Performance of cone-beam computed tomography (CBCT) in comparison to conventional computed tomography (CT) and magnetic resonance imaging (MRI) for the detection of bone invasion in oral squamous cell cancer (OSCC): a prospective study.

BACKGROUND: Oral squamous carcinoma (OSCC) is often diagnosed at late stages and bone erosion or invasion of the jawbone is frequently present. Computed tomography (CT) and magnetic resonance imaging (MRI) are known to have high diagnostic sensitivities, specificities, and accuracies in detecting these bone affections in patients suffering from OSCC. To date, the existing data regarding the impact of cone-beam computed tomography (CBCT) have been weak. Therefore, this study aimed to investigate whether CBCT is a suitable tool to detect bone erosion or invasion in patients with OSCC. METHODS: We investigated in a prospective trial the impact of CBCT in the diagnosis of bone erosion or invasion in patients with OSCC who underwent surgery. Every participant received a CBCT, CT, and MRI scan during staging. Imaging modalities were evaluated by two specialists in oral and maxillofacial surgery (CBCT) and two specialists in radiology (CT and MRI) in a blinded way, to determine whether a bone affection was present or not. Reporting used the following 3-point system: no bony destruction ("0"), cortical bone erosion ("1"), or medullary bone invasion ("2"). Histological examination or a follow-up served to calculate the sensitivities, specificities, and accuracies of the imaging modalities. RESULTS: Our results revealed high diagnostic sensitivities (95.6%, 84.4%, and 88.9%), specificities (87.0%, 91.7%, and 91.7%), and accuracies (89.5%, 89.5%, and 90.8%) for CBCT, CT, and MRI. A pairwise comparison found no statistical difference between CBCT, CT, and MRI. CONCLUSION: Our data support the routine use of CBCT in the diagnosis of bone erosion and invasion in patients with OSCC as diagnostic accuracy is equal to CT and MRI, the procedure is cost-effective, and it can be performed during initial contact with the patient.

PKM2 Modulation in Head and Neck Squamous Cell Carcinoma.

The enzyme pyruvate kinase M2 (PKM2) plays a major role in the switch of tumor cells from oxidative phosphorylation to aerobic glycolysis, one of the hallmarks of cancer. Different allosteric inhibitors or activators and several posttranslational modifications regulate its activity. Head and neck squamous cell carcinoma (HNSCC) is a common disease with a high rate of recurrence. To find out more about PKM2 and its modulation in HNSCC, we examined a panel of HNSCC cells using real-time cell metabolic analysis and Western blotting with an emphasis on phosphorylation variant Tyr105 and two reagents known to impair PKM2 activity. Our results show that in HNSCC, PKM2 is commonly phosphorylated at Tyrosine 105. Its levels depended on tyrosine kinase activity, emphasizing the importance of growth factors such as EGF (epidermal growth factor) on HNSCC metabolism. Furthermore, its correlation with the expression of CD44 indicates a role in cancer stemness. Cells generally reacted with higher glycolysis to PKM2 activator DASA-58 and lower glycolysis to PKM2 inhibitor Compound 3k, but some were more susceptible to activation and others to inhibition. Our findings emphasize the need to further investigate the role of PKM2 in HNSCC, as it could aid understanding and treatment of the disease.

HGF-Induced PD-L1 Expression in Head and Neck Cancer: Preclinical and Clinical Findings.

Head and neck squamous cell carcinoma (HNSCC) is a widespread disease with a low survival rate and a high risk of recurrence. Nowadays, immune checkpoint inhibitor (ICI) treatment is approved for HNSCC as a first-line treatment in recurrent and metastatic disease. ICI treatment yields a clear survival benefit, but overall response rates are still unsatisfactory. As shown in different cancer models, hepatocyte growth factor/mesenchymal-epithelial transition (HGF/Met) signaling contributes to an immunosuppressive microenvironment. Therefore, we investigated the relationship between HGF and programmed cell death protein 1 (PD-L1) expression in HNSCC cell lines. The preclinical data show a robust PD-L1 induction upon HGF stimulation. Further analysis revealed that the HGF-mediated upregulation of PD-L1 is MAP kinase-dependent. We then hypothesized that serum levels of HGF and soluble programmed cell death protein 1 (sPD-L1) could be potential markers of ICI treatment failure. Thus, we determined serum levels of these proteins in 20 HNSCC patients before ICI treatment and correlated them with treatment outcomes. Importantly, the clinical data showed a positive correlation of both serum proteins (HGF and sPD-L1) in HNSCC patient's sera. Moreover, the serum concentration of sPD-L1 was significantly higher in ICI non-responsive patients. Our findings indicate a potential role for sPD-L1 as a prognostic marker for ICI treatment in HNSCC.

