Glioblastoma (GBM) is the most common malignant brain tumor of adults, with a median survival of less than 1 year (1
). The disease is characterized by invasion of the tumor into the adjacent brain parenchyma and by the abnormal activation of receptor tyrosine kinase (RTK) signaling pathways.
However, despite the testing of a number of chemotherapeutic modalities targeting known GBM signaling pathways, only limited clinical success has been achieved. One explanation for the limited efficacy of targeted therapeutics may be that GBM is driven by the summation of multiple signaling inputs (2
). Thus, effective therapeutic strategies may require a more comprehensive understanding of tumor signaling, including modulation by its microenvironment (3
), a known regulator of lethal characteristics of other cancers (4
). The identification of distinct GBM subtypes, based on expression and genomic and proteomic data (3
), supports the notion that GBM is a heterogeneous disease with different patterns of abnormal signaling.
RTK signaling pathways regulate many aspects of tumorigenesis, including cell growth and proliferation. In GBM, abnormal activation of these pathways can be driven by altered ligand availability and altered receptor levels. Indeed, the second most commonly amplified gene in GBM is PDGFR
). Overexpression of its ligand, PDGF-B, mediates the oncogenic influence of TGF-β in human GBM (12
) and can drive tumorigenesis in murine models for glioma (13
). Once released from the cell, growth factors can be sequestered by the extracellular microenvironment. Mechanisms regulating their postsynthetic availability are just beginning to be elucidated, but include enzymatic release from the extracellular matrix (ECM), thus allowing the growth factors to be available for signaling or alternatively to diffuse away. Because GBMs diffusely invade into the surrounding brain parenchyma, ligand availability in the tumor microenvironment, an underappreciated factor, may be a critical determinant of RTK signaling pathway activity and tumor growth.
Heparan sulfate proteoglycans (HSPGs) are ubiquitously produced by most animal cells and are a major component of the extracellular environment in normal brain and GBM (16
). Present on the cell surface and in the ECM, HSPGs play a key role in a number of biological processes based on their ability to bind and regulate the activity of diverse molecules including chemokines and growth factors, such as PDGF, VEGF, and FGF, in many tissues, including the brain (18
). Indeed, HSPG binding can sequester ligands and decrease signaling, such as with the Wnts, or it can act as a coreceptor and actually promote receptor signaling, such as with FGF2 and VEGF (23
). HSPGs consist of a protein core and heparan sulfate (HS) chains containing repeating disaccharide units of glucuronic/iduronic acid and glucosamine. The fine structure of HSPG is highly modified through a combination of posttranslational modifications, including sulfation on the N, 3-O, and 6-O positions of glucosamine and the 2-O position of the uronic acid units (26
). The sulfation pattern of the HS chains is a major determinant of the specificity and the affinity of ligand interactions (27
). Changes in this pattern can alter growth factor bioavailability and thus influence cell signaling during development and disease. Pertinent to the present study, 6-O–sulfation is pivotal for the binding of many growth factors to HSPGs and is critical for normal development (29
Although a number of enzymes control HSPG biosynthesis, the extracellular sulfatases, or SULFs, are unique because they modify the sulfation status of HSPG postsynthetically in the cellular microenvironment. By removing internal 6-O–sulfates of glucosamine on HS chains in regions of high overall sulfation, the SULFs can regulate the activity of HSPG-interacting ligands dynamically in the extracellular environment (23
). In humans, 2 extracellular sulfatases, SULF1
, have been characterized (32
). First identified for their role in Wnt-dependent signaling during muscle development in quail (23
), the SULFs regulate the in vivo activity of a number of HSPG-interacting ligands. In addition to Wnts, these include glial cell line–derived neurotrophic factor (GDNF), sonic hedgehog (SHH), VEGF, FGF2, and Noggin (23
). Both Sulf1
are highly expressed in the central nervous system and help regulate SHH signaling and neurite outgrowth (36
In tumorigenesis, SULFs may serve either tumor-promoting or tumor-inhibiting functions, depending on the dominant signaling pathway(s) active in a given tumor (42
). In human hepatocellular, breast, pancreatic, and non–small cell lung carcinoma, SULF2 is upregulated and promotes tumorigenesis (43
). In the latter 2 cases, SULF2 exerts its growth-promoting effects via increased Wnt signaling. In contrast, in SULF-negative human ovarian adenocarcinoma cell lines, overexpression of SULF1 results in decreased FGF2 and heparin-binding EGF-like growth factor (HB-EGF) signaling (46
GBM is driven by the abnormal activation of RTK signaling pathways. We hypothesized that GBM uses the extracellular SULFs to manipulate the tumor microenvironment and affect tumorigenesis. We tested this hypothesis in human GBM cell lines and in an orthotopic murine model for high-grade glioma (47
) by altering SULF2 expression and examining the effects on tumor growth and activation of critical growth factor signaling pathways. In this study, we also explored SULF expression levels in human GBM. Our findings indicate SULF2 expression may contribute to the pathogenesis of an important subset of human GBM.