Possible pathogenetic role of ammonia in liver cirrhosis without hyperammonemia of venous blood: The so-called latency period of abnormal ammonia metabolism.

Ammonia plays a crucial role in the pathogenesis of hepatic encephalopathy. Ammonia is also involved in many other pathological conditions seen in cirrhosis, such as sarcopenia, liver fibrosis, hepatocellular injury, immune dysfunction, and hyperammonemia. Furthermore, the ammonia level of the veins is a useful prognostic factor for cirrhosis. In cirrhosis without hyperammonemia of the vein, however, covert hepatic encephalopathy has been reported. This discrepancy is because of the anatomical features of ammonia metabolism. There are two systems in the body for detoxifying ammonia: one is the urea cycle in the liver, and the other is the glutamine synthesis pathway in skeletal muscle and other tissues. The blood processed in the liver's urea cycle is then transported via arteries to various organs. Further processing occurs in the brain and skeletal muscle's glutamine synthesis pathway before entering the veins. When the urea cycle function decreases in cirrhosis, the ammonia levels in the artery increase. In response, the glutamine synthesis pathway compensates by increasing the capacity to process ammonia. Therefore, the ammonia concentration in the veins downstream of skeletal muscles does not increase immediately. However, the brain and skeletal muscles, which receive arterial blood, might be exposed to high ammonia concentrations. In addition, branched-chain amino acids in venous blood decrease. This period is the transition phase from early- to late-phase cirrhosis, and understanding the pathophysiology during this stage is extremely important for preventing the progression of cirrhosis.

Kaposi's sarcoma-associated herpesvirus replication and transcription activator protein activates CD274/PD-L1 gene promoter.

Kaposi's sarcoma-associated herpesvirus (KSHV) is responsible for the pathogenesis of Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman disease. The expression of immunosuppressive genes, such as IL-10 and CD274/PD-L1 is observed during KSHV-associated pathogenesis, and the modulation of the host immune system by KSHV contributes to establishing viral persistence in the host. Understanding the mechanism that allows the virus to evade host cell immunity would be helpful in order to develop therapeutic strategies for KSHV malignancy. In this study, we show that KSHV replication and transcriptional activator (K-RTA), an essential activator of the viral lytic cycle, transactivates the CD274/PD-L1 gene promoter. Mechanistically, we demonstrate that the binding of K-RTA to the cellular specificity protein 1 (SP1) is critical for K-RTA-mediated CD274/PD-L1 promoter activation. These findings suggest that K-RTA cooperates with intracellular SP1 to activate the expression of CD274/PD-L1, which helps the virus regulate immune checkpoints to escape and survive.

Clinical significance of the latency period of abnormal ammonia metabolism in chronic liver disease: Proposal of a new concept.

The liver is a key organ in regulating metabolism, and chronic liver disease is associated with several metabolic disorders. In the later stages of liver cirrhosis, the urea cycle is impaired, which disrupts of ammonia detoxification and eventually causes hyperammonemia and hepatic encephalopathy. Although hyperammonemia is not detected during the period between the late stage of chronic hepatitis and the early stage of liver cirrhosis, hepatic albumin synthesis capacity decreases as the fibrosis progresses. Increased ammonia levels are associated with a decreased capacity of the liver to synthesize albumin as well as activation of hepatic stellate cells, which promote fibrosis. Herein, we discuss the possibility that abnormal ammonia metabolism might play an important role in the pathogenesis of liver diseases even without hyperammonemia. We consider the disease period without hyperammonemia as the latency period of abnormal ammonia metabolism and discuss its clinical significance.

RSK Inhibition Induces Apoptosis by Downregulating Protein Synthesis in a Variety of Acute Myeloid Leukemia Cell Lines.

