Department of Neurology


Deimmunization of human T-cell reactive dystrophin epitopes

A significant hurdle for DMD gene therapy for many patients is the likelihood of a T-cell mediated immune response against micro-dystrophin (µDys).  We seek to avoid this problem via a Rosetta-based protein immuno-silencing & redesign protocol that integrates human immune epitope data, MHC epitope or antigen prediction tools (structure-based design) & proteomic data (sequence-based design) to derive µDys proteins with highly reduced immunogenicity while retaining stability & function.  Our proposal builds on preliminary data demonstrating µDys can be redesigned to eliminate immunogenicity of a human T-cell epitope, (previously identified in a clinical trial), while retaining functional properties in dmd mice. Importantly, the algorithm used in our studies predicted T-cell responses to the same epitope, independently of its detection in the clinical trial. The application of T-cell epitope redesign to internally truncated dystrophins also applies to novel junctions in dystrophin resulting from exon-skipping or Crispr/Cas9-based approaches.

Combinatorial therapies toward alleviating cardiopulmonary deficits in an advanced model of DMD

DMD cardiomyopathy is a common feature developing in approximately the second decade of life.   As supportive therapies for respiratory function improve, cardiomyopathy, resulting in heart failure, has become a primary cause of death in an ever increasing number of patients. The goal of this project is to provide important information towards a potentially translatable cardiac therapy for DMD patients with advanced disease; likely not responsive toward skeletal muscle gene replacement therapies alone.  As such, we aim to comprehensively evaluate and characterize the potential synergy between two promising gene therapy approaches on ameliorating cardiopulmonary performance, independently in young and advanced aged dystrophic dmd mice (a model of end-stage Duchenne dilated cardiomyopathy).  Addressing the underlying genetic defect of DMD (the lack of dystrophin), thereby protecting myofibers from further injury, while simultaneously improving the relative magnitude & rate of cardiac contractility via alternative nucleotide therapy (ribonucleotide reductase) with ionotropic enhancement, potentially provides for a unique therapeutic advantage.

The potential role of dystrophin family protein members within the DGC

Alpha dystrobrevin (α-Db), a dystrophin superfamily member, is a component of the dystrophin-glycoprotein comlex (DGC) within striated muscles. Decreasing amounts of α-Db from the sarcolemma contributes to severity of disease in several muscular dystrophies including Duchenne muscular dystrophy (DMD). We have shown by whole-body recombinant adeno-associated viral vector (rAAV) gene transfer into α-Db null mice, that a formerly understudied isoform, α-DB3, becomes integrated within the DGC, localizes to costameres, prevents muscle degeneration, & restores diaphragm function. The association of α-Db3 within the DGC appears essential for its function, whose absence likely contributes to sarcolemma fragility & susceptibility to injury in multiple muscular dystrophies.  Ongoing efforts are aimed at elucidating novel protein-protein interactions that facilitate retention of α-Db3 to the DGC &/or costameres, which in turn lead to functional benefits and prevention of myopathy in α-Db null mice, even in the presence of numerous key DGC components.

Utrophin (Utrn), an additional dystrophin superfamily member, holds a high degree of sequence similarity to that of dystrophin; so much so that over 70 exons spanning > 900 kb reveals intron/exon structural identity.  In normal adult skeletal muscle, Utrn can be found concentrated at the neuromuscular and myotendinous junctions; whereas in newly regenerated myofibers it is also localized to the sarcolemma.  Further, in early fetal development Utrn can also be found localized to the sarcolemma in skeletal muscle at ~11 weeks and wanes at ~23 weeks postnatal when expression of dystrophin ensues. This difference in developmental expression resulted in the proposal that Utrn may be the autosomal fetal form of dystrophin.  Thus, leading to the hypothesis that Utrn may be able to functionally compensate for dystrophin in DMD patients. Implying a gene therapy approach focusing on the exogenous delivery of µUtrn to DMD muscle fibers could be a viable approach for treating DMD.  We have been developing truncated µUtrn vectors capable of being packaged within rAAV that are designed to increase efficacy.  Our lab is continuing studies to develop & refine µUtrn function with the goal of providing a long lasting alternative therapeutic for DMD.