Targeting Polθ Helicase with RP-3467: Preclinical Validation and Clinical Response in a gBRCA1 ovarian cancer patient with an acquired PARPi resistance mutation in TP53BP1

Homologous recombination (HR) deficiency underlies a range of cancers, and PARP inhibitors (PARPi) targeting this vulnerability have demonstrated clinical efficacy in BRCA1/2-mutant tumors. However, resistance mechanisms and tolerability issues limit their long-term clinical effectiveness. Here, we characterize RP-3467, a novel, selective small-molecule inhibitor targeting the ATPase domain of DNA polymerase theta (Pol{theta}), the key mediator of the error-prone microhomology-mediated end joining (MMEJ) pathway. We demonstrate that RP-3467 potently suppresses Pol{theta} enzymatic activity, abrogates MMEJ, and disrupts mitotic DNA repair in HR-deficient (HRD) models, inducing synthetic lethality. In preclinical studies, we show that RP-3467 synergizes strongly with PARPi (olaparib and rucaparib), achieving sustained tumor regression across multiple xenograft and patient-derived xenograft (PDX) models without added systemic toxicity. Mechanistically, we find that RP-3467 increases mitotic DNA damage in vivo, as evidenced by elevated micronuclei and CIP2A foci. RP-3467 was evaluated in a phase 1 clinical trial (NCT06560632) and has shown preliminary signs of clinical activity. A heavily pretreated germline BRCA1-mutant ovarian cancer patient achieved a confirmed partial response to RP-3467 plus olaparib (RECIST v1.1), with retrospective profiling revealing a TP53BP1 loss-of-function mutation previously linked to both PARPi resistance and POLQ dependency. Although broader clinical validation is needed, these early findings suggest that Pol{theta} inhibition has the potential to address PARPi resistance and offer therapeutic benefit in HRD cancers.