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c-Jun is a proto-oncoprotein that is commonly overexpressed in many types of cancer and is believed to regulate cell proliferation, the cell cycle, and apoptosis by controlling AP-1 activity. Understanding the c-Jun regulation is important to develop treatment strategy for cancer. The COP9 signalosome subunit 6 (CSN6) plays a critical role in ubiquitin-mediated protein degradation. MEKK1 is a serine/threonine kinase and E3 ligase containing PHD/RING domain involved in c-Jun ubiquitination. Here, we show that CSN6 associates with MEKK1 and reduces MEKK1 expression level by facilitating the ubiquitin-mediated degradation of MEKK1. Also we show that CSN6 overexpression diminishes MEKK1-mediated c-Jun ubiquitination, which is manifested in mitigating osmotic stress-mediated c-Jun downregulation. Thus, CSN6 is involved in positively regulating the stability of c-Jun. Overexpression of CSN6 correlates with the upregulation of c-Jun target gene expression in cancer. These findings provide new insight into CSN6-MEKK1-c-Jun axis in tumorigenesis.

Cervical cancer is one of the leading causes of cancer death in women. Human papillomaviruses (HPVs) are the major cause in almost 99.7% of cervical cancer. E6 oncoprotein of HPV and E6-associated protein (E6AP) are critical in causing p53 degradation and malignancy. Understanding the E6AP regulation is critical to develop treating strategy for cervical cancer patients. The COP9 signalosome subunit 6 (CSN6) is involved in ubiquitin-mediated protein degradation. We found that both CSN6 and E6AP are overexpressed in cervical cancer. We characterized that CSN6 associated with E6AP and stabilized E6AP expression by reducing E6AP poly-ubiquitination, thereby regulating p53 activity in cell proliferation and apoptosis. Mechanistic studies revealed that CSN6-E6AP axis can be regulated by EGF/Akt signaling. Furthermore, inhibition of CSN6-E6AP axis hinders cervical cancer growth in mice. Taken together, our results indicate that CSN6 is a positive regulator of E6AP and is important for cervical cancer development.

Biomarkers for predicting chemotherapy response are important to the treatment of colorectal cancer patients. Cryptochrome 2 (CRY2) is a circadian clock protein involved in cell cycle, but the biologic consequences of this activity in cancer are poorly understood. We set up biochemical and cell biology analyses to analyze CRY2 expression and chemoresistance. Here, we report that CRY2 is overexpressed in chemoresistant colorectal cancer samples, and CRY2 overexpression is correlated with poor patient survival. Knockdown of CRY2 increased colorectal cancer sensitivity to oxaliplatin in colorectal cancer cells. We also identify FBXW7 as a novel E3 ubiquitin ligase for targeting CRY2 through proteasomal degradation. Mechanistic studies show that CRY2 is regulated by FBXW7, in which FBXW7 binds directly to phosphorylated Thr300 of CRY2. Furthermore, FBXW7 expression leads to degradation of CRY2 through enhancing CRY2 ubiquitination and accelerating the CRY2's turnover rate. High FBXW7 expression downregulates CRY2 and increases colorectal cancer cells' sensitivity to chemotherapy. Low FBXW7 expression is correlated with high CRY2 expression in colorectal cancer patient samples. Also, low FBXW7 expression is correlated with poor patient survival. Taken together, our findings indicate that the upregulation of CRY2 caused by downregulation of FBXW7 may be a novel prognostic biomarker and may represent a new therapeutic target in colorectal cancer. Mol Cancer Ther; 14(6); 1476–87. ©2015 AACR.

p27 is a critical CDK inhibitor involved in cell cycle regulation, and its stability is critical for cell proliferation. Constitutive photomorphogenic 1 (COP1) is a RING-containing E3 ubiquitin ligase involved in regulating important target proteins for cell growth, but its biological activity in cell cycle progression is not well characterized. Here, we report that p27Kip1 levels are accumulated in G1 phase, with concurrent reduction of COP1 levels. Mechanistic studies show that COP1 directly interacts with p27 through a VP motif on p27 and functions as an E3 ligase of p27 to accelerate the ubiquitin-mediated degradation of p27. Also, COP1-p27 axis deregulation is involved in tumorigenesis. These findings define a new mechanism for posttranslational regulation of p27 and provide insight into the characteristics of COP1-overexpressing cancers.

// Hyun Ho Choi 1, 2 , Chun-Hui Su 1 , Lekun Fang 1 , Jin Zhang 3 , Sai-Ching J. Yeung 4, 5 , Mong-Hong Lee 1, 2, 6, 7 1 Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA 2 Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, TX 77030, USA 3 Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center of Cancer, Tianjin, People's Republic of China 4 Department of Emergency Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA 5 Departiment of Endocrine Neoplasia and Hormonal Disorders, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA 6 Program in Cancer Biology, The University of Texas Graduate School of Biomedical Sciences at Houston, Houston, TX 77030, USA 7 Program in Genes and Development, The University of Texas Graduate School of Biomedical Sciences at Houston, Houston, TX 77030, USA Correspondence to: Mong-Hong Lee, e-mail: mhlee@mdanderson.org Keywords: p27, COP1, 14-3-3σ, ubiquitination, DNA damage Received: September 22, 2014 Accepted: January 17, 2015 Published: January 31, 2015 ABSTRACT Understanding genome integrity and DNA damage response are critical to cancer treatment. In this study, we identify CSN6's biological function in regulating genome integrity. Constitutive photomorphogenic 1 (COP1), an E3 ubiquitin ligase regulated by CSN6, is downregulated by DNA damage, but the biological consequences of this phenomenon are poorly understood. p27 Kip1 is a critical CDK inhibitor involved in cell cycle regulation, but its response to DNA damage remains unclear. Here, we report that p27 Kip1 levels are elevated after DNA damage, with concurrent reduction of COP1 levels. Mechanistic studies showed that during DNA damage response COP1's function as an E3 ligase of p27 is compromised, thereby reducing the ubiquitin-mediated degradation of p27 Kip1 . Also, COP1 overexpression leads to downregulation of p27 Kip1 , thereby promoting the expression of mitotic kinase Aurora A. Overexpression of Aurora A correlates with poor survival. These findings provide new insight into CSN6-COP1-p27 Kip1 -Aurora A axis in DNA damage repair and tumorigenesis.

Background: Insulin/insulin-like growth factor-1 signalling may underlie the promoting effect of type 2 diabetes on cancer. This study examined the association of diabetes, including steroid-induced diabetes (SID), and the impact of anti-diabetic medication on clinical outcomes of multiple myeloma (MM). Methods: A retrospective review was conducted of 1240 MM patients. Overall survival (OS) and MM disease status prior to death were analysed. Results: Diabetic patients had a significantly shorter OS than non-diabetic patients (median: 65.4 vs 98.7 months). In multivariate analysis, SID was a significant predictor of decreased OS, along with age, comorbidity, MM stage, and cytogenetic abnormalities. Analyzing only the diabetic MM patients, Cox regression showed that metformin predicted an increased OS, whereas use of insulin/analogues predicted a decreased OS. Competing risk analysis showed that DM was associated with increased cumulative incidence of death with progressive MM. Among the diabetics, multivariate regression showed that insulin/analogues were associated with increased, but metformin with decreased death with progressive MM. Potential immortal time bias was evaluated by landmark analyses. Conclusions: DM, SID in particular, is associated with poor clinical outcomes in MM. Insulin/analogues are associated with poor outcomes, whereas metformin is associated with improved outcomes. No conclusion about causal relationships can be made at this time. Managing hyperglycaemia with non-insulin regimens should be investigated in randomised trials.