Fms-like tyrosine kinase 3 (FLT3) and isocitrate dehydrogenase 1/2 (IDH1/2) mutations drive malignancy in acute myeloid leukemia (AML), which accounts for approximately 40% of AML cases. Treatment with FLT3 or IDH1/2 inhibitors is used for such patients; however, it is not considered for most patients with AML who lack mutations on the respective genes. In this study, p90 ribosomal S6 kinase (RSK) was found to serve as a new therapeutic target in various AMLs with or without FLT3 mutations. BI-D1870, a potent inhibitor of RSK, significantly suppressed the proliferation of AML cell lines, among which three encoded wild-type FLT3 and three contained FLT3 driver mutations, compared with chronic myeloid leukemia K562 cells or other adherent cancer cells. BI-D1870 inhibited protein synthesis by dephosphorylating the p70 S6 kinase and eukaryotic initiation factor 4E-binding protein 1 in all AML cells except KG-1a cells. Meanwhile, the expression of microtubule-associated protein light chain 3B-I and -II increased in KG-1a cells treated with BI-D1870. BI-D1870 induced caspase-dependent apoptosis in all AML cells, including KG-1a cells. We next investigated the synergistic effect of BI-D1870 with cytarabine, a traditional anticancer drug used in AML. Synergistic effects of BI-D1870 and cytarabine were not observed in any of the cell lines. The findings suggested that BI-D1870 alone exerts an adequate antiproliferative effect on AML with or without FLT3 mutations and serves as a novel AML therapeutic agent.

SNAIL- and SLUG-induced side population phenotype of HCT116 human colorectal cancer cells and its regulation by BET inhibitors.

Epithelial-mesenchymal transition (EMT) is associated with cancer malignancies such as invasion, metastasis, and drug resistance. In this study, HCT116 human colorectal cancer cells were transduced with SLUG or SNAIL retroviruses, and EMT cells with mesenchymal morphology were established. The EMT cells showed a high invasive activity and resistance to several anticancer agents such as methotrexate, SN-38, and cisplatin. Furthermore, they contained about 1-10% side population (SP) cells that were not stained by Hoechst 33342. This SP phenotype was not stable; the isolated SP cells generated both SP and non-SP cells, suggesting a potential for differentiation. Gene expression analysis of SP cells suggested the alteration of genes that are involved in epigenetic changes. Therefore, we examined the effect of 74 epigenetic inhibitors, and found that two inhibitors, namely I-BET151 and bromosporine, targeting the bromodomain and extra-terminal motif (BET) proteins, decreased the ratio of SP cells to <50% compared with the control, without affecting the immediate efflux of Hoechst 33342 by transporters. In addition, compared with the parental cells, the EMT cells showed a higher sensitivity to I-BET151 and bromosporine. This study suggests that EMT development and SP phenotype can be independent events but both are regulated by BET inhibitors in SLUG- or SNAIL-transducted HCT116 cells.

Japan's Practical Guidelines for Zinc Deficiency with a Particular Focus on Taste Disorders, Inflammatory Bowel Disease, and Liver Cirrhosis.

Zinc deficiency is common in Japan, yet awareness on this disorder is lacking. The Japanese Society of Clinical Nutrition recently issued the Japan's Practical Guideline for Zinc Deficiency 2018 setting forth criteria for diagnosing zinc deficiency, i.e., (a) one or more symptoms of zinc deficiency or low serum alkaline phosphatase, (b) ruling out other diseases, (c) low serum zinc, and (d) alleviation of symptoms upon zinc administration. Serum zinc <60 µg/dL and 60-80 µg/dL indicate zinc deficiency and marginal deficiency, respectively. Zinc deficiency symptoms vary and include dermatitis and taste disorders among others. Zinc administration improves taste in 50-82% of patients suffering from taste disorders (a common symptom of zinc deficiency). Effects of zinc administration do not appear immediately, and therapy should be continued for at least three months. Zinc deficiency often accompanies various diseases and conditions. Here, we focus on inflammatory bowel diseases and liver cirrhosis. As zinc deficiency enhances intestinal inflammation via macrophage activation, we discuss the pathological mechanism for inflammation and zinc deficiency in the context of IBD. Zinc deficiency can also lead to a nitrogen metabolic disorder in patients with liver cirrhosis. Zinc supplementation can improve not only the ammonia metabolism, but also the protein metabolism. We also discuss directions for future studies of zinc deficiency.

IL-10 promoter transactivation by the viral K-RTA protein involves the host-cell transcription factors, specificity proteins 1 and 3.

