Intracellular expression of RNA transcripts complementary to the human immunodeficiency virus type 1 gag gene inhibits viral replication in human CD4+ lymphocytes

Abstract

Intracellular expression of antisense transcripts was evaluated for its potential to interfere with human immunodeficiency virus type 1 (HIV-1) replication. Retroviral vectors encoding HIV-1 psi-gag complementary sequences downstream of a selectable gene (neo, puromycin gene, or Lyt2 gene) were stable and yielded high titers. Human CEMSS T cells were transduced with amphotropic retroviral vectors to express RNA complementary to the psi-gag sequence of HIV-1. Replication of laboratory-adapted HIV-1 strains was inhibited by more than 1 order of magnitude (log10) in these transduced cells even at high inoculation doses (4 x 10(4) 50% tissue culture infective doses). Antisense-mediated anti-HIV efficacy was further demonstrated by survival of CD4+ cells in these cultures relative to controls. The level of anti-HIV-1 activity of the psi-gag antisense sequence correlated with the length of the antisense transcript. Maximal anti-HIV efficacy was observed with complementary sequence more than 1,000 nucleotides long, whereas transcripts less than 400 nucleotides long failed to inhibit HIV-1 replication. Expression of psi-gag antisense RNA also reduced HIV-1 JR-CSF replication 10-fold in primary CD4+ lymphocytes. These results obtained with a T-cell line and primary peripheral blood lymphocytes indicate the potential of long antisense RNAs as an efficient anti-HIV-1 therapeutic agent for gene therapy.