RESUMEN
A series of novel aminodiol inhibitors of HIV protease based on the lead compound 1 with structural modifications at P1' were synthesized in order to reduce the cytotoxicity of 1. We have observed a high degree of correlation between the lipophilicity and cytotoxicity of this series of inhibitors. It was found that appropriate substitution at the para position of the P1' phenyl group of 1 resulted in the identification of equipotent (both against the enzyme and in cell culture) compounds (10l, 10m, 10n, and 15c) which possess significantly decreased cytotoxicity.
Asunto(s)
Aminas/síntesis química , Inhibidores de la Proteasa del VIH/síntesis química , Aminas/química , Aminas/farmacología , División Celular/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Inhibidores de la Proteasa del VIH/química , Inhibidores de la Proteasa del VIH/farmacología , Humanos , Relación Estructura-ActividadRESUMEN
The observed in vitro and in vivo benefit of combination treatment with anti-human immunodeficiency virus (HIV) agents prompted us to examine the potential of resistance development when two protease inhibitors are used concurrently. Recombinant HIV-1 (NL4-3) proteases containing combined resistance mutations associated with BMS-186318 and A-77003 (or saquinavir) were either inactive or had impaired enzyme activity. Subsequent construction of HIV-1 (NL4-3) proviral clones containing the same mutations yielded viruses that were severely impaired in growth or nonviable, confirming that combination therapy may be advantageous. However, passage of BMS-186318-resistant HIV-1 (RF) in the presence of either saquinavir or SC52151, which represented sequential drug treatment, produced viable viruses resistant to both BMS-186318 and the second compound. The predominant breakthrough virus contained the G48V/A71T/V82A protease mutations. The clone-purified RF (G48V/A71T/V82A) virus, unlike the corresponding defective NL4-3 triple mutant, grew well and displayed cross-resistance to four distinct protease inhibitors. Chimeric virus and in vitro mutagenesis studies indicated that the RF-specific protease sequence, specifically the Ile at residue 10, enabled the NL4-3 strain with the triple mutant to grow. Our results clearly indicate that viral genetic background will play a key role in determining whether cross-resistance variants will arise.
Asunto(s)
Inhibidores de la Proteasa del VIH/farmacología , Proteasa del VIH/genética , VIH-1/efectos de los fármacos , Secuencia de Aminoácidos , Carbamatos/farmacología , Células Clonales , Análisis Mutacional de ADN , ADN Recombinante/genética , ADN Viral/genética , Esquema de Medicación , Farmacorresistencia Microbiana/genética , Quimioterapia Combinada , Etanolaminas/farmacología , Inhibidores de la Proteasa del VIH/administración & dosificación , VIH-1/enzimología , VIH-1/genética , Células HeLa , Humanos , Indinavir , Isoquinolinas/farmacología , Compuestos de Metilurea/farmacología , Datos de Secuencia Molecular , Mutación Puntual , Provirus/enzimología , Provirus/genética , Piridinas/farmacología , Quinolinas/farmacología , Virus Reordenados/efectos de los fármacos , Virus Reordenados/genética , Proteínas Recombinantes de Fusión/antagonistas & inhibidores , Proteínas Recombinantes de Fusión/metabolismo , Saquinavir , Linfocitos T , Urea/análogos & derivados , Urea/farmacología , Valina/análogos & derivadosRESUMEN
Development of viral resistance to the aminodiol human immunodeficiency virus (HIV) protease inhibitor BMS 186,318 was studied by serial passage of HIV type 1 RF in MT-2 cells in the presence of increasing concentrations of compound. After 11 passages, an HIV variant that showed a 15-fold increase in 50% effective dose emerged. This HIV variant displays low-level cross-resistance to the C2 symmetric inhibitor A-77003 but remains sensitive to the protease inhibitors Ro 31-8959 and SC52151. Genetic analysis of the protease gene from a drug-resistant variant revealed an Ala-to-Thr change at amino acid residue 71 (A71T) and a Val-to-Ala change at residue 82 (V82A). To determine the effects of these mutations on protease and virus drug susceptibility, recombinant protease and proviral HIV type 1 clones containing the single mutations A71T and V82A or double mutation A71T/V82A were constructed. Subsequent drug sensitivity assays on the mutant proteases and viruses indicated that the V82A substitution was responsible for most of the resistance observed. Further genotypic analysis of the protease genes from earlier passages of virus indicated that the A71T mutation emerged prior to the V82A change. Finally, the level of resistance did not increase following continued passage in increasing concentrations of drug, and the resistant virus retained its drug susceptibility phenotype 34 days after drug withdrawal.
