Development of methodology for the synthesis of stereochemically pure Phe.psi.[CH2N]Pro linkages in HIV protease inhibitors

Abstract

One of the strategies currently being pursued for the design of potential HIV protease inhibitors involves the replacement of the cleaved amide bond in a minimum peptide substrate with an aminomethylen? unit. A commonly used method for the synthesis of these compounds involves a reductive alkylation of an amine with an aldehyde in the presence of sodium cyanoborohydride under acidic conditions. Accordingly, BOC-phenylalaninal(4) was reacted with the peptide-resin ProIleSer(OBz1)OResin (5) in the presence of acetic acid and sodium cyanoborohydride. The resulting product was found to consist of a mixture of diastereomers, which may result from the fact that the proline residue, which contains a secondary amine, reacts with the aldehyde to form an enamine 9 with loss of chirality at the modified Phe residue. Subsequent reduction of the iminium ion 10 would then result in production of the observed two diastereomers. In order to circumvent this problem, BOCPheProOBzl (12b) was synthesized and the central amide bond was reduced selectively with diborane. Hydrogenolysis of the benzyl protecting group gave BOCPhe9[CH2N]Pro (14a), which was coupled manually to the peptide resin IleSer(OBz1)OResin to give BOCPhe9[CH2N]ProIleSer(OBzl)OResin (6). Subsequent addition of amino acid residues to 6 and cleavage from the resin gave a series of stereochemically defined potential HTV protease inhibitors as single diastereomers. The most potent of these substances was ThrLeuksnPhe9[CH2N]ProIleSer (1)) which displayed an ICw of 1.1 pg/mL (1.4 pM) when tested for inhibition of HIV-1 protease. However, the epimer of 1 having the opposite configuration at the reduced Phe residue was inactive. A minimum length of seven amino acid residues appears to be necessary for effective recognition of the inhibitor by the enzyme. Further increase in chain length did not result in greater inhibitory potency. The replication competent retroviruses, including HIV, encode a protease which cleaves the gag and pol precursor polyproteins into the functional proteins of the mature viruse~.~-~ In vitro mutagenesis that produces proteasedefective virus results in the formation of uninfective, immature forms of the virus. Therefore, inhibition of the protease constitutes a rational strategy for the development of potential anti-AIDS agents. During the course of a program designed to synthesize inhibitors of HIV protease, we wished to prepare a series of pseudopeptides in which the cleaved PhePro amide bonds of substrate peptides were replaced by noncleavable aminomethylene (CH2N) units. The initial targets chosen for synthesis were the peptide derivatives 1,2, and 3.7-9 The sequence of amino acid residues in these peptides corresponds to the cleavage site between the protease and the reverse transcriptase in the HIV-1 gag-pol polyproteh2 A method in general use for the replacement of amide bonds in peptides with the CH2N bond involves reductive alkylation of an amine with an aldehyde.1° Accordingly, BOC-L-phenylalaninal 4 was