Five-year Treatment with Tenofovir Alafenamide Achieves High Rates of Viral Suppression, Alanine Aminotransferase Normalization, and Favorable Bone and Renal Safety in Chinese Chronic Hepatitis B Patients.

Background and Aims: After 3-years (144 week) of double-blind treatment in Chinese chronic hepatitis B patients in two ongoing phase 3 studies, tenofovir alafenamide (TAF) showed similar efficacy to tenofovir disoproxil fumarate (TDF), with improved renal and bone safety. In this study, we aimed to report the 5-year results from 2 years into the open-label TAF treatment phase. Methods: All participants completing the 144-week double-blind treatment were eligible to receive open-label TAF 25 mg once daily up to week 384. Serial analysis of viral suppression (hepatitis B virus DNA <29 IU/mL), alanine aminotransferase normalization, serological responses, and safety outcomes at year 5 (week 240) was performed. Results: The open-label phase included 93% (311/334) of the enrolled participants, which included 212 who switched from double-blind TAF to open-label TAF (TAF-TAF) and 99 who switched from double-blind TDF to open-label TAF (TDF-TAF). Baseline characteristics were comparable. Week 240 viral suppression rates were similar between groups [93.4% vs. 93.9%; difference: -1.5%, (95% CI: -6.4 to -3.5), p=0.857]. Alanine aminotransferase normalization and serological response rates were higher in the TAF-TAF group than in the TDF-TAF group. The frequencies of adverse events and laboratory abnormalities were low and similar between groups. Both groups had similar small numerical declines from baseline in estimated glomerular filtration rate at year 5 (week 240, -2.85 mL/min vs. -3.29 mL/min, p=0.910). The greater declines in renal and bone parameters in the TDF-TAF group through week 144 improved after switching to TAF. Conclusions: The 5-year TAF treatment efficacy was high and similar to that of 3-year TDF followed by 2-year TAF in Chinese chronic hepatitis B patients. Favorable effects on bone and renal parameters were sustained with TAF treatment alone and were observed following the switch from TDF to TAF.

A novel engineered IL-21 receptor arms T-cell receptor-engineered T cells (TCR-T cells) against hepatocellular carcinoma.

Strategies to improve T cell therapy efficacy in solid tumors such as hepatocellular carcinoma (HCC) are urgently needed. The common cytokine receptor γ chain (γc) family cytokines such as IL-2, IL-7, IL-15 and IL-21 play fundamental roles in T cell development, differentiation and effector phases. This study aims to determine the combination effects of IL-21 in T cell therapy against HCC and investigate optimized strategies to utilize the effect of IL-21 signal in T cell therapy. The antitumor function of AFP-specific T cell receptor-engineered T cells (TCR-T) was augmented by exogenous IL-21 in vitro and in vivo. IL-21 enhanced proliferation capacity, promoted memory differentiation, downregulated PD-1 expression and alleviated apoptosis in TCR-T after activation. A novel engineered IL-21 receptor was established, and TCR-T armed with the novel engineered IL-21 receptors (IL-21R-TCR-T) showed upregulated phosphorylated STAT3 expression without exogenous IL-21 ligand. IL-21R-TCR-T showed better proliferation upon activation and superior antitumor function in vitro and in vivo. IL-21R-TCR-T exhibited a less differentiated, exhausted and apoptotic phenotype than conventional TCR-T upon repetitive tumor antigen stimulation. The novel IL-21 receptor in our study programs powerful TCR-T and can avoid side effects induced by IL-21 systemic utilization. The novel IL-21 receptor creates new opportunities for next-generation TCR-T against HCC.

A spatiotemporal atlas of mouse liver homeostasis and regeneration.

The mechanism by which mammalian liver cell responses are coordinated during tissue homeostasis and perturbation is poorly understood, representing a major obstacle in our understanding of many diseases. This knowledge gap is caused by the difficulty involved with studying multiple cell types in different states and locations, particularly when these are transient. We have combined Stereo-seq (spatiotemporal enhanced resolution omics-sequencing) with single-cell transcriptomic profiling of 473,290 cells to generate a high-definition spatiotemporal atlas of mouse liver homeostasis and regeneration at the whole-lobe scale. Our integrative study dissects in detail the molecular gradients controlling liver cell function, systematically defining how gene networks are dynamically modulated through intercellular communication to promote regeneration. Among other important regulators, we identified the transcriptional cofactor TBL1XR1 as a rheostat linking inflammation to Wnt/ß-catenin signaling for facilitating hepatocyte proliferation. Our data and analytical pipelines lay the foundation for future high-definition tissue-scale atlases of organ physiology and malfunction.

