Best practices for screening, testing, diagnosing, and treating patients with hepatitis D (delta) virus based on global expert review and recent guidelines.

Around 15-20 million people develop chronic hepatitis delta virus worldwide. Hepatitis delta virus (HDV) is a defective RNA virus requiring the presence of the hepatitis B virus surface antigen (HBsAg) to complete its life cycle. HDV infects hepatocytes using the hepatitis B virus (HBV) receptor, the sodium taurocholate cotransporting polypeptide (NTCP). The HDV genome is a circular single-stranded RNA which encodes for a single hepatitis delta antigen (HDAg) that exists in two forms (S-HDAg and L-HDAg), and its replication is mediated by the host RNA polymerases. The HBsAg-coated HDV virions contain a ribonucleoprotein (RNP) formed by the RNA genome packaged with small and large HDAg. Farnesylation of the L-HDAg is the limiting step for anchoring this RNP to HBsAg, and thus for assembling, secreting and propagating virion particles. There is an important risk of morbidity and mortality caused by end-stage liver disease and hepatocellular carcinoma with HDV and current treatment is pegylated-interferon (PEG-IFN) for 48weeks with no other options in patients who fail treatment. The ideal goal for HDV treatment is the clearance of HBsAg, but a reasonably achievable goal is a sustained HDV virological response (negative HDV RNA 6months after stopping treatment). New drug development must take into account the interaction of HBV and HDV. In this review, we will present the new insights in the HDV life cycle that have led to the development of novel classes of drugs and discuss antiviral approaches in phase II and III of development: bulevirtide (entry inhibitor), lonafarnib, (prenylation inhibitor) and REP 2139 (HBsAg release inhibitor).

Interferon alpha is the only treatment option for hepatitis delta virus (HDV). Trials investigating the efficacy of pegylated interferon alpha (PEG-IFNa) showed HDV RNA negativity rates of 25-30% 24 weeks after therapy. However, the clinical and virological long-term outcome of HDV-infected patients treated with PEG-IFNa is unknown. We performed a retrospective-prospective follow-up of 77 patients treated for 48 weeks with either PEG-alfa-2a and adefovir (ADV) or either drug alone in the Hep-Net-International-Delta-Hepatitis-Intervention-Study 1 (HIDIT-1) trial. Long-term follow-up data were available for 58 out of 77 patients (75%) with a median time of follow-up of 4.5 (0.5-5.5) years and a median 3 visits per patient. Patients treated with ADV alone received retreatment with PEG-IFNa (48% versus 19%; P = 0.02) more often. Hepatitis B virus surface antigen (HBsAg) became negative in six PEG-IFNa-treated patients until the end of long-term follow-up (10%). Sixteen patients tested HDV RNA-negative 6 months after PEG-IFNa treatment who were entered in the long-term follow-up study. Out of these, nine individuals tested HDV RNA-positive at least once during further long-term follow-up, with seven patients being HDV RNA-positive at the most recent visit. Clinical endpoints (liver-related death, liver transplantation, hepatic decompensation, hepatocellular carcinoma) were observed in three PEG-IFNa-treated (8%) and three ADV-treated (14%) patients during posttreatment long-term follow-up with an overall annual event rate of 2.5% (4.9% in cirrhosis). Sequencing confirmed the reappearance of pretreatment virus strains in all cases. Late HDV RNA relapses may occur after PEG-IFNa therapy of hepatitis delta and thus the term sustained virological response should be avoided in HDV infection. The annual posttreatment rate of clinical events in hepatitis delta patients eligible for PEG-IFNa therapy is about 2.5% and 4.9% in patients with cirrhosis.

