Language：Japanese   Publisher：日本電気泳動学会  

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Our aim was to develop a serodiagnostic marker for lung cancer. Monoclonal antibodies were generated, and one antibody designated as KU-Lu-1, recognizing cytoskeleton-associated protein 4 (CKAP4), was studied further. To evaluate the utility of KU-Lu-1 antibody as a serodiagnostic marker for lung cancer, reverse-phase protein array analysis was performed with sera of 271 lung cancer patients and 100 healthy controls. CKAP4 was detected in lung cancer cells and tissues, and its secretion into the culture supernatant was also confirmed. The serum CKAP4 levels of lung cancer patients were significantly higher than those of healthy controls (P < 0.0001), and the area under the curve of receiver-operating characteristic curve analysis was 0.890, with 81.1% sensitivity and 86.0% specificity. Furthermore, the serum CKAP4 levels were also higher in patients with stage I adenocarcinoma or squamous cell carcinoma than in healthy controls (P < 0.0001). Serum CKAP4 levels may differentiate lung cancer patients from healthy controls, and they may be detected early even in stage I non-small cell lung cancer. Serum CKAP4 levels were also significantly higher in lung cancer patients than in healthy controls in the validation set (P < 0.0001). The present results provide evidence that CKAP4 may be a novel early serodiagnostic marker for lung cancer.

DOI： 10.1016/j.ajpath.2018.03.007

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Language：Japanese   Publisher：(一社)日本病理学会  

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Publishing type：Research paper (scientific journal)   Publisher：Japanese Electrophoresis Society  

DOI： 10.2198/electroph.62.9

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Language：Japanese   Publishing type：Research paper (scientific journal)   Publisher：Japanese Electrophoresis Society  

<p>To develop sero-diagnostic markers for lung cancer, surface proteins were labeled and collected, then identified by the shotgun analysis. Surface proteins were biotinylated from three each of lung cancer cell lines and collected them. Biotinylated proteins were identified by the shotgun analysis. Expression levels of identified proteins in four typical lung cancer cell lines were confirmed by immunoblotting and immunohistochemistry. And also examined CD109 levels as a sero-diagnostic marker for lung cancer, sera from 268 lung cancer patients and 100 healthy controls were analyzed by the reverse-phase protein array (RPPA) method. Serum levels of CD109 were significantly higher in patients with adenocarcinoma and squamous cell carcinoma than in healthy controls (p < 0.0001), and the area under the curve (AUC) of receiver-operating characteristic curve (ROC) analysis was 0.93, when an optimal cut-off value of 1.13 was applied, the diagnostic sensitivity and specificity for lung cancer were 84 and 88%, respectively. In addition, because CD109 was detected even in stage I disease, serum levels of CD109 antigen should be applicable early sero-diagnostic markers for lung cancer patients discriminating from healthy controls. To our knowledge, this is the first report providing evidence that CD109 may be an early sero diagnostic marker for lung cancer.</p>

DOI： 10.2198/electroph.61.23

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Language：Japanese   Publishing type：Research paper (scientific journal)   Publisher：Japanese Electrophoresis Society  

<p>To develop novel sero-diagnostic markers for membranous proteins of lung cancer, surface proteins were identified in combined with various proteomic methods. Biotinylated membranous proteins in lung cancer cell lines were collected and identified by the shot-gun analysis. Expression levels of identified proteins in four different histological types of lung cancer cell line were examined by the immunoblot and immunohistochemical analyses. Serum levels of these proteins were also examined in patients with lung cancer and healthy controls by the reverse-phase protein array (RPPA) method. From identified 92 membranous proteins including CD109, CD155 and Roundabout Guidance Receptor 1 (ROBO1), we focused on the CD155 protein. Serum levels of CD155 were significantly higher in patients with adenocarcinoma and squamous cell carcinoma than in healthy controls (p < 0.0001), and the area under the curve (AUC) of receiver-operating characteristic curve (ROC) analysis was 0.87, when an optimal cut-off value of 0.31 was applied, the diagnostic sensitivity and specificity for lung cancer were 81 and 82%, respectively. To our knowledge, this is the first report providing evidence that CD155 may be a sero-diagnostic marker for non-small cell lung cancer. This methodology was shown to be useful for acquisition of novel biomarkers for membranous proteins.</p>

