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The impact of concurrent fatty liver (FL) on response to antiviral therapy in chronic hepatitis B (CHB) patients has not been well characterized. We aimed to systematically review and analyse antiviral treatment response in CHB patients with and without FL. We searched PubMed, Embase, Web of Science and the Cochrane Library databases from inception to 31 May 2023 for relevant studies. Biochemical response (BR), complete viral suppression (CVS) and hepatitis B e antigen (HBeAg) seroconversion in CHB patients with FL (CHB-FL) and without FL (non-FL CHB) were compared. In an initial pool of 2101 citations, a total of 10 studies involving 2108 patients were included. After 12 weeks of treatment, CHB-FL patients as compared with non-FL CHB patients had lower BR rate (48.37% [108/227] vs. 72.98% [126/174], p = .04) but similar trend for CVS (36.86% [80/227] vs. 68.81% [114/174], p = .05) and similar rates of HBeAg seroconversion (6.59% [7/103] vs. 7.40% [7/110], p = .89). However, at week 48, there were no statistically significant differences between CHB-FL and non-FL CHB patients in any of the outcomes, including BR (60.03% [213/471] vs. 69.37% [314/717], p = .67), CVS (65.63% [459/746] vs. 73.81% [743/1132], p = .27) and HBeAg seroconversion (10.01% [30/275] vs. 14.06% [65/453], p = .58) with similar findings for week 96. BR rate was lower in CHB-FL patients after 12 weeks of antiviral treatment. However, after a longer follow-up of either 48 or 96 weeks, no statistically significant differences were observed in BR, CVS or HBeAg seroconversion rates between CHB patients with and without FL., (© 2024 John Wiley & Sons Ltd.)

We examined whether the hepatitis B virus (HBV) pregenomic RNA (pgRNA) status after nucleos(t)ide (NA) treatment can predict the long-time prognoses of chronic hepatitis B patients. Patients with chronic hepatitis B (98) who were treatment-naïve and had begun a 7-year NA therapy regimen were enrolled in this study. Biochemical indicators and serological markers of HBV infection were performed during therapy. HBV pgRNA was quantified by real-time quantitative PCR with specific primers. During treatment, HBV DNA undetectable rates increased. The aminotransferase (ALT) normalization (ALT < 50 IU/L) and HBeAg-negative rates also increased. After 48 weeks' NA treatment, 48.28% (28/58) of HBV DNA undetectable patients still had HBV pgRNA-positive. After 7 years of treatment, more HBV pgRNA-negative patients (n = 35) achieved HBeAg clearance than the patients who were HBV pgRNA-positive (n = 63) (19/23 vs 19/56, P < .00). HBV pgRNA-positive patients also had an increased risk of failing to achieve HBeAg clearance (OR = 9.25, 95% CI: 2.75-31.08). The median time to HBeAg clearance in the HBV pgRNA-positive patients was longer than that of the HBV pgRNA-negative patients (152 weeks vs 72 weeks). The HBV pgRNA-positive patients also required more time to achieve HBV DNA undetectable (124 weeks, 95% CI: 103.33-144.67 vs 48 weeks, 95% CI: 34.80-61.20). The HBV pgRNA status after NA treatment can predict the long-term prognoses of patients with chronic HBV. Patients who remain HBV pgRNA-positive after 48 weeks of NA treatment have an increased risk of not achieving HBeAg clearance, need more time to achieve HBeAg clearance and undetectable HBV DNA load., (© 2019 John Wiley & Sons Ltd.)

Investigational agents that reduce or eliminate covalently closed circular DNA (cccDNA) or enhance host immunity against hepatitis B virus (HBV)-infected hepatocytes are intended to induce a durable off-treatment clearance of hepatitis B surface antigen (HBsAg) (referred to as functional cure). The aim of this paper was to highlight challenges in interpreting liver safety data in clinical trials of these agents when given alone or in combination regimens. The incidence, grading and management of spontaneous serum ALT flares in untreated chronic HBV patients are reviewed along with a summary of serum ALT flares observed during the registration trials for peginterferon and nucleos(t)ide reverse transcriptase inhibitors. Recommendations regarding the detection, management and interpretation of liver safety biomarker data in future clinical trials as well as suggested inclusion and exclusion criteria for phase 1/2 vs phase 3 studies are provided. Criteria to help classify liver safety signals as being due to the intended therapeutic response, emergence of drug-resistant HBV virions, or idiosyncratic drug-induced liver injury are provided along with a review of the role of an expert hepatic adjudication panel in assessing a compound's hepatotoxicity profile. Finally, an algorithmic approach to the differential diagnosis and recommended medical evaluation and management of individual clinical trial patients that develop a liver safety signal is provided along with the rationale to collect and test research blood samples for future mechanistic studies., (© 2019 John Wiley & Sons Ltd.)

Chronic HBV infection results in various clinical manifestations due to different levels of immune response. In recent years, hepatitis B treatment has improved by long-term administration of nucleos(t)ide analogues (NUCs) and peg-interferon. Nucleic acid polymers (NAPs; REP 2139-Ca and REP 2139-Mg) are new antiviral drugs that block the assembly of subviral particles, thus preventing the release of HBsAg and allowing its clearance and restoration of functional control of infection when combined with various immunotherapies. In the REP 102 study (NCT02646189), 9 of 12 patients showed substantial reduction of HBsAg and seroconversion to anti-HBs in response to REP 2139-Ca, whereas 3 of 12 patients did not show responses (>1 log reduction of HBsAg and HBV DNA from baseline). We characterized the dynamic changes of HBV quasispecies (QS) within the major hydrophilic region (MHR) of the 'pre-S/S' open reading frame including the 'a' determinant in responders and nonresponders of the REP 102 study and four untreated matched controls. HBV QS complexity at baseline varied slightly between responders and nonresponders (P = .28). However, these responders showed significant decline in viral complexity (P = .001) as REP 2139-Ca therapy progressed but no significant change in complexity was observed among the nonresponders (P = .99). The MHR mutations were more frequently observed in responders than in nonresponders and matched controls. No mutations were observed in 'a' determinant of major QS population which may interfere with the detection of HBsAg by diagnostic assays. No specific mutations were found within the MHR which could explain patients' poor HBsAg response during REP 2139-Ca therapy., (© 2019 John Wiley & Sons Ltd.)

Ubiquitin-specific protease 18 (USP18) is an important inhibitor of interferon (IFN) antiviral activity, and the aim of this study was to investigate the association between the USP18 mRNA level change in peripheral blood mononuclear cells (PBMCs) when stimulated with IFN in vitro before initiating treatment and the treatment outcomes in HBeAg-positive chronic hepatitis B (CHB) patients treated with IFN. A total of 44 patients who received standard IFN-based anti-HBV therapy and follow-up were enrolled in the study. The in vitro IFN-induced USP18 mRNA change (USP18 IFN-N ) was measured via comparison of quantitative PCR-determined USP18 transcription levels of BPMCs cultured with and without IFN stimulation. Either for virological (VR) or serological response (SR), the baseline USP18 IFN-N was significantly higher (P = 0.018 for VR, P = 0.008 for SR) among nonresponders (n = 23 for VR, n = 33 for SR) than that of responders (n = 21 for VR, n = 11 for SR). Multivariate analyses revealed baseline USP18 IFN-N was a novel independent predictor for either VR (OR = 0.292, 95% CI = 0.102-0.835, P = 0.022) or SR (OR = 0.173, 95% CI = 0.035-0.849, P = 0.031) in our cohort. In addition, baseline USP18 IFN-N in combination with HBV DNA loads or HBeAg levels showed improved accuracy of pretreatment prediction for VR or SR responders, respectively. Baseline USP18 IFN-N levels are associated with both virological and serological response, and have the potential to become a clinical predictor for treatment outcomes in HBeAg-positive CHB patients before initiating IFN-α therapy., (© 2019 John Wiley & Sons Ltd.)

Chronic hepatitis B (CHB) infection affects over 250 millon people worldwide and 800000 are expected to die yearly due to the development of hepatocellular carcinoma (HCC). Current antiviral therapies include nucleoside analogs (NAs) that target the viral retrotranscriptase inhibiting de novo viral production. Pegylated interferon (Peg-IFN) is also effective in reducing the viral DNA load in serum. However, both treatments remain limited to control the infection, aiming for viral suppression and improving the quality of life of the infected patients. Complete cure is not possible due to the presence of the stable DNA intermediate covalently closed circular DNA (cccDNA). Attempts to achieve a functional cure are thus ongoing and novel targets and molecules, together with different combination therapies are currently in the pipeline for early clinical trials. In this review we discuss novel treatments both targeting directly and indirectly cccDNA. As we gain knowledge in the Hepatitis B virus (HBV) transcriptional control, and newer technologies emerge that could potentially allow the destruction of cccDNA, exciting new possibilities for curative therapies are discussed., (© 2019 John Wiley & Sons Ltd.)

