DECOY APPROACH USING RNA-DNA CHIMERA OLIGONUCLEOTIDES TO INHIBIT THE REGULATORY FUNCTION OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 REV PROTEIN

Human immunodeficiency virus type 1 (HIV-1) encodes two regulatory pro teins, Tat and Rev, that bind to target RNA sequences. These are the t rans-activation responsive (TAR) RNA and the Rev-responsive element (R RE), respectively. The Rev protein shifts RNA synthesis to viral late transcripts by binding to the RRE within the env gene. In the present study we prepared a RNA-DNA chimera consisting of 29 or 31 nucleotides to inhibit the Rev regulatory function by means of the decoy approach . The chimera oligonucleotides (anti-Rev oligonucleotides [AROs]) cont ained an RNA ''bubble'' structure (13 oligonucleotides; the Rev-bindin g element in RRE) that bound Rev with a high affinity in an in vitro a ssay. The controls were RNA-DNA chimera oligonucleotides (negative con trol oligonucleotides [NCOs]) similar to ARO, but without the bubble s tructure, that bound,vith considerably less affinity to Rev. When the inhibitory effects of these decoys on HIV-1 replication were examined, we found that AROs, but not NCOs, reduced more than 90% of the HIV-1 production generated by productively infected human T-cell lines, The production of primary HIV-1 isolates in healthy donor-derived peripher al blood mononuclear cells was also similarly inhibited by AROs. In ad dition, the induction of viral mRNAs and antigens in latently HTV-1-in fected ACH-2 cells by tumor necrosis factor alpha was specifically inh ibited by AROs, but not by NCOs. No apparent cytotoxicity was caused b y either decoy. Thus, the use of a Rev-binding element-based decoy, th e RNA-DNA chimera oligonucleotide, may represent a safer approach to g ene therapy for reducing the virus load in HIV-1-infected individuals.