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Ex vivo Dynamics of Human Glioblastoma Cells in a Microvasculature-on-a-Chip System Correlates with Tumor Heterogeneity and Subtypes

ABSTRACT: The perivascular niche (PVN) plays an essential role in brain tumor stem-like cell (BTSC) fate control, tumor invasion, and therapeutic resistance. We use a microvasculature-on-a-chip system as a PVN model to evaluate the ex vivo dynamics of BTSCs from ten glioblastoma patients. BTSCs were found to preferentially localize in the perivascular zone, where they exhibited either the lowest motility, as in quiescent cells, or the highest motility, as in the invasive phenotype, with migration over long distance. The degree of co-localization between tumor cells and microvessels varied significantly across patients. To validate these results from the microvasculature-on-a-chip system, single-cell transcriptome sequencing (10 patients and 21,750 single cells in total) was performed to identify tumor cell subtypes. The co-localization coefficient was found to positively correlate with proneural (stem-like) or mesenchymal (invasive) but not classical (proliferative) tumor cells. Furthermore, a gene signature profile including PDGFRA correlated strongly with the “homing” of tumor cells to the PVN. These findings demonstrate that our BTSC-on-a-chip model can recapitulate in vivo tumor cell dynamics and heterogeneity, representing a new route to study patient-specific tumor cell functions.

ORGANISM(S): Homo sapiens

PROVIDER: GSE125587 | GEO | 2019/02/15

REPOSITORIES: GEO

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Project description:Glioblastoma (GBM) is the most common primary malignant brain tumor, with patients exhibiting poor survival (median survival time: 15 months). Difficulties in treating GBM include not only the inability to resect the diffusively-invading tumor cells but also therapeutic resistance. The perivascular niche within the GBM tumor microenvironment contributes significantly to tumor cell invasion, cancer stem cell maintenance, and has been shown to protect tumor cells from radiation and chemotherapy. In this study, we describe the development of an in vitro artificial perivascular niche (PVN), culturing endothelial and stromal cells along with GBM cells within a methacrylamide-functionalized gelatin hydrogel, in order to examine how the presence of the PVN alters global genomic expression profiles. Using RNA-seq, we demonstrate that the inclusion of perivascular niche cells with GBM cells upregulates genes related to angiogenesis and remodeling, while downregulated genes are related to cell cycle and DNA damage repair. Signaling pathways and genes commonly implicated in GBM malignancy, such as MGMT, EGFR, PI3K-Akt signaling, and Ras/MAPK signaling are also upregulated in the presence of perivascular niche cells. We describe the kinetics of gene expression within the PVN hydrogels over a course of 14 days, observing the patterns associated with previously-observed GBM-mediated endothelial network co-option and regression that are altered in the presence of covalently-bound hyaluronic acid. We finally examine the effect of PVN culture on GBM cell responsiveness to temozolomide, a frontline chemotherapy used clinically against GBM. Notably, the presence of a PVN leads to significantly increased TMZ resistance compared to hydrogels containing only GBM cells. Overall, these results demonstrate that inclusion of cellular and matrix-associated elements of the PVN within in vitro models of GBM provide critical signals to regulate the phenotype and therapeutic responsiveness of GBM cells.

Project description:Aberrational epigenetic marks are believed to play a major role in establishing the abnormal features of cancer cells. Rational use and development of drugs aimed at epigenetic processes requires an understanding of the range, extent, and roles of epigenetic reprogramming in cancer cells. Using ChIP-chip and MeDIP-chip approaches, we localized well-established and prevalent epigenetic marks (H3K27me3, H3K4me3, H3K9me3, DNA methylation) on a genome scale in several lines of putative glioma stem cells (brain tumor stem cells, BTSCs) and, for comparison, normal human fetal neural stem cells (fNSCs). We determined a substantial M-bM-^@M-^\coreM-bM-^@M-^] set of promoters possessing each mark in every surveyed BTSC cell type, which largely overlapped the corresponding fNSC sets. However, there was substantial diversity among cell types in mark localization. We observed large differences among cell types in total number of H3K9me3+ positive promoters and peaks and in broad modification areas for H3K27me3 and, to a lesser extent, H3K9me3. We verified that a change in a broad modification (defined as >50 kb peak length) affected gene expression of CACNG7. We detected large numbers of bivalent promoters, but most bivalent promoters did not display direct overlap of contrasting epigenetic marks, but rather occupied nearby regions of the proximal promoter. There were significant differences in the sets of promoters bearing bivalent marks in the different cell types and few consistent differences between fNSCs and BTSCs. Overall, our M-bM-^@M-^\core setM-bM-^@M-^] data establishes sets of potential therapeutic targets, but the diversity in sets of sites and broad modifications among cell types underscores the need to carefully consider BTSC subtype variation in epigenetic therapy. Our results point toward substantial differences among cell types in the activity of the production/maintenance systems for H3K9me3 and for broad regions of modification (H3K27me3 or H3K9me3). Finally, the unexpected diversity in bivalent promoter sets among these multipotent cells indicates that bivalent promoters may play complex roles in the overall biology of these cells. These results provide key information for forming the basis for future rational drug therapy aimed at epigenetic processes in these cells. ChIP-chip and MeDIP-chip localization of histone modifications and DNA methylation in glioma stem cells and fetal neural stem cells comparison of 4 lines of brain tumor stem cells to each other and to normal fetal neural stem cells

Project description:Aberrational epigenetic marks are believed to play a major role in establishing the abnormal features of cancer cells. Rational use and development of drugs aimed at epigenetic processes requires an understanding of the range, extent, and roles of epigenetic reprogramming in cancer cells. Using ChIP-chip and MeDIP-chip approaches, we localized well-established and prevalent epigenetic marks (H3K27me3, H3K4me3, H3K9me3, DNA methylation) on a genome scale in several lines of putative glioma stem cells (brain tumor stem cells, BTSCs) and, for comparison, normal human fetal neural stem cells (fNSCs). We determined a substantial “core” set of promoters possessing each mark in every surveyed BTSC cell type, which largely overlapped the corresponding fNSC sets. However, there was substantial diversity among cell types in mark localization. We observed large differences among cell types in total number of H3K9me3+ positive promoters and peaks and in broad modification areas for H3K27me3 and, to a lesser extent, H3K9me3. We verified that a change in a broad modification (defined as >50 kb peak length) affected gene expression of CACNG7. We detected large numbers of bivalent promoters, but most bivalent promoters did not display direct overlap of contrasting epigenetic marks, but rather occupied nearby regions of the proximal promoter. There were significant differences in the sets of promoters bearing bivalent marks in the different cell types and few consistent differences between fNSCs and BTSCs. Overall, our “core set” data establishes sets of potential therapeutic targets, but the diversity in sets of sites and broad modifications among cell types underscores the need to carefully consider BTSC subtype variation in epigenetic therapy. Our results point toward substantial differences among cell types in the activity of the production/maintenance systems for H3K9me3 and for broad regions of modification (H3K27me3 or H3K9me3). Finally, the unexpected diversity in bivalent promoter sets among these multipotent cells indicates that bivalent promoters may play complex roles in the overall biology of these cells. These results provide key information for forming the basis for future rational drug therapy aimed at epigenetic processes in these cells. ChIP-chip and MeDIP-chip localization of histone modifications and DNA methylation in glioma stem cells and fetal neural stem cells

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Ex vivo Dynamics of Human Glioblastoma Cells in a Microvasculature-on-a-Chip System Correlates with Tumor Heterogeneity and Subtypes

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