Temozolomide Resistance Research Articles

Dual inhibition of CYP17A1 and HDAC6 by abiraterone-installed hydroxamic acid overcomes temozolomide resistance in glioblastoma through inducing DNA damage and oxidative stress

Glioblastoma (GBM) is a highly aggressive and treatment-resistant brain tumor, necessitating novel therapeutic strategies. In this study, we present a mechanistic breakthrough by designing and evaluating a series of abiraterone-installed hydroxamic acids as potential dual inhibitors of CYP17A1 and HDAC6 for GBM treatment. We established the correlation of CYP17A1/HDAC6 overexpression with tumor recurrence and temozolomide resistance in GBM patients. Compound 12, a dual inhibitor, demonstrated significant anti-GBM activity in vitro, particularly against TMZ-resistant cell lines. Mechanistically, compound 12 induced apoptosis, suppressed recurrence-associated genes, induced oxidative stress and initiated DNA damage response. Furthermore, molecular modeling studies confirmed its potent inhibitory activity against CYP17A1 and HDAC6. In vivo studies revealed that compound 12 effectively suppressed tumor growth in xenograft and orthotopic mouse models without inducing significant adverse effects. These findings highlight the potential of dual CYP17A1 and HDAC6 inhibition as a promising strategy for overcoming treatment resistance in GBM and offer new hope for improved therapeutic outcomes.

Transformed astrocytes confer temozolomide resistance on glioblastoma via delivering ALKBH7 to enhance APNG expression after educating by glioblastoma stem cells-derived exosomes.

Glioblastoma is the most malignant primary brain tumor in adults. Temozolomide (TMZ) stands for the first-line chemotherapeutic agent against glioblastoma. Nevertheless, the therapeutic efficacy of TMZ appears to be remarkably limited, because of low cytotoxic efficiency against glioblastoma. Besides, various mechanical studies and the corresponding strategies fail to enhancing TMZ curative effect in clinical practice. Our previous studies have disclosed remodeling of glial cells by GSCs, but the roles of these transformed cells on promoting TMZ resistance have never been explored. Exosomes were extracted from GSCs culture through standard centrifugation procedures, which can activate transformation of normal human astrocytes (NHAs) totumor-associated astrocytes (TAAs) for 3 days through detect the level of TGF-β, CD44 and tenascin-C. The secretive protein level of ALKBH7 of TAAs was determined by ELISA kit. The protein level of APNG and ALKBH7 of GBM cells were determined by Western blot. Cell-based assays of ALKBH7 and APNG triggered drug resistance were performed through flow cytometric assay, Western blotting and colony formation assay respectively. A xenograft tumor model was applied to investigate the function of ALKBH7 invivo. Finally, the effect of the ALKBH7/APNG signaling on TMZ resistance were evaluated by functional experiments. Exosomes derived from GSCs can activate transformation of normal human astrocytes (NHAs)to tumor-associated astrocytes (TAAs), as well as up-regulation of ALKBH7expression in TAAs. Besides, TAAs derived ALKBH7 can regulate APNG gene expression of GBM cells. After co-culturing with TAAs for 5 days, ALKBH7 and APNG expression in GBM cells were elevated. Furthermore, Knocking-down of APNG increased the inhibitory effect of TMZ on GBM cells survival. The present study illustrated a new mechanism of glioblastoma resistance to TMZ, which based on GSCs-exo educated TAAs delivering ALKBH7 to enhance APNG expression of GBM cells, which implied that targeting on ALKBH7/APNG regulation network may provide a new strategy of enhancing TMZ therapeutic effects against glioblastoma.

