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10.1371/journal.pone.0199361 http://scihub22266oqcxt.onion/10.1371/journal.pone.0199361 C6013203!6013203 !29928062     free     free     free Warning: file_get_contents(https://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?dbfrom=pubmed&id=29928062 &cmd=llinks): Failed to open stream: HTTP request failed! 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Novel antibody reagents for characterization of drug- and tumor microenvironment-induced changes in epithelial-mesenchymal transition and cancer stem cells |pdf=|usr=}}{{29928062 }} gab.com Text Novel antibody reagents for characterization of drug- and tumor microenvironment-induced changes in epithelial-mesenchymal transition and cancer stem cells JO: PLoS One http://www.kidney.de/pget.php?&tw=1&pmid=29928062 #FOAvip The presence of cancer stem cells (CSCs) and the induction of epithelial-to-mesenchymal transition (EMT) in tumors are associated with tumor aggressiveness, metastasis, drug resistance, and poor prognosis, necessitating the development of reagents for unambiguous detection of CSC- and EMT-associated proteins in tumor specimens. To this end, we generated novel antibodies to EMT- and CSC-associated proteins, including Goosecoid, Sox9, Slug, Snail, and CD133. Importantly, unlike several widely used antibodies to CD133, the anti-CD133 antibodies we generated recognize epitopes distal to known glycosylation sites, enabling analyses that are not confounded by differences in CD133 glycosylation. For all target proteins, we selected antibodies that yielded the expected target protein molecular weights by Western analysis and the correct subcellular localization patterns by immunofluorescence microscopy assay (IFA); binding selectivity was verified by immunoprecipitation-mass spectrometry and by immunohistochemistry and IFA peptide blocking experiments. Finally, we applied these reagents to assess modulation of the respective markers of EMT and CSCs in xenograft tumor models by IFA. We observed that the constitutive presence of human hepatocyte growth factor (hHGF) in the tumor microenvironment of H596 non-small cell lung cancer tumors implanted in homozygous hHGF knock-in transgenic mice induced a more mesenchymal-like tumor state (relative to the epithelial-like state when implanted in control SCID mice), as evidenced by the elevated expression of EMT-associated transcription factors detected by our novel antibodies. Similarly, our new anti-CD133 antibody enabled detection and quantitation of drug-induced reductions in CD133-positive tumor cells following treatment of SUM149PT triple-negative breast cancer xenograft models with the CSC/focal adhesion kinase (FAK) inhibitor VS-6063. Thus, our novel antibodies to CSC- and EMT-associated factors exhibit sufficient sensitivity and selectivity for immunofluorescence microscopy studies of these processes in preclinical xenograft tumor specimens and the potential for application with clinical samples. Twit Text FOAVip Novel antibody reagents for characterization of drug- and tumor microenvironment-induced changes in epithelial-mesenchymal transition and cancer stem cells ????PLoS One http://www.kidney.de/pget.php?&tw=1&pmid=29928062 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=29928062 #FOAvip English Wikipedia <ref>{{Cite pmid|29928062 }}</ref> Novel antibody reagents for characterization of drug- and tumor microenvironment-induced changes in epithelial-mesenchymal transition and cancer stem cells #MMPMID29928062 Navas T ; Pfister TD ; Colantonio S ; Aziz A ; Dieckman L ; Saul RG ; Kaczmarczyk J ; Borgel S ; Alcoser SY ; Hollingshead MG ; Lee YH ; Bottaro DP ; Hiltke T ; Whiteley G ; Takebe N ; Kinders RJ ; Parchment RE ; Tomaszewski JE ; Doroshow JH PLoS One 2018[]; 13 (6 ): e0199361 PMID29928062 show ga The presence of cancer stem cells (CSCs) and the induction of epithelial-to-mesenchymal transition (EMT) in tumors are associated with tumor aggressiveness, metastasis, drug resistance, and poor prognosis, necessitating the development of reagents for unambiguous detection of CSC- and EMT-associated proteins in tumor specimens. To this end, we generated novel antibodies to EMT- and CSC-associated proteins, including Goosecoid, Sox9, Slug, Snail, and CD133. Importantly, unlike several widely used antibodies to CD133, the anti-CD133 antibodies we generated recognize epitopes distal to known glycosylation sites, enabling analyses that are not confounded by differences in CD133 glycosylation. For all target proteins, we selected antibodies that yielded the expected target protein molecular weights by Western analysis and the correct subcellular localization patterns by immunofluorescence microscopy assay (IFA); binding selectivity was verified by immunoprecipitation-mass spectrometry and by immunohistochemistry and IFA peptide blocking experiments. Finally, we applied these reagents to assess modulation of the respective markers of EMT and CSCs in xenograft tumor models by IFA. We observed that the constitutive presence of human hepatocyte growth factor (hHGF) in the tumor microenvironment of H596 non-small cell lung cancer tumors implanted in homozygous hHGF knock-in transgenic mice induced a more mesenchymal-like tumor state (relative to the epithelial-like state when implanted in control SCID mice), as evidenced by the elevated expression of EMT-associated transcription factors detected by our novel antibodies. Similarly, our new anti-CD133 antibody enabled detection and quantitation of drug-induced reductions in CD133-positive tumor cells following treatment of SUM149PT triple-negative breast cancer xenograft models with the CSC/focal adhesion kinase (FAK) inhibitor VS-6063. Thus, our novel antibodies to CSC- and EMT-associated factors exhibit sufficient sensitivity and selectivity for immunofluorescence microscopy studies of these processes in preclinical xenograft tumor specimens and the potential for application with clinical samples. |AC133 Antigen/metabolism [MESH] |Animals [MESH] |Antibodies, Monoclonal/biosynthesis/*pharmacology [MESH] |Antineoplastic Agents/*pharmacology/therapeutic use [MESH] |Benzamides/pharmacology/therapeutic use [MESH] |Carcinoma, Non-Small-Cell Lung/metabolism/pathology [MESH] |Cell Line, Tumor [MESH] |Epithelial-Mesenchymal Transition/*drug effects [MESH] |Female [MESH] |Gene Knock-In Techniques [MESH] |Hepatocyte Growth Factor/genetics [MESH] |Humans [MESH] |Indicators and Reagents [MESH] |Lung Neoplasms/metabolism/pathology [MESH] |Mice, Transgenic [MESH] |Neoplastic Stem Cells/drug effects/metabolism/*pathology [MESH] |Phenotype [MESH] |Pyrazines/pharmacology/therapeutic use [MESH] |Sulfonamides/pharmacology/therapeutic use [MESH] |Triple Negative Breast Neoplasms/drug therapy/pathology [MESH] |Tumor Microenvironment/*drug effects [MESH]Novel antibody reagents for characterization of drug- and tumor micr(epmc).pdf DeepDyve Pubget Overpricing