AKT1 interacts with DHX9 to mitigate R-loop-induced replication stress in ovarian cancer

PARP inhibitor (PARPi)-resistant BRCA mutant (BRCAm) high-grade serous ovarian cancer (HGSOC) represents a new clinical challenge with unmet therapeutic needs. Quantitative high-throughput drug combination screen identifies that ATR inhibitor (ATRi) and AKT inhibitor (AKTi) combination is a rational treatment strategy for both PARPi-sensitive and PARPi-resistant BRCAm HGSOC by inducing DNA damage and R-loop-mediated replication stress (RS). Mechanistically, the kinase domain of AKT1 directly interacts with DHX9, thus facilitating recruitment of DHX9 to R-loops. AKTi increases ATRi-induced R-loop-mediated RS by mitigating recruitment of DHX9 to R-loops. Moreover, DHX9 is upregulated in tumors from PARPi-resistant BRCAm HGSOC patients and high co-expression of DHX9 and AKT1 correlates with worse survival. Our study reveals a previously unknown interaction between AKT1 and DHX9 in R-loop resolution and novel mechanisms of action of AKTi and ATRi combination. Our data also provide a rationale for the clinical development of ATRi and AKTi combination for BRCAm HGSOC irrespective of PARPi resistance status and a potential biomarker to predict the response of combination therapy. Overall design: RNA sequencing (RNAseq) data were obtained from total RNA from fresh frozen tumor biopsy samples from 14 BRCAm PARPi-resistant HGSOC patients enrolled in the clinical trial at the National Cancer Institute (ClinicalTrials.gov, NCT02203513). RNAseq was performed using a HiSeq4000 sequencing system (Illumina) at the CCR Sequencing Facility/NCI on pre-treatment biopsy samples from 14 patients. Paired-end sequencing was done. Quartile normalization and log-transformation prior to analysis were performed on datasets.