論文 - 齋藤　義正

Generation of functional human hepatocytes in vitro: Current status and future prospects

Yamaguchi T., Matsuzaki J., Katsuda T., Saito Y., Saito H., Ochiya T.

Inflammation and Regeneration 39 （ 1 ）  2019年07月

研究論文（学術雑誌）, 共著, 査読有り

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© 2019 The Author(s). Liver and hepatocyte transplantation are the only effective therapies for late-stage liver diseases, in which the liver loses its regenerative capacity. However, there is a shortage of donors. As a potential alternative approach, functional hepatocytes were recently generated from various cell sources. Analysis of drug metabolism in the human liver is important for drug development. Consequently, cells that metabolize drugs similar to human primary hepatocytes are required. This review discusses the current challenges and future perspectives concerning hepatocytes and hepatic progenitor cells that have been reprogrammed from various cell types, focusing on their functions in transplantation models and their ability to metabolize drugs.

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Circulating microRNA-1246 as a possible biomarker for early tumor recurrence of hepatocellular carcinoma

Chuma M., Toyoda H., Matsuzaki J., Saito Y., Kumada T., Tada T., Kaneoka Y., Maeda A., Yokoo H., Ogawa K., Kamiyama T., Taketomi A., Matsuno Y., Yazawa K., Takeda K., Kunisaki C., Ogushi K., Moriya S., Hara K., Nozaki A., Kondo M., Fukuda H., Numata K., Tanaka K., Maeda S., Sakamoto N.

Hepatology Research 49 （ 7 ） 810 - 822 2019年07月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  13866346

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© 2019 The Japan Society of Hepatology Aims: Early tumor recurrence (ETR) after hepatic resection is a crucial predictor of poor prognosis in patients with hepatocellular carcinoma (HCC). The aim of this study was to identify clinically significant serum microRNAs (miRNAs) involved in the ETR of HCC. Methods: We compared expression profiles of circulating miRNAs from serum samples between five HCC patients with ETR (recurrence within 12 months after hepatectomy) and five HCC patients without recurrence using microarray analysis of miRNA. The identified miRNA associated with ETR was further verified in 121 HCC patients, 73 liver disease patients, and 15 health controls by real-time quantitative reverse transcription–polymerase chain reaction (PCR). Results: Of the approximately 2000 miRNAs analyzed, we identified 15 miRNAs for which expression levels correlated significantly with ETR. Of these miRNAs, we further investigated expression of miRNA-1246 (miR-1246). Quantitative PCR confirmed that miR-1246 was upregulated in HCC with ETR, compared to the level in HCC without ETR (P < 0.001). Serum miR-1246 showed a receiver operating characteristic curve area of 0.762, with 77.4% specificity and 54.1% sensitivity in discriminating HCC patients with ETR from HCC patients without ETR. Altered expression of miR-1246 was associated with aggressive tumor characteristics, including tumor–node–metastasis classification (P = 0.0413), tumor differentiation (P = 0.0419), and portal vein invasion (P = 0.0394). Moreover, multivariate Cox regression analysis identified serum miR-1246 level as an independent risk factor for overall survival (hazard ratio, 2.784; 95% confidence interval, 1.528–5.071; P = 0.0008). Conclusion: Circulating miR-1246 in serum has strong potential as a novel ETR and prognostic biomarker for HCC.

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Establishment of Patient-Derived Organoids and Drug Screening for Biliary Tract Carcinoma

Saito Y., Muramatsu T., Kanai Y., Ojima H., Sukeda A., Hiraoka N., Arai E., Sugiyama Y., Matsuzaki J., Uchida R., Yoshikawa N., Furukawa R., Saito H.

Cell Reports 27 （ 4 ） 1265 - 1276.e4 2019年04月

研究論文（学術雑誌）, 共著, 筆頭著者, 査読有り

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© 2019 The Author(s) Biliary tract carcinomas (BTCs) are among the most aggressive malignancies and have a poor prognosis. Here, we successfully established organoid lines derived from intrahepatic cholangiocarcinoma, gallbladder cancer, and neuroendocrine carcinoma of the ampulla of Vater. These organoids derived from BTCs were cultured stably for >1 year and closely recapitulated the histopathology, gene expression, and genetic alterations evident in the primary tumors. Gene expression profiling of the organoids revealed that SOX2 could be a potential prognostic biomarker for patients with BTC. We screened a compound library consisting of drugs used clinically for their ability to suppress organoids derived from BTCs and found that the antifungal drugs amorolfine and fenticonazole significantly suppressed the growth of organoids derived from BTCs with minimal toxicity to normal biliary epithelial cells. Patient-derived organoids may be a powerful research tool for the clarification of molecular pathogenesis and the discovery of biomarkers and therapeutic drugs for refractory cancers.

