl CoA mutase (MUT); armadillo repeat containing, X-linked 4 (ARMCX4); Integrin alpha-M (ITGAM); Calcium channel, voltage dependent, R-type alpha-1E subunit (CACNA1E); T-cell lymphoma invasion and metastasis 2 (TIAM2); HIR histone cell cycle regulation defective homolog A (S. cerevisiae) (HIRA); dopey family member 2 (DOPEY2); Sodium channel, voltage-gated, type VII, alpha (SCN7A); chromosome 9 open reading frame 79 (C9orf79); UDP-glucose glycoprotein glucosyltransferase 2 (UGGT2); Cornulin (CRNN); coiled-coil domain-containing 18 (CCDC18); alpha-2-macroglobulin (A2M); complement component 3 (C3); hemoglobin, beta (HBB); transferrin (TF); hemoglobin, alpha 1 (HBA1/HBA2); lactotransferrin (LTF); ceruloplasmin (ferroxidase) (CP); catalase (CAT); fibrinogen gamma chain (FGG); S100 calcium binding protein A8 (S100A8); ferritin, light polypeptide (FTL); fibronectin 1 (FN1); defensin, alpha 1 (DEFA1 (includes others)); serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1); retinol binding protein 4, plasma (RBP4); peroxiredoxin 2 (PRDX2); serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2); lactate dehydrogenase (LDHA); vitronectin (VTN); kininogen-1 (KNG1); leucine-rich alpha-2-glycoprotein 1 (LRG1); gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 7 (SERPINA7); alpha-1-microglobulin/bikunin precursor (AMBP); thyroid hormone receptor interactor 11 (TRIP11); complement component 4 binding protein, alpha (C4BPA); filamin A, alpha (FLNA); hemopexin (HPX); S100 calcium binding protein A9 (S100A9); complement component 5 (C5); solute carrier family 26, member 3 (SLC26A3); complement component 9 (C9); amyloid P component, serum (APCS); complement C3-like (LOC100133511); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 (SERPINA3); Cathepsin S (CTSS); Cytidine deaminase (CDA); Shugoshin-like 2 (SGOL2); serpin peptidase inhibitor, clade B (ovalbumin), member 3 (SERPINB3); hect domain and RLD 2 (HERC2); triosephosphate isomerase 1 (TPI1); Isoform 1 of collagen, type IV, alpha 3 (Goodpasture antigen) binding protein (COL4A3BP); ribonuclease T2 (RNASET2); Glutathione reductase (GSR); spinster homolog 3 (Drosophila) (SPNS3); cDNA FLJ60317, highly similar to Aminoacylase-1 (ACY1); serpin peptidase inhibitor, clade B (ovalbumin), member 8 (SERPINB8); Branched-chain-amino-acid aminotransferase (BCAT2); sialic acid acetylesterase (SIAE); peptidase D (PEPD); major histocompatibility complex, class II, DR beta 5 (HLA-DRB5); SET domain containing 2 (SETD2); hect (homologous to the E6-AP (UBE3A) carboxyl terminus) domain and RCC1 (CHC1)-like domain (RLD) 1 (HERC1); isocitrate dehydrogenase 1 (NADP+), soluble (IDH1); cathepsin C (CTSC); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 (SERPINA1); spectrin repeat containing, nuclear envelope 1 (SYNE1); Ankyrin repeat domain-containing protein 35 (ANKRD35); NIMA (never in mitosis gene a)-related kinase 10 (NEK10); inter-alpha (globulin) inhibitor H2 (ITIH2); acyl-CoA dehydrogenase, very long chain (ACADVL); Nebulin (NEB); Zymogen granule membrane protein 16 (ZG16); Vinculin (VCL); Isoform 2 of Dedicator of cytokinesis protein 4 (DOCK4); hypothetical protein LOC643677 (C13orf40); Uncharacterized protein KIAA1797 (KIAA1797); baculoviral IAP repeat-containing 6 (BIRC6); Transaldolase 1 (TALDO1); Taste receptor type 2 member 42 (TAS2R42); chitinase 1 (chitotriosidase) (CHIT1); quiescin Q6 sulfhydryl oxidase 1 (QSOX1); bone marrow stromal cell antigen 12 (BST1); Bleomycin hydrolase (BLMH); Lysozyme C (LYZ).

In one embodiment, the method of the invention screens for one or more biomarkers having a higher discriminatory power than haemoglobin. In other words, the one or more biomarkers have a higher sensitivity and higher specificity than haemoglobin in detecting the presence of advanced adenoma or adenocarcinoma in a sample.

Thus, in one embodiment, the method of the invention screens for one or more biomarker, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers selected from the group consisting of: S100 calcium binding protein A8 (S100A8), complement component C4B (Chido blood group) 2 (C4A/C4B), transferrin (TF), alpha-2-macroglobulin (A2M), S100 calcium binding protein A9 (S100A9), proteinase 3 (PRTN3), Azurocidin (AZU1), lactotransferrin (LTF), hemopexin (HPX) and defensin, alpha 1 (DEFA1).

Thus, in one embodiment, the method of the invention screens for one or more biomarker, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers selected from the group consisting of: annexin A2 (ANXA2), azurocidin 1 (AZU1), cathepsin G (CTSG), defensin, alpha 1 (DEFA1 includes others)), elastase, neutrophil expressed (ELANE), integrin, alpha M (complement component 3 receptor 3 subunit) (ITGAM), myeloperoxidase (MPO), nuclear cap binding protein subunit 1, 80 kDa (NCBP1), proteinase 3 (PRTN3), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), annexin A1 (ANXA1), baculoviral IAP repeat containing 6 (BIRC6), minichromosome maintenance complex component 2 (MCM2), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5).

Thus, in a further embodiment, the method of the invention screens for one or more biomarker, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers selected from the group consisting of: alpha-1-B glycoprotein (A1BG), alpha-2-macroglobulin (A2M), actin, beta (ACTB), actin, alpha, ardiac muscle 1 (ACTC1), albumin (ALB), amyloid P component, serum (APCS), N-cylaminoacyl-peptide hydrolase (APEH), apolipoprotein D (APOD), azurocidin 1 (AZU1), complement component 3 (C3), complement component 4B (Chido blood group) (C4A/C4B), complement component 5 (C5), carbonic anhydrase II (CA2), carbonic anhydrase IV (CA4), catalase (CAT), ceruloplasmin (ferroxidase) (CP), cathepsin G (CTSG), defensin, alpha 1 (DEFA1(includesothers)), elastase, neutrophil expressed (ELANE), enolase 1, (alpha) (ENO1), filamin A, alpha (FLNA), fibronectin 1 (FN1), ferritin, light polypeptide (FTL), fucosidase, alpha-L-1, tissue (FUCA1), gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH), glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2), glucose-6-phosphate isomerase (GPI), glutathione synthetase (GSS), hemoglobin, alpha 1 (HBA1/HBA2), hemoglobin, beta (HBB), hemoglobin, delta (HBD), hexosaminidase B (beta polypeptide) (HEXB), histone cluster 1, H4c (HIST4H4(includesothers)), major histocompatibility complex, class I, B (HLA-B), haptoglobin (HP), hemopexin (HPX), heat shock 10 kDa protein 1 (chaperonin 10) (HSPE1), lactate dehydrogenase A (LDHA), complement C3-like (LOC100133511), leucine-rich alpha-2-glycoprotein 1 (LRG1), lactotransferrin (LTF), latexin (LXN), mannosidase, alpha, class 2B, member 1 (MAN2B1), myeloperoxidase (MPO), N-acetylglucosaminidase, alpha (NAGLU), orosomucoid 1 (ORM1/ORM2), profilin 1 (PFN1), phospholipase A2, group IIA (platelets, synovial fluid) (PLA2G2A), proteinase 3 (PRTN3), proteasome (prosome, macropain) subunit, alpha type, 4 (PSMA4), proteasome (prosome, macropain) subunit, alpha type, 6 (PSMA6), proteasome (prosome, macropain) subunit, beta type, 2 (PSMB2), retinol binding protein 4, plasma (RBP4), resistin (RETN), S100 calcium binding protein A8 (S100A8), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1), serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2), serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1), superoxide dismutase 2, mitochondrial (SOD2), transferrin (TF), transketolase (TKT), ubiquitin B (UBB), Unmapped by Ingenuity (IP100884078), Unmapped by Ingenuity (26kDaprotein, IP100942608), alpha-2-macroglobulin-like 1 (A2ML1), acyl-CoA dehydrogenase, very long chain (ACADVL), Unmapped by Ingenuity (ACY1), alkaline phosphatase, liver/bone/kidney (ALPL), bleomycin hydrolase (BLMH), bone marrow stromal cell antigen 1 (BST1), calcium channel, voltage-dependent, L type, alpha 1D subunit (CACNA1D), creatine kinase, mitochondrial 1B (CKMT1A/CKMT1B), cathepsin S (CTSS), fumarylacetoacetate hydrolase (fumarylacetoacetase) (FAH), guanine deaminase (GDA), glutathione reductase (GSR), heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1), heat shock 70 kDa protein (HSPA8), keratin 6C (KRT6C), keratin 6C (KRT6C), lipocalin 1 (tear prealbumin) (LCN1), lysozyme (LYZ), minichromosome maintenance complex component 2 (MCM2), peptidase D (PEPD), phospholipase B domain containing 1 (PLBD1), proteasome (prosome, macropain) subunit, alpha type, 2 (PSMA2), proteasome (prosome, macropain) subunit, beta type, 1 (PSMB1), proteasome (prosome, macropain) subunit, beta type, 6 (PSMB6), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), ribonuclease T2 (RNASET2), serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 (SERPINA1), serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 (SERPINA3), serpin peptidase inhibitor, clade B (ovalbumin), member 3 (SERPINB3), sialic acid acetylesterase (SIAE), triosephosphate isomerase 1 (TPI1), zymogen granule protein 16 homolog (rat) (ZG16).

In one embodiment, the method of the invention screens for one or more biomarkers, the presence of which in a sample is indicative that the individual is at risk of suffering from or is suffering from colorectal cancer, which biomarkers have been selected on the basis of correlation with secretomes of colon tumor tissue.

Thus, in one embodiment, the method of the invention screens for one or more biomarker, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers selected from the group defined in Table 7. The biomarkers provided in Table 7 represent a preferred subset of those defined in Table 6, which have been found to be expressed both in tumor issue and stool samples. Thus, the group of Table 7 represents a panel of biomarkers from which any number of biomarkers may be selected for screening. Thus, in a further embodiment, the method of the invention screens for one or more biomarker, for example two, three, four, five, six, seven, eight, nine, ten of the biomarkers selected from the group consisting of: annexin A2 (ANXA2), azurocidin 1 (AZU1), cathepsin G (CTSG), defensin, alpha 1 (DEFA1 (includes others)), elastase, neutrophil expressed (ELANE), integrin, alpha M (complement component 3 receptor 3 subunit) (ITGAM), myeloperoxidase (MPO), nuclear cap binding protein subunit 1, 80 kDa (NCBP1), proteinase 3 (PRTN3), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), annexin A1 (ANXA1), baculoviral IAP repeat containing 6 (BIRC6), minichromosome maintenance complex component 2 (MCM2), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5).

Preferably, the method of the invention has an accuracy of at least 75%, for example 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% accuracy.

Preferably, the method of the invention has a sensitivity of at least 75%, for example 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sensitivity.

Preferably, the method of the invention has a specificity of at least 75%, for example 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% specificity.

By “accuracy” it is meant the proportion of correct outcomes of a diagnosis, by “sensitivity” it is meant the proportion of all positive diagnoses that are correctly classified as positives, and by “specificity” it is meant the proportion of all negative diagnoses that are correctly classified as negatives. The methods of the invention may be verified by subsequent colonoscopy.

Screening Methods

In one embodiment, the biological sample, for example a stool sample, may be screened for the one or more biomarkers using (targeted) mass spectrometry.

Protein marker discovery and screening by LC-MS/MS and hypothesis-based protein marker detection by SRM-MS has been applied before on murine and human stool samples, respectively (Ang C S, Nice E C. J Proteome Res 2010; 9: 4346-55; Ang C S, et al. Electrophoresis 2011; 32: 1926-38; Ang C S, et al. J Chromatogr A 2010; 1217: 3330-40).

Despite the complexity of stool material, the present invention has successfully detected several of these previously reported human stool proteins. In addition, the number of included samples and corresponding identified proteins in the current study exceeds that of previous studies. Therefore the data presented here is the largest overview of the human stool proteome to date.

While sample screening by the mass spectrometric techniques described herein has been proven to be effective, a preferred screening method comprises an antibody based screening assay. The fecal immunochemical test (FIT) comprises an antibody based screening assay and so an antibody based screening assay for one or more of the biomarkers identified in the present application provides a complementary screen which can be readily incorporated into the existing FIT assay.

Thus, in one embodiment of the methods of the invention, the biological sample is screened for the one or more biomarkers using a binding agent capable of binding to the one or more biomarkers.

Binding agents (also referred to as binding molecules) can be selected from a library, based on their ability to bind a given motif, as discussed below.

In one embodiment, the first binding agent is an antibody or a fragment thereof. Thus, a fragment may contain one or more of the variable heavy (V_H) or variable light (V_L) domains. For example, the term antibody fragment includes Fab-like molecules (Better et al Science 1988; 240, 1041); Fv molecules (Skerra et al Science 1988; 240, 1038); single-chain Fv (ScFv) molecules where the V_Hand V_Lpartner domains are linked via a flexible oligopeptide (Bird et al Science 1988; 242,423; Huston et al Proc. Natl. Acad. Sci. USA 1988; 85, 5879) and single domain antibodies (dAbs) comprising isolated V domains (Ward et al Nature 1989; 341,544).

The term “antibody variant” includes any synthetic antibodies, recombinant antibodies or antibody hybrids, such as but not limited to, a single-chain antibody molecule produced by phage-display of immunoglobulin light and/or heavy chain variable and/or constant regions, or other immunointeractive molecule capable of binding to an antigen in an immunoassay format that is known to those skilled in the art.

A general review of the techniques involved in the synthesis of antibody fragments which retain their specific binding sites is to be found in Winter & Milstein Nature 1991; 349, 293-299.

In one embodiment, the antibody or fragment thereof is a recombinant antibody or fragment thereof (such as an scFv). By “ScFv molecules” it is meant molecules wherein the V_Hand V_Lpartner domains are linked via a flexible oligopeptide.

The advantages of using antibody fragments, rather than whole antibodies, are several-fold. Effector functions of whole antibodies, such as complement binding, are removed. Fab, Fv, ScFv and dAb antibody fragments can all be expressed in and secreted from E. coli, thus allowing the facile production of large amounts of the said fragments.

Whole antibodies, and F(ab′)₂fragments are “bivalent”. By “bivalent” it is meant that the said antibodies and F(ab′)₂fragments have two antigen combining sites. In contrast, Fab, Fv, ScFv and dAb fragments are monovalent, having only one antigen combining site.

The antibodies may be monoclonal or polyclonal. Suitable antibodies may be prepared by known techniques and need no further discussion.

Additionally or alternatively the binding agent may be an aptamer. Suitable aptamers may be prepared by known techniques and need no further discussion.

In one embodiment, the one or more biomarker(s) in the test sample is labelled with a detectable moiety. In one embodiment, the one or more biomarker(s) in the control sample is labelled with a detectable moiety (which may be the same or different from the detectable moiety used to label the test sample).

A “detectable moiety” is one which may be detected and the relative amount and/or location of the moiety (for example, the location on an array) determined.

Detectable moieties are well known in the art. A detectable moiety may be a fluorescent and/or luminescent and/or chemiluminescent moiety which, when exposed to specific conditions, may be detected. For example, a fluorescent moiety may need to be exposed to radiation (i.e. light) at a specific wavelength and intensity to cause excitation of the fluorescent moiety, thereby enabling it to emit detectable fluorescence at a specific wavelength that may be detected.

Alternatively, the detectable moiety may be an enzyme which is capable of converting a (preferably undetectable) substrate into a detectable product that can be visualised and/or detected. Examples of suitable enzymes are discussed in more detail below in relation to, for example, ELISA assays.

Alternatively, the detectable moiety may be a radioactive label, which may be incorporated by methods well known in the art.

Arrays

In one embodiment, the methods of the present invention may be carried out on an array.

Arrays per se are well known in the art. Typically they are formed of a linear or two-dimensional structure having spaced apart (i.e. discrete) regions (“spots”), each having a finite area, formed on the surface of a solid support. An array can also be a bead structure where each bead can be identified by a molecular code or colour code or identified in a continuous flow. Analysis can also be performed sequentially where the sample is passed over a series of spots each adsorbing the class of molecules from the solution.

The solid support is typically glass or a polymer, the most commonly used polymers being cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride or polypropylene. The solid supports may be in the form of tubes, beads, discs, silicon chips, microplates, polyvinylidene difluoride (PVDF) membrane, nitrocellulose membrane, nylon membrane, other porous membrane, non-porous membrane (e.g. plastic, polymer, perspex, silicon, amongst others), a plurality of polymeric pins, or a plurality of microtitre wells, or any other surface suitable for immobilising proteins, antibodies and other suitable molecules and/or conducting an immunoassay.

The binding processes are well known in the art and generally consist of cross-linking covalently binding or physically adsorbing a protein molecule, antibody or the like to the solid support. By using well-known techniques, such as contact or non-contact printing, masking or photolithography, the location of each spot can be defined.

Once suitable binding molecules (discussed above) have been identified and isolated, the skilled person can manufacture an array using methods well known in the art of molecular biology.

In one embodiment, the screening may comprise using an assay comprising a second binding agent capable of binding to the one or more biomarkers, the second binding agent having a detectable moiety.

In one embodiment, the second binding agent is an antibody or a fragment thereof (for example, as described above in relation to the first binding agent).

Typically, the assay is an ELISA (Enzyme Linked Immunosorbent Assay) which usually involves the use of enzymes which give a coloured reaction product, usually in solid phase assays. Enzymes such as horseradish peroxidase and phosphatase have been widely employed. A way of amplifying the phosphatase reaction is to use NADP as a substrate to generate NAD which now acts as a coenzyme for a second enzyme system. Pyrophosphatase from Escherichia coli provides a good conjugate because the enzyme is not present in tissues, is stable and gives a good reaction colour. Chemi-luminescent systems based on enzymes such as luciferase can also be used.

Conjugation with the vitamin biotin is also employed used since this can readily be detected by its reaction with enzyme-linked avidin or streptavidin to which it binds with great specificity and affinity.

It will be appreciated by persons skilled in the art that there is a degree of fluidity in the biomarker composition of the signatures of the invention. Thus, different combinations of the biomarkers may be equally useful in the diagnosis, prognosis and/or characterisation of colorectal cancer. In this way, each biomarker (either alone or in combination with one or more other biomarkers) makes a contribution to the signature.

Compounds and Methods for Treating CRC

The identification of the biomarkers as defined in Table 1 and/or Table 6 allows not only the detection of advanced colonic adenomas and colonic adenocarcinomas (colorectal cancer), but enables also methods of treating colorectal cancers as defined in the fourth aspect of the invention, and also provides for compounds for use in methods of treating colorectal cancers as defined in the fifth aspect of the invention.

While early diagnosis of colorectal cancer often allows for curative surgical removal of the tumour, later diagnosis may result in a (chemo)therapeutic treatment instead. Therapeutic agents used to treat colorectal cancer include monoclonal antibodies, small molecule inhibitors and chemotherapeutic agents.

Typical therapeutic monoclonal antibodies include but are not limited to bevacizumab, cetuximab or panitumumab. Typical small molecule inhibitors include but are not limited to erlotinib, sorafenib or alisertib. Typical chemotherapeutic agents include but are not limited to 5-FU, capecitabine, irinotecan oxaliplatin, or leucovorin or any combination thereof. Combination therapies of, for example, a therapeutic monoclonal antibody and a small molecule inhibitor may be used. Thus, any combination of two or more of a monoclonal antibody, a small molecule inhibitor and a chemotherapeutic agent is envisaged.

Kit for Performing the Method

The kit for performing the method according to the invention may be selected from any suitable assay and data processing apparatus and equipments.

The suitable selection will be well within the ability of those skilled in the art, and further description is not necessary here.

“Comprising”

The term “comprising” and related terms herein is to be interpreted as embracing “consisting essentially of” and “consisting of”, these two expressions being interchangeable with “comprising” in all definitions and discussion in this patent in order to specify alternative extents of exclusion of unspecified elements additional to the recited elements.

The term “comprising” and related expressions means “including” and therefore leaves open the option of including unspecified elements, whether essential or not. The term “consisting essentially of” and related expressions permits the presence of elements that do not materially affect the basic and novel or functional characteristic(s) of that embodiment of the invention. The term “consisting of” and related expressions means “consisting only of” and therefore excludes any element not recited in that description of the embodiment.

BRIEF DESCRIPTION OF THE FIGURES

The invention shall now be further described by the following example with reference to the attached figures. The example is provided by way of example only, without any intended limitation of the scope of the invention. All cited references are incorporated herein by reference in their entireties.

FIG. 1A shows the number of human proteins detected in stool samples from 12 CRC patients and from 10 control patients. The Venn diagram shows the number of proteins detected in CRC or control stool samples and their overlap;

FIG. 1B shows the subcellular location of the identified human proteins. The pie chart shows the percentage of different subcellular locations of the 830 human proteins that were identified in 22 stool samples. The ratios were similar for CRC patients and control patients;

FIG. 2A shows a scatterplot of protein quantifications of hemoglobin alpha as measured by LC-MS/MS compared to hemoglobin quantification by FIT of individual stool samples;

FIG. 2B shows a scatterplot of protein quantifications of hemoglobin beta as measured by LC-MS/MS compared to hemoglobin quantification by FIT of individual stool samples;

FIG. 2C shows a scatterplot of protein quantifications of hemoglobin alpha and beta as measured by LC-MS/MS of individual stool samples;

FIG. 2D shows a scatterplot of protein quantifications of S100A8 and S100A9 as measured by LC-MS/MS of individual stool samples. Filled circles represent stool samples from subjects with CRC (n=12), crosses represent stool samples from subjects without colon neoplasia (n=10); and

FIG. 3 shows a scatterplot of protein quantifications of Complement component 4B measured by LC-MS/MS in relation to FIT values of individual stool samples. The dotted line shows the cut-off of 75 ng/ml for FIT. A FIT value less than 75 ng/ml is regarded as a negative test result (Van Veen, NTG, 2009). Filled circles represent stool samples from subjects with CRC (n=12), crosses represent stool samples from subjects without colon neoplasia (n=10).

FIGS. 4 and 5 show bar graphs with the mean of the areas under the curves of all transitions of each peptide in triplicate analyses. One to five different peptides per protein are depicted. Peptides were measured in triplicate in pools from stool samples from controls (N, n=5), non-advanced adenoma patients (A, n=5), advanced adenoma patients (AA, n=5) and CRC patients (CRC, n=5). Error bars represent standard deviations.

TABLES

The following tables list the biomarkers according to the invention, and may continue over several pages.

TABLE 1

Biomarkers indicative of colorectal cancer.

Accession

Number
Gene Symbol
Entrez Gene Name

IPI00892604
C4A/C4B
complement component 4B (Chido blood

group)

IPI00018206
GOT2
glutamic-oxaloacetic transaminase 2,

mitochondrial (aspartate aminotransferase 2)

IPI00027497
GPI
glucose-6-phosphate isomerase

IPI00643920
TKT
transketolase

IPI00337741
APEH
N-acylaminoacyl-peptide hydrolase

IPI00453473
HIST4H4 (includes
histone cluster 1, H4c

others)

IPI00007797
FABP5
fatty acid binding protein 5 (psoriasis-

associated)

IPI00012585
HEXB
hexosaminidase B (beta polypeptide)

IPI00296215
EPCAM
epithelial cell adhesion molecule

IPI00604590
NME1-NME2
NME1-NME2 readthrough

IPI00022314
SOD2
superoxide dismutase 2, mitochondrial

IPI00027107
TUFM
Tu translation elongation factor, mitochondrial

IPI00010706
GSS
glutathione synthetase

IPI00418169
ANXA2
annexin A2

IPI00303476
ATP5B
ATP synthase, H+ transporting, mitochondrial

F1 complex, beta polypeptide

IPI00220362
HSPE1
heat shock 10 kDa protein 1 (chaperonin 10)

IPI00220766
GLO1
glyoxalase I

IPI00003935
HIST2H2BE
histone cluster 2, H2be

(includes others)

IPI00032313
S100A4
S100 calcium binding protein A4

IPI00013895
S100A11
S100 calcium binding protein A11

IPI00106687
LXN
latexin

IPI00034280
DHRS11
dehydrogenase/reductase (SDR family)

member 11

IPI00008787
NAGLU
N-acetylglucosaminidase, alpha

IPI00018768
TSN
translin

IPI00299155
PSMA4
proteasome (prosome, macropain) subunit,

alpha type, 4

IPI00029623
PSMA6
proteasome (prosome, macropain) subunit,

alpha type, 6

IPI00010271
RAC1
ras-related C3 botulinum toxin substrate 1 (rho

family, small GTP binding protein Rac1)

IPI00012007
AHCY
adenosylhomocysteinase

IPI00385751
FUCA1
fucosidase, alpha-L-1, tissue

IPI00017526
S100P
S100 calcium binding protein P

IPI00028006
PSMB2
proteasome (prosome, macropain) subunit,

beta type, 2

IPI00793375
XPNPEP1
X-prolyl aminopeptidase (aminopeptidase P) 1,

soluble

IPI00554788
KRT18
keratin 18

IPI00019380
NCBP1
nuclear cap binding protein subunit 1, 80 kDa

IPI00012989
MAN2B1
mannosidase, alpha, class 2B, member 1

IPI00027463
S100A6
S100 calcium binding protein A6

IPI00022774
VCP
valosin-containing protein

IPI00300086
QPRT
quinolinate phosphoribosyltransferase

IPI00472073
HLA-B
major histocompatibility complex, class I, B

IPI00453476
PGAM1
phosphoglycerate mutase 1 (brain)

IPI00020999
ENPP3
ectonucleotide

pyrophosphatase/phosphodiesterase 3

IPI00010304
SERPINB10
serpin peptidase inhibitor, clade B (ovalbumin),

member 10

IPI00007244
MPO
myeloperoxidase

IPI00877726
CKMT1A/CKMT1B
creatine kinase, mitochondrial 1B

IPI00027409
PRTN3
proteinase 3

IPI00027769
ELANE
elastase, neutrophil expressed

IPI00004362
MORC1
MORC family CW-type zinc finger 1

IPI00719280
UBB
ubiquitin B

IPI00026962
PLA2G2A
phospholipase A2, group IIA (platelets, synovial

fluid)

IPI00027466
CA4
carbonic anhydrase IV

IPI00071703
GFM2
G elongation factor, mitochondrial 2

IPI00219806
S100A7
S100 calcium binding protein A7

IPI00827847
BPI
bactericidal/permeability-increasing protein

IPI00176125
COL6A5
collagen, type VI, alpha 5

IPI00470355
LHX8
LIM homeobox 8

IPI00942117
CRISP3
cysteine-rich secretory protein 3

IPI00022246
AZU1
azurocidin 1

IPI00045512
HMCN1
hemicentin 1

IPI00300376
TGM3
transglutaminase 3 (E polypeptide, protein-

glutamine-gamma-glutamyltransferase)

IPI00640468
CDC42BPA
CDC42 binding protein kinase alpha (DMPK-

like)

IPI00028064
CTSG
cathepsin G

IPI00006988
RETN
resistin

IPI00024934
MUT
methylmalonyl CoA mutase

IPI00552983
ARMCX4
armadillo repeat containing, X-linked 4

IPI00217987
ITGAM
integrin, alpha M (complement component 3

receptor 3 subunit)

IPI00165045
CACNA1E
calcium channel, voltage-dependent, R type,

alpha 1E subunit

IPI00018363
TIAM2
T-cell lymphoma invasion and metastasis 2

IPI00217560
HIRA
HIR histone cell cycle regulation defective

homolog A (S. cerevisiae)

IPI00294653
DOPEY2
dopey family member 2

IPI00004401
ITGB1BP3
integrin beta 1 binding protein 3

IPI00300117
SCN7A
sodium channel, voltage-gated, type VII, alpha

IPI00061114
RAB3C
RAB3C, member RAS oncogene family

IPI00168442
C9orf79
chromosome 9 open reading frame 79

IPI00385215
NFATC4
nuclear factor of activated T-cells, cytoplasmic,

calcineurin-dependent 4

IPI00024467
UGGT2
UDP-glucose glycoprotein glucosyltransferase 2

IPI00297056
CRNN
cornulin

IPI00914663
KCP
kielin/chordin-like protein

IPI00418592
CD1E
CD1e molecule

IPI00642206
CCDC18
coiled-coil domain containing 18

IPI00514090
LTA4H
leukotriene A4 hydrolase

IPI00745872
ALB
albumin

IPI00478003
A2M
alpha-2-macroglobulin

IPI00783987
C3
complement component 3

IPI00654755
HBB
hemoglobin, beta

IPI00022463
TF
transferrin

IPI00410714
HBA1/HBA2
hemoglobin, alpha 1

IPI00298860
LTF
lactotransferrin

IPI00017601
CP
ceruloplasmin (ferroxidase)

IPI00465436
CAT
catalase

IPI00555812
GC
group-specific component (vitamin D binding

protein)

IPI00032179
SERPINC1
serpin peptidase inhibitor, clade C

(antithrombin), member 1

IPI00219713
FGG
fibrinogen gamma chain

IPI00007047
S100A8
S100 calcium binding protein A8

IPI00375676
FTL
ferritin, light polypeptide

IPI00021439
ACTB
actin, beta

IPI00022418
FN1
fibronectin 1

IPI00005721
DEFA1 (includes
defensin, alpha 1

others)