HER2/neu oncogene is frequently deregulated in cancers, and the (PI3K)-Akt signaling is one of the major pathways in mediating HER2/neu oncogenic signal. p57 (Kip2) , an inhibitor of cyclin-depependent kinases, is pivotal in regulating cell cycle progression, but its upstream regulators remain unclear. Here we show that the HER2-Akt axis is linked to p57 (Kip2) regulation, and that Akt is a negative regulator of p57 (Kip2) . Ectopic expression of Akt can decrease the expression of p57 (Kip2) , while Akt inhibition leads to p57 (Kip2) stabilization. Mechanistic studies show that Akt interacts with p57 (Kip2) and causes cytoplasmic localization of p57 (Kip2) . Akt phosphorylates p57 on Ser 282 or Thr310. Akt activity results in destabilization of p57 by accelerating turnover rate of p57 and enhancing p57 ubiquitination. Importantly, the negative impact of HER2/Akt on p57 stability contributes to HER2-mediated cell proliferation, transformational activity and tumorigenicity. p57 restoration can attenuate these defects caused by HER2. Significantly, Kaplan-Meier analysis of tumor samples demonstrate that in tumors where HER2 expression was observed, high expression levels of p57 (Kip2) were associated with better overall survival. These data suggest that HER2/Akt is an important negative regulator of p57 (Kip2) , and that p57 restoration in HER2-overexpressing cells can reduce breast tumor growth. Our findings indicate the applicability of employing p57 regulation as a therapeutic intervention in HER2-overexpressing cancers.

Subunit 6 of the COP9 signalosome complex, CSN6, is known to be critical to the regulation of the MDM2-p53 axis for cell proliferation and anti-apoptosis, but its many targets remain unclear. Here we show that p57Kip2 is a target of CSN6, and that CSN6 is a negative regulator of p57Kip2. CSN6 associates with p57Kip2, and its overexpression can decrease the steady-state expression of p57Kip2; accordingly, CSN6 deficiency leads to p57Kip2 stabilization. Mechanistic studies show that CSN6 associates with p57Kip2 and Skp2, a component of the E3 ligase, which, in turn, facilitates Skp2-mediated protein ubiquitination of p57Kip2. Loss of Skp2 compromised CSN6-mediated p57Kip2 destabilization, suggesting collaboration between Skp2 and CSN6 in degradation of p57Kip2. CSN6’s negative impact on p57Kip2 elevation translates into cell growth promotion, cell cycle deregulation and potentiated transformational activity. Significantly, univariate Kaplan-Meier analysis of tumor samples demonstrates that high CSN6 expression or low p57 expression is associated with poor overall survival. These data suggest that CSN6 is an important negative regulator of p57Kip2, and that overexpression of CSN6 in many types of cancer could lead to decreased expression of p57Kip2 and result in promoted cancer cell growth.

HER2/neu oncogene is frequently overexpressed in various types of cancer, and the (PI3K)-Akt signaling pathway is often activated in HER2-overexpressing cancer cells. CSN6, subunit 6 of the COP9 signalosome complex, is pivotal in regulating MDM2 to destabilize p53, but its upstream regulators remain unclear. Here we show that the HER2-Akt axis is linked to CSN6 regulation, and that Akt is a positive regulator of CSN6. Ectopic expression of Akt can increase the expression of CSN6; accordingly, Akt inhibition leads to CSN6 destabilization. Mechanistic studies show that Akt causes CSN6 phosphorylation at Ser 60, which, in turn, reduces ubiquitin-mediated protein degradation of CSN6. Significantly, Akt’s positive impact on CSN6 elevation translates into p53 degradation, potentiating transformational activity and increasing DNA damage. Akt inhibition can attenuate these defects caused by CSN6. These data suggest that Akt is an important positive regulator of CSN6, and that activation of Akt in many types of cancer could lead to abnormal elevation of CSN6 and result in downregulated p53 and increased DNA damage, which promotes cancer cell growth.

Background Insulin/insulin-like growth factor-I (IGF-I) signaling is a mechanism mediating the promoting effect of type 2 diabetes (DM2) on cancer. Human epidermal growth factor receptor (HER2), insulin receptor and IGF-I receptor involve the same PI3K/AKT/mTOR signaling, and different antidiabetic pharmacotherapy may differentially affect this pathway, leading to different prognoses of HER2+ breast cancer. Methods We reviewed 1983 consecutive patients with HER2+ breast cancer treated between 1 January 1998 and 30 September 2010. The overall survival, breast cancer-specific death rate, age, race, nuclear grade, stage, menopausal status, estrogen and progesterone receptor status, body mass index and classes of antidiabetic pharmacotherapy were analyzed. Results A Cox regression analysis showed that DM2 [P = 0.026, hazard ratio (HR) = 1.42, 95 % confidence interval (95 % CI) 1.04–1.94] predicted poor survival of stage ≥2 HER2+ breast cancer. In Kaplan–Meier analysis, metformin predicted lengthened survival and so did thiazolidinediones. Analyzing only the diabetics, Cox regression showed that metformin (P = 0.041, HR = 0.52, 95 % CI 0.28–0.97) and thiazolidinediones (P = 0.036; HR = 0.41, 95 % CI 0.18–0.93) predicted lengthened survival, and competing risk analysis showed that metformin and thiazolidinediones were associated with decreased breast cancer-specific mortality (P = 0.023, HR = 0.47, 95 % CI 0.24–0.90 and P = 0.044, HR = 0.42, 95 % CI 0.18–0.98, respectively). Conclusions Thiazolidinediones and metformin users are associated with better clinical outcomes than nonusers in diabetics with stage ≥2 HER2+ breast cancer. The choice of antidiabetic pharmacotherapy may influence prognosis of this group.

Hyperglycemia during hyper-CVAD chemotherapy is associated with poor outcomes of acute lymphoblastic leukemia (ALL) (Cancer 2004; 100: 1179-85). The optimal clinical strategy to manage hyperglycemia during hyper-CVAD is unclear. To examine whether anti-diabetic pharmacotherapy can influence chemosensitivity of ALL cells, we examined the impacts of different anti-diabetic agents on ALL cell lines and patient samples. Pharmacologically achievable concentrations of insulin, aspart and glargine significantly increased the number of ALL cells, and aspart and glargine did so at lower concentrations than human insulin. In contrast, metformin and rosiglitazone significantly decreased the cell number. Human insulin and analogs activated AKT/mTOR signaling and stimulated ALL cell proliferation (as measured by flow cytometric methods), but metformin and rosiglitazone blocked AKT/mTOR signaling and inhibited proliferation. Metformin 500 μM and rosiglitazone 10 μM were found to sensitize Reh cells to daunorubicin, while aspart, glargine and human insulin (all at 1.25 mIU/L) enhanced chemoresistance. Metformin and rosiglitazone enhanced daunorubicin-induced apoptosis, while insulin, aspart and glargine antagonized daunorubicin-induced apoptosis. In addition, metformin increased etoposide-induced and L-asparaginase-induced apoptosis; rosiglitazone increased etoposide-induced and vincristine-induced apoptosis. In conclusion, our results suggest that use of insulins to control hyperglycemia in ALL patients may contribute to anthracycline chemoresistance, while metformin and thiazolidinediones may improve chemosensitivity to anthracycline as well as other chemotherapy drugs through their different impacts on AKT/mTOR signaling in leukemic cells. Our data suggest that the choice of anti-diabetic pharmacotherapy during chemotherapy may influence clinical outcomes in ALL.