The impact of calcium-binding protein S100P on temozolomide sensitivity in glioblastoma cells

AimGlioblastoma (GBM), a highly aggressive primary brain tumor, often displays resistance to the standard chemotherapy agent Temozolomide (TMZ), presenting a formidable treatment challenge. This study aims to explore the impact of S100P, a calcium-binding protein linked to TMZ resistance in GBM cells, by investigating the consequences of S100P knockdown on TMZ resistance. The research, employing bioinformatics analysis and siRNA-mediated gene silencing, reveals a significant relationship between S100P and TRIM29 in the context of TMZ resistance. Material and methodsHuman GBM cells underwent transfection with S100P siRNA to achieve targeted suppression of S100P expression. The assessment of TMZ resistance was conducted through MTT cell viability assays, and the molecular mechanisms underlying the observed effects of S100P knockdown on TMZ resistance were explored using real-time PCR. ResultsOur findings reveal a significant reduction in TMZ resistance upon S100P knockdown in GBM cells. This effect is demonstrated by an increase in cell death and a concurrent decrease in the expression of proteins associated with TMZ resistance. ConclusionThe study establishes S100P as a promising therapeutic target to overcome TMZ resistance in GBM. Silencing S100P expression emerges as a strategy to enhance the sensitivity of GBM cells to TMZ chemotherapy. These results advocate further investigations to unveil the clinical viability of targeting S100P in the treatment of GBM.

NRP1 Induces Enhanced Stemness and Chemoresistance in Glioma Cells via YAP.

Neuropilin-1 (NRP1), a transmembrane glycoprotein, plays an important role in the malignant progression of gliomas; however, its role in chemoresistance is not fully understood. In this study, we observed the effects of NRP1 on the stemness and chemoresistance of glioma cells and the mediating role of Yes-associated protein (YAP). We constructed NRP1 overexpressing LN-229 glioma cells. Cells were treated with recombinant NRP1 protein (rNRP1) and the YAP inhibitor Super-TDU when necessary. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect the sensitivity of cells to temozolomide (TMZ). Sphere and clone formation assays were performed to detect the sphere- and clone-forming abilities of cells. Western blotting was performed to detect cellular CD133, CD44, p-LATS1, and p-YAP protein expression. Immunofluorescence and flow cytometry were used to detect the subcellular localization of YAP and apoptosis, respectively. We found that both NRP1 overexpression and rNRP1 treatment enhanced self-renewal, TMZ resistance, and CD133 and CD44 protein expression in LN-229 cells. NRP1 overexpression and rNRP1 treatment also induced LATS1 and YAP dephosphorylation and YAP nuclear translocation. Super-TDU inhibits NRP1 overexpression-induced enhanced self-renewal and TMZ resistance in LN-229 cells. Our study suggests that NRP1 induces increased stemness in glioma cells, resulting in chemoresistance, and that this effect is associated with YAP activation.

Exploring the Influence of tGLI-1 on Temozolomide Resistance in Glioblastomas: Unraveling Novel Therapeutic Targets

Background/Objective:Temozolomide (TMZ) is a standard chemotherapy treatment for patients with glioblastoma (GBM), but its effectiveness is limited, with only 50% of patients initially responding and developing resistance over time. Glioma stem cells (GSCs) have been implicated in TMZ resistance, particularly the mesenchymal subtype. The truncated form of GLI1, known as tGLI1, is highly expressed in mesenchymal GSCs and has been associated with poor patient outcomes in GBM. However, the role of tGLI1 in TMZ resistance remains unknown.
 Methods:The GBM cell line, U87MG, was utilized for this study. The IC50 of TMZ was determined using a cell viability assay. After successful transfection with vector, GLI1, and tGLI1, the cells were treated with the IC50 of TMZ to assess changes in cell viability between the groups.
 Results:The IC50 of TMZ is 290.1 μM, as averaged between replicate assays. Thus far, the results showed that tGLI1-expressing cells exhibited significantly higher cell viability (average: 39.09%) compared to Vector (average: 26.78%) and GLI1 (average: 27.11%). However, the tGLI1 group displayed higher variability in cell viability results, as evidenced by a larger standard deviation (0.2758) and standard error (0.1592) compared to vector (SD: 0.0325, SE: 0.0188) and GLI1 (SD: 0.1354, SE: 0.0781). The One-Way ANOVA, followed by Tukey's Multiple Comparison Test, results showed no statistically significant differences in cell viability between the groups.
 Conclusion/Impact:The increased cell viability observed in tGLI1-expressing cells suggests a potential association between tGLI1 and TMZ resistance, warranting additional research to fully comprehend its impact on GBM treatment response. Further investigation and replication studies are needed to establish the robustness of these results. This knowledge may contribute to the development oftargeted therapies aimed at inhibiting tGLI1 or its downstream signaling pathways, potentiallyimproving the response to TMZ and patient outcomes.

