Proliferation Of Glioma Stem Cells Research Articles

Deciphering the role of transcription factors in glioblastoma cancer stem cells.

Glioblastoma (GBM), the most aggressive and fatal brain malignancy, is largely driven by a subset of tumor cells known as cancer stem cells (CSCs). CSCs possess stem cell-like properties, including self-renewal, proliferation, and differentiation, making them pivotal for tumor initiation, invasion, metastasis, and overall tumor progression. The regulation of CSCs is primarily controlled by transcription factors (TFs) which regulate the expressions of genes involved in maintaining stemness and directing differentiation. This review aims to provide a comprehensive overview of the role of TFs in regulating CSCs in GBM. The discussion encompasses the definitions of CSCs and TFs, the significance of glioma stem cells (GSCs) in GBM, and how TFs regulate GSC self-renewal, proliferation, differentiation, and transformation. The potential for developing TF-targeted GSC therapies is also explored, along with future research directions. By understanding the regulation of GSCs by TFs, we may uncover novel diagnostic and therapeutic strategies against this devastating disease of GBM.

Effect of luteolin on glioblastoma's immune microenvironment and tumor growth suppression

BackgroundGlioblastoma is the most malignant and prevalent primary human brain tumor, and the immunological microenvironment controlled by glioma stem cells is one of the essential elements contributing to its malignancy. The use of medications to ameliorate the tumor microenvironment may give a new approach for glioma treatment. MethodsGlioma stem cells were separated from clinical patient-derived glioma samples for molecular research. Other studies, including CCK8, EdU, Transwell, and others, supported luteolin's ability to treat glioma progenitor cells. Network pharmacology and molecular docking models were used to study the drug target, and qRT-PCR, WB, and IF were used to evaluate the molecular mechanism. Intracranial xenografts were examined using HE and IHC, while macrophage polarization was examined using FC. ResultsWe originally discovered that luteolin inhibits glioma stem cells. IL6 released by glioma stem cells is blocked during medication action and inhibits glioma stem cell proliferation and invasion via the IL6/STAT3 signaling pathway. Additionally, luteolin inhibits the secretion of TGFβ1, affects the polarization function of macrophages in the microenvironment, inhibits the polarization of M2 macrophages in TAM, and further inhibits various functions of glioma stem cells by affecting the IL6/STAT3 signaling pathway, luteolin crosstalk TGFβ1/SMAD3 signaling pathway, and so on. ConclusionsThrough the suppression of the immunological microenvironment and inhibition of the IL6/STAT3 signaling pathway, our study determined the inhibitory effect of luteolin on glioma stem cells. This medication's dual inhibitory action, which has a significant negative impact on the glioma stem cells' malignant process, makes it both a viable anti-glioma medication and a candidate for targeted glioma microenvironment therapy.

Roles of lncRNA-MALAT1 in the Progression and Prognosis of Gliomas.

Long noncoding RNAs (lncRNAs) represent a large subgroup of RNA transcripts that lack the function of coding proteins and may be essential universal genes involved in carcinogenesis and metastasis. LncRNA metastasis-associated lung adenocarcinoma transcript 1 (lncRNAMALAT1) is overexpressed in various human tumors, including gliomas. However, the biological function and molecular mechanism of action of lncRNA-MALAT1 in gliomas have not yet been systematically elucidated. Accumulating evidence suggests that the abnormal expression of lncRNA-MALAT1 in gliomas is associated with various physical properties of the glioma, such as tumor growth, metastasis, apoptosis, drug resistance, and prognosis. Furthermore, lncRNAs, as tumor progression and prognostic markers in gliomas, may affect tumorigenesis, proliferation of glioma stem cells, and drug resistance. In this review, we summarize the knowledge on the biological functions and prognostic value of lncRNA-MALAT1 in gliomas. This mini-review aims to deepen the understanding of lncRNA-MALAT1 as a novel potential therapeutic target for the individualized precision treatment of gliomas.