Hepatitis B e antigen (HBeAg) seroconversion experienced during nucleo(s)tide analogue (NUC) therapy is often not sustained. We aimed to study whether hepatitis B core-related antigen (HBcrAg) levels predict sustained HBeAg seroconversion in patients treated with NUCs. We studied HBeAg-positive patients treated with NUCs for at least 6 months. We quantified HBcrAg at baseline and at the time of HBeAg seroconversion and studied the relationship with HBeAg seroconversion and subsequent relapse. HBcrAg was quantified at baseline in 196 patients; levels varied significantly by HBV genotype and correlated with HBsAg, HBV DNA and HBeAg. Baseline HBcrAg levels were lower in patients who achieved HBeAg seroconversion than in those who did not; the unadjusted hazard ratio (HR) was 0.802 (95% CI: 0.656-0.980, P = 0.031); and this association was not sustained in multivariate analysis. HBcrAg remained detectable in all patients at the time of HBeAg seroconversion. Higher HBcrAg at the time of seroconversion was an independent predictor of relapse (adjusted HR: 1.855 (95% CI: 1.099-3.133, P = 0.021), and none of the patients with HBcrAg < 4.90 log U/mL experienced relapse. Baseline HBcrAg is not an independent predictor of HBeAg seroconversion during NUC therapy. HBcrAg remains detectable in patients after HBeAg seroconversion. Patients with lower levels at the time of seroconversion have a higher probability of sustained HBeAg seroconversion., (© 2019 John Wiley & Sons Ltd.)

Hepatitis B surface antigen (HBsAg) loss is considered a functional cure in chronic hepatitis B (CHB). However, the durability of HBsAg loss after stopping treatment remains unknown. This study aimed to assess the sustained functional cure achieved by interferon therapy in hepatitis B envelope antigen (HBeAg)-negative CHB patients. In this prospective study, 176 HBeAg-negative CHB patients with functional cure were enrolled for 12 weeks of cessation treatment, and treatment information and baseline data were collected. Hepatitis B virus (HBV) biomarkers and clinical biochemical indicators were evaluated every 3 months; liver imaging examinations were performed every 3-6 months during the 48-week follow-up. The sustained functional cure was evaluated. After the 48-week follow-up, the sustained functional cure rate was 86.63%. The cumulative rates of HBsAg reversion and HBV DNA reversion were 12.79% and 2.33%, respectively. Consolidation treatment ≥ 12 weeks after HBsAg loss achieved a significantly higher rate of sustained functional cure and significantly lower rate of HBsAg reversion than consolidation treatment < 12 weeks (76.19% vs 90.00%, P = 0.022 and 23.81% vs 9.23%, P = 0.014, respectively). Patients with hepatitis B surface antibody (HBsAb) had higher rate of sustained functional cure than patients achieving HBsAg loss but without HBsAb (89.86% vs 73.53%, P = 0.012). Consolidation treatment ≥ 12 weeks (odds ratio [OR] 16.478; 95% confidence interval [CI], 2.135-127.151; P = 0.007) and high HBsAb levels (OR 8.312; 95% CI, 1.824-37.881; P = 0.006) were independent predictors of sustained functional cure. Results suggested that 12 weeks of consolidation therapy after HBsAg clearance and elevated HBsAb levels help to improve functional cure., (© 2019 John Wiley & Sons Ltd.)

The study was undertaken in order to provide a snapshot from real clinical practice of virological presentation and outcome of patients developing immunosuppression-driven HBV reactivation. Seventy patients with HBV reactivation were included (66.2% treated with rituximab, 10% with corticosteroids and 23.8% with other immunosuppressive drugs). Following HBV reactivation, patients received anti-HBV treatment for a median (IQR) follow-up of 31(13-47) months. At baseline-screening, 72.9% of patients were HBsAg-negative and 27.1% HBsAg-positive. About 71.4% had a diagnosis of biochemical reactivation [median (IQR) HBV DNA and ALT: 6.9 (5.4-7.8) log IU/mL and 359 (102-775) U/L]. Moreover, 10% of patients died from hepatic failure. Antiviral prophylaxis was documented in 57.9% and 15.7% of HBsAg-positive and HBsAg-negative patients at baseline-screening (median [IQR] prophylaxis duration: 24[15-33] and 25[17-36] months, respectively). Notably, HBV reactivation occurred 2-24 months after completing the recommended course of anti-HBV prophylaxis in 35.3% of patients. By analysing treatment outcome, the cumulative probability of ALT normalization and of virological suppression was 97% and 69%, respectively. Nevertheless, in patients negative to HBsAg at baseline-screening, only 27% returned to HBsAg-negative status during prolonged follow-up, suggesting the establishment of chronic infection. In conclusion, most patients received a diagnosis of HBV reactivation accompanied by high ALT and 10% died for hepatic failure, supporting the importance of strict monitoring for an early HBV reactivation diagnosis. Furthermore, HBV reactivation correlates with high risk of HBV chronicity in patients negative for HBsAg at baseline-screening, converting a silent into a chronic infection, requiring long-term antiviral treatment. Finally, a relevant proportion of patients experienced HBV reactivation after completing the recommended course of anti-HBV prophylaxis, suggesting the need to reconsider proper duration of prophylaxis particularly in profound immunosuppression., (© 2019 John Wiley & Sons Ltd.)

Diabetes mellitus may be a risk factor of HCC development in chronic hepatitis B infected patients and affect the all-cause mortality. This study aimed to examine whether DM was associated with the development of HCC with CHB and affected the all-cause mortality. A total of 2966 CHB patients newly diagnosed with DM in 2000 were retrieved from the Longitudinal Cohort of Diabetes Patients database and used propensity scores matching based on age, sex-gender, alcohol-related liver disease and baseline liver cirrhosis to compare with the non-DM patients from the Taiwanese National Health Insurance Research Database. The CHB patients with DM compared to the non-DM had significantly increased (3.3%) risk for HCC development and significantly increased (2.8%) risk of HCC-related mortality. Interestingly, the all-cause mortality was significantly higher in the DM cohort (16.9%) compared to the non-DM cohort (8.2%). In a multivariable transition-specific Cox model to investigate the adjusted hazard ratio of CHB patients with DM or non-DM during the transitions from start to HCC was 1.35; 95% CI (1.16-1.57) and from HCC to death was 1.31; 95% CI (1.06-1.62). All-cause mortality between CHB patients with DM or non-DM during the transitions from start to death was 2.32; 95% CI (1.84-2.92). Taken together, DM is an independent risk factor associated with increasing disease development of HCC, HCC-related mortality and all-cause mortality in CHB patients. This study may provide a clinical strategy for strict DM control in order to reduce the risk of disease development in CHB patients., (© 2019 John Wiley & Sons Ltd.)

The massive implementation of the vaccine and antiviral agents against hepatitis B virus (HBV), targeting the envelope and viral polymerase genes, induces a selection pressure that might lead to the emergence of variants that impair the effectiveness of the vaccine, diagnostic methods and antiviral therapy. The aim of this study was to evaluate the prevalence of HBV vaccine escape mutants (VEMs), diagnostic failure mutants (DFMs) and treatment resistance mutants (ARMs) among individuals from Buenos Aires, Argentina. HBV surface antigen and polymerase sequences obtained from serum samples of 530 HBV-infected individuals were analysed. Samples belonged to genotypes A (28.1%), D (13.6%) and F (58.3%). VEMs, DMFs and ARMs were present in 40 (7.5%), 57 (10.7%) and 27 (5.1%) samples within the studied population. Additionally, eight nonpreviously reported VEMs and nine DFMs were identified. VEMs and DFMs were biased by genotype, being higher in genotype D (33.3% and 33.3%) compared to genotype A (6% and 17.4%) and genotype F (2.3% and 2.3%) (P > 0.001). On the contrary, there was no association between the presence of ARMs and HBV genotype (P = 0.324). VEMs, DFMs and ARMs create public health concerns. The current study provided valuable information about mutants in surface antigen and polymerase in HBV-infected patients from Argentina where HBV-F is the most prevalent genotype. Consequently, it constitutes an important reference for Latin American clinicians in order to optimize the management of HBV-infected patients., (© 2018 John Wiley & Sons Ltd.)

The treatment of patients suffering from HBeAg-positive chronic hepatitis B with REP 2139-Ca resulted in potent reductions in HBsAg and HBV DNA, seroconversion to anti-HBs and the establishment of functional control of infection. In this cohort of 12 patients, we investigated whether differences between HBsAg sequences might explain the lack of response to REP 2139-Ca observed in 3 of 12 patients. We also assessed if the reduction or complete loss of HBsAg in serum observed during therapy were caused by mutations in the "a" determinant preventing the detection of HBsAg by standard diagnostic assays. The complete pre-S/S open reading frame (ORF) was sequenced and pre-S1, pre-S2 and S amino acid sequences were analysed. We found no major differences between pre-S1, pre-S2 and S sequences in responders and nonresponders correlated with low reduction in HBsAg. In addition, we found no mutations in the "a" determinant that would significantly affect the reactivity of HBsAg in diagnostic assays. These results demonstrate that the amino acid sequence of complete pre-S/S ORF has no direct influence on response to REP 2139-Ca therapy., (© 2018 John Wiley & Sons Ltd.)

