Guttiferone K impedes cell cycle re-entry of quiescent prostate cancer cells via stabilization of FBXW7 and subsequent c-MYC degradation.

Z Xi,Q Dong,J Holst,H Tan,C Xie,H-X Xu,Y Lao,S Cai,Y Li,T Liu,M Yao

doi:10.1038/cddis.2016.123

Abstract

Cell cycle re-entry by quiescent cancer cells is an important mechanism for cancer progression. While high levels of c-MYC expression are sufficient for cell cycle re-entry, the modality to block c-MYC expression, and subsequent cell cycle re-entry, is limited. Using reversible quiescence rendered by serum withdrawal or contact inhibition in PTENnull/p53WT (LNCaP) or PTENnull/p53mut (PC-3) prostate cancer cells, we have identified a compound that is able to impede cell cycle re-entry through c-MYC. Guttiferone K (GUTK) blocked resumption of DNA synthesis and preserved the cell cycle phase characteristics of quiescent cells after release from the quiescence. In vehicle-treated cells, there was a rapid increase in c-MYC protein levels upon release from the quiescence. However, this increase was inhibited in the presence of GUTK with an associated acceleration in c-MYC protein degradation. The inhibitory effect of GUTK on cell cycle re-entry was significantly reduced in cells overexpressing c-MYC. The protein level of FBXW7, a subunit of E3 ubiquitin ligase responsible for degradation of c-MYC, was reduced upon the release from the quiescence. In contrast, GUTK stabilized FBXW7 protein levels during release from the quiescence. The critical role of FBXW7 was confirmed using siRNA knockdown, which impaired the inhibitory effect of GUTK on c-MYC protein levels and cell cycle re-entry. Administration of GUTK, either in vitro prior to transplantation or in vivo, suppressed the growth of quiescent prostate cancer cell xenografts. Furthermore, elevation of FBXW7 protein levels and reduction of c-MYC protein levels were found in the xenografts of GUTK-treated compared with vehicle-treated mice. Hence, we have identified a compound that is capable of impeding cell cycle re-entry by quiescent PTENnull/p53WT and PTENnull/p53mut prostate cancer cells likely by promoting c-MYC protein degradation through stabilization of FBXW7. Its usage as a clinical modality to prevent prostate cancer progression should be further evaluated.

Highlights

The presence of quiescent cancer cells has been documented in many types of tumors.[5,6] These quiescent cancer cells are defined clinically as being Ki-67 negative, a protein marker of cells in cell cycle,[7,8] and reside in a reversible G0 arrest state.[9]
We have identified a compound that is capable of impeding cell cycle re-entry by quiescent PTENnull/p53WT and PTENnull/p53mut prostate cancer cells likely by promoting v-myc avian myelocytomatosis viral oncogene homolog (c-MYC) protein degradation through stabilization of F-box and WD repeat domain containing 7 (FBXW7)
Another mechanism of c-MYC regulation is through FBXW7 (F-box and WD repeat domain containing 7, E3 ubiquitin protein ligase), which plays a key role in c-MYC protein degradation in a Thr58-dependent manner,[19] and this mechanism has been shown to play a critical role in leukemia-initiating cells.[20]

Summary

Introduction

The presence of quiescent cancer cells has been documented in many types of tumors.[5,6] These quiescent cancer cells are defined clinically as being Ki-67 negative, a protein marker of cells in cell cycle,[7,8] and reside in a reversible G0 arrest state.[9]. C-MYC mRNA levels are low in quiescent cells but increase within 1–3 h upon mitogen stimulation.[13] Forced expression of c-MYC induces cell cycle re-entry of quiescent cells, but the downregulation or inactivation of c-MYC results in the impairment of cell cycle progression.[14] Amplification of Abbreviations: GUTK, Guttiferone K; c-MYC, v-myc avian myelocytomatosis viral oncogene homolog; FBXW7, F-box and WD repeat domain containing 7; PTEN, phosphatase and tensin homolog; p53, tumor suppressor protein p53; ERK, extracellular signal-regulated kinase; GSK3β, glycogen synthase kinase 3 beta; AKT, v-akt murine thymoma viral oncogene homolog; Rb, retinoblastoma protein; PI, propidium iodide; SKP2, S-phase kinase-associated protein 2; DMSO, dimethyl sulfoxide; H&E, hematoxylin–eosin staining; siRNA, small interfering RNA; CHX, cycloheximide; Cont, control cells; Qsct, quiescent cells. We describe for the first time that GUTK impedes cell cycle re-entry of quiescent PTENnull/p53WT and PTENnull/p53mut prostate cancer cells via stabilization of FBXW7 and subsequent c-MYC degradation

Methods

Results

Conclusion