IPI00291866
SERPING1
serpin peptidase inhibitor, clade G (C1

inhibitor), member 1

IPI00022420
RBP4
retinol binding protein 4, plasma

IPI00027350
PRDX2
peroxiredoxin 2

IPI00021885
FGA
fibrinogen alpha chain

IPI00029863
SERPINF2
serpin peptidase inhibitor, clade F (alpha-2

antiplasmin, pigment epithelium derived factor),

member 2

IPI00218414
CA2
carbonic anhydrase II

IPI00020091
ORM1/ORM2
orosomucoid 1

IPI00217966
LDHA
lactate dehydrogenase A

IPI00298971
VTN
vitronectin

IPI00215894
KNG1
kininogen 1

IPI00023006
ACTC1
actin, alpha, cardiac muscle 1

IPI00022417
LRG1
leucine-rich alpha-2-glycoprotein 1

IPI00023728
GGH
gamma-glutamyl hydrolase (conjugase,

folylpolygammaglutamyl hydrolase)

IPI00465248
ENO1
enolase 1, (alpha)

IPI00216691
PFN1
profilin 1

IPI00292946
SERPINA7
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 7

IPI00022426
AMBP
alpha-1-microglobulin/bikunin precursor

IPI00021405
LMNA
lamin A/C

IPI00006662
APOD
apolipoprotein D

IPI00003515
TRIP11
thyroid hormone receptor interactor 11

IPI00021727
C4BPA
complement component 4 binding protein,

alpha

IPI00010779
TPM4
tropomyosin 4

IPI00302592
FLNA
filamin A, alpha

IPI00641737
HP
haptoglobin

IPI00022488
HPX
hemopexin

IPI00473011
HBD
hemoglobin, delta

IPI00298497
FGB
fibrinogen beta chain

IPI00027462
S100A9
S100 calcium binding protein A9

IPI00032291
C5
complement component 5

IPI00031036
SLC26A3
solute carrier family 26, member 3

IPI00022395
C9
complement component 9

IPI00022391
APCS
amyloid P component, serum

IPI00022895
A1BG
alpha-1-B glycoprotein

IPI00887739
LOC100133511
complement C3-like

IPI00218192
ITIH4
inter-alpha (globulin) inhibitor H4 (plasma

Kallikrein-sensitive glycoprotein)

IPI00011252
C8A
complement component 8, alpha polypeptide

IPI00292530
ITIH1
inter-alpha (globulin) inhibitor H1

IPI00937735
ACADVL
acyl-CoA dehydrogenase, very long chain

IPI00009268
ACY1
cDNA FLJ60317, highly similar to

Aminoacylase-1

IPI00166331
ANKRD35
Ankyrin repeat domain-containing protein 35

IPI00299635
BIRC6
baculoviral IAP repeat-containing 6

IPI00219575
BLMH
Bleomycin hydrolase

IPI00026240
BST1
bone marrow stromal cell antigen 1

IPI00929313
C13orf40
hypothetical protein LOC643677

IPI00027983
CDA
Cytidine deaminase

IPI00852865
CHIT1
chitinase 1 (chitotriosidase)

IPI00022810
CTSC
Dipeptidyl-peptidase 1

IPI00299150
CTSS
Cathepsin S

IPI00006024
DOCK4
Isoform 2 of Dedicator of cytokinesis protein 4

IPI00016862
GSR
Glutathione reductase

IPI00941901
HERC1
hect (homologous to the E6-AP (UBE3A)

carboxyl terminus) domain and RCC1 (CHC1)-

like domain (RLD) 1

IPI00005826
HERC2
hect domain and RLD 2

IPI00746667
HLA-DRB5
HLA class II histocompatibility antigen, DRB1-11

beta chain

IPI00027223
IDH1
Isocitrate dehydrogenase [NADP] cytoplasmic

IPI00305461
ITIH2
inter-alpha (globulin) inhibitor H2

IPI00748360
KIAA1797
Uncharacterized protein KIAA1797

IPI00019038
LYZ
Lysozyme C

IPI00303335
NEB
Nebulin

IPI00065454
NEK10
NIMA (never in mitosis gene a)-related kinase

10 (NEK10);

IPI00257882
PEPD
peptidase D

IPI00003590
QSOX1
quiescin Q6 sulfhydryl oxidase

IPI00939604
RNASET2
Ribonuclease T2

IPI00553177
SERPINA1
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 1

IPI00550991
SERPINA3
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 3

IPI00022204
SERPINB3
serpin peptidase inhibitor, clade B (ovalbumin),

member 3

IPI00307733
SETD2
SET domain containing 2

IPI00218013
SGOL2
Shugoshin-like 2

IPI00010949
SIAE
sialic acid acetylesterase

IPI00386444
SYNE1
spectrin repeat containing, nuclear envelope 1

IPI00744692
TALDO1
Transaldolase 1

IPI00375371
TAS2R42
Taste receptor type 2 member 42

IPI00465028
TPI1
triosephosphate isomerase 1

IPI00291175
VCL
Vinculin

IPI00029647
ZG16
Zymogen granule membrane protein 16

IPI00016255
PLBD1
hypothetical protein LOC79887

IPI00477505
ANKRD28
Isoform 1 of Serine/threonine-protein

phosphatase 6 regulatory ankyrin repeat subunit A

IPI00032293
CST3
Cystatin-C

IPI00293867
DDT
D-dopachrome decarboxylase

IPI00059242
SYAP1
Synapse-associated protein 1

IPI00219622
PSMA2
Proteasome subunit alpha type-2

IPI00221222
SUB1
SUB1 homolog (S. cerevisiae)

IPI00022791
MFAP3
Microfibril-associated glycoprotein 3

IPI00011229
CTSD
Cathepsin D

IPI00025019
PSMB1
proteasome (prosome, macropain) subunit, beta

type, 1

IPI00479306
PSMB5
proteasome (prosome, macropain) subunit, beta

type, 5

IPI00234793
KCTD15
cDNA FLJ61112, highly similar to BTB/POZ

domain-containing protein KCTD15

IPI00010796
P4HB
prolyl 4-hydroxylase, beta polypeptide

IPI00927606
GPX1
glutathione peroxidase 1

IPI00783625
SERPINB5
serpin peptidase inhibitor, clade B (ovalbumin),

member 5

IPI00024701
COL4A3BP
collagen, type IV, alpha 3 (Goodpasture

antigen) binding protein

IPI00000811
PSMB6
proteasome (prosome, macropain) subunit, beta

type, 6

IPI00021298
KRT20
Keratin 20

IPI00025084
CAPNS1
Calpain small subunit 1

IPI00024919
PRDX3
peroxiredoxin 3

IPI00059930
NACC2
NACC family member 2, BEN and BTB (POZ)

domain containing

IPI00003817
ARHGDIB
Rho GDP-dissociation inhibitor 2

IPI00293276
MIF
Macrophage migration inhibitory factor

IPI00514622
RANBP6
Ran-binding protein 6

IPI00333141
SPNS3
spinster homolog 3 (Drosophila)

IPI00922181
MCM2
minichromosome maintenance complex

component 2

IPI00031708
FAH
Fumarylacetoacetase

IPI00003865
HSPA8
heat shock 70 kDa protein 8

IPI00299024
BASP1
brain abundant, membrane attached signal

protein 1

IPI00181135
BCAT2
Branched-chain-amino-acid aminotransferase

IPI00219365
MSN
Moesin

IPI00032134
SERPINB8
serpin peptidase inhibitor, clade B (ovalbumin),

member 8

IPI00853547
G6PD
glucose-6-phosphate dehydrogenase isoform a

IPI00478758
C10orf119
Isoform 1 of UPF0557 protein C10orf119

IPI00873020
PSAP
Prosaposin

IPI00000875
EEF1G
eukaryotic translation elongation factor 1

gamma

IPI00014398
FHL1
four and a half LIM domains 1

IPI00301395
CPVL
carboxypeptidase, vitellogenic-like

IPI00790262
TTLL3
tubulin tyrosine ligase-like family, member 3

IPI00942608
26kDaprotein
IPI:IPI00942608.1|ENSEMBL:ENSP0000

IPI00023038
PRB1/PRB2
proline-rich protein BstNI subfamily 2

IPI00001895
PCDH8
Protocadherin-8

IPI00419215
A2ML1
Alpha-2-macroglobulin-like protein 1

IPI00644409
GDA
Guanine deaminase

IPI00009650
LCN1
Lipocalin-1

IPI00217467
HIST1H1E
Histone H1.4

IPI00937064
ZAN
IPI:IPI00937064.1|REFSEQ:XP_002342720

IPI00396378
HNRNPA2B1
heterogeneous nuclear ribonucleoprotein A2/B1

IPI00465261
ERAP2
endoplasmic reticulum aminopeptidase 2

IPI00021263
YWHAZ
14-3-3 protein zeta/delta

IPI00011241
GPR39
G-protein coupled receptor 39

IPI00786880
KIAA1783
similar to KIAA1783 protein

IPI00168479
APOA1BP
apolipoprotein A-I binding protein

IPI00031084
PSD2
pleckstrin and Sec7 domain containing 2

IPI00001593
PRCP
prolylcarboxypeptidase (angiotensinase C)

IPI00218343
TUBA1C
Tubulin alpha-1C chain

IPI00021536
CALML5
Calmodulin-like protein 5

IPI00028091
ACTR3
ARP3 actin-related protein 3 homolog (yeast)

IPI00796366
MYL6
myosin, light chain 6, alkali, smooth muscle and

non-muscle

IPI00301058
VASP
Vasodilator-stimulated phosphoprotein

IPI00470573
ACTR2
ARP2 actin-related protein 2 homolog (yeast)

IPI00884078
RF-IP18
Rheumatoid factor RF-IP18

IPI00169383
PGK1
Phosphoglycerate kinase 1

IPI00299619
SLC35F1
Solute carrier family 35 member F1

IPI00419916
ALPL
alkaline phosphatase, liver/bone/kidney

IPI00218319
TPM3
tropomyosin 3

IPI00005118
HK3
Hexokinase-3

IPI00418471
VIM
Vimentin

IPI00218918
ANXA1
Annexin A1

IPI00930073
KRT6C
IPI:IPI00930073.1|TREMBL:B2R853

IPI00299145
KRT6C
Keratin, type II cytoskeletal 6C

IPI00007858
MYH13
myosin, heavy chain 13, skeletal muscle

IPI00297235
CCPG1
cell cycle progression 1

IPI00927726
H-INV
Hypothetical protein

IPI00009008
CACNA1D
calcium channel, voltage-dependent, L type,

alpha 1D subunit

IPI00304554
LYPD5
LY6/PLAUR domain containing 5

IPI00815807
ADCK2
aarF domain containing kinase 2

IPI00743335
MYO1C
Myosin-Ic

IPI00007393
APPBP2
amyloid beta precursor protein (cytoplasmic tail)

binding protein 2

IPI00295976
ITGA2B
integrin, alpha 2b (platelet glycoprotein IIb of

IIb/IIIa complex, antigen CD41)

IPI00641706
TUBB6
tubulin, beta 6

IPI00060201
SYTL4
synaptotagmin-like 4

IPI00908634
AQP4
aquaporin 4

IPI00016786
CDC42
cell division cycle 42 (GTP binding protein,

25 kDa)

IPI00033494
MYL12B
myosin, light chain 12B, regulatory

IPI00409756
LOC100293553
protein L-Myc-2-like

IPI00015148
RAP1B
RAP1B, member of RAS oncogene family

IPI00027502
GP9
glycoprotein IX (platelet)

IPI00473014
DSTN
Destrin

IPI00022394
C1QC
complement component 1, q subcomponent, C

chain

IPI00290337
EPS8
epidermal growth factor receptor pathway

substrate 8

IPI00018671
DUSP3
dual specificity phosphatase 3

IPI00478231
RHOA
ras homolog gene family, member A

IPI00220278
MYL9
myosin, light chain 9, regulatory

IPI00419585
PPIA
peptidylprolyl isomerase A (cyclophilin A)

IPI00012011
CFL1
Cofilin-1

TABLE 2

A subset of biomarkers from Table 1 which are significantly differentially expressed in all CRC samples relative to a control sample

mean
mean

spectral
spectral

Identified
Identified

counts
counts

in nr.
in nr.

per
per

Fold Change
P-values
CRCs
Controls

Identified
positive
positive

Accession Number
Gene Symbol
Entrez Gene Name
Control vs CRC
Control vs CRC
(n = 12)
(n = 10)
Detected in plasma
in Cancer Cell Lines
CRC
control

IPI00892604
C4A/C4B
complement component
~
2.45E−06
10
0
no
yes
11
na

4B (Chido blood group)

IPI00018206
GOT2
glutamic-oxaloacetic
~
4.93E−03
5
0
no
yes
9
na

transaminase 2,

mitochondrial (aspartate

aminotransferase 2)

IPI00027497
GPI
glucose-6-phosphate
49.74
5.44E−03
7
1
no
yes
7
1

isomerase

IPI00643920
TKT
transketolase
24.13
1.71E−02
6
1
no
yes
6
1

IPI00337741
APEH
N-acylaminoacyl-
12.45
4.72E−04
9
2
no
yes
6
2

peptide hydrolase

IPI00453473
HIST4H4
histone cluster 1, H4c
2.72
4.42E−02
9
4
no
yes
5
2

(includes

others)

IPI00007797
FABP5
fatty acid binding
~
3.48E−02
3
0
no
yes
5
na

protein 5 (psoriasis-

associated)

IPI00012585
HEXB
hexosaminidase B
14.77
1.35E−02
6
1
no
yes
5
2

(beta polypeptide)

IPI00296215
EPCAM
epithelial cell adhesion
~
3.56E−02
3
0
no
yes
4
na

molecule

IPI00604590
NME1-NME2
NME1-NME2
~
1.34E−02
4
0
no
yes
4
na

readthrough

IPI00022314
SOD2
superoxide dismutase
~
6.04E−03
5
0
no
yes
4
na

2, mitochondrial

IPI00027107
TUFM
Tu translation
2.02
7.75E−03
10
2
no
yes
3
5

elongation factor,

mitochondrial

IPI00010706
GSS
glutathione synthetase
~
1.08E−02
4
0
no
yes
3
na

IPI00418169
ANXA2
annexin A2
~
4.47E−03
5
0
no
yes
3
na

IPI00303476
ATP5B
ATP synthase, H+
7.02
2.62E−02
6
1
no
yes
3
1

transporting,

mitochondrial F1

complex, beta

polypeptide

IPI00220362
HSPE1
heat shock 10 kDa
~
9.57E−03
4
0
no
yes
3
na

protein 1 (chaperonin

10)

IPI00220766
GLO1
glyoxalase 1
~
5.24E−04
7
0
no
yes
2
na

IPI00003935
HIST2H2BE
histone cluster 2, H2be
9.25
3.83E−02
5
1
no
yes
2
1

(includes

others)

IPI00032313
S100A4
S100 calcium binding
~
1.40E−02
4
0
no
yes
2
na

protein A4

IPI00013895
S100A11
S100 calcium binding
~
3.55E−03
5
0
no
yes
2
na

protein A11

IPI00106687
LXN
latexin
4.85
1.98E−02
7
2
no
yes
2
2

IPI00034280
DHRS11
dehydrogenase/
7.23
6.98E−03
8
2
no
yes
2
1

reductase (SDR family)

member 11

IPI00008787
NAGLU
N-
11.00
4.44E−03
7
1
no
yes
2
1

acetylglucosaminidase,

alpha

IPI00018768
TSN
translin
~
1.45E−02
4
0
no
yes
2
na

IPI00299155
PSMA4
proteasome (prosome,
~
1.48E−02
4
0
no
yes
2
na

macropain) subunit,

alpha type, 4

IPI00029623
PSMA6
proteasome (prosome,
~
3.02E−02
3
0
no
yes
2
na

macropain) subunit,

alpha type, 6

IPI00010271
RAC1
ras-related C3
~
4.46E−02
3
0
no
yes
2
na

botulinum toxin

substrate 1 (rho family,

small GTP binding

protein Rac1)

IPI00012007
AHCY
Adenosyl
~
1.68E−03
6
0
no
yes
2
na

homocysteinase

IPI00385751
FUCA1
fucosidase, alpha-L-1,
14.06
2.67E−02
6
1
no
yes
2
1

tissue

IPI00017526
S100P
S100 calcium binding
~
3.57E−03
5
0
no
yes
2
na

protein P

IPI00028006
PSMB2
proteasome (prosome,
~
3.77E−03
5
0
no
yes
2
na

macropain) subunit,

beta type, 2

IPI00793375
XPNPEP1
X-prolyl
~
3.35E−02
3
0
no
yes
2
na

aminopeptidase

(aminopeptidase P) 1,

soluble

IPI00554788
KRT18
keratin 18
~
3.79E−02
3
0
no
yes
2
na

IPI00019380
NCBP1
nuclear cap binding
~
4.02E−02
3
0
no
yes
2
na

protein subunit 1, 80 kDa

IPI00012989
MAN2B1
mannosidase, alpha,
7.64
2.11E−02
7
2
no
yes
1
1

class 2B, member 1

IPI00027463
S100A6
S100 calcium binding
4.33
7.50E−03
10
3
no
yes
1
1

protein A6

IPI00022774
VCP
valosin-containing
~
3.16E−02
3
0
no
yes
1
na

protein

IPI00300086
QPRT
quinolinate
~
1.33E−02
4
0
no
yes
1
na

phosphoribosyltransferase

IPI00472073
HLA-B
major histocompatibility
~
5.15E−03
5
0
no
yes
1
na

complex, class I, B

IPI00453476
PGAM1
phosphoglycerate
~
2.06E−02
4
0
no
yes
1
na

mutase 1 (brain)

IPI00020999
ENPP3
ectonucleotide
2.77
6.30E−04
10
2
no
no
11
14

pyrophosphatase/phosphodiesterase 3

IPI00010304
SERPINB10
serpin peptidase
32.16
2.84E−02
6
1
no
no
10
4

inhibitor, clade B

(ovalbumin), member 10

IPI00007244
MPO
myeloperoxidase
22.73
5.30E−04
10
3
no
no
10
2

IPI00877726
CKMT1A/
creatine kinase,
2.08
2.15E−03
11
4
no
no
7
7

CKMT1B
mitochondrial 1B

IPI00027409
PRTN3
proteinase 3
93.49
1.90E−05
10
1
no
no
5
1

IPI00027769
ELANE
elastase, neutrophil
23.48
1.47E−02
6
1
no
no
5
1

expressed

IPI00004362
MORC1
MORC family CW-type
6.22
3.95E−02
5
1
no
no
4
1

zinc finger 1

IPI00719280
UBB
ubiquitin B
3.45
1.49E−02
11
6
no
no
4
2

IPI00026962
PLA2G2A
phospholipase A2,
6.13
4.30E−03
7
1
no
no
3
1

group IIA (platelets,

synovial fluid)

IPI00027466
CA4
carbonic anhydrase IV
4.02
3.87E−03
10
2
no
no
3
4

IPI00071703
GFM2
G elongation factor,
4.03
2.72E−02
10
5
no
no
3
1

mitochondrial 2

IPI00219806
S100A7
S100 calcium binding
3.15
4.75E−02
7
4
no
no
3
1

protein A7

IPI00827847
BPI
bactericidal/permeability-
~
1.61E−02
4
0
no
no
3
na

increasing protein

IPI00176125
COL6A5
collagen, type VI, alpha 5
2.74
2.13E−02
8
2
no
no
3
2

IPI00470355
LHX8
LIM homeobox 8
~
1.30E−02
4
0
no
no
3
na

IPI00942117
CRISP3
cysteine-rich secretory
~
1.45E−02
4
0
no
no
2
na

protein 3

IPI00022246
AZU1
azurocidin 1
~
2.27E−05
8
0
no
no
2
na

IPI00045512
HMCN1
hemicentin 1
8.30
6.80E−03
8
2
no
no
2
1

IPI00300376
TGM3
transglutaminase 3 (E
~
3.67E−02
3
0
no
no
2
na

polypeptide, protein-

glutamine-gamma-

glutamyltransferase)

IPI00640468
CDC42BPA
CDC42 binding protein
1.88
4.92E−02
8
3
no
no
2
1

kinase alpha (DMPK-

like)

IPI00028064
CTSG
cathepsin G
~
1.03E−02
4
0
no
no
2
na

IPI00006988
RETN
resistin
~
3.74E−02
3
0
no
no
2
na

IPI00024934
MUT
methylmalonyl CoA
11.42
7.07E−03
8
2
no
no
2
1

mutase

IPI00552983
ARMCX4
armadillo repeat
12.86
1.65E−02
7
1
no
no
2
2

containing, X-linked 4

IPI00217987
ITGAM
integrin, alpha M
~
1.59E−02
4
0
no
no
2
na

(complement

component 3 receptor 3

subunit)

IPI00165045
CACNA1E
calcium channel,
~
7.77E−04
7
0
no
no
1
na

voltage-dependent, R

type, alpha 1E subunit

IPI00018363
TIAM2
T-cell lymphoma
18.20
4.58E−03
7
1
no
no
1
1

invasion and metastasis 2

IPI00217560
HIRA
HIR histone cell cycle
~
8.36E−03
5
0
no
no
1
na

regulation defective

homolog A (S. cerevisiae)

IPI00294653
DOPEY2
dopey family member 2
13.89
2.46E−02
6
1
no
no
1
1

IPI00004401
ITGB1BP3
integrin beta 1 binding
~
3.38E−02
3
0
no
no
1
na

protein 3

IPI00300117
SCN7A
sodium channel,
~
3.38E−02
3
0
no
no
1
na

voltage-gated, type VII,

alpha

IPI00061114
RAB3C
RAB3C, member RAS
~
3.41E−02
3
0
no
no
1
na

oncogene family

IPI00168442
C9orf79
chromosome 9 open
~
3.89E−02
3
0
no
no
1
na

reading frame 79

IPI00385215
NFATC4
nuclear factor of
~
3.95E−02
3
0
no
no
1
na

activated T-cells,

cytoplasmic, calcineurin-

dependent 4

IPI00024467
UGGT2
UDP-glucose
~
4.02E−02
3
0
no
no
1
na

glycoprotein

glucosyltransferase 2

IPI00297056
CRNN
cornulin
~
4.04E−02
3
0
no
no
1
na

IPI00914663
KCP
kielin/chordin-like
~
9.68E−03
4
0
no
no
1
na

protein

IPI00418592
CD1E
CD1e molecule
~
1.46E−02
4
0
no
no
1
na

IPI00642206
CCDC18
coiled-coil domain
~
4.58E−03
5
0
no
no
1
na

containing 18

IPI00514090
LTA4H
leukotriene A4
~
4.50E−02
3
0
no
no
1
na

hydrolase

IPI00745872
ALB
albumin
4.71
1.41E−02
12
10
yes
yes
576
124

IPI00478003
A2M
alpha-2-macroglobulin
34.08
9.01E−06
12
6
yes
yes
141
11

IPI00783987
C3
complement component 3
43.72
2.21E−04
12
6
yes
yes
119
5

IPI00654755
HBB
hemoglobin, beta
9.87
5.89E−05
12
7
yes
yes
108
14

IPI00022463
TF
transferrin
1713.68
7.84E−06
11
1
yes
yes
96
1

IPI00410714
HBA1/HBA2
hemoglobin, alpha 1
44.44
1.77E−04
12
5
yes
yes
68
2

IPI00298860
LTF
lactotransferrin
59.22
3.14E−05
10
2
yes
yes
43
6

IPI00017601
CP
ceruloplasmin
94.27
1.28E−04
11
3
yes
yes
41
2

(ferroxidase)

IPI00465436
CAT
catalase
77.03
1.66E−03
10
3
yes
yes
24
1

IPI00555812
GC
group-specific
~
3.43E−02
3
0
yes
yes
20
na

component (vitamin D

binding protein)

IPI00032179
SERPINC1
serpin peptidase
2.17
3.20E−02
12
10
yes
yes
20
11

inhibitor, clade C

(antithrombin), member 1

IPI00219713
FGG
fibrinogen gamma chain
~
1.11E−03
6
0
yes
yes
20
na

IPI00007047
S100A8
S100 calcium binding
3.87
1.12E−06
12
9
yes
yes
17
5

protein A8

IPI00375676
FTL
ferritin, light polypeptide
2.51
2.92E−02
12
10
yes
yes
12
5

IPI00021439
ACTB
actin, beta
2.44
1.07E−02
10
3
yes
yes
12
11

IPI00022418
FN1
fibronectin 1
17.62
1.31E−02
10
4
yes
yes
11
2

IPI00005721
DEFA1
defensin, alpha 1
4.16
4.47E−05
12
9
yes
yes
8
2

(includes

others)

IPI00291866
SERPING1
serpin peptidase
~
2.88E−04
7
0
yes
yes
8
na

inhibitor, clade G (C1

inhibitor), member 1

IPI00022420
RBP4
retinol binding protein 4,
4.47
1.20E−02
8
3
yes
yes
8
6

plasma

IPI00027350
PRDX2
peroxiredoxin 2
36.98
4.57E−03
7
1
yes
yes
6
1

IPI00021885
FGA
fibrinogen alpha chain
21.98
3.30E−02
6
2
yes
yes
6
1

IPI00029863
SERPINF2
serpin peptidase
8.61
8.05E−04
9
1
yes
yes
4
8

inhibitor, clade F (alpha-

2 antiplasmin, pigment

epithelium derived

factor), member 2

IPI00218414
CA2
carbonic anhydrase II
2.87
1.12E−02
7
1
yes
yes
4
10

IPI00020091
ORM1/
orosomucoid 1
4.11
2.56E−02
9
3
yes
yes
4
3

ORM2

IPI00217966
LDHA
lactate dehydrogenase A
~
1.85E−03
6
0
yes
yes
4
na

IPI00298971
VTN
vitronectin
~
1.15E−03
6
0
yes
yes
4
na

IPI00215894
KNG1
kininogen 1
~
3.33E−04
7
0
yes
yes
4
na

IPI00023006
ACTC1
actin, alpha, cardiac
3.17
2.33E−03
10
3
yes
yes
4
2

muscle 1

IPI00022417
LRG1
leucine-rich alpha-2-
31.63
4.06E−03
7
1
yes
yes
3
1

glycoprotein 1

IPI00023728
GGH
gamma-glutamyl
6.57
1.73E−02
7
2
yes
yes
3
3

hydrolase (conjugase,

folylpolygammaglutamyl

hydrolase)

IPI00465248
ENO1
enolase 1, (alpha)
5.38
3.17E−02
7
2
yes
yes
3
1

IPI00216691
PFN1
profilin 1
~
4.15E−02
3
0
yes
yes
3
na

IPI00292946
SERPINA7
serpin peptidase
~
6.51E−04
7
0
yes
yes
3
na

inhibitor, clade A (alpha-

1 antiproteinase,

antitrypsin), member 7

IPI00022426
AMBP
alpha-1-
~
1.75E−03
6
0
yes
yes
3
na

microglobulin/bikunin

precursor

IPI00021405
LMNA
lamin A/C
~
1.47E−02
4
0
yes
yes
3
na

IPI00006662
APOD
apolipoprotein D
3.38
4.74E−03
8
1
yes
yes
2
4

IPI00003515
TRIP11
thyroid hormone
7.65
3.20E−03
9
1
yes
yes
2
2

receptor interactor 11

IPI00021727
C4BPA
complement component
~
1.23E−02
4
0
yes
yes
2
na

4 binding protein, alpha

IPI00010779
TPM4
tropomyosin 4
~
4.12E−02
3
0
yes
yes
2
na

IPI00302592
FLNA
filamin A, alpha
~
2.45E−03
6
0
yes
yes
1
na

IPI00641737
HP
haptoglobin
18.82
2.89E−04
11
6
yes
no
89
6

IPI00022488
HPX
hemopexin
~
4.42E−05
8
0
yes
no
23
na

IPI00473011
HBD
hemoglobin, delta
~
6.31E−05
8
0
yes
no
15
na

IPI00298497
FGB
fibrinogen beta chain
~
1.18E−02
4
0
yes
no
11
na

IPI00027462
S100A9
S100 calcium binding
4.41
1.82E−05
12
8
yes
no
10
3

protein A9

IPI00032291
C5
complement component 5
33.44
9.34E−03
8
2
yes
no
8
1

IPI00031036
SLC26A3
solute carrier family 26,
64.79
1.18E−02
7
1
yes
no
6
1

member 3

IPI00022395
C9
complement component 9
12.69
4.19E−04
11
5
yes
no
6
1

IPI00022391
APCS
amyloid P component,
5.75
2.53E−03
10
3
yes
no
6
5

serum

IPI00022895
A1BG
alpha-1-B glycoprotein
~
1.31E−02
4
0
yes
no
6
na

IPI00887739
LOC100133511
complement C3-like
~
4.38E−03
5
0
yes
no
5
na

IPI00218192
ITIH4
inter-alpha (globulin)
~
3.64E−02
3
0
yes
no
5
na

inhibitor H4 (plasma

Kallikrein-sensitive

glycoprotein)

IPI00011252
C8A
complement component
~
3.26E−02
3
0
yes
no
3
na

8, alpha polypeptide

IPI00292530
ITIH1
inter-alpha (globulin)
~
1.05E−02
4
0
yes
no
1
na

inhibitor H1

TABLE 3

A subset of biomarkers from Table 1 which are differentially expressed in

FIT-negative CRC samples relative to a control sample

Accession

Number
Gene Symbol
Entrez Gene Name

IPI00022426
AMBP
alpha-1-microglobulin/bikunin precursor

IPI00022246
AZU1
azurocidin 1

IPI00827847
BPI
bactericidal/permeability-increasing protein

IPI00892604
C4A/C4B
complement component 4B (Chido blood group)

