EXPERIMENTAL PATHOLOGY AND CHEMOTHERAPY DIVISION

3.EXPERIMENTAL PATHOLOGY AND CHEMOTHERAPY DIVISION

    Studies are progressing on the susceptibility of human c-Ha-ras proto-oncogene transgenic rats to various chemical carcinogens and underlying mechanisms, prevention of carcinogenesis and metastasis by bovine lactoferrin (bLF), and cancer prevention by ferulic acid and its derivatives and several other compounds.
Characterization of Carcinogen Susceptibility of Human c-Ha-ras Proto-oncogene Transgenic Rats

    A rat line carrying 3 copies of the human c-Ha-ras proto-oncogene, designated as Hras128, is highly susceptible to induction of mammary, urinary bladder, esophagus and skin tumors by chemical carcinogens.⁽⁵⁶⁾ Mutations in the transgene, but not the endogenous ras gene, and increase of active form Ras protein were observed in carcinomas developing in the mammary glands and esophagus of these rats. The transgene thus is preferentially targeted by chemical carcinogens. In mammary carcinogenesis, since carcinogen application is most effective at 49 days of age, terminal endbuds (TEBs) at the tips of developing mammary ducts appear important. Higher numbers of TEBs and with higher levels of c-myc and active MAP kinase in transgenic rats provide further support for a histogenetic role.
    The rat gene encoding neuronal leucine-rich repeat protein-3 (NLRR-3), found overexpressed in a sarcoma spontaneously developing in the mammary region of a Hras128 rat, has further been cloned. Roles of the gene in invasion and metastasis of malignant tumors are currently under investigation.⁽⁵⁷⁾
    This transgenic rat was also found to be highly susceptible to various carcinogens such as N,N'-bis(2-hydroxypropyl)nitrosamine, diethylnitrosamine, azoxymathane, NNK, IQ, MeIQx, TPA and some others, which preferentially induce tumors in organs other than mammary glands. Taking advantage of this specific carcinogen susceptibility, application of the in vivo medium-term carcinogen assay system is currently scheduled in collaboration with other research groups.
Anti-carcinogenesis and Anti-metastasis of Bovine Lactoferrin (bLF)

    Focusing on food components and derivatives as candidates because of practical use and known lack of toxicity, bovine lactoferrin (bLF), a multifunctional protein in milk with antimicrobial and immune response-enhancing activity, was shown to inhibit colon, bladder, tongue and liver carcinogenesis, along with lung metastatic colony formation in Co26Lu bearing mice. A possible mechanism of such preventive effects is due to the augmented killing of tumor cells by CD8⁺ T and NK (asialoGM1⁺) cells, which may be activated by interleukin-18 (IL-18) thought to be induced in epithelial cells of the small intestine after treatment with bLF.^(58-⁶⁰⁾
    Based on our observation of the preventive effects of bLF on colon carcinogenesis, clinical study are under way in the National Cancer Center Hospital. Clinical trials for patients with chronic hepatitis are also under way in the National Cancer Center Hospital in collaboration with other hospitals.
Cancer Chemoprevention by Ferulic Acid Derivatives and Other Compounds

    In separate research projects, 3-ethyl(4-geranyloxy-3'-methoxyphenyl)-2-propionate (EGMP), from rice bran,⁽⁶¹⁾ lycopene, a carotenoid, and unsaturated n-3 fatty acids ⁽⁶²⁾ and other compounds like lycopene⁽⁶³⁾ and DHEA⁽⁶⁴⁾ were additionally found to exert beneficial influence.
Studies on the Association Ability of Metallophthalocyanine Complexes with Polycyclic Aromatic Carcinogens

    Molecular associations occurring between pairs of carcinogenic heterocyclic amine (HCA) in aqueous solution can increase hydrophilicity and lower permeability, explaining that lack of carcinogenic summation with two or more HCAs in combination in vivo. Testing of several phthalocyanines with analogous structures allowed identification of zinc phthalocyaninetetrasulfonic acid which may act in this way to 'eliminate' HCAs.
Other Studies

    Studies on the global effects of anchorage on gene expression of mammary cell lines were analyzed by gene expression profilometry⁽⁶⁵⁾ and establishment and characterization of novel cell lines cloned from rat renal cell carcinomas were also reported.⁽⁶⁶⁾




