title: Escape Mutations, Ganciclovir Resistance, and Teratoma Formation in Human iPSCs Expressing an HSVtk Suicide Gene.
abstract: Human pluripotent stem cells (hPSCs) hold great promise for cell therapy. However, a major concern is the risk of tumor formation by residual undifferentiated cells contaminating the hPSC-derived cell product. Suicide genes could safeguard against such adverse events by enabling elimination of cells gone astray, but the efficacy of this approach has not yet been thoroughly tested. Here, we engineered a lentivirally encoded herpes simplex virus thymidine kinase (HSVtk) with expression restricted to undifferentiated hPSCs through regulation by the let7 family of miRNAs. We show that induced pluripotent stem cells (iPSCs) expressing a let7-regulated HSVtk transgene are selectively killed by ganciclovir (GCV), whereas differentiated cells are fully protected. However, in contrast to previous studies, we find that in vivo GCV administration results in longer latency but does not prevent teratoma formation by iPSCs expressing either a constitutive or a let7-regulated HSVtk, without evidence of silencing of the HSVtk. Clonal analyses of iPSCs expressing HSVtk revealed frequent emergence of GCV resistance which, at least in some cases, could be attributed to preexisting inactivating mutations in the HSVtk coding sequence, selected for upon GCV treatment. Our findings have important consequences for the future use of suicide genes in hPSC-based cell therapies.
authors.affiliation: Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Division of Hematology, Department of Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Andriana G
authors.initials: AG
authors.lastname: Kotini
authors.affiliation: Antitumor Assessment Core, Memorial Sloan Kettering Cancer Center, New York, New York, USA.
authors.forename: Elisa
authors.initials: E
authors.lastname: de Stanchina
authors.affiliation: Center for Cell Engineering, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.
authors.forename: Maria
authors.initials: M
authors.lastname: Themeli
authors.affiliation: Center for Cell Engineering, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.
authors.forename: Michel
authors.initials: M
authors.lastname: Sadelain
authors.affiliation: Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, USA.|Division of Hematology, Department of Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Eirini P
authors.initials: EP
authors.lastname: Papapetrou
references.citation: Stem Cells. 2012 May;30(5):935-45
references.pmid: 22328536
references.citation: J Clin Invest. 2009 Jan;119(1):157-68
references.pmid: 19033646
references.citation: J Clin Invest. 2013 Apr;123(4):1647-61
references.pmid: 23524971
references.citation: Stroke. 2010 Jan;41(1):153-9
references.pmid: 19940279
references.citation: Cell Stem Cell. 2012 Nov 2;11(5):615-9
references.pmid: 23084023
references.citation: Cancer Gene Ther. 2014 Nov;21(11):472-82
references.pmid: 25323693
references.citation: Stem Cells. 2008 Jun;26(6):1454-63
references.pmid: 18356574
references.citation: Stem Cells Dev. 2012 Mar 1;21(4):521-9
references.pmid: 21631153
references.citation: Nat Med. 2006 Nov;12(11):1259-68
references.pmid: 17057709
references.citation: Nucleic Acids Res. 2006 Jan 24;34(2):e9
references.pmid: 16434699
references.citation: Stem Cells. 2003;21(3):257-65
references.pmid: 12743320
references.citation: Nat Med. 2006 May;12(5):585-91
references.pmid: 16633348
references.citation: J Biol Chem. 2012 Sep 21;287(39):32338-45
references.pmid: 22865887
references.citation: Nat Rev Genet. 2009 Aug;10(8):578-85
references.pmid: 19609263
references.citation: Cell. 2003 Jun 13;113(6):803-9
references.pmid: 12809610
references.citation: Dev Biol. 2004 Jun 15;270(2):488-98
references.pmid: 15183728
references.citation: Stem Cell Reports. 2015 Mar 10;4(3):360-73
references.pmid: 25684226
references.citation: Cell Stem Cell. 2009 Apr 3;4(4):289-300
references.pmid: 19341619
references.citation: Dev Cell. 2003 Aug;5(2):351-8
references.pmid: 12919684
references.citation: Mol Ther. 2007 Jan;15(1):183-92
references.pmid: 17164790
references.citation: Nat Protoc. 2011 Aug 04;6(9):1251-73
references.pmid: 21886095
references.citation: Stem Cells Dev. 2008 Aug;17(4):619-27
references.pmid: 18393636
references.citation: Stem Cells. 2012 Jul;30(7):1424-35
references.pmid: 22570183
references.citation: Hum Gene Ther. 2007 Dec;18(12):1182-92
references.pmid: 18021021
references.citation: Nat Med. 2013 Aug;19(8):998-1004
references.pmid: 23921754
references.citation: Nat Biotechnol. 2011 Jan;29(1):73-8
references.pmid: 21151124
references.citation: Mol Ther. 2007 Jul;15(7):1248-52
references.pmid: 17505474
references.citation: Nat Biotechnol. 2011 Aug 14;29(9):829-34
references.pmid: 21841799
references.citation: Biomaterials. 2012 Apr;33(11):3195-204
references.pmid: 22269649
references.citation: Cell Stem Cell. 2013 Feb 7;12(2):167-79
references.pmid: 23318055
references.citation: Stem Cells. 2015 Jan;33(1):91-100
references.pmid: 25330775
references.citation: Nat Rev Cancer. 2011 Apr;11(4):268-77
references.pmid: 21390058
references.citation: Sci Transl Med. 2012 Dec 12;4(164):164ps25
references.pmid: 23241742
references.citation: Cell Rep. 2014 Sep 25;8(6):1677-85
references.pmid: 25242333
references.citation: Nature. 2011 Nov 06;480(7378):547-51
references.pmid: 22056989
references.citation: Mol Ther Methods Clin Dev. 2014 Nov 12;1:14053
references.pmid: 26052521
references.citation: Blood. 2004 Dec 1;104(12):3543-9
references.pmid: 15308565
references.citation: Cell Cycle. 2009 Aug 15;8(16):2608-12
references.pmid: 19597339
references.citation: Proc Natl Acad Sci U S A. 2009 Aug 4;106(31):12759-64
references.pmid: 19549847
references.citation: Mol Ther. 2011 Sep;19(9):1667-75
references.pmid: 21587213
references.citation: Mol Ther. 2015 Sep;23(9):1475-85
references.pmid: 26022733
references.citation: Nat Methods. 2009 May;6(5):370-6
references.pmid: 19404254
references.citation: Rheumatology (Oxford). 2003 Jan;42(1):162-5
references.pmid: 12509630