Radiotherapy is the primary treatment for nasopharyngeal carcinoma (NPC), but radioresistance severely reduces NPC radiocurability. Here, we have established a radio-resistant NPC cell line, CNE-2R, and investigate the role of miRNAs in radioresistance. The miRNAs microarray assay reveals that miRNAs are differentially expressed between CNE-2R and its parental cell line CNE-2. We find that miR-205 is elevated in CNE-2R. A target prediction algorithm suggests that miR-205 regulates expression of PTEN, a tumor-suppressor. Introducing miR-205 into CNE-2 cells suppresses PTEN protein expression, followed by activation of AKT, increased number of foci formation and reduction of cell apoptosis post-irradiation. On the other hand, knocking down miR-205 in CNE-2R cells compromises the inhibition of PTEN and increases cell apoptosis. Significantly, immunohistochemistry studies demonstrate that PTEN is downregulated at late stages of NPC, and that miR-205 is significantly elevated followed the radiotherapy. Our data conclude that miR-205 contributes to radioresistance of NPC by directly targeting PTEN. Both miR-205 and PTEN are potential predictive biomarkers for radiosensitivity of NPC and may serve as targets for achieve successful radiotherapy in NPC.

14-3-3σ, a gene upregulated by p53 in response to DNA damage, exists as part of a positive-feedback loop, which activates p53 and is a human cancer epithelial marker downregulated in various cancer types. 14-3-3σ levels are critical for maintaining p53 activity in response to DNA damage and regulating signal mediators such as Akt. In this study, we identify mammalian constitutive photomorphogenic 1 (COP1) as a novel E3 ubiquitin ligase for targeting 14-3-3σ through proteasomal degradation. We show for the first time that COP9 signalosome subunit 6 (CSN6) associates with COP1 and is involved in 14-3-3σ ubiquitin-mediated degradation. Mechanistic studies show that CSN6 expression leads to stabilization of COP1 through reducing COP1 self-ubiquitination and decelerating COP1's turnover rate. We also show that CSN6-mediated 14-3-3σ ubiquitination is compromised when COP1 is knocked down. Thus, CSN6 mediates 14-3-3σ ubiquitination through enhancing COP1 stability. Subsequently, we show that CSN6 causes 14-3-3σ downregulation, thereby activating Akt and promoting cell survival. Also, CSN6 overexpression leads to increased cell growth, transformation and promotes tumorigenicity. Significantly, 14-3-3σ expression can correct the abnormalities mediated by CSN6 expression. These data suggest that the CSN6-COP1 axis is involved in 14-3-3σ degradation, and that deregulation of this axis will promote cell growth and tumorigenicity.

Metastasis-associated protein 1 (MTA1), a component of the nucleosome remodeling and histone deacetylation (NuRD) complex, is widely upregulated in human cancers. However, the mechanism for regulating its protein stability remains unknown. Here we report that MTA1 is an ubiquitinated protein and targeted by the RING-finger E3 ubiquitin–protein ligase constitutive photomorphogenesis protein 1 (COP1) for degradation via the ubiquitin–proteasome pathway. Induced expression of wild-type COP1 but not its RING motif mutants promotes the ubiquitination and degradation of MTA1, indicating that the ligase activity is required for the COP1-mediated proteolysis of MTA1. Conversely, depletion of endogenous COP1 resulted in a marked decrease in MTA1 ubiquitination, accompanied by a pronounced accumulation of MTA1 protein. MTA1, in turn, destabilizes COP1 by promoting its autoubiquitination, thus creating a tight feedback loop that regulates both MTA1 and COP1 protein stability. Accordingly, disruption of the COP1-mediated proteolysis by ionizing radiation leads to MTA1 stabilization, accompanied by an increased coregulatory function of MTA1 on its target. Furthermore, we discovered that MTA1 is required for optimum DNA double-strand break repair after ionizing radiation. These findings provide novel insights into the regulation of MTA1 protein and reveal a novel function of MTA1 in DNA damage response.

Tumor hypoxia directly promotes genomic instability and facilitates cell survival, resulting in tumors with a more aggressive phenotype. The proto-oncogene pim-1 regulates apoptosis and the cell cycle by phosphorylating target proteins. Overexpression of Pim-1 can cause genomic instability and contribute to lymphomagenesis. It is not clear whether Pim-1 is involved in hypoxia-mediated tumor survival in solid tumors. Here, we show that hypoxia can stabilize Pim-1 by preventing its ubiquitin-mediated proteasomal degradation and can cause Pim-1 translocation from the cytoplasm to the nucleus. Importantly, overexpression of Pim-1 increases NIH3T3 cell transformation exclusively under hypoxic conditions, suggesting that Pim-1 expression under hypoxia may be implicated in the transformation process of solid tumors. Also, blocking Pim-1 function by introduction of dominant negative Pim-1 resensitizes pancreatic cancer cells to apoptosis induced by glucose-deprivation under hypoxia. Introduction of short interfering RNAs for Pim-1 also resensitizes cancer cells to glucose deprivation under hypoxic conditions, while forced overexpression of Pim-1 causes solid tumor cells to become resistant to glucose deprivation. Moreover, dominant negative Pim-1 reduces tumorigenicity in pancreatic cancer cells and HeLa xenograft mouse models. Together, our studies indicate that Pim-1 plays a distinct role in solid tumor formation in vivo, implying that Pim-1 may be a novel target for cancer therapy.

Cancer cells undergo mitosis frequently, and many mitotic regulators are aberrantly expressed in these cells. Members of the Aurora family of serine/threonine kinases are expressed during mitosis and carry out vital functions in chromosome alignment, segregation and cytokinesis. Here we review the functions of Aurora-B kinases in mitosis and summarize the current literature on Aurora-B kinase inhibitors. In the process of developing these inhibitors as anticancer drugs, the Aurora kinase inhibitors have also helped to advance our understanding of the role of Aurora kinases in mitosis. The mechanism of action and structure-activity relationship of a selective Aurora-B inhibitor are also discussed. The future may see mechanism guided design of chemotherapy combinations that include these cell-cycle phase-specific drugs. The therapeutic potential of Aurora-B inhibitors is promising.