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CAPZA1 determines the risk of gastric carcinogenesis by inhibiting Helicobacter pylori CagA-degraded autophagy

Tsugawa H., Mori H., Matsuzaki J., Sato A., Saito Y., Imoto M., Suematsu M., Suzuki H.

Autophagy 15 （ 2 ） 242 - 258 2019年02月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  15548627

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© 2018, © 2018 Informa UK Limited, trading as Taylor & Francis Group. Helicobacter pylori-derived CagA, a type IV secretion system effector, plays a role as an oncogenic driver in gastric epithelial cells. However, upon delivery into gastric epithelial cells, CagA is usually degraded by macroautophagy/autophagy. Hence, the induction of autophagy in H. pylori-infected epithelial cells is an important host-protective ability against gastric carcinogenesis. However, the mechanisms by which autophagosome-lysosome fusion is regulated, are unknown. Here, we report that enhancement of LAMP1 (lysosomal associated membrane protein 1) expression is necessary for autolysosome formation. LAMP1 expression is induced by nuclear translocated LRP1 (LDL receptor related protein 1) intracellular domain (LRP1-ICD) binding to the proximal LAMP1 promoter region. Nuclear translocation of LRP1-ICD is enhanced by H. pylori infection. In contrast, CAPZA1 (capping actin protein of muscle Z-line alpha subunit 1) inhibits LAMP1 expression via binding to LRP1-ICD in the nuclei. The binding of CAPZA1 to LRP1-ICD prevents LRP1-ICD binding to the LAMP1 proximal promoter. Thus, in CAPZA1-overexpressing gastric epithelial cells infected with H. pylori, autolysosome formation is inhibited and CagA escapes autophagic degradation. These findings identify CAPZA1 as a novel negative regulator of autolysosome formation and suggest that deregulation of CAPZA1 expression leads to increased risk of gastric carcinogenesis. Abbreviations: CagA: cytotoxin-associated gene A; CAPZA1: capping actin protein of muscle Z-line alpha subunit 1; ChIP: chromatin immunoprecipitation; GTF2I: general transcription factor IIi; HDAC: histone deacetylase; LAMP1: lysosomal associated membrane protein 1; LRP1: LDL receptor related protein 1; LRP1-ICD: CagA intracellular domain; qPCR: quantitative polymerase chain reaction; VacA: vacuolating cytotoxin.

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Vonoprazan-based third-line therapy has a higher eradication rate against sitafloxacin-resistant Helicobacter pylori

Saito Y., Konno K., Sato M., Nakano M., Kato Y., Saito H., Serizawa H.

Cancers 11 （ 1 ）  2019年01月

研究論文（学術雑誌）, 共著, 査読有り

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© 2019 by the authors. Licensee MDPI, Basel, Switzerland. Eradication of Helicobacter pylori (H. pylori) is an effective strategy for preventing various gastrointestinal diseases such as gastric cancer and mucosa-associated lymphoid tissue (MALT) lymphoma. However, the eradication success rate is decreasing because of a recent increase in drug-resistant strains of H. pylori. Here, we evaluated the success rate of eradication therapy with vonoprazan (VPZ), a new potassium-competitive acid blocker, against drug-resistant H. pylori. In total, 793 patients who received H. pylori eradication therapy were investigated retrospectively. All underwent esomeprazole (EPZ)-based triple therapy (n = 386) or VPZ-based triple therapy (n = 407) for first-, second- and third-line H. pylori eradication for 7 days. The overall success rates of first- and third-line H. pylori eradication were significantly higher for VPZ-based triple therapy (88.4% and 93.0%, respectively, per protocol (PP)) than for EPZ-based triple therapy (69.5% and 56.5%, respectively, PP). Moreover, the success rates of first- and third-line eradication of clarithromycin (CLR)- and sitafloxacin (STFX)-resistant H. pylori were significantly higher for VPZ-based triple therapy (72.0% and 91.7%, PP) than for EPZ-based triple therapy (38.5% and 20.0%, PP). In addition, patient age did not affect the eradication rate of VPZ-based first-line therapy, whereas the success rate of EPZ-based therapy was lower in patients under 65 years of age. Our results clearly demonstrated that VPZ-based therapy achieved a higher eradication rate even against CLR- and STFX-resistant H. pylori, and that patient age did not affect the eradication rate of VPZ-based therapy. These findings suggest that dual therapy using VPZ and amoxicillin may be sufficient for standard H. pylori eradication, and may thus also be beneficial for avoiding antibiotic misuse.

