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Tumour spheroids have the potential to be used as preclinicalchemosensitivity assays. However, the production of three dimensional (3D)tumour spheroids remains challenging as not all tumour cell lines formspheroids with regular morphologies and spheroid transfer often inducesdisaggregation. In the field of pancreatic cancer, the MiaPaCa-2 cell line isan interesting model for research but it is known for its difficulty to formstable spheroids; also, when formed, spheroids from this cell line are weak andarduous to manage and to harvest for further analyses such as multiple stainingand imaging. In this work, we compared different methods (i.e. hanging drop,round bottom wells and Matrigel embedding, each of them with or withoutmethylcellulose in the media) to evaluate which one allowed to better overpassthese limitations. Morphometric analysis indicated that hanging drop inpresence of methylcellulose leaded to well-organized spheroids; interestingly,quantitative PCR (qPCR) analysis reflected the morphometric characterization,indicating that same spheroids expressed the highest values of CD44, VIMENTIN,TGF beta1 and Ki67. In addition, we investigated the generation of MiaPaCa-2spheroids when cultured on substrates of different hydrophobicity, in order tominimize the area in contact with the culture media and to further improvespheroid formation.

A synergic approach to enhance long term culture and manipulation of MiaPaCa-2 pancreatic cancer spheroids