Abstract P2-01-13: Longitudinal circulating tumor DNA (ctDNA) monitoring by digital droplet PCR (ddPCR) in metastatic breast cancer

Abstract

Abstract Background: A common challenge in treating patients with metastatic breast cancer (MBC) is ensuring adequate therapeutic efficacy, maintenance of quality of life, and avoidance of drug toxicity by preventing exposure to ineffective treatment. Critical to balancing these goals is the ability to quickly identify progression in order to minimize exposure to ineffective therapies. Imaging is the current standard-of-care for disease response evaluation, but high cost, relative low sensitivity, and tumor flare phenomenon remain significant issues. Circulating tumor DNA (ctDNA) testing based on next generation sequencing (NGS) is emerging as an alternative to imaging-based response assessment, but remains an expensive proposition. Here we evaluate a hybrid monitoring strategy using an initial baseline NGS-based ctDNA analysis to identify mutant alleles, followed by response monitoring with a low-cost, personalized digital droplet PCR (ddPCR) assay. Methods: Blood for ddPCR ctDNA analysis was collected at the same timepoint as a sample for commercial ctDNA analysis by NGS. PCR primer and probe sets were designed against clinically relevant mutations identified from each patient’s commercial ctDNA NGS report. ddPCR was then performed. Variant allele fractions (VAFs) detected by ddPCR were compared to allelic fractions reported by commercial NGS using Bland-Altman analysis and significance was determined by paired t-test. To demonstrate the potential for longitudinal ddPCR-based ctDNA monitoring, serial samples from the same patient were analyzed. Results: In our pilot validation set, 10 paired patient samples were analyzed by NGS and ddPCR. NGS-reported VAFs ranged from 0.30% to 16.20%. High comparative performance was observed between NGS and ddPCR platforms over expected mutant fractional abundance ranges. There was no significant quantitative difference between the two approaches (p=0.45). Longitudinal monitoring of ctDNA by ddPCR was performed on a metastatic hormone-positive breast cancer patient and revealed a significant rise in the clinically-relevant PIK3CA H1047L mutation that preceded imaging-based documentation of progression by 3 months. A final timepoint was taken several months before the patient’s death with a further significant increase in ctDNA burden. Conclusions: Here we demonstrate that highly-sensitive ddPCR-based ctDNA assays provide comparable results to next generation sequencing, but at considerably lower cost. These results indicate that bespoke tracking of select alleles by ddPCR offers an inexpensive and rapid approach to serial ctDNA monitoring in metastatic breast cancer patients. A larger prospective study has been designed and is underway. Citation Format: Nicole Higashiyama, LaTerrica Williams, Bryant McCue, Alphi Kuriakose, Tiffaney Tran, Stephanie Gonzalez, Mayra Licerio, Carol Chenault, Heidi Dowst, Susan Hilsenbeck, Matthew Ellis, Bora Lim, George Miles. Longitudinal circulating tumor DNA (ctDNA) monitoring by digital droplet PCR (ddPCR) in metastatic breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-01-13.