Safety and Immunogenicity of SARS-CoV-2 Vaccines in Patients With Chronic Liver Diseases (CHESS-NMCID 2101): A Multicenter Study.

BACKGROUND & AIMS: We aimed to assess the safety and immunogenicity of inactivated whole-virion severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines in patients with chronic liver diseases (CLD) in this study. METHODS: This was a prospective, multi-center, open-label study. Participants aged over 18 years with confirmed CLD and healthy volunteers were enrolled. All participants received 2 doses of inactivated whole-virion SARS-CoV-2 vaccines. Adverse reactions were recorded within 14 days after any dose of SARS-CoV-2 vaccine, laboratory testing results were collected after the second dose, and serum samples of enrolled subjects were collected and tested for SARS-CoV-2 neutralizing antibodies at least 14 days after the second dose. RESULTS: A total of 581 participants (437 patients with CLD and 144 healthy volunteers) were enrolled from 15 sites in China. Most adverse reactions were mild and transient, and injection site pain (n = 36; 8.2%) was the most frequently reported adverse event. Three participants had grade 3 aminopherase elevation (defined as alanine aminopherase >5 upper limits of normal) after the second dose of inactivated whole-virion SARS-CoV-2 vaccination, and only 1 of them was judged as severe adverse event potentially related to SARS-CoV-2 vaccination. The positive rates of SARS-CoV-2 neutralizing antibodies were 76.8% in the noncirrhotic CLD group, 78.9% in the compensated cirrhotic group, 76.7% in the decompensated cirrhotic group (P = .894 among CLD subgroups), and 90.3% in healthy controls (P = .008 vs CLD group). CONCLUSION: Inactivated whole-virion SARS-CoV-2 vaccines are safe in patients with CLD. Patients with CLD had lower immunologic response to SARS-CoV-2 vaccines than healthy population. The immunogenicity is similarly low in noncirrhotic CLD, compensated cirrhosis, and decompensated cirrhosis.

Safety and immunogenicity of COVID-19 vaccination in patients with hepatocellular carcinoma (CHESS-NMCID 2101): A multicenter prospective study.

Data on safety and immunogenicity of coronavirus disease 2019 (COVID-19) vaccinations in hepatocellular carcinoma (HCC) patients are limited. In this multicenter prospective study, HCC patients received two doses of inactivated whole-virion COVID-19 vaccines. The safety and neutralizing antibody were monitored. Totally, 74 patients were enrolled from 10 centers in China, and 37 (50.0%), 25 (33.8%), and 12 (16.2%) received the CoronaVac, BBIBP-CorV, and WIBP-CorV, respectively. The vaccines were well tolerated, where pain at the injection site (6.8% [5/74]) and anorexia (2.7% [2/74]) were the most frequent local and systemic adverse events. The median level of neutralizing antibody was 13.5 (interquartile range [IQR]: 6.9-23.2) AU/ml at 45 (IQR: 19-72) days after the second dose of vaccinations, and 60.8% (45/74) of patients had positive neutralizing antibody. Additionally, lower γ-glutamyl transpeptidase level was related to positive neutralizing antibody (odds ratio = 1.022 [1.003-1.049], p = 0.049). In conclusion, this study found that inactivated COVID-19 vaccinations are safe and the immunogenicity is acceptable or hyporesponsive in patients with HCC. Given that the potential benefits may outweigh the risks and the continuing emergences of novel severe acute respiratory syndrome coronavirus 2 variants, we suggest HCC patients to be vaccinated against COVID-19. Future validation studies are warranted.

Safety and immunogenicity of COVID-19 vaccination in patients with non-alcoholic fatty liver disease (CHESS2101): A multicenter study.

