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The differential diagnosis of intrahepatic cholangiocarcinomas (iCCAs) from metastatic adenocarcinomas from organs adjacent to the liver (gallbladder, pancreas, and stomach) is difficult due to histopathological similarity and a lack of specific markers. This study aimed to develop a method to differentiate iCCA and adenocarcinomas originated from extrahepatic organs adjacent to the liver.We retrospectively enrolled surgically resected iCCA (n = 181) and adenocarcinomas from extrahepatic organs (n = 30, n = 28, and n = 38 from gallbladder, pancreas, and stomach, respectively) between 2007 and 2013. The albumin mRNA in situ hybridization (ISH) and immunohistochemistry (IHC) of filamin-A and cytokeratin 19 (CK19) were performed using tissue microarray. Using logistic regression analysis of three markers, iCCA-score was developed, and its diagnostic performance was evaluated.The iCCAs were more frequently positive for albumin ISH (23.2% vs. 0%), filamin-A IHC (47.5% vs. 12.5%) and CK19 (68.5% vs. 40.6%) than extrahepatic adenocarcinomas (p 0.001 for all). The iCCA-score consisting of these three markers was developed, and it showed higher diagnostic performance (area under the curve [AUC], 0.798 vs. 0.616, p 0.001). Taking an iCCA-score of 2 or higher as the threshold for iCCA, the sensitivity was substantially higher than albumin ISH alone (45.9% and 23.2%, respectively; p 0.001), but maintained high specificity (94.8% and 100%, respectively).Albumin ISH and IHC staining for filamin-A and CK19 showed distinct expression patterns between iCCA and extrahepatic adenocarcinomas from gallbladder, pancreas, and stomach. We developed iCCA-score that consisted of those three markers, and it showed better diagnostic performance than albumin ISH alone.

Background/Aim: Transforming growth factor-beta (TGF-β) has a dichotomous role, functioning as a tumor suppressor and tumor promoter. TGF-β signatures, explored in mouse hepatocytes, have been reported to predict the clinical outcomes of hepatocellular carcinoma (HCC) patients; HCCs exhibiting early TGF-β signatures showed a better prognosis than those with late TGF-β signatures. The expression status of early and late TGF-β signatures remains unclear in defined lesions of human B-viral multistep hepatocarcinogenesis.Methods: The expression of TGF-β signatures, early and late responsive signatures of TGF-β were investigated and analyzed for their correlation in cirrhosis, low-grade dysplastic nodules (DNs), high-grade DNs, early HCCs and progressed HCCs (pHCCs) by real-time PCR and immunohistochemistry.Results: The expression levels of TGF-β signaling genes (TGFB1, TGFBR1, TGFBR2 and SMAD4) gradually increased with the progression of hepatocarcinogenesis, peaking in pHCCs. The expression of early responsive genes of TGF-β (GADD45B, FBP1, CYP1A2 and CYP3A4) gradually decreased, and that of the late TGF-β signatures (TWIST and SNAI1) significantly increased according to the progression of multistep hepatocarcinogenesis. Furthermore, mRNA levels of TWIST and SNAI1 were well correlated with those of stemness markers, with upregulation of TGF-β signaling, whereas FBP1 expression was inversely correlated with that of stemness markers.Conclusions: The enrichment of the late responsive signatures of TGF-β with induction of stemness is considered to be involved in the progression of the late stage of multistep hepatocarcinogenesis, whereas the early responsive signatures of TGF-β are suggested to have tumor-suppressive roles in precancerous lesions of the early stage of multistep hepatocarcinogenesis.

