Circulating tumor DNA association with residual cancer burden after neoadjuvant chemotherapy in triple-negative breast cancer in TBCRC 030.

BACKGROUND: We aimed to examine circulating tumor DNA (ctDNA) and its association with residual cancer burden (RCB) using an ultrasensitive assay in patients with triple-negative breast cancer (TNBC) receiving neoadjuvant chemotherapy. PATIENTS AND METHODS: We identified responders (RCB 0/1) and matched non-responders (RCB 2/3) from the phase II TBCRC 030 prospective study of neoadjuvant paclitaxel versus cisplatin in TNBC. We collected plasma samples at baseline, 3 weeks and 12 weeks (end of therapy). We created personalized ctDNA assays utilizing MAESTRO mutation enrichment sequencing. We explored associations between ctDNA and RCB status and disease recurrence. RESULTS: Of 139 patients, 68 had complete samples and no additional neoadjuvant chemotherapy. Twenty-two were responders and 19 of those had sufficient tissue for whole-genome sequencing. We identified an additional 19 non-responders for a matched case-control analysis of 38 patients using a MAESTRO ctDNA assay tracking 319-1000 variants (median 1000 variants) to 114 plasma samples from 3 timepoints. Overall, ctDNA positivity was 100% at baseline, 79% at week 3 and 55% at week 12. Median tumor fraction (TFx) was 3.7 × 10-4 (range 7.9 × 10-7-4.9 × 10-1). TFx decreased 285-fold from baseline to week 3 in responders and 24-fold in non-responders. Week 12 ctDNA clearance correlated with RCB: clearance was observed in 10 of 11 patients with RCB 0, 3 of 8 with RCB 1, 4 of 15 with RCB 2 and 0 of 4 with RCB 3. Among six patients with known recurrence, five had persistent ctDNA at week 12. CONCLUSIONS: Neoadjuvant chemotherapy for TNBC reduced ctDNA TFx by 285-fold in responders and 24-fold in non-responders. In 58% (22/38) of patients, ctDNA TFx dropped below the detection level of a commercially available test, emphasizing the need for sensitive tests. Additional studies will determine whether ctDNA-guided approaches can improve outcomes.

Reversion and non-reversion mechanisms of resistance to PARP inhibitor or platinum chemotherapy in BRCA1/2-mutant metastatic breast cancer.

BACKGROUND: Little is known about mechanisms of resistance to poly(adenosine diphosphate-ribose) polymerase inhibitors (PARPi) and platinum chemotherapy in patients with metastatic breast cancer and BRCA1/2 mutations. Further investigation of resistance in clinical cohorts may point to strategies to prevent or overcome treatment failure. PATIENTS AND METHODS: We obtained tumor biopsies from metastatic breast cancer patients with BRCA1/2 deficiency before and after acquired resistance to PARPi or platinum chemotherapy. Whole exome sequencing was carried out on each tumor, germline DNA, and circulating tumor DNA. Tumors underwent RNA sequencing, and immunohistochemical staining for RAD51 foci on tumor sections was carried out for functional assessment of intact homologous recombination (HR). RESULTS: Pre- and post-resistance tumor samples were sequenced from eight patients (four with BRCA1 and four with BRCA2 mutation; four treated with PARPi and four with platinum). Following disease progression on DNA-damaging therapy, four patients (50%) acquired at least one somatic reversion alteration likely to result in functional BRCA1/2 protein detected by tumor or circulating tumor DNA sequencing. Two patients with germline BRCA1 deficiency acquired genomic alterations anticipated to restore HR through increased DNA end resection: loss of TP53BP1 in one patient and amplification of MRE11A in another. RAD51 foci were acquired post-resistance in all patients with genomic reversion, consistent with reconstitution of HR. All patients whose tumors demonstrated RAD51 foci post-resistance were intrinsically resistant to subsequent lines of DNA-damaging therapy. CONCLUSIONS: Genomic reversion in BRCA1/2 was the most commonly observed mechanism of resistance, occurring in four of eight patients. Novel sequence alterations leading to increased DNA end resection were seen in two patients, and may be targetable for therapeutic benefit. The presence of RAD51 foci by immunohistochemistry was consistent with BRCA1/2 protein functional status from genomic data and predicted response to later DNA-damaging therapy, supporting RAD51 focus formation as a clinically useful biomarker.

Whole-exome sequencing of cell-free DNA and circulating tumor cells in multiple myeloma.

Liquid biopsies including circulating tumor cells (CTCs) and cell-free DNA (cfDNA) have enabled minimally invasive characterization of many cancers, but are rarely analyzed together. Understanding the detectability and genomic concordance of CTCs and cfDNA may inform their use in guiding cancer precision medicine. Here, we report the detectability of cfDNA and CTCs in blood samples from 107 and 56 patients with multiple myeloma (MM), respectively. Using ultra-low pass whole-genome sequencing, we find both tumor fractions correlate with disease progression. Applying whole-exome sequencing (WES) to cfDNA, CTCs, and matched tumor biopsies, we find concordance in clonal somatic mutations (~99%) and copy number alterations (~81%) between liquid and tumor biopsies. Importantly, analyzing CTCs and cfDNA together enables cross-validation of mutations, uncovers mutations exclusive to either CTCs or cfDNA, and allows blood-based tumor profiling in a greater fraction of patients. Our study demonstrates the utility of analyzing both CTCs and cfDNA in MM.