INTRACELLULAR EXPRESSION OF RNA TRANSCRIPTS COMPLEMENTARY TO THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 GAG GENE INHIBITS VIRAL REPLICATION INHUMAN CD4(+) LYMPHOCYTES

Intracellular expression of antisense transcripts mas evaluated for it s potential to interfere with human immunodeficiency virus type I (HIV -1) replication. Retroviral vectors encoding HIV-1 Psi-gag complementa ry sequence downstream of a selectable gene (neo, puromycin gene, or L yt2 gene) were stable and yielded high titers. Human CEMSS T cells wer e transduced with amphotropic retroviral vectors to express RNA comple mentary to the Psi-gag sequence of HIV-1. Replication of laboratory-ad apted HIV-1 strains was inhibited by more than 1 order of magnitude (l og(10)) in these transduced cells even at high inoculation doses (4x10 (4) 50% tissue culture infective doses), Antisense-mediated anti-HN ef ficacy was further demonstrated by survival of CD4(+) cells in these c ultures 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 t han 400 nucleotides long failed to inhibit HIV-1 replication, Expressi on of Psi-gag antisense RNA also reduced HIV-1 JR-CSF replication 10-f old in primary CD4(+). lymphocytes. These results obtained with a T-ce ll line and primary peripheral blood lymphocytes indicate tile potenti al of long antisense RNAs as an efficient anti-HIV-1 therapeutic agent for gene therapy.