Taraxasterol enhanced bladder cancer cells radiosensitivity via inhibiting the COX-2/PGE2/JAK2/STAT3/MMP pathway.

PURPOSE: Radiotherapy with bladder preservation is highly acceptable among patients bearing bladder cancer (BCa), but the occurrence of secondary tolerance (ARR) during treatment is one of the important reasons for the failure of clinical radiotherapy. COX-2 has been frequently reported to be highly expressed and associated with radio-resistance in various cancers. In this study, the feasibility of Taraxasterol (Tara) as a radiosensitizer was investigated, and the target effect of Tara on COX-2 and its underlying mechanism were explored. METHODS AND MATERIALS: The toxicity of Tara toward BCa cells was detected with the MTT method and cells in response to IR or Tara + IR were compared by clone formation assay. Next, a small RNA interference system (siRNA) was employed to decrease endogenous COX-2 expression in BCa cells, and the stem cell-like features and motion abilities of BCa cells under different treatments were investigated using microsphere formation and transwell chamber assay, respectively. Meanwhile, the expression of a series of inflammation-related molecules and stem cell characteristic molecules was determined by qRT-PCR, western blot and ELISA method. In vivo studies, BCa cells were subcutaneously injected into the right flank of each male mouse. Those mice were then grouped and exposed to different treatment: Tara, IR, IR + Tara and untreated control. The volumes of each tumor were measured every two days and target proteins were detected with immunohistochemical (IHC) staining. RESULTS: The results show that COX-2 decline, due to COX-2 knocking-down or Tara treatment, could greatly enhance BCa cells' radiosensitivity and significantly decrease their migration, invasion and microsphere formation abilities, companied with the reduce of JAK2, phos-STAT3, MMP2 and MMP9 expression. However, Tara could not further reduce the expression of an above molecule of cells in COX-2-deficient BCa cells. Correspondingly, Tara treatment could not further enhance those siCOX-2 BCa cells response to IR. CONCLUSIONS: Our data support that Tara can improve the radiosensitivity of BCa cells by targeting COX-2/PGE2. The mechanism may involve regulating STAT3 phosphorylation, DNA damage response protein activation, and expression of MMP2/MMP9.

Synergetic approaches of fucoidan and trabectedin complex coated PLGA nanoparticles effectively suppresses proliferation and induce apoptosis for the treatment on non-small cell lung cancer.

Traditional methods of treating lung cancer have not been very effective, contributing to the disease's high incidence and death rate. As a result, Fn/Tn-PLGA NPs, a novel directed fucoidan and trabectedin complex loaded PLGA nanoparticle, were produced to investigate the role of developing therapeutic strategies for NSCLC and A549 cell lines. Quantitative real-time polymerase chain reaction was used to examine protein expression and mRNA expression, respectively. Protein activity was knocked down using specific inhibitors and short disrupting RNA transfection. Lastly, cancer cell lines H1299 and A549 were subjected to an in vitro cytotoxicity experiment. Commercial assays were used to assess the levels of cell viability, ROS and proliferation found that Fn/Tn-PLGA NPs effectively killed lung cancer cells. To examine cell death, annexin flow cytometry was employed. In addition, a scratch-wound assay was conducted to assess the migration effects of Fn/Tn-PLGA NPs in a laboratory setting. Finally, PLGA NPs covered with a mix of fucoidan and trabectedin could be a good vehicle for targeting cancerous tissues with chemotherapeutic drugs.

Down-regulation of autophagy-associated protein increased acquired radio-resistance bladder cancer cells sensitivity to taxol.

