Human Immunodeficiency Virus Type 1 Infected Individuals Research Articles

Genetic and functional heterogeneity of CNS-derived tat alleles from patients with HIV-associated dementia

Human immunodeficiency virus type 1 (HIV-1) demonstrates a high degree of viral diversity which has an impact on viral fitness. Genetic compartmentalization of HIV-1 proteins between central nervous system (CNS) and lymphoid tissues is well established and reflects altered requirements for HIV-1 replication in macrophages/microglia, brain-specific immune selection pressures and possibly the timing of virus invasion of the CNS. Tat-encoding mRNA has been detected in the CNS of HIV-1 infected individuals and its neurotoxic effects in the CNS are well documented. However, while CNS-derived tat sequences have demonstrated significant diversity, the effect of this molecular diversity on transcriptional regulation and its impact on the pathogenesis of HIV-associated dementia (HAD) remains unclear. In this study, we cloned and characterised 44 unique tat alleles from brain, cerebral spinal fluid, spinal cord and blood/lymphoid tissue-derived HIV-1 isolates from five subjects with HAD. While phylogenetic analyses revealed tissue-specific compartmentalization of Tat variants for two patients, broad compartmentalization across the panel of tissue-derived viruses was not observed. Despite the lack of consistent tissue-specific compartmentalization, sequence variations within patients segregated CNS and non-CNS tat alleles. These amino acid alterations predominated within the transactivation domain of Tat and could account for alterations in the ability of particular Tat proteins to transactivate the LTR. Although a subset of patients demonstrated reduced transactivation capacity among CNS-derived Tat proteins compared to those from matched lymphoid tissues, overall Tat proteins from the CNS to lymphoid compartments maintained similar levels of transactivation function. Together, these data suggest that despite the observed heterogeneity in tat alleles isolated from matched lymphoid to CNS compartments, Tat function is maintained, highlighting the importance of Tat function in HIV-1 neuropathogenesis.

Anti–HIV‐1 Activity of Weekly or Biweekly Treatment with Subcutaneous PRO 140, a CCR5 Monoclonal Antibody

PRO 140 is a humanized CCR5 monoclonal antibody that has demonstrated potent antiviral activity when it is administered intravenously to adults infected with CCR5-tropic (R5) human immunodeficiency virus type 1 (HIV-1). This study is the first to evaluate subcutaneous administration. A randomized, double-blind, placebo-controlled study was conducted among 44 subjects with HIV-1 RNA levels of >5000 copies/mL, CD4(+) cell counts of >300 cells/microL, no receipt of antiretroviral therapy for >or=12 weeks, and only R5 HIV-1 detectable. Subjects received placebo, 162 mg of PRO 140, or 324 mg of PRO 140 weekly for 3 weeks or 324 mg of PRO 140 every other week for 2 doses by means of subcutaneous infusion. Subjects were monitored for 58 days for safety, antiviral effects, and PRO 140 serum concentrations. Subcutaneous PRO 140 demonstrated potent and prolonged antiretroviral activity. Mean log(10) reductions in HIV-1 RNA level were 0.23, 0.99 (P=.009), 1.37 (P<.001), and 1.65 (P<.001) for the placebo, 162 mg weekly, 324 mg biweekly, and 324 mg weekly dose groups, respectively. Viral loads remained suppressed between successive doses. Treatment was generally well tolerated. This trial demonstrates proof of concept for a monoclonal antibody administered subcutaneously in HIV-1 infected individuals. Subcutaneous PRO 140 offers the potential for significant dose-dependent HIV-1 RNA suppression and infrequent patient self-administration. ClinicalTrials.gov identifier: NCT00642707 .

Association of HLA-A, B, DRB1 alleles and haplotypes with HIV-1 infection in Chongqing, China.

