Κωνσταντόπουλος, Κωνσταντίνος

Abstract Extracellular matrix (ECM) stiffening correlates with tumor invasion in various cancer types, including glioblastoma (GBM). Increased matrix stiffness promotes a migratory phenotype through dysregulation of cell-ECM bidirectional communication. Exposure to stiffer environments is sensed by cells, which then adapt in ways that promote invasive behavior. These adaptive changes are imprinted onto the cells and persist even after they are placed in new, softer microenvironments via a process known as “mechanical memory.” Mechanical memory is believed to be driven by mechanosensitive transcription factor activity and epigenetic remodeling. Glioblastoma recurrence is linked to the ability of cells to disperse and infiltrate the surrounding healthy tissue. Extracellular matrix stiffness in GBM is heterogeneous; it starts with a softer tumor core and becomes progressively stiffer toward the tumor’s edges, potentially promoting sustained tumor invasion through mechanical memory. This review discusses the role of ECM stiffness in cancer cell behavior and the implications of ECM stiffening in GBM. We then describe the findings associated with mechanical memory and relay underlying mechanisms currently understood to drive the preservation of mechanically primed phenotypes. Finally, we discuss how matrix stiffness can drive migratory phenotypes in GBM cells and the potential role that progressive ECM dysregulation at the tumor periphery can link the formation of invasive tumor niches to the aggressive, resistant, and mesenchymal-like phenotypes present in GBM recurrent tumors.

Abstract The extracellular matrix (ECM) plays a critical role in cancer progression, with its biophysical properties influencing cancer behavior. ECM stiffness can induce long-lasting adaptations in cancer cells, promoting sustained invasion through a process known as “mechanical memory” (MM). Glioblastoma (GBM) exhibits spatial heterogeneity in stiffness, with stiffer ECM found at the tumor periphery where invasive cells reside and promote recurrence. We hypothesize that increased stiffness at the tumor periphery primes GBM cells for sustained invasion through MM. METHODS Cell lines were established from image-guided human GBM samples from tumor margin and core (n=10 patients, 20 cell lines). Cells from random locations were conditioned in physiologically relevant extracellular matrices for two weeks. Human organotypic assay (n=3), Migration Assays, Atomic Force Microscopy (AFM), Immunocytochemistry, Western Blot, and RNA-sequencing (n=6) were performed. RESULTS GBM cells at higher stiff surfaces displayed increased migration (p<0.01). Magnetic Resonance Elastography (n=9) confirmed GBM heterogeneity, with softer cores (0.2-0.5kPa) transitioning to stiffer margins (>2kPa). Organotypic scaffolds mimicking the tumor margin promoted enhanced migration of GFP-labeled GBM cells. Margin cells and cells conditioned on stiffer matrices demonstrated higher migratory potential compared to their core and soft-conditioned counterparts (p<0.05). These trends persist over time and across multiple passages. ECM stiffness exposure causes nanomechanical, cytoskeletal, and transcriptional adaptations. AFM shows that cells from the margin (n=6) are softer (p<0.01) and more deformable (p<0.01). RNAseq identified upregulation of MM-specific genes in GBM margin cells. The mechanosensitive transcription factor YAP1 emerged as a key driver of MM. Notably, YAP1 is targetable by the FDA-approved drug Verteporfin. YAP1 expression was elevated in margin and stiff-conditioned cells (p<0.05). Stiff-conditioned cells and margin cells displayed increased sensitivity to Verteporfin treatment (p<0.05), with treatment significantly reducing margin migration compared to the core (p<0.001). CONCLUSION Stiffer GBM margins promote the formation of aggressive phenotypes through MM. Targeting YAP1 with Verteporfin, a readily available drug, presents a promising strategy to halt GBM invasion and improve patient outcomes.