Impact and Interplay of Corticosteroid Regimen and Exercise Training on DMD Muscle Function
Phase: PHASE2
Conditions: Duchenne Muscular Dystrophy (DMD)
Sponsor: University of Florida

Protocol for:  
 
Impact and interplay of corticosteroid regimen and exercise training on DMD muscle function  
 
NCT04322357 
 
Date Approved: October 17, 2024 
 
 

University of Florida IRB-01 
  
1. Title: IRB201901339 
 
Impact and Interplay of Corticosteroid Regimen and Exercise Training on DMD Muscle 
Function 
2. Investigators: 
 
Principal Investigator: 
• Tanja Taivassalo, PhD (Physiology & Functional Genomics) 
Co-investigators and study staff: 
• Warren Dixon, PhD (Mechanical & Aerospace Engineering) 
• Kimberly Stubbs, PhD student (under Warren Dixon) 
• Donovan Lott, PhD (Physical Therapy) 
• Sean Forbes, PhD (Physical Therapy) 
• John Sladky, MD (Pediatric Neurology) 
• Lee Sweeney, PhD (Pharmacology & Therapeutics) 
• Krista Vandenborne, PhD, PT (Physical Therapy) 
• David Fedel, PhD, ABPP (Child Psychology) 
• Catherine Powers (Physical Therapy) 
• Samuel Riehl (Physical Therapy) 
 
 
 

3. Abstract: 
 
In the search for effective treatments for patients with Duchenne Muscular Dystrophy (DMD), 
the question of optimal glucocorticoid steroid (GC) administration regimen persists. 
Compelling preclinical data suggests that transient low dose GC administration provides the 
same efficacy with fewer side-effects as daily use, and may have ergogenic impacts on 
muscle similar to exercise. No study has yet investigated the potential of such low-dose twice 
weekly GC use as an optimal therapeutic strategy in boys with DMD. Furthermore, the role 
of exercise is increasingly being considered for boys with DMD. Recent preclinical data 1 and 
our own data in boys with DMD2  have demonstrated that isometric strengthening exercise at 
a low to moderate intensity is not only safe but improves muscle strength and function 
whereas others 3,4 have demonstrated that dynamic cycling exercise is safe and therapeutic 
for boys with DMD. The purpose of this work is to determine the potential of twice weekly 
(2x/week) administration of low-dose GC and whether exercise training can synergize to 
delay disease progression, reverse secondary effects of disuse and induce benefit on muscle 
strength and physical function in boys with DMD. This proposal is innovative in its use of non-
invasive magnetic resonance imaging and spectroscopy to assess the impact of GC and 
exercise on lower extremity muscle remodeling and disease progression in DMD. The 
exercise program combines dynamic cycling and isometric leg strengthening performed at 
the patient’s and supervised remotely by the study team. 
 
Up to 32 steroid-naïve boys (ages 4.0-8.0 years old) will be recruited and randomized to 
undergo the 2x/week GC regimen for 6 months followed by a continuation of the regimen for 
another 6 months with or without exercise training. To compare to the standard daily GC 
dosing regimen, 1-year data from age-matched DMD patients (part of the ImagingDMD 
natural history study) will be used. To compare the interplay of GC regimen and exercise, up 
to 16 age-matched boys with DMD (ages 5.0 to 9.0 years old) on daily GC will be recruited 
to undergo 6 months of exercise. Physical function, blood and urine samples and magnetic 
resonance imaging (MRI) and spectroscopy (MRS) will be used to monitor the intervention 
impact on disease pathophysiology and muscle remodeling. This project has promise to 
impact clinical practice, DMD patient care and quality of life, by providing information on a 
GC regimen at a dose/paradigm different from what is most commonly prescribed, and by 
providing information on exercise prescription and its impact and interplay with GC on 
physical function and disease progression in boys with DMD. Current clinical advice regarding 
exercise is non-specific. Our findings will not only advance the use of exercise in the field, 
but form a foundation for future assessment of adjuvant therapies as clinical trials advance. 
The researchers have extensive experience working with DMD patients, exercise and muscle 
physiology and MRI/MRS. 

4. Background: 
 
Current standard of care in DMD: DMD is a devastating, genetically linked neuromuscular 
disease characterized by rapidly progressive muscle atrophy and weakness, resulting in loss 
of ambulation early in the second decade of life, with death from respiratory muscle or cardiac 
failure occurring in the subsequent decade 5. DMD is currently incurable and associated with 
poor quality of life, and GC are the only pharmacological intervention shown to delay disease 
progression6. DMD is caused by a deficiency of the structural sarcolemma-stabilizing protein 
dystrophin, which when absent leads to mechanical weakening, increased membrane 
damage resulting in cycles of myofiber degeneration, regeneration and eventual replacement 
of muscle with fat and fibrotic tissue 7,8. These circumstances provoke an abnormal 
persistence of inflammatory macrophages within the muscle, contributing to the dystrophic 
pathology. GC are believed to act through immune system modulation and reduction of 
fibrosis within dystrophic muscle 9,10. Treatment with prednisone or deflazacort is 
recommended to start between the ages of 4-6 years, resulting in an apparent improvement 
in muscle strength and ambulation in most boys for 6-12 months, followed by stabilization for 
about 2 years and long-term slower deterioration than untreated patients 10. Despite these 
benefits, long-term daily treatment is associated with a high incidence of clinically adverse 
effects, which include excess weight gain beginning early in disease progression, cushingoid 
features, behavioral disturbances, short stature, osteopenia and risk of fractures, insulin 
resistance/diabetes, and others 11 with the first four features being the most common side 
effects 12 and occurring in 60 to 100% of DMD boys. In a recent retrospective analysis of GC 
use in US males with DMD as part of the Duchenne Registry, 25% of patients reported never 
using GC with the primary reason being concerns about side-effects 13. Another 12% of 
patients reported discontinuing GC treatment with the primary reason being problems with 
side-effects. Concerns about the side effects have led to the use of many alternative 
regimens. Up to 29 different regimens have been identified and are in regular use in clinics 
around the world 14. However data on the efficacy of potential alternative regimens vs. 
placebo, or vs. daily GC, are limited 15.  There is a clear uncertainty concerning the best 
treatment regimen (maximizing benefit and minimizing side effects). Patients and families 
have documented a high level of frustration with the status quo, asking explicitly for more 
information to be generated to guide best practice 10,14.  
 
Previous work on weekend GC dosing: Escolar and colleagues 16 compared a regimen of 
high-dose GC pulses on weekends (5 mg/kg/day on Saturday and Sunday) vs daily (0.75 
mg/kg/day) prednisone in DMD boys aged 4–10 years, reporting that weekend was 
equivalent to daily dosing over 12 months based on similar increases in leg and arm muscle 
strength. Increases in body mass index were still detected on the weekend regimen, but linear 
growth was greater and lumbar mineral density was improved compared to the daily dosing 
group. The authors concluded that the weekend prednisone strategy provides a safe and 
effective alternate to daily dosing. However, the total weekly dose for weekend therapy (10 
mg/kg) was almost two times higher than the total daily dosage each week (5.25 mg/kg). 
There was no justification for this high weekend dose other than it was previously used in 
children with epilepsy with minimal side effects 17. In their lead-up pilot study, 30% of DMD 
boys or their parents reported irritability or sleep disturbance during 1– 2 days following 
prednisone administration, and two children discontinued medication because of the severe 

side effect time locked to 2 days following the high dose given. These findings lead to 
consideration of a lower dose strategy: one that uses the standard daily dose taken twice a 
week as a more optimal alternate GC strategy for patients with DMD.  
Data from animal studies support this lower dose 2x/week paradigm. The impact of a 
single weekly administration of a daily dose was recently compared to daily administration 
over 4 weeks in the mdx mouse model 18. Both dosing regimens improved sarcolemmal injury 
and fibrosis comparably, however weekly dosing also improved muscle performance (time to 
exhaustion on treadmill and grip strength), resistance to fatigue, myofiber cross sectional 
area (CSA) and increased expression of genes associated with muscle remodeling whereas 
daily dosed mice ran less than controls, lost body mass and demonstrated induction of 
muscle atrophy markers. Given the broad range of health outcomes that were differently 
impacted and improved by weekly compared to daily dosing, 2x/week administration of the 
GC daily dose should be considered for DMD patients.  
 
Postulated mechanism of action of weekly GC dosing based on preclinical models: GC 
are known to have both beneficial and detrimental effects on skeletal muscle, with the 
frequency and dose playing an important role in therapeutic outcome. Chronic GC exposure 
induces muscle atrophy pathways19,20, whereas transient exposure has been documented to 
enhance muscle performance and produce ergogenic effects in both animals and humans 
(i.e. doping by athletes) 21-24. The mechanism underlying the therapeutic effect is thought to 
relate to their anti-inflammatory effects, however other mechanisms may be involved. 
Recently, a distinct GC-induced gene regulatory pathway (KLF15 axis) in skeletal muscle 
was identified and reported to mediate the ergogenic effects of GC by regulating muscle 
substrate utilization (lipid metabolism), with no impact on muscle atrophy 20. Results show 
that in dystrophic muscle (both patient and mice), the expression of this key metabolic 
transcription factor (KLF15) was low relative to control, and transient GC administration in 
mdx mice increased KLF15 expression, and this was associated with improved exercise 
performance as measured by treadmill distance/time to exhaustion and grip strength.  
Interestingly, in the above-mentioned study on weekly GC dosing in mdx mice 18, genes 
associated with this KLF15 pathway were upregulated after weekly administration of a daily 
dose of GC over 4 weeks, but downregulated after daily dosing over that time period. The 
authors suggest that weekly GC dosing better harnesses the dual beneficial steroid effect on 
muscle by improving sarcolemmal repair as well as inducing ergogenic remodeling, and that 
when chronic treatment is required, weekly low dose administration may be associated with 
the benefit without driving atrophic pathways 18. 
 
Exercise training as a therapeutic strategy in DMD: Although exercise is well-known to be 
a potent inducer of signaling pathways that lead to remodeling and physiological adaptations 
improving muscle strength and cardiorespiratory fitness in healthy individuals, very little is 
known about its impact in DMD. The few studies that have assessed the impact of either 
aerobic or strength exercise are summarized in 25. Overall, the DMD community has been 
cautious in its approach to exercise training, 26,27. Longstanding clinical concerns have 
appropriately been based on the fact that high intensity and eccentric muscle actions are 
damaging to dystrophic muscle 28, however, similar to GC administration, the frequency and 
dose (intensity) of exercise play an important role in functional and therapeutic outcome. This 
is becoming more recognized in clinical application of exercise to DMD, with recent studies 

indicating that submaximal exercise may be beneficial 3,4. To this point, the “No Use is Disuse” 
study was the first randomized clinical trial (RCT), comparing the effects of assisted cycle 
training to usual care in late ambulatory or recently wheelchair-dependent DMD boys 4. In 
addition to demonstrating safety and feasibility over 24 weeks of exercise, it highlighted the 
potential of exercise to delay functional deterioration of motor abilities (as the usual care 
group demonstrated a 6% decline in that time period). However, improvements in muscle 
function were not detected, likely relating to the low intensity of the assisted cycling exercise, 
which was aimed at reducing the amount of secondary disuse.  
 
Early exploratory studies assessing strength training in DMD revealed improvements in 
muscle strength with no ill-effects and advocated for starting exercise early in the course of 
disease when there is a maximum amount of functional muscle 29-31. Despite these early 
reports, there have been no published reports on strength training in DMD until a recent 
publication by our laboratory2. We assessed a 12-week leg strength training study in seven 
boys with DMD with the aim of determining safety and feasibility of a home-based remotely 
supervised trial (Principal Investigator D. Lott, NIH grant R21 AR064949-01A1). Isometric 
(where the muscle generates force without changing length) exercise was chosen as the 
modality because of the advantage that it does not expose the muscle to potential eccentric 
actions (which are known to cause damage to dystrophic muscle). Patients underwent three 
exercise sessions of their knee extensors and knee flexors per week, ~1.5 hours per session 
at an intensity of 50% of maximal voluntary contraction. Overall compliance to training was 
high (85%) and notably, we detected no evidence of muscle damage (as measured by MRI 
transverse relaxation time, T2, a construct of muscle damage, inflammation, and edema) and 
no increase in serum creatine kinase (a blood marker of muscle damage) after 12 weeks of 
training. Interestingly, improvements in muscle strength and function were detected in the 
boys with DMD after training. Increases in mean peak torque of KE (23%, p< 0.05) and KF 
(17%, p=0.09) were detected and the time to ascend or descend 4-stairs decreased. 
Collectively, these findings demonstrate safety, feasibility and benefit of leg isometric 
Figure 1. Baseline (BL) measures compared to post-training (PT) of the knee extensors (KE) 
and flexors (KF) after 12 weeks of moderate intensity isometric exercise training in DMD boys. 
No increase in MRI T2 was noted (A), but significant improvements occurred for both strength (B) 
and functional ability (C). Differences noted between BL and PT (*p<0.05, **p< 0.01). 

resistance training in DMD but also support the safety of dynamic, aerobic exercise training, 
as aerobic exercise by definition elicits less mechanical load compared to resistance 
exercise.  
 