Expert consensus on the diagnosis and treatment of end-stage liver disease complicated by infections.

End-stage liver disease (ESLD) is a life-threatening clinical syndrome and when complicated with infection the mortality is markedly increased. In patients with ESLD, bacterial or fungal infection can induce or aggravate the occurrence or progression of liver decompensation. Consequently, infections are among the most common complications of disease deterioration. There is an overwhelming need for standardized protocols for early diagnosis and appropriate management for patients with ESLD complicated by infections. Asia Pacific region has the largest number of ESLD patients, due to hepatitis B and the growing population of alcohol and NAFLD. Concomitant infections not only add to organ failure and high mortality but also to financial and healthcare burdens. This consensus document assembled up-to-date knowledge and experience from colleagues across the Asia-Pacific region, providing data on the principles as well as evidence-based current working protocols and practices for the diagnosis and treatment of patients with ESLD complicated by infections.

Atovaquone enhances antitumor efficacy of TCR-T therapy by augmentation of ROS-induced ferroptosis in hepatocellular carcinoma.

T-cell receptor (TCR) engineered T-cell therapy has recently emerged as a promising adoptive immunotherapy approach for tumor treatment, yet hindered by tumor immune evasion resulting in poor therapeutic efficacy. The introduction of ferroptosis-targeted inducers offers a potential solution, as they empower T cells to induce ferroptosis and exert influence over the tumor microenvironment. Atovaquone (ATO) stands as a prospective pharmaceutical candidate with the potential to target ferroptosis, effectively provoking an excessive generation and accumulation of reactive oxygen species (ROS). In this study, we evaluated the effectiveness of a combination therapy comprising ATO and TCR-T cells against hepatocellular carcinoma (HCC), both in vitro and in vivo. The results of lactate dehydrogenase and cytokine assays demonstrated that ATO enhanced cytotoxicity mediated by AFP-specific TCR-T cells and promoted the release of IFN-γ in vitro. Additionally, in an established HCC xenograft mouse model, the combined therapy with low-dose ATO and TCR-T cells exhibited heightened efficacy in suppressing tumor growth, with no apparent adverse effects, comparable to the results achieved through monotherapy. The RNA-seq data unveiled a significant activation of the ferroptosis-related pathway in the combination therapy group in comparison to the TCR-T cells group. Mechanistically, the synergy between ATO and TCR-T cells augmented the release of IFN-γ by TCR-T cells, while concurrently elevating the intracellular and mitochondrial levels of ROS, expanding the labile iron pool, and impairing the integrity of the mitochondrial membrane in HepG2 cells. This multifaceted interaction culminated in the potentiation of ferroptosis within the tumor, primarily induced by an excess of ROS. In summary, the co-administration of ATO and TCR-T cells in HCC exhibited heightened vulnerability to ferroptosis. This heightened susceptibility led to the inhibition of tumor growth and the stimulation of an anti-tumor immune response. These findings suggest that repurposing atovaquone for adoptive cell therapy combination therapy holds the potential to enhance treatment outcomes in HCC.

Elevated expression of HIGD1A drives hepatocellular carcinoma progression by regulating polyamine metabolism through c-Myc-ODC1 nexus.

BACKGROUND: Hypoxia contributes to cancer progression through various molecular mechanisms and hepatocellular carcinoma (HCC) is one of the most hypoxic malignancies. Hypoxia-inducible gene domain protein-1a (HIGD1A) is typically induced via epigenetic regulation and promotes tumor cell survival during hypoxia. However, the role of HIGD1A in HCC remains unknown. METHODS: HIGD1A expression was determined in 24 pairs of human HCC samples and para-tumorous tissues. Loss-of-function experiments were conducted both in vivo and in vitro to explore the role of HIGD1A in HCC proliferation and metastasis. RESULTS: Increased HIGD1A expression was found in HCC tissues and cell lines, which was induced by hypoxia or low-glucose condition. Moreover, HIGD1A knockdown in HCC cells arrested the cell cycle at the G2/M phase and promoted hypoxia-induced cell apoptosis, resulting in great inhibition of cell proliferation, migration, and invasion, as well as tumor xenograft formation. Interestingly, these anti-tumor effects were not observed in normal hepatocyte cell line L02. Further, HIGD1A knockdown suppressed the expression of ornithine decarboxylase 1 (ODC1), a rate-limiting enzyme of polyamine metabolism under c-Myc regulation. HIGD1A was found to bind with the c-Myc promoter region, and its knockdown decreased the levels of polyamine metabolites. Consistently, the inhibitory effect on HCC phenotype by HIGD1A silencing could be reversed by overexpression of c-Myc or supplementation of polyamines. CONCLUSIONS: Our results demonstrated that HIGD1A activated c-Myc-ODC1 nexus to regulate polyamine synthesis and to promote HCC survival and malignant phenotype, implying that HIGD1A might represent a novel therapeutic target for HCC.

