Preferential and persistent impact of acute HIV-1 infection on CD4+ iNKT cells in colonic mucosa.

Acute HIV-1 infection (AHI) results in the widespread depletion of CD4+ T cells in peripheral blood and gut mucosal tissue. However, the impact on the predominantly CD4+ immunoregulatory invariant natural killer T (iNKT) cells during AHI remains unknown. Here, iNKT cells from peripheral blood and colonic mucosa were investigated during treated and untreated AHI. iNKT cells in blood were activated and rapidly depleted in untreated AHI. At the time of peak HIV-1 viral load, these cells showed the elevated expression of cell death-associated transcripts compared to preinfection. Residual peripheral iNKT cells suffered a diminished responsiveness to in vitro stimulation early into chronic infection. Additionally, HIV-1 DNA, as well as spliced and unspliced viral RNA, were detected in iNKT cells isolated from blood, indicating the active infection of these cells in vivo. The loss of iNKT cells occurred from Fiebig stage III in the colonic mucosa, and these cells were not restored to normal levels after initiation of ART during AHI. CD4+ iNKT cells were depleted faster and more profoundly than conventional CD4+ T cells, and the preferential infection of CD4+ iNKT cells over conventional CD4+ T cells was confirmed by in vitro infection experiments. In vitro data also provided evidence of latent infection in iNKT cells. Strikingly, preinfection levels of peripheral blood CD4+ iNKT cells correlated directly with the peak HIV-1 load. These findings support a model in which iNKT cells are early targets for HIV-1 infection, driving their rapid loss from circulation and colonic mucosa.

Longitudinal Analysis of Peripheral and Colonic CD161+ CD4+ T Cell Dysfunction in Acute HIV-1 Infection and Effects of Early Treatment Initiation.

CD161 expression on CD4+ T cells is associated with a Th17 functional phenotype, as well as with an innate capacity to respond to interleukin (IL)-12 and IL-18 without T cell receptor (TCR) stimulation. Chronic HIV-1 infection is associated with loss of the CD161+ CD4 T cell population, and non-human primate studies suggest that their depletion is associated with disease progression. However, the dynamics of the CD161+ CD4+ T cell population during acute HIV-1 infection remains unknown. In this study, we characterize peripheral blood CD161+ CD4+ T cells in detail, and examine how they are affected during the earliest stages of HIV-1 infection. Unbiased surface proteome screening and principal component analysis indicated that CD161+ CD4+ T cells are relatively phenotypically homogeneous between donors, and are intermediates between conventional CD4 T cells and innate-like T cells. In acute untreated HIV-1 infection, the circulating CD161+ CD4+ T cell population decreased in frequency, as did absolute cell counts starting from peak viral load, with elevated levels of activation and exhaustion markers expressed throughout acute HIV-1 infection. The capacity of these cells to respond to stimulation with IL-12 and IL-18 was also reduced. Early initiation of anti-retroviral treatment (ART) during acute HIV-1 infection restored the functionality of peripheral blood CD161+ CD4+ T cells, but not their frequency. In contrast, early ART initiation prevented the decline of colonic CD161+ CD4+ T cells that otherwise started during acute infection. Furthermore, loss of peripheral and colonic CD161+ CD4+ T cells in untreated infection was associated with levels of viral load. These results suggest that acute HIV-1 infection has profound effects on the CD161+ CD4+ T cell population that could not be completely prevented by the initiation of ART.

A randomized trial of vorinostat with treatment interruption after initiating antiretroviral therapy during acute HIV-1 infection.