IPI00021727
C4BPA
complement component 4 binding protein, alpha

IPI00168442
C9orf79
chromosome 9 open reading frame 79

IPI00165045
CACNA1E
calcium channel, voltage-dependent, R type, alpha 1E

subunit

IPI00642206
CCDC18
coiled-coil domain containing 18

IPI00297056
CRNN
cornulin

IPI00296215
EPCAM
epithelial cell adhesion molecule

IPI00219713
FGG
fibrinogen gamma chain

IPI00302592
FLNA
filamin A, alpha

IPI00220766
GLO1
glyoxalase I

IPI00018206
GOT2
glutamic-oxaloacetic transaminase 2, mitochondrial

(aspartate aminotransferase 2)

IPI00010706
GSS
glutathione synthetase

IPI00217560
HIRA
HIR histone cell cycle regulation defective homolog A

(S. cerevisiae)

IPI00472073
HLA-B
major histocompatibility complex, class I, B

IPI00022488
HPX
hemopexin

IPI00217987
ITGAM
integrin, alpha M (complement component 3 receptor

3 subunit)

IPI00215894
KNG1
kininogen 1

IPI00554788
KRT18
keratin 18

IPI00217966
LDHA
lactate dehydrogenase A

IPI00470355
LHX8
LIM homeobox 8

IPI00887739
LOC100133511
complement C3-like

IPI00019380
NCBP1
nuclear cap binding protein subunit 1, 80 kDa

IPI00604590
NME1-NME2
NME1-NME2 readthrough

IPI00299155
PSMA4
proteasome (prosome, macropain) subunit, alpha

type, 4

IPI00300117
SCN7A
sodium channel, voltage-gated, type VII, alpha

IPI00292946
SERPINA7
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 7

IPI00291866
SERPING1
serpin peptidase inhibitor, clade G (C1 inhibitor),

member 1

IPI00024467
UGGT2
UDP-glucose glycoprotein glucosyltransferase 2

IPI00298971
VTN
vitronectin

IPI00478003
A2M
alpha-2-macroglobulin

IPI00022391
APCS
amyloid P component, serum

IPI00337741
APEH
N-acylaminoacyl-peptide hydrolase

IPI00552983
ARMCX4
armadillo repeat containing, X-linked 4

IPI00783987
C3
complement component 3

IPI00032291
C5
complement component 5

IPI00022395
C9
complement component 9

IPI00027466
CA4
carbonic anhydrase IV

IPI00465436
CAT
catalase

IPI00176125
COL6A5
collagen, type VI, alpha 5

IPI00017601
CP
ceruloplasmin (ferroxidase)

IPI00005721
DEFA1(include
defensin, alpha 1

sothers)

IPI00294653
DOPEY2
dopey family member 2

IPI00020999
ENPP3
ectonucleotide pyrophosphatase/phosphodiesterase 3

IPI00022418
FN1
fibronectin 1

IPI00375676
FTL
ferritin, light polypeptide

IPI00385751
FUCA1
fucosidase, alpha-L-1, tissue

IPI00071703
GFM2
G elongation factor, mitochondrial 2

IPI00023728
GGH
gamma-glutamyl hydrolase (conjugase,

folylpolygammaglutamyl hydrolase)

IPI00027497
GPI
glucose-6-phosphate isomerase

IPI00410714
HBA1/HBA2
hemoglobin, alpha 1

IPI00654755
HBB
hemoglobin, beta

IPI00012585
HEXB
hexosaminidase B (beta polypeptide)

IPI00045512
HMCN1
hemicentin 1

IPI00022417
LRG1
leucine-rich alpha-2-glycoprotein 1

IPI00298860
LTF
lactotransferrin

IPI00106687
LXN
latexin

IPI00012989
MAN2B1
mannosidase, alpha, class 2B, member 1

IPI00007244
MPO
myeloperoxidase

IPI00024934
MUT
methylmalonyl CoA mutase

IPI00026962
PLA2G2A
phospholipase A2, group IIA (platelets, synovial fluid)

IPI00027350
PRDX2
peroxiredoxin 2

IPI00027409
PRTN3
proteinase 3

IPI00022420
RBP4
retinol binding protein 4, plasma

IPI00027463
S100A6
S100 calcium binding protein A6

IPI00219806
S100A7
S100 calcium binding protein A7

IPI00007047
S100A8
S100 calcium binding protein A8

IPI00027462
S100A9
S100 calcium binding protein A9

IPI00010304
SERPINB10
serpin peptidase inhibitor, clade B (ovalbumin),

member 10

IPI00029863
SERPINF2
serpin peptidase inhibitor, clade F (alpha-2

antiplasmin, pigment epithelium derived factor),

member 2

IPI00031036
SLC26A3
solute carrier family 26, member 3

IPI00022463
TF
transferrin

IPI00018363
TIAM2
T-cell lymphoma invasion and metastasis 2

IPI00643920
TKT
transketolase

IPI00003515
TRIP11
thyroid hormone receptor interactor 11

IPI00027107
TUFM
Tu translation elongation factor, mitochondrial

IPI00181135
BCAT2
branched chain amino-acid transaminase 2,

mitochondrial

IPI00024701
COL4A3BP
collagen, type IV, alpha 3 (Goodpasture antigen)

binding protein

IPI00032134
SERPINB8
serpin peptidase inhibitor, clade B (ovalbumin),

member 8

IPI00333141
SPNS3
spinster homolog 3 (Drosophila)

IPI00937735
ACADVL
acyl-CoA dehydrogenase, very long chain

IPI00009268
ACY1
Unmapped by Ingenuity

IPI00166331
ANKRD35
ankyrin repeat domain 35

IPI00299635
BIRC6
baculoviral IAP repeat-containing 6

IPI00219575
BLMH
bleomycin hydrolase

IPI00026240
BST1
bone marrow stromal cell antigen 1

IPI00929313
C13orf40
chromosome 13 open reading frame 40

IPI00027983
CDA
cytidine deaminase

IPI00852865
CHIT1
chitinase 1 (chitotriosidase)

IPI00022810
CTSC
cathepsin C

IPI00299150
CTSS
cathepsin S

IPI00006024
DOCK4
dedicator of cytokinesis 4

IPI00016862
GSR
glutathione reductase

IPI00941901
HERC1
hect (homologous to the E6-AP (UBE3A) carboxyl

terminus) domain and RCC1 (CHC1)-like domain

(RLD) 1

IPI00005826
HERC2
hect domain and RLD 2

IPI00746667
HLA-DRB5
major histocompatibility complex, class II, DR beta 5

IPI00027223
IDH1
isocitrate dehydrogenase 1 (NADP+), soluble

IPI00305461
ITIH2
inter-alpha (globulin) inhibitor H2

IPI00748360
KIAA1797
KIAA1797

IPI00019038
LYZ
lysozyme

IPI00303335
NEB
nebulin

IPI00065454
NEK10
NIMA (never in mitosis gene a)-related kinase 10

IPI00257882
PEPD
peptidase D

IPI00003590
QSOX1
quiescin Q6 sulfhydryl oxidase 1

IPI00939604
RNASET2
ribonuclease T2

IPI00553177
SERPINA1
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 1

IPI00550991
SERPINA3
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 3

IPI00022204
SERPINB3
serpin peptidase inhibitor, clade B (ovalbumin),

member 3

IPI00307733
SETD2
SET domain containing 2

IPI00218013
SGOL2
shugoshin-like 2

IPI00010949
SIAE
sialic acid acetylesterase

IPI00386444
SYNE1
spectrin repeat containing, nuclear envelope 1

IPI00744692
TALDO1
transaldolase 1

IPI00375371
TAS2R42
taste receptor, type 2, member 42

IPI00465028
TPI1
triosephosphate isomerase 1

IPI00291175
VCL
vinculin

IPI00029647
ZG16
zymogen granule protein 16

TABLE 4

A subset of biomarkers from Table 1 which were found to be expressed only

in CRC samples and not in control samples

Accession

Number
Gene Symbol
Entrez Gene Name

IPI00892604
C4A/C4B
complement component 4B (Chido blood group)

IPI00018206
GOT2
glutamic-oxaloacetic transaminase 2, mitochondrial

(aspartate aminotransferase 2)

IPI00007797
FABP5
fatty acid binding protein 5 (psoriasis-associated)

IPI00296215
EPCAM
epithelial cell adhesion molecule

IPI00604590
NME1-NME2
NME1-NME2 readthrough

IPI00022314
SOD2
superoxide dismutase 2, mitochondrial

IPI00010706
GSS
glutathione synthetase

IPI00418169
ANXA2
annexin A2

IPI00220362
HSPE1
heat shock 10 kDa protein 1 (chaperonin 10)

IPI00220766
GLO1
glyoxalase I

IPI00032313
S100A4
S100 calcium binding protein A4

IPI00013895
S100A11
S100 calcium binding protein A11

IPI00018768
TSN
translin

IPI00299155
PSMA4
proteasome (prosome, macropain) subunit, alpha type, 4

IPI00029623
PSMA6
proteasome (prosome, macropain) subunit, alpha type, 6

IPI00010271
RAC1
ras-related C3 botulinum toxin substrate 1 (rho family,

small GTP binding protein Rac1)

IPI00012007
AHCY
adenosylhomocysteinase

IPI00017526
S100P
S100 calcium binding protein P

IPI00028006
PSMB2
proteasome (prosome, macropain) subunit, beta type, 2

IPI00793375
XPNPEP1
X-prolyl aminopeptidase (aminopeptidase P) 1, soluble

IPI00554788
KRT18
keratin 18

IPI00019380
NCBP1
nuclear cap binding protein subunit 1, 80 kDa

IPI00022774
VCP
valosin-containing protein

IPI00300086
QPRT
quinolinate phosphoribosyltransferase

IPI00472073
HLA-B
major histocompatibility complex, class I, B

IPI00453476
PGAM1
phosphoglycerate mutase 1 (brain)

IPI00827847
BPI
bactericidal/permeability-increasing protein

IPI00470355
LHX8
LIM homeobox 8

IPI00942117
CRISP3
cysteine-rich secretory protein 3

IPI00022246
AZU1
azurocidin 1

IPI00300376
TGM3
transglutaminase 3 (E polypeptide, protein-glutamine-

gamma-glutamyltransferase)

IPI00028064
CTSG
cathepsin G

IPI00006988
RETN
resistin

IPI00217987
ITGAM
integrin, alpha M (complement component 3 receptor 3

subunit)

IPI00165045
CACNA1E
calcium channel, voltage-dependent, R type, alpha 1E

subunit

IPI00217560
HIRA
HIR histone cell cycle regulation defective homolog A (S. cerevisiae)

IPI00004401
ITGB1BP3
integrin beta 1 binding protein 3

IPI00300117
SCN7A
sodium channel, voltage-gated, type VII, alpha

IPI00061114
RAB3C
RAB3C, member RAS oncogene family

IPI00168442
C9orf79
chromosome 9 open reading frame 79

IPI00385215
NFATC4
nuclear factor of activated T-cells, cytoplasmic, calcineurin-

dependent 4

IPI00024467
UGGT2
UDP-glucose glycoprotein glucosyltransferase 2

IPI00297056
CRNN
cornulin

IPI00914663
KCP
kielin/chordin-like protein

IPI00418592
CD1E
CD1e molecule

IPI00642206
CCDC18
coiled-coil domain containing 18

IPI00514090
LTA4H
leukotriene A4 hydrolase

IPI00555812
GC
group-specific component (vitamin D binding protein)

IPI00219713
FGG
fibrinogen gamma chain

IPI00291866
SERPING1
serpin peptidase inhibitor, clade G (C1 inhibitor), member 1

IPI00217966
LDHA
lactate dehydrogenase A

IPI00298971
VTN
vitronectin

IPI00215894
KNG1
kininogen 1

IPI00216691
PFN1
profilin 1

IPI00292946
SERPINA7
serpin peptidase inhibitor, clade A (alpha-1 antiproteinase,

antitrypsin), member 7

IPI00022426
AMBP
alpha-1-microglobulin/bikunin precursor

IPI00021405
LMNA
lamin A/C

IPI00021727
C4BPA
complement component 4 binding protein, alpha

IPI00010779
TPM4
tropomyosin 4

IPI00302592
FLNA
filamin A, alpha

IPI00022488
HPX
hemopexin

IPI00473011
HBD
hemoglobin, delta

IPI00298497
FGB
fibrinogen beta chain

IPI00022895
A1BG
alpha-1-B glycoprotein

IPI00887739
LOC100133511
complement C3-like

IPI00218192
ITIH4
inter-alpha (globulin) inhibitor H4 (plasma Kallikrein-

sensitive glycoprotein)

IPI00011252
C8A
complement component 8, alpha polypeptide

IPI00292530
ITIH1
inter-alpha (globulin) inhibitor H1

IPI00016255
PLBD1
hypothetical protein LOC79887

IPI00477505
ANKRD28
Isoform 1 of Serine/threonine-protein phosphatase 6

regulatory ankyrin repeat subunit A

IPI00032293
CST3
Cystatin-C

IPI00293867
DDT
D-dopachrome decarboxylase

IPI00059242
SYAP1
Synapse-associated protein 1

IPI00219622
PSMA2
Proteasome subunit alpha type-2

IPI00221222
SUB1
SUB1 homolog (S. cerevisiae)

IPI00022791
MFAP3
Microfibril-associated glycoprotein 3

IPI00011229
CTSD
Cathepsin D

IPI00025019
PSMB1
proteasome (prosome, macropain) subunit, beta type, 1

IPI00479306
PSMB5
proteasome (prosome, macropain) subunit, beta type, 5

IPI00234793
KCTD15
cDNA FLJ61112, highly similar to BTB/POZ domain-

containing protein KCTD15

IPI00010796
P4HB
prolyl 4-hydroxylase, beta polypeptide

IPI00927606
GPX1
glutathione peroxidase 1

IPI00783625
SERPINB5
serpin peptidase inhibitor, clade B (ovalbumin), member 5

IPI00024701
COL4A3BP
collagen, type IV, alpha 3 (Goodpasture antigen) binding

protein

IPI00000811
PSMB6
proteasome (prosome, macropain) subunit, beta type, 6

IPI00021298
KRT20
Keratin 20

IPI00025084
CAPNS1
Calpain small subunit 1

IPI00024919
PRDX3
peroxiredoxin 3

IPI00059930
NACC2
NACC family member 2, BEN and BTB (POZ) domain

containing

IPI00003817
ARHGDIB
Rho GDP-dissociation inhibitor 2

IPI00293276
MIF
Macrophage migration inhibitory factor

IPI00514622
RANBP6
Ran-binding protein 6

IPI00333141
SPNS3
Isoform 1 of Protein spinster homolog 3

IPI00922181
MCM2
minichromosome maintenance complex component 2

IPI00031708
FAH
Fumarylacetoacetase

IPI00003865
HSPA8
heat shock 70 kDa protein 8

IPI00299024
BASP1
brain abundant, membrane attached signal protein 1

IPI00181135
BCAT2
Branched-chain-amino-acid aminotransferase

IPI00219365
MSN
Moesin

IPI00032134
SERPINB8
Serpin B8

IPI00853547
G6PD
glucose-6-phosphate dehydrogenase

IPI00478758
C10orf119
Isoform 1 of UPF0557 protein C10orf119

IPI00873020
PSAP
Prosaposin

IPI00000875
EEF1G
eukaryotic translation elongation factor 1 gamma

IPI00014398
FHL1
four and a half LIM domains 1

IPI00301395
CPVL
carboxypeptidase, vitellogenic-like

IPI00790262
TTLL3
tubulin tyrosine ligase-like family, member 3

IPI00942608
26kDaprotein
IPI:IPI00942608.1|ENSEMBL:ENSP0000

IPI00023038
PRB1/PRB2
proline-rich protein BstNI subfamily 2

IPI00001895
PCDH8
Protocadherin-8

IPI00419215
A2ML1
Alpha-2-macroglobulin-like protein 1

IPI00644409
GDA
Guanine deaminase

IPI00009650
LCN1
Lipocalin-1

IPI00217467
HIST1H1E
Histone H1.4

IPI00937064
ZAN
IPI:IPI00937064.1|REFSEQ:XP_002342720

IPI00396378
HNRNPA2B1
heterogeneous nuclear ribonucleoprotein A2/B1

IPI00465261
ERAP2
endoplasmic reticulum aminopeptidase 2

IPI00021263
YWHAZ
14-3-3 protein zeta/delta

IPI00011241
GPR39
G-protein coupled receptor 39

IPI00786880
KIAA1783
similar to KIAA1783 protein

IPI00168479
APOA1BP
apolipoprotein A-I binding protein

IPI00031084
PSD2
pleckstrin and Sec7 domain containing 2

IPI00001593
PRCP
prolylcarboxypeptidase (angiotensinase C)

IPI00218343
TUBA1C
Tubulin alpha-1C chain

IPI00021536
CALML5
Calmodulin-like protein 5

IPI00028091
ACTR3
ARP3 actin-related protein 3 homolog (yeast)

IPI00796366
MYL6
myosin, light chain 6, alkali, smooth muscle and non-

muscle

IPI00301058
VASP
Vasodilator-stimulated phosphoprotein

IPI00470573
ACTR2
ARP2 actin-related protein 2 homolog (yeast)

IPI00884078
RF-IP18
Rheumatoid factor RF-IP18

IPI00169383
PGK1
Phosphoglycerate kinase 1

IPI00299619
SLC35F1
Solute carrier family 35 member F1

IPI00419916
ALPL
alkaline phosphatase, liver/bone/kidney

IPI00218319
TPM3
tropomyosin 3

IPI00005118
HK3
Hexokinase-3

IPI00418471
VIM
Vimentin

IPI00218918
ANXA1
Annexin A1

IPI00930073
KRT6C
IPI:IPI00930073.1|TREMBL:B2R853

IPI00299145
KRT6C
Keratin, type II cytoskeletal 6C

IPI00007858
MYH13
myosin, heavy chain 13, skeletal muscle

IPI00297235
CCPG1
cell cycle progression 1

IPI00927726
H-INV
Hypothetical protein

IPI00009008
CACNA1D
calcium channel, voltage-dependent, L type, alpha 1D

subunit

IPI00304554
LYPD5
LY6/PLAUR domain containing 5

IPI00815807
ADCK2
aarF domain containing kinase 2

IPI00743335
MYO1C
Myosin-Ic

IPI00007393
APPBP2
amyloid beta precursor protein (cytoplasmic tail) binding

protein 2

IPI00295976
ITGA2B
integrin, alpha 2b (platelet glycoprotein IIb of IIb/IIIa

complex, antigen CD41)

IPI00641706
TUBB6
tubulin, beta 6

IPI00060201
SYTL4
synaptotagmin-like 4

IPI00908634
AQP4
aquaporin 4

IPI00016786
CDC42
cell division cycle 42 (GTP binding protein, 25 kDa)

IPI00033494
MYL12B
myosin, light chain 12B, regulatory

IPI00409756
LOC100293553
protein L-Myc-2-like

IPI00015148
RAP1B
RAP1B, member of RAS oncogene family

IPI00027502
GP9
glycoprotein IX (platelet)

IPI00473014
DSTN
Destrin

IPI00022394
C1QC
complement component 1, q subcomponent, C chain

IPI00290337
EPS8
epidermal growth factor receptor pathway substrate 8

IPI00018671
DUSP3
dual specificity phosphatase 3

IPI00478231
RHOA
ras homolog gene family, member A

IPI00220278
MYL9
myosin, light chain 9, regulatory

IPI00419585
PPIA
peptidylprolyl isomerase A (cyclophilin A)

IPI00012011
CFL1
Cofilin-1

TABLE 5

A subset of biomarkers from table 1 which were identified in an independent

(verification) set of stool samples

Accession

Number
Gene Symbol
Entrez Gene Name

IPI00022895
A1BG
alpha-1-B glycoprotein

IPI00478003
A2M
alpha-2-macroglobulin

IPI00021439
ACTB
actin, beta

IPI00023006
ACTC1
actin, alpha, cardiac muscle 1

IPI00745872
ALB
albumin

IPI00022391
APCS
amyloid P component, serum

IPI00337741
APEH
N-acylaminoacyl-peptide hydrolase

IPI00006662
APOD
apolipoprotein D

IPI00022246
AZU1
azurocidin 1

IPI00783987
C3
complement component 3

IPI00892604
C4A/C4B
complement component 4B (Chido blood group)

IPI00032291
C5
complement component 5

IPI00218414
CA2
carbonic anhydrase II

IPI00027466
CA4
carbonic anhydrase IV

IPI00465436
CAT
catalase

IPI00017601
CP
ceruloplasmin (ferroxidase)

IPI00028064
CTSG
cathepsin G

IPI00005721
DEFA1(includesothers)
defensin, alpha 1

IPI00027769
ELANE
elastase, neutrophil expressed

IPI00465248
ENO1
enolase 1, (alpha)

IPI00302592
FLNA
filamin A, alpha

IPI00022418
FN1
fibronectin 1

IPI00375676
FTL
ferritin, light polypeptide

IPI00385751
FUCA1
fucosidase, alpha-L-1, tissue

IPI00023728
GGH
gamma-glutamyl hydrolase (conjugase,

folylpolygammaglutamyl hydrolase)

IPI00018206
GOT2
glutamic-oxaloacetic transaminase 2, mitochondrial

(aspartate aminotransferase 2)

IPI00027497
GPI
glucose-6-phosphate isomerase

IPI00010706
GSS
glutathione synthetase

IPI00410714
HBA1/HBA2
hemoglobin, alpha 1

IPI00654755
HBB
hemoglobin, beta

IPI00473011
HBD
hemoglobin, delta

IPI00012585
HEXB
hexosaminidase B (beta polypeptide)

IPI00453473
HIST4H4(includesothers)
histone cluster 1, H4c

IPI00472073
HLA-B
major histocompatibility complex, class I, B

IPI00641737
HP
haptoglobin

IPI00022488
HPX
hemopexin

IPI00220362
HSPE1
heat shock 10 kDa protein 1 (chaperonin 10)

IPI00217966
LDHA
lactate dehydrogenase A

IPI00887739
LOC100133511
complement C3-like

IPI00022417
LRG1
leucine-rich alpha-2-glycoprotein 1

IPI00298860
LTF
lactotransferrin

IPI00106687
LXN
latexin

IPI00012989
MAN2B1
mannosidase, alpha, class 2B, member 1

IPI00007244
MPO
myeloperoxidase

IPI00008787
NAGLU
N-acetylglucosaminidase, alpha

IPI00020091
ORM1/ORM2
orosomucoid 1

IPI00216691
PFN1
profilin 1

IPI00026962
PLA2G2A
phospholipase A2, group IIA (platelets, synovial

fluid)

IPI00027409
PRTN3
proteinase 3

IPI00299155
PSMA4
proteasome (prosome, macropain) subunit, alpha

type, 4

IPI00029623
PSMA6
proteasome (prosome, macropain) subunit, alpha

type, 6

IPI00028006
PSMB2
proteasome (prosome, macropain) subunit, beta

type, 2

IPI00022420
RBP4
retinol binding protein 4, plasma

IPI00006988
RETN
resistin

IPI00007047
S100A8
S100 calcium binding protein A8

IPI00027462
S100A9
S100 calcium binding protein A9

IPI00010304
SERPINB10
serpin peptidase inhibitor, clade B (ovalbumin),

member 10

IPI00032179
SERPINC1
serpin peptidase inhibitor, clade C (antithrombin),

member 1

IPI00029863
SERPINF2
serpin peptidase inhibitor, clade F (alpha-2

antiplasmin, pigment epithelium derived factor),

member 2

IPI00291866
SERPING1
serpin peptidase inhibitor, clade G (C1 inhibitor),

member 1

IPI00022314
SOD2
superoxide dismutase 2, mitochondrial

IPI00022463
TF
transferrin

IPI00643920
TKT
transketolase

IPI00719280
UBB
ubiquitin B

IPI00884078

Unmapped by Ingenuity

IPI00942608
26kDaprotein
Unmapped by Ingenuity

IPI00419215
A2ML1
alpha-2-macroglobulin-like 1

IPI00937735
ACADVL
acyl-CoA dehydrogenase, very long chain

IPI00009268
ACY1
Unmapped by Ingenuity

IPI00419916
ALPL
alkaline phosphatase, liver/bone/kidney

IPI00219575
BLMH
bleomycin hydrolase

IPI00026240
BST1
bone marrow stromal cell antigen 1

IPI00009008
CACNA1D
calcium channel, voltage-dependent, L type, alpha

1D subunit

IPI00877726
CKMT1A/CKMT1B
creatine kinase, mitochondrial 1B

IPI00299150
CTSS
cathepsin S

IPI00031708
FAH
fumarylacetoacetate hydrolase

(fumarylacetoacetase)

IPI00644409
GDA
guanine deaminase

IPI00016862
GSR
glutathione reductase

IPI00396378
HNRNPA2B1
heterogeneous nuclear ribonucleoprotein A2/B1

IPI00003865
HSPA8
heat shock 70 kDa protein 8

IPI00299145
KRT6C
keratin 6C

IPI00930073
KRT6C
keratin 6C

IPI00009650
LCN1
lipocalin 1 (tear prealbumin)

IPI00019038
LYZ
lysozyme

IPI00922181
MCM2
minichromosome maintenance complex component 2

IPI00257882
PEPD
peptidase D

IPI00016255
PLBD1
phospholipase B domain containing 1

IPI00219622
PSMA2
proteasome (prosome, macropain) subunit, alpha

type, 2

IPI00025019
PSMB1
proteasome (prosome, macropain) subunit, beta

type, 1

IPI00000811
PSMB6
proteasome (prosome, macropain) subunit, beta

type, 6

IPI00003590
QSOX1
quiescin Q6 sulfhydryl oxidase 1

IPI00939604
RNASET2
ribonuclease T2

IPI00553177
SERPINA1
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 1

IPI00550991
SERPINA3
serpin peptidase inhibitor, clade A (alpha-1

antiproteinase, antitrypsin), member 3

IPI00022204
SERPINB3
serpin peptidase inhibitor, clade B (ovalbumin),

member 3

IPI00010949
SIAE
sialic acid acetylesterase

IPI00465028
TPI1
triosephosphate isomerase 1

IPI00029647
ZG16
zymogen granule protein 16 homolog (rat)

TABLE 6

Proteins that are significantly more abundant in colon tumor secretomes

compared to normal colon secretomes

Accession

Number
Gene Symbol
Entrez Gene Name

IPI00903112
LTF
lactotransferrin

IPI00329573
COL12A1
collagen, type XII, alpha 1

IPI00374563
AGRN
agrin

IPI00005024
MYBBP1A
MYB binding protein (P160) 1a

IPI00925034
TRRAP
transformation/transcription domain-associated

protein

IPI00221226
ANXA6
annexin A6

IPI00028275
CKAP5
cytoskeleton associated protein 5

IPI00018350
MCM5
minichromosome maintenance complex component 5

IPI00156374
IPO4
importin 4

IPI00894235
NBEAL2
neurobeachin-like 2

IPI00795318
MCM4
minichromosome maintenance complex component 4

IPI00002405
OAS3
2′-5′-oligoadenylate synthetase 3, 100 kDa

IPI00013214
MCM3
minichromosome maintenance complex component 3

IPI00018968
NAE1
NEDD8 activating enzyme E1 subunit 1

IPI00438229
TRIM28
tripartite motif containing 28

IPI00221354
FUS
fused in sarcoma

IPI00031820
FARSA
phenylalanyl-tRNA synthetase, alpha subunit

IPI00013163
MNDA
myeloid cell nuclear differentiation antigen

IPI00026970
SUPT16H
suppressor of Ty 16 homolog (S. cerevisiae)

IPI00017617
DDX5
DEAD (Asp-Glu-Ala-Asp) box polypeptide 5

IPI00031008
TNC
tenascin C

IPI00007175
NIP7
nuclear import 7 homolog (S. cerevisiae)

IPI00000846
CHD4
chromodomain helicase DNA binding protein 4

IPI00465044
RCC2
regulator of chromosome condensation 2

IPI00031519
DNMT1
DNA (cytosine-5-)-methyltransferase 1

IPI00028357
XPO4
exportin 4

IPI00010720
CCT5
chaperonin containing TCP1, subunit 5 (epsilon)

IPI00012340
SRSF9
serine/arginine-rich splicing factor 9

IPI00012645
SPTBN2
spectrin, beta, non-erythrocytic 2

IPI00032292
TIMP1
TIMP metallopeptidase inhibitor 1

IPI00026944
NID1
nidogen 1

IPI00013871
RRM1
ribonucleotide reductase M1

IPI00386533
EIF4G1
eukaryotic translation initiation factor 4 gamma, 1

IPI00149849
COG4
component of oligomeric golgi complex 4

IPI00002894
POLD1
polymerase (DNA directed), delta 1, catalytic subunit

125 kDa

IPI00221106
SF3B2
splicing factor 3b, subunit 2, 145 kDa

IPI00015905
EXOSC2
exosome component 2

IPI00031517
MCM6
minichromosome maintenance complex component 6

IPI00216694
PLS3
plastin 3

IPI00218407
ALDOB
aldolase B, fructose-bisphosphate

IPI00556369
SMG1
SMG1 homolog, phosphatidylinositol 3-kinase-

related kinase (C. elegans)

IPI00909083
GSPT1
G1 to S phase transition 1

IPI00479786
KHSRP
KH-type splicing regulatory protein

IPI00015953
DDX21
DEAD (Asp-Glu-Ala-Asp) box polypeptide 21

IPI00334907
PITPNB
phosphatidylinositol transfer protein, beta

IPI00297572
AQR
aquarius homolog (mouse)

IPI00011274
HNRPDL
heterogeneous nuclear ribonucleoprotein D-like

IPI00024095
ANXA3
annexin A3

IPI00293331
POP1
processing of precursor 1, ribonuclease P/MRP

subunit (S. cerevisiae)

