Human Pharyngeal Squamous Carcinoma Research Articles

Fucoxanthin Inhibits the Proliferation and Metastasis of Human Pharyngeal Squamous Cell Carcinoma by Regulating the PI3K/Akt/mTOR Signaling Pathway.

Human pharyngeal squamous cell carcinoma (HPSCC) is the most common malignancy in the head and neck region, characterized by high mortality and a propensity for metastasis. Fucoxanthin, a carotenoid isolated from brown algae, exhibits pharmacological properties associated with the suppression of tumor proliferation and metastasis. Nevertheless, its potential to inhibit HPSCC proliferation and metastasis has not been fully elucidated. This study represents the first exploration of the inhibitory effects of fucoxanthin on two human pharyngeal squamous carcinoma cell lines (FaDu and Detroit 562), as well as the mechanisms underlying those effects. The results showed dose-dependent decreases in the proliferation, migration, and invasion of HPSCC cells after fucoxanthin treatment. Further studies indicated that fucoxanthin caused a significant reduction in the expression levels of proteins in the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mechanistic target of rapamycin (mTOR) pathway, as well as the downstream proteins matrix metalloproteinase (MMP)-2 and MMP-9. Specific activators of PI3K/AKT reversed the effects of fucoxanthin on these proteins, as well as on cell proliferation and metastasis, in FaDu and Detroit 562 cells. Molecular docking assays confirmed that fucoxanthin strongly interacted with PI3K, AKT, mTOR, MMP-2, and MMP-9. Overall, fucoxanthin, a functional food component, is a potential therapeutic agent for HPSCC.

Targeting EIF3C to suppress the development and progression of nasopharyngeal carcinoma.

Nasopharyngeal carcinoma occurs in many parts of the pars nasalis pharyngis, and the pathological type is mainly squamous cell carcinoma. Because of the special position of nasopharynx, breathing, pronunciation and daily life will be seriously affected. At present, the research direction of nasopharyngeal carcinoma is mainly to explore the law of tumor cell proliferation and migration, study the molecular mechanism, master its biological behavior and clinical significance, try to find therapeutic targets, and further improve the level of tumor treatment. However, the pathologic structure and molecular mechanism of nasopharyngeal carcinoma have not been fully elucidated. In this study, the Lentivirus-mediated EIF3C shRNA vector (L.V-shEIF3C) was constructed to down-regulate the expression of EIF3C in human pharyngeal squamous carcinoma cell FaDu and the human nasopharyngeal carcinoma cell 5-8F, it was found that down-regulation of EIF3C could significantly inhibit the cell proliferation, promote cell apoptosis, induce cell cycle arrest, and inhibit the formation and growth of tumors in mouse models. This study provides strong evidence that EIF3C is a key gene driving the development and progression of head and neck cancer, which is of great significance for the diagnosis, prognosis or treatment of tumors, suggesting that EIF3C may become a valuable therapeutic development and intervention target.

Discovery of a potent cytotoxic agent that promotes G2/M phase cell cycle arrest and apoptosis in a malignant human pharyngeal squamous carcinoma cell line.

Squamous cell carcinoma is the major form of malignancy that arises in head and neck cancer. The modest improvement in the 5‑year survival rate underpins its complex etiology and provides the impetus for the discovery of new therapeutics. The present study describes the discovery of an indole‑based small molecule(24a) that was a potent cytotoxic agent with antiproliferative and pro‑apoptotic properties against a pharyngeal carcinoma cell line, Detroit562, effectively killing the cells at a half‑maximal inhibitory concentration of 0.03µM, as demonstrated using cell proliferation studies. The antiproliferative property of 24a was demonstrated by its ability to promote G2/M blockade, as assessed by cell cycle analysis using flow cytometry and the monitoring of real‑time cell cycle progression by the fluorescence ubiquitination‑based cell cycle indicator. This pro‑apoptotic property is supported by the promotion of TUNEL‑staining and increase in the activities of caspases‑3/7 and ‑6, in addition to the expression of death receptors and the cleavage of poly (ADP‑ribose) polymerase1 protein as demonstrated by western blotting. Given that Detroit562 lacks functional p53, it is suggested that 24a acts independently of the tumor suppressor.

