Prolonged viral suppression with anti-HIV-1 antibody therapy.

HIV-1 infection remains a public health problem with no cure. Anti-retroviral therapy (ART) is effective but requires lifelong drug administration owing to a stable reservoir of latent proviruses integrated into the genome of CD4+ T cells1. Immunotherapy with anti-HIV-1 antibodies has the potential to suppress infection and increase the rate of clearance of infected cells2,3. Here we report on a clinical study in which people living with HIV received seven doses of a combination of two broadly neutralizing antibodies over 20 weeks in the presence or absence of ART. Without pre-screening for antibody sensitivity, 76% (13 out of 17) of the volunteers maintained virologic suppression for at least 20 weeks off ART. Post hoc sensitivity analyses were not predictive of the time to viral rebound. Individuals in whom virus remained suppressed for more than 20 weeks showed rebound viraemia after one of the antibodies reached serum concentrations below 10 µg ml-1. Two of the individuals who received all seven antibody doses maintained suppression after one year. Reservoir analysis performed after six months of antibody therapy revealed changes in the size and composition of the intact proviral reservoir. By contrast, there was no measurable decrease in the defective reservoir in the same individuals. These data suggest that antibody administration affects the HIV-1 reservoir, but additional larger and longer studies will be required to define the precise effect of antibody immunotherapy on the reservoir.

Efficiency of bridging-sheet recruitment explains HIV-1 R5 envelope glycoprotein sensitivity to soluble CD4 and macrophage tropism.

HIV-1 R5 viruses vary extensively in their capacity to infect macrophages. R5 viruses that confer efficient infection of macrophages are able to exploit low levels of CD4 for infection and predominate in brain tissue, where macrophages are a major target for infection. HIV-1 R5 founder viruses that are transmitted were reported to be non-macrophage-tropic. Here, we investigated the sensitivities of macrophage-tropic and non-macrophage-tropic R5 envelopes to neutralizing antibodies. We observed striking differences in the sensitivities of Env(+) pseudovirions to soluble CD4 (sCD4) and to neutralizing monoclonal antibodies (MAbs) that target the CD4 binding site. Macrophage-tropic R5 Envs were sensitive to sCD4, while non-macrophage-tropic Envs were significantly more resistant. In contrast, all Envs were sensitive to VRC01 regardless of tropism, while MAb b12 conferred an intermediate neutralization pattern where all the macrophage-tropic and about half of the non-macrophage-tropic Envs were sensitive. CD4, b12, and VRC01 share binding specificities on the outer domain of gp120. However, these antibodies differ in their ability to induce conformational changes on the trimeric envelope and in specificity for residues on the V1V2 loop stem and ß20-21 junction that are targets for CD4 in recruiting the bridging sheet. These distinct specificities of CD4, b12, and VRC01 likely explain the observed differences in Env sensitivity to inhibition by these reagents and provide an insight into the envelope mechanisms that control macrophage tropism. We present a model where the efficiency of bridging-sheet recruitment by CD4 is a major determinant of HIV-1 R5 envelope sensitivity to soluble CD4 and macrophage tropism.

Susceptibility to 3BNC117 and 10-1074 in ART suppressed chronically infected persons.

OBJECTIVE: The aim of this study was to assess the susceptibility of HIV to two HIV monoclonal antibodies (bnAbs), 3BNC117 and 10-1074, in individuals with chronically antiretroviral therapy (ART) suppressed HIV infection. DESIGN: The susceptibility of bnAbs was determined using the PhenoSense mAb Assay, which is a cell-based infectivity assay designed to assess the susceptibility of luciferase-reporter pseudovirions. This assay is the only Clinical Laboratory Improvement Ammendment (CLIA)/College of American Pathologist (CAP) compliant screening test specifically developed for evaluating bnAb susceptibility in people with HIV infection. METHOD: The susceptibility of luciferase-reporter pseudovirions, derived from HIV-1 envelope proteins obtained from peripheral bloodmononuclear cells of 61 ART-suppressed individuals, to 3BNC117 and 10-1074 bnAbs was assessed using the PhenoSense mAb assay. Susceptibility was defined as an IC 90 of <2.0 µg/ml and 1.5 µg/ml for 3BNC117 and 10-1074, respectively. RESULTS: About half of the individuals who were chronically infected and virologically suppressed were found to harbor virus with reduced susceptibility to one or both of the tested bnAbs. CONCLUSIONS: The reduced combined susceptibility of 3BNC117 and 10-1074 highlights a potential limitation of using only two bnAbs for pre-exposure prophylaxis or treatment. Further studies are needed to define and validate the clinical correlates of bnAb susceptibility.