BackgroundThe human immunodeficiency virus type 1(HIV-1) epidemic in Chongqing, China, is increasing rapidly with the dominant subtype of CRF07_BC over the past 3 years. Since human leukocyte antigen (HLA) polymorphisms have shown strong association with susceptibility/resistance to HIV-1 infection from individuals with different ethnic backgrounds, a recent investigation on frequencies of HLA class I and class II alleles in a Chinese cohort also indicated that similar correlation existed in HIV infected individuals from several provinces in China, however, such information is unavailable in Chongqing, southwest China.MethodsIn this population-based study, we performed polymerase chain reaction analysis with sequence-specific oligonucleotide probes (PCR-SSOP) for intermediate-low-resolution HLA typing in a cohort of 549 HIV-1 infected individuals, another 2475 healthy subjects from the Han nationality in Chongqing, China, were selected as population control. We compared frequencies of HLA-A, B, DRB1 alleles, haplotypes and genotypes between the two groups, and analyzed their association with HIV-1 susceptibility or resistance.ResultsThe genetic profile of HLA (A, B, DRB1) alleles of HIV-1 infected individuals from Chongqing Han of China was obtained. Several alleles of HLA-B such as B*46 (P = 0.001, OR = 1.38, 95%CI = 1.13-1.68), B*1501G(B62) (P = 0.013, OR = 1.42, 95%CI = 1.08-1.88), B*67 (P = 0.022, OR = 2.76, 95%CI = 1.16-6.57), B*37 (P = 0.014, OR = 1.93, 95%CI = 1.14-3.28) and B*52 (P = 0.038, OR = 1.64, 95%CI = 1.03-2.61) were observed to have association with susceptibility to HIV-1 infection in this population. In addition, the haplotype analysis revealed that A*11-B*46, A*24-B*54 and A*01-B*37 for 2-locus, and A*11-B*46-DRB1*09, A*02-B*46-DRB1*08, A*11-B*4001G-DRB1*15, A*02-B*4001G-DRB1*04, A*11-B*46-DRB1*08 and A*02-B*4001G-DRB1*12 for 3-locus had significantly overrepresented in HIV-1 infected individuals, whereas A*11-B*1502G, A*11-B*1502G-DRB1*12 and A*33-B*58-DRB1*13 were underrepresented. However, the low-resolution homozygosity of HLA-A, B, DRB1 loci and HLA-Bw4/Bw6 genotypes did not differ significantly between the two groups.ConclusionThese results may contribute to the database of HLA profiles in HIV-1 infected Chinese population, consequently, the association of certain HLA alleles with susceptibility or resistance to HIV-1 infection would provide with clues in choosing proper preventive strategies against HIV-1 infection and developing effective HIV-1 vaccines in Chinese population, especially for those in southwest China.

Primary Antiretroviral Drug Resistance among HIV Type 1-Infected Individuals in Brazil

Infection with drug-resistant human immunodeficiency virus type 1 (HIV-1) has been documented in all countries that have surveyed for it and may result in an unfavorable response to therapy. The prevalence and characteristics of individuals with transmitted resistance to antiretroviral drugs have been scarcely described in Brazil. We performed antiretroviral resistance testing prior to initiation of therapy in 400 subjects enrolled from 20 centers in 13 Brazilian cities between March and September 2007. Genotyping was conducted using PCR-amplified HIV pol products by automated sequencing, and genotype interpretation was done according to the IAS-USA consensus. Of 400 eligible participants, 387 (95.8%) were successfully tested. Seven percent of antiretroviral-naive patients carried viruses with one or more major mutation associated with drug resistance. The prevalence of these mutations was 1.0% for protease inhibitors, 4.4% for nonnucleoside reverse transcriptase inhibitors, and 1.3% for nucleoside reverse transcriptase inhibitors. The frequency of multidrug resistance among the resistant strains was 13.6%. Among subjects infected with drug-resistant virus, the majority were infected with subtype B viruses (91%). Subjects from the city of São Paulo had higher transmitted resistance mutations compared to the rest of the country. Reporting a partner taking antiretroviral medications was associated with a higher chance of harboring HIV variants with major drug resistance mutations [odds ratio = 2.57 (95% confidence interval, 1.07-6.16); p = 0.014]. Resistance testing in drug-naive individuals identified 7% of subjects with mutations associated with reduced susceptibility to antiretroviral drugs. Continued surveillance of drug-resistant HIV-1 in Brazil is warranted when guidelines for HIV prophylaxis and treatment are updated. Resistance testing among drug-naive patients prior to treatment initiation should be considered, mainly directed at subjects whose partners are already on antiretroviral therapy.

