Effect of the Communities that HEAL intervention on receipt of behavioral therapies for opioid use disorder: A cluster randomized wait-list controlled trial.

BACKGROUND: The U.S. opioid overdose crisis persists. Outpatient behavioral health services (BHS) are essential components of a comprehensive response to opioid use disorder and overdose fatalities. The Helping to End Addiction Long-Term® (HEALing) Communities Study developed the Communities That HEAL (CTH) intervention to reduce opioid overdose deaths in 67 communities in Kentucky, Ohio, New York, and Massachusetts through the implementation of evidence-based practices (EBPs), including BHS. This paper compares the rate of individuals receiving outpatient BHS in Wave 1 intervention communities (n = 34) to waitlisted Wave 2 communities (n = 33). METHODS: Medicaid data included individuals ≥18 years of age receiving any of five BHS categories: intensive outpatient, outpatient, case management, peer support, and case management or peer support. Negative binomial regression models estimated the rate of receiving each BHS for Wave 1 and Wave 2. Effect modification analyses evaluated changes in the effect of the CTH intervention between Wave 1 and Wave 2 by research site, rurality, age, sex, and race/ethnicity. RESULTS: No significant differences were detected between intervention and waitlisted communities in the rate of individuals receiving any of the five BHS categories. None of the interaction effects used to test the effect modification were significant. CONCLUSIONS: Several factors should be considered when interpreting results-no significant intervention effects were observed through Medicaid claims data, the best available data source but limited in terms of capturing individuals reached by the intervention. Also, the 12-month evaluation window may have been too brief to see improved outcomes considering the time required to stand-up BHS. TRIAL REGISTRATION: Clinical Trials.gov http://www. CLINICALTRIALS: gov: Identifier: NCT04111939.

Development and validation of a community-level social determinants of health index for drug overdose deaths in the HEALing Communities Study.

INTRODUCTION: Social determinants of health (SDoH), such as socioeconomic status, education level, and food insecurity, are believed to influence the opioid crisis. While global SDoH indices such as the CDC's Social Vulnerability Index (SVI) and Area Deprivation Index (ADI) combine the explanatory power of multiple social factors for understanding health outcomes, they may be less applicable to the specific challenges of opioid misuse and associated outcomes. This study develops a novel index tailored to opioid misuse outcomes, tests the efficacy of this index in predicting drug overdose deaths across contexts, and compares the explanatory power of this index to other SDoH indices. METHODS: Focusing on four HEALing Communities Study (HCS) states (Kentucky, Massachusetts, New York and Ohio; encompassing 4269 ZIP codes), we identified multilevel SDoH potentially associated with opioid misuse and aggregated publicly available data for each measure. We then leveraged a random forest model to develop a composite measure that predicts age-adjusted drug overdose mortality rates based on SDoH. We used this composite measure to understand HCS and non-HCS communities in terms of overdose risk across areas of varying racial composition. Finally, we compared variance in drug overdose deaths explained by this index to variance explained by the SVI and ADI. RESULTS: Our composite measure included 28 SDoH measures and explained approximately 89 % percent of variance in age-adjusted drug overdose mortality across HCS states. Health care measures, including emergency department visits and primary care provider availability, were top predictors within the index. Index accuracy was robust within and outside of HCS communities and states. This measure identified high levels of overdose mortality risk in segregated communities. CONCLUSIONS: Existing SDoH indices fail to explain much variation in area-level overdose mortality rates. Having tailored composite indices can help us to identify places in which residents are at highest risk based on their composite contexts. A comprehensive index can also help to develop effective community interventions for programs such as HCS by considering the context in which people live.

Community-level determinants of stakeholder perceptions of community stigma toward people with opioid use disorders, harm reduction services and treatment in the HEALing Communities Study.

