Human immunoglobulin gene allelic variation impacts germline-targeting vaccine priming.

Vaccine priming immunogens that activate germline precursors for broadly neutralizing antibodies (bnAbs) have promise for development of precision vaccines against major human pathogens. In a clinical trial of the eOD-GT8 60mer germline-targeting immunogen, higher frequencies of vaccine-induced VRC01-class bnAb-precursor B cells were observed in the high dose compared to the low dose group. Through immunoglobulin heavy chain variable (IGHV) genotyping, statistical modeling, quantification of IGHV1-2 allele usage and B cell frequencies in the naive repertoire for each trial participant, and antibody affinity analyses, we found that the difference between dose groups in VRC01-class response frequency was best explained by IGHV1-2 genotype rather than dose and was most likely due to differences in IGHV1-2 B cell frequencies for different genotypes. The results demonstrate the need to define population-level immunoglobulin allelic variations when designing germline-targeting immunogens and evaluating them in clinical trials.

Ultrasound-guided lymph node fine-needle aspiration for evaluating post-vaccination germinal center responses in humans.

The lymph node (LN) is a critical biological site for immune maturation after vaccination as it includes several cell populations critical for priming the antibody response. Here, we present a protocol for sampling the LN and isolating cell populations to evaluate immunogens targeting germline cells. We describe steps for media and tube preparation and sample collection using an ultrasound-guided LN fine-needle aspiration procedure. This protocol is safe, quick, low-cost, and less invasive than excisional biopsy. For complete details on the use and execution of this protocol, please refer to Leggat et al. (2022).1.

A first-in-human germline-targeting HIV nanoparticle vaccine induced broad and publicly targeted helper T cell responses.

The engineered outer domain germline targeting version 8 (eOD-GT8) 60-mer nanoparticle was designed to prime VRC01-class HIV-specific B cells that would need to be matured, through additional heterologous immunizations, into B cells that are able to produce broadly neutralizing antibodies. CD4 T cell help will be critical for the development of such high-affinity neutralizing antibody responses. Thus, we assessed the induction and epitope specificities of the vaccine-specific T cells from the IAVI G001 phase 1 clinical trial that tested immunization with eOD-GT8 60-mer adjuvanted with AS01B. Robust polyfunctional CD4 T cells specific for eOD-GT8 and the lumazine synthase (LumSyn) component of eOD-GT8 60-mer were induced after two vaccinations with either the 20- or 100-microgram dose. Antigen-specific CD4 T helper responses to eOD-GT8 and LumSyn were observed in 84 and 93% of vaccine recipients, respectively. CD4 helper T cell epitope "hotspots" preferentially targeted across participants were identified within both the eOD-GT8 and LumSyn proteins. CD4 T cell responses specific to one of these three LumSyn epitope hotspots were observed in 85% of vaccine recipients. Last, we found that induction of vaccine-specific peripheral CD4 T cells correlated with expansion of eOD-GT8-specific memory B cells. Our findings demonstrate strong human CD4 T cell responses to an HIV vaccine candidate priming immunogen and identify immunodominant CD4 T cell epitopes that might improve human immune responses either to heterologous boost immunogens after this prime vaccination or to other human vaccine immunogens.

Human immunoglobulin gene allelic variation impacts germline-targeting vaccine priming.

Vaccine priming immunogens that activate germline precursors for broadly neutralizing antibodies (bnAbs) have promise for development of precision vaccines against major human pathogens. In a clinical trial of the eOD-GT8 60mer germline-targeting immunogen, higher frequencies of vaccine-induced VRC01-class bnAb-precursor B cells were observed in the high dose compared to the low dose group. Through immunoglobulin heavy chain variable (IGHV) genotyping, statistical modeling, quantification of IGHV1-2 allele usage and B cell frequencies in the naive repertoire for each trial participant, and antibody affinity analyses, we found that the difference between dose groups in VRC01-class response frequency was best explained by IGHV1-2 genotype rather than dose and was most likely due to differences in IGHV1-2 B cell frequencies for different genotypes. The results demonstrate the need to define population-level immunoglobulin allelic variations when designing germline-targeting immunogens and evaluating them in clinical trials. One-Sentence Summary: Human genetic variation can modulate the strength of vaccine-induced broadly neutralizing antibody precursor B cell responses.

