title: In-depth characterization of the secretome of colorectal cancer metastatic cells identifies key proteins in cell adhesion, migration, and invasion.
abstract: Liver metastasis in colorectal cancer is the major cause of cancer-related deaths. To identify and characterize proteins associated with colon cancer metastasis, we have compared the conditioned serum-free medium of highly metastatic KM12SM colorectal cancer cells with the parental, poorly metastatic KM12C cells using quantitative stable isotope labeling by amino acids in cell culture (SILAC) analyses on a linear ion trap-Orbitrap Velos mass spectrometer. In total, 1337 proteins were simultaneously identified in SILAC forward and reverse experiments. For quantification, 1098 proteins were selected in both experiments, with 155 proteins showing >1.5-fold change. About 52% of these proteins were secreted directly or using alternative secretion pathways. GDF15, S100A8/A9, and SERPINI1 showed capacity to discriminate cancer serum samples from healthy controls using ELISAs. In silico analyses of deregulated proteins in the secretome of metastatic cells showed a major abundance of proteins involved in cell adhesion, migration, and invasion. To characterize the tumorigenic and metastatic properties of some top up- and down-regulated proteins, we used siRNA silencing and antibody blocking. Knockdown expression of NEO1, SERPINI1, and PODXL showed a significant effect on cellular adhesion. Silencing or blocking experiments with SOSTDC1, CTSS, EFNA3, CD137L/TNFSF9, ZG16B, and Midkine caused a significant decrease in migration and invasion of highly metastatic cells. In addition, silencing of SOSTDC1, EFNA3, and CD137L/TNFSF9 reduced liver colonization capacity of KM12SM cells. Finally, the panel of six proteins involved in invasion showed association with poor prognosis and overall survival after dataset analysis of gene alterations. In summary, we have defined a collection of proteins that are relevant for understanding the mechanisms underlying adhesion, migration, invasion, and metastasis in colorectal cancer.
authors: Functional Proteomics Laboratory, Centro de Investigaciones Biológicas, 28040 Madrid, Spain.
authors: Rodrigo
authors: R
authors: Barderas
authors: Marta
authors: M
authors: Mendes
authors: Sofia
authors: S
authors: Torres
authors: Rubén A
authors: RA
authors: Bartolomé
authors: María
authors: M
authors: López-Lucendo
authors: Roi
authors: R
authors: Villar-Vázquez
authors: Alberto
authors: A
authors: Peláez-García
authors: Eduardo
authors: E
authors: Fuente
authors: Félix
authors: F
authors: Bonilla
authors: J Ignacio
authors: JI
authors: Casal
mesh_terms: D014408:Biomarkers, Tumor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D002465:Cell Movement; D003106:Colon / Q000378:metabolism / Q000473:pathology; D003110:Colonic Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007553:Isotope Labeling; D008099:Liver / Q000378:metabolism / Q000473:pathology; D008113:Liver Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism / Q000556:secondary; D008297:Male; D013058:Mass Spectrometry; D009361:Neoplasm Invasiveness; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D011379:Prognosis; D025941:Protein Interaction Mapping; D020543:Proteome / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism
chemical_list: D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins; D020543:Proteome; D034741:RNA, Small Interfering