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Published on: September 28, 2018
PD-1 suppresses CAR signaling by forming the inhibitory signalosome colocalizing to CAR microclusters
Yosuke Yoshida1,2, Hiroaki Machiyama3, Ei Wakamatsu1
1Department of Immunology, Tokyo Medical University, Tokyo, Japan.
Abstract:
The combination of chimeric antigen receptor (CAR)-T cell therapy with immune checkpoint blockade (ICB) using anti-PD-1 has been demonstrated to enhance antitumor CAR-T cell responses. Although, in conventional T cells, PD-1 suppresses T cell activation by binding to PD-L1 with recruitment of a phosphatase SHP2, the precise mechanisms underlying this process in CAR-T cells remain unclear. Here we show that, using real-time single-cell imaging, CD19-CAR aggregates to form "CAR microclusters," which transduce activation signals in coordination with key downstream molecules involved in TCR signaling. Concurrently, PD-1 forms inhibitory signalosomes, recruiting SHP2 into CAR microclusters by PD-L1 binding, leading to diminished CD3ζ phosphorylation, as well as reduced in cytokine production, antigen-specific cytotoxicity, and antitumor CAR-T cell effects in vivo. Importantly, anti-PD-1 treatment impairs PD-1 microcluster formation and restores CAR-T cell activities. Thus, these findings suggest that PD-1 microcluster formation can serve as a trigger for immune tolerance in CAR-T cells.
Insights
Chimeric antigen receptor (CAR)-T cell therapy combined with anti-PD-1 enhances anti-tumor responses. PD-1 forms microclusters in CAR-T cells, inhibiting their function, but anti-PD-1 therapy restores CAR-T cell activity.
Area of Science:
- Immunology
- Cell Biology
- Cancer Therapy
Background:
- Chimeric antigen receptor (CAR)-T cell therapy shows promise in cancer treatment.
- Immune checkpoint blockade (ICB) with anti-PD-1 enhances CAR-T cell responses.
- Mechanisms of PD-1 inhibition in CAR-T cells are not fully understood.
Purpose of the Study:
- To elucidate the molecular mechanisms by which PD-1 signaling inhibits CAR-T cell function.
- To investigate the role of PD-1 microcluster formation in CAR-T cell immune tolerance.
Main Methods:
- Real-time single-cell imaging to observe CAR and PD-1 dynamics.
- Analysis of downstream signaling molecules, including CD3ζ phosphorylation.
- Assessment of cytokine production and antigen-specific cytotoxicity.
- In vivo studies to evaluate anti-tumor effects.
Main Results:
- CD19-CAR forms "CAR microclusters" that transduce activation signals.
- PD-1 forms inhibitory signalosomes by binding PD-L1, recruiting SHP2 into CAR microclusters.
- PD-1 recruitment of SHP2 diminishes CD3ζ phosphorylation, cytokine production, and cytotoxicity.
- Anti-PD-1 treatment disrupts PD-1 microcluster formation, restoring CAR-T cell activity.
Conclusions:
- PD-1 microcluster formation is a key mechanism of immune tolerance in CAR-T cells.
- Targeting PD-1 microclusters can overcome immune suppression and enhance CAR-T cell therapy efficacy.
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