Dataset Information

Functional genomics of glioblastoma: from epigenetics to proteomic investigation of tumor-initiating cell secretome

ABSTRACT: The main objectives of the project are as follows: 1) Deciphering the TIC specific secretome signature. The secretome collected from four different TICs and their corresponding differentiated cell cultures will be subjected to label-free quantitative proteomic analysis to identify a TIC specific secretome signature. 2) Validation and functional characterization of selected molecules from TIC specific secretome signature. Selected molecules based on the level of regulation and literature information will be validated at the level of transcript and protein. The importance of a set of validated proteins will be studied both in vitro using neurosphere cultures and angiogenesis assays, and in vivo using an intracranial glioma model in nude mice 3) Characterizing of GBM specific serum proteome signature. Using label-free quantitative proteomics, we will compare the serum proteome of tumor bearing mice (xenotransplantated with TICs), early after tumor establisment and after the tumor has reached its maximum size.We will also explore by targeted quantitative proteomics whether the level of “specific” TIC-secreted proteins is increased in the serum of TIC initiated GBM bearing mice and verify, using the same strategy, proteins found to be differentially expressed in serum in both mice groups. These studies will reveal whether TIC-secreted proteins (and/or other GBM-derived proteins) can be retrieved in serum and thus be considered as candidate biomarkers of GBM.

INSTRUMENT(S): Q Exactive HF-X

ORGANISM(S): Homo Sapiens (human)

DISEASE(S): Glioblastoma

SUBMITTER: Functional Proteomics Platform FPP

LAB HEAD: Prof. Kumaravel Somasundaram

PROVIDER: PXD032958 | Pride | 2022-06-07

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	Oxidation__M_Sites.txt	Txt
	RefProteome_HUMAN-cano_2018_11.fasta	Fasta
	SPHERE_170414_P2_D5.raw	Raw
	SPHERE_170414_P_A1.raw	Raw
	SPHERE_170414_P_A2.raw	Raw

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Publications

Differentiated glioma cell-derived fibromodulin activates integrin-dependent Notch signaling in endothelial cells to promote tumor angiogenesis and growth.

Sengupta Shreoshi S Mondal Mainak M Prasasvi Kaval Reddy KR Mukherjee Arani A Magod Prerna P Urbach Serge S Friedmann-Morvinski Dinorah D Marin Philippe P Somasundaram Kumaravel K

eLife 20220601

Cancer stem cells (CSCs) alone can initiate and maintain tumors, but the function of non-cancer stem cells (non-CSCs) that form the tumor bulk remains poorly understood. Proteomic analysis showed a higher abundance of the extracellular matrix small leucine-rich proteoglycan fibromodulin (FMOD) in the conditioned medium of differentiated glioma cells (DGCs), the equivalent of glioma non-CSCs, compared to that of glioma stem-like cells (GSCs). DGCs silenced for FMOD fail to cooperate with co-impla ...[more]

PMID: 35642785

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Project description:Glioblastoma (GBM), the most malignant primary brain tumor, is characterized by widespread heterogeneity, leading to poor and unpredictable clinical outcomes. Recent studies have provided evidences that the tumor microenvironment has a critical role in regulating tumor growth by establishing a complex network of interactions with tumor cells. Here we investigate how GBM cells modulate glial cells, and how this modulation can influence back on the malignant phenotype of GBM cells. Primary mouse glial cultures were established and cultured in serum-free media (unprimed) or exposed to secretome derived from GL261 GBM cells (primed). Conditioned media (CM) from each glial culture were collected and a proteomic analysis was conducted. Glial cells CM (unprimed and primed) were also used in GL261 GBM cells to evaluate its impact in critical hallmarks of GBM cells, including viability, migration, and activation of tumor-related intracellular signaling pathways. The proteomic analysis revealed that the pre-exposure of glial cells to CM from GBM cells led to the upregulation of several proteins related to inflammatory response, cell adhesion and extracellular structure organization within the secretome of primed glial cells, consistent with a pattern of reactive astrogliosis. At the functional levels, CM derived from unprimed glial cells favored an increase in cell migration capacity, while CM from primed glial cells was more efficient in promoting GBM cells viability. These effects on GBM cells were accompanied by activation of particular intracellular cancer-related pathways, mainly the MAPK/ERK pathway, which is a known regulator of cell proliferation. Together, our results demonstrate that glial cells can have a different impact on the progression of GBM tumors, suggesting that the secretome of GBM cells is able to modulate the secretome of neighboring glial cells, in a way that regulates the “go-or-grow” phenotypic switch of GBM cells. Together, our results suggest that glial cells can impact on the pathophysiology of GBM tumors, and that the secretome of GBM cells is able to modulate the secretome of neighboring glial cells, in a way that regulates the “go-or-grow” phenotypic switch of GBM cells.