Abstract 3180: Suppression of the CPEB3 ribozyme modulates the progression of glioblastoma

Abstract Glioblastoma multiforme (GBM) is the most aggressive primary malignant brain tumor in adults, with a poor prognosis that highlights a dire clinical need for innovative therapeutic interventions. Despite significant advances in diagnoses and multimodality therapies, the overall prognosis for patients with GBM remains poor, with a median survival time of 15-18 months. Therefore, there is an unmet medical need to develop alternative treatment strategies to improve clinical outcomes. Dysregulation of post-transcriptional control and translational machinery have been implicated in malignant tumor development. Cytoplasmic polyadenylation element binding proteins (CPEB1-CPEB4) are RNA-binding proteins that regulate poly(A) tail elongation of target mRNAs and subsequently contribute to phenotypic changes in cancer cells. Notably, a self-cleaving ribozyme was identified in the CPEB3 gene, but its role in cancer is wholly unexplored. Considering the role of CPEB3 as a tumor suppressor gene and the promotion of cancer progression through the downregulation of CPEB3, our hypothesis is that the CPEB3 ribozyme regulates CPEB3 expression, and its activity contributes to the progression of tumors. Using antisense oligonucleotides (ASOs) as an approach, we demonstrated that inhibition of CPEB3 ribozyme resulted in an increase of CPEB3 mRNA and protein expression. Blocking the CPEB3 ribozyme led to a significant reduction in cell proliferation, migration, and invasion in GBM cell lines. Gene set enrichment analysis (GSEA) revealed the downregulation of epithelial-mesenchymal transition (EMT), angiogenesis, and hypoxia gene sets in GBM cells treated with ASO compared to Ctrl-ASO. We further measured VEGFA mRNA and protein expression and found that ASO-treated GBM cells secreted significantly less VEGF in conditioned media. Inhibition of the CPEB3 ribozyme also mitigated the EMT process in GBM cells. Subsequently, ASO strategies were applied to patient-derived glioma stem cells (GSCs), representing a clinically relevant model for pre-clinical therapeutic intervention. We found that treatment of CPEB3 ribozyme ASO up-regulated CPEB3 mRNA and inhibited cell proliferation in GSCs. Furthermore, the combination of ASO and temozolomide chemotherapy exhibited a more pronounced decrease in GSCs proliferation compared to individual treatment alone. Collectively, this study highlights the significance of the CPEB3 ribozyme in GBM and explores therapeutic approaches focused on targeting CPEB3 in cancer. Citation Format: Claire Chen, Eric Wang, Lily Tong, Mehran Nikan, Daniela A. Bota, Claudia Benavente, Andrej Luptak. Suppression of the CPEB3 ribozyme modulates the progression of glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3180.

CREB5 promotes the proliferation and self-renewal ability of glioma stem cells

Glioblastoma multiforme (GBM) is the most fatal form of brain cancer in humans, with a dismal prognosis and a median overall survival rate of less than 15 months upon diagnosis. Glioma stem cells (GSCs), have recently been identified as key contributors in both tumor initiation and therapeutic resistance in GBM. Both public dataset analysis and direct differentiation experiments on GSCs have demonstrated that CREB5 is more highly expressed in undifferentiated GSCs than in differentiated GSCs. Additionally, gene silencing by short hairpin RNA (shRNA) of CREB5 has prevented the proliferation and self-renewal ability of GSCs in vitro and decreased their tumor forming ability in vivo. Meanwhile, RNA-sequencing, luciferase reporter assay, and ChIP assay have all demonstrated the closely association between CREB5 and OLIG2. These findings suggest that targeting CREB5 could be an effective approach to overcoming GSCs.

Inhibitor of Wnt receptor 1 suppresses the effects of Wnt1, Wnt3a and β‑catenin on the proliferation and migration of C6 GSCs induced by low‑dose radiation.