Broadly neutralizing antibody treatment maintained HIV suppression in children with favorable reservoir characteristics in Botswana.

Broadly neutralizing antibodies (bNAbs) may provide an alternative to standard antiretroviral treatment (ART) for controlling HIV-1 replication and may have immunotherapeutic effects against HIV-1 reservoirs. We conducted a prospective clinical trial with two HIV-1 bNAbs (VRC01LS and 10-1074) in children (n = 25) who had previously initiated small-molecule ART treatment before 7 days of age and who continued treatment for at least 96 weeks. Both bNAbs were dosed intravenously every 4 weeks, overlapping with ART for at least 8 weeks and then continued for up to 24 weeks or until detectable viremia of HIV-1 RNA rose above 400 copies per milliliter in the absence of ART. Eleven (44%) children maintained HIV-1 RNA below 400 copies per milliliter through 24 weeks of bNAb-only treatment; 14 (56%) had detectable viremia above 400 copies per milliliter at a median of 4 weeks. Archived HIV-1 provirus susceptible to 10-1074, lower birth HIV-1 DNA reservoir in peripheral blood mononuclear cells, sustained viral suppression throughout early life, and combined negative qualitative HIV-1 DNA polymerase chain reaction and negative HIV-1 serology at entry were associated with maintaining suppression on bNAbs alone. This proof-of-concept study suggests that bNAbs may represent a promising treatment modality for infants and children living with HIV-1. Future studies using newer bNAb combinations with greater breadth and potency are warranted.

First-in-human immunoPET imaging of HIV-1 infection using 89Zr-labeled VRC01 broadly neutralizing antibody.

A major obstacle to achieving long-term antiretroviral (ART) free remission or functional cure of HIV infection is the presence of persistently infected cells that establish a long-lived viral reservoir. HIV largely resides in anatomical regions that are inaccessible to routine sampling, however, and non-invasive methods to understand the longitudinal tissue-wide burden of HIV persistence are urgently needed. Positron emission tomography (PET) imaging is a promising strategy to identify and characterize the tissue-wide burden of HIV. Here, we assess the efficacy of using immunoPET imaging to characterize HIV reservoirs and identify anatomical foci of persistent viral transcriptional activity using a radiolabeled HIV Env-specific broadly neutralizing antibody, 89Zr-VRC01, in HIV-infected individuals with detectable viremia and on suppressive ART compared to uninfected controls (NCT03729752). We also assess the relationship between PET tracer uptake in tissues and timing of ART initiation and direct HIV protein expression in CD4 T cells obtained from lymph node biopsies. We observe significant increases in 89Zr-VRC01 uptake in various tissues (including lymph nodes and gut) in HIV-infected individuals with detectable viremia (N = 5) and on suppressive ART (N = 5) compared to uninfected controls (N = 5). Importantly, PET tracer uptake in inguinal lymph nodes in viremic and ART-suppressed participants significantly and positively correlates with HIV protein expression measured directly in tissue. Our strategy may allow non-invasive longitudinal characterization of residual HIV infection and lays the framework for the development of immunoPET imaging in a variety of other infectious diseases.

Early intervention with 3BNC117 and romidepsin at antiretroviral treatment initiation in people with HIV-1: a phase 1b/2a, randomized trial.

