SPINK2 Protein Expression Is an Independent Adverse Prognostic Marker in AML and Is Potentially Implicated in the Regulation of Ferroptosis and Immune Response.

There is an urgent need for the identification as well as clinicopathological and functional characterization of potent prognostic biomarkers and therapeutic targets in acute myeloid leukemia (AML). Using immunohistochemistry and next-generation sequencing, we investigated the protein expression as well as clinicopathological and prognostic associations of serine protease inhibitor Kazal type 2 (SPINK2) in AML and examined its potential biological functions. High SPINK2 protein expression was an independent adverse biomarker for survival and an indicator of elevated therapy resistance and relapse risk. SPINK2 expression was associated with AML with an NPM1 mutation and an intermediate risk by cytogenetics and European LeukemiaNet (ELN) 2022 criteria. Furthermore, SPINK2 expression could refine the ELN2022prognostic stratification. Functionally, an RNA sequencing analysis uncovered a potential link of SPINK2 with ferroptosis and immune response. SPINK2 regulated the expression of certain P53 targets and ferroptosis-related genes, including SLC7A11 and STEAP3, and affected cystine uptake, intracellular iron levels and sensitivity to erastin, a specific ferroptosis inducer. Furthermore, SPINK2 inhibition consistently increased the expression of ALCAM, an immune response enhancer and promoter of T-cell activity. Additionally, we identified a potential small-molecule inhibitor of SPINK2, which requires further characterization. In summary, high SPINK2 protein expression was a potent adverse prognostic marker in AML and might represent a druggable target.

Deep genomic characterization highlights complexities and prognostic markers of pediatric acute myeloid leukemia.

Pediatric acute myeloid leukemia (AML) is an uncommon but aggressive hematological malignancy. The poor outcome is attributed to inadequate prognostic classification and limited treatment options. A thorough understanding on the genetic basis of pediatric AML is important for the development of effective approaches to improve outcomes. Here, by comprehensively profiling fusion genes as well as mutations and copy number changes of 141 myeloid-related genes in 147 pediatric AML patients with subsequent variant functional characterization, we unveil complex mutational patterns of biological relevance and disease mechanisms including MYC deregulation. Also, our findings highlight TP53 alterations as strong adverse prognostic markers in pediatric AML and suggest the core spindle checkpoint kinase BUB1B as a selective dependency in this aggressive subgroup. Collectively, our present study provides detailed genomic characterization revealing not only complexities and mechanistic insights into pediatric AML but also significant risk stratification and therapeutic strategies to tackle the disease.

Mutational spectrum and prognosis in Chinese patients with prefibrotic primary myelofibrosis.

Prefibrotic primary myelofibrosis (Pre-PMF) has been classified as a separate entity of myeloproliferative neoplasms (MPNs). Pre-PMF is clinically heterogeneous but a specific prognostic model is lacking. Gene mutations have emerged as useful tools for stratification of myelofibrosis patients. However, there have been limited studies comprehensively investigating the mutational spectrum and its clinicopathological significance in pre-PMF subjects. In this study, we addressed these issues by profiling the mutation status of 141 genes in 172 Chinese MPN patients including 72 pre-PMF cases. Our findings corroborated the clinical/molecular distinctiveness of pre-PMF and suggested a refined risk classification strategy for this entity.

A novel NUP98-JADE2 fusion in a patient with acute myeloid leukemia resembling acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) is a specific subtype of acute myeloid leukemia (AML) characterized by block of differentiation at the promyelocytic stage and the presence of PML-RARA fusion. In rare instances, RARA is fused with other partners in variant APL. More infrequently, non-RARA genes are rearranged in AML patients resembling APL. However, the underlying disease pathogenesis in these atypical cases is largely unknown. Here, we report the identification and characterization of a NUP98- JADE2 fusion in a pediatric AML patient showing APL-like morphology and immunophenotype. Mechanistically, we showed that NUP98-JADE2 could impair all-trans retinoic acid (ATRA)-mediated transcriptional control and myeloid differentiation. Intriguingly, NUP98-JADE2 was found to alter the subcellular distribution of wild-type JADE2, whose down-regulation similarly led to attenuated ATRA-induced responses and myeloid activation, suggesting that NUP98-JADE2 may mediate JADE2 inhibition. To our knowledge, this is the first report of a NUP98-non-RAR rearrangement identified in an AML patient mimicking APL. Our findings suggest JADE2 as a novel myeloid player involved in retinoic acid-induced differentiation. Despite lacking a rearranged RARA, our findings implicate that altered retinoic acid signaling by JADE2 disruption may underlie the APL-like features in our case, corroborating the importance of this signaling in APL pathogenesis.

