Pub Date : 2019-01-01DOI: 10.2198/JELECTROPH.63.15
Takuya Toki, Y. Kodera, Ryo Konno, Yoshiya Hirata, Tatsuya Saito, M. Shichiri
Autoimmune mechanisms have been hypothesized to underlie a number of human disorders in which both disease pathogenesis and diagnostic biomarkers remain poorly understood. This is partly due to a lack of efficient techniques for identification of plasma autoantibodies associated with specific pathophysiological conditions. We have developed a novel proteomic methodology to comprehensively identify plasma IgG-bound proteins using liquid chromatography tandem mass spectrometry (LC-MS/MS) after denaturing enriched plasma IgG to solubilize and release low molecular weight proteins. In total, we identified 44 proteins using this method that were undetectable in unprocessed plasma, 21 of which were not identified in the Human Plasma Proteome Draft of 2017. Comparison of plasma IgGbound proteins between healthy subjects and patients with isolated adrenocorticotropic hormone deficiency, a rare endocrine disorder speculated to involve autoimmune mechanisms, revealed several distinct IgG-bound proteins specifically detected in patient plasma but not in healthy subjects. Our results suggest that solubilization of low molecular weight proteins bound to enriched plasma IgG and subsequent proteomic analysis by LC-MS/MS could provide a promising strategy for identification of autoantigens in human peripheral blood.
{"title":"A novel strategy to identify autoantigens by proteomic analysis of plasma IgG-bound proteins","authors":"Takuya Toki, Y. Kodera, Ryo Konno, Yoshiya Hirata, Tatsuya Saito, M. Shichiri","doi":"10.2198/JELECTROPH.63.15","DOIUrl":"https://doi.org/10.2198/JELECTROPH.63.15","url":null,"abstract":"Autoimmune mechanisms have been hypothesized to underlie a number of human disorders in which both disease pathogenesis and diagnostic biomarkers remain poorly understood. This is partly due to a lack of efficient techniques for identification of plasma autoantibodies associated with specific pathophysiological conditions. We have developed a novel proteomic methodology to comprehensively identify plasma IgG-bound proteins using liquid chromatography tandem mass spectrometry (LC-MS/MS) after denaturing enriched plasma IgG to solubilize and release low molecular weight proteins. In total, we identified 44 proteins using this method that were undetectable in unprocessed plasma, 21 of which were not identified in the Human Plasma Proteome Draft of 2017. Comparison of plasma IgGbound proteins between healthy subjects and patients with isolated adrenocorticotropic hormone deficiency, a rare endocrine disorder speculated to involve autoimmune mechanisms, revealed several distinct IgG-bound proteins specifically detected in patient plasma but not in healthy subjects. Our results suggest that solubilization of low molecular weight proteins bound to enriched plasma IgG and subsequent proteomic analysis by LC-MS/MS could provide a promising strategy for identification of autoantigens in human peripheral blood.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"8 Pt 2 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83974663","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-01-01DOI: 10.2198/jelectroph.63.33
E. Hattori, T. Kondo
Proteogenomics is a novel approach to understand the molecular backgrounds of diseases. In cancer research, proteomic studies have been conducted without using the genome data of individual samples. For example, a common public database has always been used to identify proteins by mass spectrometry. However, tumor genomes, even tumors of the same type of cancer, can differ considerably, and such differences affect the response to treatments. Thus, genomic backgrounds should be considered when identifying proteins by mass spectrometry. In cancer proteogenomics, a virtual proteome database is generated using the genome data of identical samples for the mass spectrometric identification of proteins reflecting genetic mutations, which are not common and not cited in the commonly used databases. Such proteins are candidate biomarkers and therapeutic targets. Although previous studies have reported software capable of translating genomic data to proteomic data, a standard protocol has not been established. In addition, the utility of proteogenomics has also not been established, and it is not self-evident that proteins with mutations unique to certain groups can be exploited for innovative treatments or to provide clues for the resolution of biological problems in cancers. Collaborative efforts by cancer researchers and specialists in mass spectrometry and bioinformatics are required for fruitful advancements.
