Optimized human CYP4B1 in combination with the alkylator prodrug 4-ipomeanol serves as a novel suicide gene system for adoptive T-cell therapies.
Engineering autologous or allogeneic T cells to express a suicide gene can control potential toxicity in adoptive T-cell therapies. We recently reported the development of a novel human suicide gene system that is based on an orphan human cytochrome P450 enzyme, CYP4B1, and the naturally occurring alkylator prodrug 4-ipomeanol. The goal of this study was to systematically develop a clinically applicable self-inactivating lentiviral vector for efficient co-expression of CYP4B1 as an ER-located protein with two distinct types of cell surface proteins, either MACS selection genes for donor lymphocyte infusions after allogeneic stem cell transplantation or chimeric antigen receptors for retargeting primary T cells. The U3 region of the myeloproliferative sarcoma virus in combination with the T2A site was found to drive high-level expression of our CYP4B1 mutant with truncated CD34 or CD271 as MACS suitable selection markers. This lentiviral vector backbone was also well suited for co-expression of CYP4B1 with a codon-optimized CD19 chimeric antigen receptor (CAR) construct. Finally, 4-ipomeanol efficiently induced apoptosis in primary T cells that co-express mutant CYP4B1 and the divergently located MACS selection and CAR genes. In conclusion, we here developed a clinically suited lentiviral vector that supports high-level co-expression of cell surface proteins with a potent novel human suicide gene.
Department of Otorhinolaryngology and Head/Neck Surgery, Heinrich Heine University, Düsseldorf, Germany.
K
K
Roellecke
Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN, USA.
E L
EL
Virts
Institute for Organic Chemistry, University of Tübingen, Tübingen, Germany.
R
R
Einholz
Department of Medicinal Chemistry, School of Pharmacy, University of Washington, Seattle, WA, USA.
K Z
KZ
Edson
Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, Muenster, Germany.
B
B
Altvater
Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, Muenster, Germany.
C
C
Rossig
Institute for Virology, Innsbruck Medical University, Innsbruck, Austria.
D
D
von Laer
Department of Otorhinolaryngology and Head/Neck Surgery, Heinrich Heine University, Düsseldorf, Germany.
K
K
Scheckenbach
Department of Otorhinolaryngology and Head/Neck Surgery, Heinrich Heine University, Düsseldorf, Germany.
M
M
Wagenmann
Department of Pediatrics III, University Children's Hospital Essen, University Duisburg-Essen, Essen, Germany.
D
D
Reinhardt
Division of Pediatric Hematology and Oncology, Department of Child and Adolescent Health, University Medical Center Göttingen, Göttingen, Germany.
C M
CM
Kramm
Department of Medicinal Chemistry, School of Pharmacy, University of Washington, Seattle, WA, USA.
A E
AE
Rettie
Department of Otorhinolaryngology and Head/Neck Surgery, Heinrich Heine University, Düsseldorf, Germany.
C
C
Wiek
Department of Otorhinolaryngology and Head/Neck Surgery, Heinrich Heine University, Düsseldorf, Germany.|Department of Pediatrics III, University Children's Hospital Essen, University Duisburg-Essen, Essen, Germany.
H
H
Hanenberg
D018952:Antigens, CD34 / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D041722:Genes, Transgenic, Suicide; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D019169:Jurkat Cells; D016086:Lentivirus / Q000235:genetics; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000378:metabolism; D013729:Terpenes / Q000627:therapeutic use
D018952:Antigens, CD34; C000605956:CD19-specific chimeric antigen receptor; D011948:Receptors, Antigen, T-Cell; D013729:Terpenes; D001189:Aryl Hydrocarbon Hydroxylases; C095029:cytochrome P-450 CYP4B1; C001470:4-ipomeanol