A cryptic site in class 5 epitope of SARS-CoV-2 RBD maintains highly conservation across natural isolates.

The emergence of SARS-CoV-2 variants raises concerns about the efficacy of existing COVID-19 vaccines and therapeutics. Previously, we identified a conserved cryptic class 5 epitope of SARS-CoV-2 receptor binding domain (RBD) by two cross-neutralizing antibodies 7D6 and 6D6. Intriguingly, this site remains resistant to substantial mutations occurred in ever-changing SARS-CoV-2 subvariants. As compared to class 3 antibody S309, 6D6 maintains broad and consistent neutralizing activities against SARS-CoV-2 variants. Furthermore, 6D6 effectively protected hamster from the virulent Beta strain. Sequence alignment of approximately 6 million documented SARS-CoV-2 isolates revealed that 6D6 epitope maintains an exceptionally high conservation rate (99.92%). Structural analysis demonstrated that all 33 mutations accumulated in XBB.1.5 since the original strain do not perturb the binding 6D6 to RBD, in line with the sequence analysis throughout the antigenicity evolution of SARS-CoV-2. These findings suggest the potential of this epitope serving as a critical determinant for vaccines and therapeutic design.

Stepping forward: T-cell redirecting bispecific antibodies in cancer therapy.

T cell-redirecting bispecific antibodies are specifically designed to bind to tumor-associated antigens, thereby engaging with CD3 on the T cell receptor. This linkage between tumor cells and T cells actively triggers T cell activation and initiates targeted killing of the identified tumor cells. These antibodies have emerged as one of the most promising avenues within tumor immunotherapy. However, despite success in treating hematological malignancies, significant advancements in solid tumors have yet to be explored. In this review, we aim to address the critical challenges associated with T cell-redirecting bispecific antibodies and explore novel strategies to overcome these obstacles, with the ultimate goal of expanding the application of this therapy to include solid tumors.

Comprehensive Overview of Broadly Neutralizing Antibodies against SARS-CoV-2 Variants.

Currently, SARS-CoV-2 has evolved into various variants, including the numerous highly mutated Omicron sub-lineages, significantly increasing immune evasion ability. The development raises concerns about the possibly diminished effectiveness of available vaccines and antibody-based therapeutics. Here, we describe those representative categories of broadly neutralizing antibodies (bnAbs) that retain prominent effectiveness against emerging variants including Omicron sub-lineages. The molecular characteristics, epitope conservation, and resistance mechanisms of these antibodies are further detailed, aiming to offer suggestion or direction for the development of therapeutic antibodies, and facilitate the design of vaccines with broad-spectrum potential.

A new mechanism of antibody diversity: formation of the natural antibodies containing LAIR1 and LILRB1 extracellular domains.

The recent discovery of public antibodies targeting Plasmodium falciparum-encoded repetitive interspersed families of polypeptides (RIFINs), which contain extracellular immunoglobulin-like domains from LAIR1 or LILRB1, constitutes a significant step forward in comprehending the reactivity of the Plasmodium parasite. These antibodies arise from unique B cell clones and demonstrate extensive cross-reactivity through their interaction with P. falciparum RIFINs. LAIR1 and LILRBs are specialized type I transmembrane glycoproteins, classified as immune inhibitory receptors, restricted to primates and mainly found on hematopoietic cells. They are instrumental in modulating interactions within the tumor microenvironment and across the immune system, and are increasingly recognized as important in anti-cancer immunotherapy and pathogen defense. The presence of LAIR1/LILRB1-containing antibodies offers new insights into malaria parasite evasion strategies and the immune system's response. Additionally, the innovative method of integrating extra exons into the antibody switch region is a noteworthy advancement, enriching the strategies for the generation of a varied array of bispecific and multispecific antibodies.

Active surveillance of hepatitis E: a 10-year epidemiological analysis in a city in eastern China.