{"title":"Current status of cancer proteogenomics: a brief introduction","authors":"E. Hattori, T. Kondo","doi":"10.2198/jelectroph.63.33","DOIUrl":"https://doi.org/10.2198/jelectroph.63.33","url":null,"abstract":"Proteogenomics is a novel approach to understand the molecular backgrounds of diseases. In cancer research, proteomic studies have been conducted without using the genome data of individual samples. For example, a common public database has always been used to identify proteins by mass spectrometry. However, tumor genomes, even tumors of the same type of cancer, can differ considerably, and such differences affect the response to treatments. Thus, genomic backgrounds should be considered when identifying proteins by mass spectrometry. In cancer proteogenomics, a virtual proteome database is generated using the genome data of identical samples for the mass spectrometric identification of proteins reflecting genetic mutations, which are not common and not cited in the commonly used databases. Such proteins are candidate biomarkers and therapeutic targets. Although previous studies have reported software capable of translating genomic data to proteomic data, a standard protocol has not been established. In addition, the utility of proteogenomics has also not been established, and it is not self-evident that proteins with mutations unique to certain groups can be exploited for innovative treatments or to provide clues for the resolution of biological problems in cancers. Collaborative efforts by cancer researchers and specialists in mass spectrometry and bioinformatics are required for fruitful advancements.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"1 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"90397253","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Glioblastoma (GBM) is the most common brain tumor in adults. Although the surgical and chemoradiotherapy approaches for treatment have improved, the prognosis of patients with GBM is still poor and novel drugs are urgently required. Therefore, we investigated small molecular inhibitors to target GBM on the basis of gene expression data by using a Connectivity Map (CMAP)–based approach. Using meta-analysis performed with publically available gene expression data, we identified the gene expression signature of GBM. The CMAP analysis identified 15 candidate drugs for GBM treatment. We confirmed the anticancer cell proliferation activity of cantharidin as one of the top 15 drugs with high negative enrichment scores in CMAP analysis by using GBM cell lines. Our results indicate the potential utility of CMAP to discover the potent drugs in the GBM treatment. This approach can be applied to other malignancies than GBM.
{"title":"Identification of cantharidin as a drug candidate for glioblastoma by using a Connectivity Map–based approach","authors":"Zhiwei Qiao, T. Kondo","doi":"10.2198/JELECTROPH.63.9","DOIUrl":"https://doi.org/10.2198/JELECTROPH.63.9","url":null,"abstract":"Glioblastoma (GBM) is the most common brain tumor in adults. Although the surgical and chemoradiotherapy approaches for treatment have improved, the prognosis of patients with GBM is still poor and novel drugs are urgently required. Therefore, we investigated small molecular inhibitors to target GBM on the basis of gene expression data by using a Connectivity Map (CMAP)–based approach. Using meta-analysis performed with publically available gene expression data, we identified the gene expression signature of GBM. The CMAP analysis identified 15 candidate drugs for GBM treatment. We confirmed the anticancer cell proliferation activity of cantharidin as one of the top 15 drugs with high negative enrichment scores in CMAP analysis by using GBM cell lines. Our results indicate the potential utility of CMAP to discover the potent drugs in the GBM treatment. This approach can be applied to other malignancies than GBM.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"58 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75856298","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-01-01DOI: 10.2198/JELECTROPH.63.25
Hiroshi Kusamoto, Emiko Kinoshita-Kikuta, Tomoyo Nishimura, T. Nagai, E. Kinoshita, T. Koike
SUMMARY Phosphorylation, one of the most common post-translational modifications of proteins, plays a critical role in many biological processes. We have previously developed several analytical methods for determining the phosphorylation status of certain proteins by using a phosphate-capturing binuclear metal complex known as Phos-tag. Here, we describe a novel method for the gel-based in vitro analysis of the phosphorylation status of a protein by a simple and rapid fluorometric staining method that uses a tetramethylrhodamine (TAMRA)-labeled Phos-tag derivative (TAMRA–Phos-tag). The entire staining protocol, which requires less than 2 h to complete, uses three buffer solutions for staining, washing, and dilution, respectively, at room temperature. The gel-based analysis of phosphoproteins in a polyacrylamide gel can be conducted by using a fluorescence imaging scanner with a 532-nm excitation laser and a 580-nm longpass emission filter. As a practical example of the use of the TAMRA–Phos-tag staining method, we examined the time course of dephosphorylation of ovalbumin by an alkaline phosphatase. In addition, inhibitor profiling of a tyrosine kinase Abl was performed by using an Abl-substrate (GST-Abltide) and an Abl-inhibitor (Imatinib).
