Role of Equilibrative Nucleobase Transporter 1/SLC43A3 as a Ganciclovir Transporter in the Induction of Cytotoxic Effect of Ganciclovir in a Suicide Gene Therapy with Herpes Simplex Virus Thymidine Kinase.
A suicide gene therapy using herpes simplex virus thymidine kinase (HSV-TK) with ganciclovir (GCV) has been under development as a tumor-targeted therapy; however, the mechanism of cellular GCV uptake, which is prerequisite in the therapy, has not been clarified. In an attempt to resolve this situation and gain information to optimize HSV-TK/GCV system for cancer therapy, we found that human equilibrative nucleobase transporter 1 (ENBT1) can transport GCV with a Michaelis constant of 2.75 mM in Madin-Darby canine kidney II (MDCKII) cells stably transfected with this transporter. In subsequent experiments using green fluorescent protein (GFP)-tagged ENBT1 (GFP-ENBT1) and HSV-TK, the uptake of GCV (30 μM), which was minimal in MDCKII cells and unchanged by their transfection with HSV-TK alone, was increased extensively by their transfection with GFP-ENBT1, together with HSV-TK. Accordingly, cytotoxicity, which was assessed by the WST-8 cell viability assay after the treatment of those cells with GCV (30 μM) for 72 hours, was induced in those transfected with GFP-ENBT1, together with HSV-TK but not in those transfected with HSV-TK alone. These results suggest that ENBT1 could facilitate GCV uptake and thereby enhance cytotoxicity in HSV-TK/GCV system. We also identified Helacyton gartleri (HeLa) and HepG2 as cancer cell lines that are rich with ENBT1 and A549, HCT-15 and MCF-7 as those poor with ENBT1. Accordingly, the HSV-TK/GCV system was effective in inducing cytotoxicity in the former but not in the latter. Thus, ENBT1 was found to be a GCV transporter that could enhance the performance of HSV-TK/GCV suicide gene therapy.
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya (J.F., T.Y., Y.M., H.Y.); College of Pharmacy, Kinjo Gakuin University, Nagoya (K.O.); and Department of Biopharmaceutics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan (K.I.).
Junji
J
Furukawa
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya (J.F., T.Y., Y.M., H.Y.); College of Pharmacy, Kinjo Gakuin University, Nagoya (K.O.); and Department of Biopharmaceutics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan (K.I.) kinoue@toyaku.ac.jp.
Katsuhisa
K
Inoue
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya (J.F., T.Y., Y.M., H.Y.); College of Pharmacy, Kinjo Gakuin University, Nagoya (K.O.); and Department of Biopharmaceutics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan (K.I.).
Kinya
K
Ohta
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya (J.F., T.Y., Y.M., H.Y.); College of Pharmacy, Kinjo Gakuin University, Nagoya (K.O.); and Department of Biopharmaceutics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan (K.I.).
Tomoya
T
Yasujima
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya (J.F., T.Y., Y.M., H.Y.); College of Pharmacy, Kinjo Gakuin University, Nagoya (K.O.); and Department of Biopharmaceutics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan (K.I.).
Yoshihisa
Y
Mimura
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya (J.F., T.Y., Y.M., H.Y.); College of Pharmacy, Kinjo Gakuin University, Nagoya (K.O.); and Department of Biopharmaceutics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan (K.I.).
Hiroaki
H
Yuasa
D026905:Amino Acid Transport Systems / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D001692:Biological Transport; D002460:Cell Line; D004285:Dogs; D015774:Ganciclovir / Q000378:metabolism* / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D015316:Genetic Therapy; D006801:Humans; D018139:Simplexvirus / Q000201:enzymology* / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics*
D026905:Amino Acid Transport Systems; C000605140:SLC43A3 protein, human; D013937:Thymidine Kinase; D015774:Ganciclovir