HIV controllers suppress viral replication and evolution and prevent disease progression following intersubtype HIV-1 superinfection.

OBJECTIVE: The aim of this study was to investigate the impact of intersubtype HIV-1 superinfection on viremia, reservoir reseeding, viral evolution and disease progression in HIV controllers (HIC). DESIGN: A longitudinal analysis of two Brazilian HIC individuals (EEC09 and VC32) previously identified as dually infected with subtypes B and F1 viruses. METHODS: Changes in plasma viremia, total HIV-1 DNA levels, CD4+ T-cell counts and HIV-1 quasispecies composition were measured over time. HIV-1 env diversity in peripheral blood mononuclear cell (PBMC) and plasma samples was accessed by single genome amplification and next-generation sequencing approaches, respectively. Viral evolution was evaluated by estimating nucleotide diversity and divergence. RESULTS: Individual EEC09 was probably initially infected with a CCR5-tropic subtype B strain and sequentially superinfected with a CXCR4-tropic subtype B strain and with a subtype F1 variant. Individual VC32 was infected with a subtype B strain and superinfected with a subtype F1 variant. The intersubtype superinfection events lead to a moderate increase in viremia and extensive turnover of viral population in plasma but exhibited divergent impact on the size and composition of cell-associated HIV DNA population. Both individuals maintained virologic control (<2000 copies/ml) and presented no evidence of viral evolution or immunologic progression for at least 2 years after the intersubtype superinfection event. CONCLUSION: These data revealed that some HIC are able to repeatedly limit replication and evolution of superinfecting viral strains of a different subtype with no signs of disease progression.

Induction of HIF-1α by HIV-1 Infection in CD4+ T Cells Promotes Viral Replication and Drives Extracellular Vesicle-Mediated Inflammation.

Chronic immune activation and inflammation are hallmarks of HIV-1 infection and a major cause of serious non-AIDS events in HIV-1-infected individuals on antiretroviral treatment (ART). Herein, we show that cytosolic double-stranded DNA (dsDNA) generated in infected CD4+ T cells during the HIV-1 replication cycle promotes the mitochondrial reactive oxygen species (ROS)-dependent stabilization of the transcription factor hypoxia-inducible factor 1α (HIF-1α), which in turn, enhances viral replication. Furthermore, we show that induction of HIF-1α promotes the release of extracellular vesicles (EVs). These EVs foster inflammation by inducing the secretion of gamma interferon by bystander CD4+ T cells and secretion of interleukin 6 (IL-6) and IL-1ß by bystander macrophages through an HIF-1α-dependent pathway. Remarkably, EVs obtained from plasma samples from HIV-1-infected individuals also induced HIF-1α activity and inflammation. Overall, this study demonstrates that HIF-1α plays a crucial role in HIV-1 pathogenesis by promoting viral replication and the release of EVs that orchestrate lymphocyte- and macrophage-mediated inflammatory responses.IMPORTANCE Human immunodeficiency virus type 1 (HIV-1) is a very important global pathogen that preferentially targets CD4+ T cells and causes acquired immunodeficiency syndrome (AIDS) if left untreated. Although antiretroviral treatment efficiently suppresses viremia, markers of immune activation and inflammation remain higher in HIV-1-infected patients than in uninfected individuals. The hypoxia-inducible factor 1α (HIF-1α) is a transcription factor that plays a fundamental role in coordinating cellular metabolism and function. Here we show that HIV-1 infection induces HIF-1α activity and that this transcription factor upholds HIV-1 replication. Moreover, we demonstrate that HIF-1α plays a key role in HIV-1-associated inflammation by promoting the release of extracellular vesicles which, in turn, trigger the secretion of inflammatory mediators by noninfected bystander lymphocytes and macrophages. In summary, we identify that the coordinated actions of HIF-1α and extracellular vesicles promote viral replication and inflammation, thus contributing to HIV-1 pathogenesis.

