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An abstract of the research paper "Antibody responses to V2 loop are induced by CRF01_A E and not Clade B envelopes," by N. Karasawas and colleagues is presented.

An important approach to stopping the AIDS epidemic is the development of a vaccine that elicits antibodies that block virus capture, the initial interactions of HIV-1 with the target cells, and replication. We utilized a previously developed qRT-PCR-based assay to examine the effects of broadly neutralizing antibodies (bNAbs), plasma from vaccine trials, and monoclonal antibodies (mAbs) on virus capture and replication. A panel of bNAbs inhibited primary HIV-1 replication in PBMCs but not virus capture. Plasma from RV144 and RV305 trial vaccinees demonstrated inhibition of virus capture with the HIV-1 subtype prevalent in Thailand. Several RV305 derived V2-specific mAbs inhibited virus replication. One of these RV305 derived V2-specific mAbs inhibited both virus capture and replication, demonstrating that it is possible to elicit antibodies by vaccination that inhibit virus capture and replication. Induction of a combination of such antibodies may be the key to protection from HIV-1 acquisition., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Inc.)

The modestly efficacious HIV-1 vaccine regimen (RV144) conferred 31% vaccine efficacy at 3 years following the four-shot immunization series, coupled with rapid waning of putative immune correlates of decreased infection risk. New strategies to increase magnitude and durability of protective immunity are critically needed. The RV305 HIV-1 clinical trial evaluated the immunological impact of a follow-up boost of HIV-1-uninfected RV144 recipients after 6-8 years with RV144 immunogens (ALVAC-HIV alone, AIDSVAX B/E gp120 alone, or ALVAC-HIV + AIDSVAX B/E gp120). Previous reports demonstrated that this regimen elicited higher binding, antibody Fc function, and cellular responses than the primary RV144 regimen. However, the impact of the canarypox viral vector in driving antibody specificity, breadth, durability and function is unknown. We performed a follow-up analysis of humoral responses elicited in RV305 to determine the impact of the different booster immunogens on HIV-1 epitope specificity, antibody subclass, isotype, and Fc effector functions. Importantly, we observed that the ALVAC vaccine component directly contributed to improved breadth, function, and durability of vaccine-elicited antibody responses. Extended boosts in RV305 increased circulating antibody concentration and coverage of heterologous HIV-1 strains by V1V2-specific antibodies above estimated protective levels observed in RV144. Antibody Fc effector functions, specifically antibody-dependent cellular cytotoxicity and phagocytosis, were boosted to higher levels than was achieved in RV144. V1V2 Env IgG3, a correlate of lower HIV-1 risk, was not increased; plasma Env IgA (specifically IgA1), a correlate of increased HIV-1 risk, was elevated. The quality of the circulating polyclonal antibody response changed with each booster immunization. Remarkably, the ALVAC-HIV booster immunogen induced antibody responses post-second boost, indicating that the viral vector immunogen can be utilized to selectively enhance immune correlates of decreased HIV-1 risk. These results reveal a complex dynamic of HIV-1 immunity post-vaccination that may require careful balancing to achieve protective immunity in the vaccinated population. Trial registration: RV305 clinical trial (ClinicalTrials.gov number, NCT01435135). ClinicalTrials.gov Identifier: NCT00223080., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Objective: The aim of this study was to determine the prevalence of high-risk (HR) and vaccine-type human papillomavirus (HPV) infection among Thai schoolgirls who were not included in the national HPV immunization program., Methods: Cross-sectional surveys were conducted among grade 10 (15-16 years old) and grade 12 (17-18 years old) schoolgirls in two provinces of Thailand. Urine samples were collected using the Colli-Pee Ⓡ device from November 2018 to February 2019. The samples were initially tested using Cobas Ⓡ 4800. Subsequently, all Cobas-positive samples and 1:1 matched Cobas-negative samples were tested by Anyplex Ⓡ assay. Prevalences of any HPV, any HR HPV, vaccine-type HPV, and individual HR HPV types were estimated by school grade., Results: Prevalences of any HPV and any HR HPV were 11.6% and 8.6% for grade 10, and 18.5% and 12.4% for grade 12 schoolgirls, respectively. Prevalences of bivalent vaccine-type HPV infection in grades 10 and 12 were 3.4% and 4.5%, respectively. Prevalences of quadrivalent and nonavalent vaccine-type HPV infections were 4.0%/6.6% and 6.4%/10.4% in grades 10 and 12, respectively. HPV16 was the most common type detected, followed by HPV58, 51, and 52. Circulating HR HPV types were similar between the school grades., Conclusion: A substantial burden of HR HPV infections was found among unvaccinated high school girls in Thailand., Competing Interests: The authors declare that they have no competing interests., (© 2023 The Authors. Published by Elsevier Ltd on behalf of International Society for Infectious Diseases.)

