Using the Inducible Caspase-9 Suicide-Safeguard System with iPSC and Bioluminescent Tracking

Jonathan Villanueva; Toshinobu Nishimura; Hiromitsu Nakauchi

doi:10.1007/978-1-4939-9728-2_20

Using the Inducible Caspase-9 Suicide-Safeguard System with iPSC and Bioluminescent Tracking

Methods Mol Biol. 2019:2048:259-264. doi: 10.1007/978-1-4939-9728-2_20.

Authors

Jonathan Villanueva¹, Toshinobu Nishimura¹, Hiromitsu Nakauchi²

Affiliations

¹ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA, USA.
² Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA, USA. nakauchi@stanford.edu.

PMID: 31396943
DOI: 10.1007/978-1-4939-9728-2_20

Abstract

For scientists working within the field of induced pluripotent stem cells (iPSCs), this protocol will provide a thorough walk-through on how to conduct in vitro and in vivo experiments that validate the function of a particular safeguard system technology. In short, we provide instructions on how to generate inducible Caspase-9 (iC9) safeguard system with human iPSCs that act as normal or abnormal models of the cells for therapeutics to be tried after differentiation. These iC9-iPSCs should be modified prior to beginning this protocol by constitutively expressing luciferase, an enzyme capable of generating bioluminescent signals through the oxidation of the substrate luciferin. Monitoring the bioluminescent signal over time provides the information on whether a safeguard system is working or not.

Keywords: Bioluminescence; Cancer; Cytotoxicity; Immunotherapy; T cell.

MeSH terms

Animals
Benzothiazoles / administration & dosage
Benzothiazoles / chemistry
Caspase 9 / genetics
Caspase 9 / metabolism
Cell Culture Techniques / instrumentation
Cell Culture Techniques / methods
Cell Differentiation
Cell Line
Culture Media / metabolism
Disease Models, Animal
Gene Expression / drug effects
Genes, Reporter / drug effects
Genes, Reporter / genetics
Genes, Transgenic, Suicide*
Humans
Immunotherapy / adverse effects
Immunotherapy / methods
Induced Pluripotent Stem Cells / metabolism
Injections, Intraperitoneal
Intravital Microscopy / instrumentation
Intravital Microscopy / methods*
Luciferases, Firefly / genetics
Luciferases, Firefly / metabolism
Luminescent Measurements / instrumentation
Luminescent Measurements / methods*
Mice, Inbred NOD
Mice, SCID
Tacrolimus / administration & dosage
Tacrolimus / analogs & derivatives
Teratoma / diagnostic imaging*
Teratoma / immunology
Teratoma / pathology
Teratoma / therapy
Tumor Burden

Substances

AP20187
Benzothiazoles
Culture Media
D-luciferin
Luciferases, Firefly
CASP9 protein, human
Caspase 9
Tacrolimus