Treatment Of Human Nasopharyngeal Carcinoma Research Articles

Improved Radiotherapy Sensitivity of Nasopharyngeal Carcinoma Cells by miR-29-3p Targeting COL1A1 3’-UTR

BackgroundRadio-resistance is an obstacle to the treatment of human nasopharyngeal carcinoma (NPC). However, how microRNAs (miRNA) are involved in this process remains unclear. In the present study we explored the role and possible molecular mechanism of miR-29a-3p, formerly known as tumor suppressors, in radio-sensitivity of NPC cells.Material/MethodsA radio-resistant sub-cell line, CNE-2R, was established to detect the expression of miR-29a/b/c-3p using qRT-PCR. CCK-8 assay, colony formation assay, and single-cell gel electrophoresis (SCGE) assay were carried out to analyze the radio-sensitivity of NPC cells. qRT-PCR, luciferase reporter, and Western blot experiments were performed to validate the targeting of COL1A1 by miR-29a. Short interference RNAs (siRNAs) were used to investigate whether COL1A1 mediates the radio-sensitizer role of miR-29a. Expression of miR-29a and COL1A1 in radio-resistant NPC tissues was finally determined.ResultsmiR-29a was decreased in the radio-resistant CNE-2R cells. Following a time-course irradiation (IR) exposure, miR-29a exhibited a time-dependent decrease. Cellular experiments confirmed that miR-29a induced radio-sensitivity of CNE-2R cells via suppressing cell viability and enhancing cell apoptosis after IR. We confirmed that COL1A1 is a direct target of miR-29a and can exert radio-resistance effects in NPC cells. We also found that knockdown of COL1A1 inhibits NPC cell viability and sensitivity to IR. Finally, we observed a downregulation of miR-29a in radio-resistant NPC tissues and its decrease was associated with upregulation of COL1A1.ConclusionsmiR-29a is a critical determinant of NPC radio-response for NPC patients, and its induction provides a promising therapeutic choice to elevate NPC radio-sensitivity.

MicroRNA-19b Promotes Nasopharyngeal Carcinoma More Sensitive to Cisplatin by Suppressing KRAS.

MicroRNAs have been reported to play a vital role in diverse biological processes and tumorigenesis. MicroRNA-19b-5p has been observed to be downregulated in other cancers, but the function of microRNA-19b-5p in human nasopharyngeal carcinoma has not been well investigated. In our study, these results demonstrated that microRNA-19b-5p was significantly downregulated in 37 pairs of nasopharyngeal carcinoma tissues when compared to normal tissues. Enforced expression of microRNA-19b-5p inhibited activity of cell proliferation and cell migration of nasopharyngeal carcinoma cancer cells, CNE1 and HNE1. Furthermore, microRNA-19b-5p targeted KRAS proto-oncogene, GTPase in cancer cells. In human clinical specimens, KRAS was higher expressed in cancer tissues when compared with normal tissues, which was inversely correlated with the expression of microRNA-19b-5p. More interestingly, microRNA-19b-5p sensitizes CNE1 cells to cisplatin by inhibiting its target KRAS. Finally, microRNA-19b-5p inhibits tumorigenesis in vivo. Thus, our results investigated that microRNA-19b-5p functioned as a tumor suppressor and indicated its potential application for the treatment of human nasopharyngeal carcinoma in future.

Tetrandrine Induces Apoptosis in Human Nasopharyngeal Carcinoma NPC-TW 039 Cells by Endoplasmic Reticulum Stress and Ca2+/Calpain Pathways.

