Abstract P093: Proteomic analysis reveals a mechanism of resistance to radiation mediated by microvesicles in glioma

Abstract

Abstract Glioma accounts for 80% of malignant brain tumors. Standard treatment for glioma involves surgical resection followed by radiation therapy and chemotherapy, however, tumors recur leading to death in 12 to 18 months on average. Gliomas contain tumor-initiating cells (glioma stem cells, GSCs) that contribute causally to resistance to radiation therapy. GSCs shed large amounts of extracellular vesicles (EVs), which can potentiate growth, therapy resistance and invasiveness of less-aggressive cells present within the tumor microenvironment. However, how EVs mediate these effects is still not well-understood. To gain insights into mechanisms of resistance to radiation, we profiled the proteome of patient-derived GSCs that were either left untreated or treated with a standard therapeutic dose of ionizing radiation (IR). We identify a subset of patient-derived GSC lines that are resistant to radiation. Particularly, we find the cell line GSC-267 displaying a pronounced vesicular transport function. Microvesicles (MVs) but not exosomes shed by GSC-267 can strongly potentiate the proliferation of recipient cells. Subsequent proteomic analysis of EVs highlights elevated levels of nicotinamide phosphoribosyltransferase (NAMPT) within the MVs shed by GSC-267 as well as in the cells of origin. NAMPT is a central enzyme in NAD+ metabolism, is overexpressed in a subset of glioma patients, and correlates with poor patient survival. We find that the NAMPT protein is transferred to recipient cells via MVs derived from GSC-267. Furthermore, presence of NAMPT within MVs, and not solely overxpression within cells, is required for MVs to be able to rescue the proliferation of fibroblasts treated with a radio-mimetic compound. NAMPT inhibition ablates the proliferative gain induced by MVs derived from GSC-267. Supplementation with nicotinamide mononucleotide, the enzymatic product of NAMPT, does not enhance the effect of MVs derived from GSC-267, indicating a NAD+-dependent mechanism. MVs isolated from NAMPT knock-down GSC-267, which no longer carry the NAMPT protein, are not able to rescue the proliferation of recipient cells that were treated with IR. Finally, to more closely model the tumor microenvironment, we employ radiation-sensitive GSC-1079 as recipient cells. Transfer of MVs derived from GSC-267 is able to rescue the viability of GSC-1079 treated with a therapeutic dose of IR. In summary, MVs derived from the radiation-resistant GSC-267 are able to spread aggressive traits in the tumor microenvironment determining an overall shift towards a resistant phenotype. We identify the transfer of the enzyme NAMPT via MVs to be a causative mediator of resistance to radiation. Specific therapeutics targeting NAMPT have been tested in the context of other types of cancer but have shown a poor safety profile. Further analyses may indicate molecular mediators of the ability of NAMPT to confer resistance to radiation, which may lead to identifying novel targets for the treatment of glioma. Citation Format: Elena Panizza, Brandon D. Regalado, Fangyu Wang, Robert J. Munroe, Nathaniel M. Vacanti, Marc A. Antonyak, Richard A. Cerione. Proteomic analysis reveals a mechanism of resistance to radiation mediated by microvesicles in glioma [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P093.