The Influence of Met Receptor Level on HGF-Induced Glycolytic Reprogramming in Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is known to overexpress a variety of receptor tyrosine kinases, such as the HGF receptor Met. Like other malignancies, HNSCC involves a mutual interaction between the tumor cells and surrounding tissues and cells. We hypothesized that activation of HGF/Met signaling in HNSCC influences glucose metabolism and therefore substantially changes the tumor microenvironment. To determine the effect of HGF, we submitted three established HNSCC cell lines to mRNA sequencing. Dynamic changes in glucose metabolism were measured in real time by an extracellular flux analyzer. As expected, the cell lines exhibited different levels of Met and responded differently to HGF stimulation. As confirmed by mRNA sequencing, the level of Met expression was associated with the number of upregulated HGF-dependent genes. Overall, Met stimulation by HGF leads to increased glycolysis, presumably mediated by higher expression of three key enzymes of glycolysis. These effects appear to be stronger in Methigh-expressing HNSCC cells. Collectively, our data support the hypothesized role of HGF/Met signaling in metabolic reprogramming of HNSCC.

MAGE-A11 expression contributes to cisplatin resistance in head and neck cancer.

OBJECTIVE: The objective of this study is to investigate the roles of melanoma-associated antigens (MAGEs) in the cisplatin treatment of head and neck cancer. MATERIALS AND METHODS: We assessed the efficacy of cisplatin in a set of four head and neck cancer cell lines using a crystal violet assay. The MAGE-A expression in all cell lines was measured with RT-qPCR. The correlation between MAGE-A expression and cisplatin efficacy was investigated using Spearman's correlation analysis. Furthermore, we established a cell line with stable overexpression of MAGE-A11 and determined influence on proliferation, cisplatin efficacy and cell apoptosis. In this cell line, the effects of cisplatin were assessed using either crystal violet assays or flow cytometry (Annexin V). RESULTS: For MAGE-A11, we observed the highest correlation (r = 1.000, p = 0.0417) with low cisplatin efficacy. Stable overexpression of MAGE-A11 resulted in no changes in proliferation, but in lower cisplatin cytotoxicity and lower rates of apoptosis. Also, mouse double minute 2 homolog (MDM2) expression was induced by MAGE-A11 overexpression. CONCLUSION: We provide evidence that MAGE-A11 expression contributes to cisplatin resistance in head and neck cancer. CLINICAL RELEVANCE: Our study underscores the negative predictive role of MAGE-A11 expression in head and neck cancer.

The prognostic value of GLUT-1 staining in the detection of malignant transformation in oral mucosa.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) ranks as the sixth most common tumor entity worldwide. Unfortunately, the multimodal treatment consisting of surgery, radiation, and chemotherapy does not show the desired efficacy. The intent of this study was to evaluate the sensitivity and specificity of an oral brush biopsy in combination with glucose transporter (GLUT)-1 staining in identifying premalignant and malignant lesions. METHODS: A total of 72 patients were included in the study, divided into four diagnostic subgroups (24 healthy, 15 carcinoma, 18 leukoplakia, 15 oral lichen planus). Oral brush biopsies were taken and analyzed for GLUT-1 expression by immunocytologic staining. Incisional biopsy served as the gold standard. RESULTS: Twelve (80 %) of the 15 carcinomas, nine (50 %) of the 18 leukoplakia, nine (60 %) of the 15 oral lichen planus, and none of the healthy specimens stained positive for GLUT-1. This resulted in a sensitivity rate of 80 % and a specificity rate of 68.42 %. Diagnostic accuracy was 70.83 % based on the correct diagnoses in 51 of 72 patients. CONCLUSION: An oral brush biopsy can easily be performed throughout the entire oral cavity, is noninvasive, and shows high sensitivity and specificity rates with conventional cytology or computer-assisted analysis. CLINICAL RELEVANCE: The significance of GLUT-1-specific staining with an oral brush biopsy is more limited than expected but could be used as an additional tool in detecting malignant transformation in the oral cavity.

