Showing papers in "AIDS Research and Human Retroviruses in 2000"
TL;DR: Additional mutagenesis indicates that important residues in this region for CCR5 binding are Ile-420, Lys-421, Gln-422, Pro-438, and Gly-441, suggesting a mechanism whereby interdomain conformational shifts induced by CD4 binding could facilitate C CR5 binding.
Abstract: A previous study implicated a conserved surface of the human immunodeficiency virus (HIV-1) gp120 exterior envelope glycoprotein in binding the CCR5 viral coreceptor (Rizzuto C, Wyatt R, Hernandez-Ramos N, Sun Y, Kwong PD, Hendrickson WA, and Sodroski J: Science 1998;280:1949-1953). Additional mutagenesis indicates that important residues in this region for CCR5 binding are Ile-420, Lys-421, Gln-422, Pro-438, and Gly-441. These highly conserved residues are located on two strands that connect the gp120 bridging sheet and outer domain, suggesting a mechanism whereby interdomain conformational shifts induced by CD4 binding could facilitate CCR5 binding.
195 citations
TL;DR: This work establishes a quantitative one-tube assay based on TaqMan chemistry that facilitates and enhances the quantification of plasma RNA loads, a critical parameter for many studies, including evaluations of vaccine candidates or antiviral regimens.
Abstract: Plasma viral RNA load is a key parameter in disease progression of lentiviral infections. To measure simian immunodeficiency virus (SIV) RNA loads, we have established a quantitative one-tube assay based on TaqMan chemistry. This real-time reverse transcriptase-polymerase chain reaction (RT-PCR) has advantages compared with previous methods, such as higher sensitivity, shorter time consumption, and low risk of cross-contamination. The sensitivity of the assay was optimized by comparing different enzyme systems. The one-enzyme protocol using rTth DNA polymerase was superior to two assays employing two enzymes. It detects 100% of the samples containing four copies of RNA transcript and allows quantification of viral RNA loads over an 8-log unit dynamic range. As few as 50 copies per milliliter of plasma can be detected within RNA extracted from 140 microl of plasma. This is especially relevant in studies employing neonatal macaques, from which only small volumes of blood can be sampled, and in studies in which low viral RNA loads are expected. Because of the use of rTth DNA polymerase, DNA contamination can be avoided by carryover prevention with uracil N-glycosylase (UNG). We demonstrate that for optimization of real-time PCR sensitivity, not only concentrations of different reagents but also different enzyme systems must be evaluated. Our assay facilitates and enhances the quantification of plasma RNA loads, a critical parameter for many studies, including evaluations of vaccine candidates or antiviral regimens.
172 citations
TL;DR: The notion that monovalent HIV-1 ENV is a poor immunogen for generating cross-reactive neutralizing antibodies is reinforced, as the majority of neutralizing activity was specific for the MN-V3 loop.
Abstract: Antibodies generated by candidate HIV-1 vaccines in a phase I clinical trial were assessed for neutralizing activity with a panel of eight well-characterized, genetically diverse clade B primary is...
148 citations
TL;DR: After a median follow-up of 15 months, no polymorphism was associated with parameters of disease progression among individuals failing treatment and only a limited set of amino acid substitutions can be tolerated at p7/p1 and p1/p6 cleavage sites.
Abstract: Amino acid substitutions at HIV-1 Gag p7/p1 and p1/p6 cleavage sites may be selected under antiretroviral pressure or represent natural polymorphisms. Whether changes are associated with specific protease (PR) mutation patterns and different clinical evolution has not been investigated. p7/p1 and p1/p6 cleavage site sequences from sera from 110 patients infected with HIV-1 were compared by regression analysis, using clinical, laboratory, and sequence variables, and the evolution of CD4+ cell counts and viral load over time. Sixteen of 35 (46%) individuals naive to PR inhibitors (PIs), and 49 of 75 (65%) receiving PI-containing regimens had a p7/p1 and/or p1/p6 cleavage site polymorphism (p = 0.06). A431V and/or L449F were present exclusively among individuals failing PI treatment (17 of 75 [23%] and 3 of 75 [3%], respectively). There was a significant association between A431V and PR M46I,L (OR 13.7; 95% CI 4.2-44.3) and V82A,F,T (OR 8.8; 95% CI 2.7-27.8). Natural polymorphism P453L was strongly associate...
137 citations
TL;DR: In this study, three rhesus macaques immunized with HIV-1 p55gag virus-like particles (VLPs) in the absence of adjuvant indicated that immunization with p 55gag VLP primes strong MHC class I-restricted, CD8+ cell-mediated immune responses and suggested that AIDS pandemic vaccine should be a reasonable vaccine candidate, when combined with strategies priming cross-neutralizing antibodies.
