Geoffrey I Shapiro

Phase I study of ribociclib (CDK4/6 inhibitor) with spartalizumab (PD-1 inhibitor) with and without fulvestrant in metastatic hormone receptor-positive breast cancer or advanced ovarian cancer

Background Preclinical evidence suggests that cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitors enhance antitumor immunity. We conducted a phase I trial of ribociclib (CDK4/6 inhibitor) plus spartalizumab (PD-1 inhibitor) in patients with hormone receptor (HR)-positive/HER2-negative metastatic breast cancer (MBC) or advanced ovarian cancer (AOC). The combination was also evaluated with fulvestrant in MBC. Methods In Cohort A, ribociclib was administered on Days 1–21 (28-day cycle) starting at 400 mg, and spartalizumab at 400 mg on Day 1. Dose escalation was followed by expansion in AOC. Fulvestrant was added (Cohort B) with a safety run-in followed by expansion in MBC. Primary objectives were to determine the maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D), and safety and tolerability of the combinations. Results 33 patients enrolled (n=18, Cohort A; n=15, Cohort B). The RP2D of ribociclib in both cohorts was 600 mg. Treatment-related adverse events in >20% of patients in either cohort were neutropenia, fatigue, anemia, thrombocytopenia, hypertransaminasemia, maculopapular rash, fatigue, and nausea. Hypertransaminasemia occurred in 66.7% (AST) and 46.7% (ALT) of patients in Cohort B, including 46.7% and 40.0%, respectively, of grade 3 or 4 events. Two confirmed partial responses were observed (13.3%) in Cohort B, in patients with low baseline serum thymidine kinase activity, coupled with an increase on-treatment. Peripheral blood flow cytometry across patients demonstrated on-target drug binding with increases in PD-1 occupancy and activated CD8+ T cells during treatment, irrespective of response. PD-L1-positivity, tumor-infiltrating lymphocytes, or tumor mutational burden did not correlate with progression-free survival (PFS). Several copy-number variations detected with next-generation sequencing correlated with PFS. Conclusions Ribociclib with spartalizumab and fulvestrant showed limited efficacy and elevated hepatotoxicity, precluding further development. Correlative analyses revealed treatment-induced immunological effects, and genomic alterations associated with PFS.

PARG inhibitor sensitivity correlates with accumulation of single-stranded DNA gaps in preclinical models of ovarian cancer

Poly (ADP-ribose) glycohydrolase (PARG) is a dePARylating enzyme which promotes DNA repair by removal of poly (ADP-ribose) (PAR) from PARylated proteins. Loss or inhibition of PARG results in replication stress and sensitizes cancer cells to DNA-damaging agents. PARG inhibitors are now undergoing clinical development for patients having tumors with homologous recombination deficiency (HRD), such as cancer patients with germline or somaticBRCA1/2-mutations. PARP inhibitors kill BRCA-deficient cancer cells by increasing single-stranded DNA gaps (ssGAPs) during replication. Here, we report that, like PARP inhibitor (PARPi), PARG inhibitor (PARGi) treatment also causes an accumulation of ssGAPs in sensitive cells. PARGi exposure increased accumulation of S-phase-specific PAR, a marker for Okazaki fragment processing (OFP) defects on lagging strands and induced ssGAPs, in sensitive cells but not in resistant cells. PARGi also caused accumulation of PAR at the replication forks and at the ssDNA sites in sensitive cells. Additionally, PARGi exhibited monotherapy activity in specific HR-deficient, as well as HR-proficient, patient-derived, or patient-derived xenograft (PDX)-derived organoids of ovarian cancer, and drug sensitivity directly correlated with the accumulation of ssGAPs. Taken together, PARGi treatment results in toxic accumulation of PAR at replication forks resulting in ssGAPs due to OFP defects during replication. Regardless of theBRCA/HRD-status, the induction of ssGAPs in preclinical models of ovarian cancer cells correlates with PARGi sensitivity. Patient-derived organoids (PDOs) may be a useful model system for testing PARGi sensitivity and functional biomarkers.

Single-Stranded DNA Gap Accumulation Is a Functional Biomarker for USP1 Inhibitor Sensitivity

Abstract Recent studies suggest that PARP and POLQ inhibitors confer synthetic lethality in BRCA1-deficient tumors by accumulation of single-stranded DNA (ssDNA) gaps at replication forks. Loss of USP1, a deubiquitinating enzyme, is also synthetically lethal with BRCA1 deficiency, and USP1 inhibitors are now undergoing clinical development for these cancers. Herein, we show that USP1 inhibitors also promote the accumulation of ssDNA gaps during replication in BRCA1-deficient cells, and this phenotype correlates with drug sensitivity. USP1 inhibition increased monoubiquitinated proliferating cell nuclear antigen at replication forks, mediated by the ubiquitin ligase RAD18, and knockdown of RAD18 caused USP1 inhibitor resistance and suppression of ssDNA gaps. USP1 inhibition overcame PARP inhibitor resistance in a BRCA1-mutated xenograft model and induced ssDNA gaps. Furthermore, USP1 inhibition was synergistic with PARP and POLQ inhibition in BRCA1-mutant cells, with enhanced ssDNA gap accumulation. Finally, in patient-derived ovarian tumor organoids, sensitivity to USP1 inhibition alone or in combination correlated with the accumulation of ssDNA gaps. Assessment of ssDNA gaps in ovarian tumor organoids represents a rapid approach for predicting response to USP1 inhibition in ongoing clinical trials. Significance: USP1 inhibitors kill BRCA1-deficient cells and cause ssDNA gap accumulation, supporting the potential of using ssDNA gap detection as a functional biomarker for clinical trials on USP1 inhibitors.

