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Biomedicine & Pharmacotherapy =... Oct 2019Multidrug resistance protein 1 (MRP1/ABCC1) actively transports a variety of drugs, toxic molecules and important physiological substrates across the plasma membrane. It...
Multidrug resistance protein 1 (MRP1/ABCC1) actively transports a variety of drugs, toxic molecules and important physiological substrates across the plasma membrane. It can confer broad-spectrum multidrug resistance and can decrease the bioavailability of many important drugs. Substrates of MRP1 include anti-cancer agents, antibiotics, antivirals, antidepressants and anti-inflammatory drugs. Using calcein as a fluorescent reporter in a high content uptake assay, we recently reported the identification of 12 MRP1 inhibitors after screening an anti-cancer library of 386 compounds. Here, we describe the development of a new high content imaging-based uptake assay using doxorubicin as a fluorescent reporter. Screening the same anti-cancer library of 386 compounds, the new assay identified a total of 28 MRP1 inhibitors including 16 inhibitors that have not been previously reported as inhibitors of MRP1. Inhibition of MRP1 activity was confirmed using flow cytometry and confocal microscopy-based transport assays. Six drugs (afatinib, celecoxib, doramapimod, mifepristone, MK-2206 and rosiglitazone) were evaluated for their ability to reverse resistance of MRP1-overexpressing H69AR lung cancer cells against vincristine, doxorubicin and etoposide. Mifepristone and doramapimod were most effective in reversal of resistance against vincristine while mifepristone and rosiglitazone were most successful in resensitizing H69AR cells against doxorubicin. Furthermore, resistance towards etoposide was completely reversed in the presence of celecoxib or doramapimod. Selected drugs were also evaluated for resistance reversal in HEK cells that overexpress P-glycoprotein or breast cancer resistance protein. Our results indicate mifepristone and doramapimod as pan inhibitors of these three drug transporters while celecoxib exhibited selective MRP1 inhibition. Together, our findings signify the importance of MRP1 in drug discovery and demonstrate the effectiveness and value of doxorubicin-based high content screening approach. Anti-cancer agents that exhibit MRP1 inhibition may be used to reverse multidrug resistance or to improve the efficacy and reduce the toxicity of various cancer chemotherapies. On the other hand, anti-cancer drugs that did not interact with MRP1 carry a low risk for developing MRP1-mediated resistance.
Topics: ATP Binding Cassette Transporter, Subfamily B, Member 1; Antineoplastic Agents; Biological Assay; Cell Line, Tumor; Doxorubicin; Drug Resistance, Neoplasm; Fluoresceins; Fluorescent Dyes; HEK293 Cells; High-Throughput Screening Assays; Humans; Multidrug Resistance-Associated Proteins; Protein Transport; Reproducibility of Results
PubMed: 31401398
DOI: 10.1016/j.biopha.2019.109289 -
Cancer Apr 2019
Topics: Anthracyclines; Antibodies, Monoclonal, Humanized; Antineoplastic Agents, Immunological; Antineoplastic Combined Chemotherapy Protocols; Benzodiazepinones; Carboplatin; Clinical Decision-Making; Disease Progression; Etoposide; Humans; Immunoconjugates; Irinotecan; Liver Neoplasms; Lung Neoplasms; Molecular Targeted Therapy; Neoplasm Recurrence, Local; Nivolumab; Paclitaxel; Piperazines; Pyrimidines; Salvage Therapy; Small Cell Lung Carcinoma; Temozolomide; Thiourea; Topotecan
PubMed: 30561759
DOI: 10.1002/cncr.31849 -
Oncotarget Oct 2017Amuvatinib (MP-470) is a multi-targeted kinase inhibitor with potent activity against c-Kit, synergistic with DNA-damaging agents. We evaluated amuvatinib in combination...
BACKGROUND
Amuvatinib (MP-470) is a multi-targeted kinase inhibitor with potent activity against c-Kit, synergistic with DNA-damaging agents. We evaluated amuvatinib in combination with platinum-etoposide (EP) chemotherapy by objective response rate, survival, and tolerability in platinum-refractory small cell lung cancer (SCLC) patients.
METHODS
This study used a Simon 2-stage design requiring ≥3 centrally confirmed responses in the first 21 subjects. Subjects received EP with 300 mg amuvatinib orally three times daily in cycles of 21 days. A three-day amuvatinib run-in period before EP occurred in Cycle 1. Subjects received the same EP chemotherapy regimen given prior to progression/relapse.