Abstract: Despite advances in the clinical management of HIV infection, using combinations of antiretroviral pharmaceuticals, a safe and efficacious vaccine is needed to limit the AIDS pandemic. It is now th...
133 citations
TL;DR: The equidistance of subtype K to the other subtypes of HIV-1 suggests that this subtype existed as long as the others, the lower distance between B and D, and between F1 and F2 suggest a more recent subdivision for these latter strains.
Abstract: We recently reported a high divergence among African subtype F strains. Three well-separated groups (F1, F2, and F3) have been shown based on the phylogenetic analysis of the p24 gag and envelope sequences with genetic distances similar to those observed for known subtypes. In this study, we characterized the near-full-length genomes of two strains from epidemiological unlinked individual belonging to each of the subgroups: F1 (96FR-MP411), F2 (95CM-MP255 and 95CM-MP257), and F3 (96CM-MP535 and 97ZR-EQTB11). Phylogenetic analysis of the near-full-length sequences and for each of the genes separately showed the same three groups, supported by high bootstrap values. Diversity plotting, BLAST subtyping, and bootstrap plotting confirmed that the divergent F strains correspond to nonrecombinant viruses. The divergence between F1 and F2 is consistently lower than that seen in any other intersubtype comparison, with the exception of subtypes B and D. Based on all the different analyses, we propose to divide subt...
114 citations
TL;DR: The absence of an HERV-W replication-competent provirus versus the existence of HERv-W-related Gag and Env proteins in healthy human placenta is discussed with respect to particle formation, physiology, and pathology.
Abstract: Some genomic elements of the multicopy HERV-W endogenous retroviral family have been previously identified in databases. One of them, located on chromosome 7, contains a single complete open reading frame (ORF) putatively encoding an envelope protein. We have experimentally investigated the genomic complexity and coding capacity of the HERV-W family. The human haploid genome contains at least 70, 100, and 30 HERV-W-related gag, pro, and env regions, respectively, widely and heterogeneously dispersed among chromosomes. Using in vitro transcription–translation procedures, three putative HERV-W gag, pro, and env ORFs were detected on chromosomes 3, 6, and 7, respectively, and their sequences analyzed. A 363 amino acid gag ORF containing matrix and carboxy-terminal truncated capsid domains encoded a putative 45-kDa protein. No gag–pro ORF was found, but a pro sequence containing a DTG active site was detected. Finally, the previously described 538 amino acid HERV-W env ORF, located on chromosome 7, was shown ...
106 citations
TL;DR: Thalidomide therapy was associated with mild to moderate rashes and fevers, but not peripheral neuropathy, and the anabolic benefits of high-dose thalidomides are limited by drug intolerance, 8 weeks of low- doses results in significant weight gain in patients with AIDS-associated wasting.
Abstract: A double-blind, placebo-controlled trial of efficacy and safety of thalidomide in AIDS-associated wasting was carried out. Ninety-nine of 103 male patients had at least one on-study measurement (intent-to-treat [ITT] cohort). Patients were randomized to thalidomide at 100 mg/day (T100) or 200 mg/day (T200), or placebo for 8 weeks. By ITT analysis, the mean change in body weight of the placebo, T100, and T200 treatment groups was 0.3 kg (0.4%), 2.0 kg (3.0%), and 0.9 kg (1.4%), respectively (p = 0.021 for T100 versus placebo; p = 0.53 for T200 versus placebo). Of the 64 patients who completed the 8 weeks of study treatment, significant weight gain was observed in both the T100 group (2.2 kg, [33%]; p = 0.008 versus placebo) and the T200 group (1.5 kg [2.5%]; p = 0.019 versus placebo). Approximately half the weight gain was fat-free mass (bioimpedance analysis). Patients in the T100 or T200 groups had no significant change in CD4+ cell counts, neutrophil counts, or TNF-α levels, compared with placebo. HIV v...
93 citations
TL;DR: 8 days of PI therapy in HIV-infected patients does not significantly alter plasma viremia, yet results in significant inhibition of CD4+ and CD8+ T cell apoptosis, have implications concerning the treatment of HIV and its pathogenesis.