OBJECTIVE AND DESIGN: A randomized, open-label pilot study in individuals treated with antiretroviral therapy (ART) since acute HIV infection (AHI) with a regimen including a histone deacetylase inhibitor to induce HIV from latency and control HIV replication during subsequent treatment interruption (TI). METHODS: Fifteen participants who initiated ART at AHI were randomized to vorinostat/hydroxychloroquine/maraviroc (VHM) plus ART (n â€‹= â€‹10) or ART alone (n â€‹= â€‹5). The VHM arm received three 14-day vorinostat cycles within 10 weeks before TI. ART was resumed for plasma viral load (VL) â€‹> â€‹1,000 HIV RNA copies/mL. Primary outcome was proportion of participants on VHM â€‹+ â€‹ART versus ART only with VL â€‹< â€‹50 copies/mL for 24 weeks after TI. RESULTS: Fifteen participants on ART (median: 178 weeks: range 79-295) enrolled. Two on VHM â€‹+ â€‹ART experienced serious adverse events. Fourteen participants underwent TI; all experienced VL rebound with no difference in time between arms: VHM â€‹+ â€‹ART (n â€‹= â€‹9) median: 4 weeks and ART only (n â€‹= â€‹5) median: 5 weeks. VHM induced a 2.2-fold increase in VL (p â€‹= â€‹0.008) by single-copy HIV RNA assay after the first cycle. Neopterin levels increased significantly following the first two cycles. After VHM treatment, the frequencies of peripheral blood mononuclear cells harboring total HIV DNA and cell-associated RNA were unchanged. All participants achieved VL suppression following ART re-initiation. CONCLUSIONS: Administration of VHM increased HIV VL in plasma, but this was not sustained. VHM did not impact time to viral rebound following TI and had no impact on the size of the HIV reservoir, suggesting that HIV reservoir elimination will require alternative treatment strategies.

Pattern and Frequency of Seroreactivity to Routinely Used Serologic Tests in Early-Treated Infants With HIV.

BACKGROUND: Previous studies have shown low frequencies of seroreactivity to HIV diagnostic assays for infected infants treated with antiretroviral therapy (ART) early in infection. METHODS: Fifty-eight HIV-infected infants treated with ART at a median age of 1.9 months (range: 0.2-5.4) for up to 4 years of life were assessed for seroreactivity to 4 routinely used HIV clinical immunoassays (IA): Second-generation (2ndG) IA and 2 rapid diagnostic tests (RDT), based on third-generation principles, measuring antibody only and a fourth-generation (4thG) antigen/antibody IA. HIV Western blot assay was also performed to assess HIV-specific antibodies. RESULTS: The 2ndG IA demonstrated the highest frequency of seroreactivity in children (69%) followed by the 4thG IA (40%) and the RDT (26%) after one year of ART. Infants initiating ART during ages 3-6 months (N = 15) showed a greater frequency (range: 53%-93%) and breadth (median and range: 3 [1-4]) of reactivity across the assays compared with those treated within 3 months (N = 43):16%-61% and breadth (1 [0-4]). The 4thG IA showed significantly reduced reactivity relative to the 2ndG IA at one (P = 0.016) and 3 (P = 0.004) years of ART. Western blot profiles following 3 years of ART showed the highest frequency of reactivity to HIV Gag p24 (76%) and lowest reactivity to Env gp120 and gp41, with only 24% of children confirmed positive by the assay. CONCLUSIONS: These results suggest that the use of 4thG IA and RDT test combination algorithms with limited HIV antigen breadth may not be adequate for diagnosis of HIV-infected children following early treatment.

Estimating HIV-1 incidence in Japan from the proportion of recent infections.

The first step of the UNAIDS/WHO 90-90-90 targets to encourage early diagnosis with treatment for the control of HIV-1 epidemic is to achieve 90% HIV-1 diagnosis in infected individuals. In Japan, approximately 30% of newly reported cases have been annually identified by AIDS onset, implying that substantial numbers of HIV-1-infected individuals remain undiagnosed. However, the proportion of undiagnosed cases has not yet been determined. In the present study, the proportion of recent HIV-1 infections to newly-diagnosed cases was determined from 2006 to 2015 using a recent infection assay for three metropolitan areas in Japan: Tokyo, Osaka, and Fukuoka. Estimated median periods between infection and diagnosis were 1.0, 1.8, and 2.9 years for Tokyo, Osaka, and Fukuoka, respectively. Estimation of annual HIV-1 incidence by a back-calculation method using these data as well as HIV/AIDS national surveillance data indicated proportions of undiagnosed to new HIV-1 infections from 2006 to 2015 to be 18% in Tokyo, 22% in Osaka, 38% in Fukuoka, and 28% in Japan. This is the first report estimating HIV-1 incidence in Japan using a serological biomarker in combination with national HIV/AIDS surveillance data.