BACKGROUND & AIMS: The development of COVID-19 vaccines has progressed with encouraging safety and efficacy data. Concerns have been raised about SARS-CoV-2 vaccine responses in the large population of patients with non-alcoholic fatty liver disease (NAFLD). The study aimed to explore the safety and immunogenicity of COVID-19 vaccination in NAFLD. METHODS: This multicenter study included patients with NAFLD without a history of SARS-CoV-2 infection. All patients were vaccinated with 2 doses of inactivated vaccine against SARS-CoV-2. The primary safety outcome was the incidence of adverse reactions within 7 days after each injection and overall incidence of adverse reactions within 28 days, and the primary immunogenicity outcome was neutralizing antibody response at least 14 days after the whole-course vaccination. RESULTS: A total of 381 patients with pre-existing NAFLD were included from 11 designated centers in China. The median age was 39.0 years (IQR 33.0-48.0 years) and 179 (47.0%) were male. The median BMI was 26.1 kg/m2 (IQR 23.8-28.1 kg/m2). The number of adverse reactions within 7 days after each injection and adverse reactions within 28 days totaled 95 (24.9%) and 112 (29.4%), respectively. The most common adverse reactions were injection site pain in 70 (18.4%), followed by muscle pain in 21 (5.5%), and headache in 20 (5.2%). All adverse reactions were mild and self-limiting, and no grade 3 adverse reactions were recorded. Notably, neutralizing antibodies against SARS-CoV-2 were detected in 364 (95.5%) patients with NAFLD. The median neutralizing antibody titer was 32 (IQR 8-64), and the neutralizing antibody titers were maintained. CONCLUSIONS: The inactivated COVID-19 vaccine appears to be safe with good immunogenicity in patients with NAFLD. LAY SUMMARY: The development of vaccines against coronavirus disease 2019 (COVID-19) has progressed rapidly, with encouraging safety and efficacy data. This study now shows that the inactivated COVID-19 vaccine appears to be safe with good immunogenicity in the large population of patients with non-alcoholic fatty liver disease.

Decreased antigenicity profiles of immune-escaped and drug-resistant hepatitis B surface antigen (HBsAg) double mutants.

BACKGROUND: Selective pressure from either the immune response or the use of nucleoside analogs in antiviral therapy could be driving the emergence of HBV mutants. Because of the overlap of the open reading frame (ORF) S for the HBsAg and ORF P for viral polymerase, rtM204I and rtM204V mutations in the polymerase would produce sI195M and sW196S in the HBsAg. The combined effects of immune-escaped mutations (sT118M, sG145K, sG145R) and drug-resistant mutations (rtM204I, rtM204V) on the antigenicity profiles of HBsAg has not been widely explored. METHODS: To determine the combined effects of immune-escaped and drug-resistant mutants on the antigenicity profiles of HBsAg, recombinant plasmids encoding HBsAg double mutants were constructed using site-directed mutagenesis. The supernatant from each plasmid transfection was analyzed for HBsAg in the western-blotting and five of the most commonly used commercial ELISA kits in China. RESULTS: Western-blotting assay showed the successful expression of each HBsAg mutant. All five ELISA kits manifested similar avidity, which were demonstrated by the slope of the curves, for the sT118M mutant, and sT118M-rtM204I (sT118M-sI195M) and sT118M-rtM204V (sT118M-sW196S) double mutants, suggesting that drug-resistant YMDD mutants caused negligible losses in the antigenicity of immune-escaped sT118M HBsAg. In contrast, the presence of the rtM204I (sI195M) mutation, but not rtM204V (sW196S) in combination with the sG145K mutation significantly reduced the avidity of sG145K HBsAg. The rtM204I (sI195M) mutation also decreased the antigenicity profiles for sG145R HBsAg. CONCLUSIONS: Drug-resistant mutations rtM204I (sI195M) and rtM204V (sW196S) caused significant reduction in antigenicity for the immune-escaped HBsAg mutants sG145K and sG145R, which may hamper HBV diagnosis and disease control from HBV blood-transfusion transmissions in China. The development of ELISA kits with a greater sensitivity for drug-resistant and immune-escaped HBsAg warrants further consideration.

[Relationship between metastasis or recurrence of hepatocellular carcinoma and hepatitis B virus DNA or double mutation at 1762/1764 in the basic core promoter].