Association of HLA-B22 serotype with SARS-CoV-2 susceptibility in Hong Kong Chinese patients.

The coronavirus disease 2019 (COVID-19) is a highly infectious disease caused by SARS-CoV-2. Since its first report in December 2019, COVID-19 has evolved into a global pandemic causing massive healthcare and socioeconomic challenges. HLA system is critical in mediating anti-viral immunity and recent studies have suggested preferential involvement of HLA-B in COVID-19 susceptibility. Here, by investigating the HLA-B genotypes in 190 unrelated Chinese patients with confirmed COVID-19, we identified a significant positive association between the B22 serotype and SARS-CoV-2 infection (p = 0.002, Bonferroni-corrected p = 0.032). Notably, the B22 serotype has been consistently linked to susceptibility to other viral infections. These data not only shed new insights into SARS-CoV-2 pathogenesis and vaccine development but also guide better infection prevention/control.

Concurrent fatty liver increases risk of hepatocellular carcinoma among patients with chronic hepatitis B.

BACKGROUND AND AIMS: Concurrent fatty liver in hepatitis B virus (HBV)-infected patients without significant alcohol intake is a frequent and increasingly alarming problem because of the non-alcoholic fatty liver disease pandemic. The risk of HBV-related hepatocellular carcinoma (HCC) development was increased by concomitant obesity and diabetes. Direct evidence of the hepatocarcinogenic effect of fatty liver in chronic HBV remains elusive. We aimed to evaluate the risk of concurrent histologically proven fatty liver in HBV hepatocarcinogenesis. METHODS: We conducted a retrospective cohort study on a liver biopsy cohort of HBV-infected patients without significant alcohol intake to evaluate the prevalence of concurrent histologically proven fatty liver and its association with subsequent HCC development. We also examined nine polymorphisms on six non-alcoholic fatty liver disease-related candidate genes (ADIPOQ, APOC3, GCKR, LEPR, PNPLA3, and PPARG). RESULTS: Among 270 HBV-infected patients, concurrent fatty liver was found in 107 patients (39.6%) and was associated with metabolic risks, cirrhosis (P = 0.016) and PNPLA3 rs738409 CG/GG genotype (P = 0.002). At a median follow-up of 79.9 months, 11 patients (4.1%) developed HCC, and nine of them had concurrent fatty liver. By multivariable Cox analysis, concurrent fatty liver (HR 7.27, 95% confidence interval: 1.52-34.76; P = 0.013), age, cirrhosis, and APOC3 rs2854116 TC/CC genotype (HR 3.93, 95% confidence interval: 1.30-11.84; P = 0.013) were independent factors predicting HCC development. CONCLUSIONS: Concurrent fatty liver is common in HBV-infected patients and an independent risk factor potentiating HBV-associated HCC development by 7.3-fold. The risk of HBV-related HCC is increased by APOC3 gene polymorphism, and further characterization is required by its role.

Noninvasive assessments of liver fibrosis with transient elastography and Hui index predict survival in patients with chronic hepatitis B.

BACKGROUND AND AIMS: The prognostic role of noninvasive assessments of liver fibrosis has been evolving. Our aim was to investigate the prognostic value of liver stiffness measurement (LSM) with transient elastography and serum-based Hui index to predict hepatic events and deaths in chronic hepatitis B (CHB) patients. METHODS: The main prospective cohort included 1555 consecutive CHB patients referred for transient elastography examination; a subgroup of 980 patients underwent follow-up assessments at least 3 years later formed the serial cohort. Cox proportional hazard model was performed to determine the relationship of LSM, Hui index and other clinical variables with hepatic events and deaths. RESULTS: During a mean follow-up of 69 ± 9 months, 119 patients (7.6%) developed hepatic events or deaths. Hepatic event-free survival was significantly decreased with increasing stages of LSM and Hui index. The 5-year cumulative probability of hepatic event-free survival of patients of Stage 1-7 of LSM were 99.3%, 98.8%, 95.7%, 90.9%, 89.6%, 74.6%, and 50.0%, respectively; that of Stage 1 to 3 of Hui index were 98.2%, 93.1%, and 77.5%, respectively. Independent predictors of hepatic event-free survival were age, baseline LSM, and follow-up Hui index. Serum ALT and body mass index affected the accuracy of prediction by LSM. Patients remained early stages of LSM or Hui index at follow-up visit had better survival compared to those remained at late stages. CONCLUSION: Baseline and change in noninvasive parameters of liver fibrosis, LSM and Hui index, are accurate to predict hepatic event-free survival in CHB patients.

