Debio-025 as a candidate to reduce muscular dystrophy pathology
Doug Millay Lab of Jeff Molkentin Cincinnati Children’s Hospital Medical Center Muscular Dystrophy Pathology and Treatment
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Dystrophin (or other DGC) Wt MD Mutation Hallmarks of MD Pathology -Influx of Inflammatory Cells (white arrow) -Central Nuclei (regeneration, black arrow) -Fiber Size Variability (arrowhead) Gene Therapy -Fibrosis
? Degeneration
Inflammation Pharmacologic Agents Regeneration Stem Cell Therapy The Dystrophin-Glycoprotein Complex (DGC)
Lama2-/- laminin Severe phenotype Typically die within 2 months of age Scgd-/- Resembles human disease closely Reduced survival
αDG sarcoglycan complex extracellular β DG
CAV 3 TRPCs syn intracellular NOS db dystrophin actin
Function: Connects the cytoskeleton to the extracellular matrix, stabilizing the mdx sarcolemma The DGC, Ca2+, and Degeneration
2+ Ca Ca2+ Ca2+ 2+ Ca Ca2+
Contraction DGCX DGCX Ca2+ Entry via Sarcolemmal Lesions What are the pathways by which And/or Channel Activity elevated intracellular Ca2+ causes damage?
Ca2+ Membrane Repair Proteins
Apoptotic Calpain ROS Mitochondria Signaling
Necrosis Fiber Death Mitochondrial Calcium Overload
Adapted from Dr. Christopher Baines Muscular Dystrophy Mice Have Swollen Mitochondria
Wt Absorbance, Swelling 0.6 Scgd-/- 0.5 0.4 * 0.3 0.2
Absorbance 0.1 6 6 0 Wt Scgd-/-
Millay DP et al. Nat. Med. (2008) The Mitochondrial Pore
¾Cyclophilin D (gene=Ppif) is a critical component of the mitochondrial pore.
¾ Ca2+ and oxidative damage can lead to pathological opening and cell death, HOWEVER Ppif-/- mitochondria are resistant.
OMM
IMM
CypD Ca2+ ROS Loss of cyclophilin D Rescues Muscle Weights
Wt Scgd-/- Scgd-/- Ppif-/- Ppif-/-
8 6 wks * 12 8 Mo. # # 10 6 * 8 4 # 6 # 4 * 2 2 * MW/TL (mg/mm) MW/TL
6 7 6 3 6 7 6 3 (mg/mm) MW/TL 0 0 9 10 10 3 9 10 10 3 Gastroc. Quad. Gastroc. Quad.
Millay DP et al. Nat. Med. (2008) Genetic Ablation of cyclophilin D Reduces Pathology and 25 #
Increases Function in Scgd-/- Muscle ) 2 20 * 15 (N/cm -/- 6 6 6 6 Wt Scgd Scgd-/- Ppif-/- Specific force 10 Wt Scgd-/- Scgd-/- Diaph. Ppif-/- Ppif-/- Gastroc. Quadriceps
Millay DP et al. Nat. Med. (2008) Mechanism of Protection
Wt Scgd-/- Scgd-/- Ppif-/-
5,000X
10,000X
Millay DP et al. Nat. Med. (2008) Mechanism of Protection
Wt Scgd-/- Scgd-/- Ppif-/-
30,000X
Millay DP et al. Nat. Med. (2008) Analysis of Cardiac Defects
Wt
40 # 30 * Scgd-/- 20 FS (%)
10 11 7 11 0
Wt Scgd-/- Ppif-/- Scgd-/- Scgd-/- Ppif-/-
Millay DP et al. Nat. Med. (2008) Increased Lifespan and Body Weight in Lama2-/- Mice Lacking cyclophilin D Wt Lama2-/- Lama2-/- Ppif-/-
4
3 # 2 # * # 1 * * MW/TL (mg/mm) MW/TL 10 7 6 10 7 6 10 7 6 0
Millay DP et al. Nat. Med. (2008) movie Ca2+ Ca2+ 2+ 2+ 2+ Ca2+ Ca Ca Ca Ca2+ DGC Calcium-Induced X Ca2+ Wt -/- Degeneration through the Mitochondrial Pore
Mitochondrial Pore
CypD
Inhibitors of cyclophilin D (CyclosporineA) (Debio-025) Cyclophilin D Inhibitors
¾ Cyclosporine A ¾ Inhibitor of mitochondrial pore, but also calcineurin (regeneration). Is also an immuno-suppressant. ¾ Effective in reducing mitochondrial abnormalities in patients lacking collagen VI (Merlini L et. al. Proc Natl Acad Sci U S A. 2008 Apr 1;105(13):5225-9)
¾ Debio-025 (Db25) ¾ Cyclosporine analog and cyclophilin inhibitor (10X more potent than cyclosporine A). No effect on calcineurin.
