Background: Circulating tumor cells (CTCs) play a key role in cancer progression, and in vitro CTC models are essential for studying their biology. This study examined extracellular vesicles (EVs) from CTC lines of a metastatic colorectal cancer (mCRC) patient who progressed despite therapy. Methods and results: EVs from different CTC lines showed size and morphological variations. Proteomic analysis revealed an enrichment of proteins linked to stemness, endosomal biogenesis, and mCRC prognosis. Integrins were notably enriched in EVs from post-therapy CTC lines. In vivo, EVs accumulated in the liver, lungs, kidneys, and bones, though their influence on CTC organotropism was unclear. Conclusions: This study highlights therapy-related changes in EVs from mCRC CTCs and their role in cancer spread. These findings suggest the utility of CTC-derived EVs in understanding cancer dissemination, though further validation in mCRC patients is needed.