Adverse effects of vitamin D deficiency on outcomes of patients with chronic hepatitis B.

BACKGROUND & AIMS: Vitamin D is an immunomodulator that might be involved in the pathogenesis of viral hepatitis. We investigated the effects of vitamin D deficiency on long-term outcomes of patients with chronic hepatitis B (CHB). METHODS: We performed a prospective cohort study of 426 patients with CHB (65% male; mean age, 41 ± 13 years), who were enrolled from 1997 through 2000. Serum levels of 25-hydroxycholecalciferol (25(OH)D3) were measured on study enrollment (baseline). Patients were followed for 159 ± 46 months until last clinic visit or death; approximately 33% received antiviral therapy during the follow-up period. The primary outcome was a clinical event (hepatocellular carcinoma, complications of cirrhosis, or death). RESULTS: At baseline, the patients' mean serum level of hepatitis B virus DNA was 5.0 ± 2.1 log10 IU/mL; their mean level of 25(OH)D3 was 24.3 ± 9.4 ng/mL, and 348 patients (82%) had vitamin D deficiency (<32 ng/mL). Serum levels of 25(OH)D3 did not correlate with cirrhosis or viral load. Ninety-seven patients (22.8%) developed clinical events by a mean time of 118 ± 60 months after study enrollment. Patients who developed clinical events had lower baseline serum levels of 25(OH)D3 (23.2 ± 10.4 ng/mL) than patients who did not (28.2 ± 9.3 ng/mL, P < .001). Low baseline serum 25(OH)D3 was an independent factor associated with clinical events after adjustment for sex, age, and cirrhosis. The adjusted hazard ratio of vitamin D deficiency for clinical events was 1.90 (95% confidence interval [CI], 1.06-2.43; P = .04). The 15-year cumulative incidence rate of clinical events among patients with vitamin D deficiency was 25.5% (95% CI, 23.1%-27.9%), compared with 11.1% (95% CI, 7.4%-14.8%) in patients with normal serum levels of 25(OH)D3. CONCLUSIONS: Vitamin D deficiency is common among patients with CHB and is associated with adverse clinical outcomes.

PNPLA3 gene polymorphism and response to lifestyle modification in patients with nonalcoholic fatty liver disease.

BACKGROUND AND AIM: Lifestyle modification is the cornerstone for the management of nonalcoholic fatty liver disease (NAFLD), and patatin-like phospholipase 3 (PNPLA3) is one of the most important genetic determinants of NAFLD. We aimed to investigate the effect of PNPLA3 gene polymorphism on the response to lifestyle modification in NAFLD patients. METHODS: This was a post-hoc analysis of a randomized controlled trial on a lifestyle modification program in community NAFLD patients. The PNPLA3 rs738409 gene polymorphism was correlated with changes in metabolic profile and intrahepatic triglyceride content (IHTG) as measured by proton magnetic resonance spectroscopy. RESULTS: One hundred and fifty-four patients were equally randomized into the intervention and control groups. The presence of G allele was associated with greater reduction in IHTG (CC: 3.7 ± 5.2%, CG: 6.5 ± 3.6%), and GG: 11.3 ± 8.8% (Spearman's correlation, 0.34; P = 0.002), body weight (P = 0.030), waist-to-hip ratio (P = 0.024), total cholesterol (P = 0.031), and low-density lipoprotein cholesterol (P = 0.009) in the intervention group. In contrast, PNPLA3 polymorphism had no impact on IHTG changes in the control group. By multivariable analysis, PNPLA3 genotype and body mass index (BMI) change were independently associated with IHTG reduction in the intervention group. Only BMI change was associated with IHTG reduction in the control group. CONCLUSION: Although the PNPLA3 rs738409 GG genotype confers a higher risk of NAFLD, these patients are more sensitive to the beneficial effects of lifestyle modification and should be encouraged to do so.