The radioresistance of glioma is an important cause of treatment failure and tumor aggressiveness. In the present study, under performed with linear accelerator, the effects of 0.3 and 3.0Gy low‑dose radiation (LDR) on the proliferation and migration of C6 glioma stem cells invitro were examined by flow cytometric analysis, immunocytochemistry and western blot analysis. It was found that low‑dose ionizing radiation (0.3Gy) stimulated the proliferation and migration of these cells, while 3.0Gy ionizing radiation inhibited the proliferation of C6 glioma stem cells, which was mediated through enhanced Wnt/β‑catenin signaling, which is associated with glioma tumor aggressiveness. LDR treatment increased the expression of the DNA damage marker γ‑H2AX but promoted cell survival with a significant reduction in apoptotic and necrotic cells. When LDR cells were also treated with an inhibitor of Wnt receptor 1 (IWR1), cell proliferation and migration were significantly reduced. IWR1 treatment significantly inhibited Wnt1, Wnt3a and β‑catenin protein expression. Collectively, the current results demonstrated that IWR1 treatment effectively radio‑sensitizes glioma stem cells and helps to overcome the survival advantages promoted by LDR, which has significant implications for targeted treatment in radioresistant gliomas.

SPI1 activates TGF-β1/PI3K/Akt signaling through transcriptional upregulation of FKBP12 to support the mesenchymal phenotype of glioma stem cells.

Glioma stem cells (GSCs) exhibit diverse molecular subtypes with the mesenchymal (MES) population representing the most malignant variant. The oncogenic potential of Salmonella pathogenicity island 1 (SPI1), an oncogenic transcription factor, has been established across various human malignancies. In this study, we explored the association between the SPI1 pathway and the MES GSC phenotype. Through comprehensive analysis of the Cancer Genome Atlas and Chinese Glioma Genome Atlas glioma databases, along with patient-derived GSC cultures, we analyzed SPI1 expression. Using genetic knockdown and overexpression techniques, we assessed the functional impact of SPI1 on GSC MES marker expression, invasion, proliferation, self-renewal, and sensitivity to radiation in vitro, as well as its influence on tumor formation in vivo. Additionally, we investigated the downstream signaling cascades activated by SPI1. Our findings revealed a positive correlation between elevated SPI1 expression and the MES phenotype, which in turn, correlated with poor survival. SPI1 enhanced GSC MES differentiation, self-renewal, and radioresistance in vitro, promoting tumorigenicity in vivo. Mechanistically, SPI1 augmented the transcriptional activity of both TGF-β1 and FKBP12 while activating the non-canonical PI3K/Akt pathway. Notably, inhibition of TGF-β1/PI3K/Akt signaling partially attenuated SPI1-induced GSC MES differentiation and its associated malignant phenotype. Collectively, our results underscore SPI1's role in activating TGF-β1/PI3K/Akt signaling through transcriptional upregulation of FKBP12, thereby supporting the aggressive MES phenotype of GSCs. Therefore, SPI1 emerges as a potential therapeutic target in glioma treatment.

Tumor-specific polycistronic miRNA delivered by engineered exosomes for the treatment of glioblastoma.

Glioblastoma (GBM) has poor prognosis due to ineffective agents and poor delivery methods. MicroRNAs (miRs) have been explored as novel therapeutics for GBM, but the optimal miRs and the ideal delivery strategy remain unresolved. In this study, we sought to identify the most effective pan-subtype anti-GBM miRs and to develop an improved delivery system for these miRs. We conducted an unbiased screen of over 600 miRs against 7 glioma stem cell (GSC) lines representing all GBM subtypes to identify a set of pan-subtype-specific anti-GBM miRs and then used available TCGA GBM patient outcomes and miR expression data to hone in on miRs that were most likely to be clinically effective. To enhance delivery and expression of the miRs, we generated a polycistronic plasmid encoding 3 miRs (pPolymiR) and used HEK293T cells as biofactories to package pPolymiR into engineered exosomes (eExos) that incorporate viral proteins (Gag/VSVg) in their structure (eExos+pPolymiR) to enhance function. Our stepwise screen identified miR-124-2, miR-135a-2, and let-7i as the most effective miRs across all GBM subtypes with clinical relevance. Delivery of eExos+pPolymiR resulted in high expression of all 3 miRs in GSCs, and significantly decreased GSC proliferation in vitro. eExos+pPolymiR prolonged survival of GSC-bearing mice in vivo when compared with eExos carrying each of the miRs individually or as a cocktail. eExos+pPolymiR, which includes a pan-subtype anti-glioma-specific miR combination encoded in a polycistronic plasmid and a novel exosome delivery platform, represents a new and potentially powerful anti-GBM therapeutic.

