Carcinogenesis Division

14. Carcinogenesis Division

The carcinogenesis studies being undertaken in this division use animal model systems. These models have a number of advantages over the direct study of humans. Another major area of study is the role of serine/threonine protein phosphatase in carcinogenesis.
Heterocyclic Amine (HCA) Carcinogenesis

Although humans are exposed to the food borne carcinogen, HCA, the role of HCA in human carcinogenesis has not yet been clarified. Analysis of the mutational fingerprint should provide clues to address this question. Mutational spectra of the rat colon carcinogens, 2-amino-3,4-dimethylimidazo[4,5-f] quinoline (MeIQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and a probable colon carcinogen,⁽¹⁸⁴⁾ 2-amino-9H-pyrido[2,3-b] indole(AaC) were analyzed in the colon tissue of the Big Blue mouse. Although all these compounds preferentially induced a GC ¨TA transversion, each compound had unique preferential target sequences. PhIP preferentially targeted the run of guanine bases, and a guanine deletion from GGGA was significantly characteristic of PhIP.⁽¹⁸⁵⁾ These characteristics coincided well with the mutation spectra of the tumor-related gene in the tumors induced by these HCAs.⁽¹⁸⁶⁾ Mutation spectra in the tumors induced by other carcinogens were also studied.⁽¹⁸⁷⁾ PhIP induces colon tumors in male F344 rats but not in females. However, mutation frequencies in the colon of male and female Big Blue rats were the same. In addition, no differences in mutation spectra were detected between males and females. Thus, initiation of colon carcinogenesis might occur in the same way in females as in males.⁽¹⁸⁸⁾ One of the other important target organs of HCA carcinogenesis is the mammary gland. A high fat diet did not significantly affect mammary-tumor incidence induced by PhIP. This effect differs from other mammary carcinogens.⁽¹⁸⁹⁾ Some genotoxic effects of HCAs were inhibited by vanillin, coumarin and caffeine.⁽¹⁹⁰⁾
Rat Genome Study

Different strains of rat show different susceptibilities to carcinogenesis. The ACI rat is sensitive to and the BUF rat resistant to N-methyl-NÕ-nitro-N-nitrosoguanidine (MNNG)-induced stomach carcinogenesis.⁽²⁵⁵⁾ The gene for the resistance in stomach carcinogenesis was mapped to rat chromosome 3. For identification of the gene(s), isolation of polymorphic markers is important. Rb1 was mapped to chromosome 15⁽¹⁹¹⁾ and Brca2 to chromosome 12.⁽¹⁹²⁾ It was suggested that the polymorphism detected in Brca2 was not implicated in the susceptibility of the SD strain to mammary carcinogenesis. Oxidative stress in LEC rats was evaluated by the level of ascorbic acid in plasma.⁽¹⁹³⁾
Serine/Threonine Protein Phosphatases (PPs) in Carcinogenesis

Okadaic acid (OA), a potent and specific inhibitor of PP2A, induced minisatellite instability. To clarify the protein involved in maintaining genomes in a stable condition, fibroblast cells derived from the scid mouse were examined for minisatellite stability, and found to be unstable. Since the gene responsible for the scid phenotype is DNA-PKcs, inactivation of DNA-PK by treatment with OA is a pos sible mechanism.⁽¹⁹⁴⁾ PP1 is another major PP in mammals, and its activity is regulated by proteinaceous inhibitor-2. Three isotypes of inhibitor-2 were found in testis, two of which were specifically expressed at this site.⁽¹⁹⁵⁾ A novel B regulatory subunit cDNA for PP2A was isolated from Xenopus oocytes.⁽¹⁹⁶⁾
Analysis of Aberrant Methylation in Carcinogenesis

Epigenetic changes such as changes in methylation status may play an important role in carcinogenesis. A method to clone the genomic fragments of altered methylation status was established using a mouse liver tumor induced by MeIQ, as well as normal liver samples. By application of RDA to HpaII-digested DNA, five fragments which were hypermethylated and six frag ments which were hypomethylated in the tumor were obtained. Among these, four were LINE 1, and they were hypomethylated in all the tumors examined. All sequences which were hypermethylated in tumors were unknown. Thus, this method is useful approach for isolating unknown genomic regions where methylation status has changed.⁽¹⁹⁷⁾
Familial Breast Cancer

Twenty-two Japanese breast cancer families of which two had been found to have mutations in BRCA1 were examined for BRCA2 mutations. Five families had mutations. Three of them resulted in truncation of the encoded protein and co-segregated with breast cancer manifestations. The other two families had the same mutation which resulted in a missense mutation, and co-segregation was not clear. The mutation was suggested to be a probable cancer-prone mutation.⁽¹⁹⁸⁾ Mutation in the androgen receptor gene played a minor role in Japanese prostate cancer.⁽¹⁹⁹⁾