Attempts to reduce the human immunodeficiency virus type 1 (HIV-1) reservoir and induce antiretroviral therapy (ART)-free virologic control have largely been unsuccessful. In this phase 1b/2a, open-label, randomized controlled trial using a four-group factorial design, we investigated whether early intervention in newly diagnosed people with HIV-1 with a monoclonal anti-HIV-1 antibody with a CD4-binding site, 3BNC117, followed by a histone deacetylase inhibitor, romidepsin, shortly after ART initiation altered the course of HIV-1 infection ( NCT03041012 ). The trial was undertaken in five hospitals in Denmark and two hospitals in the United Kingdom. The coprimary endpoints were analysis of initial virus decay kinetics and changes in the frequency of CD4+ T cells containing intact HIV-1 provirus from baseline to day 365. Secondary endpoints included changes in the frequency of infected CD4+ T cells and virus-specific CD8+ T cell immunity from baseline to day 365, pre-ART plasma HIV-1 3BNC117 sensitivity, safety and tolerability, and time to loss of virologic control during a 12-week analytical ART interruption that started at day 400. In 55 newly diagnosed people (5 females and 50 males) with HIV-1 who received random allocation treatment, we found that early 3BNC117 treatment with or without romidepsin enhanced plasma HIV-1 RNA decay rates compared to ART only. Furthermore, 3BNC117 treatment accelerated clearance of infected cells compared to ART only. All groups had significant reductions in the frequency of CD4+ T cells containing intact HIV-1 provirus. At day 365, early 3BNC117 + romidepsin was associated with enhanced HIV-1 Gag-specific CD8+ T cell immunity compared to ART only. The observed virological and immunological effects of 3BNC117 were most pronounced in individuals whose pre-ART plasma HIV-1 envelope sequences were antibody sensitive. The results were not disaggregated by sex. Adverse events were mild to moderate and similar between the groups. During a 12-week analytical ART interruption among 20 participants, 3BNC117-treated individuals harboring sensitive viruses were significantly more likely to maintain ART-free virologic control than other participants. We conclude that 3BNC117 at ART initiation enhanced elimination of plasma viruses and infected cells, enhanced HIV-1-specific CD8+ immunity and was associated with sustained ART-free virologic control among persons with 3BNC117-sensitive virus. These findings strongly support interventions administered at the time of ART initiation as a strategy to limit long-term HIV-1 persistence.

Reanalysis of coreceptor tropism in HIV-1-infected adults using a phenotypic assay with enhanced sensitivity.

The enhanced-sensitivity Trofile assay (TF-ES; Monogram Biosciences) was used to retest coreceptor tropism samples from 4 different cohorts of HIV-1-infected patients. Nine percent to 26% of patients with CCR5-tropic virus by the original Trofile assay had CXCR4-using virus by TF-ES. Lower CD4 cell counts were associated with CXCR4-using virus in all cohorts.

Design of a potent D-peptide HIV-1 entry inhibitor with a strong barrier to resistance.

The HIV gp41 N-trimer pocket region is an ideal viral target because it is extracellular, highly conserved, and essential for viral entry. Here, we report on the design of a pocket-specific D-peptide, PIE12-trimer, that is extraordinarily elusive to resistance and characterize its inhibitory and structural properties. D-peptides (peptides composed of D-amino acids) are promising therapeutic agents due to their insensitivity to protease degradation. PIE12-trimer was designed using structure-guided mirror-image phage display and linker optimization and is the first D-peptide HIV entry inhibitor with the breadth and potency required for clinical use. PIE12-trimer has an ultrahigh affinity for the gp41 pocket, providing it with a reserve of binding energy (resistance capacitor) that yields a dramatically improved resistance profile compared to those of other fusion inhibitors. These results demonstrate that the gp41 pocket is an ideal drug target and establish PIE12-trimer as a leading anti-HIV antiviral candidate.

Maraviroc versus efavirenz, both in combination with zidovudine-lamivudine, for the treatment of antiretroviral-naive subjects with CCR5-tropic HIV-1 infection.

