CRISPR gene editing makes stem cells "invisible" to the immune system

Scientists at the University of San Francisco have used the CRISPR-Cas9 gene editing system to create the first pluripotent stem cells functionally "invisible" to the immune system, a feat of biological engineering that, in laboratory studies, prevented the rejection of stem cell transplants. Because these "universal" stem cells can be made more efficiently than custom-designed stem cells for each patient – the individualized approach that has dominated previous efforts – they bring the promise of regenerative medicine closer to reality.

"Scientists often tout the therapeutic potential of pluripotent stem cells, which can evolve in any adult tissue, but the immune system is a major obstacle to safe and effective stem cell therapies," Tobias Deuse said. MD, Julien IE Hoffman, MD, Chair of UCSF Cardiac Surgery and Lead Author of the New Study, published Feb. 18 in the journal Nature Biotechnology.

The immune system does not forgive. It is programmed to eliminate everything it perceives as foreign, which protects the body against infectious agents and other invaders that could wreak havoc if left untouched. But it also means that organs, tissues or grafted cells are perceived as a potentially dangerous foreign incursion, which invariably causes a vigorous immune response leading to rejection of the graft. When this occurs, the donor and the recipient are said, in medical language, to be "incompatible in terms of histocompatibility".

"We can administer drugs that suppress immune activity and make rejection less likely, but these immunosuppressive drugs make patients more susceptible to infections and cancer," said surgical professor Sonja Schrepfer, MD, Ph. D., lead author and study director, Stem Cell Transplantation and Immunobiology Laboratory (RSW) at the time of the study.

In the field of stem cell transplants, scientists once thought that the problem of rejection was solved by induced pluripotent stem cells (iPSCs), created from fully mature cells, such as skin or fat cells, which are reprogrammed in order to allow them to develop. in one of the myriad of cells that make up the tissues and organs of the body. If cells derived from iPSCs were transplanted to the same patient who had donated the cells originally, the reasoning would be that the body would consider the transplanted cells as "self" and would not trigger an immune attack .

But in practice, the clinical use of iPSCs has proven difficult. For reasons that are still unknown, the cells of many patients do not lend themselves to reprogramming. In addition, it is expensive in time and money to produce iPSCs for each patient likely to benefit from stem cell therapy.

"IPSC technology raises a lot of problems, but the main hurdle is quality control and reproducibility, we do not know what makes some cells susceptible to reprogramming, but most scientists agree that It can not be done reliably, "said Deuse. "Most individualized iPSC therapies have been discontinued because of that."

Deuse and Schrepfer wondered if it would be possible to avoid these problems by creating "universal" CSPIs that can be used by all patients who need them. In their new article, they describe how, after modifying the activity of only three genes, iPSCs were able to avoid rejection after transplantation into histocompatibility-incompatible recipients with fully functioning immune systems.

"This is the first time anyone has genetically modified cells that can be universally transplanted and can survive in immunocompetent recipients without triggering an immune response," said Deuse.

The researchers first used CRISPR to remove two genes essential for the proper functioning of a class I and II major histocompatibility complex (MHC) protein family. MHC proteins lie on the surface of almost every cell and display molecular signals that help the immune system distinguish an intruder from a native. Cells lacking MHC genes do not display these signals, so they are not recorded as foreign. However, cells lacking MHC proteins become targets of immune cells called NK (Natural Killer) cells.

In collaboration with Professor Lewis Lanier, Ph.D. – co-author of the study, director of the Department of Microbiology and Immunology of the UCSF and expert of the signals that activate and inhibit the disease. NK cell activity – Schrepfer's team discovered that CD47, a cell surface protein acts as a signal "do not eat me" against immune cells called macrophages, also has a potent inhibitory effect on NK cells .

Believing that CD47 could be the key to completely suppressing the rejection, the researchers loaded the CD47 gene into a virus, which provided additional copies to human and mouse stem cells in which the MHC proteins had been neutralized.

CD47 has indeed turned out to be the missing piece of the puzzle. When the researchers transplanted their triple engineered mouse stem cells into mice that were incompatible with a normal immune system, they observed no rejection. They then transplanted human stem cells in the same way into so-called humanized mice – mice whose immune system has been replaced by components of the human immune system to mimic human immunity – and have to again observed no rejection.

In addition, researchers derived various types of human cardiac cells from these triple engineered stem cells, which they transplanted again into humanized mice. Stem cell-derived cardiac cells have been able to ensure their long-term survival and have even begun to form rudimentary blood vessels and cardiac muscle, suggesting the possibility that triple-engineered stem cells may be used someday to repair failing hearts.

"Our technique solves the problem of rejection of stem cells and stem cell-derived tissues, and represents a major breakthrough for the field of stem cell therapy," said Deuse. "Our technique can benefit a greater number of people with lower production costs than any individualized approach, we only need to manufacture our cells once and we end up with a product that can be applied universally. "

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University of California at San Francisco