Safety and efficacy of REP 2139 and pegylated interferon alfa-2a for treatment-naive patients with chronic hepatitis B virus and hepatitis D virus co-infection (REP 301 and REP 301-LTF): a non-randomised, open-label, phase 2 trial

Large Hepatitis Delta Antigen Modulates Transforming Growth Factor-β Signaling Cascades: Implication of Hepatitis Delta Virus–Induced Liver Fibrosis

Hepatitis delta virus (HDV) is a defective RNA virus that depends on hepatitis B virus (HBV) for its lifecycle. Treatment of chronic HDV infection is difficult as it does not have an enzymatic function as a target, such as polymerases and proteases of HBV and hepatitis C virus. Recently, it has been suggested that farnesyl transferase could be an enzymatic target. Currently, interferon is the only agent against HDV infection. Virological response has risen to 20–47% with pegylated interferon. Monotherapy of nucleos(t)ide analogs are ineffective against the HDV infection, but adefovir and pegylated interferon combination therapy have had some advantages for reduction of HBV surface antigen (HBsAg) levels. Recent studies suggest that measuring HBsAg levels during treatment could be more meaningful than HDV RNA negativity to predict virological response. Prenylation inhibitors that can affect the interactions between the large HDV antigen and HBsAg in the HDV virion are expected treatments for HDV infection. More studies are needed to understand the molecular mechanisms of HDV to manage the disease.

Hepatitis delta virus (HDV) is a defective virus that requires the supply of hepatitis B virus surface antigen (HBsAg) for replication and transmission. We have previously demonstrated that co-transduction of BHK cells with Bac- GD, a recombinant baculovirus expressing large hepatitis delta antigen (L-HDAg), and Bac-GS2, another recombinant baculovirus expressing HBsAg, gives rise to the assembly and secretion of 22 nm HBsAg subviral particles and 35-37 nm HDV-like particles (HDV VLP). In this study we uncovered oversize particles (>50 nm in diameter) comprised of HBsAg and L-HDAg and the particle properties varied with the relative dosages of Bac-GD and Bac-GS2, as demonstrated by Western blot, transmission electron microscopy and immunogold labeling. At a given Bac-GS2 dosage, decreasing the Bac-GD dosage resulted in the expression of more HBsAg, elevated secretion of HBsAg subviral particles, incorporation of more HBsAg into the HDV VLP, narrower particle size distribution and lower particle density. These data collectively demonstrated that the composition, and hence the properties, of HDV VLPs could be manipulated by altering the relative expression levels of structure proteins.

Hepatitis D virus (HDV) can infect HBsAg-positive individuals, causing rapid fibrosis progression, early decompensation, increased hepatocellular carcinoma risk, and higher mortality than hepatitis B virus (HBV) mono-infection. Most countries lack high-quality HDV prevalence data, and the collection techniques employed often bias published data. In recent meta-analyses, HDV prevalence in HBsAg-positive patients reaches 5%-15% and is even significantly higher in endemic areas. Since HBV vaccination programs were implemented, HDV prevalence has decreased among younger populations. However, owing to immigrant influx, it has increased in some Western countries. The current practice of HDV screening in HBsAg-positive individuals is stepwise, based on physician's discretion, and limited to at-risk populations and may require numerous visits. Double reflex testing, which includes anti-HDV testing in all HBsAg-positive individuals and then HDV RNA testing for anti-HDV-positive ones, is uncommon. Reflex testing can identify more HDV infection cases and link identified patients to further care and follow-up. Moreover, laboratory-based double reflex screening is less biased than physician-led testing. Therefore, healthcare providers should learn about reflex testing, and federal and provincial hepatitis control programs should implement laboratory-based double reflex testing to obtain reliable HDV prevalence estimates. The test's cost-effectiveness depends on the number of HBV-positive patients screened to identify one HDV-positive patient. Such testing may be viable in areas with low HBsAg but high HDV prevalence. However, its economic impact on areas with low HDV prevalence needs further study.

Hepatitis delta: In vitro evaluation of cytotoxicity and cytokines involved in PEG-IFN therapy

Hepatitis delta virus (HDV) is a defective single negative chain RNA virus, as its envelope protein synthesis is dependent on hepatitis B virus (HBV). Studies have consistently shown that coinfection of HBV and HDV is the most serious form of viral hepatitis, with accelerated progression to liver cirrhosis and hepatocellular carcinoma. About 74 million of HBV surface antigen (HBsAg) positive patients worldwide are also co-infected with HDV. Besides, patients with intravenous drug use and high-risk sexual behavior are at higher risk of HDV infection. Therapeutic schedules for HDV are limited, and relapse of HDV has been observed after treatment with pegylated interferon alpha. To reduce the transmission of HDV, all people infected with HBV should be screened for HDV. At present, several serological and molecular detection methods are widely used in the diagnosis of HDV. However, due to the lack of international standards diagnostic results from different laboratories are often not comparable. Therefore, the true prevalence of HDV is still unclear. In this manuscript, we have analyzed various factors influencing the estimation of HDV prevalence. We have also discussed about the advantages and disadvantages of currently available HDV laboratory diagnostic methods, in order to provide some ideas for improving the detection of HDV.