Real-world implementation of a multilevel interventions program to prevent mother-to-child transmission of HBV in China.

Reducing hepatitis B virus (HBV) mother-to-child transmission (MTCT) is a fundamental step toward the HBV elimination goal. The multicentred, multilevel SHIELD program aimed to use an intense intervention package to reduce HBV MTCT in China. This study was conducted in diverse health settings across China, encompassing 30,109 pregnant women from 178 hospitals, part of the interim analysis of stage II of the SHIELD program, and 8,642 pregnant women from 160 community-level health facilities in stage III of the SHIELD program. The study found that the overall MTCT rate was 0.23% (39 of 16,908; 95% confidence interval (CI): 0.16-0.32%) in stage II and 0.23% (12 of 5,290; 95% CI: 0.12-0.40%) in stage III. The MTCT rate was lower among participants who were compliant with the interventions (stage II: 0.16% (95% CI: 0.10-0.26%); stage III: 0.03% (95% CI: 0.00-0.19%)) than among those who were noncompliant (3.16% (95% CI: 1.94-4.85%); 1.91% (95% CI: 0.83-3.73%); P < 0.001). Our findings demonstrate that the comprehensive interventions among HBV-infected pregnant women were feasible and effective in dramatically reducing MTCT.

Cell-free DNA testing for early hepatocellular carcinoma surveillance.

BACKGROUND: Liver cirrhosis (LC) is the highest risk factor for hepatocellular carcinoma (HCC) development worldwide. The efficacy of the guideline-recommended surveillance methods for patients with LC remains unpromising. METHODS: A total of 4367 LCs not previously known to have HCC and 510 HCCs from 16 hospitals across 11 provinces of China were recruited in this multi-center, large-scale, cross-sectional study. Participants were divided into Stage Ⅰ cohort (510 HCCs and 2074 LCs) and Stage Ⅱ cohort (2293 LCs) according to their enrollment time and underwent Tri-phasic CT/enhanced MRI, US, AFP, and cell-free DNA (cfDNA). A screening model called PreCar Score was established based on five features of cfDNA using Stage Ⅰ cohort. Surveillance performance of PreCar Score alone or in combination with US/AFP was evaluated in Stage Ⅱ cohort. FINDINGS: PreCar Score showed a significantly higher sensitivity for the detection of early/very early HCC (Barcelona stage A/0) in contrast to US (sensitivity of 51.32% [95% CI: 39.66%-62.84%] at 95.53% [95% CI: 94.62%-96.38%] specificity for PreCar Score; sensitivity of 23.68% [95% CI: 14.99%-35.07%] at 99.37% [95% CI: 98.91%-99.64%] specificity for US) (P < 0.01, Fisher's exact test). PreCar Score plus US further achieved a higher sensitivity of 60.53% at 95.08% specificity for early/very early HCC screening. INTERPRETATION: Our study developed and validated a cfDNA-based screening tool (PreCar Score) for HCC in cohorts at high risk. The combination of PreCar Score and US can serve as a promising and practical strategy for routine HCC care. FUNDING: A full list of funding bodies that contributed to this study can be found in Acknowledgments section.

Efficacy, safety, and pharmacokinetics of capsid assembly modulator linvencorvir plus standard of care in chronic hepatitis B patients.

