Dataset Information

A peptidomimetic proprotein convertase inhibitor is a potential anti-glioma drug as revealed by biological assays and proteomic studies

ABSTRACT: Proprotein convertases (PCs) are proteases which have an important regulatory function in a wide variety of biological processes. However, these enzymes have a key role in the activation of many cancer-related proteins and promotes the malignant phenotype of cancer cells. Here, with proteomics and biological tests, we investigated the consequences of the use of a peptidomimetic PCs inhibitor on glioma cells and macrophages in the same time and its potential application to cancer therapy. Thus, we demonstrated that the PCs inhibitor acts on macrophage activation, characterized by the secretion of pro-inflammatory and anti-tumoral factor. Also, the PCs inhibitor triggers a strong decrease of the proliferation and invasion of glioma cells, even when they are associated with macrophages in spheroids. In fact, the PCs inhibitor induces great changes in the intracellular and secreted protein profiles of the glioma cells. Many of cancerous processes are affected, such as cell growth, proliferation, angiogenesis and the immunosuppressive capacity of cancer cells. Taken together, these data, establish that the inhibitor acts both on cancer cells and macrophages and can be used as a potential anti-cancer therapeutic strategy.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Rattus Norvegicus (rat)

TISSUE(S): Cell Culture, Macrophage

DISEASE(S): Glioblastoma

SUBMITTER: Soulaimane Aboulouard

LAB HEAD: Michel Salzet

PROVIDER: PXD014679 | Pride | 2020-05-27

REPOSITORIES: Pride

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Publications

The Role of a Proprotein Convertase Inhibitor in Reactivation of Tumor-Associated Macrophages and Inhibition of Glioma Growth.

Rose Mélanie M Duhamel Marie M Aboulouard Soulaimane S Kobeissy Firas F Le Rhun Emilie E Desmons Annie A Tierny Dominique D Fournier Isabelle I Rodet Franck F Salzet Michel M

Molecular therapy oncolytics 20200331

Tumors are characterized by the presence of malignant and non-malignant cells, such as immune cells including macrophages, which are preponderant. Macrophages impact the efficacy of chemotherapy and may lead to drug resistance. In this context and based on our previous work, we investigated the ability to reactivate macrophages by using a proprotein convertases inhibitor. Proprotein convertases process immature proteins into functional proteins, with several of them having a role in immune cell ...[more]

PMID: 32300641

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Project description:Glioblastoma multiforme (GBM) is a highly aggressive and heterogeneous form of primary brain tumors, driven by a complex repertoire of oncogenic alterations, including the constitutively active epidermal growth factor receptor (EGFRvIII). EGFRvIII impacts both cell-intrinsic and non-cell autonomous aspects of GBM progression, including cell invasion, angiogenesis and modulation of the tumor microenvoiront, this is, at least in part, attributable to the release and intercellular trafficking of extracellular vesicles (EVs), heterogeneous membrane structures containing multiple bioactive macromolecules. Here we analyzed the impact of EGFRvIII on the profile of glioma EVs using a series of isogenic tumor cell lines, in which this oncogene exhibits a strong transforming activity. Thus, we observed that EGFRvIII expression alters several properties of glioma EVs, including their output and global protein composition. Using mass spectrometry, quantitative proteomic analysis and Gene Ontology terms filters, we observed that EVs released by EGFRVIII-transformed cells were enriched for extracellular exosome and focal adhesion related proteins. Among them, we validated the association of pro-invasive proteins (CD44, BSG, CD151) with EVs of EGFRvIII expressing cells, and down-regulation of exosomal markers (CD81 and CD82) in EVs of their indolent counterparts. Nano-flow cytometry revealed that the EV output from individual glioma cell lines was highly heterogeneous, such that only a fraction of EVs contained specific proteins such as EGFR, CD9, or others. Notably, cells expressing EGFRvIII released ample EVs double positive for CD44/BSG, and these proteins also co-localized in cellular filopodia. We also detected the expression of homophilic adhesion molecules and increased homologous EV uptake by EGFRvIII-positive glioma cells. These results suggest that oncogenic EGFRvIII reprograms the proteome of GBM-related EVs, a notion with considerable implications for their biological activity and properties relevant for development of EV-based cancer biomarkers.

Project description:Diffuse midline glioma (DMG) is an aggressive pediatric tumor localized in the brainstem. The median survival after diagnosis is less than one year. Surgical resection is not possible due to the critical localization of the tumor and radiation therapy alone or in combination with chemotherapy show only minimal benefit. In 80% of cases, DIPGs are harbor genetic abnormalities such as a K27M mutation in histone H3, suggesting drugs targeting epigenetic anomalies might be efficient. Indeed, Grasso et al. (Nature Medicine, 2015) have shown that the histone deacetylase inhibitor panobinostat (PS) affects glioma cell growth in vitro and in vivo. PS is currently undergoing phase 1 clinical trials. To better understand the mechanism of action of PS on tumor cells, we performed a proteomic analysis by comparing DMG cells treated with panobinostat to control cells. We identified 2 proteins in PS-treated cells: EBP50 (or NHERF-1) and IRSp53 (or BAIAP2). EBP50 protein is implicated in invasion (Kislin et al. Neoplasia, 2009) but it can also act like a tumor suppressor on glioblastoma (Molina et al. Cancer Research, 2010). There is a dual role of the protein which depends on its localization in the cell: when localized at the cell membrane, EBP50 acts as a tumor suppressor, but has oncogenic properties when present in the cytoplasm or nucleus. Using electron microscopy combined with immunofluorescence, we could validate expression of EBP50 in all cellular compartments. IRSp53 is implicated in proliferation of tumor cells (Liu P.S. Oncogene, 2010) and may have an important role during migration and invasion (Funato et al., Cancer Research 2004). To elucidate the role of the two proteins in DIPG, we investigated effects of gene knockdown by siRNA and stable overexpression using lentiviruses. For each protein, we found a significant diminution of proliferation, migration and invasion and an increase in apoptosis cells were transfected with siRNAs. In contrast, overexpression of EBP50 did not have any effect on cells. Our results suggest that the two proteins induced by PS treatment are implicated in tumor progression and might constitute novel additional therapeutic targets, especially as adjuvant treatment together with PS.

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Dataset Information

A peptidomimetic proprotein convertase inhibitor is a potential anti-glioma drug as revealed by biological assays and proteomic studies

Publications

The Role of a Proprotein Convertase Inhibitor in Reactivation of Tumor-Associated Macrophages and Inhibition of Glioma Growth.

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