Duchenne Muscular Dystrophy

1. Duchenne Muscular Dystrophy A PHM142 Presentation PHM142 Fall 2012 Instructor: Dr. Jeffrey Henderson Matthew DeianaMichael De MarcoGheorghe Mando

2. Intro, Pathogenesis, and Symptoms

3. What is DMD? • A recessive X-linked form of muscular dystrophy • Characterized by progressive proximal muscle weakness • Symptoms usually appear in male children before age 5 • By age 10, braces are usually required to aid in locomotion and by age 12, wheelchair dependence occurs in almost all patients • Intellectual impairment may be present along with this condition • Average life expectancy = 25 years http://www.uic.edu/classes/bms/bms655/gfx/pedigree7.gif (http://www.nlm.nih.gov/medlineplus/ency/article/000705.htm)

4. Pathogenesis • Caused by a mutation of the dystrophin gene at locus Xp21 • Dystrophin is a protein that connects the cytoskeleton of muscle fibres to the underlying basal lamina through a protein complex • The absence of dystrophin allows excess calcium to penetrate the sarcolemma of muscle fibres • Increased oxidative stress within the muscle fibres damages the sarcolemma and leads to cell death • The muscle fibres are replaced with adipose and connective tissue http://www.centerforhumanreprod.com/blog/wp-content/uploads/genes-3.jpg (Nowak and Davies 2004)

5. Symptoms • Awkward manner of walking and running • Frequent falls • Difficulty getting up • Fatigue • Progressive difficulty walking • Risk of neurobehavioural disorders (like ADHD) and weakness in cognitive skills • Skeletal deformities (including scoliosis in some cases) http://images.teamsugar.com/files/upl1/1/12981/19_2008/tiptoe.jpg http://www.activemotionphysio.ca/media/img/348843/scoliosis_cause02.jpg (http://www.mayoclinic.com/health/muscular-dystrophy/DS00200/DSECTION=symptoms)

6. Diagnosis and Treatment

7. Diagnostic Signs • Muscle function in everyday activity • Gower’s Sign: • Elevated plasma creatine kinase and/or transaminase levels • Family History (http://nursingcrib.com/wp-content/uploads/gowers-sign.gif?9d7bd4) (Bushby 2010)

8. Diagnostic Tests • Muscle biopsy • Genetic Tests: • 1.) Multiplex PCR and/or MLPA • 2.) Prenatal Testing – Chorionic Villus Sampling/Amniocentesis • Blood testing • Electron Microscopy • EMG • Neuromuscular/Musculoskeletal testing (Bushby 2010; NHS 2011)

9. Treatment • There is no available cure, only management of the symptoms • Glucocorticoids (Prednisone, Deflazacort) – prevent degradation, and improve muscle strength and function • Exercise – improves muscle functioning • Physiotherapy – improves muscle strength, function, and normalization of ROM • Assistive Aids (orthotics, braces) – improves posture and mobility • Surgery • Control of respiratory and heart complications – regular monitoring, assisted ventilation, ACE inhibitors/Beta-blockers, pacemaker • Proper nutrition • Pain management – intervention depends on source • Social/Psychological interventions – coping skills, education, social services (Bushby 2010; NHS 2011)

10. Ongoing and Future Research

11. Exon-Skipping • Antisense oligonucleotides • Bind faulty portions of dystrophin gene pre-mRNA  removed during splicing • Study: improvement in 12 patients with DMD • Pitfall: requires frequent re-administration into muscles http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/E/exon_skipping.png (Goemans and others 2011)

12. Stem Cell Replacement • Using multipotent pericytes • Extracted and grown in culture • Culture with growth factors  differentiate into myocytes • Transduce with dystrophin gene • Systemically administered to patients • Fuse, form myotubes http://www.biolsci.org/v06/p0796/ijbsv06p0796g05.jpg (Dellavalle and others 2007)

