Certain bis(heteroaryl)piperazines (BHAPs) are potent inhibitors of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) at concentrations lower by 2-4 orders of magnitude than that which inhibits normal cellular DNA polymerase activity. Combination of a BHAP with nucleoside analog IIV-1 RT inhibitors suggested that together these compounds inhibited RT synergistically. In three human lymphocytic cell systems using several laboratory and clinical HIV-1 isolates, the BHAPs blocked HIV-1 replication with potencies nearly identical to those of 3'-azido-2',3'-dideoxythymidine or 2',3'-dideoxyadenosine; in primary cultures of human peripheral blood mononuclear cells, concentrations of these antiviral agents were lower by at least 34 orders of magnitude than cytotoxic levels. The BHAPs do not inhibit replication of HIV-2, the simian or feline immunodeficiency virus, or Rauscher murine leukemia virus in culture. Evaluation of a BHAP in HIV-1-infected SCID-hu mice (severe combined ict mice implanted with human fetal lymph node) showed that the compound could block HIV-1 replication in vivo. The BHAPs are readily obtained synthetically and have been extensively characterized in preclinical evaluations. These compounds hold promise for the treatment of HIV-1 infection.The reverse transcriptase (RT) encoded by human immunodeficiency virus type 1 (HIV-1) catalyzes the conversion of the viral genomic RNA into proviral DNA (1, 2). Since RT is essential for virus replication and has no closely related identified cellular homolog, it has been the prime target for antiviral therapy against theacquired immunodeficiency syndrome (AIDS; refs. 3 and 4). This strategy is appropriate since 3'-azido-2',3'-dideoxythymidine (AZT), a nucleoside analog inhibitor of reverse transcription, was the first drug shown to benefit HIV-1-infected individuals (5). Other nucleoside analog RT inhibitors also show promise in clinical evaluations (6, 7). However, the administration of these drugs to patients is usually limited by serious toxicities (7,8). In addition, HIV-1 with reduced AZT-sensitivity has been obtained from AZT-treated patients, suggesting the emergence of resistant virus will limit the drug's efficacy (9, 10). Thus, effective prolonged treatment of HIV-1 infection likely requires the discovery of other, perhaps multiple, RT inhibitors. To this end, we and others (11, 12) have sought to identify other nonnucleoside HIV-1 RT inhibitors. MATERIALS AND METHODSCell Culture and Virus Infections. The cell cultures were maintained at 370C in 5% C02/95% air. The HIV-1 infectivity studies were conducted in MT-2 cells, peripheral blood mononuclear cells (PBMC), and H9 cells as described (9,(13)(14)(15). In brief, MT-2 or H9 cells were infected with HIV-1 (IlIb isolate) at a multiplicity of infection of 0.001. In MT-2 cells syncytium formation was determined 4 days after infection at the peak of the viral cytopathic effect (13). In H9 cells, growth medium with fresh drug was replaced every 3-4 days, and at 14 days-...

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Bisheteroarylpiperazines are potent inhibitors of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT). We describe a novel bisheteroarylpiperazine, U-90152 {1-(5-methanesulfonamido-IH-indol-2-yI-carbonyl)-4-[3-(1-methylethyl-amino)pyridinyllpiperazine}, which inhibited recombinant HIV-1 RT at a 50%Y inhibitory concentration (IC.) of 0.26 FLM (compared with IC50s of >440 ,uM for DNA polymerases at and B). U-90152 blocked the replication in peripheral blood lymphocytes of 25 primary HWV-i isolates, including variants that were highly resistant to 3'-azido-2',3'-dideoxythymidine (AZT) or 2',3'-dideoxyinosine, with a mean 50%o effective dose of 0.066 + 0.137 pM. U-90152 had low cellular cytotoxicity, causing less than 8% reduction in peripheral blood lymphocyte viability at 100 ,uM. In experiments assessing inhibition of the spread of HIV-MB in cell cultures, U-90152 was much more effective than AZT. When approximately 500 HIV-1l1m-infected MT-4 cells were mixed 1:1,000 with uninfected cells, 3 ,uM AZT delayed the evidence of rapid viral growth for 7 days. In contrast, 3 ,uM U-90152 totally prevented the spread of HIV-i, and death and/or dilution of the original inoculum of infected cells prevented renewed viral growth after U-90152 was removed at day 24. The combination of U-90152 and AZT, each at 0.5 ,uM, also totally prevented viral spread. Finally, although the RT amino acid substitutions K103N (lysine 103 to asparagine) and Y181C (tyrosine 181 to cysteine), which confer cross-resistance to several nonnucleoside inhibitors, also decrease the potency of U-90152, this drug retains significant activity against these mutant RTs in vitro (IC50s, approximately 8 ,uM).

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