OBJECTIVE: To study the relationship between metastasis or recurrence of hepatocellular carcinoma (HCC) and hepatitis B virus (HBV) DNA load or the presence of double mutation at 1762/1764 in the basic core promoter (BCP). METHODS: One-hundred-and-fifty-seven patients with HCC were included in the study. Events of tumor metastasis or recurrence were recorded during 120 weeks of clinical follow-up after treatment by surgery or transarterial chemoembolization (TACE). The 1-year follow-up included monthly alpha fetoprotein (AFP) measurement and abdominal ultrasonography (US), as well as helical computed tomographic (CT) scan performed every 3 months. Follow-up beyond 1-year (surveillance) included AFP measurement and abdominal US every 2 months and helical CT scan every 6 months. Suspected metastasis or recurrence was investigated by hepatic angiography and confirmed according to the combined imaging findings. Serum HBV DNA level was measured by real-time PCR. HBV genotypes were determined by PCR-restriction fragment length polymorphism analysis. RESULTS: Of the 157 HCC cases 110 experienced tumor metastasis or recurrence; the cumulative probability of post-treatment HCC metastasis or recurrence was 4 (2.55%) at week 12, 14 (8.92%) at week 24, 28 (17.83%) at week 48, 64 (40.76%) at week 72, 92 (58.60%) at week 96, and 110 (70.06%) at week 120. Multivariate analysis indicated that both the BCP 1762/1764 double mutations and HBV DNA levels were risk factors for HCC recurrence or metastasis. In particular, the incidence of HCC recurrence or metastasis increased with baseline serum HBV DNA levels in a dose-response manner, ranging from 8/19 (42.1%) for less than 3 log10 copies/ml HBV DNA to 35/61 (57.3%) for 3-5 log10 copies/ml and 67/77 (87.0%) for more than 5 log10 copies/ml. After adjusting for potential confounders, serum HBV DNA level remained independently associated with HCC metastasis or recurrence. HCC recurrence or metastasis occurred in 22/43 (51.2%) of patients without BCP 1762/1764 mutations and 88/114 (77.2%) of patients with BCP 1762/1764 mutations. The adjusted odds ratio for patients infected with BCP 1762/1764 double mutation HBV, compared with those infected with non-BCP 1762/1764 mutation HBV, was 5.264 (95% CI: 1.436-12.574, P less than 0.05). CONCLUSION: Infection with HBV carrying the BCP 1762/1764 double mutation and presence of high HBV DNA load are independent risk factors for developing HCC metastasis or recurrence after surgery or TACE.

LncRNA MORT Overexpression Inhibits Cancer Cell Migration and Invasion in Hepatocellular Carcinoma by Downregulating NOTCH1.

Background: Long noncoding RNA (lncRNA) MORT is silenced in many malignancies, but its role in cancer remains hardly known. Methods: The expression of MORT and NOTCH1 was determined by real-time quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Correlation between MORT and NOTCH1 was analyzed by Pearson's correlation analysis. To further investigate the interaction between MORT and NOTCH1, overexpression experiments were performed. Results: In our study, MORT expression was downregulated in hepatocellular carcinoma (HCC), while NOTCH1 expression was upregulated in HCC patients. Hepatitis B virus and hepatitis C virus infection and tumor size did not significantly affect MORT expression, but MORT expression was lower in metastatic HCC patients compared with nonmetastatic HCC patients. MORT and NOTCH1 were inversely correlated across HCC tissues. MORT overexpression decreased NOTCH1 expression, while NOTCH1 overexpression did not significantly affect MORT. MORT overexpression inhibited the migration and invasion of HCC cells, while NOTCH1 overexpression promoted the migration and invasion of HCC cells. In addition, NOTCH1 overexpression attenuated the effects of MORT overexpression on cell migration and invasion. Conclusion: Therefore, MORT overexpression may inhibit HCC by downregulating NOTCH1.

Pan-cancer analysis identifies ITIH1 as a novel prognostic indicator for hepatocellular carcinoma.

Although a previous pan-cancer study has reported the expression patterns of ITIHs in various tumors, their analyses have been restricted to limited cancer types. We thus comprehensively analyzed the expression profiles and clinical significances of ITIHs in a broader spectrum of cancers from TCGA. Our results showed that ITIHs were primarily down-regulated in tested cancers. The ITIH members were associated with either survival advantage or disadvantage, depending on the cancer type tested and the genes queried. Importantly, we for the first time demonstrated that ITIH1 had substantially decreased expression in liver hepatocellular carcinoma (LIHC) compared with corresponding normal tissue, and its down-regulation adversely impacted patient outcome. Moreover, ITIH1 expression was consistently declining during the progression of LIHC. Further analysis revealed that ITIH1 may be involved in cellular metabolic processes. Our findings established ITIH1 as a potential diagnostic and prognostic biomarker for LIHC, which awaits future experimental validation.

miR­498 inhibits the growth and metastasis of liver cancer by targeting ZEB2.

