HUMANIZED MONOCLONAL ANTIBODIES AND METHODS OF USE FOR THE DIAGNOSIS AND TREATMENT OF COLON AND PANCREAS CANCER

Abstract

This invention relates to humanized antibodies that selectively bind the 31.1 epitope on the A33 protein differentially expressed in cancers including, lung cancer, ovarian cancer, pancreas cancer, breast cancer, and colon cancer, and diagnostic and therapeutic usages.

Description

Claims

1. An isolated antibody that binds to an A33 antigen, or a fragment thereof, comprising: a) at least one heavy chain sequence comprising the amino acid sequence of SEQ ID NO: 74, 75, 76, or 77 or one of the variable regions contained therein;b) at least one heavy chain CDR sequence comprising at least one amino acid sequence of SEQ ID NO:78, 79, 80, 81, 82, or 83;c) at least one light chain sequence comprising the amino acid sequence of SEQ ID NO:84, 85, 86, 87:d) at least one light chain CDR sequence comprising at least one amino acid sequence of SEQ ID NO:88, 89, 90, 91, 92, or 93;e) a heavy chain comprising the amino acid sequence of SEQ ID NO:74 and the light chain amino acid sequence of SEQ ID NO:84;f) a heavy chain comprising the amino acid sequence of SEQ ID NO:75 and the light chain amino acid sequence of SEQ ID NO:85;g) a heavy chain comprising the amino acid sequence of SEQ ID NO:76 and the light chain amino acid sequence of SEQ ID NO:86:h) a heavy chain comprising the amino acid sequence of SEQ ID NO:77 and the light chain amino acid sequence of SEQ ID NO:87;i) a heavy chain comprising the CDR1 amino acid sequence of SEQ ID NO:78, the CDR2 amino acid sequence of SEQ ID NO:79, 80, 81, or 82 and the CDR3 amino acid sequence of SEQ ID NO:83;j) a heavy chain comprising the CDR1 amino acid sequence of SEQ ID NO:78, the CDR2 amino acid sequence of SEQ ID NO:79, and the CDR3 amino acid sequence of SEQ ID NO:83:k) a heavy chain comprising the CDR1 amino acid sequence of SEQ ID NO:78, the CDR2 amino acid sequence of SEQ ID NO:80 and the CDR3 amino acid sequence of SEQ ID NO:83;1) a heavy chain comprising the CDR1 amino acid sequence of SEQ ID NO:78, the CDR2 amino acid sequence of SEQ ID NO:81 and the CDR3 amino acid sequence of SEQ ID NO:83;m) a heavy chain comprising the CDR1 amino acid sequence of SEQ ID NO:78, the CDR2 amino acid sequence of SEQ ID NO:82 and the CDR3 amino acid sequence of SEQ ID NO:83;n) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:88, 89, or 90, the CDR2 amino acid sequence of SEQ ID NO:91 or 92 and the CDR3 amino acid sequence of SEQ ID NO:93;o) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:88, the CDR2 amino acid sequence of SEQ ID NO:91 and the CDR3 amino acid sequence of SEQ ID NO:93;p) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:89, the CDR2 amino acid sequence of SEQ ID NO:91 and the CDR3 amino acid sequence of SEQ ID NO:93;q) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:90, the CDR2 amino acid sequence of SEQ ID NO:91 and the CDR3 amino acid sequence of SEQ ID NO:93;r) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:88, the CDR2 amino acid sequence of SEQ ID NO:92 and the CDR3 amino acid sequence of SEQ ID NO:93;s) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO: 89, the CDR2 amino acid sequence of SEQ ID NO:92 and the CDR3 amino acid sequence of SEQ ID NO:93;t) a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:90, the CDR2 amino acid sequence of SEQ ID NO:92 and the CDR3 amino acid sequence of SEQ ID NO:93;u) a heavy chain comprising the CDR1 amino acid sequence of SEQ ID NO:78, the CDR2 amino acid sequence of SEQ ID NO:79, 80, 81, or 82 and the CDR3 amino acid sequence of SEQ ID NO:83 and a light chain comprising the CDR1 amino acid sequence of SEQ ID NO:88, 89, or 90, the CDR2 amino acid sequence of SEQ ID NO:91 or 92 and the CDR3 amino acid sequence of SEQ ID NO:93;v) a heavy chain comprising the CDRs contained in antibody cdr31.1 -HC (SEQ ID NO:74) and a light chain comprising the CDRs contained in antibody cdr31.1 -LC (SEQ ID NO:84);w) a heavy chain comprising the CDRs contained in antibody cdr31.1-HC (SEQ ID NO:74) and a light chain comprising the CDRs contained in antibody abb31.1 -LC (SEQ ID NO:85);x) a heavy chain comprising the CDRs contained in antibody cdr31.1-HC (SEQ ID NO:74) and a light chain comprising the CDRs contained in antibody sdr3 1.1-LC (SEQ ID NO:86);y) a heavy chain comprising the CDRs contained in antibody cdr3 1.1-HC (SEQ ID NO:74) and a light chain comprising the CDRs contained in antibody ven31.1 -LC (SEQ ID NO:87);z) a heavy chain comprising the CDRs contained in antibody abb31.1 -HC (SEQ ID NO:75) and a light chain comprising the CDRs contained in antibody cdr3 1.1 -LC (SEQ ID NO:84);aa) a heavy chain comprising the CDRs contained in antibody abb3 1.1 -HC (SEQ ID NO:75) and a light chain comprising the CDRs contained in antibody abb31.1 -LC (SEQ ID NO:85);bb) a heavy chain comprising the CDRs contained in antibody abb31.1 -HC (SEQ ID NO:75) and a light chain comprising the CDRs contained in antibody sdr31.1 -L C (SEQ ID NO:86);cc) a heavy chain comprising the CDRs contained in antibody abb3 1.1 -HC (SEQ ID NO:75) and a light chain comprising the CDRs contained in antibody ven31.1 -LC (SEQ ID NO:87);dd) a heavy chain comprising the CDRs contained in antibody sdr31.1 -HC (SEQ ID NO:76) and a light chain comprising the CDRs contained in antibody cdr31.1 -LC (SEQ ID NO:84);ee) a heavy chain comprising the CDRs contained in antibody sdr31.1 -HC (SEQ ID NO:76) and a light chain comprising the CDRs contained in antibody abb3 1.1-LC (SEQ ID NO:85);ff) a heavy chain comprising the CDRs contained in antibody sdr31.1 -HC (SEQ ID NO:76) and a light chain comprising the CDRs contained in antibody sdr31 .1 -LC (SEQ ID NO: 86);gg) a heavy chain comprising the CDRs contained in antibody sdr31.1 -HC (SEQ ID NO:76) and a light chain comprising the CDRs contained in antibody ven31.1-LC (SEQ 1D NO:87);hh) a heavy chain comprising the CDRs contained in antibody ven31.1-HC (SEQ ID NO:77) and a light chain comprising the CDRs contained in antibody cdr31.1-LC (SEQ ID NO:84);ii) a heavy chain comprising the CDRs contained in antibody ven31.1 -HC (SEQ ID NO:77) and a light chain comprising the CDRs contained in antibody abb31.1-LC (SEQ ID NO:85);jj) a heavy chain comprising the CDRs contained in antibody ven31.1 -HC (SEQ ID NO:77) and a light chain comprising the CDRs contained in antibody sdr31.1 -LC (SEQ ID NO:86);kk) a heavy chain comprising the CDRs contained in antibody ven31.1 -HC (SEQ ID NO:77) and a light chain comprising the CDRs contained in antibody ven31.1 -LC (SEQ ID NO:87);11) a heavy chain comprising the variable chain of antibody cdr31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:74) and a light chain comprising the variable light chain of antibody cdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO:84);mm) a heavy chain comprising the variable chain of antibody cdr31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:74) and a light chain comprising the variable light chain of antibody abb31.1 -LC (amino acids 1 to 107 of SEQ ID NO:85);nn) a heavy chain comprising the variable chain of antibody cdr31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:74) and a light chain comprising the variable light chain of antibody sdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO: 86);oo) a heavy chain comprising the variable chain of antibody cdr31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:74) and a light chain comprising the variable light chain of antibody ven31.1 -LC (amino acids 1 to 107 of SEQ ID NO:87);pp) a heavy chain comprising the variable chain of antibody abb31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:75) and a light chain comprising the variable light chain of antibody cdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO:84);qq) a heavy chain comprising the variable chain of antibody abb31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:75) and a light chain comprising the variable light chain of antibody abb31.1 -L C (amino acids 1 to 107 of SEQ ID NO:85);rr) a heavy chain comprising the variable chain of antibody abb31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:75) and a light chain comprising the variable light chain of antibody sdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO:86):ss) a heavy chain comprising the variable chain of antibody abb3 1.1-HC (amino acids 1 to 1 18 of SEQ ID NO:75) and a light chain comprising the