RESUMEN
Human immunodeficiency virus type 1 (HIV-1) tropism can be assessed using phenotypic assays, but this is quite laborious, expensive, and time-consuming and can be made only in sophisticated laboratories. More accessible albeit reliable tools for testing of HIV-1 tropism are needed in view of the prompt introduction of CCR5 antagonists in clinical practice. Bioinformatics tools based on V3 sequences might help to predict HIV-1 tropism; however, most of these methods have been designed by taking only genetic information derived from HIV-1 subtype B into consideration. The aim of this study was to evaluate the performances of several genotypic tools to predict HIV-1 tropism in non-B subtypes, as data on this issue are scarce. Plasma samples were tested using a new phenotypic tropism assay (Phenoscript-tropism; Eurofins), and results were compared with estimates of coreceptor usage using eight different genotypic predictor softwares (Support Vector Machine [SVM], C4.5, C4.5 with positions 8 to 12 only, PART, Charge Rule, geno2pheno coreceptor, Position-Specific Scoring Matrix X4R5 [PSSM(X4R5)], and PSSM(sinsi)). A total of 150 samples were tested, with 115 belonging to patients infected with non-B subtypes and 35 drawn from subtype B-infected patients, which were taken as controls. When non-B subtypes were tested, the concordances between the results obtained using the phenotypic assay and distinct genotypic tools were as follows: 78.8% for SVM, 77.5% for C4.5, 82.5% for C4.5 with positions 8 to 12 only, 82.5% for PART, 82.5% for Charge Rule, 82.5% for PSSM(X4R5), 83.8% for PSSM(sinsi), and 71.3% for geno2pheno. When clade B viruses were tested, the best concordances were seen for PSSM(X4R5) (91.4%), PSSM(sinsi) (88.6%), and geno2pheno (88.6%). The sensitivity for detecting X4 variants was lower for non-B than for B viruses, especially in the case of PSSM(sinsi) (38.4% versus 100%, respectively), SVM(wetcat) (46% versus 100%, respectively), and PART (30% versus 90%, respectively). In summary, while inferences of HIV-1 coreceptor usage using genotypic tools seem to be reliable for clade B viruses, their performances are poor for non-B subtypes, in which they particularly fail to detect X4 variants.
Asunto(s)
Biología Computacional/métodos , Proteína gp120 de Envoltorio del VIH/genética , Infecciones por VIH/virología , VIH-1/clasificación , VIH-1/patogenicidad , Fragmentos de Péptidos/genética , Receptores CCR5/metabolismo , Receptores CXCR4/metabolismo , Adulto , Antígenos CD4/metabolismo , Femenino , Genotipo , Proteína gp120 de Envoltorio del VIH/metabolismo , VIH-1/genética , VIH-1/metabolismo , Humanos , Masculino , Persona de Mediana Edad , Fragmentos de Péptidos/metabolismo , Fenotipo , Juego de Reactivos para Diagnóstico , Sensibilidad y EspecificidadRESUMEN
The predictive value of three genotypic methods to determine HIV-1 co-receptor usage was assessed in 83 plasma specimens taking as reference the results obtained using a recombinant phenotypic assay (Phenoscript). The best concordance was found for webPSSM, followed by geno2pheno and wetcat (85.9, 71.8 and 70.5%, respectively). Less than 5.1% of phenotypic X4 viruses were missed by genotypic tools. The genotypic prediction of HIV-1 co-receptor usage can thus assist therapeutic decisions for using CCR5 antagonists.
Asunto(s)
Proteína gp120 de Envoltorio del VIH/genética , Infecciones por VIH/diagnóstico , VIH-1/genética , Fragmentos de Péptidos/genética , Receptores del VIH/genética , Secuencia de Aminoácidos , Secuencia de Bases , Biología Computacional , Cartilla de ADN/genética , Femenino , Genotipo , Infecciones por VIH/genética , Humanos , Masculino , Datos de Secuencia Molecular , Fenotipo , Receptores CCR5/genética , Receptores CXCR4/genética , Sensibilidad y Especificidad , Análisis de Secuencia de ADN/métodosRESUMEN
BACKGROUND: Double-boosted protease inhibitors (PIs) are under investigation for the treatment of patients who are unable to take nucleoside reverse transcriptase inhibitors because of cross-resistance and/or intolerance. Evidence of synergistic inhibition of wild-type HIV has been reported for saquinavir with atazanavir or lopinavir. METHODS: We investigated the activity of these two combinations against a panel of six site-directed mutant HIV-1 strains and 14 clinically derived recombinant HIV-1 strains presenting a range of PI-resistance profiles. RESULTS: No evidence of synergy was observed against wild-type virus for either combination. The combination of saquinavir and lopinavir showed evidence of synergy against four viruses displaying high-level resistance to lopinavir and low-level resistance to saquinavir. Similarly, evidence of synergy between saquinavir and atazanavir was only observed in two viruses which were more susceptible to saquinavir than to atazanavir. CONCLUSIONS: We hypothesize that differences between the PIs in intracellular protein-binding behaviour or inhibition of drug transporters (P glycoprotein, MDR1 and MDR2) could result in intracellular levels of saquinavir being increased by co-administration with lopinavir or atazanavir. The effect of this increase would be masked in cases involving viruses that were susceptible to atazanavir or lopinavir. In virus resistant to lopinavir or atazanavir but susceptible to saquinavir, the majority of the antiviral effect is due to saquinavir; thus even small increases in intracellular concentration could significantly increase virus inhibition. These results confirm that in vitro synergy can be observed between PIs and suggest that the degree of synergy observed might depend on the resistance profile of the virus.