MicroRNAs (miRNAs) play critical roles in the growth, metastasis and therapeutic resistance of liver cancer. Accumulating evidence suggests that miR­498 is aberrantly expressed in several human malignancies. However, the role and underlying mechanism of miR­498 in liver cancer remain unclear. In the present study, we investigated the potential roles and clinical value of miR­498 in liver cancer. We found that the miR­498 expression level was significantly lower in liver cancer patient tissues than that in healthy control tissues. The expression of miR­498 was also decreased in liver cancer cell lines compared to that noted in a normal human normal liver cell line. miR­498 overexpression markedly inhibited liver cancer cell proliferation, migration and invasion. miR­498 overexpression induced cell cycle arrest and apoptosis while it suppressed epithelial­mesenchymal transition (EMT) in liver cancer cells. Bioinformatic analysis and luciferase reporter assay further identified zinc finger E­box binding homeobox 2 (ZEB2) as a novel target of miR­498. Furthermore, ZEB2 knockdown recapitulated the inhibitory effects of miR­498 overexpression in liver cancer cells. ZEB2 overexpression rescued the inhibition of liver cancer cell proliferation, migration, and invasion by miR­498, indicating that ZEB2 acts as a downstream effector of miR­498 in liver cancer cells. Thus, we demonstrated that miR­498 suppresses the growth and metastasis of liver cancer cells, partly at least, by directly targeting ZEB2, suggesting that miR­498 may serve as a potential biomarker for the diagnosis and therapy of liver cancer.

LINC00978 promotes the progression of hepatocellular carcinoma by regulating EZH2-mediated silencing of p21 and E-cadherin expression.

Long non-coding RNAs (lncRNAs) have been suggested as important regulators of cancer development and progression in hepatocellular carcinoma (HCC). Nevertheless, the clinical value and biological roles of LINC00978 in HCC remain unclear. In this study, we detected the expression of LINC00978 in tumor tissues and serum of HCC patients, examined the roles of LINC00978 in HCC progression and elucidated the underlying molecular mechanisms. We found that LINC00978 expression was upregulated in tumor tissues and serum of HCC patients. Higher serum levels of LINC00978 could distinguish HCC patients from hepatitis and liver cirrhosis patients and healthy controls. LINC00978 knockdown inhibited HCC cell proliferation, migration and invasion while promoted cell cycle arrest and apoptosis. Overexpression of LINC00978 led to the opposite effects. LINC00978 knockdown also inhibited HCC growth and metastasis in mouse tumor models. Mechanistically, LINC00978 bound to EZH2 and mediated its accumulation at the promoter region of p21 and E-cadherin genes, leading to the trimethylation of H27K3 and the inhibition of p21 and E-cadherin expression. Moreover, the simultaneous depletion of p21 and E-cadherin expression reversed the inhibitory effects of LINC00978 knockdown on HCC cell proliferation, migration, and invasion. Taken together, these findings suggest that LINC00978 promotes HCC progression by inhibiting p21 and E-cadherin expression via EZH2-mediated epigenetic silencing. LINC00978 may represent a novel biomarker for HCC diagnosis, prognosis, and therapy.

Ductopenia and cirrhosis in a 32-year-old woman with progressive familial intrahepatic cholestasis type 3: A case report and review of the literature.

Progressive familial intrahepatic cholestasis type 3 is caused by a mutation in the ATP-binding cassette, subfamily B, member 4 (ABCB4) gene encoding multidrug resistance protein 3. A 32-year-old woman with a history of acute hepatitis at age 9 years was found to have jaundice during pregnancy in 2008, and was diagnosed as having intrahepatic cholestasis of pregnancy. In 2009, she underwent cholecystectomy for gallstones and chronic cholecystitis. However, itching and jaundice did not resolve postoperatively. She was admitted to our hospital with fatigue, jaundice, and a recently elevated γ-glutamyl transpeptidase level. Liver biopsy led to the diagnosis of biliary cirrhosis with ductopenia. Genetic testing revealed a pathogenic heterozygous mutation, ex13 c.1531G > A (p.A511T), in the ABCB4 gene. Her father did not carry the mutation, but her mother's brother carried the heterozygous mutation. We made a definitive diagnosis of familial intrahepatic cholestasis type 3. Her symptoms and liver function improved after 3 mo of treatment with ursodeoxycholic acid.