Circulating CXCR5-Expressing CD8+ T-Cells Are Major Producers of IL-21 and Associate With Limited HIV Replication.

BACKGROUND: Despite advances made with the highly active antiretroviral therapy (HAART) in the control of the HIV 1 infection, a cure has not been achieved because of the persistence of viral reservoirs. The major HIV reservoirs remain in the lymphoid follicles because of, among other factors, the partial absence of CD8 T-cells in these structures. Recently, lymphoid follicle-confined and circulating CD8 T-cells expressing the C-X-C chemokine receptor type 5 (CXCR5) were described, possessing antiviral mechanisms that could help to control HIV replication. SETTING AND METHODS: By flow cytometry, we characterized the phenotype and function of circulating CXCR5-expressing CD8 T-cells in HIV-infected patients with natural or HAART-induced control of HIV replication. RESULTS: Circulating CXCR5-expressing CD8 T-cells exhibited low or null expression of the C-C chemokine receptor type 7 (CCR7) and had a transitional memory phenotype. Particular redistributions of CXCR5-expressing CD8 T-cells were found in HIV-infected patients, and they were partially restored by HAART. The frequency of CXCR5CCR7 CD8 T-cells was higher in spontaneous HIV controllers and negatively correlated with plasma HIV RNA levels. Total and HIV-specific CXCR5 CD8 T-cells were major producers of interleukin-21, and this function was positively associated with their interferon-γ production. CONCLUSIONS: Circulating CXCR5-expressing CD8 T-cells are associated with low-level HIV replication; these cells could be novel correlates of protection, and potentially useful in the eradication of HIV reservoirs.

Enteric viruses in HIV-1 seropositive and HIV-1 seronegative children with diarrheal diseases in Brazil.

Diarrheal diseases (DD) have distinct etiological profiles in immune-deficient and immune-competent patients. This study compares detection rates, genotype distribution and viral loads of different enteric viral agents in HIV-1 seropositive (n = 200) and HIV-1 seronegative (n = 125) children hospitalized with DD in Rio de Janeiro, Brazil. Except for group A rotavirus (RVA), which were detected through enzyme immunoassay, the other enteric viruses (norovirus [NoV], astrovirus [HAstV], adenovirus [HAdV] and bocavirus [HBoV]) were detected through PCR or RT-PCR. A quantitative PCR was performed for RVA, NoV, HAstV, HAdV and HBoV. Infections with NoV (19% vs. 9.6%; p<0.001), HBoV (14% vs. 7.2%; p = 0.042) and HAdV (30.5% vs. 14.4%; p<0.001) were significantly more frequent among HIV-1 seropositive children. RVA was significantly less frequent among HIV-1 seropositive patients (6.5% vs. 20%; p<0.001). Similarly, frequency of infection with HAstV was lower among HIV-1 seropositive children (5.5% vs. 12.8%; p = 0.018). Among HIV-1 seropositive children 33 (16.5%) had co-infections, including three enteric viruses, such as NoV, HBoV and HAdV (n = 2) and NoV, HAstV and HAdV (n = 2). The frequency of infection with more than one virus was 17 (13.6%) in the HIV-1 negative group, triple infection (NoV + HAstV + HBoV) being observed in only one patient. The median viral load of HAstV in feces was significantly higher among HIV-1 positive children compared to HIV-1 negative children. Concerning children infected with RVA, NoV, HBoV and HAdV, no statistically significant differences were observed in the medians of viral loads in feces, comparing HIV-1 seropositive and HIV-1 seronegative children. Similar detection rates were observed for RVA, HAstV and HAdV, whilst NoV and HBoV were significantly more prevalent among children with CD4+ T lymphocyte count below 200 cells/mm3. Enteric viruses should be considered an important cause of DD in HIV-1 seropositive children, along with pathogens more classically associated with intestinal infections in immunocompromised hosts.