Potential adaptive impact of exercise on dystrophic muscle: The lack of dystrophin leads 
to numerous cell and vascular abnormalities including mechanical weakening of the 
sarcolemma, abnormal calcium homeostasis, mitochondrial dysfunction, increased oxidative 
stress, apoptosis and functional ischemia 32. Based on preclinical studies, exercise training 
can benefit dystrophic muscle by 1) preventing functional decline caused by secondary 
muscle disuse and 2) modulating cellular and molecular signaling cascades that promote 
muscle adaptation and improve the underlying pathophysiology 33. Aerobic and strength 
exercises typically induce divergent molecular mechanisms and signaling pathways in 
muscle, resulting in oxidative versus hypertrophy phenotypes respectively 34. Aerobic muscle 
have more mitochondria, capillaries, anti-oxidant enzymes and express higher levels of 
utrophin along the sarcolemma, and are more resilient to injury and fatigue as compared to 
fast, glycolytic muscle. Targeting mitochondrial biogenesis and promotion of the oxidative 
phenotype is therefore an emerging and promising physiologically-relevant avenue of 
therapeutic research for DMD, with various pharmacological agents in the research pipeline 
26,33,35. Indeed, exercise in the form of voluntary wheel running can increase muscle utrophin 
levels, mitochondrial function, Type 1 fibers proportions, antioxidants enzymes and fatigue 
resistance in mdx mice 36; whether aerobic exercise induces these adaptations in DMD is 
currently unknown. Strength training increases synthesis of contractile proteins, resulting in 
fiber hypertrophy and increases in specific and absolute maximal force production. In the 
animal literature, the large majority of studies assessing exercise use voluntary wheel running 
as opposed to true strengthening exercise, and report improvements in both aerobic 
pathways (increase in mitochondrial biogenesis,37 and in hypertrophy related pathways 
(increase contractile proteins, 36; specific and absolute force 38,39). Although our recent study 
of isometric strength training resulted in increased leg muscle strength (see above), it was 
not designed to address underlying mechanisms. The detected improvements are likely a 
combination of improved neural recruitment and muscle hypertrophy. This current proposal 
will measure potential increases in muscle cross sectional area as a marker of hypertrophy.  
 
A key obstacle to studying the impact of exercise in DMD boys has been the difficulty to 
noninvasively monitor how dystrophic muscle responds in a repeatable and valid manner 40. 
To overcome this, a key strength of this study is our use of quantitative magnetic 
resonance imaging (MRI) and spectroscopy (MRS), which are emerging as powerful, non-
invasive outcome measures for assessing muscle pathophysiology and disease progression 
in DMD, as primary endpoints to monitor the muscle response to exercise in DMD. Through 
ImagingDMD, a multicenter natural history study of MR biomarkers in boys with DMD led by 
Dr. Vandenborne, they have shown that lower extremity measures are reproducible and 
standardized 41; discriminate patients from controls41, responsive to disease progression 42,43; 
responsive to GC intervention 44 and correlate with clinically meaningful outcomes suggesting 
that quantitative MR biomarkers may be appropriate surrogates of ambulatory function and 
have the potential to help demonstrate efficacy/inefficacy of therapeutic GC in clinical trials 
45. 
 

 Rationale for combined exercise and low dose 2x/week-GC: Considering that >85% of 
Figure 2: Impact and interplay of GC and exercise in skeletal muscle. Transient (low dose WE) 
GC (red) bind to the GC receptor to trigger the KLF15 axis resulting in ergogenic effects whereas 
chronic GC (blue) triggers atrophy signaling. Exercise optimizes the adaptive response; aerobic 
(cycling) induces both the KLF15 axis and promotes an oxidative muscle phenotype with increased 
ability to metabolize fat. Strength exercise stimulates hypertrophy signaling and inhibits atrophy. 
We propose the combination of low-dose 2x/week-GC and exercise will be synergistic, and protect 
the muscle membrane, attenuate muscle injury and preserve muscle function in patients with 
DMD. 
 

DMD patients are on GC16, understanding the interplay between GC dosing strategy and 
exercise is necessary for patient management and therapeutic application. Although this is 
true for both daily as well as alternate GC regimens, chronic GC have been shown to 
decrease muscle protein synthesis in human muscle 46. Given the compelling pre-clinical data 
on the ergogenic effects of weekly/transient GC administration along with findings that one 
bout of aerobic exercise markedly increases KLF15 expression in healthy human locomotor 
muscle 47 and that KLF15 levels are low in DMD patient muscle 20, we propose that the 
combination of 2x/week-GC and exercise may represent an optimal therapeutic strategy DMD 
(see Figure 1). Not only do both strategies converge on the KLF15-axis, but exercise may 
also recruit compensatory pathways increasing mitochondrial biogenesis and the oxidative 
phenotype, or promote hypertrophy and synthesis of contractile proteins and inhibit atrophy 
signaling within muscle thus enhancing the remodeling effect and leading to optimal benefit. 
Combinatorial treatment strategies are increasingly being considered for DMD, and there is 
evidence from preclinical studies that exercise training can augment the effect of 
pharmacological treatment strategies 36,48.  
5. Specific Aims: 
 
Aim 1: To determine the 12-month impact of a low dose (0.75 mg/kg x 2 days of prednisone) 
twice weekly regimen on steroid-associated changes in BMI, DMD muscle pathophysiology 
and physical function.  These data will be compared to the retrospective data of boys with 
DMD who were on daily steroids and previously enrolled in IRB201700056. We hypothesize 
that compared to the standard daily regimen, DMD boys on 2x/week dosing will present with 
less change in BMI, similar slowing of fat fraction increase, and trend for improvement in 
physical function over the 1-year. 
Aim 2: To determine whether a 6-month in-home, moderate intensity exercise training 
program modulates the GC effects on muscle pathophysiology and physical function in 
patients with DMD. The exercise intervention will involve a combination of aerobic and 
isometric leg strength training. We hypothesize that boys with DMD undergoing the exercise 
intervention will show increased physical function, strength and cross sectional area (CSA) 
of the trained muscles compared to boys who do not exercise, and that exercise will result in 
greater attenuation of disease progression and greater muscle remodeling in the 2x/week-
GC exercise group compared to the daily-GC group.  
6. Research Plan:  
This exploratory study will be conducted over 3 years and is designed to compare the effects 
of a 12-month 2x/week-GC regimen (using a dose lower than previously used for weekend 
dosing) to the standard daily GC regimen on change in BMI, disease progression and 
physical function of boys with DMD, and investigate whether lower extremity exercise training 
for 6 months will modulate the GC effects to further improve physical function (see Overview 
of Study design, Figure 3). 
Aim 1 will comprise of up to 32 steroid-naïve patients at baseline, who begin a 12-month 
treatment period with two days of prednisone per week (dose of 0.75 mg/kg per day). Half of 
the group will be randomized to continue 2x/week-GC for another 6 months (Aim 1), whereas 
the other half will begin a 6-month exercise-training program while continuing on the 2x/week-
GC regimen (Aim 2). Patients in Aim 1 will be compared to retrospective data (IRB20170056) 
from up to 32 age-matched DMD boys on daily GC, who have undergone yearly evaluations 

as part of the ImagingDMD (the largest natural history database of MRI assessments in boys 
with DMD). Analysis of this retrospective data will allow us to generate natural history control 
data for the expected yearly change in BMI, physical function and FF progression in boys on 
daily GC. For Aim 2, up to 16 patients already receiving a daily GC regimen will be recruited 
to undergo 6-months of exercise training, allowing for comparison of the impact and interplay 
of exercise and GC-dosing paradigm.  
For the majority of patients in Aim 1, the study involves 4 visits (baseline, 3, 6 and 12 months) 
to the University of Florida. A subset of patients randomized to Aim 1 who are relatively local 
to the study site may be asked to undergo one additional study visit 4 weeks after starting 
twice weekly prednisone treatment in order to explore the time course of improvement (Figure 
4). The benefits of prednisone initiation in steroid-native individuals are known to be rapid. 
Previous studies in steroid-naive boys with DMD treated with daily prednisone at a dose of 
0.75 mg/Kg show improvements in muscle strength and timed function tests as early as 10 
days after starting therapy49 typically peaking by 3 months 50. Similarly, corticosteroid benefits 
on muscle inflammation/edema occur early after initiation in boys with DMD, as suggested 
by a decrease in MRI-T2 at 3 months after treatment 44. Here, in a subset of patients, we will 
confirm whether 0.75 mg/kg administered twice weekly has an impact on supine to stand, 10 
meter walk/run, NSAA as well as MRI-T2 after the first month of treatment. In the subset of 
Figure 3: Overall study design. Orange lines indicate comparison of GC dosing regimen (Aim 1). 
Blue lines indicate comparison of exercise impact and interplay with GC regimen. The 2x/week-GC 
group will be assessed over 4 visits (baseline, 3, 6 and 12 months). After 6-months, patients will 
either continue the 2x/week-GC regimen alone (Aim 1) or in combination with exercise (Aim 2). 
Patients on daily GC  recruited into Aim 2 will undergo a total 2 to 3 visits (baseline, mid way and 6 
months.  

boys undergoing 6 months of exercise, we may ask the participant to undergo one additional 
study visit midway during the cycle training (2 to 3 months after starting) to capture 
adaptations specific to cycle training. This will be based on demonstrating a heart rate 
response to cycling exercise during the home-based, supervised exercise sessions after 
starting the intervention (where heart rate is lower at the same level of exercise). Such a heart 
rate response indicates the exercise training has been done in the right training intensity and 
heart rate zone, and suggests other physiological adaptations improving muscle and 
functional performance have occurred and should be quantified at a study visit.  
Patients who are on daily GC recruited into Aim 2 will undergo 2 to 3 visits (baseline and 6 
months). Patients who demonstrate a heart rate response to cycling exercise during the 
home-based sessions may be asked to return for a mid-study visit to capture adaptations 
specific to cycle training (for a total of 3 study visits). 
  
 
 
 
 
 
 
Prior to the initiation of the exercise intervention (Aim 2), development and finalization of the 
therapeutic exercise chair designed specifically for DMD patients as part of this overall study 
(and conceptualized below in Figure 6) will be done. The design, description and use of this 
exercise chair by patients with DMD as part of this study was deemed a non-significant risk 
by the UF IRB-01 review (see letter from IRB dated 10/2/2019). A prototype exercise chair 
Figure 4: A small group of boys randomized to Aim 1 who live within a 3-hour drive from 
Gainesville, Florida may be asked to undergo an additional study visit 4 visits after starting twice 
weekly prednisone treatment to explore the time-course of early prednisone-related 
improvements. Specifically, on the Friday after the fourth week of dosing, patients may undergo 
MRI, timed function tests and the NSAA. 

has been developed in study collaborator Dr. W. Dixon’s lab in Mechanical and Aerospace 
Engineering. The aim of this initial sub-study is to test the components of the prototype as 
they are being developed (sizing of pedal attachments, ergometer crank, seat height, motor 
resistance options) and to obtain feedback from a maximum of 10 patients with DMD 
regarding comfort and ease of use. Patients will be asked to exercise on the chair for no more 
than 30 minutes while the study team asks for their feedback, ratings of perceived exertion 
and makes measurements of leg length and heart rate using a standard wrist watch or chest 
strap monitor. Patients may be invited back for two more visits (total visits to equal 3 within 
the span of one month) to provide feedback on any adjustments made to the device. To 
monitor the muscle response to exercise, patients may undergo magnetic resonance imaging 
and spectroscopy (as described in study procedures below) after exercising on the chair on 
one of their visits. 
 
 
Study population:  The target population for this study includes ambulatory boys with 
genetically and clinically confirmed DMD. The subject selection is limited to males as DMD is 
an X-linked recessive, genetic disorder leading to progressive muscle weakness in males. 
Although females are carriers, they do not exhibit the same symptoms. Selection is also 
limited to children since the detrimental effects of DMD begin in early childhood and life 
expectancy of these boys is shortened to the early to mid-20s.  The minimum age for study 
entry is 4.0 years. This is based on the notion that Study Aim 1 focuses to determine whether 
low dose 2x/weekly-GC provides a novel, efficacious treatment strategy for DMD. As such, 
given the fact that clinical decisions for GC treatment typically occur between the ages of 4 
to 6 years and to prevent complications related to weaning from daily GC use, only boys who 
are steroid-naïve upon entry into the study (and 6 months prior) will be recruited. As study 
participation is expected to be 1 year, we have set the maximum age range as 8 years.  
 
Within Aim 1, boys who are 5 years old and above will undergo randomization to the 2x/week-
GC regimen with or without exercise (months 6-12 of study) at baseline to allocate 
participants to either group in a 1:1 ratio using the randomization feature in RedCap.   Patients 
who are 4 to 5years old enrolled into Aim 1 and patients who are enrolled into Aim 2 (on daily 
GC) will not be randomized. Because prednisone and deflazacort are the two most commonly 
prescribed GC in the US, and the current use of deflazacort is reported to be slightly higher 
than prednisone use (54 vs 46%) 13, boys on a stable regimen of either GC will be included 
in Aim 2. 
 
For the exercise chair development and feedback sub-study, the age range of boys with DMD 
will differ. We will recruit a total of 10 boys with confirmed DMD who are between 6 and 12 
years of age. The lower limit of 6 years is chosen to be able to ask questions and obtain their 
feedback. We will recruit up to age 12 years in order to get a range of patient heights as the 
intent is to make this exercise chair adjustable for future exercise studies. Boys who take part 
in this sub-study and fit the inclusion criteria below (for Aim 1 and 2) may be eligible to partake 
in the intervention studies. 
 
