CRISPR-Edited TILs for Enhanced Persistence and Expansion
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Solution Overview
Problem
Current methods for culturing tumor infiltrating lymphocytes (TILs) face challenges due to their weak persistence, expansion capacity, and limited ability to kill target cells, primarily attributed to inhibitions from the tumor microenvironment.
Innovation Solution
A method involving the reduction of target gene expression or activity in TILs, followed by co-culturing with feeder cells after exposure to T cell activators and/or growth factors, utilizing a gene regulatory system like CRISPR-Cas proteins and guide RNA to enhance TIL properties such as expansion, persistence, and tumor cell killing ability.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If tumor infiltrating lymphocytes are cultured using conventional methods, then the culture process is simple, but the TILs exhibit weak persistence, weak expansion capacity, and limited tumor cell killing ability
Solution Approach 1:
The patent applies parameter changes by modifying the genetic expression parameters of TILs through CRISPR-Cas9 technology. Specifically, it knocks out inhibitory genes (PD-1, TIM-3, LAG-3) and overexpresses activating genes (4-1BB, CD28, IL-21) to transform TIL properties, enabling them to overcome tumor microenvironment suppression and achieve robust persistence and expansion in vivo
Solution Approach 2:
The patent uses feeder cells as an intermediary component in the co-culture system. These feeder cells provide essential growth factors, cytokines, and survival signals that support TIL expansion and activation. The intermediary feeder cells bridge the gap between simple culture and complex genetic modification by providing a nurturing microenvironment that enhances TIL functionality
2Reliability
If the expression of inhibitory genes in TILs is reduced through gene editing, then tumor cell killing ability is enhanced, but the risk of off-target effects and immune rejection increases
Solution Approach 1:
The patent applies partial action by selectively knocking out only specific inhibitory genes (PD-1, TIM-3, LAG-3) rather than performing comprehensive genome-wide modifications. This targeted approach removes only the necessary suppressive elements while preserving other genomic functions, thereby reducing off-target effects. The partial modification strategy balances therapeutic efficacy with safety by avoiding excessive genetic intervention
Solution Approach 2:
The patent implements preliminary action by conducting thorough in vitro validation and characterization of gene-edited TILs before in vivo administration. This includes assessing off-target effects, ensuring proper gene editing efficiency, and verifying TIL functionality through multiple quality control checkpoints. The preliminary screening and optimization steps are performed to minimize risks before the TILs are transferred to patients
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 method significantly improves TIL properties, including increased numbers, viability, and tumor cell killing capacity, leading to enhanced anti-tumor efficacy.
Implementation Method 1
the guide nucleic acid molecule is capable of binding to the sequence of the target gene
Implementation Method 2
the gene regulatory system comprises a guide nucleic acid molecule and a zymoprotein
Data Source
AI summary
Provided are a modified tumor infiltrating lymphocyte (TIL) and a use thereof. Also provided is a method for culturing the TIL, comprising reducing the expression and/or decreasing the activity of NF-κB pathway inhibitory molecules of the TIL. Further provided is a method for preventing and/or treating tumors by using the TIL.