Hepatocellular carcinoma (HCC) undergoes a stepwise progression from liver cirrhosis to low-grade dysplastic nodule (LGDN), high-grade dysplastic nodule (HGDN), early HCC (eHCC), and progressed HCC (pHCC). Here, we profiled multilayered genomic, epigenomic, and transcriptomic aberrations in the stepwise hepatocarcinogenesis. Initial DNA methylation was observed in eHCC (e.g., DKK3, SALL3, and SOX1) while more extensive methylation was observed in pHCC. In addition, eHCCs showed an initial loss of DNA copy numbers of tumor suppressor genes in the 4q and 13q regions, thereby conferring survival benefits to cancer cells. Transcriptome analysis revealed that HGDNs expressed endoplasmic reticulum (ER) stress–related genes, while eHCC started to express oncogenes. Furthermore, integrative analysis indicated that expression of the serine peptidase inhibitor, Kazal type 1 (SPINK1), played a pivotal role in eHCC development. Significant demethylation of SPINK1 was observed in eHCC compared to HGDN. The study also demonstrated that ER stress may induce SPINK1 demethylation and expression in liver cancer cells. In conclusion, these results reveal the dynamics of multiomic aberrations during malignant conversion of liver cancer, thus providing novel pathobiological insights into hepatocarcinogenesis.Significance:Multiomics profiling and integrative analyses of stepwise hepatocarcinogenesis reveal novel mechanistic and clinical insights into hepatocarcinogenesis.

Supplementary Figures S1-S9 S1: The representative case showing CD90 expression. Morphology and character of hepatic stellate cell lines. S2: The expression change of KRT19 following S3I-201, TPA, U0126, and SCH772984. The transfection efficiency of HCC cell lines. S3: The HGF concentrations in conditioned media from hTERT-HSC, LX-2, and primary human hepatic stellate cells. Western blot analysis of hepatocellular carcinoma cell lines, showing expression of phospho-MET, MET, phsopho-ERK1/2, and KRT19. mRNA and protein expression of KRT19 in HepG2 and SNU475 cells, following transient expression of MET. S4: Morphologic changes in HepG2 and SNU423 induced by conditioned media (CM) from inducible HGF knockdown stable cell lines established from hTERT-HSC: shHGF-1 and shHGF-2. mRNA expression levels of KRT19 and morphologic changes in HepG2 and SNU423, following CM treatment combined with hepatocyte growth factor (HGF) neutralizing antibody. S5: The gene knock-down efficiencies of two pooled siRNAs targeting JUN, JUND, FOSL1, or FOSL2. The gene knock-down efficiencies of siRNAs targeting JUN or FOSL1. S6: Expression of HGF in hepatic stellate cell line of hTERT-HSC, and HCC cell lines of HepG2 and SNU423. Distribution of HGF and MET expression in HepG2 cells, at various time points after conditioned media (CM) treatment. S7: S7. Increased KRT19 expression and nuclear translocation of ERK1/2, phospho-ERK1/2, FOSL1, phospho-FOSL1 after the conditioned media treatment in HepG2. Rapid decline in phosphor-MET levels after removal of conditioned media. S8: Schematic representation of regulatory mechanism of KRT19 gene by cancer-associated fibroblast (CAF) derived HGF via MET-ERK1/2- AP1 and SP1 axis. S9: Conservation of AP1 and SP1 binding sites in KRT19 promoter.

Supplementary Table S1: Tissue specimens used for RNA-seq analysis. Supplementary Table S2: Tissues specimens used for immunohistochemical analysis. Supplementary Table S3: Gene signatures used in the study. Supplementary Table S4: Differentially methylated probes during hepatocarcinogenesis. Supplementary Table S5: Univariate and multivariate analysis for SPINK1 expression and HCC clinico-pathological features in TCGA-LIHC data.

Supplementary Materials and Methods

Keratin (KRT) 19 is a poor prognostic marker for hepatocellular carcinoma (HCC); however, regulatory mechanisms underlying its expression remain unclear. We have previously reported the presence of fibrous tumor stroma in KRT19-positive HCC, suggesting that cross-talk between cancer-associated fibroblasts (CAF) and tumor epithelial cells could regulate KRT19 expression. This was investigated in this study using an in vitro model of paracrine interaction between HCC cell lines (HepG2, SNU423) and hepatic stellate cells (HSC), a major source of hepatic myofibroblasts. HSCs upregulated transcription and translation of KRT19 in HCC cells via paracrine interactions. Mechanistically, hepatocyte growth factor (HGF) from HSCs activated c-MET and the MEK–ERK1/2 pathway, which upregulated KRT19 expression in HCC cells. Furthermore, AP1 (JUN/FOSL1) and SP1, downstream transcriptional activators of ERK1/2, activated KRT19 expression in HCC cells. In clinical specimens of human HCC (n = 339), HGF and KRT19 protein expression correlated with CAF levels. In addition, HGF or MET protein expression was associated with FOSL1 and KRT19 expression and was found to be a poor prognostic factor. Analysis of data from The Cancer Genome Atlas also revealed KRT19 expression was closely associated with CAF and MET-mediated signaling activities. These results provide insights into the molecular background of KRT19-positive HCC that display an aggressive phenotype.Significance: These findings reveal KRT19 expression in hepatocellular carcinoma is regulated by cross-talk between cancer-associated fibroblasts and HCC cells, illuminating new therapeutic targets for this aggressive disease. Cancer Res; 78(7); 1619–31. ©2018 AACR.