Kaposi's sarcoma-associated herpesvirus (KSHV)/human herpesvirus-8 (HHV-8) causes a persistent infection, presenting latent and lytic replication phases during its life cycle. KSHV-related diseases are associated with deregulated expression of inflammatory cytokines, including IL-6 and IL-10, but the mechanisms underlying this dysregulation are unclear. Herein, we report a molecular mechanism for KSHV-induced IL-10 gene expression. KSHV replication and transcription activator (K-RTA) is a molecular switch for the initiation of expression of viral lytic genes, and we describe, for the first time, that K-RTA significantly activates the promoter of the human IL-10 gene. Of note, mutations involving a basic region of K-RTA reduced the association of K-RTA with the IL-10 promoter. Moreover, the host-cell transcription factors, specificity proteins (SP) 1 and 3, play a pivotal cooperative role in K-RTA-mediated transactivation of the IL-10 promoter. K-RTA can interact with SP1 and SP3 directly in vitro, and electrophoresis mobility shift assays (EMSAs) revealed co-operative interaction involving K-RTA, SP1, and SP3 in binding to the IL-10 promoter. As DNase I footprinting assays indicated that K-RTA did not affect SP3 binding to the IL-10 promoter, SP3 can function to recruit K-RTA to the IL-10 promoter. These findings indicate that K-RTA can directly contribute to IL-10 up-regulation via a functional interplay with the cellular transcription factors SP1 and SP3.

Novel pharmacological effects of poly (ADP-ribose) polymerase inhibitor rucaparib on the lactate dehydrogenase pathway.

Poly (ADP-ribose) polymerases (PARPs) are involved in various cellular events, including DNA repair. PARP inhibitors including olaparib and rucaparib, have been specially developed against breast and ovarian cancers deficient in DNA repair systems. In this study, we found that PARP1-defective olaparib-resistant A2780 cells (ola-R cells) cells were still sensitive to two PARP inhibitors, rucaparib and veliparib. Metabolomic analysis revealed that rucaparib suppressed the lactate dehydrogenase (LDH)-mediated conversion of pyruvic acid to lactic acid in A2780 cells, although olaparib did not. The inhibition of LDH by siRNA-mediated knockdown or by LDH inhibitors suppressed the growth of ovarian cancer cells. Our results suggested that the suppression of the LDH-associated pathway contributed to the pharmacological effects of rucaparib.

Synthesis of 5-Hydroxy-3',4',7-trimethoxyflavone and Related Compounds and Elucidation of Their Reversal Effects on BCRP/ABCG2-Mediated Anticancer Drug Resistance.

3',4',7-Trimethoxyflavone (TMF) has been reported to show a potent reversal effect on drug resistance mediated by breast cancer resistance protein (BCRP)/ATP-binding cassette subfamily G member 2 (ABCG2). In this study, we designed and synthesized five derivatives with either a hydroxy group or a fluorine atom at C-5 and several kinds of capping moiety at the C-7 hydroxy group, on the same 3',4'-dimethoxy-substituted flavone skeleton. We subsequently evaluated the efficacies of these compounds against BCRP-expressing human leukaemia K562/BCRP cells. Reversal of drug resistance was expressed as the concentration of compound causing a twofold reduction in drug sensitivity (RI50 ). Of the synthesized compounds, the reversal effect of 5-hydroxy-3',4',7-trimethoxyflavone (HTMF, RI50 7.2 nm) towards 7-ethyl-10-hydroxycamptothecin (SN-38) was stronger than that of TMF (RI50 18 nm). Fluoro-substituted 5-fluoro-3',4',7-trimethoxyflavone (FTMF, RI50 25 nm) and monoglycosylated 7-(ß-glucosyloxy)-5-hydroxy-3',4'-dimethoxyflavone (GOHDMF, 91 nm) also exhibited reversal effects, whereas the di- and triglycoside derivatives did not. TMF, HTMF and FTMF at 0.01-10 µm upregulated the K562/BCRP cellular accumulation of Hoechst 33342 nuclear staining dye. In addition, western blotting revealed that treatment of K562/BCRP cells with 0.1 µm TMF, HTMF or FTMT suppressed the expression of BCRP. HTMF showed the strongest inhibition of BCRP-mediated efflux and suppression of BCRP expression of the three effective synthesized flavones.

Hypozincemia is associated with human hepatocarcinogenesis in hepatitis C virus-related liver cirrhosis.