Immunogens and vaccination regimens can influence patterns of immune-epitope recognition, steering them towards or away from epitopes of potential viral vulnerability. HIV-1 envelope (Env)-specific antibodies targeting variable region 2 (V2) or 3 (V3) correlated with protection during the RV144 trial, however, it was suggested that the immunodominant V3 region might divert antibody responses away from other relevant sites. We mapped IgG responses against linear Env epitopes in five clinical HIV vaccine trials, revealing a specific pattern of Env targeting for each regimen. Notable V2 responses were only induced in trials administering CRF01_AE based immunogens, but targeting of V3 was seen in all trials, with the soluble, trimeric CN54gp140 protein eliciting robust V3 recognition. Strong V3 targeting was linked to greater overall response, increased number of total recognised antigenic regions, and where present, stronger V2 recognition. Hence, strong induction of V3-specific antibodies did not negatively impact the targeting of other linear epitopes in this study, suggesting that the induction of antibodies against V3 and other regions of potential viral vulnerability need not be necessarily mutually exclusive., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Horvath, Rogers, Pollakis, Baranov, Pieroth, Joseph, Chachage, Heitzer, Maganga, Msafiri, Joachim, Viegas, Eller, Kibuuka, Rerks-Ngarm, Pitisuttithum, Nitayapan, Dhitavat, Premsri, Fidler, Shattock, Robb, Weber, McCormack, Munseri, Lyamuya, Nilsson, Kroidl, Hoelscher, Wagner, Geldmacher and Held.)

Human papillomavirus (HPV) is a common infection principally spread through sexual activity. Most HPV infections are asymptomatic and resolve spontaneously. However, persistent infection may progress to cervical cancer. Highly efficacious HPV vaccines have been available since 2006, yet uptake into national programs has been slow in part due to cost. WHO guidelines call for a two-dose (0,6 month) schedule for girls 9-14 years of age. Post-hoc analyses of randomized trials have found high vaccine effectiveness following a single dose of vaccine. In order to provide additional data on the potential impact of single dose HPV vaccination in a real-world setting, we are conducting an effectiveness study among Thai schoolgirls. This is an observational study of a single dose (SD) or two doses (2D) of the bivalent HPV vaccine CERVARIX® (GlaxoSmithKline plc.) administered in a school-based program to 8-9,000 Grade 8 female students in two provinces of Thailand beginning in 2018; one province is assigned the SD, and the other the standard 2D regimen. The reduction in HPV vaccine-type prevalence will be assessed in each province two and four years after vaccination by comparing HPV prevalence in urine samples obtained through cross-sectional surveys of the immunized grade cohort as they age and compared to a historical "baseline" HPV prevalence of same age students., Competing Interests: The authors have declared that no competing interests exist.

Fc-mediated immune functions have been correlated with protection in the RV144 HIV vaccine trial and are important for immunity to a range of pathogens. IgG antibodies (Abs) that form complexes with Fc receptors (FcRs) on innate immune cells can activate Fc-mediated immune functions. Genetic variation in both IgGs and FcRs have the capacity to alter IgG-FcR complex formation via changes in binding affinity and concentration. A growing challenge lies in unraveling the importance of multiple variations, especially in the context of vaccine trials that are conducted in homogenous genetic populations. Here we use an ordinary differential equation model to quantitatively assess how IgG1 allotypes and FcγR polymorphisms influence IgG-FcγRIIIa complex formation in vaccine-relevant settings. Using data from the RV144 HIV vaccine trial, we map the landscape of IgG-FcγRIIIa complex formation predicted post-vaccination for three different IgG1 allotypes and two different FcγRIIIa polymorphisms. Overall, the model illustrates how specific vaccine interventions could be applied to maximize IgG-FcγRIIIa complex formation in different genetic backgrounds. Individuals with the G1m1,17 and G1m1,3 allotypes were predicted to be more responsive to vaccine adjuvant strategies that increase antibody FcγRIIIa affinity (e.g. glycosylation modifications), compared to the G1m-1,3 allotype which was predicted to be more responsive to vaccine boosting regimens that increase IgG1 antibody titers (concentration). Finally, simulations in mixed-allotype populations suggest that the benefit of boosting IgG1 concentration versus IgG1 affinity may be dependent upon the presence of the G1m-1,3 allotype. Overall this work provides a quantitative tool for rationally improving Fc-mediated functions after vaccination that may be important for assessing vaccine trial results in the context of under-represented genetic populations., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Lemke, Theisen, Bozich, McLean, Lee, Lopez, Rerks-Ngarm, Pitisuttithum, Nitayaphan, Kratochvil, Wines, Hogarth, Kent, Chung and Arnold.)

