Project Summary/Abstract. Duchenne muscular dystrophy (DMD) is the most common, fatal, X-linked disease
worldwide. Progressive muscle weakness leads to loss of ambulation early in life, followed by death caused by
respiratory or cardiac failure. The susceptibility of dystrophic muscle to severe injury at such a young age, and
shortcomings of recent clinical trials underscore the urgent need to identify effective strategies to preserve mus-
cle function in DMD patients. Anti-inflammatory drugs provide limited, though meaningful, therapeutic impact,
but are accompanied by side effects. Separately, endoplasmic reticulum (ER) stress is emerging as a mediator
of disease progression. The double-stranded RNA-dependent protein kinase (PKR) is a potent driver of muscle
inflammation and ER stress, and is elevated in human and rodent dystrophic skeletal muscle. Inhibition of PKR
elicits powerful anti-inflammatory effects and prevents ER stress in several muscle pathologies. Consequently,
there is a critical need to investigate the role of PKR in disease progression and as a future therapeutic target
for DMD. Our long-term goal is to develop and implement strategies to successfully prevent or treat disease-
related pathologies in DMD. The objective in this application is to determine the role of PKR as a promotor of
inflammation, ER stress, and muscle dysfunction in dystrophin-deficient skeletal muscle. Our central hypothesis
is that inhibition of PKR will attenuate inflammation, ER stress, and disease parameters in dystrophin-deficient
skeletal muscle. This hypothesis was formulated based on existing literature and our own preliminary findings
demonstrating that: 1) PKR expression is increased in muscle of DMD patients and activation is elevated in
muscle from mdx mice; 2) dystrophin deficiency is accompanied by muscle inflammation and ER stress; 3) PKR
inhibition attenuates muscle inflammation, ER stress, and atrophy, including glucocorticoids; and 4) PKR inhibi-
tion protects against inflammation-induced muscle dysfunction. In Aim 1 PKR will be knocked down using adeno-
associated virus (AAV)-mediated delivery of siRNA targeting PKR mRNA. We will measure in vivo and in vitro
functional parameters followed by a biochemical and histopathological analysis in mdx and control mice with our
without glucocorticoid treatment. We hypothesize that knockdown of PKR will attenuate inflammation and ER
stress as well as muscle injury and dysfunction. In Aim 2 the PKR inhibitor imoxin will be administered, with or
without glucocorticoid, to dystrophic mice for four months followed by measures of in vivo limb muscle strength
and respiratory function, in vitro muscle function, and histopathology. We hypothesize that imoxin will attenuate
muscle injury and dysfunction in dystrophin-deficient skeletal muscle, concomitant with reductions in inflamma-
tion and ER stress. This project is innovative because it utilizes a novel approach, not previously considered, to
combat the muscle impairments caused by dystrophin deficiency. This contribution is significant because it will
it will address a critical knowledge gap regarding the mechanisms underlying the pathophysiology caused by
dystrophin deficiency and suggest a novel therapeutic target for the treatment of muscular dystrophy.