Fig 2: The HDC expression is regulated by FLI1. (A and B) ShFLI1 and control cells were subjected to RT-qPCR analysis for the expression of FLI1 (A) and HDC (B) genes. (C) ShFLI1 and scrambled control cells were examined for expression of FLI1 and HDC using Western blotting. Rd: Relative density determined using densitometer. (D) Relative expression of FLI1 protein in inducible K562-fli-1 cells after addition of Doxycycline. (E) Relative expression of HDC by RT-qPCR in K562-fli-1 cells after addition of doxycycline. (F) Chromatin immunoprecipitation (ChIp) analysis of the HDC promoter in HEL cells for binding to FLI1, by RT-qPCR (Lower panel). Upper panel represents the gel image of the immunoprecipitated PCR-amplified band relative to input. P=<0.01 (**), P=<0.001 (***) and P=<0.0001 (****) by two-tailed Student’s t-test.

Fig 3: Tetrandrine inhibits HDC through downregulation of FLI1. (A) Chemical structure of tetrandrine. (B and C) Tetrandrine inhibits FLI1 protein (B) and its transcript (C) in HEL cells in a dose dependent manner. (D) Tetrandrine inhibit HDC in a dose dependent manner, as determined by RT-qPCR. (E) Tetrandrine activated the expression of miR145 in a dose dependent manner. (F) Tetrandrine inhibits HEL cell proliferation in culture. (G) Tetrandrine induces cell death of HEL cells in a dose dependent manner. (G) Cells were photographed 24 h post-drug treatment (−50µm; magnification 20X). Black arrows shows death cells. P=<0.05 (*), P=<0.01 (**) and P=<0.001 (***) by two-tailed Student’s t-test.

Fig 4: The HDC knockdown does not affect cell proliferation in culture. (A and B) Relative expression of HDC in shHDC1-3 cells by RT-qPCR (A) and Western blot (B). (C) Cell proliferation index in scrambled and shHDC2 cells. (D and E) Cell proliferation of HEL cells in culture following addition of the indicated concentrations of famotidine (D) and histamine (E). (F) Cell proliferation in shHDC2 and control cells with or without addition of histamine. (G and H) Relative expression of globin genes HBA1 (G) and HBA2 (H) in shHDC2 and control cells, by RT-qPCR. P=<0.01 (**) and P=<0.001 (***) by two-tailed Student’s t-test.

Fig 5: Diacerein and tetrandrine inhibit leukemia progression in vivo. (A–D) Leukemic mice induced by F-MuLV were treated with 5 mg/kg histamine (A), 3 mg/kg famotidine (B), 3 mg/Kg diacerein (C) and 3 mg/Kg tetrandrine (D) for two weeks. Mice were observed for the development of leukemia and scored at the onset of death. The tumor growth was calculated by Student’s t-test and death was used to plot a Kaplan–Meier survival curve. (E) In this model, FLI1 controls HDC transcription, which itself regulates histamine production involved in allergenic reaction. HDC is also controls CXCR2 and IL1B associated with asthmatic reaction. These inflammatory factors likely effects leukemia progression through inflammation. While tetrandrine inhibits HDC through FLI1, EGC controls transcription of HDC through another mechanism.