 

Inclusion criteria: 
• Diagnosis of DMD confirmed by 1) clinical history with features before the age of five, 2) 
physical examination, 3) elevated serum creatine kinase level and 4) absence of 
dystrophin expression, as determined by immunostain or Western blot (<2%) and/or DNA 
confirmation of dystrophin mutation. 
• Aim 1 inclusion is age 4.0 to 8.0 years at enrollment: a lower age limit of 4.0 years is 
selected as clinicians are prescribing daily steroids at this age.  
• Aim 2 inclusion is age 5.0 to 9.0 at enrollment: a lower age limit of 5.0 is selected based 
on our previous experience with compliance to exercise traiing. An upper age limit of 9 
years has been set to allow comparison with boys in Aim 1 who turn 9 years old over the 
1-year intervention. 
• Ambulatory at the time of the first visit, defined as the ability to walk for at least 100 m 
without an external assistive device and able to climb four stairs. 
• Aim 1 only: GC-naïve at baseline (and prior 6 months) 
• Aim 2 only: on stable daily GC regimen for 6 months prior to baseline 
• Followed by a doctor or medical professional who coordinates Duchenne care on a regular 
basis  
 
Exclusion criteria:  
• Contraindication to an MR examination (e.g. aneurysm clip, severe claustrophobia, 
magnetic implants) 
• Presence of unstable medical problems, significant concomitant illness including 
cardiomyopathy or cardiac conduction abnormalities, physical findings, or laboratory 
abnormality (including but not limited to renal insufficiency or impaired hepatic function) 
that, in the Investigator’s opinion, could adversely affect the safety of the patient, make it 
unlikely that the course of treatment or follow up would be completed, or impair the 
assessment of study results  
• Prior history of rhabdomyolysis  
• Presence of a secondary condition that impacts muscle function or muscle metabolism 
(e.g. myasthenia gravis, endocrine disorder, mitochondrial disease) 
• Presence of a secondary condition leading to developmental delay or impaired motor 
control (e.g. cerebral palsy) 
• Presence of an unstable medical condition (e.g. uncontrolled seizure disorder) 
• Behavioral problems causing an inability to cooperate during testing or understand 
exercise instruction 
• Participation in other forms of investigational drug or gene therapy during the period of 
the study (FDA approved therapies are allowed) 
• Use of carnitine, other aminoacids, creatine, glutamine, coenzyme Q10, herbal 
supplements within 3 months prior to enrollment. Note: Vitamin D, calcium, ranitidine and 
Tums are allowed 
• Use of human growth hormone within 3 months prior to Day 1 
 
Study control subjects:  In addition to the recruitment of above-mentioned patients with 
DMD, we will leverage existing data from the ImagingDMD natural history study (led by Dr. 

Vandenborne; R01 AR056973) where patients with clinically and genetically confirmed DMD 
have undergone yearly assessments (functional testing and MRI/MRS evaluation similar to 
assessments included in this current proposal). Data from up to 32 patients who are age-
matched, ambulatory, on daily GC therapy, and similar exclusion criteria to above will be 
selected from this database (these individuals have previous enrolled in IRB201700056). 
Analysis of this retrospective data will allow us to generate natural history data for the 
expected yearly change in BMI, physical function, MRI and MRS based markers of disease 
progression.  
 
Subject recruitment: Although DMD is considered a rare muscle disease, it is one of the 
most frequent genetic conditions affecting approximately 1 in 3,500 live male births. As with 
any rare disease, having access to the population of interest and the ability to safely 
implement all experimental procedures are concerns, and a potential barrier to accrual of 
patients include competing studies for steroid-naïve patients with DMD. The University of 
Florida has a history of strong recruitment, extensive experience and successful studies in 
boys with DMD. Patient recruitment will come from various sources, as our group has 
developed an extensive collaborative network with links to the Muscular Dystrophy 
Association Clinic at University of Florida as well as nationally and internationally in Canada, 
Parent Project Muscular Dystrophy, Duchenne Connect as well as close ties with other 
investigators in the dystrophy community. In addition, we currently have over 180 patients 
enrolled in our ongoing natural history study (Imaging DMD) from which recruitment may be 
possible.  
Participants will be recruited at the University of Florida using multiple, direct 
solicitation methods. Flyers with the study information will be distributed in clinics and in 
strategic places in hospitals (i.e. waiting rooms, physical therapy areas). An invitation letter 
will be mailed to all families of boys with DMD who may have a sibling or are in the Multi-site 
study (R01 AR056973), titled “Magnetic Resonance Imaging and Biomarkers for Muscular 
Dystrophy” (PI Vandenborne). A similar recruitment letter will be mailed to all patients with 
DMD who meet the age requirements for the study and are registered with the respective 
regional offices of the Muscular Dystrophy Association, Parent Project Muscular Dystrophy, 
and DuchenneConnect. 
 
Screening procedures:  Prospective participants will be screened by the PI and study 
physician to ensure they meet the criteria for participation in the study. Parents or guardians 
who contact the PI and/or the research team will complete a telephone screening interview 
to assess eligibility. Subjects that meet initial eligibility criteria will be mailed the consent form 
and a complete description of the study as well as potential dates for the baseline 
assessment. Informed consent/assent for participation in the study will be obtained on-site 
during the first visit according to the guidelines of the Institutional Review Board. Oral and 
written information will be provided that explains in simple terms 1) the criteria for 
participation, 2) the purpose of the research and the procedures involved, 3) the subject’s 
right to withdraw at anytime without penalty, 4) potential benefit to the subject, 5) potential 
risks, and 6) assurance of anonymity. The parent and the subject will be encouraged to ask 
questions. Written, informed consent will be obtained from the child’s parent or legal guardian 
and the child will provide written assent. The study will not begin until full approval has been 
received by the Institutional Review Board. 
 

Study Variables: The primary and secondary endpoints are listed in Table 1. All study 
procedures will take place at the Clinical Research Center within the Clinical Translational 
Research Building (CTRB) with the exception of the MRI and MRS which will take place in the 
Advanced Magnetic Resonance Imaging and Spectroscopy (AMRIS) Facility at the University 
of Florida. The study drug prednisone will be dispensed by the Investigational Drug Service 
at UF Shands Hospital. During the study visits, digital photo or video recordings may be made 
depending on participant consent for educational purposes and to assess the validity of the 
physical activity bout. More specifically, we will take pictures/videos of each patient on the 
device at the study visit, and share with them so that they understand proper assembly and 
use of the the exercise device when used at home. We may also share the photo or video 
recording with clinicians and other researchers in the DMD community, or families and 
patients with DMD who inquire about participating in the study. Because no such exercise 
device currently exists for patients with DMD, and there is little information about the potential 
benefits of exercise in the DMD community, showing images and videos of the exercise 
device (in patients who have provided their consent) will be educational and advance the 
importance of understanding the role of exercise in this patient population. 
 
 
Aim 1 will consist of 4 to 5 visits (baseline, 3, 6 and 12 months with a possible visit at 8-9 
months to assess exercise training progress) and Aim 2 will involve 2 visits (baseline and 
end-study) with a possible third (mid-exercise) visit at 2-3 months after baseline to assess 
exercise training progress. The baseline and 3 month visits in Aim 1 will involve testing over 
2 days; for patients randomized to no exercise intervention, the 6 and 12-month visits will 
also involve 2 days of testing. For boys randomized to the exercise intervention (Aim 2), the 
6-month visit will require 4 days (and involves familiarization to the exercise device and 
intervention); the mid-exercise and 12-month visit may involve up to 2 days of testing. For 
boys on daily GC who will undergo the exercise intervention as part of Aim 2, the baseline 
visit will require 4 days (familiarization to exercise device and intervention) and the mid-
exercise and 6-month visit will require up to 2 days. See table 1 for schedule of procedures.  
 
Aim 1 
The purpose of this aim is to determine the impact of a low dose, twice weekly regimen of 
prednisone over the course of 1 year in initially steroid-naïve boys with DMD. Primary and 
secondary measures for side-effect and efficacy outcomes are listed in Table 1. We 
hypothesize that compared to the standard daily regimen, DMD boys on 2x/weekly dosing 
will present with less change in BMI, similar slowing of fat fraction increase, and trend for 
improvement in physical function over the 1-year. A table of study procedures relating to Aim 
1 is shown in Table 2. 
 
Following the consent/assent procedure, each patient will undergo baseline safety 
measures which will serve as comparison for development of prednisone-related side-
effects. First the Study physician will do a clinical exam of the patient which involves 
assessment of the patient’s general appearance; skin; head, ears eyes, nose, throat; lymph 
nodes; heart; lungs; abdomen; extremities/joints; and neurological and mental status (as 
needed). The Study physician or research nurse will obtain the patients’ vital signs (sitting 
blood pressure (BP), heart rate (HR) , respiratory rate and temperature) and a 12-lead 

electrocardiogram. The 12-lead ECG will be obtained only after the patient is positioned 
supine, resting, and quiet for a minimum of 15 minutes. After measurement of weight and 
height, the research nurse will perform a venipuncture in the arm. Blood samples (2 
tablespoons) will be analyzed in the Shands hospital to rule out diabetes and in our lab for 
creatine kinase (CK), myoglobin, troponin I, ALT/AST and inflammatory profile 
determination. A urine sample will also be collected by the patient using a standard urine 
collection cup for analysis of metabolites (creatine, creatinine, citrate levels amongst 
others). If available, a portion of these samples (1 tablespoon blood and 1 tablespoon urine) 
may be stored for up to 5 years for future analysis of exploratory biomarkers of disease and 
intervention impact. At the 6 and 12-month time points, patients will under a test of adrenal 
reserve (low dose ACTH stimulation test) to assess whether the twice weekly steroid 
regimen has an impact on the normal functioning of the adrenal gland cortisol secretion. 
This test is done by measuring the cortisol level in blood before and after an injection of 
ACTH (Cortrosyn). Daily dosing with steroids is known to cause adrenal insufficiency as 
measured by this test. We are performing this test to demonstrate the additional safety of 
twice weekly steroids in DMD (and expect a normal cortisol response to this test). The test 
will involve IV placement with baseline blood sampling for ACTH and cortisol levels, 
administration of 1 mcg of cortrosyn (in 2-5 ml of 0.9% sodium chloride injection over a 2-
minute period), with another blood sample taken at 30 minutes post cortrosyn 
administration for measurement of cortisol levels. This test involves less than 1 tablespoon 
of blood.  
 
 
Thereafter, the patient will have a dual-energy x-ray absorptiometry (DEXA) scan for 
measurement of bone density and lumbar spine Z score.  
 
Following these clinical measures, patients will undergo quantitative MRI and MRS for 
determination of efficacy outcomes reflecting disease progression. Characterization of thigh 
muscles will be performed using a 3T whole body MRI scanner (Philips Elition 3T) within the 
AMRIS facility. Patients will be positioned supine in the magnet for up to 60 minutes allowing 
for assessment of 3 measures in muscles of the leg: fat fraction, MR transverse relaxation 
time (T2) and cross sectional area (CSA). Specific quality assurance and standardization 
procedures have been developed for each MR sequence by our group 41,43 for determination 
of these measures. Determination of FF (FF=lipid / water + lipid) within the vastus lateralis 
(VL) will be done using single voxel localized 1H spectroscopy (1H-MRS) as previously 
described 43,51. 1H-MRS T2 will be quantified used a spectroscopic relaxometry sequence, 
and is a measure that is independent of lipid infiltration and used to reflect primarily 
inflammation and edema in muscle 43. The VL will serve as the target muscle group for MRS 
data acquisition, as we have the most longitudinal natural history data on it and it will be 
targeted for the exercise intervention. MRI 3-point Dixon imaging, which is also increasingly 
being used to quantify FF in multiple muscles simultaneously, will be used for cross sectional 
area (CSA) determinations of upper leg muscles. Regions of interest will be drawn around 
the borders of the knee extensor (KE) and knee flexor (KF) muscles (recruited and targeted 
by both exercise training modalities) from Dixon images. CSA and contractile area (CSA -( 
CSA *fat fraction) will be determined as previously reported by our group 52,53. Determination 
of CSA (highly reproducible41) will allow calculation of specific force (CSA /peak strength) of 

KE and KE. MR outcome data analyses will be performed as per standard technique in our 
laboratory 54, 41,43.  
 
Patients will be given a rest after the MRI/MRS and then undergo pulmonary, muscle and 
functional performance tests back at the CTRB which will serve as efficacy outcomes. 
Strength of the respiratory muscles will be measured through standard pulmonary functions 
testing. Spirometry will be performed using a Carefusion Microlab (San Diego, CA) portable 
spirometer in the sitting position to assess forced vital capacity (FVC) and peak expiratory 
flow (PEF). The maximal inspiratory pressure (MIP), and maximal expiratory pressure 
(MEP) will be performed in sitting using a Carefusion MicroRPM device (San Diego, 
CA).  There is no minimal threshold for FVC and FEV1 in order for the patient to proceed 
with subsequent exercise; these values are needed to calculate exercise related 
parameters that elucidate whether the respiratory system limits exercise capacity in a given 
patient. 
 