BACKGROUND: Community stigma toward people with opioid use disorder (OUD) can impede access to harm reduction services and treatment with medications for opioid use disorder (MOUD). Such community OUD stigma is partially rooted in community-level social and economic conditions, yet there remains a paucity of large-scale quantitative data examining community-level factors associated with OUD stigma. We examined whether rurality, social inequity, and racialized segregation across communities from four states in the HEALing Communities Study (HCS) were associated with 1) greater perceived community stigma toward people treated for OUD, 2) greater perceived intervention stigma toward MOUD, and 3) greater perceived intervention stigma toward naloxone by community stakeholders in the HCS. METHODS: From November 2019-January 2020, a cross-sectional survey about community OUD stigma was administered to 801 members of opioid overdose prevention coalitions across 66 communities in four states prior to the start of HCS intervention activities. Bivariate analyses assessed pairwise associations between community rural/urban status and each of the three stigma variables, using linear mixed effect modeling to account for response clustering within communities, state, and respondent sociodemographic characteristics. We conducted similar bivariate analyses to assess pairwise associations between racialized segregation and social inequity. RESULTS: On average, the perceived community OUD stigma scale score of stakeholders from rural communities was 4% higher (ß=1.57, SE=0.7, p≤0.05), stigma toward MOUD was 6% higher (ß=0.28, SE=0.1, p≤0.05), and stigma toward naloxone was 10% higher (ß=0.46, SE=0.1, p≤0.01) than among stakeholders from urban communities. No significant differences in the three stigma variables were found among communities based on racialized segregation or social inequity. CONCLUSION: Perceived community stigma toward people treated for OUD, MOUD, and naloxone was higher among stakeholders in rural communities than in urban communities. Findings suggest that interventions and policies to reduce community-level stigma, particularly in rural areas, are warranted.

Comorbid alcohol use disorder and posttraumatic stress disorder: A proof-of-concept randomized placebo-controlled trial of buprenorphine and naltrexone combination treatment.

BACKGROUND: Effective pharmacologic treatments for comorbid alcohol use disorder (AUD) and posttraumatic stress disorder (PTSD) are lacking. Kappa (κ) opioid receptor antagonists may address this unmet need. Buprenorphine is a κ-opioid antagonist and a partial agonist of mu (µ) opioid receptors. Whereas naltrexone blocks all µ-mediated effects combining it with buprenorphine yields a pharmacologic net effect of opioid receptor antagonism. Because no κ-opioid receptor antagonist it available for clinical use, we tested this combination in a proof-of-concept study. METHODS: Consenting participants were enrolled in a Phase II, multisite, double-blind, randomized, placebo-controlled trial evaluating the effectiveness of sublingual (SL) buprenorphine combined with extended-release (XR) injectable naltrexone for the treatment of comorbid AUD and PTSD. Eligible participants (n = 75) were randomized (1:1:1) to receive either buprenorphine 2 mg/day plus naltrexone-XR (n = 35), buprenorphine 8 mg/day plus naltrexone-XR (n = 6) or SL plus injectable placebo (n = 34) for 12 weeks. The buprenorphine 8 mg/day plus naltrexone-XR arm was dropped early in the trial due to the negative impact of COVID-19 on enrollment. A binary primary outcome of response at week 8 was defined as a decrease from baseline of ≥10 points on the past week Clinician-Administered PTSD Scale (CAPS-5) and a reduction of ≥1 of past month alcohol risk level, as defined by the World Health Organization (WHO) and measured by the Timeline Follow-Back. RESULTS: Based on the results of a futility analysis, enrollment was stopped prior to reaching the initial goal of 90 participants. At the week eight primary timepoint, there were no statistically significant differences between buprenorphine plus naltrexone-XR and placebo group for the primary composite outcome (OR = 0.63; p-value = 0.52), or the subcomponents of the PTSD outcome (OR = 0.76; p-value = 0.69) and AUD outcome (OR = 0.17; p-value = 0.08). The placebo arm had a significantly higher proportion of participants with ≥1 WHO risk level reduction than the buprenorphine plus naltrexone-XR arm (OR = 0.18, p value = 0.02). CONCLUSIONS: This is the first study to evaluate the potential of κ-opioid receptor antagonism for the treatment of comorbid AUD and PTSD. The combination of buprenorphine and naltrexone-XR showed no significant improvement over placebo for the composite, PTSD, or alcohol measures.

Performance of model-based vs. permutation tests in the HEALing (Helping to End Addiction Long-termSM) Communities Study, a covariate-constrained cluster randomized trial.

