19. Investigative Treatment Division

The main goal of the research conducted in this division is developing new strategies for cancer prevention, diagnosis and treatment based on a better understanding of the biology of cancer tissues and the interaction between cancer and host. Improvement of preexistent modalities of cancer diagnosis and treatment is also within the scope of this research activity.

Analysis of Precancerous Lesions

K-ras activation appears to play a crucial role in the initiation of colonic carcinogenesis in some fraction of colon cancer cases.⁽¹⁸⁵⁾ Little is known about the function of activated K-ras in epithelial cells. In order to elucidate this function, the biological effects of K-ras activation were examined in rat primary colonic epithelial cells. Expression vectors containing two types of activated K-ras genes and the wild type gene were constructed using a viral vector. When activated K-ras was introduced into colonic epithelial cells, these cells were morphologically transformed and showed anchorage independent growth and tumorigenicity in nude mice while the wild type gene produced neither transformation nor anchorage independence. As reported previously, activated K-ras alone was not sufficient for full transformation of rat primary fibroblasts. Cell fusion analysis of the effect on transformation showed that fibroblasts were not transformation suppressive. Unknown transformation dominant factors in epithelial cells are being investigated. To clarify the significance of specific neoplastic lesions in carcinogenesis, the establishment of an animal model of carcinogenesis is very important. Dogs were administered 50 mg of ENNG as a suppository daily for 5 months and, 3 months later, aberrant crypt foci were detected under magnifying colonoscopy. The number of aberrant crypt foci increased up to 12 months despite ENNG administration having been discontinued at 5 months. The number then decreased gradually through 24 months. Nineteen tumorous lesions had formed in two dogs by 20 months and one of the lesions was confirmed to be invasive cancer histologically. Seventeen of the 19 tumors were never polypoid at any point in the course of tumor formation. This canine model is a good potential model of flat-type adenocarcinoma of the colon, the significance of which is still controversial. Analysis of the clonality of minute lesions is informative for determining whether or not they are neoplastic. X-chromosome linked gene analysis has been utilized for this purpose. When the clonality of single fundic and pyrolic glands in the human stomach was analyzed by HUMARA and PGK gene methylation, most of the pyrolic glands were monoclonal, as expected. However, the majority of fundic glands were found to be polyclonal.⁽¹⁸⁶⁾ Once intestinal metaplasia had occurred in a pyloric region, individual metaplastic glands showed polyclonality suggesting either migration of stem cells to the metaplastic glands from outside of the pyrolic gland or instability of X-chromosome methylation.⁽¹⁸⁷⁾

Roles of Nitric Oxide in Carcinogenesis and the Clinical Course of Cancer

We have studied the molecular mechanisms of gene regulation of the nitric oxide synthases and the biochemical consequences of nitric oxide production.^(188-190) We analyzed the mechanisms of iNOS gene induction in the human neuroblastoma cell line NB-39-nu.⁽¹⁹¹⁾ The results showed that the iNOS gene is regulated by TNF-alpha through the TNF type II receptor. Most of the genes encoding inflammatory cytokines, regulated by TNF-alpha, are known to be regulated by NF-kappaB activation through the type I receptor. Although iNOS induction by TNF-alpha and IFN-gamma can be completely inhibited by dexamethasone in this cell line and iNOS induction by antimicrotuble anticancer drugs was shown to be dependent on TNF-alpha production by these drugs, iNOS induction by these drugs was not inhibited by dexamethasone. Promoter analysis of the iNOS gene and gel shift assay of the NF-kappaB binding site suggested that some silencing factors are present in the cells and the existence of a new pathway of iNOS induction is suspected. Among the many biological roles of nitric oxide, blood vessel dilatation is the major recognized function. With regard to the hypoxic reaction which is critical for tumor progression, we have analyzed the effect of nitric oxide on the hypoxia response of gene expression. When several tumor cell lines were treated with nitric oxide generating compounds, VEGF gene expression was remarkably elevated. Together with VEGF expression, some hypoxia response gene expressions were also enhanced. These findings suggest that nitric oxide produced by either tumor cells or surrounding tissues could be a factor determining tumor vascular formation. When several tumor cell lines were treated with nitric oxide, they were found to become more resistant to apoptosis induced by vincristine, vinblastine and paclitaxel. These observations suggest the possibility that nitric oxide is an important factor modulating the clinical course of cancer in addition to the process of carcinogenesis.^(192-195)

List of papers from this division

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