Determination of Cytisine and N-Methylcytisine from Selected Plant Extracts by High-Performance Liquid Chromatography and Comparison of Their Cytotoxic Activity

Quinolizidine alkaloids exhibit various forms of biological activity. A lot of them were found in the Leguminosae family, including Laburnum and Genista. The aim of the study was the optimization of a chromatographic system for the analysis of cytisine and N-methylcytisine in various plant extracts as well as an investigation of the cytotoxic activities of selected alkaloids and plant extracts obtained from Laburnum anagyroides, Laburnum anagyroides L. quercifolium, Laburnum alpinum, Laburnum watereri, Genista germanica, and Genista tinctoria against various cancer cell lines. The determination of investigated compounds was performed by High Performance Liquid Chromatography with Diode Array Detection (HPLC-DAD), while High Performance Liquid Chromatography coupled with Quadrupole Time-of-Flight–Mass Spectrometry (HPLC-QTOF-MS) was applied for the qualitative analysis of plant extracts. The retention, separation selectivity, peaks shape, and systems efficiency obtained for cytisine and N-methylcytisine in different chromatographic systems were compared. The application of columns with alkylbonded and phenyl stationary phases led to a very weak retention of cytisine and N-methylcytisine, even when the mobile phases containing only 5% of organic modifiers were used. The strongest retention was observed when hydrophilic interaction chromatography (HILIC) or especially when ion exchange chromatography (IEC) were applied. The most optimal system in terms of alkaloid retention, peak shape, and system efficiency containing an strong cation exchange (SCX) stationary phase and a mobile phase consisted of 25% acetonitrile and formic buffer at pH 4.0 was applied for investigating alkaloids analysis in plant extracts. Cytotoxic properties of the investigated plant extracts as well as cytisine and N-methylcytisine were examined using human tongue squamous carcinoma cells (SCC-25), human pharyngeal squamous carcinoma cells (FaDu), human triple-negative breast adenocarcinoma cell line (MDA-MB-231), and human breast adenocarcinoma cell line (MCF-7). The highest cytotoxic activity against FaDu, MCF-7, and MDA-MB cancer cell lines was observed after applying the Genista germanica leaves extract. In contrast, the highest cytotoxic activity against SCC-25 cell line was obtained after treating with the seed extract of Laburnum watereri. The investigated plant extracts exhibit significant cytotoxicity against the tested human cancer cell lines and seem to be promising for further research on its anticancer activity.

Moscatilin Induces Apoptosis in Human Head and Neck Squamous Cell Carcinoma Cells via JNK Signaling Pathway.

Dendrobii Herba is an herbal medicine that uses the stems of Dendrobium species (Orchidacea). It has been traditionally used to treat fever, hydrodipsomania, stomach disorders, and amyotrophia. In our previous study, a bibenzyl compound, moscatilin, which is isolated from Dendrobii Herba, showed potent cytotoxicity against a FaDu human pharyngeal squamous carcinoma cell line. Prompted by this finding, we performed additional studies in FaDu cells to investigate the mechanism of action. Moscatilin induced FaDu cell death by using 5 μM of concentration and by mediating apoptosis, whereas cell proliferation following treatment with 1 μM of moscatilin was not suppressed to the same levels as by the anti-cancer agent, cisplatin. Apoptosis-related protein expression (cleaved caspase-8, cleaved caspase-7, cytochrome c, cleaved caspase-9, cleaved caspase-3, and poly (ADP-ribose) polymerase (PARP) was increased by treating with 5 μM of moscatilin. This suggests that moscatilin-mediated apoptosis is associated with the extrinsic and intrinsic apoptotic signaling pathways. In addition, moscatilin-induced apoptosis was mediated by the c-Jun N-terminal kinase (JNK) signaling pathway. Overall, this study identified additional biological activity of moscatilin derived from natural products and suggested its potential application as a chemotherapeutic agent for the management of head and neck squamous cell carcinoma.

Mechanisms of different response to ionizing irradiation in isogenic head and neck cancer cell lines