BACKGROUND: The HEALing (Helping to End Addiction Long-termSM) Communities Study (HCS) is a multi-site parallel group cluster randomized wait-list comparison trial designed to evaluate the effect of the Communities That Heal (CTH) intervention compared to usual care on opioid overdose deaths. Covariate-constrained randomization (CCR) was applied to balance the community-level baseline covariates in the HCS. The purpose of this paper is to evaluate the performance of model-based tests and permutation tests in the HCS setting. We conducted a simulation study to evaluate type I error rates and power for model-based and permutation tests for the multi-site HCS as well as for a subgroup analysis of a single state (Massachusetts). We also investigated whether the maximum degree of imbalance in the CCR design has an impact on the performance of the tests. METHODS: The primary outcome, the number of opioid overdose deaths, is count data assessed at the community level that will be analyzed using a negative binomial regression model. We conducted a simulation study to evaluate the type I error rates and power for 3 tests: (1) Wald-type t-test with small-sample corrected empirical standard error estimates, (2) Wald-type z-test with model-based standard error estimates, and (3) permutation test with test statistics calculated by the difference in average residuals for the two groups. RESULTS: Our simulation results demonstrated that Wald-type t-tests with small-sample corrected empirical standard error estimates from the negative binomial regression model maintained proper type I error. Wald-type z-tests with model-based standard error estimates were anti-conservative. Permutation tests preserved type I error rates if the constrained space was not too small. For all tests, the power was high to detect the hypothesized 40% reduction in opioid overdose deaths for the intervention vs. comparison group both for the overall HCS and the subgroup analysis of Massachusetts (MA). CONCLUSIONS: Based on the results of our simulation study, the Wald-type t-test with small-sample corrected empirical standard error estimates from a negative binomial regression model is a valid and appropriate approach for analyzing cluster-level count data from the HEALing Communities Study. TRIAL REGISTRATION: ClinicalTrials.gov http://www. CLINICALTRIALS: gov ; Identifier: NCT04111939.

Development and performance of a hereditary hemorrhagic telangiectasia-specific quality-of-life instrument.

Hereditary hemorrhagic telangiectasia (HHT) is characterized by arteriovenous malformations and telangiectasia, with primary clinical manifestations of epistaxis and gastrointestinal bleeding and resultant anemia. HHT negatively affects health-related quality of life (HR-QoL); however, existing tools to measure HR-QoL are not HHT specific. Our objective was to develop an HHT-specific HR-QoL (HHT-QoL) instrument and evaluate its performance in a cross-sectional survey of individuals with HHT. Four HHT-specific questions were developed to evaluate the impact of HHT on productivity and social and personal interactions. An anonymous e-mail survey was conducted through Cure HHT. Participants also indicated their perceived HHT severity and completed 3 Patient-Reported Outcomes Measurement Information System (PROMIS) questionnaires: Discretionary Social Activities, Social Roles, and Emotional Distress. Complete data were available for 290 participants who self-identified their HHT severity as mild (29%), moderate (46%), or severe (25%). The HHT-QoL scale was reliable (Cronbach's-α, 0.83). Principal components analysis indicated the instrument was unidimensional. Participants had low levels of QoL with their ability to participate in discretionary social activities (PROMIS mean 36.4 [standard deviation 14.3]) and perform in social roles (41.5 [17.2]), and the presence of a high level of emotional distress (64.8 [24.2]). The HHT-QoL score correlated negatively with PROMIS Discretionary Social Activities (r = -0.65) and Social Roles (r = -0.68) and positively correlated with PROMIS Emotional Distress (r = 0.51). In conclusion, the 4-item HHT-QoL instrument provides valuable insight and may be a useful addition to future clinical research in HHT.

Marginal modeling in community randomized trials with rare events: Utilization of the negative binomial regression model.