Despite diversity in genetic events in oncogenesis, cancer cells exhibit a common set of functional characteristics. Otto Warburg discovered that cancer cells have consistently higher rates of glycolysis than normal cells. The underlying mechanisms leading to the Warburg phenomenon include mitochondrial changes, upregulation of rate-limiting enzymes/proteins in glycolysis and intracellular pH regulation, hypoxia-induced switch to anaerobic metabolism, and metabolic reprogramming after loss of p53 function. The regulation of energy metabolism can be traced to a "triad" of transcription factors: c-MYC, HIF-1 and p53. Oncogenetic changes involve a nonrandom set of gene deletions, amplifications and mutations, and many oncogenes and tumor suppressor genes cluster along the signaling pathways that regulate c-MYC, HIF-1 and p53. Glycolysis in cancer cells has clinical implications in cancer diagnosis, treatment and interaction with diabetes mellitus. Many drugs targeting energy metabolism are in development. Future advances in technology may bring about transcriptome and metabolome-guided chemotherapy.

The 14-3-3sigma, upregulated by p53 in response to DNA damage, can have a positive-feedback impact driving p53 activities and is a human cancer epithelial marker downregulated in various tumors. However, the precise roles of 14-3-3sigma during tumorigenesis are not well characterized. Here, we show that 14-3-3sigma is a critical regulator of murine double minute oncogene (MDM2). 14-3-3sigma interacts with MDM2 at the RING domain. The C-terminal region of 14-3-3sigma binds to MDM2 very efficiently. Importantly, 14-3-3sigma overexpression leads to destabilization of MDM2 through enhancing MDM2 self-ubiquitination and accelerating turnover rate. Conversely, loss of 14-3-3sigma results in a significant increase in MDM2 protein. Moreover, live-cell images indicated that 14-3-3sigma can affect the location of MDM2 from the nucleus to the cytoplasm, and that MDM2-mediated cytoplasmic localization of p53 can be reversed by the presence of 14-3-3sigma. Significantly, we further showed that 14-3-3sigma causes MDM2 downregulation, thereby stabilizing p53 and inhibiting tumor growth in animal tumors. Also, 14-3-3sigma blocks MDM2-mediated retinoblastoma degradation and p53 NEDDylation. Our results provide evidence that 14-3-3sigma is a pivotal MDM2 regulator involved in blocking a variety of activities of MDM2.

The understanding of how hypoxia stabilizes and activates HIF1α in the nucleus with related oncogenic signals could revolutionize targeted therapy for cancers. Here, we find that histone H2AX displays oncogenic activity by serving as a crucial regulator of HIF1α signalling. H2AX interacts with HIF1α to prevent its degradation and nuclear export in order to allow successful VHL-independent HIF1α transcriptional activation. We show that mono-ubiquitylation and phosphorylation of H2AX, which are strictly mediated by hypoxia-induced E3 ligase activity of TRAF6 and ATM, critically regulate HIF1α-driven tumorigenesis. Importantly, TRAF6 and γH2AX are overexpressed in human breast cancer, correlate with activation of HIF1α signalling, and predict metastatic outcome. Thus, TRAF6 and H2AX overexpression and γH2AX-mediated HIF1α enrichment in the nucleus of cancer cells lead to overactivation of HIF1α-driven tumorigenesis, glycolysis and metastasis. Our findings suggest that TRAF6-mediated mono-ubiquitylation and subsequent phosphorylation of H2AX may serve as potential means for cancer diagnosis and therapy.

Adrenocortical carcinoma is a rare malignancy with poor prognosis and limited response to chemotherapy. Hepatocyte growth factor (HGF) and its receptor cMET augment cancer growth and resistance to chemotherapy, but their role in adrenocortical carcinoma has not been examined. In this study, we investigated the association between HGF/cMET expression and cancer hallmarks of adrenocortical carcinoma. Transcriptomic and immunohistochemical analyses indicated that increased HGF/cMET expression in human adrenocortical carcinoma samples was positively associated with cancer-related biologic processes, including proliferation and angiogenesis, and negatively correlated with apoptosis. Accordingly, treatment of adrenocortical carcinoma cells with exogenous HGF resulted in increased cell proliferation in vitro and in vivo while short hairpin RNA–mediated knockdown or pharmacologic inhibition of cMET suppressed cell proliferation and tumor growth. Moreover, exposure of cells to mitotane, cisplatin, or radiation rapidly induced pro-cMET expression and was associated with an enrichment of genes (e.g., CYP450 family) related to therapy resistance, further implicating cMET in the anticancer drug response. Together, these data suggest an important role for HGF/cMET signaling in adrenocortical carcinoma growth and resistance to commonly used treatments. Targeting cMET, alone or in combination with other drugs, could provide a breakthrough in the management of this aggressive cancer. Cancer Res; 75(19); 4131–42. ©2015 AACR.

SummaryBiomarkers for predicting prognosis are critical to treating colorectal cancer (CRC) patients. We found that CSN6, a subunit of COP9 signalosome, is overexpressed in CRC samples and that CSN6 overexpression is correlated with poor patient survival. Mechanistic studies revealed that CSN6 is deregulated by epidermal growth factor receptor (EGFR) signaling, in which ERK2 binds directly to CSN6 Leu163/Val165 and phosphorylates CSN6 at Ser148. Furthermore, CSN6 regulated β-Trcp and stabilized β-catenin expression by blocking the ubiquitin-proteasome pathway, thereby promoting CRC development. High CSN6 expression was positively correlated with ERK2 activation and β-catenin overexpression in CRC samples, and inhibiting CSN6 stability with cetuximab reduced colon cancer growth. Taken together, our study’s findings indicate that the deregulation of β-catenin by ERK2-activated CSN6 is important for CRC development.

Summary Extensive reprogramming of cellular energy metabolism is a hallmark of cancer. Despite its importance, the molecular mechanism controlling this tumour metabolic shift remains not fully understood. Here we show that 14-3-3σ regulates cancer metabolic reprogramming and protects cells from tumourigenic transformation. 14-3-3σ opposes tumour-promoting metabolic programs by enhancing c-Myc poly-ubiquitination and subsequent degradation. 14-3-3σ demonstrates the suppressive impact on cancer glycolysis, glutaminolysis, mitochondrial biogenesis and other major metabolic processes of tumours. Importantly, 14-3-3σ expression levels predict overall and recurrence-free survival rates, tumour glucose uptake and metabolic gene expression in breast cancer patients. Thus, these results highlight that 14-3-3σ is an important regulator of tumour metabolism, and loss of 14-3-3σ expression is critical for cancer metabolic reprogramming. We anticipate that pharmacologically elevating the function of 14-3-3σ in tumours could be a promising direction for targeted anti-cancer metabolism therapy development in future.