Erlotinib and gefitinib responsiveness in head and neck cancer cell lines--a comparing analysis with cetuximab.

OBJECTIVE: The objective of this study is to examine the efficacy of erlotinib and gefitinib with respect to epidermal growth factor (EGF) and cetuximab response in head and neck cancer cell lines. MATERIALS AND METHODS: Five human head and neck carcinoma cell lines were treated with EGF, cetuximab, erlotinib, and gefitinib, and the effects were measured with a crystal violet assay. The efficacies of cetuximab, erlotinib, and gefitinib in clinically relevant concentrations were statistically analyzed. The expression of the epidermal growth factor receptor (EGFR) and phosphorylation patterns were detected with fluorescence-activated cell sorting (FACS) analysis and western blot analysis. The endogenous production of EGF by the cells was detected with an enzyme-linked immunosorbent assay. Finally, EGFR, KRAS, BRAF, and PI3K mutation analyses were performed. RESULTS: All of the cell lines had a poor or no response to EGF but exhibited distinct EGFR phosphorylation and EGFR expression. Compared to cetuximab, erlotinib and gefitinib demonstrated a greater impact on the majority of the cell lines. The only cell line that showed a concentration-dependent behavior toward EGF and strong EGFR phosphorylation was entirely resistant to cetuximab, erlotinib, and gefitinib. The production of EGF in all cell lines was very low. Mutational analysis of all cell lines revealed wild-type EGFR, KRAS, BRAF, and PI3K. CONCLUSIONS: The prediction of anti-EGFR treatment cannot be based on responsiveness to EGF or EGFR activation. CLINICAL RELEVANCE: Erlotinib and gefitinib show good response in EGF-independent cell lines and might be useful drugs in tumors that are less responsive to cetuximab.

Cytotoxic effects of SMAC-mimetic compound LCL161 in head and neck cancer cell lines.

OBJECTIVES: Head and neck squamous cell carcinoma (HNSCC) is one of the most common tumor entities worldwide. Unfortunately, recent drug developments in other fields of oncology have yielded no efficacy in the treatment of oral squamous cell carcinoma. As a new starting point, we investigated the impact of Fas ligand (FasL) and the SMAC-mimetic compound LCL161 in mono- and combination treatment in HNSCC cell lines. METHODS: Five different cell lines of HNSCC were treated with FasL and LCL161 in mono- and combination treatment. Cytotoxicity was measured via a crystal violet assay. The cell lines were characterized for CD95 (FasL receptor) expression via flow cytometry. The degradation of cellular inhibitor of apoptosis protein 1 (cIAP1) was detected via Western blot. RESULTS: Incubation with FasL led to a significant decrease in three out of five cell lines. Combination treatment with LCL161 enhanced cytotoxicity significantly. Two cell lines were FasL resistant, but one of them could be resensitized with LCL161. In all cell lines, Western blot analysis showed degradation of cIAP1 after LCL161 application. However, one cell line showed only minor vulnerability to the FasL and LCL161 combination. CONCLUSION: This is the first study investigating combination treatment of FasL and LCL161 in head and neck cancer cell lines. Pro-apoptotic effects of the combination were detected in the majority of the cell lines. Interestingly, one of two FasL-resistant cell lines was sensitive to the combination therapy with FasL and LCL161. CLINICAL RELEVANCE: SMAC-mimetic compounds show promising results in the treatment of other tumor entities in vitro and might be useful drugs to improve HNSCC therapy.

The human HECA interacts with cyclins and CDKs to antagonize Wnt-mediated proliferation and chemoresistance of head and neck cancer cells.