DOI： 10.2198/electroph.61.120

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Publishing type：Research paper (scientific journal)   Publisher：Japanese Electrophoresis Society  

DOI： 10.2198/electroph.61.39

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Language：Japanese   Publisher：Japanese Electrophoresis Society  

<p>We have generated several tumor markers for lung cancer using various proteomic methods including generation of monoclonal antibodies by using the random immunization method, the two-dimensional electrophoresis, and autoantibody analysis using lung cancer tissues, cell lines and patients' sera. Although the adjuvant platinum-based chemotherapy has been shown to improve survival of patients with completely resected stage II and IIIA non-small cell lung cancer (NSCLC), its effect is limited. In addition, the prior clinical examination whether this therapy is effective has not been performed. In the present study, we investigated the utility as the therapeutic effect prediction markers about the candidate marker proteins for lung cancer that we obtained previously. We review mainly on our recent study about the utility of nestin, S100A16, and myosin-9 proteins as the effect prediction factor for the adjuvant platinum-based chemotherapy.</p>

DOI： 10.2198/electroph.61.128

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：SCIENTIFIC PUBLISHERS INDIA  

To identify diagnostic markers of psoriasis vulgaris and psoriatic arthritis, autoantibodies in sera from patients with psoriasis were screened by two-dimensional immunoblotting (2D-IB). We previously found 22 autoantigens each in psoriasis sera. In this study, serum levels of valosin-containing protein (VCP) in patients, one of the identified autoantigens in psoriasis patients, were studied by reverse-phase protein array analysis using sera from patients with psoriasis and healthy controls. Serum levels of VCP were significantly higher in patients with psoriatic arthritis than in controls or patients with psoriasis vulgaris (P&lt;0.05 each). The area under the curve for VCP between patients with psoriatic arthritis and controls or psoriatic vulgaris patients was 0.88 and 0.77, respectively. The serum level of VCP was weakly correlated with the disease activity of psoriasis patients and psoriasis area severity index score. These data suggest that VCP is a novel and differential sero-diagnostic marker of psoriatic arthritis.

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Language：English   Publishing type：Research paper (scientific journal)  

To investigate the relationships between the expression of MUC5B and clinicopathological parameters, the expression of MUC5B was immunohistochemically studied. MUC5B expression was observed in 129 of 198 (65.2%) adenocarcinomas and in 4 of 49 (8.2%) squamous cell carcinomas (P < 0.00001). MUC5B expression was significantly associated with poorer differentiation (P = 0.0303), higher pathological TNM stage (p = 0.0153) and poorer prognosis of adenocarcinoma patients (P = 0.0017). Multivariable analysis with Cox proportional hazards models confirmed that MUC5B expression increased the hazard of death after adjusting for other clinicopathological factors (HR = 2.66; 95%CI, 1.26-5.61). We also immunohistochemically evaluated TTF-1 expression and found that the combination of MUC5B with TTF-1 is a useful marker for adenocarcinomas. The diagnostic accuracies of TTF-1 and MUC5B for adenocarcinoma were 83.8% and 70.4%, respectively. The accuracy increased to 94.3% when the two factors were combined. In survival analysis, the MUC5B(High)/TTF-1(-) group was significantly associated with a poorer outcome compared with the MUC5B(Low)/TTF-1(+) group (p < 0.0001). The present study suggested that the combination of MUC5B and TTF-1 expression is useful for discriminating adenocarcinomas from squamous cell carcinomas, yielding prognostic significance in patients with lung adenocarcinoma.

DOI： 10.1038/srep08649

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Language：English   Publishing type：Research paper (scientific journal)  

BACKGROUND: Identification of predictive markers for the efficacy of platinum-based chemotherapy is necessary to improve the quality of the life of cancer patients. MATERIALS AND METHODS: We detected proteins recognized by autoantibodies in pretreated sera from patients with lung adenocarcinoma (AC) evaluated as showing progressive disease (PD) or a partial response (PR) after cisplatin-based chemotherapy by proteomic analysis. Then, the levels of the candidate autoantibodies in the pretreated serum were validated by dot-blot analysis for 22 AC patients who received platinum-based chemotherapy, and the expression of identified proteins was immunohistochemically analyzed in 40 AC biopsy specimens. RESULTS: An autoantibody against galectin-3 (Gal-3) was detected in pretreated sera from an AC patient with PD. Serum IgG levels of anti-Gal-3 autoantibody were significantly higher in patients evaluated with PD than in those with PR and stable disease (SD) (p = 0.0084). Furthermore, pretreated biopsy specimens taken from patients evaluated as showing PD following platinum- based chemotherapy showed a tendency to have a higher positive rate of Gal-3 than those with PR and SD (p = 0.0601). CONCLUSIONS: These results suggest that serum IgG levels of anti-Gal-3 autoantibody may be useful to predict the efficacy of platinum-based chemotherapy for patients with lung AC.