The COP9 signalosome subunit 6 (CSN6), which is involved in ubiquitin-mediated protein degradation, is overexpressed in many types of cancer. CSN6 is critical in causing p53 degradation and malignancy, but its target in cell cycle progression is not fully characterized. Constitutive photomorphogenic 1 (COP1) is an E3 ubiquitin ligase associating with COP9 signalosome to regulate important target proteins for cell growth. p27 is a critical G1 CDK inhibitor involved in cell cycle regulation, but its upstream regulators are not fully characterized. Here, we show that the CSN6-COP1 link is regulating p27(Kip1) stability, and that COP1 is a negative regulator of p27(Kip1). Ectopic expression of CSN6 can decrease the expression of p27(Kip1), while CSN6 knockdown leads to p27(Kip1) stabilization. Mechanistic studies show that CSN6 interacts with p27(Kip1) and facilitates ubiquitin-mediated degradation of p27(Kip1). CSN6-mediated p27 degradation depends on the nuclear export of p27(Kip1), which is regulated through COP1 nuclear exporting signal. COP1 overexpression leads to the cytoplasmic distribution of p27, thereby accelerating p27 degradation. Importantly, the negative impact of COP1 on p27 stability contributes to elevating expression of genes that are suppressed through p27 mediation. Kaplan-Meier analysis of tumor samples demonstrates that high COP1 expression was associated with poor overall survival. These data suggest that tumors with CSN6/COP1 deregulation may have growth advantage by regulating p27 degradation and subsequent impact on p27 targeted genes.

© 2015 Macmillan Publishers Limited. Tripartite motif 24 protein (TRIM24) is a plant homeodomain/bromodomain histone reader, recently associated with poor overall survival of breast-cancer patients. At a molecular level, TRIM24 is a negative regulator of p53 levels and a co-activator of estrogen receptor. However, the role of TRIM24 in breast tumorigenesis remains largely unknown. We used an isogenic human mammary epithelial cell (HMEC) culture model, derived from reduction mammoplasty tissue, and found that ectopic expression of TRIM24 in immortalized HMECs (TRIM24 iHMECs) greatly increased cellular proliferation and induced malignant transformation. Subcutaneous injection of TRIM24 iHMECs in nude mice led to growth of intermediate to high-grade tumors in 60-70% of mice. Molecular analysis of TRIM24 iHMECs revealed a glycolytic and tricarboxylic acid cycle gene signature, alongside increased glucose uptake and activated aerobic glycolysis. Collectively, these results identify a role for TRIM24 in breast tumorigenesis through reprogramming of glucose metabolism in HMECs, further supporting TRIM24 as a viable therapeutic target in breast cancer.

The 14-3-3 sigma (sigma) protein, a unique member of 14-3-3 family, is a negative regulator of the cell cycle and is induced by p53 to initiate cell cycle checkpoint control after DNA damage. Among the 14-3-3 family members, 14-3-3 sigma is uniquely induced by p53 and has a positive feedback effect on p53 activity in response to DNA damage. Although 14-3-3 sigma is linked to p53-regulated cell cycle checkpoint control, the detailed mechanisms of cell cycle regulation by 14-3-3 sigma remain unclear. Decreased expression of 14-3-3 sigma was reported in several types of carcinomas, suggesting that the negative regulatory role of 14-3-3 sigma in the cell cycle is compromised during tumorigenesis. Given the fact that p53's tumor suppressive function is lost in almost half of all human cancers and that 14-3-3 sigma's activity is linked to the p53 network, a perspective regarding the p53/14-3-3 sigma relationship is needed for cancer research. Here we discuss the mechanisms by which 14-3-3 sigma-stabilizes p53 with the hope that these insights may be applied to develop targeted therapeutic strategies for cancer treatment.

14-3-3 σ is induced by tumor suppressor protein p53 in response to DNA damage. p53 can directly transactivate the expression of 14-3-3 σ to cause a G2 cell cycle arrest when cell DNA is damaged. The expression of 14-3-3 σ protein is down-regulated in various tumors, but its function has not been fully established. Protein kinase B/Akt, a crucial regulator of oncogenic signal involved in cell survival and proliferation, is deregulated in many types of cancer. Akt activation can enhance p53 degradation, but its role in DNA damage response is not clear. Here, we show that Akt activation is diminished when p53 and 14-3-3 σ is up-regulated in response to DNA damage. Evidence is provided that 14-3-3 σ binds and inhibits Akt. In keeping with this concept, Akt-mediated cell survival is inhibited by 14-3-3 σ. Significantly, we show that 14-3-3 σ inhibits Akt-mediated cell growth, transformation, and tumorigenesis. Low expression of 14-3-3 σ in human primary breast cancers correlates with Akt activation. These data provide an insight into Akt regulation and rational cancer gene therapy by identifying 14-3-3 σ as a molecular regulator of Akt and as a potential anticancer agent for Akt-activated cancers. (Cancer Res 2006; (66)6: 3096-105)

The 200-amino-acid repeat (HIN-200) gene family with the hematopoietic interferon (IFN)-inducible nuclear protein encodes highly homologous proteins involved in cell growth, differentiation, autoimmunity, and tumor suppression. IFIX is the newest member of the human HIN-200 family and is often downregulated in breast tumors and breast cancer cell lines. The expression of the longest isoform of IFIX gene products, IFIXalpha1, is associated with growth inhibition, suppression of transformation, and tumorigenesis. However, the mechanism underlying the tumor suppression activity of IFIXalpha1 is not well understood. Here, we show that IFIXalpha1 downregulates HDM2, a principal negative regulator of p53, at the posttranslational level. IFIXalpha1 destabilizes HDM2 protein and promotes its ubiquitination. The E3 ligase activity of HDM2 appears to be required for this IFIXalpha1 effect. Importantly, HDM2 downregulation is required for the IFIXalpha1-mediated increase of p53 protein levels, transcriptional activity, and nuclear localization, suggesting that IFIXalpha1 positively regulates p53 by acting as a negative regulator of HDM2. We found that IFIXalpha1 interacts with HDM2. Interestingly, the signature motif of the HIN-200 gene family, i.e., the 200-amino-acid HIN domain of IFIXalpha1, is sufficient not only for binding HDM2 but also for downregulating it, leading to p53 activation. Finally, we show that IFIX mediates HDM2 downregulation in an IFN-inducible system. Together, these results suggest that IFIXalpha1 functions as a tumor suppressor by repressing HDM2 function.

The 14-3-3 sigma (sigma) protein, a negative regulator of the cell cycle, is a human mammary epithelium-specific marker that is downregulated in transformed mammary carcinoma cells. It has also been identified as a p53-inducible gene product involved in cell cycle checkpoint control after DNA damage. Although 14-3-3 sigma is linked to p53-regulated cell cycle checkpoint control, detailed mechanisms of how cell cycle regulation occurs remain unclear. Decreased expression of 14-3-3 sigma was recently reported in several types of carcinomas, further suggesting that the negative regulatory role of 14-3-3 sigma in the cell cycle is compromised during tumorigenesis. However, this possible tumor-suppressive role of 14-3-3 sigma has not yet been characterized. Here, we studied the link between 14-3-3 sigma activities and p53 regulation. We found that 14-3-3 sigma interacted with p53 in response to the DNA-damaging agent adriamycin. Importantly, 14-3-3 sigma expression led to stabilized expression of p53. In studying the molecular mechanism of this increased stabilization of p53, we found that 14-3-3 sigma antagonized the biological functions of Mdm2 by blocking Mdm2-mediated p53 ubiquitination and nuclear export. In addition, we found that 14-3-3 sigma facilitated the oligomerization of p53 and enhanced p53's transcriptional activity. As a target gene of p53, 14-3-3 sigma appears to have a positive feedback effect on p53 activity. Significantly, we also showed that overexpression of 14-3-3 sigma inhibited oncogene-activated tumorigenicity in a tetracycline-regulated 14-3-3 sigma system. These results defined an important p53 regulatory loop and suggested that 14-3-3 sigma expression can be considered for therapeutic intervention in cancers.