GARNL3 identified as a crucial target for overcoming temozolomide resistance in EGFRvIII-positive glioblastoma.

Amplification of the epidermal growth factor receptor (EGFR) and its active mutant type III (EGFRvIII), frequently occurr in glioblastoma (GBM), contributing to chemotherapy and radiation resistance in GBM. Elucidating the underlying molecular mechanism of temozolomide (TMZ) resistance in EGFRvIII GBM could offer valuable insights for cancer treatment. To elucidate the molecular mechanisms underlying EGFRvIII-mediated resistance to TMZ in GBM, we conducted a comprehensive analysis using Gene Expression Omnibus and The cancer genome atlas (TCGA) databases. Initially, we identified common significantly differentially expressed genes (DEGs) and prioritized those correlating significantly with patient prognosis as potential downstream targets of EGFRvIII and candidates for drug resistance. Additionally, we analyzed transcription factor expression changes and their correlation with candidate genes to elucidate transcriptional regulatory mechanisms. Using estimate method and databases such as Tumor IMmune Estimation Resource (TIMER) and CellMarker, we assessed immune cell infiltration in TMZ-resistant GBM and its relationship with candidate gene expression. In this study, we examined the expression differences of candidate genes in GBM cell lines following EGFRvIII intervention and in TMZ-resistant GBM cell lines. This preliminary investigation aimed to verify the regulatory impact of EGFRvIII on candidate targets and its potential involvement in TMZ resistance in GBM. Notably, GTPase Activating Rap/RanGAP Domain Like 3 (GARNL3) emerged as a key DEG associated with TMZ resistance and poor prognosis, with reduced expression correlating with altered immune cell profiles. Transcription factor analysis suggested Epiregulin (EREG) as a putative upstream regulator of GARNL3, linking it to EGFRvIII-mediated TMZ resistance. In vitro experiments confirmed EGFRvIII-mediated downregulation of GARNL3 and decreased TMZ sensitivity in GBM cell lines, further supported by reduced GARNL3 levels in TMZ-resistant GBM cells. GARNL3 downregulation in EGFRvIII-positive and TMZ-resistant GBM implicates its role in TMZ resistance, suggesting modulation of EREG/GARNL3 signaling as a potential therapeutic strategy.

CRISPR-Cas9 library screening combined with an exosome-targeted delivery system addresses tumorigenesis/TMZ resistance in the mesenchymal subtype of glioblastoma.

