Correlation of HBV Core-Related Antigen with Conventional Serologic Indicators of Hepatitis B and Disease Stage.

BACKGROUND: Hepatitis B caused by hepatitis B virus (HBV) infection is a serious global public health issue. Currently, serological indicators serve as important markers for the diagnosis of hepatitis B. It has been found that HBV core-related antigen (HBcrAg) correlates well with intrahepatic cccDNA, intrahepatic HBV DNA, serum HBV DNA, and hepatitis B e antigen (HBeAg). To provide a more reliable basis for the diagnosis and treatment of hepatitis B, we explored the correlation between HBcrAg and conventional serologic testing indicators and disease staging. METHODS: Five hundred forty-two patient serum samples were collected at the First Affiliated Hospital of Soochow University from November 2021 to March 2022. The serum HBcrAg was measured by the magnetic particle chemiluminescence method in addition with other serum indicators. RESULTS: HBcrAg statistically correlated with HBV DNA level (r = 0.655, p < 0.001) and HBeAg level (r = 0.945, p < 0.001. The mean HBcrAg levels in the immune-tolerant and immune-clearance phases were significantly higher than those in the immunologic-control phase and the reactivation phase. This study demonstrated that serum HBcrAg positively correlated with serum HBV DNA and HBeAg. Even in cases where HBV DNA and HBeAg are negative, there is still a higher positivity rate of HBcrAg in hepatitis B patients. CONCLUSIONS: HBcrAg is a reliable serum marker to avoid underdiagnosis of occult HBV infection.

Impact of Hepatitis B Surface and Core Antibody Levels on Hepatitis B Virus Reactivation.

Hepatitis B virus reactivation (HBV-R) is a serious complication that can occur in patients with resolved HBV infection during cancer chemotherapy. We examined the levels of HBV surface antibody (HBsAb) and HBV core antibody (HBcAb) to assess the incidence of HBV-R in cancer patients including hematopoietic stem cell transplantation (HSCT) and rituximab administration. This retrospective cohort study included 590 patients with resolved HBV infection. The incidence of HBV-R was evaluated 761.5 (range, 90-3898) days after the inititiation of chemotherapy. Of the patients, 13 (2.2%) developed HBV-R after the start of chemotherapy. All 13 patients exhibited lower HBsAb (<100 mIU/mL) levels at baseline. A higher level of HBcAb (≥100 cut off index (C.O.I.)) was a possible risk factor for HBV-R as well as HSCT and rituximab administration. The simultaneous presence of HBsAb <100 mIU/mL and HBcAb ≥100 C.O.I. increased the risk of HBV-R by 18.5%. Patients treated with rituximab were at a higher risk of HBV-R (18.4%) despite having HBcAb <100 C.O.I. Our results suggest that assessment of HBsAb and HBcAb levels prior to the chemotherapy is important for identifying patients at high risk of HBV-R, especially in solid cancers without HSCT and rituximab administration.

A potential antiviral role for CCR5+CD8+ T cells in children with hepatitis B.

While dysfunctional exhausted CD8+ T cells hamper viral control when children acquire hepatitis B virus (HBV) infection, it's crucial to recognize that CD8+ T cells have diverse phenotypes and functions. This study explored a subset of CD8+ T cells expressing C-C chemokine receptor type 5 (CCR5) in children with HBV infection. Thirty-six patients in the immune tolerant group, 33 patients in the immune active group, 55 patients in the combined response group, and 22 healthy control children were enrolled. The frequency, functional molecules, and effector functions of the CCR5+CD8+ T cell population in different groups were evaluated. The frequency of CCR5+CD8+ T cells correlated positively with the frequency of CCR5+CD4+ T cells and patient age, and it correlated negatively with alanine aminotransferase, aspartate transaminase, HBV DNA, hepatitis B surface antigen, and lactic dehydrogenase levels. CCR5+CD8+ T cells had higher levels of inhibitory and activated receptors and produced higher levels of IFN-γ, IL-2, and TNF-α than CCR5-CD8+ T cells. CCR5+CD8+ T cells were partially exhausted but possessed a stronger antiviral activity than CCR5-CD8+ T cells. The identification of this subset increases our understanding of CD8+ T cell functions and serves as a potential immunotherapeutic target for children with HBV infection.