Targeting EZH2 regulates the biological characteristics of glioma stem cells via the Notch1 pathway.

Glioma is the most common malignant brain tumor, and its behavior is closely related to the presence of glioma stem cells (GSCs). We found that the enhancer of zeste homolog 2 (EZH2) is highly expressed in glioma and that its expression is correlated with the prognosis of glioblastoma multiforme (GBM) in two databases: The Cancer Genome Atlas and the Chinese Glioma Genome Atlas. Additionally, EZH2 is known to regulate the stemness-associated gene expression, proliferation, and invasion ability of GSCs, which may be achieved through the activation of the STAT3 and Notch1 pathways. Furthermore, we demonstrated the effect of the EZH2-specific inhibitor GSK126 on GSCs; these results not only corroborate our hypothesis, but also provide a potential novel treatment approach for glioma.

SPOPL induces tumorigenicity and stemness in glioma stem cells by activating Notch signaling.

Recent studies have increasingly shown that glioma stem cells (GSCs) are extremely important for developing and treating glioblastoma multiforme (GBM). The Broad-complex, Tram-track, and Bric-a-brac protein family is functionally related to a variety of tumor stem cells, and the role of SPOPL as a member of this family in GSCs deserves to be investigated. To investigate the expression of SPOPL in GSCs and its impact on the prognosis of GBM patients by using clinical specimens, patient-derived primary GSCs and public databases. In vivo and in vitro, the effect of SPOPL on the proliferation, self-renewal, and differentiation ability of GSCs was explored. Probing the mechanism by which SPOPL affects the biological function of GSCs using RNA sequencing (RNA-seq) and rescue experiments. The expression of SPOPL was significantly upregulated in GSCs and GBM, and patients with high SPOPL expression had a poorer prognosis. SPOPL enhanced the proliferation and self-renewal ability of GSCs and enhanced the tumorigenicity of GSCs. The Notch signaling pathway was significantly inhibited in SPOPL knockdown GSCs. Activation or inhibition of the Notch signaling pathway rescued changes in the biological function of GSCs caused by altered SPOPL expression. SPOPL can be used as a potential prognostic biomarker for GBM in clinical work and promotes the proliferation and stemness of GSCs by activating the Notch signaling pathway, which may be a potential molecule for targeting GSCs to treat GBM.

CAMK2D serves as a molecular scaffold for RNF8-MAD2 complex to induce mitotic checkpoint in glioma.

MAD2 is a spindle assembly checkpoint protein that participates in the formation of mitotic checkpoint complex, which blocks mitotic progression. RNF8, an established DNA damage response protein, has been implicated in mitotic checkpoint regulation but its exact role remains poorly understood. Here, RNF8 proximity proteomics uncovered a role of RNF8-MAD2 in generating the mitotic checkpoint signal. Specifically, RNF8 competes with a small pool of p31comet for binding to the closed conformer of MAD2 via its RING domain, while CAMK2D serves as a molecular scaffold to concentrate the RNF8-MAD2 complex via transient/weak interactions between its p-Thr287 and RNF8's FHA domain. Accordingly, RNF8 overexpression impairs glioma stem cell (GSC) mitotic progression in a FHA- and RING-dependent manner. Importantly, low RNF8 expression correlates with inferior glioma outcome and RNF8 overexpression impedes GSC tumorigenicity. Last, we identify PLK1 inhibitor that mimics RNF8 overexpression using a chemical biology approach, and demonstrate a PLK1/HSP90 inhibitor combination that synergistically reduces GSC proliferation and stemness. Thus, our study has unveiled a previously unrecognized CAMK2D-RNF8-MAD2 complex in regulating mitotic checkpoint with relevance to gliomas, which is therapeutically targetable.