Nucleoside and nucleotide analogues (NUCs) targeting hepatitis B virus are capable of selecting resistant viruses upon long-term administration as monotherapies. The prevalence of resistance-associated substitutions (RASs) and fitness-associated substitutions at baseline of NUC therapy and their impact on treatment responses remain unknown. A total of 232 treatment-naïve patients chronically infected with hepatitis B virus (HBV) consecutively referred for the first time to one of French reference centres were included. The nearly full-length HBV reverse transcriptase was sequenced by means of deep sequencing, and the sequences were analysed. RASs were detected in 25% of treatment-naïve patients, generally representing low proportions of the viral quasispecies. All amino acid positions known to be associated with HBV resistance to currently approved NUCs or with increased fitness of resistant variants were affected, except position 80. RASs at positions involved in lamivudine, telbivudine and adefovir resistance were the most frequently detected. All patients with RASs detectable by next-generation sequencing at baseline who were treatment-eligible and treated with currently recommended drugs achieved a virological response. The presence of pre-existing HBV RASs has no impact on the outcome of therapy if potent drugs with a high barrier to resistance are used., (© 2018 John Wiley & Sons Ltd.)

HBV surface antigen (HBsAg) reduction is well observed in chronic hepatitis B (CHB) patients treated with pegylated interferon alpha-2a (PegIFNα). However, the mechanism of HBsAg suppression has not been fully elucidated. Twenty-seven of 55 entecavir-treated CHB e antigen positive patients were switched to PegIFNα treatment (Group A) whereas 28 patients continued entecavir treatment (Group B). The percentage or absolute number of CD56 bright /CD56 dim NK cells, expression of receptors and cytokines were evaluated by flow cytometry for 48 weeks and correlated with treatment efficacy. In vitro, purified NK cells were co-cultured with HepAD38 cells for measurement of HBsAg, apoptosis and covalently closed circular DNA (cccDNA). In association with a reduction of HBsAg, the percentage and absolute number of CD56 bright NK cells was significantly elevated in patients in group A, especially in Virologic Responders (VRs, HBsAg decreased). Furthermore, the percentage of NKp30 + , NKp46 + , TRAIL + , TNF-α + and IFNγ + CD56 bright NK cells were significantly expanded in Group A, which were positively correlated with the decline of HBsAg at week 48. In vitro, peripheral NK cells from Group A induced a decline of HBsAg in comparison with NK cells from Group B which was significantly inhibited by anti-TRAIL, anti-TNF-α and anti-IFNγ antibodies. Furthermore, apoptosis of HepAD38 cells and levels of cccDNA, were significantly reduced by TRAIL + and TNF-α + /IFNγ + NK cells from Group A, respectively. A functional restoration of CD56 bright NK cells in entecavir-treated patients who were switched to PegIFNα contributes to HBsAg and cccDNA clearance through TRAIL-induced cytolysis and TNF-α/IFNγ-mediated noncytolytic pathways., (© 2018 John Wiley & Sons Ltd.)

Entecavir 0.5 mg (ETV) is widely used among treatment-naïve chronic hepatitis B (CHB) patients. However, 10%-30% of patients show partial virologic response (PVR) to the drug. If the hepatitis B virus (HBV) continues to replicate, the underlying liver disease may progress. Herein, we compared the efficacy of switching to tenofovir disoproxil fumarate (TDF) with that of continuing ETV in CHB patients with PVR to ETV. This was an open-label randomized controlled trial including CHB patients who had been receiving 0.5 mg of ETV for >12 months, but who still had detectable HBV DNA levels of >60 IU/mL without known resistance to ETV. Sixty patients were enrolled and 45 qualified for the study: Twenty-two patients were randomly assigned into the TDF group and 23 into the ETV group. After 12 months of treatment, the virologic response rate (HBV DNA <20 IU/mL) was significantly higher in the TDF group than in the ETV group, as measured using per-protocol analysis (55% vs 20%; P = .022) and intention-to-treat analysis (50% vs 17.4%; P = .020). The reduction in HBV DNA was greater (-1.13 vs -0.67 log 10 IU/mL; P = .024), and the mean HBV DNA level was lower (1.54 vs 2.01 log 10 IU/mL; P = .011) in the TDF group than in the ETV group. In conclusion, to achieve optimal response in CHB patients with PVR to ETV, switching to TDF would be a better strategy than continuing ETV. Appropriate modification of therapy would further improve the outcome of chronic HBV infection., (© 2018 John Wiley & Sons Ltd.)

Vesatolimod is an oral agonist of toll-like receptor 7 designed to minimize systemic exposure and side effects. We assessed the safety and efficacy of vesatolimod in viremic chronic hepatitis B (CHB) patients not currently on oral antiviral treatment (OAV) in a phase 2, multicentre, double-blind, randomized, placebo-controlled study. A total of 192 patients stratified by HBeAg status and alanine aminotransferase level were randomized 2:2:2:1 to receive oral vesatolimod (1-, 2- or 4-mg) or placebo once weekly for 12 weeks; tenofovir disoproxil fumarate (300-mg daily) was administered daily for 48 weeks. Efficacy was assessed by quantitative serum HBsAg decline at Week 24 from baseline. In addition to safety assessments, changes in whole-blood interferon-stimulated gene (ISG) transcripts and serum cytokines were explored. Most patients were male (64.1%) and HBeAg-negative (60.9%) at baseline. Among vesatolimod-treated patients, most (60.4%-69.1%) experienced ≥1 treatment-emergent adverse event; the majority were mild or moderate in severity. No clinically meaningful differences in HBsAg changes from baseline were observed between treatment groups. No patients experienced HBsAg loss, while 3 patients experienced HBeAg loss and hepatitis B e-antibody seroconversion at week 48. HBV DNA suppression rates were similar across all treatment arms at Week 24. ISG15 induction was dose-dependent and did not correlate with HBsAg changes. A small proportion of patients exhibited dose-dependent interferon-α induction that correlated with grade of influenza-like adverse events. Overall, vesatolimod is safe and well tolerated in CHB patients. Although consistent dose-dependent pharmacodynamic induction of ISGs was demonstrated, it did not result in clinically significant HBsAg decline., (© 2018 John Wiley & Sons Ltd.)

The management of chronic hepatitis B virus (HBV) infection is challenged by its varying natural course and its stealthy nature. Not all HBV-infected patients will develop complications of infection; however, it is of utmost importance to identify patients who are at risk and require antiviral treatment and/or close surveillance. Hepatic inflammation and quantification of HBV DNA have guided treatment decisions in the last decade, and these guided interventions have been shown to reduce liver-related complications and death. Data on the quantification of additional HBV markers such as hepatitis B surface antigen (HBsAg), hepatitis B core-related antigen (HBcrAg) and hepatitis B virus RNA (HBV RNA) have accumulated in recent years. Here, we review the current evidence of how to use these markers and discuss open issues that require additional research., (© 2018 John Wiley & Sons Ltd.)

This study evaluated the long-term efficacy and safety of an 18-month lamivudine prophylaxis in 68 HBsAg-negative/anti-HBc-positive patients with oncohaematological disease. All 68 consecutive HBsAg-negative/anti-HBc-positive patients with an oncohaematological disease and naïve for chemotherapy observed from April 2008 to December 2012 at 2 Hematology Units in Naples were treated with lamivudine for 18 months after stopping chemotherapy and monitored for HBsAg at months 1 and 3 during chemotherapy and then every 3 months after its discontinuation. During follow-up, 13 (19.1%) of the 68 patients died of complications related to their oncohaematological disease, and 3 (4%) showed a virological HBV reactivation (retroconversion to HBsAg positivity) 1-7 months after the discontinuation of lamivudine prophylaxis (2 treated for chronic lymphocytic leukaemia and one for Waldenstrom's disease); of these, 2 showed a biochemical reactivation. Comparing the demographic and clinical characteristics of the 3 patients with a virological HBV reactivation to the 65 without, the former were older (median age and range: 67 years [75-78] vs. 61 [24-88]; P = .05) and were less frequently treated for B-cell non-Hodgkin lymphoma (B-NHL) (0 vs. 70.7%, P = .03). In conclusion, a 18 months of lamivudine prophylaxis was effective in preventing HBV reactivation in HBsAg-negative/anti-HBc-positive patients treated for B-NHL. However, in patients with chronic and severe immunodepression, such as those with chronic lymphocytic leukaemia and Waldenstrom's disease, prophylaxis should be continued for an indefinite period., (© 2017 John Wiley & Sons Ltd.)

Recently, cases of hepatitis B virus reactivation (HBVr) with direct-acting antiviral therapy (DAAs) for HCV have been reported. However, few data exist from large, Western cohorts. The study objectives were to evaluate the incidence of alanine aminotransferase (ALT) flares, clinically significant hepatic events, and HBVr among a national cohort of US veterans with prior exposure to HBV (anti-HBc+) treated with DAAs. We used a national administrative database to identify patients treated with DAAs from January 2014 through November 2016 and obtained clinical and demographic as well as HBV and HCV treatment data. HBVr was defined as an at least 1-log increase in HBV DNA titre. Among 17 779 anti-HBc+ patients, 17 400 were HIV- and 379 were HIV+. Among the HIV- patients, 17 266 (99%) were HBsAg- prior to DAA therapy and 134 were HBsAg+. Among HIV-, HBsAg- patients, ALT elevations greater than 10 times the upper limit of normal (ULN; ≥300 IU/mL) were rare and occurred more frequently after treatment completion: 31 cases (<0.1%) during vs 85 (0.6%) following treatment. Clinically significant hepatic events defined as ALT increases >100 IU/L with total bilirubin >2.5 mg/dL occurred in 39 cases (0.3%), most often following DAA completion (n = 35 cases, 3/35 in setting of HCV relapse). Among 31 patients with post-DAA hepatic events without HCV relapse, 10 (32%) were confirmed unrelated to HBVr by HBsAg and/or HBV DNA testing, 1 (3%) confirmed due to HBVr, and 20 (65%) did not have documented HBV-related testing. One additional case of HBsAg- to + seroreversion was identified. Among HBsAg+ DAA recipients, 2/97 (2%), both with cirrhosis, experienced ALT elevations ≥300 IU/mL in the setting of HBVr. In conclusion, clinically significant hepatic events and HBVr were rare and much more likely among HBsAg-positive individuals. Anti-HBc + patients should be monitored for ALT flares and HBVr during and possibly for up to 6 months post-DAA therapy., (© 2017 John Wiley & Sons Ltd.)

