Accurate identification of tumor-derived somatic variants in plasma circulating cell-free DNA (cfDNA) requires understanding the various biologic compartments contributing to the cfDNA pool. We sought to define the technical feasibility of a high-intensity sequencing assay of cfDNA and matched white-blood cell (WBC) DNA covering a large genomic region (508 genes, 2Mb, >60,000X raw-depth) in a prospective study of 124 metastatic cancer patients, with contemporaneous matched tumor tissue biopsies, and 47 non-cancer controls. The assay displayed a high sensitivity and specificity, allowing for de novo detection of tumor-derived mutations and inference of tumor mutational burden, microsatellite instability, mutational signatures and sources of somatic mutations identified in cfDNA. The vast majority of cfDNA mutations (81.6% in controls and 53.2% in cancer patients) had features consistent with clonal hematopoiesis (CH). This cfDNA sequencing approach revealed that CH constitutes a pervasive biological phenomenon emphasizing the importance of matched cfDNA-WBC sequencing for accurate variant interpretation.

SUMMARY CDK4/6 inhibitors (CDK4/6i) are effective in breast cancer, however drug resistance is frequently encountered and poorly understood. We conducted a genomic analysis of 348 estrogen receptor-positive breast cancers treated with CDK4/6i and identified loss of function mutations affecting FAT1 and RB1 linked to drug resistance. FAT1 loss led to marked elevations in CDK6 whose suppression restored sensitivity to CDK4/6i. The induction of CDK6 was mediated by the Hippo pathway with accumulation of YAP and TAZ transcription factors on the CDK6 promoter. Genomic alterations in other Hippo pathway components were also found to promote CDK4/6i resistance. These findings uncover a tumor suppressor function of Hippo signaling in ER+ breast cancer and establish FAT1 loss as a mechanism of resistance to CDK4/6i.

This study demonstrates that we can now begin to individualize the treatment of ILBC, with HER2, HER3, and AKT1 mutations representing high-prevalence therapeutic targets and FOXA1 mutations and ESR1 gains deserving urgent dedicated clinical investigation, especially in the context of endocrine treatment.

SummaryLittle is known about how RNA editing operates in cancer. Transcriptome analysis of 68 normal and cancerous breast tissues revealed that the editing enzyme ADAR acts uniformly, on the same loci, across tissues. In controlled ADAR expression experiments, the editing frequency increased at all loci with ADAR expression levels according to the logistic model. Loci-specific “editabilities,” i.e., propensities to be edited by ADAR, were quantifiable by fitting the logistic function to dose-response data. The editing frequency was increased in tumor cells in comparison to normal controls. Type I interferon response and ADAR DNA copy number together explained 53% of ADAR expression variance in breast cancers. ADAR silencing using small hairpin RNA lentivirus transduction in breast cancer cell lines led to less cell proliferation and more apoptosis. A-to-I editing is a pervasive, yet reproducible, source of variation that is globally controlled by 1q amplification and inflammation, both of which are highly prevalent among human cancers.