The drug-metabolizing enzyme CYP is mainly involved in the metabolism of various substances in the liver, such as drugs, endogenous substances, and carcinogens. Recent reports have also revealed that CYP1B1 plays a major role in the developmental process. Because the level of CYP expression is markedly high in the liver, we hypothesize that CYP plays a role in the developmental process of the liver. To verify this hypothesis, we analyzed the expression patterns of various CYP molecular species and their functions during the differentiation of embryonic stem cells (ES cells) into hepatocytes and the developmental process in mice. The results demonstrated that CYP2R1 and CYP26A1 are expressed at an earlier stage of the differentiation of ES cells into hepatocytes than hepatoblast-specific markers. Additionally, during the development of the mouse liver, CYP2R1 and CYP26A1 were mostly up-regulated during the stage when hepatoblasts appeared. In addition, when CYP2R1 and CYP26A1 expressions were forced in ES cells and liver of adult mice, they differentiated into hepatoblast marker positive cells. These results suggest that CYP2R1 and CYP26A1 may play a major role in hepatoblast cell differentiation during the development of the liver.Key words embryonic stem cell; liver; CYP; hepatoblast The mature liver contains a variety of cell types, such as hepatocytes, sinusoidal endothelial cells, bile duct epithelial cells, stellate cells and Kupffer cells. The hepatocyte account for 80% of the volume of the liver and play a major role in liver functions, such as metabolism and excretion of drugs. Additionally, the fetal liver is known to act as a hematopoietic organ.The expression of CYP is significantly up-regulated in the liver, and CYP converts fat-soluble drugs into a water-soluble state so that they can be easily excreted. In addition to drug metabolism, CYP is also known to be involved in the oxidative metabolism of endogenous substances, including steroids, bile acid, hormones, and eicosanoids. To date, 58 types of CYP molecular species have been identified in humans, and 108 types have been identified in mice.1-3) As the CYP molecular species share high amino acid sequence homology in several substrate-recognition sites, in addition to the abovementioned functions of CYP, it may also play a major role in development and differentiation in the body. Because CYP begins to be expressed close to the time of hematopoiesis initiation during the fetal stage and CYP requires heme for its structure, CYP is considered to be involved in the development of the liver during the fetal stage. Moreover, it has been reported that the metabolites of endogenous substances and the intermediate metabolites of chemical substances have an effect on the development of individuals and on homeostasis.4-6) Therefore, CYP, which transiently appears during the process of development, is thought to possibly play an important role in the development of the liver.It has been reported that the knockout of either the CYP26A1 gene in mice causes abn...

Phenotypic screening in drug discovery has been revived with the expectation of providing promising lead compounds and drug targets and improving the success rate of drug approval. However, target identification remains a major bottleneck in phenotype-based drug discovery. We identified the lead compounds K542 and K405 with a selective inhibition of cell viability against sphingosine-1-phosphate lyase 1 (SGPL1)-transduced ES-2 cells by phenotypic screening. We therefore performed an in vivo pharmacological examination and observed the antitumor activity of K542 in an HT-1080 tumor-bearing mouse xenograft model. SGPL1 was expected to be a therapeutic target in some cancers, suggesting that these lead molecules might be promising candidates; however, their mechanisms of action still remain unexplained. We therefore synthesized the affinity probe Ind-tag derived from K542 and identified the proteins binding to Ind-tag via a pull-down experiment. Proteomics and biochemical analyses revealed that the target molecule of these lead compounds was Nicotinamide phosphoribosyltransferase (NAMPT). We established K542-resistant DLD-1 and HT-1080 cells, and genetic analyses of these cells identified a missense mutation in the NAMPT-encoding gene. This enzymatic experiment clearly showed that K393 exerts enzymatic inhibition against NAMPT. These proteomics, genetics and biochemical analyses clarified that compounds K542 and K405 were NAMPT inhibitors.

Cytochrome P450 enzymes (CYPs) are involved in the metabolism of various substances in the liver and small intestine and show markedly higher expression levels in the liver compared to other organs. The liver exhibits a remarkable capacity to regenerate. After excision of 70% of the liver, the organ can regenerate to its original size in approximately 1 week. Unlike the normal liver, in the injured liver, hepatic stem cells known as oval cells are considered to play an important role in regeneration. However, the role of CYPs in liver regeneration remains unclear. In the present study, we investigated the role of CYPs in the regeneration of injured liver. Liver injury was induced by 4-week repeated doses of 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) in the diet. Next, both DDC-fed mice and control diet (containing no DDC)-fed mice were subjected to 70% hepatectomy, and the hepatic gene expression patterns measured during regeneration were analyzed. Mice with DDC-induced liver injury expressed the oval cell markers cytokeratin 19 (CK19) and epithelial cell adhesion molecule (EpCAM), and partial hepatectomy increased the expression levels of CYP2R1 and CYP26A1 as well as the hepatoblast marker alpha-fetoprotein (AFP) in these mice. The results of this study suggest that CYP2R1 and CYP26A1 are important in the differentiation of oval cells into hepatoblastlike cells in the injured liver. Key words injured liver CYP26A1; CYP2R1; liver regenerationThe adult liver consists of hepatic parenchymal cells (hepatocytes), hepatic sinusoidal endothelial cells, biliary epithelial cells, stellate cells, and Kupffer cells. Hepatic parenchymal cells occupy approximately 80% of the liver volume and play important roles in various liver functions, including nutrient storage, detoxification, metabolism, and excretion. The fetal liver, in contrast, is the major site of hematopoiesis.The liver has the highest regenerative capacity of any organ in the body. After excision of 70% of the liver, the remnant liver regenerates to its original size in approximately 1 week through the hypertrophy or proliferation of the remaining tissue. Two major mechanisms have been proposed for liver generation: hypertrophy and proliferation of hepatocytes and differentiation of undifferentiated cells into hepatocytes.1) In the first mechanism, in a normal liver that has undergone partial hepatectomy, the remaining hepatocytes retain the potential for hypertrophy and proliferation for liver regeneration. In the second mechanism, in a liver that has been injured by drugs, the proliferative capacity of hepatocytes is reduced, and thus the hypertrophy or proliferation of hepatocytes is unlikely. 1)Instead, some biliary epithelial cells dedifferentiate, likely into undifferentiated oval cells, which then proliferate and differentiate into hepatocytes to contribute to liver regeneration.In rats in which hepatocyte proliferation is inhibited by 2-acethylaminofluorene, partial hepatectomy or administration of carbon tetrachloride results in the emergence of...