Supplementary materials and methods about the generation of conditioned media and ELISA, quantitative real time PCR, western blot analysis and phospho-receptor tyrosine kinase array, plasmids, siRNA, and transfection, dual luciferase assay, oligo pull-down and ChIP, immunofluorescence staining. Supplementary tables S1-S10. S1: Clinicopathologic characteristics of hepatocellular carcinoma cohort (n=339). S2: List of antibodies used for the immunohistochemistry. S3: Criteria for qualitative immunohistochemistry analysis. S4: Criteria for semiquantitative immunohistochemistry analysis. S5: Primer sequences for PCR reaction. S6: List of antibodies. S7: shRNA/siRNA target sequence for gene knockdown. S8: Clinicopathologic characteristics of hepatocellular carcinoma with HGF or MET expression. S9: Univariate and multivariate analyses of disease-specific survival. S10: Univariate and multivariate analyses of disease-free survival.

Supplementary Figure S1: Overall study design for multi-omic data analysis. Supplementary Figure S2: The distribution of beta-values in DNA methylation data. Supplementary Figure S3: The distribution of differentially methylated probes (DMPs). Supplementary Figure S4: Differentially methylated probes in the integrated data of GSE44970 and YSHCC. Supplementary Figure S5: Distribution of CNAs during stepwise hepatocarcinogenesis. Supplementary Figure S6: DNA methylation of oncogenes and TSGs during multi-step hepatocarcinogenesis. Supplementary Figure S7: Frequencies of nonsynonymous or missense mutations in each patient. Supplementary Figure S8: Functionally enriched expression in each step of hepatocarcinogenesis. Supplementary Figure S9: Validation of the three genes including SPINK1, CAP2, and RRAGD. Supplementary Figure S10: SPINK1 overexpression increases proliferation and invasion of Huh7 cells. Supplementary Figure S11: Correlation between methylation and expression. Supplementary Figure S12: Western blot of DNMT1 expression. Supplementary Figure S13: GRP78 expression under ER stress. Supplementary Figure S14: Effects of knockdown of p65 expression in the expression of GRP78 and SPINK1.

Background/aims Intrahepatic cholangiocarcinoma (iCCA) with a ductal plate malformation (DPM) pattern is a recently recognized rare variant. The genomic profile of iCCA with DPM pattern needs to be elucidated. Methods Cases of iCCA with DPM pattern were retrospectively reviewed based on the medical records, pathology slides, and magnetic resonance imaging (MRI) reports collected between 2010 to 2019 at a single center. Massive parallel sequencing was performed for >500 cancer-related genes. Results From a total of 175 iCCAs, five (2.9%) cases of iCCA with DPM pattern were identified. All cases were of the small duct type, and background liver revealed chronic B viral or alcoholic hepatitis. Three iCCAs with DPM pattern harbored MRI features favoring the diagnosis of hepatocellular carcinoma, whereas nonspecific imaging features were observed in two cases. All patients were alive without recurrence during an average follow-up period of 57 months. Sequencing data revealed 64 mutated genes in the five cases, among which FGFR and PTPRT were most frequently mutated (three cases each) including an FGFR-TNC fusion in one case. Mutations in ARID1A and CDKN2A were found in two cases, and mutations in TP53, BAP1, ATM, NF1, and STK11 were observed in one case each. No IDH1, KRAS, or PBRM1 mutations were found. Conclusions iCCAs with DPM pattern have different clinico-radio-pathologic and genetic characteristics compared to conventional iCCAs. Moreover, FGFR and ARID1A variants were identified. Altogether, these findings further suggest that iCCA with DPM pattern represents a specific subtype of small duct type iCCA.