Diagnostic value of FIB-4, aspartate aminotransferase-to-platelet ratio index and liver stiffness measurement in hepatitis B virus-infected patients with persistently normal alanine aminotransferase.

AIM: To assess the diagnostic value of FIB-4, aspartate aminotransferase-to-platelet ratio index (APRI), and liver stiffness measurement (LSM) in patients with hepatitis B virus infection who have persistently normal alanine transaminase (PNALT). METHODS: We enrolled 245 patients with chronic hepatitis B: 95 in PNALT group, 86 in intermittently elevated alanine transaminase (PIALT1) group [alanine transaminase (ALT) within 1-2 × upper limit of normal value (ULN)], and 64 in PIALT2 group (ALT > 2 × ULN). All the patients received a percutaneous liver biopsy guided by ultrasonography. LSM, biochemical tests, and complete blood cell counts were performed. RESULTS: The pathological examination revealed moderate inflammatory necrosis ratios of 16.81% (16/95), 32.56% (28/86), and 45.31% (28/64), and moderate liver fibrosis of 24.2% (23/95), 33.72% (29/86), and 43.75% (28/64) in the PNALT, PIALT1, and PIALT2 groups, respectively. The degrees of inflammation and liver fibrosis were significantly higher in the PIALT groups than in the PNALT group (P < 0.05). No significant difference was found in the areas under the curve (AUCs) between APRI and FIB-4 in the PNALT group; however, significant differences were found between APRI and LSM, and between FIB-4 and LSM in the PNALT group (P < 0.05 for both). In the PIALT1 and PIALT2 groups, no significant difference (P > 0.05) was found in AUCs for all comparisons (P > 0.05 for all). In the overall patients, a significant difference in the AUCs was found only between LSM and APRI (P < 0.05). CONCLUSION: APRI and FIB-4 are not the ideal noninvasive hepatic fibrosis markers for PNALT patients. LSM is superior to APRI and FIB-4 in PNALT patients because of the influence of liver inflammation and necrosis.

Serum MiRNA panel as potential biomarkers for chronic hepatitis B with persistently normal alanine aminotransferase.

BACKGROUND: Circulating miRNAs, a family of miRNAs existing in plasma and serum, have a great potential to serve as novel biomarkers in body fluids for non-invasive diagnosis and prognosis of many diseases. METHODS: A multistage, case-control study was designed to establish a panel of serum miRNAs that could be surrogate markers for chronic hepatitis B with persistently normal alanine aminotransferase (ALT). A total of 295 CHB patients presenting persistently normal ALT levels with significant histological features (SPNALT group), 243 CHB patients presenting persistently normal ALT levels with no significant histological features (NSPNALT group), and 178 healthy controls (healthy group) were enrolled in the study. An initial screening of miRNAs was performed by Illumina sequencing using serum samples pooled from SPNALT patients and controls. Quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) assay was performed to evaluate the expression of selected miRNAs. A logistic regression model was constructed using a training cohort (n=380) and validated using a cohort (n=258). The area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. RESULTS: We identified 9 miRNAs (hsa-miR-885-5p, hsa-miR-122-5p, hsa-miR-10a-5p, hsa-miR-511-5p, hsa-miR-574-5p, hsa-miR-98-5p, hsa-miR-26a-5p, hsa-miR-192-5p, hsa-miR-30b-5p) and established 3 miRNA panels that provided high diagnostic accuracy for SPNALT. The AUC of miRNA panels for SPNALT vs. healthy was 0.882 (95% CI=0.839 to 0.925), for SPNALT vs. NSPNALT was 0.894 (95% CI=0.857 to 0.930), and for SPNALT vs. control was 0.860 (95% CI=0.821 to 0.899). CONCLUSIONS: We constructed serum miRNA panels with considerable clinical value in diagnosing PNALT.