BackgroundTreatment options for recurrent head and neck tumours in the previously irradiated area are limited, including re-irradiation due to radioresistance of the recurrent tumour and previous dose received by surrounding normal tissues. As an in vitro model to study radioresistance mechanisms, isogenic cells with different radiosensitivity can be used. However, they are not readily available. Therefore, our objective was to establish and characterize radioresistant isogenic human pharyngeal squamous carcinoma cells and to evaluate early radiation response in isogenic parental, radioresistant and radiosensitive cells.MethodsRadioresistant cells were derived from parental FaDu cells by repeated exposure to ionizing radiation. Radiosensitivity of the established isogenic radioresistant FaDu-RR cells was evaluated by clonogenic assay and compared to isogenic parental FaDu and radiosensitive 2A3 cells. Additional phenotypic characterization of these isogenic cells with different radiosensitivity included evaluation of chemosensitivity, cell proliferation, cell cycle, radiation-induced apoptosis, resolution of DNA double-strand breaks, and DNA damage and repair signalling gene expression before and after irradiation.ResultsIn the newly established radioresistant cells in response to 5 Gy irradiation, we observed no alteration in cell cycle regulation, but delayed induction and enhanced resolution of DNA double-strand breaks, lower induction of apoptosis, and pronounced over-expression of DNA damage signalling genes in comparison to parental cells. On the other hand, radiosensitive 2A3 cells were arrested in G2/M-phase in response to 5 Gy irradiation, had a prominent accumulation of and slower resolution of DNA double-strand breaks, and no change in DNA damage signalling genes expression.ConclusionsWe concluded that the emergence of the radioresistance in the established radioresistant isogenic cells can be at least partially attributed to the enhanced DNA double-strand break repair, altered expression of DNA damage signalling and repair genes. On the other hand, in radiosensitive isogenic cells the reduced ability to repair a high number of induced DNA double-strand breaks and no transcriptional response in DNA damage signalling genes indicate on a lack of adaptive response to irradiation. Altogether, our results confirmed that these isogenic cells with different radiosensitivity are an appropriate model to study the mechanisms of radioresistance.

Comparison of Anticancer Activity and HPLC-DAD Determination of Selected Isoquinoline Alkaloids from Thalictrum foetidum, Berberis sp. and Chelidonium majus Extracts.

Background: Plants are an important origin of natural substances that the raw material for various pharmaceutical and therapeutic applications due to the presence of phytochemicals, such as alkaloids. Alkaloids, which are found in different plant species, possess numerous biological activities. Some alkaloids have strong cytotoxic effects on various cancer cells. The search for new drugs to treat various cancers is one of the most important challenges of modern scientific research. Objective: This study aimed to investigate of cytotoxic activity of extracts that were obtained from Chelidonium Majus; Berberis sp.; Thalictrum foetidum containing various alkaloids on selected cancer cell lines. The aim was also the quantification of selected alkaloids in the investigated extracts by HPLC. Methods: The analysis of alkaloids contents were performed while using HPLC in reversed phase (RP) mode using Polar RP column and mobile phase containing acetonitrile, water, and ionic liquid. The cytotoxic effect of the tested plant extracts and respective alkaloids’ standards were examined while using human pharyngeal squamous carcinoma cells (FaDu), human tongue squamous carcinoma cells (SCC-25), human breast adenocarcinoma cell line (MCF-7), and human triple-negative breast adenocarcinoma cell line (MDA-MB-231). Conclusion: All of the investigated plant extracts possess cytotoxic activity against cancer cell lines: FaDu, SCC-25, MCF-7, and MDA-MB-231. The highest cytotoxic activity against FaDu and MDA-MB-231 cells was observed for Chelidonium majus root extract, while the highest cytotoxic activity against SCC-25 and MCF-7 cells was estimated for the Thalictrum foetidum root extract. There obtained significant differences in the cytotoxic activity of extracts that were obtained from the roots and herbs of Chelidonium majus and Thalictrum foetidum. Based on these results, investigated plant extracts can be recommended for further investigations of anticancer activity.

Citrate-Induced p85α⁻PTEN Complex Formation Causes G2/M Phase Arrest in Human Pharyngeal Squamous Carcinoma Cell Lines.

Citrate is a key intermediate of the tricarboxylic acid cycle and acts as an allosteric signal to regulate the production of cellular ATP. An elevated cytosolic citrate concentration inhibits growth in several types of human cancer cells; however, the underlying mechanism by which citrate induces the growth arrest of cancer cells remains unclear. The results of this study showed that treatment of human pharyngeal squamous carcinoma (PSC) cells with a growth-suppressive concentration of citrate caused cell cycle arrest at the G2/M phase. A coimmunoprecipitation study demonstrated that citrate-induced cell cycle arrest in the G2/M phase was associated with stabilizing the formation of cyclin B1–phospho (p)-cyclin-dependent kinase 1 (CDK1) (Thr 161) complexes. The citrate-induced increased levels of cyclin B1 and G2/M phase arrest were suppressed by the caspase-3 inhibitor Ac-DEVD-CMK and caspase-3 cleavage of mutant p21 (D112N). Ectopic expression of the constitutively active form of protein kinase B (Akt1) could overcome the induction of p21 cleavage, cyclin B1–p-CDK1 (Thr 161) complexes, and G2/M phase arrest by citrate. p85α–phosphatase and tensin homolog deleted from chromosome 10 (PTEN) complex-mediated inactivation of Akt was required for citrate-induced G2/M phase cell cycle arrest because PTEN short hairpin RNA or a PTEN inhibitor (SF1670) blocked the suppression of Akt Ser 473 phosphorylation and the induction of cyclin B1–p-CDK1 (Thr 161) complexes and G2/M phase arrest by citrate. In conclusion, citrate induces G2/M phase arrest in PSC cells by inducing the formation of p85α–PTEN complexes to attenuate Akt-mediated signaling, thereby causing the formation of cyclin B1–p-CDK1 (Thr 161) complexes.