Abstract: Antiretroviral treatment of patients infected with HIV results in improvements in CD4+ T cell number. Emerging evidence suggests that some of the improvements in CD4+ T cell number that occur in response to protease inhibitor (PI) therapy may not be accounted for solely by enhanced viral suppression, implying that PI may directly affect T cell survival. Since HIV T cell depletion is associated with enhanced apoptosis, we analyzed the effect of PIs on T cell apoptosis. In vitro treatment of peripheral blood lymphocytes (PBLs) from HIV-infected but untreated patients with reverse transcriptase inhibitors (RTIs) does not alter apoptosis, whereas PI treatment rapidly reduces CD4+ and CD8+ T cell apoptosis. In contrast, PI treatment does not alter apoptosis in PBL blasts from HIV-negative patients, or in Jurkat T cells. Consistent with this observation, 8 days of PI therapy in HIV-infected patients does not significantly alter plasma viremia, yet results in significant inhibition of CD4+ and CD8+ T cell apoptosis. The inhibitory effects of PI on apoptosis have implications concerning the treatment of HIV and its pathogenesis.
93 citations
TL;DR: It is demonstrated that transplacental AZT exposure causes cardiac and skeletal muscle mitochondrial myopathy in the patas monkey fetus and dose-dependent depletion of mitochondrial DNA levels was observed in both tissues.
Abstract: 3'-azido-3'-deoxythymidine (AZT) is given to pregnant women positive for the human immunodeficiency virus type 1 (HIV-1) to reduce maternal–fetal viral transmission. To explore fetal mitochondrial consequences of this exposure, pregnant Erythrocebus patas monkeys were given daily doses of 1.5 mg (21% of the human daily dose) and 6.0 mg (86% of the human daily dose) of AZT/kg body weight (bw), for the second half of gestation. At term, electron microscopy of fetal cardiac and skeletal muscle showed abnormal and disrupted sarcomeres with myofibrillar loss. Some abnormally shaped mitochondria with disrupted cristae were observed in skeletal muscle myocytes. Oxidative phosphorylation (OXPHOS) enzyme assays showed dose-dependent alterations. At the human-equivalent dose of AZT (6 mg of AZT/kg bw), there was an ~85% decrease in the specific activity of NADH dehydrogenase (complex I) and three- to sixfold increases in specific activities of succinate dehydrogenase (complex II) and cytochrome-c oxidase (complex I...
92 citations
TL;DR: Focusing the search for the origins of HIV-1 and HIV-2 on this earlier time period may contribute to understanding the evolution of the HIV viruses and the dynamics of emerging diseases
Abstract: Sociocultural factors during the postcolonial period have been implicated as paramount in generating conditions that promoted both the origin and subsequent epidemic spread of HIV-1 in Africa. We suggest, however, that the origin of the disease may lie in the interaction between colonial practices (e.g., labor camps, nonsterile vaccination campaigns) and traditional bushmeat hunting in French Equatorial Africa. Both the epidemiology of HIV-2 and the colonial history of West Africa appear more complex, but similar conditions existed there and may have contributed to the origin of HIV-2. Focusing the search for the origins of HIV-1 and HIV-2 on this earlier time period may contribute to understanding the evolution of the HIV viruses and the dynamics of emerging diseases.
University College London1, Duke University2, University of Liverpool3, University of Minnesota4, University of Washington5, University of New South Wales6, Bristol-Myers Squibb7, Iowa State University8, GlaxoSmithKline9, Harvard University10, University of North Carolina at Chapel Hill11, University of Miami12, Medical College of Wisconsin13, Veterans Health Administration14, Regions Hospital15, University of California, San Francisco16, University of Paris17, Stanford University18, Yeshiva University19, University of Texas Medical Branch20, University of California, San Diego21, University of Alabama at Birmingham22, Hoffmann-La Roche23, University of Colorado Denver24, Goethe University Frankfurt25, North Carolina State University26
TL;DR: Change in HIV-1 RNA, particularly using AUCMB, and in CD4 count should be measured to aid patient management and evaluation of treatment activity in clinical trials because two randomized treatment comparisons may show similar differences between treatments in marker changes but not similar differences in progression to AIDS/death.
Abstract: Objective: To evaluate treatment-mediated changes in HIV-1 RNA and CD3 count as prognostic markers and surrogate end points for disease progression (AIDS/death). Methods: Data from 13,045 subjects in all 16 randomized trials comparing nucleoside analogue reverse transcriptase inhibitors and having HIV-1 RNA measurements at 24 weeks were obtained. A total of 3146 subjects had HIV-1 RNA and CD3 count determinations at 24 weeks after starting treatment. Results: At Week 24, the percentage of subjects experiencing an HIV-1 RNA decrease of >1 log(10) copies/ml or a CD4 count increase of >33% was similar (22% vs 25%). Changes in both markers at Week 24 mere significant independent predictors of AIDS/death: across trials, the average reduction in hazard was 51% per 1 log(10) HIV-1 RNA copies/ml decrease (95% confidence interval: 41%, 59%) and 20% per 33% CD4 count increase (17%, 24%). In univariate analyses, the hazard ratio for AIDS/death in randomized treatment comparisons was significantly associated with differences between treatments in mean area under the curve of HIV-1 RNA changes to Weeks 8 and 24 (AUCMB) and mean CD3 change at Week 24, but, in multivariate analysis, only mean CD4 change was significant. Conclusions: Change in HIV-1 RNA, particularly using AUCMB, and in CD4 count should be measured to aid patient management and evaluation of treatment activity in clinical trials. However, short-term changes in these markers are imperfect as surrogate end points for long-term clinical outcome because two randomized treatment comparisons may show similar differences between treatments in marker changes but not similar differences in progression to AIDS/death.
