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

Smart overview of the Invention

Genome editing has advanced significantly with the development of CRISPR-Cas technology, which allows for precise alterations in mammalian genomes. By utilizing guide RNA, CRISPR-Cas can target specific DNA sequences for double-strand cleavage, leading to cellular repair processes that can introduce mutations or insert new genetic material. However, the efficiency of this method is limited by the need for donor templates with extensive homology and the activation of cellular stress responses.

Challenges in Current Gene Therapy Approaches

Existing gene therapy methods face obstacles, particularly in treating human monogenetic diseases due to the vast array of patient-specific mutations. While CRISPR-Cas offers potential solutions through exon-skipping strategies, these approaches are not universally effective. Additionally, traditional gene editing methods struggle with delivering large genes due to size constraints, hindering their application in clinical settings.

Advantages of Retroviral Integrase

Retroviral integrase (IN) presents a promising alternative for genome editing as it facilitates the insertion of large DNA sequences into host genomes without requiring target sequence homology. This non-specific integration capability allows for more flexible delivery of genetic material, although it raises concerns about off-target effects and potential mutations that could lead to disease.

Proposed Method for Treating Friedreich's Ataxia

A novel method for treating Friedreich's Ataxia involves administering a fusion protein that combines retroviral integrase with a CRISPR-associated protein and a nuclear localization signal. This method utilizes a guide nucleic acid targeting specific genomic regions and a donor template encoding frataxin to promote effective gene editing and correction.

Fusion Proteins and Genetic Editing Systems

The invention includes various fusion proteins and nucleic acids designed for efficient genome editing. These systems incorporate components such as CRISPR-associated proteins and nuclear localization signals, enhancing the delivery and integration of therapeutic genes. The approach aims to improve the precision and effectiveness of genetic modifications for treating genetic disorders.