(B) Normalized apoptosis percentages in main AML cells with or without MSC co-culture. molecules that induce apoptosis through inhibiting the expression or activities of multiple targets. studies showed that this intravenous or intraperitoneal administration of NSC-743380 caused total tumor regression or P005672 HCl (Sarecycline HCl) significant growth suppression in xenograft tumor models derived from renal malignancy cell A498 and lung malignancy cell H157 [11, 12]. Upon correlation analysis of the anti-cancer activity of NSC-743380 and the gene expression levels in NCI-60 cell lines as well as functional characterization of the top genes associated with NSC-743380Cmediated anticancer activity, we recently reported that expression of the sulfotransferase SULT1A1, a biotransformation enzyme that bioactivates a number of pro-carcinogens [17], is usually causally associated with the selective anti-cancer activity of NSC-743380 [18]. Moreover, we found that a subset of leukemia cell lines, mostly AML cells, express SULT1A1 and are highly sensitive to NSC-743380. Nevertheless, whether NSC-743380 has similar mechanisms of action in AML cells as that observed in solid malignancy cells is unknown. The purpose of this study was to characterize molecular alterations induced by NSC-743380 in NSC-743380Csusceptible AML cell lines and measure the activity of NSC-743380 against main leukemia cell samples. Our P005672 HCl (Sarecycline HCl) results showed that NSC-743380 induces strong apoptosis, abrogates the expression of cFLIP, and inhibits the activity of the PI3K/AKT/mTOR pathway in AML cells and induces strong apoptosis in main AML samples that express SULT1A1. RESULTS NSC-743380 induces strong apoptosis in SULT1A1-expressing AML cells We previously reported that leukemia cells expressing SULT1A1, mostly AML cells, are susceptible to NSC-743380 treatment [18]. P005672 HCl (Sarecycline HCl) In a repeated cell viability assay, we found that treatment of cells from your SULT1A1-expressing AML cell lines U937, THP-1, and MV4-11 with NSC743380 P005672 HCl (Sarecycline HCl) resulted in growth inhibition, with IC50 values between 0.1 and 0.8 M (data not shown). In contrast, SULT1A1-unfavorable HL60 cells were resistant to NSC-743380, with IC50 above 10 M, the highest concentration tested. To determine whether NSC-743380Cmediated growth inhibition in these cells is usually caused by cell cycle arrest or apoptosis, we analyzed cell cycle profiles after the cells were treated with 0.1C3 M NSC-743380 for 24 h. Cells treated with DMSO were used as controls. In U937, THP-1, and MV4-11 cells (NSC-743380 sensitive), treatment with NSC-743380 led to dose-dependent apoptosis (sub-G1), which is usually significant when compared with control cells (P 0.05) (Figure ?(Figure1).1). Apoptosis was induced in 53%-94% of THP-1, MV4-11, and U937 cells with 3 M NSC-743380, whereas the control cells experienced background level of apoptotic cells ( 5%). In contrast, no significant increase in apoptosis occurred in HL-60 cells (NSC-743380 resistant) relative to the control cells. We also performed Western blot analysis to validate the apoptosis induction by NSC-743380. Treatment of THP-1 and U937 cells with NSC-743380 at 0.1 to 3 M triggered obvious cleavage of caspase 3, caspase 8, and poly(ADP-ribose) polymerase (PARP), demonstrating that NSC-743380 induced strong apoptosis in AML cells. Open in a separate window Physique 1 Apoptosis induction by NSC-743380U937, THP-1, MV4-11, and HL-60 cells were treated with different concentrations of NSC-743380 (0.1 to 3 M) for 24 h and then harvested and incubated with propidium iodide for 30 min. Apoptosis was detected by circulation cytometry analysis. (A) The number in CD207 each panel indicates the percentage of apoptotic cells. (B) Apoptosis percentages are shown, as means standard deviations. * P 0.05 when compared with DMSO treated cells. (C) Western blot analysis. THP-1 cells were treated with numerous concentrations of NSC-743380 as indicated for 24 h. Activation of caspase 3, caspase 8, and.