Epigenetic TRAC Repression for Safer Immune Cell Engineering
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Solution Overview
Problem
Existing genetic engineering methods for immune cells rely on permanent manipulation and are associated with risks such as chromosomal translocations, undesired insertions and deletions and off-target mutations.
Innovation Solution
The use of epigenetic editors for TRAC gene expression, comprising DNMT domain and transcriptional repressor domain, and DNA-binding domain, and transcriptional repressor domain, and transcriptional repressor domain.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If permanent genetic manipulation is used to engineer immune cells, then the desired therapeutic effect is achieved, but chromosomal translocations and off-target mutations occur
Solution Approach 1:
The patent replaces permanent genetic manipulation (mechanical cutting and joining of DNA) with epigenetic modification (chemical marking of DNA). Instead of using CRISPR-Cas9 nucleases that create double-strand breaks and risk chromosomal translocations, the invention uses fusion proteins with transcriptional repressor domains that chemically modify histone proteins or DNA methylation patterns at target loci, achieving gene silencing without DNA breaks and eliminating the risk of chromosomal translocations and off-target mutations.
Solution Approach 2:
The patent changes the mechanism of gene regulation from permanent genomic alteration to reversible epigenetic modification. By modifying chromatin structure through histone modification or DNA methylation at the TRAC locus, the invention achieves durable gene silencing that can be reversed if needed, whereas traditional genetic engineering creates permanent changes that cannot be reversed and carry high risks of harmful effects.
2Adaptability or versatility
If traditional genetic engineering methods are used, then TRAC expression can be modified, but the risk of undesired insertions and deletions increases
Solution Approach 1:
The patent substitutes mechanical DNA manipulation with chemical epigenetic modification. Instead of using nucleases that create double-strand breaks requiring repair (which leads to insertions and deletions), the invention uses fusion proteins that chemically modify chromatin structure at target loci, achieving gene expression modification without DNA breaks and eliminating the risk of undesired insertions and deletions during the editing process.
3Reliability
If epigenetic editors are used to repress TRAC transcription, then alloreactivity is reduced, but the complexity of the system increases
Solution Approach 1:
The patent divides the epigenetic editor into separate functional domains: a DNA-binding domain (such as dCas9 guided by guide RNA) that targets the TRAC locus, and an effector domain (transcriptional repressor with KRAB domain and DNMT3A) that performs epigenetic modification. This segmentation allows independent optimization of targeting specificity and epigenetic modification function, reducing overall system complexity while maintaining high reliability in reducing alloreactivity.
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
The efficacy of the TRAC gene expression is achieved by the use of transcriptional editors, which are used to generate specific cells with reduced alloreactivity.
Implementation Method 1
a DNA methyltransferase (DNMT) domain and/or a domain that recruits a DNMT
Implementation Method 2
the DNA-binding domain comprises a dead CRISPR Cas (dCas) domain
Data Source
AI summary
This invention relates to compositions and methods comprising epigenetic editors for epigenetic modification of TRAC, as well as nucleic acids and vectors encoding the same. Also disclosed are cells epigenetically modified by the epigenetic editors.