ALLOGENEIC CELL-BASED THERAPIES FOR INDIVIDUALS WITH RECESSIVE DYSTROPHIC EPIDERMOLYSIS BULLOSA

Abstract

Epidermolysis bullosa (EB) is a clinically and genetically heterogeneous group of inherited blistering skin diseases that affect 500,000 individuals worldwide. The clinically more severe recessive dystrophic (RDEB) variant affects ~5% of EB individuals with a prevalence of 8 per one million of the population. RDEB is caused by loss-of-function mutations in the type VII collagen gene, COL7A1, which leads to reduced or absent type VII collagen (C7) and structurally defective anchoring fibrils at the dermo-epidermal junction (DEJ). From a clinical perspective, individuals with RDEB have fragile skin and are susceptible to blistering following minimal trauma, which leads to poor wound healing, scarring, contractures and oesophageal strictures. The major cause of mortality in RDEB is metastatic cutaneous squamous cell carcinoma (SCC). At present, care is mainly supportive and there are no effective treatments for this debilitating disease. The main therapeutic challenge, therefore has been to develop gene, protein, cell and drug therapies that are safe, effective and affordable. The basis of this thesis is to evaluate safety and efficacy of allogeneic cell-based therapies and attempt to elucidate their mechanism of action in wound healing in RDEB. To examine if allogeneic fibroblasts promote healing of chronic wounds in RDEB I intradermally injected allogeneic fibroblasts around the wound margins in an individual with RDEB. I demonstrated that these injections result in Heparin-Binding EGF-like Growth Factor encoding gene (HBEGF) and COL7A1 upregulation followed by C7 production. These led to reduction in wound size by 30% at 8 months post injections. HB-EGF, a member of the EGF family, has been implicated in RDEB-associated SCC. I also assessed whether another growth factor, EGF, which is commercially available for human use and which is related to HB-EGF, could upregulate COL7A1 expression in RDEB epidermal cells. I demonstrated in vitro that 100ng/ml EGF at 90 minutes and 10ng/ml at 15 minutes led to 3-fold and 5-fold COL7A1 upregulation in RDEB keratinocytes and fibroblasts respectively. HB-EGF also led to COL7A1 upregulation in RDB keratinocytes (6-fold) but to a lesser extent in RDEB fibroblasts (2-fold). To evaluate the effects of intradermally injected allogeneic fibroblasts in a larger number of RDEB individuals, I conducted a prospective, randomised, vehicle-controlled, phase II clinical trial. Twenty-six wounds in 11 adults with RDEB were injected. Fourteen wounds received fibroblasts and 12 were injected with vehicle only. I showed that allogeneic fibroblasts are safe and lead to a greater reduction in erosion area compared to vehicle within the first 28 days following treatment with a single set of injections to the wound margins, but not thereafter. Finally, I explored the safety and potential of intravenously administered allogeneic bone marrow-derived mesenchymal stromal cells (BM-MSCs) in children with RDEB in an open-label trial. I showed that the infusions were well-tolerated, with no serious adverse events. Although, there was no increase in C7 deposition, children and their parents reported improved wound healing, reduction in blister numbers and pruritus, increased skin resilience to trauma, and reduced pain during dressing changes. In addition, significant reduction in circulating inflammatory cytokines (IL-10, p<0.001; IFN-γ, p=0.04 and IL-17A, p=0.03) was observed. Taken together, these data reveal new insights into the mechanisms of action of allogeneic cell-based therapies in RDEB and provide evidence for their efficacy in wound healing and reducing morbidity in the context of clinical trials.

Date of Award	2017
Original language	English
Awarding Institution	King's College London
Supervisor	John McGrath (Supervisor)