An inducible transgenic porcine model for the study of human cancer

Abstract:

The common rodent-based models are limited in their capability to mimic human cancer development. Given the more similarity in genetics and physiology between human and pig, we aim to generate a novel transgenic porcine oncogenesis model for the study of human cancer. Our previous study has shown that overexpression of several human oncogenes led to tumor development in pigs. We thus chose two major mutant oncogenes, Kras and p53, to generate transgenic pigs that can be induced to simultaneously express both genes. We first cloned and sequenced porcine Kras and p53 wild-type genes.  Protein sequence alignment of each gene among pig, human and mouse determined the pig-specific mutation sites that correspond to those commonly found in human cancers. Porcine Kras mutation occurs at the 12^th glycine (G) to aspartic acid (D), whereas p53 arginine (R) at 167^th position was mutated to histidine (H). Kras^G12D and p53^R167H mutants were linked by internal ribosome entry sites (IRES) for their simultaneous expression. The cassette was then inserted into a vector following the LoxP-polyA(STOP)-LoxP sequence. We further tested this vector construct in pig fibroblasts by co-transfection with Cre plasmid, which deletes the polyA “STOP” sequence and allows transgene transcription. We found that expression of Kras^G12D and p53^R167H mutants was significantly induced upon Cre recombinase introduction. In the future, we will use this vector construct and clone technology to generate inducible Kras^G12D and p53^R167H transgenic pig, and ultimately to create a more human-like tumor model for the study of cancer etiology, and for the development of anti-cancer therapy.