Abstract 5595: Serial monitoring of circulating tumor cells and circulating tumor DNA in metastatic lobular breast cancer identifies intra-tumor heterogeneity and precision and immuno-oncology biomarkers of therapeutic importance
Authors: Cani Andi K. , Dolce Emily M. , Hu Kevin , Liu Chia-Jen , Darga Elizabeth P. , Robinson Dan , Wu Yi-Mi , Thomas Dafydd G. , Paoletti Costanza , Tomlins Scott A. , Rae James M. , Udager Aaron M. , Chinnaiyan Arul M. , Cobain Erin F. , Hayes Daniel F. 
Published: June 2023

Abstract
Precision and immuno-oncology clinical decisions are based on predictive biomarkers. Circulating biomarkers offer a minimally invasive approach to monitor intra-patient tumor heterogeneity and detect in real-time the clinically-relevant evolving clonal architecture. Although currently underutilized, single-cell DNA next generation sequencing of circulating tumor cells (CTC) is a particularly suitable method to complement tissue and circulating tumor DNA (ctDNA). Here we analyzed 113 individual CTC, 21 ctDNA, 15 white blood cells (WBC) samples and 15 tissue biopsies, from 15 CTC-positive lobular breast cancer patients. CTC were enriched with the CellSearch® system and isolated as single cells with the DEPArray™ system. CTC, WBC, and ctDNA underwent scNGS with the Oncomine Comprehensive Assay covering ~500 genes and 1.1Mb of genomic space to detect mutations, copy number alterations, tumor mutation burden (TMB) and microsatellite instability (MSI). 99.1% of single cells and 95.2% of ctDNA samples were informative. Our CTC-based precision medicine reporting platform, MI-CTCSeq, detected CTC in 9 of 15 patients (60%) with FDA-approved actionable alterations including in the oncogenes PIK3CA and FGFR2 and HER2. 3 of these 9 (33%) harbored actionable alterations not shared between all 3 analyte types (tissue, CTC and ctDNA) including 3 actionable mutations found in CTC and ctDNA only, 1 in tissue and ctDNA only, and 1 in ctDNA only. Two of those ctDNA mutations were identified near the limit of detection and with a priori knowledge from tissue or CTC. Interestingly, 1 patient with plentiful CTC had no detectable ctDNA. Another patient’s tissue biopsy was inadequate for sequencing while both liquid biopsy analytes were abundant. 13 patients (87%) displayed intra-patient, inter-CTC genomic heterogeneity of driver mutations. 1 of 4 (25%) patients with CTC available in &gt;1 timepoint displayed fluctuations in their CTC subclonal makeup between timepoints. Data from this patient’s 2 tissue biopsies, 4 ctDNA samples, and 27 individual CTC over 6 timepoints, combined to reveal in unprecedented detail inter-metastatic lesion and inter-CTC heterogeneity and evolution in response to endocrine, chemo and immunotherapy selective pressures. In a novel approach we show detection of single-cell CTC TMB and MSI. CTC TMB was highly concordant (R 0.81) with the corresponding tissue biopsies. Further, in a novel observation, we detected intra patient, inter-CTC heterogeneity of TMB and MSI, which has potential implications for immunotherapy response and development of resistance. Taken together, these data support the non-invasive biomarker interrogation and monitoring by liquid biopsy that incorporates CTC to complement tissue in informing treatment approaches.
Citation Format: Andi K. Cani, Emily M. Dolce, Kevin Hu, Chia-Jen Liu, Elizabeth P. Darga, Dan Robinson, Yi-Mi Wu, Dafydd G. Thomas, Costanza Paoletti, Scott A. Tomlins, James M. Rae, Aaron M. Udager, Arul M. Chinnaiyan, Erin F. Cobain, Daniel F. Hayes. Serial monitoring of circulating tumor cells and circulating tumor DNA in metastatic lobular breast cancer identifies intra-tumor heterogeneity and precision and immuno-oncology biomarkers of therapeutic importance. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5595.