Risk Factors for Immune Checkpoint Inhibitor-Induced Liver Injury and the Significance of Liver Biopsy.

Immune checkpoint inhibitor (ICI)-induced liver injury (LI) is a common adverse event, but the clinical characteristics based on the classification of hepatocellular injury and cholestatic types are not fully evaluated. This study aims to analyze risk factors and histological findings in relation to the classification of ICI-induced LI. In total, 254 ICI-induced LI patients among 1086 treated with ICIs between September 2014 and March 2022 were classified according to the diagnostic criteria for drug-induced LI (DILI), and their risk factors and outcomes were evaluated. Kaplan-Meier analyses showed that overall survival in patients with hepatocellular-injury-type LI was significantly longer than others (p < 0.05). Regarding pre-treatment factors, the lymphocyte count was significantly higher in patients with ICI-induced LI, especially in hepatocellular-injury-type LI. Gamma glutamyl transferase (γGTP) and alkaline phosphatase (ALP) were also significantly lower in patients with ICI-induced LI (p < 0.05). Multivariate analyses revealed that malignant melanoma, high lymphocyte count, and low ALP levels were extracted as factors contributing to hepatocellular-injury-type LI. The histological findings among 37 patients diagnosed as ICI-induced LI via liver biopsy also revealed that the spotty/focal necrosis was significantly frequent in hepatocellular-injury-type LI, whereas ductular reactions were frequently observed in cholestatic-type LI. It is suggested that the histological inflammation pattern in patients with LI is closely correlated with the type of DILI.

Clinical Significance and Remaining Issues of Anti-HBc Antibody and HBV Core-Related Antigen.

Currently, hepatitis B virus (HBV) core antibody (anti-HBc antibody) and HBV core-related antigen (HBcrAg) are widely used as serum markers for diagnosis based on the HBV core region. This review focused on anti-HBc antibodies and HBcrAg and aimed to summarize the clinical significance of currently used assay systems and the issues involved. While anti-HBc is very significant for clinical diagnosis, the clinical significance of quantitative assay of anti-HBc antibody has been reevaluated with improvements in diagnostic performance, including its association with clinical stage and prediction of carcinogenesis and reactivation. In addition, concerning the new HBcrAg, a high-sensitivity assay method has recently been established, and its diagnostic significance, including the prediction of reactivation, is being reevaluated. On the other hand, the quantitative level of anti-HBc antibody expressed in different units among assay systems complicates the interpretation of the results. However, it is difficult to standardize assay systems as they vary in advantages, and caution is needed in interpreting the assay results. In conclusion, with the development of highly sensitive HBcrAg and anti-HBc antibody, a rapid and sensitive detection assay system has been developed and used in clinical practice. In the future, it is hoped that a global standard will be created based on the many clinical findings.

Current Roles of Ramucirumab in the Sequential Treatment of Unresectable Hepatocellular Carcinoma.

BACKGROUND/AIM: The treatment algorithm for systemic therapies for advanced hepatocellular carcinoma (HCC) has changed dramatically; however, the therapeutic landscape for sequential second-line or later-line treatments, including ramucirumab, remains controversial. This study aimed to investigate the role of ramucirumab for treating HCC. PATIENTS AND METHODS: We retrospectively analyzed data from 17 patients with advanced HCC who received ramucirumab, and 8 of them who received lenvatinib re-administration after ramucirumab treatment failure. RESULTS: The median overall survival of 17 patients treated with ramucirumab was 11.5 months. The median ratios of the 1-month post-treatment α-fetoprotein (AFP) levels and albumin-bilirubin (ALBI) scores to the pre-treatment AFP levels and ALBI scores following ramucirumab treatment were 0.880 and 0.965, respectively. The median ratios of the 1-month post-treatment AFP and ALBI levels to the pre-treatment levels were 1.587 and 0.970 for mALBI grade 1/2a, and 1.313 and 0.936 for mALBI grade 2b/3, respectively. Six of the eight patients who received lenvatinib rechallenge treatment exhibited a decrease in AFP levels one month post-lenvatinib treatment. Deterioration of liver function 3 months post-lenvatinib treatment was noted in five of the eight patients who received lenvatinib rechallenge treatment after ramucirumab. CONCLUSION: Ramucirumab may be equally useful in patients with unresectable HCC who have poor liver function or whose liver function is aggravated by other therapies. Rechallenge treatment with lenvatinib after ramucirumab may be a valid treatment option for HCC.