Hepatitis B: Model Systems and Therapeutic Approaches.

Hepatitis B virus (HBV) infection is a major global health issue and ranks among the top causes of liver cirrhosis and hepatocellular carcinoma. Although current antiviral medications, including nucleot(s)ide analogs and interferons, could inhibit the replication of HBV and alleviate the disease, HBV cannot be fully eradicated. The development of cellular and animal models for HBV infection plays an important role in exploring effective anti-HBV medicine. During the past decades, advancements in several cell culture systems, such as HepG2.2.15, HepAD38, HepaRG, hepatocyte-like cells, and primary human hepatocytes, have propelled the research in inhibiting HBV replication and expression and thus enriched our comprehension of the viral life cycle and enhancing antiviral drug evaluation efficacy. Mouse models, in particular, have emerged as the most extensively studied HBV animal models. Additionally, the present landscape of HBV therapeutics research now encompasses a comprehensive assessment of the virus's life cycle, targeting numerous facets and employing a variety of immunomodulatory approaches, including entry inhibitors, strategies aimed at cccDNA, RNA interference technologies, toll-like receptor agonists, and, notably, traditional Chinese medicine (TCM). This review describes the attributes and limitations of existing HBV model systems and surveys novel advancements in HBV treatment modalities, which will offer deeper insights toward discovering potentially efficacious pharmaceutical interventions.

Highlights from the 2023 International Meeting on the Molecular Biology of Hepatitis B virus.

Since its discovery in 1965, our understanding of the hepatitis B virus (HBV) replication cycle and host immune responses has increased markedly. In contrast, our knowledge of the molecular biology of hepatitis delta virus (HDV), which is associated with more severe liver disease, is less well understood. Despite the progress made, critical gaps remain in our knowledge of HBV and HDV replication and the mechanisms underlying viral persistence and evasion of host immunity. The International HBV Meeting is the leading annual scientific meeting for presenting the latest advances in HBV and HDV molecular virology, immunology, and epidemiology. In 2023, the annual scientific meeting was held in Kobe, Japan and this review summarises some of the advances presented at the Meeting and lists gaps in our knowledge that may facilitate the development of new therapies.

CARs derived from broadly neutralizing, human monoclonal antibodies identified by single B cell sorting target hepatitis B virus-positive cells.

To design new CARs targeting hepatitis B virus (HBV), we isolated human monoclonal antibodies recognizing the HBV envelope proteins from single B cells of a patient with a resolved infection. HBV-specific memory B cells were isolated by incubating peripheral blood mononuclear cells with biotinylated hepatitis B surface antigen (HBsAg), followed by single-cell flow cytometry-based sorting of live, CD19+ IgG+ HBsAg+ cells. Amplification and sequencing of immunoglobulin genes from single memory B cells identified variable heavy and light chain sequences. Corresponding immunoglobulin chains were cloned into IgG1 expression vectors and expressed in mammalian cells. Two antibodies named 4D06 and 4D08 were found to be highly specific for HBsAg, recognized a conformational and a linear epitope, respectively, and showed broad reactivity and neutralization capacity against all major HBV genotypes. 4D06 and 4D08 variable chain fragments were cloned into a 2nd generation CAR format with CD28 and CD3zeta intracellular signaling domains. The new CAR constructs displayed a high functional avidity when expressed on primary human T cells. CAR-grafted T cells proved to be polyfunctional regarding cytokine secretion and killed HBV-positive target cells. Interestingly, background activation of the 4D08-CAR recognizing a linear instead of a conformational epitope was consistently low. In a preclinical model of chronic HBV infection, murine T cells grafted with the 4D06 and the 4D08 CAR showed on target activity indicated by a transient increase in serum transaminases, and a lower number of HBV-positive hepatocytes in the mice treated. This study demonstrates an efficient and fast approach to identifying pathogen-specific monoclonal human antibodies from small donor cell numbers for the subsequent generation of new CARs.