variable light chain of antibody ven31.1 -LC (amino acids 1 to 107 of SEQ ID NO:87);tt) a heavy chain comprising the variable chain of antibody sdr31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:76) and a light chain comprising the variable light chain of antibody cdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO:84);uu) a heavy chain comprising the variable chain of antibody sdr31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:76) and a light chain comprising the variable light chain of antibody abb31.1 -LC (amino acids 1 to 107 of SEQ ID NO:85);vv) a heavy chain comprising the variable chain of antibody sdr31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:76) and a light chain comprising the variable light chain of antibody sdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO:86);ww) a heavy chain comprising the variable chain of antibody sdr31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:76) and a light chain comprising the variable light chain of antibody ven31.1 -LC (amino acids 1 to 107 of SEQ ID NO:87);xx) a heavy chain comprising the variable chain of antibody ven31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:77) and a light chain comprising the variable light chain of antibody cdr31.1 -LC (amino acids 1 to 107 of SEQ ID NO:84);yy) a heavy chain comprising the variable chain of antibody ven31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:77) and a light chain comprising the variable light chain of antibody abb31.1 -LC (amino acids 1 to 107 of SEQ ID NO:85);zz) a heavy chain comprising the variable chain of antibody ven31.1-HC (amino acids 1 to 1 18 of SEQ ID NO:77) and a light chain comprising the variable light chain of antibody sdr31 .1 -LC (amino acids 1 to 107 of SEQ ID NO:86);aaa) a heavy chain comprising the variable chain of antibody ven31.1 -HC (amino acids 1 to 1 18 of SEQ ID NO:77) and a light chain comprising the variable light chain of antibody ven31 .1 -LC (amino acids 1 to 107 of SEQ ID NO:87):bbb) a heavy chain polypeptide of antibody cdr31.1 -HC (SEQ ID NO:74) and a light chain polypeptide of antibody cdr3 1.1 -LC (SEQ ID NO: 84);ccc) a heavy chain polypeptide of antibody cdr31.1 -HC (SEQ ID NO:74) and a light chain polypeptide of antibody abb31.1 -LC (SEQ ID NO:85);ddd) a heavy chain polypeptide of antibody cdr31.1 -HC (SEQ ID NO: 74) and a light chain polypeptide of antibody sdr31.1 -LC (SEQ ID NO:86);eee) a heavy chain polypeptide of antibody cdr31.1-HC (SEQ ID NO:74) and a light chain polypeptide of antibody ven3 1.1-LC (SEQ ID NO:87);fff) a heavy chain polypeptide of antibody abb31.1 -HC (SEQ ID NO: 75) and a light chain polypeptide of antibody cdr31.1-LC (SEQ ID NO:84);ggg) a heavy chain polypeptide of antibody abb31.1 -HC (SEQ ID NO:75) and a light chain polypeptide of antibody abb31.1 -LC (SEQ ID NO: 85);hhh) a heavy chain polypeptide of antibody abb3 1.1 -HC (SEQ ID NO:75) and a light chain polypeptide of antibody sdr31.1 -LC (SEQ ID NO: 86);iii) a heavy chain polypeptide of antibody abb31.1 -HC (SEQ ID NO:75) and a light chain polypeptide of antibody ven31.1 -LC (SEQ ID NO:87);jjj) a heavy chain polypeptide of antibody sdr3 1.1 -HC (SEQ ID NO:76) and a light chain polypeptide of antibody cdr31.1 -LC (SEQ ID NO:84);kkk) a heavy chain polypeptide of antibody sdr31.1 -HC (SEQ ID NO:76) and a light chain polypeptide of antibody abb31.1 -L C (SEQ ID NO:85);lll) a heavy chain polypeptide of antibody sdr31.1 -HC (SEQ ID NO:76) and a light chain polypeptide of antibody sdr31.1 -LC (SEQ ID NO: 86);mmm) a heavy chain polypeptide of antibody sdr3 1.1 -HC (SEQ ID NO:76) and a light chain polypeptide of antibody ven31 .1 -LC (SEQ ID NO:87);nnn) a heavy chain polypeptide of antibody ven31.1 -HC (SEQ ID NO:77) and a light chain polypeptide of antibody cdr31.1-LC (SEQ ID NO:84);ooo) a heavy chain polypeptide of antibody ven3 1.1 -HC (SEQ ID NO:77) and a light chain polypeptide of antibody abb31.1 -L C (SEQ ID NO:85);ppp) a heavy chain polypeptide of antibody ven31.1 -HC (SEQ ID NO:77) and a light chain polypeptide of antibody sdr31.1-LC (SEQ ID NO:86); orqqq) a heavy chain polypeptide of antibody ven31.1 -HC (SEQ ID NO:77) and a light chainpolypeptide of antibody ven31.1-LC (SEQ ID NO: 87).