 
The primary outcome measure will be The North Star Ambulatory Assessment (NSAA), an 
outcome instrument to measure ambulatory function (that range from standing to running) in 
DMD 55. 12-month longitudinal data are available in patients with DMD 56 including in our 
ImagingDMD cohort. The time for the 10-meter walk/run from the NSAA will be used as a 
secondary measure of functional mobility. The time to climb a standard four stairs and 
6MWT, both standard tests to measure functional and ambulatory performance in DMD 58 
will also be assessed. As an exploratory measure, we will perform the Neuromuscular 
Gross Motor Outcome (GRO), which is designed to assess whole body strength, motor 
development and function for all levels of disability across the lifespan in individuals 
diagnosed with neuromuscular disease. It includes measures performed as part of the 
NSAA along with other simple tasks that require between 10-15 minutes to administer. The 
GRO was recently validated for patients with Spinal Muscular Atrophy {Alfano, 2021 #466} 
and is being assessed as a clinical tool to quantify function in DMD. During the walking 
physical activity tests, patients may also be asked to wear a mask that is connected to a 
portable metabolic cart (the Cosmed K-5 which is like a small backpack strapped to the 
participant’s back that weighs less than 2 pounds). This will allow us to measure the 
intervention impact on metabolic and breathing variables.  
 
Additionally, peak isometric strength of KE and KF will be tested using a quantitative Biodex 
dynamometer as previously done by our group reliably in boys with DMD aged 5-14 years 
52,57. Muscle specific force will also be determined by normalization to maximal cross-
sectional area (CSA) from MRI.  
 
Peak aerobic capacity will be measured during a cardiopulmonary exercise test performed 
on a stationary cycle ergometer. Briefly, before the exercise, we will have the patient perform 
standard breathing tests using spirometry, where we measure forced vital capacity and peak 
expirotory flow. Following this, patients will do a brief warm-up of up to 5 minutes of unloaded 
pedaling, and then asked to pedal at moderate intensity for up to 20 minutes. The exercise 
protocol consists of variable watt/minute ramp increase to volitional exhaustion. Watt 
increments will vary between 5-20 watts per minute intervals, depending on participant ability 

and fitness level. Patients will wear a mask that is connected to a standard metabolic cart to 
measure different breathing variables that include oxygen consumption, carbon dioxide 
production, heart rate, respiratory exchange rate (RER), minute ventilation and ventilatory 
(anaerobic) threshold. A 12-lead ECG and blood pressure are monitored by a research nurse 
(who is trained in reading ECG during exercise) at baseline, throughout the test, and for 10 
minutes of recovery. Throughout the exercise, we will ask the patient to rate his level of 
perceived exertion by pointing to a number on the OMNI scale of perceived exertion. Exercise 
will be terminated if the patient reaches 85% of age-predicted heart rate (calculated as 220-
age).   
 
 
Finally, questionnaires will be administered. The Child Behavior Check List (CBCL) will be 
used to monitor behavioral changes as previously done in boys with DMD 16. This is a parent-
report questionnaire 59 rating various behavioral and emotional problems in children aged 4 
through 18 years. The SIDECORT (side effect of corticosteroids), self-report questionnaire 
on perceived cognitive, behavioral and emotional side effects of GC will also be administered 
to the patient and parent 60. As health-related quality of life has become one of the most 
important goals of disease management, the PedsQL Multidimensional Fatigue Scale, which 
assesses fatigue with a total of 18 items and has a form specific to young children (5-7 years) 
will be used 61.  We will also ask 9 questions to the child’s caregiver (Caregiver Impression 
of Change) to obtain their impression of the intervention impact on their child’s overall well-
being, mood, sleep, etc.  
 
We will use Actigraph accelerometers to quantify physical activity (i.e. steps taken per day, 
percentage of time spent in varying intensities of activity, etc.). Each subject will be provided 
an accelerometer and will be asked to wear the accelerometer for all waking hours (except 
bathing and swimming) for 7 consecutive days as we have previously done48 at three time 
points over the course of the study (during the first week of the intervention, prior to the 6-
month visit, the last week of the intervention).  A self-addressed, postage paid return envelope 
for the device will be included to enable the subject an easy means to return the 
accelerometer to the testing site, or they will be asked to return it with them to the study visit. 
A member of the research team will contact the subject once before and once during the 7 
days of wearing the device (and be available for contact should the subject have questions 
regarding the device), and contact information will be provided to the subject to communicate 
with the research team should any questions or concerns arise during the time of physical 
activity monitoring.  Data on daily step count and time spent in sedentary behavior as well as 
varying intensities of activity will be assessed, and changes in any parameter will be 
correlated with changes in peak aerobic capacity, endurance and other measures obtained 
from exercise and functional testing. Aim 2 
The purpose of this aim is to determine the impact of a 6-month in-home exercise training 
program (involving a combination of aerobic and isometric leg strength exercises) in boys 
with DMD, who either take low dose twice weekly prednisone or daily GC (prednisone or 
deflazacort). We hypothesize that boys with DMD undergoing the exercise intervention will 
show increased physical function, strength and cross sectional area (CSA) of the trained 
muscles compared to boys who do not exercise, and that exercise will result in greater 

attenuation of disease progression and greater muscle remodeling in the 2x/weekly-GC 
exercise group compared to the daily-GC group.  
 
A table of study procedures relating to Aim 2 is shown in Table 3.  In addition to all the testing 
described above for Aim 1, patients will undergo an additional MR assessment after the other 
measures (fat fraction, T2 and CSA) have been obtained. This involves 31P MRS to measure 
the recovery kinetics of phosphocreatine following a brief bout of leg exercise in the magnet, 
as this reflects the mitochondrial capacity for oxidative phosphorylation and will serve as a 
marker of energetics. 31P MRS is done using a surface coil secured over the VL and after 
appropriate localization and shimming of the muscle, unlocalized spectra will be collected at 
rest, and after a brief (< 2-minutes) bout of moderate intensity exercise using the KE muscles. 
We will also conduct an interview with the caregiver(s) to obtain insight on the exercise 
training and device used for the study at the 6-month visit.  
 
As part of Aim 2, baseline testing will be done over the course of 4 days to allow for 
familiarization to the exercise intervention and the therapeutic exercise device, as well as to 
develop the individualized exercise prescription for the at-home training intervention. 
  
On day 2, a 6-minute cycle test similar to what was used by Jansen et al (2010) will be 
performed to assess muscle endurance on the custom-built exercise device. Heart rate will 
be monitored at rest and throughout cycle exercise using standard non-invasive monitoring 
techniques (6 to 10 lead electrodes or chest strap and wrist watch).  The patient will be asked 
to cycle at his maximal intensity level for 6 minutes and the rating of perceived exertion on 
the OMNI scale and distance covered will be noted. After a 5-10 minute rest period, the 
patient will be asked to cycle again for 5 minutes, at an intensity of between 3 and 6 on the 
OMNI scale and this will correspond to the exercise intensity the patient will undergo at home. 
For the strengthening, the maximal voluntary contraction for KE and KF will be determined 
and then patients will perform ~4 sets of 6 reps of isometric KE, KF at an intensity of 50% of 
MVC.  
 
On day 3, patients will not undergo any testing. On day 4 the patient will undergo another 
exercise familiarization session, where he will do cycle exercise for up to 10 minutes at an 
OMNI intensity of 3 to 6. This will be followed by a strength exercise session. Following this, 
patients and their families will receive detailed instructions relating to the exercise protocol 
and device. We will also provide detailed instructions and video of how to operate the exercise 
device on the laptop computer that will be provided to the patient. A final blood draw will be 
obtained 4 hours after the exercise sessions on day 4 to monitor serum CK.  
 
 
Table 1: Outcome measures  
 
 
 
AIM 1 
 
Side-effect outcome 
Efficacy outcome 
Primary 
BMI (weight and linear height) 
NSAA and 10 m walk/run 
time 
 
 
MRS FF (VL) 
Secondary 
Bone density, Questionnaire on 
steroid side effects (Sidecort), 
Muscle 
strength, 
4-stair 
climb, 6MWT, peak aerobic 

behavior (Child behavior check 
list), quality of life (PedsQL), 
ACTH stimulation test  
capacity, 
pulmonary 
function, blood and urine 
biomarkers 
 
 
MRS T2 
 
Exploratory  
GRO, change in physical 
activity levels 
 
 
AIM 2 
Primary 
 
MRI T2  
 
NSAA and 10 m walk/run 
time, Peak aerobic capacity 
 
 
Muscle CSA 
Secondary  BMI 
Similar to Aim 1, muscle 
endurance 
 
Aim 1 questionnaires  
Caregiver Impression of change, 
Caregiver interview 
MRS FF (VL) 
31P MRS 
 
Exploratory  
GRO, change in physical 
activity levels 
 
Table 2: Events for AIM 1: 
AIM 1 
Visit 1 
Visit 2 
Visit 3 
Visit 4 (for patients 
in exercise group 
only) 
Visit 5 
Consent 
X 
 
 
 
 
Medical history & 
exam  
X 
 
 
 
 
Vital signs, ECG 
X 
X 
X 
X 
X 
ACTH 
stimulation test 
 
 
X 
 
X 
MRI/MRS 
X 
 
X 
X 
X 
Dexa scan 
X 
 
 
 
X 
Muscle strength 
X 
 
X 
X 
X 
Functional 
testing 
X 
X 
X 
X 
X 
Pulmonary 
function testing 
X 
 
X 
X 
X 
Cardiopulmonary 
exercise test  
Xa 
Xa 
X 
X 
X 
Questionaires 
X 
X 
X 
X 
X 
Prednisone 
dispensation 
X 
X 
X 
 
X 
a = Cardiopulmonary exercise testing will be done on one of these 2 visits (baseline OR 3 
months) and used as a familiarization test prior to the pre-exercise training visit (6 months).  
 
Table 3: Events for AIM 2: 
AIM 2 
Visit 1 
Visit 2 
 
Visit 3 
Consent 
X 
 
 

Medical history & exam 
X 
 
 
Vital signs, ECG 
X 
X 
X 
MRI/MRS 
X 
X 
X 
Dexa scan 
X 
 
X 
Muscle strength 
X 
X 
X 
Functional testing 
X 
X 
X 
Pulmonary function testing 
X 
X 
X 
Cardiopulmonary 
exercise 
test  
X 
X 
X 
Exercise practice 
X 
 
 
Questionaires 
X 
X 
X 
 
Study drug prescription and administration: 
The Investigational Drug Service (IDS) at the University of Florida will oversee study drug 
related matters. The lead pharmacist is Irin Reji will be the direct contact, but all 
communication should be sent to IDS@shands.ufl.edu. 
Prednisone tablets (Manufacturer: multiple FDA-approved generic options) will be prescribed 
by the study physician at a dose of 0.75 mg/Kg and dispensed by IDS or Outpatient pharmacy 
at UF Shands Hospital to the patient. Tablets are available as 2.5, 5 or 10 mg strengths. 
Doses for prednisone will be according to patient weight using dosing tables. Throughout the 
study, patients and their parents/caregivers will complete a diary documenting prednisone 
administration and diet compliance. Specifically, parents/patients will be asked to log drug 
administration and diet history that is checked every two weeks. Upon completion of data 
entry, the database will undergo a quality review to ensure acceptable accuracy and 
completeness.  In addition, site calls, which will occur throughout the study will include a 
compliance check on prednisone and diet.   
 
Exercise prescription for boys with DMD:  

Exercise prescription in clinical populations advocates for a combination of endurance and 
strength training for optimal benefits 62. We have taken consideration of previous exercise-
based studies in DMD 3,4,29 in establishing an effective yet safe exercise intervention, which 
will involve a sequential 3-phase combination of aerobic training followed by strengthening 
followed by a mixed combination of both aerobic and strengthening exercise (Figure 4). The 
rationale is based on the notion that aerobic exercise first targets pathophysiological 
mechanisms to induce adaptation within recruited muscle that will be protective to 
subsequent strengthening exercise, and thereafter a combination of the two for maintenance 
of adaptations gained. In the “No Use is Disuse” study4, assisted bicycle training was 
compared to usual care in late ambulatory or wheelchair dependent boys with DMD, 
demonstrating the safety and feasibility of dynamic exercise using parameters of: 5x/week, 
15 min/day for 6 months using the OMNI scale (> 3 and < 6) for patient rating of perceived 
exertion (RPE) to guide exercise intensity 63. As this was the first study to ever use dynamic 
exercise in DMD and the boys were older (mean age 10.8 years), exercise was performed 
with electrical motor support. Our study will use similar exercise parameters but also 
proposes a combination of isometric strengthening with cycle ergometry, which is expected 
to lead to functional gains not detected in the No Use study. The strength training paradigm 
proposed herein is based on our pilot data (shown in Figure 1), indicating that 3 times/week 
of 4 sets/6 repetitions at 50% MVC over 12 weeks was safe and significantly improved muscle 
strength and functional ability. In the current proposal, we will have patients do bilateral KE 
and KF exercise after 8 weeks of initial aerobic cycling, and then in combination on alternate 
days to maximize potential gains. 
 
The schedule for exercise (days of the week and time of session) will be discussed at the 
baseline visit and the study investigators will provide a tentative monthly training calendar to 
the patient/parent based on their feedback and availability. The study investigator will confirm 
the weekly schedule every Friday prior to the scheduled training week. Additionally, at the 
end of every exercise session, the study team will confirm the date and time of the subsequent 
exercise session. The patient and parent will be provided with the contact information of the 
study team members responsible for leading the training session.   
 
For each exercise session, a parent or guardian must be in the home during the time of the 
exercise and in close enough proximity to hear the study investigator over the video 
call/computer. They may be needed to help set up the patient on the exercise device and/or 
laptop computer. This will be discussed at the baseline visit and factored into the training 
Figure 5: Exercise prescription parameters for the 6-month training program.  

schedule to ensure adult supervision is available at the scheduled times. The study team will 
also interact with the parent/guardian to discuss the exercise experience.  
 
