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Elevated erythropoietin (EPO) levels have been reported to predict poor survival in various populations including diabetic patients. However, data regarding its impact on renal outcomes are scarce. We conducted a single-center, prospective cohort study of 339 type 2 diabetic patients with anemia. The primary outcome was the estimated glomerular filtration rate (eGFR) slope for two years. We performed multiple linear regression and restricted cubic spline analyses to assess the association of serum EPO levels with the renal outcome. Chronic kidney disease (CKD) was defined as eGFR 30 mg/g creatinine. Median baseline EPO and eGFR level were 14.4 IU/L and 53 mL/min/1.73 m2, respectively. Inappropriately low EPO levels were observed in 73% of anemic patients and 59% of anemic patients even without CKD, suggesting that EPO deficiency precedes the onset of CKD in diabetes mellitus. Multivariable analysis revealed that iron status and hemoglobin levels were major determinants of EPO levels. Median eGFR slope was −1.3 mL/min/1.73 m2/year. We found that low EPO levels, but not low hemoglobin levels, were associated with a faster decline in eGFR, independent of clinically relevant factors. The eGFR decline was steeper, particularly when the EPO level was below the upper limit of normal. Lower EPO concentrations were associated with rapid eGFR decline, especially in patients with iron deficiency (P for interaction = 0.01). Relative EPO deficiency should be considered as a culprit in anemia of unknown etiology in diabetic patients, even those without CKD. Low EPO levels, especially when accompanied by poor iron status, are predictive of rapid loss of renal function.

Background: Empagliflozin reduces the risk of hospitalization for heart failure in patients with type 2 diabetes and cardiovascular disease. We sought to elucidate the effect of empagliflozin as an add-on therapy on decongestion and renal function in patients with type 2 diabetes admitted for acute decompensated heart failure. Methods: The study was terminated early due to COVID-19 pandemic. We enrolled 59 consecutive patients with type 2 diabetes admitted for acute decompensated heart failure. Patients were randomly assigned to receive either empagliflozin add-on (n=30) or conventional glucose-lowering therapy (n=29). We performed laboratory tests at baseline and 1, 2, 3, and 7 days after randomization. Percent change in plasma volume between admission and subsequent time points was calculated using the Strauss formula. Results: There were no significant baseline differences in left ventricular ejection fraction and serum NT-proBNP (N-terminal pro-B-type natriuretic peptide), hematocrit, or serum creatinine levels between the 2 groups. Seven days after randomization, NT-proBNP level was significantly lower in the empagliflozin group than in the conventional group ( P =0.040), and hemoconcentration (≥3% absolute increase in hematocrit) was more frequently observed in the empagliflozin group than in the conventional group ( P =0.020). The decrease in percent change in plasma volume between baseline and subsequent time points was significantly larger in the empagliflozin group than in the conventional group 7 days after randomization ( P =0.017). The incidence of worsening renal function (an increase in serum creatinine ≥0.3 mg/dL) did not significantly differ between the 2 groups. Conclusions: In this exploratory analysis, empagliflozin achieved effective decongestion without an increased risk of worsening renal function as an add-on therapy in patients with type 2 diabetes with acute decompensated heart failure. Registration: URL: https://www.umin.ac.jp/ctr/index.htm ; Unique identifier: UMIN000026315.

Objectives The ratio of glycated albumin to HbA1c (GA/HbA1c ratio) is a known indicator that reflects fluctuations in plasma glucose. In this study, the association of the GA/HbA1c ratio to diabetic nephropathy and diabetic retinopathy in patients with type 2 diabetes was investigated. Design and methods Among patients with type 2 diabetes, 613 patients (364 males and 249 females, aged 63.2 ± 12.5, body mass index (BMI) 25.4 ± 4.8 kg/m2) were enrolled. Patients with overt proteinuria, reduced renal function, or anemia were excluded. Results In a comparison between patients with and without diabetic nephropathy, significance was observed in insulin therapy, HbA1c, and GA. In addition, in a comparison between patients with and without diabetic retinopathy, the GA/HbA1c ratio along with insulin therapy, HbA1c, and GA showed significant differences. When the GA/HbA1c ratios were divided into three groups and compared, the rates of diabetic nephropathy did not show any significance, while the rate of diabetic retinopathy increased significantly as the GA/HbA1c ratio increased. In multivariable analyses, while insulin therapy and BMI were the significant independent variables for diabetic nephropathy, insulin therapy and the GA/HbA1c ratios were the significant independent variable for diabetic retinopathy. Conclusions The GA/HbA1c ratio was associated with diabetic retinopathy, but not with diabetic nephropathy in patients with type 2 diabetes. These results suggest that the development and progression of diabetic retinopathy is associated with plasma glucose fluctuations.

