Dystrophin Expressing Chimeric (DEC) Human Cells Provide a Potential Therapy for Duchenne Muscular Dystrophy

Abstract

Duchenne Muscular Dystrophy (DMD) is a progressive and lethal disease caused by mutations of the dystrophin gene. Currently no cure exists. Stem cell therapies targeting DMD are challenged by limited engraftment and rejection despite the use of immunosuppression. There is an urgent need to introduce new stem cell-based therapies that exhibit low allogenic profiles and improved cell engraftment. In this proof-of-concept study, we develop and test a new human stem cell-based approach to increase engraftment, limit rejection, and restore dystrophin expression in the mdx/scid mouse model of DMD. We introduce two Dystrophin Expressing Chimeric (DEC) cell lines created by ex vivo fusion of human myoblasts (MB) derived from two normal donors (MBN1/MBN2), and normal and DMD donors (MBN/MBDMD). The efficacy of fusion was confirmed by flow cytometry and confocal microscopy based on donor cell fluorescent labeling (PKH26/PKH67). In vitro, DEC displayed phenotype and genotype of donor parent cells, expressed dystrophin, and maintained proliferation and myogenic differentiation. In vivo, local delivery of both DEC lines (0.5 × 106) restored dystrophin expression (17.27%±8.05—MBN1/MBN2 and 23.79%±3.82—MBN/MBDMD) which correlated with significant improvement of muscle force, contraction and tolerance to fatigue at 90 days after DEC transplant to the gastrocnemius muscles (GM) of dystrophin-deficient mdx/scid mice. This study establishes DEC as a potential therapy for DMD and other types of muscular dystrophies.

Cardiac Protection after Systemic Transplant of Dystrophin Expressing Chimeric (DEC) Cells to the mdx Mouse Model of Duchenne Muscular Dystrophy

“…We applied our well-established ex-vivo PEG mediated cell fusion procedure and confirmed fusion efficacy by flow cytometry ( Fig. 1) [31,32]. To confirm fusion, suitable controls of unstained myoblasts derived from wild type mice (MB wt ) and MB (MB mdx ) and MSC (MSC mdx ) derived from the dystrophin deficient mdx mice as well as single stained controls of PKH26 labeled MB wt and PKH67 labeled MB mdx for confirmation of fusion for DEC of myoblast origin ( Fig.…”

“…Our standard polyethylene glycol (PEG) fusion protocol was applied as described before [31,32]. Parent snj (wt) and mdx myoblasts (MB wt and MB mdx ) as well as mdx MSC (MSC mdx ) were harvested and washed in serum-free DMEM culture media supplemented with 1% Antibiotic-Antimycotic solution (Gibco-ThermoFischer, Waltham, MA, USA).…”

“…Stem cell transplants based on the delivery of either autologous or allogenic stem cells have shown promise as an alternative method for DMD treatment, but limited or short-term cell engraftment, allogenic immune response, and side effects of immunosuppressive therapy have all been challenges that these treatments have faced in their path to changing the course of the disease for DMD patients [26][27][28][29][30][31][32][33][34][35][36][37][38][39].…”

“…We have previously reported in both the mdx and mdx/scid mouse models of DMD that local transplantation of Dystrophin Expressing Chimeric (DEC) cells significantly improved dystrophin expression which correlated with significant improvement of strength and functional outcomes [31,32]. Thus, in order to assess the effect of DEC therapy on cardiac function, we applied our established protocol of systemic-intraosseous administration of both DEC lines (MB wt /MB mdx and MB wt /MSC mdx ) into mdx mice [31,32,45].…”

Engraftment of skeletal progenitor cells by bone-directed transplantation improves osteogenesis imperfecta murine bone phenotype

“…One limitation of this study is that a local treatment for a systemic disorder is not an ideal strategy, although local cell therapy has shown promising results in other diseases such as in Duchenne Muscular Dystrophy . With this limitation of local transplantation in mind, several related points are worthy of consideration.…”