Endothelial cell-selective adhesion molecule (ESAM) is a member of the immunoglobulin receptor family that mediates homophilic interactions between endothelial cells. To address potential in vivo angiogenic functions of this molecule, mice lacking ESAM (ESAM-/-) were generated by gene-targeted deletion. ESAM-/- mice did not show overt morphological defects in the vasculature. To evaluate the role of ESAM in pathological angiogenesis, wild type (WT) and ESAM-/- mice were injected with melanoma and Lewis lung carcinoma cells. By 14 days after injection, tumor volumes of B16F10 and LL/2 in ESAM-/- mice were 48 and 37% smaller, respectively, compared with WT mice. Vascular density of the tumors, as determined by CD31 staining, was also decreased in the ESAM null animals. Matrigel plug assays showed less neovascularization in ESAM-/- mice than in WT mice. ESAM-/- endothelial cells exhibited less in vitro tube formation and decreased migration in response to basic fibroblast growth factor when compared with WT cells, and endothelial-like yolk sac cells engineered to overexpress ESAM showed accelerated tube formation in vitro. These in vitro and in vivo studies suggest that ESAM has a redundant functional role in physiological angiogenesis but serves a unique and essential role in pathological angiogenic processes such as tumor growth.