IPI00007927
SMC2
structural maintenance of chromosomes 2

IPI00011592
DYNC1LI2
dynein, cytoplasmic 1, light intermediate chain 2

IPI00003927
PPID
peptidylprolyl isomerase D

IPI00002926
VPS37B
vacuolar protein sorting 37 homolog B (S. cerevisiae)

IPI00012497
ADRBK1
adrenergic, beta, receptor kinase 1

IPI00746351
DIS3
DIS3 mitotic control homolog (S. cerevisiae)

IPI00031960
POLR1A
polymerase (RNA) I polypeptide A, 194 kDa

IPI00290566
TCP1
t-complex 1

IPI00026952
PKP3
plakophilin 3

IPI00856120
LARP1B
La ribonucleoprotein domain family, member 1B

IPI00449049
PARP1
poly (ADP-ribose) polymerase 1

IPI00012035
CD46
CD46 molecule, complement regulatory protein

IPI00419979
PAK2
p21 protein (Cdc42/Rac)-activated kinase 2

IPI00303207
ABCE1
ATP-binding cassette, sub-family E (OABP),

member 1

IPI00872664
USP14
ubiquitin specific peptidase 14 (tRNA-guanine

transglycosylase)

IPI00290770
CCT3
chaperonin containing TCP1, subunit 3 (gamma)

IPI00294879
RANGAP1
Ran GTPase activating protein 1

IPI00013862
DTYMK
deoxythymidylate kinase (thymidylate kinase)

IPI00329692
NMT1
N-myristoyltransferase 1

IPI00037283
DNM1L
dynamin 1-like

IPI00008922
IFITM2
interferon induced transmembrane protein 2 (1-8D)

IPI00304754
FERMT1
fermitin family member 1

IPI00396203
TBCD
tubulin folding cofactor D

IPI00009071
LOC100505793/SRSF10
serine/arginine-rich splicing factor 10

IPI00022827
SLK
STE20-like kinase

IPI00918002
MUC5AC/MUC5B
mucin 5AC, oligomeric mucus/gel-forming

IPI00008240
MARS
methionyl-tRNA synthetase

IPI00217013
SMEK1
SMEK homolog 1, suppressor of mek1

(Dictyostelium)

IPI00219097
HMGB2
high mobility group box 2

IPI00304596
NONO
non-POU domain containing, octamer-binding

IPI00018219
TGFBI
transforming growth factor, beta-induced, 68 kDa

IPI00023824
FBLN2
fibulin 2

IPI00022228
HDLBP
high density lipoprotein binding protein

IPI00306322
COL4A2
collagen, type IV, alpha 2

IPI00018452
CPNE1
copine I

IPI00018627
NAA50
N(alpha)-acetyltransferase 50, NatE catalytic subunit

IPI00007163
LSM7
LSM7 homolog, U6 small nuclear RNA associated

(S. cerevisiae)

IPI00005154
SSRP1
structure specific recognition protein 1

IPI00007401
IPO8
importin 8

IPI00429538
YPEL5
yippee-like 5 (Drosophila)

IPI00030116
PGM3
phosphoglucomutase 3

IPI00162563
RNF40
ring finger protein 40

IPI00219420
SMC3
structural maintenance of chromosomes 3

IPI00012501
REG4
regenerating islet-derived family, member 4

IPI00029764
SF3A3
splicing factor 3a, subunit 3, 60 kDa

IPI00296099
THBS1
thrombospondin 1

IPI00027626
CCT6A
chaperonin containing TCP1, subunit 6A (zeta 1)

IPI00007928
PRPF8
PRP8 pre-mRNA processing factor 8 homolog (S. cerevisiae)

IPI00023344
SYMPK
symplekin

IPI00375441
FUBP1
far upstream element (FUSE) binding protein 1

IPI00005613
U2AF1
U2 small nuclear RNA auxiliary factor 1

IPI00002335
HTT
huntingtin

IPI00299254
EIF5B
eukaryotic translation initiation factor 5B

IPI00179953
NASP
nuclear autoantigenic sperm protein (histone-

binding)

IPI00514561
HNRNPK
heterogeneous nuclear ribonucleoprotein K

IPI00031812
YBX1
Y box binding protein 1

IPI00414320
ANXA11
annexin A11

IPI00178431
RECQL
RecQ protein-like (DNA helicase Q1-like)

IPI00029601
CTTN
cortactin

IPI00013683
TUBB3
tubulin, beta 3

IPI00646105
PYCRL
pyrroline-5-carboxylate reductase-like

IPI00298057
PPL
periplakin

IPI00173346
PGM2L1
phosphoglucomutase 2-like 1

IPI00373869
C17orf49
chromosome 17 open reading frame 49

IPI00106491
MRTO4
mRNA turnover 4 homolog (S. cerevisiae)

IPI00290184
METTL1
methyltransferase like 1

IPI00006025
SART3
squamous cell carcinoma antigen recognized by T

cells 3

IPI00016179
S100A13
S100 calcium binding protein A13

IPI00100292
NPEPL1
aminopeptidase-like 1

IPI00026689
CDK1
cyclin-dependent kinase 1

IPI00217405
UBR1
ubiquitin protein ligase E3 component n-recognin 1

IPI00296353
ARHGAP18
Rho GTPase activating protein 18

IPI00293434
SRP14
signal recognition particle 14 kDa (homologous Alu

RNA binding protein)

IPI00292532
CAMP
cathelicidin antimicrobial peptide

IPI00010740
SFPQ
splicing factor proline/glutamine-rich

IPI00026262
RASA1
RAS p21 protein activator (GTPase activating

protein) 1

IPI00409601
RALGAPB
Ral GTPase activating protein, beta subunit (non-

catalytic)

IPI00013976
LAMB1
laminin, beta 1

IPI00554590
RAB3GAP2
RAB3 GTPase activating protein subunit 2 (non-

catalytic)

IPI00302925
CCT8
chaperonin containing TCP1, subunit 8 (theta)

IPI00103247
HNRPLL
heterogeneous nuclear ribonucleoprotein L-like

IPI00414127
RANBP1
RAN binding protein 1

IPI00001458
KNTC1
kinetochore associated 1

IPI00221394
DKC1
dyskeratosis congenita 1, dyskerin

IPI00016613
CSNK2A1
casein kinase 2, alpha 1 polypeptide

IPI00013455
CLIP1
CAP-GLY domain containing linker protein 1

IPI00297779
CCT2
chaperonin containing TCP1, subunit 2 (beta)

IPI00029048
TTLL12
tubulin tyrosine ligase-like family, member 12

IPI00298306
ATM
ataxia telangiectasia mutated

IPI00017451
SF3A1
splicing factor 3a, subunit 1, 120 kDa

IPI00012493
RPS20
ribosomal protein S20

IPI00783378
UBE2O
ubiquitin-conjugating enzyme E2O

IPI00742682
TPR
translocated promoter region (to activated MET

oncogene)

IPI00002203
BCCIP
BRCA2 and CDKN1A interacting protein

IPI00291783
GEMIN5
gem (nuclear organelle) associated protein 5

IPI00019196
RPP30
ribonuclease P/MRP 30 kDa subunit

IPI00056386
LOH12CR1
loss of heterozygosity, 12, chromosomal region 1

IPI00019971
STXBP2
syntaxin binding protein 2

IPI00020965
UBE2H
ubiquitin-conjugating enzyme E2H

IPI00465045
DIP2B
DIP2 disco-interacting protein 2 homolog B

(Drosophila)

IPI00607591
RAP1GDS1
RAP1, GTP-GDP dissociation stimulator 1

IPI00171903
HNRNPM
heterogeneous nuclear ribonucleoprotein M

IPI00291802
LMO7
LIM domain 7

IPI00004273
RBM25
RNA binding motif protein 25

IPI00221234
ALDH7A1
aldehyde dehydrogenase 7 family, member A1

IPI00026219
CPSF1
cleavage and polyadenylation specific factor 1,

160 kDa

IPI00015262
CNN2
calponin 2

IPI00018465
CCT7
chaperonin containing TCP1, subunit 7 (eta)

IPI00014238
KARS
lysyl-tRNA synthetase

IPI00000684
UAP1
UDP-N-acteylglucosamine pyrophosphorylase 1

IPI00295485
HSPA4L
heat shock 70 kDa protein 4-like

IPI00926528
138 kDa protein
138 kDa protein

IPI00549189
THOP1
thimet oligopeptidase 1

IPI00008552
GLRX3
glutaredoxin 3

IPI00011200
PHGDH
phosphoglycerate dehydrogenase

IPI00014197
CDV3
CDV3 homolog (mouse)

IPI00411559
SMC4
structural maintenance of chromosomes 4

IPI00024320
RBM3
RNA binding motif (RNP1, RRM) protein 3

IPI00020956
HDGF
hepatoma-derived growth factor

IPI00479217
HNRNPU
heterogeneous nuclear ribonucleoprotein U (scaffold

attachment factor A)

IPI00604756
NRBP1
nuclear receptor binding protein 1

IPI00418797
POLR1B
polymerase (RNA) I polypeptide B, 128 kDa

IPI00019812
PPP5C
protein phosphatase 5, catalytic subunit

IPI00216008
G6PD
glucose-6-phosphate dehydrogenase

IPI00291560
ARG1
arginase, liver

IPI00008475
HMGCS1
3-hydroxy-3-methylglutaryl-CoA synthase 1 (soluble)

IPI00023234
UBA2
ubiquitin-like modifier activating enzyme 2

IPI00298991
KIAA1033
KIAA1033

IPI00793199
ANXA4
annexin A4

IPI00889541
DDX17
DEAD (Asp-Glu-Ala-Asp) box polypeptide 17

IPI00165393
ANP32E
acidic (leucine-rich) nuclear phosphoprotein 32

family, member E

IPI00896504
GNE
glucosamine (UDP-N-acetyl)-2-epimerase/N-

acetylmannosamine kinase

IPI00157790
KIAA0368
KIAA0368

IPI00182728
VPS4B
vacuolar protein sorting 4 homolog B (S. cerevisiae)

IPI00020127
RPA1
replication protein A1, 70 kDa

IPI00219678
EIF2S1
eukaryotic translation initiation factor 2, subunit 1

alpha, 35 kDa

IPI00290461
EIF3J
eukaryotic translation initiation factor 3, subunit J

IPI00784161
SUPT6H
suppressor of Ty 6 homolog (S. cerevisiae)

IPI00218993
HSPH1
heat shock 105 kDa/110 kDa protein 1

IPI00640703
XPO5
exportin 5

IPI00333215
TCEA1
transcription elongation factor A (SII), 1

IPI00009032
SSB
Sjogren syndrome antigen B (autoantigen La)

IPI00745313
AEBP1
AE binding protein 1

IPI00303258
LMCD1
LIM and cysteine-rich domains 1

IPI00219330
ILF3
interleukin enhancer binding factor 3, 90 kDa

IPI00019269
WDR61
WD repeat domain 61

IPI00386189
NAA15
N(alpha)-acetyltransferase 15, NatA auxiliary subunit

IPI00000015
SRSF4
serine/arginine-rich splicing factor 4

IPI00251559
RNF20
ring finger protein 20

IPI00006952
LACTB2
lactamase, beta 2

IPI00041325
NHP2
NHP2 ribonucleoprotein homolog (yeast)

IPI00607714
C17orf28
chromosome 17 open reading frame 28

IPI00645702
CTPS2
CTP synthase II

IPI00163187
FSCN1
fascin homolog 1, actin-bundling protein

(Strongylocentrotus purpuratus)

IPI00289807
TRNT1
tRNA nucleotidyl transferase, CCA-adding, 1

IPI00375462
SREK1
splicing regulatory glutamine/lysine-rich protein 1

IPI00025158
STAG1
stromal antigen 1

IPI00024971
OSBP
oxysterol binding protein

IPI00375015
DUT
deoxyuridine triphosphatase

IPI00396174
CCDC25
coiled-coil domain containing 25

IPI00020021
DEK
DEK oncogene

IPI00006378
CCDC72
coiled-coil domain containing 72

IPI00291093
POLR2E
polymerase (RNA) II (DNA directed) polypeptide E,

25 kDa

IPI00019178
PSPH
phosphoserine phosphatase

IPI00291939
SMC1A
structural maintenance of chromosomes 1A

IPI00006725
DDX23
DEAD (Asp-Glu-Ala-Asp) box polypeptide 23

IPI00009010
TRMT112
tRNA methyltransferase 11-2 homolog (S. cerevisiae)

IPI00018813
COPS2
COP9 constitutive photomorphogenic homolog

subunit 2 (Arabidopsis)

IPI00023640
PDCD5
programmed cell death 5

IPI00031681
CDK2
cyclin-dependent kinase 2

IPI00011603
PSMD3
proteasome (prosome, macropain) 26S subunit,

non-ATPase, 3

IPI00221325
RANBP2
RAN binding protein 2

IPI00410693
SERBP1
SERPINE1 mRNA binding protein 1

IPI00005780
OGT
O-linked N-acetylglucosamine (GlcNAc) transferase

(UDP-N-acetylglucosamine:polypeptide-N-

acetylglucosaminyl transferase)

IPI00299524
NCAPD2
non-SMC condensin I complex, subunit D2

IPI00150057
SMARCC2
SWI/SNF related, matrix associated, actin

dependent regulator of chromatin, subfamily c,

member 2

IPI00032853
NOP10
NOP10 ribonucleoprotein homolog (yeast)

IPI00002255
LRBA
LPS-responsive vesicle trafficking, beach and

anchor containing

IPI00554742
API5
apoptosis inhibitor 5

IPI00385267
SRPR
signal recognition particle receptor (docking protein)

IPI00022462
TFRC
transferrin receptor (p90, CD71)

IPI00022305
BZW2
basic leucine zipper and W2 domains 2

IPI00025427
RNASE3
ribonuclease, RNase A family, 3

IPI00010182
DBI
diazepam binding inhibitor (GABA receptor

modulator, acyl-CoA binding protein)

IPI00219005
FKBP4
FK506 binding protein 4, 59 kDa

IPI00184871
C6orf130
chromosome 6 open reading frame 130

IPI00103483
COBRA1
cofactor of BRCA1

IPI00026215
FEN1
flap structure-specific endonuclease 1

IPI00296635
GBE1
glucan (1,4-alpha-), branching enzyme 1

IPI00029267
SNRPB2
small nuclear ribonucleoprotein polypeptide B

IPI00100197
NSFL1C
NSFL1 (p97) cofactor (p47)

IPI00010415
ACOT7
acyl-CoA thioesterase 7

IPI00306369
NSUN2
NOP2/Sun domain family, member 2

IPI00302927
CCT4
chaperonin containing TCP1, subunit 4 (delta)

IPI00023845
KLK6
kallikrein-related peptidase 6

IPI00003919
QPCT
glutaminyl-peptide cyclotransferase

IPI00465128
BAG6
BCL2-associated athanogene 6

IPI00016910
EIF3C/EIF3CL
eukaryotic translation initiation factor 3, subunit C

IPI00007812
ATP6V1B2
ATPase, H+ transporting, lysosomal 56/58 kDa, V1

subunit B2

IPI00027846
MMP8
matrix metallopeptidase 8 (neutrophil collagenase)

IPI00023919
PSMC5
proteasome (prosome, macropain) 26S subunit,

ATPase, 5

IPI00217253
GCHFR
GTP cyclohydrolase I feedback regulator

IPI00384028
PAPOLA
poly(A) polymerase alpha

IPI00219344
HPCAL1
hippocalcin-like 1

IPI00012442
G3BP1
GTPase activating protein (SH3 domain) binding

protein 1

IPI00024163
POLR3A
polymerase (RNA) III (DNA directed) polypeptide A,

155 kDa

IPI00647217
SKIV2L2
superkiller viralicidic activity 2-like 2 (S. cerevisiae)

IPI00031627
POLR2A
polymerase (RNA) II (DNA directed) polypeptide A,

220 kDa

IPI00873137
COL1A2
collagen, type I, alpha 2

IPI00025039
FBL
fibrillarin

IPI00923606
EPRS
glutamyl-prolyl-tRNA synthetase

IPI00301936
ELAVL1
ELAV (embryonic lethal, abnormal vision,

Drosophila)-like 1 (Hu antigen R)

IPI00183500
NCBP2
nuclear cap binding protein subunit 2, 20 kDa

IPI00001159
GCN1L1
GCN1 general control of amino-acid synthesis 1-like

1 (yeast)

IPI00024719
HAT1
histone acetyltransferase 1

IPI00470883
STAG2
stromal antigen 2

IPI00216057
SORD
sorbitol dehydrogenase

IPI00032830
REXO2
REX2, RNA exonuclease 2 homolog (S. cerevisiae)

IPI00003881
HNRNPF
heterogeneous nuclear ribonucleoprotein F

IPI00216230
TMPO
thymopoietin

IPI00902614
USP24
ubiquitin specific peptidase 24

IPI00182757
KIAA1967
KIAA1967

IPI00296165
C1R
complement component 1, r subcomponent

IPI00002460
ANXA7
annexin A7

IPI00009104
RUVBL2
RuvB-like 2 (E. coli)

IPI00651738
ADI1
acireductone dioxygenase 1

IPI00009328
EIF4A3
eukaryotic translation initiation factor 4A3

IPI00456887
HNRNPUL2
heterogeneous nuclear ribonucleoprotein U-like 2

IPI00646605
UBR4
ubiquitin protein ligase E3 component n-recognin 4

IPI00386718
SMARCA2
SWI/SNF related, matrix associated, actin

dependent regulator of chromatin, subfamily a,

member 2

IPI00008234
CYB5R2
cytochrome b5 reductase 2

IPI00300371
SF3B3
splicing factor 3b, subunit 3, 130 kDa

IPI00156282
GPS1
G protein pathway suppressor 1

IPI00012369
MAD2L1
MAD2 mitotic arrest deficient-like 1 (yeast)

IPI00016736
PLCG1
phospholipase C, gamma 1

IPI00014263
EIF4H
eukaryotic translation initiation factor 4H

IPI00031556
U2AF2
U2 small nuclear RNA auxiliary factor 2

IPI00304589
TNKS1BP1
tankyrase 1 binding protein 1, 182 kDa

IPI00294578
TGM2
transglutaminase 2 (C polypeptide, protein-

glutamine-gamma-glutamyltransferase)

IPI00027834
HNRNPL
heterogeneous nuclear ribonucleoprotein L

IPI00329213
INPP5D
inositol polyphosphate-5-phosphatase, 145 kDa

IPI00296528
ANXA10
annexin A10

IPI00013180
BUD31
BUD31 homolog (S. cerevisiae)

IPI00216048
PITPNA
phosphatidylinositol transfer protein, alpha

IPI00103994
LARS
leucyl-tRNA synthetase

IPI00027681
NNMT
nicotinamide N-methyltransferase

IPI00185374
PSMD12
proteasome (prosome, macropain) 26S subunit,

non-ATPase, 12

IPI00004838
CRK
v-crk sarcoma virus CT10 oncogene homolog

(avian)

IPI00847535
PRG2
proteoglycan 2, bone marrow (natural killer cell

activator, eosinophil granule major basic protein)

IPI00009802
VCAN
versican

IPI00306290
XPOT
exportin, tRNA (nuclear export receptor for tRNAs)

IPI00025347
EMG1
EMG1 nucleolar protein homolog (S. cerevisiae)

IPI00410091
C11orf73
chromosome 11 open reading frame 73

IPI00024364
TNPO1
transportin 1

IPI00292150
LTBP2
latent transforming growth factor beta binding

protein 2

IPI00470891
CSDE1
cold shock domain containing E1, RNA-binding

IPI00168554
SRXN1
sulfiredoxin 1

IPI00103525
PSPC1
paraspeckle component 1

IPI00419880
RPS3A
ribosomal protein S3A

IPI00375631
ISG15
ISG15 ubiquitin-like modifier

IPI00027808
POLR2B
polymerase (RNA) II (DNA directed) polypeptide B,

140 kDa

IPI00054042
GTF2I
general transcription factor IIi

IPI00026167
NHP2L1
NHP2 non-histone chromosome protein 2-like 1 (S. cerevisiae)

IPI00003565
PSMD10
proteasome (prosome, macropain) 26S subunit,

non-ATPase, 10

IPI00030781
STAT1
signal transducer and activator of transcription 1,

91 kDa

IPI00293026
EFTUD1
elongation factor Tu GTP binding domain containing 1

IPI00646226
MED23
mediator complex subunit 23

IPI00745433
EIF2C2
eukaryotic translation initiation factor 2C, 2

IPI00027838
RBM4B
RNA binding motif protein 4B

IPI00024425
KIAA0664
KIAA0664

IPI00024871
CBFB
core-binding factor, beta subunit

IPI00008524
PABPC1
poly(A) binding protein, cytoplasmic 1

IPI00018873
NAMPT
nicotinamide phosphoribosyltransferase

IPI00216088
CRABP2
cellular retinoic acid binding protein 2

IPI00032342
TRIP12
thyroid hormone receptor interactor 12

IPI00154975
DNAJC9
DnaJ (Hsp40) homolog, subfamily C, member 9

IPI00186008
STARD10
StAR-related lipid transfer (START) domain

containing 10

IPI00828098
RNF213
ring finger protein 213

IPI00011898
EIF2B5
eukaryotic translation initiation factor 2B, subunit 5

epsilon, 82 kDa

IPI00301434
BOLA2/BOLA2B
bolA homolog 2 (E. coli)

IPI00181391
MGEA5
meningioma expressed antigen 5 (hyaluronidase)

IPI00022664
RABGGTA
Rab geranylgeranyltransferase, alpha subunit

IPI00384689
PDXDC1
pyridoxal-dependent decarboxylase domain

containing 1

IPI00552920
EXOSC8
exosome component 8

IPI00001734
PSAT1
phosphoserine aminotransferase 1

IPI00010105
EIF6
eukaryotic translation initiation factor 6

IPI00745105
C16orf13
chromosome 16 open reading frame 13

IPI00306436
STAT3
signal transducer and activator of transcription 3

(acute-phase response factor)

IPI00220815
EFEMP1
EGF containing fibulin-like extracellular matrix

protein 1

IPI00008301
DEFA6
defensin, alpha 6, Paneth cell-specific

IPI00015018
PPA1
pyrophosphatase (inorganic) 1

IPI00298176
GPX2
glutathione peroxidase 2 (gastrointestinal)

IPI00456635
UNC13D
unc-13 homolog D (C. elegans)

IPI00218604
PTPN6
protein tyrosine phosphatase, non-receptor type 6

IPI00760846
MYO18A
myosin XVIIIA

IPI00296534
FBLN1
fibulin 1

IPI00025329
RPL19
ribosomal protein L19

IPI00030876
DIAPH1
diaphanous homolog 1 (Drosophila)

IPI00154451
MMS19
MMS19 nucleotide excision repair homolog (S. cerevisiae)

IPI00646917
NUDT21
nudix (nucleoside diphosphate linked moiety X)-type

motif 21

IPI00010404
SF3B5
splicing factor 3b, subunit 5, 10 kDa

IPI00014938
SARNP
SAP domain containing ribonucleoprotein

IPI00002186
ARFGEF2
ADP-ribosylation factor guanine nucleotide-

exchange factor 2 (brefeldin A-inhibited)

IPI00028564
GBP1
guanylate binding protein 1, interferon-inducible

IPI00179298
HUWE1
HECT, UBA and WWE domain containing 1

IPI00027142
POP7
processing of precursor 7, ribonuclease P/MRP

subunit (S. cerevisiae)

IPI00006504
EIF2B3
eukaryotic translation initiation factor 2B, subunit 3

gamma, 58 kDa

IPI00013184
NAA10
N(alpha)-acetyltransferase 10, NatA catalytic subunit

IPI00015947
DNAJB1
DnaJ (Hsp40) homolog, subfamily B, member 1

IPI00019463
EIF2AK2
eukaryotic translation initiation factor 2-alpha kinase 2

IPI00003519
EFTUD2
elongation factor Tu GTP binding domain containing 2

IPI00004524
GCA
grancalcin, EF-hand calcium binding protein

IPI00333776
NRCAM
neuronal cell adhesion molecule

IPI00552073
DHX16
DEAH (Asp-Glu-Ala-His) box polypeptide 16

IPI00013949
SGTA
small glutamine-rich tetratricopeptide repeat (TPR)-

containing, alpha

IPI00304742
STK10
serine/threonine kinase 10

IPI00305833
SMU1
smu-1 suppressor of mec-8 and unc-52 homolog (C. elegans)

IPI00150269
PRPF4
PRP4 pre-mRNA processing factor 4 homolog

(yeast)

IPI00028888
HNRNPD
heterogeneous nuclear ribonucleoprotein D (AU-rich

element RNA binding protein 1, 37 kDa)

IPI00290142
CTPS
CTP synthase

IPI00015195
CSTF3
cleavage stimulation factor, 3′ pre-RNA, subunit 3,

77 kDa

IPI00011528
CSTF1
cleavage stimulation factor, 3′ pre-RNA, subunit 1,

50 kDa

IPI00013070
HNRNPUL1
heterogeneous nuclear ribonucleoprotein U-like 1

IPI00008433
RPS5
ribosomal protein S5

IPI00298347
PTPN11
protein tyrosine phosphatase, non-receptor type 11

IPI00514234
LAD1
ladinin 1

IPI00000057
COG2
component of oligomeric golgi complex 2

IPI00014311
CUL2
cullin 2

IPI00221093
RPS17/RPS17L
ribosomal protein S17

IPI00022694
PSMD4
proteasome (prosome, macropain) 26S subunit,

non-ATPase, 4

IPI00218918
ANXA1
annexin A1

IPI00418169
ANXA2
annexin A2

IPI00289499
ATIC
5-aminoimidazole-4-carboxamide ribonucleotide

formyltransferase/IMP cyclohydrolase

IPI00022246
AZU1
azurocidin 1

IPI00299635
BIRC6
baculoviral IAP repeat containing 6

IPI00002147
CHI3L1
chitinase 3-like 1 (cartilage glycoprotein-39)

IPI00011062
CPS1
carbamoyl-phosphate synthase 1, mitochondrial

IPI00028064
CTSG
cathepsin G

IPI00005721
DEFA1 (includes
defensin, alpha 1

others)

IPI00099110
DMBT1
deleted in malignant brain tumors 1

IPI00027769
ELANE
elastase, neutrophil expressed

IPI00217987
ITGAM
integrin, alpha M (complement component 3

receptor 3 subunit)

IPI00299547
LCN2
lipocalin 2

IPI00023673
LGALS3BP
lectin, galactoside-binding, soluble, 3 binding protein

IPI00922181
MCM2
minichromosome maintenance complex component 2

IPI00027509
MMP9
matrix metallopeptidase 9 (gelatinase B, 92 kDa

gelatinase, 92 kDa type IV collagenase)

IPI00007244
MPO
myeloperoxidase

IPI00103397
MUC5AC/MUC5B
mucin 5AC, oligomeric mucus/gel-forming

IPI00019380
NCBP1
nuclear cap binding protein subunit 1, 80 kDa

IPI00299512
NF1
neurofibromin 1

IPI00022255
OLFM4
olfactomedin 4

IPI00303063
PDS5A
PDS5, regulator of cohesion maintenance, homolog

A (S. cerevisiae)

IPI00021085
PGLYRP1
peptidoglycan recognition protein 1

IPI00027409
PRTN3
proteinase 3

IPI00003590
QSOX1
quiescin Q6 sulfhydryl oxidase 1

IPI00009027
REG1A
regenerating islet-derived 1 alpha

IPI00027462
S100A9
S100 calcium binding protein A9

IPI00010304
SERPINB10
serpin peptidase inhibitor, clade B (ovalbumin),

member 10

IPI00783625
SERPINB5
serpin peptidase inhibitor, clade B (ovalbumin),

member 5

IPI00072534
UNC45A
unc-45 homolog A (C. elegans)

TABLE 7

A subset of biomarkers from Table 1 which were also significantly more

secreted by colon tumor samples than by normal colon (Table 6).

Gene

Accession
Symbol
Entrez Gene Name

IPI00418169
ANXA2
annexin A2

IPI00022246
AZU1
azurocidin 1

IPI00028064
CTSG
cathepsin G

IPI00005721
DEFA1
defensin, alpha 1

(includes

others)

IPI00027769
ELANE
elastase, neutrophil expressed

IPI00217987
ITGAM
integrin, alpha M (complement component 3

receptor 3 subunit)

IPI00007244
MPO
myeloperoxidase

IPI00019380
NCBP1
nuclear cap binding protein subunit 1,

80 kDa

IPI00027409
PRTN3
proteinase 3

IPI00027462
S100A9
S100 calcium binding protein A9

IPI00010304
SERPINB10
serpin peptidase inhibitor, clade B

(ovalbumin), member 10

IPI00218918
ANXA1
annexin A1

IPI00299635
BIRC6
baculoviral IAP repeat containing 6

IPI00922181
MCM2
minichromosome maintenance complex

component 2

IPI00003590
QSOX1
quiescin Q6 sulfhydryl oxidase 1

IPI00783625
SERPINB5
serpin peptidase inhibitor, clade B

(ovalbumin), member 5

The invention is further described by the following, illustrative examples. These are not intended to limit the scope of the invention.

Example 1
Materials & Methods

Stool Samples and Protein Extraction

Written informed consent was obtained from all subjects who provided stool samples and this study was approved by the Medical Ethical Committee of the VU University Medical Center, The Netherlands.

Partial stool samples were collected from referral subjects who underwent colonoscopy between November 2003 and June 2006 at the VU University Medical Center in Amsterdam, The Netherlands. Partial stool samples were collected from before colonoscopy or following diagnosis at colonoscopy and prior to surgical resection of their tumors (see Table 8 for patient characteristics).

