Abstract Afro-Brazilian women suffer from higher excess breast cancer mortality after adjusting for age, level of education and stage at diagnosis. Limited studies have researched disease characteristics in this population, which can benefit from the identification of molecular markers that can provide an insight into who is at greater risk of mortality. Epigenetic biomarkers represent the combined effect of environmental and genetic factors. Particularly DNA methylation biomarkers can be valuable as biomarkers of breast cancer risk and prognosis. While studies in other populations have identified DNA methylation biomarkers associated with breast cancer and its clinicopathological outcomes, research on Brazilian cohorts has been limited in scope and focused on women of European descent. The goal of this work was to determine the epigenetic makeup of breast cancer in Brazilian women of African descent. To identify epigenetic biomarkers in Brazilian women who self-identified as Black or Brown, we conducted a case-series analysis investigating DNA methylation patterns in tumor (T) and adjacent non-tumor (NT) tissue. For this, we used the Illumina EPIC (850k) arrays to carry out DNA methylation profiling in 48 frozen tissue paired T and NT samples of breast cancer patients treated at the National Cancer Institute Breast Cancer Hospital in Rio de Janeiro, Brazil. We identified a total of 7,749 probes with Benjamini-Hochberg adjusted p-value < 0.0001 and differences between T and NT tissue above |0.3|. Of these, 5,976 or 77.12% had lower DNA methylation in T when compared to NT tissue; while 1,773 (22.88%) had higher DNA methylation. These were used to identify 21 differentially methylated regions (DMRs) that contained a minimum of seven cg sites, a mean difference larger than |0.2| and a maximum difference above |0.4|. DMRs were mostly (12) hypermethylated in T, and included PCDHGA, NKAPL, ZKSCAN4, SPAG6, CA3, DPP6, RGS22, NXPH1, ANK3, OTX2, SOX17, and CDO1. We used pyrosequencing to validate the levels of SOX17 in eighty-eight tumor and non-adjacent tumor tissue DNAs. We found SOX17 to be hypermethylated in T (44.5±19.1% vs 10.2±10.6, p<0.0001). Differences in DNA methylation in T and NT tissue we still observed by race/ethnicity categories, 43.9±18.6% vs 11.0.±13.2% (p<0.0001) for Brown, and 45.1±20.7% vs 8.8±3.0% for Black women (p<0.0001), respectively. Similarly to previously reported, SOX17 hypermethylation was found to be associated with Luminal A and B molecular subtypes (p<0.0001). To our knowledge, this is the first study to use a discovery approach to identify epigenetic biomarkers in Black and Brown Brazilian women. Our findings confirming previously reported data on SOX17 hypermethylation in breast cancer are encouraging. We expect to validate other epigenetic markers unique to this population that can ultimately aid in the molecular characterization of Brazilian breast cancers. Citation Format: Lissette Delgado-Cruzata, Diego J. Gomes de Paula, Jennifer G. Vieira, André L. Christianes, Tatiana Simao, Leonor B. Gusmao, Luis Felipe Ribeiro Pinto, Sheila Coelho Soares Lima. Hypermethylation of SOX17 is an important epigenetic mark in Afro-Brazilian breast cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5207.