Kaempferol and Biomodified Kaempferol from Sophora japonica Extract as Potential Sources of Anti-Cancer Polyphenolics against High Grade Glioma Cell Lines.

The enzymatic hydrolysis of the extract of Sophora japonica by two glycosyl hydrolases (hesperidinase and galactosidase) was performed in order to obtain kaempferol (KPF)-enriched extract with an enhanced anticancer activity. The current study examined the effectiveness of both Sophora japonica extracts (before (KPF-BBR) and after (KPF-ABR) bioconversion reactions) in reducing cell viability and inducing apoptosis in human high-degree gliomas in vitro. Cytotoxicity was determined using an MTT assay. The effects of both compounds on the proliferation of glioma cell lines were measured using trypan blue exclusion, flow cytometry for cell cycle, wound healing (WH), and neurosphere formation assays. Cellular apoptosis was detected by DNA fragmentation and phosphatidylserine exposure. qPCR and luciferase assays evaluated NF-kB pathway inhibition. The survival rate of NG-97 and U-251 cells significantly decreased in a time- and dose-dependent manner after the addition of KPF-BBR or KPF-ABR. Thus, a 50% reduction was observed in NG-97 cells at 800 µM (KPF-BBR) and 600 µM (KPF-ABR) after 72 h. Both compounds presented an IC50 of 1800 µM for U251 after 72 h. The above IC50 values were used in all of the following analyses. Neither of the KPF presented significant inhibitory effects on the non-tumoral cells (HDFa). However, after 24 h, both extracts (KPF-BBR and KPF-ABR) significantly inhibited the migration and proliferation of NG-97 and U-251 cells. In addition, MMP-9 was downregulated in glioma cells stimulated by 12-O-tetradecanoylphorbol-13-acetate (TPA) plus KPF-BBR and TPA+KPF-ABR compared with the TPA-treated cells. Both KPF-BBR and KPF-ABR significantly inhibited the proliferation of glioma stem cells (neurospheres) after 24 h. DNA fragmentation assays demonstrated that the apoptotic ratio of KPF-ABR-treated cell lines was significantly higher than in the control groups, especially NG-97, which is not TMZ resistant. In fact, the flow cytometric analysis indicated that KPF-BBR and KPF-ABR induced significant apoptosis in both glioma cells. In addition, both KPF induced S and G2/M cell cycle arrest in the U251 cells. The qPCR and luciferase assays showed that both KPFs downregulated TRAF6, IRAK2, IL-1β, and TNF-α, indicating an inhibitory effect on the NF-kB pathway. Our findings suggest that both KPF-BBR and KPF-ABR can confer anti-tumoral effects on human cell glioma cells by inhibiting proliferation and inducing apoptosis, which is related to the NF-κB-mediated pathway. The KPF-enriched extract (KPF-ABR) showed an increased inhibitory effect on the cell migration and invasion, characterizing it as the best antitumor candidate.

Deciphering the molecular mechanism of the THBS1 gene in the TNF signaling axis in glioma stem cells

Glioma stem cells (GSCs) are thought to be responsible for the initiation and progression of glioblastoma (GBM). GBM presents highly invasive growth with a very high recurrence rate, so it has become a clinical problem to be solved urgently. RNAseq demonstrates that thrombospondin 1 (THBS1) acts not only in the angiogenic core of glioma but also with a high degree of invasiveness and infiltration. Nevertheless, defects in the signaling pathway research lead to a poor prognosis in glioma patients. To investigate the relevant molecular mechanism and signal pathway of glioma stem cell behavior mediated by THBS1, U251 astroglioma cells and GSCs were taken as model cells for in vitro experiments. The biological effects of THBS1 on glioma proliferation, migration, and adhesion were evaluated using Cell Counting Kit-8(CCK8) assays, EdU incorporation assays, migration assays, Transwell assays, Western blotting, and RNAseq. We found that the knockout of the THBS1 gene by CRISPR/Cas9 promoted proliferation and migration in U251 cells and GSCs, as well as influencing cell cycle progression by regulating the TNF/MAPK/NF-κB and TGF-β/Smad signaling pathways. Moreover, U251 cells and GSCs showed different responses to THBS1 knockout, suggesting specific and potential targets for GSCs in signaling pathways mediated by THBS1.

