Abstract B019: Clinical and molecular correlates of circulating tumor DNA (ctDNA) dynamics in breast tumors resistant to neoadjuvant therapy (NAT)

Abstract Purpose: We examined the utility of serial ctDNA testing for refining risk stratification of NAT- resistant tumors (RTs, defined as residual cancer burden, RCB-II/III) and predicting early treatment response. We also characterized the molecular correlates of ctDNA dynamics in RTs to elucidate the mechanisms associated with increased metastatic potential. Methods: We performed serial ctDNA testing using a tumor-informed personalized assay (SignateraTM, Natera Inc.) in 723 patients (pts) with high-risk early-stage breast cancer receiving NAT (I-SPY2). Tumors were stratified according to RCB class and ctDNA dynamics (clearance patterns), and the distant recurrence-free survival (DRFS) between groups was compared. We examined the predictive value of ctDNA dynamics using the RCB index (the continuous measure of RCB) as the response endpoint. Serial whole exome sequencing (WES), gene expression profiling (GEP), and reverse-phase protein analysis (RPPA) data were analyzed to elucidate the molecular biology of RTs. Results: Of the 723 pts, 300 (41%) had hormone receptor (HR)+HER2-, 237 (33%) triple-negative (TN), and 186 (26%) HER2+ breast cancers with pretreatment ctDNA-positivity rates of 76%, 92%, and 77%, respectively. 321 (45%) pts had responding tumors (RCB-0/I), and 55% had RTs: RCB-II (n=255) and RCB-III (n=132). CtDNA dynamics revealed heterogeneity in the metastatic potential of RTs, including tumors with a reduced propensity to metastasize (persistent ctDNAneg and early clearance). Failure to clear ctDNA after NAT was associated with an increased risk of metastatic recurrence and death (3-yr DRFS rates: no clearance=35% vs. persistent ctDNAneg=98%, p<0.001). Early clearance of ctDNA, 3 weeks after treatment initiation, skewed the distributions toward lower RCB indices (favorable response), including in arms containing immune checkpoint inhibitors. Of the 387 RTs, 316 had ctDNA data post-NAT, of which only 51 (16%) were ctDNA+. We hypothesized that changes in the mutational landscape during NAT could have impacted the detection of patient-specific ctDNA targets selected from WES of pretreatment tumors. However, serial WES (n=100) of RTs showed that these targets remained detectable in solid tissue over time, even in persistent ctDNAneg. To investigate the biology of persistent ctDNA positivity, we used logistic regression to analyze serial GEP (n=174) and RPPA (n=98) data in RTs that were ctDNA+ at pretreatment, comparing those with ctDNA clearance by surgery vs. those that remained ctDNA+. We found subtype- specific associations: In HR+HER2-, failure to clear ctDNA during NAT was associated with low pretreatment ER/AR, high early on-treatment proliferation, and low HER2-signaling after NAT, while in TN, fewer associations were observed; these included high pretreatment levels of MYC and proliferation. Conclusions: ctDNA dynamics refined risk stratification of RTs and enabled early prediction of treatment response. Understanding the molecular profiles of RTs based on ctDNA dynamics could aid treatment selection to overcome resistance to NAT. Citation Format: Mark Jesus M. Magbanua, Denise M. Wolf, Samuel Rivera-Hinojosa, Ziad Ahmed, Nayelis A. Manon, Rosalyn Sayaman, Antony Tin, Ekaterina Kalashnikova, Lamorna Brown Swigart, Gillian L. Hirst, Christina Yau, Wen Li, Claudine Isaacs, Rebecca A. Shatsky, Amy L. Delson, Charuta C. Palsuledesai, Angel Rodriguez, Minetta C. Liu, Paula R. Pohlmann, Laura J. Esserman, Hope S. Rugo, Angela DeMichele, Laura van 't Veer. Clinical and molecular correlates of circulating tumor DNA (ctDNA) dynamics in breast tumors resistant to neoadjuvant therapy (NAT) [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr B019.