Comparison of treatment efficacy of very long chain acyl-CoA (VLCAD) deficiency with an AAV9.hVLCAD vector, synthetic VLCAD mRNA, and triheptanoin
Biochemical/Metabolic and Therapeutics
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Secondary Categories:
Introduction:
Very long chain acyl-CoA dehydrogenase (VLCAD) deficiency (VLCADD) is the most prevalent long chain fatty acid oxidation disorder, characterized by hypoglycemia, cardiomyopathy, and recurrent rhabdomyolysis. Previously we have reported the creation of a robust muscle specific long chain acyl-CoA dehydrogenase on a complete VLCAD deficient backgroung (mAcadl-/-:Acadvl-/-; MDKO) mouse model exhibiting the major clinical manifestations in adolescents and adults, recurrent e rhabdomyolysis and progressive cardiomyopathy. We used this model to compare the treatment efficacy of triheptanoin, a synthetic human VLCAD mRNA previously reported to improve symptoms in another VLCADD mouse model, and an AAV9 directed human VLCAD gene replacement therapy.
Methods:
The MDKO mice were fed with regular diet supplemented with 12.5% triheptanoin for 7 days followed by characterization of exercise tolerance, measurement of lactate and creatine phosphokinase (CPK), evaluation of plasma acylcarnitine profiles, and cold tolerance. Histopathological studies in liver, heart and muscle of MDKO were performed. A second group of MDKO mice were injected with mRNA sequence via tail vein (1mg/Kg body weight) and all tests described above repeated. A third group was treated with an AAV9 vector containing the full length human ACADVL (AAV9.hVl) cDNA (1x1012 vector genomes/mouse) and all characterization repeated at 8 weeks, 6months and 1 year post treatment. Cardiac status of these animal was followed by serial cardiac magnetic resonance imaging (cMRI) during the one-year period.
Results:
Adult MDKO mice demonstrate elevated resting CPK (2650 vs 120 U/L) and ran 75 vs 500 meters (the test maximum), respectively, compared with wild type animals on a treadmill. Blood lactate (3.5 vs 7.5 µM) and CPK levels (150 vs 5680 u/L) were higher in MDKO mice pre vs post exercise. The animals also exhibited extreme cold intolerance compared to wild type. Cardiac MRI revealed impaired cardiac function compared to wild type by 6 months of age (43 vs 73% left ventricular ejection fraction). Treatment with triheptanoin and VLCAD mRNA led to some improvement in the treadmill running of the animals (126 and 289 meters, respectively) whereas the AAV9.hVL treated mice were able to complete the full exercise goal of 500 m. The post exercise levels of lactate (5.6, 7.9, and 7.0 µM, respectively) and CPK (567, 3603, 702 U/L, respectively) were lower in AAV9.hVL treated mice than in the triheptanoin and mRNA treated animals. The AAV9.hVl treated mice demonstrated showed no hepatic steatosis or cardiac fibrosis on histology, both dramatically abnormal in untreated animals, one year post treatment. Cardiac left ventricular ejection fraction in AAV9.hVl animals was 59% one year post treatment compared to 43% in untreated animals as measured by cMRI. Of note, cardiac fibrosis was demonstrated by cMRI to be an early finding in MDKO (visible at 2 months of age), predating the functional deficit in cardiac output, and was prevented by AAV9.hVI treatment. All other physiologic parameters in AAV9.hVl treated animals remained stable 1 year post treatment.
Conclusion:
Treatment of MDKO animals with an AAV9-VLCAD vector led to significant improvement in functional testing over a year with better outcome compared to triheptanoin and human VLCAD mRNA, improving exercise tolerance, reducing resting and post exercise rhabdomyolysis and lactic acidosis, and preventing cardiomyopathy. Gene therapy effects were long lasting (1 year). Our results serve as a support for advancing the AAV9 directed VLCAD gene therapy towards human clinical trials. They also demonstrate the potential value of multiple therapeutic modalities in this disease. Recognition that cardiac fibrosis the precedes functional deficits in MDKO animals suggests that this pathophysiology should be explored in patients as it indicates a potential benefit for antifibrosis therapy.