Hepatitis E virus (HEV) is an important cause of acute hepatitis, however, is highly neglected and largely underreported. This study aimed to describe the detailed epidemiology of hepatitis E (HE) through a 10-year surveillance. A community-based active hepatitis surveillance was conducted between November 2007 and October 2017 in 11 townships of Dongtai City in China, involving 355,673 residents. Serum samples were obtained from patients presenting with hepatitis symptoms for more than 3 days. Serum alanine aminotransferase (ALT) levels greater than 2.5 times the upper limit of normal (ULN) were considered acute hepatitis. Samples were subsequently tested for IgG and IgM anti-HEV antibodies, HEV RNA, and hepatitis B surface antigen (HBsAg). The data indicated the incidence of HE fluctuated downward from 2007 to 2017, with an average annual age-standardized incidence of 17.50 per 100,000, exceeding the 10.26 per 100,000 in the National Notifiable Disease Report System (NNDRS). The incidence was notably higher among males (20.95 per 100,000) and individuals aged 50-69 years (37.47 per 100,000). Genotype 4 (HEV-4) was the predominantly circulating genotype during the study period. Furthermore, the study revealed the incidence of hepatitis with HEV and hepatitis B virus (HBV) co-infection was 4.99 per 100,000. The active surveillance system identified a higher incidence of HE compared to NNDRS, with a decreased prevalence over a 10-year period. While efforts are still needed to prevent HE in high-risk populations, including individuals with hepatitis B and the elderly.

A Chymotrypsin-Dependent Live-Attenuated Influenza Vaccine Provides Protective Immunity against Homologous and Heterologous Viruses.

Influenza virus is one of the main pathogens causing respiratory diseases in humans. Vaccines are the most effective ways to prevent viral diseases. However, the limited protective efficacy of current influenza vaccines highlights the importance of novel, safe, and effective universal influenza vaccines. With the progress of the COVID-19 pandemic, live-attenuated vaccines delivered through respiratory mucosa have shown robustly protective efficacy. How to obtain a safe and effective live-attenuated vaccine has become a major challenge. Herein, using the influenza virus as a model, we have established a strategy to quickly obtain a live-attenuated vaccine by mutating the cleavage site of the influenza virus. This mutated influenza virus can be specifically cleaved by chymotrypsin. It has similar biological characteristics to the original strain in vitro, but the safety is improved by at least 100 times in mice. It can effectively protect against lethal doses of both homologous H1N1 and heterologous H5N1 viruses post mucosal administration, confirming that the vaccine generated by this strategy has good safety and broad-spectrum protective activities. Therefore, this study can provide valuable insights for the development of attenuated vaccines for respiratory viruses or other viruses with cleavage sites.

Combination therapy with oncolytic virus and T cells or mRNA vaccine amplifies antitumor effects.

Antitumor therapies based on adoptively transferred T cells or oncolytic viruses have made significant progress in recent years, but the limited efficiency of their infiltration into solid tumors makes it difficult to achieve desired antitumor effects when used alone. In this study, an oncolytic virus (rVSV-LCMVG) that is not prone to induce virus-neutralizing antibodies was designed and combined with adoptively transferred T cells. By transforming the immunosuppressive tumor microenvironment into an immunosensitive one, in B16 tumor-bearing mice, combination therapy showed superior antitumor effects than monotherapy. This occurred whether the OV was administered intratumorally or intravenously. Combination therapy significantly increased cytokine and chemokine levels within tumors and recruited CD8+ T cells to the TME to trigger antitumor immune responses. Pretreatment with adoptively transferred T cells and subsequent oncolytic virotherapy sensitizes refractory tumors by boosting T-cell recruitment, down-regulating the expression of PD-1, and restoring effector T-cell function. To offer a combination therapy with greater translational value, mRNA vaccines were introduced to induce tumor-specific T cells instead of adoptively transferred T cells. The combination of OVs and mRNA vaccine also displays a significant reduction in tumor burden and prolonged survival. This study proposed a rational combination therapy of OVs with adoptive T-cell transfer or mRNA vaccines encoding tumor-associated antigens, in terms of synergistic efficacy and mechanism.