Anecdotal reports suggest that patients with chronic hepatitis C virus (HCV) hepatitis and overt or occult hepatitis B virus (HBV) coinfection may reactivate HBV when HCV is suppressed or cleared by direct-acting antivirals (DAAs). We assessed the prevalence of overt or previous HBV coinfection and the risk of HBV reactivation in patients with HCV cirrhosis treated with DAAs. This was a retrospective cohort of 104 consecutive patients with HCV cirrhosis treated with DAAs. Serum HCV-RNA and HBV-DNA were tested at weeks 4, 8 and 12 of DAAs therapy and at week 12 of follow-up. At the start of DAAs, eight patients (7.7%) were HBsAg positive/HBeAg negative with undetectable HBV-DNA and low levels of quantitative HBsAg (four on nucleos(t)ide analogues [NUCs] and four inactive carriers), 37 patients (35.6%) had markers of previous HBV infection (25 anti-HBc positive, 12 anti-HBc/anti-HBs positive) and 59 (56.7%) had no evidence of HBV infection. Sixty-seven patients (64.4%) were HCV-RNA negative at week 4 and 98 (94.2%) achieved sustained virological response. All four HBsAg-positive patients treated with NUCs remained HBV-DNA negative, but three of four untreated patients showed an increase in HBV-DNA of 2-3 log without a biochemical flare and achieved HBV-DNA suppression when given NUCs. During or after DAAs, by conventional assay, HBV-DNA remained not detectable in all 37 anti-HBc-positive patients but in three of them (8.1%) HBV-DNA became detectable with a highly sensitive PCR. HBV reactivation is likely to occur in untreated HBV/HCV-coinfected cirrhotic patients when they undergo HCV treatment with DAAs. Pre-emptive therapy with NUCs should be considered in this setting. Anti-HBc-positive patients rarely reactivate HBV without clinical or virological outcomes., (© 2017 John Wiley & Sons Ltd.)

Antiviral drug resistance hepatitis B virus (HBV) variants (HBV-DR) occur spontaneously in chronic hepatitis B (CHB) patients and after exposure to nucleos(t)ide analogues (NUCs). We determined the prevalence of HBV-DR variants among participants of the Hepatitis B Research Network (HBRN) Cohort Study conducted at 21 sites in the United States (US) and Canada. Samples obtained from 1342 CHB participants aged ≥18 years, and who were currently not receiving NUCs, were tested for HBV-DR variants by Sanger sequencing. In addition, next generation sequencing (NGS) was used to characterize HBV-DR variants from 66 participants with and 66 participants with no prior NUC exposure matched for HBV genotype and HBV DNA level. Half the participants were men, 75% Asian, 26% HBeAg positive. Primary HBV-DR variants were detected by Sanger sequencing in 16 (1.2%) participants: 2/142 (1.4%) with and 14/1200 (1.2%) without prior NUC exposure; only 1 of these 16 had a secondary variant. In total, 23 (1.7%) participants had secondary variants, including 1 with prior NUC experience. In the subset of 132 participants, NGS detected HBV-DR variants in a higher proportion of participants: primary variants in 18 (13.6%) (8 [12.1%] with, and 10 [15.2%] without prior NUC therapy) and secondary variants in 10 (7.6%) participants. Based on Sanger sequencing, prevalence of primary HBV-DR variants is low (1.2%) among adults with CHB in US/Canada. The similar low prevalence of HBV-DR variants in participants with and without NUC treatment suggests transmission of these variants is uncommon., (© 2017 John Wiley & Sons Ltd.)

We compared the viral suppressive efficacy of tenofovir disoproxil fumarate (TDF) mono-rescue therapy (TDF group) and TDF plus entecavir (ETV) combination-rescue therapy (TDF + ETV group) in chronic hepatitis B (CHB) patients with lamivudine resistance and entecavir resistance. One hundred and thirty-three CHB patients with lamivudine and entecavir resistance were investigated. Ninety-six patients were treated with TDF and 37 with TDF + ETV for at least 6 months. We compared the virologic response rate (HBV DNA level <20 IU/mL) between the two groups and identified the predictive factors of treatment outcome. There were no significant differences between the two groups in demographic characteristics. Up to 24 months [median: 18 (range 6-24) months], 85.4% and 89.2% of the TDF group and TDF + ETV group, respectively, achieved a virologic response (P=.068). Only the HBV DNA level at baseline was significantly associated with a virologic response in the multivariate analysis. In a subanalysis of patients with HBV DNA levels ≥4 log (IU/mL) at baseline, a higher proportion of patients in the TDF + ETV group than the TDF group achieved a virologic response (92.9% vs 68.3%; P<.001), while 90% of patients with HBV DNA (IU/mL) levels <4 log in all both TDF and TDF + ETV groups achieved a virologic response. TDF mono-rescue therapy is a reasonable option in patients with lamivudine resistance and entecavir resistance. However, the combination strategy should be considered in patients with high baseline HBV DNA levels., (© 2016 John Wiley & Sons Ltd.)

A major hurdle in the long-term treatment of chronic hepatitis B (CHB) patients is to maintain viral suppression in the absence of drug resistance. To date, no evidence of resistance to tenofovir disoproxil fumarate (TDF) has been observed. A cumulative evaluation of CHB patients who qualified for resistance surveillance over 8 years of TDF treatment was conducted. Patients in studies GS-US-174-0102 (HBeAg-) and GS-US-174-0103 (HBeAg+) were randomized 2:1 to receive TDF or adefovir dipivoxil (ADV) for 48 weeks followed by open-label TDF through year 8. Population sequencing of HBV pol/RT was attempted for all TDF-treated patients at baseline and, annually if viremic, at discontinuation, or with addition of emtricitabine. Overall, 88/641 (13.7%) patients qualified for sequence analysis at one or more time points. The percentage of patients qualifying for sequence analysis declined over time, from 9 to 11% in years 1-2 to <4% over years 3-8. Forty-one episodes of virologic breakthrough (VB) occurred throughout the study, with most (n=29, 70%) associated with nonadherence to study medication. Fifty-nine per cent of VB patients with an opportunity to resuppress HBV achieved HBV DNA resuppression. A minority of patients who qualified for sequencing had polymorphic (41/165, 24.8%) or conserved (17/165, 10.3%) site changes in pol/RT, with six patients developing lamivudine and/or ADV resistance-associated mutations. No accumulation of conserved site changes was detected. The long-term treatment of CHB with TDF monotherapy maintains effective suppression of HBV DNA through 8 years, with no evidence of TDF resistance or accumulation of conserved site changes., (© 2016 John Wiley & Sons Ltd.)

In the treatment of chronic hepatitis B virus (HBV) infection, polymerase inhibitors successfully suppress HBV DNA production. However, the production of viral proteins continues unhindered, which hampers viral clearance. Here, we screen for compounds that suppress HBV transcription, which would prevent viral protein production. A total of 640 FDA-approved drugs were evaluated for their ability to inhibit HBV transcription in a transfection-based HBV reporter assay. The assay was performed in the presence and absence of the HBV accessory protein X (HBx), which is essential for in vivo HBV RNA transcription. We observed that in the absence of HBx 47, and in the presence of HBx 24 compounds suppressed transcription by more than 20%. We selected the 24 most potent compounds in each condition for further analysis. On average, the selected compounds reduced transcription by 33.9% (range: 24.1-65.8%) in the absence of HBx expression, and 30.6% (range: 20.4-48.9%) in the presence of HBx. The two selections of 24 compounds had 12 compounds in common, resulting in a final selection of 36 compounds, which were evaluated for their capacity to suppress HBV replication in constitutively HBV-replicating HepG2.2.15 cells. Twenty-three of these compounds reduced HBV replication by interfering with RNA transcription. Further analysis revealed that one of the compounds, terbinafine, potently and specifically suppressed HBx-mediated HBV RNA transcription in HepG2 cells. Inhibition of HBV protein production is a promising step towards HBV clearance. In combination with an HBV polymerase inhibitor, the added suppression of HBV RNA transcription may markedly improve antiviral treatment outcome., (© 2015 John Wiley & Sons Ltd.)