BACKGROUND/AIMS: Four-week treatment of linvencorvir (RO7049389) was generally safe and well tolerated, and showed anti-viral activity in chronic hepatitis B (CHB) patients. This study evaluated the efficacy, safety, and pharmacokinetics of 48-week treatment with linvencorvir plus standard of care (SoC) in CHB patients. METHODS: This was a multicentre, non-randomized, non-controlled, open-label phase 2 study enrolling three cohorts: nucleos(t)ide analogue (NUC)-suppressed patients received linvencorvir plus NUC (Cohort A, n=32); treatment-naïve patients received linvencorvir plus NUC without (Cohort B, n=10) or with (Cohort C, n=30) pegylated interferon-α (Peg-IFN-α). Treatment duration was 48 weeks, followed by NUC alone for 24 weeks. RESULTS: 68 patients completed the study. No patient achieved functional cure (sustained HBsAg loss and unquantifiable HBV DNA). By Week 48, 89% of treatment-naïve patients (10/10 Cohort B; 24/28 Cohort C) reached unquantifiable HBV DNA. Unquantifiable HBV RNA was achieved in 92% of patients with quantifiable baseline HBV RNA (14/15 Cohort A, 8/8 Cohort B, 22/25 Cohort C) at Week 48 along with partially sustained HBV RNA responses in treatment-naïve patients during follow-up period. Pronounced reductions in HBeAg and HBcrAg were observed in treatment-naïve patients, while HBsAg decline was only observed in Cohort C. Most adverse events were grade 1-2, and no linvencorvir-related serious adverse events were reported. CONCLUSION: 48-week linvencorvir plus SoC was generally safe and well tolerated, and resulted in potent HBV DNA and RNA suppression. However, 48-week linvencorvir plus NUC with or without Peg-IFN did not result in the achievement of functional cure in any patient.

A genetic variant of CXCR4 predicts pegylated interferon-alpha treatment response in HBeAg-positive chronic hepatitis B patients.

Chemokine receptor 4 (CXCR4) plays a vital role in immunoregulation during hepatitis B virus (HBV) infection. This study aimed to screen single-nucleotide polymorphisms (SNPs) of CXCR4 for predicting pegylated interferon-alpha (PegIFNα) therapy response in chronic hepatitis B (CHB) patients. This retrospective cohort study enrolled a total of 945 CHB patients in two cohorts (Cohort 1, n = 238; Cohort 2, n = 707), and all the patients were hepatitis B e antigen (HBeAg)-positive and treated with PegIFNα for 48 weeks and followed up for 24 weeks. Twenty-two tag SNPs were selected in CXCR4 and its flanking region. A polygenic score (PGS) was utilized to evaluate the cumulative effect of multiple SNPs. The relationships between CXCR4 SNPs and PGS and PegIFNα treatment response were explored in the two cohorts. Among the 22 candidate SNPs of CXCR4, rs28367495 (T > C) was significantly linked to PegIFNα treatment response in both cohorts. In patients with more number of rs28367495 C allele, a higher rate of combined response (CR, defined as HBeAg seroconversion and HBV DNA level < 3.3 log10 IU/mL; P = 1.51 × 10-4), a lower mean hepatitis B surface antigen (HBsAg) level (P = 4.76 × 10-4), and a higher mean HBsAg decline (P = 3.88 × 10-4) at Week 72 were achieved. Moreover, a PGS integrating CXCR4_rs28367495 and five previously reported SNPs was strongly correlated with CR (P = 1.26 × 10-13), HBsAg level (P = 4.90 × 10-4), and HBsAg decline (P = 0.005) in all the patients of the two cohorts. CXCR4_rs28367495 is a promising indicator for predicting the responsiveness to PegIFNα treatment for HBeAg-positive CHB patients. The new PGS may further improve the prediction performance.

Serum hepatitis B virus spliced RNA proportion increases with liver disease progression in patients with chronic hepatitis B.

