Visualising the dynamics of live pancreatic microtumours self-organised through cell-in-cell invasion (original) (raw)

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. Despite accumulating evidence on the pathological features exhibited by cancer cells in various carcinomas, recent in vitro cancer cell studies have focused on the behavior of single cells in isolation. In contrast, in analyses conducted at the tumor tissue level, in vivo methodologies still largely depend on observational diagnostic methods such as histopathological analysis and in vivo imaging systems using fluorescent imaging probes. Consequently, the underlying pathophysiological tumor dynamics in tissue remains mostly unclear. Thus, in current cancer research, directly linking in vitro cell-based studies with in vivo tissue-based pathological studies could result in a huge gap in our understanding. Indeed, we have been confronted with numerous unexpected difficulties in clinical trials of molecular-targeted anticancer agents for which there should be near perfect evidence for targets from conventional evaluations undertaken both in vivo and in vitro. Recently, the importance of using three-dimensional (3D) cell culture systems has remarkably increased in the field of anticancer drug development 1-4 ; 3D cell culture systems are expected to provide more physiologically relevant information to the in vivo setting as compared with traditional two-dimensional culture systems 5-8. PDAC, which constitutes approximately 90% of pancreatic cancers, is still one of the most lethal malignant tumours 9. KRAS mutation is the initiating genetic event for pancreatic intraepithelial neoplasia (PanINs), premalignant lesions of PDACs 10. Recent work has shown that once PDACs become detectable, they progress from T1 stage to T4 stage in approximately 14 months 11. PDACs rapidly progress through highly frequent DNA damage and mitotic abnormalities through unknown catastrophic events 12,13. Generally, the epithelial-mesenchymal