Rationale: The large-scale genomic analysis classifies glioblastoma (GBM) into three major subtypes, including classical (CL), proneural (PN), and mesenchymal (MES) subtypes. Each of these subtypes exhibits a varying degree of sensitivity to the temozolomide (TMZ) treatment, while the prognosis corresponds to the molecular and genetic characteristics of the tumor cell type. Tumors with MES features are predominantly characterized by the NF1 deletion/alteration, leading to sustained activation of the RAS and PI3K-AKT signaling pathways in GBM and tend to acquire drug resistance, resulting in the worst prognosis compared to other subtypes (PN and CL). Here, we used the CRISPR/Cas9 library screening technique to detect TMZ-related gene targets that might play roles in acquiring drug resistance, using overexpressed KRAS-G12C mutant GBM cell lines. The study identified a key therapeutic strategy to address the chemoresistance against the MES subtype of GBM. Methods: The CRISPR-Cas9 library screening was used to discover genes associated with TMZ resistance in the U87-KRAS (U87-MG which is overexpressed KRAS-G12C mutant) cells. The patient-derived GBM primary cell line TBD0220 was used for experimental validations in vivo and in vitro. Chromatin isolation by RNA purification (ChIRP) and chromatin immunoprecipitation (ChIP) assays were used to elucidate the silencing mechanism of tumor suppressor genes in the MES-GBM subtype. The small-molecule inhibitor EPIC-0412 was obtained through high-throughput screening. Transmission electron microscopy (TEM) was used to characterize the exosomes (Exos) secreted by GBM cells after TMZ treatment. Blood-derived Exos-based targeted delivery of siRNA, TMZ, and EPIC-0412 was optimized to tailor personalized therapy in vivo. Results: Using the genome-wide CRISPR-Cas9 library screening, we found that the ERBIN gene could be epigenetically regulated in the U87-KRAS cells. ERBIN overexpression inhibited the RAS signaling and downstream proliferation and invasion effects of GBM tumor cells. EPIC-0412 treatment inhibited tumor proliferation and EMT progression by upregulating the ERBIN expression both in vitro and in vivo. Genome-wide CRISPR-Cas9 screening also identified RASGRP1(Ras guanine nucleotide-releasing protein 1) and VPS28(Vacuolar protein sorting-associated protein 28) genes as synthetically lethal in response to TMZ treatment in the U87-KRAS cells. We found that RASGRP1 activated the RAS-mediated DDR pathway by promoting the RAS-GTP transformation. VPS28 promoted the Exos secretion and decreased intracellular TMZ concentration in GBM cells. The targeted Exos delivery system encapsulating drugs and siRNAs together showed a powerful therapeutic effect against GBM in vivo. Conclusions: We demonstrate a new mechanism by which ERBIN is epigenetically silenced by the RAS signaling in the MES subtype of GBM. Restoration of the ERBIN expression with EPIC-0412 significantly inhibits the RAS signaling downstream. RASGRP1 and VPS28 genes are associated with the promotion of TMZ resistance through RAS-GDP to RAS-GTP transformation and TMZ efflux, as well. A quadruple combination therapy based on a targeted Exos delivery system demonstrated significantly reduced tumor burden in vivo. Therefore, our study provides new insights and therapeutic approaches for regulating tumor progression and TMZ resistance in the MES-GBM subtype.

Revisiting Temozolomide's role in solid tumors: Old is gold?

Temozolomide is an imidazotetrazine with a long history in oncology especially for the high grade malignant glioma and metastatic melanoma. However, last year's new indications for its use are added. Its optimum pharmacodynamic profile, its ability to penetrate the blood-brain barrier, the existence of methylation of MGMT in solid tumors which enhances its efficacy, the identification of new agents that can overcome temozolomide's resistance, the promising role of temozolomide in turning immune cold tumors to hot ones, are leading to expand its use in other solid tumors, giving oncologists an additional tool for the treatment of advanced and aggressive neoplasms.

RNASE4 promotes malignant progression and chemoresistance in hypoxic glioblastoma via activation of AXL/AKT and NF-κB/cIAPs signaling pathways.

Glioblastoma (GBM) is the most malignant brain tumor frequently characterized by a hypoxic microenvironment. In this investigation, we unveiled unprecedented role of Ribonuclease 4 (RNASE4) in GBM pathogenesis through integrative methodologies. Leveraging The Cancer Genome Atlas (TCGA) dataset and clinical specimens from normal brain tissues, low- and high-grade gliomas, alongside rigorous in vitro and in vivo functional analyses, we identified a consistent upregulation of RNASE4 correlating with advanced GBM pathological stages and poor clinical survival outcomes. Functional assays corroborated the pivotal influences of RNASE4 on key tumorigenic processes such as cell proliferation, migration, invasion, stemness properties and temozolomide (TMZ) resistance. Further, Gene Set Enrichment Analysis (GSEA) illuminated the involvement of RNASE4 in modulating epithelial-mesenchymal transition (EMT) via activation of AXL, AKT and NF-κB signaling pathways. Furthermore, recombinant human RNASE4 (hRNASE4)-mediated NF-κB activation through IκBα phosphorylation and degradation could result in the upregulation of inhibitors of apoptosis proteins (IAPs), such as cIAP1, cIAP2, and SURVIVIN. Notably, treating RNASE4-induced TMZ-resistant cells with the SURVIVIN inhibitor YM-155 significantly restored cellular sensitivity to TMZ therapy. Herein, this study positions RNASE4 as a potent prognostic biomarker and therapeutic target, offering new insights into molecular pathogenesis of GBM and new avenues for future therapeutic interventions.

