Year 3

This project has developed a new method for making stem cells out of ordinary adult cells, such as skin cells. Our project utilizes the discovery that ordinary skin cells can be “reprogrammed” into stem cells that are similar in potential to embryonic stem cells. Such cells can become any type of cell and are useful for understanding and treating many diseases. The reprogramming process is done by introducing four genes that can change the pattern of expression of the genes in a skin cell to that of an embryonic cell.

The reprogramming genes are typically introduced into cells by putting them into viruses that can incorporate, or integrate, themselves into the chromosomes. This process is effective to bring about reprogramming, but it leaves behind viruses embedded in the chromosomes, which can activate genes that cause cancer. To get around this problem, my laboratory has developed a safer method for reprogramming, in which no viruses are used. Instead, we utilize an enzyme that can place a single copy of the reprogramming genes into a safe place in the chromosomes.

In our method, the reprogramming genes are present on small circles of DNA that are easily made and purified from bacteria grown in the laboratory. The circles of DNA, along with DNA that codes for the integration enzyme, are introduced into skin cells. The enzyme causes the reprogramming genes to incorporate into a chromosome at a single, safe location. After the cells are reprogrammed in this way, the reprogramming genes are completely and precisely removed from the chromosomes by introducing another enzyme. The result is reprogrammed cells whose chromosomes are very similar to the chromosomes of the starting cell and contain no added reprogramming genes. These cells appear to be safe to use in the clinic.

In addition, in the project, we developed a method to add a therapeutic gene to the reprogrammed cells. The therapeutic gene is a correct copy of the gene that is mutated in patients with a genetic disease. By adding a correct copy of the mutated gene, the stem cells now make a functional gene product and are healthy. The corrected stem cells can be used for transplantation into the patient to generate healthy tissues.

We demonstrated that cells reprogrammed by our method can be grown in such a way that they form muscle precursor cells that have the capacity to become healthy muscle fibers. We added a therapeutic gene to the reprogrammed cells that provides a correct copy of the gene that is mutated in the limb girdle form of muscular dystrophy. These corrected stem cells will be used in a strategy to repair the muscle damage present in these muscular dystrophy patients