For heavy-duty gas-turbine engines, one of the promising approaches to reducing NOX emissions is the adoption of lean premixed combustion. This technique could be combined with the conventional technique of exhaust gas recirculation (EGR). However, the reduction in the oxygen concentration will influence the burning velocity and reaction zone characteristics of the lean premixed flame. To elucidate this effect, in this study, we measured the lean premixed flame temperature and OH concentration distributions for various oxygen concentrations instantaneously and simultaneously using laser imaging techniques. Based on the results, we investigated the characteristics of a lean premixed flame under various oxygen concentrations and found that the OH laser-induced fluorescence (LIF) intensity in the reaction zone decreased with the oxygen concentration, as did the flame temperature at a given axial distance from the exit nozzle. The characteristics of the premixed flame changed from a small-scale convexo-concav...

Polypyridine grafted with poly (ethylene glycol) copolymers (Py-g-PEG) with varying composition were prepared through radical copolymerization of methyl-terminated PEG macromonomer and 4-pyridylmethyl methacrylate, and the obtained copolymer spontaneously adsorbs from aqueous solution onto gold surfaces, where the pyridine parts act as the multipoint anchor to the surface and the PEG parts provide the strong steric repulsion between the chains. As a result, the highly protein repellent and stable surface was constructed through multipoint pyridine attachment as compared with singlepoint pyridine attachment. Furthermore, the property of multimolecular micellization for the obtained copolymer in aqueous milieu was examined to estimate the relation between the surface properties on the substrate and the self-assembling ability in aqueous milieu. Py-g-PEGs thus prepared are promising materials to functionalize metal and semiconductor materials and to self-assemble into micelles in biotechnological and biomedical fields.

Polypyridine grafted with poly(ethylene glycol) (Py-g-PEG) have been synthesized. Radical copolymerization of methyl-terminated PEG macromonomer with 4-pyridylmethyl methacrylate homogeneously proceeded and the obtained copolymer spontaneously adsorbs from aqueous solution onto gold surfaces, where the pyridine parts act as the multipoint anchor to the surface and the PEG parts provide the strong steric repulsion between the chains. As a result, the highly protein repellent and stable surface was constructed through multipoint pyridine attachment as compared with singlepoint pyridine attachment. Py-g-PEGs thus synthesized are promising material to function-alize metal and semiconductor material and to self-assemble into micelle in biotechnological and biomedical field.

Hydroxyapatite (HAp)-20vol% β-tricalcium phosphate (β-TCP) composites were prepared. The effect of β-TCP size on bone-like layer growth and adhesion of osteoblast-like cells on the composites was systematically studied in vitro. When the composites were soaked in a simulated body fluid, the formation and growth rates of the bone-like layer increased with increasing β-TCP size, even if the volume fraction of β-TCP was constant. Moreover, selective dissolution of β-TCP phase and formation of the bone-like layer around the phase were frequently observed. Higher Ca 2+ concentration due to the fast dissolution of β-TCP beneath the sample surface resulted in faster formation and growth of the bone-like layer, especially in the samples containing β-TCP powders larger than 100 μm. MC3T3-E1 osteoblast-like cells preferentially adhered to β-TCP phase in HAp/β-TCP composites because of the enrichment of Ca 2+ ions around β-TCP.

