Related Experiment Video
Updated: Apr 12, 2026
A Nonviral Approach to Generate Transient Chimeric Antigen Receptor T Cells Using mRNA for Cancer Immunotherapy
Published on: February 21, 2025
Rapid CAR screening and circRNA-driven CAR-NK cells for persistent shed-resistant immunotherapy
Joo-Yoon Chung1,2, Jisu Hong3, Su-Min Yee1
1Medicinal Materials Research Center, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul, 02792, Republic of Korea.
Abstract:
Chimeric antigen receptor (CAR)-based immunotherapies against solid tumors face two major hurdles, the "decoy effect" of shedding antigens that sequester CARs, and the limited persistence of immune effectors within the immunosuppressive tumor microenvironment. Here, we present a mechanistic approach to overcome these barriers by integrating a physiologically relevant screening platform with circular RNA (circRNA) engineering. Unlike conventional screens using immortalized cell lines, we performed rapid functional screening directly in human primary natural killer (NK) cells to identify a novel scFv, CLMS10. Structural modeling revealed that CLMS10 targets a membrane-proximal epitope that overlaps the proteolytic cleavage site, thereby evading inhibition by soluble mesothelin (solMSLN). Furthermore, we demonstrated that circRNA-mediated CAR expression, when codelivered with interleukin-21 (IL-21), confers sufficient stability to withstand the continuous antigen shedding induced by cancer-associated fibroblasts (CAFs), resulting in reduced CAR downregulation. In an in vivo metastatic pancreatic cancer model, IL-21-augmented circCAR-MS10-NK cells exhibited potency comparable to that of lentivirally engineered CAR-NK cells while offering superior manufacturability. Overall, this study establishes a paradigm for generating shed-resistant CAR therapeutics through the strategic integration of epitope-specific functional selection and enhanced RNA stability.
Insights
This study engineered circular RNA (circRNA) to create shed-resistant chimeric antigen receptor (CAR)-NK cells. This novel approach enhances CAR-NK cell persistence and efficacy against solid tumors, overcoming key immunotherapy challenges.
Area of Science:
- Immunology
- Biotechnology
- Oncology
Background:
- Chimeric antigen receptor (CAR)-based immunotherapies face challenges in solid tumors, including antigen shedding and limited immune cell persistence.
- The immunosuppressive tumor microenvironment and decoy effects from shedding antigens hinder CAR T-cell efficacy.
Purpose of the Study:
- To develop a novel CAR-NK cell therapy overcoming antigen shedding and enhancing immune cell persistence in solid tumors.
- To engineer shed-resistant CAR therapeutics using circular RNA (circRNA) technology and functional screening.
Main Methods:
- Functional screening in human primary natural killer (NK) cells to identify a novel scFv, CLMS10, targeting a membrane-proximal epitope.
- Utilizing circular RNA (circRNA) for CAR expression, co-delivered with interleukin-21 (IL-21), to enhance stability and persistence.
- Evaluating the efficacy of engineered CAR-NK cells in an in vivo metastatic pancreatic cancer model.
Main Results:
- Identified CLMS10 scFv that evades inhibition by soluble mesothelin (solMSLN) by targeting a shed-resistant epitope.
- circRNA-mediated CAR expression with IL-21 demonstrated stability against antigen shedding induced by cancer-associated fibroblasts (CAFs).
- Engineered IL-21-augmented circCAR-MS10-NK cells showed comparable potency to lentiviral CAR-NK cells with superior manufacturability in vivo.
Conclusions:
- Established a paradigm for generating shed-resistant CAR therapeutics by integrating epitope-specific selection and circRNA technology.
- circRNA-based CAR expression offers enhanced stability and persistence, crucial for overcoming solid tumor microenvironment challenges.
- This approach provides a promising strategy for developing more effective CAR-based immunotherapies against solid tumors.