METTL3 knockdown promotes temozolomide sensitivity of glioma stem cells via decreasing MGMT and APNG mRNA stability

Chemo-resistance hinders the therapeutic efficacy of temozolomide (TMZ) in treating glioblastoma multiforme (GBM). Recurrence of GBM even after combination of maximal tumor resection, concurrent radio-chemotherapy, and systemic TMZ applocation is inevitable and attributed to the high therapeutic resistance of glioma stem cells (GSCs), which can survive, evolve, and initiate tumor tissue remodeling, the underlying mechanisms of GSCs chemo-resistance, have not been fully elucidated up-to-now. Emerging evidence showed that METTL3-mediated N6-methyladenosine (m6A) modification contributed to the self-renew and radio-resistance in GSCs, however, its role on maintenance of TMZ resistance of GSCs has not been clarified and need further investigations. We found that the cell viability and half-maximal inhibitory concentration (IC50) of GSCs against TMZ significantly decreased after GSCs underwent serum-induced differentiation to adherent growth of tumor cells. Besides, METTL3 expression and total m6A modification declined dramatically in consistence with GSCs differentiation. Knockdown of METTL3 weakened self-renew, proliferation and TMZ IC50 of GSCs, whereas enhanced TMZ induced γH2AX level, indicating upregulation of double-strand DNA damage. We also found that mRNA stability of two critical DNA repair genes (MGMT and APNG) was regulated by METTL3-mediated m6A modification. In conclusion, we speculated that METTL3-mediated m6A modification of MGMT and APNG mRNAs played crucial roles on suppression of TMZ sensitivity of GSCs, which suggest a potential new therapeutic target of METTL3 against GBM.

The U2AF65/circNCAPG/RREB1 feedback loop promotes malignant phenotypes of glioma stem cells through activating the TGF-β pathway

Glioma is the most aggressive and common malignant neoplasms in human brain tumors. Numerous studies have showed that glioma stem cells (GSCs)drive the malignant progression of gliomas. Recent studies have revealed that circRNAs can maintain stemness and promote malignant progression of glioma stem cells. We used bioinformatics analysis to identify circRNAs and potential RNA-binding proteins (RBPs) in glioma. qRT-PCR, western blotting, RNA FISH, RNA pull-down, RNA immunoprecipitation assay, ChIP, immunohistochemistry, and immunofluorescence methods were used to quantified the expression of circNCAPG, U2AF65, RREB1 and TGF-β1, and the underlying mechanisms between them. MTS, EDU, neurosphere formation, limiting dilution neurosphere formation and transwell assays examined the proliferation and invasive capability of GSCs, respectively. We identified a novel circRNA named circNCAPG was overexpressed and indicated the poor prognosis in glioma patients. Upregulating circNCAPG promoted the malignant progression of GSCs. RNA binding protein U2AF65 could stabilize circNCAPG by direct binding. Mechanically, circNCAPG interacted with and stabilized RREB1, as well as stimulated RREB1 nuclear translocation to activate TGF-β1 signaling pathway. Furthermore, RREB1 transcriptionally upregulated U2AF65 expression to improve the stability of circNCAPG in GSCs, which established a feedback loop involving U2AF65, circNCAPG and RREB1. Since circRNA is more stable than mRNA and can execute its function continuously, targeting circNCAPG in glioma may be a novel promising therapeutic.

A pan-cancer landscape of IGF2BPs and their association with prognosis, stemness and tumor immune microenvironment.