Case report: Recombinant human type II tumour necrosis factor receptor-antibody fusion protein induced occult hepatitis B virus reactivation leading to liver failure.

Recombinant human type II tumour necrosis factor receptor-antibody fusion protein (rh TNFR:Fc) is an immunosuppressant approved for treating rheumatoid arthritis (RA). This case report describes a case of hepatitis B reactivation in a patient with drug-induced acute-on-chronic liver failure. A 58-year-old woman with a history of RA was treated with rh TNFR:Fc; and then subsequently received 25 mg rh TNFR:Fc, twice a week, as maintenance therapy. No anti-hepatitis B virus (HBV) preventive treatment was administered. Six months later, she was hospitalized with acute jaundice. HBV reactivation was observed, leading to acute-on-chronic liver failure. After active treatment, the patient's condition improved and she recovered well. Following careful diagnosis and treatment protocols are essential when treating RA with rh TNFR:Fc, especially in anti-hepatitis B core antigen antibody-positive patients, even when the HBV surface antigen and the HBV DNA are negative. In the case of HBV reactivation, liver function parameters, HBV surface antigen and HBV DNA should be closely monitored during treatment, and antiviral drugs should be used prophylactically when necessary, as fatal hepatitis B reactivation may occur in rare cases. A comprehensive evaluation and medication should be administered in a timely manner after evaluating the patient's physical condition and closely monitoring the patient.

Comparison of the effectiveness four years after Homo/Hetero prime-boost with 10 µg HP and 20 µg CHO recombinant hepatitis B vaccine at 1 and 6 months in maternal HBsAg-negative children.

Introduction: Limited data were available on the effectivenessfour years after Homo or Hetero prime-boost with 10 µg Hansenulapolymorpha recombinant hepatitis B vaccine (HepB-HP) and 20 µgChinese hamster ovary cell HepB (HepB-CHO). Methods: A crosssectional study was performed in maternalhepatitis B surface antigen (HBsAg)-negative children whoreceived one dose of 10 µg HepB-HP at birth, Homo or Heteroprime-boost with 10 µg HepB-HP and 20 µg HepB-CHO at 1 and 6months. HBsAg and hepatitis B surface antibody (anti-HBs) fouryears after immunization were quantitatively detected by achemiluminescent microparticle immunoassay (CMIA). Results: A total of 359 children were included; 119 childrenreceived two doses of 10 µg HepB-HP and 120 children receivedtwo doses of 20 µg HepB-CHO, called Homo prime-boost; 120children received Hetero prime-boost with 10 µg HepB-HP and 20µg HepB-CHO. All children were HBsAg negative. The geometricmean concentration (GMC) and overall seropositivity rate (SPR) ofanti-HBs were 59.47 (95%CI: 49.00 - 72.16) mIU/ml and 85.51%(307/359). Nearly 15% of the study subjects had an anti-HBsconcentration < 10 mIU/ml and 5.01% had an anti-HBsconcentration ≤ 2.5 mIU/ml. The GMC of the 20 µg CHO Homoprime-boost group [76.05 (95%CI: 54.97 - 105.19) mIU/ml] washigher than that of the 10 µg HP Homo group [45.86 (95%CI:31.94 - 65.84) mIU/ml] (p = 0.035). The GMCs of the Heteroprime-boost groups (10 µg HP-20 µg CHO and 20 µg CHO-10 µgHP) were 75.86 (95% CI: 48.98 - 107.15) mIU/ml and 43.65(95%CI: 27.54 - 69.18) mIU/ml, respectively (p = 0.041). Aftercontrolling for sex influence, the SPR of the 20 µg CHO Homoprime-boost group was 2.087 times than that of the 10 µg HPHomo group. Discussion: The HepB booster was not necessary in the generalchildren, Homo/Hetero prime-boost with 20 µg HepB-CHO wouldincrease the anti-HBs concentration four years after immunization,timely testing and improved knowledge about the self-pay vaccinewould be good for controlling hepatitis B.