Association between tumor morphology and efficacy of atezolizumab plus bevacizumab for advanced hepatocellular carcinoma.

AIM: The IMbrave150 trial revealed that atezolizumab plus bevacizumab (AtezoBv) showed a higher objective response rate (ORR) in patients with advanced hepatocellular carcinoma (HCC). Although conversion therapy after AtezoBv has been recently reported, markers predictive of its efficacy, particularly radiological imaging markers, have not yet been identified. The present study focused on tumor morphological appearance on radiological imaging and evaluated whether it could be associated with AtezoBv efficacy. METHODS: Ninety-five intrahepatic lesions in 74 patients who were given AtezoBv for advanced HCC were recruited for evaluation. The lesions were divided into two groups, simple nodular (SN group) and non-simple nodular (non-SN group), based on the gross morphology on pretreatment imaging, and retrospectively evaluated for treatment response and other relevant clinical outcomes. RESULTS: Assessing the size of individual tumors after treatment, waterfall plots showed that tumor shrinkage in the non-SN group including 56 lesions was higher than that in the SN group comprising 39 lesions. The ORR was significantly higher in the non-SN group (39.3% vs. 15.4%, p = 0.012). Additionally, the median time to nodular progression was longer in the non-SN group (21.0 months vs. 8.1 months, p = 0.119) compared to the SN group. Six patients with non-SN lesions underwent sequential local therapy. CONCLUSIONS: Atezolizumab plus bevacizumab may show increased therapeutic efficacy in patients with tumors with a higher potential for aggressive oncological behavior, such as non-SN lesions. Treatment strategies focusing on conversion therapy may be crucial in patients with non-SN lesions.

An Analysis of 10,000 Cases of Living Donor Liver Transplantation in Japan: Special Reference to the Graft-Versus-Recipient Weight Ratio and Donor Age.

OBJECTIVE: To analyze 10,000 cases of living donor liver transplantation (LDLT) recipient data to elucidate outcomes with special reference to the graft-versus-recipient weight ratio (GRWR), based on the Japanese Liver Transplantation Society (JLTS) registry. BACKGROUND: The JLTS registry has been accurate and complete in characterizing and following trends in patient characteristics and survival of all patients with LDLT. METHODS: Between November 1989 and August 2021, 10,000 patients underwent LDLT in Japan. The procedures performed during the study period included pediatric liver transplantation (age <18 years, n = 3572) and adult liver transplantation (age ≥18 years, n=6428). Factors related to patient survival (PS) and graft survival (GS) were also analyzed. RESULTS: The GRWR was <0.7, 0.7 to <0.8, 0.8 to <3, 3 to <5, and ≥5 in 0.2%, 2.0%, 61.8%, 31.8%, and 2.6% of pediatric patients and <0.6, 0.6 to <0.7, 0.7 to <0.8, and ≥0.8 in 8.0%, 12.7%, 17.7%, and 61.5% of adult patients, respectively. Among pediatric recipients, the PS rate up to 5 years was significantly better in cases with a GRWR ≤5 than in those with a GRWR >5. When the GRWR and donor age were combined, among adult recipients 50 to 60 years old, the early PS and GS up to 5 years were significantly better in cases with a GRWR ≥0.7, than in those with a GRWR <0.7. (P = 0.02). In adults, a multivariate analysis showed that GRWR <0.6, transplant era (<2011), donor age (>60 years), recipient age (>60 years), model for end-stage liver disease score (≥20), and center volume (<10) were significant prognostic factors for long-term PS. CONCLUSION: Although a satisfactory long-term PS and GS, especially in the recent era (2011-2021), was achieved in the JLTS series, a GRWR ≥5 in pediatric cases and relatively old donors with a GRWR <0.7 in adult cases should be managed with caution.