Intranasal influenza-vectored COVID-19 vaccines confer broad protection against SARS-CoV-2 XBB variants in hamsters.

The XBB.1.5 subvariant has garnered significant attention due to its exceptional immune evasion and transmissibility. Significantly, the evolutionary trajectory of SARS-CoV-2 has shown continual progression, with a recent global shift observed from XBB to BA.2.86, exemplified by the emergence of the predominant JN.1 subvariant. This phenomenon highlights the need for vaccines that can provide broad-spectrum antigenic coverage. In this study, we utilized a NS1-deleted (dNS1) influenza viral vector to engineer an updated live-attenuated vectored vaccine called dNS1-XBB-RBD. This vaccine encodes the receptor-binding domain (RBD) protein of the XBB.1.5 strain. Our findings demonstrate that the dNS1-XBB-RBD vaccine elicits a similar systemic and mucosal immune response compared to its prototypic form, dNS1-RBD. In hamsters, the dNS1-XBB-RBD vaccine provided robust protection against the SARS-CoV-2 immune-evasive strains XBB.1.9.2.1 and Beta. Remarkably, nasal vaccination with dNS1-RBD, which encodes the ancestor RBD gene, also effectively protected hamsters against both the XBB.1.9.2.1 and Beta strains. These results provide valuable insights about nasal influenza-vectored vaccine and present a promising strategy for the development of a broad-spectrum vaccine against COVID-19 in the future.

Design of a recombinant asparaginyl ligase for site-specific modification using efficient recognition and nucleophile motifs.

Asparaginyl ligases have been extensively utilized as valuable tools for site-specific bioconjugation or surface-modification. However, the application is hindered by the laborious and poorly reproducible preparation processes, unstable activity and ambiguous substrate requirements. To address these limitations, this study employed a structure-based rational approach to obtain a high-yield and high-activity protein ligase called OaAEP1-C247A-aa55-351. It was observed that OaAEP1-C247A-aa55-351 exhibits appreciable catalytic activities across a wide pH range, and the addition of the Fe3+ metal ion effectively enhances the catalytic power. Importantly, this study provides insight into the recognition and nucleophile peptide profiles of OaAEP1-C247A-aa55-351. The ligase demonstrates a higher recognition ability for the "Asn-Ala-Leu" motif and an N-terminus "Arg-Leu" as nucleophiles, which significantly increases the reaction yield. Consequently, the catalytic activity of OaAEP1-C247A-aa55-351 with highly efficient recognition and nucleophile motif, "Asn-Ala-Leu" and "Arg-Leu" under the buffer containing Fe3+ is 70-fold and 2-fold higher than previously reported OaAEP1-C247A and the most efficient butelase-1, respectively. Thus, the designed OaAEP1-C247A-aa55-351, with its highly efficient recognition and alternative nucleophile options, holds promising potential for applications in protein engineering, chemo-enzymatic modification, and the development of drugs.

Orchestrated Codelivery of Peptide Antigen and Adjuvant to Antigen-Presenting Cells by Using an Engineered Chimeric Peptide Enhances Antitumor T-Cell Immunity.

The intrinsic pharmacokinetic limitations of traditional peptide-based cancer vaccines hamper effective cross-presentation and codelivery of antigens (Ag) and adjuvants, which are crucial for inducing robust antitumor CD8+ T-cell responses. In this study, we report the development of a versatile strategy that simultaneously addresses the different pharmacokinetic challenges of soluble subunit vaccines composed of Ags and cytosine-guanosine oligodeoxynucleotide (CpG) to modulate vaccine efficacy via translating an engineered chimeric peptide, eTAT, as an intramolecular adjuvant. Linking Ags to eTAT enhanced cytosolic delivery of the Ags. This, in turn, led to improved activation and lymph node-trafficking of Ag-presenting cells and Ag cross-presentation, thus promoting Ag-specific T-cell immune responses. Simple mixing of eTAT-linked Ags and CpG significantly enhanced codelivery of Ags and CpG to the Ag-presenting cells, and this substantially augmented the adjuvant effect of CpG, maximized vaccine immunogenicity, and elicited robust and durable CD8+ T-cell responses. Vaccination with this formulation altered the tumor microenvironment and exhibited potent antitumor effects, with generally further enhanced therapeutic efficacy when used in combination with anti-PD1. Altogether, the engineered chimeric peptide-based orchestrated codelivery of Ag and adjuvant may serve as a promising but simple strategy to improve the efficacy of peptide-based cancer vaccines.

