Post by David on Feb 25, 2007 10:02:48 GMT -5
A Long-term Blinded Controlled Efficacy Study of SNT-MC17/Idebenone in the Dystrophin-Deficient MDX Mouse
Gunnar M. Buyse, Leuven, Belgium, Gerry Van der Mieren, Michael Erb, Jan D'Hooge, Paul Herijgers, Eric Verbeken, Alejandro Jara, Luc Mertens, Leuven, Belgium, Isabelle Courdier-Fruh, Patrizia Barzaghi, Thomas Meier, Liestal, Switzerland
BACKGROUND: The identification of a disease-modifying or protective compound for a certain disease may require early-started and long-term administration, but such type of therapeutic trial is not evident in human patients. We have performed such a trial of SNT-MC17/idebenone in the mdx mouse model of Duchenne muscular dystrophy (DMD), based on the drugs potential to improve mitochondrial respiratory chain function and reduce oxidative stress. DESIGN/METHODS: 200 mg/kg bodyweight of either SNT-MC17/idebenone (ide) or vehicle/placebo (veh) was given from the age of 4 weeks until the age of 10 months in mdx mice (mdx-ide, n=18; mdx-veh, n=13) and wild-type mice (WT-veh, n=9). In vivo cardiac assessments at the age of 10 months included transthoracic echocardiography and strain rate imaging, and haemodynamic pressure-volume loop measurements for determination of intrinsic contractile properties in basal state and during dobutamine stress. Cardiac morphology was evaluated for degree of inflammation and fibrosis. Voluntary wheel running performance was recorded from week 5 to week 12. All evaluators were blinded to mouse type and treatment groups. RESULTS: 10 month old mdx mice showed cardiac hypertrophy and diastolic dysfunction, the latter significantly improving with SNT-MC17/idebenone treatment. During physiological low dose dobutamine stress, placebo-treated mdx mice showed a 58% mortality due to systolic contractile failure, which was significantly reduced to 19% in the SNT-MC17/idebenone treated mdx group. SNT-MC17/idebenone treatment significantly improved long-term voluntary wheel running performance in the mdx mouse. CONCLUSIONS/RELEVANCE: We show that the mdx animal model provides opportunities for long-term controlled preclinical therapeutic studies. We have identified a novel potential therapeutic strategy for DMD, as early-started and long-term treatment with SNT-MC17/idebenone was cardioprotective and improved exercise performance in the dystrophin-deficient mdx mouse. Encouraged by these animal data, SNT-MC17/idebenone is currently under investigation in a randomised controlled trial in DMD patients.
Compartmentalized Vascular Delivery of Micro-dystrophin as a Gene Therapy Treatment for Duchenne Muscular Dystrophy
Louise R. Rodino-Klapac, Chrystal L. Montgomery, K. Reed Clark, Brian Coley, Ryan Jensen, Louis G. Chicoine, Jerry Mendell, Columbus, OH
OBJECTIVE: Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder with monogenic mutations setting the stage for successful gene therapy treatment for patients. Long-term therapeutic goals include systemic delivery of a small dystrophin transgene, micro-dystrophin, delivered by adeno-associated virus (AAV) via the vasculature. For safety reasons, we have proceeded in a stepwise fashion, where the transgene is delivered via the femoral artery to an isolated lower limb or muscle thereof. BACKGROUND: Isolated limb perfusion (ILP) is more likely to the be next step in gene therapy for the following reasons: 1) delivery of vector to lower limb muscles can produce clinically meaningful results; 2) the lower limb can be compartmentalized to prevent spread of virus to other organ systems providing an important measure of safety; 3) delivery of virus in a compartmentalized system provides safe passage for the virus since pre-existing immunity to AAV precludes muscle transduction. DESIGN/METHODS: We first tested ILP of micro-dystrophin in the mdx mouse model of DMD with the goal of comparing the efficiency of AAV serotypes [AAV1, 6, or AAV8] in crossing the vascular barrier leading to widespread gene expression in muscle. For this study we used a micro-dystrophin construct which deletes the untranslated regions and C-terminus, while retaining spectrin repeats 1-3 and 24, and hinges 1, 2, and 4 of full-length dystrophin. RESULTS: Comparative studies suggest that AAV6 and AAV8 deliver and transduce micro-dystrophin by ILP more efficiently than AAV1. In addition, we have been successful in translating the ILP vascular delivery approach to a larger animal model by specifically targeting the gastrocnemius muscle with a fluoroscopy guided catheter to deliver AAV8.GFP (green fluorescent protein). CONCLUSIONS/RELEVANCE: These findings and ongoing studies set the stage for a future clinical trial in DMD patients with vascular delivery of the micro-dystrophin transgene.
