Absence of p.R50X Pygm read-through in McArdle disease cellular models

Area of Science:

• Genetics and Molecular Biology
• Rare Diseases
• Biochemistry

Background:

• McArdle disease is an autosomal recessive disorder characterized by impaired muscle glycogen breakdown due to a deficiency in muscle glycogen phosphorylase.
• This deficiency leads to exercise intolerance and muscle pain, highlighting the need for effective therapeutic strategies.

Purpose of the Study:

• To evaluate the efficacy of various read-through agents in restoring functional muscle glycogen phosphorylase in cellular models of McArdle disease.
• To investigate the influence of the premature termination codon (PTC) context on read-through efficiency in the PYGM gene.

Main Methods:

• Utilized three distinct cellular models: HeLa cells with GFP-PYGM constructs, HEK293T cells stably expressing a GFP-PYGM construct, and primary skeletal muscle cultures from a McArdle mouse model.
• Tested the efficiency of amlexanox, Ataluren, RTC13, and G418 as read-through agents.
• Analyzed the nucleotide sequences surrounding the PYGM p.R50X mutation's stop codon (TGA) and compared them with known read-through influencing contexts.

Main Results:

• No detectable levels of read-through were observed with any of the tested agents across all evaluated cellular models.
• The PYGM p.R50X mutation context (TGA stop codon, G at -1 and -9, C at -3) presented potentially favorable features for read-through.
• However, the presence of C at position -2 and absence of a nucleotide at +4 may counteract efficient read-through induction.

Conclusions:

• The evaluated read-through agents demonstrated no efficacy in restoring PYGM gene function in McArdle disease cellular models.
• The specific PTC context of the PYGM p.R50X mutation may not be sufficiently permissive for read-through induction by current agents.
• Further research into PTC context elements and novel read-through strategies is warranted for potential McArdle disease therapies.