A multi-institutional phase I study of acetazolamide with temozolomide in adults with newly diagnosed MGMT-methylated malignant glioma.

A significant unmet need exists for the treatment of glioblastoma, IDH-wildtype (GBM). Preclinical work shows that acetazolamide sensitizes GBM to temozolomide (TMZ) by overcoming TMZ resistance due to BCL-3-dependent upregulation of carbonic anhydrase. Acetazolamide is Food and Drug Administration-approved for the treatment of altitude sickness. Drug repurposing enables the application of drugs to diseases beyond initial indications. This multi-institutional, open-label, phase I trial examined a combination of acetazolamide and TMZ in patients with MGMT promoter-methylated high-grade glioma. A total of 24 patients (GBM, IDH-wildtype = 22; Grade 4 astrocytoma, IDH-mutant = 1; Grade 3 astrocytoma, IDH-mutant = 1) were accrued over 17 months. All patients received oral acetazolamide (250mg BID for 7 days increased to 500mg BID for Days 8-21 of each 28-day cycle) during the adjuvant phase of TMZ for up to 6 cycles. No patient had a dose-limiting toxicity. Adverse events were consistent with known sequelae of acetazolamide and TMZ. In the 23 WHO Grade 4 patients, the median overall survival (OS) was 30.1 months and the median progression-free survival was 16.0 months. The 2-year OS was 60.9%. In total 37% of the study population had high BCL-3 staining and trended toward shorter OS (17.2 months vs N.R., P = .06). The addition of acetazolamide is safe and tolerable in GBM patients receiving standard TMZ. Survival results compare favorably to historical data from randomized trials in patients with MGMT promoter-methylated GBM and support examination of acetazolamide in a randomized trial. BCL-3 expression is a potential biomarker for prognosis in GBM or for patients more likely to benefit from TMZ.

A chloromethyl-triazole fluorescent chemosensor for O6-methylguanine DNA methyltransferase.

Fluorescent chemosensors offer a direct means of measuring enzyme activity for cancer diagnosis, predicting drug resistance, and aiding in the discovery of new anticancer drugs. O6-methylguanine DNA methyltransferase (MGMT) is a predictor of resistance towards anticancer alkylating agents such as temozolomide. Using the fluorescent molecular rotor, 9-(2-carboxy-2-cyanovinyl)julolidine (CCVJ), we synthesized, and evaluated a MGMT fluorescent chemosensor derived from a chloromethyl-triazole covalent inhibitor, AA-CW236, a non-pseudosubstrate of MGMT. Our fluorescence probe covalently labelled the MGMT active site C145, producing a 18-fold increase in fluorescence. Compared to previous fluorescent probes derived from a substrate-based inhibitor, our probe had improved binding and reaction rate. Overall, our chloromethyl triazole-based fluorescence MGMT probe is a promising tool for measuring MGMT activity to predict temozolomide resistance.

Diterpenoid honatisine overcomes temozolomide resistance in glioblastoma by inducing mitonuclear protein imbalance through disruption of TFAM-mediated mtDNA transcription