Histological diagnosis of polyploidy discriminates an aggressive subset of hepatocellular carcinomas with poor prognosis.

BACKGROUND: Although genome duplication, or polyploidization, is believed to drive cancer evolution and affect tumor features, its significance in hepatocellular carcinoma (HCC) is unclear. We aimed to determine the characteristics of polyploid HCCs by evaluating chromosome duplication and to discover surrogate markers to discriminate polyploid HCCs. METHODS: The ploidy in human HCC was assessed by fluorescence in situ hybridization for multiple chromosomes. Clinicopathological and expression features were compared between polyploid and near-diploid HCCs. Markers indicating polyploid HCC were explored by transcriptome analysis of cultured HCC cells. RESULTS: Polyploidy was detected in 36% (20/56) of HCCs and discriminated an aggressive subset of HCC that typically showed high serum alpha-fetoprotein, poor differentiation, and poor prognosis compared to near-diploid HCCs. Molecular subtyping revealed that polyploid HCCs highly expressed alpha-fetoprotein but did not necessarily show progenitor features. Histological examination revealed abundant polyploid giant cancer cells (PGCCs) with a distinct appearance and frequent macrotrabecular-massive architecture in polyploid HCCs. Notably, the abundance of PGCCs and overexpression of ubiquitin-conjugating enzymes 2C indicated polyploidy in HCC and efficiently predicted poor prognosis in combination. CONCLUSIONS: Histological diagnosis of polyploidy using surrogate markers discriminates an aggressive subset of HCC, apart from known HCC subgroups, and predict poor prognosis in HCC.

Factors associated with the response to atezolizumab/bevacizumab combination therapy for hepatocellular carcinoma.

Background and Aim: The purpose of this study was to analyze factors associated with the overall survival (OS) of atezolizumab/bevacizumab combination therapy for advanced hepatocellular carcinoma (aHCC). We also assessed the OS of patients with ineffective therapy and those who discontinued treatment owing to adverse events (AEs). Methods: This retrospective multicenter study involved 139 patients with aHCC who received atezolizumab/bevacizumab combination therapy between November 2020 and September 2022. Results: The median duration of treatment was 136.5 days, and the median observation period was 316 days. The overall response rate was 40%, and the disease control rate was 78% according to mRECIST criteria. Grade ≥2 AEs occurred in 63 patients (43%) and led to treatment discontinuation in 16 patients. Multivariate analysis revealed that treatment response and occurrence of grade ≥2 AEs after therapy, as well as low level of albumin-bilirubin (ALBI) grade and low level of des-gamma carboxy prothrombin (DCP) before therapy, were extracted as factors that contributed to OS. Log-rank tests with the Kaplan-Meier method showed significant differences in OS among these factors. The OS of patients who discontinued owing to AEs was significantly shorter than that of other patients. Conclusion: Not only factors before therapy but also treatment response and the appearance of AEs are involved in OS for atezolizumab/bevacizumab combination therapy. Although the development of AEs also contributed to OS, appropriate management of AEs is important to avoid discontinuing treatment with this combination.

Anti-integrin αvß6 autoantibodies in patients with primary sclerosing cholangitis.

BACKGROUND: Patients with primary sclerosing cholangitis (PSC) possess autoantibodies against biliary epithelial cells. However, the target molecules remain unknown. METHODS: The sera of patients with PSC and controls were subjected to enzyme-linked immunosorbent assays to detect autoantibodies using recombinant integrin proteins. Integrin αvß6 expression in the bile duct tissues was examined using immunofluorescence. The blocking activity of the autoantibodies was examined using solid-phase binding assays. RESULTS: Anti-integrin αvß6 antibodies were detected in 49/55 (89.1%) patients with PSC and 5/150 (3.3%) controls (P < 0.001), with a sensitivity and specificity of 89.1% and 96.7%, respectively, for PSC diagnosis. When focusing on the presence or absence of IBD, the proportion of the positive antibodies in PSC with IBD was 97.2% (35/36) and that in PSC alone was 73.7% (14/19) (P = 0.008). Integrin αvß6 was expressed in bile duct epithelial cells. Immunoglobulin (Ig)G from 15/33 patients with PSC blocked integrin αvß6-fibronectin binding through an RGD (Arg-Gly-Asp) tripeptide motif. CONCLUSIONS: Autoantibodies against integrin αvß6 were detected in most patients with PSC; anti-integrin αvß6 antibody may serve as a potential diagnostic biomarker for PSC.

