Polyvalent immunization elicits a synergistic broadly neutralizing immune response to hypervariable region 1 variants of hepatitis C virus.

A hepatitis C virus (HCV) vaccine is urgently needed. Vaccine development has been hindered by HCV's genetic diversity, particularly within the immunodominant hypervariable region 1 (HVR1). Here, we developed a strategy to elicit broadly neutralizing antibodies to HVR1, which had previously been considered infeasible. We first applied a unique information theory-based measure of genetic distance to evaluate phenotypic relatedness between HVR1 variants. These distances were used to model the structure of HVR1's sequence space, which was found to have five major clusters. Variants from each cluster were used to immunize mice individually, and as a pentavalent mixture. Sera obtained following immunization neutralized every variant in a diverse HCVpp panel (n = 10), including those resistant to monovalent immunization, and at higher mean titers (1/ID50 = 435) than a glycoprotein E2 (1/ID50 = 205) vaccine. This synergistic immune response offers a unique approach to overcoming antigenic variability and may be applicable to other highly mutable viruses.

Neutralizing antibodies to interferon alfa arising during peginterferon therapy of chronic hepatitis B in children and adults: Results from the HBRN Trials.

BACKGROUND AIMS: Pegylated interferon-α (PegIFNα) is of limited utility during immunotolerant or immune active phases of chronic hepatitis B infection but is being explored as part of new cure regimens. Low/absent levels of IFNα found in some patients receiving treatment are associated with limited/no virological responses. The study aimed to determine if sera from participants inhibit IFNα activity and/or contain therapy-induced anti-IFNα antibodies. APPROACH RESULTS: Pre-treatment, on-treatment, and post-treatment sera from 61 immunotolerant trial participants on PegIFNα/entecavir therapy and 88 immune active trial participants on PegIFNα/tenofovir therapy were screened for anti-IFNα antibodies by indirect ELISA. The neutralization capacity of antibodies was measured by preincubation of sera±recombinant human IFNα added to Huh7 cells with the measurement of interferon-stimulated gene (ISG)-induction by qPCR. Correlations between serum-induced ISG inhibition, presence, and titer of anti-IFNα antibodies and virological responses were evaluated. Preincubation of on-treatment serum from 26 immunotolerant (43%) and 13 immune active (15%) participants with recombinant-human IFNα markedly blunted ISG-induction in Huh7 cells. The degree of ISG inhibition correlated with IFNα antibody titer ( p < 0.0001; r = 0.87). On-treatment development of anti-IFNα neutralizing antibodies (nAbs) was associated with reduced quantitative HBsAg and qHBeAg declines ( p < 0.05) and inhibited IFNα bioactivity to 240 weeks after PegIFNα cessation. Children developed anti-IFNα nAbs more frequently than adults ( p = 0.004) but nAbs in children had less impact on virological responses. CONCLUSIONS: The development of anti-IFNα nAbs during PegIFNα treatment diminishes responses to antiviral therapy. Understanding how and why anti-IFNα antibodies develop may allow for the optimization of IFN-based therapy, which is critical given its renewed use in HBV-cure strategies.

Role of HVR1 sequence similarity in the cross-genotypic neutralization of HCV.

Despite available treatments, a prophylactic HCV vaccine is needed to achieve elimination targets. HCV vaccine development has faltered largely because the extreme diversity of the virus limits the protective breadth of vaccine elicited antibodies. It is believed that the principle neutralizing epitope in natural infection, HVR1, which is the most variable epitope in HCV, mediates humoral immune escape. So far, efforts to circumvent HVR1 interference in the induction and function of conserved targeting Ab have failed. Efforts to understand factors contributing to cross-neutralization of HVR1 variants have also been limited. Here, following mouse immunizations with two patient-derived HVR1 peptides, we observe cross-genotype neutralization of variants differing at 15/21 positions. Surprisingly, sequence similarity was not associated with cross-neutralization. It appeared neutralization sensitivity was an intrinsic feature of each variant, rather than emergent from the immunogen specific Ab response. These findings provide novel insight into HVR1-mediated immune evasion, with important implications for HCV vaccine design.

Impacting Management of Chronic Kidney Disease Through Primary Care Practice Audits: A Quality Improvement Study.

