In vitro resistance profile of the human immunodeficiency virus type 1 protease inhibitor BMS-232632.

BMS-232632 is an azapeptide human immunodeficiency virus (HIV) type 1 (HIV-1) protease inhibitor that displays potent anti-HIV-1 activity (50% effective concentration [EC(50)], 2.6 to 5.3 nM; EC(90), 9 to 15 nM). In vitro passage of HIV-1 RF in the presence of inhibitors showed that BMS-232632 selected for resistant variants more slowly than nelfinavir or ritonavir did. Genotypic and phenotypic analysis of three different HIV strains resistant to BMS-232632 indicated that an N88S substitution in the viral protease appeared first during the selection process in two of the three strains. An I84V change appeared to be an important substitution in the third strain used. Mutations were also observed at the protease cleavage sites following drug selection. The evolution to resistance seemed distinct for each of the three strains used, suggesting multiple pathways to resistance and the importance of the viral genetic background. A cross-resistance study involving five other protease inhibitors indicated that BMS-232632-resistant virus remained sensitive to saquinavir, while it showed various levels (0. 1- to 71-fold decrease in sensitivity)-of cross-resistance to nelfinavir, indinavir, ritonavir, and amprenavir. In reciprocal experiments, the BMS-232632 susceptibility of HIV-1 variants selected in the presence of each of the other HIV-1 protease inhibitors showed that the nelfinavir-, saquinavir-, and amprenavir-resistant strains of HIV-1 remained sensitive to BMS-232632, while indinavir- and ritonavir-resistant viruses displayed six- to ninefold changes in BMS-232632 sensitivity. Taken together, our data suggest that BMS-232632 may be a valuable protease inhibitor for use in combination therapy.

Reverse transcription takes place within extracellular HIV-1 virions: potential biological significance.

Extracellular HIV-1 virions purified from cell culture supernatants have been found to contain viral DNA that is the result of partial reverse transcription within the virus particles. Our data supported these observations and further indicated that the ratio of genomic RNA to viral DNA was approximately 10(3):1 for the "strong stop" (R-U5) region and 10(5):1 for the gag region. We have shown that, in the absence of detergent, large amounts of DNase-resistant viral DNA can be synthesized within intact HIV-1 virions, indicating that this phenomenon is not dependent on perturbation of the viral envelope. Nascent viral DNA synthesis also occurred in purified virions incubated at 37 degrees C in cell-free human physiological fluids including seminal plasma, blood plasma, breast milk, and fecal fluid. In vitro HIV-1 infection assays, in which HIV-1 DNA synthesis was initiated in HIV-1 virions by prior incubation with deoxyribonucleoside triphosphates, demonstrated that virus particles so treated had an increased infectious titer over untreated virions when incubated with target human T cells. Our data suggest that HIV-1 virion-associated DNA synthesis may occur in vivo and may impact on the efficiency of intra- and interhost virus transmission. If so, this phenomenon should prove to be an important target for antiviral therapeutic strategies.

Sequence analysis of an immunogenic and neutralizing domain of the human T-cell lymphoma/leukemia virus type I gp46 surface membrane protein among various primate T-cell lymphoma/leukemia virus isolates including those from a patient with both HTLV-I-associated myelopathy and adult T-cell leukemia.

Human T-cell lymphoma/leukemia virus type I (HTLV-I) causes adult T-cell leukemia/lymphoma and HTLV-I-associated myelopathy. Specific regions within the outer envelope proteins of other retroviruses, e.g., human immunodeficiency virus type 1, are highly immunogenic and, because of the selective pressure of the host immune system, quite variable. Mutations in the external envelope protein gene of murine retroviruses and human immunodeficiency virus type 1 influence cellular tropism and disease pathogenesis. By contrast, no disease-specific viral mutations have been identified in HTLV-I-infected patients. However, all isolates studied thus far have originated from leukemic cell lines, peripheral blood mononuclear cells, or cerebrospinal fluid lymphocytes from patients with HTLV-I-associated myelopathy and adult T-cell leukemia/lymphoma and, therefore, may not truly reflect tissue-associated variation. The midregion of the HTLV-I gp46 external envelope glycoprotein (amino acids 190-209) induces an antibody response in 90% of infected individuals, and a hexapeptide in this region (amino acids 191-196) elicits antibodies in rabbits which inhibit syncytia formation and infection of target lymphocytes. Because of the above, we expected the neutralizing domain of the gp46 env gene of HTLV-I to possess disease or organ-associated mutations selected by the infected host's immune system. Hence, we amplified, cloned, and sequenced HTLV-I DNA directly from in vivo central nervous system, spleen, and kidney specimens, and a leukemic cell line from a patient (M. J.) with both HTLV-I-associated myelopathy and adult T-cell leukemia/lymphoma to discern the possibility of tissue- and/or disease-specific variants. In addition, we sequenced several HTLV-I isolates from different regions of the world, including Papua New Guinea, Bellona, and Liberia, and compared them to other previously published HTLV-I and related retroviral sequences. The 239-base pair sequence corresponding to amino acids 178 to 256 in gp46 displayed minor tissue-specific variation in clones derived from central nervous system tissues from patient M. J., but overall was highly conserved at both the DNA and amino acid levels. Variation was observed in this region among the other HTLV-I, simian T-cell lymphoma virus type I, and HTLV-II isolates in a pattern that was consistent with their known phylogenetic relationship. No consistent disease-related changes were observed.(ABSTRACT TRUNCATED AT 400 WORDS)