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Decoding of the surfaceome and endocytome in primary glioblastoma cells identifies potential target antigens in the hypoxic tumor niche.

ABSTRACT: Immunotherapies with antibody-drug-conjugates (ADC) and CAR-T cells, targeted at tumor surface antigens (surfaceome), currently revolutionize clinical oncology. However, target identification warrants a better understanding of the surfaceome and how it is modulated by the tumor microenvironment. Here, we decode the surfaceome and endocytome and its remodeling by hypoxic stress in glioblastoma (GBM), the most common and aggressive brain tumor in adults. We employed a comprehensive approach for global and dynamic profiling of the surfaceome and endocytosed (endocytome) proteins and their regulation by hypoxia in patient-derived GBM cultures. We found a heterogeneous surface-endocytome profile and a divergent response to hypoxia across GBM cultures. We provide a quantitative ranking of more than 600 surface resident and endocytosed proteins, and their regulation by hypoxia, serving as a resource to the cancer research community. As proof-of-concept, the established target antigen CD44 was identified as a commonly and abundantly expressed surface protein with high endocytic activity. Among hypoxia induced proteins, we reveal CXADR, CD47, CD81, BSG, and FXYD6 as potential targets of the stressed GBM niche. We could validate these findings by immunofluorescence analyses in patient tumors and by increased expression in the hypoxic core of GBM spheroids. Selected candidates were finally confronted by treatment studies, showing their high capacity for internalization and ADC delivery. Importantly, we highlight the limited correlation between transcriptomics and proteomics, emphasizing the critical role of membrane protein enrichment strategies and quantitative mass spectrometry. Our findings provide a comprehensive understanding of the surface-endocytome and its remodeling by hypoxia in GBM as a resource for exploration of targets for immunotherapeutic approaches in GBM.

SUBMITTER: de Oliveira KG

PROVIDER: S-EPMC10898066 | biostudies-literature | 2024 Feb

REPOSITORIES: biostudies-literature

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Decoding of the surfaceome and endocytome in primary glioblastoma cells identifies potential target antigens in the hypoxic tumor niche.

de Oliveira Kelin Gonçalves KG Bång-Rudenstam Anna A Beyer Sarah S Boukredine Axel A Talbot Hugo H Governa Valeria V Johansson Maria C MC Månsson Ann-Sofie AS Forsberg-Nilsson Karin K Bengzon Johan J Malmström Johan J Welinder Charlotte C Belting Mattias M

Acta neuropathologica communications 20240227 1

Immunotherapies with antibody-drug-conjugates (ADC) and CAR-T cells, targeted at tumor surface antigens (surfaceome), currently revolutionize clinical oncology. However, target identification warrants a better understanding of the surfaceome and how it is modulated by the tumor microenvironment. Here, we decode the surfaceome and endocytome and its remodeling by hypoxic stress in glioblastoma (GBM), the most common and aggressive brain tumor in adults. We employed a comprehensive approach for gl ...[more]

PMID: 38414005

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Project description:BackgroundGlioblastoma multiforme (GBM) is a highly lethal, stage IV brain tumour with a prevalence of approximately 2 per 10,000 people globally. The cell surface proteins or surfaceome serve as information gateway in many oncogenic signalling pathways and are important in modulating cancer phenotypes. Dysregulation in surfaceome expression and activity have been shown to promote tumorigenesis. The expression of GBM surfaceome is a case in point; OMICS screening in a cell-based system identified that this sub-proteome is largely perturbed in GBM. Additionally, since these cell surface proteins have 'direct' access to drugs, they are appealing targets for cancer therapy. However, a comprehensive GBM surfaceome landscape has not been fully defined yet. Thus, this study aimed to define GBM-associated surfaceome genes and identify key cell-surface genes that could potentially be developed as novel GBM biomarkers for therapeutic purposes.MethodsWe integrated the RNA-Seq data from TCGA GBM (n = 166) and GTEx normal brain cortex (n = 408) databases to identify the significantly dysregulated surfaceome in GBM. This was followed by an integrative analysis that combines transcriptomics, proteomics and protein-protein interaction network data to prioritize the high-confidence GBM surfaceome signature.ResultsOf the 2381 significantly dysregulated genes in GBM, 395 genes were classified as surfaceome. Via the integrative analysis, we identified 6 high-confidence GBM molecular signature, HLA-DRA, CD44, SLC1A5, EGFR, ITGB2, PTPRJ, which were significantly upregulated in GBM. The expression of these genes was validated in an independent transcriptomics database, which confirmed their upregulated expression in GBM. Importantly, high expression of CD44, PTPRJ and HLA-DRA is significantly associated with poor disease-free survival. Last, using the Drugbank database, we identified several clinically-approved drugs targeting the GBM molecular signature suggesting potential drug repurposing.ConclusionsIn summary, we identified and highlighted the key GBM surface-enriched repertoires that could be biologically relevant in supporting GBM pathogenesis. These genes could be further interrogated experimentally in future studies that could lead to efficient diagnostic/prognostic markers or potential treatment options for GBM.

Project description:High-density lipoprotein (HDL) is a mixture of complex particles mediating reverse cholesterol transport (RCT) and several cytoprotective activities. Despite its relevance for human health, many aspects of HDL-mediated lipid trafficking and cellular signaling remain elusive at the molecular level. During HDL's journey throughout the body, its functions are mediated through interactions with cell surface receptors on different cell types. To characterize and better understand the functional interplay between HDL particles and tissue, we analyzed the surfaceome-residing receptor neighborhoods with which HDL potentially interacts. We applied a combination of chemoproteomic technologies including automated cell surface capturing (auto-CSC) and HATRIC-based ligand-receptor capturing (HATRIC-LRC) on four different cellular model systems mimicking tissues relevant for RCT. The surfaceome analysis of EA.hy926, HEPG2, foam cells, and human aortic endothelial cells (HAECs) revealed the main currently known HDL receptor scavenger receptor B1 (SCRB1), as well as 155 shared cell surface receptors representing potential HDL interaction candidates. Since vascular endothelial growth factor A (VEGF-A) was recently found as a regulatory factor of transendothelial transport of HDL, we next analyzed the VEGF-modulated surfaceome of HAEC using the auto-CSC technology. VEGF-A treatment led to the remodeling of the surfaceome of HAEC cells, including the previously reported higher surfaceome abundance of SCRB1. In total, 165 additional receptors were found on HAEC upon VEGF-A treatment representing SCRB1 co-regulated receptors potentially involved in HDL function. Using the HATRIC-LRC technology on human endothelial cells, we specifically aimed for the identification of other bona fide (co-)receptors of HDL beyond SCRB1. HATRIC-LRC enabled, next to SCRB1, the identification of the receptor tyrosine-protein kinase Mer (MERTK). Through RNA interference, we revealed its contribution to endothelial HDL binding and uptake. Furthermore, subsequent proximity ligation assays (PLAs) demonstrated the spatial vicinity of MERTK and SCRB1 on the endothelial cell surface. The data shown provide direct evidence for a complex and dynamic HDL receptome and that receptor nanoscale organization may influence binding and uptake of HDL.

Dataset Information

Decoding of the surfaceome and endocytome in primary glioblastoma cells identifies potential target antigens in the hypoxic tumor niche.

Publications

Decoding of the surfaceome and endocytome in primary glioblastoma cells identifies potential target antigens in the hypoxic tumor niche.

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