To allow for both exercise modalities to be performed in the home-setting, the Department of 
Engineering (Co-investigator Dr. Dixon) will modify and repurpose components from the 
exercise device used in Dr. Lott’s previous IRB approved study (IRB201400874). As shown 
in Figure 5, this will consist of a stable chair for postural support, a motorized-assisted cycling 
ergometer, a cuff and strain gauge cable system, and a live computer monitoring system. 
Similar to the previous study by Jansen and colleagues (2013), the ergometer will allow 
cycling to be assisted with motor support. However, because our patient cohort will be less 
clinically affected compared to the Jansen study, the ergometer will also have options of no 
load or low resistance to pedaling once muscles become accustomed to the exercise. 
Customizable, set thresholds of workload and 3 levels of safety checks are built into the 
device. As standard in commercially available devices, at all times the participant will have 
access to an emergency stop button that will immediately halt the motor should it become at 
all uncomfortable. Additionally, hardware and software limits will be imposed that ensure the 
minimal risks of the session are mitigated. The device will be fairly compact for in-home use 
and can be easily configured for exercise modality. For strengthening exercise, the lower leg 
will be secured to a padded bar and the patient will perform kicking of pulling type exercise 
isometrically with the hip at ~90˚ of flexion and the knee positioned at 60˚ of flexion recruiting 
the KE and KF muscles as in our previous study. For the cycling, the padded bar will swivel 
away 
and 
the 
ergometer will slide 
towards the patient. 
The 
intensity 
of 
exercise 
will 
be 
guided by the level of 
patient 
exertion 
using 
the 
OMNI 
scale of rating of 
perceived 
exertion 
(RPE). Heart rate will 
be monitored and 
recorded on a wrist 
watch provided to the 
patient We will use 
the real-time video 
feature 
(Skype 
or 
Zoom) on the laptop 
to monitor all training 
sessions 
and 
provide 
encouragement and 
verbal 
feedback 
regarding any concerns. The computer will be equipped with custom-written software that will 
provide visual and auditory guidance and instructions for the subject and his parent(s) to 
Figure 6: Exercise modalities used for leg cycle and isometric strength 
training in boys with DMD.  

complete the exercise sessions at home. Patients and their families will be familiarized to the 
exercise device and instruction during the baseline visit of Aim 2. We will use and repurpose 
7 exercise chairs from our previous study for this current project. 
 
Data Analysis 
After the aforementioned procedures are completed, the collected data will be compiled for 
analysis. The participants will provide data on primary and secondary outcome measures 
specifically for research purposes. Only the PI and research staff involved with subject 
recruitment and data collection will have access to the individual, identifiable private data 
collected during the study. The participants of this study are children and as such, written 
informed consent will be obtained from the parent or legal guardian and assent will be 
obtained from the child. Subject data will be coded to ensure confidentiality and no personal 
health information will be released. All electronic data will be secured on password protected, 
encrypted computers and hard copies will be kept in locked filing cabinets. In case of any 
adverse events from the study protocol that result in injury, medical attention will be sought. 
Both adverse events and/or breeches of confidentiality will be reported immediately to the PI, 
who will immediately report the event to the IRB. If an adverse event occurs, the investigators 
will reassess the risk/benefit ratio of the study and submit any modifications deemed 
necessary to the IRB for approval. Dr. Taivassalo will inform the funding agency of any actions 
taken by the IRB in response to any adverse event reports. 
Access to linked identifiers is limited to research personnel intimately involved with the human 
participants. All data and records acquired from participants is for research purposes only 
and will be kept confidential and maintained in a secure database identifiable only by 
participant code. The results of the study may be published for scientific purposes; however, 
participants’ identities will not be revealed and data will not be traceable to any individuals in 
any resultant publications. The information gathered during this study will be kept confidential 
to the extent permitted by law. 
Given the similarities in testing protocols between this study and PI Dr. Taivassalo’s study 
(IRB202301491), data collected from participants will be shared, integrated, and analyzed as 
part of the Vaso Rex study, contingent upon participants enrolling and providing 
consent/assent. This integration will apply only to participants who have consented to the 
Vaso Rex study and have completed identical tests related to this project within three months 
prior to their enrollment in IRB202301491. This approach aims to streamline testing protocols, 
enabling the research team to reduce the schedule for the Vaso Rex study while still 
leveraging available data from this study within the specified timeframe. Ultimately, this will 
help minimize testing and alleviate the burden on participants. 
 
 

7. Possible Discomforts and Risks: 
 
As the safety of the boys with DMD is paramount, we are in the unique position to have the 
personnel, experience, and the necessary equipment to monitor the proposed interventions 
and conduct all aspects of the proposed protocol. Members of this proposed research team 
have working on projects involving boys with DMD over the past 10 years, with more than 
180 boys enrolled in our ongoing studies. With regards to safely implementing the 
methodology proposed in this study, we have performed MRI/MRS, strength, and functional 
assessments hundreds of times on boys with DMD without incident and with high confidence 
and subject compliance. We have also implemented the proposed isometric strength training 
portion of the exercise intervention in a previous study (Dr. Lott, IRB201400874) in 8 boys 
with DMD with no adverse events. As such, the research team has extensive experience to 
conduct the assessments following special precautions to minimize risks or hazards. The 
potential risks for each assessment and intervention proposed in this study are described, 
along with the measures we undertake to protect the patient against the risk. 
 
Study parameters will be assessed at baseline, 3, 6 and 12 months and include physical 
examinations, vital signs, clinical laboratory tests (blood and urine collection), ECGs, 
concomitant medication monitoring and treatment-emergent adverse events (AEs) within in 
the Clinical Research Unit at every study visit. If clinically significant deterioration is noted in 
vital signs or physical examination, the change will be documented as a treatment-emergent 
adverse event (AEs). Clinical significance is defined as any finding that has medical relevance 
that results in an alteration in medical care. The Study physician will continue to monitor the 
patient until the parameter returns to its Baseline status. The Study Physician will review the 
results of the ECG report and designate the findings as normal or abnormal; if abnormal, the 
Study Physician will indicate whether the ECG tracing is not clinically significant or clinically 
significant. Clinically significant deterioration should be reported as an AE. Any indication of 
heart abnormality will be followed up by a cardiologist.  
Adverse events will be recorded from when the Informed Consent Form (ICF) has been 
signed until the End-of Study visit. AEs before study drug first intake will be considered as 
“unrelated”. The study investigators are responsible for reporting all serious and unexpected 
adverse events (regardless of relatedness) to the UF-IRB within 5 working days after learning 
of the event. Any AE or other unwanted response which occurs in the course of the study will 
be monitored and followed up until: it has resolved or stabilized according to the judgment of 
the study physician; there is a return to normal or baseline values, it has been shown to be 
unrelated to the study intervention. 
 
MRI/MRS: Current guidelines from the US Food and Drug Administration (FDA) indicate that 
there are no known risks for human exposure to magnetic fields of 3.0 Tesla, unless metallic 
implants or cardiac pacemakers are introduced into the magnetic field or fringe field. The 
participants and those individuals assisting during the MRI will be screened using 
standardized procedures for metal or magnetic implants and other incompatible devices prior 
to entering the MRI scanner area. Anyone with a contraindication to MRI will not be permitted 
to enter the MRI scanner area. The MRI produces a loud noise, which has the potential to 
produce hearing loss in a very small number of patients when hearing protection is not used. 

All persons present in the MRI during the scan will be provided with ear protection (ear plugs 
and/or insulated headphones) and instructed to wear these for the entire duration of the scan, 
which nearly eliminates the risk of hearing loss.  
 
Physical function and muscle strength assessment: Safety precautions will be taken during 
the muscle strength and functional assessments. The 10-meter walk/run and six-minute walk 
tests will be administered in an unobstructed hallway. There is a minimal risk of falls with the 
stair climbing test. To minimize this risk, the child will be asked to wear suitable footwear and 
will be supervised at all times by a physical therapist. During the six minute walk test, safety 
precautions will be taken during the test by applying standardized stopping criteria. If the 
participant reports pain, tightness or pressure in the chest, significant shortness of breath, 
feeling faint, lightheaded or dizzy, or significant other medical problems the test will be 
stopped. Blood pressure will be assessed before and after this test. This measurement is 
non-invasive, however, cuff inflations may pose momentary discomfort. Research nurses are 
immediately available in the CRC if needed.  The exercise assessment (both on cycle 
ergometer and determination of peak muscle strength may cause muscle fatigue and/or 
soreness in some children. If the participant becomes tired during testing, he/she may request 
a break to rest. To decrease the risk of these undesirable effects, participants will be attended 
to at all times while participating at UF. 
 
Blood draws: There is a slight risk of minimal bleeding or infection (< 1% chance) during the 
blood draw. To minimize the risk, a qualified nurse in the CRC will perform the blood draw at 
UF, and only nurses/physicians will perform the blood draw in the subjects’ own community. 
The blood draw is done using sterile techniques to reduce the risk of infection. For the ACTH 
stimulation test, other than venipuncture, there are no additional risks.  
 
Study drug interventions:  
Upon initiation of prednisone, the study physician and/or PI will be in contact with the patient 
and family by phone or videoconference once every 2 weeks for the first 3 months and 
monthly thereafter to ask the patient about any new signs or symptoms, review medication 
and diet history and inquire about any specific concerns they may have. If the patient has 
evidence of developing any side effects, we may require the patient to see his primary care 
physician at home for follow up. If these changes are persistent and/or the patient has non-
manageable side-effects, the dose will be adjusted appropriately by the study physician. Also, 
during the entire course of the study, the patients and their parents will be encouraged to 
contact the investigator at any time to report any safety concerns that may have emerged 
between the scheduled visits or phone communications.  
 
The study visits are 3 months apart in the first half of the intervention. This time frame was 
based on standard clinical practice whereby steroid naïve patients are typically seen by their 
physician 3 months after initiating of glucocorticoids. For Aim 2, study visits are 6 months 
apart. Safety parameters for both study Aims include physical examinations, growth 
parameters, vital signs, clinical laboratory tests (including chemistry, hematology, urinalysis, 
and tests for adrenal function), ECGs, and concomitant medication monitoring and AEs.   
 
Prednisone: 

There are several well-established risks associated with chronic daily use of glucocorticoids. 
Despite this, glucocorticoid treatment is standard of care for DMD and prednisone has been 
used to treat boys with DMD off-label for the past 30 years. Prednisone has shown to improve 
or stabilize muscle strength and delay the loss of ambulation compared to boys with DMD 
who remain steroid-naïve. The evidence-based dose recommendation for prednisone is 
0.75mg/kg/day 13. Because of the numerous risks/side effects associated with this daily dose, 
the search for an optimum dosing regimen exists. In patients with DMD, the most commonly 
reported risks are weight gain (mean of 3.2 kg), central obesity, cushingoid features, 
behavioral changes, growth delay and fractures 12,64. In boys with DMD treated on an 
alternate high-dose weekend regimen (weekend dosing using 5 mg/kg/2 days on weekend 
for 6 to 12 months) compared to untreated boys with DMD, linear growth was maintained and 
obesity rates did not differ from untreated patients 17 Furthermore, Cushinoid features such 
as hirsuitism, acne, and hypertension did not occur in the weekend dose patients. In a 
subsequent study comparing the weekend dosing regimen described above to daily dosing 
of prednisone over 12 months 16, increases in body mass index were detected with both 
regimens (although tended to be more with the daily dosing). The weekend dosing was 
associated with greater linear growth than daily dosing. Although femoral bone density was 
note assessed in this study, lumbar spine Z score showed small changes with both dosing 
regimens. The aim of the current study is to demonstrate that low dose (0.75 mg/kg) twice 
weekly use will lessen the side effects all together. 
 
We submitted an Investigational New Drug application to the Food and Drug Administration 
for the use of prednisone in boys with DMD for this specific study and received an exemption 
(IND 144718) in June 2019. The current ‘standard of care’ prednisone regimen commonly 
prescribed to patients with DMD poses a greater risk than the low-dose twice weekly regimen 
proposed in this study.  Therefore, given the dosing regimen, we do not expect adverse side 
effects but will take precautions to safely monitor each patient. At the same time, we will also 
take precautions to monitor any potential inadequacy of the low-dose prednisone to delay 
onset of muscle pathology. Based on the preclinical data presented by Quattrocelli et al 
(2017) in the mdx mouse showing that weekly prednisone dosing enhanced sarcolemmal 
repair and reduced fibrosis and immune cell infiltration to a comparable extent as daily dosing, 
we expect the low-dose twice weekly prednisone administration to delay the onset of disease 
in boys with DMD similar to the established daily regimen. This, along with the notion that the 
twice weekly dosing will lessen the side effects associated with chronic daily dosing while 
also inducing ergogenic remodeling is the main premise being studied in Aim 1. 
 
To monitor the risk of steroid inadequacy, muscle pathology (fat accumulation) and strength 
will be assessed at the 3 and 6-month visits. Based on data from our natural history cohort 
within ImagingDMD who were corticosteroid naïve, we can expect a 5% increase in MRS-
measured vastus lateralis fat fraction in a 6-month time frame. Therefore, we will take a 5% 
or greater increase in muscle fat fraction in a boy with DMD on low-dose twice weekly 
prednisone dosing after 6 months as an indication that the glucocorticoid is delaying benefit. 
In addition, if we detect a decrement in strength measured in the knee extensor or knee flexor 
muscles at this time point (along with an increase in MRS fat fraction), we will assume the 
dosing paradigm is inadequate and will adjust the dose. The current recommended weekend 
dose based on work done by Ann Connolly’s group is 5 mg/Kg on each of two weekend days. 

