title: A new class of pluripotent stem cell cytotoxic small molecules.
abstract: A major concern in Pluripotent Stem Cell (PSC)-derived cell replacement therapy is the risk of teratoma formation from contaminating undifferentiated cells. Removal of undifferentiated cells from differentiated cultures is an essential step before PSC-based cell therapies can be safely deployed in a clinical setting. We report a group of novel small molecules that are cytotoxic to PSCs. Our data indicates that these molecules are specific and potent in their activity allowing rapid eradication of undifferentiated cells. Experiments utilizing mixed PSC and primary human neuronal and cardiomyocyte cultures demonstrate that up to a 6-fold enrichment for specialized cells can be obtained without adversely affecting cell viability and function. Several structural variants were synthesized to identify key functional groups and to improve specificity and efficacy. Comparative microarray analysis and ensuing RNA knockdown studies revealed involvement of the PERK/ATF4/DDIT3 ER stress pathway. Surprisingly, cell death following ER stress induction was associated with a concomitant decrease in endogenous ROS levels in PSCs. Undifferentiated cells treated with these molecules preceding transplantation fail to form teratomas in SCID mice. Furthermore, these molecules remain non-toxic and non-teratogenic to zebrafish embryos suggesting that they may be safely used in vivo.
authors.affiliation: School of Chemical & Life Sciences, Nanyang Polytechnic, Singapore.
authors.forename: Mark
authors.initials: M
authors.lastname: Richards
authors.affiliation: School of Chemical & Life Sciences, Nanyang Polytechnic, Singapore.
authors.forename: Chee Wee
authors.initials: CW
authors.lastname: Phoon
authors.affiliation: Singapore Eye Research Institute, 11 Third Hospital Avenue, Singapore.
authors.forename: Gwendoline Tze Wei
authors.initials: GT
authors.lastname: Goh
authors.affiliation: School of Chemical & Life Sciences, Nanyang Polytechnic, Singapore.
authors.forename: Eng Khuan
authors.initials: EK
authors.lastname: Seng
authors.affiliation: School of Chemical & Life Sciences, Nanyang Polytechnic, Singapore.
authors.forename: Xu Ming
authors.initials: XM
authors.lastname: Guo
authors.affiliation: School of Chemical & Life Sciences, Nanyang Polytechnic, Singapore.
authors.forename: Cherine Mei Fong
authors.initials: CM
authors.lastname: Tan
authors.affiliation: Department of Biological Sciences, National University of Singapore, Singapore.
authors.forename: Woon-Khiong
authors.initials: WK
authors.lastname: Chan
authors.affiliation: School of Chemical & Life Sciences, Nanyang Polytechnic, Singapore; Food & Human Nutrition Programme (Singapore), School of Agriculture, Food and Rural Development, Agriculture Building, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: Joel Mun Kin
authors.initials: JM
authors.lastname: Lee
mesh_terms: D051701:Activating Transcription Factor 4 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003603:Cytotoxins / Q000138:chemical synthesis / Q000494:pharmacology*; D004625:Embryo, Nonmammalian / Q000187:drug effects / Q000502:physiology; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects / Q000235:genetics; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009928:Organ Specificity; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000037:antagonists & inhibitors / Q000378:metabolism; D015398:Signal Transduction; D054852:Small Molecule Libraries / Q000138:chemical synthesis / Q000494:pharmacology*; D033581:Stem Cell Transplantation; D013329:Structure-Activity Relationship; D013724:Teratoma / Q000517:prevention & control*; D051742:Transcription Factor CHOP / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015027:Zebrafish; D019892:eIF-2 Kinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D051701:Activating Transcription Factor 4; D003603:Cytotoxins; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D054852:Small Molecule Libraries; D051742:Transcription Factor CHOP; D019892:eIF-2 Kinase; C081452:Atf4 protein, mouse; C497250:Ddit3 protein, mouse