On-treatment alpha-fetoprotein is a specific tumor marker for hepatocellular carcinoma in patients with chronic hepatitis B receiving entecavir.

UNLABELLED: Alpha-fetoprotein (AFP) is the most widely used biomarker for hepatocellular carcinoma (HCC) surveillance, which is criticized as neither sensitive nor specific in active hepatitis and liver cirrhosis. The aim of this study was to determine the performance of AFP as a tumor marker for HCC in entecavir-treated patients with chronic hepatitis B (CHB). This was a retrospective-prospective cohort study of 1,531 entecavir-treated patients under regular HCC surveillance with AFP and ultrasonography. Mean age was 52 ± 12 years; 1,099 (72%) patients were male and 332 (21.7%) had clinical evidence of cirrhosis. At a mean follow-up of 51 ± 13 months, 57 (2.9%) patients developed HCC (median size: 3.3 cm). AFP fluctuated with alanine aminotransferase (ALT) and peaked at the time of starting entecavir, then gradually decreased after. AFP started to increase 6 months before the diagnosis of HCC. The receiver operator characteristic curve (AUROC) of AFP was highest at the time of HCC diagnosis (0.85; 95% confidence interval [CI]: 0.73-0.98) and remained satisfactory at 3 (0.82; 95% CI: 0.73-0.91) and 6 months (0.79; 95% CI: 0.69-0.89) before the diagnosis. Using the conventional AFP cut-off (20 µg/L) at month 0, the sensitivity and specificity to diagnose HCC were 38.6% and 98.9%, respectively. Adopting the lower cut-off value (6 µg/L) of AFP level at month 0, sensitivity was increased to 80.7%, whereas specificity was decreased to 80.4%. CONCLUSION: On-treatment AFP is a specific tumor marker for HCC in CHB patients receiving entecavir therapy. Adopting a lower cut-off value of AFP level at 6 µg/L would significantly increase the sensitivity for HCC detection.

Liver fibrosis progression in chronic hepatitis B patients positive for hepatitis B e antigen: a prospective cohort study with paired transient elastography examination.

BACKGROUND AND AIM: Chronic hepatitis B patients in immune-reactive hepatitis B e antigen (HBeAg)-positive phase may have more rapid progression than those in immune-tolerant phase. We aimed to evaluate the risk of liver fibrosis progression in HBeAg-positive patients at different phases. METHODS: Two hundred forty-seven HBeAg-positive patients without advanced fibrosis at baseline underwent liver stiffness measurement (LSM) by transient elastography in 2006-2008 and again in 2010-2012. Liver fibrosis progression was defined as increase in LSM by 30% or more to levels suggestive of advanced fibrosis at the second assessment. RESULTS: At baseline, the mean age was 38 ± 11 years, 58% were males, alanine aminotransferase (ALT) was 65 ± 52 IU/L, hepatitis B virus DNA was 4.2 ± 1.2 log IU/mL, and LSM was 6.3 ± 2.1 kPa. At an interval of 42 ± 6 months, 13 patients (5.2%) developed liver fibrosis progression, and 106 patients (42.9%) required antiviral therapy. None of the clinical parameters (e.g., gender, age, ALT, hepatitis B virus DNA, hepatitis B surface antigen level, etc.) was associated with liver fibrosis progression. Among 74 and 137 patients in immune-tolerant and immune-reactive phase, 4.1% and 6.6% had liver fibrosis progression, and 12.2% and 67.2% received antiviral therapy respectively (P = 0.45 and P < 0.001). Immune-tolerant patients with low-normal (< 0.5× upper limit of normal) or high-normal ALT (0.5-1× upper limit of normal) also had similar risk of liver fibrosis progression (5.7% vs. 2.6%; P = 0.49). CONCLUSIONS: Liver fibrosis progression is uncommon in HBeAg-positive patients. Patients in immune-reactive phase treated with antiviral therapy did not have increased risk of liver fibrosis progression.

Liver fibrosis progression is uncommon in patients with inactive chronic hepatitis B: a prospective cohort study with paired transient elastography examination.

