[Effect and Mechanism of Radiosensitization of Poly (ADP-Ribose) Polymerase Inhibitor n Lewis Cells and Xenografts].

ABSTRACT

BACKGROUND AND OBJECTIVE: The DNA damage of the irradiated tumor cells is mainly single strand breaks (SSBs) and double strand breaks (DSBs), in which the frequency of occurrence of SSBs is dozens of times than DSBs. However, most of the SSBs could be repaired by the Poly (ADP-Ribose) Polymerase (PARP) and other related factors. Recently listed drug-Olaparib (PARP1/PARP2/PARP3 inhibitor) could target the repair pathways of single strand breaks, and recent clinical trials of PARP inhibitors combined with chemotherapy obtained encouraging results. The aim of this study is to investigate the effect and potential mechanism of radiosensitization of Poly (ADP-Ribose) polymerase inhibitor-Olaparib on lewis cells and xenografts. METHODS: The inhibition concentration 10% inhibitory concentration (IC10) of Olaparib to Lewis cells was detected by methyl thiazolyltetrazolium (MTT) assay. The radiosensitization effect of Olaparib on Lewis cells was determined by classical colony forming assay. Lewis xenografts models were established, and the mice were randomly divided into four groups Control group, Olaparib group, Radiotherapy group (RT, 2 Gy × 5 d), Olaparib combined with RT group. Xenograft volume was measured during the treatment. Flow cytometry was used to analyze the apoptosis rate of the Lewis cells in each group, and the apoptosis of xenograft tissues was observed by TUNEL stain. The ralative protein levels of γH2AX (associated with DNA strand breaks repair), Bax/Bcl-2, Caspase-3 (apoptosis-associated protein) were detected by Western blot in vitro and in vivo. RESULTS: The IC10 value of Olaparib was 4.4 µmol/L. The radio-sensitivity enhancement ratio (SER) of Olaparib combined with RT was 1.211 in vitro. Compared with RT (2 Gy × 5 d) alone, the combination of Olaparib with fractionated radiotherapy significantly increased the growth delay of Lewis xenografts (P<0.001). Flow cytometry and TUNEL analysis indicated that the apoptosis rate in the combination group was significantly higher than in RT group in vitro and in vivo (P<0.05). Furthermore, Western blot results confirmed that in the combination group the expression levels of γH2AX, Bax, Caspase-3 were increased, while that of Bcl-2 was decreased as opposed to RT group (P<0.05). CONCLUSION: The combination of Olaprib and fractionated radiotherapy can markedly improve the radiobiological effects on lewis cells and xenografts, which may be induced by promoting the formation of DNA double strand break and upregulating the expression of Bax/Bcl-2 pro-apoptotic proteins.