Background: Risk prediction tools are important in chronic disease management, but their implementation into clinical workflow is often limited by lack of electronic health record (EHR)-linked solutions. Objective: To implement the Khure Health (KH) clinical decision support platform with an artificial intelligence (AI)-enabled algorithm for chronic kidney disease (CKD) risk detection in 201 primary care provider practices across Ontario. Design: Multi-practice quality improvement study. Setting: The study was conducted in Ontario, Canada. Participants: 201 primary care practices. Measurements: Per-practice CKD risk stratification and clinician action. Methods: Data for estimated glomerular filtration rate (eGFR), albuminuria, demographics, and comorbid conditions were extracted from the EHR using KH's natural language processing (NLP) algorithms. Patients already on dialysis, visiting a nephrologist, older than 85, or already on a sodium-glucose cotransporter 2 inhibitor (SGLT2i) were excluded. The remaining individuals were risk stratified using the kidney failure risk equation, presence or absence of cardiovascular disease (CVD), or other comorbid conditions. A dashboard with disease-specific educational information and links to the EHRs of the identified patients was created. Results: We screened 361 299 individuals and identified 8194 patients with CKD Stage 3 at risk for progression or cardiovascular events. A total of 620 individuals were at high risk for CKD progression or CVD, and 2592 were at intermediate risk. A total of 2010 individuals (10 patients per practice) at high or moderate risk were selected for a chart audit, and appropriate additional testing (repeat eGFR or albuminuria) or prescription of disease-modifying therapy occurred in 24.32% of these patients. Limitations: Data on comorbidities, medications, or demographic variables are not available for presentation or statistical analysis due to privacy legislation and primary care provider (PCP) custodianship over EHR data. Conclusion: An AI-enabled EHR clinical decision support application that can detect and risk stratify patients with CKD can enable improved laboratory testing and management. Larger trials of clinical decision support and practice audit applications will be needed to impact CKD management nationally.

Contexte: Les outils de prédiction des risques sont importants dans la gestion des maladies chroniques, mais leur intégration dans le flux de travail clinique est souvent limitée par le manque de solutions liées aux dossiers de santé informatisés. Objectif: Mettre en œuvre, dans 201 cabinets de soins primaires de l'Ontario, la plateforme d'aide à la décision clinique Khure Health (KH) dotée d'un algorithme basé sur l'intelligence artificielle (IA) pour détecter les risques d'insuffisance rénale chronique (IRC). Conception: Étude d'amélioration de la qualité dans plusieurs cabinets. Cadre: Étude menée en Ontario (Canada). Sujets: 201 cabinets de soins primaires. Mesures: Stratification du risque d'IRC par cabinet et actions du clinicien. Méthodologie: Les données relatives au débit de filtration glomérulaire estimé (DFGe), à l'albuminurie, à la démographie et aux maladies concomitantes ont été extraites des dossiers de santé informatisés (DSI) à l'aide des algorithmes de traitement du langage naturel (TLN) de KH. Ont été exclus les patients sous dialyse, suivis par un néphrologue, âgés de plus de 85 ans ou traités avec un inhibiteur du cotransporteur sodium-glucose de type 2 (SGLT2i). Les autres individus ont été stratifiés selon le risque évalué par l'équation prédictive du risque d'évolution vers l'insuffisance rénale, et par la présence ou l'absence de maladie cardiovasculaire ou d'autres affections concomitantes. Un tableau de bord contenant des informations éducatives sur la maladie et des liens vers les DSI des patients identifiés a été créé. Résultats: Nous avons examiné les dossiers de 361 299 personnes et identifié 8 194 patients atteints d'IRC de stade 3 présentant un risque de progression de l'IRC ou d'événements cardiovasculaires. De ces 8 194 patients, 620 présentaient un risque jugé élevé et 2 592 un risque modéré. En tout, 2 010 personnes (10 patients par cabinet) présentant un risque élevé ou modéré ont été sélectionnées pour une vérification des dossiers. Les tests supplémentaires appropriés (répétition des mesures du DFGe ou de l'albuminurie) ou la prescription de traitement modifiant la maladie ont été ordonnés chez 24,32 % de ces patients. Limites: Les données sur les maladies concomitantes, la médication ou les variables démographiques n'étaient pas disponibles pour la présentation ou l'analyse statistique en raison de la loi sur la protection de la vie privée et du fait que les DSI sont sous la garde du médecin de soins primaires. Conclusion: Une application d'aide à la prise de décisions cliniques basée sur l'IA pour les DSI, qui est capable de détecter et de stratifier les risques chez les patients atteints d'IRC, pourrait permettre d'améliorer la gestion de la maladie et les tests de laboratoire. Des essais à plus grande échelle portant sur les applications d'aide à la décision clinique et de vérification des pratiques seront nécessaires pour avoir une incidence sur la gestion de l'IRC à l'échelle nationale.