The prevalence of non-alcoholic fatty liver disease-related hepatocellular carcinoma (NAFLD-HCC) has increased parallelly with that of metabolic syndrome. This study aimed to compare the clinical and survival outcomes of NAFLD-HCC and HBV-related HCC(HBV-HCC).The medical records of patients who underwent hepatectomy for HCC at Severance Hospital between 2005 and 2015 were retrospectively reviewed. Occult HBV infection was identified by nested PCR. Propensity score matching (PSM) was conducted to minimize lead-time bias caused by the lack of surveillance in NAFLD patients. Surgical and oncologic outcomes were compared between the two groups.There were 32 patients (7%) with NAFLD-HCC, 200 (46%) with HBV-HCC, and 194 (44%) with HBV/NAFLD-HCC (HBV and NAFLD). Before PSM, cirrhosis was more frequently detected in HBV-HCC patients (55% vs 15%, p 0.001) and the average tumor size was larger in the NAFLD-HCC group than in the HBV-HCC group (4.4 ± 3.3 cm vs 3.4 ± 1.8 cm, p = 0.014). After a median follow-up of 74 months (range 0-157 months), survival analyses before PSM showed better 5-year overall survival (OS) in HBV-HCC patients than in NAFLD-HCC patients (80% vs 63%, p = 0.041). After PSM, 5-year OS rates were similar (60% vs 63%, p = 0.978). There were no differences between the groups in recurrence-free or disease-specific survival before and after PSM.Patients with NAFLD-HCC were less likely to have underlying cirrhosis but more likely to have larger tumors at the time of diagnosis than patients with HBV-HCC. The OS of patients with NAFLD-HCC appeared to be worse than that of patients with HBV-HCC. Therefore, active HCC surveillance is recommended in patients with metabolic syndrome for the early detection of HCC.

Intrahepatic cholangiocarcinoma (iCCA) is subclassified into mass-forming (MF), periductal-infiltrative (PI), and mixed types grossly; however, their clinicopathological significance remains controversial.Clinicopathological characteristics of iCCA gross types were analysed according to histopathological type (small-duct, large-duct, indeterminate) or cholangiolocellular differentiation trait (CDT) in 108 iCCAs. The expression levels of inflammation-marker (CRP, FGB) and proliferation-marker (phospho-ERK1/2, Ki-67) were evaluated by immunohistochemistry.There were 87 MF, 8 PI, and 13 mixed-gross type. Small-duct-type (39, 44.8%) and CDT (19, 21.8%) were found only in MF-gross type. The inflammation-marker expression was higher in MF-type than in PI- and mixed-gross types (P = 0.023). It was high in small-duct-type, middle in indeterminate-type, and low in large-duct-type (P = 0.015), and iCCAs with CDT showed higher inflammation-marker expression compared to those without (P 0.001). Proliferation-marker expression did not differ according to gross type; however it was lower in iCCA with CDT compared to those without (P = 0.004). Subgrouping of the gross type according to histopathological type or CDT revealed that MF-type with small-duct-type or CDT had better overall survival compared to the others (P 0.05).MF-type iCCA is more heterogeneous than other gross types. High inflammation-marker/low proliferation-marker expression in MF-type with CDT or small-duct-type may be related to a good outcome.

