Longitudinal profiling identifies co-occurring BRCA1/2 reversions, TP53BP1, RIF1 and PAXIP1 mutations in PARP inhibitor-resistant advanced breast cancer

BackgroundResistance to therapies that target homologous recombination deficiency (HRD) in breast cancer limits their overall effectiveness. Multiple preclinically-validated mechanisms of resistance have been proposed, but their existence and relative frequency in clinical disease is unclear, as is how to target resistance.Patients and MethodsLongitudinal mutation and methylation profiling of circulating tumour (ct)DNA was performed in 47 patients with metastatic BRCA1, BRCA2 or PALB2 mutant breast cancer treated with HRD-targeted therapy and developed progressive disease – 18 patients had primary resistance, 29 exhibited response followed by resistance. ctDNA isolated at multiple timepoints in the patient treatment course (before, on-treatment, and at progression) was sequenced using a novel >750 gene intron/exon targeted sequencing panel. Where available, matched tumour biopsies were whole exome and RNA sequenced and also used to assess functional RAD51 status.ResultsBRCA1/2 reversion mutations were present in 60% of patients as the most prevalent form of resistance. In 10 cases, reversions were detected in ctDNA prior to clinical progression. Two new reversion-based mechanisms were identified: (i) intragenic BRCA1/2 deletions with intronic breakpoints; and (ii) intragenic BRCA1/2 secondary mutations that formed novel splice acceptor sites, the latter being confirmed by in vitro minigene reporter assays. When seen prior to commencing subsequent treatment, reversions were associated with significantly shorter time to progression. Tumours with reversions retained HRD mutational signatures but had functional HRD based on RAD51 status. Although less frequent than reversions, non-reversion mechanisms (loss of function mutations in TP53BP1, RIF1 or PAXIP1) were evident in patients with acquired resistance and occasionally co-existed with reversions, challenging the notion that singular resistance mechanisms emerge in each patient.ConclusionsThese observations map the prevalence of candidate drivers of resistance across time in a clinical setting, information with implications for clinical management and trial design in HRD breast cancers.