RESULTS
Among 23 subjects treated, we observed four PRs (17.4%) per RECIST 1.1, only two of which were centrally confirmed (8.7%, response duration 119, 151 days). Three subjects (13%) had confirmed stable disease. c-Kit H-score was ≥100 in two subjects whose respective durations of disease control were 151 and 256 days.
CONCLUSIONS
The addition of amuvatinib to EP chemotherapy in unselected, platinum-refractory SCLC did not meet the primary endpoint of ≥3 confirmed responses in stage 1. However, high c-Kit expression in two subjects with durable disease control suggests the potential for further study of amuvatinib in SCLC patients with high c-Kit expression.
PubMed: 29113403
DOI: 10.18632/oncotarget.19888 -
Oncotarget Jun 2017Resistance to docetaxel is a major clinical problem in advanced prostate cancer. The overexpression of AXL receptor tyrosine kinase (AXL) has been correlated with...
Resistance to docetaxel is a major clinical problem in advanced prostate cancer. The overexpression of AXL receptor tyrosine kinase (AXL) has been correlated with chemotherapeutic drug resistance. However, the role of AXL expression in docetaxel resistance in prostate cancer is yet unclear. In this study, we demonstrate that AXL is overexpressed and activated independent of Gas6 in docetaxel-resistant prostate cancer cells (PC3-DR and DU145-DR). Moreover, we show that forced overexpression of AXL in PC3 and DU145 cells is sufficient to induce resistance to docetaxel in these cell lines. Notably, genetic or pharmacologic inhibition of AXL in the resistant models suppressed cell proliferation, migration, invasion, and tumor growth, and these effects were significantly augmented when AXL inhibition was combined with docetaxel treatment. Mechanistically, we found that AXL inhibition led to reversion of the epithelial-mesenchymal transition (EMT) phenotype and decreased the expression of ATP-binding cassette B1 (ABCB1). Overall, our results identify AXL as an important mediator of docetaxel resistance in prostate cancer. We propose that AXL-targeted therapy, in combination with docetaxel, has the potential to improve the response to docetaxel therapy and reduce resistance induced by prolonged docetaxel therapy in prostate cancer.
Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Benzocycloheptenes; Cell Line, Tumor; Docetaxel; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Humans; Male; Mice, Nude; Piperazines; Prostatic Neoplasms; Protein Kinase Inhibitors; Proto-Oncogene Proteins; Pyrimidines; RNA Interference; Receptor Protein-Tyrosine Kinases; Taxoids; Thiourea; Triazoles; Xenograft Model Antitumor Assays; Axl Receptor Tyrosine Kinase
PubMed: 28455956
DOI: 10.18632/oncotarget.17026 -
Journal of Hematology & Oncology Oct 2016
PubMed: 27737688
DOI: 10.1186/s13045-016-0335-5 -
Oncotarget Feb 2015c-Kit/α-PDGFR targeted therapies are effective for gastrointestinal stromal tumors (GIST), but, >50% develop drug resistance.
BACKGROUND
c-Kit/α-PDGFR targeted therapies are effective for gastrointestinal stromal tumors (GIST), but, >50% develop drug resistance.
METHODS
RTK expression (c-Kit, c-Met, AXL, HER-1, HER-2, IGF-1R) in pre-/post-imatinib (IM) GIST patient samples (n=16) and 4 GIST cell lines were examined for RTK inhibitor activity. GIST-882 cells were cultured in IM every other day, cells collected (1 week to 6 months) and analyzed by qRT-PCR and Western blotting.
RESULTS
Immunohistochemistry pre-/post-IM demonstrated continued expression of c-Kit and HER1, while a subset expressed IGF-1R, c-Met and AXL. In GIST cells (GIST-882, GIST430/654, GIST48) c-Kit, HER1 and c-Met are co-expressed. Acute IM over-express c-Kit while chronic IM, lose c-Kit and HER-1 in GIST882 cells. GIST882 and GIST430/654 cells have an IC50 0.077 and 0.59 µM to IM respectively. GIST48 have an IC50 0.66 µM to IM, 0.91 µM to amuvatinib [AMU] and 0.67 µM to erlotinib (Erl). Synergistic combinations: GIST882, AMU + Erl (CI 0.20); IM + AMU (CI 0.50), GIST430/654, IM + afatinib (CI 0.39); IM + AMU (CI 0.42), GIST48, IM + afatinib (CI 0.03); IM + AMU (CI 0.04); AMU + afatinib (CI 0.36); IM + Erl (CI 0.63).
CONCLUSION
Targeting c-Kit plus HER1 or AXL/c-Met abrogates IM resistance in GIST.
