PHARMACOLOGICAL RESCUE OF MISFOLDED PROTEINS: INNOVATIVE APPROACHES FOR THE CURE OF MUSCULAR DISEASES

The key pathogenetic event of an extremely heterogeneous group of genetic diseases, collectively called Unfolded Protein Diseases (UPDs), is the presence of gene mutations that cause either unfolding or misfolding of a coded protein. This usually leads to either toxic gain of function, because of mutated protein aggregation, or loss of function because of the premature disposal of the defective protein by the cell’s quality control system (QCS). Sarcoglycanopathies, in particular type 2D Limb Girdle Muscular Dystrophy (LGMD2D), Brody’s disease (BD) and its bovine counterpart pseudomyotonia (PMT) and recessive Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT) are genetic diseases of striated muscles showing characteristics of UPDs. Despite presenting largely different clinical phenotypes, these disorders share a common feature, i.e., normal levels of the mutated transcripts and almost undetectable levels of the coded products (alpha-sarcoglycan, SERCA1a and calsequestrin2, respectively). Evidence from our laboratory demonstrates that mutants causing LGMD2D (Gastaldello et al Am. J. Path. 2008) and PMT (ongoing results) are substrate of the ER associated protein degradation (ERAD). Unfortunately, at present, no effective therapies are available for these disorders. Aim of this project is to prove the feasibility of a new pharmacological therapy designed “to cure” the mutated proteins of such pathologies either by preventing their degradation (protein rescue strategy) or promoting their folding and trafficking (protein repair strategy). We have identified several ERAD components responsible for the degradation of the V247M alpha-sarcoglycan mutant that might be potential drug targets in the so called “protein rescue strategy”. Knock down of selected ERAD components by using small interfering RNAs, in fact, permits the rescue of the alpha-sarcoglycan mutant. On the other hand, inhibition of proteasome has positive effects also on the recovery of SERCA1a mutant in both in vivo experiments on cellular model and ex vivo experiments on skeletal muscle explants isolated from a PMT affected cattle. Regarding the “protein repair strategy”, we are testing several different small molecules that, by assisting folding, promote the mutant proteins’ maturation and trafficking. First experiments are very promising because drug treatments preserve both SERCA1a and alpha-sarcoglycan mutants from degradation and permit their proper localization. The present project constitutes the proof of principle for the development of innovative pharmacological therapies for the cure of rare muscle diseases for which neither classical nor innovative therapies, i.e., gene therapy, are efficacious or applicable.