Pattern of multiple human papillomavirus infection and type competition: An analysis in healthy Chinese women aged 18-45 years.

To assess the pattern of multiple human papillomavirus infection to predict the type replacement postvaccination. A total of 7372 women aged 18-45y from a phase III trial of an Escherichia coli-produced HPV-16/18 vaccine were analyzed at enrollment visit before vaccination. Hierarchical multilevel logistic regression was used to evaluate HPV vaccine type and nonvaccine-type interactions with age as a covariate. Binary logistic regression was construed to compare multiple infections with single infections to explore the impact of multiple-type infections on the risk of cervical disease. Multiple HPV infections were observed in 25.2% of HPV-positive women and multiple infections were higher than expected by chance. Statistically significant negative associations were observed between HPV16 and 52, HPV18 and HPV51/52/58, HPV31 and HPV39/51/52/53/54/58, HPV33 and HPV52/58, HPV58 and HPV52, HPV6 and HPV 39/51/52/53/54/56/58. Multiple HPV infections increased the risk of CIN2+ and HSIL+, with the ORs of 2.27(95%CI: 1.41, 3.64) and 2.26 (95%CI: 1.29, 3.95) for multiple oncogenic HPV infection separately. However, no significant evidence for the type-type interactions on risk of CIN2+ or HSIL+. There is possibility of type replacement between several pairs of vaccine and nonvaccine HPV type. Multiple HPV infection increased the risk of cervical disease, but coinfection HPV types seem to follow independent disease processes. Continued post-vaccination surveillance for HPV 51/52/58 types and HPV 39/51 types separately was essential after the first and second generation of HPV vaccination implementation in China.

A non-viral DNA delivery system consisting of multifunctional chimeric peptide fused with zinc-finger protein.

Non-viral gene delivery systems have received sustained attention as a promising alternative to viral vectors for disease treatment and prevention in recent years. Numerous methods have been developed to enhance gene uptake and delivery in the cytoplasm; however, due to technical difficulties and delivery efficiency, these systems still face challenges in a range of biological applications, especially in vivo. To alleviate this challenge, we devised a novel system for gene delivery based on a recombinant protein eTAT-ZF9-NLS, which consisted of a multifunctional chimeric peptide and a zinc-finger protein with sequence-specific DNA-binding activity. High transfection efficiency was observed in several mammalian cells after intracellular delivery of plasmid containing ZF9-binding sites mediated by eTAT-ZF9-NLS. Our new approach provides a novel transfection strategy and the transfection efficiency was confirmed both in vitro and in vivo, making it a preferential transfection reagent for possible gene therapy.

Immunogenicity and safety of an ORF7-deficient skin-attenuated and neuro-attenuated live vaccine for varicella: a randomised, double-blind, controlled, phase 2a trial.