Female sex workers in Kigali, Rwanda: a key population at risk of HIV, sexually transmitted infections, and unplanned pregnancy.

Female sex workers (FSWs) were recruited from known hotspots in Kigali, Rwanda, and offered free, anonymous human immunodeficiency virus (HIV) counseling and testing, diagnosis and treatment of sexually transmitted infections (STIs) and long-acting reversible contraception (LARC). From September 2012 to March 2015, 1168 FSWs sought services, including 587 (50%) who were HIV-positive. More than 90% had previously tested for HIV, and 26% who reported previously testing negative had seroconverted. Of the 349 who already knew their HIV-positive status, 74% were on antiretroviral treatment. The prevalence of serologic syphilis was 43% in HIV-positive and 19% in HIV-negative FSWs (p < 0.0001), and Trichomonas vaginalis was found in vaginal wet mounts in 21% of HIV-positive and 13% of HIV-negative FSWs (p < 0.0001). Signs and symptoms of STIs were found in 35% of HIV-positive compared with 21% of HIV-negative FSWs (p < 0.0001). Only one-third reported consistent condom use in the last month. Modern contraceptive use was reported by 43% of HIV-positive and 56% of HIV-negative FSWs (p < 0.0001). Current pregnancy was reported by 4% of HIV-positive and 6% of HIV-negative FSWs (p = 0.0409). Despite Rwanda's successes with preventing 70% of new infections in the general population through nationwide couples' testing in antenatal clinics, prevention and timely treatment in key populations including FSWs are lacking. The prevalence of HIV - including many new cases - and STIs among FSWs in Kigali is high and condom and contraceptive use are low. Tailored and integrated HIV/STIs and family planning programs are urgently needed for FSWs.

First-in-Human Evaluation of the Safety and Immunogenicity of an Intranasally Administered Replication-Competent Sendai Virus-Vectored HIV Type 1 Gag Vaccine: Induction of Potent T-Cell or Antibody Responses in Prime-Boost Regimens.

BACKGROUND: We report the first-in-human safety and immunogenicity assessment of a prototype intranasally administered, replication-competent Sendai virus (SeV)-vectored, human immunodeficiency virus type 1 (HIV-1) vaccine. METHODS: Sixty-five HIV-1-uninfected adults in Kenya, Rwanda, and the United Kingdom were assigned to receive 1 of 4 prime-boost regimens (administered at 0 and 4 months, respectively; ratio of vaccine to placebo recipients, 12:4): priming with a lower-dose SeV-Gag given intranasally, followed by boosting with an adenovirus 35-vectored vaccine encoding HIV-1 Gag, reverse transcriptase, integrase, and Nef (Ad35-GRIN) given intramuscularly (SLA); priming with a higher-dose SeV-Gag given intranasally, followed by boosting with Ad35-GRIN given intramuscularly (SHA); priming with Ad35-GRIN given intramuscularly, followed by boosting with a higher-dose SeV-Gag given intranasally (ASH); and priming and boosting with a higher-dose SeV-Gag given intranasally (SHSH). RESULTS: All vaccine regimens were well tolerated. Gag-specific IFN-γ enzyme-linked immunospot-determined response rates and geometric mean responses were higher (96% and 248 spot-forming units, respectively) in groups primed with SeV-Gag and boosted with Ad35-GRIN (SLA and SHA) than those after a single dose of Ad35-GRIN (56% and 54 spot-forming units, respectively) or SeV-Gag (55% and 59 spot-forming units, respectively); responses persisted for ≥8 months after completion of the prime-boost regimen. Functional CD8+ T-cell responses with greater breadth, magnitude, and frequency in a viral inhibition assay were also seen in the SLA and SHA groups after Ad35-GRIN boost, compared with those who received either vaccine alone. SeV-Gag did not boost T-cell counts in the ASH group. In contrast, the highest Gag-specific antibody titers were seen in the ASH group. Mucosal antibody responses were sporadic. CONCLUSIONS: SeV-Gag primed functional, durable HIV-specific T-cell responses and boosted antibody responses. The prime-boost sequence appears to determine which arm of the immune response is stimulated. CLINICAL TRIALS REGISTRATION: NCT01705990.