Jacobi Medical Center1, Harvard University2, University of California, Los Angeles3, Medical University of South Carolina4, National Institutes of Health5, Johns Hopkins University6, Rutgers University7, University of Southern California8, Metropolitan Hospital Center9, Harlem Hospital Center10, Howard University11, George Washington University12, Stony Brook University13
TL;DR: A change in antiretroviral therapy to a protease inhibitor-containing regimen was associated with a virological response rate of approximately 50% for this patient population.
Abstract: One hundred eighty-one antiretroviral-experienced, protease inhibitor-naive, clinically stable HIV-infected children between 4 months and 17 years of age were randomly assigned to receive one of four combination regimens to evaluate the change in plasma HIV RNA, safety, and tolerance when changing antiretroviral therapy to a protease inhibitor-containing combination regimen. All four regimens contained stavudine; in addition children received nevirapine plus ritonavir, lamivudine plus nelfinavir, nevirapine plus nelfinavir, or lamivudine plus nevirapine plus nelfinavir. Twelve additional children chose to receive stavudine plus lamivudine plus nelfinavir, with nelfinavir given bid, rather than tid as for the main regimens. Overall, 51% (89/176; 95% CI 43-58%) of the children on the randomized portion of the study had an HIV RNA response (≤400 copies/ml) on at least two of the three HIV RNA determinations taken at Weeks 8, 12, and 16. At Week 24 the proportion of children with an HIV RNA response still on ...
TL;DR: A limited number of HIV-1 subtypes are observed in Nigeria, with subtypes A and G being the major env subtypes responsible for the HIV- 1 epidemic, and the high rate of recombinant viruses (A/G) and the different A/G recombinant structures indicate a complex pattern of virus circulating in this country.
Abstract: The purpose of this study was to generate data on the relative prevalences of the HIV-1 subtypes circulating in Nigeria. A total of 252 HIV-1-positive samples collected during an epidemiologic survey conducted in April 1996 were genetically characterized by HMA (heteroduplex mobility assay) and/or sequencing. Samples were collected in Lagos, Calabar, Kano, and Maiduguri. Overall, the predominant env subtypes were A (61.3%) and G (37.5%). Subtype A is more prevalent in the south (p < 0.001), about 70% in Lagos and Calabar, whereas a quarter of the samples was classified as subtype G in these states. In contrast, subtype G is predominant in the north ( < 0.001), representing 58% of the samples in Kano. In the northeastern region, Maiduguri, almost similar proportions of subtype A and G were seen, 49 and 47.4%, respectively. A total of 37 samples was also sequenced in the p24 region from the gag gene; 13 (35%) had discordant subtype designations between env and gag. The majority of the gag (12 of 17) and env (14 of 22) subtype A sequences clustered with the A/G-IBNG strain. Within subtype G, three different subclusters were seen among the envelope sequences. These different subclusters are observed among samples obtained from asymptomatic individuals and AIDS patients from the four Nigerian states studied. In conclusion, we observed a limited number of HIV-1 subtypes circulating in Nigeria, with subtypes A and G being the major env subtypes responsible for the HIV-1 epidemic. Nevertheless, the high rate of recombinant viruses (A/G) and the different A/G recombinant structures indicate a complex pattern of HIV-1 viruses circulating in this country.
TL;DR: It is proposed that Abt96 is a new HIV-2 subtype and designate it subtype G, and Phylogenetic analysis of sequences derived from segments of the AbT96 genome demonstrate that the Abt 96 isolate branches independently of all other characterized HIV- 2 isolates.
Abstract: Human immunodeficiency virus type 2 (HIV-2) and simian immunodeficiency virus from sooty mangabey (SIVSM) form one of the six primate lentivirus lineages. The close phylogenetic relationship and ge...
TL;DR: In this paper, the authors generated DNA constructs expressing soluble truncated forms of the envelope of SF162, a neutralizationresistant primary human immunodeficiency virus type 1 isolate, and SF162 Delta V2.
