Self-Assembling eVLPs for Transient Base Editor Delivery
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for delivering gene editing agents like base editors (BEs) in vivo face challenges with viral delivery systems, leading to off-target editing, prolonged expression, and integration risks, while non-viral strategies lack efficient delivery to multiple tissues and organs.
Innovation Solution
Development of engineered virus-like particles (eVLPs) that package and deliver therapeutic ribonucleoproteins, such as Cas9 and BEs, with optimized architecture to achieve efficient on-target editing and minimal off-target effects by iteratively addressing cargo packaging, release, and localization bottlenecks.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If viral delivery systems (AAV or LV) are used to deliver base editor encoding DNA, then delivery efficiency to multiple tissues and organs is improved, but off-target editing frequency increases and integration risks occur
Solution Approach 1:
The patent extracts the base editor protein and guide RNA from the viral DNA delivery system and delivers them as pre-assembled ribonucleoprotein (RNP) complexes. This removes the need for viral replication and prolonged expression, thereby reducing off-target editing while maintaining delivery efficiency to multiple tissues and organs.
Solution Approach 2:
The patent employs non-integrating viral vectors or viral-like particles that deliver RNP complexes with transient activity. These short-living delivery vehicles perform their function of delivering the base editor RNP to target cells and then degrade without integrating into the genome, thus avoiding prolonged expression and off-target effects.
2Productivity
If viral delivery systems are used to deliver base editor encoding DNA, then delivery efficiency to multiple tissues and organs is improved, but integration into genome and oncogenesis risk increase
Solution Approach 1:
The patent extracts the base editor as a protein-RNA complex rather than delivering it as encoded DNA. This eliminates the risk of viral integration and subsequent oncogenesis while maintaining the ability to deliver the editing machinery to multiple tissues and organs through optimized viral or viral-like particle vectors.
Solution Approach 2:
The patent employs non-integrating delivery vehicles that transiently deliver the base editor RNP. These delivery systems complete their function and are cleared without integrating into the host genome, thereby eliminating integration risks and associated oncogenesis concerns.
3Manufacturing precision
If base editor DNA is delivered to enable gene editing, then on-target editing efficiency is achieved, but prolonged expression increases off-target editing frequency
Solution Approach 1:
The patent performs preliminary assembly of the base editor protein with its guide RNA into functional RNP complexes before delivery. This pre-assembled state ensures immediate on-target editing activity upon cellular uptake while the complexes remain transient, naturally limiting the duration of action and reducing off-target editing opportunities.
Solution Approach 2:
The patent employs transiently active base editor RNPs delivered via non-integrating vectors. These RNPs perform their editing function and are subsequently degraded by cellular mechanisms, limiting the duration of expression and thereby reducing the window for off-target editing while maintaining high on-target efficiency.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
eVLPs enable highly efficient base editing with minimal off-target editing across various cell types and multiple organs, demonstrating therapeutic potential in mouse models by knocking down serum Pcsk9 levels and restoring visual function.
Implementation Method 1
VLPs, assemblies of viral proteins that can infect cells but lack viral genetic material
Data Source
AI summary
The present disclosure provides virus-like particles for delivering gene editing agents such as nucleic acid-programmable DNA-binding proteins (napDNAbps) and base editor fusion proteins (“BE-VLPs” or “eVLPs”), and systems comprising such eVLPs. The present disclosure also provides polynucleotides encoding the eVLPs described herein, which may be useful for producing said eVLPs. Also provided herein are methods for editing the genome of a target cell by introducing the presently described eVLPs into the target cell. The present disclosure also provides fusion proteins that make up a component of the eVLPs described herein, as well as polynucleotides, vectors, cells, and kits.