Engineered Treg Cryopreservation to Preserve CD62L Expression
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Solution Overview
Problem
Cryopreservation of regulatory T cells (Tregs) leads to a loss of CD62L expression, impairing their migratory and homing functions and suppressive capabilities, necessitating lengthy post-thawing culture and expansion steps that are impractical in clinical settings.
Innovation Solution
Transducing Tregs with an exogenous FOXP3 polynucleotide prior to cryopreservation maintains CD62L expression, allowing the cells to retain immunosuppressive functions post-thawing without additional culture or expansion.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If Tregs are cryopreserved, then they can be stored and transported for clinical use, but CD62L expression is lost and suppressive function is impaired
Solution Approach 1:
The patent applies preliminary action by transducing Tregs with FOXP3-expressing vectors before cryopreservation. This pre-treatment ensures that FOXP3 is already present and functional in the cells prior to freezing, allowing them to maintain CD62L expression and suppressive function after thawing without requiring post-thaw culture or expansion steps.
2Manufacturing precision
If Tregs undergo lengthy post-thawing culture and expansion, then CD62L expression and suppressive function can be restored, but clinical implementation becomes impractical
Solution Approach 1:
The patent performs the necessary cellular recovery action in advance by transducing Tregs with FOXP3 before cryopreservation. This eliminates the need for lengthy post-thaw culture and expansion steps, allowing cells to be used immediately after thawing while maintaining high CD62L expression and suppressive function.
Solution Approach 2:
The transduced Tregs possess self-service capability through constitutive FOXP3 expression, which enables them to autonomously maintain CD62L expression and functional phenotype after cryopreservation without requiring external support from prolonged culture conditions or additional stimulation steps.
3Reliability
If FOXP3 is overexpressed in Tregs, then immunosuppressive function is enhanced, but cell proliferation and expansion are reduced
Solution Approach 1:
The patent applies preliminary action by performing FOXP3 transduction before cryopreservation, allowing sufficient time for expression establishment prior to freezing. This timing strategy ensures high immunosuppressive function in the final product while minimizing the impact on expansion, as the cells are not required to undergo extensive post-thaw proliferation.
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
Cryopreserved Tregs with FOXP3 transduction maintain CD62L expression, enabling immediate clinical use with enhanced viability, purity, and potency as off-the-shelf therapeutics for various therapeutic applications.
Implementation Method 1
Transducing Tregs with an exogenous FOXP3 polynucleotide prior to cryopreservation maintains CD62L expression
Implementation Method 2
The effects of freezing and thawing on the Treg cell population are not well defined
Data Source
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AI summary
The present invention relates to a method of preserving CD62L expression in a regulatory T cell (Treg) population that has been cryopreserved, comprising introducing a polynucleotide encoding a FOXP3 polypeptide into the Treg population prior to cryopreservation. The present invention also relates to a method of preserving CD62L in a Treg population after cryopreservation, comprising introducing a polynucleotide encoding a FOXP3 polypeptide into the Treg population and cryopreserving said Treg population. Furthermore, the present invention relates to the use of an exogenous polynucleotide encoding FOXP3 for the preservation of CD62L expression in a Treg population after cryopreservation, and to a cryopreserved engineered Treg, pharmaceutical compositions comprising the cryopreserved engineered Treg and to therapeutic uses thereof.