American Journal of Respiratory Cell and Molecular Biology. Vol. 33, pp. 541-548, 2005. Published ahead of print on August 25, 2005. [Link]
Lung Biology Center or Department of Pathology, San Francisco General Hospital, University of California San Francisco; and Cancer Research Institute, UCSF Comprehensive Cancer Center, University of California San Francisco, San Francisco, California
Correspondence and requests for reprints should be addressed to V. Courtney Broaddus, M.D., Lung Biology Center, Box 0854 UCSF, San Francisco, CA 94143-0854. E-mail: email@example.com
Like many tumors, malignant mesothelioma exhibits significant chemoresistance and resistance to apoptosis in vivo that is not seen in current in vitro models. To study the mechanisms of this multicellular resistance, biologically relevant in vitro models are necessary. Therefore, we characterized and tested human mesothelioma tissue grown in vitro as tumor fragment spheroids. After 5–10 d in culture, fragments from each of 15 human mesothelioma tumors rounded into spheroids. The tumor fragment spheroids maintained multiple characteristics of the original tumors for up to 3 mo including the presence of viable mesothelioma cells, macrophages, and a collagen- rich stroma. In 14-d-old spheroids, mesothelioma cells showed the same proliferation rate and expression of a death receptor, DR5, as in the original tumor. To determine responses to treatment, we treated tumor fragment spheroids grown from three separate tumors with agents, TNF-related apoptosis-inducing ligand (TRAIL) plus cycloheximide, that induced near total apoptosis in three human mesothelioma cell lines (M28, REN, MS-1) grown as monolayers (94 ± 6% apoptosis; mean ± SEM). Compared with mesothelioma cells in monolayers, mesothelioma cells in the spheroids were resistant to TRAIL plus cycloheximide (32 ± 4% apoptosis; mean ± SEM). Apoptotic resistance of mesothelioma cells was significantly reduced by inhibiting either the PI3K/Akt pathway with LY294002 (47 ± 6% apoptosis) or the mTOR pathway with rapamycin (50 ± 17% apoptosis). We conclude that human mesothelioma can be maintained in vitro in a biologically relevant model that exhibits apoptotic resistance, thereby permitting study of its tumor biology and of novel approaches to therapy.
Key Words: collagen • death receptor DR5 • mTOR • multicellular resistance • PI3K/Akt survival pathway • TNF-related apoptosis-inducing ligand (TRAIL) • tumor-associated macrophage • tumor fragment spheroid