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-9719080-B2
Publication Date
2017-08-01
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 relates to synthetic polynucleotides encoding human methylmalonyl-CoA mutase (synMUT) that exhibit augmented expression in eukaryotic cells compared to the natural human MUT gene. The synMUT polynucleotides are codon-optimized to enhance expression upon administration, enabling improved transcriptional, translational, and protein refolding efficacy.
Methylmalonic acidemia (MMA) is a severe autosomal recessive disorder caused by defects in the mitochondrial enzyme methylmalonyl-CoA mutase (MUT), which catalyzes the conversion of L-methylmalonyl-CoA to succinyl-CoA. Current treatment consists of dietary restrictions, but patients still experience metabolic instability and progression of disease, leading to hospitalizations and potential fatality. The invention provides a synthetic human MUT transgene that can be used in gene therapy or enzyme replacement to restore MUT function, prevent metabolic instability, and ameliorate disease progression.
Claims Coverage
The patent includes multiple independent claims covering synthetic polynucleotides, expression vectors, and methods of treatment using the synthetic polynucleotide. The inventive features focus on the synthetic polynucleotide design, enhanced expression, and therapeutic application.
Synthetic codon-optimized methylmalonyl-CoA mutase polynucleotide
A synthetic methylmalonyl-CoA mutase (MUT) polynucleotide comprising the nucleic acid sequence of SEQ ID NO:1 or a codon-optimized polynucleotide with at least about 80% identity to SEQ ID NO:1 encoding a polypeptide according to SEQ ID NO:2, having equivalent expression in a host to either SEQ ID NO:1 or SEQ ID NO:3 expression, wherein the codon-optimized polynucleotide does not have the natural MUT nucleic acid sequence (SEQ ID NO:3).
Expression vector comprising synthetic polynucleotide
An expression vector comprising the synthetic methylmalonyl-CoA mutase polynucleotide, including specific vectors such as AAV2/8-HCR-hAAT-RBG designed for liver-specific expression.
Method of treating diseases mediated by methylmalonyl-CoA mutase insufficiency
Methods of treating diseases or conditions mediated by methylmalonyl-CoA mutase, including methylmalonic acidemia (MMA), by administering a therapeutically effective amount of the synthetic methylmalonyl-CoA mutase polynucleotide or variants thereof with high identity to SEQ ID NO:1.
Use of genome editing to insert synthetic polynucleotide
Administration of the synthetic polynucleotide via genome editing techniques employing nucleases such as zinc finger nucleases (ZFNs), TALENs, CRISPR/Cas systems, or engineered homing endonucleases to genetically modify subject cells.
The independent claims encompass the synthetic codon-optimized MUT polynucleotide with augmented expression, its use in expression vectors particularly AAV-based liver-specific vectors, therapeutic methods for treating MMA and related conditions, and genome editing approaches to introduce the synthetic sequences into subject cells.
Stated Advantages
The synthetic codon-optimized MUT gene exhibits enhanced expression compared to the natural human MUT gene, improving therapeutic efficacy.
Gene therapy using the synthetic MUT sequence rescues lethal phenotype in MMA mouse models and lowers circulating methylmalonic acid levels, demonstrating functional correction of the disease.
Use of liver-specific promoters in AAV vectors restricts transgene expression and facilitates persistent expression with a better safety profile, reducing risks such as hepatocellular carcinoma compared to ubiquitous promoters.
Documented Applications
Gene therapy for methylmalonic acidemia (MMA) using synthetic methylmalonyl-CoA mutase polynucleotide delivered via viral vectors (e.g., AAV) targeting the liver or other tissues.
Enzyme replacement therapy by producing synthetic MUT protein through in vitro expression systems for administration by various routes to treat MMA.
Ex vivo gene therapy where patient cells are genetically modified with synMUT via integrating vectors or genome editing for use as cellular therapy.
Production of transgenic animals incorporating the synthetic MUT gene for research or recombinant protein production.
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