Asunto(s)
Inhibidores de la Proteasa del VIH/farmacología , VIH-1/efectos de los fármacos , Oligopéptidos/farmacología , Piridinas/farmacología , Pirimidinonas/farmacología , Saquinavir/farmacología , Sulfato de Atazanavir , Línea Celular Tumoral , Farmacorresistencia Viral , Sinergismo Farmacológico , VIH-1/genética , Humanos , Concentración 50 Inhibidora , Lopinavir , Pruebas de Sensibilidad Microbiana , Recombinación Genética , TransfecciónRESUMEN
BACKGROUND: Data on susceptibility of HIV-1 non-B subtypes to Enfuvirtide (ENF) is rather limited. OBJECTIVE: To determine if ENF could be active in vitro against HIV-1 non-B subtypes and how the gp41 genetic variability across variants may influence ENF susceptibility. METHODS: Using PHENOSCRIPT Env, a recombinant envelope virus assay, ENF susceptibility was investigated in isolates from 19 drug-naive HIV-1-infected individuals harboring non-B subtypes. RESULTS: Using phylogenetic analyses of the gp41 gene, distinct HIV-1 subtypes were recognized: A (2), C (5), D (1), F (2), G (1), J (1), CRF02_AG (6) and CRF06 (1). Susceptibility to ENF and IC(50) values in vitro could be obtained in only 13 (68.4%) specimens, most likely due to the high genetic variability in HR1 and HR2 regions in the remaining cases. A wide range of IC(50) values with a median of 0.013 microg/ml was observed (range, 0.005-0.180 microg/ml). Natural polymorphisms, but not classical ENF resistance associated mutations within HR1 (residues 36-45) were identified in most non-B viruses. CONCLUSION: This study provides information about the baseline susceptibility to ENF in antiretroviral-naive subjects infected with different HIV-1 non-B subtypes. Susceptibility to ENF seems to be preserved despite high genetic variability in HR1 and HR2 regions.
Asunto(s)
Proteína gp41 de Envoltorio del VIH/genética , Inhibidores de Fusión de VIH/farmacología , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Adulto , Secuencia de Aminoácidos , Enfuvirtida , Femenino , Proteína gp41 de Envoltorio del VIH/inmunología , Proteína gp41 de Envoltorio del VIH/farmacología , VIH-1/genética , VIH-1/aislamiento & purificación , Humanos , Masculino , Persona de Mediana Edad , FilogeniaRESUMEN
Enfuvirtide (ENF) is the first of a novel class of drugs that blocks HIV fusion to host cells. We analyzed the dynamics of genotypic and phenotypic resistance to ENF during and after long-term ENF therapy and its clinical implications in eight heavily treatment-experienced HIV-infected patients who underwent salvage therapy with enfuvirtide along with other antiretroviral agents. All patients showed a rapid decline in plasma HIV-RNA followed by viral rebound. Changes at codons 36, 42, 43 and/or 44 within the HR1 region of gp41 were selected in all cases, resulting in high-level phenotypic resistance to ENF, ranging from 15- to 445-fold. Both genotypic and phenotypic resistance to ENF rapidly disappeared after discontinuation of the drug, suggesting that ENF-resistant viruses may have an impaired replicative capacity.
Asunto(s)
Proteína gp41 de Envoltorio del VIH/farmacología , Inhibidores de Fusión de VIH/farmacología , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Secuencia de Aminoácidos , Antirretrovirales/uso terapéutico , Farmacorresistencia Viral/genética , Enfuvirtida , Evolución Molecular , Proteína gp41 de Envoltorio del VIH/genética , Infecciones por VIH/tratamiento farmacológico , VIH-1/genética , Humanos , Datos de Secuencia Molecular , Terapia Recuperativa , Alineación de Secuencia , Especificidad de la EspecieRESUMEN
Four heavily antiretroviral-experienced HIV-infected patients had significant plasma HIV-RNA reductions (>1 log) after beginning an Enfuvirtide (ENF)-based rescue regimen. However, all had viral rebound shortly thereafter, sustaining high levels of plasma viremia over 80 weeks. These patients developed rapidly genotypic and phenotypic resistance to ENF. Mutations within the HR1 env region were selected (N43D in three and G36V/D in one), resulting in high-level phenotypic resistance to ENF. Interestingly, two patients had a sustained CD4+ T-cell increase and two maintained stable CD4+ T-cell counts despite virologic failure under ENF. The possible mechanisms involved in this response were examined. Changes in virus tropism from R5 to R5/X4 were observed in two patients, in parallel with increases in ENF phenotypic resistance. Low levels of T-cell activation, T-cell turnover, and cytotoxic T lymphocyte (CTL) activity were found in all four patients. An overall increase in the proportion of viruses released from cells of the macrophage lineage was observed. In summary, single mutations at the HR1 env region result in significant loss of susceptibility to ENF. Despite virologic failure, these patients may maintain elevated CD4+ counts through a reduction in their overall immune activation.