A gene signature was previously found to be correlated with mosaic adenovirus 26 vaccine protection in simian immunodeficiency virus and simian-human immunodeficiency virus challenge models in non-human primates. In this report, we investigated the presence of this signature as a correlate of reduced risk in human clinical trials and potential mechanisms of protection. The absence of this gene signature in the DNA/rAd5 human vaccine trial, which did not show efficacy, strengthens our hypothesis that this signature is only enriched in studies that demonstrated protection. This gene signature was enriched in the partially effective RV144 human trial that administered the ALVAC/protein vaccine, and we find that the signature associates with both decreased risk of HIV-1 acquisition and increased vaccine efficacy (VE). Total RNA-seq in a clinical trial that used the same vaccine regimen as the RV144 HIV vaccine implicated antibody-dependent cellular phagocytosis (ADCP) as a potential mechanism of vaccine protection. CITE-seq profiling of 53 surface markers and transcriptomes of 53,777 single cells from the same trial showed that genes in this signature were primarily expressed in cells belonging to the myeloid lineage, including monocytes, which are major effector cells for ADCP. The consistent association of this transcriptome signature with VE represents a tool both to identify potential mechanisms, as with ADCP here, and to screen novel approaches to accelerate the development of new vaccine candidates., Competing Interests: SS, PE, AG, LY, GK, KM, SF, KN, LW, SS, EL, PP, SN, SC, SR, MR, DD, PG, GT, NM, SV, RT No competing interests declared

Immunoglobulin G (IgG) antibodies that activate Fc-mediated immune functions have been correlated with vaccine efficacy, but it is difficult to unravel the relative roles of multiple IgG and Fc receptor (FcR) features that have the capacity to influence IgG-FcR complex formation but vary on a personalized basis. Here, we develop an ordinary differential-equation model to determine how personalized variability in IgG subclass concentrations and binding affinities influence IgG-FcγRIIIa complex formation and validate it with samples from the HIV RV144 vaccine trial. The model identifies individuals who are sensitive, insensitive, or negatively affected by increases in HIV-specific IgG1, which is validated with the addition of HIV-specific IgG1 monoclonal antibodies to vaccine samples. IgG1 affinity to FcγRIIIa is also prioritized as the most influential parameter for dictating activation broadly across a population. Overall, this work presents a quantitative tool for evaluating personalized differences underlying FcR activation, which is relevant to ongoing efforts to improve vaccine efficacy., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

Identifying whether viral features present in acute HIV-1 infection predetermine the development of neutralization breadth is critical to vaccine design. Incorporating such features in vaccine antigens could initiate cross-reactive antibody responses that could sufficiently protect vaccinees from HIV-1 infection despite the uniqueness of each founder virus. To understand the relationship between Env determinants and the development of neutralization breadth, we focused on 197 individuals enrolled in two cohorts in Thailand and East Africa (RV144 and RV217) and followed since their diagnosis in acute or early HIV-1 infection. We analyzed the distribution of variable loop lengths and glycans, as well as the predicted density of the glycan shield, and compared these envelope features to the neutralization breadth data obtained 3 years after infection ( n   = 121). Our study revealed limited evidence for glycan shield features that associate with the development of neutralization breadth. While the glycan shield tended to be denser in participants who subsequently developed breadth, no significant relationship was found between the size of glycan holes and the development of neutralization breadth. The parallel analysis of 3,000 independent Env sequences showed no evidence of directional evolution of glycan shield features since the beginning of the epidemic. Together, our results highlight that glycan shield features in acute and early HIV-1 infection may not play a role determinant enough to dictate the development of neutralization breadth and instead suggest that the glycan shield's reactive properties that are associated with immune evasion may have a greater impact. IMPORTANCE A major goal of HIV-1 vaccine research is to design vaccine candidates that elicit potent broadly neutralizing antibodies (bNAbs). Different viral features have been associated with the development of bNAbs, including the glycan shield on the surface of the HIV-1 Envelope (Env). Here, we analyzed data from two cohorts of individuals who were followed from early infection to several years after infection spanning multiple HIV-1 subtypes. We compared Env glycan features in HIV-1 sequences obtained in early infection to the potency and breadth of neutralizing antibodies measured 1 to 3 years after infection. We found limited evidence of glycan shield properties that associate with the development of neutralization breadth in these cohorts. These results may have important implications for antigen design in future vaccine strategies and emphasize that HIV-1 vaccines will need to rely on a complex set of properties to elicit neutralization breadth.

The scale of the HIV-1 epidemic underscores the need for a vaccine. The multitude of circulating HIV-1 strains together with HIV-1's high evolvability hints that HIV-1 could adapt to a future vaccine. Here, we wanted to investigate the effect of vaccination on the evolution of the virus post-breakthrough infection. We analyzed 2,635 HIV-1 env sequences sampled up to a year post-diagnosis from 110 vaccine and placebo participants who became infected in the RV144 vaccine efficacy trial. We showed that the Env signature sites that were previously identified to distinguish vaccine and placebo participants were maintained over time. In addition, fewer sites were under diversifying selection in the vaccine group than in the placebo group. These results indicate that HIV-1 would possibly adapt to a vaccine upon its roll-out., (© The Author(s) 2021. Published by Oxford University Press.)

