Drug Screening for Hepatitis A Virus (HAV): Nicotinamide Inhibits c-Jun Expression and HAV Replication.

Hepatitis A virus (HAV) infection often causes acute hepatitis, which results in a case fatality rate of 0.2% and fulminant hepatitis in 0.5% of cases. However, no specific potent anti-HAV drug is available on the market to date. In the present study, we focused on inhibition of HAV internal ribosomal entry site (IRES)-mediated translation and investigated novel therapeutic drugs through drug repurposing by screening for inhibitors of HAV IRES-mediated translation and cell viability using a reporter assay and cell viability assay, respectively. The initial screening of 1,158 drugs resulted in 77 candidate drugs. Among them, nicotinamide significantly inhibited HAV HA11-1299 genotype IIIA replication in Huh7 cells. This promising drug also inhibited HAV HM175 genotype IB subgenomic replicon and HAV HA11-1299 genotype IIIA replication in a dose-dependent manner. In the present study, we found that nicotinamide inhibited the activation of activator protein 1 (AP-1) and that knockdown of c-Jun, which is one of the components of AP-1, inhibited HAV HM175 genotype IB IRES-mediated translation and HAV HA11-1299 genotype IIIA and HAV HM175 genotype IB replication. Taken together, the results showed that nicotinamide inhibited c-Jun, resulting in the suppression of HAV IRES-mediated translation and HAV replication, and therefore, it could be useful for the treatment of HAV infection. IMPORTANCE Drug screening methods targeting HAV IRES-mediated translation with reporter assays are attractive and useful for drug repurposing. Nicotinamide (vitamin B3, niacin) has been shown to effectively inhibit HAV replication. Transcription complex activator protein 1 (AP-1) plays an important role in the transcriptional regulation of cellular immunity or viral replication. The results of this study provide evidence that AP-1 is involved in HAV replication and plays a role in the HAV life cycle. In addition, nicotinamide was shown to suppress HAV replication partly by inhibiting AP-1 activity and HAV IRES-mediated translation. Nicotinamide may be useful for the control of acute HAV infection by inhibiting cellular AP-1 activity during HAV infection processes.

Clinical significance of CDT1 mRNA expression in chronic hepatitis C or liver cirrhosis.

We have previously reported that chromatin licensing and DNA replication factor 1 (CDT1) is associated with the postoperative recurrence of hepatocellular carcinoma (HCC). Based on this fact, we verified whether CDT1 mRNA expression is also associated with HCC development from chronic hepatitis C (CHC) and liver cirrhosis (LC). There were 142 cases with CHC or LC who underwent liver biopsy. Detection of CDT1 mRNA in liver was performed by RT-qPCR using frozen liver biopsy tissues. We examined the association between the CDT1 mRNA expression and clinical conditions and long-term outcome. We then examined the association between serum cytokine/chemokine levels and CDT1 mRNA expression in 58 cases. The cumulative incidence rates of HCC development in cases with CDT1 mRNA in the low expression group showed significantly lower than those in the high expression group (p = 0.0391). A significant correlation was found between CDT1 mRNA expression and the extent of proliferation of atypical hepatocytes in hematoxylin and eosin-stained sections (p<0.0001). CDT1 mRNA expression has been associated with cytokines involved in tumorigenesis in experimental and human cancers. We found that cases with high CDT1 mRNA expression were at risk for developing HCC, even if they were CHC or LC.

Amantadine and Rimantadine Inhibit Hepatitis A Virus Replication through the Induction of Autophagy.

