u-0126 and Nasopharyngeal-Neoplasms

Nasopharyngeal carcinoma (NPC) is a common malignancy in southern China and Southeast Asia. NPC frequently metastasizes to the bone in advanced patients resulting in high mortality. The molecular mechanisms for NPC development and cancer-induced bone lesions are unclear. In this study, we firstly determined chemokine receptor CCR2 and CXCR6 expressions in clinical specimens and CNE2, SUNE1, CNE1, and HK1 cell lines. Then, we measured chemokine CCL2 and CXCL16 production in these NPC cell lines by ELISA. Expression levels of these chemokines and their receptors were observed to positively correlate with tumor aggressiveness. Furthermore, U0126 (MEK inhibitor) was used to treat these NPC cell lines. CCL2 and CXCL16 expression levels and cell proliferation were significantly inhibited by U0126 in a dose- and time-dependent manner. Finally, we collected conditioned medium (CM) from NPC cell cultures in the presence of U0126 treatment. When mouse bone marrow non-adherent cells were treated with the CM, the numbers of multinucleated osteoclast formation were dramatically diminished. These results indicate that MEK inhibitor diminishes NPC cell proliferation and NPC-induced osteoclastogenesis via modulating CCL2 and CXCL16 expressions. This study provides novel therapeutic targets such as CCL2/CCR2 and CXCL16/CXCR6 for advanced NPC patients.

Topics: Animals; Bone Marrow Cells; Butadienes; Carcinoma; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Chemokine CCL2; Chemokine CXCL16; Chemokines, CXC; Culture Media, Conditioned; Gene Expression Regulation, Neoplastic; Humans; MAP Kinase Kinase Kinases; Mice; Nasopharyngeal Carcinoma; Nasopharyngeal Neoplasms; Nitriles; Osteoclasts; Protein Kinase Inhibitors; Receptors, Scavenger

Nedaplatin, a cisplatin analog, was developed to reduce the toxicity of cisplatin, whereas it can be cross-resistant with cisplatin in some circumstances. This study aimed to investigate the role of autophagy in nedaplatin induced cell death in cisplatin-resistant nasopharyngeal carcinoma cells. Here, we showed that HNE1/DDP and CNE2/DDP cells were resistant to nedaplatin-induced cell death with reduced apoptotic activity. Nedaplatin treatment resulted in autophagosome accumulation and increased expression of LC3-II, indicating the induction of autophagy by nedaplatin in HNE1/DDP and CNE2/DDP cells. Inhibition of autophagy by Bafilomycin A1 (Baf A1) and 3-Methyladenine (3-MA) remarkably enhanced the antitumor efficacy of nedaplatin in HNE1/DDP and CNE2/DDP cells, suggesting that the resistance to nedaplatin-induced cell death was caused by enhanced autophagy in nedaplatin-resistant NPC cells. Additionally, Baf A1 enhanced reactive oxygen species (ROS) generation and apoptosis induced by nedaplatin in HNE1/DDP cells. Mechanistically, nedaplatin treatment caused activation of ERK1/2 and suppression of Akt/mTOR signaling pathways. While inhibition of ERK1/2 by MEK1/2 inhibitor, U0126, could reduce the expression of LC3-II in nedaplatin-resistant NPC cells. Furthermore, suppression of ROS could inhibit nedaplatin-induced ERK activation in HNE1/DDP cells, indicating that ROS and ERK were involved in nedaplatin-induced autophagy. Together, these findings suggested that autophagy played a cytoprotective role in nedaplatin-induced cytotoxicity of HNE1/DDP and CNE2/DDP cells. Furthermore, our results highlighted a potential approach to restore the sensitivity of cisplatin-resistant nasopharyngeal cancer cells to nedaplatin in combination with autophagy inhibitors.

