FOXP3 And Helios eTregs for Stable T Cell Immunosuppression
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Solution Overview
Problem
Current therapies using regulatory T cells (Tregs) for conditions like graft-versus-host disease (GVHD) face challenges such as the need for large cell numbers, instability of the Treg phenotype, and contamination with conventional T cells, leading to ineffective immunosuppression and increased infection risk.
Innovation Solution
Engineered regulatory T cells (eTregs) with ectopic overexpression of FOXP3 and Helios, generated from total T cell populations using sequential transduction of nucleic acid constructs, ensuring stable expression and homogeneity, and allowing for rapid expansion and purification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If Tregs are isolated by selecting CD4+ CD25+ T cells from collected samples, then Tregs can be obtained for therapy, but the population is contaminated with activated conventional T cells that also express these markers
Solution Approach 1:
The patent extracts the specific Treg population from the mixed T cell population by using targeted selection markers (CD4+CD25+CD127-) and purification techniques, separating the desired Tregs from contaminating conventional T cells that express similar markers
Solution Approach 2:
The patent uses intermediate purification steps including magnetic bead separation and flow cytometry sorting as mediators to achieve high purity Treg populations, where these intermediate technologies facilitate the separation without requiring direct manual isolation
2Quantity of substance
If Tregs are expanded ex vivo to required numbers, then sufficient cell doses can be obtained for therapy, but the Treg phenotype becomes unstable and cells may convert to conventional T cells
Solution Approach 1:
The patent performs preliminary characterization of Treg phenotype markers and expansion conditions before initiating large-scale expansion, establishing optimal culture conditions that maintain phenotype stability throughout the expansion process
Solution Approach 2:
The patent implements monitoring of Treg phenotype markers (FOXP3 expression, CD25, CD127 levels) during expansion to detect early signs of phenotype conversion, allowing adjustment of expansion conditions to maintain Treg stability while achieving required cell numbers
3Productivity
If FOXP3 is ectopically expressed in conventional T cells to generate engineered Tregs, then large numbers of immunosuppressive cells can be produced, but the cells may not fully acquire stable Treg phenotype and function
Solution Approach 1:
The patent combines multiple genetic elements including FOXP3 transgene with Treg-specific promoter sequences and regulatory elements to create a composite genetic construct that ensures both high expression levels and stable Treg phenotype maintenance in engineered cells
Solution Approach 2:
The patent optimizes multiple parameters including transgene copy number, promoter strength, and culture conditions to achieve the optimal balance between high FOXP3 expression levels and stable Treg phenotype, transforming the cellular state from conventional T cell to functional eTreg
Data Source
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AI summary
Cell therapy compositions comprising engineered human regulatory T cells (eTregs) characterized by ectopic overexpression of FOXP3 and Helios protein, produced via introduction of separate nucleic acid constructs respectively encoding FOXP3 and Helios (FOXP3+Helios+ eTregs). Cell therapy compositions comprising mixed populations of CD4+ and CD8+ Treg cells each with ectopic overexpression of FOXP3 and Helios. Methods of making and use the same for therapies involving inflammation and/or a disorder of the immune system.