Topics: Afatinib; Aged; Aged, 80 and over; Cell Line, Tumor; Drug Resistance, Neoplasm; Drug Synergism; ErbB Receptors; Erlotinib Hydrochloride; Female; Gastrointestinal Stromal Tumors; Gene Expression Regulation, Neoplastic; Humans; Imatinib Mesylate; Immunoblotting; Immunohistochemistry; Male; Middle Aged; Molecular Targeted Therapy; Piperazines; Protein Kinase Inhibitors; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-kit; Proto-Oncogene Proteins c-met; Pyrimidines; Quinazolines; Receptor Protein-Tyrosine Kinases; Reverse Transcriptase Polymerase Chain Reaction; Thiourea; Axl Receptor Tyrosine Kinase
PubMed: 25557174
DOI: 10.18632/oncotarget.3021 -
Melanoma Research Oct 2014Effective targeted therapy strategies are still lacking for the 15-20% of melanoma patients whose melanomas are driven by oncogenic NRAS. Here, we report on the...
Effective targeted therapy strategies are still lacking for the 15-20% of melanoma patients whose melanomas are driven by oncogenic NRAS. Here, we report on the NRAS-specific behavior of amuvatinib, a kinase inhibitor with activity against c-KIT, Axl, PDGFRα, and Rad51. An analysis of BRAF-mutant and NRAS-mutant melanoma cell lines showed the NRAS-mutant cohort to be enriched for targets of amuvatinib, including Axl, c-KIT, and the Axl ligand Gas6. Increasing concentrations of amuvatinib selectively inhibited the growth of NRAS-mutant, but not BRAF-mutant melanoma cell lines, an effect associated with induction of S-phase and G2/M-phase cell cycle arrest and induction of apoptosis. Mechanistically, amuvatinib was noted to either inhibit Axl, AKT, and MAPK signaling or Axl and AKT signaling and to induce a DNA damage response. In three-dimensional cell culture experiments, amuvatinib was cytotoxic against NRAS-mutant melanoma cell lines. Thus, we show for the first time that amuvatinib has proapoptotic activity against melanoma cell lines, with selectivity observed for those harboring oncogenic NRAS.
Topics: Apoptosis; Cell Culture Techniques; Cell Cycle; Cell Line, Tumor; Cohort Studies; DNA Damage; Genes, ras; Humans; Inhibitory Concentration 50; Intercellular Signaling Peptides and Proteins; Ligands; Melanoma; Mutation; Piperazines; Proteomics; Proto-Oncogene Proteins; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-kit; Pyrimidines; Receptor Protein-Tyrosine Kinases; Signal Transduction; Skin Neoplasms; Thiourea; Axl Receptor Tyrosine Kinase
PubMed: 24950457
DOI: 10.1097/CMR.0000000000000103 -
Journal of Hematology & Oncology Dec 2013MET is a receptor tyrosine kinase that is activated by the ligand HGF and this pathway promotes cell survival, migration, and motility. In accordance with its oncogenic...
BACKGROUND
MET is a receptor tyrosine kinase that is activated by the ligand HGF and this pathway promotes cell survival, migration, and motility. In accordance with its oncogenic role, MET is constitutively active, mutated, or over-expressed in many cancers. Corollary to its impact, inhibition of MET kinase activity causes reduction of the downstream signaling and demise of cells. In myeloma, a B-cell plasma malignancy, MET is neither mutated nor over-expressed, however, HGF is increased in plasma or serum obtained from myeloma patients and this was associated with poor prognosis. The small-molecule, amuvatinib, inhibits MET receptor tyrosine kinase. Based on this background, we hypothesized that targeting the HGF/MET signaling pathway is a rational approach to myeloma therapy and that myeloma cells would be sensitive to amuvatinib.
METHODS
Expression of MET and HGF mRNAs in normal versus malignant plasma cells was compared during disease progression. Cell death and growth as well as MET signaling pathway were assessed in amuvatinib treated primary myeloma cells and cell lines.
RESULTS
There was a progressive increase in the transcript levels of HGF (but not MET) from normal plasma cells to refractory malignant plasma cells. Amuvatinib readily inhibited MET phosphorylation in primary CD138+ cells from myeloma patients and in concordance, increased cell death. A 48-hr amuvatinib treatment in high HGF-expressing myeloma cell line, U266, resulted in growth inhibition. Levels of cytotoxicity were time-dependent; at 24, 48, and 72 h, amuvatinib (25 μM) resulted in 28%, 40%, and 55% cell death. Consistent with these data, there was an amuvatinib-mediated decrease in MET phosphorylation in the cell line. Amuvatinib at concentrations of 5, 10, or 25 μM readily inhibited HGF-dependent MET, AKT, ERK and GSK-3-beta phosphorylation. MET-mediated effects were not observed in myeloma cell line that has low MET and/or HGF expression.
