B7-H3 expression in Merkel cell carcinoma-associated endothelial cells correlates with locally aggressive primary tumor features and increased vascular density

Purpose: Merkel cell carcinoma is an aggressive cutaneous malignancy whose pathogenesis and prognosis are related to the integrity of the host immune system. Despite promising clinical responses to immune checkpoint blockade, response and resistance remain unpredictable, underscoring a critical need to delineate novel prognostic biomarkers and/or therapeutic targets for this disease. Experimental Design: Expression of immune regulatory markers (PD-L2, B7-H3, B7-H4, IDO-1, ICOS, TIM3, LAG3, VISTA and OX-40) was assessed using singlet chromogenic immunohistochemistry in 10 primary MCCs. Multiplex immunofluorescence (mIF) quantified CD31 and B7-H3 expression in 52 primary and 25 metastatic MCCs. B7-H3 and CD31 expression were tabulated as a series of independent (X,Y) cell centroids. A spatial G-function, calculated based on the distribution of distances of B7-H3+ (X,Y) cell centroids around the CD31+ (X,Y) cell centroids, was used to estimate a co-localization index equivalent to the percentage of CD31-positive cell centroids that overlap with a B7-H3-poisitive cell centroid. Results:Primary and metastatic MCC exhibit a dynamic range of co-localized CD31 and B7-H3 expression. Increasing co-localized expression of B7-H3 with CD31 significantly associated with increased tumor size (p=0.0060), greater depth of invasion (p=0.0110), presence of lymphovascular invasion (p=0.0453) and invasion beyond skin (p=0.0428) in primary MCC. Consistent with these findings, increasing co-localized expression of B7-H3 and CD31 correlated with increasing vascular density in primary MCC, but not metastatic MCC. Conclusions:Our results demonstrate that co-localized expression of B7-H3/CD31 is a poor prognostic indicator and suggest therapies targeting B7-H3 may represent an effective approach to augmenting immune-activating therapies for MCC.