BACKGROUND: The Oka varicella vaccine strain remains neurovirulent and can establish lifelong latent infection, raising safety concerns about vaccine-related herpes zoster. In this study, we aimed to evaluate the immunogenicity and safety of a skin-attenuated and neuro-attenuated varicella vaccine candidate (v7D vaccine). METHODS: We did this randomised, double-blind, controlled, phase 2a clinical trial in Jiangsu, China. Healthy children aged 3-12 years with no history of varicella infection or vaccination were enrolled and randomly assigned (1:1:1:1) to receive a single subcutaneous injection of the v7D vaccine at 3·3 log10 plaque forming units (PFU; low-dose v7D group), 3·9 log10 PFU (medium-dose v7D group), and 4·2 log10 PFU (high-dose v7D group), or the positive control varicella vaccine (vOka vaccine group). All the participants, laboratory personnel, and investigators other than the vaccine preparation and management staff were masked to the vaccine allocation. The primary outcome was assessment of the geometric mean titres (GMTs) and seroconversion rates of anti-varicella zoster virus immunoglobulin G (IgG) induced by different dose groups of v7D vaccine at 0, 42, 60, and 90 days after vaccination in the per-protocol set for humoral immune response analysis. Safety was a secondary outcome, focusing on adverse events within 42 days post-vaccination, and serious adverse events within 6 months after vaccination. This study was registered on Chinese Clinical Trial Registry, ChiCTR2000034434. FINDINGS: On Aug 18-21, 2020, 842 eligible volunteers were enrolled and randomly assigned treatment. After three participants withdrew, 839 received a low dose (n=211), middle dose (n=210), or high dose (n=210) of v7D vaccine, or the vOka vaccine (n=208). In the per-protocol set for humoral immune response analysis, the anti-varicella zoster virus IgG antibody response was highest at day 90. At day 90, the seroconversion rates of the low-dose, medium-dose, and high-dose groups of v7D vaccine and the positive control vOka vaccine group were 100·0% (95% CI 95·8-100·0; 87 of 87 participants), 98·9% (93·8-100·0; 87 of 88 participants), 97·8% (92·4-99·7; 91 of 93 participants), and 96·4% (89·8-99·2; 80 of 83 participants), respectively; the GMTs corresponded to values of 30·8 (95% CI 26·2-36·0), 31·3 (26·7-36·6), 28·2 (23·9-33·2), and 38·5 (31·7-46·7). The v7D vaccine, at low dose and medium dose, elicited a humoral immune response similar to that of the vOka vaccine. However, the high-dose v7D vaccine induced a marginally lower GMT compared with the vOka vaccine at day 90 (p=0·027). In the per-protocol set, the three dose groups of the v7D vaccine induced a similar humoral immune response at each timepoint, with no statistically significant differences. The incidence of adverse reactions in the low-dose, medium-dose, and high-dose groups of v7D vaccine was significantly lower than that in the vOka vaccine group (17% [35 of 211 participants], 20% [41 of 210 participants], and 13% [27 of 210 participants] vs 24% [50 of 208 participants], respectively; p=0·025), especially local adverse reactions (10% [22 of 211 participants], 14% [30 of 210 participants] and 9% [18 of 210 participants] vs 18% [38 of 208 participants], respectively; p=0·016). None of the serious adverse events were vaccine related. INTERPRETATION: The three dose groups of the candidate v7D vaccine exhibit similar humoral immunogenicity to the vOka vaccine and are well tolerated. These findings encourage further investigations on two-dose vaccination schedules, efficacy, and the potential safety benefit of v7D vaccine in the future. FUNDING: The National Natural Science Foundation of China, CAMS Innovation Fund for Medical Sciences, the Fundamental Research Funds for the Central Universities, and Beijing Wantai. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.

Wa-VP4* as a candidate rotavirus vaccine induced homologous and heterologous virus neutralizing antibody responses in mice, pigs, and cynomolgus monkeys.

Group A rotavirus (RVA) is the primary etiological agent of acute gastroenteritis (AGE) in children under 5 years of age. Despite the global implementation of vaccines, rotavirus infections continue to cause over 120,000 deaths annually, with a majority occurring in developing nations. Among infants, the P[8] rotavirus strain is the most prevalent and can be categorized into four distinct lineages. In this investigation, we expressed five VP4(aa26-476) proteins from different P[8] lineages of human rotavirus in E. coli and assessed their immunogenicity in rabbits. Among the different P[8] strains, the Wa-VP4 protein, derived from the MT025868.1 strain of the P[8]-1 lineage, exhibited successful purification in a highly homogeneous form and significantly elicited higher levels of neutralizing antibodies (nAbs) against both homologous and heterologous rotaviruses compared to other VP4 proteins derived from different P[8] lineages in rabbits. Furthermore, we assessed the immunogenicity of the Wa-VP4 protein in mice, pigs, and cynomolgus monkeys, observing that it induced robust production of nAbs in all animals. Interestingly, there was no significant difference between in nAb titers against homologous and heterologous rotaviruses in pigs and mankeys. Collectively, these findings suggest that the Wa-VP4* protein may serve as a potential candidate for a rotavirus vaccine.