Engineered monoclonal antibody tobevibart enhances HBsAg capture by Fc receptor-positive cells and activates HBV-specific T cells.

Potent broadly neutralizing antibody VIR-3434 controls hepatitis B and D virus infection and reduces HBsAg in humanized mice

To determine how chronic hepatitis D virus (HDV) infection affects intrahepatic hepatitis B virus (HBV) antigen expression. Ninety eight liver biopsy specimens from 68 patients seropositive for total antibody to HDV were studied by immunohistochemistry, and the amount of HBV antigens was also quantified by radioimmunoassay in 12 patients and compared with 30 patients with chronic HBV infection. Forty nine of the 68 patients were positive for intrahepatic HDV antigen and only five were positive for HBV core antigen (HBcAg). HBV surface antigen (HBsAg) was present in 55 (80.9%) patients and was always cytoplasmic in distribution. Hepatic pre-S1 and pre-S2 expressions paralleled that of HBsAg, and were detected in 53 (77.9%) and 54 (79.4%) patients, respectively. There was no relation between the intrahepatic expression of HDV antigen and HBsAg/pre-S1/pre-S2. Follow up biopsy specimens in 25 patients showed either static or deteriorating histology while intrahepatic HDV antigen remained the same or fell. The patients with intrahepatic expression of HBcAg had either absent or noticeably decreased expression of HBcAg in their follow up biopsy specimens (median two years). In contrast, HBsAg/pre-S1/pre-S2 were the same or increased (p less than 0.001). Quantification of intrahepatic HBsAg in patients with chronic HDV infection (0.61 pg/hepatocyte, range: 0.05-1.08, n = 12) showed no difference with patients with chronic HBV infection alone (0.64 pg/hepatocyte, range: 0.02-1.02, n = 30, p = NS). These data indicate that chronic HDV infection suppresses intrahepatic expression of HBcAg but not HbsAg and pre-S antigens, suggesting a differential effect of chronic HDV infection on HBV gene expression.

Background The Hepatitis B virus (HBV) affects over 250 million people worldwide and can lead to cirrhosis and hepatocellular carcinoma. HBV surface antigen (HBsAg) quantification is increasingly used to predict disease activity and treatment response. As there is no virologic cure, clinicians are seeking a “functional cure”, or HBsAg loss, as a guide to safely stopping nucleos(t)ide analogue therapy. Tenofovir Disoproxil Fumarate (TDF) and Entecavir (ETV) are first line therapy but require prolonged treatment to achieve HBsAg clearance. Aims To assess the association between nucleos(t)ide therapy and decline in quantitative HBsAg (qHBsAg) in patients with HBV from Calgary, Alberta. Methods A retrospective review of adult patients with chronic HBV, followed at the University of Calgary Liver Clinic, was conducted between January 2012 and October 2020. Patients were excluded if treatment was discontinued or changed, only had a single qHBsAg measurement, or were co-infected with hepatitis C virus, hepatitis delta virus, or HIV. Patients were stratified according to therapy with TDF, ETV, or no treatment. To identify associations between mean changes in qHBsAg by medication exposure, one-way ANOVAs and t-tests were performed. Results were reported as means and 95% confidence intervals (CI). The median time from initial qHBsAg to most-recent was calculated. Results 187 patients were included in the final analysis (Table 1). 77 were excluded for being on more than one medication over the study period, 10 were excluded due to discontinuation of treatment, and 195 were excluded for single qHBsAg measurements. The mean qHBsAg decline was -750.04 IU/mL (95% CI -1311.58, -188.50) in the TDF group (n=45) and -309.20 IU/mL (95% CI -600.90, -17.50) in the ETV group (n=35) (p=0.20). In the no treatment group (n=107), the mean qHBsAg increased by 711.29 IU/mL (95% CI -600.78, 2023.80)(Figure 1). The median time from initial qHBsAg to most-recent was 980 days. Conclusions Quantitative HBsAg levels declined in patients on TDF and ETV, but increased in untreated patients. Although HBsAg levels showed a trend for greater decline in TDF-treated patients, results failed to reach statistical significance, and may be affected by overall treatment duration. Future studies will explore medication use as a time-varying covariate to identify how change in treatment influences changes in HBsAg over time. Funding Agencies None

