HIV-1 Nef-Src Family Kinase Interaction: A Novel Target for the Inhibition of HIV-1 Pathogenesis

[摘要] Human immunodeficiency virus-1 (HIV-1) is a lentivirus responsible for development of AIDS.In addition to typical retroviral proteins (Gag, Pol and Env), primate lentiviruses like HIV-1 encode two regulatory (Tat and Rev) and four accessory proteins (Vif, Vpu, Vpr and Nef). The HIV-1 accessory factor Nef is essential for high-titer viral replication and AIDS progression.Nef function requires interaction with many host cell proteins, including specific members of the Src kinase family.In this dissertation project, I explored whether Src-family kinase (SFK) activation is a conserved property of nef alleles from a wide range of primary HIV-1 isolates and its sensitivity to selective pharmacological inhibitors.Representative Nef proteins from the major HIV-1 subtypes A1, A2, B, C, F1, F2, G, H, J and K strongly activated Hck and Lyn as well as c-Src to a lesser extent, demonstrating for the first time that SFK activation is a highly conserved property of primary M-group HIV-1 Nef isolates.Moreover, patient-derived Nef proteins also strongly activated Hck. Recently, our group identified 4-amino diphenylfuranopyrimidines (DFPs) and diphenylpyrazolyldiazene (PPD-B9) compounds that selectively inhibit Nef-dependent SFK activation as well as HIV replication. To determine whether these novel compounds exhibit broad-spectrum Nef-dependent antiretroviral activity against HIV-1, I first constructed chimeric forms of the viral strain NL4-3 expressing the same 10 primary nef alleles. The infectivity and replication of these Nef chimeras was indistinguishable from that of wild-type in three distinct cell lines (MT2, U87MG and CEM-T4). Importantly, the 4-aminopropanol and 4-aminobutanol derivatives of DFP as well as PPD-B9 potently inhibited the replication of all chimeric forms of HIV-1 in both U87MG and CEM-T4 cells in a Nef-dependent manner.The effects of these compounds against HIV replication correlated with inhibition of Nef-dependent activation of endogenous SFKs. My results demonstrate that the activation of Hck, Lyn and c-Src by Nef is highly conserved among all major clades of HIV-1 and that selective targeting of this pathway uniformly inhibits HIV-1 replication. My results have strong public health significance for developing therapeutics against current drug resistant variants of HIV-1.

[发布机构] the University of Pittsburgh