Significant reductions in GYN cancer patient mortality and m
Significant reductions in GYN cancer patient mortality and morbidity rates require treatments that proactively prevent and reverse resistance to chemotherapy and radiation. Combining chemotherapy and radiation with inhibitors of key DDR proteins targets is likely to enhance the ability of genotoxic treatments to kill a variety of cancer gpr40 agonist , , , , , , , , , . Previous studies have demonstrated the benefits of targeting DDR pathways to enhance response of a broad range of genotoxic therapies, including chemotherapy and radiation , , , , , , , , , .
Given the pivotal roles that platinum agents and radiation play in the treatment of GYN cancers, the success of targeting DDR pathways to enhance genotoxin response, including in ovarian and cervical cancers , the prevalence of defects in homologous recombination in ovarian and endometrial cancers , , , the strong link between human papilloma virus and DDR pathways , , and the availability of more selective inhibitors of ATR and ATM, and that ATR kinase inhibitors have been shown to sensitize ATM- and p53-deficient cells to cisplatin , we sought to evaluate the impact of selectively inhibiting ATR and ATM in a panel of sensitive and resistant ovarian cancer cells as well as endometrial and cervical cancer cells with wild type or mutant p53 treated with cisplatin, carboplatin, or IR. We hypothesized that targeting these DDR pathways in GYN cancer cells would synergize with platinum-based chemotherapy and radiation. For these studies, we utilized small molecule pharmacologic kinase inhibitors of ATR (ETP-46464, “ATRi”)  and ATM (KU55933, “ATMi”)  and show enhancements in the response of IR in human cervical, endometrial and ovarian carcinoma in vitro, independent of p53 status. Further, our data show that selective inhibition of ATR, but not ATM, synergizes with platinum in cell line models in all three disease sites, independent of p53 status. This divergent response is of central importance as it informs the selective use of ATR inhibitors, but not ATM inhibitors, with cisplatin- or carboplatin-based chemotherapy, and directs the combined use of ATR and ATM inhibitors with radiation or cisplatin-based chemoradiation in newly diagnosed and recurrent ovarian, endometrial and cervical cancer.
Discussion Here we show that pharmacological inhibition of ATR, but not ATM, sensitizes ovarian, endometrial and cervical cancer cells to cisplatin and carboplatin, and that the combination of either platinum agent with the ATRi was synergistic. We present data utilizing a syngeneic model of platinum-resistant ovarian cancer, A2780 (platinum-sensitive) versus A2780-CP20 (platinum-resistant), showing that inhibition of ATR produces marked enhancement in response when combined with cisplatin or carboplatin. These results demonstrate that ATRi abrogates ATR-dependent phosphorylation of Chk1 induced by platinum-mediated DNA damage. Phosphorylation of Chk1 at Ser345 has been established as a classic indicator of ATR kinase activity . Recent evidence has revealed that treatment with VE-821, another ATR inhibitor, and MK-8776, a Chk1 inhibitor, enhanced cisplatin, gemcitabine, topotecan and veliparib response in ovarian cancer cells . It should be noted that ETP-46464 is an effective inhibitor of other PIKK enzymes, most notably the mammalian target of rapamycin (mTOR) . It has been previously noted that inhibitors of PI3K, mTOR, ATM or DNAPKcs do not generate replicative stress, nor do they enhance hydroxyurea-induced replicative stress . Previously published results have documented the formation of 53BP1 foci in hydroxyurea (HU)-treated cells inhibited with ETP46464, the absence of 53BP1 foci in HU-treated cells inhibited with rapamycin and another mTORi, and that replication fork restart was suppressed by ETP-46464, and not rapamycin . Although the preponderance of the data showing enhanced cisplatin killing of ETP-46464 point to its inhibitory activity on ATR-Chk1 signaling, the known inhibitory activity on mTOR signaling may, in part, contribute to the observed enhanced cisplatin sensitivity. Cellular sensitization to cisplatin was recapitulated with VE-821, another highly selective small molecule pharmacologic inhibitor of ATR (Supplementary Fig. S5).