title: Comparative Analysis of Human Nucleoside Kinase-Based Reporter Systems for PET Imaging.
abstract: Radionuclide-based reporter gene imaging has the sensitivity to monitor gene- and cell-based therapies in human subjects. Potential immunogenicity of current viral transgenes warrants development of human-based reporter systems. We compared human nucleoside kinase reporters to a panel of nucleoside analogs of FEAU, FMAU, and FIAU, including the first in vivo assessment of L-[18F]FEAU.
  
  
  
  Human isogenic U87 cell lines were transduced to express different human reporter genes including dCK-R104M/D133A (dCKDM), dCK-R104Q/D133N (dCKep16A), dCK-A100V/R104M/D133A (dCK3M), and TK2-N93D/L109F (TK2DM), and wild-type dCK (dCK) and herpes simplex virus type-1 (HSVTK) reporter gene as references. In vitro cell uptake assays were performed with [18F]FEAU, L-[18F]FEAU, [14C]FMAU, L-[18F]FMAU, and [124I]FIAU. Micro-positron emission tomography/X-ray computed tomography imaging of xenograft-bearing nu/nu mice was conducted with [18F]FEAU, L-[18F]FEAU, L-[18F]FMAU, and [124I]FIAU on consecutive days. A cell viability assay was also performed to assess sensitivities to gemcitabine and bromovinyldeoxyuridine (BVdU).
  
  
  
  In vitro, dCKep16A and dCKDM with [18F]FEAU exhibited the highest sensitivity and selectivity of the human reporters, second only to HSVTK/[18F]FEAU. L-[18F]FEAU biodistribution in mice was on par with [18F]FEAU and L-[18F]FMAU. L-[18F]FMAU uptake in isogenic xenografts was highest for all human reporter genes. However, [18F]FEAU was the most selective of the short half-life reporter probes due to its minimal recognition by human dCK and relative sensitivity, whereas [124I]FIAU permitted imaging at a later time point, improving signal-to-background ratio. Of the human reporter genes, dCKep16A consistently outperformed the other tested reporters. Reporter genes of interest increased potency to the nucleoside analog prodrugs gemcitabine and BVdU.
  
  
  
  We demonstrate that human nucleoside kinase reporter systems vary significantly in their sensitivity and selectivity for in vivo imaging. The sufficiently high signal-to-background ratios and enhanced suicide gene potential support clinical translation.
authors.affiliation: Crump Institute for Molecular Imaging, Department of Molecular and Medical Pharmacology, David Geffen School of Medicine at UCLA, California NanoSystems Institute Rm 2151, 570 Westwood Plaza, Los Angeles, CA, 90095, USA.|Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Jason T
authors.initials: JT
authors.lastname: Lee
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Hanwen
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Maxim A
authors.initials: MA
authors.lastname: Moroz
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Yury
authors.initials: Y
authors.lastname: Likar
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Larissa
authors.initials: L
authors.lastname: Shenker
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Nikita
authors.initials: N
authors.lastname: Sumzin
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Jose
authors.initials: J
authors.lastname: Lobo
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA.
authors.forename: Juan
authors.initials: J
authors.lastname: Zurita
authors.affiliation: Crump Institute for Molecular Imaging, Department of Molecular and Medical Pharmacology, David Geffen School of Medicine at UCLA, California NanoSystems Institute Rm 2151, 570 Westwood Plaza, Los Angeles, CA, 90095, USA.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Collins
authors.affiliation: Crump Institute for Molecular Imaging, Department of Molecular and Medical Pharmacology, David Geffen School of Medicine at UCLA, California NanoSystems Institute Rm 2151, 570 Westwood Plaza, Los Angeles, CA, 90095, USA.
authors.forename: R Michael
authors.initials: RM
authors.lastname: van Dam
authors.affiliation: Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA. ponomarv@mskcc.org.|Sloan Kettering Institute, Molecular Pharmacology and Chemistry Program, Memorial Sloan Kettering Cancer Center, New York, NY, 10065, USA. ponomarv@mskcc.org.|Molecular Imaging Laboratory, Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Ave, New York, NY, 10065, USA. ponomarv@mskcc.org.
authors.forename: Vladimir
authors.identifier: 0000-0002-9119-9128
authors.initials: V
authors.lastname: Ponomarev
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D017930:Genes, Reporter; D041722:Genes, Transgenic, Suicide; D006801:Humans; D008819:Mice, Nude; D009154:Mutation / Q000235:genetics; D010770:Phosphotransferases / Q000378:metabolism*; D049268:Positron-Emission Tomography / Q000379:methods*; D013936:Thymidine / Q000378:metabolism; D014018:Tissue Distribution
chemical_list: D010770:Phosphotransferases; C021892:nucleoside phosphotransferase; D013936:Thymidine
keywords: Deoxycytidine kinase; Gene and cell therapy; Human reporter gene imaging; Immunotherapy; L-[18F]FEAU; L-[18F]FMAU; Thymidine kinase; [124I]FIAU; [14C]FMAU; [18F]FEAU