Safety and efficacy of VRC01 broadly neutralising antibodies in adults with acutely treated HIV (RV397): a phase 2, randomised, double-blind, placebo-controlled trial.

BACKGROUND: HIV-1-specific broadly neutralising antibodies such as VRC01 could promote HIV remission by halting viral replication and clearing infected cells. We investigated whether VRC01 could promote sustained viral control off antiretroviral therapy (ART) in adults who initiated ART during acute HIV infection. METHODS: We did a randomised, double-blind, placebo-controlled trial at the Thai Red Cross AIDS Research Centre in Bangkok, Thailand. Eligible participants were aged 20-50 years, had initiated ART during acute infection (ie, Fiebig stages I-III), had been taking ART for more than 24 months, had fewer than 50 HIV-1 RNA copies per mL on three consecutive measurements, had more than 400 CD4 cells per µL, had fewer than ten copies of integrated HIV-1 DNA per 106 peripheral blood mononuclear cells, and were in generally good health. Eligible participants were randomly assigned (3:1) based on computer-generated lists with a blocking factor of 4 to receive VRC01 (40 mg/kg) or placebo (saline) intravenously every 3 weeks for up to 24 weeks during analytic interruption of ART, followed by continued observation off all therapies. Randomisation was stratified by Fiebig stage (I vs II vs III) at HIV diagnosis. Participants were monitored closely and resumed ART if 1000 or more HIV-1 RNA copies were detected per mL of plasma. The primary outcomes were the frequency of serious adverse events and the proportion of participants with fewer than 50 HIV-1 RNA copies per mL 24 weeks after treatment interruption. Efficacy analyses included all participants who received at least one full dose of study product, and safety analyses included all participants exposed to any study product. The trial was registered with ClinicalTrials.gov, number NCT02664415. This trial is completed. FINDINGS: Between Aug 8, 2016, and Jan 9, 2017, 19 men were randomly assigned, 14 to the VRC01 group and five to the placebo group. One participant in the VRC01 group received a partial infusion without undergoing treatment interruption. The other 18 participants all received at least one full study infusion and underwent ART interruption. No serious adverse events were reported in either group. Only one participant in the VRC01 group achieved the primary efficacy endpoint of viral suppression 24 weeks after ART interruption. The other 17 restarted ART because of a confirmed recording of 1000 or more HIV-1 RNA copies per mL before 24 weeks. INTERPRETATION: VRC01 monotherapy in individuals who initiated ART during acute HIV infection was well tolerated but did not significantly increase the number of participants with viral suppression 24 weeks after ART interruption. Further development of VRC01 and other immunotherapies for HIV will probably occur as part of combination regimens that include several treatments directed against unique therapeutic targets. FUNDING: US Department of the Army, US National Institutes of Health, and the Thai Red Cross AIDS Research Centre.

A flow cytometry based assay that simultaneously measures cytotoxicity and monocyte mediated antibody dependent effector activity.

Antibody effector functions such as antibody dependent cellular cytotoxicity (ADCC) and antibody dependent cellular phagocytosis (ADCP) are considered important immunologic parameters following results from the RV144 clinical trial where a reduced risk of infection was associated with non-neutralizing antibody against the V1/V2 region of HIV envelope. The rapid and fluorometric ADCC (RFADCC) assay has been widely used to measure ADCC, however, the mechanism behind the activity measured remains unclear. Here, we demonstrate that monocytes acquire the PKH26 dye used in the RFADCC assay and that the commonly used RFADCC readout correlates with phagocytosis. The RFADCC assay was combined with an amine reactive dye staining to confirm target cell killing. Interestingly, the majority of RFADCC and amine indices were mutually exclusive. In fact, the amine reactive assay results correlated with results from another assays that directly measure NK cell antibody effector functions not associated with phagocytosis. Together, this combined assay offers the opportunity to discriminate monocytes and NK cell antibody effector functions simultaneously.

Transmission dynamics among participants initiating antiretroviral therapy upon diagnosis of early acute HIV-1 infection in Thailand.

