Multiplexed Immunofluorescent Characterization of <scp>T</scp> ‐Cell and Myeloid Cell Populations Reveals Histologic Subtype Differences Within the Immunosuppressive Landscape of Human Basal Cell Carcinoma

BACKGROUND: Basal cell carcinomas (BCCs) are thought to reside within an immunosuppressed tumor microenvironment, but a detailed characterization of BCC-associated immune cells was lacking. METHODS: Utilizing multiplex immunohistochemistry (IHC) (Vectra Polaris, Akoya Biosciences), we designed two staining panels, targeting either T cells (CD3, CD4, CD8, FoxP3, PD1) or myeloid cells (neutrophil elastase, CD68, CD163, HLADR, CD1c). Formalin-fixed paraffin-embedded (FFPE) BCC tumor specimens (n = 30) were analyzed using the panels. The majority of immune cells reside at the peritumoral margin; therefore, a boundary zone of 150 μm around tumor nests was used to capture them. RESULTS: A CD8/T-reg ratio and M1/M2 macrophage ratio of < 0.5 was observed across all BCC specimens, confirming an overall immunosuppressive phenotype. Among different histologic subtypes, infiltrative BCC had significantly lower CD8 T cells (p = 0.038), T-regulatory cells (p = 0.039), and CD8/T-reg ratio (p = 0.048), as well as lower M1/M2 ratio (p = 0.034) compared to non-infiltrative BCC subtypes. CONCLUSIONS: The association of infiltrative BCC with a more immunosuppressed microenvironment may contribute to more aggressive biological behavior. Overall, these data demonstrate a method to determine T cell and myeloid profiles in fixed FFPE skin tumor tissues that can identify subtle immune profile differences.