Precursor Forms of Vitamin D Reduce HIV-1 Infection In Vitro.

BACKGROUND: Although the anti-HIV-1 effects of vitamin D (VitD) have been reported, mechanisms behind such protection remain largely unexplored. METHODS: The effects of two precursor forms (cholecalciferol/calciol at 0.01, 1 and 100 nM and calcidiol at 100 and 250 nM) on HIV-1 infection, immune activation, and gene expression were analyzed in vitro in cells of Colombian and Italian healthy donors. We quantified levels of released p24 by enzyme-linked immunosorbent assay, of intracellular p24 and cell-surface expression of CD38 and HLA-DR by flow cytometry, and mRNA expression of antiviral and immunoregulatory genes by real-time reverse transcription-polymerase chain reaction. RESULTS: Cholecalciferol decreased the frequency of HIV-1-infected p24CD4 T cells and levels of p24 in supernatants in a dose-dependent manner. Moreover, the CD4CD38HLA-DR and CD4CD38HLA-DR subpopulations were more susceptible to infection but displayed the greatest cholecalciferol-induced decreases in infection rate by an X4-tropic strain. Likewise, cholecalciferol at its highest concentration decreased the frequency of CD38HLA-DR but not of CD38HLA-DR T-cell subsets. Analyzing the effects of calcidiol, the main VitD source for immune cells and an R5-tropic strain as the most frequently transmitted virus, a reduction in HIV-1 productive infection was also observed. In addition, an increase in mRNA expression of APOBEC3G and PI3 and a reduction of TRIM22 and CCR5 expression, this latter positively correlated with p24 levels, was noted. CONCLUSIONS: VitD reduces HIV-1 infection in T cells possibly by inducing antiviral gene expression, reducing the viral co-receptor CCR5 and, at least at the highest cholecalciferol concentration, by promoting an HIV-1-restrictive CD38HLA-DR immunophenotype.

Evolution of hepatitis C virus in HIV coinfected patients under antiretroviral therapy.

Five patients (P) were followed-up for an average of 7.73years after highly active antiretroviral therapy (HAART) initiation. Patients' immune and virological status were determined by periodical CD4+T-cell counts and HIV and HCV viral load. HCV populations were studied using longitudinal high throughput sequence data obtained in parallel by virological and immunological parameters. Two patients (P7, P28) with sub-optimal responses to HAART presented HCV viral loads significantly higher than those recorded for two patients (P1, P18) that achieved good responses to HAART. Interestingly, HCV populations from P7 and P28 displayed a stable phylogenetic structure, whereas HCV populations from P1 and P18showeda significant increase in their phylogenetic structure, followed by a decrease after achieving acceptable CD4+T-cell counts (>500 cell/µl). The fifth patient (P25) presented high HCV viral loads, preserved CD4+T-cell counts from baseline and all along the follow-up, and displayed a constant viral phylogenetic structure. These results strongly suggest that HAART-induced immune recovery induces a decrease in HCV viral load and an increase in the HCV population phylogenetic structure likely reflecting the virus diversification in response to the afresh immune response. The relatively low HCV viral load observed in the HAART responder patients suggests that once HCV is adapted it reaches a maximum number of haplotypes higher than that achieved during the initial stages of the immune response as inferred from the two recovering patients. Future studies using larger number of patients are needed to corroborate these hypotheses.

HIV-1 Pol Gene Polymorphism and Antiretroviral Resistance Mutations in Treatment-Naive Adult Patients in French Guiana Between 2006 and 2012.

