Di 2 pyridylketone

di(2-pyridyl)ketone 4-methyl-3-thiosemicarbazone (Dp4mT, Figure 1C) was prepared according to a previously reported method [36 (link)]. di(2-pyridyl)ketone 4,4-dimethylsemicarbazone (Dp44mS; Supplemental Figure 4A) was synthesized by the reaction of di(2-pyridyl)ketone (Sigma-Aldrich) and 4,4-dimethylsemicarbazone, as described in the Supplementary Materials and Methods. N-cyclohexyl-N′-(di(pyridin-2-yl)methylene)-N-methylformohydrazonamide (DpC-A, Figure 1C) was prepared by the oxidation of the parent drug, DpC, with hydrogen peroxide according to a procedure described previously [32 (link)]. di(2-pyridyl)ketone 4-cyclohexyl-4-methylsemicarbazone (DpC-S; Supplemental Figure 4A) was prepared by the reaction of di(2-pyridyl)ketone (Sigma-Aldrich, Germany) with 4-cyclohexyl-4-methylsemicarbazide in ethanol.
The chemical structures described above and their purities were confirmed by ¹H and ¹³C NMR and MS-ESI⁺. The details of these syntheses and the characterization of standards are described in the Supplementary Material.

MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), ethidium bromide (EB), di-2-pyridylketone, RPMI-1640, and other chemicals were purchased from Sigma-Aldrich (Shanghai, China). LC3 antibody was obtained from Proteintech Group (Wuhan, China). Antibodies against cyclin D1, caspase 3, β-actin, Bax, and Bcl-2 were purchased from Boster (Wuhan, China).

MTT, acridine orange, di-2-pyridylketone, pifithrin-α, and monodansylcadaverine (MDC) were purchased from Sigma-Aldrich. LC3 antibody was obtained from Proteintech Group (Wuhan, China), caspase-8, GAPDH, bax, cytochrome c, and bcl-2 were purchased from Boster (Wuhan, China). RPMI 1640 and fetal bovine serum were purchased from Zhejiang Tianhang Biological Technology Co. Ltd.