As this dose is often associated with irritability and sleep disturbances that are time stamped 
to the days following this high-dose administration, we will use a dose of 2 mg/Kg (which is 
in between our current dose of 0.75 mg/Kg and 5 mg/Kg) twice weekly for the remaining 6 
months of the study. Any adjustments to the dose and potential findings of drug ‘inadequacy’ 
will be reviewed by our local Safety Monitoring Committee.  
 
Exercise intervention:  
Based on the previous studies assessing exercise in DMD demonstrating safety 3,4 along with 
our pilot data on strength training in boys with DMD (see above), we do not expect our 
proposed exercise intervention to pose any risks, induce any muscle damage or cause 
significant pain. Long-standing clinical concerns of exercise in DMD are largely based on 
preclinical data using eccentric muscle actions, which are known to be damaging to muscle. 
We have designed the proposed exercise intervention to minimize any risks or potential for 
muscle damage in boys with DMD. The proposed exercise intervention involves a 
combination of aerobic cycling and isometric strength exercise modalities, performed at a low 
to moderate intensity level. As such, there is no potential for the boys to perform an eccentric 
muscle action. Dynamic forward pedaling will be performed during the cycling with assistance 
from the motor or low intensity resistance. As part of daily life, boys with DMD in this age 
group (5-8 years of age) typically engage in play and physical activity that involve repeated 
eccentric contractions. Therefore, the risks to the boys with DMD from the muscle 
contractions and exercise protocols for this study are less than the risks posed from eccentric 
contractions routinely encountered in their daily lives.  
 
To ensure safety of the patient during exercise, we will take certain precautions. First, 
throughout the exercise intervention, our study team will supervise every exercise session 
remotely over the real-time video on the laptop computer (provided by us to the patient) and 
adjust the exercise guidelines accordingly. During the exercise sessions, procedures to 
minimize discomfort of exercise include a warm-up and cool-down. The participants will also 
be introduced to the exercise in a structured way, such that they begin with lighter intensity 
and gradually increase over the course of the intervention. Prior to starting each exercise 
session, the patient will be asked to rate any pain they may be experiencing at the time of 
assessment using a Wong-Baker FACES Pain Rating Scale with faces and corresponding 
numbers ranging from 0 (No Hurt) to 10 (Hurts Worst) 65. Subjects will be asked to select one 
of the faces with its corresponding numerical rating and pain description. An increase in pain 
>4 will be considered the threshold for muscle damage and patient will not be allowed to 
exercise. We will also ask the patient if he has noticed dark urine. This will be noted and 
discussed with Study Physician. To ensure the patient exercises at low to moderate intensity, 
we will monitor the patients rating of perceived exertion using the OMNI scale, an assessment 
of perceived exertion using pictorial and verbal cues suited for healthy and clinically affected 
children 63. Patients will be instructed to exercise at an intensity corresponding to > 3 and < 
6 on the OMNI scale. If the patient rating is > 6 on the OMNI scale, the bicycle resistance 
may be lowered accordingly and noted by the investigator, or a rest break will be taken. Heart 
rate at rest and during exercise will also be monitored to gain information on perceived and 
physical exertion of exercise. Some muscle soreness may be expected initially when the 
patient starts the exercise program (given that the muscle has been unaccustomed to 
exercise) and initially if workload is increased. Patients will be asked to rate any pain they 

may experience before, during or after exercise using a Wong-Baker FACES Pain Rating 
Scale with faces and corresponding numbers ranging from 0 (No Hurt) to 10 (Hurts Worst). 
Patients will be asked to select one of the faces with its corresponding numerical rating and 
pain description. An increase in pain >4 will be considered the threshold for muscle pain and 
exercise will be stopped. The research investigator will also ask the patient and parent if there 
are any other concerns or experiences of uncomfortable feelings during or after the exercise. 
Based on the response, the exercise intensity may be lowered accordingly and noted by the 
investigator. Some muscle soreness may be expected initially when the patient starts the 
exercise program (given that the muscle has been unaccustomed to exercise) and initially if 
workload is increased. Rest breaks will be provided as needed.  
 
If the patient does experience muscle soreness and/or joint stiffness, these symptoms should 
not persist more than a few days and are comparable to typical post-workout soreness. The 
forces used during exercise for this study are well within safety limits, and the exercise device 
has numerous hardware and software constraints that limit the exercise to safe levels. For 
example, both hardware and software constraints are built in to limit the power output of the 
motor to avoid lower-limb rotations at excessive speeds. This speed is constantly measured 
so that excessive speeds will cause the test to automatically stop. The electric motor torque 
will be continuously monitored to prevent torque that may lead to muscle and/or tissue tear.  
There are also other risks that relate to soft tissue injury, falls and fractures, post-exercise 
hypotension, and cardiovascular events. Although the patient will be seated throughout thee 
exercise, there is a risk that a participant may trip, stumble, or fall getting on or off the device. 
The patient may experience shortness of breath, dizziness, rapid or irregular heartbeat, chest 
pain or discomfort, heartburn, light headedness, or feeling about to faint. If the patient 
becomes tired during exercise, he may request a break to rest. During the exercises, the 
patient will have immediate access to a stop button that can be pressed at any time for any 
reason that will halt the exercise. We will require a parent or guardian be present at home 
during every exercise session. The risks to patients participating in this study are lower than 
risks if the patient were to exercise alone and without our instruction since we are monitoring 
the exercise and providing safety precautions. If any of the above-mentioned conditions occur 
during exercise or persist after rest, the participant’s primary physician may be called and 
referred for evaluation. If the participant complains of angina at rest, loss of consciousness 
occurs, or cardiac arrest, the investigator will instruct the parent/guardian to contact 
emergency medical services immediately.  
 
For exercise performed within the CTRB at UF, research staff will be present and available 
throughout and emergency treatment will be available if it becomes necessary. In the event 
of an adverse medical event, standard facility emergency procedures will be followed and 
proper personnel notified. In the event that a cardiac event is suspected the exercise session 
will be stopped immediately, and local emergency medical technicians (EMTs) will be called 
and the participant will be taken to an emergency medical center, if recommended by the 
EMTs. 
To confirm the safety of the exercise intervention, blood for analysis of serum creatine kinase 
(CK) levels will be obtained at baseline and after 6 months on exercise. Serum CK is 

commonly elevated and variable in DMD independent of any intervention. A previous study 
using whole-body vibration used an increase in CK level by >40,000 U/L to indicate muscle 
damage relating to their intervention 66. We will use a more conservative threshold (>20,000 
U/L), and we will also assess MRI-T2 within the exercised quadriceps muscles at the study 
visits, where an elevation (>2SD) in the proportion of pixels within a muscle is considered the 
threshold for muscle damage. If at the 6-month assessment the patient has elevated serum 
CK (>20,000 UL from baseline) and elevated MRI T2, the exercise intervention will be reported 
as a serious adverse event.  
Adverse events: definitions and action plan 
An adverse event (AD) is any untoward medical occurrence in a clinical investigation patient, 
which does not necessarily have a causal relationship with the investigational product (active 
or placebo drug, biologic, or device).  An AE can, therefore, be any unfavorable and 
unintended symptom, sign, disease or condition, or test abnormality, whether considered 
related to the investigational product. 
Adverse events include: 
• Symptoms described by the patient or signs observed by the Investigator or 
medical staff 
• Test abnormalities (laboratory tests, ECG, etc.) that result in an alteration in 
medical care (diagnostic or therapeutic) 
Adverse events do not include: 
• Changes that are characteristic of disease progression in DMD 
Abnormalities present at Baseline are considered AEs only if they reoccur after resolution or 
they worsen during the study. 
Seriousness of an AE serves as a guide for defining regulatory reporting obligations.  An SAE 
is any AE that results in any of the following: death, life-threatening event, required or 
prolonged inpatient hospitalization, 30ersistent or significant disability/incapacity, important 
medical events 
     Evaluation of Adverse Events/Serious Adverse Events: Assessment of the association 
between the AE and study drug exposure is important for regulatory reporting.  For each 
AE/SAE, the Investigator and Study Physician will determine whether there is a reasonable 
possibility that the AE may have been caused by the study drug according to the following 
categories: 
• Not Related:  The AE is clearly not related to study drug. 
• Unlikely Related:  There is no evidence of a causal relationship between study drug 
and the AE; although such relationship cannot be ruled out. 
• Possibly Related:  There is some evidence supporting the possibility of a causal 
relationship between study drug and the AE. 
• Related:  There is strong evidence that there is a causal relationship between study 
drug and the AE. 

• A relationship to the investigational product must be given for each AE/SAE recorded, 
even if there is only limited information at the time.  The Investigator may change 
his/her opinion of causality in light of follow-up information, amending the AE/SAE 
report accordingly. 
Severity of an AE (Note that severity is not the same as “seriousness,”: The Investigator will 
grade the severity of all AEs/SAEs as mild, moderate, or severe, based on the following 
definitions: 
• Mild:  An AE that is usually transient and may require only minimal treatment or 
therapeutic intervention.  The event does not generally interfere with usual activities of 
daily living. 
• Moderate:  An AE that is usually alleviated with specific therapeutic intervention.  The 
event interferes with usual activities of daily living, causing discomfort, but poses no 
significant or permanent risk of harm to the research participant. 
• Severe:  An AE that interrupts usual activities of daily living, or significantly affects 
clinical status, or may require intensive therapeutic intervention. 
 
The Investigator will provide information regarding the patient outcome of each AE. Possible 
results of an AE outcome are defined as follows: 
• Fatal:  The termination of life as a result of an AE. 
• Not recovered/not resolved:  The patient has not recuperated or the AE has not 
improved. 
• Recovering/resolving:  The patient is recuperating or the AE is improving. 
• Recovered/resolved:  The patient has recuperated or the AE has resolved. 
• Recovered with sequelae/resolved with sequelae:  The AE has resolved, but the 
patient has been left with symptoms or pathology. 
• Unknown:  Not known, not observed, not recorded, or refused. 
 
The Investigator will be required to provide the action taken regarding prednisone in response 
to the AE according to the following categories: 
• Dose not changed:  No change in administration of the investigational product. 
• Dose reduced:  Reduction in the frequency, strength or amount of investigational 
product administered. 
• Drug interrupted:  Temporary interruption (termination) in administration of the 
investigational product. 
• Drug withdrawn:  Administration of the investigational product terminated (no further 
dosing). 

• Not applicable:  Determination of a value is not relevant in the current context. 
• Unknown:  Not known, not observed, not recorded, or refused. 
 
All AEs/SAEs experienced by the patient will be recorded on a Study adverse event log form. 
The following information will be recorded:  a concise description of the event; date and time 
of event onset and resolution; determination of seriousness, severity, corrective treatment, 
outcome, and relationship to investigational product; and, action taken regarding the 
investigational product.  Abnormal vital signs, laboratory results, or other abnormal safety 
assessments will be recorded as an AE if they meet the definition of an AE.  When possible, 
a diagnosis should be recorded as an AE rather than symptoms or isolated laboratory 
abnormalities related to that diagnosis.  A medical or surgical procedure is not an AE; rather 
the condition leading to the procedure should be recorded as the AE.  If the condition is not 
known, the procedure must be reported as an AE instead.  Similarly, death is not an AE, but 
rather is the outcome of the AE(s) that resulted in death.  If the AE(s) leading to death are not 
known, then death must be reported as an AE. 
The study investigators are responsible for reporting all serious and unexpected adverse 
events (regardless of relatedness) to the UF-IRB within 5 working days after learning of the 
event. Any AE or other unwanted response which occurs in the course of the study will be 
monitored and followed up until: it has resolved or stabilized according to the judgment of the 
study physician; there is a return to normal or baseline values, it has been shown to be 
unrelated to the study intervention. 
All Serious Adverse Events that are a suspected adverse reaction, serious and unexpected 
must be submitted by the investigator to the FDA using form 3500A (MedWatch). Note: 
events must meet all criteria to be submitted to the FDA. 
All AEs/SAEs documented at a previous visit/contact that are designated as not 
recovered/not resolved or recovering/resolving will be reviewed by the Investigator at 
subsequent visits/contacts. 
All AEs will be followed until the parameter returns to its Baseline status, resolution of AE, 
completion of the patient’s participation, agreement is reached between the Investigator and 
Sponsor that the event no longer requires follow-up, or study termination, whichever occurs 
first.  SAEs will be followed until resolution, the condition stabilizes, or the Investigator notes 
that follow-up is no longer necessary.  Rules for AE/SAE follow-up apply to all patients, 
including those withdrawn prematurely to the extent allowed by the patient’s consent.  The 
Investigator will ensure that follow-up includes further investigations consistent with 
appropriate medical management and patient consent to elucidate the nature and/or causality 
of the AE/SAE. 
Temporary or Persistent Dose Reductions: Temporary or persistent dose reductions may be 
made for safety or tolerability reasons in the judgment of the Study Physician and Principal 
Investigator.  If at any time after initiation of treatment the patient experiences a clinically 
significant AE indicative of drug intolerance, the patient will undergo clinical assessment as 
soon as reasonably possible.   

Monitoring of Abnormal Laboratory Values: During the study, liver function tests will be 
monitored, including ALT, AST, GGT, alkaline phosphatase, glutamate dehydrogenase, and 
bilirubin.  If increases in ALT or AST are noted, the relationship of these to CK will be 
determined, as well as to GGT and measures of liver synthetic function.  Trends in these 
parameters will be monitored.  The Investigator may stop dosing in individual patients based 
on tolerability or AEs. 
 
