B cell immunofocusing and repriming in two HIV-1 Env immunization regimens.

Diverse and rapidly mutating viruses pose challenges to immunogen and vaccine design. In this study, we evaluated the ability of memory B-cells obtained from two independent NHP trials to cross-react with individual HIV-1 vaccine components of two different multivalent immunization strategies. We demonstrated that while an HIV-1 Env multiclade, multivalent immunization regimen resulted in a dominant memory B-cell response that converged toward shared epitopes, in a sequential immunization with clonally-related non-stabilized gp140 HIV-1 Envs followed by SOSIP-stabilized gp140 trimers, the change in immunogen format resulted in repriming of the B-cell response.

In vivo delivery of synthetic DNA-encoded antibodies induces broad HIV-1-neutralizing activity.

Interventions to prevent HIV-1 infection and alternative tools in HIV cure therapy remain pressing goals. Recently, numerous broadly neutralizing HIV-1 monoclonal antibodies (bNAbs) have been developed that possess the characteristics necessary for potential prophylactic or therapeutic approaches. However, formulation complexities, especially for multiantibody deliveries, long infusion times, and production issues could limit the use of these bNAbs when deployed, globally affecting their potential application. Here, we describe an approach utilizing synthetic DNA-encoded monoclonal antibodies (dmAbs) for direct in vivo production of prespecified neutralizing activity. We designed 16 different bNAbs as dmAb cassettes and studied their activity in small and large animals. Sera from animals administered dmAbs neutralized multiple HIV-1 isolates with activity similar to that of their parental recombinant mAbs. Delivery of multiple dmAbs to a single animal led to increased neutralization breadth. Two dmAbs, PGDM1400 and PGT121, were advanced into nonhuman primates for study. High peak-circulating levels (between 6 and 34 µg/ml) of these dmAbs were measured, and the sera of all animals displayed broad neutralizing activity. The dmAb approach provides an important local delivery platform for the in vivo generation of HIV-1 bNAbs and for other infectious disease antibodies.

The Center for HIV/AIDS Vaccine Immunology (CHAVI) multi-site quality assurance program for cryopreserved human peripheral blood mononuclear cells.

The Center for HIV/AIDS Vaccine Immunology (CHAVI) consortium was established to determine the host and virus factors associated with HIV transmission, infection and containment of virus replication, with the goal of advancing the development of an HIV protective vaccine. Studies to meet this goal required the use of cryopreserved Peripheral Blood Mononuclear Cell (PBMC) specimens, and therefore it was imperative that a quality assurance (QA) oversight program be developed to monitor PBMC samples obtained from study participants at multiple international sites. Nine site-affiliated laboratories in Africa and the USA collected and processed PBMCs, and cryopreserved PBMC were shipped to CHAVI repositories in Africa and the USA for long-term storage. A three-stage program was designed, based on Good Clinical Laboratory Practices (GCLP), to monitor PBMC integrity at each step of this process. The first stage evaluated the integrity of fresh PBMCs for initial viability, overall yield, and processing time at the site-affiliated laboratories (Stage 1); for the second stage, the repositories determined post-thaw viability and cell recovery of cryopreserved PBMC, received from the site-affiliated laboratories (Stage 2); the third stage assessed the long-term specimen storage at each repository (Stage 3). Overall, the CHAVI PBMC QA oversight program results highlight the relative importance of each of these stages to the ultimate goal of preserving specimen integrity from peripheral blood collection to long-term repository storage.

Acute HIV-1 infection in the Southeastern United States: a cohort study.