Hepatitis A virus (HAV) infection is a major cause of acute viral hepatitis worldwide. Furthermore, HAV causes acute liver failure or acute-on-chronic liver failure. However, no potent anti-HAV drugs are currently available in the clinical situations. There have been some reports that amantadine, a broad-spectrum antiviral, suppresses HAV replication in vitro. Therefore, we examined the effects of amantadine and rimantadine, derivates of adamantane, on HAV replication, and investigated the mechanisms of these drugs. In the present study, we evaluated the effects of amantadine and rimantadine on HAV HM175 genotype IB subgenomic replicon replication and HAV HA11-1299 genotype IIIA replication in cell culture infection systems. Amantadine and rimantadine significantly inhibited HAV replication at the post-entry stage in Huh7 cells. HAV infection inhibited autophagy by suppressing the autophagy marker light chain 3 and reducing number of lysosomes. Proteomic analysis on HAV-infected Huh7 cells treated by amantadine and rimantadine revealed the changes of the expression levels in 42 of 373 immune response-related proteins. Amantadine and rimantadine inhibited HAV replication, partially through the enhancement of autophagy. Taken together, our results suggest a novel mechanism by which HAV replicates along with the inhibition of autophagy and that amantadine and rimantadine inhibit HAV replication by enhancing autophagy. IMPORTANCE Amantadine, a nonspecific antiviral medication, also effectively inhibits HAV replication. Autophagy is an important cellular mechanism in several virus-host cell interactions. The results of this study provide evidence indicating that autophagy is involved in HAV replication and plays a role in the HAV life cycle. In addition, amantadine and its derivative rimantadine suppress HAV replication partly by enhancing autophagy at the post-entry phase of HAV infection in human hepatocytes. Amantadine may be useful for the control of acute HAV infection by inhibiting cellular autophagy pathways during HAV infection processes.

Molecular Mechanisms, Diagnosis and Treatments in Digestive Malignancy.

In this Special Issue, "Molecular Mechanisms, Diagnosis and Treatments in Digestive Malignancy", of the International Journal of Molecular Sciences, a total of 10 impactful articles have been published [...].

Masitinib Inhibits Hepatitis A Virus Replication.

The hepatitis A virus (HAV) infection causes acute hepatitis. HAV also induces acute liver failure or acute-on-chronic liver failure; however, no potent anti-HAV drugs are currently available in clinical situations. For anti-HAV drug screening, more convenient and useful models that mimic HAV replication are needed. In the present study, we established HuhT7-HAV/Luc cells, which are HuhT7 cells stably expressing the HAV HM175-18f genotype IB subgenomic replicon RNA harboring the firefly luciferase gene. This system was made by using a PiggyBac-based gene transfer system that introduces nonviral transposon DNA into mammalian cells. Then, we investigated whether 1134 US Food and Drug Administration (FDA)-approved drugs exhibited in vitro anti-HAV activity. We further demonstrated that treatment with tyrosine kinase inhibitor masitinib significantly reduced both HAV HM175-18f genotype IB replication and HAV HA11-1299 genotype IIIA replication. Masitinib also significantly inhibited HAV HM175 internal ribosomal entry-site (IRES) activity. In conclusion, HuhT7-HAV/Luc cells are adequate for anti-HAV drug screening, and masitinib may be useful for the treatment of severe HAV infection.

Gut-Microbiota Dysbiosis in Stroke-Prone Spontaneously Hypertensive Rats with Diet-Induced Steatohepatitis.

Metabolic-dysfunction-associated fatty-liver disease (MAFLD) is the principal worldwide cause of liver disease. Individuals with nonalcoholic steatohepatitis (NASH) have a higher prevalence of small-intestinal bacterial overgrowth (SIBO). We examined gut-microbiota isolated from 12-week-old stroke-prone spontaneously hypertensive-5 rats (SHRSP5) fed on a normal diet (ND) or a high-fat- and high-cholesterol-containing diet (HFCD) and clarified the differences between their gut-microbiota. We observed that the Firmicute/Bacteroidetes (F/B) ratio in both the small intestines and the feces of the SHRSP5 rats fed HFCD increased compared to that of the SHRSP5 rats fed ND. Notably, the quantities of the 16S rRNA genes in small intestines of the SHRSP5 rats fed HFCD were significantly lower than those of the SHRSP5 rats fed ND. As in SIBO syndrome, the SHRSP5 rats fed HFCD presented with diarrhea and body-weight loss with abnormal types of bacteria in the small intestine, although the number of bacteria in the small intestine did not increase. The microbiota of the feces in the SHRSP5 rats fed HFCD was different from those in the SHRP5 rats fed ND. In conclusion, there is an association between MAFLD and gut-microbiota alteration. Gut-microbiota alteration may be a therapeutic target for MAFLD.

