Patient-Specific Stem Cells Plus Personalized Gene Therapy for Blindness


Researchers from Columbia University Medical Center (CUMC) have devised protocols to develop personalized gene therapies for patients with an eye known as retinitis pigmentosa (RP), which is a leading cause of vision loss. While RP can begin during infancy, the first symptoms typically emerge during early adulthood. Typically the disease begins with night blindness, and RP eventually progresses to rob the patients of their peripheral vision. In its later stages, RP destroys photoreceptors in the macula, that region of the retina that provides the best vision under lighted conditions. RP is estimated to affect at least 75,000 people in the United States and 1.5 million worldwide.

The approach utilized by this Columbia team utilizes induced pluripotent stem (iPS) cell technology to transform patient’s skin cells into retinal cells, which are then used as a patient-specific model for disease study and preclinical testing.

The leader of this research group, Stephen H. Tsang, MD, PhD, showed that a form of RP caused by mutations to the MFRP gene compromised the structural integrity of the retinal cells. The MFRP gene encodes a protein called the Membrane Frizzled-Related Protein, which plays an important role in eye development. Mutations in the MFRP gene are associated with small eye conditions such as nanophthalmos, posterior microphthalmia, or retinal issues such as retinitis pigmentosa, foveoschisis, or even optic disc drusen. Tsang and his group, however, showed that the effects of these MFRP mutations could be reversed with gene therapy. Thus this new approach could potentially be used to create personalized therapies for other forms of RP, or even other genetic diseases.

“The use of patient-specific cell lines for testing the efficacy of gene therapy to precisely correct a patient’s genetic deficiency provides yet another tool for advancing the field of personalized medicine,” said Dr. Tsang, the Laszlo Z. Bito Associate Professor of Ophthalmology and associate professor of pathology and cell biology. This work was recently published in the online edition of Molecular Therapy, the official journal of the American Society for Gene & Cell Therapy.

Mutations in more than 60 different genes have been linked to RP. Such a genetic disease is known as a heterogeneous trait and genetic diseases like RP or deafness or other such conditions are very difficult to develop models to study. Animal models, though useful, have significant limitations because of interspecies differences. Eye researchers have also used human retinal cells from eye banks to study RP. This eye tissue comes from the eyes of patients who suffered from the disease and donated their eye tissue to research after death. Unfortunately, despite their usefulness, donated eye tissues typically illustrate the end stage of the disease process. Despite their usefulness, they reveal little about how RP develops. Also, there are no human tissue culture models of RP, since it is dangerous to harvest retinal cells from patients. Finally, human embryonic stem cells could be useful in RP research, but they are fraught with ethical, legal, and technical issues.

However, the Tsang group used iPS technology to transform skin cells from RP patients, each of whom harbored a different MFRP mutation, into retinal cells. Thus they created patient-specific models for studying the disease and testing potential therapies. Because they used iPS technology, Tsang found a way around the limitations and concerns and dog embryonic stem cells. Thus researchers can induce the patient’s own skin cells and de-differentiated them to a more basic, embryonic stem cell–like state. Such cells are “pluripotent,” which means that they can be transformed into specialized cells of various types.

When Tsang and others analyzed these patient-specific cells, they discovered that the primary effect of MFRP mutations is to disrupt the regulation of a cytoskeletal protein called actin, the scaffolding that gives the cell its structural integrity. “Normally, the cytoskeleton looks like a series of connected hexagons,” said Dr. Tsang. “If a cell loses this structure, it loses its ability to function.” They also found that MFRP works in tandem with another gene, CTRP5, and that a balance between the two genes is required for normal actin regulation.

In the next phase of the study, the CUMC team used adeno-associated viruses (AAVs) to introduce normal copies of MFRP into the iPS-derived retinal cells. This successfully restored the cells’ function. Tsang and others used gene therapy to “rescue” mice with RP due to MFRP mutations. According to Dr. Tsang, the mice showed long-term improvement in visual function and restoration of photoreceptor numbers.

“This study provides both in vitro and in vivo evidence that vision loss caused by MFRP mutations could potentially be treated through AAV gene therapy,” said coauthor Dieter Egli, PhD, an assistant professor of developmental cell biology (in pediatrics) at CUMC, who is also affiliated with the New York Stem Cell Foundation.

Dr. Tsang thinks this approach could potentially be used to study other forms of RP. “Through genome-sequencing studies, hundreds of novel genetic spelling mistakes have been associated with RP,” he said. “But until now, we’ve had very few ways to find out whether these actually cause the disease. In principle, iPS cells can help us determine whether these genes do, in fact, cause RP, understand their function, and, ultimately, develop personalized treatments.”

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