Congenital Muscular Dystrophy

Biomarker Discovery

James Collins MD, PhDAssistant Professor

Division of NeurologyCincinnati Children’s Hospital Medical Center

Disclosures

Research Foundation Grant: Cure Congenital Muscular Dystrophies

partnered with S.A.M. (www.curecmd.org)

“A characteristic that is objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes, or pharmacologic responses to a therapeutic intervention.”

Biomarkers Definition Working Group, NIH Clin Pharmacol Ther 2001;69:89-95

Biomarker

http://curecmd.org/

Biomarker

Clinical practice• identify risk for or diagnose a disease

• assess disease severity or progression

• predict prognosis

• or guide treatment

http://www.biomarkersconsortium.org/index.php?option=com_content&task=view&id=132&Itemid=184

Biomarker

http://www.prostateuk.org/psa/psa.htm

Liotta, et al. Nature, 2003.

CMD Biomarker Discovery

Taniguchi, M. et al., Biochem Biophys Res Commun, 2006. 342(2): p. 489-502.

Biomarker Drug development

• How does a drug work in the body• Is the drug safe or effective• What dose of the drug is effective• Response to a treatment• Treatment trial - FDA regulatory

approval process

http://www.biomarkersconsortium.org/index.php?option=com_content&task=view&id=132&Itemid=184

Therapeutic

Intervention BiomarkerClinical

Endpoint

Beneficial or Harmful Effects Not Measured by a

Biomarker

Measured to/ Substitute for

Effects of therapeutic interventions on biomarkers and clinical endpoints in clinical

trials.

Biomarkers Definition Working Group, NIH. Clin Pharmacol Ther 2001;69:89-95

affects

Drug development

Biomarker development Discovery phase

• Proximal fluids, cell lines, animal models, tissue of interest• candidates

Qualification phase• Human plasma• Confirm candidate molecules

Verification phase• Population-based (specificity)

Validation and clinical assay development• Sensitivity and specificity

Rifai, N., M.A. Gillette, and S.A. Carr, Protein biomarker discovery and validation: the long and uncertain path to clinical utility. Nat Biotechnol, 2006. 24(8): p. 971-83.

Biomarker discovery

Genomics• Relevant disease genes, expression profiles,

signaling pathways Proteomics

• Protein expression and post-translational modifications

Metabolomics• small molecule metabolites specific to disease

Imaging• Imaging changes reflect disease state

RNA Expression Profiling of Blood from Humans

GeneSpring, Partek and NCI public software to analyze data

DAVID, KEGG and others for pathways

Apply RNA to MICROARRAYS

Whole Blood

Isolate RNA

STORE -700C

Cluster analysis of genes [with ≥ 2.5-fold change]

blood of DMD compared to healthy age matched males)

Wong, B., et al., Gene expression in blood of subjects with Duchenne muscular dystrophy. Neurogenetics, 2009. 10(2): p. 117-25.

DMD Gene expression profile Steroid treatment effect

Lit L. et al., Pharmacogenomics J, 2009. 9(6): p. 411-8.

http://biogratech.com/resources/From+blood+withdrawal+to+RBC+proteomics.PNG

Proteomics approach

Merosin-deficient mice models (dy/dy) muscle

contractile proteins • shift towards embryonic and perinatal

myosin forms

genes involved in cellular adhesion procollagen genes genes related to immune response and

complement activation•van Lunteren, E. et al., Physiol Genomics, 2006. 25(1): p. 85-95.

Gene expression profiling CMD

up-regulation• extracellular matrix and • basement membrane component genes

unique expression pattern • dystrophin-deficient muscle• Unique profile of FCMD compared to MDC1A

Taniguchi, M., et al.Biochem Biophys Res Commun, 2006. 342(2): p. 489-502

Fukuyama-type CMD and Merosin - deficient patients expression profile in muscle

Gene expression profiling CMD

Proteomics - CMD no published proteomic studies 14th international congress WMS society 2009, Geneva,

Switzerland Abstract

• Col6a1-/- mice vs WT using 2D-DGE showed 37 proteins differentially expressed in diaphragm

Bovelenta et al. NMD 2009 EM.P.5.01; doi:10.1016/j.nmd.2009.06.268

Going Forward: “Bench to Bedside”

merosin deficient CMD (proteomic and gene expression profiling)

CMD subtypes Verification: animal models / biobank tissues Qualification: therapeutic interventions Validation Teaming up with multiple academic centers, private

and governmental agencies• Biomarker correlation with natural history outcome

measures, imaging, and functional mobility scales • Goal: biomarker profile assay use as surrogate end point

Challenges

Access to patients and samples Heterogeneity Progressive disorders defining functional endpoints Funding / incentives

Acknowledgements• Carsten Bonnemann

• Kate Bushby

• Ton DeGrauw

• Prasad Devarajan

• Andrew Hershey

• Anne Rutkwoski

• Brenda Wong

• CMD families

References1. Biomarkers Definition Working Group, NIH Clin Pharmacol Ther 2001;69:89-952. Rodland K. Systems biology and biomarker discovery. Disease Markers 2010;28:195-73. http://www.biomarkersconsortium.org/index.php?

option=com_content&task=view&id=132&Itemid=1844. Sorani et al. Clinical and biological data integration for biomarker discovery Drug Discovery

Today 2010, doi:10.1016/j.drudis.2010.06.0055. Fuller, H.R., et al., Valproate and Bone Loss: iTRAQ Proteomics Show that Valproate Reduces

Collagens and Osteonectin in SMA Cells. J Proteome Res, 2010.6. Hampel, H., et al., Biomarkers for Alzheimer's disease: academic, industry and regulatory

perspectives. Nat Rev Drug Discov, 2010. 9(7): p. 560-74.7. Rifai, N., M.A. Gillette, and S.A. Carr, Protein biomarker discovery and validation: the long and

uncertain path to clinical utility. Nat Biotechnol, 2006. 24(8): p. 971-83.8. Wong, B., et al., Gene expression in blood of subjects with Duchenne muscular dystrophy.

Neurogenetics, 2009. 10(2): p. 117-25.9. Lit, L., et al., Corticosteroid effects on blood gene expression in Duchenne muscular dystrophy.

Pharmacogenomics J, 2009. 9(6): p. 411-8.10. van Lunteren, E., M. Moyer, and P. Leahy, Gene expression profiling of diaphragm muscle in

alpha2-laminin (merosin)-deficient dy/dy dystrophic mice. Physiol Genomics, 2006. 25(1): p. 85-95

11. Taniguchi, M., et al., Expression profiling of muscles from Fukuyama-type congenital muscular dystrophy and laminin-alpha 2 deficient congenital muscular dystrophy; is congenital muscular dystrophy a primary fibrotic disease? Biochem Biophys Res Commun, 2006. 342(2): p. 489-502.

12. Bovelenta et al. Gene expression and proteome profiles in Col6a1-/- mice, a model of Ullrich congenital muscular dystrophy. NMD 2009 EM.P.5.01; doi:10.1016/j.nmd.2009.06.268

13. Liotta, L.A., M. Ferrari, and E. Petricoin, Clinical proteomics: written in blood. Nature, 2003. 425(6961): p. 905.