Olaparib mediated DNA damage response and its resistant mechanism in cervical cancer cells

Abstract

Cervical cancer is still the most common gynecologic cancer in developing countries. Olaparib is a poly-(ADR-ribose)-polymerase 1 (PARP1) inhibitor targeting at DNA strand break repair. However its efficacy in cervical cancer is unclear. This study aimed to evaluate the therapeutic potential of olaparib in cervical cancer. Olaparib remarkably reduced the PARylation activity and arrested cervical cancer cells at G2/M phase, and was associated with reduced level of a mitosis marker, phosphor-histone H3 as indicated by propidium iodide DNA staining and immunofluorescence. Cytotoxicity of olaparib against cervical cancer cells was determined by XTT assay with IC50 ranged from 35.49µM in C4I to 87.17µM in SiHa. The difference in olaparib sensitivity between C4I and SiHa was consistently demonstrated in annexin V based apoptotic assay, where a higher level of apoptotic response was found in C4I (21.12%) compared to SiHa (6.37%). Immunofluorescent staining and western blot for gH2AX clearly showed DNA double-strand break accumulation in both cell lines upon PARP1 blockade. Lower expression level of RAD51, a homologous recombination (HR) repair factor, was also observed in C4I, when compared to SiHa. Besides, gH2AX/RAD51 foci co-immunofluorescent line scan image analysis showed that there was reduced level of DNA damage recognition by RAD51 (26.80%), compared to SiHa (65.31%). These findings suggested that there was a reduced HR activity in C4I and might explain the higher sensitivity to olaparib compared to SiHa. We also demonstrated the therapeutic potential of olaparib in cervical cancer, and the rationale of using RAD51 as a predictive biomarker for olaparib sensitivity.