Hepatitis B surface antigen levels of cessation of nucleos(t)ide analogs associated with virological relapse in hepatitis B surface antigen-negative chronic hepatitis B patients.

AIM: To investigate the virological relapse rate in hepatitis B e antigen (HBeAg)-negative patients after antiviral therapy discontinuation and analyze the factors associated with virological relapse. METHODS: Among patients diagnosed with chronic hepatitis B infection between May 2005 and July 2010, 204 were eligible for analysis. The Kaplan-Meier method and log-rank test were used to calculate the cumulative rate of relapse and compare cumulative relapse rates between groups. The Cox proportional hazards regression model was used to evaluate the predictive factor of virological relapse. RESULTS: The 2 and 1 year cumulative risks of virological relapse after antiviral therapy discontinuation were 79.41% (162/204) and 43.82% (71/162), respectively. Multivariate analysis revealed that only post treatment hepatitis B surface antigen (HBsAg) level was associated with virological relapse (P = 0.011). The cumulative risk of virological relapse was higher in the patients with HBsAg levels ≥ 1500 IU/L than in those with HBsAg levels < 1500 IU/L (P = 0.0013). The area under the curve was 0.603 (P = 0.033). The cutoff HBsAg value for predicting virological relapse was 1443 IU/L. CONCLUSION: We found that the virological relapse rate remained high after antiviral therapy discontinuation in the HBeAg-negative patients and that the post treatment HBsAg levels predicted virological relapse.

Natural YMDD-motif mutants affect clinical course of lamivudine in chronic hepatitis B.

AIM: To investigate the prevalence of nature tyrosine-methionine-aspartic acid-aspartic acid motif mutations in chronic hepatitis B (CHB) patients and to evaluate the efficacy of lamivudine. METHODS: A total of 1268 CHB patients were recruited from the outpatient and inpatient departments of six centers. Tyrosine-methionine-aspartic acid-aspartic acid (YMDD) mutations were analyzed using the hepatitis B virus (HBV) drug resistance line probe assay. Forty voluntary patients were selected from those with positive or negative natural YMDD mutations to undergo treatment with lamivudine. RESULTS: YMDD mutations were detected in 288 (22.71%) of the 1268 CHB patients. Multivariate analysis revealed that the patients' HBV DNA level (P=0.0282) and hepatitis B e antigen status (P=0.0133) were also associated with natural YMDD mutations. The rates of normalization of alanine aminotransferase levels and HBV DNA nondetection at 6, 24, 36, and 48 wk were compared between the patients with natural YMDD mutations and those without, and the differences were not significant. However, there was a significant difference in the cumulative emergence rates of virological breakthrough at 48 wk in the patients with natural YMDD mutations and those without (32.5% vs 12.5%, P=0.032). CONCLUSION: Naturally occurring YMDD mutations are detectable in a large proportion of CHB patients; breakthrough hepatitis tended to occur in patients with natural YMDD mutations.

A pilot study of serum microRNAs panel as potential biomarkers for diagnosis of nonalcoholic fatty liver disease.

BACKGROUND: The invasive nature of liver biopsy makes the histopathological diagnosis of non-alcoholic fatty liver disease (NAFLD) difficult and its diagnostic performance unsatisfactory. The present study aimed to identify a serum microRNA (miRNA) expression profile that could serve as a novel diagnostic biomarker for NAFLD. METHODS: Serum miRNA expression was investigated using three cohorts comprising 465 participants (healthy controls and NAFLD patients) recruited between August 2010 and June 2013. miRNA expression was initially screened by Illumina sequencing using serum samples pooled from 20 patients and 20 controls. Quantitative reverse transcriptase polymerase chain reaction assay was then used to evaluate the expression of selected miRNAs. A logistic regression model was constructed using a training cohort (n = 242) and validated using another cohort (n = 183). The area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. RESULTS: We identified an miRNA panel (hsa-miR-122-5p, hsa-miR-1290, hsa-miR-27b-3p, and hsa-miR-192-5p) with a high diagnostic accuracy for NAFLD. The satisfactory diagnostic performance of the miRNA panel remained regardless of the NAFLD activity score (NAS) status. There was significant difference between the AUC values of the miRNA panel and those of ALT (AUC = 0.786, 95% CI = 0.717-0.855; P = 0.142) and FIB-4 (AUC = 0.795, 95% CI = 0.730-0.860; sensitivity = 69.9%, specificity = 83.7%. CONCLUSION: We identified a serum microRNA panel with considerable clinical value in NAFLD diagnosis. The results indicate that the miRNA panel is a more sensitive and specific biomarker for NAFLD than ALT and FIB-4.

