Establishment and Molecular Characterization of an In Vitro Model for PARPi-Resistant Ovarian Cancer

SIMPLE SUMMARY: We established an in vitro model for PARPi-resistant ovarian cancer by long-term olaparib exposure of either BRCA1-proficient or BRCA1-deficient cell lines. We describe that PARPi-resistant cells show a broad spectrum of cross-resistance toward other clinically relevant PARPis and chemotherapeutic drugs independently of the BRCA1-status of the parental cell lines. Using co-culture experiments, we studied the clonal dynamics of our in vitro model and showed that PARPi-resistant cells have a proliferative disadvantage over PARPi-sensitive cells under drug-free conditions, while they rapidly gained clonal dominance under selection pressure with olaparib, which can be blocked by pharmacological inhibition of ABC-transporter proteins. Conclusively, we present a well-characterized in vitro model of PARPi resistance that could be useful in dissecting mechanisms of PARPi resistance derived from a BRCA1-proficient or BRCA1-deficient background. Furthermore, our model will allow studying how experimental therapeutic drugs, such as novel PARPi-sensitizers, affect the clonal dynamics of PARPi-resistant cells. ABSTRACT: Overcoming PARPi resistance is a high clinical priority. We established and characterized comparative in vitro models of acquired PARPi resistance, derived from either a BRCA1-proficient or BRCA1-deficient isogenic background by long-term exposure to olaparib. While parental cell lines already exhibited a certain level of intrinsic activity of multidrug resistance (MDR) proteins, resulting PARPi-resistant cells from both models further converted toward MDR. In both models, the PARPi-resistant phenotype was shaped by (i) cross-resistance to other PARPis (ii) impaired susceptibility toward the formation of DNA-platinum adducts upon exposure to cisplatin, which could be reverted by the drug efflux inhibitors verapamil or diphenhydramine, and (iii) reduced PARP-trapping activity. However, the signature and activity of ABC-transporter expression and the cross-resistance spectra to other chemotherapeutic drugs considerably diverged between the BRCA1-proficient vs. BRCA1-deficient models. Using dual-fluorescence co-culture experiments, we observed that PARPi-resistant cells had a competitive disadvantage over PARPi-sensitive cells in a drug-free medium. However, they rapidly gained clonal dominance under olaparib selection pressure, which could be mitigated by the MRP1 inhibitor MK-751. Conclusively, we present a well-characterized in vitro model, which could be instrumental in dissecting mechanisms of PARPi resistance from HR-proficient vs. HR-deficient background and in studying clonal dynamics of PARPi-resistant cells in response to experimental drugs, such as novel olaparib-sensitizers.