Molecular–Genetic PET Imaging Using an HSV1-tk Mutant Reporter Gene with Enhanced Specificity to Acycloguanosine

Imaging 2 different molecular–genetic events in a single subject by PET is essential in a variety of in vivo applications. Using her-pes simplex virus-1 thymidine kinase (HSV1-tk) mutants with narrower substrate specificities in combination with wild-type HSV1-tk (wtHSV1-tk) would enable differential imaging with corresponding radiotracers, namely 29-deoxy-29-18F-fluoro-5-ethyl-1-b-D-arabinofuranosyl-uracil (18F-FEAU) and the acyclo-guanosine derivative 9-(4-18F-fluoro-3-[hydroxymethyl]butyl)-guanine (18F-FHBG). In this study, we evaluated wtHSV1-tk and the A168H mutant, which has been reported to exhibit en-hanced acycloguanosine substrate catalytic activity and dimin-ished pyrimidine phosphorylating activity, as PET reporter genes. Methods: Computational analysis was performed to as-sess the binding mode of FHBG and FEAU to wtHSV1-tk and the A168H variant. U87 cells were stably transduced with wtHSV1-t