Inhibition of the HIV-1 nuclear regulatory protein tat could potentially yield particularly useful drugs because it functions as an activator of transcription. It has no known cellular counterpart, and deletions in the tat gene destroy the ability of HIV-1 to replicate. We recently reported that a structurally unique class of tat inhibitors, 3-keto/enol 4,5-alpha-epoxy steroids bearing electron-withdrawing substituents at position 2, specifically inhibit tat-induced gene expression in virus free transfected SW480 cells. In this paper, we report on additional SAR (structure-activity relationships) for the steroid series and the localization of the pharmacophore to the A-ring functionality. There is a weak enantioselective preference for the natural steroid stereochemistry and hints of additional SAR in the electron-withdrawing group. Compound 34a is of particular interest in that it inhibits HIV replication in H9 cells at a concentration equivalent to its inhibitory level in the primary tat assay.