Modified Cas9 Polyribonucleotides for Gene Editing Delivery

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Solution Overview

Problem

The delivery of Cas9 proteins and RNA components for CRISPR-Cas9 technology is challenging, limiting its therapeutic applications due to inefficient methods of delivery.

Innovation Solution

The use of modified polyribonucleotides and polynucleotides encoding Cas9 proteins, which can be optimized for expression in mammalian cells and combined with single guide RNAs, are provided in various compositions and methods for in vitro, ex vivo, and in vivo applications, enhancing delivery and activity.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If conventional delivery methods are used for Cas9 proteins and RNA components, then the CRISPR-Cas9 system can be delivered, but the delivery efficiency is insufficient, limiting therapeutic applications

Engineering Contradiction:

Improvedelivery efficiencyVSAvoidtherapeutic application reliability

Core Design Contradiction:

ProductivityVSReliability

Solution Approach 1:

The patent modifies the nucleotide composition of the polyribonucleotide encoding Cas9 by incorporating specific ratios of modified uridines (30-45%) and modified cytidines (5-10%), which changes the chemical parameters of the molecule to improve cellular uptake and stability, thereby enhancing delivery efficiency while maintaining therapeutic reliability

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The invention creates a composite polyribonucleotide structure combining multiple types of modified nucleotides (uridine analogs and cytidine analogs) with natural nucleotides in specific proportions, resulting in a hybrid molecule that exhibits improved delivery properties compared to unmodified sequences

Inventive Principle:

Principle #40Composite materials

2Productivity

If modified polyribonucleotides with high levels of modified nucleotides are used, then translational efficiency is improved, but immunogenicity increases

Engineering Contradiction:

Improvetranslational efficiencyVSAvoidimmunogenicity

Core Design Contradiction:

ProductivityVSObject-affected harmful factors

Solution Approach 1:

The patent optimizes the concentration parameters of modified nucleotides by limiting uridine analogs to 30-45% and cytidine analogs to 5-10%, a balance that maintains high translational efficiency while keeping immunogenicity at acceptable levels for therapeutic use

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The invention applies different modification strategies to different regions of the polyribonucleotide, with specific modification ratios applied throughout the sequence, creating localized optimization that enhances translation without uniformly increasing immunogenicity across the entire molecule

Inventive Principle:

Principle #3Local quality

3Productivity

If unmodified polyribonucleotides are used, then immunogenicity is lower, but expression efficiency in mammalian cells is insufficient

Engineering Contradiction:

Improveexpression efficiencyVSAvoidimmunogenicity

Core Design Contradiction:

ProductivityVSObject-affected harmful factors

Solution Approach 1:

The patent introduces controlled modifications to the polyribonucleotide sequence, changing the chemical composition parameters by incorporating specific ratios of modified nucleotides that enhance expression efficiency in mammalian cells while attempting to minimize immunogenicity through controlled modification levels

Inventive Principle:

Principle #35Parameter changes

Data Source

PatentUS10927383B2Cas9 mRNAs

Publication Date: 2021.02.23 ETHRIS

AI summary

In certain aspects, the disclosure relates to compositions comprising modified Cas9 polyribonucleotides and methods of use, as well as Cas9 polynucleotides and polyribonucleotides.