BACKGROUND: The MERIT (Maraviroc versus Efavirenz in Treatment-Naive Patients) study compared maraviroc and efavirenz, both with zidovudine-lamivudine, in antiretroviral-naive patients with R5 human immunodeficiency virus type 1 (HIV-1) infection. METHODS: Patients screened for R5 HIV-1 were randomized to receive efavirenz (600 mg once daily) or maraviroc (300 mg once or twice daily) with zidovudine-lamivudine. Coprimary end points were proportions of patients with a viral load <400 and <50 copies/mL at week 48; the noninferiority of maraviroc was assessed. RESULTS: The once-daily maraviroc arm was discontinued for not meeting prespecified noninferiority criteria. In the primary 48-week analysis (n = 721), maraviroc was noninferior for <400 copies/mL (70.6% for maraviroc vs 73.1% for efavirenz) but not for <50 copies/mL (65.3% vs 69.3%) at a threshold of -10%. More maraviroc patients discontinued for lack of efficacy (11.9% vs 4.2%), but fewer discontinued for adverse events (4.2% vs 13.6%). In a post hoc reanalysis excluding 107 patients (15%) with non-R5 screening virus by the current, more sensitive tropism assay, the lower bound of the 1-sided 97.5% confidence interval for the difference between treatment groups was above -10% for each end point. CONCLUSIONS: Twice-daily maraviroc was not noninferior to efavirenz at <50 copies/mL in the primary analysis. However, 15% of patients would have been ineligible for inclusion by a more sensitive screening assay. Their retrospective exclusion resulted in similar response rates in both arms Trial registration. ClinicalTrials.gov identifier: (NCT00098293) .

Hepatitis C Resistance-Associated Substitutions Among People Who Inject Drugs Treated With Direct-Acting Antiviral-Containing Regimens.

BACKGROUND: Resistance-associated substitutions (RASs) to HCV direct-acting antivirals (DAAs) can contribute to virologic failure and limit retreatment options. People who inject drugs (PWID) are at highest risk for transmission of resistant virus. We report on RASs at baseline and after virologic failure in DAA-naive and protease inhibitor-experienced PWID. METHODS: We sequenced the NS3/4A, NS5A, and NS5B regions from 150 PWID with genotype 1 (GT1) viruses; 128 (85.3%) GT1a, 22 (14.7%) GT1b. RESULTS: Among the 139 (92.7%) DAA-naive PWID, 85 of 139 (61.2%) had baseline RASs-67 of 139 (48.2%) in NS3 (predominantly Q80K/L); 25 of 139 (18.0%) in NS5A; and 8 of 139 (5.8%) in NS5B. Of the 11 protease inhibitor-experienced participants, 9 had baseline NS3 RASs (V36L N = 1, Q80K N = 9) and 4 had baseline NS5A RASs (M28V N = 2, H58P N = 1, A92T N = 1). Among the 11 participants who had posttreatment samples with detectable virus (7 treatment failures, 1 late relapse, 3 reinfections), 1 sofosbuvir/ledipasvir failure had a baseline H58P. Two sofosbuvir/ledipasvir-treated participants developed new NS5A mutations (Q30H, Y93H, L31M/V). Otherwise, no RASs were detected. CONCLUSIONS: Our results demonstrate RAS prevalence among DAA-naive PWID is comparable to that in the general population. Only 2 of 150 (1.3%) in our longitudinal cohort developed treatment-emergent RASs. Concern for transmission of resistant virus may therefore be minimal.

Use of a small molecule CCR5 inhibitor in macaques to treat simian immunodeficiency virus infection or prevent simian-human immunodeficiency virus infection.