OBJECTIVE: To assess transmission characteristics in a predominantly MSM cohort initiating antiretroviral therapy (ART) immediately following diagnosis of acute HIV-1infection (AHI). METHODS: A longitudinal study (2009-2017) was performed in participants with AHI (n = 439) attending a single clinic in Bangkok. Plasma samples obtained prior to ART were used to obtain HIV-1 pol sequences and combined with clinical and epidemiologic data to assess transmission dynamics (cluster formation and size) using phylogenetic analysis. Clusters were estimated using maximum likelihood, genetic distance of 1.5% and visual inspection. The potential transmitter(s) in a cluster was determined using time to viral suppression and interview data. RESULTS: The cohort was predominantly MSM (93%) and infected with HIV-1 CRF01_AE (87%). Medians (ranges) for age and viral load prior to ART were 26 (18-70) years and 5.9 (2.5-8.2) log10 HIV-1 RNA copies/ml. Median time from history of HIV-1 exposure to diagnosis was 19 (3-61) days. Viral suppression was observed in 388 of 412 (94%) participants at a median time of 12 weeks following ART. Twenty-six clusters with median cluster size of 2 (2-5) representing 62 of 439 (14%) participants were observed. Younger age was associated with cluster formation: median 28 versus 30 years for unique infections (P = 0.01). A potential transmitter was identified in 11 of 26 (42%) clusters. CONCLUSION: Despite high rates of viral suppression following diagnosis and treatment of AHI within a cohort of young Thai MSM, HIV-1 transmission continued, reflecting the need to expand awareness and treatment access to the entire MSM population.

Characterization of Cellular Immune Responses in Thai Individuals With and Without HIV-Associated Neurocognitive Disorders.

HIV-associated neurocognitive disorder (HAND) remains a challenge despite antiretroviral therapy (ART), and has been linked to monocyte/macrophage (M/M) migration to the brain. Due to the potential impact of T cell effector mechanisms in eliminating activated/HIV-infected M/M, T cell activation may play a role in the development of HAND. We sought to investigate the relationship between cognition and both CD8+ T cell activation (HLA-DR+/CD38+) and HIV-specific CD8+ T cell responses at the time of HIV diagnosis and 12 months postinitiation of ART. CD8+ T cell activation was increased in HAND compared to cognitive normal (NL) individuals and correlated directly with plasma viral load and inversely with the cognitive status. In addition, Gag-specific cytolytic activity (CD107a/b+) was decreased in HAND compared with NL individuals and correlated with their neurological testing, suggesting a potential role of cytotoxic CD8+ T cells in the mechanism of HAND development.

Characterization of HIV-1 gp120 antibody specificities induced in anogenital secretions of RV144 vaccine recipients after late boost immunizations.

Sexual transmission is the principal driver of the human immunodeficiency virus (HIV) pandemic. Understanding HIV vaccine-induced immune responses at mucosal surfaces can generate hypotheses regarding mechanisms of protection, and may influence vaccine development. The RV144 (ClinicalTrials.gov NCT00223080) efficacy trial showed protection against HIV infections but mucosal samples were not collected, therefore, the contribution of mucosal antibodies to preventing HIV-1 acquisition is unknown. Here, we report the generation, magnitude and persistence of antibody responses to recombinant gp120 envelope and antigens including variable one and two loop scaffold antigens (gp70V1V2) previously shown to correlate with risk in RV144. We evaluated antibody responses to gp120 A244gD and gp70V1V2 92TH023 (both CRF01_AE) and Case A2 (subtype B) in cervico-vaginal mucus (CVM), seminal plasma (SP) and rectal secretions (RS) from HIV-uninfected RV144 vaccine recipients, who were randomized to receive two late boosts of ALVAC-HIV/AIDSVAX®B/E, AIDSVAX®B/E, or ALVAC-HIV alone at 0 and 6 months. Late vaccine boosting increased IgG geometric mean titers (GMT) to gp120 A244gD in AIDSVAX®B/E and ALVAC-HIV/AIDSVAX®B/E CVM (28 and 17 fold, respectively), followed by SP and RS. IgG to gp70V1V2 92TH023 increased in AIDSVAX®B/E and ALVAC-HIV/AIDSVAX®B/E CVM (11-17 fold) and SP (2 fold) two weeks post first boost. IgG to Case A2 was only detected in AIDSVAX®B/E and ALVAC-HIV/AIDSVAX®B/E CVM. Mucosal IgG to gp120 A244gD (CVM, SP, RS), gp70V1V2 92TH023 (CVM, SP), and Case A2 (CVM) correlated with plasma IgG levels (p<0.001). Although the magnitude of IgG responses declined after boosting, anti-gp120 A244gD IgG responses in CVM persisted for 12 months post final vaccination. Further studies in localization, persistence and magnitude of envelope specific antibodies (IgG and dimeric IgA) in anogenital secretions will help determine their role in preventing mucosal HIV acquisition.

