ABSTRACT
This corrects the article DOI: 10.1038/onc.2014.445.
ABSTRACT
The SET protein is a potent inhibitor of protein phosphatase 2A (PP2A). Here, we report the oncogenic role of SET in hepatocarcinogenesis, clinical aggressiveness and anti-hepatocellular carcinoma (HCC) therapeutics. By analyzing samples obtained from 147 HCC patients, we found that SET overexpression was detected specifically in 30.6% HCC tumor samples, and was significantly associated with worse clinical features and high p-Akt expression in HCC tumors. Co-expression of SET and Akt predicted shorter post-operative recurrence-free survival in this cohort (P=0.045). Furthermore, SET was significantly associated with cell growth and hepatosphere formation. To elucidate the anti-HCC potential of targeting SET, we generated a novel SET antagonist, EMQA (N(4)-(3-ethynylphenyl)-6,7-dimethoxy-N(2)-(4-phenoxyphenyl) quinazoline-2,4-diamine). EMQA enhanced PP2A activity via disrupting SET-PP2Ac (catalytic domain of PP2A) binding in HCC cells, which restored PP2A-mediated p-Akt downregulation and promoted HCC cell death. In HCC cells or recombinant proteins expressing the N- and C- truncated forms of SET, only the C-terminal SET was required for EMQA targeting. Furthermore, combining sorafenib and EMQA showed good synergism in inhibiting HCC survival. Our findings suggested the oncogenic role of SET and the adverse prognostic value of SET overexpression in HCC. This alteration defines a subgroup of HCC patients who could benefit from SET antagonists, such as EMQA.
Subject(s)
Carcinoma, Hepatocellular/drug therapy , Histone Chaperones/genetics , Liver Neoplasms/drug therapy , Prognosis , Quinazolines/administration & dosage , Transcription Factors/genetics , Animals , Apoptosis/drug effects , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Cell Proliferation/drug effects , DNA-Binding Proteins , Female , Gene Expression Regulation, Neoplastic/drug effects , Histone Chaperones/antagonists & inhibitors , Humans , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Male , Mice , Niacinamide/administration & dosage , Niacinamide/analogs & derivatives , Phenylurea Compounds/administration & dosage , Quinazolines/chemical synthesis , Sorafenib , Transcription Factors/antagonists & inhibitors , Xenograft Model Antitumor AssaysABSTRACT
Epithelial-to-mesenchymal transition (EMT) is well known to involve in tumor invasion and metastasis. Src homology region 2 domain-containing phosphatase 1 (SHP-1) functions as a potent tumor suppressor and also acts as a negative regulator of p-STAT3(Tyr705) oncogenic signaling. However, little is known about the molecular mechanism(s) through which SHP-1 regulates EMT during hepatocellular carcinoma (HCC) progression. Here we first reported that endogenous SHP-1 protein levels were significantly downregulated in cells with mesenchymal characteristics and negatively correlated with p-STAT3(Tyr705) and vimentin but positively correlated with E-cadherin. SHP-1 overexpression abolished transforming growth factor-ß1 (TGF-ß1)-induced p-STAT3(Tyr705) and EMT, as well inhibited migration and invasion but further rescued by signal transducer and activator of transcription factor 3 (STAT3) overexpression. Depletion of SHP-1 could induce a more increase in TGF-ß1-induced p-STAT3(Tyr-705) and EMT characteristics, further supporting the mechanism that suppression of TGF-ß1-induced EMT is dependent on SHP-1-mediated STAT3 inactivation. Constitutively overexpressed SHP-1 tyrosine phosphatase activity by D61A-mutated SHP-1 markedly reduced TGF-ß1-induced p-STAT3(Tyr705) and EMT features but was not altered by C453S catalytic-dead mutant SHP-1. Consequently, SHP-1 acted as a powerful suppressor in preventing EMT by exerting its tyrosine phosphatase activity that directly downregulated p-STAT3(Tyr705). Most notably, we discovered a novel SHP-1 agonist SC-43 better than sorafenib to exert more potent anti-EMT effects in vitro as well as anti-metastatic growth in vivo. In conclusion, SHP-1 is a potent suppressor of HCC EMT and metastasis, thus highlighting that SC-43-SHP-1 axis may serve as a potential therapeutic target that antagonized p-STAT3(Tyr705) and thereby prevented HCC EMT and metastasis.