Human immunodeficiency virus type 1 (HIV-1) fuses with cells after sequential interactions between its envelope glycoproteins, CD4 and a coreceptor, usually CC chemokine receptor 5 (CCR5) or CXC receptor 4 (CXCR4). CMPD 167 is a CCR5-specific small molecule with potent antiviral activity in vitro. We show that CMPD 167 caused a rapid and substantial (4-200-fold) decrease in plasma viremia in six rhesus macaques chronically infected with simian immunodeficiency virus (SIV) strains SIVmac251 or SIVB670, but not in an animal infected with the X4 simian-human immunodeficiency virus (SHIV), SHIV-89.6P. In three of the SIV-infected animals, viremia reduction was sustained. In one, there was a rapid, but partial, rebound and in another, there was a rapid and complete rebound. There was a substantial delay (>21 d) between the end of therapy and the onset of full viremia rebound in two animals. We also evaluated whether vaginal administration of gel-formulated CMPD 167 could prevent vaginal transmission of the R5 virus, SHIV-162P4. Complete protection occurred in only 2 of 11 animals, but early viral replication was significantly less in the 11 CMPD 167-recipients than in 9 controls receiving carrier gel. These findings support the development of small molecule CCR5 inhibitors as antiviral therapies, and possibly as components of a topical microbicide to prevent HIV-1 sexual transmission.

Response to vicriviroc in treatment-experienced subjects, as determined by an enhanced-sensitivity coreceptor tropism assay: reanalysis of AIDS clinical trials group A5211.

The enhanced-sensitivity Trofile assay (Monogram Biosciences) was used to retest coreceptor use at both study screening and study entry for 118 treatment-experienced subjects in AIDS Clinical Trials Group A5211 who had CCR5-tropic (R5) virus detected by the original Trofile assay at study screening. Among 90 recipients of vicriviroc, a significantly (P< .001) greater mean reduction in HIV-1 RNA was observed in 72 subjects with R5 virus versus 15 subjects reclassified as having dual/mixed-tropic viruses at screening: -1.11 versus -0.09 log(10) copies/mL at day 14 and -1.91 versus -0.57 log(10) copies/mL at week 24, respectively. Results suggest that the enhanced-sensitivity assay is a better screening tool for determining patient eligibility for CCR5 antagonist therapy.

Comparison of human immunodeficiency virus type 1 tropism profiles in clinical samples by the Trofile and MT-2 assays.

The recent availability of CCR5 antagonists as anti-human immunodeficiency virus (anti-HIV) therapeutics has highlighted the need to accurately identify CXCR4-using variants in patient samples when use of this new drug class is considered. The Trofile assay (Monogram Biosciences) has become the method that is the most widely used to define tropism in the clinic prior to the use of a CCR5 antagonist. By comparison, the MT-2 assay has been used since early in the HIV epidemic to define tropism in clinical specimens. Given that there are few data from direct comparisons of these two assays, we evaluated the performance of the plasma-based Trofile assay and the peripheral blood mononuclear cell (PBMC)-based MT-2 assay for the detection of CXCR4 use in defining the tropism of HIV isolates derived from clinical samples. The various samples used for this comparison were derived from participants of the Amsterdam Cohort Studies on HIV infection and AIDS who underwent consecutive MT-2 assay testing of their PBMCs at approximately 3-month intervals. This unique sample set was specifically selected because consecutive MT-2 assays had demonstrated a shift from negative to positive in PBMCs, reflecting the first emergence of CXCR4-using virus in PBMCs above the level of detection of the assay in these individuals. Trofile testing was performed with clonal HIV type 1 (HIV-1) variants (n = 21), MT-2 cell culture-derived cells (n = 20) and supernatants (n = 42), and plasma samples (n = 76). Among the clonal HIV-1 variants and MT-2 cell culture-derived samples, the results of the Trofile and MT-2 assays demonstrated a high degree of concordance (95% to 98%). Among consecutive plasma samples, detection of CXCR4-using virus was at or before the time of first detection by the MT-2 assay in 5/10 patients by the original Trofile assay and in 9/10 patients by the enhanced-sensitivity Trofile assay. Differences in the time to the first detection of CXCR4 use between the MT-2 assay (PBMCs) and the original Trofile assay (plasma) were greatly reduced by the enhanced-sensitivity Trofile assay, suggesting that sensitivity for the detection of minor CXCR4-using variants may be a more important determinant of discordant findings than compartmentalization. The similarities in performance of the enhanced-sensitivity Trofile and MT-2 assays suggest that either may be an appropriate methodology to define tropism in patient specimens.

