title: Activating the expression of human K-rasG12D stimulates oncogenic transformation in transgenic goat fetal fibroblast cells.
abstract: Humane use of preclinical large animal cancer models plays a critical role in understanding cancer biology and developing therapeutic treatments. Among the large animal candidates, goats have great potentials as sustainable sources for large animal cancer model development. Goats are easier to handle and cheaper to raise. The genome of the goats has been sequenced recently. It has been known that goats develop skin, adrenal cortex, breast and other types of cancers. Technically, goats are subject to somatic cell nuclear transfer more efficiently and exhibit better viability through the cloning process. Towards the development of a goat cancer model, we created a transgenic goat fetal fibroblast (GFF) cell as the donor cell for SCNT. Human mutated K-ras (hK-rasG12D) was chosen as the transgene, as it is present in 20% of cancers. Both hK-rasG12D and a herpes simplex viral thymidine kinase (HSV1-tk) reporter genes, flanked by a pair of LoxP sites, were knocked in the GFF endogenous K-ras locus through homologous recombination. Following Cre-mediated activation (with a 95% activation efficiency), hK-rasG12D and HSV1-tk were expressed in the transgenic GFF cells, evidently through the presence of corresponding mRNAs, and confirmed by HSV1-tk protein function assay. The hK-rasG12D expressing GFF cells exhibited enhanced proliferation rates and an anchorage-independent growth behavior. They were able to initiate tumor growth in athymic nude mice. In conclusion, after activating hK-rasG12D gene expression, hK-rasG12D transgenic GFF cells were transformed into tumorgenesis cells. Transgenic goats via SCNT using the above-motioned cells as the donor cells have been established.
authors.affiliation: Department of Chemistry and Biochemistry, Utah State University, Logan, Utah, United States of America.
authors.forename: Jianhua
authors.initials: J
authors.lastname: Gong
authors.affiliation: Department of Animal, Dairy, and Veterinary Sciences, Utah State University, Logan, Utah, United States of America.
authors.forename: Zhongde
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Department of Animal, Dairy, and Veterinary Sciences, Utah State University, Logan, Utah, United States of America.
authors.forename: Irina
authors.initials: I
authors.lastname: Polejaeva
authors.affiliation: Department of Radiology, University of Chicago, Chicago, Illinois, United States of America.
authors.forename: Ravi
authors.initials: R
authors.lastname: Salgia
authors.affiliation: Department of Radiology, University of Chicago, Chicago, Illinois, United States of America.
authors.forename: Chien-Min
authors.initials: CM
authors.lastname: Kao
authors.affiliation: Department of Radiology, University of Chicago, Chicago, Illinois, United States of America.
authors.forename: Chin-Tu
authors.initials: CT
authors.lastname: Chen
authors.affiliation: Department of Chemistry and Biochemistry, Utah State University, Logan, Utah, United States of America.
authors.forename: Guangchun
authors.initials: G
authors.lastname: Chen
authors.affiliation: Department of Chemistry and Biochemistry, Utah State University, Logan, Utah, United States of America.
authors.forename: Liaohai
authors.initials: L
authors.lastname: Chen
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D063646:Carcinogenesis / Q000378:metabolism / Q000473:pathology; D002471:Cell Transformation, Neoplastic / Q000378:metabolism* / Q000473:pathology*; D005333:Fetus / Q000473:pathology*; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology*; D005822:Genetic Vectors / Q000378:metabolism; D006041:Goats / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D018259:Herpesvirus 1, Human / Q000378:metabolism; D006801:Humans; D019426:Integrases / Q000378:metabolism; D008819:Mice, Nude; D050505:Mutant Proteins / Q000378:metabolism*; D010641:Phenotype; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras); D011995:Recombination, Genetic / Q000235:genetics; D013937:Thymidine Kinase / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism*
chemical_list: C117307:KRAS protein, human; D050505:Mutant Proteins; D011518:Proto-Oncogene Proteins; D049452:Green Fluorescent Proteins; D013937:Thymidine Kinase; C045073:Cre recombinase; D019426:Integrases; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins