Sulfamoylation of 2-methoxyestrone (2-MeOEI) was shown previously to enhance its potency as an anti-proliferative agent against breast cancer cells. We have examined the ability of a series of 2-methoxyestradiol (2-MeOE2) and 2-ethylestradiol (2-EtE2) sulfamates to inhibit angiogenesis in vitro. 2-MeOE2 bis-sulfamate and 2-EtE2 sulfamate were potent inhibitors of human umbilical vein endothelial cell (HUVEC) proliferation with IC50 values of 0.05 muM and 0.01 muM, respectively. A novel co-culture system, in which endothelial cells were cultured in a matrix of human dermal fibroblasts, was also used to assess the anti-angiogenic potential of these drugs. In this system endothelial cells proliferate and migrate through the culture matrix to form tubule structures. Whereas 2-MeOE2 (1.0 muM) caused a small reduction in tubule formation, both 2-MeOE2 bis-sulfamate (0.1 muM) and 2-EtE2 sulfamate (0.1 muM) almost completely abolished tubule formation. 2-MeOE2 bis-sulfamate and 2-EtE2 sulfamate both induced BCL-2 phosphorylation, p53 protein expression and apoptosis in HUVECs. Microarray analysis of a limited number of genes known to be involved in the angiogenic process did not show any gross changes in cells treated with the 2-substituted estrogens. The sulfamoylated derivatives of 2-MeOE2 and 2-EtE2 are potent inhibitors of in vitro angiogenesis and both compounds should have therapeutic potential. (C) 2004 Wiley-Liss, Inc.