Expression of S1P metabolizing enzymes and receptors correlate with survival time and regulate cell migration in glioblastoma multiforme

Abstract

A signaling molecule which is involved in proliferation and migration of malignant cells is the lipid mediator sphingosine-1-phosphate (S1P). There are hints for a potential role of S1P signaling in malignant brain tumors such as glioblastoma multiforme (GBM) which is characterized by a poor prognosis. Therefore, a comprehensive expression analysis of S1P receptors (S1P1-S1P5) and S1P metabolizing enzymes in human GBM (n = 117) compared to healthy brain (n = 10) was performed to evaluate their role for patient's survival. Furthermore, influence of S1P receptor inhibition on proliferation and migration were studied in LN18 GBM cells. Compared to control brain, mRNA levels of S1P1, S1P2, S1P3 and S1P generating sphingosine kinase-1 were elevated in GBM. Kaplan-Meier analyses demonstrated an association between S1P1 and S1P2 with patient's survival times. In vitro, an inhibitory effect of the SphK inhibitor SKI-II on viability of LN18 cells was shown. S1P itself had no effect on viability but stimulated LN18 migration which was blocked by inhibition of S1P1 and S1P2. The participation of S1P1 and S1P2 in LN18 migration was further supported by siRNA-mediated silencing of these receptors. Immunoblots and inhibition experiments suggest an involvement of the PI3-kinase/AKT1 pathway in the chemotactic effect of S1P in LN18 cells.In summary, our data argue for a role of S1P signaling in proliferation and migration of GBM cells. Individual components of the S1P pathway represent prognostic factors for patients with GBM. Perspectively, a selective modulation of S1P receptor subtypes could represent a therapeutic approach for GBM patients and requires further evaluation.

Sphingosine-1-Phosphate in the Tumor Microenvironment: A Signaling Hub Regulating Cancer Hallmarks

“…The proliferative role of extracellular S1P in GBM is further substantiated by several reports demonstrating that human GBM cells respond mitogenically to nanomolar concentrations of S1P [57,96,121,182], and that extracellular S1P antagonized the inhibition of cell proliferation induced by inhibition of the S1P transporter ABCA1 [121]. The proliferating effect of S1P was found to involve its binding to, and activation of, different S1P receptors, including S1P1-3 and S1P5, which are expressed in different GBM cell lines and human GBM specimens [54,57,58,96,171,182]. The S1P binding to all these receptors is able to affect GBM proliferation due to the ability of these receptors to activate G i (all S1P receptors) and/or G 12/13 (S1P2, S1P3 and S1P5) [121].…”

“…In particular, several studies reported that human GBM displays a high expression level of SphK1, and SphK1 mRNA positively correlates with S1P level [50,[54][55][56][57]. High SphK1 expression has been found to be correlated with a significant poor prognosis in patients with GBM by some studies [54,55], but such a relation was not detected in more recent reports [50,57,58]. Despite these contradictory findings in tumor heterogeneity, the mechanisms underlying SphK1 gene overexpression in GBM remain unclarified.…”

“…It was shown that different human GBM cell lines and GBM specimens express the genes of S1P1-3 and S1P5 receptors. Increased levels of S1P1, S1P2, and S1P3 were observed in GBM tissue specimens, and signaling of S1P1 and S1P2 were markedly correlated with patient survival rates [57]. Despite Yoshida et al [58,171] reporting that S1P1 expression is downregulated in GBM specimens, which enhances tumor cell proliferation and correlates with shorter survival of GBM patients, recent investigations demonstrated an inverse correlation between S1P1 expression and GBM patient survival [50,56,57].…”

“…In particular, S1P emerged as a key signal in the regulation of GBM invasion, being able to stimulate different events involved in this complex process, including reduction of cells and of extracellular matrix (ECM) adhesion, cytoskeletal remodeling, matrix metalloprotease (MMP) secretion, and then ECM degradation ( Figure 5). Overall, G protein signaling of the S1P1-3 subtypes results in: (1) Rac activation [198], which regulates cell morphology and actin dynamics, and stimulates cell squeezing through the narrow extracellular spaces that are typical of the brain parenchyma; (2) increased secretion of the matricellular protein CCN1/Cyr61 that, once secreted, binds to integrin αV-β3 to enhance ECM adhesion [185,200,201]; (3) activation of the signaling cascades MEK1/2, PI3-kinase/AKT1 and Rhokinase, which leads to enhanced gene and protein expression of the plasminogen activator system proteins, including the urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor 1 (PAI-1), and a receptor for uPA (uPAR) [57,183,202].…”

“…(3) activation of the signaling cascades MEK1/2, PI3-kinase/AKT1 and Rho-kinase, which leads to enhanced gene and protein expression of the plasminogen activator system proteins, including the urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor 1 (PAI-1), and a receptor for uPA (uPAR) [57,183,202].…”