Serum hepatitis B virus (HBV) spliced RNAs (spRNAs) are ubiquitous in HBV-infected patients; however, their clinical significance remains unknown. Therefore, we aimed to explore the relationship between HBV spRNAs and liver disease progression in chronic hepatitis B (CHB) patients; in vitro cell line assessment was also performed. The serum HBV wild-type RNA (wtRNA) and spRNA levels were individually quantified in a cohort of 279 treatment-naïve, hepatitis B e antigen positive CHB patients with or without cirrhosis. The spRNA proportion was determined as (spRNA × 100%)/(spRNAs + wtRNA). 20 patients' serum samples underwent spRNA species profiling using next-generation sequencing. Serum spRNA species 1, 2, 3, 4, and 5 were the most common variants. The spRNA proportion varied from 0.00% to 19.02%, with higher levels in HBV genotype C patients than in those with genotype B (1.76% vs. 0.84%, p < 0.001). The spRNA proportion was positively associated with the alanine aminotransferase levels (r = 0.144, p = 0.053) and significantly higher in cirrhotic than in non-cirrhotic patients (1.69% vs. 1.04%, p = 0.001). Multivariate analysis revealed a 2.566-fold higher risk of cirrhosis in patients with elevated spRNA proportion (p = 0.024). In vitro experiments confirmed that spRNAs contributed to hepatic stellate cell activation, which is critical in liver fibrosis development. Therefore, increased HBV spRNA expression poses a risk for liver disease progression. Quantifying serum HBV spRNAs can aid in monitoring liver disease progression. Furthermore, the therapeutic targeting of spRNAs may improve the prognosis of patients with CHB.

Higher BST2 Expression Promotes the Anti-HBV Effect of IFN-α and BST2 Genetic Variant Predicts PegIFNα Treatment Response of HBeAg-Positive Chronic Hepatitis B Patients.

We previously reported that an interferon (IFN)-inducible protein, BST2, was regulated by the JAK-STAT pathway activated by CD40, and subsequently suppressing hepatitis B virus (HBV) repliaction and transcription. The current research attempted to assess the impact of BST2 on the IFN-treated anti-HBV effect, and explore BST2 variants for predicting pegylated IFN alpha (PegIFNα) therapy response of patients with hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB). Using an HBV-transfected cell model, the function of BST2 on HBV DNA replication and transcription driven by IFN was studied. The potentially functional BST2 variants were selected through a strategy of gene-wide screening. The associations of BST2 variants and polygenic score (PGS) model, which was used to quantify the combined influence of several genetic variants, with treatment response were examined in 2 separate PegIFNα-treated cohorts of 238 and 707 patients with CHB, respectively. We found that overexpression of BST2 improved the anti-HBV activity triggered by IFN-α. Among PegIFNα-treated patients with CHB, BST2_rs9576 was screened out to be significantly correlated with combined response (CR; i.e., HBeAg seroconversion along with HBV DNA level <3.3log10 IU/mL, P = 7.12 × 10-5 ). Additionally, there was a strong correlation between the PGS incorporating BST2_rs9576 and other 5 genetic variations (previously described predictors of therapy response to PegIFNα) and CR (P = 1.81 × 10-13 ), hepatitis B surface antigen (HBsAg) level (P = 0.004), as well as HBsAg decline (P = 0.017). In conclusion, higher BST2 expression responded better to IFN-α treatment. BST2_rs9576 is an effective indicator to forecast therapy response of PegIFNα-treated patients with CHB. The PGS possesses the potential to boost the ability of PegIFNα therapy response.

Machine-learning model comprising five clinical indices and liver stiffness measurement can accurately identify MASLD-related liver fibrosis.

BACKGROUND & AIMS: aMAP score, as a hepatocellular carcinoma risk score, is proven to be associated with the degree of chronic hepatitis B-related liver fibrosis. We aimed to evaluate the ability of aMAP score for metabolic dysfunction-associated steatotic liver disease (MASLD; formerly NAFLD)-related fibrosis diagnosis and establish a machine-learning (ML) model to improve the diagnostic performance. METHODS: A total of 946 biopsy-proved MASLD patients from China and the United States were included in the analysis. The aMAP score, demographic/clinical indices and liver stiffness measurement (LSM) were included in seven ML algorithms to build fibrosis diagnostic models in the training set (N = 703). The performance of ML models was evaluated in the external validation set (N = 125). RESULTS: The AUROCs of aMAP versus fibrosis-4 index (FIB-4) and aspartate aminotransferase-platelet ratio (APRI) in cirrhosis and advanced fibrosis were (0.850 vs. 0.857 [P = 0.734], 0.735 [P = 0.001]) and (0.759 vs. 0.795 [P = 0.027], 0.709 [P = 0.049]). When using dual cut-off values, aMAP had a smaller uncertainty area and higher accuracy (26.9%, 86.6%) than FIB-4 (37.3%, 85.0%) and APRI (59.0%, 77.3%) in cirrhosis diagnosis. The seven ML models performed satisfactorily in most cases. In the validation set, the ML model comprising LSM and 5 indices (including age, sex, platelets, albumin and total bilirubin used in aMAP calculator), built by logistic regression algorithm (called LSM-plus model), exhibited excellent performance. In cirrhosis and advanced fibrosis detection, the LSM-plus model had higher accuracy (96.8%, 91.2%) than LSM alone (86.4%, 67.2%) and Agile score (76.0%, 83.2%), respectively. Additionally, the LSM-plus model also displayed high specificity (cirrhosis: 98.3%; advanced fibrosis: 92.6%) with satisfactory AUROC (0.932, 0.875, respectively) and sensitivity (88.9%, 82.4%, respectively). CONCLUSIONS: The aMAP score is capable of diagnosing MASLD-related fibrosis. The LSM-plus model could accurately identify MASLD-related cirrhosis and advanced fibrosis.

