title: Elucidation of a novel phenformin derivative on glucose-deprived stress responses in HT-29 cells.
abstract: Recently, we developed a variety of phenformin derivatives as selective antitumor agents. Based on previous findings, this study evaluated a promising compound, 2-(2-chlorophenyl)ethylbiguanide (2-Cl-Phen), on the basis of stress responses in the human colon cancer cell line HT-29 under a serum- and glucose-deprived condition. 2-Cl-Phen triggered morphological changes such as shrinkage and plasma membrane disintegration, as well as a decrease in mitochondrial activity and an increase in LDH leakage. To understand intracellular issues relating to 2-Cl-Phen, this study focused on the expression levels of ER stress-inducible genes and several oncogenic genes. Serum and glucose deprivation significantly induced a variety of ER stress-inducible genes, but a 12-h treatment of 2-Cl-Phen down-regulated expression of several ER stress-related genes, with the exception of GADD153. Interestingly, the expression levels of ATF6α, GRP78, MANF, and CRELD2 mRNA were almost completely decreased by 2-Cl-Phen. This study also observed that a 24-h treatment of 2-Cl-Phen attenuated the expression levels of GRP78, GADD153, and c-Myc protein. The decrease in c-Myc protein occurred before the fluctuation of GRP78 protein, while the expression of c-Myc mRNA showed little change with cotreatment of serum and glucose deprivation with 2-Cl-Phen. To further understand the 2-Cl-Phen-induced down-regulation of ATF6-related genes, this study investigated the stability of ATF6α and GRP78 proteins using NanoLuc-tagged constructs. The expression levels of NanoLuc-tagged ATF6α and GRP78 were significantly down-regulated by 2-Cl-Phen in the presence or absence of the translation inhibitor cycloheximide. Taken together, our novel phenformin derivative 2-Cl-Phen has the unique characteristic of diminishing tumor adaptive responses, especially the expression of ATF6-related genes, as well as that of c-Myc protein, in a transcriptional and posttranscriptional manner under a serum- and glucose-deprived condition. Further characterization of cytotoxic mechanisms related to phenformin derivatives may give new insights into developing additional promising anticancer agents.
authors.affiliation: Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan. oohashi@gifu-u.ac.jp.|United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan. oohashi@gifu-u.ac.jp.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Oh-Hashi
authors.affiliation: Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan.
authors.forename: Nao
authors.initials: N
authors.lastname: Irie
authors.affiliation: Laboratory of Pharmaceutical and Medicinal Chemistry, Gifu Pharmaceutical University, 1-25-4 Daigakunishi, Gifu, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Sakai
authors.affiliation: Laboratory of Bioorganic and Natural Products Chemistry, Kobe Pharmaceutical University, 4-19-1 Motoyama-kita, Higashinada, Kobe, 658-8558, Japan.
authors.forename: Kensuke
authors.initials: K
authors.lastname: Okuda
authors.affiliation: Laboratory of Pharmaceutical and Medicinal Chemistry, Gifu Pharmaceutical University, 1-25-4 Daigakunishi, Gifu, Japan.
authors.forename: Hideko
authors.initials: H
authors.lastname: Nagasawa
authors.affiliation: Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan.|United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Hirata
authors.affiliation: Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan.|United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan.
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Kiuchi
mesh_terms: D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000378:metabolism* / Q000473:pathology; D000091342:Endoplasmic Reticulum Chaperone BiP; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D005947:Glucose; D006801:Humans; D009363:Neoplasm Proteins / Q000096:biosynthesis*; D010629:Phenformin / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D009363:Neoplasm Proteins; D010629:Phenformin; D005947:Glucose
keywords: ATF6; ER stress; GRP78; Phenformin; c-Myc