Patent Application
20170360863
The present invention relates generally to methods for suppressing myeloid-derived suppressor cells.
Owing to the recent advancement in precision surgeries, early diagnosis of cancer, and adjuvant therapies with chemotherapeutic drugs, cancer death rate now is mainly reflecting the degree and pattern of residual or circulating tumor cells metastasizing from the primary tumor site to the secondary tissue target sites. Initiation of metastatic process was evidenced in 60% to 70% of patients by the time of diagnosis or thereafter. Control, blockage and prevention of such metastasis have hence been recognized as the key steps for successful intervention with cancer metastasis. Currently, therapy for metastatic disease still encounters great challenges.
Myeloid-derived suppressor cells (MDSCs) are main immunosuppressive cells that have been shown to negatively regulate immune responses against cancers. MDSCs are shown to be largely responsible for inhibiting host antitumor immunities and consequently impairing the effectiveness of anticancer immunosuppressive therapeutic approaches, MDSCs are a heterogeneous population of cells that consists of myeloid progenitor cells and immature myeloid cells (IMCs) present during tumor progression, tissue inflammation and pathogen infection. Two different subtypes of MDSCs, namely monocytic MDSCs and granulocytic MDSCs (mMDSCs and gMDSCs, respectively), have been identified based on their morphology, biomarkers and functions. Various MDSCs have therefore been recognized to play a hierarchical role in tumor-induced immunosuppression activity. As a result, the strategy of preventing or blocking the development of MDSCs in cancer patients is being considered as a prime approach for cancerous diseases.
In one aspect, the invention relates to a pharmacological composition comprising: (i) a therapeutically effective amount of Bidens pilosa extract or more than one polyacetylenic compounds purified or isolated from the B. pilosa extract; and (ii) a pharmaceutically acceptable carrier, for use in suppressing, blocking and/or preventing tumor metastasis in a subject in need thereof.
Alternatively, the invention relates to use of the aforementioned pharmacological composition in the manufacture of a medicament for suppressing, reducing, blocking and/or preventing tumor metastasis in a subject in need thereof.
The invention also relates to a method for suppressing, blocking and/or preventing tumor metastasis in a subject in need thereof, comprising administering to the subject in need thereof the aforementioned pharmacological composition.
In another aspect, the invention relates to a pharmacological composition comprising: (i) a therapeutically effective amount of Bidens pilosa extract, or more than one polyacetylenic compounds purified or isolated from the B. pilosa extract; and (ii) a pharmaceutically acceptable carrier, for use in inhibiting differentiation, functional activities, and population of granulocytic myeloid-derived suppressor cells (gMDSCs) and/or suppressing metastatic cancer or cancer metastasis in a subject in need thereof.
Alternatively, the invention relates to use of the aforementioned pharmacological composition in the manufacture of a medicament for inhibiting differentiation, functional activities, and population of granulocytic myeloid-derived suppressor ceils (gMDSCs) and/or suppressing metastatic cancer or cancer metastasis in a subject in need thereof.
The invention relates to a method for inhibiting differentiation, functional activities, and population of granulocytic myeloid-derived suppressor cells (gMDSCs) and/or suppressing metastatic cancer or cancer metastasis in a subject in need thereof, comprising: administering to the subject in need thereof the aforementioned pharmacological composition.
In another embodiment of the invention, the pharmacological composition comprises at least 80% or no less than 89% (wt/wt) of compounds 2-β-D-glucopyranosyloxy-1-5(E)-tridecene-7,9,11-triyne, 2-D-glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne, and 3-β-D-glucopyranosyloxy-1-hydroxy-6(E)-tetradecene-8,10,12-triyne.
In another embodiment of the invention, the pharmacological composition comprises: (a) 2-β-D-glucopyranosyloxy-1-hydroxy-5(E)-tridecene-7,9,11-triyne, (b) 2-D-glucopyranosyloxy-1-hydroxytrideca-5,7,9,11 -tetrayne, and (c) 3-β-D-glucopyranosyloxy-1hydroxy-6(E)-tetradecene-8,10,12-triyne at a ratio ranging from 1:1:2 to 1:2:4, or from 1:1:1 to 1:2:4.
In another embodiment of the invention, the subject has breast cancer, or is a post-operative cancer surgery patient, or in need for control, blockage and prevention of cancer metastasis.
In another embodiment of the invention, the pharmaceutical composition inhibits differentiation, functional activities, and population of granulocytic myeloid-derived suppressor ceils (gMDSCs) and suppresses tumor metastasis without causing cytotoxicity or apoptosis to the gMDSCs.
