1235 Endothelial AMBRA1 and Claudin5 as potential biomarkers of metastases in early-stage melanoma

The loss of AMBRA1 and Loricrin (AMBLor) in the epidermis overlying nonulcerated early-stage primary melanomas has recently been identified as a prognostic biomarker and is mediated by tumoral secretion of TGFβ2 (Ellis et al 2020, Cosgarea et al 2021). Pilot data shows loss of epidermal AMBRA1 overlying primary melanomas is also associated with loss of expression in the surrounding tumor endothelium, suggesting endothelial AMBRA1 expression may define tumors at risk of metastasis. The aim of the present study was to evaluate endothelial AMBRA1 expression as a potential companion biomarker for personalized adjuvant treatment, and to define the mechanisms mediating its loss. Semi-quantitative automated immunohistochemical analysis of AMBRA1 expression in FFPE sections derived from a cohort of 47 non-ulcerated AJCC stage I/II melanomas, with loss of AMBLor in the peri-tumoral epidermis, revealed variable AMBRA1 expression levels in the endothelium of vessel sub-populations within intra- and/or peri-tumoral locations, possibly reflecting tumor and microenvironment heterogeneity. Treatment of human umbilical vein endothelial cells (HUVEC) cultured on an extra-cellular matrix with recombinant TGFβ2 resulted in reduced tubule formation and expression of the tight junction protein Claudin5, but no change in AMBRA1 expression. Further investigation of TGFβ family ligands revealed treatment of HUVECs with BMP7 for 72 hours resulted in a small reduction in AMBRA1 expression. Collectively these data suggest the reduction of endothelial AMBRA1 and Claudin5 expression contribute to the loss of vascular integrity but are mediated through different ligands of the TGFβ superfamily. Endothelial AMBRA1/Claudin5 expression may therefore serve as early biomarkers of metastasis for high-risk early-stage cutaneous melanomas.