	14. Carcinogenesis Division



		The carcinogenesis studies being undertaken in this division use animal model systems. These models have a number of advantages over the direct study of humans. Another major area of study is the role of serine/threonine protein phosphatase in carcinogenesis. Heterocyclic Amine (HCA) Carcinogenesis Although humans are exposed to the food borne carcinogen, HCA, the role of HCA in human carcinogenesis has not yet been clarified. Analysis of the mutational fingerprint should provide clues to address this question. Mutational spectra of the rat colon carcinogens, 2-amino-3,4-dimethylimidazo[4,5-f] quinoline (MeIQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and a probable colon carcinogen,⁽¹⁸⁴⁾ 2-amino-9H-pyrido[2,3-b] indole(AaC) were analyzed in the colon tissue of the Big Blue mouse. Although all these compounds preferentially induced a GC ¨TA transversion, each compound had unique preferential target sequences. PhIP preferentially targeted the run of guanine bases, and a guanine deletion from GGGA was significantly characteristic of PhIP.⁽¹⁸⁵⁾ These characteristics coincided well with the mutation spectra of the tumor-related gene in the tumors induced by these HCAs.⁽¹⁸⁶⁾ Mutation spectra in the tumors induced by other carcinogens were also studied.⁽¹⁸⁷⁾ PhIP induces colon tumors in male F344 rats but not in females. However, mutation frequencies in the colon of male and female Big Blue rats were the same. In addition, no differences in mutation spectra were detected between males and females. Thus, initiation of colon carcinogenesis might occur in the same way in females as in males.⁽¹⁸⁸⁾ One of the other important target organs of HCA carcinogenesis is the mammary gland. A high fat diet did not significantly affect mammary-tumor incidence induced by PhIP. This effect differs from other mammary carcinogens.⁽¹⁸⁹⁾ Some genotoxic effects of HCAs were inhibited by vanillin, coumarin and caffeine.⁽¹⁹⁰⁾ Rat Genome Study Different strains of rat show different susceptibilities to carcinogenesis. The ACI rat is sensitive to and the BUF rat resistant to N-methyl-NÕ-nitro-N-nitrosoguanidine (MNNG)-induced stomach carcinogenesis.⁽²⁵⁵⁾ The gene for the resistance in stomach carcinogenesis was mapped to rat chromosome 3. For identification of the gene(s), isolation of polymorphic markers is important. Rb1 was mapped to chromosome 15⁽¹⁹¹⁾ and Brca2 to chromosome 12.⁽¹⁹²⁾ It was suggested that the polymorphism detected in Brca2 was not implicated in the susceptibility of the SD strain to mammary carcinogenesis. Oxidative stress in LEC rats was evaluated by the level of ascorbic acid in plasma.⁽¹⁹³⁾ Serine/Threonine Protein Phosphatases (PPs) in Carcinogenesis Okadaic acid (OA), a potent and specific inhibitor of PP2A, induced minisatellite instability. To clarify the protein involved in maintaining genomes in a stable condition, fibroblast cells derived from the scid mouse were examined for minisatellite stability, and found to be unstable. Since the gene responsible for the scid phenotype is DNA-PKcs, inactivation of DNA-PK by treatment with OA is a pos sible mechanism.⁽¹⁹⁴⁾ PP1 is another major PP in mammals, and its activity is regulated by proteinaceous inhibitor-2. Three isotypes of inhibitor-2 were found in testis, two of which were specifically expressed at this site.⁽¹⁹⁵⁾ A novel B regulatory subunit cDNA for PP2A was isolated from Xenopus oocytes.⁽¹⁹⁶⁾ Analysis of Aberrant Methylation in Carcinogenesis Epigenetic changes such as changes in methylation status may play an important role in carcinogenesis. A method to clone the genomic fragments of altered methylation status was established using a mouse liver tumor induced by MeIQ, as well as normal liver samples. By application of RDA to HpaII-digested DNA, five fragments which were hypermethylated and six frag ments which were hypomethylated in the tumor were obtained. Among these, four were LINE 1, and they were hypomethylated in all the tumors examined. All sequences which were hypermethylated in tumors were unknown. Thus, this method is useful approach for isolating unknown genomic regions where methylation status has changed.⁽¹⁹⁷⁾ Familial Breast Cancer Twenty-two Japanese breast cancer families of which two had been found to have mutations in BRCA1 were examined for BRCA2 mutations. Five families had mutations. Three of them resulted in truncation of the encoded protein and co-segregated with breast cancer manifestations. The other two families had the same mutation which resulted in a missense mutation, and co-segregation was not clear. The mutation was suggested to be a probable cancer-prone mutation.⁽¹⁹⁸⁾ Mutation in the androgen receptor gene played a minor role in Japanese prostate cancer.⁽¹⁹⁹⁾