The lack of HBsAg secretion does neither facilitate induction of antiviral T cell responses nor Hepatitis B Virus clearance in mice.

Immune tolerance to the hepatitis B virus (HBV) is crucial for developing chronic hepatitis B, and the HBV surface antigen (HBsAg) produced and secreted in high amounts is regarded as a key contributor. HBsAg is expressed in HBV-infected hepatocytes and those carrying an HBV integration. Whether either HBsAg secretion or the high antigen amount expressed in the liver determines its immunomodulatory properties, however, remains unclear. We, therefore, developed a novel HBV animal model that allowed us to study the role of secreted HBsAg. We introduced a previously described HBs mutation, C65S, abolishing HBsAg secretion into a replication-competent 1.3-overlength HBV genome and used adeno-associated virus vectors to deliver it to the mouse liver. The AAV-HBV established a carrier state of wildtype and C65S mutant HBV, respectively. We investigated antiviral B- and T-cell immunity in the HBV-carrier mice after therapeutic vaccination. Moreover, we compared the effect of a lacking HBsAg secretion with that of an antiviral siRNA. While missing HBsAg secretion allowed for higher levels of detectable anti-HBs antibodies after therapeutic vaccination, it did neither affect antiviral T-cell responses nor intrahepatic HBV gene expression, irrespective of the starting level. A treatment with HBV siRNA restricting viral antigen expression within hepatocytes, however, improved the antiviral efficacy of therapeutic vaccination, irrespective of the ability of HBV to secrete HBsAg. Our data indicate that clearing HBsAg from blood cannot significantly impact HBV persistence or T-cell immunity. This indicates that a restriction of hepatic viral antigen expression will be required to break HBV immunotolerance.

Current and future use of antibody-based passive immunity to prevent or control HBV/HDV infections.

With the increasing momentum and success of monoclonal antibody therapy in conventional medical practices, there is a revived emphasis on the development of monoclonal antibodies targeting the hepatitis B surface antigen (anti-HBs) for the treatment of chronic hepatitis B (HBV) and hepatitis D (HDV). Combination therapies of anti-HBs monoclonal antibodies, and novel anti-HBV compounds and immunomodulatory drugs presenting a promising avenue to enhanced therapeutic outcomes in HBV/HDV cure regimens. In this review, we will cover the role of antibodies in the protection and clearance of HBV infection, the association of anti-HBV surface antigen antibodies (anti-HBs) in protection against HBV and how antibody effector functions, beyond neutralization, are likely necessary. Lastly, we will review clinical data from previous and ongoing clinical trials of passive antibody therapy to provide a state-of-the-are perspective on passive antibody therapies in combinations with additional novel agents.

Dismantling Barriers to Hepatitis B and Delta Screening, Prevention, and Linkage to Care among the PWUD Community in Philadelphia.

