Extended Data Fig. 2: Anticancer activity of HDAC6 inhibitors.
(a) The western blot shows the titration of ricolinostat to identify an effective dose (accumulation of Ac-α-Tubulin) without off-target effects in class-I HDACs (accumulation of Ac-H3K27). SAHA is used as a control Pan-HDAC inhibitor. WT-blot results were reproduced n = 3 times from independent experiments. (b) Chemical structure of the different HDAC6 inhibitors used in Fig. 1e. (c) Normalized cell number 6 days after transfection with individualized siRNAs targeting HDAC6 or non-targeting control (NTC) (n = 3 independent independent experiments per siRNA). The WT-blots show the silencing efficiency. WT-blot results were reproduced n = 3 times from independent experiments. All error bars represent Mean±SD. P value was estimated by two-tailed t test. (d) The graphic shows the lack of synergistic activity between ricolinostat and commonly used chemotherapy (paclitaxel and doxorubicin) in MDA-MB-436 cells. In contrast, cells sensitive to ricolinostat MDA-MB-453, SK-BR-3 and MDA-MB-474 show synergistic activity between ricolinostat and chemotherapy. R and S indicate ricolinostat resistance and sensitivity respectively. N = 3 independent replicate experiments per drug combination and concentration. (e) Histological intratumoral evaluation of H&E, Caspase-3, and Ki-67 in tumor samples from Fig. 1b. Quantification is also shown in bar graphs. Notice that the combo treatment (Pac+Ric) is not shown because all tumors regressed with this treatment. The white asterisks indicate necrotic areas and the white arrows indicate Caspase-3 positive stained cells. All error bars represent Mean±SD. P value was estimated by two-tailed t test. N = 6 samples from individual tumors. (f) The list shows all the transgenic mouse models evaluated by the HDAC6 score in Fig. 2c, d and indicates their correlation with human BCs. (g) Kaplan–Meier graphic showing the survival of the MMTV tumors in Fig. 2e. Control n = 7; ricolinostat (Ric) n = 8; paclitaxel (Pac) n = 8; Ric+Pac n = 8. P value was estimated using two-tailed Log-Rank test.