Correction: Antigen-Specific Antibody Glycosylation Is Regulated via Vaccination.

[This corrects the article DOI: 10.1371/journal.ppat.1005456.].

Assessment of Anti-HIV-1 Antibodies in Oral and Nasal Compartments of Volunteers From 3 Different Populations.

In this study, we assessed the feasibility of collecting standardized nasal and salivary samples at centers in Nairobi (Kenya), Kigali (Rwanda), and London (United Kingdom) using different collection devices and media (synthetic absorptive matrices versus flocked swabs, and Salimetrics oral swabs versus whole oral fluid collection). We detected anti-Gag (p24) and envelope (gp140) antibodies in both nasal fluid and salivary collections from all HIV-infected individuals, and cross-reactive anti-p24 antibodies were detected in 10% of HIV-uninfected individuals enrolled at one site. Collections from the nasal turbinates were comparable with samples collected deeper in the nasopharyngeal tract, and the yield of anti-p24 IgA in the whole oral fluid samples was higher than in samples collected from the parotid gland. We noted a trend toward reduced levels of anti-HIV antibody in the volunteers receiving anti-retroviral therapy. Levels of antibodies were stable over multiple collection visits. Overall, this study shows that nasal and salivary samples can be collected in a standardized manner over repeated visits in both low- and high-resource settings. These methods may be used in support for future HIV vaccine clinical trials.

Antigen-Specific Antibody Glycosylation Is Regulated via Vaccination.

Antibody effector functions, such as antibody-dependent cellular cytotoxicity, complement deposition, and antibody-dependent phagocytosis, play a critical role in immunity against multiple pathogens, particularly in the absence of neutralizing activity. Two modifications to the IgG constant domain (Fc domain) regulate antibody functionality: changes in antibody subclass and changes in a single N-linked glycan located in the CH2 domain of the IgG Fc. Together, these modifications provide a specific set of instructions to the innate immune system to direct the elimination of antibody-bound antigens. While it is clear that subclass selection is actively regulated during the course of natural infection, it is unclear whether antibody glycosylation can be tuned, in a signal-specific or pathogen-specific manner. Here, we show that antibody glycosylation is determined in an antigen- and pathogen-specific manner during HIV infection. Moreover, while dramatic differences exist in bulk IgG glycosylation among individuals in distinct geographical locations, immunization is able to overcome these differences and elicit antigen-specific antibodies with similar antibody glycosylation patterns. Additionally, distinct vaccine regimens induced different antigen-specific IgG glycosylation profiles, suggesting that antibody glycosylation is not only programmable but can be manipulated via the delivery of distinct inflammatory signals during B cell priming. These data strongly suggest that the immune system naturally drives antibody glycosylation in an antigen-specific manner and highlights a promising means by which next-generation therapeutics and vaccines can harness the antiviral activity of the innate immune system via directed alterations in antibody glycosylation in vivo. .

Assessment of the Safety and Immunogenicity of 2 Novel Vaccine Platforms for HIV-1 Prevention: A Randomized Trial.

