Humanized ACE2 and TMPRSS Loci in Non-Human Animal Models

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Solution Overview

Problem

Current models for studying coronavirus infections, such as SARS-COV and SARS-COV-2, in non-human animals do not effectively replicate human coronavirus infections due to sequence differences between human and non-human animal ACE2 proteins, limiting the assessment of coronavirus infection and therapy efficacy.

Innovation Solution

Development of non-human animals, cells, or genomes that comprise humanized ACE2 and TMPRSS genes, allowing for the expression of recombinant ACE2 and TMPRSS proteins. These models are designed to mimic human coronavirus infections by expressing human ACE2, which is the primary entry point for coronaviruses, and TMPRSS proteins, which are essential for viral infection.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Ease of operation

If non-human animals with endogenous ACE2 and TMPRSS genes are used for coronavirus infection studies, then the model is simpler to maintain and operate, but the model accuracy for replicating human coronavirus infections deteriorates due to sequence differences between human and non-human animal ACE2 proteins

Engineering Contradiction:

Improveease of maintaining animal modelsVSAvoidmodel accuracy for coronavirus infection

Core Design Contradiction:

Ease of operationVSReliability

Solution Approach 1:

The patent applies local quality by humanizing only the critical ACE2 and TMPRSS protein regions that are essential for coronavirus binding and entry, while maintaining the rest of the animal genome as native. This selective humanization of specific loci provides the necessary human-relevant interactions without requiring complete humanization of the animal model, thus balancing model accuracy with operational simplicity.

Inventive Principle:

Principle #3Local quality

Solution Approach 2:

The patent creates an intermediary model system where non-human animals serve as hosts for humanized ACE2 and TMPRSS genes. This intermediary approach allows studying human coronavirus infections in a living animal system that is more accessible and ethically feasible than human subjects, while still providing human-relevant biological responses through the expressed human proteins.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Reliability

If humanized ACE2 and TMPRSS genes are introduced into non-human animals, then the model accuracy for studying coronavirus infections improves, but the genetic modification complexity and manufacturing difficulty worsen

Engineering Contradiction:

Improvemodel accuracy for coronavirus infectionVSAvoidgenetic modification complexity

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

The patent segments the humanization process into distinct loci modifications - separately targeting the ACE2 locus and TMPRSS locus for humanization. This segmentation allows for modular genetic modification strategies, where each locus can be modified independently using targeted approaches such as CRISPR/Cas9 or homologous recombination, reducing the overall complexity compared to simultaneous whole-genome humanization.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The patent uses copying by introducing human ACE2 and TMPRSS gene sequences into the non-human animal genome at appropriate loci. Rather than attempting to recreate the entire human genome in the animal, the approach copies only the essential human gene sequences needed for coronavirus susceptibility, significantly simplifying the genetic modification process while maintaining model accuracy.

Inventive Principle:

Principle #26Copying

3Measurement precision

If human ACE2 protein is used for coronavirus entry, then the viral ligand binding accuracy improves, but the sequence differences between human and non-human animal ACE2 proteins worsen the applicability of standard animal models

Engineering Contradiction:

Improveviral ligand binding accuracyVSAvoidapplicability of standard animal models

Core Design Contradiction:

Measurement precisionVSAdaptability or versatility

Solution Approach 1:

The patent changes the critical parameter of ACE2 protein sequence by introducing human ACE2 gene sequences into the non-human animal. This parameter change ensures that the ACE2 protein structure and binding properties match human ACE2, providing accurate viral ligand binding characteristics essential for studying human coronavirus infections while maintaining the animal model framework.

Inventive Principle:

Principle #35Parameter changes

Data Source

PatentUS20250049006A1Non-human animals comprising humanized ace2 and tmprss loci

Publication Date: 2025.02.13 REGENERON PHARMACEUTICALS INC

AI summary

Non-human animal cells and non-human animals comprising a humanized ACE2 locus and a humanized TMPRSS locus, and methods of using such non-human animal cells and non-human animals are provided. Non-human animal cells or non-human animals comprising a humanized ACE2 locus and a humanized TMPRSS locus express a human ACE2 protein or a chimeric ACE2 protein, fragments of which are from human ACE2; and a human TMPRSS or chimeric TMPRSS protein, fragments of which are from human TMPRSS. Methods are also provided for using such non-human animals comprising a humanized ACE2 locus and a humanized TMPRSS locus to assess in vivo ACE2 activity, e.g., coronavirus infection and/or the treatment or prevention thereof.