PubMed

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BACKGROUND: Uroplakins have been widely investigated as potential markers in patients with bladder cancer because these proteins are specific to the urothelium. However, the role of uroplakin proteins in bladder cancer remains unknown. In this study, preoperative serum levels of uroplakin III were measured in patients with urothelial carcinoma of the urinary bladder and examined for possible association with clinicopathological features and clinical outcomes. MATERIALS AND METHODS: This study included 52 bladder cancer patients at various stages and 28 healthy controls. Uroplakin III levels were detected in preoperative sera using an automated dot blot system and a micro-dot blot array. RESULTS: There was a significant increase in serum uroplakin III levels in patients with bladder cancer as compared to healthy controls (p<0.05). In addition, serum uroplakin III levels were associated with muscle-invasive status, high grade and lymphovascular invasion (p<0.02). Log-rank tests indicated high serum uroplakin III to be significantly associated with cancer-specific mortality. CONCLUSIONS: Determination of serum uroplakin III level could be valuable for identifying patients with biologically aggressive bladder cancer.

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Language：Japanese   Publisher：Japanese Electrophoresis Society  

DOI： 10.2198/electroph.59.129

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Other Link： https://jlc.jst.go.jp/DN/JLC/20016223408?from=CiNii

Language：English   Publishing type：Research paper (scientific journal)  

Cancer tissues are comprised of various components including tumor cells and the surrounding tumor stroma, which consists of the extracellular matrix and inflammatory cells. Since the tumor stroma plays critical roles in tumor development, investigation of the tumor stroma in addition to tumor cells is important to identify useful tumor-associated markers. To discover novel and useful sero-diagnostic markers, a comparative study of tumor-associated autoantibodies (AAbs) in sera from lung adenocarcinoma (AC) patients was investigated by two-dimensional immunoblotting with AC cell lines or each autologous AC tissues. Autoantigens identified from tissue and cell line samples comprised 58 (45 antigens) and 53 spots (41 antigens), respectively. Thirty-six proteins including Transforming growth factor-beta-induced protein ig-h3 (BIGH3) and Hyaluronan and proteoglycan link protein 1 (HAPLN1) were detected only from tissues, 32 proteins only from cell lines, and 9 proteins from both. BIGH3 and HAPLN1 expressions were confirmed in the tumor stroma, but not in AC cell lines by immunostaining and immunoblotting. These data suggest that autologous tumor tissue and serum are important to coincidently detect AAbs derived from the tumor stroma in addition to tumor cells.

PubMed

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To identify diagnostic markers for psoriasis vulgaris and psoriatic arthritis, autoantibodies in sera from psoriasis vulgaris and psoriatic arthritis patients were screened by two-dimensional immunoblotting (2D-IB). Based on 2D-IB and MADLI TOF/TOF-MS analyses, eleven proteins each in psoriasis vulgaris and psoriatic arthritis were identified as autoantigens. Furthermore, serum levels of moesin, keratin 17 (K17), annexin A1 (ANXA1), and stress-induced phophoprotein-1 (STIP1), which were detected as autoantigens, were studied by dot blot analysis with psoriasis patients and healthy controls. The levels of moesin and STIP1 were significantly higher in sera from patients with psoriasis vulgaris than in the controls (moesin: P<0.05, STIP1: P<0.005). The area under the curve (AUC) for moesin and STIP1 between patients with psoraisis vulgaris and controls was 0.747 and 0.792, respectively. STIP1 and K17 levels were significantly higher in sera from patients with psoriatic arthritis than in those with psoriasis vulgaris (P<0.05 each). The AUC for STIP1 and K17 between patients with psoriatic arthritis and psoriasis vulgaris was 0.69 and 0.72, respectively. The STIP1 or moesin, CK17 serum level was not correlated with disease activity of psoriasis patients. These data suggest that STIP1 and moesin may be novel and differential sero-diagnostic markers for psoriasis vulgaris and psoriatic arthritis.