The human insulin-like growth factor 2 mRNA binding proteins 1-3 (IGF2BP1-3, also called IMP1-3) play essential roles in mRNA regulation, including its splicing, translocation, stability, and translation. However, knowledge regarding the involvement of IGF2BPs in tumor immunity and stemness across cancer types is still lacking. In this study, we comprehensively analyzed pan-cancer multi-omic data to determine the correlation of IGF2BPs mRNA and protein expression with various cancer parameters such as mutation frequency, prognostic value, the tumor microenvironment (TME), checkpoint blockade, tumor immune infiltration, stemness and drug sensitivity. Validation of the expression of IGF2BPs in cancer samples and glioma cells were performed by quantitative real-time (qRT)-PCR, and immunofluorescence staining. Investigation of the functional role of IGF2BP3 in glioma stem cells(GSCs) were performed by sphere formation, cytotoxicity, transwell, and wound healing assays. We found that IGF2BP1 and 3 are either absent or expressed at very low levels in most normal tissues. However, IGF2BP1-3 can be re-expressed in a broad range of cancer types and diverse cancer cell lines, where their expression often correlates with poor prognosis. Immunofluorescence staining and qRT-PCR analyses also showed that the expression of IGF2BP2 and IGF2BP3 were higher in cancer tissues than that in adjacent normal tissues. Moreover, IGF2BPs are associated with TME and stemness in human pan-cancer. Remarkably, IGF2BP3 participated in the maintenance and self-renewal of glioma stem cell (GSCs). Knockdown of IGF2BP3 attenuated GSC and glioma cell proliferation, invasion, and migration. Our systematic pan-cancer study confirmed the identification of IGF2BPs as therapeutic targets and highlighted the need to study their association with stemness, and the TME, which contribute to the cancer drug-discovery research. Especially, preliminary studies demonstrate the IGF2BP3 as a potential negative regulator of glioma tumorigenesis by modulating stemness.

ABTC-0904: targeting glioma stem cells in GBM: a phase 0/II study of hedgehog pathway inhibitor GDC-0449.

Gliomagenesis and resistance of glioblastoma (GBM) are believed to be mediated by glioma stem cells (GSC). Evidence suggests that SHH signaling promotes GSC proliferation and self-renewal. ABTC-0904 was a two-arm, multicenter phase 0/II study of GDC-0449, an oral inhibitor of Smoothened (SMO) in patients undergoing resection for recurrent GBM. All patients (Arms I and II) had surgery and received drug post-operatively. Only patients in Arm I received drug prior to surgery. The primary objective was to determine 6-month progression free survival (PFS-6). Secondary endpoints include median PFS (mPFS) and overall survival (mOS), response rate, and toxicity. Correlative studies included bioanalysis of GDC-0449, and inhibition of SHH signaling, GSC proliferation and self-renewal. Forty-one patients were enrolled. Pharmacokinetics of GDC-0449 in plasma demonstrated levels within expected therapeutic range in 75% of patients. The proportion of tumorcells producing CD133+ neurospheres, neurosphere proliferation, self-renewal, and expression of the SHh downstream signaling was significantly decreased in Arm I following GDC-0449 treatment (p < 0.005; p < 0.001 respectively) compared to Arm II (no drug pre-op). Treatment was well tolerated. There were no objective responders in either arm. Overall PFS-6 was 2.4% (95% CI 0.9-11.1%). Median PFS was 2.3months (95% CI 1.9-2.6) and mOS was 7.8months (95% CI 5.4-10.1). GDC-0449 was well tolerated, reached tumor, and inhibited CD133+ neurosphere formation, but had little clinical efficacy as a single agent in rGBM. This suggests growth and maintenance of rGBM is not solely dependent on the SHH pathway thus targeting SMO may require combined approaches.

Novel INHAT repressor drives glioblastoma growth by promoting ribosomal DNA transcription in glioma stem cells.