Calcium-deficient hydroxyapatite (DAp) was synthesized by hydrolysis of alpha-TCP at different pH. Ca/P ratio of DAp increased with increasing pH during the hydrolysis. In addition, DAp which was synthesized at lower pH, showed higher solubility in the simulated body fluid (SBF) and had a more negatively charged surface than that at higher pH. Bone-like crystals were formed on DAp which have been soaked in SBF at 309.5 K in vitro. DAp demonstrated faster formation of a bone-like layer than stoichiometric hydroxyapatite (HAp). Moreover, the formation of the crystals on DAp synthesized at lower pH was much faster than that at higher pH. The adhesion of MC3T3-E1 osteoblast-like cells was also influenced by pH during the synthesis of DAp; the number of adhering cells increased with a decrease in the pH. Therefore, high solubility and negative charge of DAp, especially that synthesized at lower pH resulted in fast formation of bone-like crystals and good cell adhesion.

Purpose We investigated whether serum interleukin (IL)-8 reflects the tumor microenvironment and has prognostic value in patients with oral squamous cell carcinoma (OSCC). Experimental Design Fifty OSCC patients who received radical resection of their tumor(s) were enrolled. Preoperative sera were measured for IL-8 by ELISA. Expression of IL-8 and the infiltration of immune cells in tumor tissues were analyzed by an immunohistochemical staining of surgical specimens. Results We found that disease-free survival (DFS) was significantly longer in the Stage I/II OSCC patients with low serum IL-8 levels compared to those with high levels (p = 0.001). The tumor expression of IL-8, i.e., IL-8(T) and the density of CD163-positive cells in the tumor invasive front, i.e., CD163(IF) were correlated with the serum IL-8 level (p = 0.033 and p = 0.038, respectively), and they were associated with poor clinical outcome (p = 0.007 and p = 0.002, respectively, in DFS) in all patients. A multivariate analysis revealed that N status, IL-8(T) and CD163(IF) significantly affected the DFS of the patients. Further analysis suggested that combination of N status with serum IL-8, IL-8(T) or CD163(IF) may be a new criterion for discriminating between OSCC patients at high and low risk for tumor relapse. Interestingly, the in vitro experiments demonstrated that IL-8 enhanced generation of CD163-positive M2 macrophages from peripheral blood monocytes, and that the cells produced IL-10. Conclusions These findings indicate that IL-8 may be involved in poor clinical outcomes via generation of CD163-positive M2 macrophages, and that these factors in addition to N status may have prognostic value in patients with resectable OSCSS.

The fabrication of quasi-phase-matching (QPM) crystals by selective-area growth on the two asymmetrically polar surfaces of GaN is examined. We attempted the fabrication of GaN-QPM crystals by one-time growth using a carbon mask. For GaN double-polarity selective-area growth (DP-SAG), we investigated the effect of varied nitriding times of the Al2O3 templates patterned with the carbon mask. We optimized the nitriding conditions for the DP-SAG process, and evaluated the substrate fabricated by the optimized DP-SAG process. In addition, we examined the interface formation mechanism of DP-GaN fabricated by GaN DP-SAG process. We determined that it is possible to fabricate DP-GaN with a sharp interface by optimizing the growth conditions.

Summary Objectives Oncogene addiction has provided therapeutic opportunities in many human malignancies, but molecular targeted therapy for oral squamous cell carcinoma (OSCC) is not yet available. In this study, we attempted to identify an appropriate target molecule for treatment of patients with OSCC. Materials and methods Microarray analysis was performed to determine the gene expression profiles in nine human OSCC cell lines and a non-neoplastic keratinocyte cell line. The expression levels of Aurora kinase A (AURKA) mRNA and protein in human OSCC cells and tissues were examined. We investigated the effect of small interfering RNAs specific for AURKA (siAURKAs) and MLN8237, an AURKA selective inhibitor on the growth of OSCC cells in vitro and in vivo. We also analyzed clinical significance in AURKA mRNA expression levels in OSCC. Results AURKA was overexpressed in human OSCC cell lines and tissues. All siAURKAs almost completely suppressed the expression of AURKA protein, and significantly inhibited the growth of OSCC cells by 31–89%. MLN8237 also reduced the cellular growth rate by 38–74%. Both siAURKA and MLN8237 significantly reduced the size of subcutaneously xenografted OSCC tumors by 66% and 40%. Knockdown of AURKA expression and MLN8237 induced the growth inhibition of primary cultured cells established from patients’ OSCC tumors. Furthermore, we found a significant association between AURKA mRNA expression levels and histological differentiation and lymph node metastasis. Conclusions AURKA plays a critical role in the growth of human OSCC cells and targeting AURKA may be a useful therapeutic strategy for OSCC.