Interim Analysis of a Phase 2 Study of PTC124 for Nonsense Mutation Suppression Therapy of Duchenne Muscular Dystrophy (DMD)
Richard Finkel, Samit Hirawat, Brenda Wong, Cincinnati, OH, Kevin Flanigan, Carsten Bonnemann, H. Lee Sweeney, Elisabeth R. Barton, Philadelphia, PA, Michael Howard, Salt Lake City, UT, Valerie J. Northcut, Gary L. Elfring, Langdon L. Miller, S. Plainfield, NJ
OBJECTIVE: To evaluate the safety, compliance, pharmacokinetics (PK) and pharmacological activity of PTC124 in patients with DMD due to a nonsense mutation (premature stop codon). BACKGROUND: PTC124 is a novel, oral, nonantibiotic, small molecule that promotes ribosomal readthrough of nonsense mutations. PTC124 induces production of full-length dystrophin protein and decreases muscle injury in mdx mice, which have a nonsense mutation in the dystrophin encoding DMD gene. DESIGN/METHODS: 26 patients, ages 5 to 13 years, each received PTC124 for 28 days at a low-dose, 16-mg/kg/day [n=6] or a high-dose 40-mg/kg/day [n=20]. Blood PK samples, extensor digitorum brevis muscle biopsies, serum muscle enzymes, and myometry and timed function test data were obtained at baseline and on Day 27. Immunofluorescence expression of dystrophin is being evaluated using a C-terminal antibody. RESULTS: PTC124 was well tolerated with few potentially drug-related adverse events (mild, transient, abdominal pain/diarrhea in 4 patients), no clinically relevant drug-related laboratory abnormalities, and excellent compliance (>98%). PK assessments indicated stable drug exposures over time, although plasma trough concentrations were lower than predicted from PTC124 studies in adult subjects. In vitro assessments demonstrated dose-dependent PTC124-induced dystrophin expression by immunofluorescence in biopsy-derived cultured myotubes. In vivo assessments suggested qualitative increases in muscle biopsy dystrophin protein by immunofluorescence in a number of patients over the 28 days of PTC124 treatment; more definitive quantitative measurements are underway. Serum muscle enzyme levels declined significantly during treatment. Evaluations of dystrophin mRNA expression by polymerase chain reaction and analyses of myometry and functional test data are in progress. CONCLUSIONS/RELEVANCE: These interim data indicate that PTC124 nonsense-suppression therapy is safe and suggest evidence of pharmacodynamic activity over a 28-day treatment. Because the doses tested did not achieve the target plasma concentrations associated with maximal preclinical activity, further dose escalation in an additional cohort of boys with DMD is warranted.
Vascular Endothelial Growth Factor Gene Transfer Using Adeno-Associated Viral Vectors Stimulates Skeletal Muscle Regeneration and Enhances Muscle Function in mdx Mice
Sonia Messina, Mohammed Aguennouz, Anna Mazzeo, Alessandra Bitto, Mauro Giacca, Trieste, Italy, Francesco Squadrito, Messina, Italy, Giuseppe Vita, Messina, Italy
OBJECTIVE: Vascular endothelial growth factor (VEGF) is a major regulator of physiological and pathological angiogenesis. We tested VEGF effect on muscle function, histopathology, immunohistochemistry and biochemical parameters in mdx and wild type mice. BACKGROUND: Several studies support its role in myogenesis and in myoblast migration and survival. Recently it has been reported that the delivery of VEGF using adeno-associated-virus (AAV) vectors reduces muscle damage and promotes muscle regeneration in experimental models of muscle necrosis. DESIGN/METHODS: We performed intramuscular administration into the biceps and tibialis anterior muscles of AAV-VEGF or AAV-LacZ as controls. RESULTS: One month after injection, AAV-VEGF treated muscles showed augmented expression of VEGF (p<0.05) and VEGFR-2 (p<0.01) compared to controls. VEGF treated mdx mice showed higher forelimb strength (+19,5%, p<0.05) as well as higher strength normalised to weight (+ 14,9%, p<0.05). At quantitative morphological evaluation of mdx mice, VEGF-treated muscles showed a reduction of necrotic fibers area (p<0.05) and an increase of small centrally nucleated fibers area (p<0.05). VEGF-treated mdx mice muscle showed an increased number of cells positive for markers of early and late regeneration, respectively myogenin and developmental myosin heavy chain (p<0.05), and also an augmented capillary density in regenerating fibers area (p<0.05). CONCLUSIONS/RELEVANCE: We report the novel observation of a beneficial effect of VEGF in mdx mice exerted mainly by a pro-regenerative and angiogenic effect. Further studies are needed to better clarify the mechanisms of action of VEGF and therefore the possible therapeutic implications in Duchenne muscular dystrophy.