{"title":"Gel-based analysis of protein phosphorylation status by rapid fluorometric staining using TAMRA-labeled Phos-tag","authors":"Hiroshi Kusamoto, Emiko Kinoshita-Kikuta, Tomoyo Nishimura, T. Nagai, E. Kinoshita, T. Koike","doi":"10.2198/JELECTROPH.63.25","DOIUrl":"https://doi.org/10.2198/JELECTROPH.63.25","url":null,"abstract":"SUMMARY Phosphorylation, one of the most common post-translational modifications of proteins, plays a critical role in many biological processes. We have previously developed several analytical methods for determining the phosphorylation status of certain proteins by using a phosphate-capturing binuclear metal complex known as Phos-tag. Here, we describe a novel method for the gel-based in vitro analysis of the phosphorylation status of a protein by a simple and rapid fluorometric staining method that uses a tetramethylrhodamine (TAMRA)-labeled Phos-tag derivative (TAMRA–Phos-tag). The entire staining protocol, which requires less than 2 h to complete, uses three buffer solutions for staining, washing, and dilution, respectively, at room temperature. The gel-based analysis of phosphoproteins in a polyacrylamide gel can be conducted by using a fluorescence imaging scanner with a 532-nm excitation laser and a 580-nm longpass emission filter. As a practical example of the use of the TAMRA–Phos-tag staining method, we examined the time course of dephosphorylation of ovalbumin by an alkaline phosphatase. In addition, inhibitor profiling of a tyrosine kinase Abl was performed by using an Abl-substrate (GST-Abltide) and an Abl-inhibitor (Imatinib).","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"12 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74092630","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
There is a need for novel drugs for sarcoma treatment. In the present study, to identify inhibitors with potential therapeutic utility in sarcomas, we screened the growth inhibitory effects of 361 inhibitors, including experimental reagents and anti-cancer drugs approved for use in non-sarcoma malignancies and those under clinical trials. The inhibitors were initially tested using 10 osteosarcoma cell lines. The half-maximal inhibitory concentration (IC50) of leptomycin B, actinomycin D, chetomin, and staurosporine was <100 nM in all the cell lines. As the promiscuous effects of staurosporine on kinases make it unsuitable for clinical applications, the other three inhibitors were tested in an additional 15 sarcoma cell lines derived from synovial sarcoma, fibrosarcoma, liposarcoma, rhabdomyosarcoma, malignant peripheral nerve sheath tumor, leiomyosarcoma, and Ewing’s sarcoma. The IC50 of leptomycin B and actinomycin D was <100 nM in all cell lines and that of chetomin was <100 nM in all but three synovial sarcoma cell lines. Although the clinical development of leptomycin B, a chromosomal region maintenance (CRM)1/exportin (XPO)1 inhibitor, was discontinued because of toxicity, a previous clinical trial revealed that other CRM1/XPO1 inhibitors, such as selinexor, have anti-tumor effects in sarcomas. Actinomycin D has proven clinical utility in the treatment of sarcomas. Chetomin disrupts the interaction of hypoxia-inducible factor-1 with the transcriptional coactivator p300 and its clinical utility has not been established in sarcomas. Chetomin exhibited growth inhibitory effects on sarcoma cells with different histological subtypes. Library screening is a powerful approach to detect the potential utility of anti-cancer drugs in sarcoma treatment.
