Abstract Use of prognostic assays and clinical features to further risk stratify patients with hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) breast cancer has become standard of care. Stratification of patients into low/mid risk of disease recurrence enables physicians to decide which patients can safely forgo chemotherapy. Yet there exists a subpopulation of patients who tend to have recurrences following endocrine therapy alone. Complicating the issue, recent data has shown these proliferation-based markers assessed from a single site tissue biopsy may not be reliable for minority populations, potentially owing to spatial heterogeneity of tumors. To address this, we used a novel 2-paramenter pharmacokinetic modeling framework that allows biosignatures to be extracted from dynamic contrast enhanced (DCE) magnetic resonance imaging (MRI) studies that contain 3-6 timepoints spaced 60-90 seconds apart. These parameters, referred to as P1 and P2, represent leakiness from vessels into the extravascular space and vice versa. This approach was previously developed in a study of 111 breast cancer patients where the 21-gene recurrence score and DCE-MRIs were available. Low P1 showed better outcomes in low- and mid- patients (n=88, p≤0.028; log-rank test). Patients with a high P1 had a 20.2% chance recurrence at six years. The same trend was observed in mid-recurrence score patients only (n=23, p≤0.058). No recurrences were observed in patients with low P1 in either the RS-low or RS-mid categories. There were no recurrences in the high P1, RS-low/-mid category that received chemotherapy, suggesting that chemotherapy could be beneficial in this category of patients, although the trend was not statistically significant (n=10, p=0.46). Here we present an independent, single site validation of these prognostic markers in patients who received only neoadjuvant endocrine therapy and had corresponding pre-treatment MRIs. Two hundred ninety-eight patients with early-stage breast cancer and pre-treatment MRIs were identified via chart review. Of the patients assessed, 33 patients were treated with neoadjuvant endocrine therapy and qualified for the analysis. Consistent with the previous analysis, the 5-year event-free survival rate in low P1 population was 100% while that in the high P1 population was 45.8% (p=0.053). Overall, we find strong support that these markers could help physicians further fine tune their decision-making when determining who to forgo chemotherapy. Citation Format: Joseph Peterson, Jose Rubio-Romera, Georgia Giakoumis Spear, Michele Britto, Bradley Hack, Tushar Pandey, Poornima Saha. Validation of prognostic platform to further refine identification High Risk Patients indicated for Chemotherapy Free Treatment in Early-Stage Breast Cancer abstract. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P6-01-36.