Background: The differential diagnosis of intrahepatic cholangiocarcinomas (iCCAs) from metastatic adenocarcinomas from organs adjacent to the liver is difficult due to histopathological similarity and a lack of specific markers. This study evaluates the performance of albumin mRNA in situ hybridization (ISH) and immunohistochemistry (IHC) of filamin-A and cytokeratin 19 (CK19) for differential diagnosis.Methods: A total of 181 iCCAs and 116 adenocarcinomas from extrahepatic organs (30 gallbladder, 28 pancreas, and 58 stomach) were collected, and albumin ISH and IHC for filamin-A and CK19 were performed. The sensitivity, specificity, and the area under the curve (AUC) on the receiver operating characteristic (ROC) curve analysis of each marker were evaluated for the diagnosis of iCCA over extrahepatic adenocarcinoma.Results: Albumin ISH showed 23.2% sensitivity and 100% specificity for iCCAs, including both the small-duct and large-duct types. The expression of filamin-A and CK19 in albumin ISH-negative iCCAs was compared with that in extrahepatic adenocarcinomas. Filamin-A showed 71.1% sensitivity, 87.5% specificity, and 0.79 AUC on the ROC in small-duct-type iCCAs, whereas it had 27.5% sensitivity, 87.5% specificity, and 0.58 AUC on the ROC in large-duct-type iCCAs. CK19 showed 87.9% sensitivity, 59.4% specificity, and 0.74 AUC on the ROC in large-duct-type iCCAs, whereas it had 44.7% sensitivity, 59.4% specificity, and 0.52 AUC on the ROC in small-duct-type iCCAs.Conclusion: Albumin ISH and IHC for filamin-A and CK19 showed distinct expression patterns according to iCCA type. Albumin ISH positivity supports the possibility of iCCA, and filamin-A IHC is helpful for differentiating small-duct-type iCCA from extrahepatic adenocarcinomas.

Primary liver cancers (LCs), including HCC and intrahepatic cholangiocarcinoma (iCCA), are derived from a common developmental lineage, conferring a molecular spectrum between them. To elucidate the molecular spectrum, we performed an integrative analysis of transcriptome profiles associated with patients' radiopathologic features.We identified four LC subtypes (LC1-LC4) from RNA-sequencing profiles, revealing intermediate subtypes between HCC and iCCA. LC1 is a typical HCC characterized by active bile acid metabolism, telomerase reverse transcriptase promoter mutations, and high uptake of gadoxetic acid in MRI. LC2 is an iCCA-like HCC characterized by expression of the progenitor cell-like trait, tumor protein p53 mutations, and rim arterial-phase hyperenhancement in MRI. LC3 is an HCC-like iCCA, mainly small duct (SD) type, associated with HCC-related etiologic factors. LC4 is further subclassified into LC4-SD and LC4-large duct iCCAs according to the pathological features, which exhibited distinct genetic variations (e.g., KRAS, isocitrate dehydrogenase 1/2 mutation, and FGF receptor 2 fusion), stromal type, and prognostic outcomes.Our integrated view of the molecular spectrum of LCs can identify subtypes associated with transcriptomic, genomic, and radiopathologic features, providing mechanistic insights into heterogeneous LC progression.

In the present work, we report a preparation strategy for hematite phase-pure photoanodes consisting of Sn-doped hematite nanoflakes/hematite thin film bilayer nanostructure (Sn-HB). This approach is based on a two-step annealing process of pure iron films deposited on fluorine doped tin oxide (FTO) substrates by advanced magnetron sputtering. While the high density hematite ultrathin nanoflakes (HNs) with detrimental iron oxide layers (Fe3O4 and/or FeO) are generated during the first annealing step at 400 degrees C for two hours, the second thermal treatment at 800 degrees C for 15 minutes oxidises all the undesired iron oxide phases to a photoactive hematite layer as well as is providing efficient Sn doping of a drop-casted SnCl4 in order to increase the conductivity. The optimized Sn-HB shows an around 11 times higher photocurrent density (0.71 mA cm(-2) at 1.23 V-RHE) compared with a reference hematite photoanode produced from iron foil under the same conditions. Web of Science 9 11 art. no. E202200066

Yeosinsan, an herbal formula composed of roots of Paeonia lactiflora and tubers of Cyperus rotundus, was reported as a possible anti-inflammatory and pro-fertility drug. However, the safety of Yeosinsan has not yet been previously investigated. The possible acute and chronic oral toxicity of Yeosinsan was estimated using female and male Sprague Dawley rats. During the administration period, rats were monitored for mortality, body weight changes, food intake, clinical signs, and gross findings. Hematological analysis, serum biochemistry analysis, urinalysis, organ weight measurement, and histopathological examination were also conducted after sacrifice. Acute and chronic oral administration of Yeosinsan did not result in any signs of toxicity in the animals during the observation period. However, in the histopathological examination, several significant changes were observed in the stomach and spleen. In the high-dose group, extramedullary hematopoiesis and increased pigmentation in the spleen and squamous cell hyperplasia in the forestomach were observed. In conclusion, the no observed adverse effect level (NOAEL) of the test material, Yeosinsan, was estimated at 1000 mg∙kg−1∙d−1 for both male and female rats. Therefore, our data suggest that Yeosinsan might be safe to use for treating female infertility.

