CD7-Knockout CAR-T Cells to Prevent Self-Killing
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Solution Overview
Problem
In CAR-T cell immunotherapy, the expression of CD7 in T cells poses a risk of mutual killing, necessitating its knockout to avoid self-reactivity while maintaining T cell functionality.
Innovation Solution
An engineered immune cell with a CD7 gene knockout using sgRNA, combined with Cas nuclease, and introduction of a chimeric antigen receptor for targeted therapy, including specific ligand binding, transmembrane, and signaling domains, to treat cancer and autoimmune diseases.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If CD7 is used as a target antigen for CAR-T cell immunotherapy, then therapeutic efficacy against CD7-expressing cancers is improved, but CAR-T cells may undergo mutual killing due to self-reactivity
Solution Approach 1:
The patent applies the extraction principle by removing the CD7 gene from the T cell genome through CRISPR/Cas9-mediated gene knockout. This extracts the harmful self-reactive element (CD7 expression) from the CAR-T cell system, allowing the cells to target CD7 on cancer cells without undergoing mutual killing. The sgRNA sequences provided in the patent are specifically designed to target and eliminate the CD7 gene, resolving the contradiction between maintaining therapeutic efficacy and preventing self-killing.
2Object-affected harmful factors
If CD7 is knocked out in T cells to prevent mutual killing, then self-reactivity is reduced, but T cell development and function may be compromised
Solution Approach 1:
The patent applies the local quality principle by performing CD7 knockout specifically in the CAR-T cell population rather than in all T cells. The CRISPR/Cas9 system is introduced into cultured T cells that will become CAR-T cells, so only these engineered cells lose CD7 expression. This localized modification prevents self-reactivity in the therapeutic cells while leaving normal T cell development and function in the patient unchanged.
Solution Approach 2:
The patent applies the preliminary action principle by knocking out the CD7 gene before introducing the CAR construct into T cells. This preliminary CD7 elimination is performed in the T cell culture before CAR transfection, ensuring that the subsequently generated CAR-T cells will not have self-reactivity. The sgRNA-mediated CD7 knockout is completed prior to CAR expression, preventing any potential self-killing during the therapeutic effect phase.
3Manufacturing precision
If CD7 knockout is performed using CRISPR/Cas9 system, then CD7 expression is eliminated, but off-target effects and gene editing safety may be concerns
Solution Approach 1:
The patent applies the parameter changes principle by providing multiple sgRNA sequences with different target locations and designs within the CD7 gene. The patent lists several sgRNA options (SEQ ID NOs: 1-9) that target different regions of the CD7 gene, allowing optimization of the CRISPR/Cas9 system parameters to achieve high knockout efficiency while minimizing off-target effects. Different sgRNAs can be selected based on their specific binding characteristics and potential off-target profiles.
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 CD7 knockout strategy efficiently prevents CAR-T cell self-killing and enables universal CAR-T cells, enhancing therapeutic efficacy by targeting various antigens while reducing graft-versus-host disease risk.
Implementation Method 1
introducing a Cas nuclease and the above sgRNA into the cell
Data Source
AI summary
Disclosed herein are an engineered immune cell with CD7 gene knock-out and use thereof. According to the present invention, an sgRNA specifically targeting CD7 gene is designed and synthesized, which can accurately target CD7 gene to achieve gene knock-out with high knock-out efficiency. The provided sgRNA can be used for preparing a CD7-targeting engineered immune cell, and can be further used for preparing a CD7-targeting universal CAR-T cell