Introduction: Hepatitis delta virus (HDV) is associated with the most severe forms of viral hepatitis with rapid progression to cirrhosis and hepatocellular carcinoma. The prevalence of HDV may be significantly higher than formerly acknowledged in the United States, making it doubly important to identify at-risk individuals. Case Description/Methods: A recently emigrated 29-year-old Afghani male with chronic hepatitis B virus (HBV) infection presented with abdominal pain, nausea and vomiting to an outside facility. He underwent a laparoscopic cholecystectomy for symptomatic cholelithiasis. He developed postoperative abdominal distension with imaging initially concerning for a large bile leak however ERCP and a diagnostic laparoscopy were unrevealing. Four liters of fluid were drained, and a surgical drain was placed which continued to drain up to two liters of fluid daily. Upon transfer to our hospital, diagnostic workup showed AST 583 U/L, ALT 452 U/L, alkaline phosphatase 88 U/L, total bilirubin 1.8 mg/dL, and albumin 2.8 g/dL. HBV DNA PCR was < 10 IU/mL. HBV core antibodies, HBV e antibody, and HBV surface antigen were reactive. HBV e antigen and HBV surface antibody were non-reactive. Ascites fluid studies showed a high serum ascites albumin gradient and low protein, prompting a liver biopsy that showed cirrhosis (image 1). The patient was ultimately found to be positive for HDV antibody, confirming a diagnosis of cirrhosis due to HDV superinfection. The patient had his drains removed and was discharged on oral diuretics and entecavir for treatment of HBV infection. Diuretic uptitration is ongoing with plans to enroll in a trial in the future pending clinical status. Discussion: In patients with chronic HBV who acquire HDV as a superinfection, 70-80% develop cirrhosis or hepatocellular carcinoma within 5-10 years. Superinfection is the likely cause of cirrhosis in our patient, as cirrhosis in a 29-year-old would not be expected with HBV infection alone. Early recognition of at-risk individuals can aid in faster diagnosis and earlier treatment. Screening should be considered in all individuals with a reactive HBV surface antigen, and especially in migrants from endemic areas, hemodialysis patients, healthcare employees, and IV drug users. Total HDV antibodies should be obtained for screening, with diagnosis confirmation by serum RT-PCR. Currently, therapeutic options are only available for compensated patients, with pegylated interferon alpha approved in the United States and buleviritide in Europe.Figure 1.: (A) Trichrome stain showing markedly active chronic hepatitis consistent with cirrhosis. (B) Immunohistochemical stains demonstrate patchy staining with HBV surface antigen but (C) negative for HBV core antigen.

Hepatitis delta virus (HDV) is a defective satellite virus and propagates in the presence of Hepatitis B virus (HBV) surface antigen (HBsAg). Approximately 5% of the people who infected with HBV are also infected with HDV. Chronic hepatitis caused by delta is the most severe form of chronic viral hepatitis including accelerated fibrosis, liver decompensation and development of hepatocellular carcinoma. Interferon-based therapies still remain the only treatment option of the hepatitis delta. The beneficiary effects of the interferon-based therapies, however, stop frequently with termination of the given therapy and relapse rate is very high. Accordingly, the efficiency rate of this treatment does not exceed 30%. On the other hand, serious side effects of interferons are another troublesome leading to withdrawal of the therapy. The main goal of the current treatments is clearance of HBsAg. There is no available drug acting directly against the HDV. New therapies interacting with HDV life cycle are under investigation. While prenylation inhibitors act on merely HDV, viral entry inhibitors and HBsAg release inhibitors would be used in the treatment of both HBV and HDV. We hope that in the future, the use of novel therapies and HBV vaccination provide to clinicians to cope with this troublesome agent.