2. The isolated antibody or fragment thereof of claim 1, which comprises at least one heavy chain or heavy chain variable region and at least one light chain or light chain variable region, comprises at least one light chain CDR and at least one heavy chain CDR, comprises at least two light chain CDRs and at least two heavy chain CDRs, or comprises at least three light chain CDRs and three heavy chain CDRs.

3. The isolated antibody or fragment thereof of claim 1 which specifically binds the amino acid sequence of one or more of SEQ ID NOs: 10, 11, 12, or 13.

4. The isolated antibody or fragment thereof of claim 1 which has anti-tumor activity, is a human IgG 1 or human IgG3 antibody, or is a Fab, Fab′, F(ab′)2, Fv, CDR, paratope, or portion of an antibody that is capable of binding the antigen, is directly or indirectly conjugated to a label, cytotoxic agent, therapeutic agent, or an immunosuppressive agent, and/or said antibody potentiates ADCC or CDC.

5. The isolated antibody or fragment thereof of claim 1, which is admixed in a composition with an antibody, label, cytotoxic agent, therapeutic agent, or an immunosuppressive agent, is administered in combination with an antibody, label, cytotoxic agent, therapeutic agent, or an immunosuppressive agent, simultaneously or sequentially, wherein said label is optionally a chemiluminescent label, paramagnetic label, an MRI contrast agent, fluorescent label, bioluminescent label, or radioactive label, wherein said paramagnetic label is optionally paramagnetic label is aluminum, manganese, platinum, oxygen, lanthanum, lutetium, scandium, yttrium, or gallium, wherein the cytotoxic agent is optionally a moiety that inhibits DNA, RNA, or protein synthesis, a radionuclide, ribosomal inhibiting protein, 212Bi, 131l, 188Re, 90Y, vindesine, methotrexate, adriamycin, cisplatin, pokeweed antiviral protein, Pseudomonas exotoxin A, ricin, diphtheria toxin, ricin A chain, or cytotoxic phospholipase enzyme, wherein optionally said therapeutic agent is a lymphokine or growth factor, and/or wherein optionally wherein said immunosuppressive agent is cyclosporine, leflunomide, methotrexate, azothiprine, mercaptopurine, dactinomycin, tacrolimus, or sirolimus.

