Biology Division

4. Biology Division

　To understand the molecular basis of human carcinogenesis, the positional as well as functional cloning of cancer-related genes are underway in the Biology Division. To date, several genes including the human OGG1 gene involved in DNA repair and the mouse Elm1 gene involved in metastasis, have been isolated in the Division. Functional analyses of those genes and other cancer-related genes are also in progress.
Genetic Alterations in Human Cancers

　The identification of homozygous deletions in sporadic cancers has been pivotal in the positional cloning of several tumor suppressor genes. By a deletion mapping and candidate gene screening, homozygous deletions were identified in novel regions on chromosomes 9p, 18q and 21q in lung cancers. ^(64-67) These results suggest that novel tumor suppressor genes are harbored in those chromosomal regions. Micro-cell transfer studies indicated the presence of a telomerase repressor gene in the 3p14.2-p21.1 region.⁽⁶⁸⁾ CpG islands hypermethylated in human lung cancer were identified by a modification of the arbitrarily primed-PCR method.⁽⁶⁹⁾ Expression of the Fhit protein was associated with prognosis of patients with lung cancer,⁽⁷⁰⁾ while expression of p16 and DCC proteins correlated with progression of neuroblastoma.^(71,72) The Smad2 gene was not mutated in colon cancer, while the PTEN gene was mutated in a small subset of lung cancers.^(73,74) The ABI-1 gene was identified as being fused with the MLL gene in acute myeloid leukemia with t(10;11)(p11.2;q23),⁽⁷⁵⁾ and a novel variant of chronic myelomonocytic leukemia carrying t(3;12)(q26;p13) with characteristics of 3q21q26 syndrome was identified.⁽⁷⁶⁾
Functional Analysis of Cancer-related Genes

　There are several projects in the Division to elucidate the pathogenetic significance of tumor suppressor gene inactivation in human cancer. Phosphorylation mechanisms of the p53 protein were investigated in association with its function,^(77-80) and it was found that the ATM protein phosphorylates the amino terminus of the p53 protein. A tetracycline-dependent inducible system for the expression of the p53 gene revealed that cell cycle arrest and apoptosis were differentially regulated by the level of p53 expression in a small cell lung carcinoma cell line.⁽⁸¹⁾ Sp1-mediated transcription of the Werner helicase gene was modulated by RB and p53.⁽⁸²⁾
Characterization of a Gene Involved in the Repair of Oxidative DNA Damage, 8-hydroxyguanine

　To elucidate the involvement of DNA repair system for 8-hydroxyguanine, a major form of oxygen-free radical-induced DNA damage, in human carcinogenesis, molecular analyses of the human OGG1 (oxoguanine DNA glycosylase) gene were performed.^(83-85) Although OGG1 mutations rarely occur in lung and gastric cancers, a genetic polymorphism resulting in different enzymatic activity of the OGG1 protein was identified. The genotype with a low enzymatic activity was more frequently detected in patients with lung and gastric cancers, suggesting that the OGG1 activity is involved in human carcinogenesis.
Identification of a Metastasis Suppressor Gene and Construction of a Model for the Pleural Metastasis of Lung Cancer

　Eight genes were identified as being differentially expressed in association with the metastatic potential of K-1735 mouse melanoma cells by the mRNA differential display method. Molecular cloning and sequence analysis revealed that one of genes expressed in low metastatic cells, Elm1, is a novel member of the CCN family genes.⁽⁸⁶⁾ Expression of the Elm1 gene in highly metastatic K-1735 cells resulted in the suppression of in vivo growth and metastasis, suggesting that Elm1 is a metastasis suppressor gene. Injection of human non-small cell carcinoma cells into the pleural cavity of nude mice led to frequent intrapleural growth and metastasis.⁽⁸⁷⁾ Since this method is easy to perform and the results are reproducible, it can be applied to biological studies of human lung cancer cells in vivo.
Molecular Epidemiology for Cancer Susceptibility

　Since it has been shown that genomic instability is present in a variety of cancer cells and in cancer-prone families, the prevalence of microsatellite instability was investigated in gastric cancer-prone families. Microsatellite instability was infrequently detected in gastric cancer of patients with family histories of gastric cancer aggregation, suggesting that inherited disorders in mismatch repair systems do not largely contribute to high susceptibility to gastric cancers.⁽⁸⁸⁾ Germline mutations of the PTEN gene were identified in three Japanese patients with Cowden disease.⁽⁸⁹⁾ Families with aggregation of lung cancer were not identified in the analysis of 1068 lung cancer patients, suggesting that familial lung cancer is rare in the Japanese population.⁽⁹⁰⁾




