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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV230029	1/2	Homo sapiens	MIA PaCa-2	(d)(U)C EX01-25L Exo-spin Standard Kit	Nannan, Lise	2024	67%
Study summary Full title Dysregulation of intercellular communication in vitro and in vivo via extracellular vesicles secreted by pancreatic duct adenocarcinoma cells and generated under the influence of the AG9 elastin peptide-conditioned microenvironment All authors Lise Nannan, Salomé Decombis, Christine Terryn, Sandra Audonnet, Jean Michel, Sylvie Brassart-Pasco, Willy Gsell, Uwe Himmelreich, Bertrand Brassart Journal J Extracell Biol Abstract Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with poor prognosis due to its h (show more...)Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with poor prognosis due to its highly metastatic profile. Intercellular communication between cancer and stromal cells via extracellular vesicles (EVs) is crucial for the premetastatic microenvironment preparation leading to tumour metastasis. This study shows that under the influence of bioactive peptides derived from the extracellular matrix microenvironment, illustrated here by the AG-9 elastin-derived peptide (EDP), PDAC cells secrete more tumour-derived EVs. Compared to PDAC-derived EVs, tumour-derived EVs resulting from AG-9 treatment (PDAC AG-9-derived EVs) significantly stimulated cell proliferation. At constant amount, tumour-derived EVs were similarly taken up by PDAC and HMEC-1 cells. Tumour-derived EVs stimulated cell proliferation, migration, proteinase secretion, and angiogenesis. Bioluminescence imaging allowed tumour-derived EV/FLuc+ tracking in vivo in a PDAC mouse model. The biodistribution of PDAC AG-9-derived EVs was different to PDAC-derived EVs. Our results demonstrate that the microenvironment, through EDP release, may not only influence the genesis of EVs but may also affect tumour progression (tumour growth and angiogenesis), and metastatic homing by modifying the in vivo biodistribution of tumour-derived EVs. They are potential candidates for targeted drug delivery and modulation of tumour progression, and they constitute a new generation of therapeutic tools, merging oncology and genic therapy. (hide) EV-METRIC 67% (94th 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 eGFP/FLuc overexpression 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 (Differential) (ultra)centrifugation Commercial method Protein markers EV: CD9/ CD63/ CD81 non-EV: GM130 Proteomics no Show all info Study aim Function/Biomarker/Mechanism of uptake/transfer/New methodological development Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MIA PaCa-2 EV-harvesting Medium Serum free medium Cell viability (%) 97 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: rotor type Type 70 Ti Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 0.1 Wash: time (min) 70 Wash: Rotor Type TLA-100.4 Wash: speed (g) 100000 Commercial kit EX01-25L Exo-spin Standard Kit Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD9/ CD63/ CD81 Not detected contaminants GM130 Flow cytometry aspecific beads Detected EV-associated proteins CD9/ CD63/ CD81 Flow cytometry specific beads Selected surface protein(s) CD9/ CD63/ CD81 Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Proteinase treatment No RNAse treatment Yes RNAse type RNase A RNAse concentration 0.004 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 120 EV concentration Yes Particle yield particles per milliliter of starting sample: 1.50E+08 EM EM-type Transmission-EM/ Immuno-EM EM protein CD63 Image type Close-up Report size (nm) 120

	EV230029	2/2	Homo sapiens	MIA PaCa-2	(d)(U)C EX01-25L Exo-spin Standard Kit	Nannan, Lise	2024	67%
Study summary Full title Dysregulation of intercellular communication in vitro and in vivo via extracellular vesicles secreted by pancreatic duct adenocarcinoma cells and generated under the influence of the AG9 elastin peptide-conditioned microenvironment All authors Lise Nannan, Salomé Decombis, Christine Terryn, Sandra Audonnet, Jean Michel, Sylvie Brassart-Pasco, Willy Gsell, Uwe Himmelreich, Bertrand Brassart Journal J Extracell Biol Abstract Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with poor prognosis due to its h (show more...)Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with poor prognosis due to its highly metastatic profile. Intercellular communication between cancer and stromal cells via extracellular vesicles (EVs) is crucial for the premetastatic microenvironment preparation leading to tumour metastasis. This study shows that under the influence of bioactive peptides derived from the extracellular matrix microenvironment, illustrated here by the AG-9 elastin-derived peptide (EDP), PDAC cells secrete more tumour-derived EVs. Compared to PDAC-derived EVs, tumour-derived EVs resulting from AG-9 treatment (PDAC AG-9-derived EVs) significantly stimulated cell proliferation. At constant amount, tumour-derived EVs were similarly taken up by PDAC and HMEC-1 cells. Tumour-derived EVs stimulated cell proliferation, migration, proteinase secretion, and angiogenesis. Bioluminescence imaging allowed tumour-derived EV/FLuc+ tracking in vivo in a PDAC mouse model. The biodistribution of PDAC AG-9-derived EVs was different to PDAC-derived EVs. Our results demonstrate that the microenvironment, through EDP release, may not only influence the genesis of EVs but may also affect tumour progression (tumour growth and angiogenesis), and metastatic homing by modifying the in vivo biodistribution of tumour-derived EVs. They are potential candidates for targeted drug delivery and modulation of tumour progression, and they constitute a new generation of therapeutic tools, merging oncology and genic therapy. (hide) EV-METRIC 67% (94th 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 eGFP/FLuc overexpression, AG-9 treated 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 (Differential) (ultra)centrifugation Commercial method Protein markers EV: CD9/ CD63/ CD81 non-EV: GM130 Proteomics no Show all info Study aim Function/Biomarker/Mechanism of uptake/transfer/New methodological development Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MIA PaCa-2 EV-harvesting Medium Serum free medium Cell viability (%) 97 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: rotor type Type 70 Ti Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 0.1 Wash: time (min) 70 Wash: Rotor Type TLA-100.4 Wash: speed (g) 100000 Commercial kit EX01-25L Exo-spin Standard Kit Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD9/ CD63/ CD81 Not detected contaminants GM130 Flow cytometry aspecific beads Detected EV-associated proteins CD9/ CD63/ CD81 Flow cytometry specific beads Selected surface protein(s) CD9/ CD63/ CD81 Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Proteinase treatment No RNAse treatment Yes RNAse type RNase A RNAse concentration 0.004 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 120 EV concentration Yes Particle yield particles per milliliter of starting sample: 1.50E+08 EM EM-type Transmission-EM Image type Close-up Report size (nm) 120

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EV-TRACK ID	EV230029
species	Homo sapiens
sample type	Cell culture
cell type	MIA PaCa-2
condition	eGFP/FLuc overexpression	eGFP/FLuc overexpression AG-9 treated
separation protocol	dUC/ EX01-25L Exo-spin Standard Kit	dUC/ EX01-25L Exo-spin Standard Kit
Exp. nr.	1	2
EV-METRIC %	67	67