Adeno-associated virus gene therapy for 21-hydroxylase deficiency
Inventors
Bougneres, Pierre • Gao, Guangping
Assignees
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Publication Number
US-12195767-B2
Publication Date
2025-01-14
Expiration Date
Abstract
Disclosed herein are recombinant adeno-associated viral vectors expressing 21-hydroxylase (21OH) protein and related uses for treating 21OH deficiency.
Core Innovation
The invention provides recombinant adeno-associated virus (rAAV) particles for delivering a non-AAV 21-hydroxylase (21OH) protein in a form having an amino acid sequence with at least 95% identity to SEQ ID NO: 1. The rAAV particles include an rAAV vector having at least one AAV inverted terminal repeat (ITR) and a non-AAV nucleotide sequence encoding the 21OH protein operably linked to a CAG promoter, and the particles are defined as an AAV5 serotype.
The invention further defines sequence and regulatory features for the non-AAV nucleotide sequence and its expression control. Example configurations described include a human 21OH (CYP21A2) coding sequence, a codon-optimized CYP21, and additional regulatory sequence elements such as a Kozak sequence and an miR-122 binding site for detargeting. The construct design is described as incorporating additional regulatory elements alongside the AAV and promoter elements to support expression from the rAAV vector.
The invention addresses the need to treat 21-hydroxylase deficiency (21OHD) and congenital adrenal hyperplasia (CAH) by targeting expression in adrenal tissue. The described embodiments indicate intended expression in adrenal cortex/adrenal medulla and related adrenal cell populations, along with therapeutic and pharmaceutical-composition uses.
Claims Coverage
The independent claims cover four main claim categories, comprising rAAV particle composition with defined vector/serotype architecture, a constraint on the selectable ITR serotype sources while maintaining AAV5 particle serotype, a production method for generating the AAV5 rAAV particles, and an rAAV particle variant that adds an miR-122 binding site to the human 21OH expression cassette. Across these independent claims, the inventive features are concentrated on four features: the defined 21OH protein sequence identity, operable linkage to a CAG promoter, inclusion of at least one AAV ITR, and defining the produced/packaged rAAV particle as an AAV5 serotype, with additional sequence-regulatory constraints in the variant claim.
AAV5 rAAV particle with CAG-driven 21OH at least 95% identity to SEQ ID NO:1
An rAAV particle having an rAAV vector comprising at least one AAV inverted terminal repeat (ITR) and a non-AAV nucleotide sequence encoding a 21-hydroxylase (21OH) protein with an amino acid sequence having at least 95% identity to SEQ ID NO: 1, the non-AAV nucleotide sequence operably linked to a CAG promoter, wherein the rAAV particle is an AAV5 serotype.
rAAV particle with selectable ITR serotype origins and CAG-driven 21OH
An rAAV particle having an rAAV vector with a non-AAV nucleotide sequence encoding a 21-hydroxylase (21OH) protein with an amino acid sequence having at least 95% identity to SEQ ID NO: 1, the non-AAV nucleotide sequence operably linked to a CAG promoter, and an rAAV vector comprising at least one AAV inverted terminal repeat (ITR) wherein the ITR is from an AAV of serotype AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, rh10, or rh74, wherein the rAAV particle is an AAV5 serotype.
Method of producing an AAV5 rAAV particle with CAG-driven 21OH
A method of producing an rAAV particle by culturing a host cell containing an rAAV vector with at least one AAV inverted terminal repeat (ITR) and a non-AAV nucleotide sequence encoding a 21-hydroxylase (21OH) protein with an amino acid sequence having at least 95% identity to SEQ ID NO: 1, the non-AAV nucleotide sequence operably linked to a CAG promoter, together with a nucleic acid encoding an AAV rep protein, nucleic acid encoding at least one AAV capsid protein, and sufficient helper functions for packaging, wherein the produced rAAV particle is an AAV5 serotype.
AAV5 rAAV particle with miR-122 binding site in the human 21OH cassette
An rAAV particle having an rAAV vector comprising a non-AAV nucleotide sequence encoding a human 21-hydroxylase (21OH) protein with an amino acid sequence having at least 95% identity to SEQ ID NO: 1 and an miR-122 binding site, wherein the non-AAV nucleotide sequence encoding the human 21OH protein is operably linked to a CAG promoter, and wherein the rAAV particle is an AAV5 serotype.
Overall, the independent claim set is centered on AAV5 rAAV particles whose vectors include AAV ITRs and a CAG promoter-operably linked non-AAV 21OH coding sequence defined by at least 95% identity to SEQ ID NO: 1, with additional claim coverage for selectable ITR serotype origins, for culturing-based production of the AAV5 particles using rep/capsid/helper functions, and for an miR-122 binding site variant in a human 21OH expression cassette.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Treating a subject with 21-hydroxylase deficiency (21OHD) by providing a therapeutically effective amount of the rAAV particle.
Recombinant AAV particle and pharmaceutical-composition uses for 21-hydroxylase deficiency (21OHD) and congenital adrenal hyperplasia (CAH).
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