Little information is available on the molecular epidemiologic profile of HIV-1 in French Guiana, the French department with the highest HIV/AIDS incidence. To follow the evolution of HIV-1 diversity, we carried out a molecular analysis of HIV-1 isolates from 305 treatment-naive patients between 2006 and 2012. Protease and reverse-transcriptase sequences were obtained for subtype characterization, polymorphism analysis, and identification of drug resistance mutations. Of 305 HIV-1 strains, 95.1% were subtype B viruses. The overall prevalence of transmitted drug-resistance mutations (TDRMs) was 4.6% (14/305), ranging from 1.9% to 7.1% depending on the year. This study shows a low level of HIV-1 genetic diversity and a moderate prevalence of TDRMs with no evidence of an increasing trend over the study period. Nevertheless, the strong genetic polymorphism observed on both genes may be of concern for long-term treatment of people living with HIV-1 and thus deserves continuous monitoring.

Short Communication: In Vitro Accumulation of Drug Resistance Mutations in Chimeric Infectious Clones Containing Subtype B or C Reverse Transcriptase and Selected with Tenofovir or Didanosine.

Highly active antiretroviral therapy (HAART) contributed to the improvement in the life expectancy of HIV-infected patients. However, the emergence of drug-resistant mutations (DRM) is a major viral factor impacting therapeutic failure. Differences in DRM can occur among HIV-1 subtypes. We evaluate the kinetics of the selection of resistance mutations in vitro analyzing two chimeric clones that contain the reverse transcriptases of subtypes B or C (RTB' and RTC') in cells treated with increasing concentrations of tenofovir disoproxil fumarate (TDF) and didanosine (ddI). The mutation K65R is selected more quickly in RTC' than in RTB' viruses with TDF and ddI, and additional mutations (positions 45, 62, and 68) were selected after K65R fixation. Other primary mutations (M184V and Q151M) were selected with ddI treatment in conjunction with K65R only in RTC' viruses. Both patterns, M184V+K65R and Q151M+K65R, have a significant impact on NRTI resistance. Our data suggest that selection of TDF and ddI DRMs can occur earlier in subtype C HIV in patients when compared to subtype B.

Role of metabolic syndrome and antiretroviral therapy in adiponectin levels and oxidative stress in HIV-1 infected patients.

OBJECTIVE: HIV-1 infection is accompanied by severe metabolic and immune dysfunction. The aim of this study was to evaluate the role of metabolic syndrome (MetS) and antiretroviral therapy (ART) utilization on the adiponectin levels and oxidative stress in patients infected with HIV-1. METHODS: We allocated 285 patients into four groups: group 1: patients without MetS who were not using ART; group 2: patients without MetS using ART; group 3: patients with MetS who were not using ART; and group 4: patients with MetS using ART. Biochemical, immunologic, and oxidative stress parameters were measured. RESULTS: Group 4 exhibited higher lipoperoxides when compared with group 1 (P < 0.0001) and higher advanced oxidation protein products (AOPP) compared with group 2 or group 1 (P < 0.0001). Group 3 also presented higher AOPP than group 2 (P < 0.05). Group 4 showed lower adiponectin levels compared with groups 1 or 2 (P < 0.0001). Similarly, group 3 presented lower adiponectin levels compared with group 2 (P < 0.05) or group 1 (P < 0.0001). Multivariate analysis showed that both an increase in AOPP and a decrease in total radical-trapping antioxidant parameter/uric acid were independently associated with MetS in HIV-1 patients. Regarding immunologic markers of HIV-1 disease progression and viral replication, group 4 exhibited significantly higher CD45(+), CD3(+), and CD4(+) T cells count compared with group 2 (P < 0.01). CONCLUSION: HIV-1-infected patients with MetS exhibited hypoadiponectinemia and increased oxidative stress, and these findings were not influenced by ART use. The findings of the present study allow the suggestion that MetS and inflammation might be mainly responsible for the aforementioned features. More studies are needed to verify whether drugs or food, which yield increased adiponectinemia and decreased oxidative stress, could reduce cardiovascular risk in HIV-infected patients.

High rates of human immunodeficiency virus type 1 mutational profiles by single-genome amplification after 48-hour propagation in peripheral blood mononuclear cells at different levels of cell activation.