BACKGROUND AND AIMS: The European Association for the Study of the Liver (EASL) defines the inactive hepatitis B virus (HBV) carrier state based on HBV DNA and alanine aminotransferase (ALT) levels. This study aimed to evaluate the risk of disease progression in such patients. METHODS: Three hundred sixty-one patients negative for hepatitis B e antigen (HBeAg) with HBV DNA levels < 20,000 IU/mL and normal ALT and without advanced fibrosis at baseline underwent liver stiffness measurement (LSM) by transient elastography between 2006 and 2008 and again between 2010 and 2012. Liver fibrosis progression was defined as an increase in LSM by 30% or more at the second assessment to levels suggestive of advanced fibrosis. RESULTS: At baseline, the mean age was 48 ± 11 years and 51% were males; ALT level was 28 ± 11 IU/L, HBV DNA level was 2.7 ± 1.0 log10 IU/mL, and LSM was 5.4 ± 1.5 kPa. After an interval of 44 ± 7 months, liver fibrosis progression was observed in 10 (2.8%) patients, and 49 (13.6%) started antiviral therapy. Gender, age, and levels of ALT, HBV DNA, and HBsAg were shown not to be associated with liver fibrosis progression. Among 244 patients with baseline HBV DNA < 2000 IU/mL, 2.9% had liver fibrosis progression, 8.2% started antiviral therapy, and 4.1% had HBV DNA ≥ 20,000 IU/mL during follow-up. Corresponding figures in 117 patients with baseline HBV DNA levels of 2000-20,000 IU/mL were 2.6%, 24.8%, and 7.7%, respectively (P = 1.0, < 0.001 and = 0.21 respectively). CONCLUSIONS: Liver fibrosis progression within 3-4 years is rare in HBeAg-negative patients with HBV DNA <20,000 IU/mL and normal ALT, but a significant proportion of patients develop treatment indications during follow-up. The study supports the EASL's definition of inactive carriers and its recommendation of regular monitoring.

Serum hepatitis B surface antigen kinetics in severe reactivation of hepatitis B e antigen negative chronic hepatitis B patients receiving nucleoside/nucleotide analogues.

BACKGROUND: Kinetics of serum hepatitis B surface antigen (HBsAg) level in hepatitis B e antigen (HBeAg)-negative chronic hepatitis B patients presented with severe reactivation and received oral antiviral therapy is unknown. We aimed to investigate the kinetics of HBsAg level among these patients. METHODS: HBeAg-negative patients on antiviral therapy with follow-up for 2 years were studied. Those presented with severe reactivation (alanine aminotransferase [ALT] ≥5 times of normal) were compared to those with mild hepatitis. Serum HBsAg level was measured by Elecsys HBsAg II Quant assay (Roche) at baseline and 6-monthly. RESULTS: A total of 192 (74 severe reactivation) patients were studied. Eighty-one (42%), 74 (39%) and 37 (19%) patients were on lamivudine, entecavir and telbivudine, respectively. Forty-four (23%) patients had early HBsAg decline, that is, ≥0.5 log10 reduction, at month 6. Patients with severe reactivation had higher serum baseline ALT (1,415 ±897 versus 73 ±39 IU/l), HBV DNA (6.4 ±1.6 versus 5.2 ±1.2 log10 IU/ml) and HBsAg (3.3 ±1.0 versus 2.9 ±0.6 log10 IU/ml), as well as an earlier HBsAg decline (50% versus 6%; all P<0.001) than those without. The HBsAg change of patients with severe reactivation was higher at months 0-6 (-0.58 ±-1.26 versus -0.01 ±-0.26 log10 IU/ml; P<0.001) but then became comparable from months 6-24 (-0.19 ±-0.60 versus -0.13 ±-0.19 log10 IU/ml; P=0.85), compared to those presented with mild hepatitis. CONCLUSIONS: Patients who presented with severe reactivation of HBeAg-negative hepatitis were more likely to develop early HBsAg decline during antiviral therapy. It may indicate a transient strong immune clearance with rapid initial reduction in serum HBsAg, which cannot be sustained due to a faster clearance of serum HBsAg.

Efficacy of entecavir switch therapy in chronic hepatitis B patients with incomplete virological response to telbivudine.