The Apoptosis-Inducing Effects of HIV vpr Recombinant Eukaryotic Expression Vectors with Different Mutation Sites on Transfected Hela Cells

The vpr gene of human immunodeficiency virus type 1 (HIV-1) plays an important role in the pathogenicity of viruses. Previous studies showed that mutation of certain sites in HIV-1 Vpr amino acid sequences might influence the clinical process of infected individuals and attenuate its apoptosis-inducing capacity on infected cells. The present study was designed to transfect hela cells with several HIV-1 vpr fragments carrying specific mutation sites, and to observe the distinction of apoptosis-inducing effects of different HIV-1 vpr variant fragments on host cells and explore the possible mechanisms. According to the previous results, 14 typical vpr variant fragments were chosen from Chinese HIV-1 infected individuals. After PCR amplification of vpr gene, the products were purified and double digested with Hind and BamHI. Then pcDNA3.1 (+) eukaryotic expression plasmids were used for the ligation and transduction experiments. The recombinant plasmids were transiently transfected into Hela cells with liposomes, and meanwhile the blank cell and empty vector cell were established as control. RT-PCR was used to detect the mRNA expression of target genes; fluorescent microscope for observing apoptotic cells with Hoechst staining; DNA agarose electrophoresis for detecting apoptotic gradient band; Annexin assay for detecting cell apoptosis; and Caspase activity test for exploring the pathway of cell apoptosis. After transfected with 14 vpr gene segments, the Hela cells exhibited various apoptosis-inducing capabilities. We found that the HIV-1 Vpr segments with mutation at the 70(th), 85(th), 86(th), or 94(th) site showed lower apoptosis-inducing capabilities than other segments on Hela cells, and the apoptosis-inducing ability of HIV-1 vpr gene on host cells might be related to its subtype. Meanwhile, we found that the Caspase3 activity of transfected cell carrying these mutated fragment sites was decreased as compared to the cell with other fragments. This study firstly found that the HIV-1 Vpr fragments with the 70(th), 85(th), 86(th), or 94(th) site mutation might attenuate their apoptosis-inducing abilities on Hela cells. One of the mechanisms might be the attenuation of Caspase-3 activity.

HIV-1 Superinfection in the Antiretroviral Therapy Era: Are Seroconcordant Sexual Partners at Risk?

BackgroundAcquisition of more than one strain of human immunodeficiency virus type 1 (HIV-1) has been reported to occur both during and after primary infection, but the risks and repercussions of dual and superinfection are incompletely understood. In this study, we evaluated a longitudinal cohort of chronically HIV-infected men who were sexual partners to determine if individuals acquired their partners' viral strains.MethodologyOur cohort of HIV-positive men consisted of 8 couples that identified themselves as long-term sexual partners. Viral sequences were isolated from each subject and analyzed using phylogenetic methods. In addition, strain-specific PCR allowed us to search for partners' viruses present at low levels. Finally, we used computational algorithms to evaluate for recombination between partners' viral strains.Principal Findings/ConclusionsAll couples had at least one factor associated with increased risk for acquisition of new HIV strains during the study, including detectable plasma viral load, sexually transmitted infections, and unprotected sex. One subject was dually HIV-1 infected, but neither strain corresponded to that of his partner. Three couples' sequences formed monophyletic clusters at the entry visit, with phylogenetic analysis suggesting that one member of the couple had acquired an HIV strain from his identified partner or that both had acquired it from the same source outside their partnership. The 5 remaining couples initially displayed no evidence of dual infection, using phylogenetic analysis and strain-specific PCR. However, in 1 of these couples, further analysis revealed recombinant viral strains with segments of viral genomes in one subject that may have derived from the enrolled partner. Thus, chronically HIV-1 infected individuals may become superinfected with additional HIV strains from their seroconcordant sexual partners. In some cases, HIV-1 superinfection may become apparent when recombinant viral strains are detected.