Golgi integral membrane protein 4 manipulates cellular proliferation, apoptosis, and cell cycle in human head and neck cancer.

The incidence of global head and neck cancer has increased markedly in the last 10 years, and its prognosis is poor, which seriously endangers people’s life and health. At present, there are few studies on its pathogenesis. Golgi integral membrane protein 4 (GOLIM4) is a major member of the Golgi apparatus transporter complex, and its role in tumor is unclear. The present study found that GOLIM4 was the key target protein downstream of stromal interaction molecule 1 (STIM1), which can inhibit the proliferation of head and neck cancer cells FaDu (human pharyngeal squamous carcinoma cell) and Tca-8113 (human tongue squamous carcinoma cell) with knockdown of GOLIM4 by lentivirus. And the decreased expression of GOLIM4 induced cellular apoptosis. Further experiments revealed that FaDu cell cycle progression was changed after GOLIM4 silence, G1 phase arrest and the number of G2/M cells decreased significantly. It was also found that the cells in S-phase decreased markedly after GOLIM4 was knocked down compared with the control group by 5-bromo-2′-deoxyuridine (BrdU) incorporation experiment. In conclusion, we found that GOLIM4, as the target gene downstream of STIM1, inhibited the proliferation of head and neck cancer, promoted apoptosis, and regulated cell cycle progression, and GOLIM4 is a novel oncogene in head and neck cancer and might help in developing promising targetted therapies for head and neck cancer patients.

Adenosine induces intrinsic apoptosis via the PI3K/Akt/mTOR signaling pathway in human pharyngeal squamous carcinoma FaDu cells.

Adenosine has been identified to occur abundantly intra-and extracellularly, and to exert diverse biological functions, including the suppression of cell proliferation and the induction of apoptosis. Adenosine has been reported to induce apoptosis in several cancer cell lines; however, to the best of our knowledge, the effect of adenosine on head and neck cancer cells has not been investigated. Therefore, the purpose of the present study was to evaluate whether adenosine exerts any anticancer effect via induction of apoptosis in human pharyngeal squamous carcinoma FaDu cells. An MTT assay demonstrated that adenosine-treated FaDu cells inhibited a dose-dependent rate of cell growth, whereas human oral keratinocytes cells were unaffected by adenosine treatment. In addition, A1 and A2a adenosine receptor mRNA was detected in FaDu cells by reverse transcription-polymerase chain reaction, and adenosine-induced FaDu cell death was significantly suppressed by treatment with ATL-444, an antagonist of these receptors. Furthermore, adenosine-induced cell growth inhibition was exerted via apoptosis, as confirmed by the analysis of DNA fragmentation, Hoechst nuclear staining and flow cytometry with Annexin V-fluorescein isothiocyanate and propidium iodide staining. Adenosine was also demonstrated to induce an increase in Bcl-associated X expression, a decrease in B-cell lymphoma 2 expression, the release of cytochrome c from mitochondria, and the activation of caspase-3, -9 and poly(ADP-ribose) polymerase in FaDu cells. Finally, phosphoinositide 3-kinase (PI3K), RAC serine/threonine-protein kinase (Akt) and mechanistic target of rapamycin (mTOR) phosphorylation was found to be significantly inhibited in adenosine-treated FaDu cells, as was phosphorylation of the mTOR downregulators, S6 kinase β1, eukaryotic translation initiation factor 4E-binding protein 1, and eukaryotic translation initiation factor 4 γ1. Taken together, these results indicate that adenosine induces apoptosis via the mitochondrial intrinsic pathway, and activates caspase-3 and -9 activity via the PI3K/Akt/mTOR signaling pathway.

Licochalcone-E induces caspase-dependent death of human pharyngeal squamous carcinoma cells through the extrinsic and intrinsic apoptotic signaling pathways.

