BACKGROUND: Temozolomide (TMZ) resistance is a significant challenge in treating glioblastoma (GBM). Collagen remodeling has been shown to be a critical factor for therapy resistance in other cancers. This study aimed to investigate the mechanism of TMZ chemoresistance by GBM cells reprogramming collagens. METHODS: Key extracellular matrix components, including collagens, were examined in paired primary and recurrent GBM samples as well as in TMZ-treated spontaneous and grafted GBM murine models. Human GBM cell lines (U251, TS667) and mouse primary GBM cells were used for in vitro studies. RNA-sequencing analysis, chromatin immunoprecipitation, immunoprecipitation-mass spectrometry, and co-immunoprecipitation assays were conducted to explore the mechanisms involved in collagen accumulation. A series of in vitro and in vivo experiments were designed to assess the role of the collagen regulators prolyl 4-hydroxylase subunit alpha 1 (P4HA1) and yes-associated protein (YAP) in sensitizing GBM cells to TMZ. RESULTS: This study revealed that TMZ exposure significantly elevated collagen type I (COL I) expression in both GBM patients and murine models. Collagen accumulation sustained GBM cell survival under TMZ-induced stress, contributing to enhanced TMZ resistance. Mechanistically, P4HA1 directly binded to and hydroxylated YAP, preventing ubiquitination-mediated YAP degradation. Stabilized YAP robustly drove collagen type I alpha 1 ( COL1A1) transcription, leading to increased collagen deposition. Disruption of the P4HA1-YAP axis effectively reduced COL I deposition, sensitized GBM cells to TMZ, and significantly improved mouse survival. CONCLUSION P4HA1 maintained YAP-mediated COL1A1 transcription, leading to collagen accumulation and promoting chemoresistance in GBM.
Temozolomide ; Humans ; Glioblastoma/drug therapy* ; Animals ; Mice ; Cell Line, Tumor ; Drug Resistance, Neoplasm/genetics* ; YAP-Signaling Proteins ; Hydroxylation ; Dacarbazine/pharmacology* ; Adaptor Proteins, Signal Transducing/metabolism* ; Transcription Factors/metabolism* ; Collagen/biosynthesis* ; Collagen Type I/metabolism* ; Prolyl Hydroxylases/metabolism* ; Antineoplastic Agents, Alkylating/therapeutic use*

Objective: To investigate the function of primary cilia in regulating the cellular response to temozolomide (TMZ) and ionizing radiation (IR) in glioblastoma (GBM). Methods: GBM cells were treated with TMZ or X-ray/carbon ion. The primary cilia were examined by immunostaining with Arl13b and γ-tubulin, and the cellular resistance ability was measured by cell viability assay or survival fraction assay. Combining with cilia ablation by IFT88 depletion or chloral hydrate and induction by lithium chloride, the autophagy was measured by acridine orange staining assay. The DNA damage repair ability was estimated by the kinetic curve of γH2AX foci, and the DNA-dependent protein kinase (DNA-PK) activation was detected by immunostaining assay. Results: Primary cilia were frequently preserved in GBM, and the induction of ciliogenesis decreased cell proliferation. TMZ and IR promoted ciliogenesis in dose- and time-dependent manners, and the suppression of ciliogenesis significantly enhanced the cellular sensitivity to TMZ and IR. The inhibition of ciliogenesis elevated the lethal effects of TMZ and IR via the impairment of autophagy and DNA damage repair. The interference of ciliogenesis reduced DNA-PK activation, and the knockdown of DNA-PK led to cilium formation and elongation. Conclusion Primary cilia play a vital role in regulating the cellular sensitivity to TMZ and IR in GBM cells through mediating autophagy and DNA damage repair.
Antineoplastic Agents, Alkylating/therapeutic use* ; Brain Neoplasms/metabolism* ; Cell Line, Tumor ; DNA/therapeutic use* ; Glioblastoma/metabolism* ; Humans ; Radiation, Ionizing ; Temozolomide/therapeutic use*