BACKGROUND/AIMS: This work is motivated by the HEALing Communities Study, which is a post-test only cluster randomized trial in which communities are randomized to two different trial arms. The primary interest is in reducing opioid overdose fatalities, which will be collected as a count outcome at the community level. Communities range in size from thousands to over one million residents, and fatalities are expected to be rare. Traditional marginal modeling approaches in the cluster randomized trial literature include the use of generalized estimating equations with an exchangeable correlation structure when utilizing subject-level data, or analogously quasi-likelihood based on an over-dispersed binomial variance when utilizing community-level data. These approaches account for and estimate the intra-cluster correlation coefficient, which should be provided in the results from a cluster randomized trial. Alternatively, the coefficient of variation or R coefficient could be reported. In this article, we show that negative binomial regression can also be utilized when communities are large and events are rare. The objectives of this article are (1) to show that the negative binomial regression approach targets the same marginal regression parameter(s) as an over-dispersed binomial model and to explain why the estimates may differ; (2) to derive formulas relating the negative binomial overdispersion parameter k with the intra-cluster correlation coefficient, coefficient of variation, and R coefficient; and (3) analyze pre-intervention data from the HEALing Communities Study to demonstrate and contrast models and to show how to report the intra-cluster correlation coefficient, coefficient of variation, and R coefficient when utilizing negative binomial regression. METHODS: Negative binomial and over-dispersed binomial regression modeling are contrasted in terms of model setup, regression parameter estimation, and formulation of the overdispersion parameter. Three specific models are used to illustrate concepts and address the third objective. RESULTS: The negative binomial regression approach targets the same marginal regression parameter(s) as an over-dispersed binomial model, although estimates may differ. Practical differences arise in regard to how overdispersion, and hence the intra-cluster correlation coefficient is modeled. The negative binomial overdispersion parameter is approximately equal to the ratio of the intra-cluster correlation coefficient and marginal probability, the square of the coefficient of variation, and the R coefficient minus 1. As a result, estimates corresponding to all four of these different types of overdispersion parameterizations can be reported when utilizing negative binomial regression. CONCLUSION: Negative binomial regression provides a valid, practical, alternative approach to the analysis of count data, and corresponding reporting of overdispersion parameters, from community randomized trials in which communities are large and events are rare.

Co-immunization of DNA and Protein in the Same Anatomical Sites Induces Superior Protective Immune Responses against SHIV Challenge.

We compare immunogenicity and protective efficacy of an HIV vaccine comprised of env and gag DNA and Env (Envelope) proteins by co-administration of the vaccine components in the same muscles or by separate administration of DNA + protein in contralateral sites in female rhesus macaques. The 6-valent vaccine includes gp145 Env DNAs, representing six sequentially isolated Envs from the HIV-infected individual CH505, and matching GLA-SE-adjuvanted gp120 Env proteins. Interestingly, only macaques in the co-administration vaccine group are protected against SHIV CH505 acquisition after repeated low-dose intravaginal challenge and show 67% risk reduction per exposure. Macaques in the co-administration group develop higher Env-specific humoral and cellular immune responses. Non-neutralizing Env antibodies, ADCC, and antibodies binding to FcγRIIIa are associated with decreased transmission risk. These data suggest that simultaneous recognition, processing, and presentation of DNA + Env protein in the same draining lymph nodes play a critical role in the development of protective immunity.

Neonatal Rhesus Macaques Have Distinct Immune Cell Transcriptional Profiles following HIV Envelope Immunization.

HIV-1-infected infants develop broadly neutralizing antibodies (bnAbs) more rapidly than adults, suggesting differences in the neonatal versus adult responses to the HIV-1 envelope (Env). Here, trimeric forms of HIV-1 Env immunogens elicit increased gp120- and gp41-specific antibodies more rapidly in neonatal macaques than adult macaques. Transcriptome analyses of neonatal versus adult immune cells after Env vaccination reveal that neonatal macaques have higher levels of the apoptosis regulator BCL2 in T cells and lower levels of the immunosuppressive interleukin-10 (IL-10) receptor alpha (IL10RA) mRNA transcripts in T cells, B cells, natural killer (NK) cells, and monocytes. In addition, immunized neonatal macaques exhibit increased frequencies of activated blood T follicular helper-like (Tfh) cells compared to adults. Thus, neonatal macaques have transcriptome signatures of decreased immunosuppression and apoptosis compared with adult macaques, providing an immune landscape conducive to early-life immunization prior to sexual debut.

Intrahost Dynamics of Human Cytomegalovirus Variants Acquired by Seronegative Glycoprotein B Vaccinees.