8. Possible Benefits: 
 
Boys with DMD who participate in this study may experience direct, tangible benefits, 
although any benefit cannot be guaranteed. Anticipated benefits of taking the low dose twice 
weekly prednisone alone or in combination with exercise training include improved muscle 
strength and slowing of disease progression whereby patients may experience a stabilization 
or improvement in performance of daily activities including walking and stair climbing. The 
patient may experience improved endurance and resistance to fatigue. Together these 
improvements may be reflected as a benefit in their daily mood and/or quality of life 
(measured using the Pediatric Quality of life inventory (PedsQL). Boys who undergo low dose 
twice weekly prednisone alone(without exercise training) may also benefit from the anti-
inflammatory and ergogenic effects of prednisone without experiencing the side effects 
(weight gain, central adiposity, behavioral issues) associated with daily use. 
Other potential benefits for boys with DMD and their families include the educational 
experience during their interaction with investigators who are highly interested in muscular 
dystrophy and treatment. The boys and families may gain information on their condition from 
the functional assessments and the MR studies, as well as information on disease 
progression within their lower extremity muscles and the intervention effect. Individual data 
obtained in the study will be summarized for each participant and shared with him and his 
family. All study participants will be encouraged to communicate the results from the study to 
their primary care providers. The risks of participating in this study are low especially with the 
careful monitoring that will be done throughout the intervention and study visits. Therefore, 
the benefits both to the study participants and to the wider muscular dystrophy community 
outweigh the potential risks. There are no alternative approaches to the procedures outlined 
in the study protocol; the subject may withdraw (or his parents may withdraw him) at any time 
without any consequences if he no longer wishes to participate, as will be indicated in the 
informed consent form. 
 
This study has significant promise to benefit scientific knowledge and have both short-term 
and long-term impact in the field of DMD. The knowledge gained on low dose twice weekly 
prednisone dosing has far reaching implications particularly given that more than 50% of boys 
treated with daily glucocorticoids develop significant enough side effects that medication is 
decreased or discontinued.  The development and validation of exercise paradigm guidelines 
that may be beneficial dystrophic muscle remains an ever-present need in the field. This study 
strives to develop exercise prescription parameters that will have important implications for 
future comprehensive clinical care and management of patients with DMD. 
  
Short-term impact: This study provides fundamental new knowledge relating to an alternate 

GC regimen and impact of exercise in DMD. It will be the first to reveal whether a low-dose 
twice weekly regimen represents a more optimal frequency and dosage to maximize benefit 
and minimize risk than the current standard, and is expected to have less impact on BMI 
compared to standard daily dosing, along with few behavioral disturbances. The exercise 
training is anticipated to provide additive benefit to the GC regimen by improving muscle 
strength and physical function, and potentially delaying the progression of disease that 
characterizes this population of patients. Collectively, these novel interventions are expected 
to markedly improve quality of life and physical function in our cohort of young boys with 
DMD. 
 
Long-term impact: By providing information on an alternate GC strategy in DMD, this study 
has real potential to impact clinical practice and patient care for DMD as well as other 
muscular dystrophies for whom GC are currently part of patient management. New 
knowledge on the interplay between GC regimen and exercise is expected to form a 
comprehensive clinical care approach, which needs to be confirmed in older patients. 
Through the use of non-invasive imaging, our study will provide new understanding regarding 
the impact of exercise on muscle pathophysiology in dystrophic muscle, thereby helping to 
shift the current paradigm in the field to promote rather than avoid exercise for patients with 
DMD. This proposed research is expected to add to the growing recognition that appropriate 
exercise can be amongst the most potent therapies for a variety of diseases and may work 
synergistically with pharmacological agents, extending the health span of a boy with DMD. 
Particularly as molecular and pharmacological treatments advance to allow patients to live a 
more active lifestyle, the role of exercise as an adjuvant therapy needs to be considered 67, 
and our findings provide a foundation for these future trial considerations. Finally, the exercise 
intervention proposed herein may be relevant to patients with other muscular dystrophies.  
 
9. Data Safety Plan: 
A local safety committee will be formed consisting of the PI, Study Physician Dr. Sladky, Co-
investigator Dr. Sweeney, and a paediatric neurologist. The committee will review the 
progress of each patient enrolled in the study, review source documents including 
concomitant medications, compliance to intervention and adverse events. The adverse 
events include mood changes, hyperglycemia, hypertension, muscle soreness, weight gain 
and chest pain, all of which have a low likelihood of occurring. The committee will have a 
predetermined plan to deal with each event if it occurs. The committee will meet at least once 
every 3 months. In the case of adverse events, an ad hoc meeting may be called by the PI 
and/or Study physician. The Principal Investigator and study physician, with the assistance 
of study staff, will treat any adverse events to the extent that they are able and is necessary. 
If adverse events are serious enough and warrant emergency medical services, 911 will be 
called and the Principal Investigator and the study staff will ensure maximal comfort and 
safety of the subject until emergency medical services arrive. If adverse events occur at the 
patients' home, the PI and study staff will request the parent of the patient to seek medical 
attention. 
A safety endpoint has been established for this study. With respect to the study drug 
prednisone, the side effects of chronic daily steroids have been acknowledged and although 
not expected given the low dose twice weekly use of prednisone, they will be monitored in 

this study. Any adverse events associated with steroid side effects will be discussed by the 
local safety committee and a decision will be made to discontinue the patient from the study 
(described above under “Adverse events”. 
There may also be a risk of inadequacy of the low-dose twice weekly prednisone 
administration, as chronic daily use is known to result in an apparent improvement in muscle 
strength and ambulation in most boys for 6-12 months, followed by stabilization for about 2 
years and long-term slower deterioration than untreated patients. Based on the preclinical 
data presented by Quattrocelli et al (2017) showing that weekly prednisone dosing enhanced 
sarcolemmal repair and reduced fibrosis and immune cell infiltration to a comparable extent 
as daily dosing, we expect the low-dose twice weekly prednisone administration to delay the 
onset of disease in boys with DMD similar to the established daily regimen. This, along with 
the notion that the twice weekly dosing will lessen the side effects associated with chronic 
daily dosing while also inducing ergogenic remodeling is the main premise being studied. 
However, we will monitor the risk of drug inadequacy at the 3 and 6-month visits when 
patients return to the University of Florida by assessing muscle pathology (using magnetic 
resonance spectroscopy, MRS, to measure fat accumulation within the vastus lateralis 
muscle) and muscle strength (using Biodex dynamometry). Based on data from our natural 
history cohort within ImagingDMD who were corticosteroid naïve, we can expect a 5% 
increase in MRS-measured vastus lateralis fat fraction in a 6-month time frame. Therefore, 
we will take a 5% or greater increase in muscle fat fraction in a boy with DMD on low-dose 
twice weekly prednisone dosing after 6 months as an indication that the glucocorticoid is 
delaying benefit. In addition, if we detect a decrement in strength measured in the knee 
extensor or knee flexor muscles at this time point (along with an increase in MRS fat fraction), 
we will assume the dosing paradigm is inadequate and will adjust the dose and revise the 
IRB. The current recommended weekend dose based on work done by Ann Connolly’s group 
is 5 mg/Kg on each of two weekend days. As this dose is often associated with irritability and 
sleep disturbances that are time stamped to the days following this high-dose administration, 
we would plan to use a dose of 2 mg/Kg (which is in between our current dose of 0.75 mg/Kg 
and 5 mg/Kg) for the remaining 6 months of the study. 
 
An efficacy endpoint has also been established. Similar to above description, if the low-dose  
(0.75 mg/kg) twice weekly prednisone appears to be inadequate (based on increases in 
muscle fat fraction and decreases in strength over 6 months) in terms of inducing the benefits 
normally associated with daily prednisone, we will seek permission (amendment to IRB) to 
increase the dose to 2 mg/Kg (which is in between our current dose of 0.75 mg/Kg and the 
high dose 5 mg/Kg previously used on weekends and shown to induce irritability) for the 
remaining 6 months of the study. 
 
The safety and monitoring plan also includes close supervision of every patient throughout 
the study.  At the study visits, vital signs will be monitored. If the participant displays signs of 
altered vital signs, procedures will be adjusted or halted and the participant will be closely 
monitored. The study physician will be contacted if the participant's condition persists. After 
initiation of the study drug (Prednisone) when the patient is back home, the study physician 
will be in contact with the patient and family by phone once every 2 weeks for the first 3 

months and as needed thereafter. The patients and their parents will be encouraged to 
contact the investigator at any time to report any safety concerns that may have emerged 
between the scheduled visits or phone communications. Regular monitoring includes asking 
about changes in medications, pain ratings, overall function. 
 
To ensure safety of exercise, we will take certain precautions. First, throughout the exercise 
intervention, our study team will supervise every exercise session remotely over the real-time 
video on the laptop computer and adjust the exercise guidelines accordingly. We will also be 
monitoring the patients rating of perceived exertion using the OMNI scale (assessment of 
perceived exertion using pictorial and verbal cues suited for healthy and clinically affected 
children, where patients will be encouraged to exercise at an intensity corresponding to > 3 
and < 6 on the OMNI scale.  Patients will be asked to rate any pain they are experiencing 
during or after exercise using a Wong-Baker FACES Pain Rating Scale with faces and 
corresponding numbers ranging from 0 (No Hurt) to 10 (Hurts Worst). They will select one of 
the faces with its corresponding numerical rating and pain description during the real-time 
Skype monitoring for the research investigator to note. The research investigator will also ask 
the patient and parent if there are any other concerns or experiences of excessive pain/ 
uncomfortable feelings during or after the exercise. If the patient rating is > 4 on the Pain 
Rating scale, the exercise intensity may be lowered accordingly and noted by the investigator. 
Some muscle soreness may be expected initially when the patient starts the exercise 
program (given that the muscle has been unaccustomed to exercise) and initially if workload 
is increased. Rest breaks will be provided as needed.  
 
The accuracy and quality of data will be checked. All data collection will be supervised by the 
Principal Investigator or a physical therapist or exercise physiologist who is a study staff 
member. Accuracy and quality of the physiological and functional data will be checked for 
accuracy by the PI. Demographic and functional data will be entered via the secure, web-
based application Research Electronic Data Capture (REDCap). Heart rate will be obtained 
from a pulse oximeter. Heart rate may also be assessed manually by study staff with 
experience assessing heart rate using this method to double-check for accuracy. Blood 
pressure will be assessed manually using a blood pressure cuff by study staff with experience 
taking blood pressures. Blood pressure may also be monitored with an automatic blood 
pressure cuff in order to double-check for accuracy.  For the MRI/MRS data, Dr. Mueller will 
serve as the Informatics Manager for the MRI related projects in the ImagingDMD laboratory. 
He will be responsible for MR data transfer, data management and data security as per 
protocol standards set forth by ImagingDMD. He will maintain the patient data base, check 
data for integrity and establish quality control procedures, produce regular reports 
summarizing the status of data acquisition and subject enrollment, and convert the data base 
for entry into the statistical analysis package. He is also responsible for reviewing the master 
database and flagging any missing or questionable data.Standard, validated assessments of 
pain will be used. Study staff have experience with boys with DMD and these assessments. 
10. Conflict of Interest: 
 
There is no conflict of interest relating to any of the investigators and this protocol beyond the 

professional benefit from academic publication or presentation of the results. 
 
 
Referred Citations:  
 
1. 
Lindsay A, Larson AA, Verma M, Ervasti JM, Lowe DA. Isometric resistance training increases 
strength and alters histopathology of dystrophin-deficient mouse skeletal muscle. J Appl Physiol 
(1985). 2019;126(2):363-375. 
2. 
Lott DJ, Taivassalo T, Cooke KD, et al. Safety, feasibility, and efficacy of strengthening exercise 
in Duchenne muscular dystrophy. Muscle Nerve. 2020. 
3. 
Alemdaroglu I, Karaduman A, Yilmaz OT, Topaloglu H. Different types of upper extremity 
exercise training in Duchenne muscular dystrophy: effects on functional performance, strength, 
endurance, and ambulation. Muscle Nerve. 2015;51(5):697-705. 
4. 
Jansen M, van Alfen N, Geurts AC, de Groot IJ. Assisted bicycle training delays functional 
deterioration in boys with Duchenne muscular dystrophy: the randomized controlled trial "no use 
is disuse". Neurorehabil Neural Repair. 2013;27(9):816-827. 
5. 
Bushby K, Finkel R, Birnkrant DJ, et al. Diagnosis and management of Duchenne muscular 
dystrophy, part 1: diagnosis, and pharmacological and psychosocial management. Lancet Neurol. 
2010;9(1):77-93. 
6. 
Bushby K, Finkel R, Birnkrant DJ, et al. Diagnosis and management of Duchenne muscular 
dystrophy, part 2: implementation of multidisciplinary care. Lancet Neurol. 2010;9(2):177-189. 
7. 
Petrof BJ, Shrager JB, Stedman HH, Kelly AM, Sweeney HL. Dystrophin protects the sarcolemma 
from stresses developed during muscle contraction. Proc Natl Acad Sci U S A. 1993;90(8):3710-
3714. 
8. 
Hoffman EP, Brown RH, Jr., Kunkel LM. Dystrophin: the protein product of the Duchenne 
muscular dystrophy locus. Cell. 1987;51(6):919-928. 
9. 
Moxley RT, 3rd, Ashwal S, Pandya S, et al. Practice parameter: corticosteroid treatment of 
Duchenne dystrophy: report of the Quality Standards Subcommittee of the American Academy of 
Neurology and the Practice Committee of the Child Neurology Society. Neurology. 2005;64(1):13-
20. 
10. 
Matthews E, Brassington R, Kuntzer T, Jichi F, Manzur AY. Corticosteroids for the treatment of 
Duchenne muscular dystrophy. Cochrane Database Syst Rev. 2016(5):CD003725. 