AIM: Hypozincemia is associated with the progression of chronic liver diseases, but it is unknown whether hypozincemia promotes human hepatocarcinogenesis. Our aim is to evaluate the serum zinc levels in liver cirrhosis (LC) patients and clarify the relationship between the serum zinc levels and the development of hepatocellular carcinoma (HCC). METHODS: Cirrhotic patients without HCC (n = 299) were enrolled from 14 medical institutes in Japan as a multicenter prospective study (No. 2028). Of the 299 patients, 157 were included in the present study based on reliable and consistent serum zinc levels and no history of oral zinc supplementation. Clinical parameters associated with the development of HCC were determined. Furthermore, the cumulative incidence of HCC was analyzed using Kaplan-Meier methods and was calculated using the log-rank test. A Cox regression analysis was utilized for the multivariate analysis to evaluate the predictors of hepatocarcinogenesis. RESULTS: Thirty of 157 patients (19.1%) developed HCC during an observation period of 3 years. Serum zinc levels were significantly decreased in hepatitis C virus-related LC (C-LC) patients with HCC (0.0180). The risk factors for incidence of HCC were hypozincemia (0.0014), high α-fetoprotein (0.0080), low branched chain amino acids-to-tyrosine ratio (0.0128), or female sex (0.0228). Hypozincemia (hazard ratio 1.61, 0.0324) was the only significant predictor of hepatocarcinogenesis by multivariate Cox regression analysis. CONCLUSIONS: Hypozincemia is associated with hepatocarcinogenesis in C-LC patients.

Incidence and risk factors of hepatocellular carcinoma change over time in patients with hepatitis C virus infection who achieved sustained virologic response.

AIM: In patients with chronic hepatitis C, hepatocellular carcinoma (HCC) occurs at a certain frequency, even if a sustained virologic response (SVR) is achieved by antiviral treatment. Old age, liver fibrosis, and high post-treatment α-fetoprotein (AFP) level are typical risk factors of post-SVR HCC. We examined whether the frequencies and factors of HCC in patients with an SVR achieved from interferon treatment changed. Methods Among patients prospectively registered for pegylated interferon and ribavirin treatment, 2021 with an SVR without HCC development during the treatment period were followed up. The mean observation period was 49.5 ± 26.2 months. RESULTS: The multivariable Cox regression analysis showed that older age, diabetes mellitus, advanced liver disease, and higher post-treatment AFP level were the independent risk factors throughout the observation period. The annual occurrence rate of HCC was 0.74% in the third year, 0.54% in the fourth year, and 0.40% in the fifth year; it gradually decreased from the third year. Because the time course hazards for HCC changed at 48 months, we separately analyzed its risk factors before and after this change point. The multivariable Cox regression analysis showed that the four above-mentioned factors were significantly related to HCC development within 4 years. Conversely, the univariable Cox regression analysis only identified diabetes mellitus as a significant factor for HCC development after 4 years. CONCLUSION: The frequency of HCC in hepatitis C patients who achieved an SVR from interferon treatment decreased during the observation period, and its risk factors changed between the early and late periods.

Functional Effects of AKT3 on Aurora Kinase Inhibitor-induced Aneuploidy.

The suppression of mitotic Aurora kinases (AURKs) by AURK inhibitors frequently causes cytokinetic failure, leading to polyploidy or aneuploidy, indicating the critical role of AURK-mediated phosphorylation during cytokinesis. We demonstrate the deregulated expression of AKT3 in Aurora kinase inhibitor (AURKi)-resistant cells, which we established from human colorectal cancer HCT 116 cells. The AKT family, which includes AKT1, -2, and -3, plays multiple roles in antiapoptotic functions and drug resistance and is involved in cell growth and survival pathways. We found that an AKT inhibitor, AZD5363, showed synergistic effect with an AURKi, VX-680, on two AKT3-expressing AURKi-resistant cell lines, and AKT3 knockdown sensitized cells to VX-680. Consistent with these activities, AKT3 expression suppressed AURKi-induced apoptosis and conferred resistance to AURKi. Thus, AKT3 expression affects cell sensitivity to AURKi. Moreover, we found that AKT3 expression suppressed AURKi-induced aneuploidy, and inversely AKT3 knockdown enhanced it. In addition, partial co-localization of AKT3 with AURKB was observed during anaphase. Overall, this study suggests that AKT3 could repress the antiproliferative effects of AURKi, with a novel activity particularly suppressing the aneuploidy induction.

Protein kinase C alpha-mediated phosphorylation of PIM-1L promotes the survival and proliferation of acute myeloid leukemia cells.