BACKGROUND: As a bladder-preserving therapy, radiation therapy (RT) has been widely used in the treatment of bladder cancer (BCa) and made great progress in the past few decades. However, some BCa patients have low RT responsiveness and local recurrence rate after RT could reach 50%. Acquired radio-resistance (ARR) is one of the important reasons for the failure of RT. Unfortunately, these ARR cells also lack sensitivity to chemotherapy and cause tumor recurrence and metastasis. PURPOSE: To build ARR-phenotype BCa cell model, discuss the possible molecular mechanism of ARR and find effective target molecules to overcome ARR. MATERIALS AND METHODS: Five thousand six hundred and thirty-seven cells were subjected 30 times to 2 Gy of γ-rays and the surviving cells were called 5637R. Colony formation and MTT assay were applied to evaluate cells sensitivity to ionizing radiation (IR) and anti-neoplastic agents, respectively. Cells abilities of migration and invasion were determined using transwell method. Quantitative real-time polymerase chain reaction (RT-qPCR) and western blot (WB) were respectively utilized to compare the difference of gene and protein expression between 5637 and 5637R cells. Molecule inhibitors and small interfering RNA (siRNA) systems were employed to decrease the expression of target proteins, respectively. RESULTS: BCa cells survived from fractionated irradiation (FI) exhibited tolerance to both IR and chemotherapy drugs. These ARR cells (5637R) had elevated migration and invasion abilities, accompanied by increased expression of epithelial mesenchymal transition (EMT)-related transcription factors (ZEB1/Snail/Twist). Moreover, 5637R cells showed enhanced cancer stem cell (CSC)-like characteristics with activated KMT1A-GATA3-STAT3 circuit, a newly reported self-renewal pathway of human bladder cancer stem cell (BCSC). Combined with Kaplan-Meier's analysis, we speculated that GATA3/MMP9/STAT3 could be an effective molecular panel predicting poor prognosis of BCa. In order to enhance the sensitivity of resistant cells to radiation, we introduced ERK inhibitor (FR 180204) and STAT3 inhibitor (S3I-201). However, both of them could not enhance ARR cells response to IR. On the other hand, siRNAs were respectively implemented to inhibit the expression of endogenous Beclin1 and Atg5, two important autophagy-related genes, in BCa cells, which significantly increased 5637R cells death upon taxol exposing. Similarly, chloroquine (CQ), a classic autophagy inhibitor, enhanced the cytotoxicity of taxol only on 5637R cells. CONCLUSIONS: Long-term FI treatment is an effective method to establish the ARR-phenotype BCa cell model, by enriching BCSCs and enhancing cells migration and invasion. Both inhibiting the expression of autophagy-related proteins and using autophagy inhibitor can increase the sensitivity of ARR cells to taxol, suggesting that autophagy may play an important role in ARR cells chemical tolerance.

Clinicopathological Characteristics of Breast Ductal Carcinoma In Situ: An Analysis of Chinese Population of 617 Patients.

BACKGROUND: The purpose of this study was to describe the clinicopathological characteristics of breast DCIS in Chinese women and compare with that of patients in western countries. METHOD: From December 2005 to December 2015, 617 women diagnosed with pure DCIS after surgery at our institution were enrolled, and the clinicopathological characteristics were described. RESULTS: In this study, the percentage of patients detected on screening, diagnosed at ≤50 years of age, with tumor size ≤2.0 cm, and with low-intermediate grade was 39.4%, 56.7%, 72.6%, and 77.4%, respectively, as compared to 50-80%, 20-30%, 70-90%, and 40-60% in published reports from western countries. The percentage of ER-positive patients was 76.3% in this study, which is similar to the mean expression rate of ER (mean: 68.7%, range: 49-96.6%) reported previously. CONCLUSIONS: The clinicopathological characteristics of Chinese DCIS patients include less detection on screening, younger age at diagnosis, and more low-intermediate nuclear grade.

STAT3 enhances radiation-induced tumor migration, invasion and stem-like properties of bladder cancer.

Bladder cancer (BCa) is the most common cancer of the human urinary system, and is associated with poor patient prognosis and a high recurrence rate. Cancer stem cells (CSCs) are the primary cause of tumor recurrence and metastasis, possessing self­renewal properties and resistance to radiation therapy. Our previous studies indicated that phosphorylated signal transduction and transcription activator 3 (STAT3) may be a potential biomarker to predict radiation tolerance and tumor recurrence in patients with BCa, following conventional radiotherapy. The aim of the present study was to investigate the underlying mechanism of STAT3 in the radio­resistance of BCa cells. It was found that fractionated irradiation promoted the activation of two STAT3­associated CSCs signaling pathways in BCa cells, namely suppressor of variegation 3­9 homolog 1/GATA binding protein 3/STAT3 and Janus kinase 2/STAT3. Surviving cells exhibited elevated migratory and invasive abilities, enhanced CSC­like characteristics and radio­resistance. Furthermore, knockdown of STAT3 expression or inhibition of STAT3 activation markedly decreased the self­renewal ability and tumorigenicity of radiation­resistant BCa cells. Kaplan­Meier analysis revealed that decreased STAT3 mRNA levels were associated with increased overall survival times in patients with BCa. Taken together, these data indicated that STAT3 may be an effective therapeutic target for inhibiting the progression, metastasis and recurrence of BCa in patients receiving radiotherapy.