Patients with chronic hepatitis B (HBsAg-positive) are at risk of viral reactivation if rituximab is administered without antiviral treatment, a potentially fatal complication of treatment. Patients with so-called 'resolved hepatitis B virus infection' (HBsAg-negative/cAb-positive) may also be at risk. We performed a systematic review of the English and Chinese language literature to estimate the risk of hepatitis B virus (HBV) reactivation in HBsAg-negative/cAb-positive patients receiving rituximab for lymphoma. A pooled risk estimate was calculated for HBV reactivation. The impact of HBsAb status and study design on reactivation rates was explored. Data from 578 patients in 15 studies were included. 'Clinical HBV reactivation', (ALT >3 × normal and either an increase in HBV DNA from baseline or HBsAg seroreversion), was estimated at 6.3% (I(2) = 63%, P = 0.006). Significant heterogeneity was detected. Reactivation rates were higher in prospective vs retrospective studies (14.2% vs 3.8%; OR = 4.39, 95% CI 0.83-23.28). Exploratory analyses found no effect of HBsAb status on reactivation risk (OR = 0.083; P = 0.151). Our meta-analysis confirms a measurable and potentially substantial risk of HBV reactivation in HBsAg-negative/cAb-positive patients exposed to rituximab. However, heterogeneity in the existing literature limits the generalizability of our findings. Large, prospective studies, with uniform definitions of HBV reactivation, are needed to clarify the risk of HBV reactivation in HBsAg-negative/cAb-positive patients., (© 2015 John Wiley & Sons Ltd.)

The study aimed to clarify clinical significance of hepatitis B virus (HBV) rtA181S mutation in Chinese HBV-infected patients. A total of 18 419 patients with chronic HBV infection from Beijing 302 Hospital were investigated. HBV complete reverse transcriptase region of polymerase was screened by direct sequencing, and the results were verified by clonal sequencing. Replication-competent mutant and wild-type HBV genomic amplicons were constructed and transfected into the HepG2 cells and cultured in the presence or absence of serially diluted nucleos(t)ide analogues. Intracellular HBV replicative intermediates were quantitated for calculating the 50% effective concentration of the drug (EC(50)). The rtA181S was detected in 98 patients with 12 kinds of mutational patterns. Genotype C and genotype B HBV infection occupied 91.8% and 8.2% in rtA181S-positive patients, in contrast to 84.6% and 15.4% in rtA181S-negative patients (P < 0.01). All rtA181S-positive patients had received nucleos(t)ide analogues. rtA181S was detected in multiple patients with virologic breakthrough. Phenotypic analysis of patient-derived viral strains showed that rtA181S, rtA181S+N236T, rtN236T and rtA181V strains had 68.5%, 49.9%, 71.4% and 66.2% of natural replication capacity of wild-type strain, and 3.7-fold, 9.8-fold, 7.9-fold and 5.6-fold increased EC(50) to adefovir dipivoxil (ADV). The rtA181S strain remained susceptible to lamivudine, entecavir and tenofovir, and ADV susceptibility was restored after the mutation was eliminated through site-directed mutagenesis. Rescue therapy with entecavir or combination therapy was effective in rtA181S-related ADV-refractory patients. The rtA181S mutation confers moderate resistance to ADV. It could be induced by either lamivudine or ADV and contribute ADV treatment failure., (© 2014 John Wiley & Sons Ltd.)

This study sought to assess the antiviral efficacy of lamivudine (LMV) administered during third trimester to reduce maternal viraemia and to identify the emergence of LMV resistance. A prospective observational analysis was performed on 26 mothers with high viral load (>10⁷ IU/mL). Twenty-one women received LMV (treated group) for an average of 53 days (range 22-88 days), and the remaining five formed the untreated control group. Serum samples from two time points were used to measure HBV DNA levels and antiviral drug resistance. The LMV-treated women achieved a median HBV DNA reduction of 2.6-log10 IU/mL. Although end-of-treatment (EOT) HBV DNA in four (18%) LMV-treated women remained at >10(7) IU/mL (± 0.5 log IU/mL), no mother-to-baby transmission was observed. In contrast, a baby from the untreated mother was HBsAg positive at 9 months postpartum. Four technologies were used for drug resistance testing. Only ultra-deep pyrosequencing (UDPS) was sufficiently sensitive to detect minor viral variants down to <1%. UDPS showed that LMV therapy resulted in increased viral quasispecies diversity and positive selection of HBV variants with reverse transcriptase amino acid substitutions at sites associated with primary LMV resistance (rtM204I/V and rtA181T) in four (19%) women. These viral variants were detected mostly at low frequencies (0.63-5.92%) at EOT, but one LMV-treated mother had an rtA181T variant that increased from 2.2% pretherapy to 25.59% at EOT. This mother was also infected with the vaccine escape variant (sG145R), which was inhibited by LMV treatment. LMV therapy during late pregnancy only reduced maternal viraemia moderately, and drug-resistant viral variants emerged., (© 2013 John Wiley & Sons Ltd.)

Hepatitis B virus surface antigen (HBsAg) plays an important role in maintaining the tolerance and may interfere with host innate and adaptive immune responses; therefore, novel therapeutic strategies to reduce HBsAg loads in patients infected with hepatitis B virus (HBV) are emerging as an attractive but challenging issue. Metformin could regulate hepatic metabolism while the latter interacts with HBV infection. We hypothesized that metformin could affect HBsAg expression and HBV replication and may work synergistically when combined with current antivirals. In our study, a notably inhibitory effect on HBsAg production, as well as a moderate inhibition in HBV replication and HBeAg expression was observed following metformin treatment. The 50% effective concentration (EC50) for extracellular HBsAg and intracellular HBsAg in HBV-producing HepG2.2.15 cells was 2.85 mm and 2.75 mm, respectively, with a similarly selective index of about 18. When administered in combination, metformin enhanced the inhibitory effects of interferon-α2b on HBsAg expression and HBV replication and provided a complimentary role in HBsAg expression for lamivudine (LMV). This novel action of metformin derives partially from its inhibition on multiple HBV cis-acting elements. By the virtues of preferably hepatocyte distribution and safety profile, collectively, our results suggest that metformin would be potentially clinically helpful as an HBsAg production inhibitor., (© 2013 John Wiley & Sons Ltd.)

This single-arm, open-label, descriptive study assessed the efficacy and safety of entecavir (ETV) in nucleos(t)ide-naïve Black/African American patients with chronic hepatitis B (CHB), a patient population underrepresented in ETV registration trials. Forty patients with HBeAg(+) or HBeAg(-) compensated CHB of self-described Black/African American race received ETV 0.5 mg daily for 52 weeks; 37 patients completed 52 weeks of treatment. At Week 48, 29/40 (72.5%, noncompleter = failure) patients achieved the primary endpoint of HBV DNA <50 IU/mL. Rates for HBeAg loss (11/22; 50%) and HBeAg seroconversion (9/22; 41%) were high, possibly due to the high HBV genotype A prevalence (70%). No patient experienced virological breakthrough. Samples for resistance testing were available in 6/8 patients with HBV DNA >50 IU/mL at Week 48 or last on-treatment visit. No ETV resistance was detected. The safety profile of ETV was consistent with that observed in ETV registration trials. This study shows that in Black/African American patients with CHB, ETV was well tolerated and demonstrated comparable antiviral efficacy to that observed in White and Asian patients in ETV Phase III studies., (© 2013 John Wiley & Sons Ltd.)

Traditional Chinese herbal medicine (TCHM) has been widely used in the treatment of chronic hepatitis B (CHB) in China. The systematic analysis of clinical research of TCHM against CHB revealed its potential but not confirmed its therapeutic effect. To understand the detailed antiviral effect of TCHM against HBV infection, we systematically analysed the anti-HBV effect of individual Chinese herbs on the basis of the research on individual TCHM in vitro and in vivo, which were published from 1995 to 2012. Among 171 herbal components isolated from 76 Chinese herbs, we found 13 compounds and 9 extracts isolated from 18 Chinese herbs showing strong inhibitory effect on HBV DNA, HBeAg or HBsAg release with low cytotoxicity in HepG2.2.15 cells, and agents from 12 Chinese herbs showing the highest inhibition rates of plasma DHBV DNA of more than 50% in DHBV-infected ducks. In addition, the two compounds chrysophanol 8-O-beta-D-glucoside isolated from Rheum palmatum and wogonin isolated from Scutellaria baicalensis were found to display strong anti-HBV activity. Interestingly, compounds isolated from 5 of these effective anti-HBV Chinese herbs were found to show strong antibacterial or antifungal activity also. This review summarizes and analyses the studies on the anti-HBV effect of individual TCHM in cell and animal models, providing potential perspective in the understanding of TCHM in the treatment of hepatitis B and the development of new anti-HBV drugs from TCHM., (© 2013 John Wiley & Sons Ltd.)

Telbivudine, one of the oral anti-hepatitis B virus (HBV) nucleoside analogues, has been used for more than 5 years to treat HBeAg-positive and -negative chronic hepatitis B (CHB) patients. This includes not only global Phase II and III trials, but also Chinese Phase III trials and several real-world clinical practice studies from China. The present review will first focus on 1-, 2- and 3-year data of telbivudine therapy on HBV DNA suppression, alanine aminotransferase (ALT) normalization, hepatitis B e antigen (HBeAg) seroconversion, viral resistance and safety in HBeAg-positive and -negative CHB patients. Second, telbivudine treatment predictors, including HBV DNA undetectability, ALT level and other immune-related markers at 12 and 24 weeks will be summarized to optimize therapy. Besides several retrospective studies, the Chinese EFFORT prospective study adapted from telbivudine virological response at 24 weeks has shown very promising results. Finally, the Chinese experience of using telbivudine in the second and third trimesters of pregnancy to prevent perinatal transmission of HBV infection will be touched on citing the latest studies., (© 2012 Blackwell Publishing Ltd.)

