Promoter-Engineered CD4 T-Cells for Stable FOXP3 Expression
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
The instability of FOXP3 expression in peripheral regulatory T-cells (pTregs) due to epigenetic regulation poses a significant barrier for their therapeutic use in autoimmune diseases and organ transplantation, as it can lead to a conversion to pro-inflammatory CD4 T-cells, worsening autoimmune symptoms.
Innovation Solution
Engineer CD4 T-cells to achieve stable FOXP3 expression by inserting a heterologous promoter upstream of the FOXP3 gene using gene editing techniques, such as CRISPR/Cas9 or TALEN, to bypass epigenetic control and ensure consistent suppressive function.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If peripheral regulatory T-cells (pTregs) are used for therapy, then immune suppression and tolerance induction are achieved, but FOXP3 expression instability due to epigenetic regulation causes conversion to pro-inflammatory cells
Solution Approach 1:
The patent changes the epigenetic state of the FOXP3 gene by demethylating its promoter region, transforming pTregs into cells with stable FOXP3 expression comparable to natural Tregs. This parameter change (methylation status) directly addresses the reliability issue by preventing FOXP3 silencing under inflammatory conditions.
Solution Approach 2:
The patent performs preliminary demethylation treatment of the FOXP3 promoter before therapeutic use of pTregs. This preliminary action establishes stable FOXP3 expression in advance, preventing subsequent conversion to pro-inflammatory phenotypes during therapy.
2Reliability
If FOXP3 expression is enhanced in pTregs, then suppressive function is improved, but epigenetic silencing under inflammatory conditions reduces therapeutic reliability
Solution Approach 1:
The patent changes the epigenetic parameter (DNA methylation status) of the FOXP3 promoter from a silenced state to an active, demethylated state. This parameter change ensures that FOXP3 expression remains stable and resistant to inflammatory-induced silencing, thereby improving both reliability and compositional stability.
3Reliability
If gene editing techniques are used to stabilize FOXP3 expression, then therapeutic potential is improved, but genetic modification complexity increases
Solution Approach 1:
The patent extracts and removes the problematic epigenetic regulation mechanism (methylation-prone promoter region) by demethylating it, thereby isolating the FOXP3 gene from the source of instability. This extraction of the epigenetic control issue simplifies the overall system by eliminating the need for complex continuous regulation.
Solution Approach 2:
The patent uses demethylation agents as intermediary substances to modify the FOXP3 promoter region. These intermediaries chemically alter the DNA methylation status, serving as a bridge between the desired stable expression state and the existing epigenetic landscape, thereby achieving stability without requiring complex genetic engineering.
Data Source
Figure 1
Figure 2
Figure 3
AI summary
Disclosed are methods of making a genetically cell that expressed FOXP3 and methods of treatment. In some embodiments, the method can providing a first nucleotide sequence, wherein the first nucleotide sequence comprises a coding strand, the coding strand comprising one or more regulatory elements and a FOXP3 gene or portion thereof providing a nuclease and performing a gene editing process on the first nucleotide sequence, which edits said one or more regulatory elements, and optionally edits the FOXP3 gene or portion thereof. Methods of treating a subject suffering from an autoimmune disease and subjects suffering the effects of organ transplantation are also provided.