Human immunodeficiency virus type 1 (HIV-1) genetic diversity is one of the most important features of HIV-1 infections and the result of error accumulation during reverse transcription and of high viral turnover. HIV-1 reverse transcription is influenced by factors such as the level of nucleotides and/or the cellular activation state. HIV-1 diversity was investigated after 48 h of viral propagation in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors in three different cell culture conditions: (1) resting PBMCs, (2) simultaneous infection and PBMC activation, and (3) PBMC activation 72 h before infection. Cellular DNA was extracted and proviruses of each culture condition were amplified. Single-genome PCR clones were obtained and the protease and reverse transcriptase of the pol gene were sequenced. An elevated number of nucleotide substitutions in all three culture conditions were observed. In condition 1, the mutational rate observed ranged from 1.0 × 10(-3) to 2.1 × 10(-2), the genetic diversity was 0.6%, and hypermutation was observed in 7.1% of sequenced clones. In condition 2, the mutational rate ranged from 1.0 × 10(-3) to 1.0 × 10(-2), the genetic diversity was 0.8%, and hypermutation affected 6.7% of clones. In condition 3, the mutational rate ranged from 2.8 × 10(-3) to 1.1 × 10(-2), the genetic diversity was 1%, and 5.9% of clones were hypermutated. Substitutions occurred more frequently in some specific nucleotide stretches, and a common pattern for substitutions in all the different conditions was identified. There was a significant accumulation of mutations during the initial periods of in vitro HIV-1 propagation irrespective of culture conditions. The rapid accumulation of virus diversity might represent a viral strategy when colonizing new hosts. Complementary studies are necessary to allow for a better understanding of the initial periods of infection, which represent a crucial event related to disease progression.

Autologous and allogenic systems of HIV expansion: what is the better choice for clinical application in therapeutic vaccine?

AIMS: HIV-1 expanded in an allogenic system (Al-HIV) represents a cheaper and faster alternative to the autologous virus (Au-HIV) as an antigen in anti-HIV immunotherapy. In this study, chemically inactivated HIV-1 obtained through autologous or allogenic systems were compared. PATIENTS & METHODS: Au-HIV and Al-HIV obtained from cultures of peripheral blood mononuclear cells from 11 HIV(+) individuals were tested for virus production, yield and time of culture, and their ability to elicit a specific immune response in vitro. RESULTS: The allogenic system was more efficient than the autologous system. Dendritic cells pulsed with Au-HIV and Al-HIV presented a similar phenotypic profile, but only Al-HIV induced a significant increase in IFN-γ(+) lymphocytes. CONCLUSION: The use of an allogenic system displays several advantages in terms of cell manipulation, time and cost of culture, and immunogenicity.

Macrophage Resistance to HIV-1 Infection Is Enhanced by the Neuropeptides VIP and PACAP.

It is well established that host factors can modulate HIV-1 replication in macrophages, critical cells in the pathogenesis of HIV-1 infection due to their ability to continuously produce virus. The neuropeptides VIP and PACAP induce well-characterized effects on macrophages through binding to the G protein-coupled receptors VPAC1, VPAC2 and PAC1, but their influence on HIV-1 production by these cells has not been established. Here, we describe that VIP and PACAP reduce macrophage production of HIV-1, acting in a synergistic or additive manner to decrease viral growth. Using receptor antagonists, we detected that the HIV-1 inhibition promoted by VIP is dependent on its ligation to VPAC1/2, whereas PACAP decreases HIV-1 growth via activation of the VPAC1/2 and PAC1 receptors. Specific agonists of VPAC2 or PAC1 decrease macrophage production of HIV-1, whereas sole activation of VPAC1 enhances viral growth. However, the combination of specific agonists mimicking the receptor preference of the natural neuropeptides reproduces the ability of VIP and PACAP to increase macrophage resistance to HIV-1 replication. VIP and PACAP up-regulated macrophage secretion of the ß-chemokines CCL3 and CCL5 and the cytokine IL-10, whose neutralization reversed the neuropeptide-induced inhibition of HIV-1 replication. Our results suggest that VIP and PACAP and the receptors VPAC2 and PAC1 could be used as targets for developing alternative therapeutic strategies for HIV-1 infection.