Gunnar M. Buyse, Leuven, Belgium, Gerry Van der Mieren, Michael Erb, Jan D'Hooge, Paul Herijgers, Eric Verbeken, Alejandro Jara, Luc Mertens, Leuven, Belgium, Isabelle Courdier-Fruh, Patrizia Barzaghi, Thomas Meier, Liestal, Switzerland
BACKGROUND: The identification of a disease-modifying or protective compound for a certain disease may require early-started and long-term administration, but such type of therapeutic trial is not evident in human patients. We have performed such a trial of SNT-MC17/idebenone in the mdx mouse model of Duchenne muscular dystrophy (DMD), based on the drugs potential to improve mitochondrial respiratory chain function and reduce oxidative stress. DESIGN/METHODS: 200 mg/kg bodyweight of either SNT-MC17/idebenone (ide) or vehicle/placebo (veh) was given from the age of 4 weeks until the age of 10 months in mdx mice (mdx-ide, n=18; mdx-veh, n=13) and wild-type mice (WT-veh, n=9). In vivo cardiac assessments at the age of 10 months included transthoracic echocardiography and strain rate imaging, and haemodynamic pressure-volume loop measurements for determination of intrinsic contractile properties in basal state and during dobutamine stress. Cardiac morphology was evaluated for degree of inflammation and fibrosis. Voluntary wheel running performance was recorded from week 5 to week 12. All evaluators were blinded to mouse type and treatment groups. RESULTS: 10 month old mdx mice showed cardiac hypertrophy and diastolic dysfunction, the latter significantly improving with SNT-MC17/idebenone treatment. During physiological low dose dobutamine stress, placebo-treated mdx mice showed a 58% mortality due to systolic contractile failure, which was significantly reduced to 19% in the SNT-MC17/idebenone treated mdx group. SNT-MC17/idebenone treatment significantly improved long-term voluntary wheel running performance in the mdx mouse. CONCLUSIONS/RELEVANCE: We show that the mdx animal model provides opportunities for long-term controlled preclinical therapeutic studies. We have identified a novel potential therapeutic strategy for DMD, as early-started and long-term treatment with SNT-MC17/idebenone was cardioprotective and improved exercise performance in the dystrophin-deficient mdx mouse. Encouraged by these animal data, SNT-MC17/idebenone is currently under investigation in a randomised controlled trial in DMD patients.
Compartmentalized Vascular Delivery of Micro-dystrophin as a Gene Therapy Treatment for Duchenne Muscular Dystrophy
Louise R. Rodino-Klapac, Chrystal L. Montgomery, K. Reed Clark, Brian Coley, Ryan Jensen, Louis G. Chicoine, Jerry Mendell, Columbus, OH
OBJECTIVE: Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder with monogenic mutations setting the stage for successful gene therapy treatment for patients. Long-term therapeutic goals include systemic delivery of a small dystrophin transgene, micro-dystrophin, delivered by adeno-associated virus (AAV) via the vasculature. For safety reasons, we have proceeded in a stepwise fashion, where the transgene is delivered via the femoral artery to an isolated lower limb or muscle thereof. BACKGROUND: Isolated limb perfusion (ILP) is more likely to the be next step in gene therapy for the following reasons: 1) delivery of vector to lower limb muscles can produce clinically meaningful results; 2) the lower limb can be compartmentalized to prevent spread of virus to other organ systems providing an important measure of safety; 3) delivery of virus in a compartmentalized system provides safe passage for the virus since pre-existing immunity to AAV precludes muscle transduction. DESIGN/METHODS: We first tested ILP of micro-dystrophin in the mdx mouse model of DMD with the goal of comparing the efficiency of AAV serotypes [AAV1, 6, or AAV8] in crossing the vascular barrier leading to widespread gene expression in muscle. For this study we used a micro-dystrophin construct which deletes the untranslated regions and C-terminus, while retaining spectrin repeats 1-3 and 24, and hinges 1, 2, and 4 of full-length dystrophin. RESULTS: Comparative studies suggest that AAV6 and AAV8 deliver and transduce micro-dystrophin by ILP more efficiently than AAV1. In addition, we have been successful in translating the ILP vascular delivery approach to a larger animal model by specifically targeting the gastrocnemius muscle with a fluoroscopy guided catheter to deliver AAV8.GFP (green fluorescent protein). CONCLUSIONS/RELEVANCE: These findings and ongoing studies set the stage for a future clinical trial in DMD patients with vascular delivery of the micro-dystrophin transgene.