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Establishment of an organoid bank of biliary tract and pancreatic cancers and its application for personalized therapy and future treatment

Saito Y.

Journal of Gastroenterology and Hepatology 34 （ 11 ） 1906 - 1910 2019年

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  08159319

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© 2019 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd Biliary tract cancers and pancreatic cancers are aggressive malignancies that are difficult to diagnose early and have a poor prognosis. Patients with inoperable biliary tract and pancreatic cancers generally receive chemotherapy regimens including gemcitabine. However, the effects of these drugs are limited, and the 5-year survival rates of patients are very low. The newly developed three-dimensional culture system known as “organoid culture” allows long-term expansion of stem cells into cyst-like structures (organoids) with properties resembling those of the original tissues. We and other groups have successfully established long-term in vitro cultures of organoids derived from biliary tract and pancreatic cancers. Organoids derived from biliary tract and pancreatic cancers closely recapitulate the properties of the original tumors including genetic alterations, gene expression profiles, and histopathological structures. These patient-derived cancer organoids can be applied for drug sensitivity testing, drug screening, epigenetic therapy, and differentiation-inducing therapy to identify therapeutic agents optimal for each patient. We intend to further establish organoids derived from various cancer cases and construct an organoid bank of biliary tract and pancreatic cancers. These powerful in vitro preclinical models of refractory cancers may bridge the gap between basic research and clinical trials and allow personalized therapy for patients.

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Anticancer Drug Prescription Patterns in Japan: Future Directions in Cancer Therapy

Shibata S., Matsushita M., Saito Y., Suzuki T.

Therapeutic Innovation and Regulatory Science 52 （ 6 ） 718 - 723 2018年11月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  21684790

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© The Author(s) 2018. Background: Despite their benefits, the rapid development of new cancer treatments has been a significant driver of increasing health care expenditures in the face of limited health care budgets. In this study, we analyzed the prescribing trends for anticancer drugs from 2010 through 2016 in Japan and sought to identify unique trends that could provide a basis for future medical economic research aiming to develop more efficacious and cost-effective cancer therapies. Methods: We used publicly available marketing data for anticancer drugs in Japan for 2010-2016. The drugs selected for this research were categorized according to the Anatomical Therapeutic Chemical Classification System. We investigated the overall anticancer drug market size, the number of anticancer drugs, the top 30 selling anticancer categories, sales and prescription volumes, and changes in sales and prescription volumes between 2010 and 2016 in the country. Results: The anticancer agent market expanded each year from 2010 to 2016, with sales exceeding 1 trillion yen in 2015. The proportion of molecular targeted drugs (antineoplastic mAbs and protein kinase inhibitors) among the top 30 selling anticancer categories has continued to increase, and both the sales and prescription volumes of these drugs exceeded those of drugs in other categories, suggesting that these treatments play a dominant role in cancer pharmacotherapy. Conclusion: The availability and increasing use of innovative but more expensive targeted therapies were major drivers of increases in pharmaceutical expenditures for cancer treatment in Japan. Therefore, the effective use of genetic testing can mitigate these rising costs.

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Nrf2mediated antioxidant effects contribute to suppression of nonalcoholic steatohepatitis associated hepatocellular carcinoma in murine model

Yamada S., Kimura M., Saito Y., Saito H.

Journal of Clinical Biochemistry and Nutrition （Journal of Clinical Biochemistry and Nutrition）  63 （ 2 ） 123 - 128 2018年09月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  09120009

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©2018 JCBN. The exact mechanisms of hepatocellular carcinoma development in nonalcoholic steatohepatitis remain unclear. In this study, we used a new class of highfat diet, which could induce hepatocellular carcinoma development without the use of general chemical carcinogens or knockout mice. We investigated the correlation between hepatocellular carcinoma and oxidative stress/anti oxidant effects after depletion of the gut microbiota by treatment with antibiotics. Mice fed with the steatohepatitisinducing high fat diet (STHD01) for 41 weeks developed hepatocellular carci noma. Antibiotictreatment in mice fed with STHD01 significantly depleted the gut microbiota and significantly ameliorated liver injury/histology. The tumor numbers of hepatocellular carcinoma were dramatically decreased by the antibioticstreatment. We analyzed the factors involved in oxidative stress and antioxidant effects. Oxidative stress was elevated in mice fed with STHD01, whereas some antioxidant factors were significantly elevated after antibiotics treatment. These results suggest that the gut microbiota is a key factor in improving oxidative stress induced by STHD01 feeding.

