Loss of SPINT2 expression frequently occurs in glioma, leading to increased growth and invasion via MMP2
High-grade gliomas (HGG) remain one of the most aggressive tumors, which is primarily due to its diffuse infiltrative nature. Serine proteases and metalloproteases are known to play key roles in cellular migration and invasion mechanisms. SPINT2, also known as HAI-2, is an important serine protease inhibitor that can affect MET signaling. SPINT2 has been found to be frequently downregulated in various tumors, whereby hypermethylation of its promoter appears to serve as a common mechanism. Here, we assessed the clinical relevance of SPINT2 expression and promoter hypermethylation in pediatric and adult HGG and explored its functional role.
A series of 371 adult and 77 pediatric primary HGG samples was assessed for SPINT2 protein expression (immunohistochemistry) and promoter methylation (methylation-specific PCR) patterns. After SPINT2 knockdown and knock-in in adult and pediatric HGG cell lines, a variety of in vitro assays was carried out to determine the role of SPINT2 in glioma cell viability and invasion, as well as their mechanistic associations with metalloprotease activities.
We found that SPINT2 protein expression was frequently absent in adult (85.3%) and pediatric (100%) HGG samples. The SPINT2 gene promoter was found to be hypermethylated in approximately half of both adult and pediatric gliomas. Through functional assays we revealed a suppressor activity of SPINT2 in glioma cell proliferation and viability, as well as in their migration and invasion. These functions appear to be mediated in part by MMP2 expression and activity.
We conclude that dysregulation of SPINT2 is a common event in both pediatric and adult HGG, in which SPINT2 may act as a tumor suppressor.
KeywordsGlioma SPINT2 Hypermethylation Metalloproteases
The authors thank Dr. Scholm for providing the 4/T0 vector.
This work was funded by the Fundação para a Ciência e Tecnologia (FCT), Portugal (www.fct.pt) (grants PTDC/SAU-ONC/115513/2009_FCOMP-01-0124-FEDER-015949 and PTDC/SAU-TOX/114549/2009 to RMR). This article has been developed under the scope of project NORTE-01-0145-FEDER-000013, supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER). This work has been funded by FEDER funds, through the Competitiveness Factors Operational Programme (COMPETE), and by National funds, through FCT, under the scope of project UID/Multi/50026. MVP was recipient of a FCT Post-Doctoral fellowship (SFRH/BPD/104290/2014). SPC was recipient of a BOLSA LIGA PORTUGUESA CONTRA CANCRO (LPCC)/Fundação PT 2018 grant. AMC was recipient of fellow DL 57/2016/CP1360/CT0009. FP was recipient of a FCT Post-Doctoral fellowship (SFRH/BPD/115730/2016); CJ acknowledges NHS funding to the NIHR Biomedical Research Centre at The Royal Marsden and the ICR.
Compliance with ethical standards
Conflict of interests
The authors disclose no potential conflicts of interest.
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