The RV144 vaccine efficacy clinical trial showed a reduction in HIV-1 infections by 31%. Vaccine efficacy was associated with stronger binding antibody responses to the HIV Envelope (Env) V1V2 region, with decreased efficacy as responses wane. High levels of Ab-dependent cellular cytotoxicity (ADCC) together with low plasma levels of Env-specific IgA also correlated with decreased infection risk. We investigated whether B cell priming from RV144 vaccination impacted functional antibody responses to HIV-1 following infection. Antibody responses were assessed in 37 vaccine and 63 placebo recipients at 6, 12, and 36 months following HIV diagnosis. The magnitude, specificity, dynamics, subclass recognition and distribution of the binding antibody response following infection were different in RV144 vaccine recipients compared to placebo recipients. Vaccine recipients demonstrated increased IgG1 binding specifically to V1V2, as well as increased IgG2 and IgG4 but decreased IgG3 to HIV-1 Env. No difference in IgA binding to HIV-1 Env was detected between the vaccine and placebo recipients following infection. RV144 vaccination limited the development of broadly neutralizing antibodies post-infection, but enhanced Fc-mediated effector functions indicating B cell priming by RV144 vaccination impacted downstream antibody function. However, these functional responses were not associated with clinical markers of disease progression. These data reveal that RV144 vaccination primed B cells towards specific binding and functional antibody responses following HIV-1 infection., Competing Interests: The authors have declared that no competing interests exist.

While the RV144 HIV vaccine trial led to moderately reduced risk of HIV acquisition, emerging data from the HVTN702 trial point to the critical need to reexamine RV144-based correlates of reduced risk of protection. While in RV144, the induction of V2-binding, non-IgA, IgG3 antibody responses with nonneutralizing functions were linked to reduced risk of infection, the interactions between these signatures remain unclear. Thus, here we comprehensively profile the humoral immune response in 300 RV144 vaccinees to decipher the relationships between humoral biomarkers of protection. We found that vaccine-specific IgG1, IgG3, and IgA were highly correlated. However, ratios of IgG1:IgG3:IgA provided insights into subclass/isotype polyclonal functional regulation. For instance, in the absence of high IgG1 levels, IgG3 antibodies exhibited limited functional activity, pointing to IgG3 as a critical contributor, but not sole driver, of effective antiviral humoral immunity. Higher IgA levels were linked to enhanced antibody effector function, including neutrophil phagocytosis (ADNP), complement deposition (ADCD), and antibody-dependent NK degranulation (CD107a), some of which were increased in infected vaccinees in a case/control data set, suggesting that IgA-driven functions compromised immunity. These data highlight the interplay between IgG1, IgG3, and IgA, pointing to the need to profile the relationships between subclass/isotype selection.

The RV144 HIV-1 vaccine trial results showed moderate reduction in viral infections among vaccinees as well as induction of antibody-dependent cellular cytotoxicity and vaccine-specific IgG and IgG3 responses directed at variable loop regions 1 and 2 of the HIV envelope protein. However, with the recent failure of the HVTN 702 clinical trial, comprehensive profiling of humoral immune responses may provide insight for these disappointing results. One of the changes included in the HVTN 702 study was the addition of a late boost, aimed at augmenting peak immunity and durability. The companion vaccine trial RV305 was designed to permit the evaluation of the immunologic impact of late boosting with either the boosting protein antigen alone, the canarypox viral vector ALVAC alone, or a combination of both. Although previous data showed elevated levels of IgG antibodies in both boosting arms, regardless of ALVAC-HIV vector incorporation, the effect on shaping antibody effector function remains unclear. Thus, here we analyzed the antibody and functional profile induced by RV305 boosting regimens and found that although IgG1 levels increased in both arms that included protein boosting, IgG3 levels were reduced compared with the original RV144 vaccine strategy. Most functional responses increased upon protein boosting, regardless of the viral vector-priming agent incorporation. These data suggest that the addition of a late protein boost alone is sufficient to increase functionally potent vaccine-specific antibodies previously associated with reduced risk of infection with HIV.

Induction of protective antibodies is a critical goal of HIV-1 vaccine development. One strategy is to induce nonneutralizing antibodies (NNAbs) that kill virus-infected cells, as these antibody specificities have been implicated in slowing HIV-1 disease progression and in protection. HIV-1 Env constant region 1 and 2 (C1C2) monoclonal antibodies (MAbs) frequently mediate potent antibody-dependent cellular cytotoxicity (ADCC), making them an important vaccine target. Here, we explore the effect of delayed and repetitive boosting of RV144 vaccine recipients with AIDSVAX B/E on the C1C2-specific MAb repertoire. It was found that boosting increased clonal lineage-specific ADCC breadth and potency. A ligand crystal structure of a vaccine-induced broad and potent ADCC-mediating C1C2-specific MAb showed that it bound a highly conserved Env gp120 epitope. Thus, boosting to affinity mature these types of IgG C1C2-specific antibody responses may be one method by which to make an improved HIV vaccine with higher efficacy than that seen in the RV144 trial. IMPORTANCE Over one million people become infected with HIV-1 each year, making the development of an efficacious HIV-1 vaccine an important unmet medical need. The RV144 human HIV-1 vaccine regimen is the only HIV-1 clinical trial to date to demonstrate vaccine efficacy. An area of focus has been on identifying ways by which to improve upon RV144 vaccine efficacy. The RV305 HIV-1 vaccine regimen was a follow-up boost of RV144 vaccine recipients that occurred 6 to 8 years after the conclusion of RV144. Our study focused on the effect of delayed boosting in humans on the vaccine-induced Env constant region 1 and 2 (C1C2)-specific antibody repertoire. It was found that boosting with an HIV-1 Env vaccine increased C1C2-specific antibody-dependent cellular cytotoxicity potency and breadth., (Copyright © 2020 Easterhoff et al.)

