Fig. 1: Metabolomic characterization of temozolomide resistance.

Growth of U87MG and NSP in the absence of TMZ (a), in the presence of 10 μM TMZ (b), 100 μM TMZ (c), and 700 μM TMZ (d). The growth rates of U87MG and NSP were calculated using the Gompertz function (lines inside the dot plots) using GraphPad Prism Software. e Extracellular lactate (Lac) to Pyruvate (Pyr) ratio and f Intracellular Lac/Pyr ratio. Absolute concentration ratios (shown as bar plots) were calculated and plotted to delineate Warburg effect. g Intracellular succinate (Succ) to a-keto glutarate (AKG) ratio. The absolute levels were normalized to cell number to estimate the values plotted. h Extracellular ornithine levels. NSP cells show higher levels of Ornithine irrespective of TMZ treatment, a phenotype that is inherent in NSP. The boxes in the boxplot indicate the upper and lower quartiles of the data and the middle line is the median with the whiskers extending to 1.5× interquartile range. Dots are the sample data points (n = 5 for time points 0 h, 24 h, 48 h, 72 h, and 96 h). i Exo-metabolome profile and j Endo-metabolome profile comparison across U87MG and NSP cells. The heatmap is plotted using absolute concentration (C) differences between 96 h (t96) and 0 h (t0), and calculated as C_t96–C_t0. These values are processed to indicate Z-score (color scale) using Euclidean clustering. k Exo-metabolome PCA and (l) endo-metabolome PCA showing the clusters according to the TMZ concentrations used for treatment as 0 μM, 10 μM, 100 μM, and 700 μM. m and n. VIP scores for growth essential metabolites according to Exo-metabolome (m) and Endo-metabolome (n) profiles. Error bars indicate mean ± s.d.