Acute HIV infection detection and immediate treatment estimated to reduce transmission by 89% among men who have sex with men in Bangkok.

INTRODUCTION: Antiretroviral treatment (ART) reduces HIV transmission. Despite increased ART coverage, incidence remains high among men who have sex with men (MSM) in many places. Acute HIV infection (AHI) is characterized by high viral replication and increased infectiousness. We estimated the feasible reduction in transmission by targeting MSM with AHI for early ART. METHODS: We recruited a cohort of 88 MSM with AHI in Bangkok, Thailand, who initiated ART immediately. A risk calculator based on viral load and reported behaviour, calibrated to Thai epidemiological data, was applied to estimate the number of onwards transmissions. This was compared with the expected number without early interventions. RESULTS: Forty of the MSM were in 4th-generation AHI stages 1 and 2 (4thG stage 1, HIV nucleic acid testing (NAT)+/4thG immunoassay (IA)-/3rdG IA-; 4thG stage 2, NAT+/4thG IA+/3rdG IA-) while 48 tested positive on third-generation IA but had negative or indeterminate western blot (4thG stage 3). Mean plasma HIV RNA was 5.62 log10 copies/ml. Any condomless sex in the four months preceding the study was reported by 83.7%, but decreased to 21.2% by 24 weeks on ART. After ART, 48/88 (54.6%) attained HIV RNA <50 copies/ml by week 8, increasing to 78/87 (89.7%), and 64/66 (97%) at weeks 24 and 48, respectively. The estimated number of onwards transmissions in the first year of infection would have been 27.3 (95% credible interval: 21.7-35.3) with no intervention, 8.3 (6.4-11.2) with post-diagnosis behaviour change only, 5.9 (4.4-7.9) with viral load reduction only and 3.1 (2.4-4.3) with both. The latter was associated with an 88.7% (83.8-91.1%) reduction in transmission. CONCLUSIONS: Disproportionate HIV transmission occurs during AHI. Diagnosis of AHI with early ART initiation can substantially reduce onwards transmission.

Comparison of Antibody Responses Induced by RV144, VAX003, and VAX004 Vaccination Regimens.

The RV144 prime-boost regimen demonstrated efficacy against HIV acquisition while VAX003 and VAX004 did not. Although these trials differed by risk groups, immunization regimens, and immunogens, antibody responses may have contributed to the differences observed in vaccine efficacy. We assessed HIV-specific IgG, both total and subclass, and IgA binding to HIV envelope (Env): gp120 proteins and Cyclic V2 (CycV2) and CycV3 peptides and gp70 V1 V2 scaffolds in these 3 HIV vaccine trials. After two protein immunizations, IgG responses to 92TH023 gp120 (contained in ALVAC-HIV vaccine) were significantly higher in RV144 but responses to other Env were higher in the VAX trials lacking ALVAC-HIV. IgG responses declined significantly between vaccinations. All trials induced antibodies to gp70 V1 V2 but VAX004 responses to 92TH023 gp70 V1 V2 were weak. All CycV2 responses were undetectable in VAX004 while 92TH023 gp70 V1 V2 was detected in both RV144 and VAX003 but MN CycV2 was detected only in VAX003. Multiple protein vaccinations in VAX trials did not improve magnitude or durability of V1 V2 and CycV2 antibodies. Herpes simplex virus glycoprotein D (gD) peptide at the N terminus of AIDSVAX® B/E and B/B gp120 proteins induced antibodies in all trials, although significantly higher in VAX trials. gD peptide induced IgA, IgG1, IgG2, and IgG3 but not IgG4. Multiple protein vaccinations decreased IgG3 and increased IgG4 changing subclass contribution to total IgG. Although confounded by different modes of HIV transmission, higher Env-specific IgA and IgG4 binding antibodies induced in the VAX trials compared to RV144 raises the hypothesis that these differences may have contributed to different vaccine efficacy results.