Tetrandrine is an alkaloid extracted from a traditional China medicine plant, and is considered part of food therapy as well. In addition, it has been widely reported to induce apoptotic cell death in many human cancer cells. However, the mechanism of Tetrandrine on human nasopharyngeal carcinoma cells (NPC) is still questioned. In our study, we examined whether Tetrandrine can induce apoptosis of NPC-TW 039 cells. We found that cell morphology was changed after treatment with different concentrations of Tetrandrine. Further, we indicated that the NPC-TW 039 cells viability decreased in a Tetrandrine dose-dependent manner. We also found that tetrandrine induced cell cycle arrest in G0/G1 phase. Tetrandrine induced DNA condensation by DAPI staining as well. In addition, we found that Tetrandrine induced Ca2+ release in the cytosol. At the same time, endoplasmic reticulum (ER) stress occurred. Then we used western blotting to examine the protein expression which is associated with mitochondria-mediated apoptotic pathways and caspase-dependent pathways. To further examine whether Ca2+ was released or not with Tetrandrine induced-apoptosis, we used the chelator of Ca2+ and showed that cell viability increased. At the same time, caspase-3 expression was decreased. Furthermore, confocal microscopy examination revealed that Tetrandrine induced expression of ER stress-related proteins GADD153 and GRP78. Our results indicate that Tetrandrine induces apoptosis through calcium-mediated ER stress and caspase pathway in NPC-TW 039 cells. In conclusion, Tetrandrine may could be used for treatment of human nasopharyngeal carcinoma in future.

Low temperature plasma induced apoptosis in CNE‐2Z cells through endoplasmic reticulum stress and mitochondrial dysfunction pathways

This study is designed to investigate the effect of low temperature plasma (LTP) on the apoptosis of CNE‐2Z cell (human nasopharyngeal carcinoma cell line) and the potential mechanisms. We find out that LTP provoked the generation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in CNE‐2Z cells and significantly inhibit the cell viability and induce cell apoptosis. Furthermore, the antioxidant N‐acetyl cysteine (NAC) almost completely blocked LTP‐induced apoptosis. LTP triggered mitochondrial and endoplasmic reticulum (ER) stress‐induced apoptotic pathways indicated by significant upregulation of CHOP, p53, and Bax‐to‐Bcl‐2 ratio. This work will provide the molecular theoretical basis of clinical application of LTP for the treatment of human nasopharyngeal carcinoma.

Silver nanoparticles coupled to anti‑EGFR antibodies sensitize nasopharyngeal carcinoma cells to irradiation.

Radiotherapy is the major form of treatment for head and neck carcinoma, a malignant tumour of epithelial origin. The identification of agents, which can be co‑administered in order to sensitize these tumours to radiotherapy, has become a major focus of investigations. In the present study, a novel 20nm nanocomposite, Ag/C225, was constructed, which consisted of silver nanoparticles (AgNPs) conjugated to an epidermal growth factor receptor‑specific antibody (C225). Physical characterization demonstrated that the Ag/C225 nanoparticles were spherical and dispersed well in water. Enzyme‑linked immunosorbent assays showed that the activity of C225 was preserved in the Ag/C225 nanoparticles. The results of 3‑(4,5‑dimethylthiazol‑2‑yl)‑2,5‑diphenyltetrazolium bromide analysis revealed that AgNPs and Ag/C225 inhibited the proliferation of nasopharyngeal carcinoma epithelial (CNE) cells in a dose‑ and time‑dependent manner. Flow cytometry revealed that AgNPs and Ag/C225 induced the apoptosis of CNEs, and abrogated G2 arrest; the latter effect was more marked with Ag/C225 than with AgNPs. Clonogenic assays indicated that AgNPs and Ag/C225 increased the sensitivity of CNEs to irradiation. The sensitizer enhancement ratios were 1.610±0.012 and 1.405±0.033Gy for AgNPs and Ag/C225, respectively. Western blot analysis revealed that combining X‑ray irradiation with either AgNPs or Ag/C225 reduced the expression levels of DNA damage/repair proteins Ku‑70, Ku‑80 and Rad51; Ag/C225 was also more effective than AgNPs in this context. These results indicated that AgNPs and Ag/C225 effectively enhanced CNE cell radiosensitivity invitro. Therefore, these potent agents may be considered for use as radiosensitizers during the treatment of human nasopharyngeal carcinoma.

Inhibition of Notch-1 pathway is involved in rottlerin-induced tumor suppressive function in nasopharyngeal carcinoma cells.