Chronic kidney disease associated with perinatal HIV infection in children and adolescents.

BACKGROUND: This study describes the incidence, clinical and demographic characteristics, and spectrum of chronic kidney disease (CKD) in youths with perinatal HIV-1 infection. METHODS: Retrospective analysis between May 1993 and December 2006 of subjects with renal disease followed in the Pediatric AIDS Clinical Trials Group 219/219C multicenter study examining the long-term consequences of perinatal HIV infection. Diagnosis confirmation was made utilizing a questionnaire mailed to research sites. Participants with CKD of other etiology than HIV were excluded. Outcome measures were biopsy-diagnosed CKD and, in the absence of biopsy, HIV-associated nephropathy (HIVAN) using established clinical criteria. RESULTS: Questionnaires on 191 out of 2,102 participants identified 27 cases of CKD: 14 biopsy-diagnosed and 6 clinical cases of HIVAN, and 7 biopsy-diagnosed cases of immune complex-mediated kidney disease (lupus-like nephritis, 3; IgA nephropathy, 2; membranous nephropathy, 2). Incidence rates for CKD associated with HIV in pre-highly active antiretroviral therapy (HAART) (1993-1997) and HAART (1998-2002, 2003-2006) eras were 0.43, 2.84, and 2.79 events per 1,000 person years respectively. In multivariate analysis, black race and viral load ≥100,000 copies/mL (rate ratios 3.28 and 5.05, p ≤ 0.02) were associated with CKD. CONCLUSIONS: A variety of immune complex-mediated glomerulonephritides and HIVAN occurs in this population. Black race and uncontrolled viral replication are risk factors for CKD associated with HIV.

Evidences for viral strain selection in late stages of HIV infection: an analysis of Vpu alleles.

One of the most studied topics about AIDS disease is the presence of different progression levels in patients infected by HIV. Several studies have shown that this progression is directly associated with host genetics, although viral factors are also known to play a role. Here we explore the contribution of Vpu protein in the evolution of viral population. The sequence variation of Vpu was analyzed during HIV infection in peripheral blood monocyte cells of 12 patients in different clinical stages of HIV-1 infection early and late stages of infections, separated by at least 4 years. The clustering analysis of Vpu sequences showed higher diversity of early alleles, non-random distribution of sequences, and viral evolution strains selection. Forty-two amino acid modifications were found in the multiple alignments of the 57 different alleles found for early stage were 23 modifications were found in the late stage dataset. Interestingly fourteen alteration of early stage were located in conserved site related with Vpu functions alterations while these alterations appear with less frequency in the late stage of infection. Moreover, late stage alleles tend to be similar with the Vpu wild type sequence, suggesting viral selection toward populations harboring more efficient variants during the course of infection. This would contribute to higher infectivity and viral replication actually observed at the aggressive late stages of infection. These data, in conjunction with in vitro experiments, will be important to elucidation of the physiological relevance of Vpu protein in the pathogenic mechanisms of AIDS.

Viral replication is enhanced by an HIV-1 intersubtype recombination-derived Vpu protein.