Cell cycle dysregulation is a critical feature of tumour cells. Numerous cell cycle regulators act either as oncogenes or tumour suppressors and their aberrations result in proliferative advantage for cancer cells. Many molecular targets and their use in either abrogating the growth advantage of oncogenic mediators or enhancing the growth suppressive activity of tumour suppressors, have been filed for patents. The molecular targets associated with cell cycle inhibition are of particular interest because they are potential therapeutic agents of promise in the control of inappropriate cellular proliferation. This review focuses on the recent discovery of potential molecular targets involved in cell cycle inhibition and their evaluation as therapeutic agents for cancers. In this review, the strategies employed to control oncogenesis and their possible clinical applications are discussed.

The mammalian cell cycle can be divided into four phases: G1 (gap phase 1), S (DNA synthesis), G2 (gap phase 2), and M (mitosis). Progression through each phase of the cell cycle is delicately controled by the activity of different cyclin-dependent kinases (CDKs) and their regulatory subunits known as cyclins. CDK2, CDK4, CDK6 and their associated cyclins control the G1 to S phase transition. The association of CDK4 or CDK6 with D-type cyclins is critical for G1 phase progression, whereas the CDK2-cyclin E complex is important for initiation of the S phase. Cancer can originate from dysregulation of these regulators. A variety of intrinsic and extrinsic signals were recently identified to regulate these G1 or G1/S CDKs and cyclins. Here we discuss the regulators of these protein kinases at different mechanistic level with a hope that these insights can be applied to develop therapeutic strategies for cancer treatment.

Hyperpolarized [1-13C]-pyruvate has shown tremendous promise as an agent for imaging tumor metabolism with unprecedented sensitivity and specificity. Imaging hyperpolarized substrates by magnetic resonance is unlike traditional MRI because signals are highly transient and their spatial distribution varies continuously over their observable lifetime. Therefore, new imaging approaches are needed to ensure optimal measurement under these circumstances. Constrained reconstruction algorithms can integrate prior information, including biophysical models of the substrate/target interaction, to reduce the amount of data that is required for image analysis and reconstruction. In this study, we show that metabolic MRI with hyperpolarized pyruvate is biased by tumor perfusion and present a new pharmacokinetic model for hyperpolarized substrates that accounts for these effects. The suitability of this model is confirmed by statistical comparison with alternates using data from 55 dynamic spectroscopic measurements in normal animals and murine models of anaplastic thyroid cancer, glioblastoma, and triple-negative breast cancer. The kinetic model was then integrated into a constrained reconstruction algorithm and feasibility was tested using significantly undersampled imaging data from tumor-bearing animals. Compared with naïve image reconstruction, this approach requires far fewer signal-depleting excitations and focuses analysis and reconstruction on new information that is uniquely available from hyperpolarized pyruvate and its metabolites, thus improving the reproducibility and accuracy of metabolic imaging measurements. Cancer Res; 75(22); 4708–17. ©2015 AACR.

Metformin is derived from galegine, a natural ingredient, and recent studies have suggested that metformin could enhance the antitumor effects of hormone ablative therapy or chemotherapy and reduce prostate cancer-specific mortality. Zyflamend is a combination of herbal extracts that reduces inflammation and comprises turmeric, holy basil, green tea, oregano, ginger, rosemary, Chinese goldthread, hu zhang, barberry, and basil skullcap. We propose a maintenance regimen with metformin and/or Zyflamend that targets cancer stem cells and the tumor microenvironment to keep the cancer dormant and prevent it from activation from dormancy. Herein, we report the clinical course of four patients who experienced a clinical response after treatment with metformin and/or Zyflamend.

The constitutive photomorphogenesis 9 signalosome (CSN) regulates the stability of tumor suppressor and oncogenic proteins via proteasome-mediated protein degradation. The detailed regulatory mechanisms of CSN subunit expression in tumorigenesis remain to be illustrated. We demonstrated several important biological functions of CSN6: p53 signal transduction,neddylation regulation of Cullin, F-box protein ubiquitination, and Myc stability regulation. Biochemical studies demonstrated that CSN6 is involved in cell cycle regulator stability function and various cancer signaling pathways, including epidermal growth factor receptor (EGFR) pathway. We show that CSN6 overexpression is frequently observed in human colorectal cancers, which leads to drug resistance in anti-EGFR treatment and is correlated with poor clinical survival. We present amechanism for the role of CSN6 in ERK signaling cascade and provide new insights into the functional role of CSN6 deregulation during tumorigenesis.

On behalf of the entire the international conference Innovations in Cancer Research and Regenerative Medicine 2017 (CRRM2017) Organizing Committee, we would like to extend to you a warm welcome to Ho Chi Minh city, Vietnam to join the 3rd international conference of CRRM. We are very glad to announce that this time, CRRM has made some exciting changes with more plenary sessions, more sessions, more oral invited speakers and especially a day of pre-conference. With two main topics of cancer research and regenerative medicine, this conference has many sessions organized into some parallel sessions with different topics of cancer research and regenerative medicine. By doing so, the audience can arrange their time for networking and speaking to our commercial partners. From the basic science to clinical trials, all sessions at the conference will cover from gene to human body. Therefore, we believe that every session will provide exciting presentations for attendees at all levels from students to scientists/researchers. With this conference, all authors not only share their latest results, but also can publish their works in the reputed journals. That is a reason why the keywords for this conference is Share & Publish. Like many developing countries, Vietnam has no shortage of health challenges, from infectious diseases such as annual flu epidemics to rising rates of unmet and chronic illnesses such as cancers and diabetes. Vietnam also has to contend with increasing drug resistance for killer diseases including AIDS, tuberculosis, and malaria. There is, however, increasing evidence that Vietnamese researchers are tackling the challenges of harnessing biotechnology to improve health care for healthier society as well as advancing the sciences. The biomedicine progresses in Cancer Research, and Regenerative Medicine in Vietnam include gene diagnostics for genetic disorders of human diseases, infectious pathogens; in-vitro production of therapeutic proteins including insulin, IFNs or biosimilar forms of monoclonal antibodies for targeted therapies; biomaterials; cell therapy; stem cell therapy and tissue engineering. In this conference, with 14 sessions of 100 oral presentations and 100 poster presentations, all latest topics of cancer research and regenerative medicine will be covered. New biomarkers of cancers, immunotherapy strategy for cancers and stem cells as well as tissue engineering are hot topics of CRRM2017. In addition to the conference program, there is an extensive set of pre-conference and lunch-on innovation showcases for on 10th and 11th Sept, respectively. We are very grateful for the support from sponsors for the CRRM2017. More vents will be taking place in the exhibition area, and we encourage attendees to take advantage of the opportunities to visit the exhibit booths and corporate symposia. We wish to thank everyone who helped so enthusiastically in the organization of the conference. Our thanks also go out to all of the speakers who have generously agreed to share their research results and experiences at this conference. We hope that you will enjoy this meeting and have a wonderful time here in Ho Chi Minh city, Vietnam.

