CAR scFv Framework Humanization to Prevent Tonic Signaling

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Solution Overview

Problem

Chimeric antigen receptors (CARs) in immune cells often experience tonic signaling due to self-aggregation of scFvs, leading to T cell exhaustion and impaired antitumor activity, which cannot be effectively corrected by stabilizing the scFv through costimulation or modifying other CAR components.

Innovation Solution

Engineering CARs with humanized framework regions and specific amino acid substitutions to stabilize the scFv, preventing self-aggregation and tonic signaling, while maintaining antigen specificity and allowing for enhanced antitumor activity.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If scFv is stabilized through costimulation or modifying other CAR components, then T cell exhaustion is reduced, but tonic signaling caused by self-aggregation cannot be effectively corrected

Engineering Contradiction:

ImproveT cell exhaustion reductionVSAvoidtonic signaling

Core Design Contradiction:

ReliabilityVSObject-generated harmful factors

Solution Approach 1:

The patent applies parameter changes by modifying the physical and chemical properties of the scFv through amino acid substitutions in the framework regions. Specifically, it replaces murine framework residues with human equivalents (humanization) and introduces stabilizing mutations that alter the scFv's conformational stability, thereby preventing self-aggregation and eliminating tonic signaling while preserving antigen recognition

Inventive Principle:

Principle #35Parameter changes

2Stability of the object's composition

If scFv framework regions are humanized with amino acid substitutions, then scFv stability is improved and self-aggregation is prevented, but antigen specificity must be maintained

Engineering Contradiction:

ImprovescFv stabilityVSAvoidantigen specificity

Core Design Contradiction:

Stability of the object's compositionVSReliability

Solution Approach 1:

The patent applies local quality by making differentiated modifications to different regions of the scFv. The framework regions (FR1-FR4) are humanized with specific amino acid substitutions to enhance stability, while the complementarity-determining regions (CDRs) are preserved to maintain antigen specificity. This localized approach ensures that stability improvements do not compromise binding function

Inventive Principle:

Principle #3Local quality

3Object-generated harmful factors

If CARs are engineered to prevent self-aggregation, then tonic signaling is abrogated, but antitumor activity must be enhanced

Engineering Contradiction:

Improvetonic signaling eliminationVSAvoidantitumor efficacy

Core Design Contradiction:

Object-generated harmful factorsVSProductivity

Solution Approach 1:

The patent converts the harmful effect of scFv instability and self-aggregation into a benefit by using the framework region modifications to achieve dual outcomes: eliminating tonic signaling (harm removal) and enhancing antitumor efficacy (benefit creation). The stabilized scFv prevents T cell exhaustion, allowing CAR-T cells to maintain persistent antitumor activity

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

Data Source

PatentUS20230242666A1Methods and Compositions for the Reduction of Chimeric Antigen Receptor Tonic Signaling

Publication Date: 2023.08.03 CELL MEDICA INC

AI summary

The present disclosure relates to methods and compositions related to Chimeric Antigen Receptors (“CARs”) and modifications to the framework sequences to eliminate tonic signaling. The compositions include modified binding members having a binding specificity to chondroitin sulfate proteoglycan 4 (CSPG4) and are stable when prepared as single chain antibody (scFv) and incorporated into CARs. The methods further include nucleic acid constructs for the expression of CSPG4 CARs, and the application of the CSPG4 CAR to therapeutic methods for the treatment of cancer. The present disclosure also relates to the modification of humanized framework sequences.