BACKGROUND: A prophylactic HIV-1 vaccine is a global health priority. OBJECTIVE: To assess a novel vaccine platform as a prophylactic HIV-1 regimen. DESIGN: Randomized, double-blind, placebo-controlled trial. Both participants and study personnel were blinded to treatment allocation. (ClinicalTrials.gov: NCT01215149). SETTING: United States, East Africa, and South Africa. PATIENTS: Healthy adults without HIV infection. INTERVENTION: 2 HIV-1 vaccines (adenovirus serotype 26 with an HIV-1 envelope A insert [Ad26.EnvA] and adenovirus serotype 35 with an HIV-1 envelope A insert [Ad35.Env], both administered at a dose of 5 × 1010 viral particles) in homologous and heterologous combinations. MEASUREMENTS: Safety and immunogenicity and the effect of baseline vector immunity. RESULTS: 217 participants received at least 1 vaccination, and 210 (>96%) completed follow-up. No vaccine-associated serious adverse events occurred. All regimens were generally well-tolerated. All regimens elicited humoral and cellular immune responses in nearly all participants. Preexisting Ad26- or Ad35-neutralizing antibody titers had no effect on vaccine safety and little effect on immunogenicity. In both homologous and heterologous regimens, the second vaccination significantly increased EnvA antibody titers (approximately 20-fold from the median enzyme-linked immunosorbent assay titers of 30-300 to 3000). The heterologous regimen of Ad26-Ad35 elicited significantly higher EnvA antibody titers than Ad35-Ad26. T-cell responses were modest and lower in East Africa than in South Africa and the United States. LIMITATIONS: Because the 2 envelope inserts were not identical, the boosting responses were complex to interpret. Durability of the immune responses elicited beyond 1 year is unknown. CONCLUSION: Both vaccines elicited significant immune responses in all populations. Baseline vector immunity did not significantly affect responses. Second vaccinations in all regimens significantly boosted EnvA antibody titers, although vaccine order in the heterologous regimen had a modest effect on the immune response. PRIMARY FUNDING SOURCE: International AIDS Vaccine Initiative, National Institutes of Health, Ragon Institute, Crucell Holland.

A Phase I Double Blind, Placebo-Controlled, Randomized Study of the Safety and Immunogenicity of an Adjuvanted HIV-1 Gag-Pol-Nef Fusion Protein and Adenovirus 35 Gag-RT-Int-Nef Vaccine in Healthy HIV-Uninfected African Adults.

BACKGROUND: Sequential prime-boost or co-administration of HIV vaccine candidates based on an adjuvanted clade B p24, RT, Nef, p17 fusion protein (F4/AS01) plus a non-replicating adenovirus 35 expressing clade A Gag, RT, Int and Nef (Ad35-GRIN) may lead to a unique immune profile, inducing both strong T-cell and antibody responses. METHODS: In a phase 1, double-blind, placebo-controlled trial, 146 healthy adult volunteers were randomized to one of four regimens: heterologous prime-boost with two doses of F4/AS01E or F4/AS01B followed by Ad35-GRIN; Ad35-GRIN followed by two doses of F4/AS01B; or three co-administrations of Ad35-GRIN and F4/AS01B. T cell and antibody responses were measured. RESULTS: The vaccines were generally well-tolerated, and did not cause serious adverse events. The response rate, by IFN-γ ELISPOT, was greater when Ad35-GRIN was the priming vaccine and in the co-administration groups. F4/AS01 induced CD4+ T-cells expressing primarily CD40L and IL2 +/- TNF-α, while Ad35-GRIN induced predominantly CD8+ T-cells expressing IFN-γ +/- IL2 or TNF-α. Viral inhibition was induced after Ad35-GRIN vaccination, regardless of the regimen. Strong F4-specific antibody responses were induced. Immune responses persisted at least a year after the last vaccination. The complementary response profiles, characteristic of each vaccine, were both expressed after co-administration. CONCLUSION: Co-administration of an adjuvanted protein and an adenovirus vector showed an acceptable safety and reactogenicity profile and resulted in strong, multifunctional and complementary HIV-specific immune responses. TRIAL REGISTRATION: ClinicalTrials.gov NCT01264445.

Acceptability and feasibility of repeated mucosal specimen collection in clinical trial participants in Kenya.