Cancer cells including cancer stem cells exhibit a higher rate of ribosome biogenesis than normal cells to support rapid cell proliferation in tumors. However, the molecular mechanisms governing the preferential ribosome biogenesis in glioma stem cells (GSCs) remain unclear. In this work, we show that the novel INHAT repressor (NIR) promotes ribosomal DNA (rDNA) transcription to support GSC proliferation and glioblastoma (GBM) growth, suggesting that NIR is a potential therapeutic target for GBM. Immunoblotting, immunohistochemical and immunofluorescent analysis were used to determine NIR expression in GSCs and human GBMs. Using shRNA-mediated knockdown, we assessed the role and functional significance of NIR in GSCs and GSC-derived orthotopic GBM xenografts. We further performed mass spectrometry analysis, chromatin immunoprecipitation, and other biochemical assays to define the molecular mechanisms by which NIR promotes GBM progression. Our results show that high expression of NIR predicts poor survival in GBM patients. NIR is enriched in the nucleoli of GSCs in human GBMs. Disrupting NIR markedly suppresses GSC proliferation and tumor growth by inhibiting rDNA transcription and pre-ribosomal RNA synthesis. In mechanistic studies, we find that NIR activates rDNA transcription to promote GSC proliferation by cooperating with Nucleolin (NCL) and Nucleophosmin 1 (NPM1), 2 important nucleolar transcription factors. Our study uncovers a critical role of NIR-mediated rDNA transcription in the malignant progression of GBM, indicating that targeting this axis may provide a novel therapeutic strategy for GBM.

Downregulation of ADAMTS3 Suppresses Stemness and Tumorigenicity in Glioma Stem Cell.

Glioblastoma multiforme (GBM) is the most aggressive type of human brain tumor, with a poor prognosis and a median overall survival of fewer than 15 months. Glioma stem cells (GSCs) have recently been identified as a key player in tumor initiation and therapeutic resistance in GBM. ADAMTS family of metalloproteinases is known to cleave a wide range of extracellular matrix substrates and has been linked to tissue remodeling events in tumor development. Here, we investigate that ADAMTS3 regulates GSC proliferation and self-renewal activities, and tumorigenesis in orthotopic xenograft models. ADAMTS3 mRNA expression levels in normal human astrocyte (NHA), glioma, and GSCs cell lines were compared. After knockdown of ADAMTS3, alamarBlue assay, in vitro limiting dilution, and orthotopic xenograft assays were performed. To investigate the tumor-associated roles of ADAMTS3, several statistical assays were conducted using publicly available datasets. ADAMTS3 level was remarkably higher in GSCs than in NHA, glioma cell lines, and their matched differentiated tumor cells. Interestingly, knockdown of ADAMTS3 disrupted GSC's proliferation, self-renewal activity, and tumor formation in vivo. Furthermore, ADAMTS3 could be used as an independent predictor of malignancy progression in GBM. We identified ADAMTS3 as a potential therapeutic target for GBM.

Cyanidin inhibits glioma stem cells proliferation through the Wnt signaling pathway

Background Cyanidin has a protective effect on the nervous system and has been reported to treat tumor effectively. However, its impact on glioma stem cells (GSC) is unknown. Methods Using seven GSC lines, the anti-tumor effect of cyanidin is tested. The effect of cyanidin on the cell viability in each cell line is evaluated. Wnt signaling pathway-related genes are checked after treatment of cyanidin. Cytoplasmic/nuclear β-catenin protein levels post cyanidin treatment is detected. Protein levels of c-Myc after cyanidin treatment are determined. Twist1 and Snail1 protein levels after cyanidin treatment are checked as well. Results Cyanidin significantly reduces the cell viability of all GSCs, and exhibited the most substantial effect in GBM2 but no apparent effect in 293T cells. It can regulate the Wnt signaling pathway of all GSC lines. In the GBM2, GBM7, G166, and G179 cell lines, there is upregulation of WNT1 and MYC genes, while in the G144 and GliNS2 cell line, these two genes are down-regulated after cyanidin treatment. Cytoplasmic and nuclear protein levels of β-catenin in all cell lines are down-regulated. Cyanidin treatment significantly decreases the protein level for c-Myc in the GBM2 cell line compared with untreated cells, not in G144 or GliNS2 cells. Furthermore, cyanidin strongly reduces the expression of Twist1 and Snail1 in GBM2, G179, and G144 cell lines, while the GliNS2 cells show an opposite change in the cytoplasm and no change in nuclear. Conclusion Cyanidin exerts an anti-tumor effect in glioma stem cell lines, probably through the Wnt signaling pathway.