Recent studies have revealed that rottlerin is a natural chemical drug to exert its anti-cancer activity. However, the molecular mechanisms of rottlerin-induced tumor suppressive function have not been fully elucidated. Notch signaling pathway has been characterized to play a crucial role in tumorigenesis. Therefore, regulation of Notch pathway could be beneficial for the treatment of human cancer. The aims of our current study were to explore whether rottlerin could suppress Notch-1 expression, which leads to inhibition of cell proliferation, migration and invasion in nasopharyngeal carcinoma cells. We performed several approaches, such as CTG, Flow cytometry, scratch healing assay, transwell and Western blotting. Our results showed that rottlerin treatment inhibited cell growth, migration and invasion, and triggered apoptosis, and arrested cell cycle to G1 phase. Moreover, the expression of Notch-1 was obvious decreased in nasopharyngeal carcinoma cells after rottlerin treatment. Importantly, overexpression of Notch-1 promoted cell growth and invasion, whereas down-regulation of Notch-1 inhibited cell growth and invasion in nasopharyngeal carcinoma cells. Notably, we found the over-expression of Notch-1 could abrogate the anti-cancer function induced by rottlerin. Strikingly, our study implied that Notch-1 could be a useful target of rottlerin for the prevention and treatment of human nasopharyngeal carcinoma.

Physcion, a naturally occurring anthraquinone derivative, induces apoptosis and autophagy in human nasopharyngeal carcinoma.

Physcion is a major bioactive ingredient in the traditional Chinese medicine Radix et Rhizoma Rhei, which has an anthraquinone chemical structure and exhibits a variety of pharmacological activities including laxative, hepatoprotective, anti-inflammatory, anti-microbial and anti-proliferative effects. In this study we investigated the effect of physcion on human nasopharyngeal carcinoma in vitro and in vivo, as well as the mechanisms underlying the anti-tumor action. The nasopharyngeal carcinoma cell line CNE2 was treated with physcion, and cell viability was detected using MTT and colony formation assays. Flow cytometry was used to assess the cell cycle arrest, mitochondrial membrane potential loss, apoptosis, autophagy and intracellular ROS generation. Apoptotic cell death was also confirmed by a TUNEL assay. The expression of target or marker molecules was determined using Western blotting. The activity of caspase-3, 8, and 9 was detected with an ELISA kit. A xenograft murine model was used to evaluate the in vivo anti-tumor action of physcion, the mice were administered physcion (10, 20 mg·kg-1·d-1, ip) for 30 d. Treatment with physcion (5, 10, and 20 μmol/L) dose-dependently suppressed the cell viability and colony formation in CNE2 cells. Physcion (10 and 20 μmol/L) dose-dependently blocked cell cycle progression at G1 phase and induced both caspase-dependent apoptosis and autophagy in CNE2 cells. Furthermore, physcion treatment induced excessive ROS generation in CNE2 cells, and subsequently disrupted the miR-27a/ZBTB10 axis, resulting in repression of the transcription factor Sp1 that was involved in physcion-induced apoptosis and autophagy. Moreover, physcion-induced autophagy acted as a pro-apoptotic factor, and possibly contributed to physcion-induced apoptosis. In the xenograft murine model, administration of physcion dose-dependently suppressed the tumor growth without affecting the body weight. Furthermore, the anti-tumor effects of physcion were correlated with downregulation of Sp1 and suppression of miR-27a in the tumor tissues. Physcion induces apoptosis and autophagy in human nasopharyngeal carcinoma by targeting Sp1, which was mediated by ROS/miR-27a/ZBTB10 signaling. The results suggest that physcion is a promising candidate for the treatment of human nasopharyngeal carcinoma.

Effusanin E suppresses nasopharyngeal carcinoma cell growth by inhibiting NF-κB and COX-2 signaling.

