brivudine and Stomach-Neoplasms

Deoxyribonucleoside kinase of Drosophila melanogaster (Dm-dNK) mutants have been reported to exert suicide gene effects in combined gene/chemotherapy of cancer. Here, we aimed to further evaluate the capacity of the mutanted enzyme and its potential for inhibiting cancer cell growth.. We altered the sequence of the last 10 amino acids of Dm-dNK to perform site-directed mutagenesis and constructed active site mutanted Dm-dNK (Dm-dNKmut), RT-PCR and western bloting studies were used to reveal the expression of lentivirus mediated Dm-dNKmut in a breast cancer cell line (Bcap37), a gastric cancer cell line (SGC7901) and a colorectal cancer cell line (CCL187). [3H]-labeled substrates were used for enzyme activity assays, cell cytotoxicity was assessed by MTT assays, cell proliferation using a hemocytometer and apoptosis induction by thenannexin-V-FITC labeled FACS method. In vivo, an animal study was set out in which BALB/C nude mice bearing tumors were treated with lentivirus mediated expression of Dm-dNKmut with the pyrimidine nucleoside analog brivudine (BVDU, (E)-5-(2-bromovinyl)-(2-deoxyuridine).. The Dm-dNKmut could be stably expressed in the cancer cell lines and retained its enzymatic activity. Moreover, the cells expressing Dm-dNKmut exhibited increased sensitivity in combination with BVDU, with induction of apoptosis in vitro and in vivo.. These findings underlined the importance of BVDU phosphorylated by Dm-dNKmut in transduced cancer cells and the potential role of Dm-dNKmut as a suicide gene, thus providing the basis for future intensive research for cancer therapy.

Topics: Amino Acid Sequence; Animals; Antiviral Agents; Apoptosis; Blotting, Western; Breast Neoplasms; Bromodeoxyuridine; Cell Proliferation; Colorectal Neoplasms; Combined Modality Therapy; Drosophila melanogaster; Female; Flow Cytometry; Genetic Therapy; Genetic Vectors; Humans; Lentivirus; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutant Proteins; Mutation; Phosphotransferases (Alcohol Group Acceptor); Prohibitins; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sequence Homology, Amino Acid; Stomach Neoplasms; Tumor Cells, Cultured

We developed a rapid and simple method for the screening of antiviral agents against herpes simplex virus (HSV) in a model of gastrointestinal herpetic infection in vitro. This method was based on inhibition of HSV-induced cytopathogenicity in gastric adenocarcinoma MKN-28 cells, as monitored by an MTT colorimetric assay. From the various compounds that were evaluated for their activity against HSV-1 and HSV-2, brivudine (BVDU) emerged as the most effective. When the 50% effective concentration (EC50) values of the antiherpes agents in MKN-28 cells were compared with those in human embryo lung MRC-5 cells, all compounds, except for BVDU, showed higher EC50 values in MKN-28 cells. For BVDU the EC50 values in MKN-28 cells were 0.8 (HSV-1) and 0.036 (HSV-2) times the EC50 values in MRC-5 cells. Thus BVDU was 27.5 times more active against HSV-2 in MKN-28 cells than in MRC-5 cells. The MKN-28 gastric cancer cells may be useful for the rapid screening of anti-HSV agents and, in particular, those that may be useful in therapy of gastrointestinal HSV infections in gastrointestinal herpetic infection.

Topics: Adenocarcinoma; Antiviral Agents; Bromodeoxyuridine; Colorimetry; Coloring Agents; Drug Evaluation, Preclinical; Herpesvirus 1, Human; Herpesvirus 2, Human; Humans; Stomach Neoplasms; Tetrazolium Salts; Thiazoles; Tumor Cells, Cultured; Virus Replication