Direct Sorting of CD8+ T Cells for Low Cross-Reactivity Expansion
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Solution Overview
Problem
Current methods for manufacturing T cells for adoptive immunotherapy are inefficient, costly, and time-consuming, particularly in generating tumor-specific CD8+ T cell clones without prior activation or stimulation.
Innovation Solution
A method involving direct sorting of CD8+ T cells using multimers labeled with different detectable agents to reduce cross-reactivity, followed by expansion in the presence of cytokines, eliminating the need for prior activation and reducing the reliance on antigen-presenting cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Loss of time
If conventional methods are used to generate tumor-specific CD8+ T cell clones, then T cells can be produced, but the process is time-consuming and requires prior activation or stimulation
Solution Approach 1:
The patent applies preliminary action by pre-labeling multimers with detectable agents before the sorting process. This allows direct identification and sorting of antigen-specific T cells without requiring prior activation or stimulation steps, thereby reducing the time required for T cell generation while maintaining high productivity
Solution Approach 2:
The patent extracts and eliminates unnecessary intermediate steps (prior activation or stimulation) from the conventional T cell generation process. By directly sorting T cells based on their affinity for peptide-MHC multimers, the method removes redundant procedural elements, achieving both time reduction and enhanced productivity
2Manufacturing precision
If conventional sorting methods are used, then T cells can be isolated, but cross-reactivity against similar peptides occurs
Solution Approach 1:
The patent applies local quality by using multimers with specific detectable agents that bind to T cells with high affinity for the target peptide-MHC complex. The detectable agents are strategically positioned on the multimer structure to provide localized recognition, enabling precise differentiation between target-specific and cross-reactive T cells, thereby improving manufacturing precision while reducing harmful cross-reactivity
Solution Approach 2:
The patent introduces multimers with detectable agents as intermediaries between the T cells and the sorting system. These multimers serve as mediators that specifically recognize target peptide-MHC complexes, allowing for precise identification and sorting of desired T cells while excluding those with cross-reactivity, thus enhancing both specificity and reducing harmful effects
3Ease of manufacture
If traditional manufacturing processes are used, then T cells can be produced, but the process is costly and complex
Solution Approach 1:
The patent applies segmentation by dividing the manufacturing process into distinct functional modules: multimer preparation, T cell sorting, and expansion. This segmentation simplifies each individual step and makes the overall process more manageable and cost-effective, reducing device complexity while maintaining ease of manufacture
Solution Approach 2:
The patent uses detectable agents as copies or proxies that bind to T cells based on their affinity for target peptides. This copying mechanism simplifies the sorting process by providing a detectable signal that can be read by flow cytometry, eliminating the need for complex functional assays and reducing overall manufacturing complexity while maintaining ease of production
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
This approach yields T cells highly specific to their target peptide with decreased cross-reactivity, achieving rapid production and high purity in a significantly shorter timeframe, suitable for clinical applications.
Implementation Method 1
contacting CD8+ T cells with a first multimer comprising a target peptide in complex with an MHC molecule and with a second multimer comprising an irrelevant peptide in complex with an MHC molecule, wherein the first multimer is labelled with a first detectable agent and the second multimer is labelled with a second detectable agent
Implementation Method 2
sorting the contacted CD8+ T cells to collect sorted CD8+ T cells that are detected positive for the first detectable agent and detected negative for the second detectable agent
Data Source
Figure 1
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Figure 2C~2D
AI summary
Described herein are methods for preparing T cells, including isolating CD8+ T cells from a blood sample obtained from a patient or a donor, culturing the isolated CD8+ T cells in the presence of at least one cytokine, contacting the cultured CD8+ T cells with a multimer containing a target peptide in a complex with an MHC molecule and with at least one binding agent that binds to a T cell surface molecule, in which the multimer is labelled with a first detectable agent and the binding agent is labelled with a second detectable agent, sorting the contacted CD8+ T cells to collect the sorted CD8+ T cells that are detected positive for the first and the second detectable agents, and expanding the collected CD8+ T cells.