BACKGROUND: Mucosal specimens are essential to evaluate compartmentalized immune responses to HIV vaccine candidates and other mucosally targeted investigational products. We studied the acceptability and feasibility of repeated mucosal sampling in East African clinical trial participants at low risk of HIV and other sexually transmitted infections. METHODS AND FINDINGS: The Kenya AIDS Vaccine Initiative (KAVI) enrolled participants into three Phase 1 trials of preventive HIV candidate vaccines in 2011-2012 at two clinical research centers in Nairobi. After informed consent to a mucosal sub-study, participants were asked to undergo collection of mucosal secretions (saliva, oral fluids, semen, cervico-vaginal and rectal), but could opt out of any collection at any visit. Specimens were collected at baseline and two additional time points. A tolerability questionnaire was administered at the final sub-study visit. Of 105 trial participants, 27 of 34 women (79%) and 62 of 71 men (87%) enrolled in the mucosal sub-study. Nearly all sub-study participants gave saliva and oral fluids at all visits. Semen was collected from about half the participating men (47-48%) at all visits. Cervico-vaginal secretions were collected by Softcup from about two thirds of women (63%) at baseline, increasing to 78% at the following visits, with similar numbers for cervical secretion collection by Merocel sponge; about half of women (52%) gave cervico-vaginal samples at all visits. Rectal secretions were collected with Merocel sponge from about a quarter of both men and women (24%) at all 3 visits, with 16% of men and 19% of women giving rectal samples at all visits. CONCLUSIONS: Repeated mucosal sampling in clinical trial participants in Kenya is feasible, with a good proportion of participants consenting to most sampling methods with the exception of rectal samples. Experienced staff members of both sexes and trained counselors with standardized messaging may improve acceptance of rectal sampling.

Safety and immunogenicity of DNA prime and modified vaccinia ankara virus-HIV subtype C vaccine boost in healthy adults.

A randomized, double-blind, placebo-controlled phase I trial was conducted in 32 HIV-uninfected healthy volunteers to assess the safety and immunogenicity of 3 doses of DNA vaccine (Advax) plus 1 dose of recombinant modified vaccinia virus Ankara (MVA) (TBC-M4) or 3 doses of TBC-M4 alone (groups A and B, respectively). Both vaccine regimens were found to be safe and well tolerated. Gamma interferon (IFN-γ) enzyme-linked immunosorbent spot (ELISPOT) assay responses were detected in 1/10 (10%) individuals in group A after three Advax primes and in 9/9 individuals (100%) after the MVA boost. In group B, IFN-γ ELISPOT responses were detected in 6/12 (50%) and 7/11 (64%) individuals after the second and third MVA vaccinations, respectively. Responses to all vaccine components, but predominantly to Env, were seen. The breadth and magnitude of the T cell response and viral inhibition were greater in group A than in group B, indicating that the quality of the T-cell response was enhanced by the DNA prime. Intracellular cytokine staining indicated that the T-cell responses were polyfunctional but were skewed toward Env with a CD4(+) phenotype. At 2 weeks after the last vaccination, HIV-specific antibody responses were detected in all (100%) group B and 1/11 (9.1%) group A vaccinees. Vaccinia virus-specific responses were detected in all (100%) group B and 2/11 (18.2%) group A vaccinees. In conclusion, HIV-specific T-cell responses were seen in the majority of volunteers in groups A and B but with a trend toward greater quality of the T-cell response in group A. Antibody responses were better in group B than in group A.

Human immunodeficiency virus type 1 elite neutralizers: individuals with broad and potent neutralizing activity identified by using a high-throughput neutralization assay together with an analytical selection algorithm.