BackgroundGlioblastoma (GBM) represents as the most formidable intracranial malignancy. The systematic exploration of natural compounds for their potential applications in GBM therapy has emerged as a pivotal and fruitful avenue of research. PurposeIn the present study, a panel of 96 diterpenoids was systematically evaluated as a repository of potential antitumour agents. The primary objective was to discern their potency in overcoming resistance to temozolomide (TMZ). Through an extensive screening process, honatisine, a heptacyclic diterpenoid alkaloid, emerged as the most robust candidate. Notably, honatisine exhibited remarkable efficacy in patient-derived primary and recurrent GBM strains. Subsequently, we subjected this compound to comprehensive scrutiny, encompassing GBM cultured spheres, GBM organoids (GBOs), TMZ-resistant GBM cell lines, and orthotopic xenograft mouse models of GBM cells. ResultsOur investigative efforts delved into the mechanistic underpinnings of honatisine's impact. It was discerned that honatisine prompted mitonuclear protein imbalance and elicited the mitochondrial unfolded protein response (UPRmt). This effect was mediated through the selective depletion of mitochondrial DNA (mtDNA)-encoded subunits, with a particular emphasis on the diminution of mitochondrial transcription factor A (TFAM). The ultimate outcome was the instigation of deleterious mitochondrial dysfunction, culminating in apoptosis. Molecular docking and surface plasmon resonance (SPR) experiments validated honatisine's binding affinity to TFAM within its HMG-box B domain. This binding may promote phosphorylation of TFAM and obstruct the interaction of TFAM bound to heavy strand promoter 1 (HSP1), thereby enhancing Lon-mediated TFAM degradation. Finally, in vivo experiments confirmed honatisine's antiglioma properties. Our comprehensive toxicological assessments underscored its mild toxicity profile, emphasizing the necessity for a thorough evaluation of honatisine as a novel antiglioma agent. ConclusionIn summary, our data provide new insights into the therapeutic mechanisms underlying honatisine's selective inducetion of apoptosis and its ability to overcome chemotherapy resistance in GBM. These actions are mediated through the disruption of mitochondrial proteostasis and function, achieved by the inhibition of TFAM-mediated mtDNA transcription. This study highlights honatisine's potential as a promising agent for glioblastoma therapy, underscoring the need for further exploration and investigation.

Cytoplasmic Shotgun Proteomic Points to Key Proteins and Pathways in Temozolomide-Resistant Glioblastoma Multiforme.

Temozolomide (TMZ) is the first line of chemotherapy to treat primary brain tumors of the type glioblastoma multiforme (GBM). TMZ resistance (TMZR) is one of the main barriers to successful treatment and is a principal factor in relapse, resulting in a poor median survival of 15 months. The present paper focuses on proteomic analyses of cytosolic fractions from TMZ-resistant (TMZR) LN-18 cells. The experimental workflow includes an easy, cost-effective, and reproducible method to isolate subcellular fraction of cytosolic (CYTO) proteins, mitochondria, and plasma membrane proteins for proteomic studies. For this study, enriched cytoplasmic fractions were analyzed in replicates by nanoflow liquid chromatography tandem high-resolution mass spectrometry (nLC-MS/MS), and proteins identified were quantified using a label-free approach (LFQ). Statistical analysis of control (CTRL) and temozolomide-resistant (TMZR) proteomes revealed proteins that appear to be differentially controlled in the cytoplasm. The functions of these proteins are discussed as well as their roles in other cancers and TMZ resistance in GBM. Key proteins are also described through biological processes related to gene ontology (GO), molecular functions, and cellular components. For protein-protein interactions (PPI), network and pathway involvement analyses have been performed, highlighting the roles of key proteins in the TMZ resistance phenotypes. This study provides a detailed insight into methods of subcellular fractionation for proteomic analysis of TMZ-resistant GBM cells and the potential to apply this approach to future large-scale studies. Several key proteins, protein-protein interactions (PPI), and pathways have been identified, underlying the TMZ resistance phenotype and highlighting the proteins' biological functions.

Gene signatures associated with prognosis and chemotherapy resistance in glioblastoma treated with temozolomide.