Background: HIV vaccine trials routinely measure multiple vaccine-elicited immune responses to compare regimens and study their potential associations with protection. Here we employ unsupervised learning tools facilitated by a bidirectional power transformation to explore the multivariate binding antibody and T-cell response patterns of immune responses elicited by two pox-protein HIV vaccine regimens. Both regimens utilized a recombinant canarypox vector (ALVAC-HIV) prime and a bivalent recombinant HIV-1 Envelope glycoprotein 120 subunit boost. We hypothesized that within each trial, there were participant subgroups sharing similar immune responses and that their frequencies differed across trials., Methods and Findings: We analyzed data from three trials-RV144 (NCT00223080), HVTN 097 (NCT02109354), and HVTN 100 (NCT02404311), the latter of which was pivotal in advancing the tested pox-protein HIV vaccine regimen to the HVTN 702 Phase 2b/3 efficacy trial. We found that bivariate CD4+ T-cell and anti-V1V2 IgG/IgG3 antibody response patterns were similar by age, sex-at-birth, and body mass index, but differed for the pox-protein clade AE/B alum-adjuvanted regimen studied in RV144 and HVTN 097 (PAE/B/alum) compared to the pox-protein clade C/C MF59-adjuvanted regimen studied in HVTN 100 (PC/MF59). Specifically, more PAE/B/alum recipients had low CD4+ T-cell and high anti-V1V2 IgG/IgG3 responses, and more PC/MF59 recipients had broad responses of both types. Analyses limited to "vaccine-matched" antigens suggested that some of the differences in responses between the regimens could have been due to antigens in the assays that did not match the vaccine immunogens. Our approach was also useful in identifying subgroups with unusually absent or high co-responses across assay types, flagging individuals for further characterization by functional assays. We also found that co-responses of anti-V1V2 IgG/IgG3 and CD4+ T cells had broad variability. As additional immune response assays are standardized and validated, we anticipate our framework will be increasingly valuable for multivariate analysis., Conclusions: Our approach can be used to advance vaccine development objectives, including the characterization and comparison of candidate vaccine multivariate immune responses and improved design of studies to identify correlates of protection. For instance, results suggested that HVTN 702 will have adequate power to interrogate immune correlates involving anti-V1V2 IgG/IgG3 and CD4+ T-cell co-readouts, but will have lower power to study anti-gp120/gp140 IgG/IgG3 due to their lower dynamic ranges. The findings also generate hypotheses for future testing in experimental and computational analyses aimed at achieving a mechanistic understanding of vaccine-elicited immune response heterogeneity., Competing Interests: The authors of this manuscript have read the journal's policy and have the following competing interests: LC, ZM, YF, NG, GEG, HEJ, MJM, NR, SCR, NF, GDT, and PBG are recipients of funding from the National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health (NIH), and this publication is a result of activities funded by the NIAID. PBG and GDT are recipients of funding from the Bill and Melinda Gates Foundation, and this publication is a result of activities funded by the BMGF. SR-N is an employee of the Department of Disease Control in the Ministry of Public Heath (Thailand), and this publication is a result of activities conducted through facilities of the Thailand MPH. NLM was an employee of the U.S. Army and MLR is an employee of the Henry Jackson Foundation components of the Military HIV Research Program (MHRP)/Walter Reed Army Institute of Research, which is supported by the US Department of Defense, and this publication is a result of activities partially funded by the DOD. HEJ is a recipient of funding from the National Cancer Institute of the NIH. YH is a recipient of funding from the National Institute of General Medical Sciences of the NIH. AFG is the recipient of funding from the Infectious Disease Research Institute. LC has received grants from Gilead, Sanofi Pasteur Biologics and Immune Design Corp. PBG is the recipient of a contract from Sanofi Pasteur and has served as an unpaid consultant at advisory meetings to Sanofi Pasteur. LNC, ZM, YH, YF, and PBG have received salary support from the contract with Sanofi Pasteur and ZM, YH, YF, and PBG have received support for travel to meetings from Sanofi Pasteur. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

In the RV144 HIV-1 phase III trial, vaccine efficacy directly correlated with the magnitude of the variable region 2-specific (V2-specific) IgG antibody response, and in the presence of low plasma IgA levels, with the magnitude of plasma antibody-dependent cellular cytotoxicity. Reenrollment of RV144 vaccinees in the RV305 trial offered the opportunity to define the function, maturation, and persistence of vaccine-induced V2-specific and other mAb responses after boosting. We show that the RV144 vaccine regimen induced persistent V2 and other HIV-1 envelope-specific memory B cell clonal lineages that could be identified throughout the approximately 11-year vaccination period. Subsequent boosts increased somatic hypermutation, a critical requirement for antibody affinity maturation. Characterization of 22 vaccine-induced V2-specific mAbs with epitope specificities distinct from previously characterized RV144 V2-specific mAbs CH58 and CH59 found increased in vitro antibody-mediated effector functions. Thus, when inducing non-neutralizing antibodies, one method by which to improve HIV-1 vaccine efficacy may be through late boosting to diversify the V2-specific response to increase the breadth of antibody-mediated anti-HIV-1 effector functions.