The significance of CDT1 expression in non-cancerous and cancerous liver in cases with hepatocellular carcinoma.

We previously reported that chromatin licensing and DNA replication factor 1 (CDT1) expression was associated with the extent of proliferation of atypical hepatocytes and the time to postoperative recurrence in cases of hepatocellular carcinoma (HCC). This study aimed to clarify the clinical significance or pathogenesis of CDT1 expression in both non-cancerous and cancerous liver in HCC cases, including previously published data. We investigated the association between the expression of CDT1 in non-cancerous or cancerous liver tissues and histologic findings or biochemical examination results in 62 cases. We also examined the dual localization between CDT1 and FbxW7, P57kip2, P53 and c-Myc by confocal laser scanning microscopy. CDT1 mRNA expression was significantly higher in cancerous liver than in non-cancerous liver (p<0.0001). Elevated CDT1 mRNA expression indicates a significantly degree of inflammatory cell infiltration within lobules, along with elevated serum transaminase levels, and hepatic spare decline. CDT1 mRNA was highly expressed in a group of poorly differentiated cancer cells. CDT1 co-localized with P57kip2, Fbwx7, P53 and c-Myc in the nucleus or cytoplasm of hepatocytes and cancer cells. We found that CDT1 mRNA expression could represent the degree of hepatic spare ability and the high carcinogenic state.

Involvement of proliferation of atypical hepatocytes and CDT 1 in the liver cancer of rats administered the diethylnitrosamine.

We have reported that extent of proliferation of atypical hepatocytes (POAH) in non-cancerous liver in hepatocellular carcinoma and chromatin licensing and DNA replication factor 1 (CDT1) are associated with postoperative recurrence. Here, we investigated whether extent of POAH and expression of CDT1 in liver are also associated with chemically induced liver cancer in rats. Male Fisher strain rats were orally administered diethylnitrosamine (DEN) in their drinking water and sacrificed at 6, 8, 12, or 14 weeks after start of DEN administration. We serially monitored changes in extent of POAH, CDT1 expression by immunohistochemistry (IHC), and CDT1 mRNA expression in liver by real-time quantitative PCR. The extent of POAH in liver progressed in a time-dependent manner after start of DEN administration. CDT1 expression was higher at 8 weeks than at 6 weeks by IHC, suggesting that CDT1 expression may be a marker of POAH severity. CDT1 mRNA expression in liver was significantly higher at 12 weeks than at 6 weeks (p<0.0001). We found that extent of POAH and the expression of CDT1 are also important factors in the development of chemical carcinogen-induced hepatocarcinogenesis. Furthermore, the association with POAH and CDT1 expression in carcinogenic process is important regardless of the cause of hepatocarcinogenesis.

Evaluation of Potential Anti-Hepatitis A Virus 3C Protease Inhibitors Using Molecular Docking.

Hepatitis A virus (HAV) infection is a major cause of acute hepatitis worldwide and occasionally causes acute liver failure and can lead to death in the absence of liver transplantation. Although HAV vaccination is available, the prevalence of HAV vaccination is not adequate in some countries. Additionally, the improvements in public health reduced our immunity to HAV infection. These situations motivated us to develop potentially new anti-HAV therapeutic options. We carried out the in silico screening of anti-HAV compounds targeting the 3C protease enzyme using the Schrodinger Modeling software from the antiviral library of 25,000 compounds to evaluate anti-HAV 3C protease inhibitors. Additionally, in vitro studies were introduced to examine the inhibitory effects of HAV subgenomic replicon replication and HAV HA11-1299 genotype IIIA replication in hepatoma cell lines using luciferase assays and real-time RT-PCR. In silico studies enabled us to identify five lead candidates with optimal binding interactions in the active site of the target HAV 3C protease using the Schrodinger Glide program. In vitro studies substantiated our hypothesis from in silico findings. One of our lead compounds, Z10325150, showed 47% inhibitory effects on HAV genotype IB subgenomic replicon replication and 36% inhibitory effects on HAV genotype IIIA HA11-1299 replication in human hepatoma cell lines, with no cytotoxic effects at concentrations of 100 µg/mL. The effects of the combination therapy of Z10325150 and RNA-dependent RNA polymerase inhibitor, favipiravir on HAV genotype IB HM175 subgenomic replicon replication and HAV genotype IIIA HA11-1299 replication showed 64% and 48% inhibitory effects of HAV subgenomic replicon and HAV replication, respectively. We identified the HAV 3C protease inhibitor Z10325150 through in silico screening and confirmed the HAV replication inhibitory activity in human hepatocytes. Z10325150 may offer the potential for a useful HAV inhibitor in severe hepatitis A.