Raltegravir: The First HIV Type 1 Integrase Inhibitor

Raltegravir is the first approved human immunodeficiency virus type 1 (HIV-1) integrase inhibitor; it targets the strand transfer step of HIV-1 integration. Clinical trials have demonstrated that raltegravir-containing regimens have potent antiretroviral activity and are well tolerated in HIV-1-infected individuals. In antiretroviral treatment-experienced persons with drug-resistant HIV infection, raltegravir-containing treatment with an optimized background regimen was superior to an optimized background regimen alone. In treatment-naive persons, raltegravir was not inferior to efavirenz when the drugs were administered with tenofovir and lamivudine or emtricitabine. Raltegravir is metabolized by glucuronidation, not hepatically; thus, the potential for drug-drug interactions is decreased. Drug resistance, conferred by substitutions in the gene coding for the HIV-1 integrase enzyme, develops relatively frequently after virologic failure. As an antiretroviral drug with a novel mechanism of action, raltegravir is an important advancement in HIV-1 treatment options.

Incidence of Human Immunodeficiency Virus Type 1 Dual Infections in Amsterdam, The Netherlands, during 2003–2007

The occurrence of human immunodeficiency virus type 1 (HIV-1) dual infections in Amsterdam, The Netherlands, was examined during 2003-2007 to investigate whether the number of HIV-1 dual infections increased as the number of HIV-1 infected individuals increased during the same period. All first HIV-1 genotyping sequences obtained from 2003 through 2007 were retrieved and examined for the number of degenerate base codes in the reverse-transcriptase fragment. A total of 72 patients had >or=34 degenerate base codes; for these patients, a fragment of the V3-V4 region of the env gene was amplified, cloned, and sequenced to verify the presence of an HIV-1 dual infection. The number of dual infections were counted for each year investigated. No significant change in the incidence of dual infections was observed in our population of patients, who were selected on the basis of the number of degenerate base codes in each patient's first HIV-1 sequence obtained from 2003 through 2007. The frequency of HIV-1 dual infections varied between 1.0% and 2.4% each year, with no significant trend over time (P = .49). Patients with HIV-1 dual infections were similar to patients with single HIV-1 infections in The Netherlands with regard to distribution of risk group, sex, and HIV subtype. The proportion of HIV-1 dual infections in The Netherlands did not increase from 2003 through 2007, although the HIV-1-infected population expanded in this period.

Profile of HIV Type 1 Infection and Genotypic Resistance Mutations to Antiretroviral Drugs in Treatment-Naive HIV Type 1-Infected Individuals in Hai Phong, Viet Nam

We evaluated the prevalence and profile of antiretroviral treatment (ART)-associated resistance mutations among HIV-1 strains in northern Vietnam by genotypically analyzing strains isolated from ART-naive individuals in Hai Phong, a city in which HIV-1 is highly prevalent. Plasma samples were collected from injecting drug users (IDU, n = 760), female sex workers (FSW, n = 91), seafarers (n = 94), pregnant women (n = 200), and blood donors (n = 210), and screened for HIV-1 antibodies. Plasma viral RNA was extracted from HIV-1-positive samples, amplified by reverse transcriptase (RT)-PCR of protease and RT genes, and analyzed for genotypes and ART-associated resistance mutations. HIV-1 prevalence among IDU, FSW, seafarers, pregnant women, and blood donors was 35.9%, 23.1%, 0%, 0.5%, and 2.9%, respectively. Phylogenetic analyses revealed that the most prevalent HIV-1 subtype was CRF01_AE (98.3%), similar to strains prevalent in southern China. Four (1.4%) subtype B strains and one (0.3%) unique recombinant between subtypes B and C were also identified. We found protease inhibitor-associated major resistance mutations in one of the 294 cases analyzed (0.3%; mutation M46I). We found RT inhibitor-associated major resistance mutations in 7/273 cases (2.6%; one occurrence each of L74I, M184I, and K219E; three cases of K103N; and two cases of G190E). One CRF01_AE strain harboring a protease codon 35 insertion was first identified in Vietnam. Thus, monitoring of drug-resistant HIV-1 and establishment of a database are required for the proper selection of ART in Vietnam.

