All the TNBC cell lines exhibited high levels of Chk1 phosphorylated on Ser296 compared to the ER positive breast cancer cell lines. In the sensitive luminal breast selleck catalog cancer cell line SKBr3, the en dogenous levels of H2AX phosphorylated on Ser139 was much higher compared to all other cell lines. In the ovar ian cell lines, the sensitive A2780 and SKOV 3 as well as the resistant ES 2 cell line exhibited high levels of en dogenous pChk1. Chk1 was expressed in variable amounts across all eleven cell lines examined whilst the levels of Chk1 phosphorylated on the ATM ATR sites Ser317 and 345 was virtually undetectable. The increased levels of Chk1 and H2AX phosphorylation are consistent with underlying defects in DNA repair and or replication.

Analysis of other proteins associated with DNA replication or the DDR response did not identify a consistent mechanism for Chk1 activation. V158411 potentiates cytotoxic chemotherapy in TNBC and ovarian cancer cell lines The ability of V158411 to potentiate the cytotoxicity of a variety of cytotoxic chemotherapeutic drugs was assessed across a panel of luminal breast cancer and TNBC cell lines. V158411 effectively potentiated the growth inhibi tory activity of gemcitabine and cisplatin in the panel of p53 defective but not p53 proficient cell lines. As has been seen with other Chk1 inhibitors, the most robust potentiation was observed with gemcitabine across the range of cell lines. For gemcitabine, not only did V158411 reduce the EC50 of the chemotherapeutic agent but it also increased the fraction of cells killed.

In the ovarian carcinoma cell line SKOV 3, V158411 mod estly potentiated the cytotoxic activity of carboplatin and cisplatin but not oxaliplatin. Western blotting analysis revealed that all three platinum drugs increased the phosphorylation of Chk1 on Ser296 but only Carfilzomib the combination of cisplatin with V158411 robustly induced H2AX phosphorylation on Ser139. As well as exhibiting single agent activity against TNBC and ovarian cancer cell lines, V158411 potentiated the cytotoxicity of chemotherapeutic drugs in these tumor types suggesting that Chk1 inhibitors either alone or in combination could be a viable treatment option in these tumor types. Discussion Multiple Chk1 inhibitors are currently undergoing clinical testing in combination with a variety of cytotoxic chemo therapeutic agents for their ability to potentiate the anti V158411 there may be subsets of cancers for which a Chk1 inhibi tor, administered as a single agent, could be a useful thera peutic option. We postulated that cancers with underlying defects in tumor efficacy of the chemotherapy drugs whilst not in creasing the systemic toxicity of these drugs.