In another embodiment of the invention, the pharmaceutical composition is in a dosage form selected from the group consisting of oral, intravenous, intramuscular, and subcutaneous.
In another embodiment of the invention, the amount of the Bidens pilosa extract or the more than one polyacetylenic compounds purified or isolated from the B. pilosa extract is effective in inhibiting
tumor metastasis into lung, and accumulation of granulocytic MDSCs in lung, peripheral blood and spleen of the subject in need thereof.
In another embodiment of the invention, the Bidens pilosa extract is: (i) an ethanol extract of B. pilosa; or (ii) a first fraction eluted from an HPLC column loaded with a mixture containing the ethanol extract of B. pilosa, or (iii) a repeatedly re-chromatographed fraction of the ethanol extract of B. pilosa.
In another embodiment of the invention, the B. pilosa extract comprises no less than 89% (w/w) of polyacetylenic compounds.
In another embodiment of the invention, the pharmaceutical composition comprises a human equivalent dose of: (a) 10-1000 mg of the ethanol extract of B. pilosa/Kg body weight×(0.025 Kg/human body weight in Kg)0.33, or (b) 0.5-1000 mg of the first fraction/Kg body weight×(0.025 Kg/human body weight in Kg)0.33.
In one embodiment of the invention, the pharmacological composition comprises compounds of formula (I), (II) and (III):
These and other aspects will become apparent from the following description of the preferred embodiment taken in conjunction with the following drawings. The accompanying drawings illustrate one or more embodiments of the invention and, together with the written description, serve to explain the principles of the invention. Wherever possible, the same reference numbers are used throughout the drawings to refer to the same or like elements of an embodiment.
Growing body of evidence suggests now that chemotherapy, performed as a systemic therapy for metastatic cancer, does not benefit to all cancer patients, but impairs host immunity resulting in the promotion of tumor growth and spread. The invention relates to the discovery that oral administration of BP-E or F1 fraction of BP-E significantly suppressed metastasis. The efficacy of F1 fraction in inhibition of metastasis and MDSC accumulation was as good as docetaxel treatment. Moreover, Mice fed F1 fraction showed better general health than docetaxel-treated mice. F1 fraction, unlike docetaxel, did not induce body weight loss or hair loss in our murine mammary tumor resection model.
Comparing the efficacy, drug administration and side effects of F1 fraction and the current clinical drag docetaxel, this invention is based on an unexpected discovery that phytochemicals prepared from B. pilosa (including BP-E and F1 fractions) can suppress differentiation and functions of MDSC and inhibit mammary tumor metastasis. These extracts can be used as anti-cancer agent against MDSC and tumor metastasis of breast cancers.
As used in the description herein and throughout the claims that follow, the meaning of “a”, “an” and “the” includes plural reference unless the context clearly dictates otherwise. Also, as used in the description herein and throughout the claims that follow, the meaning of “in” includes “in” and “on” unless the context clearly dictates otherwise.
The terms used in this specification generally have their ordinary meanings in the art, within the context of the invention, and in the specific context where each term is used. Certain terms that are used to describe the invention are discussed below, or elsewhere in the specification, to provide additional guidance to the practitioner regarding the description, of the invention. For convenience, certain terms may be highlighted, for example using italics and/or quotation marks. The use of highlighting has no influence on the scope and meaning of a term; the scope and meaning of a term is the same, in the same context, whether or not it is highlighted. It will be appreciated that same thing can be said in more than one way. Consequently, alternative language and synonyms may be used for any one or more of the terms discussed herein, nor is any special significance to be placed upon whether or not a term is elaborated or discussed herein. Synonyms for certain terms are provided. A recital of one or more synonyms does not exclude the use of other synonyms. The use of examples anywhere in this specification including examples of any terms discussed herein is illustrative only.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. In the case of conflict, the present document, including definitions will control
The term “treating” or “treatment” refers to administration of an effective amount of a therapeutic agent to a subject in need thereof, who has a disease (such as tumor and/or tumor metastasis), or a symptom or predisposition toward such a disease, with the purpose of cure, alleviate, relieve, remedy, ameliorate, or prevent the disease, the symptoms of it, or the predisposition towards it, or reduce incidence of symptoms. Such a subject can be identified by a health care professional based on results from any suitable diagnostic method.