Summary Cullin-RING ubiquitin ligases (CRL) are critical in ubiquitinating Myc, while COP9 signalosome (CSN) controls neddylation of Cullin in CRL. The mechanistic link between Cullin neddylation and Myc ubiquitination/degradation is unclear. Here we show that Myc is a target of the CSN subunit 6 (CSN6)–Cullin signaling axis and that CSN6 is a positive regulator of Myc. CSN6 enhanced neddylation of Cullin-1 and facilitated auto-ubiquitination/degradation of Fbxw7, a component of CRL involved in Myc ubiquitination, thereby stabilizing Myc. Csn6 haplo-insufficiency decreased Cullin-1 neddylation but increased Fbxw7 stability to compromise Myc stability and activity in an Eµ-Myc mouse model, resulting in decelerated lymphomagenesis. We found that CSN6 overexpression, which leads to aberrant expression of Myc target genes, is frequent in human cancers. Together, these results define a mechanism for the regulation of Myc stability through the CSN-Cullin-Fbxw7 axis and provide insights into the correlation of CSN6 overexpression with Myc stabilization/activation during tumorigenesis.

Background Obesity increases the risk of cancer death among postmenopausal women with estrogen receptor–positive (ER+) breast cancer, but the direct evidence for the mechanisms is lacking. The purpose of this study is to demonstrate direct evidence for the mechanisms mediating this epidemiologic phenomenon. Methods We analyzed transcriptomic profiles of pretreatment biopsies from a prospective cohort of 137 ER+ breast cancer patients. We generated transgenic (MMTV-TGFα;A y /a) and orthotopic/syngeneic (A y /a) obese mouse models to investigate the effect of obesity on tumorigenesis and tumor progression and to determine biological mechanisms using whole-genome transcriptome microarrays and protein analyses. We used a coculture system to examine the impact of adipocytes/adipokines on breast cancer cell proliferation. All statistical tests were two-sided. Results Functional transcriptomic analysis of patients revealed the association of obesity with 59 biological functional changes (P < .05) linked to cancer hallmarks. Gene enrichment analysis revealed enrichment of AKT-target genes (P = .04) and epithelial–mesenchymal transition genes (P = .03) in patients. Our obese mouse models demonstrated activation of the AKT/mTOR pathway in obesity-accelerated mammary tumor growth (3.7- to 7.0-fold; P < .001; n = 6–7 mice per group). Metformin or everolimus can suppress obesity-induced secretion of adipokines and breast tumor formation and growth (0.5-fold, P = .04; 0.3-fold, P < .001, respectively; n = 6–8 mice per group). The coculture model revealed that adipocyte-secreted adipokines (eg, TIMP-1) regulate adipocyte-induced breast cancer cell proliferation and invasion. Metformin suppress adipocyte-induced cell proliferation and adipocyte-secreted adipokines in vitro. Conclusions Adipokine secretion and AKT/mTOR activation play important roles in obesity-accelerated breast cancer aggressiveness in addition to hyperinsulinemia, estrogen signaling, and inflammation. Metformin and everolimus have potential for therapeutic interventions of ER+ breast cancer patients with obesity.

In the past decade, the discovery and characterization of cyclin-dependent kinases (CDKs), the engine cores of the cell cycle machinery, have advanced our understanding of the cell cycle. Both positive and negative regulators of CDKs have been characterized, accelerating the important research to unravel the mechanisms of the cell cycle disease - cancer. Cancer can originate from overexpression of positive regulators, such as cyclins, or from underexpression of negative regulators, such as CDK inhibitors (CKIs). CKIs are the focus of much cancer research because they are capable of controlling cell cycle proliferation - the Holy Grail for cancer treatment. CDKs can be inactivated by several mechanisms:, (i) by association with CKIs including p16 (INK4a), p15 (INK4b), p21 (Cip1), p27 (Kip1), and p57 (Kip2), (ii) by disassociation from their cyclin regulatory unit, (iii) by dephosphorylation of a conserved threonine residue in the T-loop, and (iv) by adding inhibitory phosphate. Here we discuss what is known about each mechanism with a hope that these insights will become useful in developing strategies to eliminate cancer in the future.

Amplification or overexpression of HER-2/neu in cancer cells confers resistance to apoptosis and promotes cell growth. The cellular localization of p21Cip1/WAF1 has been proposed to be critical either in promoting cell survival or in inhibiting cell growth. Here we show that HER-2/neu-mediated cell growth requires the activation of Akt, which associates with p21Cip1/WAF1 and phosphorylates it at threonine 145, resulting in cytoplasmic localization of p21Cip1/WAF1. Furthermore, blocking the Akt pathway with a dominant-negative Akt mutant restores the nuclear localization and cell-growth-inhibiting activity of p21Cip1/WAF1. Our results indicate that HER-2/neu induces cytoplasmic localization of p21Cip1/WAF1 through activation of Akt to promote cell growth, which may have implications for the oncogenic activity of HER-2/neu and Akt.

BACKGROUND Women with unilateral breast carcinoma are at increased risk for developing contralateral disease. The clinical significance of bilateral breast carcinoma has not been fully defined, and the subset of patients who may benefit from medical or surgical risk-reduction intervention has not yet been characterized. The purpose of this study was to evaluate risk factors and outcomes for bilateral breast carcinoma. METHODS A subject group of 70 bilateral breast carcinoma patients (62% metachronous) was matched by age and survival interval with a control group of 70 unilateral breast carcinoma patients. Median follow-up was 103 months. RESULTS Eighty-two percent of the unilateral patients and 80% of the bilateral patients had Stage I or II disease at diagnosis. Median age at presentation was 53 years. In the bilateral group, the contralateral cancer was diagnosed at the same or earlier stage than the first cancer in 87% of cases. Bilateral patients were significantly more likely to have multicentric disease and to have a positive family history for breast carcinoma compared with the unilateral group. There were no significant differences regarding history of exogenous hormone exposure, lobular histology, hormone-receptor status, or HER-2/neu expression. Five-year disease-free survival was 94% for the unilateral breast carcinoma patients and 91% for the bilateral breast carcinoma patients (P = 0.16). CONCLUSIONS Survival for patients with bilateral breast carcinoma is similar to that of patients with unilateral disease; however, prophylactic risk-reduction intervention for the contralateral breast should be considered in patients who have multicentric unilateral disease or a positive family history for breast carcinoma. Cancer 2001;91:1845–53. © 2001 American Cancer Society.

Strong expression of human epidermal growth factor receptor 2 (HER-2)/neu in breast cancer has been associated with poor prognosis. Reduced expression of p27(Kip1), a cyclin-dependent kinase inhibitor, correlates with poor clinical outcome in breast cancer. In this study, we provide a correlation between these two important prognostic markers in patients with breast cancer. Breast tumor screening using immunohistochemistry indicated that downregulation of p27 correlated with HER-2/neu overexpression in studying 11 normal breast tissues and 51 primary breast carcinomas. We found HER-2/neu protein overexpression in 20 (41%) of 49 breast cancers and low p27 protein expression in 47 (92%) of 51 breast cancers. All 20 (100%) of the tumors that overexpressed HER-2/neu had low levels of p27 protein product; this correlation was statistically significant (P = 0.035). Decreasing p27 expression correlated with increasing HER-2/neu activity. Our results suggest that one function of the HER-2/neu product is to downregulate p27 expression in breast cancer. This study may be significant in selecting patients for HER-2/neu antibody therapy in the future. Mol. Carcinog. 30:169--175, 2001.