Development and clinical validation of a novel algorithmic score (GAAD) for detecting HCC in prospective cohort studies.

BACKGROUND: Alpha-fetoprotein (AFP) and des-gamma carboxyprothrombin (DCP), also known as protein induced by vitamin K absence-II (PIVKA-II [DCP]) are biomarkers for HCC with limited diagnostic value when used in isolation. The novel GAAD algorithm is an in vitro diagnostic combining PIVKA-II (DCP) and AFP measurements, age, and gender (biological sex) to generate a semi-quantitative result. We conducted prospective studies to develop, implement, and clinically validate the GAAD algorithm for differentiating HCC (early and all-stage) and benign chronic liver disease (CLD), across disease stages and etiologies. METHODS: Patients aged ≥18 years with HCC or CLD were prospectively enrolled internationally into algorithm development [n = 1084; 309 HCC cases (40.7% early-stage) and 736 controls] and clinical validation studies [n = 877; 366 HCC cases (47.6% early-stage) and 303 controls]. Serum samples were analyzed on a cobas® e 601 analyzer. Performance was assessed using receiver operating characteristic curve analyses to calculate AUC. RESULTS: For algorithm development, AUC for differentiation between early-stage HCC and CLD was 90.7%, 84.4%, and 77.2% for GAAD, AFP, and PIVKA-II, respectively. The sensitivity of GAAD for the detection of early-stage HCC was 71.8% with 90.0% specificity. Similar results were shown in the clinical validation study; AUC for differentiation between early-stage HCC and CLD was 91.4% with 70.1% sensitivity and 93.7% specificity. GAAD also showed strong specificity, with a lower rate of false positives regardless of disease stage, etiology, or region. CONCLUSIONS: The GAAD algorithm significantly improves early-stage HCC detection for patients with CLD undergoing HCC surveillance. Further phase III and IV studies are warranted to assess the utility of incorporating the algorithm into clinical practice.

Guidelines for the Diagnosis and Treatment of Primary Liver Cancer (2022 Edition).

Background: Primary liver cancer, of which around 75-85% is hepatocellular carcinoma in China, is the fourth most common malignancy and the second leading cause of tumor-related death, thereby posing a significant threat to the life and health of the Chinese people. Summary: Since the publication of Guidelines for Diagnosis and Treatment of Primary Liver Cancer in China in June 2017, which were updated by the National Health Commission in December 2019, additional high-quality evidence has emerged from researchers worldwide regarding the diagnosis, staging, and treatment of liver cancer, that requires the guidelines to be updated again. The new edition (2022 Edition) was written by more than 100 experts in the field of liver cancer in China, which not only reflects the real-world situation in China but also may reshape the nationwide diagnosis and treatment of liver cancer. Key Messages: The new guideline aims to encourage the implementation of evidence-based practice and improve the national average 5-year survival rate for patients with liver cancer, as proposed in the "Health China 2030 Blueprint."

Noncanonical regulation of HOIL-1 on cancer stemness and sorafenib resistance identifies pixantrone as a novel therapeutic agent for HCC.