Human cytomegalovirus (HCMV) is the most common congenital infection worldwide and a frequent cause of hearing loss and debilitating neurologic disease in newborn infants. Thus, a vaccine to prevent HCMV-associated congenital disease is a public health priority. One potential strategy is vaccination of women of child bearing age to prevent maternal HCMV acquisition during pregnancy. The glycoprotein B (gB) plus MF59 adjuvant subunit vaccine is the most efficacious tested clinically to date, demonstrating 50% protection against primary HCMV infection in a phase 2 clinical trial. Yet, the impact of gB/MF59-elicited immune responses on the population of viruses acquired by trial participants has not been assessed. In this analysis, we employed quantitative PCR as well as multiple sequencing methodologies to interrogate the magnitude and genetic composition of HCMV populations infecting gB/MF59 vaccinees and placebo recipients. We identified several differences between the viral dynamics in acutely infected vaccinees and placebo recipients. First, viral load was reduced in the saliva of gB vaccinees, though not in whole blood, vaginal fluid, or urine. Additionally, we observed possible anatomic compartmentalization of gB variants in the majority of vaccinees compared to only a single placebo recipient. Finally, we observed reduced acquisition of genetically related gB1, gB2, and gB4 genotype "supergroup" HCMV variants among vaccine recipients, suggesting that the gB1 genotype vaccine construct may have elicited partial protection against HCMV viruses with antigenically similar gB sequences. These findings suggest that gB immunization had a measurable impact on viral intrahost population dynamics and support future analysis of a larger cohort.IMPORTANCE Though not a household name like Zika virus, human cytomegalovirus (HCMV) causes permanent neurologic disability in one newborn child every hour in the United States, which is more than that for Down syndrome, fetal alcohol syndrome, and neural tube defects combined. There are currently no established effective measures to prevent viral transmission to the infant following HCMV infection of a pregnant mother. However, the glycoprotein B (gB)/MF59 vaccine, which aims to prevent pregnant women from acquiring HCMV, is the most successful HCMV vaccine tested clinically to date. Here, we used viral DNA isolated from patients enrolled in a gB vaccine trial who acquired HCMV and identified several impacts that this vaccine had on the size, distribution, and composition of the in vivo viral population. These results have increased our understanding of why the gB/MF59 vaccine was partially efficacious, and such investigations will inform future rational design of a vaccine to prevent congenital HCMV.

IDLV-HIV-1 Env vaccination in non-human primates induces affinity maturation of antigen-specific memory B cells.

HIV continues to be a major global health issue. In spite of successful prevention interventions and treatment methods, the development of an HIV vaccine remains a major priority for the field and would be the optimal strategy to prevent new infections. We showed previously that a single immunization with a SIV-based integrase-defective lentiviral vector (IDLV) expressing the 1086.C HIV-1-envelope induced durable, high-magnitude immune responses in non-human primates (NHPs). In this study, we have further characterized the humoral responses by assessing antibody affinity maturation and antigen-specific memory B-cell persistence in two vaccinated macaques. These animals were also boosted with IDLV expressing the heterologous 1176.C HIV-1-Env to determine if neutralization breadth could be increased, followed by evaluation of the injection sites to assess IDLV persistence. IDLV-Env immunization was associated with persistence of the vector DNA for up to 6 months post immunization and affinity maturation of antigen-specific memory B cells.

RAB11FIP5 Expression and Altered Natural Killer Cell Function Are Associated with Induction of HIV Broadly Neutralizing Antibody Responses.

HIV-1 broadly neutralizing antibodies (bnAbs) are difficult to induce with vaccines but are generated in ∼50% of HIV-1-infected individuals. Understanding the molecular mechanisms of host control of bnAb induction is critical to vaccine design. Here, we performed a transcriptome analysis of blood mononuclear cells from 47 HIV-1-infected individuals who made bnAbs and 46 HIV-1-infected individuals who did not and identified in bnAb individuals upregulation of RAB11FIP5, encoding a Rab effector protein associated with recycling endosomes. Natural killer (NK) cells had the highest differential expression of RAB11FIP5, which was associated with greater dysregulation of NK cell subsets in bnAb subjects. NK cells from bnAb individuals had a more adaptive/dysfunctional phenotype and exhibited impaired degranulation and cytokine production that correlated with RAB11FIP5 transcript levels. Moreover, RAB11FIP5 overexpression modulated the function of NK cells. These data suggest that NK cells and Rab11 recycling endosomal transport are involved in regulation of HIV-1 bnAb development.

Polyclonal HIV envelope-specific breast milk antibodies limit founder SHIV acquisition and cell-associated virus loads in infant rhesus monkeys.

