The autonomous notch signal pathway is activated by baicalin and baicalein but is suppressed by niclosamide in K562 cells

Abstract

The Notch signaling pathway plays important roles in a variety of cellular processes. Aberrant transduction of Notch signaling contributes to many diseases and cancers in humans. The Notch receptor intracellular domain, the activated form of Notch receptor, is extremely difficult to detect in normal cells. However, it can activate signaling at very low protein concentration to elicit its biological effects. In the present study, a cell based luciferase reporter gene assay was established in K562 cells to screen drugs which could modulate the endogenous CBF1-dependent Notch signal pathway. Using this system, we found that the luciferase activity of CBF1-dependent reporter gene was activated by baicalin and baicalein but suppressed by niclosamide in both dose- and time-dependent manners. Treatment with these drugs modulated endogenous Notch signaling and affected mRNA expression levels of Notch1 receptor and Notch target genes in K562 cells. Additionally, erythroid differentiation of K562 cells was suppressed by baicalin and baicalein yet was promoted by niclosamide. Colony-forming ability in soft agar was decreased after treatment with baicalin and baicalein, but was not affected in the presence of niclosamide. Thus, modulation of Notch signaling after treatment with any of these three drugs may affect tumorigenesis of K562 cells suggesting that these drugs may have therapeutic potential for those tumors associated with Notch signaling. Taken together, this system could be beneficial for screening of drugs with potential to treat Notch signal pathway-associated diseases.

Rhamnetin and Cirsiliol Induce Radiosensitization and Inhibition of Epithelial-Mesenchymal Transition (EMT) by miR-34a-mediated Suppression of Notch-1 Expression in Non-small Cell Lung Cancer Cell Lines

“…There have been many investigations for an application of flavonoids as anti-carcinogenic and radiosensitizing agents, because flavonoids display anti-inflammatory, anti-oxidant, anti-proliferative, and anti-bacterial activities (33,34). Some of the flavonoids, such as genistein, quercetin, baicalin, baicalein, and niclosamide, have been reported to regulate the Notch-1 pathway in various cancer cell lines (35)(36)(37)(38). Thus, we performed flavonoid screening and selected rhamnetin and cirsiliol as Notch-1-regulating candidates, which have been poorly studied in cancer.…”

MBP-1 Suppresses Growth and Metastasis of Gastric Cancer Cells through COX-2

“…Total RNA was isolated using Trizol reagent (Invitrogen, Carlsbad, CA), and cDNA was synthesized using Moloney murine leukemia virus reverse transcriptase (New England BioLabs, Beverly, MA) with an oligo (dT) 18 primer (Wang et al, 2009). The 305-base pairs COX-2 cDNA was amplified with primers 5Ј-TTCAAATGAGATTGTGGGAAAAT-3Ј and 5Ј-AGATCATCTCT-GCCTGAGTATCTT-3Ј.…”

“…cler FastStart DNA MasterPLUS SYBR Green I (Roche, Indianapolis, IN) (Wang et al, 2009). All data are shown as mean values and standard deviations from at least 3 independent experiments.…”

“…For transient transfection of luciferase reporter assay, K562 cells (5 ϫ 10 5 ) were seeded onto 6-well plates and transfected using SuperFect transfection reagent, (Qiagen, Valencia, CA) for two days. Luciferase activities were measured using the Dual-Luciferase reporter assay system (Promega, Madison, WI) and Renilla luciferase activity was used to normalize transfection efficiency (Wang et al, 2009).…”

“…The 176-base pairs GAPDH cDNA was amplified with primers 5Ј-AAATC-CCATCACCATCTTCC-3Ј and 5Ј-TCACACCCATGACGAACA-3Ј. Quantitative real-time PCR was performed using a LightCycler system with LightCy-cler FastStart DNA MasterPLUS SYBR Green I (Roche, Indianapolis, IN) (Wang et al, 2009). All data are shown as mean values and standard deviations from at least 3 independent experiments.…”