Development of a biologically relevant in vitro tumor model is a complex undertaking which requires culturing heterogeneous cell populations in a realistic 3D arrangement such that interactions normally occurring between different cell types (e.g., stromal cells and cancer cells) may be promoted in vitro. This level of complexity requires establishing and maintaining varied microenvironments in the culture that simulate the various microenvironmental niches present within the original tumor. Here, we demonstrate significant progress to this end by reporting results of successful efforts to co-culture heterogeneous cell populations derived from a metastatic pancreatic carcinoma tumor for 3 months using a novel, 3D perfusion culture system. The design of the culture system incorporates low-sheer, tangential flow of nutrient medium above and below an open synthetic 3D cell scaffold and exchange of metabolic gasses via gas-permeable membranes which separate the core of the culture chamber from gassing chambers located above and below the nutrient flow compartments. This design allows for the establishment of realistic gas and nutrient gradients across the culture scaffold that mimic nutrient, waste and gas gradients present in vivo. Primary tumor cultures were initiated in the 3D perfusion culture system and T-75 flasks (2D control cultures) after mechanical/enzymatic dissociation of a mouse xenograft tumor originating from a human pancreatic carcinoma that had metastasized to the liver. Each culture was seeded with an aliquot of the total tumor dispersion and maintained in culture for up to 90 days. The 3D perfusion cultures were maintained by performing partial medium exchanges as needed based on glucose concentration levels measured in the circulating medium while the 2D control cultures were maintained using traditional subculturing techniques as necessary based on confluence of the cell monolayers. A trend of increasing glucose consumption rate was observed throughout the duration of the study in the 3D tumor cultures suggesting continuous expansion of the cultured cell population without the need for subculturing. The cell populations observed in the 3D perfusion culture remained diverse throughout the 3 month culture period and the cells were found to arrange in a manner resembling the cellular organization of a corresponding tumor in vivo. 2D cultures established using cells removed from the 3D perfusion culture after 3 months closely resemble early passages of the 2D control cultures with respect to the composition and arrangement of cells. In contrast, the diversity of cell types present in the 2D control cultures decreased noticeably during the course of the study. These data suggest that the novel 3D perfusion culture system represents significant advancement over traditional culture technologies towards recapitulating human tumors in vitro.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4239. doi:1538-7445.AM2012-4239