5'-Nitro-indirubinoxime induces G1 cell cycle arrest and apoptosis in salivary gland adenocarcinoma cells through the inhibition of Notch-1 signaling.
BACKGROUND
5'-Nitro-indirubinoxime (5'-NIO) is a new derivative of indirubin that exhibits anti-cancer activity in a variety of human cancer cells. However, its mechanism has not been fully clarified.


METHODS
Human salivary gland adenocarcinoma (SGT) cells were used in this study. Western blot and RT-PCR analyses were performed to determine cellular Notch levels. The cell cycle stage and level of apoptosis were analyzed using flow cytometry analysis.


RESULTS
5'-NIO significantly inhibited the mRNA levels of Notch-1 and Notch-3 and their ligands (Delta1, 2, 3, and Jagged-2) in SGT cells. Immunocytochemistry analysis showed that 5'-NIO specifically decreased the level of Notch-1 in the nucleus. In addition, 5'-NIO induced G1 cell cycle arrest by reducing levels of CDK4 and CDK6 in SGT cells. Using flow cytometry and immunoblotting analysis, we found that 5'-NIO induces apoptosis following the secretion of cytochrome c and the activation of caspase-3 and caspase-7. Intracellular Notch-1 overexpression led to a decrease in G1 phase arrest and an inhibition of 5'-NIO-induced apoptosis.


CONCLUSIONS
These observations suggest that 5'-NIO induces cell cycle arrest and apoptosis by down-regulating Notch-1 signaling.


CONCLUSIONS
This study identifies a new mechanism of 5'-NIO-mediated anti-tumor properties. Thus, 5'-NIO could be used as a candidate for salivary gland adenocarcinoma therapeutics.
Yoon
Ji-Hye
JH
Department of Pathology, School of Dentistry, Chosun University, 375 Seosuk-dong, Dong-gu, Gwangju 501-759, Republic of Korea. jhyoon@chosun.ac.kr
Kim
Soo-A
SA
Kwon
Seong-Min
SM
Park
Jong-Hwan
JH
Park
Hee-Sae
HS
Kim
Yong-Chul
YC
Yoon
Jung-Hoon
JH
Ahn
Sang-Gun
SG
D000230:Adenocarcinoma / Q000473:pathology*; D017209:Apoptosis / Q000187:drug effects*; D002453:Cell Cycle / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D017931:DNA Primers; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D010091:Oximes / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D051881:Receptor, Notch1 / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012468:Salivary Gland Neoplasms / Q000473:pathology*
C534713:5'-nitroindirubinoxime; D017931:DNA Primers; D007211:Indoles; C497761:NOTCH1 protein, human; D010091:Oximes; D051881:Receptor, Notch1