Jaime Rivera, Guangping Gao develop new genome-editing technology to model human disease

	Jaime Rivera, PhD
	Guangping Gao, PhD

UMass Chan Medical School scientists have devised a simplified strategy for developing genetically modified mice for disease research that combines two important laboratory tools: adeno-associated viruses (AAVs) such as those used in gene therapy and CRISPR/Cas9 technology, a gene editing tool. Using this new strategy, researchers believe they will be able to develop mice strains that model human disease more quickly and effectively than current techniques.

“This is a major leap in transgenic technology since it greatly simplifies the generation of genetically modified mice,” said Jaime Rivera, PhD, associate professor of pediatrics.

An expert in early mammalian development, Dr. Rivera collaborated with AAV pioneer Guangping Gao, PhD, the Penelope Booth Rockwell Professor in Biomedical Research, professor of microbiology & physiological systems, director of the Horae Gene Therapy Center and co-director of the Li Weibo Institute for Rare Diseases Research, to create technology that utilizes recombinant AAVs to introduce CRISPR/Cas9 components into one-cell stage mice embryos.

Making this technology possible is the discovery that AAVs can penetrate the zona pellucida, a proteinaceous layer that envelops pre-implantation embryos, which normally act as a barrier to virus infection. Taking advantage of this discovery, Dr. Gao, Rivera and colleagues exposed intact one-cell mice embryos to AAV particles coding for CRISPR/Cas9 components, generating mice born with mutations in the tyrosinase gene, which lead to albino mice. They were also able to induce specific mutations by a process known as “homology directed repair” (HDR). Using HDR, they generated mutations in the fumarylacetoacetate hydrolase gene “Fah.” In humans, mutations in this gene result in type I tyrosinemia, an inborn error of metabolism.

They also showed that it is possible to genetically modify pre-implantation embryos in vivo. To conduct these experiments, they delivered AAV-CRISPR particles into the oviduct of pregnant females to create transgenic offspring.

“Traditionally, transgenic mice have been generated using microinjection techniques to deliver DNA or other molecules into one-cell stage embryos,” said Rivera. “This is a highly sophisticated technique that requires expensive equipment and dexterous personnel. The AAV-CRISPR technique eliminates the need for microinjection and if done in vivo also avoids the need to isolate the one-cell embryos and conduct embryo culture.”

These advantages make the AAV-CRISPR technology an attractive tool to generate animal models that are suited to study human disease.

“This collaboration is a classic example of how the combination of expertise in apparently disparate fields of research can be used to advance a research field. It also illuminates the collaborative atmosphere that characterizes the University of Massachusetts Medical School,” said Rivera.

The study was published in Nature Communications.