The development of a rapid and efficient system to identify human immunodeficiency virus type 1 (HIV-1)-infected individuals with broad and potent HIV-1-specific neutralizing antibody responses is an important step toward the discovery of critical neutralization targets for rational AIDS vaccine design. In this study, samples from HIV-1-infected volunteers from diverse epidemiological regions were screened for neutralization responses using pseudovirus panels composed of clades A, B, C, and D and circulating recombinant forms (CRFs). Initially, 463 serum and plasma samples from Australia, Rwanda, Uganda, the United Kingdom, and Zambia were screened to explore neutralization patterns and selection ranking algorithms. Samples were identified that neutralized representative isolates from at least four clade/CRF groups with titers above prespecified thresholds and ranked based on a weighted average of their log-transformed neutralization titers. Linear regression methods selected a five-pseudovirus subset, representing clades A, B, and C and one CRF01_AE, that could identify top-ranking samples with 50% inhibitory concentration (IC(50)) neutralization titers of >or=100 to multiple isolates within at least four clade groups. This reduced panel was then used to screen 1,234 new samples from the Ivory Coast, Kenya, South Africa, Thailand, and the United States, and 1% were identified as elite neutralizers. Elite activity is defined as the ability to neutralize, on average, more than one pseudovirus at an IC(50) titer of 300 within a clade group and across at least four clade groups. These elite neutralizers provide promising starting material for the isolation of broadly neutralizing monoclonal antibodies to assist in HIV-1 vaccine design.

Immunogenicity and safety of measles-mumps-rubella, varicella and Haemophilus influenzae type b vaccines administered concurrently with a fourth dose of heptavalent pneumococcal conjugate vaccine compared with the vaccines administered without heptavalent pneumococcal conjugate vaccine.

BACKGROUND: Prevnar [heptavalent pneumococcal conjugate vaccine (PCV7)] is licensed in the United States for routine administration in infants and may be coadministered with other infant vaccines. Safety and immunogenicity data on the coadministration of the fourth dose of PCV7 with measles-mumps-rubella (MMR), varicella and Haemophilus influenzae type b (Hib) vaccines are limited. METHODS: Children 12-15 months of age received either MMR with PCV7 (group 1) or MMR without PCV7 (group 2). All subjects received Hib and varicella vaccines. Group 2 received PCV7 6-9 weeks after MMR vaccination. Sera for analysis of all non-PCV7 antibodies were collected just before administration of MMR vaccine and 6 weeks later. Optimal antigen responses were assessed with the use of predetermined antibody titers. The primary end point was >90% response rate (all antigens). Noninferiority was defined as <10% difference between groups. Local and systemic reactions and postvaccination adverse events were monitored and compared between groups. RESULTS: A total of 694 subjects (347 per group) were enrolled. After immunization with MMR plus PCV7 concurrently, or MMR followed 6 weeks later by PCV7, the percentages of subjects seroconverting were significantly greater than 90% for all antigens. The difference between the 2 groups was significantly less than 10%. CONCLUSION: The immune response to MMR, Hib and varicella vaccines, when administered concurrently with a 4th (booster) dose of PCV7, was noninferior to that of these vaccines when given without PCV7. These results support concomitant administration of PCV7 with MMR, varicella and Hib as part of the recommended immunization schedule for children 12-15 months of age.

Pneumococcal conjugate vaccines: proceedings from an interactive symposium at the 41st Interscience Conference on Antimicrobial Agents and Chemotherapy.

Globally, Streptococcus pneumoniae is a leading cause of invasive and noninvasive disease in infants and young children. The emergence of antibiotic-resistant strains has increased interest in prevention through immunization. Currently, the only available conjugate pneumococcal vaccine is a seven-valent formulation, PNCRM7. This paper presents excerpts from a symposium that provided an update of ongoing surveillance data and clinical trials evaluating pneumococcal conjugate vaccines. The topics addressed included: (1) PNCRM7 postmarketing safety data; (2) the impact of PNCRM7 in premature infants; (3) the direct and indirect effect of pneumococcal conjugate vaccines on colonization; (4) the effect of pneumococcal conjugate vaccines on replacement disease and the rate of resistance among replacement serotypes; (5) the current recommendations for the use of PNCRM7; and (6) the potential impact of conjugate vaccines in Europe and the Asia-Pacific region.