A serum microRNA panel as potential biomarkers for hepatocellular carcinoma related with hepatitis B virus.

BACKGROUND: The identification of new high-sensitivity and high-specificity markers for HCC are essential. We aimed to identify serum microRNAs (miRNAs) as biomarkers to be used in diagnosing hepatitis B virus (HBV) -related hepatocellular carcinoma (HCC). METHODS: We investigated serum miRNA expression in (261 HCC patients, 233 cirrhosis patients, and 173 healthy controls), recruited between August 2010 and June 2013. An initial screening of miRNA expression by Illumina sequencing was performed using serum samples pooled from HCC patients and controls. Quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) was used to evaluate the expression of selected miRNAs. A logistic regression model was constructed using a training cohort (n = 357) and then validated using an independent cohort (n = 241). The area under the receiver operating characteristic curve (AUC) was used to evaluate the accuracy of the use of the biomarkers for disease diagnosis. RESULTS: We identified 8 miRNAs (hsa-miR-206, hsa-miR-141-3p, hsa-miR-433-3p, hsa-miR-1228-5p, hsa-miR-199a-5p, hsa-miR-122-5p, hsa-miR-192-5p, and hsa-miR-26a-5p) and constructed an miRNA set that provided high diagnostic accuracy for HCC (AUC = 0.887 and 0.879 for training and validation sets, respectively). The miRNAs could also be used to differentiate HCC patients from healthy (AUC = 0.893) and cirrhosis (AUC = 0.892) patients. CONCLUSIONS: We identified a serum of miRNA panel that has considerable clinical value in HCC diagnosis.

Serum microRNAs as potential biomarkers of primary biliary cirrhosis.

BACKGROUND: Circulating microRNAs (miRNAs), which are extremely stable and protected from RNAse-mediated degradation in body fluids, have emerged as candidate biomarkers for many diseases. The present study aimed to identify a serum microRNA (miRNA) expression profile that could serve as a novel diagnostic biomarker for primary biliary cirrhosis (PBC). METHODS: Serum miRNA expression was investigated using four cohorts comprising 380 participants (healthy controls and patients with PBC) recruited between August 2010 and June 2013. miRNA expression was initially analyzed by Illumina sequencing using serum samples pooled from 3 patients and 3 controls. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was then used to evaluate the expression of selected miRNAs in a screening set (n = 40). A logistic regression model was then constructed using a training cohort (n = 192) and validated using another cohort (n = 142). The area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. RESULTS: We identified a miRNA panel (hsa-miR-122-5p, hsa-miR-141-3p, and hsa-miR-26b-5p) with a high diagnostic accuracy for PBC (AUC = 0.905, 95% confidence interval (CI) = 0.857 to 0.953; sensitivity = 80.5%, specificity = 88.3%). There was a significant difference between AUC values of the miRNA panel and those of alkaline phosphatase (ALP) (AUC = 0.537, difference between areas = 0.314, 95% CI = 0.195 to 0.434, P<0.001), and those of antinuclear antibody (ANA) (AUC = 0.739, difference between areas = 0.112, 95% CI = 0.012 to 0.213, P = 0.0282). CONCLUSION: We identified a serum microRNA panel with considerable clinical value in PBC diagnosis. The results indicate that the miRNA panel is a more sensitive and specific biomarker for PBC than ALP and ANA.

Removing N-terminal sequences in pre-S1 domain enhanced antibody and B-cell responses by an HBV large surface antigen DNA vaccine.