BACKGROUND AND AIMS: Cancer stem cells (CSCs) contribute to therapy resistance in HCC. Linear ubiquitin chain assembly complex (LUBAC) has been reported to accelerate the progression of cancers, yet its role in the sorafenib response of HCC is poorly defined. Herein, we investigated the impact of LUBAC on sorafenib resistance and the CSC properties of HCC, and explored the potential targeted drugs. APPROACH AND RESULTS: We found that HOIL-1, but not the other components of LUBAC, played a contributing role in LUBAC-mediated HCC sorafenib resistance, independent of its ubiquitin ligase activity. Both in vitro and in vivo assays revealed that the upregulated HOIL-1 expression enhanced the CSC properties of HCC. Mechanistically, HOIL-1 promoted sorafenib resistance and the CSC properties of HCC through Notch1 signaling. Mass spectrometry, co-immunoprecipitation, western blot, and immunofluorescence were used to determine that the A64/Q65 residues of HOIL-1 bound with the K78 residue of Numb, resulting in impaired Numb-mediated Notch1 lysosomal degradation. Notably, pixantrone was screened out by Autodock Vina, which was validated to disrupt HOIL-1/Numb interaction to inhibit Notch1 signaling and CSC properties by targeting the Q65 residue of HOIL-1. Moreover, pixantrone exerted synergistic effects with sorafenib for the treatment of HCC in different HCC mouse models. CONCLUSIONS: HOIL-1 is critical in promoting sorafenib resistance and CSC properties of HCC through Notch1 signaling. Pixantrone targeting HOIL-1 restrains the sorafenib resistance and provides a potential therapeutic intervention for HCC.

Tenofovir alafenamide and tenofovir disoproxil fumarate reduce incidence of hepatocellular carcinoma in patients with chronic hepatitis B.

Background & Aims: Antiviral therapy may attenuate the risk of hepatocellular carcinoma (HCC) in patients with chronic hepatitis B (CHB). We aimed to explore how tenofovir alafenamide (TAF) and tenofovir disoproxil fumarate (TDF) affect HCC risk in patients with CHB. Methods: The REACH-B, aMAP, and mPAGE-B models were utilized to assess HCC risk in patients with CHB from two global randomized-controlled trials evaluating the impact of TAF vs. TDF treatment. Standard incidence ratios (SIRs) were calculated using data from the REACH-B model as a ratio of observed HCC cases in the TAF- or TDF-treated patients vs. predicted HCC cases for untreated historical controls. Proportions of treated patients shifting aMAP and mPAGE-B risk categories between baseline and Week 240 were calculated. Results: Of the 1,632 patients (TAF, n = 1,093; TDF, n = 539) followed for up to 300 weeks, 22 HCC cases developed. Those receiving TAF had an SIR that was lower compared to the SIR of individuals receiving TDF: 0.32 (p <0.001) vs. 0.56 (p = 0.06). In the general study population, individuals without cirrhosis at baseline had an SIR that was lower compared to the SIR of individuals with cirrhosis at baseline: 0.37 (p <0.001) vs. 0.58 (p = 0.15). Of the patients at low risk of HCC at baseline, the majority (97%) remained low risk by mPAGE-B and aMAP scoring at Week 240. Among those at medium or high risk at baseline, substantial portions shifted to a lower risk category by Week 240 (mPAGE-B: 22% and 42%; aMAP: 39% and 63%, respectively). Conclusions: This evaluation provides evidence that treatment with TAF or TDF can reduce HCC risk in patients with CHB, particularly in patients without cirrhosis. Impact and implications: Despite the substantial impact of HCC on long-term outcomes of patients with CHB, the differential risk of HCC development among those receiving treatment with TAF vs. TDF has not been well elucidated. Using three validated risk prediction models, we found that TAF is at least as effective as TDF in reducing HCC risk in patients with CHB. While TDF is well-studied in the context of HCC risk reduction, our novel findings underscore the effectiveness of TAF as a treatment option for patients with CHB. Clinical trial numbers: NCT01940341; NCT02836249; NCT01940471; NCT02836236.

Guidelines for the Prevention and Treatment of Chronic Hepatitis B (version 2022).

To facilitate the achieving of the goal of "eliminating viral hepatitis as a major public health threat by 2030" set by the World Health Organization, the Chinese Society of Hepatology together with the Chinese Society of Infectious Diseases (both are branches of the Chinese Medical Association) organized a panel of experts and updated the guidelines for prevention and treatment of chronic hepatitis B in China (version 2022). With the support of available evidence, this revision of the guidelines focuses on active prevention, large scale testing, and expansion of therapeutic indication of chronic hepatitis B with the aim of reducing the hepatitis B related disease burden.

CXCL13 variant predicts pegylated-interferon α treatment response in HBeAg-positive chronic hepatitis B patients.