Coreceptor tropism can be influenced by amino acid substitutions in the gp41 transmembrane subunit of human immunodeficiency virus type 1 envelope protein.

Many studies have demonstrated that the third variable region (V3) of the human immunodeficiency virus type 1 (HIV-1) envelope protein (Env) is a major determinant of coreceptor tropism. Other regions in the surface gp120 subunit of Env can modulate coreceptor tropism in a manner that is not fully understood. In this study, we evaluated the effect of env determinants outside of V3 on coreceptor usage through the analysis of (i) patient-derived env clones that differ in coreceptor tropism, (ii) chimeric env sequences, and (iii) site-directed mutants. The introduction of distinct V3 sequences from CXCR4-using clones into an R5-tropic env backbone conferred the inefficient use of CXCR4 in some but not all cases. Conversely, in many cases, X4- and dual-tropic env backbones containing the V3 sequences of R5-tropic clones retained the ability to use CXCR4, suggesting that sequences outside of the V3 regions of these CXCR4-using clones were responsible for CXCR4 use. The determinants of CXCR4 use in a set of dual-tropic env sequences with V3 sequences identical to those of R5-tropic clones mapped to the gp41 transmembrane (TM) subunit. In one case, a single-amino-acid substitution in the fusion peptide of TM was able to confer CXCR4 use; however, TM substitutions associated with CXCR4 use varied among different env sequences. These results demonstrate that sequences in TM can modulate coreceptor specificity and that env sequences other than that of V3 may facilitate efficient CXCR4-mediated entry. We hypothesize that the latter plays an important role in the transition from CCR5 to CXCR4 coreceptor use.

Prevalence and impact of baseline resistance-associated substitutions on the efficacy of ledipasvir/sofosbuvir or simeprevir/sofosbuvir against GT1 HCV infection.

Baseline resistance-associated substitutions (RASs) have variable impacts in clinical trials but their prevalence and impact in real-world patients remains unclear. We performed baseline resistance testing using a commercial assay (10% cutoff) for 486 patients treated with LDV/SOF or SMV/SOF, with or without ribavirin, in the multi-center, observational HCV-TARGET cohort. Linkage of RASs was evaluated in selected samples using a novel quantitative single variant sequencing assay. Our results showed that the prevalence of NS3, NS5A, NS5B RASs was 45%, 13%, and 8%, respectively, and 10% of patients harbored RASs in 2 or more drug classes. Baseline LDV RASs in GT1a, TE, and cirrhosis LDV/SOF subgroup was associated with 2-4% lower SVR12 rates. SMV RASs was associated with lower SVR12 rates in GT1a, treatment-experienced, cirrhotics SMV/SOF subgroup. Pooled analysis of all patients with baseline RASs revealed that SVR12 was 100% (19/19) in patients treated for longer than 98 days but was 87% (81/93) in patients treated for shorter than 98 days. These results demonstrate that RASs prevalence and their impact in real world practice are in general agreement with registration trials, and suggest that longer treatment duration may overcome the negative impact of baseline RASs on SVR12 rates in clinical practice.

High-Sequence Diversity and Rapid Virus Turnover Contribute to Higher Rates of Coreceptor Switching in Treatment-Experienced Subjects with HIV-1 Viremia.

Coreceptor switching from CCR5 to CXCR4 is common during chronic HIV-1 infection, but is even more common in individuals who have failed antiretroviral therapy (ART). Prior studies have suggested rapid mutation and/or recombination of HIV-1 envelope (env) genes during coreceptor switching. We compared the functional and genotypic changes in env of viruses from viremic subjects who had failed ART just before and after coreceptor switching and compared those to viruses from matched subjects without coreceptor switching. Analysis of multiple unique functional env clones from each subject revealed extensive diversity at both sample time points and rapid diversification of sequences during the 4-month interval in viruses from both 9 subjects with coreceptor switching and 15 control subjects. Only two subjects had envs with evidence of recombination. Three findings distinguished env clones from subjects with coreceptor switching from controls: (1) lower entry efficiency via CCR5; (2) longer V1/V2 regions; and (3), lower nadir CD4 T cell counts during prior years of infection. Most of these subjects harbored virus with lower replicative capacity associated with protease (PR) and/or reverse transcriptase inhibitor resistance mutations, and the extensive diversification tended to lead either to improved entry efficiency via CCR5 or the gain of entry function via CXCR4. These results suggest that R5X4 or X4 variants emerge from a diverse, low-fitness landscape shaped by chronic infection, multiple ART resistance mutations, the availability of target cells, and reduced entry efficiency via CCR5.