A 55-year-old woman consulted our hospital for an epulis-like small mass in the anterior region of the mandible. A biopsy of the tumor was performed. Histological analysis showed that the tumor consisted of spindle-shaped and polygonal cells with hyperchromatic nuclei, and intracytoplasmic vacuoles and mitotic figures were scattered. Immunohistochemical staining revealed that the tumor cells were positive for factor VIII-related antigen, CD31, αSMA, and vimentin, but negative for pancytokeratins, S100 protein, neuron-specific enolase, and CD56. The Ki-67 labeling index was more than 50%. Based on these findings, a final pathological diagnosis of angiosarcoma was made. The tumor did not invade into the surrounding tissue. The operation was performed with about a 20-mm surgical margin that was negative for tumor invasion. After a 4-year follow-up, no metastatic lesions were found, and the primary site was covered with a partial denture.

Slow/no-reflow phenomenon during emergent percutaneous coronary intervention in patients with ST-elevation myocardial infarction (STEMI) results in a poor prognosis. A high thrombus burden was an independent predictor of angiographic slow/no-reflow phenomenon. We experienced a case of a STEMI patient with massive intracoronary thrombus. In our case, a massive red thrombus was aspirated easily by adjunctive pulse infusion thrombolysis (PIT) after failed aspiration. Adjunctive pulse infusion thrombolysis after failed aspiration might be a useful strategy to prevent the slow/no-reflow phenomenon in STEMI patients with massive intracoronary thrombus.

We describe a simulation model based on an avian ecosystem for determining what causes birdsong evolution. It is already known that songbirds communicate with a "birdsong." This birdsong is used in territorial and courtship behaviors. Some previous researches have suggested that songs related to territorial behaviors should have simple structures while those related to courtship behaviors should have complex ones. We suspect that birdsongs are constantly evolving to achieve a suitable balance between the two behaviors while considering the surrounding environment. We consider avian habitat density to be one of the most important environmental factors influencing birdsong evolution and therefore created different densities in a simulation model. In this paper, we propose a birdsong acquisition model by sexual selection that contains both territorial and courtship behaviors. We conducted simulations with the proposed model and determined that the evolution of birdsongs differs depending on a bird's habitat density.

Biomarkers are essential for improving the effectiveness of treatments for patients with cancer. The immunological status in a tumor microenvironment is closely associated with the prognosis of cancer patients. Here we investigated whether serum interleukin (IL)-8 reflects the tumor microenvironment and has prognostic value in 50 patients with resectable oral squamous cell carcinoma (OSCC). We found that the relapse-free survival (RFS) was significantly longer in the Stage I/II OSCC patients with low serum IL-8 levels compared to those with high levels (p = 0.001). The tumor expression of IL-8, the infiltration of CD163-positive cells, and of CD66b-positive cells in the tumor tissues were correlated with the serum IL-8 level (p = 0.033, p = 0.038 and p = 0.044, respectively), and they were associated with poor clinical outcome (p = 0.007, p = 0.002 and p = 0.001 respectively, in DFS) in all patients. A multivariate analysis revealed that N status, tumor IL-8 expression, CD163 and CD66b infiltration significantly affected the DFS of the patients. A further analysis suggested that the combination of N status with serum IL-8, tumor IL-8, CD163 and CD66b infiltration in the tumor tissues may be a new criterion for discriminating between OSCC patients at high and low risk for tumor relapse. Finally, the in vitro experiments clearly demonstrated that IL-8 enhanced the generation of CD163-positive M2 macrophages from peripheral blood monocytes, and that the cells produced IL-10 which suppress anti-tumor immunity. Furthermore, IL-8 increased the generation of arginase-producing, CD66b-overexpressing cells from peripheral blood polymorphonuclear cells. These findings indicate that IL-8 may be involved in poor clinical outcomes via the generation of CD163-positive M2 macrophages and CD66b-overexpressing, arginase-producing granulocytes, and that these factors may have prognostic value as well as may be new targets for treatment in patients with resectable OSCC. Citation Format: Masato Okamoto, Yohei Fujita, Hiroyuki Goda, Koh-ichi Nakashiro, Hiroyuki Hamakawa. Interleukin-8 promotes generation of M2 macrophages and arginase-producing granulocytes: Prognostic significance of interleukin-8 and infiltration of immune cells positive for CD163 and CD66b in tumor tissues in patients with oral squamous cell carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-219. doi:10.1158/1538-7445.AM2015-LB-219

