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-9944918-B2
Publication Date
2018-04-17
Expiration Date
2034-03-14
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
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 naturally occurring human MUT gene. The synthetic gene sequences are codon-optimized to enhance expression upon administration. The invention also includes adeno-associated viral (AAV) gene therapy vectors encoding synMUT under the control of liver-specific promoters, which successfully rescue neonatal lethal phenotypes in methylmalonyl-CoA mutase-deficient mice, lower circulating methylmalonic acid levels, and provide prolonged hepatic expression in vivo.
Methylmalonic acidemia (MMA) is an autosomal recessive disorder caused by defects in the enzyme methylmalonyl-CoA mutase (MUT), leading to accumulation of methylmalonic acid and related metabolites. The disease is currently managed with dietary restrictions but lacks definitive therapy, and patients experience metabolic instability, seizures, strokes, kidney failure, and death despite management. The invention aims to address the lack of effective treatment options for MUT class MMA by providing synthetic, codon-optimized MUT transgenes capable of restoring MUT function and ameliorating disease progression.
The synthetic MUT transgene can be delivered by viral or non-viral gene transfer methods to restore MUT function in patients, prevent metabolic instability, and slow disease progression. Additionally, synMUT can be used for in vitro production of MUT protein for enzyme replacement therapy via various administration routes. The invention includes synthetic polynucleotides selected from defined sequences and variants sharing high sequence identity with augmented expression and encoded protein identical or substantially identical to natural human MUT.
Claims Coverage
The patent includes four main inventive features related to synthetic methylmalonyl-CoA mutase polynucleotides, expression vectors, methods of treating conditions mediated by MUT, and genome editing approaches applying synMUT.
Synthetic methylmalonyl-CoA mutase polynucleotides with codon optimization
The invention covers synthetic methylmalonyl-CoA mutase (synMUT) polynucleotides comprising nucleic acid sequences selected from SEQ ID NO:1, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6, and variants with at least about 80% identity encoding polypeptides with 100% identity to SEQ ID NO:2. These polynucleotides exhibit at least equivalent or increased expression compared to native human MUT (SEQ ID NO:3), achieved by codon optimization replacing less commonly used codons with more frequently used ones.
Expression vectors incorporating synthetic polynucleotides
Vectors comprising the synthetic polynucleotides are included, with a specific example being the AAV2/8-HCR-hAAT-RBG vector that drives liver-specific expression of synMUT.
Methods of treating diseases mediated by methylmalonyl-CoA mutase
Methods involve administering therapeutically effective amounts of the synthetic polynucleotide to subjects in need, specifically targeting methylmalonic acidemia (MMA) and other conditions mediated by low MUT activity.
Genome editing using synMUT polynucleotides
Methods of treating disease by genome editing cells from a subject using engineered nucleases (ZFNs, TALENs, CRISPR/cas, meganucleases) to insert synMUT polynucleotides into cells followed by administering those cells to the subject are claimed.
The claims cover synthetic MUT polynucleotides optimized for expression, expression vectors comprising them, therapeutic methods of administering synMUT to treat MUT-related diseases including MMA, and genome editing strategies utilizing synMUT.
Stated Advantages
The synthetic polynucleotides exhibit enhanced expression levels compared to naturally occurring MUT sequences.
SynMUT gene delivery rescues neonatal lethal phenotypes in MMA mouse models and lowers circulating methylmalonic acid, demonstrating therapeutic efficacy.
The AAV8-hAAT-synMUT vector demonstrates a better safety profile with reduced hepatocellular carcinoma incidence compared to other AAV constructs.
SynMUT sequences facilitate detection and identification using nucleic acid-based assays due to their novel sequences.
Documented Applications
Treatment of methylmalonic acidemia (MMA) via synthetic MUT gene delivery to restore enzyme function and prevent metabolic instability.
In vitro production of MUT enzyme using synMUT for enzyme replacement therapy administered via oral, subcutaneous, intramuscular, or intravenous routes.
Use of AAV viral vectors expressing synMUT for gene therapy in animal models and potential human clinical applications targeting liver or other tissues.
Genome editing of patient cells ex vivo or in vivo using synMUT sequences with engineered nucleases for correction of MUT deficiency.
Creation of transgenic animals encoding synMUT for research, production of recombinant enzyme, or therapeutic testing.
Interested in licensing this patent?