The aim of this study was to evaluate the efficacy and safety of telbivudine 600 mg/day compared with lamivudine 100 mg/day for 48 weeks of treatment in patients with hepatitis B-related cirrhosis. Data were reviewed retrospectively from 165 hepatitis B-related cirrhotic patients (55 compensated patients and 110 decompensated) who received antiviral therapy with telbivudine or lamivudine. Serum alanine aminotransferase (ALT) and hepatitis B virus (HBV) DNA levels, hepatitis B e antigen (HBeAg) loss and seroconversion, histological improvement and various adverse events (AEs) were evaluated. Baseline characteristics were comparable. ALT levels declined but showed no significant difference in treatment with telbivudine or lamivudine (P > 0.05). Reduction in serum HBV DNA levels was evident by week 4 in compensated HBV-related cirrhosis patients (telbivudine, 2.34 log10 copies/mL; lamivudine, 2.07 log10 copies/mL; P = 0.02) and persisted by week 8. Patients administrated with telbivudine had slightly greater HBeAg loss and seroconversion than patients with lamivudine, but the difference was not statistically significant (P > 0.05). Accumulative HBeAg loss was seen at week 48 (25.0% vs 25.0% and 13.3% vs 10.0% for telbivudine vs lamivudine in compensated and decompensated cirrhotic groups, respectively), as well as HBeAg seroconversion (15.0% vs 8.3% and 8.9% vs 6.7%). Mean Knodell Histologic Activity Index scores decreased in both compensated and decompensated cirrhotic patients (3.92 vs 3.64, 3.85 vs 3.73, for telbivudine vs lamivudine). Telbivudine and lamivudine were both well tolerated with minor AEs. The results of this study support telbivudine as an effective therapy for patients with both compensated and decompensated HBV-related cirrhosis., (© 2012 Blackwell Publishing Ltd.)

The aim of this study was to investigate the efficacy of sequential use of telbivudine in hepatitis B e antigen (HBeAg)-positive chronic hepatitis B patients with partial responses to pegylated interferon. Patients with partial responses to 48 weeks of pegylated interferon treatment were divided into two groups. In group A, patients stopped pegylated interferon directly without sequential treatment. In group B, patients received sequential treatment with telbivudine 600 mg/day. HBeAg, HBeAb, hepatitis B virus (HBV) DNA, alanine aminotransferase (ALT) and creatine kinase levels were determined at baseline and at weeks 12, 24, 36 and 48. Responses and safety were assessed after 48 weeks of telbivudine treatment. Thirty-six patients were recruited. Eighteen of these patients stopped pegylated interferon without sequential treatment (group A). After 48 weeks of follow-up, five patients (28%) had undergone HBeAg seroconversion, nine patients (50%) had undetectable levels of HBV DNA, and 11 patients (61%) achieved normal alanine aminotransferase (ALT) levels. The other 18 patients received sequential telbivudine treatment (group B). After 48 weeks of treatment, 11 patients (61%) had undergone HBeAg seroconversion, and all patients had undetectable levels of HBV DNA and normal ALT levels. All patients tolerated sequential telbivudine treatment, and only slightly elevated creatine kinase levels were observed. Switching to telbivudine therapy was efficient and safe in HBeAg-positive chronic hepatitis B patients with partial responses to 48 weeks of pegylated interferon. Sequential treatment with telbivudine resulted in an HBeAg seroconversion rate of 61% and an HBV DNA loss rate of 100% after 48 weeks. This promising strategy warrants further investigation., (© 2012 Blackwell Publishing Ltd.)

Telbivudine (LdT) has demonstrated potent antiviral activity in nucleos(t)ide analogue (NA)-naïve chronic hepatitis B patients (CHB), but data on its efficacy in NA-experienced patients are limited. The aim of this study was to investigate the effect of LdT in hepatitis B e antigen-positive CHB patients with poor response to initial adefovir dipivoxil (ADV). Forty-two CHB patients with HBV DNA > 4 log10  copies/mL after 12 months of ADV monotherapy were enroled in the study and thereafter treated with LdT 600 mg daily for 18 months. Telbivudine led to a rapid decrease in viral load, and viral replication was persistently suppressed with a reduction of 2.26 log10  copies/mL 18 months after LdT treatment. The rates corresponding to virological and biochemical response at the end of observation were 97.6% (41/42) and 65.8% (25/38), respectively. HBeAg loss was found in 30.8% (12/39) of patients, while HBeAg/anti-HBe seroconversion was found in 17.9% (7/39). Only one patient was detected to have LdT-associated mutation, and no severe adverse events were reported. Optimization therapy with LdT monotherapy may be a good choice for CHB patients with poor response to ADV, and switching to LdT may be the most cost-effective rescue therapeutic strategy for patients with poor response to initial ADV monotherapy., (© 2012 Blackwell Publishing Ltd.)

α-Galactosylceramide (α-GalCer)-activated natural killer T (NKT) cells have antiviral properties against hepatitis B virus (HBV). However, α-GalCer activation of NKT cells can induce anergy. We hypothesized that this effect may be overcome by a treatment strategy that includes manipulation of CD28/CD80 costimulatory and PD-1/PDL1 coinhibitory signals of NKT cells, thereby enhancing the anti-HBV effect of α-GalCer. We established a transgenic mouse model of chronic HBV infection and investigated hepatic NKT cell frequencies, functions and expression of immunomodulatory factors. Our results showed that compared with uninfected control mice, hepatic NKT cells from HBV transgenic mice displayed lower frequencies (7.91% vs 16.74%, P < 0.05), impaired capabilities to produce interferon (IFN)-γ (5.6% vs 1.4%, P < 0.05) and interleukin (IL)-4 (6.8% vs 0.3%, P < 0.05), higher expression of PD-1 (9.64% vs 6.36%, P < 0.05) and lower expression of CD28 (5.05% vs 28.88%, P < 0.05). However, when hepatic mononuclear cells (MNCs) were isolated from HBV transgenic mice, α-GalCer exposure in culture remarkably upregulated both PD-1(+) NKT cells (P < 0.05) and CD28(+) NKT cells (P < 0.05). Furthermore, when HBV transgenic mice were treated with combination therapies consisting of α-GalCer and anti-PDL1 monoclonal antibody (mAb) and/or anti-CD80/anti-CD28 mAbs, IFN-γ(+) NKT cell frequency was selectively increased (P < 0.05) and HBV replication was suppressed; these effects were accompanied by varying degrees and types of liver damage. Surprisingly, activating CD28/CD80 signal in HBV transgenic mice was more effective but caused less liver injury than blocking PD-1/PDL1 signal in modulating αGalCer-activated NKT cell function to inhibit HBV infection. Our findings also show that combined therapy with blocking PD-1/PDL1 and activating CD28/CD80 signal in the presence of aGalCer cannot superimpose the effect of antivirus. α-GalCer combination therapy that modulates the CD28/CD80 pathways of NKT cells may represent a promising approach to inhibit HBV replication in chronically infected patients., (© 2012 Blackwell Publishing Ltd.)

The dynamics of programmed death-1 (PD-1) as well as cytokine/chemokine expression and its correlation with virological response in patients with chronic hepatitis B (CHB) is unclear. This study was conducted in 29 treatment-naïve patients undergoing telbivudine treatment for 52 weeks. The results showed that PD-1 expression on both CD4+ and CD8+ T cells was positively correlated with hepatitis B virus (HBV) DNA levels (r = 0.621, P < 0.0001; r = 0.548, P = 0.002, respectively), and in virological responders, this decrease was directly correlated with a decrease in HBV DNA levels (r = 0.664, P = 0.002; r = 0.572, P = 0.01, respectively). Furthermore, at the end of 52 weeks, in virological responders, the decreased rate in the frequency of PD-1+ CD8+ T cells was significantly higher than in non-virological responders (58.3% vs 25.7%, P = 0.001), and at weeks 24 and 52, in virological responders, PD-1 expression on CD4+ and CD8+ T cells was lower than in non-virological responders (P = 0.01 and P = 0.035; P < 0.0001 and P < 0.0001, respectively). In 34 cytokines/chemokines detected in serum, IP-10 expression was positively correlated with viral load, level of ALT and PD-1 expression on CD8+ and CD4+ T cells at baseline (r = 0.36, P = 0.055, r = 0.635, P < 0.0001, r = 0.414, P = 0.026, and r = 0.402, P = 0.030, respectively). Moreover, the decrease in IP-10 in serum directly correlated with a decrease in ALT levels (r = 0.751, P < 0.0001). At weeks 24 and 25, IP-10 expression was significantly lower than baseline in virological responders (both P = 0.005); however, this was not observed in nonresponders. Based on the above findings, PD-1 and IP-10 may be used as predictors for virological response, and blockade of their pathway may improve the outcome of patients with CHB., (© 2012 Blackwell Publishing Ltd.)