Independent (Verification) Set of Stool Samples

Homogenized whole stool samples were collected from subjects referred for and underwent colonoscopy between July 2009 and April 2011 at the VU University Medical Center in Amsterdam, The Netherlands. Whole stool samples were collected before colonoscopy (see table 1 for patient characteristics). The study participants immediately added stabilization buffer to the stool samples after defecation, the samples were processed in the lab with a final stool:buffer w/v ratio of 1:7 within 72 hours, and stored at −80° C. until use. Protein extracts were prepared as described above using 2 ml homogenized stool sample as starting material. Equal amounts of protein extracts were mixed and further treated as a single sample. Four pools from different categories were composed i.e. controls (n=5), and individuals with non-advanced adenomas (n=5), advanced adenomas (n=5) and CRC (n=5) (see Table 8 for patient characteristics).

TABLE 8

Clinicopathological characteristics of patients

Tumor

Patient

Size
Dukes

Positive

number¹
Age
Gender
Category
(mm)
Stage
Tumor location
FIT²

A. Discovery set

1
74
f
CRC
40
C
coecum
+

2
85
m
CRC
35
A
right sided
−

3
86
m
CRC
30
A
descendens/sigmoid
−

4
82
f
CRC
59
B
right sided
−

5
60
m
CRC
15
B
rectosigmoid
+

6
85
m
CRC
60
B
right sided
+

7
83
m
CRC
70
B
flexura hepatica
−

8
71
f
CRC
35
A
sigmoid
+

9
63
f
CRC
50
C
rectosigmoid
+

10
59
m
CRC
52
C
coecum
+

11
84
f
CRC
50
C
ileocoecaal
+

12
86
m
CRC
68
C
ascendens
+

13
69
m
control
na
na
na
−

14
75
m
control
na
na
na
−

15
68
f
control
na
na
na
−

16
73
f
control
na
na
na
−

17
75
f
control
na
na
na
−

18
67
f
control
na
na
na
−

19
62
m
control
na
na
na
−

20
60
f
control
na
na
na
−

21
60
m
control
na
na
na
−

22
58
m
control
na
na
na
−

B. Verification set

23
43
f
control
na
na
na
−

24
59
f
control
na
na
na
−

25
44
f
control
na
na
na
−

26
48
m
control
na
na
na
−

27
78
m
control
na
na
na
−

28
78
m
A
2
na
sigmoid
−

29
68
m
A
3
na
ascendens
NA

30
55
f
A
2
na
sigmoid
NA

31
72
m
A
8
na
sigmoid
−

32
66
f
A
7
na
Sigmoid
+

33
84
f
AA
8
na
Sigmoid
NA

34
78
f
AA
10
na
Sigmoid
−

35
87
m
AA
2
na
descendens
−

36
78
f
AA
10
na
ascendens
+

37
78
m
AA
4
na
sigmoid
+

38
89
f
CRC
30
C
rectosigmoid
NA

39
58
m
CRC
50
B
Rectum
NA

40
71
m
CRC
35
C
ascendens/flex hepatica
+

41
72
m
CRC
40
A
Sigmoid
−

42
70
f
CRC
15
C
ascendens
−

^aThis column represents an anonymization number code

^bAs per the Modified Astler Coller classification (Compton and Greene, CA Cancer J Clin 2004)

^ccut-off >75 ng/ul (van Veen, Ned tijdschriftGeneeskd, 2009)

At collection, stool samples were immediately stored at 4° C. and transferred to −20° C. within 36 hours. Stool samples were thawed and after performing FIT (fecal immunochemical test; FIT), ˜1 g stool was sampled from each stool sample for protein extraction. For this study stool samples from 10 subjects without colon neoplasia and 4 stool samples from CRC patients with a negative FIT score were selected next to 8 stool samples from CRC patients with a positive FIT score.

Stool proteins were extracted as described before (Ang C S, Nice E C. Targeted in-gel MRM: a hypothesis driven approach for colorectal cancer biomarker discovery in human feces. J Proteome Res 2010; 9: 4346-55) with few adaptations. In short, samples were homogenized in a two-fold excess volume of PBS by vortexing and centrifuged at 4° C. for 15 minutes at 16.000 G. The supernatants were centrifuged once more at 4° C. for 10 minutes at full speed. Following the last spin cycle the supernatants were cleaned from remaining particles by filtering through a 0.22 μM PVDF filter (Millipore, Billerica, Mass., USA). Finally, the samples were concentrated to approximately 200 μl using a 3 kDa cut-off filter (Amicon Ultra, Millipore Corporation, Billerica, Mass., USA).

1D-SDS Gel Electrophoresis and Sample Processing for Proteomics Analysis

Equal protein amounts (˜30 μg) were loaded and separated on precast 4-12% gradient gels (Invitrogen, Carlsbad, USA), fixed in 50% ethanol containing 3% phosphoric acid washed and stained overnight with Coomassie R-250. After staining the gels were washed in MilliQ water and stored at 4° C. until processing for in-gel digestion. Each lane was cut in 10 equal individual bands and each band was further processed into tryptic peptides as described before (Albrethsen J, et al. Mol Cell Proteomics 2010; 9: 988-1005; Piersma S R, et al. J Proteome Res 2010; 9: 1913-22).

For samples from an independent stool collection (verification experiment) equal amounts of the samples (20 μl) were loaded on a 12.5% SDS-PAGE gel and run into the gel until the proteins entered the running gel. Then the gel was fixed and stained as described above. The samples were cut out of the gel as a single band and further processed to tryptic peptides as described before (Albrethsen J, et al. Mol Cell Proteomics 2010; 9: 988-1005; Piersma S R, et al. J Proteome Res 2010; 9: 1913-22). The peptides were extracted and the volume of the desalted peptide fractions was reduced to 50 μl in a vacuum centrifuge.

nanoLC-MS/MS Proteomics Analysis

Peptides were separated by an Ultimate 3000 nanoLC system (Dionex LC-Packings, Amsterdam, The Netherlands) equipped with a 20 cm×75 μm ID fused silica column custom packed with 3 μm 120 Å ReproSil Pur C18 aqua (Dr Maisch GMBH, Ammerbuch-Entringen, Germany) as described previously (Piersma S R, et al. J Proteome Res 2010; 9: 1913-22). Peptides were trapped on a 5 mm×300 μm ID Pepmap C18 cartridge (Dionex LC-Packings, Amsterdam, The Netherlands) and separated at 300 nl/min in a 60 min gradient. Intact peptide mass spectra and fragmentation spectra (top 5) were acquired on a LTQ-FT hybrid mass spectrometer (Thermo Fisher, Bremen, Germany). Dynamic exclusion was applied with a repeat count of 1 and an exclusion time of 30 seconds. Peptides from the four pooled samples (verification experiment) were separated in triplicate on a 75 μm×50 cm custom packed Reprosil C18 aqua column (1.9 μm, 120 Å) in a 240 min. gradient (5-32% Acetonitrile+0.5% Acetic acid at 300 nl/min) using a U3000 RSLC high pressure nanoLC (Dionex). Eluting peptides were measured on-line by a Q Exactive mass spectrometer (ThermoFisher Scientific) operating in data-dependent acquisition mode. Peptides were ionized using a stainless-steel emitter at a potential of +2 kV (ThermoScientific). Intact peptide ions were detected at a resolution of 35000 and fragment ions at a resolution of 17500; the MS mass range was 350-1500 Da. AGC Target settings for MS were 3E6 charges and for MS/MS 2E5 charges. Peptides were selected for HCD fragmentation at an underfill ratio of 1% and a quadrupole isolation window of 1.5 Da, peptides were fragmented at normalized collision energy of 30 eV.

SRM Analysis

LC-SRM analysis was performed on the four pools from the verification set. Chromatographic separation of peptides was performed by an Ultimate 3000 RSCL Nano LC system (Dionex) equipped with custom packed nano-LC columns consisting of 20 cm×75 μm ID fused silica custom packed with 3 μm 100 Å ReproSil Pur C18 aqua (Dr Maisch GMBH, Ammerbuch-Entringen, Germany) as described before¹⁶. Samples were analyzed in triplicate on a QTRAP 5500 instrument (AB SCIEX, Foster City, Calif.) operated in positive SRM mode and equipped with a nano-electrospray source with applied voltage of 2.2 kV and a capillary heater temperature of 225° C. The scheduled SRM mode comprised the following parameters: SRM detection window of 900 sec, target scan time of 3.0 s, curtain gas of 15, ion source gas 1 of 25, declustering potential of 100, entrance potential of 10. Q1 resolution was set to unit and Q3 resolution to unit. Pause between mass ranges was set to 1 ms. Collision cell exit potentials was set to 36 for all transitions.

SRM Assay Development

An SRM assay for the target proteins was developed using the MRMPilot™ software version 2.1 (AB SCIEX, Concord, ON, Canada). For each included protein, 5 peptides were selected and purchased from JPT Peptide Technologies (Berlin, Germany) as non-purified ‘SpikeTides’. For each protein one μl of a mixture of the 5 selected peptides was injected at a concentration of approximately 50 fmol per peptide. For each of the peptides, the MRMPilot™ software was set to generate up to 20 theoretically possible SRM transitions, each consisting of the calculated m/z of the precursor ion (at any charge state predicted by the software) in combination with the predicted fragment ions for each predicted precursor. The highest responding peptides/transitions at a theoretically calculated optimum collision energy were determined, as well as the identity of the peptide via SRM triggered MS/MS. Each verification analysis was set up to detect 100 of all theoretically predicted transitions and their theoretically predicted optimum collision energy corresponding to the 5 peptides assessed for each candidate protein. The combined information from each SRM—Information Dependent Acquisition experiment was used to perform Mascot searches against the human Swiss-prot protein database and MultiQuant™ software version 2.1 (AB SCIEX). MultiQuant™ software was also used to generate method files for peptide verification and collision energy optimization and to integrate the results of all optimization cycles. MRMPilot was used to schedule three transitions at the experimentally found optimum and the experimentally found retention time for each peptide. The final assay contained 114 scheduled transitions, 3 for each peptide (1-5 peptides for each of the candidate proteins) and 6 for 2 external control peptides. SRM analyses on each pool were carried out in triplicate.

SRM Data Analysis

The area under the curve (AUC) and retention time of each transition was determined using Multiquant software (AB SCIEX). Obtained transitions were then verified by two characteristics; firstly a retention time check was carried out to reveal false positives or false negatives in the assignment of transitions. If the retention time of three transitions belonging to 1 peptide was >3 seconds, the three transitions were manually inspected to assess correctness of assignment of the three transitions by the Multiquant software. If this occurred in more than one of the technical replicates then the peptide for that sample was disregarded. Secondly, the consistency of the ratio between two areas under the curve of two transitions (Transition1/Transition2, Transition2/Transition3 and Transition1/Transition3) for each peptide in the CRC pool was verified. A relative standard deviation (RSD) percentage was calculated of these ratios in the triplicate analysis. If the one transition caused the RSD percentage for two ratio's to be >20%, the transitions were manually inspected and if only one transition appeared to be incorrect, the two transitions resulting in <20% RSD over all analyses were selected for further analysis. Again if all three transition ratios had a RSD percentage above 20 then the peptide for that sample was disregarded.

Database Searching and Statistical Analysis

MS/MS spectra were searched against the human IPI database 3.62 (83.947 entries) using Sequest (version 27, rev 12). Scaffold version 3.00 (Proteomesoftware, Portland, Oreg.) was used to organize the data and to validate peptide identifications using the PeptideProphet (probability >95%) and ProteinProphet (probability of >99% with 2 peptides or more in one sample) (Keller A, et al. Anal Chem 2002; 74: 5383-92; Nesvizhskii A I, et al. Anal Chem 2003; 75: 4646-58). The software programs SecretomeP and SignalP were used for prediction of non-classical secretion and presence of a signal peptide (Bendtsen J D, et al. Protein Eng Des Sel 2004; 17: 349-56; Bendtsen J D, et al. J Mol Biol 2004; 340: 783-95). Q Exactive MS/MS spectra were searched against the human IPI database 3.68 using MaxQuant 1.2.2.5. Maximum allowed mass deviation was set to 20 ppm for MS and MS/MS. Cysteine carboxamide methylation was set as fixed modification and methionine oxidation and protein N-terminal acetylation were set as variable modifications. Identifications were filtered to 1% FDR at both the peptide and protein level. For each protein the number of assigned MS/MS spectra was summed across the 10 fractions and exported to Excel. Normalized counts were calculated by dividing the counts per protein by the sum of all counts per sample and multiciplication by the average sum across all samples. Differential analysis of proteins present in stool samples from CRC patients versus stool samples from healthy controls was performed using the beta-binomial test. The beta-binomial test takes into account the within-sample variation and the between-sample variation in a single statistical model (Pham T V, et al. Bioinformatics 2010; 26: 363-9).

Fecal Immunochemical Test Analysis

FIT samples (OC-Sensor®, Eiken Chemical Co., Tokyo, Japan) were processed with the OC sensor MICRO desktop analyser (Eiken Chemical) and analyzed according to the manufacturer's instructions. A cut-off of 75 ng/ml was used to determine a positive test result (van Veen W, Mali W P. [Colorectal cancer screening: advice from the Health Council of the Netherlands]. Ned Tijdschr Geneeskd 2009; 153:A1441).

Results

Human Protein Identification in Stool Samples

In total 830 human proteins were identified in at least one of the 22 stool samples. Of these, 624 proteins were identified both in CRC and control stool samples, 164 proteins were detected only in CRC stool samples, and 42 proteins were detected only in control stool samples (see FIG. 1A). On average the number of identified proteins was consistent across all samples and did not differ significantly between CRC and control samples (326 and 296 proteins, respectively).

The primary annotation of subcellular localization of the proteins was mainly the cytoplasm (35%) and the extracellular space (16%) and did not differ between CRC and control samples (see FIG. 1B). When the protein sequences were examined for secretion signals with the software tools signalP indicating signal peptides and secretomeP indicating non-classical secretion, 38% of proteins were predicted to have a signal peptide and the majority of proteins 64% were predicted to be a secreted protein. These data indicate that a high percentage of human proteins in stool samples consists of secreted proteins, e.g. in comparison to only 13% of proteins with a signal peptide in a cell line lysate (Piersma S R, et al. J Proteome Res 2010; 9: 1913-22).

Verification of LC-MS/MS Protein Quantification

The fecal immunochemical test (FIT) is used in many countries as a non-invasive test for the early detection of CRC, and is based on the detection of hemoglobin. To verify the results obtained by LC-MS/MS, hemoglobin spectral counts were compared with FIT values (ng/ml) determined in the same stool samples. The adult hemoglobin protein is a heterodimer consisting of two α and two β chains (Schechter A N. Blood 2008; 112: 3927-38). As can be seen in FIGS. 2A and 2B, LC-MS/MS data on both the α and β chain of hemoglobin significantly correlated to FIT values (both p=0.04). In addition, a very strong correlation was observed between the spectral counts of the α and β chain (see FIG. 2C; Pearson correlation of 0.98, p=1.1*10⁻¹⁴)

Another protein which has been frequently detected in stool and associated to CRC is Calprotectin (Bosch L J, et al. Molecular tests for colorectal cancer screening. Clin Colorectal Cancer 2011; 10:8-23). Calprotectin is a heterodimer as well, consisting of S100A8 and S100A9 subunits (Yui S, et al. Biol Pharm Bull 2003; 26: 753-60). A strong correlation was observed between spectral counts of S100A8 and S100A9 (see FIG. 2D; Pearson correlation of 0.92, p=1.2*10⁻⁹), further validating the LC-MS/MS data obtained from stool.

These results on hemoglobin and calprotectin show that LC-MS/MS on stool samples provides us with robust quantification of proteins, and is a valid approach for protein marker discovery.

Origin of Human Stool Proteins

The mechanisms of how tumor-derived biomarkers end up in stool can be broadly divided in leaked markers, secreted markers and exfoliated markers (Osborn N K, Ahlquist DA. Gastroenterology 2005; 128: 192-206). Hemoglobin is a typical example of a leaked marker from disturbed blood vessels in a neoplastic lesion. Secreted and exfoliated markers can be derived from the epithelial cells lining the colorectal lumen, but can also originate from cells in the surrounding stroma, such as immune cells. An overlap analysis with a previously obtained dataset of plasma proteins (unpublished data) together with the public available Human Proteome Organisation (HUPO), Human Plasma Proteome Project (HPPP) database (www.hupo.org/research/hppp; Omenn G S, et al. Proteomics 2005; 5: 3226-45; States D J, et al. Nat Biotechnol 2006; 24: 333-8) (high confidence list) revealed that 21.6% of the 830 identified proteins possibly originate from blood. To estimate how many of the 830 identified human proteins originate from epithelial cells, an overlap analysis was performed with proteins detected in CRC cell lines (unpublished data). Almost half of these proteins were also identified in the CRC cell lines suggesting an epithelial origin.

Stool Proteins that Discriminate CRC Patients from Control Subjects

Unsupervised hierarchical cluster analysis of human proteins identified in all CRC and control stool samples revealed two clusters. Cluster one grouped nine CRC stool samples together, and cluster two grouped all ten control stool samples and three CRC stool samples together (data not shown). Stool samples from CRC patients thus show a specific protein expression pattern as compared to stool from control subjects. This protein expression pattern can therefore be applied to discriminate most of the CRC stool samples from control stool samples without further selection of specific proteins. The complete stool protein dataset was analyzed with beta binomial statistics to identify potential cancer-associated proteins (Pham T V, et al. Bioinformatics 2010; 26: 363-9). From the 830 human stool proteins, 221 proteins were differentially detected, of which the levels of 134 proteins were significantly higher in CRC compared to control stool samples (p<0.05; Table 2), while 87 proteins were significantly lower in controls compared to CRC stool samples.

Cluster analysis based on the 221 differentially detected proteins in CRC and control stool samples revealed two clusters as well (data not shown). Again, the nine CRC stool samples clustered together in one cluster, and the same three CRC stool samples mentioned above clustered together with the control stool samples in the second cluster.

Within the second cluster, the three CRC stool samples grouped together with one control sample. The protein expression pattern of these three CRC stool samples contained aspects of both the control stool samples and the other nine CRC stool samples, indicating that a selection of these proteins would be able to discriminate these CRC samples from controls.

In addition, three out of four FIT negative CRC patients clustered together with FIT positive CRC patients. Therefore this group of proteins contains potential candidate biomarkers that complement FIT.

Verification by Semi-High Throughput nanoLC-MS/MS

In order to verify these initial results, four pools were created from an independent set of 20 stool samples. Next to CRC cases and negative controls, also samples from patients with adenomas were included. In fact, since most adenomas do not progress to cancer, for the purpose of CRC screening, only the detection of adenomas with high risk for progression besides CRC matters (Levin B et al., Gastroenterology 2008; 134:1570-1595). Therefore, pools of stool samples from patients with non-advanced adenoma as well as advanced adenomas (generally regarded as of higher risk to progression) were included. In these verification samples, 414 human proteins were identified. Out of the 830 human proteins identified in the discovery set, 331 (40%) were also detected in the verification set (see table 5). From the 134 human proteins significantly enriched in CRC stool samples, 63 (47%) were also detected in the verification set. The proteins detected in both the discovery and the verification set represented proteins with a significantly higher abundance (average spectral count of 15) compared to proteins that were not found back in the verification set (average spectral count of 4) (p=3.5*10⁻²⁷).

Proof of Concept of Candidate Protein Biomarkers for Validation by Targeted MS

From the 134 proteins significantly enriched in CRC stool samples, a subset of proteins with a potential epithelial origin (i.e. detected in CRC cell lines), that were significantly enriched in FIT-negative CRC patients compared to controls, and were recovered in the verification set in the advanced adenoma and/or CRC pools (n=29) were selected for further validation. First results for 13 of these proteins confirmed the results obtained by LC-MS/MS in the verification set. These included 6 proteins detected in CRC and/or advanced adenoma pools while not detected in controls or adenomas (e.g. C4B and Glucose-6-phosphate isomerase (GPI) (FIG. 4); and 7 proteins detected in low levels in the controls and adenomas while being more abundantly present in advanced adenomas and CRC, e.g. PRTN3 and A2M (FIG. 4).

Proteins that Complement FIT for Detecting CRC

Novel biomarkers for early detection of CRC should perform significantly better than FIT, or should complement FIT, in order to increase diagnostic accuracy. As expected from their correlation with FIT, hemoglobin α and β were both significantly more abundantly present in CRC compared to control stool samples. The data revealed several proteins of which levels were significantly higher in CRC stool samples compared to control stool samples with a higher discriminative power than hemoglobin. It is of interest to see if these proteins detect the same CRC samples as hemoglobin does, or if they detect other CRC samples. FIG. 3 shows an example of one of these proteins (C4A/C4B) that detects all FIT-negative CRC patients, without detecting control subject, thereby reaching a 100% sensitivity and specificity in this setting. However, not only proteins with a higher discriminative power than hemoglobin have potential to complement FIT.

For this reason, proteins that showed significantly higher levels in the FIT negative CRC stool samples compared to all controls were investigated. Indeed, out of the 134 proteins with significantly higher levels in CRC versus control stool samples, around 90% also showed significantly higher levels in the FIT negative CRC stool samples. This indicates that these candidates have high potential to be of added value to the current detection of hemoglobin.

The present study has delivered in-depth proteomic analysis on human stool samples, providing a list of 134 human proteins that were significantly enriched in stool samples from CRC patients, of which several showed highly significant discriminative power. Thus, discriminative markers for improving current FIT tests have been identified.

Example 2
Generation of Dataset of Biomarkers from CRC Tissue and Patient-Matched Normal Colon Tissue, for Detection in Biofluids

Material and Methods

Patients

A total of four patients that underwent surgical resection at the VU University medical center (Amsterdam, the Netherlands) were included in this study. Collection, storage, and use of tissue and patient data were performed in accordance with the Code for Proper Secondary Use of Human Tissue in the Netherlands (Societies DFOBS. http://www.federa.org/). A pathologist inspected all samples to obtain information on tumor size, tumor and nodal stage, differentiation grade, mucinous differentiation. For an overview of the clinicopathological characteristics, see table 1.

Tissue Handling and Tissue Secretome Collection

The tissue secretome collection was performed as described before (Celis J E. et al. Mol. Cell Proteomics 2004; 3:327-4). In short, following surgical resection, the specimen was immediately transferred to the pathology department, were a pathologist excised a representative part of the tumor and adjacent normal colon mucosa (near an unaffected resection margin). These pieces of tissue were cut into cubes of approximately 1 mm³and rinsed in PBS to remove blood and stool particles. Subsequently the tissue particles were incubated in 100 μl PBS for 1 hour at 37° C. Following this incubation the samples were briefly centrifuged (2000 rpm at 4° C. for 2 minutes) and the supernatant was transferred to a new eppendorf tube. The supernatants were centrifuged at maximum speed (13.200 rpm at 4° C. for 20 minutes) to remove all remaining cells and debris. The soluble fractions further, referred to as the ‘tissue secretomes’, were stored at −80° C. until further use. The tissues were processed by standard formalin fixation and paraffin embedded for histological evaluation (supplementary FIG. 4). Microsatellite instability (MSI) status was determined using the MSI Analysis System version 1.2 (Promega Corporation, Madison, USA) as described before (de with M. et al. Gut 2012; 61:855-64).

Gel Electrophoresis and Sample Preparation for Proteomics Analysis

Protein concentrations were determined using the BCA protein assay (Pierce, Thermo Fisher Scientific, Rockford, USA). Twenty μg of proteins were separated by gel electrophoresis using a pre-cast 1D 4-12% gradient SDS-PAGE gel (Invitrogen, Carlsbad, USA). For gel images see supplementary FIG. 2a-c. The gels were fixed in 50% ethanol containing 3% phosphoric acid and stained with Coomassie R-250. Gels were washed in MilliQ water and stored at 4° C. until processing for in-gel digestion. Each lane was cut in 10 gel bands and further processed into tryptic peptides as described before (Piersma S R, et al. J. Proteome Res. 2010; 9:1913-22). The volume of the desalted peptide fractions was reduced to 50 μl in a vacuum centrifuge.

nanoLC-MS/MS Proteomics Analysis

Peptides were separated by an Ultimate 3000 nanoLC system (Dionex LC-Packings, Amsterdam, The Netherlands) equipped with a 20 cm×75 μm ID fused silica column custom packed with 3 μm 120 Å ReproSil Pur C18 aqua (Dr Maisch GMBH, Ammerbuch-Entringen, Germany). After injection, peptides were trapped at 6 μl/min on a 1 cm×100 μm ID trap column packed with 5 μm 120 Å ReproSil C18aqua at 2% buffer B (buffer A: 0.05% formic acid in MQ; buffer B: 80% acetonitrile+0.05% formic acid in MQ) and separated at 300 nl/min in a 10-40% buffer B gradient in 60 min. Eluting peptides were ionized at 1.7 kV in a Nanomate Triversa Chip-based nanospray source using a Triversa LC coupler (Advion, Ithaca, N.J.). Intact peptide mass spectra and fragmentation spectra were acquired on a LTQ-FT hybrid mass spectrometer (Thermo Fisher, Bremen, Germany). Intact masses were measured at resolution 50.000 in the ICR cell using a target value of 1×10⁶charges. In parallel, following an FT pre-scan, the top 5 peptide signals (charge-states 2+ and higher) were submitted to MS/MS in the linear ion trap (3 amu isolation width, 30 ms activation, 35% normalized activation energy, Q value of 0.25 and a threshold of 5000 counts). Dynamic exclusion was applied with a repeat count of 1 and an exclusion time of 30 seconds.

Database Searching

To identify proteins from the acquired data MS/MS spectra were searched against the human IPI database v3.59 (80128 entries) using Sequest (version 27, rev 12), which is part of the BioWorks 3.3 data analysis package (Thermo Fisher, San Jose, Calif.). Following database searching the DTA and OUT files were imported into Scaffold Scaffold_—2_—06_—01 (Proteome software, Portland, Oreg.). Scaffold was used to organize the gel-band data and to validate peptide identifications using the Peptide Prophet algorithm (Nesvizhskii A I, et al., Anal. Chem. 2003; 75:4646-58). Only identifications with a probability >95% were retained. Subsequently, the Protein Prophet algorithm (Keller A, et al., Anal. Chem. 2002; 74:5383-92) was applied and protein identifications with a probability of >99% with 2 peptides or more in at least one of the samples were retained. Proteins that contained similar peptides and could not be differentiated based on MS/MS analysis alone were grouped.

Data Mining and Statistical Analysis

For each protein identified, the number of assigned spectra was exported to Excel. Differential analysis of samples was performed using a paired Beta-Binominal test as described previously (Pham T V et al., Expert Rev. Mol. Diagn. 2012; 12:343-59). (Szklarczyk D et al., Nucleic Acids Res. 2011; 39:D561-8). This modification resulted in a paired test thereby taking into account the origin of these samples e.g., comparing protein signatures between two tissues derived from the same patient. Additional general protein information was retrieved using Ingenuity Pathway Analysis (Ingenuity® Systems, www.ingenuity.com). Known and predicted protein-protein interactions were investigated using STRING version 9.0 (www.string-db.org) (Szklarczyk D, et al., Nucleic Acids Res. 2011; 39:D561-8). For cluster and gene ontology analyses we used the Cytoscape platform for network analysis (www.cytoscape.org) (Smoot M E et al., Bioinformatics 2011; 27:431-2), using the plug-ins ClusterONE version 0.93 for clustering and BINGO version 2.44 for the analysis of biological processes within our obtained networks based on GO annotations Maere S et al., Bioinformatics 2005; 21:3448-9 and Nepusz T et al., Nat Meth 2012; 9:471-2). Additionally, transmembrane domains and signal peptide sequences were investigated using secretomeP server 2.0 (Bendtsen J D et al., Protein Eng. Des. Sel. 2004; 17:349-56).

Results

Identification of Secreted Proteins in CRC and Normal Tissue Secretomes

Tissue secretomes were collected from four CRCs and matched adjacent normal colon tissue as described before by Celis at al. (Celis J E et al., Mol. Cell Proteomics 2004; 3:327-44) and processed for mass spectrometry. The tumor of patient 4 was found to be MSI and the tumors of patients 1-3 were microsatellite stable (patient details in table 8. A total of 2703 non-redundant proteins were identified in the tumor secretomes of the four CRCs and their matched normal tissue secretomes. On average 1986 proteins were identified per sample, ranging from 1264 to 2292 proteins.

Of the total of 2703 proteins, 2366 were identified in the tumor secretomes as well as in the normal tissue secretomes, 283 only in the tumor secretomes and 54 only in the normal tissue secretomes. The number of identified proteins was significantly higher in the CRC secretomes than in the normal tissue secretomes (2198 and 1775 on average, respectively, p=0.03), while the number of unique identifications in the samples did not differ significantly (supplementary FIGS. 2b and c, P=0.2). In total, 961 out of 2420 (40%) of proteins were identified in all four normal tissue secretomes and 1627 out of 2649 (61%) were detected in all four cancer secretomes.

Proteins Enriched in the Cancer Secretomes

To obtain an overview of the dataset, an unsupervised hierarchical clustering was performed using the normalized spectral count data from all 2703 identified proteins. Two major clusters were identified; one containing the normal tissue sample of patient 3 and the other containing all other samples, this could be explained by the lower amount of proteins identified in this sample compared to the others (N=1264). In the second cluster again two sub clusters were formed; one containing the other three normal tissue samples and the other all cancer samples. Within the cancer cluster the samples of patients 1-3 formed a separate cluster from the sample of patient 4, which can be explained by the different molecular background of this sample i.e. this latter tumor was MSI whereas the tumors of patients 1-3 were microsatellite stable (table 8). Overall the unsupervised cluster analysis indicated that the protein expression pattern was more similar among the four tumor samples than between paired tumor and normal tissues from the same patient. Potential biomarkers should be enriched in the tumor secretomes compared to the normal tissue secretomes. To identify such proteins the complete protein dataset was further analyzed with a statistical test for paired samples, i.e. taking into account the relationship between normal and cancer tissue samples from individual patients, to identify CRC associated proteins (Pham T V et al., Expert Rev. Mol. Diagn. 2012; 12:343-59). From the 2703 proteins, 522 proteins were significantly differentially present (P<0.05), 409 of which were more abundant in tumor secretomes compared to normal tissue secretomes, thus representing the proteins of likely highest value for discrimination between CRC patients and normal controls (see table 6).