To date, six vaccine strategies have been evaluated in clinical trials for their efficacy at inducing protective immune responses against HIV infection. However, only the ALVAC-HIV/AIDSVAX B/E vaccine (RV144 trial) has demonstrated protection, albeit modestly (31%; P = 0.03). One potential correlate of protection was a low-frequency HIV-specific CD4 T cell population with diverse functionality. Although CD4 T cells, particularly T follicular helper (Tfh) cells, are critical for effective antibody responses, most studies involving HIV vaccines have focused on humoral immunity or CD8 T cell effector responses, and little is known about the functionality and frequency of vaccine-induced CD4 T cells. We therefore assessed responses from several phase I/II clinical trials and compared them to responses to natural HIV-1 infection. We found that all vaccines induced a lower magnitude of HIV-specific CD4 T cell responses than that observed for chronic infection. Responses differed in functionality, with a CD40 ligand (CD40L)-dominated response and more Tfh cells after vaccination, whereas chronic HIV infection provoked tumor necrosis factor alpha (TNF-α)-dominated responses. The vaccine delivery route further impacted CD4 T cells, showing a stronger Th1 polarization after dendritic cell delivery than after intramuscular vaccination. In prime/boost regimens, the choice of prime and boost influenced the functional profile of CD4 T cells to induce more or less polyfunctionality. In summary, vaccine-induced CD4 T cell responses differ remarkably between vaccination strategies, modes of delivery, and boosts and do not resemble those induced by chronic HIV infection. Understanding the functional profiles of CD4 T cells that best facilitate protective antibody responses will be critical if CD4 T cell responses are to be considered a clinical trial go/no-go criterion. IMPORTANCE Only one HIV-1 candidate vaccine strategy has shown protection, albeit marginally (31%), against HIV-1 acquisition, and correlates of protection suggested that a multifunctional CD4 T cell immune response may be important for this protective effect. Therefore, the functional phenotypes of HIV-specific CD4 T cell responses induced by different phase I and phase II clinical trials were assessed to better show how different vaccine strategies influence the phenotype and function of HIV-specific CD4 T cell immune responses. The significance of this research lies in our comprehensive comparison of the compositions of the T cell immune responses to different HIV vaccine modalities. Specifically, our work allows for the evaluation of vaccination strategies in terms of their success at inducing Tfh cell populations., (Copyright © 2018 American Society for Microbiology.)

The phase III RV144 human immunodeficiency virus (HIV) vaccine trial conducted in Thailand remains the only study to show efficacy in decreasing the HIV acquisition risk. In Thailand, circulating recombinant forms of HIV clade A/E (CRF01_AE) predominate; in such viruses, env originates from clade E (HIV-E). We constructed a simian-human immunodeficiency virus (SHIV) chimera carrying env isolated from an RV144 placebo recipient in the SHIV-1157ipd3N4 backbone. The latter contains long terminal repeats (LTRs) with duplicated NF-κB sites, thus resembling HIV LTRs. We devised a novel strategy to adapt the parental infectious molecular clone (IMC), R5 SHIV-E1, to rhesus macaques: the simultaneous depletion of B and CD8 + cells followed by the intramuscular inoculation of proviral DNA and repeated administrations of cell-free virus. High-level viremia and CD4 + T-cell depletion ensued. Passage 3 virus unexpectedly caused acute, irreversible CD4 + T-cell loss; the partially adapted SHIV had become dual tropic. Virus and IMCs with exclusive R5 tropism were reisolated from earlier passages, combined, and used to complete adaptation through additional macaques. The final isolate, SHIV-E1p5, remained solely R5 tropic. It had a tier 2 neutralization phenotype, was mucosally transmissible, and was pathogenic. Deep sequencing revealed 99% Env amino acid sequence conservation; X4-only and dual-tropic strains had evolved independently from an early branch of parental SHIV-E1. To conclude, our primate model data reveal that SHIV-E1p5 recapitulates important aspects of HIV transmission and pathobiology in humans. IMPORTANCE Understanding the protective principles that lead to a safe, effective vaccine against HIV in nonhuman primate (NHP) models requires test viruses that allow the evaluation of anti-HIV envelope responses. Reduced HIV acquisition risk in RV144 has been linked to nonneutralizing IgG antibodies with a range of effector activities. Definitive experiments to decipher the mechanisms of the partial protection observed in RV144 require passive-immunization studies in NHPs with a relevant test virus. We have generated such a virus by inserting env from an RV144 placebo recipient into a SHIV backbone with HIV-like LTRs. The final SHIV-E1p5 isolate, grown in rhesus monkey peripheral blood mononuclear cells, was mucosally transmissible and pathogenic. Earlier SHIV-E passages showed a coreceptor switch, again mimicking HIV biology in humans. Thus, our series of SHIV-E strains mirrors HIV transmission and disease progression in humans. SHIV-E1p5 represents a biologically relevant tool to assess prevention strategies., (Copyright © 2018 American Society for Microbiology.)