13. Gene Therapy • Meganucleases: sequence-specific endonucleases • Induce site-specific double stranded breaks • Repaired by non-homologous end-joining • Results in indels • Corrected reading frame • Benefit: permanent • Challenge: engineer MGNs to target specific sequence http://www.nature.com/nrmicro/journal/v5/n11/images/nrmicro1768-f1.jpg (Chapdelaine and others 2010)

14. Summary • Intro, Pathogenesis, and Symptoms • A recessive, X-linked form of MD – proximal muscle weakness • Mutation of the dystrophin gene at locus Xp21 • Excess Ca2+ in muscle fibre  oxidative stress  fiber death • Replacement by adipose and connective tissue • Awkward walk, fatigue, skeletal deformities • Diagnosis and Treatment • Diagnostic signs: Gower’s sign, elevated creatine kinase, transaminase • Diagnosis: muscle biopsy, mulitplex PCR, MLPA, chorionic villus testing, amniocentesis, neuromuscular/musculoskeletal testing • Treatment: glucocorticoids (prednisone), assistive aids, control respiratory and heart complications (assisted ventilation, ACE inhibitors, Beta-blockers), exercise/nutrition • Ongoing and Future Research • Exon Skipping: antisense oligonucleotides on mutated pre-mRNA  splice out mutated exons  shortened, but functional dystrophin gene product • Stem Cell Replacement: pericytes, cultured w/ growth factors and muscle cells, transduced w/ dystrophin gene  transplanted into patient  form myotubes • Gene Therapy: meganucleases induce double-stranded breaks  insertion/deletion by non-homologous end-joining  correction of reading frame

15. References Bushby K, Finkel R, Birnkrant DJ, Case LE, Clemens PR, Cripe L, Kaul A, Kinnett K, McDonald C, Pandya S, Poysky J, Shapiro F, Tomezsko J, Constantin C. 2010. Diagnosis and management of duchenne muscular dystrophy, part 1: diagnosis, and pharmacological and psychosocial management. Lancet Neurology, 9(1),77-93. Bushby K, Finkel R, Birnkrant DJ, Case LE, Clemens PR, Cripe L, Kaul A, Kinnett K, McDonald C, Pandya S, Poysky J, Shapiro F, Tomezsko J, Constantin C. 2010. Diagnosis and management of duchenne muscular dystrophy, part 2: implementation of multidisciplinary care. Lancet Neurology, 9(2),177-89. Chapdelaine P, Pichavant C, Rousseau J, Paques F, Tremblay JP. 2010. Meganucleases can restore the reading frame of a mutated dystrophin. Gene Ther 17(7):846-58. Dellavalle A, Sampaolesi M, Tonlorenzi R, Tagliafico E, Sacchetti B, Perani L, Innocenzi A, Galvez BG, Messina G, Morosetti R, et al. 2007. Pericytes of human skeletal muscle are myogenic precursors distinct from satellite cells. Nat Cell Biol 9(3):255-67. Goemans NM, Tulinius M, van dA, Burm BE, Ekhart PF, Heuvelmans N, Holling T, Janson AA, Platenburg GJ, Sipkens JA, et al. 2011. Systemic administration of PRO051 in duchenne's muscular dystrophy. N Engl J Med 364(16):1513-22. Mayo Clinic, (2012). Muscular dystrophy: Symptoms – MayoClinic.com. Retrieved from website: http:// www.mayoclinic.com/health/muscular-dystrophy/DS00200/DSECTION=symptoms MedlinePlus, (2012). Duchenne muscular dystrophy: MedlinePlus Medical Encyclopedia. Retrieved from website: http://www.nlm.nih.gov/medlineplus/ency/article/000705.htm

16. National Health Service, (2011). Muscular dystrophy - diagnosis. Retrieved from website: http://www.nhs.uk/ Conditions/ Muscular-dystrophy/Pages/Diagnosis.aspx National Health Service, (2011). Muscular dystrophy - treatment. Retrieved from website: http://www.nhs.uk/ Conditions/ Muscular-dystrophy/Pages/Treatment.aspx Nowak KJ and Davies KE. 2004. Duchenne muscular dystrophy and dystophin: pathogenesis and oppourtinities for treatment.” Embo Reports 5(9): 872-876.