Breast milk HIV-1 transmission is currently the predominant contributor to pediatric HIV infections. Yet, only ~10% of breastfeeding infants born to untreated HIV-infected mothers become infected. This study assessed the protective capacity of natural HIV envelope-specific antibodies isolated from the milk of HIV-infected women in an infant rhesus monkey (RM), tier 2 SHIV oral challenge model. To mimic placental and milk maternal antibody transfer, infant RMs were i.v. infused and orally treated at the time of challenge with a single weakly neutralizing milk monoclonal antibody (mAb), a tri-mAb cocktail with weakly neutralizing and ADCC functionalities, or an anti-influenza control mAb. Of these groups, the fewest tri-mAb-treated infants had SHIV detectable in plasma or tissues (2/6, 5/6, and 7/8 animals infected in tri-mAb, single-mAb, and control-mAb groups, respectively). Tri-mAb-treated infants demonstrated significantly fewer plasma transmitted/founder variants and reduced peripheral CD4+ T cell proviral loads at 8 weeks post-challenge compared to control mAb-treated infants. Abortive infection was observed as detectable CD4+ T cell provirus in non-viremic control mAb- and single mAb-, but not in tri-mAb-treated animals. These results suggest that polyfunctional milk antibodies contribute to the natural inefficiency of HIV-1 transmission through breastfeeding and infant vaccinations eliciting non-neutralizing antibody responses could reduce postnatal HIV transmission.

HCMV glycoprotein B subunit vaccine efficacy mediated by nonneutralizing antibody effector functions.

Human cytomegalovirus (HCMV) is the most common congenital infection worldwide, frequently causing hearing loss and brain damage in afflicted infants. A vaccine to prevent maternal acquisition of HCMV during pregnancy is necessary to reduce the incidence of infant disease. The glycoprotein B (gB) + MF59 adjuvant subunit vaccine platform is the most successful HCMV vaccine tested to date, demonstrating ∼50% efficacy in preventing HCMV acquisition in multiple phase 2 trials. However, the mechanism of vaccine protection remains unknown. Plasma from 33 postpartum women gB/MF59 vaccinees at peak immunogenicity was tested for gB epitope specificity as well as neutralizing and nonneutralizing anti-HCMV effector functions and compared with an HCMV-seropositive cohort. gB/MF59 vaccination elicited IgG responses with gB-binding magnitude and avidity comparable to natural infection. Additionally, IgG subclass distribution was similar with predominant IgG1 and IgG3 responses induced by gB vaccination and HCMV infection. However, vaccine-elicited antibodies exhibited limited neutralization of the autologous virus, negligible neutralization of multiple heterologous strains, and limited binding responses against gB structural motifs targeted by neutralizing antibodies including AD-1, AD-2, and domain I. Vaccinees had high-magnitude IgG responses against AD-3 linear epitopes, demonstrating immunodominance against this nonneutralizing, cytosolic region. Finally, vaccine-elicited IgG robustly bound membrane-associated gB on the surface of transfected or HCMV-infected cells and mediated virion phagocytosis, although were poor mediators of NK cell activation. Altogether, these data suggest that nonneutralizing antibody functions, including virion phagocytosis, likely played a role in the observed 50% vaccine-mediated protection against HCMV acquisition.

Predominant envelope variable loop 2-specific and gp120-specific antibody-dependent cellular cytotoxicity antibody responses in acutely SIV-infected African green monkeys.