HepG2-NTCP Subclones Exhibiting High Susceptibility to Hepatitis B Virus Infection.

HepG2 cells reconstituted with Hepatitis B virus (HBV) entry receptor sodium taurocholate co-transporting polypeptide (NTCP) are widely used as a convenient in vitro cell culture infection model for HBV replication studies. As such, it is pertinent that HBV infectivity is maintained at steady-state levels for an accurate interpretation of in vitro data. However, variations in the HBV infection efficiency due to imbalanced NTCP expression levels in the HepG2 cell line may affect experimental results. In this study, we performed single cell-cloning of HepG2-NTCP-A3 parental cells via limiting dilution and obtained multiple subclones with increased permissiveness to HBV. Specifically, one subclone (HepG2-NTCP-A3/C2) yielded more than four-fold higher HBV infection compared to the HepG2-NTCP-A3 parental clone. In addition, though HBV infectivity was universally reduced in the absence of polyethylene glycol (PEG), subclone C2 maintained relatively greater permissiveness under PEG-free conditions, suggesting the functional heterogeneity within parental HepG2-NTCP-A3 may be exploitable in developing a PEG-free HBV infection model. The increased viral production correlated with increased intracellular viral antigen expression as evidenced through HBcAg immunofluorescence staining. Further, these subclones were found to express different levels of NTCP, albeit with no remarkable morphology or cell growth differences. In conclusion, we isolated the subclones of HepG2-NTCP-A3 which support efficient HBV production and thus provide an improved in vitro HBV infection model.

Implications of viral transmitted/founder (T/F) dynamics on vaccine development.

Viral infection typically originates from a limited number of virions known as transmitted/founder (T/F) viruses. Studies of cross-species transmission, and intra-species transmission of antigenically variable viruses, indicates T/F variants may express distinct, transmissibility enhancing phenotypes. However, with evidence that transmissibility is associated with not only intrinsic virological features, such as virion composition, but also extrinsic factors, such as viral population structure, the challenge of resolving T/F signatures that can be targeted by rational vaccine or antiviral design is substantial. Nonetheless, failure to develop vaccines for antigenically variable viruses, such as HIV/HCV, and the ongoing risk of cross-species transmission with pandemic potential, recommends development of T/F targeting vaccines. In this commentary, the T/F phenomena is introduced, explored in both the classical (HIV) and non-canonical (coronaviruses) instances, and discussed in relation to rational and preemptive vaccine design.

Antigenic Variability.

Protective vaccines for hypervariable pathogens are urgently needed. It has been proposed that amputating highly variable epitopes from vaccine antigens would induce the production of broadly protective antibodies targeting conserved epitopes. However, so far, these approaches have failed, partially because conserved epitopes are occluded in vivo and partially because co-localizing patterns of immunodominance and antigenic variability render variable epitopes the primary target for antibodies in natural infection. In this Perspective, to recast the challenge of vaccine development for hypervariable pathogens, I evaluate convergent mechanisms of adaptive variation, such as intrahost immune-mediated diversification, spatiotemporally defined antigenic space, and infection-enhancing cross-immunoreactivity. The requirements of broadly protective immune responses targeting variable pathogens are formulated in terms of cross-immunoreactivity, stoichiometric thresholds for neutralization, and the elicitation of antibodies targeting physicochemically conserved signatures within sequence variable domains.

A bivalent HCV peptide vaccine elicits pan-genotypic neutralizing antibodies in mice.

Vaccine development for antigenically variable pathogens has faltered because extreme genetic diversity precludes induction of broadly neutralizing antibodies (nAB) with classical vaccines. Here, using the most variable epitope of any known human pathogen (HVR1 of HCV), we describe a novel approach capable of eliciting broadly neutralizing antibodies targeting highly variable epitopes. Our proof-of-concept vaccine elicited pan-genotypic nAB against HCV variants differing from the immunogen sequences by more than 70% at the amino acid level. These findings suggest broadly nAB to highly variable pathogens can be elicited by vaccines designed to target physicochemically conserved residues within hypervariable epitopes.