The prevalence of hepatitis B and delta viruses (HBV/HDV) among people who use drugs (PWUD) remains largely unknown. In the context of one Philadelphia-based harm reduction organization (HRO), this study aimed to assess HBV/HDV prevalence and facilitate linkage to care. Participants completed a demographic HBV/HDV risk factor survey and were screened for HBV and reflexively for HDV if positive for HBV surface antigen or isolated core antibody. Fisher's exact tests and regression were used to understand relationships between risks and HBV blood markers. Of the 498 participants, 126 (25.3%) did not have hepatitis B immunity, 52.6% had been vaccinated against HBV, and 17.9% had recovered from a past infection. Eleven (2.2%) participants tested positive for isolated HBV core antibody, 10 (2.0%) for HBV surface antigen, and one (0.2%) for HDV antibody. History of incarceration was associated with current HBV infection, while transactional sex and experience of homelessness were predictive of previous exposure. This study found high rates of current and past HBV infection, and a 10% HBV/HDV co-infection rate. Despite availability of vaccine, one quarter of participants remained vulnerable to infection. Findings demonstrate the need to improve low-threshold HBV/HDV screening, vaccination, and linkage to care among PWUD. The study also identified gaps in the HBV/HDV care cascade, including lack of point-of-care diagnostics and lack of support for HROs to provide HBV services.

Class A capsid assembly modulator apoptotic elimination of hepatocytes with high HBV core antigen level in vivo is dependent on de novo core protein translation.

Capsid assembly is critical in the hepatitis B virus (HBV) life cycle, mediated by the viral core protein. Capsid assembly is the target for new anti-viral therapeutics known as capsid assembly modulators (CAMs) of which the CAM-aberrant (CAM-A) class induces aberrant shaped core protein structures and leads to hepatocyte cell death. This study aimed to identify the mechanism of action of CAM-A modulators leading to HBV-infected hepatocyte elimination where CAM-A-mediated hepatitis B surface antigen (HBsAg) reduction was evaluated in a stable HBV replicating cell line and in AAV-HBV-transduced C57BL/6, C57BL/6 SCID, and HBV-infected chimeric mice with humanized livers. Results showed that in vivo treatment with CAM-A modulators induced pronounced reductions in hepatitis B e antigen (HBeAg) and HBsAg, associated with a transient alanine amino transferase (ALT) increase. Both HBsAg and HBeAg reductions and ALT increase were delayed in C57BL/6 SCID and chimeric mice, suggesting that adaptive immune responses may indirectly contribute. However, CD8+ T cell depletion in transduced wild-type mice did not impact antigen reduction, indicating that CD8+ T cell responses are not essential. Transient ALT elevation in AAV-HBV-transduced mice coincided with a transient increase in endoplasmic reticulum stress and apoptosis markers, followed by detection of a proliferation marker. Microarray data revealed antigen presentation pathway (major histocompatibility complex class I molecules) upregulation, overlapping with the apoptosis. Combination treatment with HBV-specific siRNA demonstrated that CAM-A-mediated HBsAg reduction is dependent on de novo core protein translation. To conclude, CAM-A treatment eradicates HBV-infected hepatocytes with high core protein levels through the induction of apoptosis, which can be a promising approach as part of a regimen to achieve functional cure. IMPORTANCE: Treatment with hepatitis B virus (HBV) capsid assembly modulators that induce the formation of aberrant HBV core protein structures (CAM-A) leads to programmed cell death, apoptosis, of HBV-infected hepatocytes and subsequent reduction of HBV antigens, which differentiates CAM-A from other CAMs. The effect is dependent on the de novo synthesis and high levels of core protein.

Modulation of CD8+T cells, NK cells and Th1cytokines by metabolic milieu in decline of HBV-viremia in pregnant women treated with tenofovir-disoproxil from second trimester of pregnancy.

High HBV DNA levels predispose to mother to child transmission (MTCT) of HBV. Early nucleotide analogue (NA) therapy can reduce HBV DNA and minimize MTCT. We analysed immune-metabolic profile in pregnant mothers who received NA from 2nd trimester compared with untreated mothers. In 2nd trimester, there was no difference in immune profiles between Gr.1 and Gr.2 but high viral load women had downregulated pyruvate, NAD+ metabolism but in 3rd trimester, Gr.1 had significant reduction in HBV-DNA, upregulated pyruvate and NAD with increased IFN-2αA, CD8Tcells, NK cells and decreased Tregs, IL15, IL18, IL29, TGFß3 compared to Gr.2. In Gr.1, three eAg-ve women showed undetectable DNA and HBsAg. At delivery, Gr.1 showed no MTCT, with undetectable HBV DNA, HBsAg, high CD8 and NK cells in two women. We conclude, that starting NA from second trimester, reduces HBV load and MTCT, modulates NAD, induces immunity and suggest use of NA in early gestation in future trials.