BACKGROUND: The roadmap concept suggests the use of on-treatment HBV DNA to guide treatment strategy of chronic hepatitis B patients treated by telbivudine. Our aim was to validate the roadmap approach of entecavir switch therapy in patients with incomplete response to telbivudine. METHODS: Consecutive chronic hepatitis B patients on telbivudine monotherapy were studied. Incomplete virological response was defined as detectable HBV DNA after 6-12 months of treatment. Maintained virological response was defined as undetectable HBV DNA until the last follow-up. RESULTS: Among the 79 patients on telbivudine, 39 (49%) had undetectable HBV DNA after 6-12 months of telbivudine treatment and 40 (51%) had incomplete virological response. In total, 33 incomplete responders switched to entecavir at 11 months (6-23), and 26 (79%) achieved maintained virological response after 25 months (4-46). Low HBV DNA level before switch therapy was the independent factor associated with maintained virological response to entecavir (P=0.01). A total of 24 of 25 (96%) patients with HBV DNA<2,000 IU/ml, versus 2 of 8 (25%) patients with HBV DNA≥2,000 IU/ml, had maintained virological response after switching to entecavir. Although rtM204I and/or rtL180M was detected in 3 of 7 patients with incomplete virological response to entecavir, none of the patients with HBV DNA<2,000 IU/ml during telbivudine treatment harboured these amino acid substitutions. CONCLUSIONS: Roadmap approach using entecavir switch at month 6-12 among incomplete responders to telbivudine is recommended if the HBV DNA is <2,000 IU/ml at the time of switching.

Accuracy of risk scores for patients with chronic hepatitis B receiving entecavir treatment.

BACKGROUND & AIMS: Little is known about the validity of hepatocellular carcinoma (HCC) risk scores derived from treatment-naïve patients with chronic hepatitis B for patients treated with entecavir. METHODS: We performed a retrospective-prospective cohort study of 1531 patients with chronic hepatitis B (age, 51 ± 12 years; 1099 male; 332 with clinical cirrhosis) who were treated with entecavir 0.5 mg daily for at least 12 months at Prince of Wales Hospital in Hong Kong from December 2005 to August 2012. The patients were assessed once every 3 to 6 months for symptoms, drug history, and adherence; blood samples were collected for biochemical analyses. We validated 3 HCC risk scores (CU-HCC, GAG-HCC, and REACH-B scores) based on data collected when patients began treatment with entecavir and 2 years later. RESULTS: After 42 ± 13 months of follow-up, 47 patients (2.9%) developed HCC. The 5-year cumulative incidence of HCC was 4.3% (95% confidence interval [CI], 3.6%-5.0%). Older age, presence of cirrhosis, and virologic remission after 24 months or more of therapy were independently associated with HCC in the entire cohort; advanced age and hypoalbuminemia were associated with HCC in patients without cirrhosis. The area under the receiver operating characteristic curves (AUCs) for baseline CU-HCC, GAG-HCC, and REACH-B scores for HCC were 0.80 (95% CI, 0.75-0.86), 0.76 (95% CI, 0.70-0.82), and 0.71 (95% CI, 0.62-0.81), respectively; the time-dependent AUCs 1 to 4 years after patients started treatment were comparable to those at baseline. The cutoff value of the baseline CU-HCC score identified patients who would develop HCC with 93.6% sensitivity and 47.8% specificity, the baseline GAG-HCC score with 55.3% sensitivity and 78.9% specificity, and the baseline REACH-B score with 95.2% sensitivity and 16.5% specificity. Compared with patients with CU-HCC scores <5 at baseline, those with CU-HCC scores that either decreased from ≥5 to <5 or remained ≥5 had a higher risk of HCC (5-year cumulative incidences, 0% vs 3.9% and 7.3%; P = .002 and P < .001, respectively). CONCLUSIONS: The CU-HCC, GAG-HCC, and REACH-B HCC risk scores accurately predict which patients with chronic hepatitis B treated with entecavir will develop HCC.

Entecavir treatment reduces hepatic events and deaths in chronic hepatitis B patients with liver cirrhosis.