Human Herpesvirus 8 DNA Load in the Leukocytes Correlates with the Platelet Counts in HIV Type 1-Infected Individuals

Abstract Human herpes virus 8 (HHV-8) is known to be reactivated in immunocompromised situations and it is associated with Kaposi's sarcoma (KS) and some hematological diseases. The aim of this study was to analyze the effect of HHV-8 on HIV-1 infection, especially on thrombocytopenia complicated with HIV infection. The HHV-8 DNA load was determined by a quantitative real-time PCR, using leukocytes from 125 HIV-1-infected individuals. HHV-8 DNA was detected in 37 individuals. The increased HIV-1 load and reduced percentage of CD4-positive T cells were significantly associated with the presence of HHV-8. The prevalence and load for HHV-8 are higher in patients with KS than in patients without KS, but the difference is not significant. The increased HHV-8 DNA load was significantly correlated with thrombocytopenia, and platelet counts were significantly lower in individuals with HHV-8 than in individuals without HHV-8. We also obtained the negative correlations between changes in platelet counts and changes in HHV-8 DNA loads. The association between thrombocytopenia and HHV-8 has never been reported previously, apart from some case reports of Castleman's disease and KS. Various cytokines or chemokines are produced by HHV-8-infected cells, some of which have been reported to inhibit hematopoiesis. This may be one of the mechanisms by which HHV-8 infection induces thrombocytopenia. These results indicate that HHV-8 DNA in leukocytes may provide useful information for the assessment of the clinical appearance of HIV-1 infection.

Vitamin D Deficiency among HIV Type 1-Infected Individuals in the Netherlands: Effects of Antiretroviral Therapy

Vitamin D regulates bone metabolism but has also immunoregulatory properties. In HIV-infected patients bone disorders are increasingly observed. Furthermore, low 1,25(OH)(2)D(3) levels have been associated with low CD4(+) counts, immunological hyperactivity, and AIDS progression rates. Few studies have examined the vitamin D status in HIV-infected patients. This study will specifically focus on the effects of antiretroviral agents on vitamin D status. Furthermore, the effect of vitamin D status on CD4 cell recovery after initiation of HAART will be evaluated. Among 252 included patients the prevalence of vitamin D deficiency (<35 nmol/liter from April to September and <25 nmol/liter from October to March) was 29%. Female sex, younger age, dark skin, and NNRTI treatment were significant risk factors in univariate analysis, although in multivariate analyses skin pigmentation remained the only independent risk factor. Median 25(OH)D(3) levels were significantly lower in white NNRTI-treated patients [54.5(27.9-73.8) nmol/liter] compared to white PI-treated patients [77.3 (46.6-100.0) nmol/liter, p = 0.007], while among nonwhites no difference was observed. Both PI- and NNRTI-treated patients had significantly higher blood PTH levels than patients without treatment. Moreover, NNRTI treatment puts patients at risk of elevated PTH levels (>6.5 pmol/liter). Linear regression analysis showed that vitamin D status did not affect CD4 cell recovery after initiation of HAART. In conclusion, 29% of the HIV-1-infected patients had vitamin D deficiency, with skin color as an independent risk factor. NNRTI treatment may add more risk for vitamin D deficiency. Both PI- and NNRTI-treated patients showed higher PTH levels and might therefore be at risk of bone problems. Evaluation of 25(OH)D(3) and PTH levels, especially in NNRTI-treated and dark skinned HIV-1-infected patients, is necessary to detect and treat vitamin D deficiency early.