11. 
Angelini C. The role of corticosteroids in muscular dystrophy: a critical appraisal. Muscle Nerve. 
2007;36(4):424-435. 
12. 
McDonald CM, Henricson EK, Abresch RT, et al. Long-term effects of glucocorticoids on 
function, quality of life, and survival in patients with Duchenne muscular dystrophy: a prospective 
cohort study. Lancet. 2018;391(10119):451-461. 
13. 
Cowen L, Mancini M, Martin A, Lucas A, Donovan JM. Variability and trends in corticosteroid 
use by male United States participants with Duchenne muscular dystrophy in the Duchenne 
Registry. BMC Neurol. 2019;19(1):84. 
14. 
Guglieri M, Bushby K, McDermott MP, et al. Developing standardized corticosteroid treatment 
for Duchenne muscular dystrophy. Contemp Clin Trials. 2017;58:34-39. 
15. 
Griggs RC, Herr BE, Reha A, et al. Corticosteroids in Duchenne muscular dystrophy: major 
variations in practice. Muscle Nerve. 2013;48(1):27-31. 
16. 
Escolar DM, Hache LP, Clemens PR, et al. Randomized, blinded trial of weekend vs daily 
prednisone in Duchenne muscular dystrophy. Neurology. 2011;77(5):444-452. 
17. 
Connolly AM, Schierbecker J, Renna R, Florence J. High dose weekly oral prednisone improves 
strength in boys with Duchenne muscular dystrophy. Neuromuscul Disord. 2002;12(10):917-925. 
18. 
Quattrocelli M, Barefield DY, Warner JL, et al. Intermittent glucocorticoid steroid dosing 
enhances muscle repair without eliciting muscle atrophy. J Clin Invest. 2017;127(6):2418-2432. 
19. 
Schakman O, Gilson H, Kalista S, Thissen JP. Mechanisms of muscle atrophy induced by 
glucocorticoids. Horm Res. 2009;72 Suppl 1:36-41. 
20. 
Morrison-Nozik A, Anand P, Zhu H, et al. Glucocorticoids enhance muscle endurance and 
ameliorate Duchenne muscular dystrophy through a defined metabolic program. Proc Natl Acad 
Sci U S A. 2015;112(49):E6780-6789. 
21. 
Arlettaz A, Portier H, Lecoq AM, Labsy Z, de Ceaurriz J, Collomp K. Effects of acute prednisolone 
intake on substrate utilization during submaximal exercise. Int J Sports Med. 2008;29(1):21-26. 
22. 
Arlettaz A, Portier H, Lecoq AM, Rieth N, De Ceaurriz J, Collomp K. Effects of short-term 
prednisolone intake during submaximal exercise. Med Sci Sports Exerc. 2007;39(9):1672-1678. 
23. 
Casuso RA, Melskens L, Bruhn T, Secher NH, Nordsborg NB. Glucocorticoids improve high-
intensity exercise performance in humans. Eur J Appl Physiol. 2014;114(2):419-424. 
24. 
Gorostiaga EM, Czerwinski SM, Hickson RC. Acute glucocorticoid effects on glycogen 
utilization, O2 uptake, and endurance. J Appl Physiol (1985). 1988;64(3):1098-1106. 
25. 
Hyzewicz J, Ruegg UT, Takeda S. Comparison of Experimental Protocols of Physical Exercise 
for mdx Mice and Duchenne Muscular Dystrophy Patients. J Neuromuscul Dis. 2015;2(4):325-
342. 
26. 
Leung DG, Wagner KR. Therapeutic advances in muscular dystrophy. Ann Neurol. 
2013;74(3):404-411. 
27. 
Spaulding HR, Selsby JT. Is Exercise the Right Medicine for Dystrophic Muscle? Med Sci Sports 
Exerc. 2018;50(9):1723-1732. 
28. 
Lovering RM, Brooks SV. Eccentric exercise in aging and diseased skeletal muscle: good or bad? 
J Appl Physiol (1985). 2014;116(11):1439-1445. 
29. 
Vignos PJ, Jr., Watkins MP. The effect of exercise in muscular dystrophy. JAMA. 
1966;197(11):843-848. 
30. 
de Lateur BJ, Giaconi RM. Effect on maximal strength of submaximal exercise in Duchenne 
muscular dystrophy. Am J Phys Med. 1979;58(1):26-36. 
31. 
Scott OM, Hyde SA, Goddard C, Jones R, Dubowitz V. Effect of exercise in Duchenne muscular 
dystrophy. Physiotherapy. 1981;67(6):174-176. 

32. 
Markert CD, Ambrosio F, Call JA, Grange RW. Exercise and Duchenne muscular dystrophy: 
toward evidence-based exercise prescription. Muscle Nerve. 2011;43(4):464-478. 
33. 
Ljubicic V, Burt M, Jasmin BJ. The therapeutic potential of skeletal muscle plasticity in Duchenne 
muscular dystrophy: phenotypic modifiers as pharmacologic targets. FASEB J. 2014;28(2):548-
568. 
34. 
Egan B, Zierath JR. Exercise metabolism and the molecular regulation of skeletal muscle 
adaptation. Cell Metab. 2013;17(2):162-184. 
35. 
Bell EL, Shine RW, Dwyer P, et al. PPARdelta modulation rescues mitochondrial fatty acid 
oxidation defects in the mdx model of muscular dystrophy. Mitochondrion. 2018. 
36. 
Call JA, Voelker KA, Wolff AV, et al. Endurance capacity in maturing mdx mice is markedly 
enhanced by combined voluntary wheel running and green tea extract. J Appl Physiol (1985). 
2008;105(3):923-932. 
37. 
Hulmi JJ, Oliveira BM, Silvennoinen M, et al. Exercise restores decreased physical activity levels 
and increases markers of autophagy and oxidative capacity in myostatin/activin-blocked mdx 
mice. Am J Physiol Endocrinol Metab. 2013;305(2):E171-182. 
38. 
Selsby JT, Acosta P, Sleeper MM, Barton ER, Sweeney HL. Long-term wheel running 
compromises diaphragm function but improves cardiac and plantarflexor function in the mdx 
mouse. J Appl Physiol (1985). 2013;115(5):660-666. 
39. 
Hayes A, Williams DA. Beneficial effects of voluntary wheel running on the properties of 
dystrophic mouse muscle. J Appl Physiol (1985). 1996;80(2):670-679. 
40. 
Palmieri B, Sblendorio V, Ferrari A, Pietrobelli A. Duchenne muscle activity evaluation and 
muscle function preservation: is it possible a prophylactic strategy? Obes Rev. 2008;9(2):121-139. 
41. 
Forbes SC, Walter GA, Rooney WD, et al. Skeletal muscles of ambulant children with Duchenne 
muscular dystrophy: validation of multicenter study of evaluation with MR imaging and MR 
spectroscopy. Radiology. 2013;269(1):198-207. 
42. 
Willcocks RJ, Arpan IA, Forbes SC, et al. Longitudinal measurements of MRI-T2 in boys with 
Duchenne muscular dystrophy: effects of age and disease progression. Neuromuscul Disord. 
2014;24(5):393-401. 
43. 
Willcocks RJ, Rooney WD, Triplett WT, et al. Multicenter prospective longitudinal study of 
magnetic resonance biomarkers in a large duchenne muscular dystrophy cohort. Ann Neurol. 
2016;79(4):535-547. 
44. 
Arpan I, Willcocks RJ, Forbes SC, et al. Examination of effects of corticosteroids on skeletal 
muscles of boys with DMD using MRI and MRS. Neurology. 2014;83(11):974-980. 
45. 
Barnard AM, Willcocks RJ, Finanger EL, et al. Skeletal muscle magnetic resonance biomarkers 
correlate with function and sentinel events in Duchenne muscular dystrophy. PLoS One. 
2018;13(3):e0194283. 
46. 
Sato AY, Peacock M, Bellido T. Glucocorticoid Excess in Bone and Muscle. Clin Rev Bone Miner 
Metab. 2018;16(1):33-47. 
47. 
Haldar SM, Jeyaraj D, Anand P, et al. Kruppel-like factor 15 regulates skeletal muscle lipid flux 
and exercise adaptation. Proc Natl Acad Sci U S A. 2012;109(17):6739-6744. 
48. 
Lee I, Huttemann M, Kruger A, Bollig-Fischer A, Malek MH. (-)-Epicatechin combined with 8 
weeks of treadmill exercise is associated with increased angiogenic and mitochondrial signaling 
in mice. Front Pharmacol. 2015;6:43. 
49. 
Griggs RC, Moxley RT, 3rd, Mendell JR, et al. Prednisone in Duchenne dystrophy. A randomized, 
controlled trial defining the time course and dose response. Clinical Investigation of Duchenne 
Dystrophy Group. Arch Neurol. 1991;48(4):383-388. 

50. 
Mendell JR, Moxley RT, Griggs RC, et al. Randomized, double-blind six-month trial of 
prednisone in Duchenne's muscular dystrophy. N Engl J Med. 1989;320(24):1592-1597. 
51. 
Forbes SC, Willcocks RJ, Triplett WT, et al. Magnetic resonance imaging and spectroscopy 
assessment of lower extremity skeletal muscles in boys with Duchenne muscular dystrophy: a 
multicenter cross sectional study. PLoS One. 2014;9(9):e106435. 
52. 
Mathur S, Lott DJ, Senesac C, et al. Age-related differences in lower-limb muscle cross-sectional 
area and torque production in boys with Duchenne muscular dystrophy. Arch Phys Med Rehabil. 
2010;91(7):1051-1058. 
53. 
Akima H, Lott D, Senesac C, et al. Relationships of thigh muscle contractile and non-contractile 
tissue with function, strength, and age in boys with Duchenne muscular dystrophy. Neuromuscul 
Disord. 2012;22(1):16-25. 
54. 
Triplett WT, Baligand C, Forbes SC, et al. Chemical shift-based MRI to measure fat fractions in 
dystrophic skeletal muscle. Magn Reson Med. 2014;72(1):8-19. 
55. 
Scott E, Eagle M, Mayhew A, et al. Development of a functional assessment scale for ambulatory 
boys with Duchenne muscular dystrophy. Physiother Res Int. 2012;17(2):101-109. 
56. 
Pane M, Mazzone ES, Sivo S, et al. Long term natural history data in ambulant boys with 
Duchenne muscular dystrophy: 36-month changes. PLoS One. 2014;9(10):e108205. 
57. 
Lott DJ, Forbes SC, Mathur S, et al. Assessment of intramuscular lipid and metabolites of the 
lower leg using magnetic resonance spectroscopy in boys with Duchenne muscular dystrophy. 
Neuromuscul Disord. 2014;24(7):574-582. 
58. 
Arora H, Willcocks RJ, Lott DJ, et al. Longitudinal timed function tests in Duchenne muscular 
dystrophy: ImagingDMD cohort natural history. Muscle Nerve. 2018;58(5):631-638. 
59. 
Achenbach TM, Edelbrock CS. Behavioral problems and competencies reported by parents of 
normal and disturbed children aged four through sixteen. Monogr Soc Res Child Dev. 
1981;46(1):1-82. 
60. 
Hendriksen RGF, Lionarons JM, Hendriksen JGM, Vles JSH, McAdam LC, Biggar WD. 
Development of a New Self-Reporting Instrument Measuring Benefits and Side Effects of 
Corticosteroids in Duchenne Muscular Dystrophy: Report from a Pilot Study. J Neuromuscul Dis. 
2017;4(3):217-236. 
61. 
Wei Y, Speechley K, Campbell C. Health-Related Quality of Life in Children with 
DuchenneMuscular Dystrophy: A Review. J Neuromuscul Dis. 2015;2(3):313-324. 
62. 
Krivickas LS. Exercise in neuromuscular disease. J Clin Neuromuscul Dis. 2003;5(1):29-39. 
63. 
Robertson RJ, Goss FL, Aaron DJ, et al. Concurrent muscle hurt and perceived exertion of children 
during resistance exercise. Med Sci Sports Exerc. 2009;41(5):1146-1154. 
64. 
Griggs RC, Miller JP, Greenberg CR, et al. Efficacy and safety of deflazacort vs prednisone and 
placebo for Duchenne muscular dystrophy. Neurology. 2016;87(20):2123-2131. 
65. 
McRae ME, Rourke DA, Imperial-Perez FA, Eisenring CM, Ueda JN. Development of a research-
based standard for assessment, intervention, and evaluation of pain after neonatal and pediatric 
cardiac surgery. Pediatr Nurs. 1997;23(3):263-271. 
66. 
Vry J, Schubert IJ, Semler O, Haug V, Schonau E, Kirschner J. Whole-body vibration training in 
children with Duchenne muscular dystrophy and spinal muscular atrophy. Eur J Paediatr Neurol. 
2014;18(2):140-149. 
67. 
Kostek MC, Gordon B. Exercise Is an Adjuvant to Contemporary Dystrophy Treatments. Exerc 
Sport Sci Rev. 2018;46(1):34-41.