Proteomic profiling of the neuroblastoma secretome identifies extracellular vesicles as drivers of T cell suppression

Strijker, J. G. M.; Van Berkum, R. E.; Kalaitsidou, E.; Boltjes, A.; Hiddink Verberne, N.; Damen, M. A.; Van den Ham, F.; Baaij, L. E.; Anderson, J.; Molenaar, J. J.; Wu, W.; Wienke, J.

doi:10.64898/2025.12.02.690421

bioRxiv · 10.64898/2025.12.02.690421

Proteomic profiling of the neuroblastoma secretome identifies extracellular vesicles as drivers of T cell suppression

Strijker, J. G. M.·Van Berkum, R. E.·Kalaitsidou, E.·Boltjes, A.·Hiddink Verberne, N.·Damen, M. A.·Van den Ham, F.·Baaij, L. E.·Anderson, J.·Molenaar, J. J.·Wu, W.·Wienke, J.

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Abstract

Immunosuppressive tumor microenvironments in solid tumors hamper the efficacy of immunotherapies, including CAR T-cell therapy. To identify immunosuppressive secreted factors in high-risk neuroblastoma, a childhood solid tumor with a 5-year overall survival of below 60%, we mapped secreted factors of 16 genetically diverse patient-derived neuroblastoma tumoroids by LC-MS. The secretomes suppressed T cell activation, proliferation and cytotoxicity to varying extents, suggesting that this panel constituted a valuable discovery foundation to identify clinically relevant immunosuppressive factors. 29 proteins were significantly enriched in highly suppressive secretomes, of which Neuropeptide Y (NPY) had the highest correlation with functionally-determined immunosuppression. No direct or indirect effect of NPY on (CAR) T cell activation could be validated. Instead, NPY proved to be a biomarker for extracellular vesicle (EV) secretion. Finally, we demonstrated that neuroblastoma-derived EVs potently suppress T cell activation. Altogether, this study provides an atlas of the neuroblastoma secretome and reveals immunosuppressive effects of EVs on T cells. These insights contribute to the understanding of the immunosuppressive neuroblastoma TME.

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Strijker, J. G. M., Van Berkum, R. E., Kalaitsidou, E., Boltjes, A., Hiddink Verberne, N., Damen, M. A., Van den Ham, F., Baaij, L. E., Anderson, J., Molenaar, J. J., Wu, W., Wienke, J.. 2025-12-03. Proteomic profiling of the neuroblastoma secretome identifies extracellular vesicles as drivers of T cell suppression. https://doi.org/10.64898/2025.12.02.690421

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