Evolutional transition of HBV genome during the persistent infection determined by single-molecule real-time sequencing.

BACKGROUND: Although HBV infection is a serious health issue worldwide, the landscape of HBV genome dynamics in the host has not yet been clarified. This study aimed to determine the continuous genome sequence of each HBV clone using a single-molecule real-time sequencing platform, and clarify the dynamics of structural abnormalities during persistent HBV infection without antiviral therapy. PATIENTS AND METHODS: Twenty-five serum specimens were collected from 10 untreated HBV-infected patients. Continuous whole-genome sequencing of each clone was performed using a PacBio Sequel sequencer; the relationship between genomic variations and clinical information was analyzed. The diversity and phylogeny of the viral clones with structural variations were also analyzed. RESULTS: The whole-genome sequences of 797,352 HBV clones were determined. The deletion was the most common structural abnormality and concentrated in the preS/S and C regions. Hepatitis B e antibody (anti-HBe)-negative samples or samples with high alanine aminotransferase levels have significantly diverse deletions than anti-HBe-positive samples or samples with low alanine aminotransferase levels. Phylogenetic analysis demonstrated that various defective and full-length clones evolve independently and form diverse viral populations. CONCLUSIONS: Single-molecule real-time long-read sequencing revealed the dynamics of genomic quasispecies during the natural course of chronic HBV infections. Defective viral clones are prone to emerge under the condition of active hepatitis, and several types of defective variants can evolve independently of the viral clones with the full-length genome.

Integrated analysis of tertiary lymphoid structures in relation to tumor-infiltrating lymphocytes and patient survival in pancreatic ductal adenocarcinoma.

BACKGROUND: Tertiary lymphoid structure (TLS) reflects an intense immune response against cancer, which correlates with favorable patient survival. However, the association of TLS with tumor-infiltrating lymphocytes (TILs) and clinical outcomes has not been investigated comprehensively in pancreatic ductal adenocarcinoma (PDAC). METHODS: We utilized an integrative molecular pathological epidemiology database on 162 cases with resected PDAC, and examined TLS in relation to levels of TILs, patient survival, and treatment response. In whole-section slides, we assessed the formation of TLS and conducted immunohistochemistry for tumor-infiltrating T cells (CD4, CD8, CD45RO, and FOXP3). As confounding factors, we assessed alterations of four main driver genes (KRAS, TP53, CDKN2A [p16], and SMAD4) using next-generation sequencing and immunohistochemistry, and tumor CD274 (PD-L1) expression assessed by immunohistochemistry. RESULTS: TLSs were found in 112 patients with PDAC (69.1%). TLS was associated with high levels of CD4+ TILs (multivariable odds ratio [OR], 3.50; 95% confidence interval [CI] 1.65-7.80; P = 0.0002), CD8+ TILs (multivariable OR, 11.0; 95% CI 4.57-29.7, P < 0.0001) and CD45RO+ TILs (multivariable OR, 2.65; 95% CI 1.25-5.80, P = 0.01), but not with levels of FOXP3+ TILs. TLS was associated with longer pancreatic cancer-specific survival (multivariable hazard ratio, 0.37; 95% CI 0.25-0.56, P < 0.0001) and favorable outcomes of adjuvant S-1-treatment. TLS was not associated with driver gene alterations but tumor CD274 negative expression. CONCLUSIONS: Our comprehensive data supports the surrogacy of TLS for vigorous anti-tumor immune response characterized by high levels of helper and cytotoxic T cells and their prognostic role.