FMS-like tyrosine kinase 3 (FLT3)-internal tandem duplication (ITD) is a constitutively active mutant of FLT3 and causes 20%-30% of acute myeloid leukemia (AML) cases. FLT3-ITD upregulates the proviral integration site for Moloney murine leukemia virus 1 (PIM-1) expression and promotes the proliferation of AML cells. In this study, we investigated the role of protein kinase C (PKC)-mediated phosphorylation on the expression and function of PIM-1L. Drug screening in leukemia cell lines revealed that sotrastaurin (a PKC inhibitor) suppressed the proliferation of the FLT3-ITD-positive AML cell line MV4-11 but not of K562, HL60, or KG-1a cells, similar to SGI-1776 (a PIM-1/FLT3 inhibitor) and quizartinib (an FLT3 inhibitor). Sotrastaurin decreased the expression of pro-survival protein myeloid cell leukemia (MCL-1) and the phosphorylation of eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), both of which are downstream effectors of PIM-1. PKCα directly phosphorylated Ser65 of PIM-1L, which is a long isoform of PIM-1. The PKCα-mediated phosphorylation stabilized PIM-1L. The phosphorylation-mimicked mutant, PIM-1L-S65D, was more stable and showed higher kinase activity than PIM-1L-S65A. Expression of PIM-1L-wildtype or -S65D reduced sotrastaurin-mediated apoptosis and growth inhibition in MV4-11 transfectants. These results suggest that PKCα directly upregulates PIM-1L, resulting in promotion of the survival and proliferation of AML cells.

Phase I/II study of first-line combination therapy with sorafenib plus resminostat, an oral HDAC inhibitor, versus sorafenib monotherapy for advanced hepatocellular carcinoma in east Asian patients.

PURPOSE: Resminostat is an oral inhibitor of class I, IIB, and IV histone deacetylases. This phase I/II study compared the safety and efficacy of resminostat plus sorafenib versus sorafenib monotherapy as first-line therapy for advanced hepatocellular carcinoma (HCC). EXPERIMENTAL DESIGN: In phase I, resminostat (400 mg or 600 mg/day on days 1 to 5 every 14 days) was administered with sorafenib (800 mg/day for 14 days) to determine the recommended dose for phase II. In phase II, patients were randomized (1:1) to sorafenib monotherapy or resminostat plus sorafenib. The primary endpoint was time-to-progression (TTP). RESULTS: Nine patients (3: 400 mg, 6: 600 mg) were enrolled in phase I, and the recommended dose of resminostat was determined to be 400 mg/day. Then 170 patients were enrolled in phase II. Median TTP/overall survival (OS) were 2.8/14.1 months with monotherapy versus 2.8/11.8 months with combination therapy (Hazard Ratio [HR]: 0.984, p = 0.925/HR: 1.046, p = 0.824). The overall incidence of adverse events was similar in both groups (98.8% versus 100.0%). However, thrombocytopenia ≥ Grade 3 was significantly more frequent in the combination therapy group (34.5% versus 2.4%, p < 0.001). Subgroup analysis revealed that median TTP/OS was 1.5/6.9 months for monotherapy versus 2.8/13.1 months for combination therapy (HR: 0.795, p = 0.392/HR: 0.567, p = 0.065) among patients with a normal-to-high baseline platelet count (≥ 150 × 103/mm3). CONCLUSIONS: In patients with advanced HCC, first-line therapy with resminostat at the recommended dose plus sorafenib showed no significant efficacy advantage over sorafenib monotherapy.

Ultra-deep sequencing analysis of resistance-associated variants during retreatment with simeprevir-based triple therapy after failure of telaprevir-based triple therapy in patients with genotype 1 hepatitis C virus infection.

AIM: Simeprevir (SMV)-based triple therapy is an effective retreatment option following failure of telaprevir (TVR)-based triple therapy. However, it is unclear whether the persistence of resistance-associated variants (RAVs) induced by TVR-based therapy may reduce the treatment effect of SMV-based therapy. METHODS: The factors associated with the treatment effect, including RAVs in the NS3 region, were examined in 21 patients with genotype 1b HCV infection who were treated with SMV-based therapy after failure of TVR-based therapy. Ultra-deep sequencing was carried out to detect RAVs. RESULTS: With the exception of one patient who discontinued treatment owing to adverse events, the sustained virologic response (SVR) rate was 50% (10/20). Ultra-deep sequencing at the start of SMV-based therapy revealed that TVR-resistant variants were detected in six patients (29%), and no variants were observed at position 168. Cross-resistance between TVR and SMV with low frequency was detected in only one patient, and this patient achieved SVR. Higher SVRs for SMV-based therapy were attained in patients who discontinued treatment owing to the adverse effects of prior TVR-based therapy (discontinuation 100% vs. non-discontinuation 29%, P = 0.005), and patients who relapsed following prior pegylated interferon plus ribavirin therapy (relapse 100% vs. non-response 20%, P = 0.007). CONCLUSIONS: In this study, ultra-deep sequencing analysis revealed that TVR and/or SMV-resistant variants may have no influence on the effect of SMV-based therapy after failure of TVR-based therapy. Patients who discontinued treatment owing to adverse effects of TVR-based therapy and relapsers to previous pegylated interferon/ribavirin therapy would be good candidates for retreatment with SMV-based therapy.