Sequencing of hepatitis C virus for detection of resistance to direct-acting antiviral therapy: A systematic review.

The significance of the clinical impact of direct-acting antiviral (DAA) resistance-associated substitutions (RASs) in hepatitis C virus (HCV) on treatment failure is unclear. No standardized methods or guidelines for detection of DAA RASs in HCV exist. To facilitate further evaluations of the impact of DAA RASs in HCV, we conducted a systematic review of RAS sequencing protocols, compiled a comprehensive public library of sequencing primers, and provided expert guidance on the most appropriate methods to screen and identify RASs. The development of standardized RAS sequencing protocols is complicated due to a high genetic variability and the need for genotype- and subtype-specific protocols for multiple regions. We have identified several limitations of the available methods and have highlighted areas requiring further research and development. The development, validation, and sharing of standardized methods for all genotypes and subtypes should be a priority. (Hepatology Communications 2017;1:379-390).

Kinetic studies of HIV-1 and HIV-2 envelope glycoprotein-mediated fusion.

BACKGROUND: HIV envelope glycoprotein (Env)-mediated fusion is driven by the concerted coalescence of the HIV gp41 N-helical and C-helical regions, which results in the formation of 6 helix bundles. Kinetics of HIV Env-mediated fusion is an important determinant of sensitivity to entry inhibitors and antibodies. However, the parameters that govern the HIV Env fusion cascade have yet to be fully elucidated. We address this issue by comparing the kinetics HIV-1IIIB Env with those mediated by HIV-2 from two strains with different affinities for CD4 and CXCR4. RESULTS: HIV-1 and HIV-2 Env-mediated cell fusion occurred with half times of about 60 and 30 min, respectively. Binding experiments of soluble HIV gp120 proteins to CD4 and co-receptor did not correlate with the differences in kinetics of fusion mediated by the three different HIV Envs. However, escape from inhibition by reagents that block gp120-CD4 binding, CD4-induced CXCR4 binding and 6-helix bundle formation, respectively, indicated large difference between HIV-1 and HIV-2 envelope glycoproteins in their CD4-induced rates of engagement with CXCR4. CONCLUSION: The HIV-2 Env proteins studied here exhibited a significantly reduced window of time between the engagement of gp120 with CD4 and exposure of the CXCR4 binding site on gp120 as compared with HIV-1IIIB Env. The efficiency with which HIV-2 Env undergoes this CD4-induced conformational change is the major cause of the relatively rapid rate of HIV-2 Env mediated-fusion.

Cellular entry of HIV: Evaluation of therapeutic targets.

In the absence of a vaccine which could stop the HIV/AIDS pandemic, the development of therapeutic options is of utmost interest. The combined use of inhibitors of reverse transcriptase and protease as highly active antiretroviral therapy (HAART) provided the first effective treatment of HIV/AIDS and significantly decreased the number of AIDS related deaths in industrialized countries. However, the emergence of resistant viruses and the toxic side effects of HAART highlights that novel therapies are urgently required. The inhibition of HIV-1 entry is a promising option. Entry of HIV-1 into target cells involves interactions of the viral envelope protein (Env) with CD4 and a coreceptor, usually CCR5 or CXCR4. Env binding to receptor triggers several conformational rearrangements in Env, which involve the creation and/or exposure of structural intermediates pivotal to fusion of the viral and cellular membranes. Both, cellular receptors and structures in Env associated with membrane fusion are targets for therapeutic intervention. Here, we will discuss how HIV-1 enters cells and introduce strategies how this process can be inhibited.