Subject(s)
Carcinoma, Hepatocellular/enzymology , Epithelial-Mesenchymal Transition , Liver Neoplasms/enzymology , Lung Neoplasms/enzymology , Protein Tyrosine Phosphatase, Non-Receptor Type 6/physiology , Animals , Carcinoma, Hepatocellular/secondary , Cell Line, Tumor , Humans , Liver Neoplasms/pathology , Lung Neoplasms/secondary , Mice, Nude , Neoplasm Transplantation , STAT3 Transcription Factor/metabolismABSTRACT
Protein phosphatase 2A (PP2A) is a tumor suppressor, which is functionally defective in various cancers. Previously, we found that PP2A activity determined the anticancer effect of bortezomib and erlotinib in hepatocellular carcinoma (HCC) cells. Here, we tested a novel erlotinib derivative, TD52, in four HCC cell lines, PLC5, Huh-7, Hep3B and Sk-Hep1. Using MTT and flow cytometry, we showed that TD52 had more potent apoptotic effects than erlotinib in HCC cells. TD52-induced apoptosis was associated with dose- and time- dependent reactivation of PP2A and downregulation of cancerous inhibitor of protein phosphatase 2A (CIP2A) and p-Akt. Inhibition of PP2A or ectopic expression of CIP2A or Akt in PLC5 cells abolished the effects of TD52. Furthermore, we demonstrated that TD52 affected the binding of Elk-1 to the proximal promoter of the CIP2A gene, thus downregulating transcription of CIP2A. Importantly, TD52-induced tumor inhibition was associated with reactivation of PP2A and downregulation of CIP2A and p-Akt in vivo. In conclusion, we found that enhancement of PP2A activity by inhibition of CIP2A determines the apoptotic effect induced by TD52. Our findings disclose the therapeutic mechanism of this novel targeted agent, and suggest the therapeutic potential and feasibility of developing PP2A enhancers as a novel anticancer strategy.
Subject(s)
Antineoplastic Agents/pharmacology , Autoantigens/metabolism , Carcinoma, Hepatocellular/enzymology , Liver Neoplasms/enzymology , Membrane Proteins/metabolism , Protein Phosphatase 2/metabolism , Quinazolines/pharmacology , Animals , Antineoplastic Agents/chemistry , Apoptosis/drug effects , Autoantigens/genetics , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/physiopathology , Cell Line, Tumor , Erlotinib Hydrochloride , Gene Expression Regulation, Neoplastic/drug effects , Humans , Intracellular Signaling Peptides and Proteins , Liver Neoplasms/genetics , Liver Neoplasms/physiopathology , Male , Membrane Proteins/genetics , Mice , Mice, Nude , Protein Phosphatase 2/genetics , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Quinazolines/chemistry , Xenograft Model Antitumor AssaysABSTRACT
We investigated the molecular mechanisms underlying the effect of sorafenib and SC-59, a novel sorafenib derivative, on hepatocellular carcinoma (HCC). Sorafenib activated autophagy in a dose- and time-dependent manner in the HCC cell lines PLC5, Sk-Hep1, HepG2 and Hep3B. Sorafenib downregulated phospho-STAT3 (P-STAT3) and subsequently reduced the expression of myeloid cell leukemia-1 (Mcl-1). Inhibition of Mcl-1 by sorafenib resulted in disruption of the Beclin 1-Mcl-1 complex; however, sorafenib did not affect the amount of Beclin 1, suggesting that sorafenib treatment released Beclin 1 from binding with Mcl-1. Silencing of SHP-1 by small interference RNA (siRNA) reduced the effect of sorafenib on P-STAT3 and autophagy. Ectopic expression of Mcl-1 abolished the effect of sorafenib on autophagy. Knockdown of Beclin 1 by siRNA protected the cells from sorafenib-induced autophagy. Moreover, SC-59, a sorafenib derivative, had a more potent effect on cancer cell viability than sorafenib. SC-59 downregulated P-STAT3 and induced autophagy in all tested HCC cell lines. Furthermore, our in vivo data showed that both sorafenib and SC-59 inhibited tumor growth, downregulated P-STAT3, enhanced the activity of SHP-1 and induced autophagy in PLC5 tumors, suggesting that sorafenib and SC-59 activate autophagy in HCC. In conclusion, sorafenib and SC-59 induce autophagy in HCC through a SHP-1-STAT3-Mcl-1-Beclin 1 pathway.