“An effective amount” refers to the amount of an active compound that is required to confer a therapeutic effect on the treated subject. Effective doses will vary, as recognized by those skilled in the art, depending on route of administration, excipient usage, and the possibility of co-usage with other therapeutic treatment.
The terms “ethanol extract of B. pilosa” and “BP-E phytoextract” are interchangeable. An ethanol extract of B. pilosa refers to “the phytochemicals extracted from fresh or dried tissues of whole plant of Bidens pilosa Linn var. radiata (Asteraceae) by using ethanol (e.g., 95% EtOH)”.
The terms “F1 fraction” refers to “BP-E derived F1 phytochemicals”. The “F1 fraction” is a sub-fraction of BP-E phytoextract which was isolated by fractionation with HPLC. For example, using a PR-18 preparative HPLC column (e.g., COSMOSIL™ C18, 4.6 mm×250 mm) and a UV 235 nm detector, and a MeOH/H2O gradient at a flow rate of 0.5 ml/min, the elute fraction was collected at the retention time of 40 min to 46 min.
The “Guidance for Industry and Reviewers Estimating the Safe Starting Dose in Clinical Trials for Therapeutics in Adult Healthy Volunteers” published by the U.S. Department of Health and Human Services Food and Drug Administration discloses “a human equivalent dose” may be obtained by calculations from the following formula:
HED=animal dose in mg/kg×(animal weight in kg/human weight in kg)0.33.
As used herein, when a number or a range is recited, ordinary skill in the art understand it intends to encompass an appropriate, reasonable range for the particular field related to the invention.
By 0.5-1000 mg it meant that all tenth and integer unit amounts within the range are specifically disclosed as part of the invention. Thus, 0.5, 0.6, 0.7 and 1, 2, 3, 4 . . . 999.7, 999.8, 999.9 and 1000 unit amounts are included as embodiments of this invention.
The current study investigated the immune-regulatory and antitumor activities of the ethanol extract of B. pilosa (BP-E) on MDSC expansion and tumor metastasis. The results show that BP-E can effectively suppress the metastasis of 4T1 tumors and increase animal survival in a mouse mammary tumor-resection model. BP-E significantly decreased the tumor-induced splenomegaly and, mechanically, it specifically inhibited the differentiation and functional activities of granulocytic MDSCs and reduced the population of these cells in test mice. Bio-organic chemistry-analysis shows that specific polyacetylenic glycosides from the F1 fraction of BP-E are the major principle phytochemicals responsible for the detected MDSC and anti-metastatic activities. Our findings suggest that specific polyacetylene compounds from B. pilosa be readily and highly purified or F1 fraction and they may have useful application for future development as botanical drug(s).
It was discovered that high level expressions of G-CSF and gMDSC populations were detected with a pattern of different stages of a murine 4T1 mammary carcinoma model in tumor-bearing mice. The ethanol extract of B. pilosa (BP-E) exhibited a strong immunomodulatory capacity that can effectively suppress the G-CSF-induced differentiation of gMDSCs from bone marrow cells ex vivo, and can suppress with high potency 4T1 tumor metastasis in a tumor-resection model. The ethanol extract of B. pilosa (BP-E) can effectively suppress metastasis and increase animal survival in a mouse mammary tumor-resection model BP-E significantly decreased tumor-induced splenomegaly and, mechanically, it specifically inhibited the differentiation and functional activities of granulocytic MDSCs and reduced the population of these cells in test mice.
We further demonstrated that oral delivery of BP-E can suppress tumor metastasis via inhibiting the differentiation and function of gMDSCs in test mice. Bio-organic chemistry analysts showed that a specific group of polyacetylenic glycosides, as the great majority constituents (≧89%) of the F1 fraction of BP-E, apparently act as active phytochemicals responsible for the effect on MDSC activities ex vivo and in vivo, and the resultant anti-metastatic activities in vivo. This indicates that phytochemicals in BP plant extracts or the derived ethanol fraction may have therapeutic or other clinical applications.
Exemplary instruments, apparatus, methods and their related results according to the embodiments of the present invention are given below.
Bidens pilosa Linn. Var radiate (Asteracear) plants were grown in farms of Sanxia district, New Taipei city, Taiwan, in 2013. Air-dried shoot, leaf and root tissues of whole plants, weighting 228.2 g, were imbibed, extracted in 2.28 liters of 95% ethanol (EtOH) at room temperature for three days. This total crude extract was evaporated in vacuum to yield a dried residue (6.3955 g), that then resuspended in methanol (MeOH) and eluted with a water-MeOH mixture of decreasing polarity using a PR-18 preparative HPLC column [COSMOSIL™ C18, 4.6 mm×250 mm] with a flow rate of 0.5 ml/mm and detected at UV 235 nm to give a total of 4 sub-fractions (F1-F4). F1 (eluent of 73.5% MeOH/water from the PR-18 column) was collected at the retention time of 40 min to 46 min and identified as a bioactive fraction. The F1 was also repeatedly separated by the same eluted with 70% to 72% MeOH in water for further using in vitro and in vivo.