CONCLUSIONS
These data suggest that at the cellular level MET/HGF pathway inclines with myeloma disease progression. Amuvatinib, a small molecule MET kinase inhibitor, is effective in inducing growth inhibition and cell death in myeloma cell lines as well as primary malignant plasma cells. These cytostatic and cytotoxic effects were associated with an impact on MET/HGF pathway.
Topics: Aged; Apoptosis; Cell Line, Tumor; Disease Progression; Female; Humans; Male; Middle Aged; Multiple Myeloma; Piperazines; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-met; Pyrimidines; Signal Transduction; Thiourea
PubMed: 24326130
DOI: 10.1186/1756-8722-6-92 -
Oncogene Mar 2014Despite significant progress in the treatment of breast cancer, particularly through the use of targeted therapy, relapse and chemoresistance remain a major hindrance to...
Despite significant progress in the treatment of breast cancer, particularly through the use of targeted therapy, relapse and chemoresistance remain a major hindrance to the fight to minimize the burden of the disease. It is becoming increasingly clear that a rare subpopulation of cells known as cancer stem cells (CSC), able to be generated through epithelial-to-mesenchymal transition (EMT) and capable of tumor initiation and self-renewal, contributes to treatment resistance and metastases. This means that a more effective therapy should target both the chemoresistant CSCs and the proliferating epithelial cells that give rise to them to reverse EMT and to attenuate their conversion to CSCs. Here, we demonstrate a novel function of AXL in acting upstream to induce EMT in normal and immortalized human mammary epithelial cells in an apparent positive feedback loop mechanism and regulate breast CSC (BCSC) self-renewal and chemoresistance. Downregulation of AXL using MP470 (Amuvatinib) reversed EMT in mesenchymal normal human mammary epithelial cells and murine BCSCs attenuating self-renewal and restored chemosensitivity of the BCSCs. AXL expression was also found to be associated with the expression of stem cell genes, regulation of metastases genes, increased tumorigenicity and was important for BCSC invasion and migration. Inactivation of AXL also led to the downregulation of nuclear factor-κB pathway and reduced tumor formation in vivo. Taken together, our data suggest that targeted therapy against AXL, in combination with systemic therapies, has the potential to improve response to anticancer therapies and to reduce breast cancer recurrence and metastases.
Topics: Animals; Antineoplastic Agents; Breast Neoplasms; Carcinogenesis; Cell Line, Tumor; Cell Movement; Cell Proliferation; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Female; Humans; Mice; Mice, Transgenic; Molecular Targeted Therapy; NF-kappa B; Neoplasm Transplantation; Neoplastic Stem Cells; Piperazines; Proto-Oncogene Proteins; Pyrimidines; Receptor Protein-Tyrosine Kinases; Signal Transduction; Thiourea; Tumor Burden; Up-Regulation; Axl Receptor Tyrosine Kinase
PubMed: 23474758
DOI: 10.1038/onc.2013.57 -
Translational Lung Cancer Research Dec 2012MET and its ligand hepatocyte growth factor/scatter factor (HGF) influence cell motility and lead to tumor growth, invasion, and angiogenesis. Alterations in MET have... (Review)
Review
MET and its ligand hepatocyte growth factor/scatter factor (HGF) influence cell motility and lead to tumor growth, invasion, and angiogenesis. Alterations in MET have been observed in non-small cell lung cancer (NSCLC) tumors, with increased expression associated with more aggressive cancer, as well as acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI). MET inhibitors act via two basic mechanisms. Small molecule inhibitors antagonize ATP in the intracellular tyrosine kinase domain of MET, with studies on the following agents reviewed here: tivantinib (ARQ-197), cabozantinib (XL-184), crizotinib (PF-02341066), amuvatinib (MP470), MGCD265, foretinib (EXEL-2880), MK2461, SGX523, PHA665752, JNJ-38877605, SU11274, and K252A. The monoclonal monovalent antibody fragment onartuzumab (MetMAb) is also discussed here, which binds to and prevents the extracellular activation of the receptor by ligand. MET inhibition may both overcome the negative prognostic effect of MET tumor expression as well as antagonize MET-dependent acquired resistance to EGFR inhibitors. Here we discuss MET inhibitors in combination with other therapies in lung cancer.
PubMed: 25806189
DOI: 10.3978/j.issn.2218-6751.2012.10.08