{"title":"Screening of a growth inhibitor library of sarcoma cell lines to identify potent anti-cancer drugs","authors":"Zhiwei Qiao, T. Kondo","doi":"10.2198/JELECTROPH.63.1","DOIUrl":"https://doi.org/10.2198/JELECTROPH.63.1","url":null,"abstract":"There is a need for novel drugs for sarcoma treatment. In the present study, to identify inhibitors with potential therapeutic utility in sarcomas, we screened the growth inhibitory effects of 361 inhibitors, including experimental reagents and anti-cancer drugs approved for use in non-sarcoma malignancies and those under clinical trials. The inhibitors were initially tested using 10 osteosarcoma cell lines. The half-maximal inhibitory concentration (IC50) of leptomycin B, actinomycin D, chetomin, and staurosporine was <100 nM in all the cell lines. As the promiscuous effects of staurosporine on kinases make it unsuitable for clinical applications, the other three inhibitors were tested in an additional 15 sarcoma cell lines derived from synovial sarcoma, fibrosarcoma, liposarcoma, rhabdomyosarcoma, malignant peripheral nerve sheath tumor, leiomyosarcoma, and Ewing’s sarcoma. The IC50 of leptomycin B and actinomycin D was <100 nM in all cell lines and that of chetomin was <100 nM in all but three synovial sarcoma cell lines. Although the clinical development of leptomycin B, a chromosomal region maintenance (CRM)1/exportin (XPO)1 inhibitor, was discontinued because of toxicity, a previous clinical trial revealed that other CRM1/XPO1 inhibitors, such as selinexor, have anti-tumor effects in sarcomas. Actinomycin D has proven clinical utility in the treatment of sarcomas. Chetomin disrupts the interaction of hypoxia-inducible factor-1 with the transcriptional coactivator p300 and its clinical utility has not been established in sarcomas. Chetomin exhibited growth inhibitory effects on sarcoma cells with different histological subtypes. Library screening is a powerful approach to detect the potential utility of anti-cancer drugs in sarcoma treatment.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"238 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77627883","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-01-01DOI: 10.2198/JELECTROPH.63.39
H. Maeda, Kana Kobayashi, Toshifumi Watanabe, M. Satoh, F. Nomura, K. Sogawa
SUMMARY Chronic kidney disease (CKD) is a common disorder and cause of death in cats. In the classification proposed by the International Renal Interest Society (IRIS), stage I and II CKD are difficult to diagnose accurately using markers, in comparison with normal controls. We recently described a simple and highly reproducible two-step method for identifying potential disease-marker candidates among low-abundance urine proteins. Urine samples were taken from 56 normal control cats as the control group and from 56 cats with CKD (stage I). A carboxylesterase 5A fragment and filaggrin-2 fragment were identified as two proteins with higher levels in normal control cats. The performance of the ELISA of urine carboxylesterase 5A fragment was satisfactory in terms of recovery (97.2–102.4%) and within-run (1.3–3.6%) and between-day (1.5–4.1%) reproducibility. Urine carboxylesterase 5A fragment levels were significantly greater in normal cats (3.4±0.6 mg/dL) than in CKD (stage I) (1.9±0.5 mg/dL) (p<0.001). A carboxylesterase 5A fragment may be useful as a complementary marker to P-Cre and BUN for detection of CKD (stage I).
{"title":"Urinary carboxylesterase 5A fragment as an early diagnostic marker of cat chronic kidney disease","authors":"H. Maeda, Kana Kobayashi, Toshifumi Watanabe, M. Satoh, F. Nomura, K. Sogawa","doi":"10.2198/JELECTROPH.63.39","DOIUrl":"https://doi.org/10.2198/JELECTROPH.63.39","url":null,"abstract":"SUMMARY Chronic kidney disease (CKD) is a common disorder and cause of death in cats. In the classification proposed by the International Renal Interest Society (IRIS), stage I and II CKD are difficult to diagnose accurately using markers, in comparison with normal controls. We recently described a simple and highly reproducible two-step method for identifying potential disease-marker candidates among low-abundance urine proteins. Urine samples were taken from 56 normal control cats as the control group and from 56 cats with CKD (stage I). A carboxylesterase 5A fragment and filaggrin-2 fragment were identified as two proteins with higher levels in normal control cats. The performance of the ELISA of urine carboxylesterase 5A fragment was satisfactory in terms of recovery (97.2–102.4%) and within-run (1.3–3.6%) and between-day (1.5–4.1%) reproducibility. Urine carboxylesterase 5A fragment levels were significantly greater in normal cats (3.4±0.6 mg/dL) than in CKD (stage I) (1.9±0.5 mg/dL) (p<0.001). A carboxylesterase 5A fragment may be useful as a complementary marker to P-Cre and BUN for detection of CKD (stage I).","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"42 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88461555","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-01-01DOI: 10.2198/JELECTROPH.62.21
Zhiwei Qiao, Cuneyd Parlayan, Shigeru Saito, T. Kondo
SUMMARY Sarcomas are rare mesenchymal malignancies and comprise over 50 histological subtypes. Sarcomas are not well studied because the number of cases of individual sarcoma is low. The utilization of public data, such as gene expression data, may allow for improvement in the novel discovery of sarcoma. In this study, to gain insight into histological subtypes of sarcoma from a public database, we performed a meta-analysis of the gene-expression profiles by survey-ing the data deposited in the Gene Expression Omnibus database from 2001 to 2014. The gene-expression data for 10 sarcoma subtypes and the gene-expression profiles for 1002 cases were selected for comparative analysis. Genes with histology-oriented molecular signatures were identified, and the results were verified by functional validation using gene oncology analysis. Pathway analysis suggested the existence of differential biological processes among sarcoma subtypes. Furthermore, as an application of the sarcoma gene expression datasets used in this study, we investigated the gene expression patterns of the targets of pazopanib to predict the response of sarcoma to pazopanib. We found that the gene expression distribution patterns of targets of pazopanib were without distinction among 10 subtypes of sarcoma. Taken together, we identified the tissue-specific genes of 10 subtypes of sarcoma by bioinformatics analysis; our results demonstrated the utility of sarcoma datasets in public databases and provide valuable information for future rare cancer research.