Favipiravir Inhibits Hepatitis A Virus Infection in Human Hepatocytes.

Hepatitis A virus (HAV) is a causative agent of acute hepatitis and can occasionally induce acute liver failure. However, specific potent anti-HAV drug is not available on the market currently. Thus, we investigated several novel therapeutic drugs through a drug repositioning approach, targeting ribonucleic acid (RNA)-dependent RNA polymerase and RNA-dependent deoxyribonucleic acid polymerase. In the present study, we examined the anti-HAV activity of 18 drugs by measuring the HAV subgenomic replicon and HAV HA11-1299 genotype IIIA replication in human hepatoma cell lines, using a reporter assay and real-time reverse transcription polymerase chain reaction, respectively. Mutagenesis of the HAV 5' untranslated region was also examined by next-generation sequencing. These specific parameters were explored because lethal mutagenesis has emerged as a novel potential therapeutic approach to treat RNA virus infections. Favipiravir inhibited HAV replication in both Huh7 and PLC/PRF/5 cells, although ribavirin inhibited HAV replication in only Huh7 cells. Next-generation sequencing demonstrated that favipiravir could introduce nucleotide mutations into the HAV genome more than ribavirin. In conclusion, favipiravir could introduce nucleotide mutations into the HAV genome and work as an antiviral against HAV infection. Provided that further in vivo experiments confirm its efficacy, favipiravir would be useful for the treatment of severe HAV infection.

Molecular Changes in Relation to Alcohol Consumption and Hepatocellular Carcinoma.

Alcohol is the one of the major causes of liver diseases and promotes liver cirrhosis and hepatocellular carcinoma (HCC). In hepatocytes, alcohol is converted to acetaldehyde, which causes hepatic steatosis, cellular apoptosis, endoplasmic reticulum stress, peroxidation, production of cytokines and reduces immune surveillance. Endotoxin and lipopolysaccharide produced from intestinal bacteria also enhance the production of cytokines. The development of hepatic fibrosis and the occurrence of HCC are induced by these alcohol metabolites. Several host genetic factors have recently been identified in this process. Here, we reviewed the molecular mechanism associated with HCC in alcoholic liver disease.

Left Gastric Vein Width Is an Important Risk Factor for Exacerbation of Esophageal Varices Post Balloon-Occluded Retrograde Transvenous Obliteration for Gastric Varices in Cirrhotic Patients.

