Multi-inlet spheroid generator for modeling immune microenvironment of glioblastoma (GBM)
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE271170 | GEO | 2024/07/06
REPOSITORIES: GEO
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Project description:Background. Although several immunotherapies against glioblastoma (GBM) have been investigated for long time, only limited effective results are acquired. Therefore, we developed immunotherapy based on genome edited NK cells knocking out the checkpoint receptor, which would overcome the immunosuppressive tumor microenvironment in GBM. Methods. We generated T cell immunoglobulin and ITIM domain (TIGIT), an inhibitory receptor expressed on lymphocytes, knockout (KO) human primary NK cells using the clustered regularly interspaced short palindromic repeats (CRISPR)/ CRISPR-associated protein9 (Cas9), each single guide RNA targeting different genome sites on TIGIT coding exons. To detect anti-tumor activity of genome edited NK cells against GBM, we utilized 2D adherent model and spheroids derived from GBM cell lines, U87, T98G, LN18, and U251. Subsequently, we performed real time cell growth assays, flow cytometry based apoptosis assays, and ELISA for investigating anti-tumor activity. Result. We established TIGIT KO human primary NK cells using CRISPR/Cas9. Flow cytometry indicated effective knockout of TIGIT and unchanged expression of immune checkpoint receptors other than TIGIT. T7 endonuclease I mutation detection assays showed that RNPs disrupted the intended genome sites. Using real time cell growth assays, we revealed enhanced anti-tumor activity of genome edited NK cells against 2D adherent GBM cells. The genome edited NK cells also exhibits enhanced anti-tumor effect against GBM spheroids derived from GBM cell lines.Conclusion. Our founding suggests that immunotherapy based on TIGIT KO NK cells using CRISPR/Cas9 is a promising therapy against GBM.
2022-09-11 | GSE212941 | GEO
Project description:GBM is the most common and aggressive primary brain tumor with dismal prognosis. NK cells are large granular lymphocytes with natural cytotoxicity against tumor cells, and they should be established for the novel treatment against patients with GBM. We have previously reported highly activated and ex vivo-expanded NK cells derived from human peripheral blood, which is designated as Genuine-induced natural killer cell (GiNK), induced by our specific culture conditions exerted the cytotoxic effect on GBM cells via apoptosis. First, we comprehensively summarized the molecular characteristics, especially focusing on the expression of stem cell markers, extracellular matrix markers, chemokine, chemokine receptors, and the NK receptors’ ligands, of GBM spheroids compared with 2D adherent GBM cells by using microarray. In the spheroid derived from GBM cell lines, gene expression of stem cell markers, extracellular matrix markers, chemokine, chemokine receptor, NK cell inhibitory receptor’ ligand were up-regulated compared with 2D adherent GBM cells. A preclinical evaluation of the NK cells was performed by ex vivo 3D spheroid model derived from GBM cell lines. In 3D spheroid model, the NK cells accumulated and infiltrated around the spheroids and induced the GBM cell death. Flow cytometry-based apoptosis detection assay clearly showed that the NK cells induced the GBM cell death via apoptosis. Our findings could provide pivotal information for the NK cell based-immunotherapy to patients with GBM.
2021-08-19 | GSE182374 | GEO
Project description:GBM is the most common and aggressive primary brain tumor with dismal prognosis. NK cells are large granular lymphocytes with natural cytotoxicity against tumor cells, and they should be established for the novel treatment against patients with GBM. We have previously reported highly activated and ex vivo-expanded NK cells derived from human peripheral blood, which is designated as Genuine-induced natural killer cell (GiNK), induced by our specific culture conditions exerted the cytotoxic effect on GBM cells via apoptosis. First, we comprehensively summarized the molecular characteristics, especially focusing on the expression of stem cell markers, extracellular matrix markers, chemokine, chemokine receptors, and the NK receptors’ ligands, of GBM spheroids compared with 2D adherent GBM cells by using microarray. In the spheroid derived from GBM cell lines, gene expression of stem cell markers, extracellular matrix markers, chemokine, chemokine receptor, NK cell inhibitory receptor’ ligand were up-regulated compared with 2D adherent GBM cells. A preclinical evaluation of the NK cells was performed by ex vivo 3D spheroid model derived from GBM cell lines. In 3D spheroid model, the NK cells accumulated and infiltrated around the spheroids and induced the GBM cell death. Flow cytometry-based apoptosis detection assay clearly showed that the NK cells induced the GBM cell death via apoptosis. Our findings could provide pivotal information for the NK cell based-immunotherapy to patients with GBM.
2021-08-19 | GSE182373 | GEO
Project description:Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids. Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids.
2012-08-31 | E-GEOD-36666 | biostudies-arrayexpress
Project description:Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids. Keywords: Expression profiling by array Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids.
2014-02-17 | E-GEOD-53631 | biostudies-arrayexpress
Project description:Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids.