The Level of APOBEC3G (hA3G)-Related G-to-A Mutations Does Not Correlate with Viral Load in HIV Type 1-Infected Individuals

The APOBEC family of mammalian cytidine deaminases, such as APOBEC3G (hA3G), has been demonstrated to function as a host viral restriction factor against HIV-1. hA3G has been shown to cause extensive G-to-A mutations in the HIV-1 genome, which may play a role in viral restriction. To investigate the role of G-to-A mutations in HIV-1 pathogenesis, we isolated, amplified, and sequenced HIV-1 sequences (vif, gag, and env) from 29 therapy-naive HIV-1-infected individuals. The levels of G-to-A mutations correlated with the expression levels of hA3G in the vif (rho = 0.438, p = 0.041) and the env regions (rho = 0.392, p = 0.038), but not in the gag region (rho = 0.131, p = 0.582). There is no correlation between viral load and the level of G-to-A mutations in the vif (rho = 0.144, p = 0.522), env (rho = 0.168, p = 0.391), or gag regions (rho = -0.254, p = 0.279). Taken together, these findings suggest that the hA3G-induced G-to-A mutations may not be the mechanism by which hA3G restricts or controls viral replication. Thus, hA3G might be restricting viral growth in infected individuals through a mechanism that is independent of the cytidine deaminase activities of hA3G.

Human Immunodeficiency Virus (HIV) gag Antigen-Specific T-Helper and Granule-Dependent CD8 T-Cell Activities in Exposed but Uninfected Heterosexual Partners of HIV Type 1-Infected Individuals in North India

Human Immunodeficiency Virus (HIV) <i>gag</i> Antigen-Specific T-Helper and Granule-Dependent CD8 T-Cell Activities in Exposed but Uninfected Heterosexual Partners of HIV Type 1-Infected Individuals in North India

Drug-associated changes in amino acid residues in Gag p2, p7NC, and p6Gag/p6Pol in human immunodeficiency virus type 1 (HIV-1) display a dominant effect on replicative fitness and drug response

Regions of HIV-1 gag between p2 and p6Gag/p6Pol, in addition to protease (PR), develop genetic diversity in HIV-1 infected individuals who fail to suppress virus replication by combination protease inhibitor (PI) therapy. To elucidate functional consequences for viral replication and PI susceptibility by changes in Gag that evolve in vivo during PI therapy, a panel of recombinant viruses was constructed. Residues in Gag p2/p7NC cleavage site and p7NC, combined with residues in the flap of PR, defined novel fitness determinants that restored replicative capacity to the posttherapy virus. Multiple determinants in Gag have a dominant effect on PR phenotype and increase susceptibility to inhibitors of drug-resistant or drug-sensitive PR genes. Gag determinants of drug sensitivity and replication alter the fitness landscape of the virus, and viral replicative capacity can be independent of drug sensitivity. The functional linkage between Gag and PR provides targets for novel therapeutics to inhibit drug-resistant viruses.

A splenic marginal zone-like peripheral blood CD27+B220- B cell population is preferentially depleted in HIV type 1-infected individuals.

Peripheral blood CD27(+) B cells are reduced in HIV-1-infected individuals. In healthy individuals, the human peripheral blood CD27(+) B cell pool consists of two subsets defined by the expression, or lack thereof, of the CD45 isoform B220. We investigated the presence of circulating B220(+) and B220(-) memory B cells in HIV(+) individuals and found that the reduction in CD27(+) memory B cells occurs primarily among CD27(+)B220(-) B cells. Studies conducted using healthy controls indicate that CD27(+)B220(-) B cells have a splenic marginal zone like the immunophenotype IgM(hi)IgD(lo)CD21(+)CD23(-), express TLR9, and proliferate and secrete IgG and IgM in response to B cell-specific ODN. CD27(+)B220(+) B cells have the immunophenotype IgM(lo)IgD(hi)CD21(+)CD23(+), express activation-induced cytidine deaminase, and proliferate in response to SAC but do not secrete immunoglobulin. The AICD expression, along with CD86 expression, by CD27(+)B220(+) suggests these cells are of germinal center origin. The preferential depletion of CD27(+)B220(-) B cells mirrors alterations in spleen morphology and resident B cell populations due to HIV infection reported by other investigators and may play an important role in the defective B cell immunity against T-independent pathogens such as pneumococcus observed in HIV-1-infected individuals.

