Recessive dystrophic epidermolysis bullosa (RDEB) is caused by loss-of-function mutations in the collagen type VII (C7) gene (COL7A1), and results in a potentially fatal and terribly debilitating disorder. Individuals with severe generalied RDEB develop blisters and erosions involving large portions of the body surface area, mutilating scarring, joint contractures, and strictures of the esophagus, with opportunistic infections and aggressive squamous cell carcinoma as principal causes of early death. We have shown that hematopoietic stem cell transplantation (HSCT) can increase C7 protein in skin and mucous membranes, and ameliorate many of the disease manifestations. Unfortunately not all mucocutaneous lesions heal, and the toxicities associated with HSCT are significant. Our overarching goal is to solve these problems by establishing a reliable method for delivering C7-expressing cells that home to the dermal-epidermal junction and secrete C7 protein. In order to accomplish this goal and to overcome the limitations of allogeneic HSCT, we will: 1) identify the optimal cellular vector and homing signal for targeting injured skin in RDEB, using human mesenchymal stromal/stem cells and induced pluripotent stem cells (iPSCs), 2) define the efficacy of systemic RDEB gene therapy by C7 gene augmentation, and 3) use nucleases for gene editing of individual mutations, as well as whole regions of the C7 gene. We propose to define the conditions conducive to wound healing in this severe blistering genodermatosis, but the impact of these studies may not be limited to RDEB. The function of C7 depends on extracellular polymerization, which makes RDEB an ideal model for understanding the mechanisms of cross- correction of structural protein deficiency in the extracellular matrix. In addition, wound healing integrates skin extracellular matrix and skin cells into a dynamic system, which in RDEB results in the depletion of skin stem cell niches and, in turn, provides a unique model of skin tissue repair with donor regenerative cells. By using powerful tools for studying and manipulating the information basis of biological systems-engineered restriction enzymes (nucleases), skin- targeting, and induced cell lineage conversion (iPSCs)-we will focus on developing personalized cell therapy for individuals with RDEB with the idea that our findings may have broad implications for understanding other connective tissue extracellular matrix diseases. Our proposal is equally motivated by wanting a better understanding of the biological mechanisms in injured skin and by needing to improve the lives of people with RDEB through maximizing the benefits and reducing the risks of potential novel therapies.

Public Health Relevance

More than 3,000 new cases of genetic inborn errors are found in U.S. newborns every year. The complex and life-long medical care for these patients represents a significant challenge to the health care system. Additionally, a deeper understanding of the basic biology of skin blistering disorders and the proof-of-concept for clinical translation in these relatively rare conditions is likely to have a profound impact on muc more common ones, such as chemical and thermal burns.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
1R01AR063070-01A1
Application #
8502074
Study Section
Arthritis, Connective Tissue and Skin Study Section (ACTS)
Program Officer
Baker, Carl
Project Start
2013-04-01
Project End
2018-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
1
Fiscal Year
2013
Total Cost
$458,962
Indirect Cost
$127,618
Name
University of Minnesota Twin Cities
Department
Pediatrics
Type
Schools of Medicine
DUNS #
555917996
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Agrawal, Pranav; Ingle, Nilesh P; Boyle, William S et al. (2016) Fast, Efficient, and Gentle Transfection of Human Adherent Cells in Suspension. ACS Appl Mater Interfaces 8:8870-4
Perdoni, Christopher; Osborn, Mark J; Tolar, Jakub (2016) Gene editing toward the use of autologous therapies in recessive dystrophic epidermolysis bullosa. Transl Res 168:50-8
Osborn, Mark J; Webber, Beau R; Knipping, Friederike et al. (2016) Evaluation of TCR Gene Editing Achieved by TALENs, CRISPR/Cas9, and megaTAL Nucleases. Mol Ther 24:570-81
Watt, Stephen A; Dayal, Jasbani H S; Wright, Sheila et al. (2015) Lysyl Hydroxylase 3 Localizes to Epidermal Basement Membrane and Is Reduced in Patients with Recessive Dystrophic Epidermolysis Bullosa. PLoS One 10:e0137639
Dickson, Patricia I; Tolar, Jakub (2015) The individual (single patient) IND for inborn errors of metabolism. Mol Genet Metab 116:1-3
Wyvekens, Nicolas; Topkar, Ved V; Khayter, Cyd et al. (2015) Dimeric CRISPR RNA-Guided FokI-dCas9 Nucleases Directed by Truncated gRNAs for Highly Specific Genome Editing. Hum Gene Ther 26:425-31
Tolar, Jakub; McGrath, John A (2015) The three-body problem of therapy with induced pluripotent stem cells. Genome Med 7:15
Osborn, Mark J; Gabriel, Richard; Webber, Beau R et al. (2015) Fanconi anemia gene editing by the CRISPR/Cas9 system. Hum Gene Ther 26:114-26
Tolarova, Marketa; Tolar, Jakub (2015) From Mesoderm to Mesodermatology: Bone Marrow Mesenchymal Cells Heal Skin Wounds. Mol Ther 23:1283-4
Tolar, Jakub; Wagner, John E (2015) A biologic Velcro patch. N Engl J Med 372:382-4

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