Abstract 931: Extracellular matrix stimuli regulate cancer stem cell population and migratory potential in glioblastoma
Authors: Chen Jee-wei E. , Lumibao Jan C. , Gaskins H Rex , Sarkaria Jann N. , Harley Brendan A. 
Published: July 2017

Introduction Glioblastoma (GBM) is the most common and lethal form of brain cancer. Poor survival is linked to diffuse infiltration throughout the brain, suggesting the need to examine the effect of the tissue environment at the margins of the GBM tumor on processes of GBM invasion. Further, the presence of GBM cancer stem cells (GSCs) in primary tumor specimens is believe to play a significant role in GBM spreading, recurrence, and therapeutic resistance. While biophysical properties of the tumor microenvironment, such as spatial gradients in matrix stiffness and hyaluronic acid (HA) content as well as regional hypoxia, may affect GBM invasive phenotypes, limited tools are available to rigorously examine the relationship between tissue microenvironment and tumor growth, invasion, and therapeutic outcomes. This work focuses on the integration of HA-functionalized gelatin hydrogels with 3D cell invasion assays to investigate the impact of tissue microenvironment on GBM invasion and GSC population.

Methods Hydrogels are generated from methacrylamide functionalized gelatin (GelMA) and methacrylated HA. Prepolymer mixture was crosslinked under UV light in the presence of a photoinitiator. GBM cells, either U87 cell lines or patient-derived xenografts (PDX), were mixed with prepolymer solution prior to polymerization and collected for up to 7 days in culture. To assess invasion, cells were seeded onto dextran beads overnight, with populational invasion metrics calculated after 3 and 7 days, while live cell tracing via confocal microscopy was employed for single cell invasion metrics. Samples were cultured with 5% CO2 and ~20% O2 (normoxia) or 1% O2(hypoxia).

Results The invasion of U87 and U87viii (constitutively active EGFR) cells increased significantly in GelMA hydrogels lacking HA, consistent with our previous findings that compensatory secretion of soluble HA in the absence of matrix-bound HA was associated with upregulated invasion. Heightened U87 invasion was observed in the presence of hypoxia for all hydrogel compositions, and was associated with upregualted markers of invasion (gene expression: MMP-2, VEGF; protein expression: ERK1/2) while PI3K was downregulated, suggesting cells respond to hypoxia by alternating invasion and proliferation behaviors. We subsequently examined the effect of matrix biophysical properties and hypoxia on the expansion of GSC populations within PDX cells (a gift of Jann Sarkaria, Mayo Clinic). Here, the expansion of the GSC population in EGFRviii mutant GBM6 was a function of hypoxia as well as matrix-bound HA.

Conclusions This work demonstrates that GBM cells respond dynamically to their matrix biophysical (Stiffness, HA content, hypoxia) properties. Using PDX cells, we provide evidence that the expansion of GSC is related with both metabolic environment (O2%) and matrix composition (+/- HA). Ongoing work seeks to understand how GSCs contribute to the invasive phenotype of GBM.

Citation Format: Jee-wei E. Chen, Jan C. Lumibao, H Rex Gaskins, Jann N. Sarkaria, Brendan A. Harley. Extracellular matrix stimuli regulate cancer stem cell population and migratory potential in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 931. doi:10.1158/1538-7445.AM2017-931