There is a growing evidence that the human homologue of the Drosophila headcase (HECA) plays an important role in human carcinogenesis. So far specific protein interaction partners and affected signaling pathways of HECA are still elusive. In a recent study we showed that HECA overexpression in oral squamous-cell carcinoma (OSCC) keratinocytes has tumor suppressive effects resulting in a recuperation of cell cycle control concerning the entry and progression of S-phase, G2- and M-phase. Currently, quantitative RT-PCR and immunohistochemical analysis of primary tumor tissue from OSCC patients demonstrate that HECA expression is markedly decreased compared to normal control patients with abundant HECA expression. Additionally, there is nearly no HECA expression in OSCC metastases. Here, we show that HECA expression is negatively controlled by the Wnt-pathway and TCF4, a Wnt related transcription factor, binds to the HECA promoter. Furthermore, immunocytochemistry reveals colocalization of HECA with the cyclin dependent kinase CDK9. Immunoprecipitation experiments and proximity ligation assays further reveal an interaction of HECA with CDK2, CDK9, Cyclin A and Cyclin K, a direct transcriptional target of the p53 tumor suppressor. Silencing HECA in OSCC cell lines leads to a significant increase of cell division and a markedly increased resistance against the chemotherapeutic cisplatin. On the contrary, HECA overexpressing OSCC cell lines show decreased resistance of OSCC cells against cisplatin. Therefore, HECA could be considered as future therapeutic agent against Wnt-dependent tumor progression.

MAGE-A antigens in lesions of the oral mucosa.

Oral squamous cell carcinoma develops continuously out of predamaged oral mucosa. For the physician and pathologist, difficulties arise in distinguishing precancerous from cancerous lesions. MAGE-A antigens are tumor antigens that are found solely in malignant transformed cells. These antigens might be useful in distinguishing precancerous from cancerous lesions. The aim of this study was to verify this assumption by comparing MAGE-A expression in benign, precancerous, and cancerous lesions of the oral mucosa. Retrospectively, biopsies of different oral lesions were randomly selected. The lesions that were included are 64 benign oral lesions (25 traumatic lesions (oral ulcers), 13 dental follicles, and 26 epulis), 26 oral lichen planus, 123 epithelial precursor lesions (32 epithelial hyperplasia found in leukoplakias, 24 epithelial dysplasia found in leukoplakias, 26 erythroplasia with oral epithelial dysplasia, and 41 carcinomas in situ in erythroleukoplakias). The lesions were immunohistochemically stained with the poly-MAGE-A antibody 57B, and the results were compared. Biopsies of oral lichen planus, oral ulcers, dental follicles, epulis, and leukoplakia without dysplasia showed no positive staining for MAGE-A antigens. Leukoplakia with dysplasia, dysplasia, and carcinomata in situ displayed positive staining in 33%, 65%, and 56% of the cases, respectively. MAGE-A antigens were not detectable via immunohistochemistry in benign lesions of the oral mucosa. The staining rate of dysplastic precancerous lesions or malignant lesions ranged from 33% to 65%. The MAGE-A antigens might facilitate better differentiation between precancerous and cancerous lesions of the oral mucosa.

MAGE-A antigens in patients with primary oral squamous cell carcinoma.

MAGE-A antigens are only expressed on tumor cells. The aim of this study was to identify their expression in patients with oral squamous cell carcinoma (OSCC). Forty-seven patients with primary OSCC was selected retrospectively. Histo-pathological sections were stained immunohistochemically with MAGE-A antibody 57B. The results were evaluated regarding tumor size (T), lymph-node metastasis (N), blood vessel infiltration (V), lymph vessel infiltration (L), grading (G), and sex. MAGE-A antigens were expressed in 55% of all patients. Expression increased with tumor size (T1 = 56%; T2 = 44%; T3 = 67%; T4 = 71%). Lymph-node metastasis had no influence (N0 and N1 about 50%). Tumors with blood and lymph vessel infiltration had higher expression (V0 = 50%; V1 = 100%; L0 = 46%; L1 = 71%). Less-differentiated tumors showed higher rates (G1 = 50%; G2 = 45%; G3 = 83%). OSCC in men were positive in 62% and in women in 38%. MAGE-A antigens are frequently expressed in OSCC. Their expression seems to increase with tumor dedifferentiation.

Sensitization of head and neck squamous cell carcinoma to apoptosis by combinational SMAC mimetic and Fas ligand-Fc treatment in vitro.