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&#95;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.

Induction of broadly cross-reactive antiviral humoral responses with the capacity to target globally diverse circulating strains is a key goal for HIV-1 immunogen design. A major gap in the field is the identification of diverse HIV-1 envelope antigens to evaluate vaccine regimens for binding antibody breadth. In this study, we define unique antigen panels to map HIV-1 vaccine-elicited antibody breadth and durability. Diverse HIV-1 envelope glycoproteins were selected based on genetic and geographic diversity to cover the global epidemic, with a focus on sexually acquired transmitted/founder viruses with a tier 2 neutralization phenotype. Unique antigenicity was determined by nonredundancy (Spearman correlation), and antigens were clustered using partitioning around medoids (PAM) to identify antigen diversity. Cross-validation demonstrated that the PAM method was better than selection by reactivity and random selection. Analysis of vaccine-elicited V1V2 binding antibody in longitudinal samples from the RV144 clinical trial revealed the striking heterogeneity among individual vaccinees in maintaining durable responses. These data support the idea that a major goal for vaccine development is to improve antibody levels, breadth, and durability at the population level. Elucidating the level and durability of vaccine-elicited binding antibody breadth needed for protection is critical for the development of a globally efficacious HIV vaccine. IMPORTANCE The path toward an efficacious HIV-1 vaccine will require characterization of vaccine-induced immunity that can recognize and target the highly genetically diverse virus envelope glycoproteins. Antibodies that target the envelope glycoproteins, including diverse sequences within the first and second hypervariable regions (V1V2) of gp120, were identified as correlates of risk for the one partially efficacious HIV-1 vaccine. To build upon this discovery, we experimentally and computationally evaluated humoral responses to define envelope glycoproteins representative of the antigenic diversity of HIV globally. These diverse envelope antigens distinguished binding antibody breadth and durability among vaccine candidates, thus providing insights for advancing the most promising HIV-1 vaccine candidates., (Copyright © 2018 Yates et al.)

Vaccine-elicited humoral immune responses comprise an array of antibody forms and specificities, with only a fraction contributing to protective host immunity. Elucidation of antibody effector functions responsible for protective immunity against human immunodeficiency virus type 1 (HIV-1) acquisition is a major goal for the HIV-1 vaccine field. Immunoglobulin A (IgA) is an important part of the host defense against pathogens; however, little is known about the role of vaccine-elicited IgA and its capacity to mediate antiviral functions. To identify the antiviral functions of HIV-1-specific IgA elicited by vaccination, we cloned HIV-1 envelope-specific IgA monoclonal antibodies (MAbs) by memory B cell cultures from peripheral blood mononuclear cells from an RV144 vaccinee and produced two IgA clonal cell lines (HG129 and HG130) producing native, nonrecombinant IgA MAbs. The HG129 and HG130 MAbs mediated phagocytosis by monocytes, and HG129 blocked HIV-1 Env glycoprotein binding to galactosylceramide, an alternative HIV-1 receptor. These findings elucidate potential antiviral functions of vaccine-elicited HIV-1 envelope-specific IgA that may act to block HIV-1 acquisition at the portal of entry by preventing HIV-1 binding to galactosylceramide and mediating antibody Fc receptor-mediated virion phagocytosis. Furthermore, these findings highlight the complex and diverse interactions of vaccine-elicited IgA with pathogens that depend on IgA fine specificity and form (e.g., multimeric or monomeric) in the systemic circulation and mucosal compartments. IMPORTANCE Host-pathogen interactions in vivo involve numerous immune mechanisms that can lead to pathogen clearance. Understanding the nature of antiviral immune mechanisms can inform the design of efficacious HIV-1 vaccine strategies. Evidence suggests that both neutralizing and nonneutralizing antibodies can mediate some protection against HIV in animal models. Although numerous studies have characterized the functional properties of HIV-1-specific IgG, more studies are needed on the functional attributes of HIV-1-specific IgA, specifically for vaccine-elicited IgA. Characterization of the functional properties of HIV-1 Env-specific IgA monoclonal antibodies from human vaccine clinical trials are critical toward understanding the capacity of the host immune response to block HIV-1 acquisition., (Copyright © 2018 Wills et al.)