Overlap Analysis of Proteins Enriched in the Cancer Secretomes and Stool Proteins

Protein biomarkers that can be detected in blood or stool are of interest since they could be applied in a standard clinical setting alongside the routinely used markers such as CEA and CA19-9 or haemoglobin. Measuring molecules directly in the biological sample that will be used for screening, i.e. stool, can yield biomarkers that are stable and can reliably be detected in stool samples. An overlap analysis with a previously obtained dataset of stool proteins (unpublished data) revealed that 383 of the 2703 identified proteins are possibly detectable in stool. Out of the 409 significantly more abundant proteins in CRC secretomes compared to normal secretomes, 16 proteins of which were also detected in stool (see table 7).

INDUSTRIAL APPLICABILITY

The present invention thus provides a set of biomarkers for screening for colorectal cancer, or susceptibility to colorectal cancer. The biomarkers enable early diagnosis such that early stage surgical removal of a tumor before metastasis is possible.

The foregoing broadly describes the present invention without limitation to particular embodiments. Variations and modifications as will be readily apparent to those skilled in the art are intended to be within the scope of the invention as defined by the following claims.

Claims

1. An assay comprising: (i) adding to a biological sample obtained from an individual at least four protein binding agents, wherein each protein binding agent is capable of binding to a different biomarker selected from the group of:complement component C4B (Chido blood group) 2 (C4A/C4B); glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2); glucose-6-phosphate isomerase (GPI); transketolase (TKT); N-acylaminoacyl-peptide hydrolase (APEH); histone cluster 1, H4c (HIST4H4 (includes others)); Fatty acid-binding protein 5 (psoriasis-associated) (FABP5); hexosaminidase B (beta polypeptide) (HEXB); epithelial cell adhesion molecule (EPCAM); NME1-NME2 walkthrough (NME1-NME2); Superoxide dismutase 2, mitochondrial (SOD2); Tu translation elongation factor, mitochondrial (TUFM); Glutathione synthetase (GSS); annexin A2 (ANXA2); ATP synthase, H+ transporting, mitochondrial F1 complex, beta polypeptide (ATP5B); 10 kDa heat shock protein (chaperonin 10) (HSPE1); glyoxalase I (GL01); histone cluster 2, H2be (HIST2H2BE (includes others)); S100 calcium binding protein A4 (S100A4); S100 calcium binding protein A11 (S100A11); latexin (LXN); dehydrogenase/reductase (SDR family) member 11 (DHRS1 1); N-acetylglucosaminidase, alpha (NAGLU); Translin (TSN); Proteasome (prosome, macropain) subunit alpha type-4 (PSMA4); Proteasome (prosome, macropain) subunit alpha type-6 (PSMA6); ras-related C3 botulinum toxin substrate 1 (rho family, small GTP binding protein Rac1) (RAC1); Adenosylhomocysteinase (AHCY); fucosidase, alpha-L-1, tissue (FUCA1); S100 calcium binding protein P (S100P); Proteasome (prosome, macropain) subunit beta type-2 (PSMB2); X-prolyl aminopeptidase (aminopeptidase P) 1 (XPNPEP1); Keratin 18 (KRT18); Nuclear cap-binding protein subunit 1 80 kDa (NCBP1); mannosidase, alpha, class 2B, member 1 (MAN2B1); S100 calcium binding protein A6 (S100A6); valosin containing protein (VCP); quinolinate phosphoribosyltransferase (QPRT); major histocompatibility complex, class I, B (HLA-B); phosphoglycerate mutase 1 (brain) (PGAM1); ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3); serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10); myeloperoxidase (MPO); creatine kinase, mitochondrial 1 B (CKMT1 A/CKMT1 B); proteinase 3 (PRTN3); elastase, neutrophil expressed (ELANE); MORC family CW-type zinc finger 1 (MORC1); ubiquitin B (UBB); phospholipase A2, group HA (platelets, synovial fluid) (PLA2G2A); carbonic anhydrase IV (CA4); G elongation factor, mitochondrial 2 (GFM2); S100 calcium binding protein A7 (S100A7); Bactericidal permeability-increasing protein (BPI); collagen, type VI, alpha 5 (COL6A5); LIM homeobox 8 (LHX8); cysteine-rich secretory protein 3 (CRISP3); Azurocidin (AZU1); hemicentin 1 (HMCN 1); Transglutaminase 3 (E polypeptide, protein-glutamine gamma-glutamyltransferase) (TGM3); CDC42 binding protein kinase alpha (DMPK-like) (CDC42BPA); Cathepsin G (CTSG); Resistin (RETN); methylmalonyl CoA mutase (MUT); armadillo repeat containing, X-linked 4 (ARMCX4); Integrin alpha-M (complement component 3 receptor 3 subunit) (ITGAM); Calcium channel, voltage dependent, R-type alpha-I E subunit (CACNA1 E); T-cell lymphoma invasion and metastasis 2 (TIAM2); HIR histone cell cycle regulation defective homolog A (S. cerevisiae) (HIRA); dopey family member 2 (DOPEY2); integrin beta 1 binding protein 3 (ITGB1 BP3); Sodium channel, voltage-gated, type VII, alpha (SCN7A); Rab3C, member RAS oncogene family (RAB3C); chromosome 9 open reading frame 79 (C9orf79); nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 4 (NFATC4); UDP-glucose glycoprotein glucosyltransferase 2 (UGGT2); Cornulin (CRNN); kielin/chordin-like protein (KCP); CD1 E molecule (CD1 E); coiled-coil domain-containing 18 (CCDC18); leukotriene A-4 hydrolase (LTA4H); albumin (ALB); alpha-2-macroglobulin (A2M); complement component 3 (C3); hemoglobin, beta (HBB); transferrin (TF); hemoglobin, alpha 1 (HBA1/HBA2); lactotransferrin (LTF); ceruloplasmin (ferroxidase) (CP); catalase (CAT); group-specific component (vitamin D-binding protein) (GC); serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1); fibrinogen gamma chain (FGG); S100 calcium binding protein A8 (S100A8); ferritin, light polypeptide (FTL); actin, beta (ACTB); fibronectin 1 (FN1); defensin, alpha 1 (DEFA1 (includes others)); serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1); retinol binding protein 4, plasma (RBP4); peroxiredoxin 2 (PRDX2); fibrinogen alpha chain (FGA); serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2); carbonic anhydrase II (CA2); orosomucoid 1 (ORM1/ORM2); lactate dehydrogenase A (LDHA); vitronectin (VTN); kininogen-1 (KNG1); actin, alpha, cardiac muscle 1 (ACTC1); leucine-rich alpha-2-glycoprotein 1 (LRG1); gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH); enolase 1, (alpha) (EN01); profilin 1 (PFN1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 7 (SERPINA7); alpha-1-microglobulin/bikunin precursor (AMBP); lamin A/C (LMNA); apolipoprotein D (APOD); thyroid hormone receptor interactor 11 (TRIP1 1); complement component 4 binding protein, alpha (C4BPA); tropomyosin 4 (TPM4); filamin A, alpha (FLNA); haptoglobin (HP); hemopexin (HPX); hemoglobin, delta (HBD); fibrinogen beta chain (FGB); S100 calcium binding protein A9 (S100A9); complement component 5 (C5); solute carrier family 26, member 3 (SLC26A3); complement component 9 (C9); amyloid P component, serum (APCS); alpha-1-B glycoprotein (A1 BG); complement C3-like (LOC10013351 1); inter-alpha (globulin) inhibitor H4 (plasma Kallikrein-sensitive glycoprotein) (ITIH4); complement component C8, alpha polypeptide (C8A); inter-alpha (globulin) inhibitor H1 (ITIH1); acyl-CoA dehydrogenase, very long chain (ACADVL); cDNA F1160317, highly similar to Aminoacylase-1 (ACY1); Ankyrin repeat domain-containing protein 35 (ANKRD35); baculoviral IAP repeat-containing 6 (BIRC6); Bleomycin hydrolase (BLMH); bone marrow stromal cell antigen 12 (BST1); hypothetical protein LOC643677 (C13orf40); Cytidine deaminase (CDA); chitinase 1 (chitotriosidase) (CHIT1); cathepsin C (CTSC); Cathepsin S (CTSS); Isoform 2 of Dedicator of cytokinesis protein 4 (DOCK4); Glutathione reductase (GSR); hect (homologous to the E6-AP (UBE3A) carboxyl terminus) domain and RCC1 (CHCI)-like domain (RLD) 1 (HERC1); hect domain and RLD 2 (HERC2); major histocompatibility complex, class II, DR beta 5 (HLA-DRB5); isocitrate dehydrogenase 1 (NADP+), soluble (IDH1); inter-alpha (globulin) inhibitor H2 (ITIH2); Uncharacterized protein KIAA1797 (KIAA1797); Lysozyme C (LYZ); Nebulin (NEB); NIMA (never in mitosis gene a)-related kinase 10 (NEK10); peptidase D (PEPD); quiescin Q6 sulfhydryl oxidase 1 (QSOX1); ribonuclease T2 (RNASET2); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 (SERPINA1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 (SERPINA3); serpin peptidase inhibitor, clade B (ovalbumin), member 3 (SERPINB3); SET domain containing 2 (SETD2); Shugoshin-like 2 (SGOL2); sialic acid acetylesterase (SIRE); spectrin repeat containing, nuclear envelope 1 (SYNE1); Transaldolase 1 (TALDO1); Taste receptor type 2 member 42 (TAS2R42); triosephosphate isomerase 1 (TPI 1); Vinculin (VCL); Zymogen granule membrane protein 16 (ZG16); hypothetical protein LOC79887 (PLBD1); Isoform 1 of Serine/threonine-protein phosphatase 6 regulatory ankyrin repeat subunit A (ANKRD28); Cystatin-C(CST3); D-dopachrome decarboxylase (DDT); Synapse-associated protein 1 (SYAP1); Proteasome subunit alpha type-2 (PSMA2); SUB1 homolog (S. cerevisiae) (SUB1); Microfibril-associated glycoprotein 3 (MFAP3); Cathepsin D (CTSD); proteasome (prosome, macropain) subunit, beta type, 1 (PSMB1); proteasome (prosome, macropain) subunit, beta type, 5 (PSMB5); cDNA F1161 112, highly similar to BTB/POZ domain-containing protein KCTD15 (KCTD15); prolyl 4-hydroxylase, beta polypeptide (P4HB); glutathione peroxidase 1 (GPX1); serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5); Isoform 1 of collagen, type IV, alpha 3 (Goodpasture antigen) binding protein (COL4A3BP); proteasome (prosome, macropain) subunit, beta type, 6 (PSMB6); Keratin 20 (KRT20); Calpain small subunit 1 (CAPNS1); peroxiredoxin 3 (PRDX3); NACC family member 2, BEN and BTB (POZ) domain containing (NACC2); Rho GDP-dissociation inhibitor 2 (ARHGDIB); Macrophage migration inhibitory factor (MIF); Ran-binding protein 6 (RANBP6); spinster homolog 3 (Drosophila) (SPNS3); minichromosome maintenance complex component 2 (MCM2); Fumarylacetoacetase (FAH); heat shock 70 kDa protein 8 (HSPA8); brain abundant, membrane attached signal protein 1 (BASP1); Branched-chain-amino-acid aminotransferase (BCAT2); Moesin (MSN); serpin peptidase inhibitor, clade B (ovalbumin), member 8 (SERPINB8); glucose-6-phosphate dehydrogenase (G6PD); Isoform 1 of UPF0557 protein C10orf1 19 (C10orf1 19); Prosaposin (PSAP); eukaryotic translation elongation factor 1 gamma (EEF1 G); four and a half LIM domains 1 (FHL1); carboxypeptidase, vitellogenic-like (CPVL); tubulin tyrosine ligase-like family, member 3 (TTLL3); IPI:IPI00942608.1|ENSEMBL:ENSP0000 (unmapped; 26 kDa protein); proline-rich protein BstNI subfamily 2 (PRB1/PRB2); Protocadherin-8 (PCDH8); Alpha-2-macroglobulin-like protein 1 (A2ML1); Guanine deaminase (GDA); Lipocalin-1 (LCN1); Histone H1.4 (HIST1 H1 E); IPI:IPI00937064.1|REFSEQ:XP—002342720 (ZAN); heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1);); Endoplasmic reticulum aminopeptidase 2 (ERAP2); 14-3-3 protein zeta/delta (YWHAZ); G-protein coupled receptor 39 (GPR39); similar to KIAA1783 protein (KIAA1783 protein); apolipoprotein A-1 binding protein (APOA1 BP); pleckstrin and Sec7 domain containing 2 (PSD2); prolylcarboxypeptidase (angiotensinase C) (PROP); Tubulin alpha-1 C chain (TUBA1 C); Calmodulin-like protein 5 (CALML5); ARP3 actin-related protein 3 homolog (yeast) (ACTR3); myosin, light chain 6, alkali, smooth muscle and non-muscle (MYL6); Vasodilator-stimulated phosphoprotein (VASP); ARP2 actin-related protein 2 homolog (yeast) (ACTR2); Rheumatoid factor (RF-IP18); Phosphoglycerate kinase 1 (PGK1); Solute carrier family 35 member F1 (SLC35F1); Solute carrier family 35 member F1 (SLC35F1); alkaline phosphatase, liver/bone/kidney (ALPL); I tropomyosin 3 (TPM3); Hexokinase-3 (HK3); Vimentin (VIM); Annexin A1 (ANXA1); IPI:IPI00930073.1|TREMBL:B2R853 (KRT6C); Keratin, type II cytoskeletal 6C (KRT6C); myosin, heavy chain 13, skeletal muscle (MYH13); cell cycle progression 1 (CCPG1); Hypothetical protein (H-INV); calcium channel, voltage-dependent, L type, alpha 1 D subunit (CACNA1 D); LY6/PLAUR domain containing 5 (LYPD5); aarF domain containing kinase 2 (ADCK2); Myosin-lc (MY01C); amyloid beta precursor protein (cytoplasmic tail) binding protein 2 (APPBP2); integrin, alpha 2b (platelet glycoprotein lib of 11b/111a complex, antigen CD41) (ITGA2B); tubulin, beta 6 (TUBB6); synaptotagmin-like 4 (SYTL4); aquaporin 4 (AQP4); cell division cycle 42 (GTP binding protein, 25 kDa) (CDC42); myosin, light chain 12B, regulatory (MYL12B); protein L-Myc-2-like (LOC 100293553); RAP1 B, member of RAS oncogene family (RAP1 B); glycoprotein IX (platelet) (GP9); Destrin (DSTN); complement component 1, q subcomponent, C chain (C1 QC); epidermal growth factor receptor pathway substrate 8 (EPS8); dual specificity phosphatase 3 (DUSP3); ras homolog gene family, member A (RHOA); myosin, light chain 9, regulatory (MYL9); peptidylprolyl isomerase A (cyclophilin A) (PPIA); Cofilin-1 (CFL1); and/or lactotransferrin (LTF), collagen, type XII, alpha 1 (COL12A1), agrin (AGRN), +MYB binding protein (P160) 1 a (MYBBP1A), transformation/transcription domain-associated protein (TRRAP), annexin A6 (ANXA6), cytoskeleton associated protein 5 (CKAP5), minichromosome maintenance complex component 5 (MCM5), importin 4 (IPO4), neurobeachin-like 2 (NBEAL2), minichromosome maintenance complex component 4 (MCM4), 2′-5′-oligoadenylate synthetase 3, 100 kDa (OAS3), minichromosome maintenance complex component 3 (MCM3), NEDD8 activating enzyme E1 subunit 1 (NAE1), tripartite motif containing 28 (TRIM28), fused in sarcoma (FUS), phenylalanyl-tRNA synthetase, alpha subunit (FARSA), myeloid cell nuclear differentiation antigen (MNDA), suppressor of Ty 16 homolog S. cerevisiae) (SUPT16H), DEAD (Asp-Glu-Ala-Asp) box polypeptide 5 (DDX5), tenascin C (TNC), nuclear import 7 homolog (S. cerevisiae) (NIP7), chromodomain helicase DNA binding protein 4 (CHD4), regulator of chromosome condensation 2 (RCC2), DNA (cytosine-5-)-methyltransferase 1 (DNMT1), exportin 4 (XPO4), chaperonin containing TCP1, subunit 5 (epsilon) (CCT5), serine/arginine-rich splicing factor 9 (SRSF9), spectrin, beta, non-erythrocytic 2 (SPTBN2), TIMP metallopeptidase inhibitor 1 (TIMP1), nidogen 1 (NID1), ribonucleotide reductase M1 (RRM1), eukaryotic translation initiation factor 4 gamma, 1 (EIF4G1), component of oligomeric golgi complex 4 (COG4), polymerase (DNA directed), delta 1, catalytic subunit 125 kDa (POLD1), splicing factor 3b, subunit 2, 145 kDa (SF3B2), exosome component 2 (EXOSC2), minichromosome maintenance complex component 6 (MCM6), plastin 3 (PLS3), aldolase B, fructose-bisphosphate (ALDOB), SMG1homolog, phosphatidylinositol 3-kinase-related kinase (C. elegans) (SMG1), G1 to S phase transition 1 (GSPT1), KH-type splicing regulatory protein (KHSRP), DEAD (Asp-Glu-Ala-Asp) box polypeptide 21 (DDX21), phosphatidylinositol transfer protein, beta (PITPNB), aquarius homolog (mouse) (AQR), heterogeneous nuclear ribonucleoprotein D-like (HNRPDL), annexin A3 (ANXA3), processing of precursor 1, ribonuclease P/MRP subunit (S. cerevisiae) (POP1), structural maintenance of chromosomes 2 (SMC2), dynein, cytoplasmic 1, light intermediate chain 2 (DYNC1 LI2), peptidylprolyl isomerase D (PPID), vacuolar protein sorting 37 homolog B (S. cerevisiae) (VPS37B), adrenergic, beta, receptor kinase 1 (ADRBK1), DIS3 mitotic control homolog (S. cerevisiae) (DIS3), polymerase (RNA) I polypeptide A, 194 kDa (POLR1A), t-complex 1 (TCP1), plakophilin 3 (PKP3), La ribonucleoprotein domain family, member 1 B (LARP1 B), poly (ADP-ribose) polymerase 1 (PARP1), CD46 molecule, complement regulatory protein (CD46), p21 protein (Cdc42/Rac)-activated kinase 2 (PAK2), ATP-binding cassette, sub-family E (OABP), member 1 (ABCE1), ubiquitin specific peptidase 14 (tRNA-guanine transglycosylase) (USP14), chaperonin containing TCP1, subunit 3 (gamma) (CCT3), Ran GTPase activating protein 1 (RANGAP1), deoxythymidylate kinase (thymidylate kinase) (DTYMK), N-myristoyltransferase 1 (NMT1), dynamin 1-like (DNM1 L), interferon induced transmembrane protein 2 (1-8D) (IFITM2), fermitin family member 1 (FERMT1), tubulin folding cofactor D (TBCD), serine/arginine-rich splicing factor 10 (LOC100505793/SRSF10), STE20-like kinase (SLK), mucin 5 AC, oligomeric mucus/gel-forming (MUC5AC/MUC5B), methionyl-tRNA synthetase (MARS), SMEK homolog 1, suppressor of mek1 (Dictyostelium) (SMEK1), high mobility group box 2 (HMGB2), non-POU domain containing, octamer-binding (NONO), transforming growth factor, beta-induced, 68 kDa (TGFBI), fibulin 2 (FBLN2), high density lipoprotein binding protein (HDLBP), collagen, type IV, alpha 2 (COL4A2), copine I (CPNE1), N(alpha)-acetyltransferase 50, NatE catalytic subunit (NAA50), LSM7 homolog, U6 small nuclear RNA associated (S. cerevisiae) (LSM7), structure specific recognition protein 1 (SSRP1), importin 8 (IP08), yippee-like 5 (Drosophila) (YPEL5), phosphoglucomutase 3 (PGM3), ring finger protein 40 (RNF40), structural maintenance of chromosomes 3 (SMC3), regenerating islet-derived family, member 4 (REG4), splicing factor 3a, subunit 3, 60 kDa (SF3A3), thrombospondin 1 (THBS1), chaperonin containing TCP1, subunit 6A (zeta 1) (CCT6A), PRP8 pre-mRNA processing factor 8 homolog (S. cerevisiae) (PRPF8), symplekin (SYMPK), far upstream element (FUSE) binding protein 1 (FUBP1), U2 small nuclear RNA auxiliary factor 1 (U2AF1), huntingtin (HTT), eukaryotic translation initiation factor 5B (EIF5B), nuclear autoantigenic sperm protein (histone-binding) (NASP), heterogeneous nuclear ribonucleoprotein K (HNRNPK), Y box binding protein 1 (YBX1), annexin A11 (ANXA1 1), RecQ protein-like (DNA helicase Q1-like) (RECQL), cortactin (CTTN), tubulin, beta 3 (TUBB3), pyrroline-5-carboxylate reductase-like (PYCRL), periplakin (PPL), phosphoglucomutase 2-like 1 (PGM2L1), chromosome 17 open reading frame 49 (C17orf49), mRNA turnover 4 homolog (S. cerevisiae) (MRT04), methyltransferase like 1 (METTL1), squamous cell carcinoma antigen recognized by T cells 3 (SART3), S100 calcium binding protein A12 (S100A13), aminopeptidase-like 1 (NPEPL1), cyclin-dependent kinase 1 (CDK1), ubiquitin protein ligase E3 component n-recognin 1 (UBR1), Rho GTPase activating protein 18 (ARHGAP18), signal recognition particle 14 kDa (homologous Alu RNA binding protein) (SRP14), cathelicidin antimicrobial peptide (CAMP), splicing factor proline/glutamine-rich (SFPQ), RAS p21 protein activator (GTPase activating protein) 1 (RASA1), Ral GTPase activating protein, beta subunit (non-catalytic) (RALGAPB), laminin, beta 1 (LAMB1), RAB3 GTPase activating protein subunit 2 (non-catalytic) (RAB3GAP2), chaperonin containing TCP1, subunit 8 (theta) (CCT8), heterogeneous nuclear ribonucleoprotein L-like (HNRPLL), RAN binding protein 1 (RANBP1), kinetochore associated 1 (KNTC1), dyskeratosis congenita 1, dyskerin (DKC1), casein kinase 2, alpha 1 polypeptide (CSNK2A1), CAP-GLY domain containing linker protein 1 (CLIP1), chaperonin containing TCP1, subunit 2 (beta) (CCT2), tubulin tyrosine ligase-like family, member 12 (TTLL12), ataxia telangiectasia mutated (ATM), splicing factor 3a, subunit 1, 120 kDa (SF3A1), ribosomal protein S20 (RPS20), ubiquitin-conjugating enzyme E20 (UBE20), translocated promoter region (to activated MET oncogene) (TPR), BRCA2 and CDKN1A interacting protein (BCCIP), gem (nuclear organelle) associated protein 5 (GEMIN5), ribonuclease P/MRP 30 kDa subunit (RPP30), loss of heterozygosity, 12, chromosomal region 1 (LOH 12CR1), syntaxin binding protein 2 (STXBP2), ubiquitin-conjugating enzyme E2H (UBE2H), DIP2 disco-interacting protein 2 homolog B (Drosophila) (DIP2B), RAPT, GTP-GDP dissociation stimulator 1 (RAP1GDS1), heterogeneous nuclear ribonucleoprotein M (HNRNPM), LIM domain 7 (LM07), RNA binding motif protein 25 (RBM25), aldehyde dehydrogenase 7 family, member A1 (ALDH7A1), cleavage and polyadenylation specific factor 1, 160 kDa (CPSF1), calponin 2 (CNN2), chaperonin containing TCP1, subunit 7 (eta) (CCT7), lysyl-tRNA synthetase (KARS), UDP-N-acteylglucosamine pyrophosphorylase 1 (UAP1), heat shock 70 kDa protein 4-like (HSPA4L), 138 kDa protein (138 kDa protein), thimet oligopeptidase 1 (THOP1), glutaredoxin 3 (GLRX3), phosphoglycerate dehydrogenase (PHGDH), CDV3 homolog (mouse) (CDV3), structural maintenance of chromosomes 4 (SMC4), RNA binding motif (RNP1, RRM) protein 3 (RBM3), hepatoma-derived growth factor (HDGF), heterogeneous nuclear ribonucleoprotein U (scaffold attachment factor A) (HNRNPU), nuclear receptor binding protein 1 (NRBP1), polymerase (RNA) I polypeptide B, 128 kDa (POLR1 B), protein phosphatase 5, catalytic subunit (PPP5C), glucose-6-phosphate dehydrogenase (G6PD), arginase, liver (ARG1), 3-hydroxy-3-methylglutaryl-CoA synthase 1 (soluble) (HMGCS1), ubiquitin-like modifier activating enzyme 2 (UBA2), KIAA1033 (KIAA1033), annexin A4 (ANXA4), DEAD (Asp-Glu-Ala-Asp) box polypeptide 17 (DDX17), acidic (leucine-rich) nuclear phosphoprotein 32 family, member E (ANP32E), glucosamine (UDP-N-acetyl)-2-epimerase/N-acetylmannosamine kinase (GNE), KIAA0368 (KIAA0368), vacuolar protein sorting 4 homolog B (S. cerevisiae) (VPS4B), replication protein A1, 70 kDa (RPA1), eukaryotic translation initiation factor 2, subunit 1 alpha, 35 kDa (EIF2S1), eukaryotic translation initiation factor 3, subunit J (EIF3J), suppressor of Ty 6 homolog (S. cerevisiae) (SUPT6H), heat shock 105 kDa/110 kDa protein 1 (HSPH1), exportin 5 (XP05), transcription elongation factor A (S11), 1 (TCEA1), Sjogren syndrome antigen B (autoantigen La) (SSB), AE binding protein 1 (AEBP1), LIM and cysteine-rich domains 1 (LMCD1), interleukin enhancer binding factor 3, 90 kDa (ILF3), WD repeat domain 61 (WDR61), N(alpha)-acetyltransferase 15, NatA auxiliary subunit (NAA15), serine/arginine-rich splicing factor 4 (SRSF4), ring finger protein 20 (RNF20), lactamase, beta 2 (LACTB2), NHP2 ribonucleoprotein homolog (yeast) (NHP2), chromosome 17 open reading frame 28 (C17orf28), CTP synthase II (CTPS2), fascin homolog 1, actin-bundling protein (Strongylocentrotus purpuratus) (FSCN1), tRNA nucleotidyl transferase, CCA-adding, 1 (TRNT1), splicing regulatory glutamine/lysine-rich protein 1 (SREK1), stromal antigen 1 (STAG1), oxysterol binding protein (OSBP), deoxyuridine triphosphatase (DUT), coiled-coil domain containing 25 (CCDC25), DEK oncogene (DEK), coiled-coil domain containing 72 (CCDC72), polymerase (RNA) II (DNA directed) polypeptide E, 25 kDa (POLR2E), phosphoserine phosphatase (PSPH), structural maintenance of chromosomes 1A (SMC1A), DEAD (Asp-Glu-Ala-Asp) box polypeptide 23 (DDX23), tRNA methyltransferase 11-2 homolog (S. cerevisiae) (TRMT112), COP9 constitutive photomorphogenic homolog subunit 2 (Arabidopsis) (COPS2), programmed cell death 5 (PDCD5), cyclin-dependent kinase 2 (CDK2), proteasome (prosome, macropain) 26S subunit, non-ATPase, 3 (PSMD3), RAN binding protein 2 (RANBP2), SERPINE1 mRNA binding protein 1 (SERBP1), O-linked N-acetylglucosamine (GlcNAc) transferase (UDP-N-acetylglucosamine:polypeptide-N-acetylglucosaminyl transferase) (OGT), non-SMC condensin I complex, subunit D2 (NCAPD2), SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily c, member 2 (SMARCC2), NOP10 ribonucleoprotein homolog (yeast) (NOP10), LPS-responsive vesicle trafficking, beach and anchor containing (LRBA), apoptosis inhibitor 5 (API5), signal recognition particle receptor (docking protein) (SRPR), transferrin receptor (p90, CD71) (TFRC), basic leucine zipper and W2 domains 2 (BZW2), ribonuclease, RNase A family, 3 (RNASE3), diazepam binding inhibitor (GABA receptor modulator, acyl-CoA binding protein) (DBI), FK506 binding protein 4, 59 kDa (FKBP4), chromosome 6 open reading frame 130 (C6orf130), cofactor of BRCA1 (COBRA1), flap structure-specific endonuclease 1 (FEN1), glucan (1,4-alpha-), branching enzyme 1 (GBE1), small nuclear ribonucleoprotein polypeptide B (SNRPB2), NSFL1 (p97) cofactor (p47) (NSFL1 C), acyl-CoA thioesterase 7 (ACOT7), NOP2/Sun domain family, member 2 (NSUN2), chaperonin containing TCP1, subunit 4 (delta) (CCT4), kallikrein-related peptidase 6 (KLK6), glutaminyl-peptide cyclotransferase (QPCT), BCL2-associated athanogene 6 (BAG6), eukaryotic translation initiation factor 3, subunit C (EIF3C/EIF3CL), ATPase, H+ transporting, lysosomal 56/58 kDa, V1 subunit B2 (ATP6V1 B2), matrix metallopeptidase 8 (neutrophil collagenase) (MMP8), proteasome (prosome, macropain) 26S subunit, ATPase, 5 (PSMC5), GTP cyclohydrolase I feedback regulator (GCHFR), poly(A) polymerase alpha (PAPOLA), hippocalcin-like 1 (HPCAL1), GTPase activating protein (SH3 domain) binding protein 1 (G3BP1), polymerase (RNA) III (DNA directed) polypeptide A, 