Activation of AIM2 by hepatitis B virus results in antiviral immunity that suppresses hepatitis C virus during coinfection.

IMPORTANCE: Clinical data suggest that Hepatitis C virus (HCV) levels are generally lower in Hepatitis B virus (HBV) co-infected patients, but the mechanism is unknown. Here, we show that HBV, but not HCV, activated absent in melanoma-2. This in turn results in inflammasome-mediated cleavage of pro-IL-18, leading to an innate immune activation cascade that results in increased interferon-γ, suppressing both viruses.

Biologic Therapy Carries a Very Low Risk of Reactivation in Hepatitis B Surface Antigen-Negative Phase of Hepatitis B.

BACKGROUND: The risk of hepatitis B reactivation in hepatitis B surface antigen-negative phase of hepatitis B virus-infected patients exposed to biologic agents is not clear. We aimed to investigate the reactivation rate in hepatitis B surface antigen-negative phase of hepatitis B virus-infected patients after biologic therapy. METHODS: Patients followed at gastroenterology, rheumatology, and dermatology clinics with a diagnosis of immune-mediated inflam matory diseases were screened. Immune-mediated inflammatory diseases patients exposed to biologic agents with a negative hepatitis B surface antigen and positive hepatitis B core immunoglobulin G antibody were included in the study. RESULTS: We screened 8266 immune-mediated inflammatory disease patients, and 2484 patients were identified as exposed to biologic agents. Two hundred twenty-one patients were included in the study. The mean age was 54.08 ± 11.69 years, and 115 (52.0%) patients were female. The median number of different biologic subtype use was 1 (range: 1-6). The mean biologic agent exposure time was 55 (range: 2-179) months. One hundred and fifty-two (68.8%) patients used a concomitant immunomodulatory agent, and 84 (38.0%) patients were exposed to corticosteroids during biologic use. No hepatitis B reactivation with a reverse seroconversion of hepatitis B surface antigen positivity was seen. Antiviral prophylaxis for hepatitis B was applied to 48 (21.7%) patients. Hepatitis B virus-DNA was screened in 56 (25.3%) patients prior to the biologic exposure. Two patients without antiviral prophylaxis had hepatitis B virus-DNA reactivation with a negative hepatitis B surface antigen during exposure to the biologic agent. CONCLUSION: We found 2 reactivations and no hepatitis B surface antigen seroconversion in our cohort. Antiviral prophylaxis for patients exposed to biologic agents may need to be discussed in more detail.

Interleukin-37 suppresses the cytotoxicity of hepatitis B virus peptides-induced CD8+ T cells in patients with acute hepatitis B.

