Three human MPM cell lines (H28, H2452 and MSTO-211H) were obtained from ATCC (Manassas, VA) and cultured as previously described.26 (link) Human primary DMPM cultures (MesoII and STO) were derived from tumour samples of patients who underwent surgery, and were maintained in DMEM/F12 (Gibco) under standard culture conditions for less than 20 passages.24 (link) Cells were routinely tested for mycoplasma. Analyses of mitochondrial function and glycolysis of MSTO-211H and H2452 cells were performed with the Seahorse XFp Metabolic Flux Analyzer (Agilent Technologies, Santa Clara, CA), showing that these cells have both a normal aerobic metabolism and aerobic glycolysis, though to a different extent, as reported in the Supplemental Methods and Supplementary Fig. S1 .
Co-transduction of MesoII cells with luciferase vectors and Firefly-luciferase (F-luc) activity assessment were performed according to previously established methods.23 (link),24 (link) Pemetrexed and gemcitabine were gifts from Eli Lilly (Indianapolis, IN), while the LDH-A inhibitors NHI-2 and NHI-Glc-2 were synthesised as described previously.17 (link),27 (link) Drugs were dissolved in sterile water (gemcitabine) or dimethyl sulfoxide (DMSO, Pemetrexed and LDH-A inhibitors) and diluted in culture medium immediately before use. In each experiment, we did not use concentrations higher than 0.1% DMSO.
Co-transduction of MesoII cells with luciferase vectors and Firefly-luciferase (F-luc) activity assessment were performed according to previously established methods.23 (link),24 (link) Pemetrexed and gemcitabine were gifts from Eli Lilly (Indianapolis, IN), while the LDH-A inhibitors NHI-2 and NHI-Glc-2 were synthesised as described previously.17 (link),27 (link) Drugs were dissolved in sterile water (gemcitabine) or dimethyl sulfoxide (DMSO, Pemetrexed and LDH-A inhibitors) and diluted in culture medium immediately before use. In each experiment, we did not use concentrations higher than 0.1% DMSO.
Free full text: Click here