We characterized the early dynamics of hepatitis B virus (HBV) quasispecies evolution during the first weeks of antiviral therapy with low-to-moderate genetic barrier antiviral drugs and associated these data with antiviral response patterns. Fifteen chronic hepatitis B patients (men, 10; mean age, 34; HBeAg positive, 6) who received lamivudine or telbivudine for at least 52 weeks were included. HBV DNA was extracted from serum, and a 910-bp fragment covering domains A-F of the reverse transcriptase region was amplified, cloned and sequenced. Parameters of quasispecies heterogeneity, genetic diversity and complexity were calculated and were correlated with complete virologic response, defined as undetectable HBV DNA at week 52. Nine patients achieved complete virologic response during the observational period. While baseline HBV DNA levels and HBeAg status were associated with virologic response, baseline quasispecies complexity and diversity of responders showed no significant difference to those of nonresponders (P > 0.05). However, at week 4, quasispecies complexity of nonresponders was significantly higher compared with that of responders on the nucleotide level (P = 0.01) and the aa level (P = 0.04). The number of synonymous substitutions per synonymous site dropped significantly in responders at week 4 (P = 0.04), while there was no difference in nonresponders. The HBV quasispecies complexity at the early stage of antiviral therapy (week 4) with the low-to-moderate genetic barrier nucleoside analogs lamivudine or telbivudine was associated with subsequent virologic response. Further studies are needed to confirm HBV quasispecies evolution as additional predictive marker for beneficial treatment outcome., (© 2012 Blackwell Publishing Ltd.)

Orally administered nucleus(t)ide analogues (NA) have brought about a simple, safe and effective therapeutic approach for chronic hepatitis B (CHB). However, treatment duration is long and some female patients become pregnant during treatment. In recent years, there have been gradually increasing reports on the safety of telbivudine (LdT) treatment for chronic hepatitis B virus (HBV) infection in the third trimester of pregnancy to block mother-to-infant transmission (MTIT) of HBV; however, the safety of LdT treatment for chronic HBV infection for the entire pregnancy has not been reported. The aim of the present study was to evaluate the safety of LdT treatment for chronic HBV infection for the entire pregnancy and provide a reference for HBV-infected fertile women on how to block MTIT of HBV. Eighty-six pregnant women who received LdT treatment either before or in early pregnancy were enrolled in the study. Adverse events were prospectively observed for the entire pregnancy and perinatal period, and short-term and long-term follow-up of infants was conducted, monitoring the abnormalities of infants and blocking rate of MTIT with LdT treatment. Eighty-six pregnant women treated with LdT had a total of 89 pregnancies: 6 (6.7%) had early embryonic death or spontaneous abortion, 1 (1.1%) had ectopic pregnancy and three had a second pregnancy after initial abortion. Fifty-one mothers completed pregnancy: one had induction of labour at 24 weeks of pregnancy for cleft lip and palate of the foetus and 50 delivered 52 full-term live infants. One infant had right ear accessories, and the total occurrence of congenital abnormality was 3.8%. Thirty-nine infants were followed up for more than 6 months and completed all examinations for MTIT. None of the infants were HBsAg positive, resulting in a 100% success rate of blocking MTIT. All mothers maintained good liver function during the third trimester of pregnancy; 86% maintained complete virological response (HBV DNA <500 copies/mL) prior to delivery, and none developed progression of liver disease. Factors leading to increased adverse effects and drug resistance were not found. LdT treatment is safe and effective in chronic HBV-infected pregnant mothers for the entire pregnancy., (© 2012 Blackwell Publishing Ltd.)

The objective of this study was to compare the efficacy and safety of two rescue strategies for hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) patients with resistance to adefovir dipivoxil (ADV). This prospective study included 58 HBeAg-positive CHB patients with resistance to ADV; 30 patients underwent telbivudine (LdT) plus ADV combination therapy and 28 patients switched to entecavir (ETV) monotherapy. After 48 weeks of treatment, the rates of hepatitis B virus (HBV) DNA <3 log10 copies/mL in the LdT plus ADV group and the ETV group were not significantly different (73.3% vs 57.1%, P = 0.195). Six patients receiving LdT plus ADV had HBeAg seroconversion, while none of the patients receiving ETV alone had HBeAg seroconversion (20% vs 0%, P = 0.039). During the 48-week treatment period, two patients in the ETV monotherapy group had viral breakthrough and the strains were confirmed to be of a variant associated with ETV resistance (rtM204V+ rtL180M+ rtT184G), while one patient receiving LdT plus ADV had viral breakthrough and an LdT-associated resistance mutation (rtM204I) was detected. For the majority of the patients, both LdT plus ADV combination treatment or ETV monotherapy were generally well tolerated, and no serious side effects were observed. Both LdT plus ADV combination therapy and ETV monotherapy led to significant decreases in serum HBV DNA in HBeAg-positive CHB patients with resistance to ADV, and LdT plus ADV combination therapy exhibited a significantly higher rate of HBeAg seroconversion compared with ETV monotherapy., (© 2012 Blackwell Publishing Ltd.)

Hepatitis B virus (HBV) pol/RT mutations that confer clinical resistance to tenofovir disoproxil fumarate (TDF) have not been detected to date. In vitro, the rtN236T adefovir dipivoxil (ADV)-associated resistance mutation confers low-level cross-resistance to tenofovir: 3- to 13-fold changes in EC(50) from wild type. This study evaluated the clinical response of rtN236T mutant viruses by comparing their early viral load decay kinetics to wild-type viruses in chronic HBV monoinfected patients harbouring rtN236T prior to initiating TDF or emtricitabine (FTC)/TDF therapy. Baseline samples (n = 105) from adefovir refractory patients were tested for the presence of rtN236T using a highly sensitive allele-specific PCR assay with an rtN236T detection cut-off of 0.5%. The rtN236T mutation was detected at baseline in 14.3% (14/98) of analysable patient samples (0.5-93.2%, rtN236T percentage range). The median change in total HBV DNA at week 24 was comparable for patients with rtN236T detected at baseline (-3.7 log(10) copies/mL, n = 14) as compared to patients with wild-type HBV (-3.2 log(10) copies/mL, n = 90). In patients with rtN236T, wild-type and rtN236T mutant virus showed similar rates of HBV DNA decline with no statistically significant difference observed at week 4. Moreover, the proportion of rtN236T remained unchanged in patients in either arm of the study during treatment. In conclusion, the rtN236T mutant virus showed similar HBV DNA decline kinetics to wild-type virus in adefovir refractory patients who switched to TDF or FTC/TDF. Despite low levels of cross-resistance in vitro, TDF similarly suppresses wild-type and rtN236T mutant viruses in vivo., (© 2012 Blackwell Publishing Ltd.)

Patients with chronic hepatitis B virus (HBV) infection are at an increased risk for a severe and a potentially fatal viral reactivation following anti-cancer therapy. The molecular mechanism for this induction of HBV expression is still unclear. Here, we show that treating hepatoma cell line expressing HBV with various anti-cancer cytotoxic agents results in a significant up-regulation of HBV expression. This HBV induction is at the transcriptional level and is time dependent. Interestingly, treating hepatoma cells with anti-cancer cytotoxic agents results in a robust induction of peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α), a metabolic and energy regulator that is normally induced in the liver under starvation conditions and that has been previously shown to strongly coactivate HBV transcription. Most importantly, HBV up-regulation following anti-cancer therapy depends on PGC-1α induction, because PGC-1α knock-down abolishes HBV induction. Finally, pretreatment of HBV-expressing cells with the antioxidant agent N-acetylcysteine attenuates the induction of both PGC-1α and HBV in response to anti-cancer treatment, suggesting that chemotherapy-associated PGC-1α induction is mediated by cellular oxidative stress that ultimately leads to HBV up-regulation. We conclude that cytotoxic anti-cancer chemotherapy has a direct and an immune system-independent effect on HBV gene expression, which is mediated by PGC-1α. Our results attribute to this metabolic regulator an unexpected role in linking anti-cancer treatment to HBV reactivation and make PGC-1α a potential target for future anti-HBV therapy, especially under conditions in which it is robustly induced, such as following anti-cancer treatment., (© 2012 Blackwell Publishing Ltd.)

Data on the efficacy of lamivudine (LAM)-, tenofovir (TDF)- and emtricitabine (FTC)-based antiretroviral therapy (HAART) in HBV-HIV coinfection are limited. We completed a retrospective analysis of HBV-HIV-coinfected patients treated at the Medical University of Vienna. One-hundred and ten coinfected patients were included, with 57% being initially HBV e-Antigen (HBeAg) positive. Baseline HBV load was significantly higher in HBeAg+ than in HBeAg- patients (5962 ± 3663 vs 20 ± 19 × 10(6) IU/mL; P < 0.0001). Over a median observation period of 83 month (range: 26-183), 87% received HAART and 91% showed a suppression of HBV replication. After 5 years of continuous treatment, HBeAg seroconversion was achieved in 21% of LAM-, 50% of TDF- (P = 0.042 vs LAM) and in 57% of TDF + FTC (P = 0.008 vs LAM)-treated patients, respectively. HBsAg loss after 5 years was found in 8% (LAM), 25% (TDF; P = 0.085 vs LAM) and 29% (TDF + FTC; P = 0.037 vs LAM) of HBeAg+ patients. In HBeAg- patients, HBsAg loss was achieved in 11% (LAM), 27% (TDF; P = 0.263 vs LAM) and 36% (TDF + FTC; P = 0.05 vs LAM), respectively. Pretreatment CD4+ counts did not influence rates of HBeAg seroconversion and of HBsAg loss. Patients with HBsAg loss had lower baseline HBV-DNA levels and higher AST/ALT levels than patients without HBsAg loss. Transient HAART-related hepatotoxicity was found in 32% (Grade I: 21%; II:7%; III:2%; IV:0%). Most HBV-HIV-coinfected patients achieve complete suppression of HBV replication despite high baseline viremia. TDF-based HAART leads to high rates of HBeAg seroconversion and HBsAg loss after 5 years of continuous exposure. One-third of HBV-HIV-coinfected patients may experience transient HAART-related hepatotoxicity., (© 2012 Blackwell Publishing Ltd.)