3.EXPERIMENTAL PATHOLOGY AND CHEMOTHERAPY DIVISION



	Studies are progressing on the susceptibility of human c-Ha-ras proto-oncogene transgenic rats to various chemical carcinogens and underlying mechanisms, prevention of carcinogenesis and metastasis by bovine lactoferrin (bLF), and cancer prevention by ferulic acid and its derivatives and several other compounds. Characterization of Carcinogen Susceptibility of Human c-Ha-ras Proto-oncogene Transgenic Rats A rat line carrying 3 copies of the human c-Ha-ras proto-oncogene, designated as Hras128, is highly susceptible to induction of mammary, urinary bladder, esophagus and skin tumors by chemical carcinogens.⁽⁵⁶⁾ Mutations in the transgene, but not the endogenous ras gene, and increase of active form Ras protein were observed in carcinomas developing in the mammary glands and esophagus of these rats. The transgene thus is preferentially targeted by chemical carcinogens. In mammary carcinogenesis, since carcinogen application is most effective at 49 days of age, terminal endbuds (TEBs) at the tips of developing mammary ducts appear important. Higher numbers of TEBs and with higher levels of c-myc and active MAP kinase in transgenic rats provide further support for a histogenetic role. The rat gene encoding neuronal leucine-rich repeat protein-3 (NLRR-3), found overexpressed in a sarcoma spontaneously developing in the mammary region of a Hras128 rat, has further been cloned. Roles of the gene in invasion and metastasis of malignant tumors are currently under investigation.⁽⁵⁷⁾ This transgenic rat was also found to be highly susceptible to various carcinogens such as N,N'-bis(2-hydroxypropyl)nitrosamine, diethylnitrosamine, azoxymathane, NNK, IQ, MeIQx, TPA and some others, which preferentially induce tumors in organs other than mammary glands. Taking advantage of this specific carcinogen susceptibility, application of the in vivo medium-term carcinogen assay system is currently scheduled in collaboration with other research groups. Anti-carcinogenesis and Anti-metastasis of Bovine Lactoferrin (bLF) Focusing on food components and derivatives as candidates because of practical use and known lack of toxicity, bovine lactoferrin (bLF), a multifunctional protein in milk with antimicrobial and immune response-enhancing activity, was shown to inhibit colon, bladder, tongue and liver carcinogenesis, along with lung metastatic colony formation in Co26Lu bearing mice. A possible mechanism of such preventive effects is due to the augmented killing of tumor cells by CD8⁺ T and NK (asialoGM1⁺) cells, which may be activated by interleukin-18 (IL-18) thought to be induced in epithelial cells of the small intestine after treatment with bLF.^(58-⁶⁰⁾ Based on our observation of the preventive effects of bLF on colon carcinogenesis, clinical study are under way in the National Cancer Center Hospital. Clinical trials for patients with chronic hepatitis are also under way in the National Cancer Center Hospital in collaboration with other hospitals. Cancer Chemoprevention by Ferulic Acid Derivatives and Other Compounds In separate research projects, 3-ethyl(4-geranyloxy-3'-methoxyphenyl)-2-propionate (EGMP), from rice bran,⁽⁶¹⁾ lycopene, a carotenoid, and unsaturated n-3 fatty acids ⁽⁶²⁾ and other compounds like lycopene⁽⁶³⁾ and DHEA⁽⁶⁴⁾ were additionally found to exert beneficial influence. Studies on the Association Ability of Metallophthalocyanine Complexes with Polycyclic Aromatic Carcinogens Molecular associations occurring between pairs of carcinogenic heterocyclic amine (HCA) in aqueous solution can increase hydrophilicity and lower permeability, explaining that lack of carcinogenic summation with two or more HCAs in combination in vivo. Testing of several phthalocyanines with analogous structures allowed identification of zinc phthalocyaninetetrasulfonic acid which may act in this way to 'eliminate' HCAs. Other Studies Studies on the global effects of anchorage on gene expression of mammary cell lines were analyzed by gene expression profilometry⁽⁶⁵⁾ and establishment and characterization of novel cell lines cloned from rat renal cell carcinomas were also reported.⁽⁶⁶⁾