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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV180001	1/2	Homo sapiens	Serum	Exiqon miRCURY exosome isolation kit Filtration	Kwok HH	2018	12%
Study summary Full title Transfer of Extracellular Vesicle-Associated-RNAs Induces Drug Resistance in ALK-Translocated Lung Adenocarcinoma. All authors Kwok HH, Ning Z, Chong PW, Wan TS, Ng MH, Ho GYF, Ip MS, Lam DC Journal Cancer (basel) Abstract Anaplastic lymphoma kinase (ALK) translocation is an actionable mutation in lung adenocarcino (show more...)Anaplastic lymphoma kinase (ALK) translocation is an actionable mutation in lung adenocarcinoma. Nonetheless tumour consists of heterogeneous cell subpopulations with diverse phenotypes and genotypes, and cancer cells can actively release extracellular vesicles (EVs) to modulate the phenotype of other cells in the tumour microenvironment. We hypothesized that EVs derived from a drug-resistant subpopulation of cells could induce drug resistance in recipient cells. We have established ALK-translocated lung adenocarcinoma cell lines and subclones. The subclones have been characterized and the expression of EV-RNAs determined by quantitative polymerase chain reaction. The effects of EV transfer on drug resistance were examined in vitro. Serum EV-RNA was assayed serially in two patients prescribed ALK-tyrosine kinase inhibitor (ALK-TKI) treatment. We demonstrated that the EVs from an ALK-TKI-resistant subclone could induce drug resistance in the originally sensitive subclone. EV-RNA profiling revealed that miRNAs miR-21-5p and miR-486-3p, and lncRNAs MEG3 and XIST were differentially expressed in the EVs secreted by the resistant subclones. These circulating EV-RNA levels have been found to correlate with disease progression of EML4-ALK-translocated lung adenocarcinoma in patients prescribed ALK-TKI treatment. The results from this study suggest that EVs released by a drug-resistant subpopulation can induce drug resistance in other subpopulations and may sustain intratumoural heterogeneity. (hide) EV-METRIC 12% (43rd percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Serum Sample origin Control condition Focus vesicles extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture Exiqon miRCURY exosome isolation kit Filtration Protein markers EV: CD63 non-EV: GP96 Proteomics no Show all info Study aim Function, Biomarker Sample Species Homo sapiens Sample Type Serum Separation Method Filtration steps 0.22µm or 0.2µm Commercial kit Exiqon miRCURY exosome isolation kit Other Name other separation method Exiqon miRCURY exosome isolation kit Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD63 Not detected contaminants GP96 Characterization: RNA analysis Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Mean Reported size (nm) 84

	EV180001	2/2	Homo sapiens	FA34	Exiqon miRCURY exosome isolation kit Filtration	Kwok HH	2018	12%
Study summary Full title Transfer of Extracellular Vesicle-Associated-RNAs Induces Drug Resistance in ALK-Translocated Lung Adenocarcinoma. All authors Kwok HH, Ning Z, Chong PW, Wan TS, Ng MH, Ho GYF, Ip MS, Lam DC Journal Cancer (basel) Abstract Anaplastic lymphoma kinase (ALK) translocation is an actionable mutation in lung adenocarcino (show more...)Anaplastic lymphoma kinase (ALK) translocation is an actionable mutation in lung adenocarcinoma. Nonetheless tumour consists of heterogeneous cell subpopulations with diverse phenotypes and genotypes, and cancer cells can actively release extracellular vesicles (EVs) to modulate the phenotype of other cells in the tumour microenvironment. We hypothesized that EVs derived from a drug-resistant subpopulation of cells could induce drug resistance in recipient cells. We have established ALK-translocated lung adenocarcinoma cell lines and subclones. The subclones have been characterized and the expression of EV-RNAs determined by quantitative polymerase chain reaction. The effects of EV transfer on drug resistance were examined in vitro. Serum EV-RNA was assayed serially in two patients prescribed ALK-tyrosine kinase inhibitor (ALK-TKI) treatment. We demonstrated that the EVs from an ALK-TKI-resistant subclone could induce drug resistance in the originally sensitive subclone. EV-RNA profiling revealed that miRNAs miR-21-5p and miR-486-3p, and lncRNAs MEG3 and XIST were differentially expressed in the EVs secreted by the resistant subclones. These circulating EV-RNA levels have been found to correlate with disease progression of EML4-ALK-translocated lung adenocarcinoma in patients prescribed ALK-TKI treatment. The results from this study suggest that EVs released by a drug-resistant subpopulation can induce drug resistance in other subpopulations and may sustain intratumoural heterogeneity. (hide) EV-METRIC 12% (33rd percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture Exiqon miRCURY exosome isolation kit Filtration Protein markers EV: CD63 non-EV: GP96 Proteomics no Show all info Study aim Function, Biomarker Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells FA34 EV-harvesting Medium EV-depleted serum Preparation of EDS Commercial EDS Separation Method Filtration steps 0.22µm or 0.2µm Commercial kit Exiqon miRCURY exosome isolation kit Other Name other separation method Exiqon miRCURY exosome isolation kit Characterization: Protein analysis Protein Concentration Method Bradford Protein Yield (µg) 50 Western Blot Antibody details provided? No Detected EV-associated proteins CD63 Not detected contaminants GP96 Characterization: RNA analysis Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Mean Reported size (nm) 109

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EV-TRACK ID	EV180001
species	Homo sapiens
sample type	Serum	Cell culture
cell type	NA	FA34
medium	NA	EV-depleted serum
condition	Control condition	Control condition
separation protocol	Exiqon miRCURY exosome isolation kit Filtration	Exiqon miRCURY exosome isolation kit Filtration
Exp. nr.	1	2
EV-METRIC %	12	12