This study aimed to investigate the in vitro efficacy of three different SMAC mimetics for pro-apoptotic sensitization of HNSCC cells. We evaluated BV-6 in comparison to Birinapant and LCL161, for which pro-apoptotic sensitization effects have been demonstrated. Concentration-dependent response was measured for BV-6 in each cell line with an average IC50 value 8-fold lower than of aforementioned SMAC mimetics. Combination treatment of FasL (log2) and BV-6 (IC10) showed highly significant cell count reductions even in the lowest applied concentration in five cell lines (PCI-1: p = 0.0002, PCI-13: p = 0.0002, Detroit 562: p: p < 0.0001, FaDu: p < 0.0001, SCC-25: p = 0.0047). Synergistic effects (y < 1) were evident in eight out of 10 cell lines (PCI-1, PCI-9, PCI-13, PCI-68, Detroit 562, FaDu, SCC-25 and HaCaT). Annexin V assays revealed in nine cell lines very highly significant (p < 0.001) pro-apoptotic effects of BV-6. Western blots showed a heterogeneous IAP expression following SMAC mimetic treatment. Except for two cell lines, at least the cellular inhibitor of apoptosis protein 1 (cIAP1) was degraded in response to BV-6. For prospective targeted HNSCC therapy, this study identifies SMAC mimetics, particularly BV-6 as the compound with the highest pro-apoptotic potency, as promising antitumor agents.

The human homolog of the Drosophila headcase protein slows down cell division of head and neck cancer cells.

The human homolog of the Drosophila headcase (HECA) belongs to a new class of cell differentiation regulators. In Drosophila, the HECA protein regulates the proliferation and differentiation of cells during adult morphogenesis. There is growing evidence that HECA plays an important role in human carcinogenesis. In different tumor entities, an altered HECA expression was found (colorectal, pancreatic and renal cancer). Colorectal cancer studies also suggested HECA as a marker for early disease stages. Therefore, we speculated whether human HECA affects cell cycle progression and proliferation in head and neck cancer cells. In vivo, we found a distinct HECA protein expression in basal and superficial cells of a healthy oral epithelium via immunohistochemistry, whereas in tissues of oral squamous cell carcinoma (OSCC), a weaker staining was observed, particularly in basal cells. In vitro, mRNA and protein expression analyses of OSCC cell lines exhibited that HECA expression correlates with the state of cellular differentiation. In further investigations, we overexpressed HECA in the OSCC cell line PCI 13 and performed functional assays. HECA-overexpressing OSCC cells revealed a significant extended doubling time (up to 45%, 17 h) and yielded a lower number of proliferating cells (up to 30%) than controls. Flow cytometry analyses have shown that HECA-overexpressing OSCC cells forced to hold in the G(2)/M-Phase. In summary, our results show that human HECA slows down cell division of OSCC cells and may therefore act as a tumor suppressor in head and neck cancer.

Multi-kinase inhibitors and cisplatin for head and neck cancer treatment in vitro.

Multidrug resistance (MDR) remains one of the major causes of suboptimal outcome following therapy in head and neck squamous cell carcinoma (HNSCC). ATP-binding cassette (ABC) transporters are overexpressed in HNSCC, which contributes to the limited effect of chemotherapeutic treatment. In addition to their named function, tyrosine kinase inhibitors (TKIs) have been revealed to impact on ABC transporter activity and expression. Therefore, the present study aimed to investigate the effects of combination therapy using different TKIs combined with cisplatin. Reverse transcription-quantitative PCR was used to characterize ABC transporter and receptor expression in 5 HNSCC cell lines treated with 3 different TKIs (pazopanib, dovitinib, nintedanib) and cisplatin. Treatment efficacy was analyzed using a crystal violet staining assay. Analysis of ABC transporter (ABCB1, ABCC1 and ABCG2) genetic alterations was performed using The Cancer Genome Atlas. Statistical analysis was conducted to evaluate the effects of mono- and combination treatment. With the exception of ABCB1, all of the investigated ABC transporters were expressed in each cell line. The additive effects of TKI + cisplatin combination treatment were observed for pazopanib in three cell lines, nintedanib in four cell lines, and were not observed for dovitinib in any of the cell lines investigated. The combination of multi-kinase inhibitors and conventional chemotherapy in HNSCC may strengthen the use of current therapeutic strategies; nintedanib appears to be the most suitable TKI for combination therapy. Further efforts are required to classify TKI efficacy with regard to cisplatin resistance.

Targeting inhibitors of apoptosis in oral squamous cell carcinoma in vitro.