6. A composition comprising an antibody or fragment thereof according to claim 1, optionally comprising a pharmaceutically acceptable carrier.

7. A method of preventing or treating cancer comprising administering an effective amount of the antibody or fragment thereof of claim 1, optionally in combination simultaneously or sequentially with another antibody, a lymphokine, or a hematopoietic growth factor, wherein optionally said antibody slows the growth or replication of cancer cells in the patient, kills cancer cells in the patient, and/or promotes tumor regression in the patient, wherein optionally said cancer is lung, breast, pancreas, uterine, esophageal, colorectal, or liver cancer, wherein optionally said cancer is stage 1, 2, 3, or 4, wherein optionally said cancer has metastasized, wherein optionally said patient expresses detectable levels of a 3 1.1 epitope, wherein optionally said patient is at risk of cancer or is optionally without symptoms.

8. The method of claim 7, further comprising detecting the 31.1 epitope in a tumor sample, lymph, urine, cerebrospinal fluid, amniotic fluid, inflammatory exudate, blood, serum, stool, or liquid collected from the colorectal tract, thereby determining that said tumor contains cells that bind to said antibody.

9. A method for detecting a 31.1 epitope, comprising: (a) contacting a test sample with the antibody or antibody fragment of claim 1, that binds an 31.1 epitope, wherein said antibody or fragment is optionally conjugated to a label, chemiluminescent label, paramagnetic label, an MRI contrast agent, fluorescent label, bioluminescent label, radioactive label, wherein optionally said paramagnetic label is aluminum, manganese, platinum, oxygen, lanthanum, lutetium, scandium, yttrium, or gallium and(b) detecting antibody-epitope complexes,wherein the presence of said epitope is indicative of a carcinoma, optionally lung, breast, pancreas, uterine, esophageal, colorectal, or liver cancer,wherein step (a) optionally comprises administering the antibody to said patient,wherein optionally said test sample is obtained from a patient at risk of cancer and/or without symptoms.

10. The method of claim 9, which detects colorectal polyps, benign tumors, malignant tumors, metastatic tumors, non-metastatic tumors, and/or pre-cancerous cells.

11. The method of claim 9, wherein the antibody-epitope complex is detected in vitro by an assay selected from the group consisting of Western blot, radioimmunoassays, EL1SA (enzyme linked immunosorbent assay), “sandwich” immunoassays, lateral flow assay, immunoprecipitation assays, precipitation reactions, gel diffusion precipitation reactions, immunodiffusion assays, agglutination assays, complement-fixation assays, immunohistochemical assays, fluorescent immunoassays, and protein A immunoassays, or is detected in vivo by a method comprising positron emission tomography (PET), CCD low-light monitoring system, x-ray, CT scanning, scintigraphy, photo acoustic imaging, single photon emission computed tomography (SPECT), magnetic resonance imaging (MRI), ultrasound, paramagnetic imaging, and endoscopic optical coherence tomography.

12. An isolated nucleic acid encoding an antibody or fragment thereof according to claim 1, or a heavy chain or light chain thereof, wherein optionally said nucleic acid is contained in a vector or host cell.

13. A method of making the antibody or fragment thereof of claim 1, comprising expressing a nucleic acid encoding said antibody or fragment thereof in a cell or a cell-free translation system.

14. The method of making the antibody or fragment thereof of claim 13,which further includes isolating or purifying the expressed antibody or fragment thereof.