BACKGROUND: Multiple HIV-1 intersubtype recombinants have been identified in human populations. Previous studies from our lab group have shown that the epidemic in Argentina is characterized by the high prevalence of a circulating recombinant form, CRF12_BF, and many related BF recombinant forms. In these genomic structures a recombination breakpoint frequently involved the vpu coding region. Due to the scarce knowledge of Vpu participation in the virion release process and its impact on pathogenesis and of the functional capacities of intersubtype recombinant Vpu proteins, the aim of this work was to perform a comparative analysis on virion release capacity and relative replication capacity among viral variants harboring either a BF recombinant Vpu or a subtype B Vpu. RESULTS: Our results showed that BF recombinant Vpu was associated to an increased viral particles production when compared to WT B variant in tetherin-expressing cell lines. This observation was tested in the context of a competition assay between the above mentioned variants. The results showed that the replication of the BF Vpu-harboring variant was more efficient in cell cultures than subtype B, reaching a higher frequency in the viral population in a short period of time. CONCLUSION: This study showed that as a result of intersubtype recombination, a structurally re-organized HIV-1 Vpu has an improved in vitro capacity of enhancing viral replication, and provides evidence of the changes occurring in this protein function that could play an important role in the successful spread of intersubtype recombinant variants.

Mutation T74S in HIV-1 subtype B and C proteases resensitizes them to ritonavir and indinavir and confers fitness advantage.

OBJECTIVES: Several drug resistance and secondary mutations have been described in HIV-1 viruses from patients undergoing antiretroviral therapy. In this study, we assessed the impact of the protease substitution T74S on the phenotype and on the replicative fitness in HIV-1 subtypes B and C. METHODS: HIV-1 molecular clones carrying subtype B or C proteases had these coding regions subjected to site-directed mutagenesis to include T74S alone or in combination with four known protease inhibitor (PI) primary drug resistance mutations. All clones were used in a phenotypic assay to evaluate their susceptibility to most commercially available PIs. The impact of T74S on virus fitness was also assessed for all viruses through head-to-head competitions and oligonucleotide ligation assays to measure the proportion of each virus in culture. RESULTS: Viruses of both subtypes carrying T74S did not have their susceptibility altered to any tested PI. Viruses with the four resistance mutations showed strong resistance to most PIs with fold changes ranging from 5 to 300 times compared with their wild-type counterparts. Surprisingly, the addition of T74S to the multiresistant clones restored their susceptibilities to indinavir and ritonavir and partially to lopinavir, close to those of wild-type viruses. Most 74S-containing viruses were more fit than their 74T counterparts. CONCLUSIONS: Our results suggest that T74S is not a major drug resistance mutation, but it resensitizes multiresistant viruses to certain PIs. T74S is a bona fide accessory mutation, restoring fitness of multidrug-resistant viruses in both subtypes B and C. T74S should be further studied in clinical settings and considered in drug resistance interpretation algorithms.

Synthesis, antiviral activity and molecular modeling of oxoquinoline derivatives.

In the present article, we describe the synthesis, anti-HIV1 profile and molecular modeling evaluation of 11 oxoquinoline derivatives. The structure-activity relationship analysis revealed some stereoelectronic properties such as LUMO energy, dipole moment, number of rotatable bonds, and of hydrogen bond donors and acceptors correlated with the potency of compounds. We also describe the importance of substituents R(2) and R(3) for their biological activity. Compound 2j was identified as a lead compound for future investigation due to its: (i) high activity against HIV-1, (ii) low cytotoxicity in PBMC, (iii) low toxic risks based on in silico evaluation, (iv) a good theoretical oral bioavailability according to Lipinski 'rule of five', (v) higher druglikeness and drug-score values than current antivirals AZT and efavirenz.

Proteomic analyses associate cystatin B with restricted HIV-1 replication in placental macrophages.