F1 was subsequently chromatographed by a RP-18 UPLC column [Acquity UPLC HSS C-18 column 2.1×150 mm, 1.8 um] eluted with 30% to 32% Acetonitrile (ACN) with 0.2% Trifluoroacetic acid (TFA) to give a total of four 2nd sub-fractions, FF. A-FF. D. These 2nd sub-fractions were further separated from Fr.1 (40 mg) by a PR-18 preparative HPLC column [COSMOSIL™ CIS, 10 mm×250 mm] eluted with 31.2% ACN with 0.05% TFA to afford compound A (FF. A. 7 mg), compound B (FF. B, 10 mg), and compound C (FF. C+D, 18.79 mg). Those structures, 2-β-D-glucopyranosyloxy-1-hydroxy-5(E)-tridecene-7,9,11-triyne (A), 2-D-glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne (B) and 3-β-D-glucopyranosyloxy-1-hydroxy-6(E)-tetradecene-8,10,12-triyne (C), were compared and confirmed by their NMR and MS/MS data.
4T1-luc2 cells (5×105 cells/100 μl PBS) were orthotopically implanted into mammary fat-pad of BALB/c mice. Primary tumor growth was evaluated by measuring tumor weight and by monitoring bioluminescent imaging of mammary tumors (BL1) every 7 days. For tumor resection mouse model, 4T1-luc2 cells (5×105 cells/100 μl PBS) were orthotopically implanted into mammary fat pad of test mice. At day 21 post tumor implantation, the tumor mass was gently surgically removed. Bioluminescent imaging of metastatic tumor was monitored by using Non-invasive in vivo imaging system (IVIS). The body weight of the test mouse was approximately 25 g.
Construction of 4T1-luc2 Cells
293T cells were transfected with pMD.G, pCMV ΔR8.91, and pIF4g.As2.luc.bla to construct lentivirus with luc2 gene. After 24 hour, cell medium were collected and added to transfect 4T1 cells with constructed virus. 10 μg/mL blasticidin S was applied to select the single clone of 4T1-luc2 cells. 4T1-luc2 cells were cultured and maintained in RPMI-1640 supplemented with 10 μg/mL Blasticidin S, 10% fetal bovine serum, 1 mM Penicillin/Streptomycin, and 1 mM sodium pyruvate at 37° C. in 5% CO2 and 95% humidity.
Lung tissues of test mice were harvested and minced with 150 U/mL Type I Collagenase in tissue grinders for 20 to 50 times. After digestion and lysed with ACK buffer, grinded tissues were collected and filtered through 40 μm cell strainer. Spleen tissues were minced gently with PBS in tissue grinders. After lysed with ACK buffer, cells were collected for further analysis. Blood were lysed with ACK buffer for 3 times and harvested for further analysis. All cells were collected and stained with anti-CD11b, and anti-Ly6G/Ly-6C for flow cytometry analysis.
gMDSCs Isolation
To purify Ly-6G+ MDSCs, splenocytes of tumor-bearing mice were harvested and depleted erythrocytes by ACK buffer. Then, splenocytes were incubated with anti-Ly-6G-biotin Abs for 20 minutes and followed by positive selection using anti-biotin microbeads, following the manufacturer's instructions (MiltenyiBiotec).
BALB/c mice bone marrow cells from femorae and tibiae were depleted of RBCs with ACK lysis buffer and cultured in RPMI 1640 medium supplemented with 20 ng/ml GM-CSF, 10% fetal bovine serum, 50 μM 2-mercaptoethanol, 100 unit/ml penicillin and 100 μg/ml streptomycin in a humidified 5% CO2 incubator at 37° C.
