Microenvironmental Cues Driving CD44-mediated Glioblastoma Invasion through Hyaluronic Acid-rich Matrix
Interactions between tumor cells and the extracellular matrix (ECM) play a critical role in tumor invasion. Glioblastoma (GBM) is a highly malignant primary brain cancer characterized by diffuse infiltration, with tumor cells invading slowly through the hyaluronic acid (HA)-rich parenchyma toward vascular beds and then migrating rapidly along microvasculature rich in collagen, fibronectin, and laminin. Nonetheless, little is known about how cells navigate nonfibrillar 3D matrices, such as the HA-rich brain parenchyma, or how plasticity in cell migration modes between intraparenchymal and perivascular spaces influences invasion. Progress in understanding local infiltration, vascular homing, and perivascular invasion is further limited by the absence of culture models that recapitulate these hallmark processes. The transmembrane receptor CD44 directly facilitates tumor cell invasion by engaging HA in brain matrix and is therefore a likely candidate for mediating invasive plasticity.
In this dissertation, we investigate how topographical cues in the perivascular niche instruct cell invasion modality and how CD44 coordinates with the cytoskeleton to facilitate invasion through HA-rich matrices. We first describe the development of a platform for GBM invasion consisting of a tumor-like cell reservoir and a parallel open channel “vessel” embedded in the 3D HA matrix. We show that this simple paradigm is sufficient to capture multi-step invasion as well as transitions in cell morphology and speed reminiscent of those in GBM. Tumor cells within the model grow into multicellular masses that expand and invade the surrounding HA-rich matrices while extending long (10–100 µm), thin protrusions before encountering the open channel. We then assess how HA signals arise in vivo and how these signals are generally incorporated into in vitro models to mechanistically investigate HA-mediated motility. Finally, we employ our HA hydrogel platform to probe the role of CD44 in cytoskeletal motility. We show that continuous and patient-derived GBM tumor cells interacting with both 2D and 3D HA substrates exploit CD44-based microtentacles (McTNs) to support cell migration. These McTNs are stabilized by a balance between microtubule-driven protrusion and actomyosin-driven retraction, which are mechanistically coupled by an IQGAP1-CLIP170 complex. Cells approaching vascular structures transition from McTN-based motility to a quasi-2D mesenchymal motility involving actomyosin bundles. McTN-driven motility and/or the transition to mesenchymal motility may represent an important new target for GBM discovery and therapeutics.