OBJECTIVE: To establish a mouse model bearing orthotopic temozolomide (TMZ)-resistant glioma that mimics the development of drug resistance in gliomas METHODS: Seventy-eight adult C57BL/6 mice were randomly divided into 6 groups ( RESULTS: The mouse models bearing TMZresistant glioma was successfully established. The cells from the high-dose induced group showed a significantly higher colony-forming rate than those from the high-dose control group ( CONCLUSIONS Progressive increase of TMZ doses in mice bearing orthotopic gliomas can effectively induce TMZ resistance of the gliomas.
Animals ; Antineoplastic Agents, Alkylating/pharmacology* ; Brain Neoplasms/drug therapy* ; Cell Line, Tumor ; Disease Models, Animal ; Drug Resistance, Neoplasm ; Glioma/drug therapy* ; Mice ; Mice, Inbred C57BL ; Temozolomide/therapeutic use*

Animals ; Antineoplastic Agents, Alkylating/pharmacology* ; Breast Neoplasms/drug therapy* ; Cell Line ; Cell Line, Tumor ; Cyclophosphamide/pharmacology* ; Ginkgo biloba/chemistry* ; Mammary Neoplasms, Animal/drug therapy* ; Mice ; Plant Extracts/administration & dosage*

Animals ; Antineoplastic Agents, Alkylating ; Busulfan ; Cell Transplantation ; Chimera ; Coturnix/physiology* ; Female ; Male ; Spermatogonia/transplantation* ; Testicular Diseases/therapy* ; Testis/transplantation*

OBJECTIVE: To investigate the effect of triptolide (TP) on oxidative stress and apoptosis in TM4 sertoli cells and related molecular mechanism. METHODS: TM4 cells were incubated with different concentrations of triptolide for 24 h, then collected for further experiments. Cell proliferation analysis was used to measure the inhibitive effect of triptolide on proliferation of TM4 cells; DCFH-DA (6-carboxy-2',7'-dichlorofluorescein diacetate) probe was used to stain the TM4 cells, the level change of intracellular ROS was discovered through flow cytometry; the TM4 cells were stained by Annexin V-FITC/PI to detect whether triptolide induced apoptosis in the TM4 cells; Protein was extracted from the TM4 cells in control and triptolide group. Western blot was performed to determine the expression of apoptosis marker protein cleaved-PARP and PI3K/Akt signaling pathway-related proteins [p-Akt (Ser473), Akt, p-mTOR (Ser2448), mTOR, p-p70S6K (Thr389), p70S6K]. RESULTS: Cell proliferation analysis revealed that triptolide reduced the TM4 cells viability significantly compared with control group in a dosage-dependent manner [10 nmol/L: (73.77±20.95)%, 100 nmol/L: (51.60±10.43)%, 500 nmol/L: (44.34±5.78)%]; The level of intracellular ROS in the TM4 cells was significantly induced in a dosage-dependent manner (P<0.01); triptolide remarkably induced early-stage and late-stage apoptosis in the TM4 cells [control: (3.84±1.50)%, 100 nmol/L: (13.04±2.03)%, 200 nmol/L: (16.24±1.34)%, 400 nmol/L: (18.76±3.45)%]; The expression of cleaved-PARP was significantly upregulated in the TM4 cells after incubation with triptolide (P<0.01); The expression levels of p-Akt/Akt and p-p70S6K/p70s6k were significantly increased compared with control group (P<0.01). No significant change was observed among the expression levels of p-mTOR/mTOR (P>0.05). CONCLUSION In vitro studies showed that triptolide could effectively suppress the proliferation and induce apoptosis of TM4 sertoli cells. The oxidative stress was upregulated after incubation with triptolide, which may be one of the mechanisms of cytotoxicity in TM4 cells. Treatment of triptolide led to activation of Akt and p70S6K, indicating that the PI3K/Akt signaling pathway may be involved in response to oxidative stress in TM4 cells. The activation of PI3K/Akt signaling pathway was one of the molecular mechanisms involved in triptolide-mediated oxidative stress in TM4 cells. Our study provides insight into alleviating reproductive toxicity of triptolide in clinical and developing male contraceptive.
Antineoplastic Agents, Alkylating/pharmacology* ; Apoptosis/drug effects* ; Diterpenes/pharmacology* ; Epoxy Compounds/pharmacology* ; Humans ; Male ; Oxidative Stress/drug effects* ; Phenanthrenes/pharmacology* ; Phosphatidylinositol 3-Kinases ; Proto-Oncogene Proteins c-akt/drug effects* ; Sertoli Cells/drug effects* ; Signal Transduction/drug effects*