Outcomes of the Sequential Treatment of Unresectable Hepatocellular Carcinoma Using Lenvatinib.

BACKGROUND/AIM: The chemotherapeutic landscape for hepatocellular carcinomas (HCCs) has changed dramatically with the availability of several treatment options. This study aimed to assess the long-term outcomes of lenvatinib treatment and analyze its feasibility in the sequential treatment of HCCs. PATIENTS AND METHODS: Eighty-five consecutive patients who received lenvatinib for unresectable HCCs were investigated retrospectively. Survival was assessed based on when the patients were first radiologically diagnosed with progressive disease. Among those with radiologically diagnosed stable or progressive disease at 3 months after lenvatinib administration, the cutoff α-fetoprotein (AFP) ratio (ratio of the AFP level after lenvatinib treatment to the pretreatment AFP level) that was predictive of survival was determined using receiver operating characteristic analysis. RESULTS: The median survival time (MST) was significantly worse among patients diagnosed with progressive disease at 1 month after treatment than among those diagnosed at 2-3 or 3-4 months after treatment [MSTs at 1, 2-3, and 3-4 months: 2.2, 10.2, and 17.3 months, respectively (p<0.001)]. An AFP ratio of 1.36 (computed using the AFP level at 3 months after lenvatinib treatment) was significantly predictive of survival in patients with stable or progressive disease (26.3 vs. 11.3 months, p=0.0024). CONCLUSION: The prognosis of patients on lenvatinib who develop early progressive disease is dismal. Thus, their treatment should be ceased or switched. The 3-month AFP ratio of 1.36 may be a potentially useful cutoff for considering a switch to other treatments in patients radiologically diagnosed with stable or progressive disease.

Population Dynamics of Community Cats Living in a Tourist Area of Onomichi City, Japan, before and after the Trap-Test-Vaccinate-Alter-Return-Monitor Event.

The community cat program (CCP) was recommended by the Ministry of the Environment to reduce cats in local animal shelters and improve stray cat welfare in Japan. It is a non-lethal control measure with stray cats cared for as free-roaming cats for their lifetime in the community, while Trap-Neuter-Return (TNR) or Trap-Test-Vaccinate-Alter-Return-Monitor (TTVARM) activities are carried out. In the CCP, community cat colonies are hypothesized to be closed and static populations. However, it remains unknown whether the cats stay in the colonies, without migration of non-neutered cats following TNR/TTVARM events. We examined the population dynamics of cats before and after a TTVARM event using route censuses (107 days), fixed-point observations, and GPS-tracking in a tourist area in Onomichi. Eleven out of the 30 cats remained in the CCP areas, whereas 13 non-neutered cats immigrated into the CCP areas, within a year, suggesting the CCP program has limited efficacy. Besides, the program cannot support the lifetime management of the cats due rapid turnover of cats. Our results reject the CCP hypothesis, so that the program neither restricts cat breeding nor enhances cat welfare.

Constitution of mucosa-associated microbiota in the lower digestive tract does not change in early stage of non-alcoholic fatty liver disease with fecal dysbiosis.

Background and Aim: Regarding the gut-liver axis, fecal dysbiosis is implicated in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). The significance of mucosa-associated microbiota (MAM, which is present in the mucin layer covering the intestinal mucosa) has not been well explored. We aimed to clarify the characteristics of MAM in patients with NAFLD. Methods: MAM were obtained from seven patients with early-stage NAFLD and seven controls by colonoscopy in five locations (terminal ileum, cecum, ascending and sigmoid colon, and rectum) using mucosal brushes. The microbial 16S rDNA profiles of the MAM and fecal microbiota of patients in the NAFLD and control groups were analyzed. Results: α-diversities of fecal microbiota were decreased in patients with NAFLD (observed species, Shannon index, and Chao1: 174.57 vs 134.86, 5.51 vs 4.65, and 206.34 vs 167.91; P = 0.048, 0.067, and 0.087, respectively), and microbial composition analyses by principal coordinate analysis differed between the fecal microbiota of patients with NAFLD and those of controls (permutational analysis of variance [PERMANOVA] of weighted and unweighted: Pseud-F: 1.4179/P-value: 0.05 and Pseud-F: 2.1497/P-value: 0.049, respectively). However, α-diversities or microbial composition of MAM in most parts of the intestine did not differ significantly between the NAFLD and control groups. Unclassified Rikenellaceae, Oscillospira, Odoribacter, unclassified clostridiales, and Holdemania were decreased in the feces of patients with NAFLD (determined by linear discriminant analysis effect size), but five (except Holdemania) of the six genera were not decreased in the MAM of these patients. Conclusion: In early-stage NAFLD, MAM was uniform and relatively stable throughout the intestine, even when fecal dysbiosis appeared.

