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. 2024 Sep 17;38(12):2561–2572. doi: 10.1038/s41375-024-02405-5

Fig. 5. BIM regulates MA49- and MA50-mediated FLT3-ITD degradation and heat shock protein levels.

Fig. 5

A MV4-11 cells were transfected with siRNA against the BCL2L11 mRNA to knock down the BIM protein. Cells without and with BIM knockdown were treated with 50 nM MA49 or MA50 for 24 h and analyzed by immunoblot for FLT3, BIM, pY591-FLT3, pY694-STAT5, cleaved (cl.) caspase-3, HSP110, HSP90, and HSP27. The protein levels of vinculin and β-actin were determined to verify the equal loading of samples. The data are representative for the outcome of four independent experiments; +, treated; -, untreated; cl., cleaved; p-, phosphorylated; kDa, molecular weight in kilodalton; siCtrl, siRNA control; siBCL2L11, siRNA targeting BIM, arrows point to the cleavage fragments of active caspase-3. B Quantification of FLT3 expression from A; siCtrl, siRNA control; siBCL2L11, BIM knockdown. One-way ANOVA statistical test was used to determine statistical significance; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. C MV4-11 cells transfected with siRNA against siBCL2L11 +/− 1 h 250 nM Onalespib pre-treatment were treated with 50 nM MA49 or MA50 for 24 h and analyzed by immunoblot for FLT3, BIM, pY591-FLT3, and HSP90 expression; siCtrl, siRNA control; siBCL2L11, BIM knockdown; kDa, molecular weight in kilodalton. The protein levels of α-tubulin and β-actin were determined to verify the equal loading of samples. The data are representative for the outcome of two independent experiments; +, treated; -, untreated; cl., cleaved; p-, phosphorylated; kDa, molecular weight in kilodalton; siCtrl, siRNA control; siBCL2L11, siRNA targeting BIM. D Model showing the pharmacological mechanisms of the TPDs described in this manuscript and the associated molecular mechanisms in the upper panel; the lower panel serves as legend.