This study was performed to validate the effectiveness and safety of concurrent chemoradiotherapy and adjuvant therapy with temozolomide for newly diagnosed glioblastoma multiforme as a standard treatment protocol. Between 2004 and 2011, patients newly diagnosed with glioblastoma who were treated with temozolomide during concurrent chemoradiotherapy and adjuvant chemotherapy were included from a single institution and analyzed retrospectively. The primary endpoint was overall survival, and the secondary endpoints were progression-free survival, response, and safety. A total of 71 patients were enrolled in this study. The response rate was 41% (29/71), and the tumor control rate was 80% (57/71). In the 67 patients who completed the concurrent chemoradiotherapy with temozolomide, the median overall survival was 19 months and the 1- and 2-yr overall survival rates were 78.3% and 41.7%, respectively. The median progression free survival was 9 months, and the 1- and 2-yr progression free survival rates were 33.8% and 14.3%, respectively. The mean duration of survival after progression of disease in salvage treatment group was 11.9 (1.3-53.2) months. Concurrent chemoradiotherapy with temozolomide resulted in grade 3 or 4 hematologic toxic effects in 2.8% of the patients. The current protocol of temozolomide during and after radiation therapy is both effective and safe and is still appropriate as the standard protocol for treatment of glioblastoma. An active salvage treatment might be required for a better prognosis.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Antineoplastic Agents, Alkylating/administration & dosage ; Brain Neoplasms/diagnosis/*mortality/*therapy ; Chemoradiotherapy, Adjuvant/methods/mortality ; Comorbidity ; Dacarbazine/administration & dosage/*analogs & derivatives ; Female ; Glioblastoma/diagnosis/*mortality/*therapy ; Hematologic Diseases/*mortality ; Humans ; Longitudinal Studies ; Male ; Middle Aged ; Prevalence ; Radiotherapy, Conformal/mortality ; Republic of Korea/epidemiology ; Risk Factors ; Survival Rate ; Treatment Outcome ; Young Adult

Epithelial ovarian cancer (OC) is a common gynecologic malignancy in women. The standard treatment for OC is maximal cytoreductive surgical debulking followed by platinum-based chemotherapy. Despite the high response rate to primary therapy, approximately 85% of patients will develop recurrent ovarian cancer (ROC). This review identifies the clinical use of trabectedin in the treatment algorithm for ROC, with specific emphasis on platinum-sensitive ROC, for which trabectedin in combination with pegylated liposomal doxorubicin has been approved as a treatment protocol. The main mechanisms of action of trabectedin at the cellular level and in the tumor microenvironment is also discussed as bases for identifying biomarkers for selecting patients who may largely benefit from trabectedin-based therapies.
Antineoplastic Agents, Alkylating ; therapeutic use ; Clinical Trials as Topic ; DNA Damage ; Dioxoles ; administration & dosage ; pharmacology ; therapeutic use ; Doxorubicin ; administration & dosage ; analogs & derivatives ; Female ; Humans ; Neoplasm Recurrence, Local ; drug therapy ; Neoplasms, Glandular and Epithelial ; drug therapy ; Ovarian Neoplasms ; drug therapy ; Polyethylene Glycols ; administration & dosage ; Tetrahydroisoquinolines ; administration & dosage ; pharmacology ; therapeutic use ; Tumor Microenvironment

