Discovery of a Novel Microtubule Targeting Agent as an Adjuvant for Cancer Immunotherapy

Figure 4

Tubulin polymerization inhibiting compounds induce mitochondrial stress and show cytotoxicity in WT CEM cells. (a) Analysis of mitochondrial depolarization. WT CEM cells and TUBB mutant CEM-178 cells were treated with 0.1 μM #1 or colchicine for 72 h and stained with MitoTracker Red, which stains mitochondria in a membrane potential dependent manner. Cells were analyzed by flow cytometry and representative histograms are shown. (b) Cell numbers of WT CEM cells and TUBB mutant CEM-178 cells were measured by MTT assay. Cells were cultured with 0.1 μM compounds or 0.01 μM colchicine for 72 h. Each dot indicates an individual compound. Compounds were categorized into two clusters, cluster 1 (< 50% viability; n=13; closed circles) and cluster 2 (> 50% viability; n=10; open circles). #1 is the closed circle in black. Individual compound IDs are shown in Supplemental Figure 5. (b) NF-B activation shown in Figure 1(c) was reanalyzed according to the viability clusters. (c) ELISA for KC, IL-1β, IL-6, and IL-12 in BMDCs. BMDCs were cultured with 5 μM compounds overnight. P<0.0001 by Mann-Whitney U test for comparison of cluster 1 versus cluster 2. Data presented are representative of two independent experiments showing similar results.

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