Initiation of Antiretroviral Therapy During Acute HIV-1 Infection Leads to a High Rate of Nonreactive HIV Serology.

BACKGROUND: Third- and fourth-generation immunoassays (IAs) are widely used in the diagnosis of human immunodeficiency virus (HIV) infection. Antiretroviral therapy (ART) during acute HIV infection (AHI) may impact HIV-specific antibodies, with failure to develop antibody or seroreversion. We report on the ability of diagnostic tests to detect HIV-specific antibodies in Thai participants initiating ART during AHI. METHODS: Participants with detectable plasma HIV RNA but nonreactive HIV-specific immunoglobulin G, enrolled in an AHI study, were offered immediate initiation of ART. Participants were tested at initiation and at 12 and 24 weeks following treatment using standard second-, third-, and fourth-generation IAs and Western blot (WB). RESULTS: Participants (N = 234) initiating ART at a median of 19 days (range, 1-62 days) from HIV exposure demonstrated different frequencies of reactivity prior to and following 24 weeks of ART depending on the IA. Third-generation IA nonreactivity prior to ART was 48%, which decreased to 4% following ART (P < .001). Fourth-generation IA nonreactivity was 18% prior to ART and 17% following ART (P = .720). Negative WB results were observed in 89% and 12% of participants prior to and following 24 weeks of ART, respectively (P < .001). Seroreversion to nonreactivity during ART was observed to at least one of the tests in 20% of participants, with fourth-generation IA demonstrating the highest frequency (11%) of seroreversion. CONCLUSIONS: HIV-specific antibodies may fail to develop and, when detected, may decline when ART is initiated during AHI. Although fourth-generation IA was the most sensitive at detecting AHI prior to ART, third-generation IA was the most sensitive during treatment. CLINICAL TRIALS REGISTRATION: NCT00796146 and NCT00796263.

Prospective Study of Acute HIV-1 Infection in Adults in East Africa and Thailand.

BACKGROUND: Acute human immunodeficiency virus type 1 (HIV-1) infection is a major contributor to transmission of HIV-1. An understanding of acute HIV-1 infection may be important in the development of treatment strategies to eradicate HIV-1 or achieve a functional cure. METHODS: We performed twice-weekly qualitative plasma HIV-1 RNA nucleic acid testing in 2276 volunteers who were at high risk for HIV-1 infection. For participants in whom acute HIV-1 infection was detected, clinical observations, quantitative measurements of plasma HIV-1 RNA levels (to assess viremia) and HIV antibodies, and results of immunophenotyping of lymphocytes were obtained twice weekly. RESULTS: Fifty of 112 volunteers with acute HIV-1 infection had two or more blood samples collected before HIV-1 antibodies were detected. The median peak viremia (6.7 log10 copies per milliliter) occurred 13 days after the first sample showed reactivity on nucleic acid testing. Reactivity on an enzyme immunoassay occurred at a median of 14 days. The nadir of viremia (4.3 log10 copies per milliliter) occurred at a median of 31 days and was nearly equivalent to the viral-load set point, the steady-state viremia that persists durably after resolution of acute viremia (median plasma HIV-1 RNA level, 4.4 log10 copies per milliliter). The peak viremia and downslope were correlated with the viral-load set point. Clinical manifestations of acute HIV-1 infection were most common just before and at the time of peak viremia. A median of one symptom of acute HIV-1 infection was recorded at a median of two study visits, and a median of one sign of acute HIV-1 infection was recorded at a median of three visits. CONCLUSIONS: The viral-load set point occurred at a median of 31 days after the first detection of plasma viremia and correlated with peak viremia. Few symptoms and signs were observed during acute HIV-1 infection, and they were most common before peak viremia. (Funded by the Department of Defense and the National Institute of Allergy and Infectious Diseases.).

