Patients with high-grade serous ovarian cancer (HGSC) have experienced little improvement in overall survival, and standard treatment has not advanced beyond platinum-based combination chemotherapy, during the past 30 years. To understand the drivers of clinical phenotypes better, here we use whole-genome sequencing of tumour and germline DNA samples from 92 patients with primary refractory, resistant, sensitive and matched acquired resistant disease. We show that gene breakage commonly inactivates the tumour suppressors RB1, NF1, RAD51B and PTEN in HGSC, and contributes to acquired chemotherapy resistance. CCNE1 amplification was common in primary resistant and refractory disease. We observed several molecular events associated with acquired resistance, including multiple independent reversions of germline BRCA1 or BRCA2 mutations in individual patients, loss of BRCA1 promoter methylation, an alteration in molecular subtype, and recurrent promoter fusion associated with overexpression of the drug efflux pump MDR1.

The pan-cancer analysis of whole genomes The expansion of whole-genome sequencing studies from individual ICGC and TCGA working groups presented the opportunity to undertake a meta-analysis of genomic features across tumour types. To achieve this, the PCAWG Consortium was established. A Technical Working Group implemented the informatics analyses by aggregating the raw sequencing data from different working groups that studied individual tumour types, aligning the sequences to the human genome and delivering a set of high-quality somatic mutation calls for downstream analysis (Extended Data Fig. 1). Given the recent meta-analysis

Summary Intra-tumor heterogeneity (ITH) is a mechanism of therapeutic resistance and therefore an important clinical challenge. However, the extent, origin, and drivers of ITH across cancer types are poorly understood. To address this, we extensively characterize ITH across whole-genome sequences of 2,658 cancer samples spanning 38 cancer types. Nearly all informative samples (95.1%) contain evidence of distinct subclonal expansions with frequent branching relationships between subclones. We observe positive selection of subclonal driver mutations across most cancer types and identify cancer type-specific subclonal patterns of driver gene mutations, fusions, structural variants, and copy number alterations as well as dynamic changes in mutational processes between subclonal expansions. Our results underline the importance of ITH and its drivers in tumor evolution and provide a pan-cancer resource of comprehensively annotated subclonal events from whole-genome sequencing data.

Most women diagnosed with high-grade serous ovarian cancer (HGSC) develop recurrent disease and chemotherapy resistance, despite initially responding to treatment. The genomic characteristics of HGSC samples collected at initial surgery have been extensively studied. However, due to challenges of sample collection following treatment, much less is known about the molecular features of recurrent disease. Our recent studies have identified mechanisms of acquired resistance and biomarkers in recurrent HGSCs that could lead to improved treatment approaches.

ABCB1 encodes Multidrug Resistance protein (MDR1), an ATP-binding cassette member involved in the cellular efflux of chemotherapeutic drugs. Here we report that ovarian and breast samples from chemotherapy treated patients are positive for multiple transcriptional fusions involving ABCB1, placing it under the control of a strong promoter while leaving its open reading frame intact. We identified 15 different transcriptional fusion partners involving ABCB1, as well as patients with multiple distinct fusion events. The partner gene selected depended on its structure, promoter strength, and chromosomal proximity to ABCB1. Fusion positivity was strongly associated with the number of lines of MDR1-substrate chemotherapy given. MDR1 inhibition in a fusion positive ovarian cancer cell line increased sensitivity to paclitaxel more than 50-fold. Convergent evolution of ABCB1 fusion is therefore frequent in chemotherapy resistant recurrent ovarian cancer. As most currently approved PARP inhibitors (PARPi) are MDR1 substrates, prior chemotherapy may precondition resistance to PARPi.

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Purpose Germline BRCA1 or BRCA2 mutations in patients with high-grade serous ovarian cancer (HGSC) are associated with favorable responses to chemotherapy. However, secondary intragenic (reversion) mutations that restore protein function lead to clinically significant rates of acquired resistance. The goal of this study was to determine whether reversion mutations could be found in an unbiased manner in circulating cell-free DNA (cfDNA) to predict treatment response in HGSC. Patients and Methods Plasma and tumor samples were obtained from 30 patients with HGSC with either BRCA1 or BRCA2 germline mutation. Two cohorts were ascertained: patients with a malignancy before undergoing primary HGSC debulking surgery (n = 14) or patients at disease recurrence (n = 16). Paired tumor and plasma samples were available for most patients (24 of 30). Targeted amplicon, next-generation sequencing was performed using primers that flanked germline mutations, whose design did not rely on prior knowledge of reversion sequences. Results Five patients were identified with intragenic mutations predicted to restore BRCA1/2 open reading frames, including two patients with multiple independent reversion alleles. Reversion mutations were only detected in tumor samples from patients with recurrent disease (five of 16) and only in cfDNA from patients with a tumor-detected reversion (three of five). Findings from a rapid autopsy of a patient with multiple independent reversions indicated that reversion-allele frequency in metastatic sites is an important determinant of assay sensitivity. Abundance of tumor-derived DNA in total cell-free DNA, as measured by TP53 mutant allele frequency, also affected assay sensitivity. All patients with reversions detected in tumor-derived DNA were resistant to platin- or poly ADP ribose polymerase inhibitor-based chemotherapy. Conclusion Reversion mutations can be detected in an unbiased analysis of cfDNA, suggesting clinical utility for predicting chemotherapy response in recurrent HGSC.

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