Synthetic methylmalonyl-CoA mutase transgene for the treatment of MUT class methylmalonic acidemia (MMA)
Inventors
Venditti, Charles P. • CHANDLER, Randy J.
Assignees
US Department of Health and Human Services
Publication Number
US-10894077-B2
Publication Date
2021-01-19
Expiration Date
2034-03-14
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Abstract
Synthetic polynucleotides encoding human methylmalonyl-CoA mutase (synMUT) and exhibiting augmented expression in cell culture and/or in a subject are described herein. An adeno-associated viral (AAV) gene therapy vector encoding synMUT under the control of a liver-specific promoter (AAV2/8-HCR-hAAT-synMUT-RBG) successfully rescued the neonatal lethal phenotype displayed by methylmalonyl-CoA mutase-deficient mice, lowered circulating methylmalonic acid levels in the treated animals, and resulted in prolonged hepatic expression of the product of synMUT transgene in vivo, human methylmalonyl-CoA mutase (MUT).
Core Innovation
The invention concerns synthetic polynucleotides encoding human methylmalonyl-CoA mutase (synMUT) that exhibit augmented expression in cell culture and in subjects. These synthetic sequences, codon-optimized relative to the natural human MUT gene, enhance expression upon administration. Specifically, an AAV gene therapy vector (AAV2/8-HCR-hAAT-synMUT-RBG) encoding synMUT under the control of a liver-specific promoter successfully rescued neonatal lethal phenotypes in methylmalonyl-CoA mutase-deficient mice, lowered circulating methylmalonic acid levels, and resulted in prolonged hepatic expression of MUT in vivo.
Methylmalonic acidemia (MMA) is a severe autosomal recessive disorder caused by defects in the mitochondrial enzyme methylmalonyl-CoA mutase (MUT), leading to the accumulation of methylmalonic acid and related metabolites. Current management is limited to dietary restrictions of amino acid precursors and cofactors, which are insufficient to prevent metabolic instability, disease progression, and potentially fatal outcomes. There is a significant unmet need for more effective therapeutic interventions.
The synthetic synMUT transgene offers a novel therapeutic approach by restoring MUT function through viral- or non-viral-mediated gene delivery, thereby preventing metabolic instability and ameliorating disease progression in MMA patients. Furthermore, synMUT can be utilized for in vitro production of MUT enzyme for enzyme replacement therapy administered via various routes, expanding potential treatment modalities beyond dietary management.
Claims Coverage
The claims include two independent claims covering methods of treating MUT-mediated diseases using synthetic polynucleotides and compositions thereof.
Synthetic methylmalonyl-CoA mutase polynucleotide for treatment
Administering a therapeutic amount of a synthetic methylmalonyl-CoA mutase (synMUT) polynucleotide selected from sequences comprising SEQ ID NO:1 or codon-optimized polynucleotides with at least about 70% identity to SEQ ID NO:1 that encode a polypeptide of SEQ ID NO:2 and have equivalent expression in a host but do not have the natural SEQ ID NO:3 nucleic acid sequence.
Use of vectors and promoters to deliver synMUT
Incorporating the synthetic polynucleotide into viral vectors (adenoviral, retroviral, lentiviral, adeno-associated viral) or non-viral vectors (naked DNA, liposomes, nanoparticles, cationic polymers), placed under transcriptional control of ubiquitous, tissue-specific, or regulable promoters including specific named promoters to achieve expression in target tissues.
Delivery methods covering various administration routes
Delivering the therapeutic synthetic polynucleotide by diverse administration routes such as intradermal, subcutaneous, intravenous, intraperitoneal, intraocular, subretinal, renal artery, hepatic vein, intramuscular injections, and physical methods including electroporation, ultrasound-mediated transfection, gene gun, and oral administration.
Genome editing to incorporate synMUT
Employing genome editing techniques using engineered nucleases including Zinc finger nucleases (ZFNs), TALENs, CRISPR/Cas systems, and meganuclease re-engineered homing endonucleases to insert, replace or remove synMUT sequences in subject cells either in vivo or ex vivo for therapeutic purposes.
Treatment with mature MUT enzyme produced using synMUT polynucleotide
Administering a mature methylmalonyl-CoA mutase enzyme produced using the synthetic polynucleotide (comprising residues 33-750 of SEQ ID NO:2) to treat diseases mediated by methylmalonyl-CoA mutase including MMA.
The claims cover administering synthetic codon-optimized methylmalonyl-CoA mutase polynucleotides or their protein products for treating MUT-mediated conditions, utilizing various vectors, promoters, delivery routes, and genome editing technologies to optimize therapeutic application.
Stated Advantages
The synthetic polynucleotide exhibits augmented expression relative to the natural human MUT gene, enabling more efficient production of methylmalonyl-CoA mutase.
Gene therapy using synMUT in animal models rescues neonatal lethal phenotypes and lowers toxic metabolite levels, demonstrating functional efficacy in vivo.
The use of tissue-specific promoters restricts transgene expression, facilitating persistent and safer therapeutic gene expression.
AAV8-hAAT-synMUT demonstrates a better safety profile and lower genotoxicity compared to other vectors, suggesting suitability for human clinical trials.
Documented Applications
Treatment of methylmalonic acidemia (MMA) by restoring MUT enzymatic activity through gene therapy or enzyme replacement.
In vitro production of synthetic MUT enzyme for use in enzyme replacement therapies via oral, subcutaneous, intramuscular, intravenous, or other routes.
Use of synMUT in genome editing to correct MUT deficiencies in patient cells for cellular therapies.
Preclinical treatment of MUT-deficiency in mouse models via administration of AAV vectors delivering synMUT transgene to liver.
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