Although the use of recombinant hepatitis B virus surface (HBsAg) protein vaccine has successfully reduced global hepatitis B infection, there are still a number of vaccine recipients who do not develop detectable antibody responses. Various novel vaccination approaches, including DNA vaccines, have been used to further improve the coverage of vaccine protection. Our previous studies demonstrated that HBsAg-based DNA vaccines could induce both humoral and CMI responses in experimental animal models. However, one form of the the HBsAg antigen, the large S antigen (HBs-L), expressed by DNA vaccine, was not sufficiently immunogenic in eliciting antibody responses. In the current study, we produced a modified large S antigen DNA vaccine, HBs-L(T), which has a truncated N-terminal sequence in the pre-S1 region. Compared to the original HBs-L DNA vaccine, the HBs-L(T) DNA vaccine improved secretion in cultured mammalian cells and generated significantly enhanced HBsAg-specific antibody and B cell responses. Furthermore, this improved HBsL DNA vaccine, along with other HBsAg-expressing DNA vaccines, was able to maintain predominantly Th1 type antibody responses while recombinant HBsAg protein vaccines produced in either yeast or CHO cells elicited mostly Th2 type antibody responses. Our data indicate that HBsAg DNA vaccines with improved immunogenicity offer a useful alternative choice to recombinant protein-based HBV vaccines, particularly for therapeutic purposes against chronic hepatitis infection where immune tolerance led to poor antibody responses to S antigens.

YMDD motif mutations in chronic hepatitis B antiviral treatment naïve patients: a multi-center study.

OBJECTIVE: This study aimed to determine the natural prevalence of variants of tyrosine-methionine-aspartic acid-aspartic acid (YMDD) motif in patients with chronic hepatitis B (CHB), and to explore its relation with demographic and clinical features, hepatitis B virus (HBV) genotypes, and HBV DNA levels. METHODS: A total of 1,042 antiviral treatment naïve CHB patients (including with lamivudine [LAM]) in the past year were recruited from outpatient and inpatient departments of six centers from December 2008 to June 2010. YMDD variants were analyzed using the HBV drug resistance line probe assay (Inno-Lipa HBV-DR). HBV genotypes were detected with polymerase chain reaction (PCR) microcosmic nucleic acid cross-ELISA, and HBV deoxyribonucleic acid (DNA) was quantitated with real-time PCR. All serum samples underwent tests for HBV, HCV, and HDV with ELISA. RESULTS: YMDD variants were detected in 23.3% (243/1042) of CHB patients. YMDD mutation was accompanied by L180M mutation in 154 (76.9%) patients. Both wild-type HBV and YMDD variant HBV were present in 231 of 243 patients. Interestingly, 12 patients had only YIDD and/or YVDD variants without wild YMDD motif. In addition, 27.2% (98/359) of HbeAg-positive patients had YMDD mutations, which was higher than that in HbeAg-negative patients (21.2%, 145/683). The incidence of YMDD varied among patients with different HBV genotypes, but the difference was not significant. Moreover, the incidence of YMDD in patients with high HBV DNA level was significantly higher than that in those with low HBV DNA level. CONCLUSION: Mutation of YMDD motif was detectable at a high rate in CHB patients in this study. The incidence of YMDD may be correlated with HBeAg and HBV DNA level.

Profiles of B and T cell immune responses elicited by different forms of the hepatitis B virus surface antigen.

Gene-based hepatitis B virus (HBV) vaccines have been proposed as a novel approach to improve the immunogenicity toward non-responders and to allow for protection against potential viral escape mutants. Furthermore, there is significant interest in using DNA or viral vector vaccines to serve as therapeutic agents to treat chronic HBV infections that are resistant to existing drug therapies. However, the key protective antigen of HBV, the surface protein (HBsAg), can be expressed in three different sizes due to its multiple translational initiation sites: small, middle, and large forms of HBsAg. It is not clear whether the immunogenicity of these HBsAg is same, especially their ability to elicit HBsAg-specific B cell and T cell immune responses in addition to the traditional serum HBsAg-specific antibody responses. In the current study, the immunogenicity of three forms of HBsAg DNA vaccines was analyzed individually in a mouse model. Our results indicated that different forms of the HBsAg have unique immunogenicity profiles and this information is useful for the selection of optimal gene-based HBV vaccines for further improved prophylactic and therapeutic applications.