Background: Nectin-4, a cell adhesion molecule found in a frequently amplified region of the bladder cancer (BC) genome, has been recently used as target for the systemic treatment of locally advanced or metastatic BC. BC with variant histology (BCVH) is associated with poor clinical outcomes, and there is an unmet need for novel treatments in this population. Little is known about Nectin-4 expression and other targets. We investigated Nectin-4 and programmed death-ligand 1 (PD-L1) expression in BCVH. Methods: We performed a retrospective analysis of 117 patients with BCVH whose samples were collected at Winship Cancer Institute of Emory University from 2011-2021. Immunohistochemistry (IHC) was performed for Nectin-4 and PD-L1 expression on formalin-fixed, paraffin-embedded (FFPE) tissue. Nectin-4 staining was performed by Q2 Solutions using non-commercially available anti-Nectin-4 antibody (clone M22-312B41.1). Expression was measured using the H-score method (H-score = [% of strong positive tumor cells × 3] + [% of moderate positive tumor cells × 2] + [% of weak positive tumor cells × 1]). PD-L1 levels were assessed in tumor-infiltrating immune cells using DAKO 22C3 IHC to determine PD-L1 high, defined by a Combined Positive Score (CPS) of ≥10. Results: Median age was 70 years (mean=67.2, range 22-91), with 61.5% male, 54.7% white, and 32.5% black. Nearly all samples were taken from the bladder (95.7%), with 4.3% from sites of metastasis. The most common histologic variant was squamous cell carcinoma (26.5%), and the distribution of variants is summarized in table. Nectin-4 staining was successfully performed on 111 samples, and PD-L1 staining on 116. Nectin-4 expression was seen across different histological subtypes (mean H-score: 153.1, median [range]: 157 [0-300]), with highest expression in plasmacytoid and squamous, and lowest expression in small cell and sarcomatoid (Table). More than one-third (35.3%) were PD-L1 high, and greater in sarcomatoid and squamous cell histology (70.8% and 50% positivity, respectively). Only 5% of plasmacytoid samples were PD-L1 high. Conclusions: Although there is heterogeneity, our results showed expression of Nectin-4 and PD-L1 in this cohort of patients with BCVH. These findings highlight the therapeutic potential of antibody-drug conjugates against Nectin-4- and anti PD1/PD-L1-based therapy in this population. Additional analysis including clinical outcomes and genomic analyses are ongoing, and a prospective clinical trial is planned.