Rechallenge With Lenvatinib After Atezolizumab Plus Bevacizumab Treatment for Hepatocellular Carcinoma.

BACKGROUND/AIM: Atezolizumab plus bevacizumab and lenvatinib are the key drugs in the current systemic chemotherapeutic regimen for hepatocellular carcinoma (HCC). Studies have reported the potential effectiveness of lenvatinib introduction after an atezolizumab plus bevacizumab treatment; however, the therapeutic effectiveness of a lenvatinib rechallenge after an atezolizumab plus bevacizumab treatment remains unclear. PATIENTS AND METHODS: Thirteen consecutive patients who were rechallenged with lenvatinib after clinical failure following treatments with lenvatinib and atezolizumab plus bevacizumab were included. A comparative study was conducted on the duration and treatment efficacy of the first and second lenvatinib treatments and on the pre- and post-treatment liver function. RESULTS: The median ratios of the 1-month post-treatment alpha-fetoprotein (AFP) levels to the pretreatment AFP levels were 0.750 and 0.667 for the first and second lenvatinib treatments, respectively, without significant difference (p=0.9327). Meanwhile, the median ratios of the 1-month post-treatment albumin-bilirubin (ALBI) scores to the pretreatment ALBI scores were 1.063 and 0.827 for the first and second lenvatinib treatments, respectively, with significant difference (p=0.015). The median duration of the second lenvatinib treatment was significantly shorter than that of the first lenvatinib treatment [2.8 months (range=0.9-4.7 months) vs. 8.7 months (range=3.1-29.7 months)]. CONCLUSION: Lenvatinib re-administration after atezolizumab plus bevacizumab treatment can act as a double-edged sword, as it exerts an anti-tumor effect while being associated with potential liver function deterioration. However, this treatment sequence can be useful, and requires careful monitoring of the transitions in the liver function and the patient's performance status.

Significance of post-progression therapy after tyrosine kinase inhibitors for advanced hepatocellular carcinoma.

Background and Aim: Molecular-targeted therapies such as sorafenib and lenvatinib have long been used as first-line treatment for advanced hepatocellular carcinoma (aHCC). However, adverse events or limited therapeutic effects may necessitate the change to another therapeutic option, known as post-progression therapy. To investigate the significance of post-progression therapy, we analyzed the outcomes of aHCC patients following first-line molecular-targeted therapy in a real-world study. Methods: This retrospective, multicenter study involved patients with aHCC who received sorafenib or lenvatinib as first-line therapy between January 2011 and September 2021. Results: In total, 513 patients were analyzed: 309 treated with sorafenib and 204 with lenvatinib. The overall response and disease control rates were 15 and 50%, respectively, in the sorafenib group and 30 and 75%, respectively, in the lenvatinib group (P < 0.001). Kaplan-Meier analysis revealed no significant differences in progression-free survival and overall survival (OS) between the two treatments. Multivariate analysis revealed that fibrosis-4 index, disease control rate, post-progression therapy, and use of an immune checkpoint inhibitor (ICI) were significantly associated with OS. OS was significantly longer in patients who received post-progression therapy than in those who did not (log-rank P < 0.001). Most patients who received an ICI as post-progression therapy had previously received lenvatinib. Among lenvatinib-treated patients, OS was significantly longer in patients who received an ICI than in patients received another or no post-progression therapy (P = 0.004). Conclusion: The introduction of newer drugs for post-progression therapy is expected to prolong survival. ICI-based regimens appear to be effective after lenvatinib.