Cells lysates were prepared by using M-PER Mammalian Protein Extraction Reagent [5 mM bicine buffer, 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF 0.3 mM), leupeptin (10 μg/ml) and aprotonin (2 μg/ml)]. Lysates were run on 5% to 20% gradient polvacrylamide-sodium dodecyl sulphate (SDS) gels (20 μg protein per lane), proteins transferred onto Hybond-ECL membranes (GE-Healthcare; Amersham, UK) and immunoblotted with anti-G-CSFR antibody, anti-stat3 antibody, and anti-phosphorylated stat3 antibody. Protein bands were detected by enhanced chemiluminescence (Clarity Western ECL Substrate, BioRad) and developed by autoradiography.
Serum from test mice and conditional medium were collected and stored at −80° C. until assayed. Samples were checked for expression levels of G-CSF (R&D Systems) and quantified at a wavelength of 450 nm using a BiotekPowerWave HT spectrophotometer.
Anti-stat3 antibody and anti-phosphorylated stat3 antibody were purchased from Cell Signaling Technology. Anti-G-CSFR antibody was purchased from Abcam.
Data are presented in fold changes or in percentages with mean±s.e.m. indicated in figure legends. All statistical analyses were determined using GraphPad Software. As comparison between multiple data sets, a one-way ANOVA analysis with Tukey-Kramer method was performed.
MDSCs have been shown to expand in cell population in cancer patients. Granulocyte colony-stimulating factor (G-CSF) was shown as one of the key cytokines secreted by tumor cells that mediate MDSC production. To characterize the dynamic change of MDSC population and G-CSF expression in 4T1 tumor-bearing mice, transgenic 4T1luc2 cells were orthotopically implanted into mammary fat pad of test mice. The representative bioluminescent imaging on growth of orthotopic 4T1-luc2 tumor was recorded weekly (
Correlation Between Expression Levels of gMDSCs, G-CSF and the Rate of Tumor Growth and Metastasis
Expression levels of gMDSCs and G-CSF were previously shown to be closely associated with the progression of tumor growth in mouse models. To investigate the role of gMDSCs and G-CSF in growth and metastasis of mouse mammary tumors, we orthotopically implanted 4T1-luc2 cells into mammary fat pad of test mice. At day 21 post tumor implantation, the primary tumor mass was gently removed surgically. The level of tumor bioluminescence intensity and expression of G-CSF were weekly measured (
Effect of the Ethanol-extracted Fraction of Bidens pilosa (BP-E) on the Functional and Differentiation Activities of MDSCs and on G-CSF Expression
To develop therapeutic agents against tumor metastasis, a number of phyto-extracts or the derived phytochemicals were evaluated for their inhibitory effects on the function and differentiation of MDSCs. It was found that an ethanol partitioned fraction of the Bidens pilosa (BP-E) plant extract significantly suppressed the G-CSF-induced differentiation of gMDSCs from bone marrow cells ex vivo (
Effect of Ethanol-fractionated Phytochemicals from Bidens pilosa (BP-E) on Tumor Metastasis
To evaluate a potential inhibitory effect of oral feeding of BP-E on tumor growth, 4T1-luc2 mouse mammary carcinoma cells were orthotopically implanted into the mammary fat pad of test mice, and subsequently examined in a tumor-resection model. At 7 days post tumor implantation, test mice were divided randomly into BP-E untreated group and treated groups (supplemented via force feeding, an oral dose of 100 mg BP-E/kg body weight/day).
It is important to note that in only 7 days, a very short period of time, BP-E feeding was able to effectively suppress tumor metastasis in the tumor resection model. It is also important to point out that the current tumor-resection model was designed to mimic the present human breast cancer patients for treatment following surgery. At 80 days post tumor implantation, the metastasis rate and death rate of the control group mice had reached the level of 100%. The results again strongly suggested that the early onset of the anti-metastatic effect can be successfully maintained tor a prolonged period of time. In contrast, the metastasis rate and the death rate of BP-E treated mice were maintained at 25% and 12.5%, respectively (
For subsequent experiment, mice were sacrificed on day 42 post tumor implantation, based on the differentials seen in
The myeloid derived suppressor cell (MDSC) populations in spleen tissue of each test group were investigated. Growth of 4T1 tumor strongly induced an accumulation of granulocytic MDSCs in spleen and BP-E effectively reduced (with 50% inhibition) the tumor-induced accumulation of gMDSCs in spleen. In addition, 4T1 tumor cells also slightly increased the monocytic MDSC population in spleen, and BP-E treatment inhibited such an effect in spleen, which indicated that BP-E not only can effectively suppress gMDSC production, but also can inhibit the mMDSC production.