Background and Objectives: Balloon-occluded retrograde transvenous obliteration (BRTO) could be currently one of the best therapies for patients with gastric varices. This study examined the exacerbation rates for esophageal varices following BRTO for gastric varices in patients with hepatic cirrhosis. Materials and Methods: We enrolled 91 cirrhotic patients who underwent BRTO for gastric varices. In total, 50 patients were examined for exacerbation rates of esophageal varices following BRTO. Esophageal varices and their associated exacerbation were evaluated by upper gastrointestinal endoscopy. Patients were allocated into two groups according to the main inflow tract for gastric varices: (1) 37 patients in the left gastric vein (LGV) group with an LGV width of more than 3.55 mm, and (2) 13 patients in the non-LGV group who had short gastric vein or posterior gastric vein. Moreover, treatment outcomes were retrospectively analyzed. Results: LGV width (p < 0.01) was the major risk factor for the deterioration of esophageal varices post BRTO. In addition, LGV was the most common inflow tract, and the LGV group contained 74% (37/50) of patients. The exacerbation rates of esophageal varices at 1, 2, 3, and 4 years post BRTO were 40%, 62%, 65%, and 68%, respectively. The comparison of the exacerbation rates for esophageal varices following BRTO according to inflow tract showed that the exacerbation rates were significantly higher in the LGV group than those of the non-LGV group (p = 0.03). In more than half of the subjects, LGV was the main inflow tract for gastric varices, and this group experienced more frequent exacerbations of esophageal varices following BRTO compared to patients with different inflow tract sources. Conclusion: Careful attention should be paid to the LGV width when BRTO is performed for gastric varices.

Endoscopic biliary drainage using a 4-Fr catheter for biliary obstruction: a pilot study.

INTRODUCTION: Stent or endoscopic nasobiliary drainage (ENBD) catheter placement for a tight, complicated biliary stricture is still technically challenging. A thin, 4-Fr ENBD catheter (4-Fr catheter) has been developed to overcome this difficulty. The study aimed to evaluate the feasibility of the 4-Fr catheter for endoscopic biliary drainage (EBD). MATERIAL AND METHODS: We performed a retrospective review of 51 patients who underwent EBD with the 4-Fr catheter because placement of a conventional drainage catheter (CDC) had failed. RESULTS: The success rate of 4-Fr catheter placement was 96.1% (49/51). The median patency period of the catheter was 114 days (95% CI, 53-200). Among the 49 patients with successful placement of the catheter, adverse events occurred in five (10.2%) patients: post-endoscopic retrograde cholangiopancreatography pancreatitis (PEP), two patients; catheter dislocation, one patient; and kinking of the 4-Fr catheter, two patients. Both cases of PEP improved with conservative treatment, but all cases of catheter dislocation and kinking required reintervention with a 4-Fr catheter. Forty-three (87.8%) patients achieved clinical remission after EBD with a 4-Fr catheter. CONCLUSIONS: The newly developed 4-Fr catheter is safe and feasible for EBD in patients in whom CDC placement is difficult due to a tight, complicated biliary stricture.

Hepatocellular adenoma, approximately half and predominantly inflammatory subtype, in 38 Japanese patients with several differences in age, gender, and clinical background factors from Western populations.

AIM: Hepatocellular adenoma (HCA) has a lower prevalence in Japan than in Western countries and HCA subtypes have been reported for only a few Japanese patients. We analyzed HCA subtype data 38 patients from 23 hospitals in Japan in order to examine character and difference between Western countries. METHODS: To confirm HCA and to analyze subtypes, we performed immunohistochemical examinations. RESULTS: Thirty-eight cases were found to have HCA without cirrhosis. The male/female ratio was 18/20. Ages ranged from 15 to 79 (average, 43.2) years. Male and elder patients are not rare, furthermore, most of elder patients are male. Glycogen storage disease, past history of medicament use, hepatitis B virus surface antigen-positivity, antihepatitis C virus -positivity, diabetes mellitus, obesity, lipid metabolism disorder and alcoholism were present in of 6, 8, 1, 1, 6, 6, 4, and 6 cases, respectively. As to HCA subtypes, HNF1alpha-inactivated HCA, beta-catenin activated HCA (b-HCA), inflammatory HCA (IHCA) and unclassified HCA (U-HCA) accounted for nine (23.7%), four (10.5%), 17 (44.7%) and eight (21.1%) cases, respectively. Two cases showed coexistence of HCA and hepatocellular carcinoma (HCC) at surgery, and another had HCC which had been detected 23 years after HCA diagnosis. The HCA subtype of one of the former cases was U-HCA, while the remaining two had b-HCA and U-HCA. CONCLUSIONS: In Japanese HCA cases, the proportions of U-HCA, male and elder cases were slightly higher than in Western countries, and most of elder patients were male. IHCA was however common regardless of race, and was assumed to be the predominant subtype of HCA.

