GSK3β-dependent cyclin D1 and cyclin E1 degradation is indispensable for NVP-BEZ235 induced G0/G1 arrest in neuroblastoma cells

ABSTRACT

Cyclin D1 and cyclin E1, as vital regulatory factors of G1-S phase cell cycle progression, are frequently constitutive expressed and associated with pathogenesis and tumorigenesis in most human cancers and they have been regarded as promising targets for cancer therapy. In this study, we established NVP-BEZ235, a potent dual kinase inhibitor, could induce neuroblastoma cells proliferation inhibition without apoptosis activation. Moreover, we showed NVP-BEZ235 could induce neuroblastoma cells arrested at G0/G1 phase accompanied with significant reduction of the cyclin D1 and E1 proteins in a dose dependent manner at nanomole concentration. Additionally we found that GSK3β was dephosphorylated and activated by NVP-BEZ235 and then triggered cyclin D1 and cyclin E1 degradation through ubiquitination proteasome pathway, based on the evidences that NVP-BEZ235 induced downregulation of cyclin D1 and cyclin E1 were obviously recovered by proteasome inhibitor and the blockade of GSK3β contributed to remarkable rescue of cyclin D1 and cyclin E1. Analogous results about its anti-proliferation effects and molecular mechanism were observed on neuroblastoma xenograft mouse model in vivo. Therefore, these results indicate that NVP-BEZ235-induced cyclin D1 and cyclin E1 degradation, which happened through activating GSK3β, and GSK3β-dependent down-regulation of cyclin D1 and cyclin E1 should be available for anticancer therapeutics.

KEYWORDS:

• cyclin D1
• cyclin E1
• NVP-BEZ235
• GSK3β
• neuroblastoma

Abbreviations

4E-BP1	=	eukaryotic initiation factor 4E-binding protein 1
AKT	=	protein kinase B
ALK	=	anaplastic lymphoma kinase
CHX	=	Chlorhexidine
CQ	=	chloroquine
DMSO	=	dimethyl sulfoxide;GSK3β, glycogen synthase kinase-3β
GAPDH	=	glyceraldehyde-3-phosphate dehydrogenase
MCM2	=	Minichromosome maintenance protein2
mTOR	=	mammalian target of rapamycin
NB cells	=	neuroblastoma cells
p70s6k	=	ribosomal protein S6 kinase, 70kDa
PARP	=	Poly (ADP-ribose) polymerase
PI-3K	=	phosphatidylinositol 3 kinase

Declaration of interest statement

The authors enounce no conflict of interest in this work.

Additional information

Funding

This work was supported in part by the National Natural Science Foundation of China (81172322, 81302006), Shanghai Municipal Education Committee (13ZZ089), Science and Technology Committee of Shanghai (14401901500), Science and Technology Committee of Baoshan District (12-E-2), NO.3 People's Hospital affiliated to Shanghai Jiao-Tong University School of Medicine (sy2013-008).