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Optimal Anti-cancer Drug Profiles for Effective Penetration of the Anti-cancer Drug Market by Generic Drugs in Japan

Shibata S., Matsushita M., Saito Y., Suzuki T.

Therapeutic Innovation and Regulatory Science 52 （ 4 ） 442 - 448 2018年07月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  21684790

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© The Author(s) 2017. Background: The increased use of generic drugs is a good indicator of the need to reduce the increasing costs of prescription drugs. Since there are more expensive drugs compared with other therapeutic areas, “oncology” is an important one for generic drugs. The primary objective of this article was to quantify the extent to which generic drugs in Japan occupy each level of the Anatomical Therapeutic Chemical (ATC) classification system. Methods: The dataset used in this study was created from publicly available information obtained from the IMS Japan Pharmaceutical Market database. Data on the total amount of sales and number of prescriptions for anti-cancer drugs between 2010 and 2016 in Japan were selected. The data were categorized according to the third level of the ATC classification system. Results: All categories of the ATC classification system had increased market shares in Japan between 2010 and 2016. The barriers to market entry were relatively low in L01F (platinum anti-neoplastics), L01C (plant-based neoplastics), L02B (cytostatic hormone antagonists), and L01D (anti-neoplastic antibiotics) but were high in L02A (cytostatic hormones), L01H (protein kinase inhibitors), and L01B (anti-metabolites). Conclusions: Generic cancer drugs could bring savings to Japanese health care systems. Therefore, their development should be directed toward niche markets, such as L02A, L01H, and L01B, and not competitive markets.

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Dual effects of the Nrf2 inhibitor for inhibition of hepatitis C virus and hepatic cancer cells

Murakami Y., Sugiyama K., Ebinuma H., Nakamoto N., Ojiro K., Chu P., Taniki N., Saito Y., Teratani T., Koda Y., Suzuki T., Saito K., Fukasawa M., Ikeda M., Kato N., Kanai T., Saito H.

BMC Cancer 18 （ 1 ）  2018年06月

研究論文（学術雑誌）, 共著, 査読有り

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© 2018 The Author(s). Background: We previously showed that knockdown of nuclear factor E2-related factor 2 (Nrf2) resulted in suppression of hepatitis C virus (HCV) infection. In this study, whether brusatol, an Nrf2 inhibitor, has dual anti-HCV and anticancer effects was explored. Methods: The anti-HCV effect of brusatol was investigated by analyzing HCV RNA and proteins in a hepatic cell line persistently-infected with HCV, HPI cells, and by analyzing HCV replication in a replicon-replicating hepatic cell line, OR6 cells. Then, dual anti-HCV and anticancer effects of brusatol and enhancement of the effects by the combination of brusatol with anticancer drugs including sorafenib, which has been reported to have the dual effects, were then investigated. Results: Brusatol suppressed the persistent HCV infection at both the RNA and protein levels in association with a reduction in Nrf2 protein in the HPI cells. Analysis of the OR6 cells treated with brusatol indicated that brusatol inhibited HCV persistence by inhibiting HCV replication. Combination of brusatol with an anticancer drug not only enhanced the anticancer effect but also, in the case of the combination with sorafenib, strongly suppressed HCV infection. Conclusions: Brusatol has dual anti-HCV and anticancer effects and can enhance the comparable effects of sorafenib. There is therefore the potential for combination therapy of brusatol and sorafenib for HCV-related hepatocellular carcinoma.

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Bile acid metabolism regulated by the gut microbiota promotes non-alcoholic steatohepatitis-associated hepatocellular carcinoma in mice

Yamada S., Takashina Y., Watanabe M., Nagamine R., Saito Y., Kamada N., Saito H.

Oncotarget 9 （ 11 ） 9925 - 9939 2018年06月

研究論文（学術雑誌）, 共著, 査読有り

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© Yamada et al. Gut microbiota plays a significant role in the development of hepatocellular carcinoma (HCC) in non-alcoholic steatohepatitis (NASH). However, understanding of the precise mechanism of this process remains incomplete. A new class steatohepatitis-inducing high-fat diet (HFD), namely STHD-01, can promote the development of HCC without the administration of chemical carcinogens. Using this diet, we comprehensively analyzed changes in the gut microbiota and its metabolic functions during the development of HCC in NASH. Mice fed the STHD-01 developed NASH within 9 weeks. NASH further progressed into HCC by 41 weeks. Treatment with antibiotics significantly attenuated liver pathology and suppressed tumor development, indicating the critical role of the gut microbiota in tumor development in this model. Accumulation of cholesterol and bile acids in the liver and feces increased after feeding the mice with STHD-01. Treatment with antibiotics did not reverse these phenotypes. In contrast, accumulation of secondary bile acids was dramatically reduced after the treatment with antibiotics, suggesting the critical role of the gut microbiota in the conversion of primary bile acids to secondary bile acids. Secondary bile acids such as deoxycholic acid activated the mTOR, pathway in hepatocytes. Activation of mTOR was observed in the liver of mice fed STHD-01, and the activation was reduced when mice were treated with antibiotics. Collectively, bile acid metabolism by the gut microbiota promotes HCC development in STHD-01- induced NASH.