Background: Glioblastoma (GBM) prognosis remains extremely poor despite standard treatment that includes temozolomide (TMZ) chemotherapy. To discover new GBM drug targets and biomarkers, genes signatures associated with survival and TMZ resistance in GBM patients treated with TMZ were identified. Methods: GBM cases in The Cancer Genome Atlas who received TMZ (n = 221) were stratified into subgroups that differed by median overall survival (mOS) using network-based stratification to cluster patients whose somatic mutations affected genes in similar modules of a gene interaction network. Gene signatures formed from differentially mutated genes in the subgroup with the longest mOS were used to confirm their association with survival and TMZ resistance in independent datasets. Somatic mutations in these genes also were assessed for an association with OS in an independent group of 37 GBM cases. Results: Among the four subgroups identified, subgroup four (n = 71 subjects) exhibited the longest mOS at 18.3months (95% confidence interval: 16.2, 34.1; p = 0.0324). Subsets of the 86 genes that were differentially mutated in this subgroup formed 20-gene and 8-gene signatures that predicted OS in two independent datasets (Spearman's rho of 0.64 and 0.58 between actual and predicted OS; p < 0.001). Patients with mutations in five of the 86 genes had longer OS in a small, independent sample of 37 GBM cases, but this association did not reach statistical significance (p = 0.07). Thirty-one of the 86 genes formed signatures that distinguished TMZ-resistant GBM samples from controls in three independent datasets (area under the curve ≥ 0.75). The prognostic and TMZ-resistance signatures had eight genes in common (ANG, BACH1, CDKN2C, HMGA1, IFI16, PADI4, SDF4, and TP53INP1). The latter three genes have not been associated with GBM previously. Conclusion: PADI4, SDF4, and TP53INP1 are novel therapy and biomarker candidates for GBM. Further investigation of their oncologic functions may provide new insight into GBM treatment resistance mechanisms.

Neovascularization directed by CAVIN1/CCBE1/VEGFC confers TMZ-resistance in glioblastoma

Acquisition of resistance to temozolomide (TMZ) poses a significant challenge in glioblastoma (GBM) therapy. Neovascularization, a pivotal process in tumorigenesis and development, remains poorly understood in its contribution to chemoresistance in GBMs. This study unveils aberrant vascular networks within TMZ-resistant (TMZ-R) GBM tissues and identifies the extracellular matrix (ECM) protein CCBE1 as a potential mediator. Through in vivo and in vitro experiments involving gain and loss of function assessments, we demonstrate that high expression of CCBE1 promotes hyper-angiogenesis and orchestrates partial endothelial-to-mesenchymal transition (EndMT) in human microvascular endothelial cells (HCMEC/d3) within GBM. This is likely driven by VEGFC/Rho signaling. Intriguingly, CCBE1 overexpression substantially fails to promote tumor growth, but endows resistance to GBM cells in a vascular endothelial cell-dependent manner. Mechanically, the constitutive phosphorylation of SP1 at Ser101 drives the upregulation of CCBE1 transcription in TMZ resistant tumors, and the excretion of CCBE1 depends on caveolae associated protein 1 (CAVIN1) binding and assembling. Tumor cells derived CCBE1 promotes VEGFC maturation, activates VEGFR2/VEGFR3/Rho signaling in vascular endothelial cells, and ultimately results in hyper-angiogenesis in TMZ-R tumors. Collectively, the current study uncovers the cellular and molecular basis of abnormal angiogenesis in a chemo resistant microenvironment, implying that curbing CCBE1 is key to reversing TMZ resistance.

Carmofur prevents cell cycle progression by reducing E2F8 transcription in temozolomide-resistant glioblastoma cells

Sphingolipid metabolism is dysregulated in many cancers, allowing cells to evade apoptosis through increased sphingosine-1-phosphate (S1P) and decreased ceramides. Ceramidases hydrolyze ceramides to sphingosine, which is phosphorylated by sphingosine kinases to generate S1P. The S1P allows cells to evade apoptosis by shifting the equilibrium away from ceramides, which favor cell death. One tumor type that exhibits a shift in the sphingolipid balance towards S1P is glioblastoma (GBM), a highly aggressive brain tumor. GBMs almost always recur despite surgical resection, radiotherapy, and chemotherapy with temozolomide (TMZ). Understanding sphingolipid metabolism in GBM is still limited, and currently, there are no approved treatments to target dysregulation of sphingolipid metabolism in GBM. Carmofur, a derivative of 5-fluorouracil, inhibits acid ceramidase (ASAH1), a key enzyme in the production of S1P, and is in use outside the USA to treat colorectal cancer. We find that the mRNA for ASAH1, but not other ceramidases, is elevated in recurrent GBM. When TMZ-resistant GBM cells were treated with carmofur, decreased cell growth and increased apoptosis were observed along with cell cycle perturbations. RNA-sequencing identified decreases in cell cycle control pathways that were specific to TMZ-resistant cells. Furthermore, the transcription factor and G1 to S phase regulator, E2F8, was upregulated in TMZ-resistant versus parental GBM cells and inhibited by carmofur treatment in TMZ-resistant GBM cells, specifically. These data suggest a possible role for E2F8 as a mediator of carmofur effects in the context of TMZ resistance. These data suggest the potential utility of normalizing the sphingolipid balance in the context of recurrent GBM.