As a key immune cytokine, C-X-C motif chemokine ligand 13 (CXCL13) has been reported to play critical roles in immune control of hepatitis B virus (HBV) infection. We aimed to screen single-nucleotide polymorphisms (SNPs) of CXCL13 for predicting response to pegylated interferon-alpha (PegIFNα) therapy of chronic hepatitis B (CHB) patients. Two independent cohorts with a total of 945 (Cohort 1, n = 238; Cohort 2, n = 707) hepatitis B e antigen (HBeAg)-positive CHB patients treated with PegIFNα were enrolled in this retrospective cohort study. Eight candidate SNPs were selected through gene-wide SNP mining within or flanking CXCL13. A polygenic score (PGS) was utilized to assess the cumulative effects of multiple SNPs. The associations of candidate SNPs and PGS with combined response (CR, defined as the combination of HBeAg seroconversion and HBV DNA level <3.3log10 IU/mL) and hepatitis B surface antigen (HBsAg) level were evaluated. Among the eight candidate SNPs, rs76084459 which is located at upstream of CXCL13 was significantly associated with both CR (p = 0.002) and HBsAg level (p = 0.015). A PGS integrating CXCL13_rs76084459 and five other SNPs, which were previously identified as predictors of PegIFNα treatment response, was further strongly correlated with CR (p = 1.759 × 10-10 ) and HBsAg level (p = 0.004). This study demonstrated that CXCL13_rs76084459 can predict response to PegIFNα treatment of HBeAg-positive CHB patients. A PGS composed of six SNPs including CXCL13_rs76084459 predicts PegIFNα treatment response better.

Novel, high accuracy models for hepatocellular carcinoma prediction based on longitudinal data and cell-free DNA signatures.

BACKGROUND & AIMS: Current hepatocellular carcinoma (HCC) risk scores do not reflect changes in HCC risk resulting from liver disease progression/regression over time. We aimed to develop and validate two novel prediction models using multivariate longitudinal data, with or without cell-free DNA (cfDNA) signatures. METHODS: A total of 13,728 patients from two nationwide multicenter prospective observational cohorts, the majority of whom had chronic hepatitis B, were enrolled. aMAP score, as one of the most promising HCC prediction models, was evaluated for each patient. Low-pass whole-genome sequencing was used to derive multi-modal cfDNA fragmentomics features. A longitudinal discriminant analysis algorithm was used to model longitudinal profiles of patient biomarkers and estimate the risk of HCC development. RESULTS: We developed and externally validated two novel HCC prediction models with a greater accuracy, termed aMAP-2 and aMAP-2 Plus scores. The aMAP-2 score, calculated with longitudinal data on the aMAP score and alpha-fetoprotein values during an up to 8-year follow-up, performed superbly in the training and external validation cohorts (AUC 0.83-0.84). The aMAP-2 score showed further improvement and accurately divided aMAP-defined high-risk patients into two groups with 5-year cumulative HCC incidences of 23.4% and 4.1%, respectively (p = 0.0065). The aMAP-2 Plus score, which incorporates cfDNA signatures (nucleosome, fragment and motif scores), optimized the prediction of HCC development, especially for patients with cirrhosis (AUC 0.85-0.89). Importantly, the stepwise approach (aMAP -> aMAP-2 -> aMAP-2 Plus) stratified patients with cirrhosis into two groups, comprising 90% and 10% of the cohort, with an annual HCC incidence of 0.8% and 12.5%, respectively (p <0.0001). CONCLUSIONS: aMAP-2 and aMAP-2 Plus scores are highly accurate in predicting HCC. The stepwise application of aMAP scores provides an improved enrichment strategy, identifying patients at a high risk of HCC, which could effectively guide individualized HCC surveillance. IMPACT AND IMPLICATIONS: In this multicenter nationwide cohort study, we developed and externally validated two novel hepatocellular carcinoma (HCC) risk prediction models (called aMAP-2 and aMAP-2 Plus scores), using longitudinal discriminant analysis algorithm and longitudinal data (i.e., aMAP and alpha-fetoprotein) with or without the addition of cell-free DNA signatures, based on 13,728 patients from 61 centers across mainland China. Our findings demonstrated that the performance of aMAP-2 and aMAP-2 Plus scores was markedly better than the original aMAP score, and any other existing HCC risk scores across all subsets, especially for patients with cirrhosis. More importantly, the stepwise application of aMAP scores (aMAP -> aMAP-2 -> aMAP-2 Plus) provides an improved enrichment strategy, identifying patients at high risk of HCC, which could effectively guide individualized HCC surveillance.