Chenping Zhang

MicroRNAs contribute to the chemoresistance of cisplatin in tongue squamous cell carcinoma lines

MicroRNAs (miRNAs) are small non-coding RNAs that function as negative regulators of gene expression. They are strongly implicated in human cancers, including oral squamous cell carcinoma (OSCC). Evidence for the involvement of miRNAs as important regulators of chemosensitivity and chemoresistance in OSCC is not well understood. In this study, miRNA microarray was firstly used to compare the differential miRNAs levels between the cisplatin-sensitive tongue squamous cell carcinoma line (Tca8113) and its cisplatin-resistant subline (Tca/cisplatin). Three miRNAs of miR-21, -214, and -23a were validated by miRNAs real-time PCR, and intervened by anti-miRNA oligonucleotides (miR-214 and -23a) and pre-miRNA plasmid transfection (miR-21). Further relationship between miR-23a and DNA topoisomerase II beta (TOP2B) on the chemoresistance against cisplatin was studied. There were 19 out of 480 differential miRNAs between the Tca8113 and Tca/cisplatin cells. miR-214 and -23a were found increased as with chemoresistance against cisplatin in the Tca/cisplatin cells while miR-21 was found decreased as with chemosensitivity for cisplatin in the Tca/cisplatin cells. Intervention of these three miRNAs could decrease the chemoresistance against cisplatin in Tca/cisplatin cells. Transfection of anti-miR-23a into the Tca/cisplatin cells could increase the TOP2B protein expression. Our results suggest the existence of differential miRNAs with chemosensitivity and chemoresistance between the cisplatin-sensitive and resistant tongue squamous cell carcinoma lines. miR-21 serves as a chemosensitive miRNA, while miR-214 and -23a serve as chemoresistant miRNAs in the tongue squamous cell carcinoma lines. miR-23a is an up-stream regulator of TOP2B to realize the chemoresistance of cisplatin.

Prognostic significance of Bcl-2 and Bax protein expression in the patients with oral squamous cell carcinoma.

BACKGROUND The aim of this study was to investigate the expression of the apoptosis-inhibitory Bcl-2 protein and the apoptosis-promoting Bax protein and to identify their association with the clinical parameters and prognosis of the patients with oral squamous cell carcinoma (OSCC). METHODS The expression of Bcl-2 and Bax proteins was evaluated by immunohistochemical staining in specimens from 110 patients with OSCC. Every section was scored according to both the percentage of positive staining tumor cells and the staining intensity. The Kaplan-Meier test and Cox proportional hazards regression analysis were performed to assess the correlation between the protein levels and the long survival rate of patients. The association between Bax, Bcl-2 immunoexpression and clinicopathologic variables was analyzed with the chi-square test and non-parametric analysis. The Bcl-2 and Bax immunoexpression in 20 oral mucosa samples were also investigated as normal control. RESULTS The results showed that the 5-year survival rate was significantly higher in the patients with the ratio of Bcl-2/Bax <or= 1 than in those with Bcl-2/Bax > 1 (76.79 +/- 6.69% vs. 59.26 +/- 6.69%, P = 0.0489). Bax immunoreactivity was significantly correlated with histological grading and lymph node metastasis. Univariate analysis indicated that the ratio of Bcl-2/Bax and lymph node metastasis were two independent factors related to the prognosis. CONCLUSION The ratio of Bcl-2/Bax could be used as an effective biomarker to predict the prognosis of OSCC.