¾Reduced response to apoptosis in cells from patients with collagen VI deficiency (Angelin A et. al. Proc Natl Acad Sci U S A. 2007 Jan 16;104(3):991-6)
¾ Already proved to be safe in humans for treatment of Hepatitis C virus (Currently in Stage II Clinical Trials). Debio-025 Reduces Disease Progression in mdx Mice
12 0.10 * 10 0.08 Wt-Veh * # Wt-Db25 8 0.06 # mdx-Veh 6 # 0.04 mdx-Db25 *
absorbance 4 0.02 # Δ 7 7 9 9 (mg/mm) MW/TL 2 * 0 7 7 7 9 9 PEG 0 7 7 9 9 7 9 9 Gastroc. Quad. TA
Millay DP et al. Nat. Med. (2008) Debio-025 Reduces Disease Progression in mdx Mice vehicle Debio-025
Wt Wt
mdx mdx
Millay DP et al. Nat. Med. (2008) Debio-025 Reduces Disease Progression in mdx Mice
Wt-Veh Wt-Db25
mdx-Veh mdx-Db25 Millay DP et al. Nat. Med. (2008) Conclusions
¾ Inhibition of cyclophilin D may serve as a novel approach for treating MDs.
¾ Debio-025 may be a drug that can be used in humans to reduce MD disease progression and prolong life. Future Directions (Transition of Debio-025 into Clinic)
Gene Replacement (Cure) Gene Therapy Stem Cell Therapy
RNA Manipulations (Subset of Patients)
Pharmacologic Agents
Combinatorial Therapy Future Directions (Transition of Debio-025 into Clinic)
Dystrophin Deficient Golden Retriever October 2008—Puppies Born
Mid-November 2008—Analysis of CK levels and quadriceps biopsy
Late November 2008—Separation of affected puppies into control and treatment groups based on disease severity
December 2008—Begin treatment at 2 months of age
Collaborators: Monthly Tests: -Blood Draws (CK levels, toxicity) -Muscle Biopsy (assessment of pathology) -Human Genome Research Center of the University of São Paulo, -Video Recording (functional analysis) São Paulo Brazil
Echocardiography at 6 and 10 months of age -DEBIOPHARM GROUP (Debio-025) (cardiac function)
-Parent Project Muscular Dystrophy October 2009—End treatment at 10 months of age (8 months of Debio-025 treatment) Jeff Molkentin Scott Blair Acknowledgements Jop VanBerlo John Elrod Joerg Heineke Chris Baines Jennifer Kwong Allen York Xu Wu Majorie Maillet Hiro Nakayama Mannix Auger-Messier Matt Benard Michelle Sargent Sanjeewa Goonasekera Jeff Lynch Qinghang Liu Izhak Kehat Xu Wu Robbins Lab (EM) Erin Wissing Functional Analysis Jeffrey Robbins H. Lee Sweeney- UPenn Angela Lorts Hanna Osinska Elisabeth Barton- UPenn Jennifer Schwanekamp Jason Karch Funding NIH Jain Foundation PPMD