Molecular therapy for peritoneal dissemination of xenotransplanted human MKN-45 gastric cancer cells with adenovirus mediated Bax gene transfer.

BACKGROUND: Gene therapy is an innovative therapeutic approach for cancer. An adenoviral vector expressing the tumour suppressor p53 gene (Ad/p53) is currently under clinical evaluation for various cancers. We recently developed a binary adenoviral vector system that can express the strong proapoptotic gene Bax (Ad/PGK-GV16+Ad/GT-Bax: Ad/Bax). AIMS: To evaluate the potential of Bax gene therapy for gastric cancer, we assessed its antitumour effect in comparison with that of p53. METHODS: The human gastric cancer cell lines MKN-1, MKN-7, MKN-28, and MKN-45 were treated with Ad/Bax or Ad/p53, and cell viability, transgene expression, and caspase activation were assessed in vitro. To compare the antitumour effects of Ad/Bax and Ad/p53 treatment in vivo, subcutaneous tumours and peritoneal dissemination of MKN-45 cells were generated in nude mice. Each mouse underwent intratumoral or intraperitoneal administration of viruses and the growth of implanted tumours was observed after treatment. RESULTS: Treatment with Ad/Bax and Ad/p53 resulted in marked Bax and p53 protein expression and effective apoptosis induction in MKN-1, MKN-7, and MKN-28 cells in vitro. In contrast, MKN-45 cells showed resistance to Ad/p53 and only treatment with Ad/Bax resulted in activation of caspase 3 expression and massive apoptosis. Ad/Bax treatment was more effective in suppressing both subcutaneous and peritoneally disseminated MKN-45 tumours compared with Ad/p53 treatment. CONCLUSION: Ad/Bax treatment significantly inhibited the growth of even p53 resistant gastric cancer in vitro and in vivo. Therefore, adenovirus mediated Bax gene transfer may be useful in gene therapy for gastric cancers.

A novel method for gene delivery and expression in esophageal epithelium with fibrin glues containing replication-deficient adenovirus vector.

BACKGROUND: Gene transfer to the esophageal epithelium holds the potential for the therapy of malignant as well as premalignant lesions in the upper gastrointestinal tract. Replication-deficient recombinant adenoviruses represent an efficient means of introducing genes in vivo into cells in a variety of organs. The majority of in vivo studies utilize direct submucosal injection for delivery of the viral vectors into the locoregional area of the gut; transferring genes into epithelial cells, however, is difficult because viruses are retained in the subepithelial space. To establish the efficient method for gene transfer into the epithelial cells, we have developed a multiluminal spray catheter that can be passed through the accessory channel of an endoscope, and we have evaluated the feasibility of fibrin glues as a vehicle of recombinant adenoviruses in a porcine model. METHODS: The fibrinogen solution and the thrombion solution containing an E1/E3 deleted recombinant adenovirus expressing the bacterial lacZ gene (Ad-lacZ) were endoscopically sprayed on the porcine esophagus through the catheter attached to the dual-barrel syringe. Twenty-four hours after gene delivery, beta-galactosidase activity of the esophagus was determined under the microscope following X-gal staining. RESULTS: The fibrin glue could be locally sprayed on the porcine esophagus by using the multichannel catheter through the endoscope. Attachment of the fibrin glue comtining Ad-lacZ caused strong beta-galactosidase staining on epithelial cells in the mucosal surface, but not in the basal cell layer. CONCLUSION: Endoscopic local delivery of recombinant adenoviruses in aerosolized fibrin glues through a multiluminal catheter could provide an optimal technique for gene transfer into epithelial cells in the mucosal surfece, which may have important implications for the treatment of human esophageal premalignant diseases.

A family of multiple endocrine neoplasia type 2A with the RET proto-oncogene mutation in codon 618 (Cys-->Arg).

Multiple endocrine neoplasia type 2 (MEN-2) is a hereditary syndrome characterized by medullary thyroid carcinoma (MTC), pheochromocytoma and hyperplasia or adenoma of the parathyroid gland with hyperparathyroidism. Recent genetic studies have identified the presence of germline missense mutations in the RET proto-oncogene in almost 100% of MEN-2 patients. We report here three generations of one MEN-2 family with rare missense mutation at codon 618 (Cys-->Arg) of the RET proto-oncogene. The first patient was surgically treated at the age of 63 years but died of bone metastasis. His two children (29-year-old daughter and 25-year-old son) were treated surgically for MTC and neck lymph node metastasis. Germline mutations of the RET proto-oncogene of these three MTC patients and two children of the 29-year-old daughter (9-year-old female and 7-year-old male) were examined. Three MTC patients and the 9-year-old female possessed the mutation. The phenotype of the family with this rare point mutation of the RET proto-oncogene is reported.

Primary structure and phylogenetic analysis of the coat protein of a Toyama isolate of tobacco necrosis virus.

The amino acid sequence of the coat protein (CP) of a tobacco necrosis virus (TNV) strain, Toyama isolate, was determined by a combination of peptide and cDNA sequencing. The deduced sequence of 276 residues was compared with CPs of other TNV isolates and other plant virus isolates of Tombusviridae. It showed the highest similarity to the TNV Nebraska isolate with 92% identity and moderate similarity to the TNV strain A with 51% identity, confirming the previous serological analysis. It also showed overall similarity with CPs of mostly genera Necrovirus and Sobemovirus, and partial similarity with CPs of genera Tombusvirus and Carmovirus. Among 13 CPs that showed overall similarity, there were 10 completely conserved residues. These included three residues that participate in Ca2+ ligation at the interfaces of virion subunits in TNV crystal structure, suggesting that similar metal binding occur in the viruses of genera Necrovirus and Sobemovirus.