Allograft liver failure awaiting liver transplantation in Japan.

BACKGROUND: Following liver transplantation (LT), allograft liver failure can be developed by various causes and requires re-LT. Hence, this study aimed to clarify the characteristics and prognostic factors of patients with allograft liver failure awaiting deceased donor LT (DDLT) in Japan. METHODS: Of the 2686 DDLT candidates in Japan between 2007 and 2016, 192 adult patients listed for re-LT were retrospectively enrolled in this study. Factors associated with waitlist mortality were assessed using the Cox proportional hazards model. The transplant-free survival probabilities were evaluated using the Kaplan-Meier analysis and log-rank test. RESULTS: The median period from the previous LT to listing for re-LT was 1548 days (range, 4-8449 days). Primary sclerosing cholangitis (PSC), which was a primary indication, showed a higher listing probability for re-LT as compared with other primary etiologies. Recurrent liver disease was a leading cause of allograft failure and was more frequently observed in the primary indication of hepatitis C virus (HCV) infection and PSC in contrast with other etiologies. Multivariate analysis identified the following independent risk factors associated with waitlist mortality: age, Child-Turcotte-Pugh (CTP) score, mode for end-stage liver disease (MELD) score, alanine aminotransferase (ALT), and causes of allograft failure. CONCLUSIONS: Recurrent HCV and PSC were major causes of allograft liver failure in Japan. In addition to CTP and MELD scores, either serum ALT levels or causes of allograft failure should be considered as graft liver allocation measures.

Comprehensive Analysis of Molecular Biologic Characteristics of Pancreatic Ductal Adenocarcinoma Concomitant with Intraductal Papillary Mucinous Neoplasm.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) concomitant with intraductal papillary mucinous neoplasm (IPMN) is defined as PDAC occurring apart from IPMN. This study comprehensively investigated the molecular biologic characteristics of PDAC concomitant with IPMN in major genetic alterations, tumor microenvironment, and prognosis by contrast with those of conventional PDAC. METHODS: The study retrospectively reviewed the data of 158 surgically resected PDAC patients. The driver gene alteration status (KRAS, TP53, CDKN2A, SMAD4, and GNAS) together with the immune and fibrotic status in tumor was evaluated. The prognosis of PDAC concomitant with IPMN and that of conventional PDAC also were compared. RESULTS: No statistically significant difference was found between PDAC concomitant with IPMN and conventional PDAC in the alteration frequency analysis of the major driver genes and the immune and fibrotic status in the tumor microenvironment. Overall survival and disease-free survival between patients who had PDAC concomitant with IPMN and those who had conventional PDAC did not show statistically significant differences in propensity-matched subjects. Furthermore, the co-existence of IPMN was not a poor prognostic factor in the multivariable-adjusted Cox proportional hazards model (hazard ratio, 0.95; 95 % confidence interval, 0.51-1.78). CONCLUSIONS: In this study, PDAC concomitant with IPMN had tumor characteristics similar to those of conventional PDAC in terms of the major driver gene alterations, tumor microenvironment, and prognosis.

Expansion of Gastric Intestinal Metaplasia with Copy Number Aberrations Contributes to Field Cancerization.

