Subject(s)
Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Mutation/genetics , Nuclear Proteins/genetics , Repressor Proteins/genetics , Adult , Aged , Aged, 80 and over , Comparative Genomic Hybridization , Female , Humans , Karyotyping , Male , Middle Aged , Nucleophosmin , Oligonucleotide Array Sequence Analysis , Polymorphism, Genetic , Prognosis , Young Adult , fms-Like Tyrosine Kinase 3/genetics , ras Proteins/geneticsSubject(s)
Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Mutation, Missense , NFI Transcription Factors/genetics , Neoplasm Proteins/genetics , Point Mutation , Polycythemia Vera/genetics , Sequence Deletion , Amino Acid Substitution , Base Sequence , Chromosomes, Human, Pair 1/genetics , Diagnostic Errors , Disease Progression , Female , Genes, Tumor Suppressor , Humans , Janus Kinase 2/genetics , Leukemia, Monocytic, Acute/genetics , Male , Middle Aged , Molecular Sequence Data , NFI Transcription Factors/chemistry , Neoplasm Proteins/chemistry , Polycythemia Vera/diagnosis , Protein Structure, Tertiary/genetics , Thrombocythemia, Essential/diagnosisSubject(s)
Gene Expression Profiling , Gene Expression Regulation, Leukemic/genetics , Leukemia, Myelomonocytic, Chronic/diagnosis , Leukemia, Myelomonocytic, Chronic/genetics , Adolescent , Adult , Aged , Aged, 80 and over , Female , Humans , Leukocytes/cytology , Male , Middle Aged , Models, Genetic , Mutation , Oligonucleotide Array Sequence AnalysisABSTRACT
We developed a recombinant virus technique to determine the coreceptor usage of human immunodeficiency virus type 1 (HIV-1) from plasma samples, the source expected to represent the most actively replicating virus population in infected subjects. This method is not subject to selective bias associated with virus isolation in culture, a step required for conventional tropism determination procedures. The addition of a simple subcloning step allowed semiquantitative evaluation of virus populations with a different coreceptor (CCR5 or CXCR4) usage specificity present in each plasma sample. This procedure detected mixtures of CCR5- and CXCR4-exclusive virus populations as well as dualtropic viral variants, in variable proportions. Sequence analysis of dualtropic clones indicated that changes in the V3 loop are necessary for the use of CXCR4 as a coreceptor, but the overall context of the V1-V3 region is important to preserve the capacity to use CCR5. This convenient technique can greatly assist the study of virus evolution and compartmentalization in infected individuals.
Subject(s)
Acquired Immunodeficiency Syndrome/virology , HIV-1/physiology , Receptors, CCR5/analysis , Receptors, CXCR4/analysis , Viremia/virology , Amino Acid Sequence , Cells, Cultured , Genetic Vectors , Humans , Molecular Sequence Data , Phenotype , Recombination, Genetic , Virus ReplicationABSTRACT
Human immunodeficiency virus type 1 (HIV-1) resistance to protease inhibitors (PI) is a major obstacle to the full success of combined antiretroviral therapy. High-level resistance to these compounds is the consequence of stepwise accumulation of amino acid substitutions in the HIV-1 protease (PR), following pathways that usually differ from one inhibitor to another. The selective advantage conferred by resistance mutations may depend upon several parameters: the impact of the mutation on virus infectivity in the presence or absence of drug, the nature of the drug, and its local concentration. Because drug concentrations in vivo are subject to extensive variation over time and display a markedly uneven tissue distribution, the parameters of selection for HIV-1 resistance to PI in treated patients are complex and poorly understood. In this study, we have reconstructed a large series of HIV-1 mutants that carry single or combined mutations in the PR, retracing the accumulation pathways observed in ritonavir-, indinavir-, and saquinavir-treated patients. We have then measured the phenotypic resistance and the drug-free infectivity of these mutant viruses. A deeper insight into the evolutionary value of HIV-1 PR mutants came from a novel assay system designed to measure the replicative advantage of mutant viruses as a function of drug concentration. By tracing the resultant fitness profiles, we determined the range of drug concentrations for which mutant viruses displayed a replicative advantage over the wild type and the extent of this advantage. Fitness profiles were fully consistent with the order of accumulation of resistance mutations observed in treated patients and further emphasise the key importance of local drug concentration in the patterns of selection of drug-resistant HIV-1 mutants.
Subject(s)
Evolution, Molecular , HIV Protease Inhibitors/pharmacology , HIV Protease/genetics , HIV-1/genetics , Amino Acid Substitution , Cell Line , Drug Resistance, Microbial , Gene Products, gag/genetics , HIV-1/drug effects , HIV-1/pathogenicity , HeLa Cells , Humans , Indinavir/pharmacology , Mutation, Missense , Ritonavir/pharmacology , Saquinavir/pharmacologyABSTRACT
To investigate the genetic and biologic features of HIV-1 strains circulating in Cambodia, viruses from 95 HIV-1-seropositive individuals were subtyped by heteroduplex mobility assay (HMA) and 23 were further analyzed for their biologic characteristics. Eighty-nine individuals were clearly infected by HIV-1 subtype E. The other six samples were sequenced, together with 17 HMA subtype E samples. All but one of the 23 Cambodian env sequences clustered with previously described Thai and Vietnamese subtype E sequences, bearing a GPGQ motif at the tip of the V3 loop; the last had a GPGR motif and was phylogenetically equidistant from Asian and African subtype E viruses. Nonsyncytium-inducing, CCR5-dependent viruses predominated in patients of clinical stage B even in some with a high viral load and were detected in about 50% of the patients of stage C. All syncytium-inducing strains, mostly from AIDS patients, used both CCR5 and CXCR4. The presence of syncytium-inducing viruses did not correlate with the plasma viral load. These data show that CCR5-dependent HIV-1 subtype E is currently predominant in Cambodia. The analysis of clinical and virologic markers strongly supports the idea that dynamics of the viral population during subtype E infection in Southeast Asia is similar to that of subtype B infection in Europe and the United States.
PIP: The National AIDS Control and Prevention Program of the Cambodian ministry of health has reported that the prevalence of HIV-1 infection among blood donors in Cambodia increased from less than 1% in 1991 to 4% in 1996, and that 39.3% of prostitutes, 7.1% of military personnel, 3.2% of pregnant women, and 5.2% of tuberculosis patients were infected in 1997. Findings are presented from an investigation of the genetic and biological features of HIV-1 strains in Cambodia. Viruses from 95 HIV-1-seropositive individuals were subtyped by heteroduplex mobility assay (HMA) and 23 were further analyzed for their biologic characteristics. 89 people were clearly infected with HIV-1 subtype E. The other 6 samples, however, were sequenced together with 17 HMA subtype E samples. All but 1 of these latter 23 Cambodian env sequences clustered with previously described Thai and Vietnamese subtype E sequences bearing a GPGQ motif at the tip of the V3 loop, with the last having a GPGR motif and being phylogenetically equidistant from Asian and African subtype E viruses. Nonsyncytium-inducing, CCR5-dependent viruses predominated in patients of clinical stage B, and were detected in about half of the stage C patients. All syncytium-inducing strains, mostly from AIDS patients, used both CCR5 and CXCR4. The presence of syncytium-inducing viruses did not correlate with the plasma viral load. These data show that CCR5-dependent HIV-1 subtype E currently predominates in Cambodia. The dynamics of the viral population during subtype E infection in Southeast Asia appear to be similar to that of subtype B infection in Europe and the US.