OBJECTIVE: To determine whether histogram values of the normalized apparent diffusion coefficient (nADC) and normalized cerebral blood volume (nCBV) maps obtained in contrast-enhancing lesions detected on immediate post-operative MR imaging can be used to predict the patient response to concurrent chemoradiotherapy (CCRT) with temozolomide (TMZ). MATERIALS AND METHODS: Twenty-four patients with GBM who had shown measurable contrast enhancement on immediate post-operative MR imaging and had subsequently undergone CCRT with TMZ were retrospectively analyzed. The corresponding histogram parameters of nCBV and nADC maps for measurable contrast-enhancing lesions were calculated. Patient groups with progression (n = 11) and non-progression (n = 13) at one year after the operation were identified, and the histogram parameters were compared between the two groups. Receiver operating characteristic (ROC) analysis was used to determine the best cutoff value for predicting progression. Progression-free survival (PFS) was determined with the Kaplan-Meier method and the log-rank test. RESULTS: The 99th percentile of the cumulative nCBV histogram (nCBV C99) on immediate post-operative MR imaging was a significant predictor of one-year progression (p = 0.033). ROC analysis showed that the best cutoff value for predicting progression after CCRT was 5.537 (sensitivity and specificity were 72.7% and 76.9%, respectively). The patients with an nCBV C99 of < 5.537 had a significantly longer PFS than those with an nCBV C99 of ≥ 5.537 (p = 0.026). CONCLUSION: The nCBV C99 from the cumulative histogram analysis of the nCBV from immediate post-operative MR imaging may be feasible for predicting glioblastoma response to CCRT with TMZ.
Adult ; Aged ; Antineoplastic Agents, Alkylating/*therapeutic use ; Brain/pathology/radiography ; Brain Neoplasms/*drug therapy/mortality/radiography ; Chemoradiotherapy ; Dacarbazine/*analogs & derivatives/therapeutic use ; Diffusion Magnetic Resonance Imaging ; Disease Progression ; Disease-Free Survival ; Female ; Glioblastoma/*drug therapy/mortality/radiography ; Humans ; Kaplan-Meier Estimate ; Male ; Middle Aged ; Proportional Hazards Models ; ROC Curve ; Retrospective Studies

Although methyltransferase has been recognized as a major element that governs the epigenetic regulation of the genome during temozolomide (TMZ) chemotherapy in glioblastoma multiforme (GBM) patients, its regulatory effect on glioblastoma chemoresistance has not been well defined. This study investigated whether DNA methyltransferase (DNMT) expression was associated with TMZ sensitivity in glioma cells and elucidated the underlying mechanism. DNMT expression was analyzed by western blotting. miR-20a promoter methylation was evaluated by methylation-specific PCR. Cell viability and apoptosis were assessed using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and TdT-mediated dUTP-biotin nick end labeling assays, respectively. The results showed that compared with parental U251 cells, DNMT1 expression was downregulated, miR-20a promoter methylation was attenuated and miR-20a levels were elevated in TMZ-resistant U251 cells. Methyltransferase inhibition by 5-aza-2\'-deoxycytidine treatment reduced TMZ sensitivity in U251 cells. In U251/TM cells, DNMT1 expression was negatively correlated with miR-20a expression and positively correlated with TMZ sensitivity and leucine-rich repeats and immunoglobulin-like domains 1 expression; these effects were reversed by changes in miR-20a expression. DNMT1 overexpression induced an increase in U251/TM cell apoptosis that was inhibited by the miR-20a mimic, whereas DNMT1 silencing attenuated U251/TM cell apoptosis in a manner that was abrogated by miR-20a inhibitor treatment. Tumor growth of the U251/TM xenograft was inhibited by pcDNA-DNMT1 pretreatment and boosted by DNMT1-small hairpin RNA pretreatment. In summary, DNMT1 mediated chemosensitivity by reducing methylation of the microRNA-20a promoter in glioma cells.
Animals ; Antineoplastic Agents, Alkylating/*pharmacology/therapeutic use ; Apoptosis/drug effects ; Brain/drug effects/metabolism/pathology ; Brain Neoplasms/drug therapy/*genetics/pathology ; DNA (Cytosine-5-)-Methyltransferase/antagonists & inhibitors/*genetics/metabolism ; DNA Methylation ; Dacarbazine/*analogs & derivatives/pharmacology/therapeutic use ; Drug Resistance, Neoplasm ; Female ; Gene Expression Regulation, Neoplastic ; Glioma/drug therapy/*genetics/pathology ; Humans ; Mice, Inbred C57BL ; MicroRNAs/*genetics ; Promoter Regions, Genetic