Clinical BRCA1/2 Reversion Analysis Identifies Hotspot Mutations and Predicted Neoantigens Associated with Therapy Resistance

Abstract

Reversion mutations in BRCA1 or BRCA2 are associated with resistance to PARP inhibitors and platinum. To better understand the nature of these mutations, we collated, codifi ed, and analyzed more than 300 reversions. This identifi ed reversion "hotspots" and "deserts" in regions encoding the N and C terminus, respectively, of BRCA2, suggesting that pathogenic mutations in these regions may be at higher or lower risk of reversion. Missense and splice-site pathogenic mutations in BRCA1/2 also appeared less likely to revert than truncating mutations. Most reversions were <100 bp deletions. Although many deletions exhibited microhomology, this was not universal, suggesting that multiple DNA-repair processes cause reversion. Finally, we found that many reversions were predicted to encode immunogenic neopeptides, suggesting a route to the treatment of reverted disease. As well as providing a freely available database for the collation of future reversion cases, these observations have implications for how drug resistance might be managed in BRCA -mutant cancers. SIGNIFICANCE: Reversion mutations in BRCA genes are a major cause of clinical platinum and PARP inhibitor resistance. This analysis of all reported clinical reversions suggests that the position of BRCA2 mutations affects the risk of reversion. Many reversions are also predicted to encode tumor neoantigens, providing a potential route to targeting resistance.

A meta-analysis of reversion mutations in BRCA genes identifies signatures of DNA end-joining repair mechanisms driving therapy resistance

“…58 Notwithstanding, BRCA reversions are the only confirmed mechanism of resistance identified in clinical samples and their exact prevalence will be better defined with the acquisition of more clinical data from patients progressing on platinum drugs or PARPi. Of note, while this work was under review, an article by Pettitt and colleagues 59 was published elsewhere reporting similar findings to ours.…”

Treatment-emergent neuroendocrine prostate cancer with a germline BRCA2 mutation: identification of a candidate reversion mutation associated with platinum/PARP-inhibitor resistance

“…The case presented also has clinical relevance regarding the uses of cfDNA analysis vs tumor profiling to detect BRCA2 reversion mutations in patients with prostate cancer. The lack of detection of our reversion mutation by Guardant360 may relate to its large size (228 bp) relative to the typical reversion mutation, which is <100 bp (Pettitt et al 2020). Although some groups recommend cfDNA analysis over tumor biopsy for detection of the full spectrum of tumor heterogeneity (Carneiro et al 2018), others have found that the assay has limitations in detecting clinically relevant alterations (including large somatic deletions) in a significant percentage of patients (Taavitsainen et al 2019).…”

“…This novel mutation could be considered a reversion mutation, if occurring in cis with the germline mutation. It has not been reported in the IRC Database of Reversion Mutations (https ://reversions.icr.ac.uk/), one of the largest of such databases (Pettitt et al 2020). Because BRCA2 allelic loss was not reported by MSK-IMPACT, we attempted to determine this by performing qualitative polymerase chain reaction (qPCR) and Sanger sequencing on microdissected tumor tissue (Fig.…”

“…TaqMan qPCR genotyping assay was performed to target the germline point mutation but showed the same heterogeneity of wild-type and mutant alleles; this was in part due to the inability to eliminate all normal cells from the biopsy samples. Although we could not confirm that the somatic BRCA2 deletion was cis to the germline mutation, the concurrent emergence of resistance to platinum and PARPi therapies as well as several lines of evidence support this as a reversion mutation: (1) The affected region is frequently deleted in reversions; (2) the deletion includes the germline mutation and restores the open reading frame (ORF), an event unlikely to occur at the same location on the normal allele; and (3) the deletion is flanked by microhomology (CAA), a characteristic of many reversion mutations (Pettitt et al 2020).…”

Long-term survival of an ovarian cancer patient harboring a RAD51C missense mutation

“…In a long-term follow-up study of a large number of BRCA1/2 ovarian cancer patients, the 5-yr disease-free interval treated with platinum drugs is only ∼15% (Jorge et al 2019). The major mode of verified therapy resistance is reversion of the mutated gene (Chen et al 2018), with approximately 300 reversion alleles reported thus far from 91 patients (Pettitt et al 2020). Although reversion to wild type can occur, most often a secondary mutation restores the reading frame from a primary truncation mutation.…”