Increased expression of Toll-like receptor-9 has close relation with tumour cell proliferation in oral squamous cell carcinoma

OBJECTIVE Toll-like receptor-9 (TLR-9), a new member of the interleukin-1 receptor superfamily, was recently found to have a high level of expression in many carcinoma specimens. The objective of this study was to examine the TLR-9 expression and its role in tumour cell proliferation in oral squamous cell carcinoma. MATERIALS AND METHODS Western blot and immunohistochemistry were used to detect TLR-9 protein in oral squamous cell carcinoma (OSCC) Tca-8113 cell lines and clinical specimens (n=60). The relationship between TLR-9 expression and clinicopathologic features was analysed. Cell proliferation and inflammatory chemokines secretion were tested by MTT and ELISA methods respectively. RESULTS Results showed that TLR-9 expression level was higher in OSCC tissues than in paired adjacent normal tissues (P<0.01), and the expression level of TLR-9 was significantly associated with tumour size (P=0.001), tumour clinical stage (P=0.003) and Ki-67 expression (P<0.01). In vitro results also suggested that stimulation of Tca-8113 cells with TLR-9 agonist CpG-ODN could significantly increase tumour cell proliferation as well as subsequent IL-1α and IL-6 secretions (P<0.01), which could be partially inhibited by usage of anti-TLR-9 protein. CONCLUSIONS It was therefore hypothesized that increased expression of TLR-9 may be of great value in assessing the development of OSCC, and could be used as a new target for OSCC prevention and therapy in future.

GDF15 is a potential predictive biomarker for TPF induction chemotherapy and promotes tumorigenesis and progression in oral squamous cell carcinoma

BACKGROUND Randomized trials have not shown major survival benefits when induction chemotherapy plus standard therapy is compared with standard therapy alone in patients with oral squamous cell carcinoma (OSCC). Induction chemotherapy is likely to be effective for biologically distinct subgroups and biomarker development may lead to identification of patients whose tumors are likely to respond to a particular treatment. PATIENTS AND METHODS We evaluated immunohistochemical staining for GDF15 in pretreatment biopsy specimens of 230 of 256 OSCC patients who were treated in a prospective, randomized, phase III trial on induction chemotherapy including docetaxel, cisplatin and 5-fluorouracil (TPF). Relationship between GDF15 intervention and cell proliferation, migration, invasion, colony formation and tumorigenicity was analyzed using in vitro and in vivo OSCC models. RESULTS Low GDF15 expression predicted a better survival in OSCC patients, especially overall survival [P = 0.049, hazard ratio (HR) = 0.597] and distant metastasis-free survival (DMFS; P = 0.031, HR = 0.562). cN+ patients with low GDF15 expression benefitted from induction TPF in overall survival (P = 0.039, HR = 0.247) and DMFS (P = 0.039, HR = 0.247), cN- patients with high GDF15 expression benefitted from induction TPF in overall survival (P = 0.019, HR = 0.231), disease-free survival (P = 0.011, HR = 0.281), locoregional recurrence-free survival (P = 0.035, HR = 0.347) and DMFS (P = 0.009, HR = 0.197). Decreased GDF15 expression in OSCC lines significantly inhibited cell proliferation, migration, invasion, colony formation and tumorigenesis through increased phosphorylation of AKT and ERK1/2 (P < 0.05). Likewise, overexpression of GDF15 significantly promoted cell proliferation, migration, invasion and colony formation through decreased phosphorylation of AKT and ERK1/2 (P < 0.05). CONCLUSIONS GDF15 expression can be used as a prognostic biomarker for OSCC, and as a predictive biomarker for benefitting from TPF induction chemotherapy. GDF15 promotes tumorigenesis and progression through phosphorylation of AKT and ERK1/2 in OSCC. The clinical trial in this study was registered with www.ClinicalTrials.gov (NCT01542931).