2012-08-31 | GSE36666 | GEO
Project description:Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids. Keywords: Expression profiling by array
2014-02-17 | GSE53631 | GEO
Project description:Summary: Melanoma spheroids grown under neural crest cell conditions are highly plastic migratory/invasive tumor cells endowed with immunomodulator function Background: The aggressiveness of melanoma tumors is likely to rely on their well-recognized heterogeneity and plasticity. Melanoma comprises multi subpopulations of cancer cells some of which may possess stem cell-like properties supporting the notion of plasticity. Although useful for certain tumors, the use of the sphere-formation assay to identify stem cell-like or tumor initiating cells subpopulations in human melanoma has been recently challenged. Our study reveals that this assay predicts a functional phenotype associated with aggressive behavior of tumor cells. Methodology/Principal Findings: We analyzed the molecular and functional phenotypes of melanoma spheroids formed in neural crest cell medium. Whether from metastatic (SLM8) or advanced primary (Mela1) tumors, spheroid cells expressed melanoma-associated markers. They displayed higher capacity to differentiate along mesenchymal lineages, and showed enhanced expression of SOX2, NANOG, KLF4, and/or OCT4 transcription factors, but not extensive self-renewal or enhanced tumorigenicity when compared to their adherent counterparts. To determine whether melanoma spheroids in our model could predict a molecular or functional phenotype, we performed gene expression profiling experiments using Affymetrix microarrays. Gene expression profiling attributed a neural crest cell signature to these spheroids and indicated that a migratory/invasive and immune-function modulating program could be associated with these cells. In vitro assays confirmed that these spheroids are endowed with enhanced migratory/invasive capacities. In immune activation assays, spheroid cells elicited a poorer allogenic response from immune cells and inhibited mitogen-dependent T cells activation and proliferation more efficiently than their adherent counterparts. Thus, our findings reveal novel immune-modulator function of melanoma spheroid cells and suggest specific roles for these spheroids in invasion and in evasion of antitumor immunity. Conclusion/Significance: The association of a more plastic, invasive and evasive, thus a more aggressive tumor phenotype with melanoma spheroid cells reveals a previously unrecognized aspect of tumor cells expanded as spheroid cultures. While of limited efficiency for melanoma initiating cell identification, our melanoma spheroid model predicted aggressive phenotype and could therefore, be constructive to investigate melanoma aggressiveness, relevant to patients and clinical transferability. 12 Total samples were analyzed: SLM8 adherent (SLMA) and spheroids (SLMS) cells, and Mela1 adherent (MelaA) and spheroid (MelaS) cells, all performed in triplicates. Paired statistical analyses were performed using Student's paired t-test on the gene signal intensities (gene level) and results were considered statistically significant at p-values <=0.05 and fold-change >=1.5.
2011-03-28 | E-GEOD-26980 | biostudies-arrayexpress
Project description:Glioblastoma multiforme (GBM) is the most common and aggressive primary malignant brain tumor, but the current therapeutic approaches have very limited impact on increase of the patients’s survival with GBM. Our recent clinical trial with autologous activated nature killer (NK) cells (AKCs) including 14 patients with recurrent GBM found that the progression-free and overall survival times were significantly increased in some patients in the treatment group. In the present study, we identified TNFRSF18, TNFSF4, and IL12RB2 as biomarkers that predict response to NK cell therapeutics by studying the tumor-immune microenvironment using NanoString and qRT-PCR analyses. In addition, our results show that responders to NK therapy have an immune-activated tumor microenvironment, whereas non-responders do not. Considering that immune and inflammatory response-enrichment is related to the high-risk group in this tumor type, our findings highlight a new treatment strategy of NK cell therapeutics for patients with highly aggressive recurrent GBM.
2023-02-01 | GSE207753 | GEO
Project description:Cancer immunotherapy is gaining increasing attention. However, immune checkpoints are exploited by cancer cells to evade anti-tumor immunotherapy. Here, we knocked out NKG2A, an immune checkpoint expressed on natural killer (NK) cells, in human pluripotent stem cells (hPSCs) and differentiated these hPSCs into NK (PSC-NK) cells. We show that NKG2A knockout (KO) enhances the anti-tumor and anti-viral capabilities of PSC-NK cells. NKG2A KO endows PSC-NK cells with higher cytotoxicity against HLA-E-expressing glioblastoma (GBM) cells, leukemia cells, and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected cells in vitro. The NKG2A KO PSC-NK cells also exerted potent anti-tumor activity in vivo, leading to substantially suppressed tumor progression and prolonged survival of tumor-bearing mice in a xenograft GBM mouse model. These findings underscore the potential of PSC-NK cells with immune checkpoint KO as a promising cell-based immunotherapy. The unlimited supply and ease of genetic engineering of hPSCs makes genetically engineered PSC-NK an attractive option for easily accessible "off-the-shelf" cancer immunotherapy.
2024-10-18 | GSE241039 | GEO