Revealing the fate of cell surface human P-glycoprotein (ABCB1): The lysosomal degradation pathway.

P-glycoprotein (P-gp) transports a variety of chemically dissimilar amphipathic compounds including anticancer drugs. Although mechanisms of P-gp drug transport are widely studied, the pathways involving its internalization are poorly understood. The present study is aimed at elucidating the pathways involved in degradation of cell surface P-gp. The fate of P-gp at the cell surface was determined by biotinylating cell surface proteins followed by flow cytometry and Western blotting. Our data shows that the half-life of endogenously expressed P-gp is 26.7±1.1 h in human colorectal cancer HCT-15 cells. Treatment of cells with Bafilomycin A1 (BafA1) a vacuolar H+ ATPase inhibitor increased the half-life of P-gp at the cell surface to 36.1±0.5 h. Interestingly, treatment with the proteasomal inhibitors MG132, MG115 or lactacystin alone did not alter the half-life of the protein. When cells were treated with both lysosomal and proteasomal inhibitors (BafA1 and MG132), the half-life was further prolonged to 39-50 h. Functional assays done with rhodamine 123 or calcein-AM, fluorescent substrates of P-gp, indicated that the transport function of P-gp was not affected by either biotinylation or treatment with BafA1 or proteasomal inhibitors. Immunofluorescence studies done with the antibody against lysosomal marker LAMP1 and the P-gp-specific antibody UIC2 in permeabilized cells indicated that intracellular P-gp is primarily localized in the lysosomal compartment. Our results suggest that the lysosomal degradation system could be targeted to increase the sensitivity of P-gp- expressing cancer cells towards chemotherapeutic drugs.

Effect of AKT3 expression on MYC- and caspase-8-dependent apoptosis caused by polo-like kinase inhibitors in HCT 116 cells.

Polo-like kinase (PLK) is a cell-cycle regulator that is overexpressed in several cancer cell types. Polo-like kinase is considered a novel target for cancer therapies, and several PLK inhibitors (PLKis), including BI 2536, BI 6727, and GSK461364, have been developed. In this study, we established five BI 2536-resistant cell lines from human colorectal cancer HCT 116 cells, to explore the resistance mechanism and identify predictable biomarkers of PLKis. We showed that PLKi-induced caspase-8 activation was attenuated in the BI 2536-resistant cell lines. We also showed that the expression of P-glycoprotein (P-GP) and AKT3 was upregulated, whereas that of MYC was downregulated in some BI 2536-resistant cell lines. Expression of P-GP conferred resistance to PLKis, and PLKi-induced apoptosis was dependent on MYC and caspase-8 in HCT 116 cells. We also showed for the first time that AKT3 suppressed BI 6727-induced caspase-8 activation and conferred resistance to PLKis. Collectively, these results indicate that MYC, caspase-8, P-GP, and AKT3 play critical roles in PLKi-induced apoptosis. Therefore, they are candidate biomarkers of the pharmacological efficacy of PLKis.

Impact of ribavirin dosage in chronic hepatitis C patients treated with simeprevir, pegylated interferon plus ribavirin combination therapy.

