Abstract 2152: PJ34-mediated PARP-1 enzymatic activity inhibition in cervical cancer modulates the cellular response to cisplatin

Abstract

Abstract BACKGROUND: Manifestation of chemotherapy (Cisplatin) resistance is an important cause of treatment failure in patients of locally advanced (FIGO stage, IIB to IVA) cervix cancer. We hypothesised that Cisplatin (CDDP) resistance might be associated with PARP-1 hyper-activation and inhibition of PARP-1 enzyme through PJ34 could sensitise cells to cisplatin mediated cytotoxicity. METHODS: We studied combined effect of “PJ34 (PARP-1 inhibitor) and CDDP” in vitro in SiHa and HeLa cell line. Cytotoxicity and cell cycle progression was determined with MTT assay and FACS analysis, respectively. Poly(ADP-ribosyl)ation was assessed by western blotting to determine PARP-1 enzymatic activity inhibition by PJ34. Statistical analysis was done by two-tailed Student t tests using GraphPad software and P-values were considered significant at <0.05. RESULTS: In both cell lines, poly(ADP-ribosyl)ation was inhibited by PJ34 when used at very low concentration (≤ 3μM) and negligible band was observed at concentrations selected for further experiments. In SiHa cells, PJ34 significantly potentiated CDDP mediated cytotoxicity as assessed by MTT assay (p<0.05). Compared to CDDP alone, treatment with combination of PJ34 and CDDP lead to decrease in CDDP IC50 value from 10.8μM to 3.6μM; 7.7μM to 3.2μM; 5.3μM to 2.2μM; 3.9μM to 1.3μM at 48 h, 72 h, 96 h and 120 h treatment, respectively (p <0.01). In HeLa cells, treatment with low concentration of PJ34 (for 24 h) increased survival as well as CDDP IC50 value compared to CDDP alone. However, increase in incubation time decreased both cell survival and CDDP IC50 value. At its higher concentration, PJ34 significantly decreased cell survival (p<0.01), also decrease in CDDP IC50 value was observed from 10μM to 3.4μM; 8.9μM to 3.75μM; 6.1uM to 2μM and 5.2μM to 1.1μM at 24 h, 48 h, 72 h and 96 h treatment respectively (p <0.005). It is reported that CDDP causes cell cycle arrest at late S, early G2/M phase and death at even higher dose, cell cycle analysis revealed that PJ34 increased this late S, early G2/M block and an increased cell death as evaluated by sub G0 peak, in both the cell lines. PJ34, itself had minimum cytotoxic effect at concentrations used in the study; IC50 value was determined to be 24.8μM (24 h, p = 0.0068) for HeLa and 60μM (48 h, p = 0.00067) for SiHa. CONCLUSION: Suppression of PARP-1 enzymatic activity makes cervix cancer cells more sensitive toward CDDP induced toxicity. It seems to be a plausible mechanism for CDDP-resistant cancer cells to develop a dependency to PARP-1. Thus, PARP-1 enzyme inhibition could be explored as a potential therapy for treating CDDP resistance in cancer cervix. Citation Format: Minakshi Mann, S. S. Chauhan, Sunesh Kumar, Neerja Bhatla, Sameer Bakhshi, Ritu Gupta, Sobuhi Iqbal, Lalit Kumar. PJ34-mediated PARP-1 enzymatic activity inhibition in cervical cancer modulates the cellular response to cisplatin. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2152.