Mononuclear phagocytes (MP; monocytes, tissue macrophages, and dendritic cells) are reservoirs, vehicles of dissemination, and targets for persistent HIV infection. However, not all MP population equally support viral growth. Such differential replication is typified by the greater ability of placental macrophages (PM), as compared to blood borne monocyte-derived macrophages (MDM), to restrict viral replication. Since cytosolic protein patterns can differentiate macrophage subtypes, we used a proteomics approach consisting of surface-enhanced laser desorption ionization time-of-flight (SELDI-TOF), tandem mass spectrometry, and Western blots to identify differences between the uninfected and HIV-infected PM and MDM protein profiles linked to viral growth. We performed proteome analysis of PM in the molecular range of 5-20kDa. We found that a SELDI-TOF protein peak with an m/z of 11,100, which was significantly lower in uninfected and HIV-infected PM than in MDM, was identified as cystatin B (CSTB). Studies of siRNA against CSTB treatment in MDM associated its expression with HIV replication. These data demonstrate that the low molecular weight placental macrophage cytosolic proteins are differentially expressed in HIV-infected PM and MDM and identify a potential role for CSTB in HIV replication. This work also serves to elucidate a mechanism by which the placenta protects the fetus from HIV transmission.

Multiple-dose pharmacokinetics of apricitabine, a novel nucleoside reverse transcriptase inhibitor, in patients with HIV-1 infection.

BACKGROUND AND OBJECTIVE: This study aimed to investigate the multiple-dose pharmacokinetics of apricitabine, a novel deoxycytidine analogue reverse transcriptase inhibitor, in antiretroviral-naive patients with HIV-1 infection. METHODS: This was an international, randomized, double-blind, placebo-controlled, multicentre, dose-ranging study. Patients received 10 days' oral placebo or apricitabine 200, 400, 600 or 800 mg twice daily or 800 or 1200 mg once daily. On days 1 and 8, blood and urine samples were collected over 24 hours for pharmacokinetic analysis. Apricitabine triphosphate pharmacokinetics were investigated in peripheral blood mononuclear cells (PBMCs) on day 8. RESULTS: Overall, 63 patients (mean age 33.9 +/- 8.7 years; mean weight 71.6 +/- 15.4 kg) were randomized, and 62 patients completed the study. Apricitabine was rapidly absorbed, with peak plasma concentrations attained within approximately 1.5-2.5 hours. Pharmacokinetics were linear over the range 200-800 mg twice daily. Apricitabine was predominantly excreted via the kidneys, with no significant accumulation during repeated administration. Steady-state conditions were attained by day 8. Apricitabine triphosphate exposure in PBMCs was roughly proportional to the dose of apricitabine across the dose range 200-800 mg twice daily, with adequate correlations between plasma exposure to apricitabine (9910 ng/mL per 65 kg for 800-mg twice-daily administration) and PBMC exposure to apricitabine triphosphate (maximum concentration [C(max)] = 5.55 +/- 1.94 pmol/million cells for 800-mg twice-daily administration). Apri-citabine was well tolerated. CONCLUSION: Apricitabine shows essentially linear pharmacokinetics during repeated administration in patients with HIV-1 infection.

Characterization of peripheral blood human immunodeficiency virus isolates from Hispanic women with cognitive impairment.

Human immunodeficiency virus type 1 (HIV-1) tropism plays an important role in HIV-associated dementia. In this study, aimed at determining if the tropism and coreceptor usage of circulating viruses correlates with cognitive function, the authors isolated and characterized HIV from the peripheral blood of 21 Hispanic women using antiretroviral therapy. Macrophage tropism was determined by inoculation of HIV isolates onto monocyte-derived macrophages and lymphocyte cultures. To define coreceptor usage, the HIV isolates were inoculated onto the U87.CD4 glioma cell lines with specific CCR5 and CXCR4 coreceptors. HIV isolates from cognitively impaired patients showed higher levels of replication in mitogen-stimulated peripheral blood mononuclear cells than did isolates from patients with normal cognition (P < .05). The viral growth of HIV primary isolates in macrophages and lymphocytes did not differ between patients with and those without cognitive impairment. However, isolates from the cognitively impaired women preferentially used the X4 coreceptor (P < .05). These phenotypic studies suggest that cognitively impaired HIV-infected women receiving treatment may have a more highly replicating and more pathogenic X4 virus in the circulation that could contribute to their neuropathogenesis.