{"title":"Meta-analysis of global gene-expression profiles identify molecular signatures for histological subtypes of sarcomas","authors":"Zhiwei Qiao, Cuneyd Parlayan, Shigeru Saito, T. Kondo","doi":"10.2198/JELECTROPH.62.21","DOIUrl":"https://doi.org/10.2198/JELECTROPH.62.21","url":null,"abstract":"SUMMARY Sarcomas are rare mesenchymal malignancies and comprise over 50 histological subtypes. Sarcomas are not well studied because the number of cases of individual sarcoma is low. The utilization of public data, such as gene expression data, may allow for improvement in the novel discovery of sarcoma. In this study, to gain insight into histological subtypes of sarcoma from a public database, we performed a meta-analysis of the gene-expression profiles by survey-ing the data deposited in the Gene Expression Omnibus database from 2001 to 2014. The gene-expression data for 10 sarcoma subtypes and the gene-expression profiles for 1002 cases were selected for comparative analysis. Genes with histology-oriented molecular signatures were identified, and the results were verified by functional validation using gene oncology analysis. Pathway analysis suggested the existence of differential biological processes among sarcoma subtypes. Furthermore, as an application of the sarcoma gene expression datasets used in this study, we investigated the gene expression patterns of the targets of pazopanib to predict the response of sarcoma to pazopanib. We found that the gene expression distribution patterns of targets of pazopanib were without distinction among 10 subtypes of sarcoma. Taken together, we identified the tissue-specific genes of 10 subtypes of sarcoma by bioinformatics analysis; our results demonstrated the utility of sarcoma datasets in public databases and provide valuable information for future rare cancer research.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"53 1","pages":"21-29"},"PeriodicalIF":0.0,"publicationDate":"2018-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"72949290","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Tomohito Ayabe, Y. Motofuji, Asako Saito, Shinya Ayabe, M. Koike, Y. Kodera, T. Maeda, Y. Eishi, H. Komatsu
1 Department of Human Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan 2 Department of Clinical Laboratory Medicine, Faculty of Health Science Technology, Bunkyo Gakuin University, Tokyo, Japan 3 Department of Pathology, Shuuwa General Hospital, Saitama, Japan 4 Department of Physics, School of Science, Kitasato University, Kanagawa, Japan
{"title":"Characterization of panel antibodies for classification of cancer type using novel antibody-based phosphoproteomics","authors":"Tomohito Ayabe, Y. Motofuji, Asako Saito, Shinya Ayabe, M. Koike, Y. Kodera, T. Maeda, Y. Eishi, H. Komatsu","doi":"10.2198/JELECTROPH.62.1","DOIUrl":"https://doi.org/10.2198/JELECTROPH.62.1","url":null,"abstract":"1 Department of Human Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan 2 Department of Clinical Laboratory Medicine, Faculty of Health Science Technology, Bunkyo Gakuin University, Tokyo, Japan 3 Department of Pathology, Shuuwa General Hospital, Saitama, Japan 4 Department of Physics, School of Science, Kitasato University, Kanagawa, Japan","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"17 1","pages":"1-10"},"PeriodicalIF":0.0,"publicationDate":"2018-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74666847","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-01-01DOI: 10.2198/JELECTROPH.62.11
Yoko Ino, H. Kagawa, Tomoko Akiyama, Yusuke Nakai, Sakura Ito, M. Shimoda, Makiko Kawamura, H. Hirano, Y. Kimura
SUMMARY The SAINOME-plate consists of a 384-well plate and a cover that contains a cutter, which can cut a polyacryl-amide gel into approximately 4.5-mm square pieces following electrophoresis. In this study, we applied SDS-PAGE and the SAINOME-plate to fractionation of protein mixtures from cell extracts or serum for proteomic approaches. Compared with gel-fractionation using a cutter or a scalpel, SAINOME-plate gel-fractionation is simpler and higher-throughput. In terms of reproducibility of proteomic profiling, SAINOME-plate gel-fractionation was comparable to scalpel gel-fractionation. Additionally, human keratin contamination was lower with the SAINOME-plate than with a scalpel. In serum protein fractionation, the number of proteins identified increased approximately 2-fold and 3.7-fold relative to non-fractionation when the gel was divided into 8 and 96 fractions, respectively. The results demonstrate that the SAINOME-plate gel-fractionation will be a useful method in mass spectrometry-based proteomics.