Hepatocellular carcinoma (HCC) undergoes a stepwise progression from liver cirrhosis to low-grade dysplastic nodule (LGDN), high-grade dysplastic nodule (HGDN), early HCC (eHCC), and progressed HCC (pHCC). Here, we profiled multilayered genomic, epigenomic, and transcriptomic aberrations in the stepwise hepatocarcinogenesis. Initial DNA methylation was observed in eHCC (e.g., DKK3, SALL3, and SOX1) while more extensive methylation was observed in pHCC. In addition, eHCCs showed an initial loss of DNA copy numbers of tumor suppressor genes in the 4q and 13q regions, thereby conferring survival benefits to cancer cells. Transcriptome analysis revealed that HGDNs expressed endoplasmic reticulum (ER) stress–related genes, while eHCC started to express oncogenes. Furthermore, integrative analysis indicated that expression of the serine peptidase inhibitor, Kazal type 1 (SPINK1), played a pivotal role in eHCC development. Significant demethylation of SPINK1 was observed in eHCC compared to HGDN. The study also demonstrated that ER stress may induce SPINK1 demethylation and expression in liver cancer cells. In conclusion, these results reveal the dynamics of multiomic aberrations during malignant conversion of liver cancer, thus providing novel pathobiological insights into hepatocarcinogenesis. Significance: Multiomics profiling and integrative analyses of stepwise hepatocarcinogenesis reveal novel mechanistic and clinical insights into hepatocarcinogenesis.

Patients with Fontan circulation exhibit a high incidence of liver fibrosis and cirrhosis. Transient elastography (TE) and the enhanced liver fibrosis (ELF) test have proven useful as noninvasive surrogate markers of liver fibrosis for other chronic liver diseases. We evaluated whether TE and the ELF score can predict the degree of liver fibrosis in patients with Fontan circulation.We retrospectively reviewed the medical records of 45 adult patients with at least 10 years of Fontan duration who had undergone liver biopsy and investigated the relation between the fibrosis stage and TE and the ELF test results. Additionally, the association of these variables and other biochemical and hemodynamic parameters was assessed.The mean age was 25.9 years and the mean Fontan duration was 20.8 years. Advanced liver fibrosis was present in 36 (80.0%) patients. TE or ELF score are comparable for patients with and without advanced liver fibrosis (mean 23.3 vs 24.8 kPa [P = .85] for TE; mean 8.94 vs 9.25 [P = .44] for the ELF score). However, N-terminal pro-brain natriuretic peptide level and ventricular end-diastolic pressure were higher in patients with advanced liver fibrosis (mean 224 vs 80 pg/mL [P .01]; and mean 12 vs 9 mm Hg [P = .04], respectively). No independent predictor of advanced liver fibrosis was found in multivariate analysis.TE and the ELF score were unable to predict the degree of liver fibrosis in Fontan patients. Liver biopsy remains as the only valid method to assess fibrotic burden in this population.

Background Danggwijagyaksan (DJS) has been one of the most widely used herbal medicines for gynecological disorders in traditional East Asian medicine. Several clinical studies about DJS have shown improvement in menopausal symptoms. This pilot study aimed to evaluate the efficacy, safety and feasibility of DJS for treating climacteric syndrome with a blood-deficiency-dominant pattern. Methods This was a randomized, double-blind, placebo-controlled pilot trial. A group of 45-to 60-year-old women with climacteric syndrome were registered for the trial. The participants received treatment over a 4-week period and were then followed for 4 weeks. The primary outcome measure was the mean change in the Menopause Rating Scale (MRS). Secondary outcome measures included the World Health Organization Quality of Life-BREF (WHOQOL-BREF), the Blood-Deficiency Scoring System (BDSS), lean body mass, and serum hormone levels, including follicle-stimulating hormone (FSH) and estradiol (E2) levels. Results The MRS and BDSS scores decreased significantly in both groups, but the differences between two groups were not significant. The WHOQOL-BREF scores increased in the control group. No statistically meaningful differences in serum hormone levels or lean body mass were observed in both groups. There were no serious adverse events, and the laboratory tests were within the normal range. The recruitment rate, completion rate and medication adherence rate were over 90% in both groups, indicating high feasibility. Conclusions DJS showed clinical effectiveness in the treatment of climacteric syndrome with a blood-deficiency-dominant pattern. Additionally, DJS was shown to be safe and feasible for a large-scale study to confirm the efficacy of the treatment. Trial registration: Clinical Research Information Service (CRIS, https://cris.nih.go.kr ): KCT0002387.

