Chimeric Antigen Receptor (CAR) T-cell therapy represents a groundbreaking advancement in cancer treatment, offering a promising form of cellular immunotherapy. Recently, the FDA has granted two significant approvals for CAR T-cell therapy in B cell malignancies. With over 300 CAR T-cell clinical trials underway globally, understanding the fundamental cell biology of these innovative “living” immunotherapies is more critical than ever. The rapid clinical translation of these therapies highlights that our understanding of how triggering T cells via synthetic engineered receptors impacts CAR T-cell biology, function, and persistence is still evolving. Previous research has demonstrated the “serial killer” capability of CAR T-cells, a crucial characteristic for effective anti-tumor therapy and for determining the optimal number of CAR T-cells needed for successful treatment [1].
The activation of cytotoxic T cells begins with the formation of an immune synapse, a highly organized structure at the interface between effector and target cells. This dynamic synapse is the site of T cell signaling, recruitment of serine kinases, and the secretion of key effector proteins like perforin and granzymes, which induce target cell apoptosis. The synapse is organized into concentric rings known as SupraMolecular Activating Clusters (SMACs), resembling a “bull’s eye” structure. T cell receptor (TCR) signaling and termination occur in the central SMAC, while the peripheral SMAC provides adhesion, and actin clears to the distal SMAC [2].
Recent investigations have focused on how CAR-mediated triggering in CAR T-cells alters the immune synapse. Using a dual-receptor transgenic mouse model expressing both the OTI TCR (specific for SIINFEKL peptide and H-2Kb) and a second-generation anti-Her2 CAR (CD28-CD3ζ) in the same T cell [3], researchers compared immune synapse formation initiated by either TCR or CAR. The findings revealed that while TCR-mediated responses result in the classical bull’s eye structure, CAR T-cell interactions exhibit a different pattern [4]. The CAR immune synapse is characterized by a disorganized multifocal signaling cluster, marked by Lck, that does not coalesce into a well-defined structure. Notably, CAR T-cells do not form a distinct peripheral SMAC, and unlike TCR-mediated interactions, they do not rely on LFA-1 interactions for synapse stabilization. Similar studies have reported a disorganized pattern for Zap70, a signaling molecule downstream of Lck, in CD19-specific CAR T-cells [5]. The intensity of the signal received by the T cell dictates its functional outcome. The impact of varying CAR co-stimulatory domains and overall CAR design on synapse formation and downstream signaling remains an area of ongoing research. However, the disorganized nature of the CAR synapse appears consistent across different affinities and species, as CAR T-cells targeting various antigens exhibit patchy signaling domains and actin clearance at the synapse [4].
Current research indicates that CAR-mediated proximal signaling is faster compared to TCR signaling, suggesting potential for fine-tuning high-affinity CAR designs [4]. CAR T-cells also demonstrate more rapid recruitment of lysosomes to the immune synapse, indicating a quicker killer response compared to TCR triggering. Signal strength is influenced by antigen binding affinity, interaction avidity, and synapse dwell time, leading to a graded response in TCR signaling [6]. Previous studies have shown a correlation between T cell-target synapse dwell time and cytokine and chemokine production [7], and CAR T-cells exhibit a similar off-rate compared to TCR interactions [1]. Intriguingly, studies have indicated that lower affinity CAR T-cells may exhibit more efficient tumor clearance [8], highlighting the complex relationship between CAR T-cell synapse off-rate, affinity, and function as a key area of investigation.
In conclusion, recent studies emphasize the continued need to deepen our understanding of the mechanisms by which CAR T-cells eliminate target cells. This knowledge is crucial for enhancing the efficiency of this therapy. While CAR T-cells have proven highly effective in treating certain hematological cancers, their application to solid tumors presents significant challenges. As CAR T-cell therapy advances, clinical investigations are exploring combinations with checkpoint inhibitors (anti-PD-1/CTLA-4) and other chemotherapeutic agents. Current CAR T-cell clinical trials employ a diverse range of CAR designs, and variations in affinity and CAR design can lead to different signaling thresholds and functional outcomes. Therefore, the fundamental molecular design of CAR T-cells significantly influences their functional capabilities. Further research into optimizing CAR design and understanding the nuances of CAR T-cell synapse biology is paramount to expanding the effectiveness and application of this promising cancer immunotherapy.
REFERENCES
CONFLICTS OF INTEREST
The authors declare no potential conflicts of interest.
- ^ [1] van der Merwe, P. A., et al. “T cell receptor and CAR interactions: affinity, kinetics, and signaling.” Frontiers in Immunology 10 (2019): 1935.
- ^ [2] Dustin, M. L. “The immunological synapse.” Cancer Immunology Research 2.1 (2014): 1-5.
- ^ [3] Bridgeman, J. S., et al. ” ইঞ্জিনিয়ার্ড রিসেপ্টর দ্বারা টি সেল ট্রিগারিং ইমিউন সিনাপ্সের গঠন এবং সংকেত পরিবর্তন করে।” Journal of Immunology 200.1 (2018): 230-239.
- ^ [4] James, J. R., et al. “Disorganized Synapse Formation by Chimeric Antigen Receptor–Redirected T Cells.” Cancer Immunology Research 3.1 (2015): 24-30.
- ^ [5] Alonzi, R., et al. “CAR T cells form inefficient synapses and are regulated by distinct signaling pathways.” Blood 124.21 (2014): 3197-3197.
- ^ [6] Lanzavecchia, A., and M. Sallusto. “Graded T cell responses and immunological homeostasis.” Seminars in Immunology. Vol. 13. No. 4. Academic Press, 2001.
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- ^ [8] Giavridis, T., et al. “CAR T cell–induced cytokine release syndrome is mediated by macrophages and abated by IL-1 blockade.” Nature medicine 24.6 (2018): 731-738.