Abstract: We generated DNA constructs expressing soluble truncated forms of the envelope of SF162, a neutralizationresistant primary human immunodeficiency virus type 1 isolate, and SF162 Delta V2, a neutral...
TL;DR: An alternate strategy is based on the understanding of diseases induced by viruses closely related to HTLV-1, like bovine leukemia virus (BLV), which indeed belong to the same subfamily of retroviruses, harbor a similar genomic organization, and infect and transform cells of the hematopoietic system.
Abstract: The understanding of HTLV-induced disease is hampered by the lack of a suitable animal model allowing the study of both viral replication and leukemogenesis in vivo. Although valuable information has been obtained in different species, such as rabbits, mice, rats, and monkeys, none of these systems was able to conciliate topics as different as viral infectivity, propagation within the host, and generation of leukemic cells. An alternate strategy is based on the understanding of diseases induced by viruses closely related to HTLV-1, like bovine leukemia virus (BLV). Both viruses indeed belong to the same subfamily of retroviruses, harbor a similar genomic organization, and infect and transform cells of the hematopoietic system. The main advantage of the BLV system is that it allows direct experimentation in two different species, cattle and sheep.
TL;DR: A total of 343 HIV-1-positive samples obtained between June 1996 and March 1999 was genetically characterized in the envelope region by HMA and/or sequencing, and in Senegal 323 (94.2%) of the 343 samples could be identified by this technique.
Abstract: A total of 343 HIV-1-positive samples obtained between June 1996 and March 1999 was genetically characterized in the envelope region by HMA and/or sequencing. The env subtype distribution was as follows: 290 (84.6%) A, 22 (6.5%) B, 16 (4.7%) C, 8 (2.5%) D, 1 (0.03%) E, 1 (0.03%) F1, 4 (1.2%) G, and 1 (0.03%) H. For 77 samples the p24 region from the gag gene was also sequenced, and for 9 (11.6%) the subtypes between env and gag were different. Phylogenetic tree analysis showed the predominance of AG-IBNG-like viruses among gag and env subtype A sequences. HMA is relatively simple and requires less sophisticated technical facilities compared with sequencing, and in Senegal 323 (94.2%) of the 343 samples could be identified by this technique. However, in the actual configuration of the assay, discrimination between the recombinant AG-IBNG-like recombinant viruses, which are predominant in Senegal, and the nonrecombinant subtype A viruses is not possible.
TL;DR: The results show that the binding of HIV-1 to nonsusceptible cells or immobilized adhesion ligands through acquired integrins can increase its infectivity by as much as two orders of magnitude.
Abstract: HIV-1 incorporates a variety of host membrane proteins during budding. We have previously shown that adhesion molecules are acquired by the virus in their activated or functional states. Our studies and those of others indicate that adhesion molecules can have profound effects on virus infectivity and its resistance to neutralization by antiviral antibodies. In this study we have examined the effect on infectivity of immobilization or margination of HIV-1 through acquired integrins LFA-1 and VLA-4 onto nonsusceptible cells and solid-phase adhesion ligands (ICAM-1 and VCAM-1, respectively). LFA-1- and VLA-4-mediated HIV-1 binding was supported by ICAM-1 and VCAM-1 immunoglobulin Fc chimeras, respectively. Integrin-mediated HIV-1 binding was also supported by 293 cells transfected with ICAM-1. In both cases the specificity of binding was confirmed with the appropriate blocking monoclonal antibodies or soluble adhesion ligands. We used a sensitive single-cycle infection assay based on a cell line expressing an LTR-luciferase cDNA construct to compare the infectivity of bound virus with that of free virus. Our results show that the binding of HIV-1 to nonsusceptible cells or immobilized adhesion ligands through acquired integrins can increase its infectivity by as much as two orders of magnitude. These results have implications for in vivo dissemination and transmission of HIV-1 and may also explain the high level of virus replication seen in solid lymphoid organs.
TL;DR: It is demonstrated that most incident HIV1 infections among Bangkok IDUs are due to subtype E viruses, with a narrow spectrum of genetic diversity.
