Proteomic analysis of extracellular matrix produced by pericyte-like cells isolated from human glioblastomas

Glioblastoma (also known as glioblastoma multiforme, GBM), is the most common and the most aggressive type of primary brain cancer with a worldwide prevalence between 2 to 5 individuals per 100,000 population. GBM mostly affects people between 45 to 70 years with a median age at diagnosis of 64 years and higher incidence in men. The overall prognosis is very poor as life expectancy for GBM patients is 12-18 months (when treatment is provided) and only four months without treatment. Less than 10% of patients survive more than five years.
GBMs contain significantly higher amounts of extracellular matrix (ECM) in comparison with normal brain tissue. This ECM is enriched in fibrillar proteins such as collagen I and fibronectin and the abundance of these proteins is associated with poor prognosis of GBM patients. The highest levels of fibrillar proteins are found in perivascular niches, resided by fibroblast activation protein (FAP)-expressing pericyte-like cells. We therefore focused our proteomic analysis on the ECM produced by glioma cells and patient-derived FAP+ pericyte-like cells.
Firstly, we employed data-dependent acquisition (DDA) to create sample-specific spectral library of proteins present in decellularized ECM derived from glioma cells, human brain vascular pericytes, and patient-derived FAP+ pericyte-like cells cultivated in vitro. Then, we performed quantitative measurements using data-independent acquisition (DIA) approach, followed by data analysis in Mascot, Skyline, and MSstats. In total, we quantified 3,232 distinct proteins, including about 30% of known human core matrisome proteins. The ECM produced by patient-derived FAP+ pericyte-like cells contained significantly higher amounts of fibrillar proteins, such as collagen I and fibronectin, than ECM from glioma cells. Additionally, the composition of the ECM produced by these FAP+ pericyte-like cells was markedly different from that produced by glioma cells and human brain vascular pericytes, particularly in its higher amount of basement membrane proteins. Follow-up experiments demonstrated that the ECM produced by patient-derived FAP+ pericyte-like cells supports the migration and adhesion of glioma cells in vitro, likely via activation of focal adhesion kinase (FAK) signalling. Therefore, the FAP+ pericyte-like cells seem to be critical players in GBM development and clinical outcome.

This work was supported by the Ministry of Education, Youth and Sports of the Czech Republic projects National Institute for Cancer Research (Programme EXCELES, ID Project No. LX22NPO5102) funded by the European Union—Next Generation EU, EATRIS-CZ (LM2023053), the project Center for Tumor ecology (reg. n. CZ.02.1.01/0.0/0.0/16_019/0000785) supported by the Operational Program Research, Development and Education, Ministry of Health of the Czech Republic (grant NV19-03-00501), and Charles University Cooperatio Program, research area" "Oncology and Haematology" and project GA UK no. 1084120.