Head and neck cancer, which predominantly arises from the oral mucosa, represents the sixth most common malignancy worldwide. These cancer cells can be resistant to programmed cell death triggered by extrinsic stimuli due to innate overexpression of inhibitor of apoptosis proteins (IAPs). The cellular protein second mitochondria-derived activator of caspases (SMAC) can antagonize IAP-induced caspase inhibition and thus trigger apoptosis. Here, we investigate the cell death-sensitizing effects of the SMAC mimetic LCL161 alone and in combination with Fas ligand (FasL) using a panel of six cell lines. Fas receptor (FasR) expression was analyzed by flow cytometry. Cells were treated with FasL and LCL161 alone or in combination, and cytotoxicity was measured using crystal violet assays. Annexin V and cell viability assays using zVAD-fmk and Necrostatin-1 (Nec-1) were carried out to assess the type of programmed cell death induced by LCL161. To demonstrate the sensitizing effects of LCL161, we employed the t-test to compare the effects of FasL alone and in combination with LCL161. Linear regression analysis was performed to determine initial and half maximal inhibitory concentrations (IC10 and IC50, respectively). Distinct FasR expression was detected in each cell line. Four of six cell lines were significantly sensitized to FasL by LCL161 (p < 0.05), and synergistic effects were observed (y < 1). Moreover, the initially resistant cell line SCC-25 was effectively sensitized to FasL by LCL161. Annexin V FACS analysis demonstrated apoptosis-sensitizing and apoptosis-inducing effects of LCL161 across all cell lines. Using specific cell death inhibitors (zVAD-fmk and Nec-1), we demonstrated that LCL161-initiated apoptosis could not be prevented, highlighting the proapoptotic potential of this mimetic in these cells. Our findings show the effectiveness of apoptotic sensitization of OSCC cells by LCL161 in combination with FasL, thus confirming the importance of an IAP-targeting therapeutic approach for oral squamous cell carcinoma.

MicroRNA expression correlates with disease recurrence and overall survival in oral squamous cell carcinoma.

OBJECTIVES: Locoregional disease recurrence and metastatic events are the leading causes of death and the most important prognostic factors in patients with head and neck squamous cell carcinoma (HNSCC). A major goal of oncology is the identification of clinical and molecular parameters to evaluate the individual risk of recurrence. MicroRNAs (miRNAs) have been shown to correlate well with tumor size and differentiation. Therefore, they are candidate biomarkers for estimating clinical outcomes. MATERIALS AND METHODS: In this study, the expression levels of distinct miRNAs extracted from formalin-fixed, paraffin-embedded (FFPE) samples of oral squamous cell carcinoma were compared. RESULTS: Statistical analysis revealed significant correlations between distinct miRNAs and disease recurrence (miR-99*, miR-194*; p < 0.05) and overall survival (miR-99*; p < 0.05). The results were then validated via data from The Cancer Genome Atlas (TCGA). CONCLUSIONS: Our data show that miR-99* and miR-194* can possibly serve as biomarkers for clinical outcome in HNSCC. These findings may help to identify high-risk patients, who could profit from a more individualized treatment and follow-up.

A new multilayered membrane for tissue engineering of oral hard- and soft tissue by means of melt electrospinning writing and film casting - An in vitro study.

Membranes that form a mechanical barrier not only for cells but also for the bacterial flora of the oral cavity may be helpful in infection-free wound healing for guided tissue regeneration (GTR) applications in the field of oral- and maxillofacial surgery. Controlled wound healing without interference from bacterial contamination appears to be achievable in combination with surface scaffolds for bone- and soft tissue regeneration. As this has not yet been realized, we developed multilayered membranes in this study consisting of specific surface scaffolds for bone- and mucosal regeneration as well as bacteria-tight core membranes. These membranes were evaluated in terms of cell growth of osteoblast- (MG63), keratinocyte- (HaCaT), and fibroblast (L929) cell lines. Scaffolds were fabricated via melt electrospinning writing (MEW), while the core membrane was produced via film casting. All constructs were made of medical-grade poly(ε-caprolactone) (PCL). The bacteria-tightness was tested via a bacterial transmigration-assay. PCL scaffolds and core membranes alone demonstrated good cytocompatibility for all cell lines, which was even enhanced by fusing both components together. The core membrane displayed complete bacteria-tightness over two weeks. These bacteria-tight, individually-designed membranes from medical-grade PCL represent a high-potential, clinically oriented method of GTR in the field of oral- and maxillofacial surgery.

