Comparison of [14C]FMAU, [3H]FEAU, [14C]FIAU, and [3H]PCV for monitoring reporter gene expression of wild type and mutant herpes simplex virus type 1 thymidine kinase in cell culture.
OBJECTIVE
To assess the optimal reporter probe/reporter gene combination for monitoring herpes simplex virus type 1 thymidine kinase (HSV1-tk) gene expression, we compared the cellular uptake of 1-(2'-fluoro-2'-deoxy-D-arabinofuranosyl)-5-methyluracil (FMAU), 2'-fluoro-2'-deoxyarabinofuranosyl-5-ethyluracil (FEAU), 2'-fluoro-2'-deoxy-beta-D-arabinofuranosyl-5-iodouracil (FIAU) and penciclovir (PCV) in both HSV1-tk and HSV1-sr39tk expressing cells.


METHODS
For stably transfected cell studies, C6 rat glioma cells, C6 HSV1-tk transfectant, C6 mutant HSV1-sr39tk transfectant, rat Morris hepatoma cells (MH3924A), and MH3924A HSV1-tk transfectant cells were used. For adenoviral infection studies, C6 rat glioma cells were exposed to serial titers of AdCMV-HSV1-tk, AdCMV-HSV1-sr39tk, or AdCMV-fluc for 24 hours. These cells were incubated with [(14)C]FMAU, [(3)H]FEAU, [(14)C]FIAU, and [(3)H]PCV, and cellular uptake of radioactivity was measured.


RESULTS
[(3)H]FEAU exhibited the highest or second highest accumulation and the most selectivity regardless of the mode of gene transfer for both HSV1-tk and mutant HSV1-sr39tk reporter genes.


CONCLUSIONS
This combination of high accumulation and high selectivity for both HSV1-tk and HSV1-sr39tk makes suitably radiolabeled FEAU a promising candidate as a radiotracer for imaging HSV1-tk/HSV1-sr39tk gene expression in living subjects.
Kang
Keon Wook
KW
Department of Radiology, Bio-X Program, Stanford University, Stanford, CA 94305, USA.
Min
Jung-Joon
JJ
Chen
Xiaoyuan
X
Gambhir
Sanjiv S
SS
D000212:Acyclovir / Q000031:analogs & derivatives* / Q000032:analysis; D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001086:Arabinofuranosyluracil / Q000031:analogs & derivatives* / Q000032:analysis; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D006147:Guanine; D018259:Herpesvirus 1, Human / Q000201:enzymology; D008156:Luciferases / Q000235:genetics; D009154:Mutation / Q000235:genetics; D011868:Radioisotopes; D051381:Rats; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection
D011868:Radioisotopes; D001086:Arabinofuranosyluracil; C053539:penciclovir; C043457:fialuridine; D006147:Guanine; C054103:2'-fluoro-5-ethylarabinosyluracil; D008156:Luciferases; D013937:Thymidine Kinase; C034935:clevudine; D000212:Acyclovir