Rabdosia serra is well known for its antibacterial, anti-inflammatory and antitumor activities, but no information has been available for the active compounds derived from this plant in inhibiting human nasopharyngeal carcinoma (NPC) cell growth. In this study, we isolated and purified a natural diterpenoid from Rabdosia serra and identified its chemical structure as effusanin E and elucidated its underlying mechanism of action in inhibiting NPC cell growth. Effusanin E significantly inhibited cell proliferation and induced apoptosis in NPC cells. Effusanin E also induced the cleavage of PARP, caspase-3 and -9 proteins and inhibited the nuclear translocation of p65 NF-κB proteins. Moreover, effusanin E abrogated the binding of NF-κB to the COX-2 promoter, thereby inhibiting the expression and promoter activity of COX-2. Pretreatment with a COX-2 or NF-κB-selective inhibitor (celecoxib or ammonium pyrrolidinedithiocarbamate) had an additive effect on the effusanin E-mediated inhibition of proliferation, while pretreatment with an activator of NF-κB/COX-2 (lipopolysaccharides) abrogated the effusanin E-mediated inhibition of proliferation. Effusanin E also significantly suppressed tumor growth in a xenograft mouse model without obvious toxicity, furthermore, the expression of p50 NF-κB and COX-2 were down-regulated in the tumors of nude mice. These data suggest that effusanin E suppresses p50/p65 proteins to down-regulate COX-2 expression, thereby inhibiting NPC cell growth. Our findings provide new insights into exploring effusanin E as a potential therapeutic compound for the treatment of human nasopharyngeal carcinoma.

SZ-685C exhibits potent anticancer activity in both radiosensitive and radioresistant NPC cells through the miR-205-PTEN-Akt pathway

Radioresistance is a major obstacle to the treatment of human nasopharyngeal carcinoma (NPC). Emerging evidence has demonstrated that miRNAs are involved in cancer therapy resistance. Our research group established the radioresistant NPC cell line CNE2R derived from the CNE2 cell line, and demonstrated that irradiation-induced miR-205 determined the resistance of NPC through directly targeting PTEN. However, specific inhibitors targeting miRNAs are largely undetermined. SZ-685C was expected to abrogate the radioresistance of CNE2 cells through the miR-205‑PTEN-Akt pathway. SZ-685C exhibited a similar cytotoxic effect on both cell lines, and we demonstrated that both intrinsic and extrinsic pathways were activated by SZ-685C in the cell lines. Importantly, the miR-205-PTEN-Akt pathway was the key cell signaling pathway activated in the CNE2R cells upon SZ-685C treatment; however, the Stat3-Jab1-p27 pathway might participate in the pro-apoptotic effect in CNE2 cells but not in CNE2R cells. SZ-685C is a promising anticancer agent for treatment of NPC, and it exhibited pro-apoptotic activity in both radiosensitive and radioresistant NPC cells. Although the mechanisms between the two cell lines were not identical, the pro-apoptotic effects were similar between the two cell lines.

Antiproliferative potential of Artemisia capillaris polysaccharide against human nasopharyngeal carcinoma cells

One water-soluble polysaccharide fraction (WACP) was purified from the Artemisia capillaris by DEAE-cellulose anion-exchange and Sephacryl S-400 gel-permeation chromatography. Based on gas chromatography (GC) analysis, WACP was an arabinogalactan (AG), consisting of arabinose and galactose in the ratio of 4:2. Its molecular weight was about 5.8×104Da. The present study demonstrated the anticancer potential of WACP on human nasopharyngeal carcinoma CNE-2 cells and elucidated its possible mechanisms. MTT and flow cytometric analysis identified that WACP had a potent anti-proliferation activity on CNE-2 cells by inducing apoptosis, which was parallel with the morphological changes of CNE-2 cells under inverted microscope. In addition, WACP triggered the apoptosis via the mitochondrial apoptotic pathway, which included the loss of mitochondrial membrane potential, release of cytochrome c and activation of caspase-3/9. Taken together, these results suggest that WACP has anticancer potential in the treatment of human nasopharyngeal carcinoma.