	4. Biology Division



		To understand the molecular basis of human carcinogenesis, the positional as well as functional cloning of cancer-related genes are underway in the Biology Division. To date, several genes including the human OGG1 gene involved in DNA repair and the mouse Elm1 gene involved in metastasis, have been isolated in the Division. Functional analyses of those genes and other cancer-related genes are also in progress. Genetic Alterations in Human Cancers 　The identification of homozygous deletions in sporadic cancers has been pivotal in the positional cloning of several tumor suppressor genes. By a deletion mapping and candidate gene screening, homozygous deletions were identified in novel regions on chromosomes 9p, 18q and 21q in lung cancers. ^(64-67) These results suggest that novel tumor suppressor genes are harbored in those chromosomal regions. Micro-cell transfer studies indicated the presence of a telomerase repressor gene in the 3p14.2-p21.1 region.⁽⁶⁸⁾ CpG islands hypermethylated in human lung cancer were identified by a modification of the arbitrarily primed-PCR method.⁽⁶⁹⁾ Expression of the Fhit protein was associated with prognosis of patients with lung cancer,⁽⁷⁰⁾ while expression of p16 and DCC proteins correlated with progression of neuroblastoma.^(71,72) The Smad2 gene was not mutated in colon cancer, while the PTEN gene was mutated in a small subset of lung cancers.^(73,74) The ABI-1 gene was identified as being fused with the MLL gene in acute myeloid leukemia with t(10;11)(p11.2;q23),⁽⁷⁵⁾ and a novel variant of chronic myelomonocytic leukemia carrying t(3;12)(q26;p13) with characteristics of 3q21q26 syndrome was identified.⁽⁷⁶⁾ Functional Analysis of Cancer-related Genes 　There are several projects in the Division to elucidate the pathogenetic significance of tumor suppressor gene inactivation in human cancer. Phosphorylation mechanisms of the p53 protein were investigated in association with its function,^(77-80) and it was found that the ATM protein phosphorylates the amino terminus of the p53 protein. A tetracycline-dependent inducible system for the expression of the p53 gene revealed that cell cycle arrest and apoptosis were differentially regulated by the level of p53 expression in a small cell lung carcinoma cell line.⁽⁸¹⁾ Sp1-mediated transcription of the Werner helicase gene was modulated by RB and p53.⁽⁸²⁾ Characterization of a Gene Involved in the Repair of Oxidative DNA Damage, 8-hydroxyguanine 　To elucidate the involvement of DNA repair system for 8-hydroxyguanine, a major form of oxygen-free radical-induced DNA damage, in human carcinogenesis, molecular analyses of the human OGG1 (oxoguanine DNA glycosylase) gene were performed.^(83-85) Although OGG1 mutations rarely occur in lung and gastric cancers, a genetic polymorphism resulting in different enzymatic activity of the OGG1 protein was identified. The genotype with a low enzymatic activity was more frequently detected in patients with lung and gastric cancers, suggesting that the OGG1 activity is involved in human carcinogenesis. Identification of a Metastasis Suppressor Gene and Construction of a Model for the Pleural Metastasis of Lung Cancer 　Eight genes were identified as being differentially expressed in association with the metastatic potential of K-1735 mouse melanoma cells by the mRNA differential display method. Molecular cloning and sequence analysis revealed that one of genes expressed in low metastatic cells, Elm1, is a novel member of the CCN family genes.⁽⁸⁶⁾ Expression of the Elm1 gene in highly metastatic K-1735 cells resulted in the suppression of in vivo growth and metastasis, suggesting that Elm1 is a metastasis suppressor gene. Injection of human non-small cell carcinoma cells into the pleural cavity of nude mice led to frequent intrapleural growth and metastasis.⁽⁸⁷⁾ Since this method is easy to perform and the results are reproducible, it can be applied to biological studies of human lung cancer cells in vivo. Molecular Epidemiology for Cancer Susceptibility 　Since it has been shown that genomic instability is present in a variety of cancer cells and in cancer-prone families, the prevalence of microsatellite instability was investigated in gastric cancer-prone families. Microsatellite instability was infrequently detected in gastric cancer of patients with family histories of gastric cancer aggregation, suggesting that inherited disorders in mismatch repair systems do not largely contribute to high susceptibility to gastric cancers.⁽⁸⁸⁾ Germline mutations of the PTEN gene were identified in three Japanese patients with Cowden disease.⁽⁸⁹⁾ Families with aggregation of lung cancer were not identified in the analysis of 1068 lung cancer patients, suggesting that familial lung cancer is rare in the Japanese population.⁽⁹⁰⁾