Asunto(s)
Carbamatos/farmacología , Etanolaminas/farmacología , Inhibidores de la Proteasa del VIH/farmacología , VIH-1/efectos de los fármacos , Secuencia de Aminoácidos , Secuencia de Bases , Línea Celular , Cartilla de ADN , Farmacorresistencia Microbiana , Variación Genética , Proteasa del VIH/metabolismo , VIH-1/enzimología , VIH-1/genética , Células HeLa , Humanos , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido , Pase SeriadoRESUMEN
A series of aminodiol inhibitors of human immunodeficiency virus type 1 (HIV-1) protease were identified by using an in vitro peptide cleavage assay. BMS 182,193, BMS 186,318, and BMS 187,071 protected cells against HIV-1, HIV-2, and simian immunodeficiency virus infections, with 50% effective doses ranging from 0.05 to 0.33 microM, while having no inhibitory effect on cells infected with unrelated viruses. These compounds were also effective in inhibiting p24 production in peripheral blood mononuclear cells infected with HIV-1 IIIB and against the zidovudine-resistant HIV-1 strain A018C. Time-of-addition studies indicated that BMS 182,193 could be added as late as 27 h after infection and still retain its antiviral activity. To directly show that the activity of these compounds in culture was due to inhibition of proteolytic cleavage, the levels of HIV-1 gag processing in chronically infected cells were monitored by Western blot (immunoblot) analysis. All compounds blocked the processing of p55 in a dose-dependent manner, with 50% effective doses of 0.4 to 2.4 microM. To examine the reversibility of BMS 186,318, chronically infected CEM-SS cells were treated with drug and virions purified from the culture medium. Incubation of HIV-1 particles in drug-free medium indicated that inhibition of p55 proteolysis was slowly reversible. The potent inhibition of HIV-1 during both acute and chronic infections indicates that these aminodiol compounds are effective anti-HIV-1 compounds.
Asunto(s)
Antivirales/farmacología , Carbamatos/farmacología , Etanolaminas/farmacología , Inhibidores de la Proteasa del VIH/farmacología , VIH-1/efectos de los fármacos , Línea Celular , Productos del Gen gag/metabolismo , VIH-1/enzimología , Humanos , Precursores de Proteínas/metabolismo , Virus de la Inmunodeficiencia de los Simios/efectos de los fármacosRESUMEN
A series of HIV protease inhibitors containing a novel C2 symmetrical "aminodiol" core structure were prepared from amino acid starting materials. The ability of the aminodiols to inhibit HIV replication in cell culture is comparable to their ability to inhibit the isolated enzyme, a result compatible with good cell membrane penetration by this class of compounds. Optimization of the structure-activity in this series led to aminodiol 9a (Ki = 100 nM; ED50 (HIV-1) = 80 nM) containing P1/P1 benzyl and P2/P2 Boc substituents. Compound 9a is a selective inhibitor of HIV protease versus other aspartyl proteases such as human renin, human cathepsin D, and porcine pepsin. In addition, 9a is equipotent against HIV-1 and HIV-2 in cell culture and demonstrates similar activity in infected T-lymphocytes and PBMCs. After i.v. and oral administration in rats, 9a displayed significant oral bioavailability (ca. 40%) and a promising plasma elimination half-life (4 h).