In 1998 a collaboration between Duke University and the University of North Carolina, Chapel Hill (UNC) was founded to enhance identification of persons with acute HIV-1 infection (AHI). The Duke-UNC AHI Research Consortium Cohort consists of patients ≥18 years old with a positive nucleic acid amplification test (NAAT) and either a negative enzyme immunoassay (EIA) test or a positive EIA with a negative/indeterminate Western blot. Patients were referred to the cohort from acute care settings and state-funded HIV testing sites that use NAAT testing on pooled HIV-1 antibody-negative samples. Between 1998 and 2010, 155 patients with AHI were enrolled: 81 (52%) African-Americans, 63 (41%) white, non-Hispanics, 137 (88%) males, 108 (70%) men who have sex with men (MSM), and 18 (12%) females. The median age was 27 years (IQR 22-38). Most (n=138/155) reported symptoms with a median duration of 17.5 days. The median nadir CD4 count was 408 cells/mm(3) (IQR 289-563); the median observed peak HIV-1 level was 726,859 copies/ml (IQR 167,585-3,565,728). The emergency department was the most frequent site of initial presentation (n=55/152; 3 missing data). AHI diagnosis was made at time of first contact in 62/137 (45%; 18 missing data) patients. This prospectively enrolled cohort is the largest group of patients with AHI reported from the Southeastern United States. The demographics reflect the epidemic of this geographic area with a high proportion of African-Americans, including young black MSM. Highlighting the challenges of diagnosing AHI, less than half of the patients were diagnosed at the first healthcare visit. Women made up a small proportion despite increasing numbers in our clinics.

Efficacy of NNRTI-based antiretroviral therapy initiated during acute HIV infection.

OBJECTIVE: Characterize responses to non-nucleoside reverse transcriptase inhibitor (NNRTI)-based antiretroviral treatment (ART) initiated during acute HIV infection (AHI). DESIGN: This was a prospective, single-arm evaluation of once-daily, co-formulated emtricitabine/tenofovir/efavirenz initiated during AHI. METHODS: The primary endpoint is the proportion of responders with HIV RNA less than 200 copies/ml by week 24. We examined time to viral suppression and CD8 cell activation in relation to baseline participant characteristics. We compared time to viral suppression and viral dynamics using linear mixed-effects models between acutely infected participants and chronically infected controls. RESULTS: Between January 2005 and May 2009, 61 AHI participants were enrolled. Of participants whose enrollment date allowed 24 and 48 weeks of follow-up, 47 of 51 (92%) achieved viral suppression to less than 200 copies/ml by week 24, and 35 of 41 (85.4%) to less than 50 copies/ml by week 48. The median time from ART initiation to suppression below 50 copies/ml was 93 days (range 14-337). Higher HIV RNA levels at ART initiation (P = 0.02), but not time from estimated date of infection to ART initiation (P = 0.86), were associated with longer time to viral suppression. The median baseline frequency of activated CD8+CD38+HLA-DR+ T cells was 67% (range 40-95), and was not significantly associated with longer time to viral load suppression (P = 0.15). Viremia declined to less than 50 copies/ml more rapidly in AHI than chronically infected participants. Mixed-model analysis demonstrated similar phase I HIV RNA decay rates between acute and chronically infected participants, and more rapid viral decline in acutely infected participants in phase II. CONCLUSION: Once-daily emtricitabine/tenofovir/efavirenz initiated during AHI achieves rapid and sustained HIV suppression during this highly infectious period.

Transmitted antiretroviral drug resistance among acute and recent HIV infections in North Carolina from 1998 to 2007.

BACKGROUND: Transmitted drug resistance (TDR) limits antiretroviral options and thus complicates the management of HIV-positive patients. HIV disproportionately affects the southern US, but available national estimates of TDR prevalence principally reflect large metropolitan centres outside this region. METHODS: The Duke/UNC Acute HIV Program has collected data on acute or recent HIV infections (ARHI) in North Carolina since 1998. Acute infections represent antibody-negative, RNA-positive patients. Recent infection was determined by history of HIV testing or concordance between detuned ELISA and antibody avidity assays. Genotypic sequence data from the earliest collected pretreatment plasma samples were analysed with the Stanford HIV Database and screened for surveillance drug resistance mutations (SDRMs). RESULTS: A total of 253 individuals with ARHI between May 1998 and May 2007 had complete genotypic sequence data for analysis; 39.5% were acute infections, 78.7% were male, 64.8% were non-White and 53.8% were men who have sex with men. The overall prevalence of TDR was 17.8%, with SDRMs for non-nucleoside/nucleotide reverse transcriptase inhibitors (NNRTIs) in 9.5% of the cohort. Mutations for nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) were detected in 7.5% and for protease inhibitors (PIs) in 3.2%. K103N was the most common mutation (7.5%). Thymidine analogue mutations were found in 4.7% of samples; the most common PI SDRM was L90M (2.4%). Dual- or triple-class antiretroviral resistance was rare, encountered in only six (2.4%) samples. CONCLUSIONS: The prevalence of TDR in North Carolina is similar to estimates from the US metropolitan areas. These findings have implications for initial regimen selection and secondary prevention efforts outside of large, metropolitan HIV epicentres.