Combined PARP and HSP90 inhibition: preclinical and Phase 1 evaluation in patients with advanced solid tumours

PARP inhibitor resistance may be overcome by combinatorial strategies with agents that disrupt homologous recombination repair (HRR). Multiple HRR pathway components are HSP90 clients, so that HSP90 inhibition leads to abrogation of HRR and sensitisation to PARP inhibition. We performed in vivo preclinical studies of the HSP90 inhibitor onalespib with olaparib and conducted a Phase 1 combination study. Tolerability and efficacy studies were performed in patient-derived xenograft(PDX) models of ovarian cancer. Clinical safety, tolerability, steady-state pharmacokinetics and preliminary efficacy of olaparib and onalespib were evaluated using a standard 3 + 3 dose-escalation design. Olaparib/onalespib exhibited anti-tumour activity against BRCA1-mutated PDX models with acquired PARPi resistance and PDX models with RB-pathway alterations(CDKN2A loss and CCNE1 overexpression). Phase 1 evaluation revealed that dose levels up to olaparib 300 mg/onalespib 40 mg and olaparib 200 mg/onalespib 80 mg were safe without dose-limiting toxicities. Coadministration of olaparib and onalespib did not appear to affect the steady-state pharmacokinetics of either agent. There were no objective responses, but disease stabilisation ≥24 weeks was observed in 7/22 (32%) evaluable patients including patients with BRCA-mutated ovarian cancers and acquired PARPi resistance and patients with tumours harbouring RB-pathway alterations. Combining onalespib and olaparib was feasible and demonstrated preliminary evidence of anti-tumour activity.

Phase 1 Combination Study of the CHK1 Inhibitor Prexasertib and the PARP Inhibitor Olaparib in High-grade Serous Ovarian Cancer and Other Solid Tumors

Abstract Purpose: Checkpoint kinase 1 (CHK1) plays a central role in the response to replication stress through modulation of cell-cycle checkpoints and homologous recombination (HR) repair. In BRCA-deficient cancers with de novo or acquired PARP inhibitor resistance, the addition of the CHK1 inhibitor prexasertib to the PARP inhibitor olaparib compromises replication fork stability, as well as HR proficiency, allowing for sensitization to PARP inhibition. Patients and Methods: This study followed a 3+3 design with a 7-day lead-in of olaparib alone, followed by 28-day cycles with prexasertib administered on days 1 and 15 in combination with an attenuated dose of olaparib on days 1–5 and 15–19. Pharmacokinetic blood samples were collected after olaparib alone and following combination therapy. Patients enrolled to the expansion phase of the study underwent paired tumor biopsies for pharmacodynamic (PD) assessments. Results: Twenty-nine patients were treated. DLTs included grade 3 neutropenia and grade 3 febrile neutropenia. The MTD/recommended phase 2 dose (RP2D) was prexasertib at 70 mg/m2 i.v. with olaparib at 100 mg by mouth twice daily. Most common treatment-related adverse events included leukopenia (83%), neutropenia (86%), thrombocytopenia (66%), and anemia (72%). Four of 18 patients with BRCA1-mutant, PARP inhibitor–resistant, high-grade serous ovarian cancer (HGSOC) achieved partial responses. Paired tumor biopsies demonstrated reduction in RAD51 foci and increased expression of γ-H2AX, pKAP1, and pRPA after combination exposure. Conclusions: Prexasertib combined with olaparib has preliminary clinical activity in BRCA-mutant patients with HGSOC who have previously progressed on a PARP inhibitor. PD analyses show that prexasertib compromises HR with evidence of induction of DNA damage and replication stress.

Randomized Phase II Study of Gemcitabine With or Without ATR Inhibitor Berzosertib in Platinum-Resistant Ovarian Cancer: Final Overall Survival and Biomarker Analyses

PURPOSE The multicenter, open-label, randomized phase 2 NCI-9944 study ( NCT02595892 ) demonstrated that addition of ATR inhibitor (ATRi) berzosertib to gemcitabine increased progression-free survival (PFS) compared to gemcitabine alone (hazard ratio [HR]=0.57, one-sided log-rank P = .044, which met the one-sided significance level of 0.1 used for sample size calculation). METHODS We report here the final overall survival (OS) analysis and biomarker correlations (ATM expression by immunohistochemistry, mutational signature 3 and a genomic biomarker of replication stress) along with post-hoc exploratory analyses to adjust for crossover from gemcitabine to gemcitabine/berzosertib. RESULTS At the data cutoff of January 27, 2023 (>30 months of additional follow-up from the primary analysis), median OS was 59.4 weeks with gemcitabine/berzosertib versus 43.0 weeks with gemcitabine alone (HR 0.79, 90% CI 0.52 to 1.2, one-sided log-rank P = .18). An OS benefit with addition of berzosertib to gemcitabine was suggested in patients stratified into the platinum-free interval ≤3 months (N = 26) subgroup (HR, 0.48, 90% CI 0.22 to 1.01, one-sided log-rank P =.04) and in patients with ATM-negative/low (N = 24) tumors (HR, 0.50, 90% CI 0.23 to 1.08, one-sided log-rank P = .06). CONCLUSION The results of this follow-up analysis continue to support the promise of combined gemcitabine/ATRi therapy in platinum resistant ovarian cancer, an active area of investigation with several ongoing clinical trials.