Genome Editing by Directed Non-Homologous DNA Insertion Using a Retroviral Integrase-Cas Fusion Protein and Methods of Treatment

Smart overview of the Invention

The invention addresses limitations in current genome editing technologies, particularly those using CRISPR-Cas systems. While CRISPR-Cas has advanced genome editing by inducing double-strand DNA breaks and facilitating Homology Directed Repair (HDR), this approach struggles with efficiency and specificity, especially for large DNA sequences. Current methods can trigger unwanted cellular responses and are constrained by the need for homology templates, limiting their effectiveness in treating genetic disorders.

Innovative Approach

This invention introduces a novel method for genome editing by utilizing a fusion protein combining retroviral integrase (IN) and CRISPR-associated (Cas) proteins. This approach leverages the ability of retroviral integrase to insert large DNA sequences without requiring homology, thereby overcoming the size limitations of traditional methods. The fusion protein also includes a nuclear localization signal (NLS) to enhance its delivery to the target site within the genome.

Applications and Benefits

The method is particularly promising for treating genetic disorders such as Friedreich's Ataxia. By using a guide nucleic acid to direct the fusion protein to specific genomic regions, and a donor template nucleic acid for precise insertion, this technology could enable more effective and specific genome modifications. The integration of large genes into host genomes could potentially prevent or reverse disease outcomes, offering a new avenue for gene therapy.

Components of the Invention

The fusion protein comprises several components: a retroviral integrase from various sources such as HIV or RSV, a CRISPR-associated protein like Cas9 or Cas14, and an NLS. These components work together to achieve targeted genome integration. The invention also provides nucleic acid sequences encoding these proteins, enabling their production in biological systems.

Potential Impact

This genome editing system offers significant improvements over existing technologies by providing efficient and precise insertion of large DNA sequences into host genomes. It holds potential for treating a wide range of genetic disorders by addressing patient-specific mutations more effectively than current methods. This innovation could lead to breakthroughs in gene therapy, expanding treatment options for previously untreatable conditions.