Intestinal metaplasia (IM) is a risk factor for gastric cancer following infection with Helicobacter pylori. To explore the susceptibility of pure gastric IM to cancer development, we investigated genetic alterations in single IM gastric glands. We isolated 50 single IM or non-IM glands from the inflamed gastric mucosa of 11 patients with intramucosal gastric carcinoma (IGC) and 4 patients without IGC; 19 single glands in the noninflamed gastric mucosa of 11 individuals from our cohort and previous dataset were also included as controls. Whole-exome sequencing of single glands revealed significantly higher accumulation of somatic mutations in various genes within IM glands compared with non-IM glands. Clonal ordering analysis showed that IM glands expanded to form clusters with shared mutations. In addition, targeted-capture deep sequencing and copy number (CN) analyses were performed in 96 clustered IM or non-IM gastric glands from 26 patients with IGC. CN analyses were also performed on 41 IGC samples and The Cancer Genome Atlas-Stomach Adenocarcinoma datasets. These analyses revealed that polyclonally expanded IM commonly acquired CN aberrations (CNA), including amplification of chromosomes 8, 20, and 2. A large portion of clustered IM glands typically consisted of common CNAs rather than other cancer-related mutations. Moreover, the CNA patterns of clustered IM glands were similar to those of IGC, indicative of precancerous conditions. Taken together, these findings suggest that, in the gastric mucosa inflamed with H. pylori infection, IM glands expand via acquisition of CNAs comparable with those of IGC, contributing to field cancerization. SIGNIFICANCE: This study contributes to our understanding of gastric intestinal metaplasia as a risk factor for gastric adenocarcinoma via their multifocal expansion and acquisition of CNAs and somatic mutations.

Mutational spectrum of hepatitis C virus in patients with chronic hepatitis C determined by single molecule real-time sequencing.

The emergence of hepatitis C virus (HCV) with resistance-associated substitution (RAS), produced by mutations in the HCV genome, is a major problem in direct acting antivirals (DAA) treatment. This study aimed to clarify the mutational spectrum in HCV-RNA and the substitution pattern for the emergence of RASs in patients with chronic HCV infection. HCV-RNA from two HCV replicon cell lines and the serum HCV-RNA of four non-liver transplant and four post-liver transplant patients with unsuccessful DAA treatment were analyzed using high-accuracy single-molecule real-time long-read sequencing. Transition substitutions, especially A>G and U>C, occurred prominently under DAAs in both non-transplant and post-transplant patients, with a mutational bias identical to that occurring in HCV replicon cell lines during 10-year culturing. These mutational biases were reproduced in natural courses after DAA treatment. RASs emerged via both transition and transversion substitutions. NS3-D168 and NS5A-L31 RASs resulted from transversion mutations, while NS5A-Y93 RASs was caused by transition substitutions. The fidelity of the RNA-dependent RNA polymerase, HCV-NS5B, produces mutational bias in the HCV genome, characterized by dominant transition mutations, notably A>G and U>C substitutions. However, RASs are acquired by both transition and transversion substitutions, and the RASs-positive HCV clones are selected and proliferated under DAA treatment pressure.

Acid-treated high-amylose corn starch suppresses high-fat diet-induced steatosis.

Resistant starch (RS) has been reported to improve steatosis as well as obesity. Type 4 resistant starch (RS4), a chemically modified starch, is particularly hard to digest and suggesting higher efficacy. However, because the effects of RS4 on steatosis are not yet fully understood, the effects of RS4 on steatosis were examined using a murine high-fat diet model. Seven-week-old male mice were divided into three groups and fed a normal diet, a high-fat diet (HFD), or a high-fat diet with added RS (HFD + RS). Amylofiber SH® produced from acid-treated corn starch was used as the dietary RS. At 22 weeks old, hepatic steatosis and short chain fatty acid (SCFA) content and gut microbiota in cecum stool samples were analyzed. The ratio of body weight to 7 weeks was significantly suppressed in the HFD + RS group compared to the HFD group (132.2 ± 1.4% vs. 167.2 ± 3.9%, p = 0.0076). Macroscopic and microscopic steatosis was also suppressed in the HFD + RS group. Analysis of cecum stool samples revealed elevated SCFA levels in the HFD + RS group compared with the HFD group. Metagenome analysis revealed that Bifidobacterium (17.9 ± 1.9% vs. 3.6 ± 0.7%, p = 0.0019) and Lactobacillus (14.8 ± 3.4% vs. 0.72 ± 0.23%, p = 0.0045), which degrade RS to SCFA, were more prevalent in the HFD + RS group than the HFD group. In conclusion, RS4 suppressed steatosis, and increased Bifidobacterium and Lactobacillus, and SCFAs. RS4 may prevent steatosis by modulating the intestinal environment.