Neck dissection and post-operative chemotherapy with dimethyl triazeno imidazole carboxamide and cisplatin protocol are useful for oral mucosal melanoma

BackgroundOral mucosal melanoma (OMM) is a clinically rare disease with poor prognosis. Various treatment methods have been investigated with the aim of improving the prognosis. This study aimed to analyze the data of a single institution in the management of OMM.MethodsA total of 78 consecutive OMM patients were included in this retrospective study. The intraoral lesion was treated either by cryotherapy, surgery or both; the neck was treated by neck dissection or observation; post-operative chemotherapy with dimethyl triazeno imidazole carboxamide and cisplatin was performed in some patients. The Kaplan-Meier method was used for statistical analysis.ResultsAmong the 78 patients, there were 50 males and 28 females with an average age of 53.8 years (ranging from 27 to 85 years). The most common sites of OMM were the hard palate and gingiva. The main cause of death in OMM was distant metastasis. No significant difference was found between the intraoral/cervical lesion recurrence/post-operative distant metastasis and the intraoral lesion site/biopsy method/treatment method. The metastasis rate of cervical lymph node was high in the OMM patients, even in the patients with clinically negative necks. Cervical lesion recurrence was correlated with N stage and intraoral lesion recurrence. The survival period was longer in the patients with T3 staging, clinical stage III disease, with post-operative chemotherapy and without post-operative distant metastasis when compared to those patients with T4a staging, clinical stage IV disease, without post-operative chemotherapy and with post-operative distant metastasis.ConclusionsRadical surgery including wide intraoral resection and neck dissection is recommended for OMM patients. Post-operative chemotherapy may also be beneficial for both primary and recurrent OMM patients.

Clinical analysis of Castleman disease (hyaline vascular type) in parotid and neck region

OBJECTIVE The aim of this study was to analyze a single institutions experience in clinical diagnosis, treatment, and prognosis of Castleman disease (hyaline vascular type) in the parotid and neck region. STUDY DESIGN From 2004 to 2008, a total of 10 consecutive patients with Castleman disease (hyaline vascular type) in the parotid and neck region underwent surgery were included in this retrospective study. The preoperative examinations, clinical diagnosis, surgical treatment, and prognosis were recorded and analyzed. RESULTS Of the 10 patients, 4 were males and 6 female; their age ranged from 13 to 54 years with a mean of 26.6 years. The lesion occurred in the parotid region in 3 patients, in the neck region in 5 patients, and in both the parotid and neck regions in 2 patients. Their course of disease ranged from 3 months to 48 months with a mean of 12.5 months; 70% of the patients (7 out of 10) had a course of disease of <12 months. The patients always had no obvious complaint, and the laboratory examinations were almost within the normal limits. Magnetic resonance imaging/angiography were valuable on clinical diagnosis and differential diagnosis. All patients underwent surgical removal of the masses completely. During the follow-up period, which ranged from 9 months to 60 months with a mean of 38.9 months, no recurrence of the lesion occurred, and the quality of life of each patient was good. CONCLUSIONS Castleman disease (hyaline vascular type) in the parotid and neck region is rare, with clinical manifestation and physical examination the same as benign lesions. There is no specific indication in the laboratory tests and imaging examinations; however, magnetic resonance imaging/angiography has potential value on clinical diagnosis and differential diagnosis. Surgical resection is the choice of treatment with good prognosis.

Biomechanical three-dimensional finite element analysis of prostheses retained with/without zygoma implants in maxillectomy patients

The objective of this study was to analyze the stress distribution on zygoma implants and their supporting bones, superstructures and abutments under occlusal loads after maxilla reconstruction with prostheses. Four three-dimensional (3D) finite element models (FEMs) were constructed based on computed tomography (CT) data. One model of normal structure was constructed (Model 1) and three models of defects were simulated and restored with conventional prosthesis, one and two zygoma-implant-retained prostheses respectively (Models 2-4). Vertical and lateral loads of 150 N were loaded and the stresses distribution were observed and compared. The stresses distributed along the three mechanical pillars of maxillofacial parts in Model 1, which concentrated at the superstructure at Model 2. The zygoma implant can reduce the stresses at clasps and abutments in Model 3, while two zygoma implants can well share the stresses of affected side in Model 4. The distribution of stresses on prostheses were more rational with the help of zygoma implants which can share the stresses on the affected side adequately, so that it is fit for the reconstruction of unilateral maxilla defects.

Survivin as a potential early marker in the carcinogenesis of oral submucous fibrosis.