UNLABELLED: Entecavir is a potent antiviral agent with high genetic barrier to resistance, hence it is currently recommended as first-line antiviral therapy for chronic hepatitis B (CHB). The aim of this study was to investigate the efficacy of entecavir on clinical outcomes and deaths. It was a retrospective-prospective cohort study based on two cohorts of patients. The entecavir cohort included consecutive CHB patients who had received entecavir 0.5 mg/day for at least 12 months. The historical control cohort included untreated patients recruited since 1997 who underwent routine clinical care. The primary outcome was the 5-year cumulative probability of hepatic events, defined as any cirrhotic complications, hepatocellular carcinoma (HCC), and/or liver-related mortality. A total of 1,446 entecavir-treated patients (72% men; age, 51 ± 12 years; follow-up, 36 ± 13 months) and 424 treatment-naïve patients (65% men; age, 41 ± 13 years; follow-up, 114 ± 31 months) were studied. Overall, there was no difference in hepatic events between the entecavir and control cohorts. Among patients with liver cirrhosis (482 entecavir-treated, 69 treatment-naïve), entecavir-treated patients had reduced risks of all clinical outcomes when compared with treatment-naïve patients with cirrhosis after adjusted for model for end-stage liver disease score: hepatic events (hazard ratio [HR], 0.51; 95% confidence interval [CI], 0.34-0.78; P = 0.002), HCC (HR, 0.55; 95% CI, 0.31-0.99; P = 0.049), liver-related mortality (HR, 0.26; 95% CI, 0.13-0.55; P < 0.001), and all-cause mortality (HR, 0.34; 95% CI, 0.18-0.62; P < 0.001). Entecavir-treated patients with cirrhosis who failed to achieve undetectable hepatitis B virus DNA (105/482 [22%]) had comparable risk of hepatic events as the untreated patients. CONCLUSION: Entecavir therapy reduces the risks of hepatic events, HCC, liver-related and all-cause mortality of CHB patients with liver cirrhosis in 5 years, particularly among those who had maintained viral suppression.

Non-invasive diagnosis of non-alcoholic steatohepatitis by combined serum biomarkers.

BACKGROUND & AIMS: The diagnosis of non-alcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH) is limited by the need for liver biopsy. We aimed at testing the accuracy of cytokeratin-18 fragment (CK-18), adipocyte fatty acid binding protein (AFABP) and fibroblast growth factor 21 (FGF21) for the diagnosis of NAFLD and NASH. METHODS: 146 patients with biopsy-proven NAFLD and 74 age- and gender-matched healthy controls were included. Serum CK-18, AFABP and FGF21 levels were determined by enzyme-linked immunosorbent assay. RESULTS: Serum CK-18, AFABP, and FGF21 increased in a stepwise fashion in control subjects (median 103 U/L, 15.4 ng/ml, and 104 pg/ml), patients with non-NASH NAFLD (263 U/L, 18.9 ng/ml, and 249 pg/ml) and NASH (418 U/L, 19.4 ng/ml, and 354 pg/ml) (p<0.001, 0.060, and 0.016, respectively). The area under receiver-operating characteristics curve to diagnose NAFLD and NASH was 0.91 and 0.70 for CK-18, 0.66 and 0.59 for AFABP, and 0.84 and 0.62 for FGF21. At cut-offs of 203 and 670 U/L, CK-18 had 71% negative predictive value (NPV) and 77% positive predictive value (PPV) to exclude and diagnose NASH. A 2-step approach measuring CK-18 followed by FGF21 further improved the NPV to 74% and PPV to 82%. In a validation cohort of 51 patients with paired liver biopsies, the NPV and PPV of the 2-step approach were 67% and 78%, respectively. CONCLUSIONS: CK-18 is the most accurate biomarker for NAFLD and NASH. A two-step approach using CK-18 and FGF21 further improves the accuracy in diagnosing NASH.

Hepatitis B virus infection and fatty liver in the general population.