Molecular target drugs are used in the treatment of various malignant tumors. However, the development of molecular targeted therapy for oral squamous cell carcinoma (OSCC) is lagging behind other cancers. In this study, we have attempted to identify an appropriate molecular target for treatment of patients with OSCC. We determined the gene expression profiles of human 9 OSCC cell lines and an immortalized non-neoplastic human keratinocyte cell line by microarray analysis, and then found cell division cycle associated 5 (CDCA5) as one of genes which were commonly overexpressed in OSCC cells. CDCA5 was initially identified as substrate of anaphase-promoting complex and as a regulator of sister chromatid cohesion in HeLa. In addition, it has been reported that CDCA5 plays a critical role in human lung carcinogenesis. However, function of CDCA5 in OSCC remains still unknown. First, we confirmed the overexpression of CDCA5 in 5 human OSCC cells by quantitative RT-PCR (qRT-PCR) and Western blotting. The expression levels of CDCA5 mRNA and protein were markedly elevated in all human OSCC cell lines compared to a non-neoplastic human epithelial cell line. Next, we examined the effect of synthetic small interfering RNAs specific for CDCA5 (siCDCA5) on the growth of human OSCC cells. RNAi effect of siCDCA5 was confirmed by Western blotting. Transfection of siCDCA5 suppressed the expression of its target protein in all types of cells. The growth inhibitory effect of siCDCA5 in OSCC cells was evaluated by WST-8 assay. Knockdown of CDCA5 significantly inhibited the growth of OSCC cells in vitro. Subsequently, we analyzed the influence of siCDCA5 on cell cycle by flow cytometry. Suppression of CDCA5 resulted in the decrease in percentage of cells in G0/G1 phase and increase in G2 phase. This indicated that the anti-proliferative effect of siCDCA5 was due to G2 arrest. Furthermore, we investigated the clinical significance of CDCA5 expression in OSCC. The expression of CDCA5 mRNA in tumor and adjacent normal tissues derived from the patient with OSCC was examined by qRT-PCR. Expression levels of CDCA5 mRNA in the OSCC tissues were significantly higher than in normal tissues. We also examined the expression of CDCA5 protein in 80 OSCC cases by immunohistochemistry with anti-CDCA5 antibodies. Tissue expression of CDCA5 was divided into high or low group at 50% stained tumor cells, resulting in 36 cases as high expression and 44 cases as low expression. We then examined the association of CDCA5 expression with various clinicopathological parameters of OSCC patients, and found a significant association between CDCA5 expression levels and pathological staging, metastasis, and recurrence. These results suggest that CDCA5 functions as a critical gene supporting the growth of human OSCC cells and targeting CDCA5 appears a useful therapeutic strategy for the patients with OSCC. Citation Format: Norihiko Tokuzen, Koh-ichi Nakashiro, Hiroshi Tanaka, Yohei Fujita, Kazuki Iwamoto, Hiroyuki Hamakawa. Overexpression and function of CDCA5 in oral squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 476. doi:10.1158/1538-7445.AM2014-476

Squamous cell carcinoma is one of the most common malignancies in head and neck region. Primary tumors, especially derived from undifferentiated cells, tend to have aggressive phenotypes, which are mainly characterized by local recurrence and distant metastasis. In this study we found that Id-1, a negative regulator of tissue specific transcription factors, is highly expressed in head and neck squamous carcinoma cells (HNSCC), and that the expression is correlated with the malignant phenotype, i.e., cell proliferation and invasion. Id-1 knockdown dramatically reduces squamous cell invasion that is accompanied by profound morphologic changes and robust reduction in expression levels of mesenchymal markers.In addition, reduction in tumor growth and neoangiogenesis were observed in vivo, indicating that knock-down of Id-1 drastically suppresses VEGFA expression. Our results also suggested that VEGFA-autocrine loop activates tylosine kinase pathways including Srk and FAK, and therefore manifests invasive phenotype. With clinical materials obtained from HNSCC patients, significant correlation was observed between Id-1 and VEGFA. Furthermore, we demonstrated that a non toxic compound, cannabidiol, significantly downregulates Id-1 gene expression and associated squamous cell proliferation and invasiveness. Taken together, these results suggest that Id-1 controls cancer cell phenotype in culture and in vivo, and that drugs targeting Id-1 represent a novel and promising strategy for improving the therapy and outcome of patients with HNSCC. Citation Format: Ryuichi Murase, Sean McAllister, Yohei Fujita, Tomoki Sumida, Koichi Nakashiro, Pierre Desprez, Hiroyuki Hamakawa. Id-1 gene and protein as therapeutic target for head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1017. doi:10.1158/1538-7445.AM2014-1017

