Background: Human epidermal growth factor receptor 3 (HER3) has been found upregulated in a wide variety of cancers and has been associated with disease progression and resistance to EGFR-targeted therapies in HER2-positive malignancies. Several studies have reported expression of HER3 in prostate cancer (PCa). Furthermore, concurrent expression of HER2 and HER3 in PCa has been associated with androgen resistance, development of metastatic disease, and less favorable outcomes. However, due to conflicting results, the exact nature of the relationship between HER2 and HER3 in PCa requires further investigation. Methods: 194 patients diagnosed with PCa between 2000-2021 at the Washington DC Veterans Affairs Medical Center who had adequate formalin-fixed paraffin embedded prostate tissue available were selected from our previously published cohort of patients. HER3 immunostaining was performed and independently scored by two pathologists as 0 (absent), 1+ (weak), 2+ (moderate), or 3+ (marked). HER2 immunostaining data and clinical information was retrieved. Statistical analysis included Fisher’s exact test and linear regression models. Results: In our self-identified predominantly African American (85%) patient cohort, a 94% overall prevalence of HER3 positivity (1+, 2+, or 3+) was observed. Notably, the majority of positive patients was found to have HER3 expression of either 2+ (~60%; 116/194) or 3+ (~17%; 33/194). HER3 positivity was associated with a higher PSA (p=0.023), a higher Grade Group (GG) (p=0.017), and a higher AJCC disease stage (p=0.006) at time of diagnosis (Table 1). A significant association was observed between positivity for both HER2 and HER3 (p=0.031). HER3 positivity independently predicted GG (p=0.02) and AJCC disease stage (p=0.01) when adjusted for HER2 positivity. Conclusions: In this predominantly African American cohort, a high prevalence of HER3 positivity was observed across PCa disease stages. Higher expression levels of HER3 were observed in more advanced stages of the disease. Because of the observed efficacy of HER3-antibody drug conjugates in other HER3-expressing malignancies, the high prevalence of HER3 in PCa makes it an attractive therapeutic target that warrants further evaluation.