Topics: Adenine; Antineoplastic Agents; Apoptosis; Autophagy; Butadienes; Carcinoma; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cisplatin; Drug Resistance, Neoplasm; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Humans; Macrolides; Microtubule-Associated Proteins; Nasopharyngeal Carcinoma; Nasopharyngeal Neoplasms; Nitriles; Organoplatinum Compounds; Proto-Oncogene Proteins c-akt; Reactive Oxygen Species; Signal Transduction; TOR Serine-Threonine Kinases

Phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin inhibitor (mTOR) pathway is often constitutively activated in human tumor cells and thus has been considered as a promising drug target. To ascertain a therapeutical approach of nasopharyngeal carcinoma (NPC), we hypothesized NVP-BEZ235, a novel and potent imidazo[4,5-c] quinolone derivative, that dually inhibits both PI3K and mTOR kinases activities, had antitumor activity in NPC. Expectedly, we found that NVP-BEZ235 selectively inhibited proliferation of NPC cells rather than normal nasopharyngeal cells using MTT assay. In NPC cell lines, with the extended exposure, NVP-BEZ235 selectively inhibited proliferation of NPC cells harboring PIK3CA mutation, compared to cells with wild-type PIK3CA. Furthermore, exposure of NPC cells to NVP-BEZ235 resulted in G1 growth arrest by Propidium iodide uptake assay, reduction of cyclin D1and CDK4, and increased levels of P27 and P21 by Western blotting, but negligible apoptosis. Moreover, we found that cisplatin (CDDP) activated PI3K/AKT and mTORC1 pathways and NVP-BEZ235 alleviated the activation by CDDP through dually targeting PI3K and mTOR kinases. Also, NVP-BEZ235 combining with CDDP synergistically inhibited proliferation and induced apoptosis in NPC cells. In CNE2 and HONE1 nude mice xenograft models, orally NVP-BEZ235 efficiently attenuated tumor growth with no obvious toxicity. In combination with NVP-BEZ235 and CDDP, there was dramatic synergy in shrinking tumor volumes and inducing apoptosis through increasing Noxa, Bax and decreasing Mcl-1, Bcl-2. Based on the above results, NVP-BEZ235, which has entered phase I/II clinical trials in patients with advanced solid tumors, has a potential as a monotherapy or in combination with CDDP for NPC treatment.

Topics: Animals; Apoptosis; Blotting, Western; Butadienes; Carcinoma; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cisplatin; Enzyme Inhibitors; Female; Humans; Imidazoles; Male; Mice; Mice, Inbred BALB C; Nasopharyngeal Carcinoma; Nasopharyngeal Neoplasms; Nitriles; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Quinolines; Signal Transduction; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Mitotic serine/threonine kinase Aurora-A (Aur-A) plays a critical role in regulating centrosome segregation and spindle assemble. Aur-A overexpression causes excessive centrosome duplication and abnormal spindle structure, leading to tumor malignant progression. Here, we investigated Aur-A expression in nasopharyngeal carcinoma (NPC) and the association between Aur-A and NPC invasiveness. We showed that overexpression of Aur-A in tumor tissues was correlated with cranial bone invasion and clinical stage in NPC patients. Suppression of Aur-A by either selective Aurora inhibitory VX-680 or small-interfering RNA caused G(2)/M arrest and apoptotic cell death in NPC CNE-2 cells. Significantly, inhibition of Aur-A suppressed CNE-2 cell invasion and restored membrane expression of epithelial markers, E-cadherin and beta-catenin, suggesting a reversed epithelial-mesenchymal transition process in cancer cells. In addition, we found that Aur-A-regulated epithelial-mesenchymal transition and invasion were mediated by mitogen-activated protein kinase (MAPK) phosphorylation. Moreover, suppression of MAP kinase by small-interfering RNA or its upstream MEK1/2-selective inhibitor U0126 abrogated cell invasion enhanced by Aur-A overexpression. On the other hand, forced overexpression of constitutively active form of MEK1/2, MEK2DD, in CNE-2 cancer cells rescued cell invasive ability suppressed by VX-680-imposed Aur-A inhibition. Our results indicated that Aur-A acted through a downstream MAP kinase pathway to promote epithelial-mesenchymal transition and invasiveness in nasopharyngeal tumorigenesis. Small chemical inhibitor VX-680 may offer as a promising molecular targeting agent in human NPC.

Topics: Apoptosis; Aurora Kinases; beta Catenin; Butadienes; Cadherins; Cell Line, Tumor; Down-Regulation; Epithelium; Humans; MAP Kinase Signaling System; Mesoderm; Middle Aged; Mitogen-Activated Protein Kinases; Nasopharyngeal Neoplasms; Neoplasm Invasiveness; Neoplasm Staging; Nitriles; Phosphorylation; Piperazines; Protein Serine-Threonine Kinases; RNA, Small Interfering