Temperature-sensitive hydrophilic gel microcapsules have been newly prepared. That is, poly (L-lysineisopropylamide-terephthalic acid) microcapsules containing water have been obtained by an interfacial polymerization at a water/oil interface between L-lysineisopropylamide and terephthaloyldichloride. The microcapsule changes its size between 33 and 35 degrees C. Under 33 degrees C, the microcapsules are fully spherical and can be redispersed in distilled water, while are aggregated above 35 degrees C. The microcapsules, which are observed to show aggregation above 33 degrees C, can be redispersed by decreasing temperature within a few second. The thermosensitive morphological changes of the microcapsules are thus reversible. Also, it has been shown that the permeability of sodium chloride through the microcapsule membrane changes remarkably between 33 and 35 degrees C, while it is kept almost constant independent of temperature between 25 and 33 degrees C or between 35 and 55 degrees C. The permeability of solutes is higher under 33 degrees C than that above 35 degrees C. Such thermosensitive properties result from the fact that the polymer membrane has isopropylamide groups. That is, L-lysineisopropylamide has a chemical structure similar to N-isopropylacrylamide, the polymer of which, poly (N-isopropylacrylamide), is a thermosensitive hydrogel having its phase transition temperature around 33 degrees C.

For nonlinear optical applications using gallium nitride (GaN), periodic inversion of crystallographic orientation (polarity) is required for quasi-phase matching. We developed a novel procedure for designing polarity patterns in GaN using metal organic vapor phase epitaxy (MOVPE), and we used this to fabricate periodic polarity-inverted GaN films. By using a carbon mask for the formation of the selective area, substrate nitriding and mask removal of the selective area were carried out in the GaN epitaxial growth process. In this report, double-polarity selective area growth (DP-SAG) was realized by optimizing the nitriding and mask removal conditions. The interface of the Ga-polarity/N-polarity region became sharp by controlling the V/III ratio at 4700.

Recent experiments have showed that it is possible to use pharmacological agents that inactivate oncogenes to treat at least some types of human malignancies. However, the development of molecular targeted therapy in oral squamous cell carcinoma (OSCC) is lagging behind compare to other cancers. In this study, we attempted to identify an appropriate target molecule and to determine whether targeting such a molecule is plausible therapeutic approach for the treatment of patients with OSCC. We determined the gene expression profiles of 9 human OSCC cell lines and non-neoplastic keratinocyte cell line by microarray analysis, and then identified Aurora kinase A (AURKA) as a cancer-related gene. AURKA has been shown to be related to the progression, survival, histological differentiation, and metastasis in various tumors. We revealed the overexpression of AURKA mRNA and protein in human OSCC cell lines and tissues. To clarify the function of AURKA in the cell proliferation of OSCC cells, we investigated the effect of small interfering RNA (siRNA) specific for AURKA (siAURKA) and MLN8237, an AURKA selective inhibitor, on the growth of human OSCC cells in vitro and in vivo. All siAURKAs almost completely suppressed the expression of AURKA protein, and significantly inhibited the growth of these cells compared to non-targeting siRNA. MLN8237 also markedly reduced the growth rate of human OSCC cells. Next, we assessed the growth inhibitory effect of siAURKA and MLN8237 using mouse model. Synthetic siAURKA/atelocollagen complexes (40 μM) were administrated into mouse tail vain every 3 days, and MLN8237 (20 mg/kg) was received orally for 14 consecutive days. We found both siAURKA and MLN8237 significantly reduced the size of subcutaneously xenografted OSCC tumors. To confirm the usefulness of targeting AURKA in OSCC, we attempted to culture the resected tumor tissues from 3 patients with OSCC and obtained primary cultured cells. Knockdown of AURKA expression and MLN8237 induced the growth inhibition of OSCC primary cultured cells. Furthermore, the expression of AURKA mRNA in OSCC tissues resected from patients was examined. In 37 of 50 primary OSCC tissues, the expression levels of AURKA mRNA were more than 2-fold increase compared to normal oral mucosa tissues, and notably we found significant association between AURKA mRNA expression levels and histological differentiation and lymph node metastasis. These results suggest that AURKA plays a critical role in the growth of human OSCC cells, and targeting AURKA appears to be a potentially useful therapeutic approach for patients with OSCC. Citation Format: Hiroshi Tanaka, Koh-ichi Nakashiro, Kazuki Iwamoto, Norihiko Tokuzen, Yohei Fujita, Rikimaru Shirakawa, Ryota Oka, Hiroyuki Goda, Hiroyuki Hamakawa. Aurora kinase A as a novel therapeutic target in oral squamous cell carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3003. doi:10.1158/1538-7445.AM2013-3003