Project description:Glioblastoma (GBM) is the most prevalent primary malignant brain tumor in adults and remains almost invariably lethal due to its aggressive and invasive behavior. Despite many clinical trials have been put forward, the standard treatment that stands for more than a decade consists of surgical resection, followed by temozolomide (TMZ) chemotherapy and radiation treatment. However, this strategy has improved the average survival time to 14.6 months when compared to 12,1 months only with surgery and radiation1 . Therefore, due to the low prognosis, different efforts have been employed in the search for new drugs and therapeutic protocols for this malignancy. A class of drugs that has emerged as a promising candidate for GBM is Histone Deacetylase inhibitors (iHDAC). Indeed, pre-clinical studies have demonstrated the efficiency of different iHDACs as anti-tumor agents, especially when associated with other therapies, including chemotherapy and radiation2. In fact, the use of iHDACs reduced the expression of genes involved in DNA repair, formation of mitotic spindles and homologous chromosome segregation, leading to glioma apoptosis in vitro 3. In addition, it has been reported that small iHDACs molecules, such as Trichostatin (TSA), disrupts cellular signaling networks relevant to tumorigenesis, leading to an increased interest in testing iHDACs in clinical trials against different types of tumors 4 . In agreement, a high activity of HDAC has also been associated with tumor progression, corroborating the pharmacological inhibition of HDACs as an interesting anti-cancer molecular target. Previous works from our group are in line with this reasoning showing that iHDACs negatively regulate the migratory capacity of GBM in vitro, suggesting that these inhibitors may regulate the molecular signature of GBM secretome, making the tumor's microenvironment repulsive for cell migration5. In fact, the communication between tumor cells and their microenvironment may occur directly or indirectly through a variety of secreted molecules, such as growth factors, cytokines, chemokines and microRNAs6. One relevant secreted molecule described by our group is Nodal, an important morphogen involved in several cellular processes related to tumorigenesis. In this previous study, our group brings an original approach on the mechanisms that dynamically control intra and extracellular nodal availability during GBM tumorigenesis7. Thus, the heterotypic secretome that derives from the tumor cells, may be an important source of key regulators of the tumorigenic process6 . To better understand whether iHDACs can also affect the repertoire of secreted molecules, and therefore directly modulate the surrounding micro-environment displaying deleterious effects on tumor cells, we investigated the secretome of iHDAC treated GBM cells in vitro using a high throughput methodology of protein identification and quantification couple to label free mass spectrometry. Using this strategy we were able to successfully identify secreted molecules upon HDAC activity knockdown. Interestingly, we identified proteins exclusive to the iHDAC treated secretome as well as proteins found in both control and treated groups. These proteins were classified using the Matrisome classification system corroborating that the molecular signature of tumor secreted proteins, that belong to the ECM, was disrupted. In vitro experiments using 2D and 3D cell culture systems coupled to decellularized ECM analysis by confocal microscopy confirmed that HDAC activity knockdown gave rise to a disrupted pattern of ECM protein staining. Taken together our results first describe in the literature the relevance of epigenetic remodeling HDAC dependent for the molecular signature of GBM cells shedding light on putative molecular targets to approach the malignancy.