BACKGROUND & AIMS Extrahepatic metastasis from hepatocellular carcinoma (HCC) is a catastrophic event, yet organ-specific pathological characteristics of metastatic HCC remain unclear. We aimed to characterize the pathological aspects of HCC metastases to various organs. METHODS We collected intrahepatic HCC (cohort 1, n = 322) and extrahepatic metastatic HCC (cohort 2, n = 130) samples. Clinicopathological evaluation and immunostaining for K19, CD34, αSMA, fibroblast-associated protein (FAP), CAIX, VEGF, PD-L1, CD3, CD8, Foxp3, CD163 and epithelial-mesenchymal transition (EMT)-related markers were performed. RESULTS Independent factors for extrahepatic metastasis included BCLC stage B-C, microvascular invasion (MVI), vessels encapsulating tumour clusters (VETC)-HCC, K19 and FAP expression, and CD163+ macrophage infiltration (cohort 1, P

BACKGROUND Development of a pragmatic pathologic classifier of pancreatic ductal adenocarcinoma (PDAC) that reflects biological behavior is needed. METHODS The tumor epithelial and stromal features of PDAC and molecular subtype-related markers were evaluated in three independent cohorts. RESULTS In the non-neoadjuvant therapy cohort (n = 108), regarding tumor-epithelial feature, non-gland-forming type showed worse prognosis compared to gland-forming type (P

Background/aims Circulating tumor cells (CTCs) with cancer stemness have been demonstrated to be a direct cause of tumor recurrence, and only few studies have reported the role of CTCs in liver transplantation (LT) for hepatocellular carcinoma (HCC). Methods Epithelial cell adhesion molecule+ (EpCAM+), cluster of differentiation 90+ (CD90+) and EpCAM+/CD90+ CTCs were sorted via fluorescence-activated cell sorting, and transcripts level of EpCAM, K19 and CD90 in the peripheral blood were analyzed via real-time polymerase chain reaction preoperatively and on postoperative days 1 and 7 in 25 patients who underwent living donor liver transplantation (LDLT) for HCC. EpCAM protein was assessed in HCC tissue using immunohistochemical staining. The median follow-up duration was 40 months. Results HCC after LDLT recurred in four out of 25 patients. Detection of EpCAM+ or CD90+ CTCs correlated well with their messenger RNA levels (p 100 mAU/mL and postoperative day 1 EpCAM+/CD90+ CTCs were independent risk factors for HCC recurrence (hazard ratio, 14.64; 95% confidence interval, 1.08 to 198.20; p=0.043 and hazard ratio, 26.88; 95% confidence interval, 1.86 to 387.51; p=0.016, respectively). Conclusions EpCAM+/CD90+ CTCs can be used preoperatively and 1 day after LDLT as key biological markers in LT candidate selection and post-LDLT management.

