Molecular and therapeutic correction of XMEA using novel zebrafish and mouse models

PROJECT SUMMARY/ABSTRACT
The objective of this proposal is to define the molecular mechanisms and identify new therapeutic strategies for
of an understudied class of myopathies, specifically X-linked myopathy with excessive autophagy (XMEA).
XMEA is characterized by elevated levels of autophagy due to disruptions in the autolysosome function. One
MEA of interest is X-linked myopathy with excessive autophagy (XMEA), a rare autophagic vacuolar myopathy
that characterized by progressive proximal muscle weakness, high levels of serum creatine kinase and
accumulation of autophagic vacuoles. XMEA is caused by pathogenic mutations in the VMA21 gene in which N-
terminal loss-of-function variants result in early death by 10 years and milder pathogenic VMA21 splicing variants
result in a slower disease progression. Patients with VMA21 pathogenic mutations have a defective autophagy
and an impaired ability to form the autophagosomes. VMA21 is a subunit of the V-ATPase protein pump and its
disruption results in a failure to properly acidify the autolysosome resulting in the formation of vacuolar inclusions
in XMEA. No extensive biomarker studies have been performed in the XMEA population resulting in a dearth of
knowledge and the lack of suitable XMEA models is a significant barrier towards any effective treatment.
We have generated a Vma21 knock-in (Vma21 KI) mouse model based on an RNA-splice mutation identified in
a set of XMEA patients observed at our Children’s of Alabama muscular dystrophy clinic. Vma21 KI mice have
a progressive muscle weakness, impaired muscle function, and have vacuolar inclusions that form as they age,
which phenocopies the XMEA patient symptoms. In parallel, we generated vma21 mutant zebrafish that have a
severe loss-of-function (LoF) pathology resulting in muscle paralysis, vacuolar inclusion bodies, and early
lethality by 10 days post fertilization (dpf). An autophagy drug library screen of our vma21 mutant zebrafish
identified edaravone, an FDA-approved autophagy and oxidative stress inhibitor for ALS, as the most corrective
compound out of 29 leads for XMEA zebrafish pathologies. This proposal seeks to establish molecular and
therapeutic biomarkers for XMEA based on our analysis of XMEA patient cells, and VMA21-defective zebrafish
and mouse models, with an emphasis on the Vma21 KI mice. Proteomic evaluation of the muscles from Vma21
KI mice will allow us to identify VMA21-dependent factors that progress with XMEA disease status. We also seek
to evaluate the therapeutic mechanism of action for edaravone in a 6 month treatment of our Vma21 KI mice.
These studies seek to establish the XMEA/VMA21 disease processes while advancing a promising autophagy
inhibitor compound to eventually treat these XMEA patients suffering from this devastating neuromuscular
disorder.

Grant Number: 1R03TR005967-01
NIH Institute/Center: NIH
Principal Investigator: MATTHEW ALEXANDER