Immune landscape and response to oncolytic virus-based immunotherapy.

Oncolytic virus (OV)-based immunotherapy has emerged as a promising strategy for cancer treatment, offering a unique potential to selectively target malignant cells while sparing normal tissues. However, the immunosuppressive nature of tumor microenvironment (TME) poses a substantial hurdle to the development of OVs as effective immunotherapeutic agents, as it restricts the activation and recruitment of immune cells. This review elucidates the potential of OV-based immunotherapy in modulating the immune landscape within the TME to overcome immune resistance and enhance antitumor immune responses. We examine the role of OVs in targeting specific immune cell populations, including dendritic cells, T cells, natural killer cells, and macrophages, and their ability to alter the TME by inhibiting angiogenesis and reducing tumor fibrosis. Additionally, we explore strategies to optimize OV-based drug delivery and improve the efficiency of OV-mediated immunotherapy. In conclusion, this review offers a concise and comprehensive synopsis of the current status and future prospects of OV-based immunotherapy, underscoring its remarkable potential as an effective immunotherapeutic agent for cancer treatment.

Construction and characterization of an infectious cDNA clone of human rhinovirus A89.

Rhinoviruses (RVs) are major causes of the common cold and are related to severe respiratory tract diseases, leading to a considerable economic burden and impacts on public health. Available and stable viral resources of rhinoviruses for laboratory use are important for promoting studies on rhinoviruses and further vaccine or therapeutic drug development. Reverse genetic technology can be useful to produce rhinoviruses and will help to promote studies on their pathogenesis and virulence. In this study, rhinovirus A89, an RV-A species that has been found to be highly involved in hospitalization triggered by RV infections, was selected to construct an infectious clone based on its sequence as a representative. The viral mRNA produced by a T7 RNA transcript system was transfected into H1-HeLa cells, and the rescued RV-A89 viruses were harvested and confirmed by sequencing. The rescued RV-A89 induced a similar cytopathic effect (CPE) and shared almost identical growth kinetics curves with parental RV-A89. Moreover, 9A7, a prescreened monoclonal antibody against the parental RV-A89, had a good and specific reaction with the rescued RV-A89, and further characterization showed almost the same morphology and protein composition of both viruses; thus, recombinant RV-A89 with similar biological characterization and virulence to the parental virus was obtained. In summary, the infectious clone of RV-A89 was successfully established, and the development of reverse genetic technology for rhinovirus will provide a framework for further studies on rhinoviruses.

Swine pseudorabies virus attenuated vaccine reprograms the kidney cancer tumor microenvironment and synergizes with PD-1 blockade.

The global incidence rate of kidney cancer (KC) has been steadily increasing over the past 30 years. With the aging global population, kidney cancer has become an escalating concern that necessitates vigilant surveillance. Nowadays, surgical intervention remains the optimal therapeutic approach for kidney cancer, while the availability of efficacious treatments for advanced tumors remains limited. Oncolytic viruses, an emerging form of immunotherapy, have demonstrated encouraging anti-neoplastic properties and are progressively garnering public acceptance. However, research on oncolytic viruses in kidney cancer is relatively limited. Furthermore, given the high complexity and heterogeneity of kidney cancer, it is crucial to identify an optimal oncolytic virus agent that is better suited for its treatment. The present study investigates the oncolytic activity of the Pseudorabies virus live attenuated vaccine (PRV-LAV) against KC. The findings clearly demonstrate that PRV-LAV exhibits robust oncolytic activity targeting KC cell lines. Furthermore, the therapeutic efficacy of PRV-LAV was confirmed in both a subcutaneous tumor-bearing nude mouse model and a syngeneic mouse model of KC. Combined RNA-seq analysis and flow cytometry revealed that PRV-LAV treatment substantially enhances the infiltration of a diverse range of lymphocytes, including T cells, B cells, macrophages, and NK cells. Additionally, PRV-LAV treatment enhances T cell activation and exerts antitumor effects. Importantly, the combination of PRV-LAV with anti-PD-1 antibodies, an approved drug for KC treatment, synergistically enhances the efficacy against KC. Overall, the discovery of PRV-LAV as an effective oncolytic virus holds significant importance for improving the treatment efficacy and survival rates of KC patients.