BACKGROUND & AIMS: In animal studies, expression of hepatitis B virus (HBV) proteins causes hepatic steatosis. We aimed to study the prevalence of fatty liver in people with and without HBV infection in the general population. METHODS: We performed a cross-sectional population study in Hong Kong Chinese. Intrahepatic triglyceride content (IHTG) was measured by proton-magnetic resonance spectroscopy. RESULTS: One thousand and thirteen subjects (91 HBV patients and 922 controls) were recruited. The median IHTG was 1.3% (0.2-33.3) in HBV patients and 2.1% (0-44.2) in controls (p <0.001). Excluding subjects with significant alcohol consumption, the prevalence of nonalcoholic fatty liver disease was 13.5% (95% confidence interval [CI] 6.4%, 20.6%) in HBV patients and 28.3% (95% CI 25.3%, 31.2%) in controls (p=0.003). The fatty liver prevalence differed in HBV patients and controls aged 40-59 years but was similar in those aged 60 years or above. After adjusting for demographic and metabolic factors, HBV infection remained an independent factor associated with lower risk of fatty liver (adjusted odds ratio 0.42; 95% CI 0.20, 0.88; p=0.022). HBV patients also had a lower prevalence of metabolic syndrome (11.0% vs. 20.2%; p=0.034), but the difference was mainly attributed to lower triglyceride levels. Among HBV patients, viral genotypes, HBV DNA level and hepatitis B e antigen status were not associated with fatty liver. CONCLUSIONS: HBV infection is associated with a lower prevalence of fatty liver, hypertriglyceridemia and metabolic syndrome. Viral replication may affect lipid metabolism and this warrants further studies.

Antiviral drug resistance testing in patients with chronic hepatitis B.

BACKGROUND: Antiviral drugs against hepatitis B virus are limited by the emergence of drug resistance. AIMS: We aimed to study the impact of drug resistance testing on treatment decisions. METHODS: In part 1 of this study, consecutive patients with chronic hepatitis B who had antiviral drug resistance testing were studied. Part 2 was a two-step questionnaire survey including ten characteristic case scenarios. Hepatologists were asked about their treatment decisions before and after the knowledge of drug resistance results. RESULTS: Fifty-one patients underwent drug resistance testing, most of whom were on lamivudine, adefovir dipivoxil or entecavir monotherapy. Thirty-four (67%) patients had drug-resistant mutants detected, 4 (8%) had low viral load, and 13 (25%) harboured wild-type virus. Twenty-nine of 34 (85%) patients harbouring drug-resistant mutants and 9 of 17 (53%) patients with no mutants detected changed their drug regimens (P = 0.038). In part 2, 18 hepatologists completed all two questionnaires. Overall, treatment decision was modified in 52% of cases upon receiving the drug resistance testing results. The detection of rtA181V/I resulted in decision changes in most hepatologists, with the preferred treatment switching from tenofovir to entecavir. When no mutants were detected in partial responders to entecavir monotherapy, most hepatologists chose to increase the dose of entecavir. CONCLUSIONS: Drug-resistant mutations are detected in around two-thirds of chronic hepatitis B patients undergoing drug resistance testing. Drug resistance testing alters management in over half of the cases, and should be considered in all patients with virological breakthrough and suboptimal virological suppression.

Prediction of off-treatment response to lamivudine by serum hepatitis B surface antigen quantification in hepatitis B e antigen-negative patients.

BACKGROUND: The timing of antiviral therapy cessation in hepatitis B e antigen (HBeAg)-negative patients is controversial. Here, we aimed to investigate the role of HBV DNA and hepatitis B surface antigen (HBsAg) monitoring to predict off-treatment sustained response. METHODS: A total of 53 HBeAg-negative chronic hepatitis B patients who received lamivudine for 34 ±23 (range 12-76) months and had lamivudine stopped for 47 ±35 months were studied. Primary outcome was sustained response, defined as HBV DNA≤200 IU/ml, at 12 months post-treatment (SR-12). RESULTS: A total of 9 (17%) patients achieved SR-12. HBV DNA at baseline, month 6 and end of treatment had no association with SR-12. HBsAg levels tended to decrease more significantly during treatment among SR-12 responders. At the end of treatment, both HBsAg ≤2 log IU/ml and reduction by >1 log from baseline had sensitivity, specificity, positive and negative predictive values for SR-12 of 78%, 96%, 78% and 96%, respectively. All 5 patients with HBsAg≤2 log IU/ml and reduction >1 log at the end of treatment achieved SR-12 and all 40 patients with HBsAg>2 log IU/ml and reduction ≤1 log did not have SR-12. The cumulative probability of sustained response and HBsAg clearance at 5 years among patients with HBsAg≤2 log IU/ml were 88% and 72%, respectively, that among patients with HBsAg reduction >1 log were 74% and 61%, respectively. CONCLUSIONS: Monitoring of HBsAg level can guide the timing of stopping lamivudine in HBeAg-negative chronic hepatitis B.