Ileal Dieulafoy lesion arose 15 years after partial small bowel resection for meconium obstruction of the neonate: a case report.

BACKGROUND: Anastomotic or perianastomotic ulcers present with symptoms such as chronic anaemia and occult bleeding as long-term complications of bowel resection performed in infancy. CASE PRESENTATION: Herein, we describe a 15-year-old girl with a history of surgery for meconium obstruction without mucoviscidosis in infancy who was hospitalized with chief complaints of presyncope and convulsions. Seven hours after admission, she developed melena and went into shock. An emergency laparotomy was performed, and a Dieulafoy lesion was detected near the site of ileal anastomosis from the surgery that had been performed during infancy. CONCLUSIONS: Although overt massive lower gastrointestinal bleeding necessitating emergency care is rare in the long term after infant bowel resection, Dieulafoy lesions can cause serious bleeding, requiring rapid life-saving haemostatic procedures.

Quantitative Ultrasound Image Analysis Helps in the Differentiation of Hepatocellular Carcinoma (HCC) From Borderline Lesions and Predicting the Histologic Grade of HCC and Microvascular Invasion.

OBJECTIVES: Quantitative image analysis is one of the methods to overcome the lack of objectivity of ultrasound (US). The aim of this study was to clarify the correlation between the features from a US image analysis and the histologic grade and microvascular invasion (MVI) of hepatocellular carcinoma (HCC) and differentiation of HCC smaller than 2 cm from borderline lesions. METHODS: We retrospectively analyzed grayscale US images with histopathologic evidence of HCC or a precancerous lesion using ImageJ version 1.47 software (National Institutes of Health, Bethesda, MD). RESULTS: A total of 148 nodules were included (borderline lesion, n = 31; early HCC [eHCC], n = 3; well-differentiated HCC [wHCC], n = 16; moderately differentiated HCC [mHCC], n = 79; and poorly differentiated HCC [pHCC], n = 19). A multivariate analysis selected lower minimum gray values (odds ratio [OR], 0.431; P = .003) and a higher standard deviation (OR, 1.880; P = .019) as predictors of HCC smaller than 2 cm. Median (range) minimum gray values of borderline lesions, eHCC, wHCC, mHCC, and pHCC were 29 (0-103), 7 (0-47), 6 (0-60), 10 (0-53), and 2 (0-38), respectively, and gradually decreased from borderline lesions to pHCC (P < 0.001). The multivariate analysis showed a higher aspect ratio (OR, 2.170; P = .001) and lower minimum gray value (OR, 0.475; P = .043) as predictors of MVI. An anechoic area diagnosed by a subjective evaluation was correlated with the minimum gray value (P < .0001). The proportion of the anechoic area gradually increased from eHCC to pHCC (P = .031). CONCLUSIONS: In a US image analysis, HCC smaller than 2 cm had features of greater heterogeneity and a lower minimum gray value than borderline lesions. Moderately differentiated HCC was smoother than borderline lesions, and the anechoic area correlated with histologic grading. Microvascular invasion was correlated with a slender shape and a lower minimum gray value.

Knockdown of Mitogen-Activated Protein Kinase Kinase 3 Negatively Regulates Hepatitis A Virus Replication.

Zinc chloride is known to be effective in combatting hepatitis A virus (HAV) infection, and zinc ions seem to be especially involved in Toll-like receptor (TLR) signaling pathways. In the present study, we examined this involvement in human hepatoma cell lines using a human TLR signaling target RT-PCR array. We also observed that zinc chloride inhibited mitogen-activated protein kinase kinase 3 (MAP2K3) expression, which could downregulate HAV replication in human hepatocytes. It is possible that zinc chloride may inhibit HAV replication in association with its inhibition of MAP2K3. In that regard, this study set out to determine whether MAP2K3 could be considered a modulating factor in the development of the HAV pathogen-associated molecular pattern (PAMP) and its triggering of interferon-ß production. Because MAP2K3 seems to play a role in antiviral immunity against HAV infection, it is a promising target for drug development. The inhibition of MAP2K3 may also prevent HAV patients from developing a severe hepatitis A infection.