Long term exposure to γ­rays induces radioresistance and enhances the migration ability of bladder cancer cells.

In Western countries, bladder cancer (BCa) is one of the most common types of malignancy, with highest incidence rate among urinary system malignancies. At present, as a strategy to preserve the quality and function of the bladder tissues, external­beam radiotherapy has been applied to an increasing number of patients with BCa, particularly the elderly and those in poor health. The onset of acquired radioresistance (ARR) during radiotherapy notably reduces the effectiveness of radiotherapy; however, the cause of ARR is unclear and remains untreatable. In order to investigate the mechanism of ARR, clinical fractionated irradiation was mimicked in vitro to obtain radioresistant BCa cells, 5637R, in the present study. Compared with the parental cell line, 5637R cells exhibited an increased distribution of S phase cells, enhanced migration ability and elevated levels of phosphorylated­STAT3, as determined using flow cytometry, cell migration analysis and western blotting, respectively. Furthermore, 5637R cells exhibited notable increases in extracellular signal­regulated kinase activation in response to ionizing radiation (IR) compared with 5637 cells. In addition, 5637R cells showed an early G2/M arrest within 8 h post­IR (18.23±3.85 vs. 6.42±3.82%, IR cells vs. control cells), whereas 5637 cells were arrested in G2/M phase 24 h post­IR (19.26±2.21 vs. 12.36±4.45%, IR cells vs. control cells). Overall, the present study suggested that phosphorylated STAT3 may be a potential biomarker to predict radioresistance and tumor recurrence in patients with BCa following conventional radiotherapeutic intervention. In addition, co­treatment with ERK inhibitor may be a viable approach to increase the anticancer efficacy of radiotherapy in patients with ARR; however, further investigation is required.

Upregulating DAB2IP expression via EGR-1 inhibition, a new approach for overcoming fractionated-irradiation-induced cross-tolerance to ionizing radiation and mitomycin C in tumor cells.

PURPOSE: To evaluate the effect of fractionated irradiation (FI) on tumor cells' sensitivity to ionizing radiation (IR) and antineoplastic drugs, and examine the potential of early growth response-1 (EGR-1) inhibition to sensitize tumor cells to IR. MATERIALS AND METHODS: PC3 and HepG2 cells were subjected 10 times to γ-rays at 2 Gy. The surviving cells were named PC3/R and HepG2/R, respectively. The cells' sensitivity to irradiation and chemotherapeutic drugs, including cisplatin (PT), doxorubicin (DOX), mitomycin C (MMC) and 5-fluorouracil (5-FU), were identified by colony formation assay and MMT method, respectively. Quantitative real-time polymerase chain reaction (RT-qPCR) analysis was utilized to compare the difference of gene expression between radioresistant cells and parental cells. The small interfering RNA system was implemented to inhibit endogenous EGR-1 expression in radiation-resistant cells. Western blot was employed to identify the possible mechanism by which EGR-1 regulates cells' radiosensitivity. RESULTS: FI induced cross-resistant to IR and MMC in tumor cells. Along with the reduction of ovarian cancer-2/disabled homolog 2 (DOC-2/DAB2) interactive protein (DAB2IP) expression, EGR-1 gene was upregulated in FI-treated cells. On the other hand, downregulation of EGR-1 gene expression sensitized radioresistant cells to IR accompanied by DAB2IP overexpression and STAT3 inactivation. In addition, NF-κB inhibitor, BAY11-7082 enhanced resistant cells' radiosensitivity and chemosensitivity. CONCLUSIONS: Conventionally FI has a higher risk of forming acquired radioresistance (ARR) in vitro. EGR-1 gene-targeted drug design could be an effective strategy to overcome DAB2IP-dysregulation-induced ARR in tumor patients.