Spatial Dissection of Invasive Front from Tumor Mass Enables Discovery of Novel microRNA Drivers of Glioblastoma Invasion

Abstract

Diffuse invasion is the primary cause of treatment failure of glioblastoma (GBM). Previous studies on GBM invasionhave long been forced to use the resected tumor mass cells. Here, a strategy to reliably isolate matching pairs of invasive (GBM INV ) and tumor core (GBM TC ) cells from the brains of 6 highly invasive patient-derived orthotopic models is described. Direct comparison of these GBM INV and GBM TC cells reveals a significantly elevated invasion capacity in GBM INV cells, detects 23/768 miRNAs over-expressed in the GBM INV cells (miRNA INV ) and 22/768 in the GBM TC cells (miRNA TC ), respectively. Silencing the top 3 miRNAs INV (miR-126, miR-369-5p, miR-487b) successfully blocks invasion of GBM INV cells in vitro and in mouse brains. Integrated analysis with mRNA expression identifies miRNA INV target genes and discovers KCNA1 as the sole common computational target gene of which 3 inhibitors significantly suppress invasion in vitro. Furthermore, in vivo treatment with 4-aminopyridine (4-AP) effectively eliminates GBM invasion and significantly prolongs animal survival times (P = 0.035). The results highlight the power of spatial dissection of functionally accurate GBM INV and GBM TC cells in identifying novel drivers of GBM invasion and provide strong rationale to support the use of biologically accurate starting materials in understanding cancer invasion and metastasis.

Microenvironmental stiffness induces metabolic reprogramming in glioblastoma

“…In contrast to previous reports, we did not find any significant differences in mRNA expression of genes whose expression have been commonly associated with ECM degradation, including MMPs and hyaluronidases, or proneural or mesenchymal GBM subtypes, including TGFβ1, TIMP1, CHI3L1, and OLIG2. Likewise, although previous studies have associated CD133 as a marker of edge cells and CD109 as a marker of core cells in GBM tumors, 49 – 51 we did not observe differences in CD133 or CD109 expression in GBM cells cultured in scaffolds mechanically matched to the tumor edge or core. Finally, no differences were seen in expression or subcellular localization of proteins that have been widely associated with mechanical signaling, including YAP, whose translocation to the nucleus has been widely associated with a response to a stiff microenvironment.…”

“…In vivo , tumor edge cells, residing in a softer microenvironment, are likewise highly invasive. 49 – 51 In contrast, the OXPHOS-weighted metabolism observed in stiff hydrogel cultures correlated with a higher proliferation rate, similar to the hyperproliferative, pseudopalisading regions typically observed near the GBM tumor core, just outside of the necrotic area in patients. 49 This result aligns with the idea that cells, energetically, have the capacity to either “go or grow.” 61 , 62 Previous studies have indicated that migration through a denser matrix, like the stiff hydrogels used here, requires more energy, 63 , 64 a situation in which cells may instead only have sufficient ATP available to proliferate.…”

“…We posit that the heterogeneous mechanical landscape of tumors may contribute to phenotype differences between GBM cells residing in the stiffer core and softer invasive edge of the tumor. For example, while a mixture of cells exhibiting proneural or mesenchymal subtypes can be found in both the edge and core of GBM tumors, the edge cells have a higher capacity for invasion 49 – 51 and transcriptional states distinct from those of core cells. 49 , 52 Furthermore, it has been proposed that GBM recurrence relies on edge cell plasticity, where these cells are thought to transition to a more core-like state immediately prior to seeding a new tumor.…”

Silencing GMPPB Inhibits the Proliferation and Invasion of GBM via Hippo/MMP3 Pathways

“…Since the Hippo pathway has been implicated in multiple cell death modes, including apoptosis, autophagy, pyroptosis, and ferroptosis [53][54][55][56], it will be intriguing to explore the mechanisms of cell death, particularly in PDOX models, in the future. While the established cell lines do not exactly replicate the inter-and/or intra-tumoral heterogeneity as seen in cancer stem cell or stem-like GBM cell-derived cell cultures, PDOX models provide a useful resource to harvest and test glioma stem cells to support future efforts in validating GMPPB as a potential therapeutic target and testing new anti-cancer drugs against GMPPB in GBM [57]. Our study provides novel insights on the interplay progression of GMPPB regulation of the Hippo pathway to MMP3 through a series of functional validations.…”

Aquaporins and Ion Channels as Dual Targets in the Design of Novel Glioblastoma Therapeutics to Limit Invasiveness

“…Mediating upregulation of ECM-degrading metalloproteins nAChR Breast carcinoma [165] Cervical cancer [166] Facilitating nicotine-and growth factor-induced increases in redox regulator thioredoxin, overexpression of which induces:…”