BACKGROUND: The initial envelope (Env)-specific antibody response in acutely HIV-1-infected individuals and simian immunodeficiency virus (SIV)-infected rhesus monkeys (RMs) is dominated by non-neutralizing antibodies targeting Env gp41. In contrast, natural primate SIV hosts, such as African green monkeys (AGMs), develop a predominant Env gp120-specific antibody response to SIV infection. However, the fine-epitope specificity and function of SIV Env-specific plasma IgG, and their potential role on autologous virus co-evolution in SIV-infected AGMs and RMs remain unclear. RESULTS: Unlike the dominant linear gp41-specific IgG responses in RMs, SIV-infected AGMs demonstrated a unique linear variable loop 2 (V2)-specific plasma IgG response that arose concurrently with high gp120-directed antibody-dependent cellular cytotoxicity (ADCC) activity, and SIVsab-infected cell binding responses during acute infection. Moreover, SIV variants isolated from SIV-infected AGMs exhibited high amino acid mutation frequencies within the Env V1V2 loop compared to those of RMs. Notably, the linear V2-specific IgG epitope in AGMs overlaps with an analogous region of the HIV V2 loop containing the K169 mutation epitope identified in breakthrough viruses from RV144 vaccinees. CONCLUSION: Vaccine-elicited Env V2-specific IgG responses have been proposed as an immune correlate of reduced risk in HIV-1/SIV acquisition in humans and RMs. Yet the pathways to elicit these potentially-protective V2-specific IgG responses remain unclear. In this study, we demonstrate that SIV-infected AGMs, which are the natural hosts of SIV, exhibited high plasma linear V2-specific IgG binding responses that arose concurrently with SIV Env gp120-directed ADCC-mediating, and SIV-infected cell plasma IgG binding responses during acute SIV infection, which were not present in acutely SIV-infected RMs. The linear V2-specific antibody response in AGMs targets an overlapping epitope of the proposed site of vaccine-induced immune pressure defined in the moderately protective RV144 HIV-1 vaccine trial. Identifying host factors that control the early elicitation of Env V2-specific IgG and ADCC antibody responses in these natural SIV hosts could inform vaccination strategies aimed at rapidly inducing potentially-protective HIV-1 Env-specific responses in humans.

Immunogenicity of NYVAC Prime-Protein Boost Human Immunodeficiency Virus Type 1 Envelope Vaccination and Simian-Human Immunodeficiency Virus Challenge of Nonhuman Primates.

A preventive human immunodeficiency virus type 1 (HIV-1) vaccine is an essential part of the strategy to eradicate AIDS. A critical question is whether antibodies that do not neutralize primary isolate (tier 2) HIV-1 strains can protect from infection. In this study, we investigated the ability of an attenuated poxvirus vector (NYVAC) prime-envelope gp120 boost to elicit potentially protective antibody responses in a rhesus macaque model of mucosal simian-human immunodeficiency virus (SHIV) infection. NYVAC vector delivery of a group M consensus envelope, trivalent mosaic envelopes, or a natural clade B isolate B.1059 envelope elicited antibodies that mediated neutralization of tier 1 viruses, cellular cytotoxicity, and phagocytosis. None of the macaques made neutralizing antibodies against the tier 2 SHIV SF162P3 used for mucosal challenge. Significant protection from infection was not observed for the three groups of vaccinated macaques compared to unvaccinated macaques, although binding antibody to HIV-1 Env correlated with decreased viremia after challenge. Thus, NYVAC Env prime-gp120 boost vaccination elicited polyfunctional, nonneutralizing antibody responses with minimal protective activity against tier 2 SHIV mucosal challenge.IMPORTANCE The antibody responses that confer protection against HIV-1 infection remain unknown. Polyfunctional antibody responses correlated with time to infection in previous macaque studies. Determining the ability of vaccines to induce these types of responses is critical for understanding how to improve upon the one efficacious human HIV-1 vaccine trial completed thus far. We characterized the antibody responses induced by a NYVAC-protein vaccine and determined the protective capacity of polyfunctional antibody responses in an R5, tier 2 mucosal SHIV infection model.

Computational analysis of antibody dynamics identifies recent HIV-1 infection.

Accurate HIV-1 incidence estimation is critical to the success of HIV-1 prevention strategies. Current assays are limited by high false recent rates (FRRs) in certain populations and a short mean duration of recent infection (MDRI). Dynamic early HIV-1 antibody response kinetics were harnessed to identify biomarkers for improved incidence assays. We conducted retrospective analyses on circulating antibodies from known recent and longstanding infections and evaluated binding and avidity measurements of Env and non-Env antigens and multiple antibody forms (i.e., IgG, IgA, IgG3, IgG4, dIgA, and IgM) in a diverse panel of 164 HIV-1-infected participants (clades A, B, C). Discriminant function analysis identified an optimal set of measurements that were subsequently evaluated in a 324-specimen blinded biomarker validation panel. These biomarkers included clade C gp140 IgG3, transmitted/founder clade C gp140 IgG4 avidity, clade B gp140 IgG4 avidity, and gp41 immunodominant region IgG avidity. MDRI was estimated at 215 day or alternatively, 267 days. FRRs in untreated and treated subjects were 5.0% and 3.6%, respectively. Thus, computational analysis of dynamic HIV-1 antibody isotype and antigen interactions during infection enabled design of a promising HIV-1 recency assay for improved cross-sectional incidence estimation.