Evolution of the SARS-CoV-2 Omicron Variants: Genetic Impact on Viral Fitness.

Over the last three years, the pandemic of COVID-19 has had a significant impact on people's lives and the global economy. The incessant emergence of variant strains has compounded the challenges associated with the management of COVID-19. As the predominant variant from late 2021 to the present, Omicron and its sublineages, through continuous evolution, have demonstrated iterative viral fitness. The comprehensive elucidation of the biological implications that catalyzed this evolution remains incomplete. In accordance with extant research evidence, we provide a comprehensive review of subvariants of Omicron, delineating alterations in immune evasion, cellular infectivity, and the cross-species transmission potential. This review seeks to clarify the underpinnings of biology within the evolution of SARS-CoV-2, thereby providing a foundation for strategic considerations in the post-pandemic era of COVID-19.

Therapeutic vaccine-induced plasma cell differentiation is defective in the presence of persistently high HBsAg levels.

BACKGROUND & AIMS: Mechanisms behind the impaired response of antigen-specific B cells to therapeutic vaccination in chronic hepatitis B virus (HBV) infection remain unclear. The development of vaccines or strategies to overcome this obstacle is vital for advancing the management of chronic hepatitis B. METHODS: A mouse model, denominated as E6F6-B, was engineered to feature a knock-in of a B-cell receptor (BCR) that specifically recognizes HBsAg. This model served as a valuable tool for investigating the temporal and spatial dynamics of humoral responses following therapeutic vaccination under continuous antigen exposure. Using a suite of immunological techniques, we elucidated the differentiation trajectory of HBsAg-specific B cells post-therapeutic vaccination in HBV carrier mice. RESULTS: Utilizing the E6F6-B transfer model, we observed a marked decline in antibody-secreting cells 2 weeks after vaccination. A dysfunctional and atypical pre-plasma cell population (BLIMP-1+ IRF4+ CD40- CD138- BCMA-) emerged, manifested by sustained BCR signaling. By deploying an antibody to purge persistent HBsAg, we effectively prompted the therapeutic vaccine to provoke conventional plasma cell differentiation. This resulted in an enhanced anti-HBs antibody response and facilitated HBsAg clearance. CONCLUSIONS: Sustained high levels of HBsAg limit the ability of therapeutic hepatitis B vaccines to induce the canonical plasma cell differentiation necessary for anti-HBs antibody production. Employing a strategy combining antibodies with vaccines can surmount this altered humoral response associated with atypical pre-plasma cells, leading to improved therapeutic efficacy in HBV carrier mice. IMPACT AND IMPLICATIONS: Therapeutic vaccines aimed at combatting HBV encounter suboptimal humoral responses in clinical settings, and the mechanisms impeding their effectiveness have remained obscure. Our research, utilizing the innovative E6F6-B mouse transfer model, reveals that the persistence of HBsAg can lead to the emergence of an atypical pre-plasma cell population, which proves to be relevant to the potency of therapeutic HBV vaccines. Targeting the aberrant differentiation process of these atypical pre-plasma cells stands out as a critical strategy to amplify the humoral response elicited by HBV therapeutic vaccines in carrier mouse models. This discovery suggests a compelling avenue for further study in the context of human chronic hepatitis B. Encouragingly, our findings indicate that synergistic therapy combining HBV-specific antibodies with vaccines offers a promising approach that could significantly advance the pursuit of a functional cure for HBV.