Interim Analysis of a Phase 2 Study of PTC124 for Nonsense Mutation Suppression Therapy of Duchenne Muscular Dystrophy (DMD)
Richard Finkel, Samit Hirawat, Brenda Wong, Cincinnati, OH, Kevin Flanigan, Carsten Bonnemann, H. Lee Sweeney, Elisabeth R. Barton, Philadelphia, PA, Michael Howard, Salt Lake City, UT, Valerie J. Northcut, Gary L. Elfring, Langdon L. Miller, S. Plainfield, NJ
OBJECTIVE: To evaluate the safety, compliance, pharmacokinetics (PK) and pharmacological activity of PTC124 in patients with DMD due to a nonsense mutation (premature stop codon). BACKGROUND: PTC124 is a novel, oral, nonantibiotic, small molecule that promotes ribosomal readthrough of nonsense mutations. PTC124 induces production of full-length dystrophin protein and decreases muscle injury in mdx mice, which have a nonsense mutation in the dystrophin encoding DMD gene. DESIGN/METHODS: 26 patients, ages 5 to 13 years, each received PTC124 for 28 days at a low-dose, 16-mg/kg/day [n=6] or a high-dose 40-mg/kg/day [n=20]. Blood PK samples, extensor digitorum brevis muscle biopsies, serum muscle enzymes, and myometry and timed function test data were obtained at baseline and on Day 27. Immunofluorescence expression of dystrophin is being evaluated using a C-terminal antibody. RESULTS: PTC124 was well tolerated with few potentially drug-related adverse events (mild, transient, abdominal pain/diarrhea in 4 patients), no clinically relevant drug-related laboratory abnormalities, and excellent compliance (>98%). PK assessments indicated stable drug exposures over time, although plasma trough concentrations were lower than predicted from PTC124 studies in adult subjects. In vitro assessments demonstrated dose-dependent PTC124-induced dystrophin expression by immunofluorescence in biopsy-derived cultured myotubes. In vivo assessments suggested qualitative increases in muscle biopsy dystrophin protein by immunofluorescence in a number of patients over the 28 days of PTC124 treatment; more definitive quantitative measurements are underway. Serum muscle enzyme levels declined significantly during treatment. Evaluations of dystrophin mRNA expression by polymerase chain reaction and analyses of myometry and functional test data are in progress. CONCLUSIONS/RELEVANCE: These interim data indicate that PTC124 nonsense-suppression therapy is safe and suggest evidence of pharmacodynamic activity over a 28-day treatment. Because the doses tested did not achieve the target plasma concentrations associated with maximal preclinical activity, further dose escalation in an additional cohort of boys with DMD is warranted.
Vascular Endothelial Growth Factor Gene Transfer Using Adeno-Associated Viral Vectors Stimulates Skeletal Muscle Regeneration and Enhances Muscle Function in mdx Mice
Sonia Messina, Mohammed Aguennouz, Anna Mazzeo, Alessandra Bitto, Mauro Giacca, Trieste, Italy, Francesco Squadrito, Messina, Italy, Giuseppe Vita, Messina, Italy
OBJECTIVE: Vascular endothelial growth factor (VEGF) is a major regulator of physiological and pathological angiogenesis. We tested VEGF effect on muscle function, histopathology, immunohistochemistry and biochemical parameters in mdx and wild type mice. BACKGROUND: Several studies support its role in myogenesis and in myoblast migration and survival. Recently it has been reported that the delivery of VEGF using adeno-associated-virus (AAV) vectors reduces muscle damage and promotes muscle regeneration in experimental models of muscle necrosis. DESIGN/METHODS: We performed intramuscular administration into the biceps and tibialis anterior muscles of AAV-VEGF or AAV-LacZ as controls. RESULTS: One month after injection, AAV-VEGF treated muscles showed augmented expression of VEGF (p<0.05) and VEGFR-2 (p<0.01) compared to controls. VEGF treated mdx mice showed higher forelimb strength (+19,5%, p<0.05) as well as higher strength normalised to weight (+ 14,9%, p<0.05). At quantitative morphological evaluation of mdx mice, VEGF-treated muscles showed a reduction of necrotic fibers area (p<0.05) and an increase of small centrally nucleated fibers area (p<0.05). VEGF-treated mdx mice muscle showed an increased number of cells positive for markers of early and late regeneration, respectively myogenin and developmental myosin heavy chain (p<0.05), and also an augmented capillary density in regenerating fibers area (p<0.05). CONCLUSIONS/RELEVANCE: We report the novel observation of a beneficial effect of VEGF in mdx mice exerted mainly by a pro-regenerative and angiogenic effect. Further studies are needed to better clarify the mechanisms of action of VEGF and therefore the possible therapeutic implications in Duchenne muscular dystrophy.