The development of potent D-peptide antagonists of the HIV-1 envelope protein gp41 was shown to prevent viral fusion and entry into cells. A trimeric version of one of the isolated candidates could block pocket-specific viral entry with an IC50 as low as 250 pM. Further pharmacokinetic optimization and synthetic scaleup yielded the cholesterol-conjugated trimeric D-peptide CPT31, which is currently in Phase Ia clinical trials for the treatment of HIV.