AXL pathway as a target to overcome resistance to PARP inhibitors in high-grade serous ovarian (HGSOC) and triple-negative breast cancer (TNBC) cell lines.

Abstract

e17502 Background: PARP inhibitors (PARPi) as olaparib (OLA) have become the new standard in maintenance treatment in high grade serous ovarian carcinoma (HGSOC) and triple negative breast cancer (TNBC). Thus there is an important need to identify the mechanisms for PARPi resistance and the strategies to overcome it. Our aim is to intervene in vitro AXL, a receptor tyrosine kinase, linking its function to the OLA-resistance mechanism and assess its suitability as a therapeutical target to overcome PARPi-resistance. Methods: Three cell lines, 2 from HGSOC (PEO1-S and Kuramochi-S) and 1 from TNBC (MDA-MB-231-S) were selected, isogenic resistant pairs generated in vitro by pulsed exposure to OLA (PEO1-R) or by exposure to growing concentrations of OLA (Kuramochi-R and MDA-MB-231-R) were obtained. Resistance was confirm by the comparison of IC50 of OLA in -S vs -R lines. AXL expression was assessed by western blot (WB). Cells were treated with OLA before and after the downregulation of AXL expression by a specific siRNA. Cell viability was assessed by MTT assay and DNA damage by the evaluation of markers by western blot (WB). Results: AXL expression was maintained upon OLA treatment in all 3 resistant cell lines. However, AXL expression was reduced upon OLA exposure in 3 sensible parental cell lines. AXL silencing (Si) lead to a significantly decrease of OLA IC50 in all 3 sensitive and resistant cell lines. For sensitive cell lines, IC50 of OLA was in 1) PEO1S with control (scramble) was 5.79 mM vs 2.24 mM after silencing with siRNA, p < 0.0001; 2) Kuramochi-S scramble OLA IC50 was 481.8 mM vs 199.7 mM with siRNA, p < 0.0001 and 3) MDA-MB-231-S with control of scramble the OLA IC50 was 61.62 mM vs 26.21 mM with siRNA, p = 0.001. For resistant cell lines, 4) PEO1R control with scramble 91.49 mM vs siRNA 45.72 mM, p < 0.0001; 5) Kuramochi-R scramble OLA IC50 579.8 mM vs 219.8 mM with siRNA, p = 0.0008 and 6) 231-R scramble OLA IC50 68.22 mM vs 9.247 mM with siRNA, p < 0.0001. These results suggest that AXL downregulation increases sensitivity to OLA in both sensitive and resistant cell lines. Histone γ-H2AX was analysed pre and post silencing and both increased when treating with OLA, especially when AXL was downregulated in alll three resistant cell lines. Conclusions: This in vitro study suggest a potential role of AXL in OLA resistance and AXL downregulation migh represent a potential mechanism to overcome resistance to OLA in ovarian and TNBC cell lines.