Abstract PS5-08-21: Prospective breast cancer clinical validation study of an ultrasensitive, tumor-informed, whole genome, circulating tumor DNA assay to detect molecular residual disease and predict recurrence of high-risk early breast cancer treated with standard (neo)adjuvant therapy; NSABP B-64/EXActDNA/003/NCT06401421

Abstract Background: Detection of occult micrometastatic cancer—molecular residual disease (MRD)—in patients following treatment for high-risk phenotypes of early breast cancer using circulating tumor DNA (ctDNA) is associated with a high risk of recurrence. An ultra-sensitive, tumor-informed, whole-genome, minor-allele-enriched sequencing through recognition oligonucleotides (MAESTRO) assay has previously demonstrated a strong association with residual cancer burden and disease recurrence in an exploratory study in a subset of patients with triple-negative breast cancer (TNBC) treated in a Phase II study in the neoadjuvant setting.1 Here, we describe a large, prospective registry trial in progress designed to: (Part 1) assess the feasibility of using material from formalin-fixed paraffin-embedded (FFPE) diagnostic breast biopsies to create a bespoke MRD assay using the MAESTRO technology, and (Part 2) validate the association between ctDNA status and distant recurrence-free interval (dRFI) in 3 cohorts of patients with early breast cancer and high-risk phenotypes (TNBC, human epidermal growth factor receptor 2-positive [HER2+] inclusive of hormone-receptor positive and -negative [HR+ and HR−], and HR+/HER2-negative [HER2−] tumors) receiving neoadjuvant chemotherapy co-administered with standard targeted therapies, surgery, and standard adjuvant therapies post-surgery. Methods: Trial eligibility criteria include: age ≥18 years, ECOG performance status 0−1, histologically confirmed invasive breast carcinoma, primary tumor size >2.0 cm, planned neoadjuvant chemotherapy, and either clinically node-positive disease, or if node-negative, one of the following: TNBC or HER2+ subtype, or HR+/HER2− subtype with high-risk features. Target enrollment is 1,800 participants, based on the number needed in each of the high-risk subtypes to observe at least 30 events within 3 years. Analyses will be conducted across the 3 subtypes in Part 1 and by breast cancer subtype in tissue-evaluable participants in Part 2 of the trial, once at least 30 distant recurrences have been reported in each subtype. Patients will receive standard therapies per investigator discretion and will be followed for at least 5 years. Serial blood samples will be collected during treatment and follow-up, and analyzed for ctDNA by Exact Sciences, blinded to clinical data. Results of ctDNA analyses for each collection timepoint will be provided to the National Surgical Adjuvant Breast and Bowel Project (NSABP) Foundation statistician to merge with clinical outcomes data and analyze the endpoints. In Part 1, the primary endpoint is the proportion of participants for whom FFPE diagnostic biopsy material is sufficient to create a patient-specific ctDNA assay. In Part 2, the primary endpoint is dRFI with 2 primary objectives, which will be analyzed sequentially: (1) the association of the ctDNA-positive status at any time in serially collected blood samples (starting from the completion of cytotoxic and targeted therapy) and dRFI, and (2) the association of the ctDNA-positive status as a single timepoint at the end of cytotoxic and targeted therapy with dRFI. Secondary endpoints that will be analyzed for association with ctDNA status include pathologic complete response, recurrence-free interval, invasive breast cancer-free survival, event-free survival, and overall survival. Clearance of ctDNA on therapy will also be assessed and correlated with outcomes. ClinicalTrials.gov Identifier: NCT06401421. References: 1. Parsons HA et al. Ann Oncol. 2023 Oct;34(10):899-906. Citation Format: M. Basik, E. Diego, G. Tang, S. Puhalla, M. Balic, M. Mandadi, J. Foldi, S. Seaward, M. George, A. Shipstone, S. Vemulapalli, J. Jones, W. J. Irvin Jr, R. L. Mahtani, V. Valero, J. Frederick, B. Arrick, G. Young, J. A. Grigorieva, M. R. Palomares, F. Baehner, P. Rastogi, E. P. Mamounas, N. Wolmark, C. E. Geyer Jr. Prospective breast cancer clinical validation study of an ultrasensitive, tumor-informed, whole genome, circulating tumor DNA assay to detect molecular residual disease and predict recurrence of high-risk early breast cancer treated with standard (neo)adjuvant therapy; NSABP B-64/EXActDNA/003/NCT06401421 [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 PS5-08-21.