Interleukin-37 (IL-37) is a newly identified anti-inflammatory cytokine, owning immunosuppressive activity in infectious diseases. The aim of this study was to investigate the regulatory function of IL-37 on CD8+ T cells during hepatitis B virus (HBV) infection. Eighteen acute hepatitis B (AHB) patients, thirty-nine chronic hepatitis B (CHB) patients, and twenty controls were enrolled. IL-37 concentration was measured by ELISA. IL-37 receptor subunits expressions on CD8+ T cells were assessed by flow cytometry. Purified CD8+ T cells were stimulated with HBV peptides and recombinant IL-37. Perforin and granzyme B secretion was investigated by ELISPOT. Programmed death-1 (PD-1) and cytotoxic T-lymphocyte associated protein-4 (CTLA-4) mRNA expressions were semi-quantified by real-time PCR. CD8+ T cell cytotoxicity was assessed in direct contact and indirect contact coculture with HepG2.2.15 cells. Plasma IL-37 level was down-regulated and negatively correlated with aminotransferase levels in AHB patients. There were no significant differences of IL-37 receptor subunits among AHB patients, CHB patients, and controls. Exogenous IL-37 stimulation suppressed HBV peptides-induced perforin and granzyme B secretion by CD8+ T cells in AHB patients, but not in CHB patients. Exogenous IL-37 stimulation did not affect proinflammatory cytokines secretion as well as PD-1/CTLA-4 mRNA expressions in CD8+ T cells in AHB and CHB patients. Exogenous IL-37 stimulation dampened HBV peptide-induced CD8+ T cell cytotoxicity in a cell-to-cell contact manner. The current data indicated that acute HBV infection might induce down-regulation of IL-37, which might be associated with enhanced CD8+ T cell cytotoxicity and liver damage.

Tumor Suppressor p53 Inhibits Hepatitis B Virus Replication by Downregulating HBx via E6AP-Mediated Proteasomal Degradation in Human Hepatocellular Carcinoma Cell Lines.

HBx, a multifunctional regulatory protein, plays an essential role in the replication and pathogenesis of the hepatitis B virus (HBV). In this study, we found that in human hepatoma cells, the tumor suppressor p53 downregulates HBx via ubiquitin-dependent proteasomal degradation. p53 transcriptional activity that results from HBV infection was not essential for this effect. This was shown by treatment with a potent p53 inhibitor, pifithrin-α. Instead, we found that p53 facilitated the binding of E6-associated protein (E6AP), which is an E3 ligase, to HBx and induced E6AP-mediated HBx ubiquitination in a ternary complex of p53, E6AP, and HBx. The ability of p53 to induce E6AP-mediated downregulation of HBx and inhibit HBV replication was demonstrated in an in vitro HBV infection system. This study may provide insights into the regulation of HBx and HBV replication, especially with respect to p53 status, which may also help in understanding HBV-associated tumorigenesis in patients.

Interleukin-33 mediates both immune-related and non-immune-related inhibitory effects against hepatitis B virus.

Chronic infection by hepatitis B virus (HBV) is associated with high risks of liver fibrosis, cirrhosis and hepatocellular carcinoma. HBV covalently closed circular DNA (cccDNA) in the nucleus of infected hepatocyte serves as transcription template. Neither natural resolution of acute infection nor current treatment options for chronic infection are believed to cause cccDNA clearance. We previously showed that injection of IL-33-expressing plasmid facilitated clearance of intrahepatic HBV DNA in a mouse model of HBV persistence. In this work, HBV-targeting therapeutic effects of IL-33 were further explored. Murine IL-33 delivered by recombinant adeno-associated virus (AAV-mIL-33) induced clearance of both serum HBV markers and intrahepatic HBV DNA in two mouse models of HBV persistence based on replicon plasmid and recombinant cccDNA (rcccDNA) respectively. Clearance was associated with serum ALT elevations and liver infiltrations by CD4+ and CD8+ T cells, indicating IL-33-induced cellular immune responses against HBV-harboring cells. Adoptive transfer of splenocytes from AAV-mIL-33-cured mice was indeed sufficient to engender similar clearance in recipient mice. In vitro, intracellular, instead of extracellular, IL-33 was mainly responsible for repressing viral transcription, protein production and genome replication in Huh7 cells transfected with HBV replicon or rcccDNA. IL-33 was shown to be recruited onto rcccDNA minichromosome accompanied by loss of transcriptional activation epigenetic marks. Finally, transfection of IL-33 into HBV-infected HepG2/NTCP cells resulted in reduced transcription, antigen expression and genome replication, suggesting repression of canonical cccDNA. These data demonstrated diverse inhibitory effects on HBV and HBV-infected cells mediated by IL-33, and suggest IL-33 as an interesting therapeutic candidate.