NEXT-GENERATION MOUSE GENE-TARGETING TECHNOLOGY TO MODEL TUMORIGENESIS


Year of Award:
2013
Award Type:
R21
Project Number:
CA181859
RFA Number:
RFA-CA-13-001
Technology Track:
Molecular & Cellular Analysis Technologies
PI/Project Leader:
WANG, ZHENGHE
Other PI or Project Leader:
N/A
Institution:
CASE WESTERN RESERVE UNIVERSITY
AbstractThe goal of this application is to develop a very fast and cost-effective method to generate gene-targeted miceto model tumorigenesis. Gene-targeted mice are invaluable tools to determine the roles of oncogenicmutations in cancer development. However, conventional gene targeting is slow, expensive and prone tofailure. While nuclease-mediated targeting may speed the production of mutants, there remain significantconcerns about off-target mutations, relative ease of use and access to the entire genome. In preliminarystudies, we have successfully developed an innovative method to directly and efficiently target mouse fertilizedeggs using recombinant adeno-associated virus (rAAV)-mediated homologous recombination. Using thisapproach, we were able to generated germ-line-transmitting mice with at least 10% targeting frequency in amonth. We believe that our technology is superior to nuclease-mediated gene targeting approaches (e.g. ZFN,TALEN and CRISPR/Cas). In contrast to nuclease-mediated approaches, off-target mutations are infrequent,embryos can be processed en masse without individual microinjection, and all regions of the genome areaccessible to manipulation. Here we propose to further develop this technology to generate gene-targetedmice to model tumorigenesis by determining: (a) if gene-targeted mice generated by our method are suitablefor modeling tumorigenesis, (b) if our approach is generally applicable to create gene-targeted mice of varioustumor suppressors and oncogenes, and (c) if our approach can be used to generate conditional knock-out andknock-in mice. Successful development of these technologies will revolutionize generation of geneticallyengineered mice to model tumorigenesis. It will have huge impacts on basic cancer biology as well as cancerdrug development.