Metabolomics guides rational development of a simplified cell culture medium for drug screening against Trypanosoma brucei

ABSTRACT: In vitro culture methods underpin many experimental approaches to biology and drug discovery. The modification of established cell culture methods to be more biologically relevant, or to optimise growth, is traditionally a laborious task. Emerging metabolomics technology enables rapid evaluation of intra- and extra-cellular metabolites, and can be applied to the rational development of cell culture medium. In this study, untargeted semi-quantitative, and targeted quantitative, metabolomic analyses of fresh and spent media revealed the major nutritional requirements for growth of bloodstream-form Trypanosoma brucei. The standard culture medium (HMI11) contained unnecessarily high concentrations of 32 nutrients that were subsequently removed to more closely resemble concentrations normally found in blood. Our new minimal medium (CMM) supports in vitro growth equivalent to HMI11, and causes no significant perturbation of metabolite levels for 94% of the detected metabolome (< 3-fold change; a = 0.05). Importantly, improved sensitivity was observed for drug activity studies in whole cell phenotypic screens, and in metabolomics mode of action assays. 400-fold decreases in IC50 were observed for pentamidine and methotrexate, suggesting inhibition of activity by nutrients present in HMI11. CMM is suitable for routine cell culture and offers important advantages for metabolomics studies and drug activity screening.

INSTRUMENT(S): Exactive (Thermo Scientific)

PROVIDER: MTBLS49 | MetaboLights | 2015-05-14

REPOSITORIES: MetaboLights