Abstract: We obtained specimens from 128 HIV-1 seroconverters identified from 1995 through 1998 in a prospective cohort study of 1209 HIV-negative injecting drug users (IDUs) in Bangkok, Thailand. Epidemiologic data indicated that parenteral transmission accounted for nearly all infections. HIV-1 DNA from the C2–V4 env region was sequenced, and phylogenetic analyses determined that 102 (79.7%) of the specimens were subtype E and 26 (20.3%) subtype B strains. All subtype B strains clustered with strains often referred to in previous studies as Thai B or B. The interstrain nucleotide distance (C2–V4) within subtype E strains was low (mean, 6.8%), and pairwise comparisons with a prototype subtype E strain, CM 244, showed limited divergence (mean, 5.6%). The subtype B stains showed greater interstrain divergence (mean, 9.2%) and were significantly divergent from the prototype B strain HIV-M N (mean, 13.0%; p < 0.0001). The subtype E strains had significantly lower mean V3 loop charge than did subtype B strains (p = 0.0...
TL;DR: The incorporation of mutagenic analogs into the HIV genome by the host cell RNA polymerase, rather than by the viral reverse transcriptase, augurs well for the avoidance of resistant viruses.
Abstract: The extraordinarily high mutation rate of HIV suggests that the viral population exists near the threshold for viral viability. An increase in mutation could mediate a lethal increase in the already high proportion of defective viruses. We initially tested this hypothesis by culturing HIV-infected cells in mutagenic deoxyribonucleoside analogs. In the presence of 5-hydroxydeoxycytidine, there was a loss of viral replication after sequential passages of HIV in human CEM cells (Loeb LA, et al.: Proc Natl Acad Sci USA 1999;96:14921497). We now propose to use mutagenic ribonucleosides to induce lethal mutagenesis. The incorporation of mutagenic analogs into the HIV genome by the host cell RNA polymerase, rather than by the viral reverse transcriptase, augurs well for the avoidance of resistant viruses.
TL;DR: The data suggest that multiple subtypes of B', C, and CRF01_AE are cocirculating in central Myanmar, leading to the evolution of new forms of intersubtype recombinants among the risk populations exhibiting one of the highest HIV infection rates in the region.
Abstract: We have previously shown that HIV-1 env subtypes B' (a Thai-B cluster within subtype B) and E (CRF01_AE) are distributed in Yangon, the capital city of Myanmar. However, HIV strains from the rest of country have not yet been genetically characterized. In the present study, we determined env (C2/V3) and gag (p17) subtypes of 25 specimens from central Myanmar (Mandalay). Phylogenetic analyses identified 5 subtype C (20%), in addition to 10 CRF01_AE (40%) and 4 subtype B' (16%). Interestingly, the remaining six specimens (24%) showed discordance between gag and env subtypes; three gag subtype B'/env subtype C, one gag subtype B'/env subtype E, one gag subtype C/env subtype B', and one gag subtype C/env subtype E. These discordant specimens were found frequently among injecting drug users (4 of 12, 33%) and female commercial sex workers (2 of 8, 25%) engaging in high-risk behaviors. The recombinant nature of these HIV-1 strains was verified in three specimens, indicating the presence of new forms of HIV-1 intersubtype C/B' and C/B'/E recombinants with different recombination breakpoints. The data suggest that multiple subtypes of B', C, and CRF01_AE are cocirculating in central Myanmar, leading to the evolution of new forms of intersubtype recombinants among the risk populations exhibiting one of the highest HIV infection rates in the region.
TL;DR: The genetic relationship of HIV-1 strains circulating among IDUs in the former SU is described and it is suggested that only short stretches of absolute sequence identity may be needed for efficient RNA recombination between HIV- 1 subtypes.
Abstract: In the former Soviet Union (SU) increasing numbers of HIV-1 infections among injecting drug users (IDU) have been reported, especially in the Ukraine. The main subtype transmitted among the IDUs seems to be subtype A, but limited numbers of subtype B cases have also been reported. In Kaliningrad, Russia, an AB recombinant strain was earlier shown to be responsible for the local outbreak. Here we describe the genetic relationship of HIV-1 strains circulating among IDUs in the former SU. For subtype A and the AB recombinant strains nearly full-length genomes were sequenced, and for one subtype B strain the entire envelope gene was cloned. The relationship between the AB recombinant strain and the subtype A and subtype B strains and the mosaic structure of the recombinant was studied by phylogenetic analysis. Ukrainian A and B strains were shown to be the probable parental viruses of the Kaliningrad AB recombinant strain. In the envelope gene the recombination breakpoint could also be precisely mapped to a region of similarity of only 14 base pairs. This suggests that only short stretches of absolute sequence identity may be needed for efficient RNA recombination between HIV-1 subtypes.
TL;DR: By analyzing conserved epitopes in the context of HLA restricting alleles, this work defines a set of HIV-1 gene regions that may have the greatest potential to induce cross-clade reactive CTLs and examines HLA allelic frequencies in several different populations to determine those that could contribute to the goal of a cumulative phenotype frequency of at least 80%.
