Codon-optimized acid alpha-glucosidase expression cassettes and methods of using same
Inventors
Anguela, Xavier • ARMOUR, SEAN • NORDIN, JAYME
Assignees
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Publication Number
US-12084693-B2
Publication Date
2024-09-10
Expiration Date
Abstract
The invention provides nucleic acids encoding acid α-glucosidase (GAA). In certain embodiments, nucleic acids have greater than about 86% sequence identity to a sequence selected from the group consisting of any of the sequences set forth as SEQ ID NOs:1-5. In certain embodiments, nucleic acids encoding acid α-glucosidase (GAA) contain less than 127 CpG dinucleotides. Expression cassettes, vectors, cells and cell lines and methods of using such nucleic acids encoding acid α-glucosidase (GAA) are also provided.
Core Innovation
The disclosure describes secretable human acid α-glucosidase (GAA) variants and codon-optimized, CpG-reduced human acid α-glucosidase nucleic acids engineered to support liver-directed secretion in an AAV-based gene therapy context. The nucleic acids encode an acid α-glucosidase protein with α-glucosidase activity and are defined by sequence-identity relationships to specified SEQ ID sets, including sequences set forth in SEQ ID NOs:1-5 and SEQ ID NOs:1-24.
Expression cassettes are described with control elements that include an ApoE/hAAT enhancer/promoter, an intron, and polyadenylation elements using wild-type or CpG-reduced bGH polyA sequences. The nucleic acid cassette is flanked by 5′ and 3′ AAV inverted terminal repeats (ITRs), and CpG-reduced elements are described as reducing CpG content across the nucleic acid constructs.
AAV vector systems are described for delivering the nucleic acids, including AAV capsids/serotypes and AAV empty capsids as part of pharmaceutical compositions. The overall design goal described is to obtain circulating GAA with systemic cross-correction, particularly in the context of treating Pompe disease and other glycogen storage diseases.
Claims Coverage
The independent claims cover two nucleic acid groups. Across the provided claim texts, the inventive coverage centers on nucleic acids encoding an acid α-glucosidase protein with α-glucosidase activity and specified SEQ ID-based sequence identity or selection.
High-identity nucleic acid encoding active GAA
A nucleic acid encoding an acid α-glucosidase (GAA) protein, wherein said nucleic acid has greater than 90% sequence identity to a sequence selected from the group consisting of any of the sequences set forth as SEQ ID NOs:1-5, and wherein the encoded GAA protein has α-glucosidase activity.
Selected GAA sequence set
A nucleic acid encoding a GAA selected from the group consisting of SEQ ID NOs:1-24.
Overall, claim coverage is centered on nucleic acids defined by SEQ ID numbers and sequence-identity or selection, with the key functional requirement that the encoded GAA protein has α-glucosidase activity.
Stated Advantages
Improved potency versus parental constructs, including improved expression/secretion and higher plasma GAA activity across models as reported in the document.
Durable plasma/liver expression over months in NHP studies as described in the document.
Partial restoration of expression/clearance effects in the presence of immune responses, as described with rituximab/cyclosporine A and additional NHP immunosuppression experiments.
Documented Applications
Therapeutic efficacy in a Pompe model (Gaa−/− mouse model) is concluded in the document.
Study and comparison of expression potency for AAV-delivered secretable human GAA variants are reported across in vitro and in vivo models including mice, rats, and non-human primates, with plasma GAA activity and expression measured over time.
Immune response effects are evaluated in NHP studies using anti-GAA IgG, including partial restoration with rituximab/cyclosporine A and additional rapamycin co-administration experiments.
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