Direct iNKT Cell Reprogramming for Higher Production Efficiency
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Solution Overview
Problem
Existing methods for producing NKT cells are limited by low productivity, high time and cost consumption, and complexity, and there is a need for new resources and production methods to enhance their utilization as immune cell therapeutics, particularly for cancer treatment.
Innovation Solution
A method for directly reprogramming isolated somatic cells using a reprogramming factor and a CAR gene in specific media conditions to produce iNKT cells, bypassing the differentiation process, including a first medium with GSK3β inhibitor and a second medium with AHR agonist or anti-CD3 antibody.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If NKT cells are obtained through primary culture method from peripheral blood, then the cell source is available, but the production efficiency is low due to limited NKT cell amount (0.01%-0.1% in leukocytes)
Solution Approach 1:
The invention changes the production approach by using direct reprogramming of easily obtainable somatic cells (like PBMCs) into NKT cells through controlled expression of reprogramming factors (Oct4, Sox2, Klf4, Myc) under specific culture conditions, rather than relying on the limited natural abundance of NKT cells in peripheral blood
Solution Approach 2:
The invention introduces reprogramming factors as intermediaries to transform somatic cells into NKT cells, using viral vectors (Sendai virus) to deliver the reprogramming genes that mediate the cellular transformation process
2Quantity of substance
If stem cell differentiation method is used to produce NKT cells, then new NKT cell resources can be secured, but the production process becomes complicated and time-consuming
Solution Approach 1:
The invention extracts and eliminates the complicated multi-stage differentiation process by directly reprogramming somatic cells into NKT cells in a simplified two-stage culture system, removing unnecessary intermediate steps while maintaining production efficiency
Solution Approach 2:
The invention segments the reprogramming process into two distinct culture stages: first medium for initial reprogramming and second medium for maturation and expansion, allowing optimized conditions for each phase while simplifying overall process control
3Quantity of substance
If conventional reprogramming method is used, then NKT cells can be produced, but the production time and cost increase
Solution Approach 1:
The invention performs preliminary action by using Sendai virus to deliver reprogramming factors that enable direct conversion of somatic cells to NKT cells, avoiding time-consuming differentiation steps and reducing overall production time to approximately 2-3 weeks
Solution Approach 2:
The invention uses Sendai virus, a non-integrating viral vector that degrades naturally after delivering reprogramming factors, eliminating the need for complex viral vector production and integration safety measures, thereby reducing costs and simplifying the process
Data Source
AI summary
The present invention relates to: a method for producing immunocytes, specifically induced natural killer T (iNKT) cells that are induced by direct reprogramming of isolated somatic cells, and chimeric antigen receptor (CAR)-iNKT cells into which a CAR gene encoding a CAR is introduced; iNKT cells produced by the method; and a cell therapy composition and a pharmaceutical composition for preventing or treating cancer, comprising the iNKT cells.The method according to the present invention can produce, through direct reprogramming, iNKT cells or iNKT cells into which a CAR gene is introduced, from isolated cells so as to simplify the production process and shorten production time, thereby reducing costs, to have excellent NKT cell production efficiency, and to ensure safety according to the production without passing through induced pluripotent stem cells, thereby having an excellent NKT cell production effect distinguished from that of a conventional reprogramming technique. In addition, the iNKT cells or iNKT cells into which a CAR gene is introduced, which are produced by the method, have an excellent cancer cell killing ability, and thus can be effectively used as a cell therapy composition or a pharmaceutical composition for preventing or treating cancer.