HLA-Null Artificial Antigen-Presenting Cell via CRISPR-Cas9
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Solution Overview
Problem
Current methods for regulating HLA expression in cells are limited in their ability to generate artificial antigen-presenting cells (AAPCs) for transplantation and cell therapy, particularly in overcoming alloreactive immune responses and obtaining sufficient quantities of antigen-presenting cells for clinical applications.
Innovation Solution
The development of an HLA class I null cell line using a multiplex CRISPR-Cas9 system, where HLA-A, -B, and -C genes are completely eliminated, and then reintroduced along with co-stimulatory molecules such as CD80, CD83, CD54, CD32, 4-1BBL, and CD70 to create an artificial antigen-presenting cell capable of stimulating T cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional methods (γ-ray induced mutations, somatic cell hybridizations) are used to regulate HLA expression, then HLA null cell lines can be generated, but the process is time-consuming and produces limited quantities of cells
Solution Approach 1:
The patent replaces conventional mechanical/biological methods (γ-ray irradiation, somatic cell hybridization) with a molecular biology approach using CRISPR-Cas9 gene editing system. This substitution enables precise, rapid, and scalable generation of HLA null cell lines, dramatically increasing cell quantity and reducing time requirements while maintaining the desired HLA-negative phenotype
Solution Approach 2:
The patent utilizes CRISPR-Cas9 system to induce large deletions in HLA class I genes by introducing guide RNAs that target specific loci. This parameter change approach (using molecular scissors to delete specific DNA segments) achieves complete HLA elimination much more efficiently than conventional mutation methods, producing HLA null cell lines rapidly and in large quantities
2Reliability
If HLA molecules are completely eliminated to prevent alloreactive immune responses, then transplantation compatibility is improved, but the cells lose their natural antigen-presenting capability
Solution Approach 1:
The patent extracts and removes HLA class I molecules from the cell surface through CRISPR-Cas9-mediated deletion of HLA-A, HLA-B, and HLA-C genes. This extraction eliminates the source of alloreactive immune responses while the cell retains other essential functions and can be engineered to express alternative antigen-presenting molecules as needed
Solution Approach 2:
Instead of trying to modify existing HLA molecules to reduce incompatibility, the patent takes the opposite approach by completely eliminating HLA class I expression. This inversion of the problem-solving strategy (from modification to elimination) achieves complete compatibility while the cell's antigen-presenting capability is restored through alternative means such as expression of exogenous HLA molecules or use of HLA-class-II-restricted T cell stimulation
3Ease of operation
If existing HLA-expressing cell lines are used as artificial antigen-presenting cells, then antigen presentation is possible, but alloreactive immune responses are provoked
Solution Approach 1:
The patent converts the harmful effect of HLA expression (which causes alloreactivity) into a beneficial approach by using CRISPR-Cas9 to eliminate HLA class I genes. The resulting HLA null cell lines serve as ideal artificial antigen-presenting cells that can present antigens without provoking alloreactive immune responses, transforming the problem of HLA compatibility into a solution for creating universal APCs
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 enables the generation of artificial antigen-presenting cells that can effectively induce antigen-specific cytotoxic T cell responses, addressing the limitations of conventional antigen-presenting cells and providing a tool for treating tumors, pathogenic infections, and autoimmune diseases.
Implementation Method 1
a multiplex CRISPR-Cas9 system, where HLA-A, -B, and -C genes are completely eliminated
Implementation Method 2
expresses one or more HLA molecules selected from the group consisting of human leukocyte antigen (HLA)-A, -B, and -C; and a co-stimulatory molecule group including CD80, CD83, CD54, CD32, 4-1BBL, and CD70
Data Source
AI summary
The present invention relates to an artificial antigen-presenting cell prepared from an HLA-null cell line by using a multiplex CRISPR-Cas9 system and the use thereof and, more particularly, to a novel artificial antigen-presenting cell which includes the ability to present antigens of HLA class I and a co-stimulatory molecule group transferred from an HLA-A, -B, -C null cell line generated using a multiplex CRISPR-Cas9 system and to stimulate T cells, an immunotherapeutic agent using the same, and the use thereof for treating tumors, pathogenic infections, and autoimmune diseases.