Novel In Vivo Gene Editing System

SUMMARY

A engineered OgeuIscB- ωRNA for efficient and precise genome editing with increased potency and specificity while more compact in size compared to other CRISPR-Cas systems.

The Unmet Need: Method to increase the efficiency of the OgeuIscB-ωRNA genome-editing system

• OgeuIscB, an evolutioanary ancestor of Cas9, is an RNA (ωRNA)-guided endonuclease capable of generating double-stranded breaks on genomic DNA in mammalian cells. The combination of programmability and compact size renders the system highly attractive for in vivo gene-editing applications. However, the overall low editing activity and the unstable performance across different loci impede the use of OgeuIscB for gene editing.

• Practical applications of IscB–ωRNA as a genome-editing agent also require careful assessment of its editing fidelity. The OgeuIscB- ωRNA specifies to a 16-nucleotide region in target DNA which is 4-nucleotides shorter than that by Cas9. This may potentially lead to decreased specificity and a higher incidence of off-target cleavage.

The proposed solution: A programmable engineered OgeuIscB-ωRNA genome-editing platform for increased potency and specificity

• The faculty inventors engineered the OgeuIscB-ωRNA system for programmed genome editing by enhancing OGeuIscB variants with boosted DNA cleaveage activity both in vitro and in human cells. The enhanced activity of engineered OgeuIscB–ωRNA further translates to more robust base editing, and directly alters the nucleobase identity without requiring a double-stranded break. The inventors demonstrated the enhanced OgeuIscB–ωRNA systems efficiently edited the human genome across 26 target sites, attaining up to 87.3% indel and 62.2% base-editing frequencies.

• Compared to other CRISPR-Cas systems, the engineered OgeuIscB-ωRNA system is very small in size, which could make it straightforward to package into Adeno-Associated Virus vector for in vivo delivery. With the ease of programmability, high potency, target specificity, and compact size, the engineered OgeuIscB/ωRNA system has the potential of advancing therapeutic application as gene-editing technology.

FIGURE

ADVANTAGES

ADVANTAGES

• Bosted DNA cleavage activity both in vitro and in human cells
• More robust base editing
• Programmable
• Compact size compared to other CRISPR-Cas systems

APPLICATIONS

• In vivo gene editing

PUBLICATIONS

• Yan, H., Tan, X., Zou, S. et al. Assessing and engineering the IscB–ωRNA system for programmed genome editing. Nat Chem Biol (2024).

July 24, 2024

Proof of concept

Patent Pending

Licensing,Co-development

Weixin Tang