Sex and Urbanicity Contribute to Variation in Lymphocyte Distribution across Ugandan Populations.

Management of patient care and interpretation of research data require evaluation of laboratory results in the context of reference data from populations with known health status to adequately diagnose disease or make a physiological assessment. Few studies have addressed the diversity of lymphocyte subsets in rural and urban Ugandan populations. Here, 663 healthy blood bank donors from semi-urban centers of Kampala consented to participate in a study to define lymphocyte reference ranges. Whole blood immunophenotyping was performed to determine the frequency and absolute counts of T, B, and NK cells using clinical flow cytometry. Results from blood bank donors were compared to a rural cohort from the district of Kayunga and more urban clinical trial participants from the capital city, Kampala. Relationships between hematological and lymphocyte parameters were also explored. In the semi-urban blood donors, females were significantly different from males in all parameters except the frequency of CD8 T and B cells. Females had higher absolute counts of CD4 T, CD8 T and B cells, whereas males had higher NK cell counts. NK cell frequency and counts were significantly higher in semi-urban blood donors, regardless of sex, compared to more urban study participants. CD8 T cell frequency and counts were significantly higher in the blood donors compared to the rural participants, irrespective of sex. Interestingly, basophil counts were positively associated with overall T cell counts. These findings suggest that both sex and level of cohort urbanicity may influence lymphocyte subset distributions in Ugandans.

IgG Antibody Responses to Recombinant gp120 Proteins, gp70V1/V2 Scaffolds, and a CyclicV2 Peptide in Thai Phase I/II Vaccine Trials Using Different Vaccine Regimens.

RV144 correlates of risk analysis showed that IgG antibodies to gp70V1V2 scaffolds inversely correlated with risk of HIV acquisition. We investigated IgG antibody responses in RV135 and RV132, two ALVAC-HIV prime-boost vaccine trials conducted in Thailand prior to RV144. Both trials used ALVAC-HIV (vCP1521) at 0, 1, 3, and 6 months and HIV-1 gp120MNgD and gp120A244gD in alum (RV135) or gp120SF2 and gp120CM235 in MF59 (RV132) at 3 and 6 months. We assessed ELISA binding antibodies to the envelope proteins (Env) 92TH023, A244gD and MNgD, cyclicV2, and gp70V1V2 CaseA2 (subtype B) and 92TH023 (subtype CRF01_AE), and Env-specific IgG1 and IgG3. Antibody responses to gp120 A244gD, MNgD, and gp70V1V2 92TH023 scaffold were significantly higher in RV135 than in RV132. Antibodies to gp70V1V2 CaseA2 were detected only in RV135 vaccine recipients and IgG1 and IgG3 antibody responses to A244gD were significantly higher in RV135. IgG binding to gp70V1V2 CaseA2 and CRF01_AE scaffolds was higher with the AIDSVAX(®)B/E boost but both trials showed similar rates of antibody decline post-vaccination. MF59 did not result in higher IgG antibody responses compared to alum with the antigens tested. However, notable differences in the structure of the recombinant proteins and dosage used for immunizations may have contributed to the magnitude and specificity of IgG induced by the two trials.

Impact of nucleic acid testing relative to antigen/antibody combination immunoassay on the detection of acute HIV infection.

OBJECTIVE: To assess the addition of HIV nucleic acid testing (NAT) to fourth-generation (4thG) HIV antigen/antibody combination immunoassay in improving detection of acute HIV infection (AHI). METHODS: Participants attending a major voluntary counseling and testing site in Thailand were screened for AHI using 4thG HIV antigen/antibody immunoassay and sequential less sensitive HIV antibody immunoassay. Samples nonreactive by 4thG antigen/antibody immunoassay were further screened using pooled NAT to identify additional AHI. HIV infection status was verified following enrollment into an AHI study with follow-up visits and additional diagnostic tests. RESULTS: Among 74 334 clients screened for HIV infection, HIV prevalence was 10.9% and the overall incidence of AHI (N = 112) was 2.2 per 100 person-years. The inclusion of pooled NAT in the testing algorithm increased the number of acutely infected patients detected, from 81 to 112 (38%), relative to 4thG HIV antigen/antibody immunoassay. Follow-up testing within 5 days of screening marginally improved the 4thG immunoassay detection rate (26%). The median CD4 T-cell count at the enrollment visit was 353 cells/µl and HIV plasma viral load was 598 289 copies/ml. CONCLUSION: The incorporation of pooled NAT into the HIV testing algorithm in high-risk populations may be beneficial in the long term. The addition of pooled NAT testing resulted in an increase in screening costs of 22% to identify AHI: from $8.33 per screened patient to $10.16. Risk factors of the testing population should be considered prior to NAT implementation given the additional testing complexity and costs.

