CRISPR Protein-Encoding Nucleic Acids for Transient UTR-Enhanced Expression

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Solution Overview

Problem

The application of CRISPR-Cas systems in mammalian genomes is often hampered by poor expression of Cas proteins, which limits their effectiveness in therapeutic applications such as cancer and infectious disease treatment.

Innovation Solution

The use of specific combinations of 3′ and 5′ UTR elements derived from selected genes to enhance the expression of CRISPR-associated proteins like Cas9 or Cpf1, providing transient high-expression profiles to minimize off-target effects.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If CRISPR-Cas systems are used in mammalian genomes, then gene editing functionality is achieved, but protein expression is poor

Engineering Contradiction:

Improvegene editing functionalityVSAvoidCas protein expression

Core Design Contradiction:

ReliabilityVSQuantity of substance

Solution Approach 1:

The patent modifies the mRNA sequence parameters by incorporating specific UTR elements (5' UTR from ATP5A1, RPL32, HSD17B4, SLC7A3, NOSIP or NDUFA4; 3' UTR from GNAS, CASP1, PSMB3, ALB or RPS9) to change the expression level parameter of Cas proteins, achieving both high functionality and sufficient protein quantity

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent creates a composite mRNA structure combining coding regions with specific UTR elements from different genes to achieve synergistic expression enhancement, where the composite structure provides both gene editing capability and adequate protein expression

Inventive Principle:

Principle #40Composite materials

2Duration of action of stationary object

If Cas proteins are expressed continuously, then sustained gene editing capability is maintained, but off-target effects increase

Engineering Contradiction:

Improvegene editing capabilityVSAvoidoff-target effects

Core Design Contradiction:

Duration of action of stationary objectVSObject-affected harmful factors

Solution Approach 1:

The patent employs transient expression strategy where Cas proteins are expressed only during a limited window (around 24 hours) sufficient for gene editing, then expression is turned off, thereby maintaining editing capability while minimizing off-target effects through time-limited action

Inventive Principle:

Principle #19Periodic action

Solution Approach 2:

The patent optimizes the expression timeline to rush through the gene editing process quickly within 24 hours, completing the therapeutic action before potential harmful off-target effects can manifest, effectively skipping the prolonged expression phase

Inventive Principle:

Principle #21Skipping (Rushing through)

Data Source

PatentUS12371699B2Nucleic acids encoding CRISPR-associated proteins and uses thereof

Publication Date: 2025.07.29 CUREVAC SE

AI summary

The present invention relates to the field of biomedicine, and in particular to the field of therapeutic nucleic acids. The present invention provides artificial nucleic acids, in particular RNAs, encoding CRISPR-associated proteins. A (pharmaceutical) composition and kit-of-parts comprising the same are also provided. Furthermore, the present invention relates to the artificial nucleic acid, (pharmaceutical) composition, or kit-of-parts for use in medicine, and in particular in the treatment and/or prophylaxis of diseases amenable to treatment with CRISPR-associated proteins.