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Inhibition of microvesiculation sensitizes prostate cancer cells to chemotherapy and reduces docetaxel dose required to limit tumor growth in vivo.

ABSTRACT: Microvesicles shed from cells carry constituents of the cell cytoplasm, including, of importance in multidrug resistance to cancer chemotherapy, drugs that the tumor cell attempts to efflux. To see whether such drugs could be used at lower concentrations with the same efficacy, it was first shown that microvesiculation of prostate cancer (PCa) cells, PC3, could be inhibited pharmacologically with calpeptin (calpain inhibitor) and by siRNA (CAPNS1). In cells treated with docetaxel (DTX), this inhibition resulted in a third-fold increase in intracellular concentrations of DTX. As a result, 20-fold lower concentrations of DTX (5?nM) could be used, in the presence of calpeptin (20??M) inducing the same degree of apoptosis after 48?h in PC3 cells, as 100?nM of DTX alone. Inhibition of microvesiculation similarly improved combination chemotherapy (DTX and methotrexate). In a mouse xenograft model of PCa, DTX (0.1?mg/kg) together with calpeptin (10?mg/kg), administered i.p., significantly reduced tumor volumes compared to DTX alone (0.1?mg/kg) and brought about the same reductions in tumor growth as 10?mg/kg of DTX alone. As well as further reducing vascularization, it also increased apoptosis and reduced proliferation of PC3 cells in tumor xenografts.

SUBMITTER: Jorfi S

PROVIDER: S-EPMC4548233 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature

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Inhibition of microvesiculation sensitizes prostate cancer cells to chemotherapy and reduces docetaxel dose required to limit tumor growth in vivo.

Jorfi Samireh S Ansa-Addo Ephraim A EA Kholia Sharad S Stratton Dan D Valley Shaunelle S Lange Sigrun S Inal Jameel J

Scientific reports 20150825

Microvesicles shed from cells carry constituents of the cell cytoplasm, including, of importance in multidrug resistance to cancer chemotherapy, drugs that the tumor cell attempts to efflux. To see whether such drugs could be used at lower concentrations with the same efficacy, it was first shown that microvesiculation of prostate cancer (PCa) cells, PC3, could be inhibited pharmacologically with calpeptin (calpain inhibitor) and by siRNA (CAPNS1). In cells treated with docetaxel (DTX), this inh ...[more]

PMID: 26302712

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Project description:BackgroundDespite the introduction of several novel therapeutic approaches that improved survival, metastatic castration-resistant prostate cancer (mCRPC) remains an incurable disease. Herein we report the synergistic antitumor interaction between two well-known drugs used for years in clinical practice, the antiepileptic agent with histone deacetylase inhibitory activity valproic acid and the cholesterol lowering agent simvastatin, in mCRPC models.MethodsSynergistic anti-tumor effect was assessed on PC3, 22Rv1, DU145, DU145R80, LNCaP prostate cancer cell lines and EPN normal prostate epithelial cells, by calculating combination index (CI), caspase 3/7 activation and colony formation assays as well as on tumor spheroids and microtissues scored with luminescence 3D-cell viability assay. Cancer stem cells (CSC) compartment was studied evaluating specific markers by RT-PCR, western blotting and flow cytometry as well as by limiting dilution assay. Cholesterol content was evaluated by 1H-NMR. Overexpression of wild-type YAP and constitutively active YAP5SA were obtained by lipofectamine-based transfection and evaluated by immunofluorescence, western blotting and RT-PCR. 22Rv1 R_39 docetaxel resistant cells were selected by stepwise exposure to increasing drug concentrations. In vivo experiments were performed on xenograft models of DU145R80, 22Rv1 parental and docetaxel resistant cells, in athymic mice.ResultsWe demonstrated the capacity of the combined approach to target CSC compartment by a novel molecular mechanism based on the inhibition of YAP oncogene via concurrent modulation of mevalonate pathway and AMPK. Because both CSCs and YAP activation have been associated with chemo-resistance, we tested if the combined approach can potentiate docetaxel, a standard of care in mCRCP treatment. Indeed, we demonstrated, both in vitro and in vivo models, the ability of valproic acid/simvastatin combination to sensitize mCRPC cells to docetaxel and to revert docetaxel-resistance, by mevalonate pathway/YAP axis modulation.ConclusionOverall, mCRPC progression and therapeutic resistance driven by CSCs via YAP, can be tackled by the combined repurposing of two generic and safe drugs, an approach that warrants further clinical development in this disease.

Project description:DOT1L, the only known histone H3-lysine 79 (H3K79) methyltransferase, has been shown to be essential for the survival and proliferation of mixed-linkage leukemia (MLL) gene rearranged leukemia cells, which are often resistant to conventional chemotherapeutic agents. To study the functions of DOT1L in MLL-rearranged leukemia, SYC-522, a potent inhibitor of DOT1L developed in our laboratory, was used to treat MLL-rearranged leukemia cell lines and patient samples. SYC-522 significantly inhibited methylation at H3K79, but not H3K4 or H3K27, and decreased the expression of two important leukemia-relevant genes, HOXA9 and MEIS1, by more than 50%. It also significantly reduced the expression of CCND1 and BCL2L1, which are important regulators of cell cycle and anti-apoptotic signaling pathways. Exposure of MLL-rearranged leukemia cells to this compound caused cell cycle arrest and promoted differentiation of those cells, both morphologically and by increased CD14 expression. SYC-522 did not induce apoptosis, even at 10 µM for as long as 6 days. However, treatment with this DOT1L inhibitor decreased the colony formation ability of primary MLL-rearranged AML cells by up to 50%, and promoted monocytic differentiation. Notably, SYC-522 treatment significantly increased the sensitivity of MLL-rearranged leukemia cells to chemotherapeutics, such as mitoxantrone, etoposide and cytarabine. A similar sensitization was seen with primary MLL-rearranged AML cells. SYC-522 did not affect chemotherapy-induced apoptosis in leukemia cells without MLL-rearrangement. Suppression of DOT1L activity inhibited the mitoxantrone-induced increase in the DNA damage response marker, ?H2AX, and increased the level of cPARP, an intracellular marker of apoptosis. These results demonstrated that SYC-522 selectively inhibited DOT1L, and thereby altered gene expression, promoted differentiation, and increased chemosensitivity by preventing DNA damage response. Therefore, inhibition of DOT1L, in combination with DNA damaging chemotherapy, represents a promising approach to improving outcomes for MLL-rearranged leukemia.

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Inhibition of microvesiculation sensitizes prostate cancer cells to chemotherapy and reduces docetaxel dose required to limit tumor growth in vivo.

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Inhibition of microvesiculation sensitizes prostate cancer cells to chemotherapy and reduces docetaxel dose required to limit tumor growth in vivo.

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