Modified complex platform of adeno-associated virus with improved rate of expression of loaded genes and reduced genotoxicity
Inventors
Bae, Suk Chul • Lee, You Soub • Chi, Xinzi • Park, Tae Geun • Kim, Woo-Jin
Assignees
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Publication Number
US-11891617-B1
Publication Date
2024-02-06
Expiration Date
Abstract
Described herein is an adeno-associated virus (AAV) complex platform including an asymmetrically modified inverted terminal repeat (ITR). The AAV complex has advantages of increased productivity and expression efficiency of a transgene, and decreased genotoxicity, by having an asymmetric ITR in which any one of two ITRs is modified. Also, described herein is a composition comprising the adeno-associated virus complex and a method of gene therapy.
Core Innovation
The disclosed invention provides an adeno-associated virus (AAV) complex comprising a polynucleotide sequence encoding a transgene between a first inverted terminal repeat (ITR) and a second ITR. In the AAV complex, the first ITR is not modified, while the second ITR is modified.
The second ITR includes modifications to a stem-loop structure formed of rep-binding element (RBE), RBE’, A, A’, B, B’, C, C’, and D regions. The modified ITR exists as an open-end without forming any of the stem region and the loop region of the stem-loop structure.
This structural change is associated with suppressing concatemer formation, reducing genotoxicity, improving viral productivity, and increasing transgene expression efficiency. The disclosed embodiments further include gene therapy compositions in which the AAV complex is provided with pharmaceutically acceptable carriers for therapeutic gene delivery to subjects.
Claims Coverage
The claim set centers on an AAV complex with asymmetrically modified ITRs, with one independent feature set defining an unmodified first ITR and a modified second ITR that forms an open-end lacking stem and loop regions. Dependent claims further specify between-ITR elements, AAV serotype selection, second-ITR deletion patterns, and a pharmaceutical composition with a pharmaceutically acceptable carrier.
Asymmetrically modified ITRs with unmodified first ITR
An adeno-associated virus (AAV) complex comprising a polynucleotide sequence encoding a transgene between a first inverted terminal repeat (ITR) and a second ITR, wherein the first ITR is not modified and the second ITR is modified.
Open-end modified second ITR without stem or loop regions
In the second ITR, all or part of a stem-loop structure formed of rep-binding element (RBE), RBE’, A, A’, B, B’, C, C’, and D regions is modified such that the modified ITR exists as an open-end without forming any of the stem region and the loop region of the stem-loop structure.
Promoter and polyadenylation positioned between ITRs
The AAV complex further includes an operably linked promoter, a polynucleotide sequence encoding a transgene, and a polyadenylation sequence positioned between the first and second ITRs.
Second ITR modifications defined by specific deletion patterns
The second ITR has the B, B’, C, C’ and RBE’ regions deleted, and additionally deletes either the A or A’ region depending on which is closer to the 3’ end of the (+) strand of the transgene or the 5’ end of the (-) strand of the transgene.
Second ITR includes terminal resolution site and RBE sequence
The second ITR includes a terminal resolution site (trs) sequence and an RBE sequence.
AAV serotype selection from AAV1 to AAV12
An adeno-associated virus (AAV) complex made using an AAV serotype chosen from AAV1 through AAV12.
Pharmaceutical composition with pharmaceutically acceptable carrier
A pharmaceutical composition including the adeno-associated virus complex together with a pharmaceutically acceptable carrier.
Across the claim set, the core inventive concept is an AAV complex with an unmodified first ITR and a second ITR whose stem-loop structure is modified to exist as an open-end lacking stem and loop regions. Dependent claims further define between-ITR elements, specify serotype selection, narrow second-ITR region deletions or inclusion of trs and RBE, and extend to a pharmaceutical composition including a pharmaceutically acceptable carrier.
Stated Advantages
Improved viral productivity.
Increased transgene expression efficiency.
Reduced genotoxicity.
Suppressing concatemer formation.
Reduced integration/insertion consistent with reduced long-term or next-generation expression.
Documented Applications
Gene therapy compositions for therapeutic gene delivery to subjects, formulated with pharmaceutically acceptable carriers.
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