Follow-Up of Patients Who Achieved Sustained Virologic Response after Interferon-Free Treatment against Hepatitis C Virus: Focus on Older Patients.

Background and Objectives: Direct-acting antiviral agents (DAAs) have improved sustained virologic response (SVR) rates in patients with chronic hepatitis C virus (HCV) infection. Our aim was to elucidate the occurrence of hepatocellular carcinoma (HCC) and to compare the outcomes of patients aged 75 years or older (older group) with those of patients younger than 75 years (younger group) after SVR. Materials and Methods: Among 441 patients treated with interferon-free DAA combinations, a total of 409 SVR patients were analyzed. We compared the two age groups in terms of HCC incidence and mortality rates. Results: Older and younger groups consisted of 68 and 341 patients, respectively. Occurrence of HCC after SVR did not differ between the two groups of patients with a history of HCC. Occurrence of HCC after SVR was observed more in younger patients without a history of HCC (p < 0.01). Although older patients without a history of HCC had a higher mortality rate (p < 0.01), their causes of death were not associated with liver diseases. Among younger patients without a history of HCC, none died. Conclusions: After SVR, liver disease may not be a prognostic factor in older HCV patients without a history of HCC.

Spatial and temporal expansion of intrahepatic metastasis by molecularly-defined clonality in multiple liver cancers.

Multiple hepatocellular carcinoma (HCC) is divided into two categories: intrahepatic metastasis (IM), which is a true relapse of HCC, and multicentric origin (MO), which is a second primary tumor. Clinical diagnosis of multiple HCC is usually made based on tumor location and/or time to recurrence; however, it is often difficult to distinguish the two types of multiple HCC. Using 41 matched pairs of multiple HCC specimens, we confirmed the accuracy of clinical diagnoses using exome sequence data and investigated the importance of discriminating the type of multiple HCC. Genomic analysis revealed that 18 (43.9%) patients diagnosed as having genomic IM had common mutations in a pair of HCC tumors with the main tumor of these patients being more progressive compared to those with genomic MO. The accuracy of clinical diagnosis based on lobe (Definition 1) and segment (Definition 2) were 68.3% and 78.0%, respectively. Intriguingly, recurrence ≥2 years after initial surgery for 3 patients was IM. The survival of patients with clinical IM was significantly shorter than for those with clinical MO based on both Definition 1 (P = 0.045) and Definition 2 (P = 0.043). However, mean survival was not different between the patients with genomic IM and those with MO (P = 0.364). Taken together, genomic analysis elucidated that liver cancer may spread more extensively and more slowly than previously thought. In addition, distinguishing multiple HCC as IM or MC may have provided biological information but was not of clinical importance with respect to patient prognosis.

Elderly-onset Crohn's disease remarkably responsive to ustekinumab: a case report.

BACKGROUND: Crohn's disease is intractable and is frequently diagnosed in younger people. No clear policies exist regarding medical treatment for seniors with this disease, and its diagnosis and treatment are often hindered by difficulties attributable to comorbidities, complex differential diagnoses, and polypharmacy. We describe an elderly-onset Crohn's disease patient showing a marked remission-maintaining effect with no adverse events after administration of ustekinumab. METHODS AND RESULTS: A 75-year-old patient with Crohn's disease and a history of pulmonary tuberculosis had first presented to our hospital at age 64 years and was hospitalized. Based on physical examinations, colonoscopy, and blood test results, Crohn's disease was diagnosed. The patient experienced secondary losses of responsiveness to two tumor necrosis factor (TNF)-alpha inhibitors, and after repeated hospital admissions, she was administered ustekinumab. The patient's symptoms, endoscopic findings, Crohn's Disease Activity Index, serum albumin, and physical activity levels improved markedly, and disease remission has been maintained for 2 years to date. CONCLUSION: Ustekinumab is an effective treatment option for elderly patients with intractable Crohn's disease when TNF-alpha inhibitors are ineffective.