{"title":"Protein fractionation for proteomics using the SAINOME-plate","authors":"Yoko Ino, H. Kagawa, Tomoko Akiyama, Yusuke Nakai, Sakura Ito, M. Shimoda, Makiko Kawamura, H. Hirano, Y. Kimura","doi":"10.2198/JELECTROPH.62.11","DOIUrl":"https://doi.org/10.2198/JELECTROPH.62.11","url":null,"abstract":"SUMMARY The SAINOME-plate consists of a 384-well plate and a cover that contains a cutter, which can cut a polyacryl-amide gel into approximately 4.5-mm square pieces following electrophoresis. In this study, we applied SDS-PAGE and the SAINOME-plate to fractionation of protein mixtures from cell extracts or serum for proteomic approaches. Compared with gel-fractionation using a cutter or a scalpel, SAINOME-plate gel-fractionation is simpler and higher-throughput. In terms of reproducibility of proteomic profiling, SAINOME-plate gel-fractionation was comparable to scalpel gel-fractionation. Additionally, human keratin contamination was lower with the SAINOME-plate than with a scalpel. In serum protein fractionation, the number of proteins identified increased approximately 2-fold and 3.7-fold relative to non-fractionation when the gel was divided into 8 and 96 fractions, respectively. The results demonstrate that the SAINOME-plate gel-fractionation will be a useful method in mass spectrometry-based proteomics.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"93 1","pages":"11-15"},"PeriodicalIF":0.0,"publicationDate":"2018-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"79619508","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-01-01DOI: 10.2198/JELECTROPH.62.17
Zhiwei Qiao, T. Kondo
Glioblastoma (GBM) is the most common malignant primary tumor of the central nervous system in adults. Despite advances in GBM treatment, the prognosis of patients with GBM remains poor and novel drugs are urgently required. In this study, we aimed to identify novel drugs for GBM treatment by using a drug screening approach. To this end, we performed high-throughput screening with 118 drugs, including Food and Drug Administration (FDA)-approved anticancer drugs. We found high inhibition rates (more than 90%) for doxorubicin, bortezomib, and cephalomannine in 6 GBM cell lines. Furthermore, we determined the half-maximal inhibitory concentration (IC50) of cephalomannine and found that the drug has a high potential for anti-GBM activity. Moreover, we noted that cephalomannine inhibited cell proliferation by inducing autophagy. Thus, our results indicate that cephalomannine may be an effective drug candidate for GBM treatment.
{"title":"Identification of cephalomannine as a drug candidate for glioblastoma via high-throughput drug screening","authors":"Zhiwei Qiao, T. Kondo","doi":"10.2198/JELECTROPH.62.17","DOIUrl":"https://doi.org/10.2198/JELECTROPH.62.17","url":null,"abstract":"Glioblastoma (GBM) is the most common malignant primary tumor of the central nervous system in adults. Despite advances in GBM treatment, the prognosis of patients with GBM remains poor and novel drugs are urgently required. In this study, we aimed to identify novel drugs for GBM treatment by using a drug screening approach. To this end, we performed high-throughput screening with 118 drugs, including Food and Drug Administration (FDA)-approved anticancer drugs. We found high inhibition rates (more than 90%) for doxorubicin, bortezomib, and cephalomannine in 6 GBM cell lines. Furthermore, we determined the half-maximal inhibitory concentration (IC50) of cephalomannine and found that the drug has a high potential for anti-GBM activity. Moreover, we noted that cephalomannine inhibited cell proliferation by inducing autophagy. Thus, our results indicate that cephalomannine may be an effective drug candidate for GBM treatment.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"20 1","pages":"17-20"},"PeriodicalIF":0.0,"publicationDate":"2018-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86476206","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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