A COX-INHIBITING NITRIC OXIDE DONOR TO COUNTERACT FUNCTIONAL MUSCLE ISCHEMIA IN DUCHENNE/BECKER MUSCULAR DYSTROPHY: TRANSLATIONAL RESEARCH FROM MICE TO MEN

Duchenne/Becker muscular dystrophy (DMD/BMD) is a spectrum of devastating X-linked muscle wasting disease for which there is no specific treatment. Accounting for more than 80% of all cases of muscular dystrophy, DMD/BMD lead to shortened life spans with death in DMD often by age 20 due to respiratory muscle weakness or cardiomyopathy and in BMD before age 60 due to cardiomyopathy. Twenty-five years have passed since dystrophin was found to be the disease-causing gene in both conditions. Since then, basic/preclinical research has flourished, but clinical translation has not. Our previous work -- parallel translational experiments in mouse models and patients with muscle diseases including DMD -- showed that loss of sarcolemmal nitric oxide synthase (nNOSu), a dystrophin-associated protein, renders the diseased muscle fibers susceptible to functional muscle ischemia and implicated skeletal muscle-derived nitric oxide (NO) as a novel therapeutic target for these refractory conditions. Subsequently, other groups have shown that genetic and pharmacologic strategies that enhance NO signaling can ameliorate many features of the dystrophic phenotype -- at least in mice. Whether this compelling basic/preclinical research will translate into benefit for patients with DMD/BMD and which NO-boosting strategy will work best are key unanswered questions. Recent studies in mouse models of muscular dystrophy, including the mdx mouse model of DMD/BMD, have shown that long-term treatment with cyclooxygenase-inhibiting NO-donating drugs (CINODs) improved muscle morphology and reduced muscle necrosis, inflammation, and fatigue. We recently completed an initial study showing that short-term treatment with the CINOD HCT 1026, a NO-donating flurbiprofen, dramatically reversed functional muscle ischemia in mdx mice. Translation of these findings to DMD/BMD patients is hampered by flubiprofen's gastrointestinal side effects and lack of approval for pediatric patients. Thus, we now propose new studies to explore the translational potential of a safer CINOD -- naproxcinod, a NO-donating naproxen. Our major new hypothesis is that naproxcinod constitutes a highly effective novel strategy to eliminate functional muscle ischemia and thereby restore normal blood flow regulation in exercising skeletal muscle of mdx mice and of patients with DMD/BMD. We further hypothesize that the improved muscle perfusion is a major mechanism by which naproxcinod slows disease progression and improves early subclinical cardiac dysfunction in the mdx mouse. In Aim 1, we will determine if short-term treatment with naproxcinod improves muscle blood flow regulation by restoring NO signaling in mdx mice. We will perform: (a) acute dose-finding studies to determine the optimal naproxcinod treatment that will prevent muscle ischemia during hindlimb contractions; (b) acute mechanistic studies to evaluate the role of naproxcinod's NO-donating moiety to elevate skeletal muscle nitrite and nitrate, which we hypothesize are reduced back to NO in the hypoxic and acidic environment of the contracting muscles; and (c) acute functional studies to determine if the anti-ischemic effect of naproxcinod improves exercise tolerance and protects both skeletal muscle and cardiac muscle from stress-induced injury. In Aim 2, we will determine if the short-term benefits of naproxcinod persist during long-treatment without development of untoward side effects. We will conduct a 6-month preclinical trial of naproxcinod versus naproxen in mdx mice subjected to chronic exercise stress to exacerbate the typically mild mdx phenotype so that it more closely mirrors the more severe human disease. In Aim 3, we will begin to directly translate the preclinical studies in mdx mice to human patients with BMD and DMD. We will determine if short-term treatment with naproxcinod will reverse ischemia of the forearm muscles during handgrip exercise, first in adult BMD patients and then in pediatric DMD patients. In contrast, we predict that equimolar naproxen will be without effect. If the results of both the preclinical studies in the mdx mouse and the proof-of-concept studies in the BMD/DMD patients show that naproxcinod improves muscle blood flow regulation without detrimental side effects, we will quickly leverage these data to apply for funding for future placebo-controlled phase 2A trials of naproxcinod in BMD and DMD and then a multicenter Phase 2B trial in DMD. As no new drug development is required, this pharmacological approach has the potential to quickly impact clinical practice. Recognizing that naproxcinod will not cure DMD/BMD, the hope is that the drug will allow patients to perform more work with less muscle injury and fatigue, thereby slowing disease progression and improving quality of life. If cardiac protection is also afforded, preemptive naproxcinod therapy could delay heart failure onset and extend life. The potential for meaningful clinical impact is high.