Single-cell landscape of functionally cured chronic hepatitis B patients reveals activation of innate and altered CD4-CTL-driven adaptive immunity.

BACKGROUND & AIMS: Hepatitis B surface antigen (HBsAg) loss or functional cure (FC) is considered the optimal therapeutic outcome for patients with chronic hepatitis B (CHB). However, the immune-pathological biomarkers and underlying mechanisms of FC remain unclear. In this study we comprehensively interrogate disease-associated cell states identified within intrahepatic tissue and matched PBMCs (peripheral blood mononuclear cells) from patients with CHB or after FC, at the resolution of single cells, to provide novel insights into putative mechanisms underlying FC. METHODS: We combined single-cell transcriptomics (single-cell RNA sequencing) with multiparametric flow cytometry-based immune phenotyping, and multiplexed immunofluorescence to elucidate the immunopathological cell states associated with CHB vs. FC. RESULTS: We found that the intrahepatic environment in CHB and FC displays specific cell identities and molecular signatures that are distinct from those found in matched PBMCs. FC is associated with the emergence of an altered adaptive immune response marked by CD4 cytotoxic T lymphocytes, and an activated innate response represented by liver-resident natural killer cells, specific Kupffer cell subtypes and marginated neutrophils. Surprisingly, we found MHC class II-expressing hepatocytes in patients achieving FC, as well as low but persistent levels of covalently closed circular DNA and pregenomic RNA, which may play an important role in FC. CONCLUSIONS: Our study provides conceptually novel insights into the immuno-pathological control of HBV cure, and opens exciting new avenues for clinical management, biomarker discovery and therapeutic development. We believe that the discoveries from this study, as it relates to the activation of an innate and altered immune response that may facilitate sustained, low-grade inflammation, may have broader implications in the resolution of chronic viral hepatitis. IMPACT AND IMPLICATIONS: This study dissects the immuno-pathological cell states associated with functionally cured chronic hepatitis B (defined by the loss of HBV surface antigen or HBsAg). We identified the sustained presence of very low viral load, accessory antigen-presenting hepatocytes, adaptive-memory-like natural killer cells, and the emergence of helper CD4 T cells with cytotoxic or effector-like signatures associated with functional cure, suggesting previously unsuspected alterations in the adaptive immune response, as well as a key role for the innate immune response in achieving or maintaining functional cure. Overall, the insights generated from this study may provide new avenues for the development of alternative therapies as well as patient surveillance for better clinical management of chronic hepatitis B.

Antiviral therapy substantially reduces HCC risk in patients with chronic hepatitis B infection in the indeterminate phase.

BACKGROUND AND AIMS: HCC risk in chronic hepatitis B (CHB) is higher in the indeterminate phase compared with the inactive phase. However, it is unclear if antiviral therapy reduces HCC risk in this population. We aimed to evaluate the association between antiviral therapy and HCC risk in the indeterminate phase. APPROACH AND RESULTS: We analyzed 855 adult (59% male), treatment-naïve patients with CHB infection without advanced fibrosis in the indeterminate phase at 14 centers (USA, Europe, and Asia). Inverse probability of treatment weighting (IPTW) was used to balance the treated (n = 405) and untreated (n = 450) groups. The primary outcome was HCC development. The mean age was 46±13 years, the median alanine transaminase was 38 (interquartile range, 24-52) U/L, the mean HBV DNA was 4.5±2.1 log 10 IU/mL, and 20% were HBeAg positive. The 2 groups were similar after IPTW. After IPTW (n = 819), the 5-, 10-, and 15-year cumulative HCC incidence was 3%, 4%, and 9% among treated patients (n = 394) versus 3%, 15%, and 19%, among untreated patients (n = 425), respectively ( p = 0.02), with consistent findings in subgroup analyses for age >35 years, males, HBeAg positive, HBV DNA>1000 IU/mL, and alanine transaminase

Switching to or Add-on Peginterferon in Patients on Nucleos(t)ide Analogues for Chronic Hepatitis B: The SWAP RCT.

