Abstract P5-11-09: Feasibility of tracking plasma DNA mutation kinetics in estrogen receptor positive metastatic breast cancer using a novel digital PCR amplicon sequencing assay

Estrogen Receptor 1 ( ESR1 ) gene mutations in estrogen receptor positive (ER+) metastatic breast cancer (MBC) are associated with clinical resistance to endocrine therapy. A highly sensitive digital PCR (dPCR) assay was developed to evaluate the baseline abundance and kinetics of hotspot ESR1 mutations in plasma circulating tumor DNA (ctDNA) from 58 ER+ MBC patients who had progressed on at least one line of prior endocrine therapy. Consistent with prior reports, we demonstrate that detection of hotspot ESR1 mutations in plasma ctDNA is associated with shorter progression-free survival. However, after patients initiated a new course of systemic therapy, changes in plasma ESR1 mutation abundance did not correlate with the duration of disease control. To investigate whether mutational heterogeneity may account for this lack of correlation, we developed a customized dPCR multiplexed amplicon next-generation sequencing (NGS) assay to detect plasma ctDNA mutations in the complete coding sequence of ESR1 and TP53 , as well as hotspot regions in PIK3CA . Assay validation revealed a 79% sensitivity and 100% specificity for detecting plasma ctDNA mutations with a lower limit of detecting a 1.6% mutant allele fraction. The NGS assay revealed non-hotspot ESR1 mutations in plasma DNA from 14 of 31 (45%) patients. We also observed dynamic changes in plasma ctDNA mutant allele fraction of PIK3CA and TP53 during systemic therapy. Changes in the cumulative mutant allele fraction of ESR1 , PIK3CA , and TP53 in plasma ctDNA correlated with duration of clinical treatment response in a limited subset of patients (n=8) for whom NGS was performed at two or more time points. Thus, mutational heterogeneity limits the clinical utility of monitoring individual hotspot mutations in ESR1 over time in ER+ MBC patients. However, a plasma ctDNA NGS assay targeting ESR1 , PIK3CA , and TP53 may be useful as an early predictor of response to systemic therapy. Citation Format: Kumar S, Lindsay D, Chen B, Garrett AL, Tan XM, Anders CK, Carey LA, Gupta GP. Feasibility of tracking plasma DNA mutation kinetics in estrogen receptor positive metastatic breast cancer using a novel digital PCR amplicon sequencing assay [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-11-09.