Functional diversity of HIV-1 envelope proteins expressed by contemporaneous plasma viruses

BackgroundNumerous studies have shown that viral quasi-species with genetically diverse envelope proteins (Env) replicate simultaneously in patients infected with the human immunodeficiency virus type 1 (HIV-1). Less information is available concerning the extent that envelope sequence diversity translates into a diversity of phenotypic properties, including infectivity and resistance to entry inhibitors.MethodsTo study these questions, we isolated genetically distinct contemporaneous clonal viral populations from the plasma of 5 HIV-1 infected individuals (n = 70), and evaluated the infectivity of recombinant viruses expressing Env proteins from the clonal viruses in several target cells. The sensitivity to entry inhibitors (enfuvirtide, TAK-799), soluble CD4 and monoclonal antibodies (2G12, 48d, 2F5) was also evaluated for a subset of the recombinant viruses (n = 20).ResultsEven when comparisons were restricted to viruses with similar tropism, the infectivity for a given target cell of viruses carrying different Env proteins from the same patient varied over an approximately 10-fold range, and differences in their relative ability to infect different target cells were also observed. Variable region haplotypes associated with high and low infectivity could be identified for one patient. In addition, clones carrying unique mutations in V3 often displayed low infectivity. No correlation was observed between viral infectivity and sensitivity to inhibition by any of the six entry inhibitors evaluated, indicating that these properties can be dissociated. Significant inter-patient differences, independent of infectivity, were observed for the sensitivity of Env proteins to several entry inhibitors and their ability to infect different target cells.ConclusionThese findings demonstrate the marked functional heterogeneity of HIV-1 Env proteins expressed by contemporaneous circulating viruses, and underscore the advantage of clonal analyses in characterizing the spectrum of functional properties of the genetically diverse viral populations present in a given patient.

Efficient In Vitro Expansion of Human Immunodeficiency Virus (HIV)-Specific T-Cell Responses bygagmRNA-Electroporated Dendritic Cells from Treated and Untreated HIV Type 1-Infected Individuals

Developing an immunotherapy to keep human immunodeficiency virus type 1 (HIV-1) replication suppressed while discontinuing highly active antiretroviral therapy (HAART) is an important challenge. In the present work, we evaluated in vitro whether dendritic cells (DC) electroporated with gag mRNA can induce HIV-specific responses in T cells from chronically infected subjects. Monocyte-derived DC, from therapy-naïve and HAART-treated HIV-1-seropositive subjects, that were electroporated with consensus codon-optimized HxB2 gag mRNA efficiently expanded T cells, secreting gamma interferon (IFN-gamma) and interleukin 2 (IL-2), as well as other cytokines and perforin, upon restimulation with a pool of overlapping Gag peptides. The functional expansion levels after 1 week of stimulation were comparable in T cells from HAART-treated and treatment-naïve patients and involved both CD4(+) and CD8(+) T cells, with evidence of bifunctionality in T cells. Epitope mapping of p24 showed that stimulated T cells had a broadened response toward previously nondescribed epitopes. DC, from HAART-treated subjects, that were electroporated with autologous proviral gag mRNA equally efficiently expanded HIV-specific T cells. Regulatory T cells did not prevent the induction of effector T cells in this system, whereas the blocking of PD-L1 slightly increased the induction of T-cell responses. This paper shows that DC, loaded with consensus or autologous gag mRNA, expand HIV-specific T-cell responses in vitro.