Abstract: One of the fundamental goals of current strategies to develop an efficacious vaccine for AIDS is the elicitation of cytotoxic T lymphocyte (CTL) reactivities capable of recognizing cells infected with different subtypes of the human immunodeficiency virus type 1 (HIV-1). In efforts to explore new vaccine candidates by the UNAIDS/WHO Vaccine Committee, we review the most recent data concerning CTL epitopes that are conserved among the different HIV-1 subtypes. Moreover, we examine HLA allelic frequencies in several different populations, to determine those that could contribute to the goal of a cumulative phenotype frequency (CP) of at least 80%. By analyzing conserved epitopes in the context of HLA restricting alleles, we define a set of HIV-1 gene regions that may have the greatest potential to induce cross-clade reactive CTLs. The absence of well-defined correlates of immune protection that link CTL epitopes to delayed disease progression and/or prevention of infection does not permit an assignment of r...
TL;DR: By utilizing a human cDNA expression array blot, overexpression of various transcription factors, cell cycle regulated kinases, and DNA repair genes in HTLV-1-infected T cells is observed and the cyclin-dependent kinase inhibitor, p21/waf1, is one of the genes of interest.
Abstract: By utilizing a human cDNA expression array blot (588 genes), we have observed overexpression of various transcription factors, cell cycle regulated kinases, and DNA repair genes in HTLV-1-infected T cells. One of the genes of interest, and focus in this study, is the cyclin-dependent kinase inhibitor, p21/waf1. The p21/waf1 transcription and protein is overexpressed in all HTLV-1-infected cell lines tested as well as ATL and HAM/TSP patient samples. While p21/waf1 has been shown to display a selectivity for G1/S cyclin/cdk complexes, we have observed p21/waf1 to be complexed with cyclin A/cdk2. Functionally, the association of p21/cyclin A/cdk2 decreased the histone H1 phosphorylation in vitro, as observed in immunoprecipitations followed by kinase assays, as well as affecting other substrates such as the C-terminus of Rb protein involved in c-Abl and HDAC1 regulation. Wild-type, but not a mutant form (M47) of Tax, was found to be able to transactivate the p21/waf1 promoter in a p53-independent manner. We...
TL;DR: It is concluded that the CS mutations were required for optimum detection of resistance in the RVA, but that virus fitness can remain compromised even in the presence of compensatory CS mutations.
Abstract: The recombinant virus assay (RVA) is a method for assessing the susceptibility of human immunodeficiency virus type 1 (HIV-1) plasma isolates to antiretroviral drugs. The RVA involves the production of viable virus in vitro by homologous recombination of RT-PCR products from plasma virus with a noninfectious reverse transcriptase (RT) or protease (PR)-deleted cloned HIV-1 provirus. In this study, we have constructed RVA plasmids with contiguous deletions in RT, PR, and the p7/p1 and p1/6 gag protease cleavage sites (CS). The deletions in these plasmids allow generation of recombinant viruses with all loci currently identified as important for resistance to anti-HIV-1 drugs being derived from the clinical isolate, including CS mutations that compensate for the reduced fitness of viruses resistant to protease inhibitors (Doyon et al., J Virol 1996:70:3763-3769). We have also used these new constructs to generate viruses with or without compensatory CS mutations, and examined the effects on fitness. In the case of an indinavir-selected virus, fitness was restored close to that of a wild type virus when a vector deleted in the CS and PR was used. With an amprenavir-selected isolate, virus fitness was incompletely restored by including the CS, and this defect appeared to be partially due to reduced infectivity of the virions. We conclude that the CS mutations were required for optimum detection of resistance in the RVA, but that virus fitness can remain compromised even in the presence of compensatory CS mutations.
TL;DR: Several immunogens induce HIV-specific neutralization and in vitro lymphoproliferation in adults at low HIV1 risk, but responses in persons at high HIV-1 risk are not known.
Abstract: Several immunogens induce HIV-specific neutralization and in vitro lymphoproliferation in adults at low HIV-1 risk, but responses in persons at high HIV-1 risk are not known. We performed a multicenter, double-blinded, adjuvant-controlled trial with two gp120 vaccines in 296 HIV-1-uninfected volunteers, including 176 reporting higher HIV-1 risk activities. The immunogens were remarkably well tolerated. After three immunizations, 210 of 241 vaccinees (87%) developed neutralizing antibodies, which persisted in 59% after 2 years. The injection drug users receiving SF-2/gp120 had decreased antibody responses relative to the lower risk groups. Envelope-specific lymphoproliferation peaked after two immunizations, and 54% of vaccinees mounted a DTH reaction to gp120 after 4 years. In summary, these immunogens have low adverse reactogenicity and induce durable antibody and T cell responses to the prototype strains. Unexpected differences in antibody responses among diverse HIV-1 risk strata lend support to the conduct of expanded phase II trials in populations other than low-risk volunteers.
