Scientists have spent decades trying to manufacture pig tissues so that they would not be rejected by the human body , but they have persistently had to contend with PERVs . What are they? Remnants of ancient viral infections, the genes for endogenous porcine retroviruses , which are scattered throughout the pig genome and can infect a person who one day receives a pig’s heart, lung, or kidney as a substitute or temporary organ .
But now, an American company has managed to cross these viruses off the list of concerns. Using CRISPR gene editing, the company’s researchers and several academic laboratories have succeeded in creating dozens of apparently healthy pigs without a trace of PERV genes. In an article published in the journal Science , the team reports that the technique was used in live pigs with the same success rate as the cloning method without genetic modification, about one pig for every 100 implanted embryos. And when they tested the tissues of the thirty-seven piglets born so far, they all appeared to be PERV-free.
However, even with these retroviruses out of the picture, pigs would require other modifications so that their organs are not rejected by the human immune system or cause other damage.
At the moment, scientists have already implanted pig pancreas cells in people with diabetes and found no evidence of PERV infection . Large porcine organs, which will likely require immunosuppressive drugs to prevent patient bodies from attacking them, could pose an increased risk of PERV transmission.
Virologist Joachim Denner of the Robert Koch Institute in Berlin has already tried to cut PERV genes from pig cells with an editing technology called zinc nucleases finger , but the many imprecise DNA snippets proved toxic to the cells.
CRISPR, the ideal solution
Then CRISPR came along. Two of the early developers of that gene-editing technology, Harvard University geneticists George Church and Luhan Yang, suspected that CRISPR’s highly efficient duo of guide RNA and a DNA-cutting enzyme could make precise changes in the entire genome to pig cells. In 2015, they co-founded the eGenesis company to focus on transplanted organ engineering, and Yang became the company’s chief scientific officer. The same year, they showed that CRISPR could delete PERV genes in the pig genome.
For that project, the researchers used a line of “immortal” pig kidney cells , chosen for their ability to survive and divide indefinitely. To create PERV-free pigs, they needed to start with genetically normal cells from a live pig.
In the new work, conducted with a team of Danish and Chinese collaborators, the eGenesis team applied the CRISPR system to cells derived from the connective tissue of fetal pigs. These cells were more fragile when subjected to CRISPR work. But once corrected, they did not grow normally, possibly because damage to their DNA led them to stop dividing or self-destruct. But by exposing the cells to a chemical cocktail that stimulated growth and tamped down a key growth-suppressing gene, the team found 100% PERV-free cells.
To produce piglets, the researchers used a standard cloning technique : they inserted the nuclei containing DNA from these edited cells into cells taken from the ovaries of pigs in a Chinese slaughterhouse. They allowed each egg to develop into an embryo and implant it in the uterus of a surrogate mother.
But PERVs aren’t the only thing standing in the way of transplanted pig organs. Researchers will have to remove pig genes that affect the human immune system and insert others that will prevent toxic interactions with human blood.
Image: eGenesis.
References:
D. Niu et al. “Inactivation of porcine endogenous retrovirus in pigs using CRISPR-Cas9” Science August 10, 2017.