OBJECTIVE Oral submucous fibrosis (OSF) is a chronic precancerous condition. Survivin is one of the inhibitors of apoptosis protein, and is focused on owing to its unique therapeutic and prognostic potential. STUDY DESIGN To determine the potential involvement of survivin in the carcinogenesis of OSF, we analyzed the relationship between the survivin and clinical characteristic. RESULTS Immunohistochemistry was used to show that survivin expression levels were significantly higher in the oral squamous cell carcinoma transformed from OSF group compared with normal group (P < .01) and OSF group (P < .01). In the different stages of OSF, survivin expression exhibited difference as well. Furthermore, Western blotting and reverse-transcription polymerase chain reaction analysis confirmed the increased expression of survivin in the carcinogenesis of OSF. CONCLUSION These results suggest that survivin plays an important role during the malignant transformation of OSF and may provide an indication to early prevention and diagnosis in the progression of OSF.

A covalently bound inhibitor triggers EZH2 degradation through CHIP‐mediated ubiquitination

Enhancer of zeste homolog 2 (EZH2) has been characterized as a critical oncogene and a promising drug target in human malignant tumors. The current EZH2 inhibitors strongly suppress the enhanced enzymatic function of mutant EZH2 in some lymphomas. However, the recent identification of a PRC2‐ and methyltransferase‐independent role of EZH2 indicates that a complete suppression of all oncogenic functions of EZH2 is needed. Here, we report a unique EZH2‐targeting strategy by identifying a gambogenic acid (GNA) derivative as a novel agent that specifically and covalently bound to Cys668 within the EZH2‐SET domain, triggering EZH2 degradation through COOH terminus of Hsp70‐interacting protein (CHIP)‐mediated ubiquitination. This class of inhibitors significantly suppressed H3K27Me3 and effectively reactivated polycomb repressor complex 2 (PRC2)‐silenced tumor suppressor genes. Moreover, the novel inhibitors significantly suppressed tumor growth in an EZH2‐dependent manner, and tumors bearing a non‐GNA‐interacting C668S‐EZH2 mutation exhibited resistance to the inhibitors. Together, our results identify the inhibition of the signaling pathway that governs GNA‐mediated destruction of EZH2 as a promising anti‐cancer strategy.

The secretion of IL-6 by CpG-ODN-treated cancer cells promotes T-cell immune responses partly through the TLR-9/AP-1 pathway in oral squamous cell carcinoma

Increasing evidence suggests that communication between tumor and immune cells can alter the tumor microenvironment in ways that promote tumor development. The purpose of this study was to characterize the immune response elicited by TLR-9-activated OSCC cells, to identify the cytokines involved in the signaling pathway and to elucidate the molecular mechanism of this pathway in OSCC cells. MTS, flow cytometry and ELISA assay were used to evaluate T-cell immune responses, cancer cell proliferation and pro-inflammatory cytokine secretion, respectively. Western blot analysis, EMSA and ChIP assay were employed to detect the activity of the NF-κB and AP-1 signaling pathways. A marked response was observed when T-cells were co-cultured with supernatants from CpG-ODN-treated OSCC cells. This response was characterized by increased CD4+ and CD8+ T-cell proliferation and an increase in IFN-γ production by the CD4+ T-cell population. Treatment of OSCC cells with CpG-ODN resulted in an increase in IL-6 secretion as well as an increase in AP-1 binding activity to the IL-6 promoter. Moreover, blockage of the TLR-9/AP-1 pathway significantly decreased IL-6 expression and T-cell immune response. In human OSCC, the TLR-9 pathway, when stimulated by CpG-ODNs, promotes a T-cell immune response mediated by AP-1-activated IL-6 secretion. Although the complete molecular mechanism has yet to be understood, these findings provide evidence linking tumor cell activities to immune system responses. In addition, the TLR-9/AP-1/IL-6 pathway provides new therapeutic targets for the prevention and treatment of OSCC.