DOI： 10.1371/journal.pone.0101773

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Language：English   Publishing type：Research paper (scientific journal)  

To investigate the level of serum S100A6 in patients with bladder cancer and in healthy controls and compare these levels with clinicopathologic findings, we evaluated the level of serum S100A6 in 30 healthy controls and 50 patients with bladder cancer diagnosed via transurethral resection of bladder tumor and/or radical cystectomy. S100A6 in sera was detected by employing automatic dot blot systems, and the micro Dot Blot array with a 256-solid pin configuration. The normalized signal of serum S100A6 expression in bladder cancer patients was significantly higher than that of healthy controls (P = 0.001). Serum S100A6 expression of non-muscle-invasive cancer (NMIC) was significantly higher than that of healthy controls (P = 0.04). Furthermore, the S100A6 serum level in patients with muscle-invasive bladder cancer was significantly higher than that in patient with NMIC (P = 0.004). The sensitivity and specificity were 48.0% (95% CI: 0.337-0.626) and 93.3% (95% CI: 0.779-0.992), respectively. The area under the curve was 0.727. Serum S100A6 expression is a potentially effective detection marker for bladder cancer. Applying this serum marker to clinical practice would require less-invasive examinations of patients and would help to detect life-threatening cancerous lesions earlier than current modalities.

DOI： 10.2220/biomedres.35.351

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Language：Japanese   Publisher：Japanese Electrophoresis Society  

DOI： 10.2198/sbk.58.68

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Other Link： https://jlc.jst.go.jp/DN/JALC/10040886465?from=CiNii

Language：English   Publishing type：Research paper (scientific journal)  

To develop sero-diagnostic markers for lung cancer, we generated monoclonal antibodies using pulmonary adenocarcinoma (AD)-derived A549 cells as antigens by employing the random immunization method. Hybridoma supernatants were immunohistochemically screened for antibodies with AMeX-fixed and paraffin-embedded A549 cell preparations. Positive clones were monocloned twice through limiting dilutions. From the obtained monoclonal antibodies, we selected an antibody designated as KU-Lu-5 which showed intense membrane staining of A549 cells. Based on immunoprecipitation and MADLI TOF/TOF-MS analysis, this antibody was recognized as carbonic anhydrase XII (CAXII). To evaluate the utility of this antibody as a sero-diagnostic marker for lung cancer, we performed dot blot analysis with a training set consisting of sera from 70 lung cancer patients and 30 healthy controls. The CAXII expression levels were significantly higher in lung cancer patients than in healthy controls in the training set (P<0.0001), and the area under the curve of ROC was 0.794, with 70.0% specificity and 82.9% sensitivity. In lung cancers, expression levels of CAXII were significantly higher in patients with squamous cell carcinoma (SCC) than with AD (P = 0.035). Furthermore, CAXII was significantly higher in well- and moderately differentiated SCCs than in poorly differentiated ones (P = 0.027). To further confirm the utility of serum CAXII levels as a sero-diagnostic marker, an additional set consisting of sera from 26 lung cancer patients and 30 healthy controls was also investigated by dot blot analysis as a validation study. Serum CAXII levels were also significantly higher in lung cancer patients than in healthy controls in the validation set (P = 0.030). Thus, the serum CAXII levels should be applicable markers discriminating lung cancer patients from healthy controls. To our knowledge, this is the first report providing evidence that CAXII may be a novel sero-diagnostic marker for lung cancer.

DOI： 10.1371/journal.pone.0033952

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Language：Japanese   Publisher：Japanese Proteomics Society (Japan Human Proteome Organisation)  

DOI： 10.14889/jhupo.2011.0.166.0

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Language：Japanese   Publisher：Japanese Proteomics Society (Japan Human Proteome Organisation)  

DOI： 10.14889/jhupo.2011.0.167.0

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Language：Japanese   Publisher：日本プロテオーム学会（日本ヒトプロテオーム機構）  

DOI： 10.14889/jhupo.2010.0.115.1

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Event date： 2010.8

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