Mesenchymal Stromal Cells with Chimeric Antigen Receptors for Enhanced Immunosuppression

Rheumatology and Immunology Medicine Cell Biology Humanities and Others Life Sciences Immunology
mesenchymal stromal cells chimeric antigen receptors immunosuppression graft-versus-host disease cell therapy

Mesenchymal Stromal Cells with Chimeric Antigen Receptors for Enhanced Immunosuppression

Background

Mesenchymal Stromal Cells (MSCs) are pluripotent cells found in almost all tissues and possess significant immunosuppressive and regenerative properties. These features make MSCs widely studied for the treatment of immune diseases and tissue regeneration. Although clinical trials of allogeneic MSCs have demonstrated their safety, their immunosuppressive efficacy and therapeutic outcomes have so far been unsatisfactory. To enhance the immunosuppressive effect of MSCs, this study employed cell engineering techniques to modify primary MSCs derived from healthy donor adipose tissue, developing a novel therapeutic strategy.

Source of the Paper

This study was conducted by Olivia Sirpilla, R. Leo Sakemura, Mehrdad Hefazi, and others from the T Cell Engineering department and Division of Hematology at Mayo Clinic. The paper was published in the April 2024 issue of Nature Biomedical Engineering.

Research Process

Experimental Design and Procedure

  1. Optimization and Expression of Chimeric Antigen Receptors (CARs):

    • The research team successfully achieved high-efficiency transduction of CARs in MSCs derived from healthy donor adipose tissue using lentiviral vectors enhanced with polycationic enhancers. Specifically, protein sulfation optimization achieved a transduction efficiency of ≥70%.
    • Various antigen-specific CAR constructs were used in the experiments, particularly E-cadherin (E-cad)-targeting CAR-MSCs (ECCAR-MSCs) aimed at graft-versus-host disease (GVHD).

  2. Development of Ecad-directed CAR-MSCs:

    • Multiple scfv clones targeting Ecad were generated using phage display technology, and highly cross-reactive scfv (hmcecad.6) was selected through sequence optimization.
    • A CAR-CD28ζ structure containing hmcecad.6 was designed and stably expressed to enhance the immunosuppressive function of MSCs.

  3. Immunosuppressive Testing of CAR-MSCs:

    • In both in vitro and in vivo experiments, ECCAR-MSCs displayed superior antigen-specific T cell suppression. The transduction efficiency of ECCAR-MSCs was validated in various donors using flow cytometry.

  4. GVHD Model Testing:

    • In a mouse model, ECCAR-MSCs significantly suppressed T cell activity through antigen-specific stimulation, improving GVHD symptoms and survival rates.
    • In vivo optical imaging and immunofluorescence staining confirmed the specific migration of ECCAR-MSCs to Ecad+ colonic tissues in the GVHD mouse model.

  5. Exploration of ECCAR-MSC Immunosuppressive Mechanism:

    • RNA sequencing and proteomics analyses revealed that antigen-specific stimulation of CAR activated immunosuppressive genes and signaling pathways in MSCs, such as IL-6, IL-10, and NF-κB.
    • Further validation demonstrated the roles of these signaling pathways in enhancing immunosuppression.

Experimental Results

  1. T Cell Suppression Effect:

    • ECCAR-MSCs exhibited a dose-dependent T cell suppression effect through antigen-specific stimulation (Ecad+ cells) in in vitro experiments, consistently demonstrated across multiple donors.

  2. In Vivo Tumor Model Experiment:

    • In a mouse tumor model, ECCAR-MSCs significantly inhibited the anti-tumor activity of CD19-CAR T cells, promoting tumor growth and increasing survival rates compared to untreated MSCs.
    • Antigen-specific stimulation of ECCAR-MSCs significantly enhanced their immunosuppressive effect on Ecad+ tumor cells.

  3. Therapeutic Effect in GVHD Model:

    • ECCAR-MSCs significantly reduced weight loss, alleviated GVHD symptoms, and improved survival rates in the GVHD mouse model.
    • ECCAR-MSCs effectively suppressed T cell activity by specifically migrating to Ecad+ tissues.

  4. Activation of Immune Signaling Pathways:

    • RNA sequencing showed significant upregulation of immunosuppressive signaling pathways such as IL-6, IL-10, and NF-κB in ECCAR-MSCs following antigen-specific stimulation.
    • CAR constructs containing CD28ζ signaling domains exhibited stronger immunosuppressive efficacy.

Conclusion and Research Value

  1. Scientific Significance:

    • This study developed a CAR technology to effectively enhance the immunosuppressive function of MSCs, demonstrating potential in GVHD and tumor suppression.
    • ECCAR-MSCs activated immunosuppressive gene pathways through antigen-specific stimulation, improving T cell suppression, providing a scientific basis for further optimization of immune cell therapy.

  2. Application Value:

    • ECCAR-MSCs, as an “off-the-shelf” cell therapy technology, hold potential for application in immune-mediated diseases such as GVHD, Crohn’s disease, and ulcerative colitis.
    • Preliminary verification of their immunosuppressive effect and safety in mouse and canine models lays the foundation for clinical translation.

  3. Research Highlights:

    • This study is the first to demonstrate the potential application of MSCs in enhanced immunosuppression through the design of CAR constructs and optimization of signaling domains.
    • Validation of the safety and efficacy of ECCAR-MSCs in multiple in vitro and in vivo models indicates broad prospects for future clinical applications.

Summary

This study demonstrates the enhancement of mesenchymal stromal cell immunosuppressive function through chimeric antigen receptor technology, proposing a new cell therapy method with potential breakthroughs in the treatment of immune-mediated diseases. The results indicate that ECCAR-MSCs exhibit significant immunosuppressive effects both in vitro and in vivo with good safety, providing a feasible protocol for future clinical applications.