Abstract RF4-03: Evaluation of a whole-exome sequencing tumor-informed circulating tumor DNA MRD assay in patients with early triple-negative breast cancer (TNBC) receiving neoadjuvant chemotherapy (NAC) with or without atezolizumab: A prospective sub study of the NSABP-B59/GBG-96-GeparDouze Trial

Abstract Background: Detection of occult micro-metastatic cancer—molecular residual disease (MRD) —in patients following treatment for early triple-negative breast cancer (TNBC) using circulating tumor DNA (ctDNA) is associated with a high risk of recurrence. Dynamics of ctDNA clearance in response to neoadjuvant therapy (NAT) has also shown potential for predicting pathologic compete response (pCR) and potentially improving the prognostic ability of pCR status. NSABP B-59/GBG-96-GeparDouze was a phase III placebo-controlled trial which evaluated the efficacy and safety of adding atezolizumab or placebo to NAT followed by atezolizumab or placebo as adjuvant therapy to complete 1 year of therapy in 1, 550 patients with Stage II/III TNBC. A sub-study collected serial blood samples for ctDNA analysis during the first 2 years following randomization, which were analyzed with a whole-exome sequencing of the primary tumor with variants selection informed by parallel sequence analysis of the patient’s normal germline DNA obtained from peripheral blood mononuclear cells. Patient-specific bespoke assays with 50-200 variants were used to assess each available sample for presence or absence of ctDNA. Methods: This prospective sub-study included patients enrolled within the NSABP B-59/GBG-96-GeparDouze trial (Clin Cancer Res 2025 31 12Suppl: GS3-05) following an amendment to incorporate serial ctDNA collections at baseline prior to neoadjuvant therapy (NAT), immediately before surgery, 3-6 weeks post-surgery, 12-months after randomization, 24-months after randomization, and at recurrence. The primary objective of this sub-study was to demonstrate the association between a positive ctDNA result at the post-surgery timepoint and distant recurrence-free interval (DRFI), defined as time from surgery to first occurrence of distant metastasis. The log-rank test was used to compare DRFI between patients with and without ctDNA detection post surgery, with the lag between surgery and blood sample test considered as delayed entry. The Cox proportional hazards model was used to assess the strength of relationship between ctDNA detection and risk of distant recurrence. Secondary endpoints included recurrence-free interval (RFI) and pCR. Results: The mean age of the patients in the sub study was 50, 78. 9% had high tumor grade, 61. 2% were node positive at presentation, and 51% received atezolizumab. 160 patients with baseline blood samples had sufficient DNA yield to perform WES for development of the bespoke assay for ctDNA. We detected ctDNA at presentation in 153 (96%) of these patients. At the end of NAT prior to surgery, 155 blood samples were available and ctDNA was detected in 14 (9. 0%) indicating substantial clearance on the NAT. Following surgery, 147 blood samples were available to assess the primary endpoint of DRFI. Median follow-up after surgery was 37 months. Distant recurrences had developed in 14 patients (9. 5%). Following NAT and surgery, 138 (93. 8%) were ctDNA negative, and of those 131 (95%) were free of distant recurrence. Among the 9 patients who were ctDNA positive (MRD+) after surgery, 7 developed distant recurrences (log-rank p1E-16, HR=30. 3, 95% CI=10. 4, 88. 7). Similar finding were observed in RFI. Conclusions: Detection of ctDNA (MRD+) using a bespoke MRD assay was highly prognostic for distant recurrence in TNBC patients following neoadjuvant therapy and surgery. The assay also demonstrated a high prevalence of ctDNA at presentation with substantial clearance on the chemotherapy regimen +/- atezolizumab employed in the trial. Citation Format: M. Balic, G. Tang, G. Young, P. Rastogi, J. Acosta, A. Schneeweiss, C. Hilton, T. Freeman, C. Denkert, M. Reinisch, M. R. Palomares, A. A. Bradley, S. Murali, J. Boileau, D. Boudreau, P. J. Polewski, S. Hassan, J. Mouta, W. C. Darbonne, F. L. Baehner, E. P. Mamounas, N. Wolmark, S. Loibl, C. E. Geyer Jr. Evaluation of a whole-exome sequencing tumor-informed circulating tumor DNA MRD assay in patients with early triple-negative breast cancer (TNBC) receiving neoadjuvant chemotherapy (NAC) with or without atezolizumab: A prospective sub study of the NSABP-B59/GBG-96-GeparDouze Trial abstract. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32 (4 Suppl): Abstract nr RF4-03.