Different expression of MAGE-A-antigens in foetal and adult keratinocyte cell lines.

MAGE-A-antigens are an immunologic marker for many cancers. The goal of this study was to compare the expression profiles of MAGE-A2, -A3, -A4, -A6 and -A10 in foetal and adult keratinocytes with an oral squamous cell carcinoma (OSCC) cell line. Expression of MAGE-A2, -A3, -A4, -A6 and -A10-antigens were detected with PCR in foetal and adult keratinocyte cell lines and in an OSCC cell line (pT4N1M0). Quantitative expression of the single MAGE-A-antigens was measured with rtq-PCR. The results were compared to the reference value of the adult keratinocyte cell line. MAGE-A-antigens were detected in all cell lines. Expression profiles of adult and foetal keratinocyte cell lines differed significantly. Expression profiles of foetal and carcinoma cell lines differed significantly also. MAGE-A-antigens were detected in foetal keratinocyte cell line and oral squamous cell carcinoma cell line but differ in their expression profiles. Up to now MAGE-A-antigens were not detected in foetal keratinocytes. Their role is still unknown.

Apoptosis-sensitizing activity of birinapant in head and neck squamous cell carcinoma cell lines.

Inhibitor of apoptosis proteins, which are overexpressed in head and neck squamous cell carcinoma (HNSCC), may cause therapeutic resistance. Using SMAC mimetic compounds, including birinapant, to degrade and/or inhibit these proteins and sensitize apoptosis may enhance therapies in HNSCC. Fas expression was analyzed in nine HNSCC cell lines and one keratinocyte cell line via flow cytometry. These cell lines were treated with Fas ligand-Fc (FasL) and birinapant, a bivalent SMAC mimetic, in mono and combination therapies. Cytotoxicity was measured using a crystal violet assay. Annexin V assay was performed for detection of apoptosis. The treatment efficacy of mono and combination therapies was statistically analyzed. Nonlinear regression analysis was performed to determine the inhibitory concentration (IC10) of birinapant. Fas expression was detected in each cell line tested. Mono treatment with FasL revealed minor to no apoptotic effects in the majority of the cell lines. Crystal violet and Annexin V staining revealed increased apoptosis rates for all cell lines following incubation with birinapant in mono treatment. Combination treatment with FasL and birinapant (IC10) revealed additional and synergistic effects in eight out of the ten cell lines. To the best of our knowledge, the present study provided the first evidence of the apoptosis-sensitizing activity of combination treatment with FasL and birinapant in HNSCC cell lines.

MAGE-A9 in head and neck cancer: Prognostic value and preclinical findings in the context of irradiation.

Radiotherapy alone, or as an addition to surgery is important for the treatment of head and neck squamous cell carcinoma (HNSCC). In addition to their expression in germ cells, melanoma associated antigens-A (MAGE-A) are only expressed in malignant tissue. Notably, there is a known correlation between MAGE-A9 expression and poor prognosis in HNSCC patients. However, current knowledge regarding the function of MAGE-A9 expression, particularly in the context of irradiation, is limited. MAGE-A9 expression in 37 oral squamous cell carcinoma patents was immunohistochemically determined and analyzed for overall survival by the Kaplan-Meier log-rank test. Next, the expression of MAGE-A9 was determined by reverse transcription-quantitative polymerase chain reaction in HNSCC cell lines prior to and following irradiation with 2 Gray. The radiosensitivity of each cell line was determined using a clonogenic survival assay. There was a significantly (P=0.0468) longer overall survival in patients with a low level of MAGE-A9 expression. The median overall survival in patients with high MAGE-A9 expression was 47% compared to 73% in the group with low MAGE-A9 expression. The cell lines revealed a distinct expression pattern of MAGE-A9. Following irradiation of the cell lines, a significant enhancement of MAGE-A9 mRNA expression levels was observed. The most prominent alteration in MAGE-A9 expression was observed in the most radioresistant cell line. A high MAGE-A9 expression level correlates significantly with lower overall survival in HNSCC patients. Additionally, irradiation increased the MAGE-A9 mRNA levels in all five HNSCC cell lines, and the most resistant cell line demonstrated the greatest increase in MAGE-A9 expression following irradiation.