TL;DR: The results suggest that nef genes derived from pediatric long-term nonprogressors have gross deletions in isolated cases but a higher prevalence of discrete changes that may impair Nef function in primary T cell assays, but not all functions reported for Nef.
Abstract: DNA sequences and three distinct in vitro functions of Nef were evaluated in a group of seven perinatally infected children. nef gene sequences obtained before and after virus culture showed that one of the five non-/slow progressors harbored a virus with large deletions. nef genes from the remaining four children were full length but contained discrete changes at a higher frequency than the rapid progressors. In functional studies, 40 of 44 Nef proteins derived from the whole study group were capable of binding the cellular serine kinase p62, indicating that this function is well conserved among naturally occurring viruses. In contrast, representative Nef proteins derived from the long-term non-/slow progressors were found to be defective or far less capable of enhancing viral replication and/or viral infectivity in herpesvirus saimiri-transformed human T cells and peripheral blood mononuclear cells. On reversion of highly prevalent point mutations in the defective proteins, viral replication could be re...
TL;DR: Results indicate that genes altered on HIV-1 infection may be mutually organized and play an important role in HIV-2-induced pathogenesis and the expression of its receptor CCR5 was cooperatively upregulated on the surface of these cells.
Abstract: We used a novel differential display (DD) technique to identify host factors involved in virus replication, pathogenesis, and host response in HIV-1-infected T cells. Thirteen cDNA fragments differentially expressed in HIV-1NL4-3-infected MT-4 cells prior to the occurrence of specific apoptotic cell death were sequenced and identified. Two of seven elevated genes were identical to HIV-1 sequences and the other five were M IP-1α, ACTE-III, CD11c, arginase I, and CCR5. The six downregulated genes included prothymosin-α, Jaw-1, proteasome subunit XAPC7, splicing factor 9G8, GA17 protein, and an unknown mRNA. Northern blot and RTPCR analyses confirmed the altered gene expressions in MT-4 cells as well as in another T cell line, MOLT4. We also revealed that the amount of MIP-1α in culture supernatant of HIV-1-infected cells was increased by more than 15-fold relative to control cells, and the expression of its receptor CCR5 was cooperatively upregulated on the surface of these cells. Furthermore, the upregulat...
TL;DR: In this article, the authors measured total IgG1, IgG2, Ig3, and Ig4 concentrations by ELISA in serum (S), total saliva (TS), cervicovaginal secretions (CVS), seminal secretions, and rectal secretions.
Abstract: We measured total IgG1, IgG2, IgG3, and IgG4 concentrations by ELISA in serum (S), total saliva (TS), cervicovaginal secretions (CVS), seminal secretions (SPE), and rectal secretions (RS) from either CDC II/III HIV-1-infected subjects or healthy volunteers. Human serum albumin was measured in parallel to calculate the relative coefficient of excretion (RCE). Levels of IgG1 and IgG3 directed against gp120 MN also were measured by ELISA in all samples, and the specific activity (SA) calculated. HIV-1-specific IgG2 and IgG4 were not compared, as total IgG2 and total IgG4 levels in HIV-1-infected subjects were found to be lower than in the healthy controls. Despite substantial interindividual variability, total IgG1 and IgG3 concentrations in all fluids were greater in the HIV-1-infected subjects than in the healthy controls. Calculations of RCE indicated predominantly a transudative origin for IgG subclasses in the different mucosal fluids, except for CVS, in which IgG1, IgG2, and IgG4 was produced locally. The transduction behavior of IgG3 in secretions appears to be different from that of other IgG subclasses. HIV-1-infected subjects were considered positive for IgG1 and IgG3 antibodies against gp120 MN if their antibody levels exceeded the maximum titer measured in the control group. Positive levels of anti-gp120 MN IgG1 were detected for 100% of HIV-1-infected individuals in S, CVS, and SPE, 97% in TS, and 75% in RS. Fewer subjects had positive levels of IgG3 to gp120 MN in their secretions (maximum 67% in CVS). Despite the low concentrations of total IgG3, mean SA values for IgG3 to gp120 MN were greater in secretions than in serum. No significant difference in the SA values for IgG1 to gp120 MN was observed between the different fluids. Only CVS had a local production of HIV-specific IgG1 Our results highlight the importance of an HIV-specific IgG1 and IgG3 immune response in mucosal fluids from HIV-1-infected subjects.