Recent diversity of human immunodeficiency virus type 1 in individuals who visited sexually transmitted infection-related clinics in Osaka, Japan

By human immunodeficiency virus type 1 (HIV-1) antibody screening of people who visited sexually transmitted infection (STI)-related clinics (venereology, urology, and gynecology) and were considered to conduct high-risk sexual activities for HIV-1 infection in Osaka, Japan, during 1992 to 2004, a total of 54 HIV-1 infected individuals (51 Japanese males and 3 non-Japanese females) were identified. Based on the sequencing at env-C2V3 and pol regions, Japanese males were mostly of subtype B (50/51 cases), with the one remaining case being a recombinant circulating form, CRF01_AE, while 3/3 viruses in non-Japanese females were of CRF01_AE. Analysis of subtype B cases since 2001 showed that these viruses became wider in their genetic variation, including amino acid insertions and also deletions, than that of the cases before 2000. Thus, it was suggested that HIV-1 spreading in Osaka has been increasing in genetic variability. Although all these infected individuals were first recognized to be infected with HIV-1 by our screening, some of them were carriers of HIV-1 with drug-resistant pol sequences, indicating that they could be infected with drug-resistant HIV-1 mutants.

Successful Efavirenz Dose Reduction in HIV Type 1-Infected Individuals with Cytochrome P450 2B6 *6 and *26

Efavirenz (EFV) is metabolized primarily by cytochrome P450 2B6 (CYP2B6), and high plasma concentrations of the drug are associated with a G-->T polymorphism at position 516 (516G-->T) of CYP2B6 and frequent central nervous system (CNS)-related side effects. Here, we tested the feasibility of genotype-based dose reduction of EFV. CYP2B6 genotypes were determined in 456 human immunodeficiency virus type 1 (HIV-1)-infected patients who were receiving EFV treatment or were scheduled to receive EFV-containing treatment. EFV dose was reduced in CYP2B6 516G-->T carriers who had high plasma EFV concentrations while receiving the standard dosage (600 mg). EFV-naive homozygous CYP2B6 516G-->T carriers were treated with low-dose EFV. In both groups, the dose was further reduced when plasma EFV concentration remained high. CYP2B6 516G-->T was identified in the *6 allele (found in 17.9% of our subjects) and a novel allele, *26 (found in 1.3% of our patients). All EFV-treated CYP2B6 *6/*6 and *6/*26 carriers had extremely high plasma EFV concentrations (>6000 ng/mL) while receiving the standard dosage. EFV dose was reduced to 400 mg for 11 patients and to 200 mg for 7 patients with persistently suppressed HIV-1 loads. EFV-containing treatment was initiated at 400 mg in 4 CYP2B6 *6/*6 carriers and one *6/*26 carrier. Two of them still had a high plasma EFV concentration while receiving that dose, and the dose was further reduced to 200 mg, with successful HIV-1 suppression. CNS-related symptoms improved with dose reduction in 10 of the 14 patients, although some had not been aware of the symptoms at initial dosage. Genotype-based EFV dose reduction is feasible in CYP2B6 *6/*6 and *6/*26 carriers, which can reduce EFV-associated CNS symptoms.

Human Immunodeficiency Virus (HIV) gag Antigen-Specific T-Helper and Granule-Dependent CD8 T-Cell Activities in Exposed but Uninfected Heterosexual Partners of HIV Type 1-Infected Individuals in North India

Repeated exposure to human immunodeficiency virus (HIV) does not always result in HIV infection, and several cohorts of HIV-exposed but uninfected (EU) individuals have been described. We studied T-helper and granule-dependent cytotoxic T-lymphocyte (CTL) activities in a group of 30 EU partners of HIV type 1 (HIV-1)-infected individuals. HIV-1-specific helper-T-cell activity was studied by measuring the levels of interleukin 2 (IL-2) produced by peripheral blood mononuclear cells (PBMCs) and the granule-dependent CTL activity by measuring the intracellular levels of perforin and granzyme B expression in CD8+ T cells after stimulation with gag p24 antigen. Elevated IL-2 production by PBMCs after p24 stimulation occurred in EU individuals. The levels of perforin and granzyme B expression in CD8+ T cells were also higher among EU individuals than among healthy controls. HIV-specific helper-T-cell and granule-dependent CTL activities inversely correlated with the time since the last unprotected sexual exposure in these individuals. In our cohort, activation of T-helper and granule-dependent CTL activities against HIV might be due to unprotected sexual contact. These results indicate that HIV-1-specific T-cell responses could play a role in protection against acquiring infection in this cohort of EU individuals.