Fig. 2

Therapy-induced metabolic reprogramming of LNCaP PCa cells is associated with cellular quiescence and multi drug tolerance. a LNCaP cells were treated for 0, 7, 14, and 21 days with AR antagonist enzalutamide (Enz, 10 μM), and samples were analyzed by microarray analysis. The number of differentially expressed genes between indicated pairwise comparisons are shown (FDR 1.5fold, p < 0.05). b Gene set enrichment analysis of the microarray data described in a for the analysis of indicated pathways. c LNCaP cells were treated as in a, and samples were analyzed by qRT-PCR (AR and PSA mRNA expression), live-cell confluence imaging (proliferation), CellTiter-Glo assay (cellular ATP levels), PrestoBlue assay (cellular reducing power), quantitative single-cell imaging (qSCI) of MitoTracker Orange CMTMRos (mitochondrial membrane potential, MMP), and dead/live staining (cell death) for functional characterization of Enz-induced metabolic changes (n = 3, activities relative to day 0 with the exception of AR levels, where levels were calculated relative to day 21). d LNCaP cells were cultured in growth media without (D0) or with Enz (10 μM, ENZ D14) for 14 days including the final 24 h in the presence of vehicle control (DMSO) or the indicated compounds (paclitaxel 2.5 μM, docetaxel 2.5 μM, doxorubicin 2.5 μM, etoposide 5 μM, edelfosine 7.5 μM, simvastatin 30 μM, TOFA 60 μM, bortezomib 2.5 μM). The percentage of dead cells was calculated based on qSCI of Hoechst 3342 (total cell count) and propidium iodide (dead cells) staining (n = 3 wells/treatment with > 4000 cells/well; ns, not significant, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, one-way ANOVA followed by Dunnett’s multiple comparisons test compared to D0, representative result of 2 independent experiments)