The aim of the present study was to investigate licochalcone-E (Lico-E)-induced apoptosis and the associated apoptotic signaling pathway in FaDu cells, a human pharyngeal squamous carcinoma cell line. Treatment with Lico-E exhibited significant cytotoxicity on FaDu cells in a concentration-dependent manner. The IC50 value of Lico-E in FaDu cells was ~50 µM. Treatment with Lico-E increased the number of dead FaDu cells. Furthermore, chromatin condensation, which is associated with apoptotic cell death, was observed in FaDu cells treated with Lico-E for 24 h. By contrast, Lico-E did not produce cytotoxicity or increase the number of dead cells when applied to human normal oral keratinocytes (hNOKs). Furthermore, chromatin condensation was not observed in hNOKs treated with Lico-E. Treatment with Lico-E increased the expression of Fas ligand and the cleaved form of caspase-8 in FaDu cells. Furthermore, treatment with Lico-E increased the expression of pro-apoptotic factors, including apoptosis regulator BAX, Bcl-2-associated agonist of cell death, apoptotic protease-activating factor 1, caspase-9 and tumor suppressor p53, while decreasing the expression of anti-apoptotic factors, including apoptosis regulator Bcl-2 and Bcl-2-like protein 1 in FaDu cells. The expression of cleaved caspases-3 and poly (ADP-ribose) polymerase was significantly upregulated following treatment with Lico-E in FaDu cells, while Lico-E-induced apoptotic FaDu cell death was partially suppressed by treatment with Z-VAD-FMK, a pan caspase inhibitor. Therefore, Lico-E-induced oral cancer (OC) cell-specific apoptosis is mediated by the death receptor-dependent extrinsic and mitochondrial-dependent intrinsic apoptotic signaling pathways. In conclusion, these data suggested that Lico-E exhibits potential chemopreventive effects and warrants further developed as a chemotherapeutic agent against OC.

Capsaicin-Induced Apoptosis of FaDu Human Pharyngeal Squamous Carcinoma Cells

PurposeTo investigate the anti-tumor effect of capsaicin on human pharyngeal squamous carcinoma cells (FaDu).Materials and MethodsThe expression of apoptosis/cell cycle-related proteins (or genes) was examined by reverse transcriptase-polymerase chain reaction, western blotting and ELISA methods, while the apoptotic cell population, cell morphology and DNA fragmentation levels were assessed using flow cytometry, fluorescence microscopy and agarose gel electrophoresis.ResultsCapsaicin was found to inhibit the growth and proliferation of FaDu cells in a dose- and time-dependent manner. Apoptotic cell death was confirmed by observing increases in nuclear condensation, nuclear DNA fragmentation and sub-G1 DNA content. The observed increase in cytosolic cytochrome c, activation of caspase 3 and PARP (p85) levels following capsaicin treatment indicated that the apoptotic response was mitochondrial pathway-dependent. Gene/protein expression analysis of Bcl-2, Bad and Bax further revealed decreased anti-apoptotic Bcl-2 protein and increased pro-apoptotic Bad/Bax expression. Furthermore, capsaicin suppressed the cell cycle progression at the G1/S phase in FaDu cells by decreasing the expression of the regulators of cyclin B1 and D1, as well as cyclin-dependent protein kinases cdk-1, cdk-2 and cdk-4.ConclusionOur current data show that capsaicin induces apoptosis in FaDu cells and this response is associated with mitochondrial pathways, possibly by mediating cell cycle arrest at G1/S.

Erratum to "Capsaicin-Induced Apoptosis of FaDu Human Pharyngeal Squamous Carcinoma Cells" by Le TD, et al. (Yonsei Med J 2012;53:834-41.)

Erratum to "Capsaicin-Induced Apoptosis of FaDu Human Pharyngeal Squamous Carcinoma Cells" by Le TD, et al. (Yonsei Med J 2012;53:834-41.)

Cytotoxic Bibenzyls, and Germacrane- and Pinguisane-type Sesquiterpenoids from Indonesian, Tahitian and Japanese Liverworts

Cytotoxic bibenzyls, and germacrane- and pinguisane-type sesquiterpenoids have been isolated from unidentified Indonesian and Tahitian Frullania sp. and Japanese Porella perrottetiana by using a combination of chromatographic methods. The structure activity relationship (SAR) study showed that the presence of a phthalide group in bibenzyls, an alpha-methylene-gamma-lactone in germacrane-type sesquiterpenoids, and beta-hydroxycarbonyl in pinguisane-type sesquiterpenoids play an important role in providing cytotoxic activity against both human promyelocytic leukemia (HL-60) and human pharyngeal squamous carcinoma (KB) cell lines. The structure of each isolated compound was elucidated by using spectroscopic methods and the cytotoxicity was determined by using the WST-8 colorimetric assay.