Initiation of HIV neutralizing B cell lineages with sequential envelope immunizations.

A strategy for HIV-1 vaccine development is to define envelope (Env) evolution of broadly neutralizing antibodies (bnAbs) in infection and to recreate those events by vaccination. Here, we report host tolerance mechanisms that limit the development of CD4-binding site (CD4bs), HCDR3-binder bnAbs via sequential HIV-1 Env vaccination. Vaccine-induced macaque CD4bs antibodies neutralize 7% of HIV-1 strains, recognize open Env trimers, and accumulate relatively modest somatic mutations. In naive CD4bs, unmutated common ancestor knock-in mice Env+B cell clones develop anergy and partial deletion at the transitional to mature B cell stage, but become Env- upon receptor editing. In comparison with repetitive Env immunizations, sequential Env administration rescue anergic Env+ (non-edited) precursor B cells. Thus, stepwise immunization initiates CD4bs-bnAb responses, but immune tolerance mechanisms restrict their development, suggesting that sequential immunogen-based vaccine regimens will likely need to incorporate strategies to expand bnAb precursor pools.

HIV DNA-Adenovirus Multiclade Envelope Vaccine Induces gp41 Antibody Immunodominance in Rhesus Macaques.

Dominant antibody responses in vaccinees who received the HIV-1 multiclade (A, B, and C) envelope (Env) DNA/recombinant adenovirus virus type 5 (rAd5) vaccine studied in HIV-1 Vaccine Trials Network (HVTN) efficacy trial 505 (HVTN 505) targeted Env gp41 and cross-reacted with microbial antigens. In this study, we asked if the DNA/rAd5 vaccine induced a similar antibody response in rhesus macaques (RMs), which are commonly used as an animal model for human HIV-1 infections and for testing candidate HIV-1 vaccines. We also asked if gp41 immunodominance could be avoided by immunization of neonatal RMs during the early stages of microbial colonization. We found that the DNA/rAd5 vaccine elicited a higher frequency of gp41-reactive memory B cells than gp120-memory B cells in adult and neonatal RMs. Analysis of the vaccine-induced Env-reactive B cell repertoire revealed that the majority of HIV-1 Env-reactive antibodies in both adult and neonatal RMs were targeted to gp41. Interestingly, a subset of gp41-reactive antibodies isolated from RMs cross-reacted with host antigens, including autologous intestinal microbiota. Thus, gp41-containing DNA/rAd5 vaccine induced dominant gp41-microbiota cross-reactive antibodies derived from blood memory B cells in RMs as observed in the HVTN 505 vaccine efficacy trial. These data demonstrated that RMs can be used to investigate gp41 immunodominance in candidate HIV-1 vaccines. Moreover, colonization of neonatal RMs occurred within the first week of life, and immunization of neonatal RMs during this time also induced a dominant gp41-reactive antibody response.IMPORTANCE Our results are critical to current work in the HIV-1 vaccine field evaluating the phenomenon of gp41 immunodominance induced by HIV-1 Env gp140 in RMs and humans. Our data demonstrate that RMs are an appropriate animal model to study this phenomenon and to determine the immunogenicity in new HIV-1 Env trimer vaccine designs. The demonstration of gp41 immunodominance in memory B cells of both adult and neonatal RMs indicated that early vaccination could not overcome gp41 dominant responses.

Development of a recombinant yellow fever vector expressing a HIV clade C founder envelope gp120.

Development of a HIV-1 vaccine is a major global priority. The yellow fever virus (YFV) attenuated vaccine 17D is among the most effective of currently used vaccines. However, the stability of the YFV17D vector when carrying non-flavivirus genes has been problematic. We have constructed and expressed HIV-1 Env in YFV17D with either single transmembrane (STM) or double transmembrane (DTM) YFV E protein domains for the development of anti-HIV antibodies. Here we describe modifications of the YFV17D vector such that HIV-1 Env gp120 is expressed in up to 5 passages in Vero cells. Immunization with recombinant YFV17D vector prime followed by HIV-1 CH505 TF gp120 protein boosts were able to induce neutralizing antibodies for a HIV-1 tier 1 isolate in mice. This modified YFV vector may be a starting point for constructing HIV-1 vaccine candidate priming vectors.