Very long chain acyl-CoA dehydrogenase (VLCAD) deficiency (VLCADD) is the most prevalent long chain fatty acid oxidation disorder, characterized by hypoglycemia, cardiomyopathy, and recurrent rhabdomyolysis. Previously we have reported the creation of a robust muscle specific long chain acyl-CoA dehydrogenase on a complete VLCAD deficient backgroung (mAcadl-/-:Acadvl-/-; MDKO) mouse model exhibiting the major clinical manifestations in adolescents and adults, recurrent e rhabdomyolysis and progressive cardiomyopathy. We used this model to compare the treatment efficacy of triheptanoin, a synthetic human VLCAD mRNA previously reported to improve symptoms in another VLCADD mouse model, and an AAV9 directed human VLCAD gene replacement therapy.
Methods:
The MDKO mice were fed with regular diet supplemented with 12.5% triheptanoin for 7 days followed by characterization of exercise tolerance, measurement of lactate and creatine phosphokinase (CPK), evaluation of plasma acylcarnitine profiles, and cold tolerance. Histopathological studies in liver, heart and muscle of MDKO were performed. A second group of MDKO mice were injected with mRNA sequence via tail vein (1mg/Kg body weight) and all tests described above repeated. A third group was treated with an AAV9 vector containing the full length human ACADVL (AAV9.hVl) cDNA (1x1012 vector genomes/mouse) and all characterization repeated at 8 weeks, 6months and 1 year post treatment. Cardiac status of these animal was followed by serial cardiac magnetic resonance imaging (cMRI) during the one-year period.
Results:
Adult MDKO mice demonstrate elevated resting CPK (2650 vs 120 U/L) and ran 75 vs 500 meters (the test maximum), respectively, compared with wild type animals on a treadmill. Blood lactate (3.5 vs 7.5 µM) and CPK levels (150 vs 5680 u/L) were higher in MDKO mice pre vs post exercise. The animals also exhibited extreme cold intolerance compared to wild type. Cardiac MRI revealed impaired cardiac function compared to wild type by 6 months of age (43 vs 73% left ventricular ejection fraction). Treatment with triheptanoin and VLCAD mRNA led to some improvement in the treadmill running of the animals (126 and 289 meters, respectively) whereas the AAV9.hVL treated mice were able to complete the full exercise goal of 500 m. The post exercise levels of lactate (5.6, 7.9, and 7.0 µM, respectively) and CPK (567, 3603, 702 U/L, respectively) were lower in AAV9.hVL treated mice than in the triheptanoin and mRNA treated animals. The AAV9.hVl treated mice demonstrated showed no hepatic steatosis or cardiac fibrosis on histology, both dramatically abnormal in untreated animals, one year post treatment. Cardiac left ventricular ejection fraction in AAV9.hVl animals was 59% one year post treatment compared to 43% in untreated animals as measured by cMRI. Of note, cardiac fibrosis was demonstrated by cMRI to be an early finding in MDKO (visible at 2 months of age), predating the functional deficit in cardiac output, and was prevented by AAV9.hVI treatment. All other physiologic parameters in AAV9.hVl treated animals remained stable 1 year post treatment.
Conclusion:
Treatment of MDKO animals with an AAV9-VLCAD vector led to significant improvement in functional testing over a year with better outcome compared to triheptanoin and human VLCAD mRNA, improving exercise tolerance, reducing resting and post exercise rhabdomyolysis and lactic acidosis, and preventing cardiomyopathy. Gene therapy effects were long lasting (1 year). Our results serve as a support for advancing the AAV9 directed VLCAD gene therapy towards human clinical trials. They also demonstrate the potential value of multiple therapeutic modalities in this disease. Recognition that cardiac fibrosis the precedes functional deficits in MDKO animals suggests that this pathophysiology should be explored in patients as it indicates a potential benefit for antifibrosis therapy.
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