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Adult T-cell leukemia/lymphoma (ATL) is an intractable T-cell malignancy accompanied by massive invasion of lymphoma cells into various tissues. We demonstrate here that ATL cells cultured on a layer of epithelial-like feeder cells form anchorage-dependent multicellular aggregates (Ad-MCAs) and that a fraction of MCA-forming ATL cells acquire CD44 high cancer stem cell-like phenotypes. ATL cells forming Ad-MCAs displayed extracellular microvesicles with enhanced expression of CD44v9 at cell synapses, augmented expression of multidrug resistance protein 1, and increased NF-κB activity. Blockade of the NF-κB pathway dramatically reduced Ad-MCA formation by ATL cells and the emergence of CD44 high ATL cells, but left a considerable number of ATL cells adhering to the feeder layer. Disruption of vimentin cytoskeleton by treatment with withaferin A, a natural steroidal lactone, suppressed not only the adhesion of ATL cells to the feeder layer but also subsequent Ad-MCA formation by ATL cells, suggesting the involvement of vimentin in anchoring ATL cells to the feeder layer. Ad-MCA formation by ATL cells on a layer of epithelial-like feeder cells may mimic critical events that occur in metastatic colonization.

Pancreatic ductal adenocarcinoma (PDAC) reportedly progresses very rapidly through the initial carcinogenesis stages including DNA damage and disordered cell death. However, such oncogenic mechanisms are largely studied through observational diagnostic methods, partly because of a lack of live in vitro tumour imaging techniques. Here we demonstrate a simple live-tumour in vitro imaging technique using micro-patterned plates (micro/nanoplates) that allows dynamic visualisation of PDAC microtumours. When PDAC cells were cultured on a micro/nanoplate overnight, the cells self-organised into non-spheroidal microtumours that were anchored to the micro/nanoplate through cell-in-cell invasion. This self-organisation was only efficiently induced in small-diameter rough microislands. Using a time-lapse imaging system, we found that PDAC microtumours actively stretched to catch dead cell debris via filo/lamellipoedia and suction, suggesting that they have a sophisticated survival strategy (analogous to that of starving animals), which implies a context for the development of possible therapies for PDACs. The simple tumour imaging system visualises a potential of PDAC cells, in which the aggressive tumour dynamics reminds us of the need to review traditional PDAC pathogenesis.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal refractory cancers. Aggressive features in PDAC cells have been well studied, but those exhibited by a population of PDAC cells are largely unknown. We show here that coculture with epithelial-like feeder cells confers more malignant phenotypes upon PDAC cells forming anchorage-dependent multicellular aggregates (Ad-MCAs, a behavior of collective cells), in vitro. When CD44v3-10high/CD44slow PDAC cell lines, which exhibited an epithelial phenotype before the onset of epithelial–mesenchymal transition (EMT), were cocultured with a monolayer of HEK293T cells overnight, they formed Ad-MCAs on the feeder layer and acquired gemcitabine resistance. CD44v8-10 expression was dramatically increased and Ki-67 staining decreased, suggesting that PDAC cells forming Ad-MCAs acquired cancer stem cell (CSC)-like intractable properties. We found that highly downregulated genes in PDAC cells cocultured with HEK293T cells were significantly upregulated in malignant lesions from pancreatic cancer patients. Our work implies that PDAC cells forming Ad-MCAs partially return to a normal tissue gene profile before the onset of EMT. The collective cell behavior like Ad-MCA formation by PDAC cells may mimic critical events that occur in cancer cells at the very early phase of metastatic colonization.