Abstract SY43-01: Translating adaptive therapy to ovarian cancer patients
Authors: Mohammed Ateeb Khan, Helen Hockings, Maximilian Mossner, Ann-Marie Baker, Hall Amy, Robert L. Hollis, Weini Huang, Eszter Lakatos, Charlie Gourley, Trevor A. Graham, Michelle Lockley
Journal: Cancer Research (2025)

Abstract Introduction. Drug resistance results in dismal outcomes for cancer patients. Carboplatin is the backbone of treatment for high grade serous ovarian cancer (HGSOC), augmented by PARP inhibitor (PARPi) maintenance therapy for patients with tumors deficient in homologous recombination repair (HRR). However, most tumors evolve resistance to both drugs and no other agents, including immunotherapies, have improved survival. Therapeutic strategies that acknowledge and target this evolution offer an opportunity to preserve drug sensitivity and extend progression-free survival. Adaptive Therapy (AT) is one such approach. AT is based on the premise that fitness costs are incurred as resistance evolves, so that competition between drug-sensitive and resistant cells favors sensitive cells when drug is absent. We have developed AT in HGSOC from proof-of-concept to a randomized phase 2 trial called ACTOv, (Adaptive ChemoTherapy in Ovarian cancer) comparing carboplatin AT to standard carboplatin dosing in patients with relapsed, platinum sensitive high grade serous and endometrioid ovarian cancer. We are now extending our work to explore the evolutionary dynamics between PARPi-sensitive and PARPi-resistant HGSOC populations. Methods. We evolved resistance to cisplatin and carboplatin in HGSOC cell lines in vitro and in vivo via carboplatin treatment of mice with intraperitoneal xenografts. Resistance to cisplatin, carboplatin and the PARPis niraparib and olaparib was quantified by live/dead assays in the entire cell panel. Co-cultures of sensitive and resistant cells were created in vitro in low resource conditions (low serum or low glucose) to expose fitness deficits. Population growth dynamics were quantified over time using the Sartorius Incucyte™ Live Cell Analysis System. Mechanisms underpinning these dynamics were explored by flow cytometry to analyze cell cycle and expression of the standard apoptotic maker, Annexin V. HRR status was determined functionally by immunofluorescence for Rad51 and γH2Ax co-localization and for micronuclei formation, while BRCA expression was examined by western blot. In vivo co-cultures of GFP-expressing sensitive cells and RFP-expressing resistant cells enabled tracking of sensitive/resistant cell growth over time by qPCR. Sensitive/resistant cells were further examined by immunohistochemistry (IHC) in excised tumors. The influence of drug therapy on in vitro co-cultures of sensitive (GFP) and resistant (RFP) HGSOC was quantified by flow cytometry. AT was compared to standard carboplatin dosing in mice with HGSOC xenografts and excised tumors were examined by IHC. Cumulative drug dose and murine survival were compared. Cell-free DNA (cfDNA) and tumor biopsies were obtained from HGSOC patients during carboplatin and PARPi treatment and subjected to low pass whole genome sequencing and analysis with our published bioinformatic pipeline (LiquidCNA) that estimates the size of the emergent resistant population. Finally, established patient datasets were examined to adapt our pre-clinical discoveries for clinical testing in the ACTOv trial. Results. Cell viability assays confirmed platinum resistance. One murine HGSOC cell line with evolved platinum resistance (60577R3) was also resistant to olaparib and niraparib in vitro, whereas there was no difference in PARPi sensitivity in the other sensitive/resistant cell pairs. Fitness deficits of resistance in the absence of drug, were exposed by low resource co-cultures in vitro in four out of five HGSOC cell lines with evolved platinum resistance. Conversely, the resistant subline, 60577R3, exhibited greater fitness than sensitive, ancestral cells (60577). Resistant populations with reduced fitness declined due to both reduced proliferation and increased apoptosis in vitro and in vivo when drug was absent. Rad51 foci formation in response to double strand breaks was observed in platinum and PARPi-resistant 60577R3 cells but not sensitive 60577 indicating that functional HRR had been restored in 60577R3 as resistance evolved. Consistent with this, 60577R3 cells also made fewer segregation errors during mitosis. Western blot confirmed expression of BRCA1 in 60577R3 but not 60577 cells. Platinum treatment of in vitro co-cultures demonstrated that the size of sensitive and resistant HGSOC populations fluctuated with drug therapy and that fitter, sensitive cells re-grew when drug was withdrawn. In vivo, AT significantly prolonged survival of murine ovarian cancer models compared to standard carboplatin dosing without increasing mean daily drug dose or drug-related toxicity. Escape from control by AT was associated with growth of resistant cells in our in vivo models. In HGSOC patients, LiqCNA analysis of cfDNA obtained longitudinally during treatment showed sub-clonal selection through therapy and demonstrated that LiqCNA correlates with increases in the serum tumor marker, CA125, a proxy for disease progression. Existing clinical datasets of CA125 during standard carboplatin treatment defined the parameters for dose adaptations in patients receiving AT in the ACTOv trial. Conclusions. Platinum resistance can be associated with fitness deficits in HGSOC, which can be exploited by AT to preserve platinum sensitivity over the long-term and prolong survival. BRCA reversion mutations are the most well-documented cause of acquired PARPi resistance. Our data imply that this may provide a fitness advantage for HGSOC cells with implications for PARPi maintenance treatment and response to subsequent lines of therapy. These discoveries paved the way for us to launch the ACTOv trial, which is currently recruiting patients in ten NHS hospitals throughout the UK. ACTOv patients will provide biopsies and very frequent blood samples for circulating tumor DNA (total ∼1,400 samples from 80 patients), enabling us to interrogate clonal evolution during standard and adaptive therapy and relate this to clinical outcome. LiqCNA could provide a biomarker for the emergence of drug resistance to direct AT in second generation clinical trials. Citation Format: Mohammed Ateeb Khan, Helen Hockings, Maximilian Mossner, Ann-Marie Baker, Hall Amy, Robert L. Hollis, Weini Huang, Eszter Lakatos, Charlie Gourley, Trevor A. Graham, Michelle Lockley. Translating adaptive therapy to ovarian cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr SY43-01.