Retroviral transduction of cancer cell lines with the gene encoding Drosophila melanogaster multisubstrate deoxyribonucleoside kinase.
Nucleoside kinases from several species are investigated as "suicide genes" for treatment of malignant tumors by combined gene/chemotherapy. We have recently cloned a multisubstrate deoxyribonucleoside kinase of Drosophila melanogaster (Dm-dNK), and we have shown that the enzyme phosphorylates cytotoxic pyrimidine and purine nucleoside analogs. The broad substrate specificity of the enzyme, as well as its very high catalytic rate, makes it a unique member of the nucleoside kinase enzyme family. In the present study, we evaluated Dm-dNK as a suicide gene by constructing a replication-deficient retroviral vector that expresses the enzyme. The human pancreatic adenocarcinoma cell line MIA PaCa-2 and a thymidine kinase-deficient osteosarcoma cell line were transduced with the recombinant virus. We showed that Dm-dNK can be expressed in human cells, that the enzyme retained its enzymatic activity, and that it is localized in the cell nuclei due to a nuclear localization signal in its C-terminal region. The cells expressing Dm-dNK exhibited increased sensitivity to several cytotoxic nucleoside analogs, such as 1-beta-d-arabinofuranosylcytosine, 1-beta-d-arabinofuranosylthymine, (E)-5-(2-bromovinyl)-2'-deoxyuridine, 2-chloro-2'-deoxyadenosine, and 2',2'-difluorodeoxycytidine. These findings suggest that Dm-dNK may be used as a suicide gene in combined gene/chemotherapy of cancer.
Zheng
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X
Karolinska Institute, Division of Clinical Virology, Huddinge University Hospital, S-141 86 Stockholm, Sweden.
Johansson
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M
Karlsson
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A
D000230:Adenocarcinoma / Q000378:metabolism; D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology; D000998:Antiviral Agents / Q000494:pharmacology; D001087:Arabinonucleosides / Q000494:pharmacology; D001973:Bromodeoxyuridine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002384:Catalysis; D002455:Cell Division / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism; D017338:Cladribine / Q000494:pharmacology; D003561:Cytarabine / Q000494:pharmacology; D003841:Deoxycytidine / Q000031:analogs & derivatives / Q000494:pharmacology; D004331:Drosophila melanogaster / Q000235:genetics*; D006801:Humans; D020128:Inhibitory Concentration 50; D007700:Kinetics; D012516:Osteosarcoma / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000378:metabolism; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012190:Retroviridae / Q000378:metabolism*; D013379:Substrate Specificity; D013936:Thymidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013937:Thymidine Kinase / Q000378:metabolism; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured; D000093542:Gemcitabine
D000964:Antimetabolites, Antineoplastic; D000970:Antineoplastic Agents; D000998:Antiviral Agents; D001087:Arabinonucleosides; D003561:Cytarabine; D003841:Deoxycytidine; C020235:brivudine; D017338:Cladribine; D017853:Phosphotransferases (Alcohol Group Acceptor); C020018:deoxyribonucleoside kinases; D013937:Thymidine Kinase; D001973:Bromodeoxyuridine; C010032:thymine arabinoside; D013936:Thymidine; D000093542:Gemcitabine