Effect of F1 Fraction of BP-E (BP-E-F1) on ROS Expression in MDSCs and on Differentiation of MDSCs from Bone Marrow Cells
To identify active candidate components or phytochemicals from the BP-E phytoextracts that can confer anti-metastasis activity, BP-E was further fractionationed into 4 sub-fractions (F1 to F4) by using a HPLC analysis with an absorbance of UV at 235 nm (
Bio-organic chemical profiling of the F1 fraction phytochemicals was performed by using UPLC, HPLC, NMR and MS/MS assays. F1 fraction was initially chromatographed using a RP-18 UPLC column, and three major compounds (A-C) were isolated (
Since we were able to separate the phytochemicals of the BP-E extracts into four major fractions, we investigated possible inhibitory effect of the F1 fraction of BP-E, namely BP-E-F1, on tumor growth in a orthotopic mammary tumor growth/tumor resection mouse model. At 7 days post tumor implantation, test mice were randomly divided into untreated and BP-E-F1 groups (i.e., orally treated with 5 mg BP-E-F1/kg body weight/day).
We investigated the effect of BP-E-F1 on tumor metastasis in the tumor resection model. At 21 days post implantation, the tumor mass was surgically removed gently. Following surgery, each treatment group was randomly divided into control, F1 and the docetaxel groups (i.e., via iv injection with 10 mg docetaxel/kg every other 3 days).
Mice were sacrificed at 23 days post tumor resection, lung, liver and spleen of test mice were excised and tumor metastasis measured by bioluminescent imaging.
The results suggest that BP-E and its F1 fraction can effectively suppress tumor metastasis via inhibition of differentiation of MDSCs from bone marrow cells and accumulation of MDSCs in the tumor microenvironment. For subsequent experiment, we injected 4T1 cells or co-injected them with granulocytic MDSCs into the mammary fat pad of test mice. At 7 days post tumor implantation, mice were orally fed F1 (5 mg/kg) every day. At 18 days post tumor implantation, the tumor masses of test mice were gently removed surgically and measured.
In summary, we established a murine mammary 4T1-luc2 orthotopic, tumor resection, and subsequent tumor metastasis mouse model. We systemically investigated the roles of MDSCs in tumor growth and metastasis. The findings provide an immunotherapeutic strategy against metastatic cancer that involved high level activities of MDSC differentiation. Granulocytic MDSCs (gMDSCs) are the major MDSC population accumulated in the peripheral blood and spleen tissue of 4T1 tumor-bearing mice, present from early period to later stage of tumor growth (
Surgery and radiation therapy are current standard treatments tor various cancers, often effective for control at the original tumor of primary tumors site. However, therapies or treatments for metastatic diseases remain to encounter great challenges. Growing body of evidence suggests that chemotherapy, performed as a systemic therapy for metastatic cancers, most often were not able to benefit to cancer patients, instead it often impairs the host immunities, resulting in promotion of tumor growth and spread. We demonstrated that oral administration of the BP-E phytoextract and derived F1 phytochemicals can significantly suppress 4T1 mammary metastasis.
The efficacy of F1 fraction for inhibition of metastasis and MDSC accumulation is at a level just as good as the treatment with docetaxel (
For pharmacological application, the fraction of the administered dose of a test drug that reaches the systemic level in blood circulation is described as bioavailability. We first determined the absolute bioavailability of the three major polyacetylenic glycoside compounds (A, B, C) of BP-E-F1 fraction in blood of test mice. Oral administration was used to investigate the effect of F1 fraction on suppression of 4T1 metastasis. Bioavailability of the three F1 compounds (A-C) was assessed in BALB/c mice (n=12) via administration of F1 fraction by intravenous (iv) or oral delivery, both at 10 mg/kg. The area under curve (AUC) for oral administration and iv administration were experimentally obtained at 282.8 and 1268 mg·min/1, respectively (
F1 fraction significantly suppresses the activity in differentiation of MDSCs from bone marrow cells and the functionality of MDSCs, in vitro and in vivo. Tumor-derived G-CSF has been demonstrated to play a key role in promotion of gMDSC development. To investigate the mechanistic role of F1 fraction in inhibition of gMDSC differentiation, we also adopted an approach for the intravenous administration of recombinant G-CSF, aiming to elicit gMDSC activities. Intravenous administration of recombinant G-CSF significantly promotes the percentage of granulocytes in the peripheral blood of test mice, from 16.1% (level in untreated mice) to 49.1% (
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US15/64841 | 12/9/2015 | WO | 00 |
| Number | Date | Country | |
|---|---|---|---|
| 62091474 | Dec 2014 | US |