Commentary to: Effect of DNAzymes Targeting Akt1 on Cell Proliferation and Apoptosis in Nasopharyngeal CarcinomaLifang Yang, Lanbo Xiao, Xiaoqian Ma, Min Tang, Xinxiang Weng, Xue Chen, Lunquan Sun，Ya Cao

The association of COP9 signalosome (CSN) with Constitutive Photomorphogenic 1 (COP1) was characterized in plants for a role in photomorpho-genesis. However, since mammalian cells do not perform photomorphogenesis, roles of these two proteins remain enigmatic. Recently we began to unravel their roles in oncogenesis. The CSN is an evolutionarily conserved multiprotein complex. Mammalian CSN consists of eight subunits (CSN1-CSN8) and has diverse functions, including cell cycle control, signal transduction, and tumorigenesis. Interestingly, CSN6, which is involved in stabilizing MDM2 [1] and preserving Cullin neddylation [2], is overexpressed in many types of cancers, implying its oncogenic role in cancer. However, the mechanistic regulation and biological consequence of CSN6 overexpression in cancer remain to be examined. Mammalian COP1 is an evolutionarily conserved E3 ubiquitin ligase, which contains RING-finger, coiled-coil and WD40-repeat domains. Through association with the CSN, COP1 functions as a crucial mediator to block photomorphogenesis in the dark through regulating the ubiquitinated proteasomal degradation of light-induced transcription factor HY5. In mammalian cells, COP1 is overexpressed in a various types of cancers, and it causes ubiquitin-mediated degradation of tumor suppressor p53, suggesting its oncogenic role in cancer. But its other substrates through protein degradation may have roles in tumorigenesis. Recently, CSN6's interaction with COP1 is characterized in mammalian cells, and CSN6-COP1 axis is involved in 14-3-3σ degradation [3]. COP1 is an E3 ligase of 14-3-3σ. 14-3-3σ, a tumor suppressor involved in opposing cancer metabolic reprogramming [4] and Akt signaling, is known to be upregulated by p53 and has a positive feedback effect on p53 in response to DNA damage. CSN6 stabilizes COP1 through reducing COP1 self-ubiquitination and thus decelerates COP1's turnover rate. CSN6/COP1-mediated 14-3-3σ ubiquitination is compromised when COP1 is knocked down. Subsequently, CSN6-COP1 axis causes 14-3-3σ downregulation, thereby activating Akt and promoting cancer cell survival. Also, the CSN6-COP1 axis regulates the stability p27Kip1, a critical G1 CDK inhibitor involved in cell cycle brake. COP1 is an E3 ligase of p27 to enhance p27 ubiquitination and subsequent degradation, which acts independent of negative regulators (E3 ligases) of p27, such as SKP2, Jab1, Pirh2, or KPC1 [5, 6]. Ectopic expression of CSN6 decreases the expression of p27 while CSN6 knockdown leads to p27 stabilization. Mechanistic studies show that CSN6 interacts with p27 and facilitates ubiquitin-mediated degradation of p27. CSN6-mediated p27 degradation depends on the nuclear export of p27, which is regulated through COP1's nuclear exporting signal. COP1 overexpression leads to the cytoplasmic distribution of p27, thereby accelerating p27 degradation. Importantly, the negative impact of COP1 on p27 stability contributes to elevated expression of genes that are suppressed through p27 mediation. Also, COP1-mediated p27 translocation to the cytoplasm will promote cancer cell migration and invasiveness, phenomena imposed by p27 cytoplasmic distribution. Kaplan-Meier analysis of tumor samples demonstrates that high COP1 or CSN6 expression is correlated with poor overall survival. These data suggest that tumors with CSN6-COP1 axis activation may have growth advantage by regulating p27 degradation and by subsequently impacting on p27 targeted gene expression. Understanding the genome integrity and DNA damage response is critical to cancer treatment. CSN6-COP1 axis can regulate genome integrity [7]. CSN6 overexpression leads to mitotic defect and ROS production. Importantly, COP1 and Aurora A (a p27-mediated suppressed gene involved in mitosis) are involved in this process. p27 levels are elevated after DNA damage, with concurrent reduction of COP1 levels. Mechanistic studies demonstrate that during DNA damage response COP1's function as an E3 ligase of p27 is inactivated due to 14-3-3σ-enhanced COP1 degradation, thereby reducing the ubiquitin-mediated degradation of p27. This process of p27 accumulation in response to DNA damage is not p53-dependent, but it is ATM- and 14-3-3σ-dependent. 14-3-3σ collaborates with ATM activity to mediate downregulation of COP1 after DNA damage. Congruently, in 14-3-3σ-null cells, COP1 is not downregulated by DNA damage; therefore, p27 levels are not increased. Expression of 14-3-3σ facilitates the downregulation of COP1 and subsequent increase of p27 in response to DNA damage. Studies show that p27 levels accumulate in the nucleus when COP1 is excluded from the cytoplasm in the presence of the DNA-damaging agent doxorubicin, suggesting that DNA damage affects the subcellular distribution of COP1, which in turn affects p27 levels. Further, COP1 overexpression leads to downregulation of p27Kip1, thereby promoting the overexpression of Aurora A, which correlates with poor survival. These findings provide new insight into CSN6-COP1-p27-Aurora A axis in DNA damage repair and tumorigenesis. Taken together, data show that COP1 is critical in CSN6-mediated oncogenesis, conferring impacts on 14-3-3σ, p27, Aurora A, Akt, metabolic reprogramming, genome integrity, and others. Further studies should be explored to address the following questions. What are other downstream ubiquitination targets of CSN6-COP1 axis involved in cancer? What are the upstream oncogenic signals in regulating this axis to promote cancer growth? Strategy in hindering the CSN6-COP1 axis activation may be a useful therapeutic strategy for cancer intervention.

Breast cancer is a disease of cell cycle, and the dysfunction of cell cycle checkpoints plays a vital role in the occurrence and development of breast cancer. We employed multi-gene fluorescence in situ hybridization (M-FISH) to investigate gene copy number aberrations (CNAs) of 4 genes (Rb1, CHEK2, c-Myc, CCND1) that are involved in the regulation of cell cycle, in order to analyze the impact of gene aberrations on prognosis in the young breast cancer patients. Gene copy number aberrations of these 4 genes were more frequently observed in young breast cancer patients when compared with the older group. Further, these CNAs were more frequently seen in Luminal B type, Her2 overexpression, and tiple-negative breast cancer (TNBC) type in young breast cancer patients. The variations of CCND1, Rb1, and CHEK2 were significantly correlated with poor survival in the young breast cancer patient group, while the amplification of c-Myc was not obviously correlated with poor survival in young breast cancer patients. Thus, gene copy number aberrations (CNAs) of cell cycle-regulated genes can serve as an important tool for prognosis in young breast cancer patients.