Recombinase for accurate insertion of DNA sequences in eukaryotic cells

By using a recombinase gene editing system and combining HIV integrase with TALE, highly accurate DNA insertion in eukaryotic cells has been achieved, solving the problem of off-target mutations in existing technologies and improving the reliability and safety of gene therapy.

CN121712899APending Publication Date: 2026-03-20RENESSELAER POLYTECHNIC INST +2
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Patent Information

Application Number
CN202480040467.0
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Priority Date
2024-04-26
Filing Date
2024-04-29
Publication Date
2026-03-20

AI Technical Summary

Technical Problem

Existing gene editing technologies lack accuracy when inserting DNA into the body, making them prone to off-target mutations. This limits the widespread application of gene therapy, especially in treatments involving CRISPR or its derivatives.

Method used

The recombinase gene editing system, which includes HIV integrase and transcription activator-like effector (TALE), connects the HIV IN monomer subunit to TALE through a polypeptide linker to form a gene editing construct. This construct is then delivered to target cells using a lentiviral vector, achieving efficient and accurate gene editing.

Benefits of technology

It achieves highly accurate DNA insertion in eukaryotic cells, reduces off-target mutations, improves the reliability and safety of gene therapy, and is suitable for clinical applications.

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Abstract

The invention provides a construction body for specifically inserting a DNA (Deoxyribonucleic Acid) fragment and a transgene site into a genome, wherein the construction body comprises recombinase. The enzyme includes a modified HIV-1 integrase (HIV IN) tetramer having a plurality of mutations to facilitate heterodimer construction of the heterodimer wherein a first outer monomer and a second outer monomer are bound to a first inner monomer and a second inner monomer, respectively. A transcriptional activator-like effector (TALE) binds to the N-terminal region of the outer monomer via a polypeptide linker. The two TALEs confer enzyme sequence specificity and prevent non-specific interactions with the target DNA structure. The modified HIV IN monomer ensures the binding of the TALE to the outermost layer monomer and the arrangement of the TALE relative to the catalytic region of the HIV IN. A polynucleotide is bound to an internal monomer of the HIV IN. Upon transport to the nucleus alone or within the lentiviral capsid, the TALE interacts with DNA to locate the HIV IN, which then integrates the polynucleotide.
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