BACKGROUND & AIMS: The optimal therapeutic strategy in nucleoside analogue (NA) experienced chronic hepatitis B (CHB) using peginterferon is still unclear; hence we explored a switch to or add-on peginterferon strategy versus continued NA. METHODS: We conducted a randomized controlled trial of CHB patients on NA >12 months with HBV DNA(-) randomized to switch or add-on peginterferon-alpha2b (1.5 µg/kg/weekly) for 48 weeks versus continuing NA (controls) (allocation 2:2:1; Clinicaltrial.gov: NCT01928511) in tertiary Singapore hospitals. The primary composite endpoint at week 72 was hepatitis B e antigen (HBeAg) loss or quantitative HBsAg (qHBsAg) >1 log IU/mL reduction, and secondary endpoints were HBsAg loss, HBsAg seroconversion, qHBsAg <200 IU/mL, qHBsAg <100 IU/mL, HBV DNA(-), viral relapse, and safety. Analysis was by intention-to-treat (ITT). RESULTS: A total of 253 patients (controls 51, switch 103, add-on 99) were randomized. The primary ITT endpoint was achieved in 3.9% of controls, 33.3% of switch, and 26.7% of add-on (P < .0001, switch/add-on versus controls). HBsAg loss occurred in 0% of controls, 7.8% of switch, and 10.1% of add-on (ITT, P < .001, switch/add-on versus controls). HBeAg(+) patients on peginterferon had higher HBeAg loss than controls but poor HBsAg responses, whereas HBeAg(-) patients on peginterferon achieved better HBsAg responses than controls. Reduction in qHBsAg in HBeAg(+) was 0.14 log IU/mL versus 0.51 log IU/mL in HBeAg(-) (P < .0001) in peginterferon-treated patients. Clinical relapse was higher in switch (13.6% overall, 27% in HBeAg(+)) versus 1% add-on and 0% controls. Adverse events were typically interferon-related symptoms, with one death (myocardial infarction unrelated to therapy). CONCLUSIONS: ITT analysis showed that either peginterferon strategies were superior to NA for the primary endpoint and HBsAg loss, but add-on peginterferon is preferred to switch due to improved safety and similar efficacy. ClincialTrials.gov number: NCT01928511.

Longitudinal renal changes in chronic hepatitis B patients treated with entecavir versus TDF: a REAL-B study.

BACKGROUND AND AIMS: We aimed to compare the longitudinal changes in estimated glomerular filtration rate (eGFR) in chronic hepatitis B (CHB) patients treated with entecavir (ETV) vs. tenofovir disoproxil fumarate (TDF). METHODS: This is a retrospective study of 6189 adult treatment-naïve CHB patients initiated therapy with TDF (n = 2482) or ETV (n = 3707) at 25 international centers using multivariable generalized linear modeling (GLM) to determine mean eGFR (mL/min/1.73 m2) and Kaplan-Meier method to estimate incidence of renal impairment (≥ 1 chronic kidney disease [CKD] stage worsening). We also examined above renal changes in matched ETV and TDF patients (via propensity score matching [PSM] on age, sex, diabetes mellitus [DM], hypertension [HTN], cirrhosis, baseline eGFR, and follow-up duration). RESULTS: In the overall cohort (mean age 49.7 years, 66.2% male), the baseline eGFR was higher for TDF vs. ETV group (75.9 vs. 74.0, p = 0.009). PSM yielded 1871 pairs of ETV or TDF patients with baseline eGFR ≥ 60 and 520 pairs for the eGFR < 60 group. GLM analysis of the overall (unmatched) cohort and PSM cohorts revealed lower adjusted mean eGFRs in TDF (vs. ETV) patients (all p < 0.01) during 10 years of follow-up. Among PSM eGFR ≥ 60 patients, the 5-year cumulative incidences of renal impairment were 42.64% for ETV and 48.03% for TDF (p = 0.0023). In multivariable Cox regression, TDF vs. ETV (adjusted HR 1.26, 95% CI 1.11-1.43) was associated with higher risk of worsening renal function. CONCLUSION: Over the 10-year study follow-up, compared to ETV, TDF was associated with a lower mean eGFR and higher incidence of renal impairment.