Purpose: Dendritic cells (DCs) are professional antigen-presenting cells that play a central role in initiating adaptive and innate immune responses. DC-based vaccination is widely tested as a promising therapeutic cancer vaccine in multiple clinical trials. Programed cell death 1(PD-1)/PD-1 Ligands (PD-Ls) pathway are involved in the negative regulation of T cell-functions. PD-1 is expressed on T cells, and PD-Ls are expressed on DCs as well as some cancer cells. More recent studies have indicated that certain chemotherapeutic agents might enhance the cancer vaccine-mediated immune responses. In the current study, we conducted the in vitro experiments to examine whether or not chemotherapeutic drugs which are used for patients with head and neck cancer, may enhance T cell-stimulating ability of DCs via regulation of the expression of PD-Ls. Methods: Healthy volunteer-derived human peripheral blood monocyte were cultured with IL-4 and GM-CSF for 5 days to generate immature DCs, and then the DCs were matured by the stimulation with OK-432 which is a streptococcal immune adjuvant, for 24 hours, followed by the treatment with each chemotherapeutic drug such as 5-FU, CDDP, docetaxel or gemcitabine for 24 hours. Viability of DCs was assessed by WST-8. IL-10 and IL-12 in the supernatants from the DC cultures were measured by ELISA. Expression of mRNAs and proteins for PD-Ls was analyzed by quantitative real-time PCR and flow cytometry, respectively. DCs were co-cultured with allogeneic T cells (E/T ratio=1/20, 1/200 or 1/2000), then IFN-γ in the supernatants were measured by ELISA to evaluate T cell-stimulating ability of the DCs. Results: IL-12 production by DCs was significantly enhanced by OK-432 stimulation. Although CDDP, docetaxel and gemcitabine inhibited the OK-432-induced IL-12 production on the DCs, 5-FU did not. 5-FU and gemcitabine significantly inhibited the IL-10 production by OK-432-stimulated DCs. Expression of PD-L1 and PD-L2 genes as well as of their proteins on the DCs which was increased by OK-432 stimulation, was significantly suppressed by the treated with 5-FU or docetaxel. OK-432-stimulated DCs-treated with 5-FU strongly enhanced both proliferation and IFN-γ production of allogeneic T cells as compared with DCs stimulated only with OK-432. Conclusions: The findings obtained from the current study suggested the possibility that 5-FU may enhance the T cell-stimulating ability of dendritic cells via regulation of the expression of PD-L1 and PD-L2, and that the combination therapy of DC-based cancer vaccine with 5-FU may elicit better anti-tumor efficacy than only vaccination. Citation Format: Tomoyuki Tano, Masato Okamoto, Hiroyuki Goda, Yohei Fujita, Koh-ichi Nakashiro, Hiroyuki Hamakawa. Enhancement of the T cell-stimulating ability of dendritic cells by 5-FU via regulation of the expression of ligands for programed cell death 1. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4722. doi:10.1158/1538-7445.AM2013-4722