BACKGROUND & AIMS: Late recurrence of hepatocellular carcinoma (HCC) is regarded as de novo HCC from chronic hepatitis. This study investigated clinicopathological and molecular factors to develop a nomogram for predicting late HCC recurrence (> 2 years after curative resection). METHODS: The training and validation cohorts included HCC patients with a major etiology of hepatitis B who underwent curative resection. Clinicopathological features including lobular and porto-periportal inflammatory activity, fibrosis, and liver cell change were evaluated. Proteins encoded by genes related to late recurrence were identified using a reverse phase protein array of 95 non-tumorous liver tissues. Immunoexpression of phosphorylated signal transducer and activator of transcription 3 (pSTAT3), plasminogen activator inhibitor-1, phosphorylated extracellular signal-regulated kinase 1/2 (pERK1/2), and spleen tyrosine kinase (SYK) was measured. RESULTS: Late recurrence occurred in 74/402 (18%) and 47/243 (19%) in the training and validation cohorts, respectively. Cirrhosis, moderate/severe lobular inflammatory activity, and expression of pSTAT3, pERK1/2, and SYK proteins correlated to the gene signature of hepatocyte injury and regeneration were independently associated with late recurrence, with odds ratios (95% confidence intervals) of 2.0 (1.2–3.3), 21.1 (4.3–102.7), and 6.0 (2.1–17.7), respectively (p < 0.05 for all). A nomogram based on these variables (histological parameters and immunohistochemical marker combinations) showed high reliability in both the training and validation cohorts (Harrell’s C index: 0.701 and 0.716; 95% confidence intervals: 0.64–0.76 and 0.64–0.79, respectively). CONCLUSIONS: The combination of pSTAT3, pERK1/2, and SYK immunoexpression with high lobular inflammatory activity and cirrhosis (fibrosis) predicts late HCC recurrence.

The expression of estrogen receptor alpha (ERα, encoded by ESR1) has been shown to be associated with the prognostic outcomes of patients in various cancers; however, its prognostic and mechanistic significance in hepatocellular carcinoma (HCC) remain unclear. Here, we evaluated the expression of ERα and its association with clinicopathological features in 339 HCC patients. ERα was expressed in 9.4% (32/339) of HCCs and was related to better overall survival (OS; hazard ratio [HR] = 0.11, p = 0.009, 95% C.I. = 0.016–0.82) and disease-free survival (DFS, HR = 0.4, p = 0.013, 95% C.I. = 0.18–0.85). ERα expression was also associated with features related to more favorable prognosis, such as older age, lower serum alpha-fetoprotein level, and less microvascular invasion (p, Liver cancer: How estrogen receptors hinder tumor growth Estrogen receptor signaling can act as a brake preventing the progression of an often deadly form of liver cancer. Studies have shown that women are at a lower risk of developing and succumbing to hepatocellular carcinoma (HCC) than men, suggesting a potential role for sex hormones. Researchers in South Korea led by Hyun Goo Woo of the Ajou University School of Medicine, Suwon, and Young Nyun Park of Yonsei University College of Medicine, Seoul, have now shown that expression of the estrogen receptor α is a strong prognostic predictor for HCC. In a survey of patient tumor samples, they found that expression of this hormone receptor is associated with nearly a tenfold increased likelihood of survival. The researchers identified a mechanism by which estrogen receptor α signaling impedes cancerous growth, revealing potential new drug targets.

Background/Aims: Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) activation has been implicated in hepatocarcinogenesis and hepatic progenitor cell differentiation, and hypoxia has been shown to induce nuclear translocation of YAP in cancer cells. Here, we aimed to investigate the relationship between hypoxia, YAP and TAZ expression and stemness-related marker expression in human hepatocellular carcinomas (HCCs) and its clinical implications.Methods: Immunohistochemical stains were performed on tissue microarrays from 305 surgically resected HCCs, and the expression status of YAP and TAZ were correlated with CAIX, stemness markers (K19, EpCAM) and epithelial-mesenchymal transition (EMT)-related markers (uPAR, ezrin). The clinicopathological significance of YAP/TAZ expression was analyzed with relation to CAIX expression status.Results: YAP and TAZ expression were seen in 13.4 and 4.3% of HCCs, respectively. YAP/TAZ-positive HCCs frequently demonstrated higher serum AFP levels, microvascular invasion, advanced tumor stage, increased proliferative activity and expression of stemness- and EMT-related markers, CAIX, p53 and Smad2/3 (p < 0.05, all). Interestingly, YAP/TAZ-positivity was associated with microvascular invasion, higher serum AFP levels, stemness and EMT-related marker expression only in tumors expressing CAIX (p < 0.05, all), while these associations were not seen in CAIX-negative HCCs.Conclusions: YAP/TAZ expression is associated with vascular invasion, stemness and EMT in HCCs with hypoxia marker expression. The effect of Hippo signaling pathway deregulation in HCC may depend on the presence or absence of a hypoxic microenvironment, and hypoxia marker expression status should be taken into account when considering the use of YAP/TAZ as markers of aggressive biologic behavior in HCC.