Funded Grants

Proteoglycans as novel biomarkers and therapeutic targets in glioblastoma

Glioblastoma is the most common primary malignant brain tumor of adults and currently has a very challenging prognosis. There is a great need for improved therapeutic options and detection of recurrent disease. Although GBM is characterized by abnormal activation of receptor tyrosine kinase (RTK) signaling pathways, therapeutic targeting of single kinases has been largely disappointing. Two major factors may be contributing to the limited efficacy of these targeted therapeutics. First, GBM is driven by the summation of multiple growth signaling inputs [1]. Second, GBMs are heterogeneous and there are clinically distinct subtypes, based on expression, genomic and proteomic profiling [2-7]. Thus, different tumor subtypes may be dependent on different patterns of oncogenic signaling. Successful clinical responses to treatment will therefore require the development of novel therapies that target multiple signaling pathways, and an improved understanding of the mechanisms driving abnormal cell signaling in the different human tumor subtypes.

We are investigating the factors in the tumor microenvironment in the human brain that drive oncogenic signaling and promote malignant astrocytoma. Our focus on extracellular determinants of disease is driven by a desire to identify novel, druggable therapeutic targets for GBM. In our studies, we have identified a novel mechanism by which GBM alters heparan sulfate proteoglycans (HSPGs) in the tumor microenvironment to promote RTK signaling and drive tumor growth [8]. In this proposal to the James S. McDonnell Foundation, we test the novel hypothesis that alterations in HSPGs are critical drivers of RTK signaling in recurrent GBM and that these create novel biomarkers of disease and therapeutic targets.

RTK signaling pathways regulate many aspects of human gliomagenesis including cell proliferation, cell survival, invasion, and angiogenesis. In GBM, abnormal activation of these pathways can be driven by altered ligand availability and altered receptor levels. Indeed, the two most commonly amplified genes in GBM are EGFR and PDGFRA [6, 9-11]. Once released from the cell, oncogenic ligands can be sequestered by the extracellular microenvironment limiting their availability. Enzymatic release from the extracellular matrix is one important mechanism regulating their post-synthetic availability. In certain tumors, ligand availability in the tumor microenvironment may be a critical determinant of oncogenic signaling and tumorigenesis and this should be tested and translated for therapeutic benefit.

Heparan sulfate proteoglycans (HSPGs) and HSPG-modifying enzymes, present on the cell surface and in the extracellular matrix, are upregulated and constitute a major component of the extracellular environment in GBM [12, 13]. In many tissues including the brain, HSPGs play a key role in a number of biological processes based on their ability to bind and regulate the activity of diverse protein ligands including growth factors, chemokines, and morphogens, such as PDGF, VEGF and FGF [14-18]. In cancer, dysregulated HSPGs have the potential to alter the activity of multiple cell signaling pathways. Indeed, altered expression of HSPGs and HSPG modifying enzymes is common in cancer. The SULFs are extracellular enzymes that act on HSPGs to liberate ligands from sequestration and promote cell signaling. They are broadly overexpressed in many human cancers, including glioblastoma, non-small cell lung cancer (NSCLC), hepatocellular carcinoma, breast cancer, head and neck cancer, pancreatic adenocarcinoma, multiple myeloma, and gastric carcinoma [19, 20]. In human GBM we identified robust SULF2 expression in tumor cells; and, using knockdown and transgenic approaches, we demonstrated SULF2 promotes tumor cell proliferation, tumor growth in vivo , and the activity of multiple RTKs, including PDGFRα[8]. Together, these data support a critical role for HSPGs in the upstream regulation of multiple signaling pathways in GBM.

For the majority of GBM patients, current treatment is not tailored to the patient’s tumor molecular phenotype, but this will change with elucidation of subtype-specific patterns of oncogenic signaling. While robust criteria for clinical tumor stratification are still evolving, it is important to incorporate the molecular and phosphoproteomic phenotype of the tumor into the development and validation of novel biomarkers and therapeutics. Our studies suggest high-level expression of HSPG-modifying SULF2 may denote tumors that are more dependent on exogenous ligand [8]. This is a very exciting result as it suggests detection of SULF2, or alterations in HSPGs, may help identify a subset of tumors that are more amenable to specific therapies including multi-RTK inhibition.

Our overall objective is to identify patients for which HSPG-dependent RTK signaling pathways are critical in disease and for which HSPGs, and enzymes that modify them, are both sensitive biomarkers of recurrent disease and novel therapeutic targets. To do this we will first stratify recurrent human GBMs based upon phospho-proteome profiling and molecular phenotype, including expression of HSPGs and HSPGmodifying enzymes. We will then test specific associations using recurrent human GBM xenografts and gene knockdown approaches. To measure the sensitivity and specificity of plasma HSPG and HSPG-modifying enzyme levels for recurrent GBM, we will compare plasma levels in GBM patients and age-matched controls and in the same patient before and after tumor resection. Next, we will test the efficacy of therapeutic agents that target SULF2 and HSPGs using human GBM xenografts. Based on our preliminary studies, extracellular HSPGs, and the enzymes that modify them, play important roles in GBM, and their detection in patient blood may provide a robust biomarker of disease.


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