The concerns raised from adenovirus 5 (Ad5)-based HIV vaccine clinical trials, where excess HIV infections were observed in some vaccine recipients, have highlighted the importance of understanding host responses to vaccine vectors and the HIV susceptibility of vector-specific CD4 T cells in HIV vaccination. Our recent study reported that human Ad5-specific CD4 T cells induced by Ad5 vaccination (RV156A trial) are susceptible to HIV. Here we further investigated the HIV susceptibility of vector-specific CD4 T cells induced by ALVAC, a canarypox viral vector tested in the Thai trial RV144, as compared to Ad5 vector-specific CD4 T cells in the HVTN204 trial. We showed that while Ad5 vector-specific CD4 T cells were readily susceptible to HIV, ALVAC-specific CD4 T cells in RV144 PBMC were substantially less susceptible to both R5 and X4 HIV in vitro. The lower HIV susceptibility of ALVAC-specific CD4 T cells was associated with the reduced surface expression of HIV entry co-receptors CCR5 and CXCR4 on these cells. Phenotypic analyses identified that ALVAC-specific CD4 T cells displayed a strong Th1 phenotype, producing higher levels of IFN-γ and CCL4 (MIP-1β) but little IL-17. Of interest, ALVAC and Ad5 vectors induced distinct profiles of vector-specific CD8 vs. CD4 T-cell proliferative responses in PBMC, with ALVAC preferentially inducing CD8 T-cell proliferation, while Ad5 vector induced CD4 T-cell proliferation. Depletion of ALVAC-, but not Ad5-, induced CD8 T cells in PBMC led to a modest increase in HIV infection of vector-specific CD4 T cells, suggesting a role of ALVAC-specific CD8 T cells in protecting ALVAC-specific CD4 T cells from HIV. Taken together, our data provide strong evidence for distinct HIV susceptibility of CD4 T cells induced by different vaccine vectors and highlight the importance of better evaluating anti-vector responses in HIV vaccination.

The RV144 HIV vaccine trial included a recombinant HIV glycoprotein 120 (gp120) construct fused to a small portion of herpes simplex virus 1 (HSV-1) glycoprotein D (gD) so that the first 40 amino acids of gp120 were replaced by the signal sequence and the first 27 amino acids of the mature form of gD. This region of gD contains most of the binding site for HVEM, an HSV receptor important for virus infection of epithelial cells and lymphocytes. RV144 induced antibodies to HIV that were partially protective against infection, as well as antibodies to HSV. We derived monoclonal antibodies (MAbs) from peripheral blood B cells of recipients of the RV144 HIV vaccine and showed that these antibodies neutralized HSV-1 infection in cells expressing HVEM, but not the other major virus receptor, nectin-1. The MAbs mediated antibody-dependent cellular cytotoxicity (ADCC), and mice that received the MAbs and were then challenged by corneal inoculation with HSV-1 had reduced eye disease, shedding, and latent infection. To our knowledge, this is the first description of MAbs derived from human recipients of a vaccine that specifically target the HVEM binding site of gD. In summary, we found that monoclonal antibodies derived from humans vaccinated with the HVEM binding domain of HSV-1 gD (i) neutralized HSV-1 infection in a cell receptor-specific manner, (ii) mediated ADCC, and (iii) reduced ocular disease in virus-infected mice. IMPORTANCE Herpes simplex virus 1 (HSV-1) causes cold sores and neonatal herpes and is a leading cause of blindness. Despite many trials, no HSV vaccine has been approved. Nectin-1 and HVEM are the two major cellular receptors for HSV. These receptors are expressed at different levels in various tissues, and the role of each receptor in HSV pathogenesis is not well understood. We derived human monoclonal antibodies from persons who received the HIV RV144 vaccine that contained the HVEM binding domain of HSV-1 gD fused to HIV gp120. These antibodies were able to specifically neutralize HSV-1 infection in vitro via HVEM. Furthermore, we showed for the first time that HVEM-specific HSV-1 neutralizing antibodies protect mice from HSV-1 eye disease, indicating the critical role of HVEM in HSV-1 ocular infection., (Copyright © 2017 American Society for Microbiology.)

Non-neutralizing IgG to the V1V2 loop of HIV-1 gp120 correlates with a decreased risk of HIV-1 infection but the mechanism of protection remains unknown. This V1V2 IgG correlate was identified in RV144 Thai trial vaccine recipients, who were primed with a canarypox vector expressing membrane-bound gp120 (vCP1521) and boosted with vCP1521 plus a mixture gp120 proteins from clade B and clade CRF01&#95;AE (B/E gp120). We sought to determine whether the mechanism of vaccine protection might involve antibody-dependent complement activation. Complement activation was measured as a function of complement component C3d deposition on V1V2-coated beads in the presence of RV144 sera. Variable levels of complement activation were detected two weeks post final boosting in RV144, which is when the V1V2 IgG correlate was identified. The magnitude of complement activation correlated with V1V2-specific serum IgG and was stronger and more common in RV144 than in HIV-1 infected individuals and two related HIV-1 vaccine trials, VAX003 and VAX004, where no protection was seen. After adjusting for gp120 IgA, V1V2 IgG, gender, and risk score, complement activation by case-control plasmas from RV144 correlated inversely with a reduced risk of HIV-1 infection, with odds ratio for positive versus negative response to TH023-V1V2 0.42 (95% CI 0.18 to 0.99, p = 0.048) and to A244-V1V2 0.49 (95% CI 0.21 to 1.10, p = 0.085). These results suggest that complement activity may have contributed in part to modest protection against the acquisition of HIV-1 infection seen in the RV144 trial.