A GENE THERAPY APPROACH TO THE INHIBITION OF HIV-1 REPLICATION BY RESTORATION OF INNATE ANTIVIRAL DEFENSE PATHWAYS

Abstract

Since it emerged as an infectious agent in 1981, the human immunodeficiency virus type 1 (HIV-1) is continually disseminated and remain fatal to the majority of those infected. Strategies including highly active retroviral therapies (HAART) with nucleoside analogues and protease inhibitors have shown limited success in therapy due to the virus' ability to evolve rapidly at every replication cycle as a consequence of it's highly error prone reverse transcriptase, generating resistant retroviral strains and in addition to latent HIV-1 reservoirs. Thirty years of research efforts to find a cure or to generate a vaccine has been met with failure. It is, therefore, of necessity to broaden our paradigm of therapy for the treatment and eventual cure of HIV-1 infection. In this study, I look beyond the current anti-retroviral strategies and instead rely on the mammalian host immune system to inhibit HIV-1 replication through molecular genetic manipulation. Here, we approach the inhibition of HIV-1 replication by up-regulation of the innate antiviral pathway that is natural to mammalian cells. HIV-1 derived self-inactivating lentiviral (SIN) vectors were designed and constructed to deliver the antiviral payloads of two antiviral enzymes, p68 kinase (PKR) and 2'-5' oligoadenlyate synthetase (2-5OAS), to target cell, SupT1 lymphoblastoid cells and CD4+ T lymphocytes under the control of a constitutive cytomegalovirus (CMV) promoter. These data here demonstrates a significant inhibition of HIV-1 replication in cells transduced with the anti HIV-1 transgenes PKR and 2-5OAS as determined by HIV-1 induced syncytia formation and HIV-1 p24 antigen capture assay. Furthermore, here demonstrated is an increase up-regulation of PKR and 2-5OAS 96 hr post cell transduction in all the clones when compared to pHIV empty vector control. These results demonstrate that the over-expression of PKR and 2-5OAS can inhibit HIV-1 replication and also confirm the involvement of PKR and 2-5OAS in the IFN-associated antiviral pathway against HIV-1 infection.