The factors associated with sustained virologic response (SVR) in chronic hepatitis C (CH-C) genotype 1 patients treated with simeprevir (SMV), pegylated interferon (Peg-IFN) plus ribavirin (RBV) triple therapy have not been fully investigated. Two hundred and twenty-nine treatment-naïve CH-C patients treated with SMV triple therapy were enrolled in this study. The overall SVR rate was 87% in per-protocol analysis. In multivariate analysis, the interleukin (IL) 28B genotype (rs8099917, TT vs. non-TT, odds ratio [OR]: 0.044, P = 0.001) and RBV dose (< 10/10-12/ ≥ 12 mg/kg/day, OR: 4.513, P = 0.041) were significant factors associated with SVR. In patients with the IL28B non-TT genotype, RBV dose affected SVR dose-dependently in stratified analysis of RBV dose (P = 0.015); it was 44% (8/18) for patients administered <10 mg/kg/day of RBV, 78% (14/18) for those administered 10-12 mg/kg/day of RBV, and 100% (3/3) for those administered ≥12 mg/kg/day of RBV, whereas in patients with the IL28B TT genotype, a significant correlation between SVR and RBV dose was not observed (P = 0.229). Regarding RBV dose reduction of less than 10 mg/kg/day, the inosine triphosphate pyrophosphatase (ITPA) genotype (rs1127354, CC vs. non-CC, OR: 0.239, P = 0.003) and age (by 1 y.o., OR: 1.084, P = 0.002) were significant independent factors. RBV dosage affected SVR dose-dependently in patients with the IL28B non-TT genotype treated with SMV triple therapy. Special attention to anemia progression and RBV dosage should be paid to aged patients with the ITPA CC genotype. J. Med. Virol. 88:1776-1784, 2016. © 2016 Wiley Periodicals, Inc.

Quick and stable parallel puncture of hepatic tumors using a double-barreled needle direction system for ultrasound-guided bipolar radiofrequency ablation.

AIM: In bipolar radiofrequency ablation (RFA) therapy, insertion of multiple needles at appropriate points on a target is difficult. The aim of our study was to evaluate a simplified method for multi-electrode insertion using a newly developed double-barreled needle guidance system for percutaneous RFA of hepatic tumors. METHODS: RFA using two bipolar electrodes was performed in 15 consecutive patients (nine men, six women; mean age, 72.0 ± 8.2 years) with a solitary small (≤3 cm) hepatic tumor. The first five nodules were treated using the conventional puncture method with the standard attachment, then 10 nodules were ablated using the parallel puncture method with the double-barreled attachment. The times required for double-needle placement and the shapes of the ablated areas were compared between the two puncture methods. RESULTS: The parallel puncture method required a shorter time for double-needle placement than the conventional method (12 s [range, 8-24] vs 96 s [range, 50-240]; P = 0.0003), and allowed continuous observation of the tip of all needles and the size of the ablated area as it increased until completion of the ablation. The method also provided a stable ellipsoidal ablated area. The median height, width and thickness were 30 mm (range, 22-34), 30 mm (range, 21-33) and 20 mm (range, 7-25), respectively, using 20-mm electrodes, and 34 mm (range, 32-41), 36 mm (range, 35-38) and 24 mm (range, 23-24), respectively, using 30-mm electrodes. CONCLUSION: The parallel puncture method may be a feasible procedure for multi-needle RFA therapy.

Impact of branched-chain amino acid supplementation on survival in patients with advanced hepatocellular carcinoma treated with sorafenib: A multicenter retrospective cohort study.

AIM: The therapeutic efficacy of branched-chain amino acid (BCAA) when added to sorafenib has not been fully assessed in patients with advanced hepatocellular carcinoma (HCC). This multicenter study investigated whether BCAA supplementation improves prognosis in patients with advanced HCC who underwent sorafenib treatment. METHODS: This retrospective analysis included 256 patients with advanced HCC treated with sorafenib, including 55 who did and 201 who did not receive BCAA supplementation. Clinical characteristics and outcomes in relation to Child-Pugh classification were compared in the two groups. Statistical analyses of univariate, multivariate and propensity score-based procedures were used for this study. RESULTS: Assessment of 216 Child-Pugh A patients showed that median overall survival was significantly longer in patients with BCAA supplementation than in those without it (440 vs 299 days, P = 0.023). Multivariate analysis showed that BCAA supplementation (P = 0.023), low α-fetoprotein (<100 ng/mL) (P < 0.001), less progressive Barcelona Clinic Liver Cancer stage (A and B) (P = 0.007) and male sex (P = 0.018) were significant independent contributors to better overall survival. The significantly longer overall survival by BCAA supplementation was verified in the analysis using the propensity score in combination with the inverse probability of treatment weighted adjustment (P = 0.026). Assessment of the 40 Child-Pugh B patients showed no significant differences in overall survival between patients with and without BCAA supplementation. CONCLUSION: BCAA supplementation may be a valuable adjunctive therapy for improving prognosis in sorafenib-treated Child-Pugh A patients with advanced HCC.