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機能性食品とアンチエイジング　消化機能低下の予防

齋藤 義正、齋藤 英胤

モダンフィジシャン （新興医学出版社）  38 （ 4 ） 357 - 359 2018年04月

研究論文（学術雑誌）, 共著

Induction of differentiation of intrahepatic cholangiocarcinoma cells to functional hepatocytes using an organoid culture system

Saito Y., Nakaoka T., Muramatsu T., Ojima H., Sukeda A., Sugiyama Y., Uchida R., Furukawa R., Kitahara A., Sato T., Kanai Y., Saito H.

Scientific Reports 8 （ 1 ） 2821 2018年02月

研究論文（学術雑誌）, 共著, 査読有り

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© 2018 The Author(s). Intrahepatic cholangiocarcinoma (IHCC) is a highly aggressive malignancy with a poor prognosis. It is thought to originate from cholangiocytes, which are the component cells of intrahepatic bile ducts. However, as patients with viral hepatitis often develop IHCC, it has been suggested that transformed hepatocytes may play a role in IHCC development. To investigate whether IHCC cells can be converted to functional hepatocytes, we established organoids derived from human IHCC and cultured them under conditions suitable for hepatocyte differentiation. IHCC organoids after hepatocyte differentiation acquired functions of mature hepatocytes such as albumin secretion, bile acid production and increased CYP3A4 activity. Studies using a mouse model of IHCC indicate that Wnt3a derived from macrophages recruited upon inflammation in the liver may promote the malignant transformation of hepatocytes to IHCC cells. The results of the present study support the recently proposed hypothesis that IHCC cells are derived from hepatocytes.

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Gut microbiota-mediated generation of saturated fatty acids elicits inflammation in the liver in murine high-fat diet-induced steatohepatitis.

Yamada S, Kamada N, Amiya T, Nakamoto N, Nakaoka T, Kimura M, Saito Y, Ejima C, Kanai T, Saito H

BMC Gastroenterol. 17 （ 1 ） 136 2017年11月

研究論文（学術雑誌）, 共著

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Aberrant DNA Methylation as a Biomarker and a Therapeutic Target of Cholangiocarcinoma.

Nakaoka T, Saito Y, Saito H.

Int J Mol Sci. 18 （ 6 ） E1111 2017年05月

共著, 査読有り

Cluster microRNAs miR-194 and miR-215 suppress the tumorigenicity of intestinal tumor organoids.

Nakaoka T, Saito Y, Shimamoto Y, Muramatsu T, Kimura M, Kanai Y, Saito H.

Cancer Sci. 108 （ 4 ） 678 - 684 2017年04月

共著, 査読有り

A New Molecular Mechanism Underlying the Antitumor Effect of DNA Methylation Inhibitors via an Antiviral Immune Response.

Saito Y, Nakaoka T, Saito H.

Advances in Protein Chemistry and Structural Biology 106   227 - 242 2017年01月

研究論文（学術雑誌）, 共著, 査読有り

microRNA-mediated resistance to hypoglycemia in the HepG2 human hepatoma cell line.

Ueki S, Murakami Y, Yamada S, Kimura M, Saito Y, Saito H.

BMC Cancer 16 （ 1 ） 732 - 744 2016年

研究論文（学術雑誌）

Inhibition of DNA methylation suppresses intestinal Tumor Organoids by inducing an Anti-Viral response.

Saito Y, Nakaoka T, Sakai K, Muramatsu T, Toshimitsu K, Kimua M, Kanai T, Sato T, Saito H.

Sci Rep 6   25311 2016年

研究論文（学術雑誌）, 共著

First-line eradication for Helicobacter pylori-positive gastritis by esomeprazole-based triple therapy is influenced by CYP2C19 genotype.

Saito Y, Serizawa H, Kato Y, Nakano M, Nakamura M, Saito H, Suzuki H, Kanai T.

World J Gastroenterol. 21(48)   13548 - 13554 2015年12月

研究論文（学術雑誌）, 共著, 査読有り