Development of novel F KBP12 ligands as t herapeutic candidates for D uchenne and B ecker muscular dystrophies.

Abstract

DMD is an inherited X-linked disorder caused by the absence of dystrophin protein. Post-translational modifications guided by oxidative stress in RyRs result in the dissociation of FKBP proteins from the complex, causing an aberrant calcium leak from the sarcoplasmic reticulum. AHKs constitute a new family of FKBP12 ligands, specifically designed to FKBPs to enhance their binding to RyR channels. Furthermore, they have the potential to function as antioxidant molecules. The main objective of this thesis is the identification of a lead compound among AHKs. To this end, we have first implemented an in vitro platform based on impedance measurements to study the myogenesis of control and dystrophic human immortalized myoblasts. Furthermore, we have implemented HTP calcium measurements in these plates, and found characteristic alterations in calcium homeostasis in dystrophic myotubes. On the other hand, we have used impedance measurements to determine the in vitro toxicity of five AHKs. Regarding in vitro efficacy, we have studied the antioxidant capacity of AHKs to protect from H2O2-induced cytolysis in myotubes, finding that they have the ability to protect against oxidative stress. We have also studied AHKs efficay as calcium modulators, finding that they can restore the calcium content in the compartments of dystrophic myotubes. Finally, we have seen that AHK1 significantly reduces elevated basal cytosolic calcium levels, as well as the arrhythmic behavior of mdx cardiomyocytes. In the second section of this thesis, the bioavailability of five AHKs after oral administration in mice has been studied. In addition, the efficacy of AHK1 to reduce the cardiac deficits in mdx mice has been explored.