Both entecavir (ETV) and tenofovir (TDF) are potent antiviral agents for hepatitis B virus (HBV). Suboptimal response (SOR) following antiviral therapy is associated with an increased risk of subsequent treatment failure and viral resistance. It remains unclear whether switching to TDF is a reasonable approach in patients with SOR to ETV treatment. This study was aimed to determine how HBV patients with SOR to ETV respond to TDF monotherapy. Data of patients with SOR to ETV (failure to achieve >1 log(10) HBV-DNA reduction during the last 24 weeks of ETV treatment) who were switched to TDF monotherapy during 2005 and 2010 were reviewed. Treatment adherence was assessed by pill-count. Fourteen patients (2.9%) were identified from a total cohort of 482 ETV-treated patients. All 14 patients were Chinese and were infected with HBV genotype C (71%) or B (29%). Nine patients were men, and the median age was 41.5 years (19-64). Twelve were treatment naïve (one lamivudine- and one peginterferon-experienced patient); 85.7% were HBeAg positive. The median baseline HBV-DNA was 7.55 (5.30-9.40) log(10) copies/mL, and 57% had abnormal serum alanine aminotransferase (ALT) levels. Precore and/or basal core promoter mutations were detected in four patients, whereas no genotypic resistance was detected at baseline and before switching to TDF. The median duration of ETV treatment was 64.5 (26-126) weeks. The median HBV-DNA at the time of switching to TDF was 3.69 (3.00-4.90) log(10) copies/mL. The median HBV-DNA reduction from baseline and during the last 6-month observation period prior to switching to TDF was 4.04 (0.51-6.06) log(10) and 0.43 (-0.09-1.13) log(10) copies/mL, respectively. After the switching to TDF, all 14 patients (100%) achieved undetectable HBV-DNA and ALT normalization within a median duration of 30 weeks. In 12 patients who were HBeAg positive, HBeAg seroconversion was observed in two patients after TDF treatment of 75- and 84-weeks duration. There was no virological breakthrough observed after switching to TDF with a median follow-up period of 50 (24-160) weeks. TDF treatment was safe and well tolerated. In conclusion, suboptimal response to ETV is rare (approximately 3%). TDF monotherapy is safe and very effective in the management of HBV patients with SOR to ETV., (© 2011 Blackwell Publishing Ltd.)

The focus of this study was to evaluate the safety and efficacy of sequential peginterferon α-2a (Pegasys) therapy for chronic hepatitis B with acute exacerbation [ALT > 10 × upper limit of normal (ULN), bilirubin <2.0 mg/dL]. Four groups of patients categorized by HBeAg status and treatment regimens were studied since May 2007. Nineteen HBeAg-positive patients (Group 1) had received entecavir  pretreatment  (when ALT > 10 × ULN) plus Pegasys (180 μg/kg/week, when ALT was 5-10 × ULN) for 24 weeks. Thirteen HBeAg-negative patients (Group 2) had the same protocol for 48 weeks. In both groups, entecavir was then discontinued 14 days after the initiation of Pegasys. The results were compared, respectively, to 35 HBeAg-positive patients (Group 3) and 24 HBeAg-negative patients (Group 4), all with ALT > 5 × ULN, under continual entecavir monotherapy. The ALT levels of patients in Group 1 and 2 who had received entecavir pretreatment for a duration of 19.63 ± 3.34 days were below four times of ULN following 4 weeks of Pegasys treatment. At week 96, the rates of sustained virological response were 69.2% (9/13) and 80% (8/10), and the relapse rates were 23.1% (3/13) and 11.2% (1/9) for HBeAg-positive and HBeAg-negative patients with two-step Pegasys treatment, respectively. The HBeAg seroconversion rates were 46.2% in Group 1, and 42.1% in Group 3; HBsAg loss rates were 15.4% (2/13) in Group 1, and 30% (3/10) in Group 2, whereas none achieved HBsAg loss with entecavir monotherapy (Group 3 and 4). The two-step Pegasys treatment offers an alternative, other than the nucleos(t)ides, for treating chronic hepatitis B with acute exacerbation and provides a safe, efficacious, short-term and finite strategy., (© 2011 Blackwell Publishing Ltd.)

Hepatitis B e antigen (HBeAg)-negative hepatitis B commonly requires long-term treatment with nucleos(t)ide analogues aiming at persistently suppressing hepatitis B virus (HBV) replication to halt progression of liver disease and prevent complications. Entecavir (ETV) is widely used in HBeAg-negative hepatitis B, but distinct HBV polymerase mutations can confer resistance against ETV, in conjunction with lamivudine resistance. Precore (PC) and basal core promoter (BCP) mutations that underlie HBeAg-negativity enhance replication of lamivudine-resistant mutants. To comprehensively analyse the impact of PC or BCP mutations on viral replication of ETV-resistant HBV mutants, replication-competent HBV constructs were generated harbouring lamivudine resistance (rtM204V/rtL180M, rtM204I) plus ETV resistance (rtS202G, rtS202I or rtT184G) on wild-type (WT)-, PC- and BCP-backgrounds. Functional consequences on viral fitness and susceptibility to antivirals were assessed in vitro. The presence of any ETV resistance drastically reduced viral replication when compared to WT HBV. In rtS202G mutants (plus lamivudine resistance), addition of either PC or BCP mutations moderately enhanced the reduced replication, without reaching WT HBV levels. In rtS202I or rtT184G mutants, PC and BCP mutations did not significantly improve viral fitness. All ETV-resistant constructs, independently of PC or BCP mutations, showed resistance towards ETV and lamivudine, but remained susceptible to tenofovir. Our data demonstrate that HBeAg-suppressing PC or BCP mutations cannot restore the strongly reduced replicative capacity of ETV-resistant HBV mutants to WT level, although they moderately increase replication of rtS202G combination mutants. ETV resistance thereby differs from lamivudine resistance alone, corroborating that ETV is in short term a safe option for HBeAg-negative patients., (© 2010 Blackwell Publishing Ltd.)

Understanding the consequences of mutation in the tyrosine-methionine-aspartate-aspartate (YMDD) motif of hepatitis B virus (HBV) genome on replication is critical for treating chronic hepatitis B with lamivudine. Allele-specific gene silencing by RNAi (allele-specific RNAi: ASP-RNAi) is an advanced application of RNAi techniques. Use of this strategy as a means for specifically inhibiting an allele expression of interest suggested that it can specifically suppress the expression of alleles causing disease without inhibiting the expression of corresponding wild-type alleles. However, no studies have used ASP-RNAi to address the issue of HBV lamivudine resistance. In this study, we applied ASP-RNAi into two long-term eukaryotic cell lines of full-length HBV containing either lamivudine-resistant mutants (HBV-YIDD) or wild type (HBV-WT) which we generated in previously. The designed siRNAs were also used in this eukaryotic expression system together with lamivudine. ELISA and real-time PCR were performed to monitor virus-specific protein synthesis and viral DNA replication. The results showed that the base substitutions conferring marked ASP-RNAi appeared to be largely present in positions 1, 3, 6, 11, 12, 15 and 19 of the sense strand of siRNAs which were different from the most sensitive positions of this application in eukaryotes. In addition, siRNA-lamivudine combinations did not possess the prominent anti-HBV activity we expected because of some unknown mechanisms. These findings recapitulated many of the features of ASP-RNAi in hepadnaviruses which provided a new insight into the development of a potent strategy against HBV drug resistance., (© 2011 Blackwell Publishing Ltd.)

This study aimed to evaluate the long-term efficacy of entecavir (ETV) in adefovir (ADV)-refractory chronic hepatitis B (CHB) patients with prior lamivudine (LMV) resistance. A total of 55 ADV-refractory CHB patients with prior LMV resistance, who received rescue therapy with ETV 1 mg daily for at least 12 months, were consecutively enrolled and analysed. Forty-four patients were men, and their median age was 47 (25-69). Ten patients had liver cirrhosis and 46 patients were positive for hepatitis B e antigen (HBeAg). Median hepatitis B virus DNA levels were 6.6 (4.3-8.0) log(10) copies/mL, and the median duration of ETV therapy was 24 (12-47) months. Cumulative virologic response rates at 6, 12, 24 and 36 months were 18%, 29%, 58% and 75%, respectively. HBeAg loss occurred in 10 (21.7%) of 46 HBeAg-positive patients. In multivariate analysis, only initial virologic response at 3 months remained as an independent predictor for virologic response (RR 3.143; 95% CI 1.387-7.120; P = 0.006). The patients with a virological response at 3 months had not only a significantly higher probability of achieving a virologic response (P < 0.001) but also lower probability of experiencing a virologic breakthrough (P = 0.043) than the patients without an early response. Viral breakthrough was observed in 29 patients during the follow-up period. Cumulative breakthrough rates at 6, 12, 24 and 36 months were 0%, 15%, 45% and 73%, respectively. ETV monotherapy may be considerably efficacious in cases with an initial virological response but its efficacy is attenuated by frequent emergence of ETV resistance in ADV-refractory CHB patients with prior LMV resistance., (© 2011 Blackwell Publishing Ltd.)