Identification of immunodominant CD4-restricted epitopes co-located with antibody binding sites in individuals vaccinated with ALVAC-HIV and AIDSVAX B/E.

We performed fine epitope mapping of the CD4+ responses in the ALVAC-HIV-AIDSVAX B/E prime-boost regimen in the Thai Phase III trial (RV144). Non-transformed Env-specific T cell lines established from RV144 vaccinees were used to determine the fine epitope mapping of the V2 and C1 responses and the HLA class II restriction. Data showed that there are two CD4+ epitopes contained within the V2 loop: one encompassing the α4ß7 integrin binding site (AA179-181) and the other nested between two previously described genetic sieve signatures (AA169, AA181). There was no correlation between the frequencies of CD4+ fine epitope responses and binding antibody.

Initiation of ART during early acute HIV infection preserves mucosal Th17 function and reverses HIV-related immune activation.

Mucosal Th17 cells play an important role in maintaining gut epithelium integrity and thus prevent microbial translocation. Chronic HIV infection is characterized by mucosal Th17 cell depletion, microbial translocation and subsequent immune-activation, which remain elevated despite antiretroviral therapy (ART) correlating with increased mortality. However, when Th17 depletion occurs following HIV infection is unknown. We analyzed mucosal Th17 cells in 42 acute HIV infection (AHI) subjects (Fiebig (F) stage I-V) with a median duration of infection of 16 days and the short-term impact of early initiation of ART. Th17 cells were defined as IL-17+ CD4+ T cells and their function was assessed by the co-expression of IL-22, IL-2 and IFNγ. While intact during FI/II, depletion of mucosal Th17 cell numbers and function was observed during FIII correlating with local and systemic markers of immune-activation. ART initiated at FI/II prevented loss of Th17 cell numbers and function, while initiation at FIII restored Th17 cell numbers but not their polyfunctionality. Furthermore, early initiation of ART in FI/II fully reversed the initially observed mucosal and systemic immune-activation. In contrast, patients treated later during AHI maintained elevated mucosal and systemic CD8+ T-cell activation post initiation of ART. These data support a loss of Th17 cells at early stages of acute HIV infection, and highlight that studies of ART initiation during early AHI should be further explored to assess the underlying mechanism of mucosal Th17 function preservation.

Polyfunctional Fc-effector profiles mediated by IgG subclass selection distinguish RV144 and VAX003 vaccines.

The human phase 2B RV144 ALVAC-HIV vCP1521/AIDSVAX B/E vaccine trial, held in Thailand, resulted in an estimated 31.2% efficacy against HIV infection. By contrast, vaccination with VAX003 (consisting of only AIDSVAX B/E) was not protective. Because protection within RV144 was observed in the absence of neutralizing antibody activity or cytotoxic T cell responses, we speculated that the specificity or qualitative differences in Fc-effector profiles of nonneutralizing antibodies may have accounted for the efficacy differences observed between the two trials. We show that the RV144 regimen elicited nonneutralizing antibodies with highly coordinated Fc-mediated effector responses through the selective induction of highly functional immunoglobulin G3 (IgG3). By contrast, VAX003 elicited monofunctional antibody responses influenced by IgG4 selection, which was promoted by repeated AIDSVAX B/E protein boosts. Moreover, only RV144 induced IgG1 and IgG3 antibodies targeting the crown of the HIV envelope V2 loop, albeit with limited coverage of breakthrough viral sequences. These data suggest that subclass selection differences associated with coordinated humoral functional responses targeting strain-specific protective V2 loop epitopes may underlie differences in vaccine efficacy observed between these two vaccine trials.