Reactive oxygen species-mediated mitochondrial dysfunction is involved in apoptosis in human nasopharyngeal carcinoma CNE cells induced by Selaginella doederleinii extract

Ethnopharmacological relevanceA traditional Chinese medicine Selaginella doederleinii Hieron has been combined with radiotherapy for the treatment of human nasopharyngeal carcinoma in clinic in China. However, the detailed mechanism of anti-tumor effect of Selaginella doederleinii remains elusive. Aim of the studyThis study was designed to investigate the anti-tumor effect of ethanol extract of Selaginella doederleinii (SDE) on human nasopharyngeal carcinoma and its possible mechanisms. Materials and methodsViability, apoptosis and protein expression of tumor cells were analyzed by MTT, Annexin V staining and Western blot, respectively. Formation of intracellular reactive oxygen species was determined using dichlorofluorescin fluorescence. The in vivo anti-tumor effect was evaluated by measuring tumor volume changes and TUNEL staining in nude mice. ResultsSDE significantly inhibited the growth and induced apoptosis in human nasopharyngeal carcinoma CNE cells. In addition, SDE triggered the mitochondrial/caspase apoptotic pathway indicated by enhanced Bax-to-Bcl-2 ratio, loss of mitochondrial membrane potential, cytochrome c release, and caspase cascade. Moreover, SDE provoked the generation of reactive oxygen species in CNE cells, while the antioxidant N-acetyl cysteine almost completely blocked SDE-induced disruption of mitochondrial membrane potential, caspases activation and apoptosis. Furthermore, a transplantable nude mice model was utilized to estimate the effectiveness of SDE in vivo. The treated mice displayed decreased tumor size, which was associated with enhanced apoptotic cell death. ConclusionsThese results, offering solid evidence of the induction of mitochondria-related apoptosis in tumor cells, provide the molecular theoretical basis of clinical application of Selaginella doederleinii for the treatment of human nasopharyngeal carcinoma.

Polo-like kinase 1 siRNA-607 induces mitotic arrest and apoptosis in human nasopharyngeal carcinoma cells

Polo-like kinase (Plk) 1 is overexpressed in many human malignancies including nasopharyngeal carcinoma, indicating its potential as a therapeutic target. Recently, using a simple cellular morphologybased strategy, we have identified several novel effective siRNAs against Plk1 including Plk1 siRNA- 607. In this study, we further investigated the effects of Plk1 siRNA-607 in human nasopharyngeal carcinoma cell line, HNE-1. Real time RT-PCR and Western blot indicated that Plk1 siRNA-607 transfection resulted in a significant inhibition in Plk1 expression in the HNE-1 cells. Furthermore, cell cycle, cell growth and apoptosis analysis clearly indicated that Plk1 siRNA-607 caused a dramatic mitotic cell cycle arrest followed by massive apoptotic cell death, and eventually resulted in a significant decrease in growth and viability of the nasopharyngeal carcinoma cells. Given that Plk1 has been widely accepted as a novel efficient target for cancer therapy, these results suggested that Plk1 siRNA-607 could be further developed for the treatment of human nasopharyngeal carcinoma. Key words: Nasopharyngeal carcinoma, Plk1, RNA silencing, cell cycle, apoptosis.

Anti-angiogenic effects of Hypericin-photodynamic therapy in combination with Celebrex® in the treatment of human nasopharyngeal carcinoma

Photodynamic therapy (PDT) is being investigated as an alternative treatment modality in cancer treatment. It has been shown to induce tumor hypoxia and upregulation of cyclooxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF). The objective of this study was to improve in vivo tumor growth control of nasopharyngeal carcinoma (NPC), treated at a subcurative dosage by using a combination of Hypericin-PDT and COX-2 inhibitor, Celebrex (CX). The effect of an initial CX dose at 6- and 24-h post-PDT was investigated simultaneously. It was observed that hypoxic NPC/CNE2 cells upregulate both COX-2 and VEGF A genes in vitro. In vivo studies, down-regulation of COX-2 and hypoxia inducible factor-1alpha (HIF-1alpha) genes at 24-h post-PDT and bulk tumor ablation at 48-h post-PDT was observed. However, 24-28 days later regrowth was observed. In a combination treatment, 1st CX dose at 6-h post-PDT had the highest tumor control in which tumors were <or=0.52 cm(3) (64.29%, P<0.05). However, the tumors administered with a initial dose of CX at 24-h post-PDT had no tumor control. Co-suppression of COX-2, HIF-1alpha and VEGF A genes were observed in tumors with tumor control. Mice without tumor control that were subjected to therapy had increased human VEGF in serum compared to Hypericin-PDT mice. Thus, suggesting that the time of initial administration of CX post-PDT is an important factor for effective tumor control.