Affordable, abbreviated roche monitor assay for quantification of human immunodeficiency virus type 1 RNA in plasma.

The cost for the Roche Monitor assay kit can be reduced 50% by using only the 1:1, 1:25, and 1:625 human immunodeficiency virus (HIV) and the 1:1 quantitation standard dilutions. This abbreviated test applied to 1,774 mostly African samples had results nearly identical to those obtained following the package insert instructions. To make this approach feasible, Roche would have to provide additional lysis buffer and master mix.

GB virus C infection in children with perinatal human immunodeficiency virus infection.

BACKGROUND: GB virus C (GBV-C) infection occurs in 20-40% of human immunodeficiency virus (HIV)-infected adults, and coinfection is associated with improved HIV disease outcome. METHODS: To determine the prevalence of GBV-C infection in children who were perinatally infected with HIV, we conducted a cross-sectional prevalence survey in a cohort of perinatally infected HIV-positive children selected from a large, multicenter observational protocol. A blood specimen was obtained and tested for GBV-C viremia with the use of a qualitative GBV-C RNA assay and screened for past GBV-C infection with enzyme-linked immunosorbent assay to detect antibodies to the GBV-C envelope protein E2 (E2 Ab). RESULTS: The 354 children who participated in the substudy were relatively healthy, with a median CD4 of 784 cells/mm and median HIV-1 viral load of 1055 copies/mL. The prevalence of GBV-C viremia was 20 of 353 or 5.7% (95% confidence interval, 3.5-8.6%), and the prevalence of E2 Ab was 12 of 354 or 3.4% (95% confidence interval, 1.8-5.8%). GBV-C viremic patients were older than patients without past GBV-C infection (median age, 12.8 years versus 10.7 years). Median CD4 lymphocyte counts were highest in subjects without GBV-C infection and lowest in those with E2 Ab. CONCLUSIONS: GBV-C prevalence rates are lower in children with perinatal HIV infection than those reported for HIV-infected adults. With the exception of evidence that GBV-C viremic children had lower rates of Centers for Disease Control and Prevention HIV disease category C disease before GBV-C testing, we did not find evidence of improved HIV disease outcome in coinfected patients, but the number of HIV/GBV-C-coinfected children was small.

Hepatitis C prevalence in children with perinatal human immunodeficiency virus infection enrolled in a long-term follow-up protocol.

OBJECTIVE: To evaluate the prevalence of hepatitis C virus (HCV) infection in children with perinatal human immunodeficiency virus (HIV) infection. DESIGN: Cross-sectional substudy. SETTING: Multicenter study from 41 sites in the United States. PATIENTS: Children with perinatal HIV infection were randomly selected from a large, long-term, follow-up protocol. MAIN OUTCOME MEASURE: Hepatitis C infection was defined as having positive test results on both HCV antibody and HCV RNA assays. RESULTS: Five hundred thirty children enrolled in the substudy; definitive HCV test results were available for 525 children. Eighty-three percent were of a minority race or ethnicity. They were equally distributed by sex, had a median age of 10.7 years, and were relatively healthy, with 75% having CD4+ lymphocyte counts greater than 500 cells/mm3. Eight of 525 children (1.5%; 95% confidence interval [CI], 0.7%-3.0%) infected with HIV were coinfected with HCV. In contrast, the rate of HCV infection in a serosurvey of more than 2700 children aged 6 to 11 years from the National Health and Nutrition Examination Survey was 0.2% (95% CI, 0.04%-0.6%). In our study, there were no differences between children coinfected with HIV and HCV and those without HCV infection in terms of demographic characteristics, CD4+ or CD8+ T-lymphocyte counts, HIV 1 RNA levels, preterm or mode of delivery, or liver disease; however, the number of children coinfected with HIV and HCV was small. CONCLUSION: While HCV prevalence infection rates are low in children with perinatal HIV infection, they are 8 to 10 times higher than reported in HCV serosurveys of children in the United States.