155 kDa (POLR3A), superkiller viralicidic activity 2-like 2 (S. cerevisiae) (SKIV2L2), polymerase (RNA) II (DNA directed) polypeptide A, 220 kDa (POLR2A), collagen, type I, alpha 2 (COL1A2), fibrillarin (FBL), glutamyl-prolyl-tRNA synthetase (EPRS), ELAV (embryonic lethal, abnormal vision, Drosophila)-like 1 (Hu antigen R) (ELAVL1), nuclear cap binding protein subunit 2, 20 kDa (NCBP2), GCN1 general control of amino-acid synthesis 1-like 1 (yeast) (GCN1 L1), histone acetyltransferase 1 (HAT1), stromal antigen 2 (STAG2), sorbitol dehydrogenase (SORD), REX2, RNA exonuclease 2 homolog (S. cerevisiae) (REX02), heterogeneous nuclear ribonucleoprotein F (HNRNPF), thymopoietin (TMPO), ubiquitin specific peptidase 24 (USP24), KIAA1967 (KIAA1967), complement component 1, r subcomponent (C1 R), annexin A7 (ANXA7), RuvB-like 2 (E. coli) (RUVBL2), acireductone dioxygenase 1 (ADI 1), eukaryotic translation initiation factor 4A3 (EIF4A3), heterogeneous nuclear ribonucleoprotein U-like 2 (HNRNPUL2), ubiquitin protein ligase E3 component n-recognin 4 (UBR4), SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 2 (SMARCA2), cytochrome b5 reductase 2 (CYB5R2), splicing factor 3b, subunit 3, 130 kDa (SF3B3), G protein pathway suppressor 1 (GPS1), MAD2 mitotic arrest deficient-like 1 (yeast) (MAD2L1), phospholipase C, gamma 1 (PLCG1), eukaryotic translation initiation factor 4H (EIF4H), U2 small nuclear RNA auxiliary factor 2 (U2AF2), tankyrase 1 binding protein 1, 182 kDa (TNKS1 BP1), transglutaminase 2 (C polypeptide, protein-glutamine-gamma-glutamyltransferase) (TGM2), heterogeneous nuclear ribonucleoprotein L (HNRNPL), inositol polyphosphate-5-phosphatase, 145 kDa (INPP5D), annexin A10 (ANXA10), BUD31 homolog (S. cerevisiae) (BUD31), phosphatidylinositol transfer protein, alpha (PITPNA), leucyl-tRNA synthetase (LARS), nicotinamide N-methyltransferase (NNMT), proteasome (prosome, macropain) 26S subunit, non-ATPase, 12 (PSMD12), v-crk sarcoma virus CT10 oncogene homolog (avian) (CRK), proteoglycan 2, bone marrow (natural killer cell activator, eosinophil granule major basic protein) (PRG2), versican (VCAN), exportin, tRNA (nuclear export receptor for tRNAs) (XPOT), EMG1 nucleolar protein homolog (S. cerevisiae) (EMG1), chromosome 11 open reading frame 73 (C11orf73), transports 1 (TNP01), latent transforming growth factor beta binding protein 2 (LTBP2), cold shock domain containing E1, RNA-binding (CSDE1), sulfiredoxin 1 (SRXN1), paraspeckle component 1 (PSPC1), ribosomal protein S3A (RPS3A), ISG15 ubiquitin-like modifier (ISG15), polymerase (RNA) II (DNA directed) polypeptide B, 140 kDa (POLR2B), general transcription factor IN (GTF2I), NHP2 nonhistone chromosome protein 2-like 1 (S. cerevisiae) (NHP2L1), proteasome (prosome, macropain) 26S subunit, non-ATPase, 10 (PSMD10), signal transducer and activator of transcription 1, 91 kDa (STAT1), elongation factor Tu GTP binding domain containing 1 (EFTUD1), mediator complex subunit 23 (MED23), eukaryotic translation initiation factor 2C, 2 (EIF2C2), RNA binding motif protein 4B (RBM4B), KIAA0664 (KIAA0664), core-binding factor, beta subunit (CBFB), poly(A) binding protein, cytoplasmic 1 (PABPC1), nicotinamide phosphoribosyltransferase (NAMPT), cellular retinoic acid binding protein 2 (CRABP2), thyroid hormone receptor interactor 12 (TRIP12), DnaJ (Hsp40) homolog, subfamily C, member 9 (DNAJC9), StAR-related lipid transfer (START) domain containing 10 (STARD10), ring finger protein 213 (RNF213), eukaryotic translation initiation factor 2B, subunit 5 epsilon, 82 kDa (EIF2B5), bolA homolog 2 (E. coli) (BOLA2/BOLA2B), meningioma expressed antigen 5 (hyaluronidase) (MGEA5), Rab geranylgeranyltransferase, alpha subunit (RABGGTA), pyridoxal-dependent decarboxylase domain containing 1 (PDXDC1), exosome component 8 (EXOSC8), phosphoserine aminotransferase 1 (PSAT1), eukaryotic translation initiation factor 6 (EIF6), chromosome 16 open reading frame 13 (C16orf13), signal transducer and activator of transcription 3 (acute-phase response factor) (STAT3), EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1), defensin, alpha 6, Paneth cell-specific (DEFA6), pyrophosphatase (inorganic) 1 (PPA1), glutathione peroxidase 2 (gastrointestinal) (GPX2), unc-13 homolog D (C. elegans) (UNCI 3D), protein tyrosine phosphatase, non-receptor type 6 (PTPN6), myosin XVIIIA (MY018A), fibulin 1 (FBLN1), ribosomal protein L19 (RPL19), diaphanous homolog 1 (Drosophila) (DIAPH1), MMS19 nucleotide excision repair homolog (S. cerevisiae) (MMS19), nudix (nucleoside diphosphate linked moiety X)-type motif 21 (NUDT21), splicing factor 3b, subunit 5, 10 kDa (SF3B5), SAP domain containing ribonucleoprotein (SARNP), ADP-ribosylation factor guanine nucleotide-exchange factor 2 (brefeldin A-inhibited) (ARFGEF2), guanylate binding protein 1, interferon-inducible (GBP1), HECT, UBA and WWE domain containing 1 (HUWE1), processing of precursor 7, ribonuclease P/MRP subunit (S. cerevisiae) (POP7), eukaryotic translation initiation factor 2B, subunit 3 gamma, 58 kDa (EIF2B3), N(alpha)-acetyltransferase 10, NatA catalytic subunit (NAA10), DnaJ (Hsp40) homolog, subfamily B, member 1 (DNAJB1), eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2), elongation factor Tu GTP binding domain containing 2 (EFTUD2), grancalcin, EF-hand calcium binding protein (GCA), neuronal cell adhesion molecule (NRCAM), DEAH (Asp-Glu-Ala-His) box polypeptide 16 (DHX16), small glutamine-rich tetratricopeptide repeat (TPR)-containing, alpha (SGTA), serine/threonine kinase 10 (STK10), smu-1 suppressor of mec-8 and unc-52 homolog (C. elegans) (SMUT), PRP4 pre-mRNA processing factor 4 homolog (yeast) (PRPF4), heterogeneous nuclear ribonucleoprotein D (AU-rich element RNA binding protein 1, 37 kDa) (HNRNPD), CTP synthase (CTPS), cleavage stimulation factor, 3′ pre-RNA, subunit 3, 77 kDa (CSTF3), cleavage stimulation factor, 3′ pre-RNA, subunit 1, 50 kDa (CSTF1), heterogeneous nuclear ribonucleoprotein U-like 1 (HNRNPUL1), ribosomal protein S5 (RPS5), protein tyrosine phosphatase, nonreceptor type 1 1 (PTPN1 1), ladinin 1 (LAD1), component of oligomeric golgi complex 2 (COG2), cullin 2 (CUL2), ribosomal protein S17 (RPS17/RPS17L), proteasome (prosome, macropain) 26S subunit, non-ATPase, 4 (PSMD4), annexin A1 (ANXA1), annexin A2 (ANXA2), 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), azurocidin 1 (AZUL), baculoviral IAP repeat containing 6 (BIRC6), chitinase 3-like 1 (cartilage glycoprotein-39) (CHI3L1), carbamoyl-phosphate synthase 1, mitochondrial (CPS1), cathepsin G (CTSG), defensin, alpha 1 (DEFA1 (includes others)), deleted in malignant brain tumors 1 (DMBT1), elastase, neutrophil expressed (ELANE), integrin, alpha M (complement component 3 receptor 3 subunit) (ITGAM), lipocalin 2 (LCN2), lectin, galactoside-binding, soluble, 3 binding protein (LGALS3BP), minichromosome maintenance complex component 2 (MCM2), matrix metallopeptidase 9 (gelatinase B, 92 kDa gelatinase, 92 kDa type IV collagenase) (MMP9), myeloperoxidase (MPO), mucin 5AC, oligomeric mucus/gel-forming (MUC5AC/MUC5B), nuclear cap binding protein subunit 1, 80 kDa (NCBP1), neurofibromin 1 (NF1), olfactomedin 4 (OLFM4), PDS5, regulator of cohesion maintenance, homolog A (S. cerevisiae) (PDS5A), peptidoglycan recognition protein 1 (PGLYRP1), proteinase 3 (PRTN3), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), regenerating islet-derived 1 alpha (REG1A), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5), unc-45 homolog A (C. elegans) (UNC45A);(ii) measuring the level of binding of the at least four protein binding agents to at least four different biomarkers selected from the biomarkers of step (i); and(iii) identifying whether binding of the at least four protein binding agents in step (ii) is detected.
2-57. (canceled)
58. The assay of claim 1, wherein the individual is being tested to determine if that individual is at risk for colorectal cancer.
59. The assay of claim 1, wherein the one or more biomarkers are selected from the group of: complement component C4B (Chido blood group) 2 (C4A/C4B); glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2); glucose-6-phosphate isomerase (GPI); transketolase (TKT); N-acylaminoacyl-peptide hydrolase (APEH); histone cluster 1, H4c (HIST4H4 (includes others)); Fatty acid-binding protein 5 (psoriasis-associated) (FABP5); hexosaminidase B (beta polypeptide) (HEXB); epithelial cell adhesion molecule (EPCAM); Nucleoside diphosphate kinase (NME1-NME2); Superoxide dismutase 2, mitochondrial (SOD2); Tu translation elongation factor, mitochondrial (TUFM); Glutathione synthetase (GSS); annexin A2 (ANXA2); ATP synthase, H+ transporting, mitochondrial F1 complex, beta polypeptide (ATP5B); 10 kDa heat shock protein (chaperonin 10) (HSPE1); glyoxalase I (GL01); histone cluster 2, H2be (HIST2H2BE (includes others)); S100 calcium binding protein A4 (S100A4); S100 calcium binding protein A11 (S100A1 1); latexin (LXN); dehydrogenase/reductase (SDR family) member 11 (DHRS1 1); N-acetylglucosaminidase, alpha (NAGLU); Translin (TSN); Proteasome (prosome, macropain) subunit alpha type-4 (PSMA4); Proteasome (prosome, macropain) subunit alpha type-6 (PSMA6); ras-related C3 botulinum toxin substrate 1 (rho family, small GTP binding protein Rac1) (RAC1); Adenosylhomocysteinase (AHCY); fucosidase, alpha-L-1, tissue (FUCA1); S100 calcium binding protein P (S100P); Proteasome (prosome, macropain) subunit beta type-2 (PSMB2); X-prolyl aminopeptidase (aminopeptidase P) 1 (XPNPEP1); Keratin 18 (KRT18); Nuclear cap-binding protein subunit 1 80 kDa (NCBP1); mannosidase, alpha, class 2B, member 1 (MAN2B1); S100 calcium binding protein A6 (S100A6); valosin containing protein (VCP); quinolinate phosphoribosyltransferase (QPRT); major histocompatibility complex, class I, B (HLA-B); phosphoglycerate mutase 1 (brain) (PGAM1); ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3); serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10); myeloperoxidase (MPO); creatine kinase, mitochondrial 1 B (CKMT1 A/CKMT1 B); proteinase 3 (PRTN3); elastase, neutrophil expressed (ELANE); MORC family CW-type zinc finger 1 (MORC1); ubiquitin B (UBB); phospholipase A2, group HA (platelets, synovial fluid) (PLA2G2A); carbonic anhydrase IV (CA4); G elongation factor, mitochondrial 2 (GFM2); S100 calcium binding protein A7 (S100A7); Bactericidal permeability-increasing protein (BPI); collagen, type VI, alpha 5 (COL6A5); LIM homeobox 8 (LHX8); cysteine-rich secretory protein 3 (CRISP3); Azurocidin (AZUL); hemicentin 1 (HMCN1); Transglutaminase 3 (E polypeptide, protein-glutamine gamma-glutamyltransferase) (TGM3); CDC42 binding protein kinase alpha (DMPK-like) (CDC42BPA); Cathepsin G (CTSG); Resistin (RETN); methylmalonyl CoA mutase (MUT); armadillo repeat containing, X-linked 4 (ARMCX4); Integrin alpha-M (ITGAM); Calcium channel, voltage dependent, R-type alpha-I E subunit (CACNA1 E); T-cell lymphoma invasion and metastasis 2 (TIAM2); HIR histone cell cycle regulation defective homolog A (S. cerevisiae) (HIRA); dopey family member 2 (DOPEY2); integrin beta 1 binding protein 3 (ITGB1 BP3); Sodium channel, voltage-gated, type VII, alpha (SCN7A); Rab3C, member RAS oncogene family (RAB3C); chromosome 9 open reading frame 79 (C9orf79); nuclear factor of activated T-cells, calcineurin-dependent 4 (NFATC4); UDP-glucose glycoprotein glucosyltransferase 2 (UGGT2); Cornulin (CRNN); kielin/chordin-like protein (KCP); CD1 E molecule (CD1 E); coiled-coil domain-containing 18 (CCDC18); leukotriene A-4 hydrolase (LTA4H); albumin (ALB); alpha-2-macroglobulin (A2M); complement component 3 (C3); hemoglobin, beta (HBB); transferrin (TF); hemoglobin, alpha 1 (HBA1/HBA2); lactotransferrin (LTF); ceruloplasmin (ferroxidase) (CP); catalase (CAT); group-specific component (vitamin D-binding protein) (GC); serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1); fibrinogen gamma chain (FGG); S100 calcium binding protein A8 (S100A8); ferritin, light polypeptide (FTL); actin, beta (ACTB); fibronectin 1 (FN1); defensin, alpha 1 (DEFA1 (includes others)); serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1); retinol binding protein 4, plasma (RBP4); peroxiredoxin 2 (PRDX2); fibrinogen alpha chain (FGA); serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2); carbonic anhydrase II (CA2); orosomucoid 1 (ORM1/ORM2); lactate dehydrogenase A (LDHA); vitronectin (VTN); kininogen-1 (KNG1); actin, alpha, cardiac muscle 1 (ACTC1); leucine-rich alpha-2-glycoprotein 1 (LRG1); gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH); enolase 1, (alpha) (EN01); profilin 1 (PFN 1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 7 (SERPINA7); alpha-1-microglobulin/bikunin precursor (AMBP); lamin A/C (LMNA); apolipoprotein D (APOD); thyroid hormone receptor interactor 1 1 (TRIP11); complement component 4 binding protein, alpha (C4BPA); tropomyosin 4 (TPM4); filamin A, alpha (FLNA); haptoglobin (HP); hemopexin (HPX); hemoglobin, delta (HBD); fibrinogen beta chain (FGB); S100 calcium binding protein A9 (S100A9); complement component 5 (C5); solute carrier family 26, member 3 (SLC26A3); complement component 9 (C9); amyloid P component, serum (APCS); alpha-1-B glycoprotein (A1 BG); complement C3-like (LOC10013351 1); inter-alpha (globulin) inhibitor H4 (plasma Kallikrein-sensitive glycoprotein) (ITIH4); complement component C8, alpha polypeptide (C8A); inter-alpha (globulin) inhibitor H1 (ITIH1).
60. The assay of claim 1, wherein the one or more biomarkers are selected from the group consisting of: S100 calcium binding protein A8 (S100A8), complement component C4B (Chido blood group) 2 (C4A/C4B), transferrin (TF), alpha-2-macroglobulin (A2M), S100 calcium binding protein A9 (S100A9), proteinase 3 (PRTN3), Azurocidin (AZU1), lactotransferrin (LTF), hemopexin (HPX) and defensin, alpha 1 (DEFA1).
61. The assay of claim 1, wherein the one or more biomarkers are selected from the group of: C4A/C4B, GOT2, FABP5, EPCAM, NME1-NME2, SOD2, GSS, ANXA2, HSPE1, GL01, S100A4, S100A11, TSN, PSMA4, PSMA6, RAC1, AHCY, S100P, PSMB2, XPNPEP1, KRT18, NCBP1, VCP, QPRT, HLA-B, PGAM1, BPI, LHX8, CRISP3, AZU1, TGM3, CTSG, RETN, ITGAM, CACNA1E, HIRA, ITGB1BP3, SCN7A, RAB3C, C9orf79, NFATC4, UGGT2, CRNN, KCP, CD1E, CCDC18, LTA4H, GC, FGG, SERPING1, LDHA, VTN, KNG1, PFN1, SERPINA7, AMBP, LMNA, C4BPA, TPM4, FLNA, HPX, HBD, FGB, A1BG, LOC100133511, ITIH4, C8A, ITIH1, PLBD1, ANKRD28, CST3, DDT, SYAP1, PSMA2, SUB1, MFAP3, CTSD, PSMB1, PSMB5, KCTD15, P4HB, GPX1, SERPINB5, COL4A3BP, PSMB6, KRT20, CAPNS1, PRDX3, NACC2, ARHGDIB, MIF, RANBP6, SPNS3, MCM2, FAH, HSPA8, BASP1, BCAT2, MSN, SERPINB8, G6PD, C10orf119, PSAP, EEF1G, FHL1, CPVL, TTLL3, 26kDaprotein, PRB1/PRB2, PCDH8, A2ML1, GDA, LCN1, HIST1H1E, ZAN, HNRNPA2B1, ERAP2, YWHAZ, GPR39, KIAA1783, APOA1BP, PSD2, PRCP, TUBA1C, CALML5, ACTR3, MYL6, VASP, ACTR2, RF-IP18, PGK1, SLC35F1, ALPL, TPM3, HK3, VIM, ANXA1, KRT6C, KRT6C, MYH13, CCPG1, H-INV, CACNA1D, LYPD5, ADCK2, MY01C, APPBP2, ITGA2B, TUBB6, SYTL4, AQP4, CDC42, MYL12B, LOC100293553, RAP1B, GP9, DSTN, C1QC, EPS8, DUSP3, RHOA, MYL9, PPIA, CFL1, annexin A2 (ANXA2), azurocidin 1 (AZU1), cathepsin G (CTSG), defensin, alpha 1 (DEFA1 (includes others)), elastase, neutrophil expressed (ELANE), integrin, alpha M (complement component 3 receptor 3 subunit) (ITGAM), myeloperoxidase (MPO), nuclear cap binding protein subunit 1, 80 kDa (NCBP1), proteinase 3 (PRTN3), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), annexin A1 (ANXA1), baculoviral IAP repeat containing 6 (BIRC6), minichromosome maintenance complex component 2 (MCM2), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5)
62. The assay of claim 1, wherein the biological sample comprises a stool sample or a blood sample.
63. The assay of claim 1, wherein the binding agent is an antibody or fragment thereof.
64. The assay of claim 63, wherein the antibody or fragment thereof is selected from the group consisting of: a recombinant antibody or fragment thereof; scFv; Fab; a binding domain of an immunoglobulin molecule.
65. The assay of claim 1, wherein the plurality of binding agents are arranged as an array.
66. The assay of claim 65, wherein the array is a bead-based array or surface-based array.
67. The assay of claim 1, wherein the control sample comprises a biological sample from an individual known to be free from colorectal cancer and/or advanced adenomas.
68. An array comprising at least four binding agents, each protein binding agent capable of binding to at least one biomarker selected from the group of: complement component C4B (Chido blood group) 2 (C4A/C4B); glutamic-oxaloacetic transaminase 2, mitochondrial (aspartate aminotransferase 2) (GOT2); glucose-6-phosphate isomerase (GPI); transketolase (TKT); N-acylaminoacyl-peptide hydrolase (APEH); histone cluster 1, H4c (HIST4H4 (includes others)); Fatty acid-binding protein 5 (psoriasis-associated) (FABP5); hexosaminidase B (beta polypeptide) (HEXB); epithelial cell adhesion molecule (EPCAM); NME1-NME2 walkthrough (NME1-NME2); Superoxide dismutase 2, mitochondrial (SOD2); Tu translation elongation factor, mitochondrial (TUFM); Glutathione synthetase (GSS); annexin A2 (ANXA2); ATP synthase, H+ transporting, mitochondrial F1 complex, beta polypeptide (ATP5B); 10 kDa heat shock protein (chaperonin 10) (HSPE1); glyoxalase I (GL01); histone cluster 2, H2be (HIST2H2BE (includes others)); S100 calcium binding protein A4 (S100A4); S100 calcium binding protein A11 (S100A11); latexin (LXN); dehydrogenase/reductase (SDR family) member 11 (DHRS1 1); N-acetylglucosaminidase, alpha (NAGLU); Translin (TSN); Proteasome (prosome, macropain) subunit alpha type-4 (PSMA4); Proteasome (prosome, macropain) subunit alpha type-6 (PSMA6); ras-related C3 botulinum toxin substrate 1 (rho family, small GTP binding protein Rac1) (RAC1); Adenosylhomocysteinase (AHCY); fucosidase, alpha-L-1, tissue (FUCA1); S100 calcium binding protein P (S100P); Proteasome (prosome, macropain) subunit beta type-2 (PSMB2); X-prolyl aminopeptidase (aminopeptidase P) 1 (XPNPEP1); Keratin 18 (KRT18); Nuclear cap-binding protein subunit 1 80 kDa (NCBP1); mannosidase, alpha, class 2B, member 1 (MAN2B1); S100 calcium binding protein A6 (S100A6); valosin containing protein (VCP); quinolinate phosphoribosyltransferase (QPRT); major histocompatibility complex, class I, B (HLA-B); phosphoglycerate mutase 1 (brain) (PGAM1); ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3); serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10); myeloperoxidase (MPO); creatine kinase, mitochondrial 1 B (CKMT1 A/CKMT1 B); proteinase 3 (PRTN3); elastase, neutrophil expressed (ELANE); MORC family CW-type zinc finger 1 (MORC1); ubiquitin B (UBB); phospholipase A2, group HA (platelets, synovial fluid) (PLA2G2A); carbonic anhydrase IV (CA4); G elongation factor, mitochondrial 2 (GFM2); S100 calcium binding protein A7 (S100A7); Bactericidal permeability-increasing protein (BPI); collagen, type VI, alpha 5 (COL6A5); LIM homeobox 8 (LHX8); cysteine-rich secretory protein 3 (CRISP3); Azurocidin (AZUL); hemicentin 1 (HMCN 1); Transglutaminase 3 (E polypeptide, protein-glutamine gamma-glutamyltransferase) (TGM3); CDC42 binding protein kinase alpha (DMPK-like) (CDC42BPA); Cathepsin G (CTSG); Resistin (RETN); methylmalonyl CoA mutase (MUT); armadillo repeat containing, X-linked 4 (ARMCX4); Integrin alpha-M (complement component 3 receptor 3 subunit) (ITGAM); Calcium channel, voltage dependent, R-type alpha-I E subunit (CACNA1 E); T-cell lymphoma invasion and metastasis 2 (TIAM2); HIR histone cell cycle regulation defective homolog A (S. cerevisiae) (HIRA); dopey family member 2 (DOPEY2); integrin beta 1 binding protein 3 (ITGB1 BP3); Sodium channel, voltage-gated, type VII, alpha (SCN7A); Rab3C, member RAS oncogene family (RAB3C); chromosome 9 open reading frame 79 (C9orf79); nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 4 (NFATC4); UDP-glucose glycoprotein glucosyltransferase 2 (UGGT2); Cornulin (CRNN); kielin/chordin-like protein (KCP); CD1 E molecule (CD1 E); coiled-coil domain-containing 18 (CCDC18); leukotriene A-4 hydrolase (LTA4H); albumin (ALB); alpha-2-macroglobulin (A2M); complement component 3 (C3); hemoglobin, beta (HBB); transferrin (TF); hemoglobin, alpha 1 (HBA1/HBA2); lactotransferrin (LTF); ceruloplasmin (ferroxidase) (CP); catalase (CAT); group-specific component (vitamin D-binding protein) (GC); serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1); fibrinogen gamma chain (FGG); S100 calcium binding protein A8 (S100A8); ferritin, light polypeptide (FTL); actin, beta (ACTB); fibronectin 1 (FN1); defensin, alpha 1 (DEFA1 (includes others)); serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 (SERPING1); retinol binding protein 4, plasma (RBP4); peroxiredoxin 2 (PRDX2); fibrinogen alpha chain (FGA); serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epithelium derived factor), member 2 (SERPINF2); carbonic anhydrase II (CA2); orosomucoid 1 (ORM1/ORM2); lactate dehydrogenase A (LDHA); vitronectin (VTN); kininogen-1 (KNG1); actin, alpha, cardiac muscle 1 (ACTC1); leucine-rich alpha-2-glycoprotein 1 (LRG1); gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) (GGH); enolase 1, (alpha) (EN01); profilin 1 (PFN1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 7 (SERPINA7); alpha-1-microglobulin/bikunin precursor (AMBP); lamin A/C (LMNA); apolipoprotein D (APOD); thyroid hormone receptor interactor 11 (TRIP1 1); complement component 4 binding protein, alpha (C4BPA); tropomyosin 4 (TPM4); filamin A, alpha (FLNA); haptoglobin (HP); hemopexin (HPX); hemoglobin, delta (HBD); fibrinogen beta chain (FGB); S100 calcium binding protein A9 (S100A9); complement component 5 (C5); solute carrier family 26, member 3 (SLC26A3); complement component 9 (C9); amyloid P component, serum (APCS); alpha-1-B glycoprotein (A1 BG); complement C3-like (LOC10013351 1); inter-alpha (globulin) inhibitor H4 (plasma Kallikrein-sensitive glycoprotein) (ITIH4); complement component C8, alpha polypeptide (C8A); inter-alpha (globulin) inhibitor H1 (ITIH1); acyl-CoA dehydrogenase, very long chain (ACADVL); cDNA F1160317, highly similar to Aminoacylase-1 (ACY1); Ankyrin repeat domain-containing protein 35 (ANKRD35); baculoviral IAP repeat-containing 6 (BIRC6); Bleomycin hydrolase (BLMH); bone marrow stromal cell antigen 12 (BST1); hypothetical protein LOC643677 (C13orf40); Cytidine deaminase (CDA); chitinase 1 (chitotriosidase) (CHIT1); cathepsin C (CTSC); Cathepsin S (CTSS); Isoform 2 of Dedicator of cytokinesis protein 4 (DOCK4); Glutathione reductase (GSR); hect (homologous to the E6-AP (UBE3A) carboxyl terminus) domain and RCC1 (CHCI)-like domain (RLD) 1 (HERC1); hect domain and RLD 2 (HERC2); major histocompatibility complex, class II, DR beta 5 (HLA-DRB5); isocitrate dehydrogenase 1 (NADP+), soluble (IDH1); inter-alpha (globulin) inhibitor H2 (ITIH2); Uncharacterized protein KIAA1797 (KIAA1797); Lysozyme C (LYZ); Nebulin (NEB); NIMA (never in mitosis gene a)-related kinase 10 (NEK10); peptidase D (PEPD); quiescin Q6 sulfhydryl oxidase 1 (QSOX1); ribonuclease T2 (RNASET2); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 (SERPINA1); serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 (SERPINA3); serpin peptidase inhibitor, clade B (ovalbumin), member 3 (SERPINB3); SET domain containing 2 (SETD2); Shugoshin-like 2 (SGOL2); sialic acid acetylesterase (SIAE); spectrin repeat containing, nuclear envelope 1 (SYNE1); Transaldolase 1 (TALDO1); Taste receptor type 2 member 42 (TAS2R42); triosephosphate isomerase 1 (TPI 1); Vinculin (VCL); Zymogen granule membrane protein 16 (ZG16); hypothetical protein LOC79887 (PLBD1); Isoform 1 of Serine/threonine-protein phosphatase 6 regulatory ankyrin repeat subunit A (ANKRD28); Cystatin-C(CST3); D-dopachrome decarboxylase (DDT); Synapse-associated protein 1 (SYAP1); Proteasome subunit alpha type-2 (PSMA2); SUB1 homolog (S. cerevisiae) (SUB1); Microfibril-associated glycoprotein 3 (MFAP3); Cathepsin D (CTSD); proteasome (prosome, macropain) subunit, beta type, 1 (PSMB1); proteasome (prosome, macropain) subunit, beta type, 5 (PSMB5); cDNA F1161 112, highly similar to BTB/POZ domain-containing protein KCTD15 (KCTD15); prolyl 4-hydroxylase, beta polypeptide (P4HB); glutathione peroxidase 1 (GPX1); serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5); Isoform 1 of collagen, type IV, alpha 3 (Goodpasture antigen) binding protein (COL4A3BP); proteasome (prosome, macropain) subunit, beta type, 6 (PSMB6); Keratin 20 (KRT20); Calpain small subunit 1 (CAPNS1); peroxiredoxin 3 (PRDX3); NACC family member 2, BEN and BTB (POZ) domain containing (NACC2); Rho GDP-dissociation inhibitor 2 (ARHGDIB); Macrophage migration inhibitory factor (MIF); Ran-binding protein 6 (RANBP6); spinster homolog 3 (Drosophila) (SPNS3); minichromosome maintenance complex component 2 (MCM2); Fumarylacetoacetase (FAH); heat shock 70 kDa protein 8 (HSPA8); brain abundant, membrane attached signal protein 1 (BASP1); Branched-chain-amino-acid aminotransferase (BCAT2); Moesin (MSN); serpin peptidase inhibitor, clade B (ovalbumin), member 8 (SERPINB8); glucose-6-phosphate dehydrogenase (G6PD); Isoform 1 of UPF0557 protein C10orf1 19 (C10orf119); Prosaposin (PSAP); eukaryotic translation elongation factor 1 gamma (EEF1 G); four and a half LIM domains 1 (FHL1); carboxypeptidase, vitellogenic-like (CPVL); tubulin tyrosine ligase-like family, member 3 (TTLL3); IPI:IPI00942608.1|ENSEMBL:ENSP0000 (unmapped; 26 kDa protein); proline-rich protein BstNI subfamily 2 (PRB1/PRB2); Protocadherin-8 (PCDH8); Alpha-2-macroglobulin-like protein 1 (A2ML1); Guanine deaminase (GDA); Lipocalin-1 (LCN1); Histone H1.4 (HIST1 H1 E); IPI:IPI00937064.1|REFSEQ:XP—002342720 (ZAN); heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1);); Endoplasmic reticulum aminopeptidase 2 (ERAP2); 14-3-3 protein zeta/delta (YWHAZ); G-protein coupled receptor 39 (GPR39); similar to KIAA1783 protein (KIAA1783 protein); apolipoprotein A-1 binding protein (APOA1 BP); pleckstrin and Sec7 domain containing 2 (PSD2); prolylcarboxypeptidase (angiotensinase C) (PRCP); Tubulin alpha-1 C chain (TUBA1 C); Calmodulin-like protein 5 (CALML5); ARP3 actin-related protein 3 homolog (yeast) (ACTR3); myosin, light chain 6, alkali, smooth muscle and non-muscle (MYL6); Vasodilator-stimulated phosphoprotein (VASP); ARP2 actin-related protein 2 homolog (yeast) (ACTR2); Rheumatoid factor (RF-IP18); Phosphoglycerate kinase 1 (PGK1); Solute carrier family 35 member F1 (SLC35F1); Solute carrier family 35 member F1 (SLC35F1); alkaline phosphatase, liver/bone/kidney (ALPL); I tropomyosin 3 (TPM3); Hexokinase-3 (HK3); Vimentin (VIM); Annexin A1 (ANXA1); IPI:IPI00930073.1|TREMBL:B2R853 (KRT6C); Keratin, type II cytoskeletal 6C (KRT6C); myosin, heavy chain 13, skeletal muscle (MYH13); cell cycle progression 1 (CCPG1); Hypothetical protein (H-INV); calcium channel, voltage-dependent, L type, alpha 1 D subunit (CACNA1 D); LY6/PLAUR domain containing 5 (LYPD5); aarF domain containing kinase 2 (ADCK2); Myosin-lc (MY01C); amyloid beta precursor protein (cytoplasmic tail) binding protein 2 (APPBP2); integrin, alpha 2b (platelet glycoprotein lib of 11b/111a complex, antigen CD41) (ITGA2B); tubulin, beta 6 (TUBB6); synaptotagmin-like 4 (SYTL4); aquaporin 4 (AQP4); cell division cycle 42 (GTP binding protein, 25 kDa) (CDC42); myosin, light chain 12B, regulatory (MYL12B); protein L-Myc-2-like (LOC 100293553); RAP1 B, member of RAS oncogene family (RAP1 B); glycoprotein IX (platelet) (GP9); Destrin (DSTN); complement component 1, q subcomponent, C chain (C1 QC); epidermal growth factor receptor pathway substrate 8 (EPS8); dual specificity phosphatase 3 (DUSP3); ras homolog gene family, member A (RHOA); myosin, light chain 9, regulatory (MYL9); peptidylprolyl isomerase A (cyclophilin A) (PPIA); Cofilin-1 (CFL1); and/or lactotransferrin (LTF), collagen, type XII, alpha 1 (COL12A1), agrin (AGRN), +MYB binding protein (P160) 1 a (MYBBP1A), transformation/transcription domain-associated protein (TRRAP), annexin A6 (ANXA6), cytoskeleton associated protein 5 (CKAP5), minichromosome maintenance complex component 5 (MCM5), importin 4 (IPO4), neurobeachin-like 2 (NBEAL2), minichromosome maintenance complex component 4 (MCM4), 2′-5′-oligoadenylate synthetase 3, 100 kDa (OAS3), minichromosome maintenance complex component 3 (MCM3), NEDD8 activating enzyme E1 subunit 1 (NAE1), tripartite motif containing 28 (TRIM28), fused in sarcoma (FUS), phenylalanyl-tRNA synthetase, alpha subunit (FARSA), myeloid cell nuclear differentiation antigen (MNDA), suppressor of Ty 16 homolog (S. cerevisiae) (SUPT16H), DEAD (Asp-Glu-Ala-Asp) box polypeptide 5 (DDX5), tenascin C (TNC), nuclear import 7 homolog (S. cerevisiae) (NIP7), chromodomain helicase DNA binding protein 4 (CHD4), regulator of chromosome condensation 2 (RCC2), DNA (cytosine-5-)-methyltransferase 1 (DNMT1), exportin 4 (XPO4), chaperonin containing TCP1, subunit 5 (epsilon) (CCT5), serine/arginine-rich splicing factor 9 (SRSF9), spectrin, beta, non-erythrocytic 2 (SPTBN2), TIMP metallopeptidase inhibitor 1 (TIMP1), nidogen 1 (NID1), ribonucleotide reductase M1 (RRM1), eukaryotic translation initiation factor 4 gamma, 1 (EIF4G1), component of oligomeric golgi complex 4 (COG4), polymerase (DNA directed), delta 1, catalytic subunit 125 kDa (POLD1), splicing factor 3b, subunit 2, 145 kDa (SF3B2), exosome component 2 (EXOSC2), minichromosome maintenance complex component 6 (MCM6), plastin 3 (PLS3), aldolase B, fructose-bisphosphate (ALDOB), SMG1homolog, phosphatidylinositol 3-kinase-related kinase (C. elegans) (SMG1), G1 to S phase transition 1 (GSPT1), KH-type splicing regulatory protein (KHSRP), DEAD (Asp-Glu-Ala-Asp) box polypeptide 21 (DDX21), phosphatidylinositol transfer protein, beta (PITPNB), aquarius homolog (mouse) (AQR), heterogeneous nuclear ribonucleoprotein D-like (HNRPDL), annexin A3 (ANXA3), processing of precursor 1, ribonuclease P/MRP subunit (S. cerevisiae) (POP1), structural maintenance of chromosomes 2 (SMC2), dynein, cytoplasmic 1, light intermediate chain 2 (DYNC1 LI2), peptidylprolyl isomerase D (PPID), vacuolar protein sorting 37 homolog B (S. cerevisiae) (VPS37B), adrenergic, beta, receptor kinase 1 (ADRBK1), DIS3 mitotic control homolog (S. cerevisiae) (DIS3), polymerase (RNA) I polypeptide A, 194 kDa (POLR1A), t-complex 1 (TCP1), plakophilin 3 (PKP3), La ribonucleoprotein domain family, member 1 B (LARP1 B), poly (ADP-ribose) polymerase 1 (PARP1), CD46 molecule, complement regulatory protein (CD46), p21 protein (Cdc42/Rac)-activated kinase 2 (PAK2), ATP-binding cassette, sub-family E (OABP), member 1 (ABCE1), ubiquitin specific peptidase 14 (tRNA-guanine transglycosylase) (USP14), chaperonin containing TCP1, subunit 3 (gamma) (CCT3), Ran GTPase activating protein 1 (RANGAP1), deoxythymidylate kinase (thymidylate kinase) (DTYMK), N-myristoyltransferase 1 (NMT1), dynamin 1-like (DNM1 L), interferon induced transmembrane protein 2 (1-8D) (IFITM2), fermitin family member 1 (FERMT1), tubulin folding cofactor D (TBCD), serine/arginine-rich splicing factor 10 (LOC100505793/SRSF10), STE20-like kinase (SLK), mucin 5 AC, oligomeric mucus/gel-forming (MUC5AC/MUC5B), methionyl-tRNA synthetase (MARS), SMEK homolog 1, suppressor of mek1 (Dictyostelium) (SMEK1), high mobility group box 2 (HMGB2), non-POU domain containing, octamer-binding (NONO), transforming growth factor, beta-induced, 68 kDa (TGFBI), fibulin 2 (FBLN2), high density lipoprotein binding protein (HDLBP), collagen, type IV, alpha 2 (COL4A2), copine I (CPNE1), N(alpha)-acetyltransferase 50, NatE catalytic subunit (NAA50), LSM7 homolog, U6 small nuclear RNA associated (S. cerevisiae) (LSM7), structure specific recognition protein 1 (SSRP1), importin 8 (IP08), yippee-like 5 (Drosophila) (YPEL5), phosphoglucomutase 3 (PGM3), ring finger protein 40 (RNF40), structural maintenance of chromosomes 3 (SMC3), regenerating islet-derived family, member 4 (REG4), splicing factor 3a, subunit 3, 60 kDa (SF3A3), thrombospondin 1 (THBS1), chaperonin containing TCP1, subunit 6A (zeta 1) (CCT6A), PRP8 pre-mRNA processing factor 8 homolog (S. cerevisiae) (PRPF8), symplekin (SYMPK), far upstream element (FUSE) binding protein 1 (FUBP1), U2 small nuclear RNA auxiliary factor 1 (U2AF1), huntingtin (HTT), eukaryotic translation initiation factor 5B (EIFSB), nuclear autoantigenic sperm protein (histone-binding) (NASP), heterogeneous nuclear ribonucleoprotein K (HNRNPK), Y box binding protein 1 (YBX1), annexin A1 1 (ANXA1 1), RecQ protein-like (DNA helicase Q1-like) (RECQL), cortactin (CTTN), tubulin, beta 3 (TUBB3), pyrroline-5-carboxylate reductase-like (PYCRL), periplakin (PPL), phosphoglucomutase 2-like 1 (PGM2L1), chromosome 17 open reading frame 49 (C17orf49), mRNA turnover 4 homolog (S. cerevisiae) (MRT04), methyltransferase like 1 (METTL1), squamous cell carcinoma antigen recognized by T cells 3 (SART3), S100 calcium binding protein A13 (S100A13), aminopeptidase-like 1 (NPEPL1), cyclin-dependent kinase 1 (CDK1), ubiquitin protein ligase E3 component n-recognin 1 (UBR1), Rho GTPase activating protein 18 (ARHGAP18), signal recognition particle 14 kDa (homologous Alu RNA binding protein) (SRP14), cathelicidin antimicrobial peptide (CAMP), splicing factor proline/glutamine-rich (SFPQ), RAS p21 protein activator (GTPase activating protein) 1 (RASA1), Ral GTPase activating protein, beta subunit (non-catalytic) (RALGAPB), laminin, beta 1 (LAMB1), RAB3 GTPase activating protein subunit 2 (non-catalytic) (RAB3GAP2), chaperonin containing TCP1, subunit 8 (theta) (CCT8), heterogeneous nuclear ribonucleoprotein L-like (HNRPLL), RAN binding protein 1 (RANBP1), kinetochore associated 1 (KNTC1), dyskeratosis congenita 1, dyskerin (DKC1), casein kinase 2, alpha 1 polypeptide (CSNK2A1), CAP-GLY domain containing linker protein 1 (CLIP1), chaperonin containing TCP1, subunit 2 (beta) (CCT2), tubulin tyrosine ligase-like family, member 12 (TTLL12), ataxia telangiectasia mutated (ATM), splicing factor 3a, subunit 1, 120 kDa (SF3A1), ribosomal protein S20 (RPS20), ubiquitin-conjugating enzyme E20 (UBE20), translocated promoter region (to activated MET oncogene) (TPR), BRCA2 and CDKN1A interacting protein (BCCIP), gem (nuclear organelle) associated protein 5 (GEMIN5), ribonuclease P/MRP 30 kDa subunit (RPP30), loss of heterozygosity, 12, chromosomal region 1 (LOH 12CR1), syntaxin binding protein 2 (STXBP2), ubiquitin-conjugating enzyme E2H (UBE2H), DIP2 disco-interacting protein 2 homolog B (Drosophila) (DIP2B), RAPT, GTP-GDP dissociation stimulator 1 (RAP1GDS1), heterogeneous nuclear ribonucleoprotein M (HNRNPM), LIM domain 7 (LM07), RNA binding motif protein 25 (RBM25), aldehyde dehydrogenase 7 family, member A1 (ALDH7A1), cleavage and polyadenylation specific factor 1, 160 kDa (CPSF1), calponin 2 (CNN2), chaperonin containing TCP1, subunit 7 (eta) (CCT7), lysyl-tRNA synthetase (KARS), UDP-N-acteylglucosamine pyrophosphorylase 1 (UAP1), heat shock 70 kDa protein 4-like (HSPA4L), 138 kDa protein (138 kDa protein), thimet oligopeptidase 1 (THOP1), glutaredoxin 3 (GLRX3), phosphoglycerate dehydrogenase (PHGDH), CDV3 homolog (mouse) (CDV3), structural maintenance of chromosomes 4 (SMC4), RNA binding motif (RNP1, RRM) protein 3 (RBM3), hepatoma-derived growth factor (HDGF), heterogeneous nuclear ribonucleoprotein U (scaffold attachment factor A) (HNRNPU), nuclear receptor binding protein 1 (NRBP1), polymerase (RNA) I polypeptide B, 128 kDa (POLR1 B), protein phosphatase 5, catalytic subunit (PPP5C), glucose-6-phosphate dehydrogenase (G6PD), arginase, liver (ARG1), 3-hydroxy-3-methylglutaryl-CoA synthase 1 (soluble) (HMGCS1), ubiquitin-like modifier activating enzyme 2 (UBA2), KIAA1033 (KIAA1033), annexin A4 (ANXA4), DEAD (Asp-Glu-Ala-Asp) box polypeptide 17 (DDX17), acidic (leucine-rich) nuclear phosphoprotein 32 family, member E (ANP32E), glucosamine (UDP-N-acetyl)-2-epimerase/N-acetylmannosamine kinase (GNE), KIAA0368 (KIAA0368), vacuolar protein sorting 4 homolog B (S. cerevisiae) (VPS4B), replication protein A1, 70 kDa (RPA1), eukaryotic translation initiation factor 2, subunit 1 alpha, 35 kDa (EIF2S1), eukaryotic translation initiation factor 3, subunit J (EIF3J), suppressor of Ty 6 homolog (S. cerevisiae) (SUPT6H), heat shock 105 kDa/110 kDa protein 1 (HSPH1), exportin 5 (XP05), transcription elongation factor A (S11), 1 (TCEA1), Sjogren syndrome antigen B (autoantigen La) (SSB), AE binding protein 1 (AEBP1), LIM and cysteine-rich domains 1 (LMCD1), interleukin enhancer binding factor 3, 90 kDa (ILF3), WD repeat domain 61 (WDR61), N(alpha)-acetyltransferase 15, NatA auxiliary subunit (NAA15), serine/arginine-rich splicing factor 4 (SRSF4), ring finger protein 20 (RNF20), lactamase, beta 2 (LACTB2), NHP2 ribonucleoprotein homolog (yeast) (NHP2), chromosome 17 open reading frame 28 (C17orf28), CTP synthase II (CTPS2), fascin homolog 1, actin-bundling protein (Strongylocentrotus purpuratus) (FSCN1), tRNA nucleotidyl transferase, CCA-adding, 1 (TRNT1), splicing regulatory glutamine/lysine-rich protein 1 (SREK1), stromal antigen 1 (STAG1), oxysterol binding protein (OSBP), deoxyuridine triphosphatase (DUT), coiled-coil domain containing 25 (CCDC25), DEK oncogene (DEK), coiled-coil domain containing 72 (CCDC72), polymerase (RNA) II (DNA directed) polypeptide E, 25 kDa (POLR2E), phosphoserine phosphatase (PSPH), structural maintenance of chromosomes 1A (SMC1A), DEAD (Asp-Glu-Ala-Asp) box polypeptide 23 (DDX23), tRNA methyltransferase 11-2 homolog (S. cerevisiae) (TRMT112), COP9 constitutive photomorphogenic homolog subunit 2 (Arabidopsis) (COPS2), programmed cell death 5 (PDCD5), cyclin-dependent kinase 2 (CDK2), proteasome (prosome, macropain) 26S subunit, non-ATPase, 3 (PSMD3), RAN binding protein 2 (RANBP2), SERPINE1 mRNA binding protein 1 (SERBP1), O-linked N-acetylglucosamine (G1cNAc) transferase (UDP-N-acetylglucosamine:polypeptide-N-acetylglucosaminyl transferase) (OGT), non-SMC condensin I complex, subunit D2 (NCAPD2), SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily c, member 2 (SMARCC2), NOP10 ribonucleoprotein homolog (yeast) (NOP10), LPS-responsive vesicle trafficking, beach and anchor containing (LRBA), apoptosis inhibitor 5 (API5), signal recognition particle receptor (docking protein) (SRPR), transferrin receptor (p90, CD71) (TFRC), basic leucine zipper and W2 domains 2 (BZW2), ribonuclease, RNase A family, 3 (RNASE3), diazepam binding inhibitor (GABA receptor modulator, acyl-CoA binding protein) (DBI), FK506 binding protein 4, 59 kDa (FKBP4), chromosome 6 open reading frame 130 (C6orf130), cofactor of BRCA1 (COBRA1), flap structure-specific endonuclease 1 (FEN1), glucan (1,4-alpha-), branching enzyme 1 (GBE1), small nuclear ribonucleoprotein polypeptide B (SNRPB2), NSFL1 (p9′7) cofactor (p4′7) (NSFL1 C), acyl-CoA thioesterase 7 (ACOT7), NOP2/Sun domain family, member 2 (NSUN2), chaperonin containing TCP1, subunit 4 (delta) (CCT4), kallikrein-related peptidase 6 (KLK6), glutaminyl-peptide cyclotransferase (QPCT), BCL2-associated athanogene 6 (BAG6), eukaryotic translation initiation factor 3, subunit C (EIF3C/EIF3CL), ATPase, H+ transporting, lysosomal 56/58 kDa, V1 subunit B2 (ATP6V1 B2), matrix metallopeptidase 8 (neutrophil collagenase) (MMP8), proteasome (prosome, macropain) 26S subunit, ATPase, 5 (PSMC5), GTP cyclohydrolase I feedback regulator (GCHFR), poly(A) polymerase alpha (PAPOLA), hippocalcin-like 1 (HPCAL1), GTPase activating protein (SH3 domain) binding protein 1 (G3BP1), polymerase (RNA) III (DNA directed) polypeptide A, 155 kDa (POLR3A), superkiller viralicidic activity 2-like 2 (S. cerevisiae) (SKIV2L2), polymerase (RNA) II (DNA directed) polypeptide A, 220 kDa (POLR2A), collagen, type I, alpha 2 (COL1A2), fibrillarin (FBL), glutamyl-prolyl-tRNA synthetase (EPRS), ELAV (embryonic lethal, abnormal vision, Drosophila)-like 1 (Hu antigen R) (ELAVL1), nuclear cap binding protein subunit 2, 20 kDa (NCBP2), GCN1 general control of amino-acid synthesis 1-like 1 (yeast) (GCN1 L1), histone acetyltransferase 1 (HAT1), stromal antigen 2 (STAG2), sorbitol dehydrogenase (SORD), REX2, RNA exonuclease 2 homolog (S. cerevisiae) (REX02), heterogeneous nuclear ribonucleoprotein F (HNRNPF), thymopoietin (TMPO), ubiquitin specific peptidase 24 (USP24), KIAA1967 (KIAA1967), complement component 1, r subcomponent (C1 R), annexin A7 (ANXA7), RuvB-like 2 (E. coli) (RUVBL2), acireductone dioxygenase 1 (ADI 1), eukaryotic translation initiation factor 4A3 (EIF4A3), heterogeneous nuclear ribonucleoprotein U-like 2 (HNRNPUL2), ubiquitin protein ligase E3 component n-recognin 4 (UBR4), SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 2 (SMARCA2), cytochrome b5 reductase 2 (CYB5R2), splicing factor 3b, subunit 3, 130 kDa (SF3B3), G protein pathway suppressor 1 (GPS1), MAD2 mitotic arrest deficient-like 1 (yeast) (MAD2L1), phospholipase C, gamma 1 (PLCG1), eukaryotic translation initiation factor 4H (EIF4H), U2 small nuclear RNA auxiliary factor 2 (U2AF2), tankyrase 1 binding protein 1, 182 kDa (TNKS1 BP1), transglutaminase 2 (C polypeptide, protein-glutamine-gamma-glutamyltransferase) (TGM2), heterogeneous nuclear ribonucleoprotein L (HNRNPL), inositol polyphosphate-5-phosphatase, 145 kDa (INPP5D), annexin A10 (ANXA10), BUD31 homolog (S. cerevisiae) (BUD31), phosphatidylinositol transfer protein, alpha (PITPNA), leucyl-tRNA synthetase (LARS), nicotinamide N-methyltransferase (NNMT), proteasome (prosome, macropain) 26S subunit, non-ATPase, 12 (PSMD12), v-crk sarcoma virus CT10 oncogene homolog (avian) (CRK), proteoglycan 2, bone marrow (natural killer cell activator, eosinophil granule major basic protein) (PRG2), versican (VCAN), exportin, tRNA (nuclear export receptor for tRNAs) (XPOT), EMG1 nucleolar protein homolog (S. cerevisiae) (EMG1), chromosome 11 open reading frame 73 (C11orf73), transports 1 (TNP01), latent transforming growth factor beta binding protein 2 (LTBP2), cold shock domain containing E1, RNA-binding (CSDE1), sulfiredoxin 1 (SRXN1), paraspeckle component 1 (PSPC1), ribosomal protein S3A (RPS3A), ISG15 ubiquitin-like modifier (ISG15), polymerase (RNA) II (DNA directed) polypeptide B, 140 kDa (POLR2B), general transcription factor IN (GTF2I), NHP2 nonhistone chromosome protein 2-like 1 (S. cerevisiae) (NHP2L1), proteasome (prosome, macropain) 26S subunit, non-ATPase, 10 (PSMD10), signal transducer and activator of transcription 1, 91 kDa (STAT1), elongation factor Tu GTP binding domain containing 1 (EFTUD1), mediator complex subunit 23 (MED23), eukaryotic translation initiation factor 2C, 2 (EIF2C2), RNA binding motif protein 4B (RBM4B), KIAA0664 (KIAA0664), core-binding factor, beta subunit (CBFB), poly(A) binding protein, cytoplasmic 1 (PABPC1), nicotinamide phosphoribosyltransferase (NAMPT), cellular retinoic acid binding protein 2 (CRABP2), thyroid hormone receptor interactor 12 (TRIP12), DnaJ (Hsp40) homolog, subfamily C, member 9 (DNAJC9), StAR-related lipid transfer (START) domain containing 10 (STARD10), ring finger protein 213 (RNF213), eukaryotic translation initiation factor 2B, subunit 5 epsilon, 82 kDa (EIF2B5), bolA homolog 2 (E. coli) (BOLA2/BOLA2B), meningioma expressed antigen 5 (hyaluronidase) (MGEA5), Rab geranylgeranyltransferase, alpha subunit (RABGGTA), pyridoxal-dependent decarboxylase domain containing 1 (PDXDC1), exosome component 8 (EXOSC8), phosphoserine aminotransferase 1 (PSAT1), eukaryotic translation initiation factor 6 (EIF6), chromosome 16 open reading frame 13 (C16orf13), signal transducer and activator of transcription 3 (acute-phase response factor) (STAT3), EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1), defensin, alpha 6, Paneth cell-specific (DEFA6), pyrophosphatase (inorganic) 1 (PPA1), glutathione peroxidase 2 (gastrointestinal) (GPX2), unc-13 homolog D (C. elegans) (UNCI 3D), protein tyrosine phosphatase, non-receptor type 6 (PTPN6), myosin XVIIIA (MY018A), fibulin 1 (FBLN1), ribosomal protein L19 (RPL19), diaphanous homolog 1 (Drosophila) (DIAPH1), MMS19 nucleotide excision repair homolog (S. cerevisiae) (MMS19), nudix (nucleoside diphosphate linked moiety X)-type motif 21 (NUDT21), splicing factor 3b, subunit 5, 10 kDa (SF3B5), SAP domain containing ribonucleoprotein (SARNP), ADP-ribosylation factor guanine nucleotide-exchange factor 2 (brefeldin A-inhibited) (ARFGEF2), guanylate binding protein 1, interferon-inducible (GBP1), HECT, UBA and WWE domain containing 1 (HUWE1), processing of precursor 7, ribonuclease P/MRP subunit (S. cerevisiae) (POP7), eukaryotic translation initiation factor 2B, subunit 3 gamma, 58 kDa (EIF2B3), N(alpha)-acetyltransferase 10, NatA catalytic subunit (NAA10), DnaJ (Hsp40) homolog, subfamily B, member 1 (DNAJB1), eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2), elongation factor Tu GTP binding domain containing 2 (EFTUD2), grancalcin, EF-hand calcium binding protein (GCA), neuronal cell adhesion molecule (NRCAM), DEAH (Asp-Glu-Ala-His) box polypeptide 16 (DHX16), small glutamine-rich tetratricopeptide repeat (TPR)-containing, alpha (SGTA), serine/threonine kinase 10 (STK10), smu-1 suppressor of mec-8 and unc-52 homolog (C. elegans) (SMUT), PRP4 pre-mRNA processing factor 4 homolog (yeast) (PRPF4), heterogeneous nuclear ribonucleoprotein D (AU-rich element RNA binding protein 1, 37 kDa) (HNRNPD), CTP synthase (CTPS), cleavage stimulation factor, 3′ pre-RNA, subunit 3, 77 kDa (CSTF3), cleavage stimulation factor, 3′ pre-RNA, subunit 1, 50 kDa (CSTF1), heterogeneous nuclear ribonucleoprotein U-like 1 (HNRNPUL1), ribosomal protein S5 (RPS5), protein tyrosine phosphatase, nonreceptor type 1 1 (PTPN1 1), ladinin 1 (LAD1), component of oligomeric golgi complex 2 (COG2), cullin 2 (CUL2), ribosomal protein S17 (RPS17/RPS17L), proteasome (prosome, macropain) 26S subunit, non-ATPase, 4 (PSMD4), annexin A1 (ANXA1), annexin A2 (ANXA2), 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), azurocidin 1 (AZU1), baculoviral IAP repeat containing 6 (BIRC6), chitinase 3-like 1 (cartilage glycoprotein-39) (CHI3L1), carbamoyl-phosphate synthase 1, mitochondrial (CPS1), cathepsin G (CTSG), defensin, alpha 1 (DEFA1 (includes others)), deleted in malignant brain tumors 1 (DMBT1), elastase, neutrophil expressed (ELANE), integrin, alpha M (complement component 3 receptor 3 subunit) (ITGAM), lipocalin 2 (LCN2), lectin, galactoside-binding, soluble, 3 binding protein (LGALS3BP), minichromosome maintenance complex component 2 (MCM2), matrix metallopeptidase 9 (gelatinase B, 92 kDa gelatinase, 92 kDa type IV collagenase) (MMP9), myeloperoxidase (MPO), mucin 5AC, oligomeric mucus/gel-forming (MUC5AC/MUC5B), nuclear cap binding protein subunit 1, 80 kDa (NCBP1), neurofibromin 1 (NF1), olfactomedin 4 (OLFM4), PDS5, regulator of cohesion maintenance, homolog A (S. cerevisiae) (PDS5A), peptidoglycan recognition protein 1 (PGLYRP1), proteinase 3 (PRTN3), quiescin Q6 sulfhydryl oxidase 1 (QSOX1), regenerating islet-derived 1 alpha (REG1A), S100 calcium binding protein A9 (S100A9), serpin peptidase inhibitor, clade B (ovalbumin), member 10 (SERPINB10), serpin peptidase inhibitor, clade B (ovalbumin), member 5 (SERPINB5), unc-45 homolog A (C. elegans) (UNC45A);
69. The array of claim 68, wherein the binding agent is an antibody or antibody fragment.
70. The array of claim 69, wherein the antibody or fragment thereof is selected from the group consisting of: a recombinant antibody; scFv; Fab; a binding domain of an immunoglobulin molecule.
71. A method for treating colorectal cancer, the method comprising: (i) performing the assay of claim 67; and(ii) administering a therapeutically active amount of a cancer therapeutic agent to the individual identified in step (iii) as having at least a predetermined level of binding of the binding agent to at least four markers to be at risk of, or suffering from colorectal cancer.
72. The method of claim 71, wherein the biological sample comprises a stool sample.
73. The method of claim 71, wherein the cancer therapeutic agent comprises one or more monoclonal antibody, one or more small molecule inhibitor or one or more chemotherapeutic agent, or any combination thereof.
74. The method of claim 71, wherein the one or more therapeutic agents is selected from the group of: bevacizumab, cetuximab, panitumumab, erlotinib, sorafenib, alisertib, 5-FU, capecitabine, irinotecan oxaliplatin, or leucovorin or any combination thereof.
75. A kit for screening for colorectal cancer in an individual, the kit comprising: (a) vials containing at least four of the protein binding agents added to the assay of claim 1;(b) a specimen container; and(c) instructions for performing the assay set forth in claim 1.
76. The assay of claim 1, wherein the at least four biomarkers are selected from secretomes.

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Number	Date	Country	Kind
2008707	Apr 2012	NL	national
2010276	Feb 2013	NL	national

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Filing Document	Filing Date	Country	Kind
PCT/NL13/50316	4/26/2013	WO	00

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