Professor and Director of Research
Keck School of Medicine
- Gene Therapy
- Extracellular Matrix
- Stem Cells
- Wound Healing
- Regenerative Medicine
- Autoimmune Disease
Research OverviewOur current specific research involves structures in the skin called anchoring fibrils. These structures are critical for the adherence of the outer skin epidermal layer to the inner dermal connective tissue layer. Anchoring fibrils are composed of type VII collagen. When children are born with defects in the gene that encodes for type VII collagen, they develop a disease of the skin called dystrophic epidermal bullosa (DEB). DEB is characterized by skin fragility, chronic blistering of the skin, scarring and aggressive squamous cell carcinomas. It is incurable. In the last few years, we made significant progress in developing various therapeutic strategies for DEB. First, in an effort to develop ex vivo gene therapy for DEB, we applied a highly efficient lentiviral gene delivery approach to restore type VII collagen expression in RDEB keratinocytes and fibroblasts. This corrected the RDEB cell phenotype in vitro. We then used these gene-corrected cells to regenerate a human skin equivalent transplanted onto immunodeficient mice. Human skin regenerated by gene-corrected RDEB cells demonstrated restoration of type VII collagen expression and anchoring fibril formation at the dermal-epidermal junction (DEJ) in vivo.
Second, in an effort to develop a cell-based therapy for DEB, we showed that intradermal injection of normal human or gene-corrected RDEB fibroblasts into mouse skin resulted in the stable expression of human type VII collagen at the mouse DEJ. Third, in an effort to develop an in vivo gene therapy, we engineered a self-inactivating lentiviral vector expressing human type VII collagen and injected this vector intradermally into hairless, immunodeficient mice and into human DEB composite skin equivalents grafted onto immunodeficient mice. A single lentiviral vector injection provided stable type VII collagen at the BMZ for at least 3 months and reversed the DEB phenotype. Lastly, in an effort to develop a protein-based therapy for DEB, we intradermally injected human recombinant type VII collagen into mice. The injected human type VII collagen stably incorporated into the mouse’s BMZ and formed anchoring fibrils. Further, intradermal injection of recombinant type VII collagen into transplanted human DEB skin equivalents also stably restored type VII collagen expression at the BMZ in vivo and reversed RDEB disease features. Our studies provide the first evidence for using protein therapy to correct a skin disease due to a gene defect in a structural protein. All these studies resulted in the several publications in the highest caliber biomedical journals including Nature Genetics, Journal for Investigative Dermatology, Molecular Therapy and Nature Medicine. In summary, studies from last few years provide strong in vitro and in vivo evidence for potential therapeutic strategies for DEB in an intact mouse model or in mice transplanted with a human DEB skin equivalent. Prior to testing any of these approaches for DEB in humans, we need to utilize a preclinical animal model to determine the safety and efficacy of these approaches and address potential immune responses. The aims of our current studies are: 1) To verify the feasibility of protein-based therapy for DEB using DEB mouse and dog models, 2) To determine the safety and efficacy of lentiviral vector-based in vivo gene therapy in DEB animal models, 3) To validate the fibroblast-based approach for correction of RDEB defects in these animal models, 4) To characterize immune responses to a neo-antigen and develop strategies to blunt these responses in the DEB mouse model, and 5) To evaluate the feasibility of intravenous injection of gene-corrected fibroblasts that home to skin for DEB treatment. These studies will advance the prospects for therapy for DEB patients and bring therapy for DEB one step closer to reality.
Anchoring fibrils are also involved in an adult acquired disease called epidermolysis bullosa acquisita (EBA). In this disease, the patient makes IgG autoantibodies against his or her own type VII collagen in the anchoring fibrils. The result of this autoimmune disease is the same as genetic DEB, namely skin fragility, blister formation, scarring and chronic skin wounds. We has also developed animal model of EBA by passively transferring the affinity purified EBA patients’ anti-type VII collagen autoantibodies into the mice and inducing the blistering disease in the animals. This murine model, with features similar to the clinical, histological and immunological features of EBA, will be useful for the fine dissection of immunopathogenic mechanisms in EBA and for development of new therapeutic intervention