Background: Recently, the Notch signaling pathway has gained attention as a potential therapeutic
target for chemotherapeutic intervention. However, the efficacy of previously known Notch inhibitors in colon
cancer is still unclear. The purpose of this study was to investigate the effect of andrographolide on aberrantly
activated Notch signaling in SW-480 cells in vitro.
Methods: The cytostatic potential of andrographolide on SW-480 cells was evaluated by 3-(4,5-dimethylthiazol-
2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay, morphology assessment, and colony formation assay. The
apoptotic activity was evaluated by FITC Annexin V assay, 4′,6-diamidino-2-phenylindole (DAPI), Hoechst,
Rhodamine 123, and Mito Tracker CMXRos staining. Scratch assay was conducted for migratory potential
assessment. 7’-Dichlorodihydrofluorescein Diacetate (DCFH-DA) staining was used to evaluate the Reactive
Oxygen Species (ROS) generation. Relative mRNA expression of Bax, Bcl2, NOTCH 1, and JAGGED 1 was
estimated by Real-Time Quantitative Reverse Transcription PCR (qRT-PCR). Cell cycle phase distribution was
evaluated by Annexin V-FITC/PI staining.
Results: MTT assay demonstrated the dose and time-dependent cytotoxicity of andrographolide on SW-480
cells. It also inhibited the migratory and colony forming potential of SW-480 cells. Furthermore, andrographolide
also showed disruption of mitochondrial membrane potential and induced apoptosis through nuclear condensation.
Flow cytometric evaluation showed that andrographolide enhanced early and late apoptotic cells and
induced upregulation of pro-apoptotic (Bax and Bad) and downregulation of anti-apoptotic Bcl2 in treated SW-
480 cells. Andrographolide augmented intracellular ROS generation and induced G0/G1 phase cell cycle arrest in
colon cancer SW-480 cells. Furthermore, andrographolide repressed the Notch signaling by decreasing the expression
of NOTCH 1 and JAGGED 1.
Conclusion: The findings suggested that andrographolide constraint the growth of SW-480 cells through the
inhibition of the Notch signaling pathway.