Identification of potential glioma drug resistance target proteins based on ultra-performance liquid chromatography-mass spectrometry differential proteomics.

In this study, to screen for candidate markers of temozolomide (TMZ) resistance in glioblastoma, we artificially established TMZ drug-resistant glioblastoma (GBM) cell lines, U251-TMZ and U87-TMZ. In the U251-TMZ and U87-TMZ cell lines, we screened and analyzed differentially expressed proteins using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) differential proteomics. Compared with the U251 and U87 control cell lines, 95 differential proteins were screened in the U251-TMZ and U87-TMZ cell lines, of which 28 proteins were upregulated and 67 proteins were down-regulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of the co-upregulated proteins showed that most of the differentially expressed proteins were located in the cytoplasm and were significantly upregulated in the biological processes related to vesicular transport in the intimal system and inflammatory response mediated by myeloid leukocytes. Seven candidates were identified as potential GBM markers of TMZ resistance. Combined with existing research findings, our study supports that UAP1L1 and BCKDK are promising potential markers of TMZ resistance in GBM. This is important for further understanding the molecular mechanisms that drive the development and enhancement of TMZ resistance.

Unveiling the Influence of Tumor Microenvironment and Spatial Heterogeneity on Temozolomide Resistance in Glioblastoma Using an Advanced Human In Vitro Model of the Blood‐Brain Barrier and Glioblastoma (Small 52/2023)

Unveiling the Influence of Tumor Microenvironment and Spatial Heterogeneity on Temozolomide Resistance in Glioblastoma Using an Advanced Human In Vitro Model of the Blood‐Brain Barrier and Glioblastoma (Small 52/2023)

Interaction of SENP6 with PINK1 Promotes Temozolomide Resistance in Neuroglioma Cells via Inducing the Mitophagy

Interaction of SENP6 with PINK1 Promotes Temozolomide Resistance in Neuroglioma Cells via Inducing the Mitophagy

CREB-induced LINC00473 promotes chemoresistance to TMZ in glioblastoma by regulating O6-methylguanine-DNA-methyltransferase expression via CEBPα binding

Temozolomide (TMZ) offers substantial therapeutic benefits for glioblastoma (GB), yet its efficacy is hindered the development of chemoresistance. The role of long non-coding RNAs (lncRNAs) in tumorigenesis and chemoresistance has garnered great attention in studies on TMZ resistance. This study aimed to reveal the role of LINC00473 in TMZ chemoresistance and the underlying mechanism in GB. The expression of LINC00473 in TMZ-resistant and TMZ-sensitive GB cells was investigated using qPCR analysis. The role of LINC00473 in regulating TMZ resistance in GB cells was analyzed using the CCK-8 assay, colony formation assay, and flow cytometry. The next steps included assessing if LINC00473 is regulated by CREB and whether LINC00473 promotes chemoresistance through MGMT regulation via CEBPα. Further, chemoresistance delivery between cells via exosomal LINC00473 was validated in vitro and in vivo. Results showed that LINC00473 levels were elevated in TMZ-resistant cells upon CREB activation, and the lncRNA promoted the chemoresistance of GB cells through the upregulation of MGMT expression. Mechanistically, LINC00473 regulated the MGMT expression by binding to CEBPα. The highly-expressed LINC00473 packaged in exosomes transferred chemoresistance to the adjacent TMZ-sensitive GB cells. In conclusion, a novel CREB/LINC00473/CEBPα/MGMT pathway was revealed in the GB TMZ-resistance formation. In addition, an exosome-based mechanism of chemoresistance transmission was revealed, suggesting that LINC00473 could be used as a novel therapeutic target for GB.