DEK Overexpression Research Articles

Abstract S4-05: Expression of the DEK oncogene promotes M2 polarization and iron recycling in tumor associated macrophages

Abstract The chromatin-remodeling DEK oncogene is highly expressed in over 60% of breast cancers, regardless of sub-type. DEK over-expression correlates with poor clinical outcome, including decreased survival and chemotherapy resistance. Previously, using human breast cancer cell lines and the MMTV-Ron mouse breast cancer model, we have demonstrated that DEK expression promotes proliferation and metastasis and supports the breast cancer stem cell population. These functions were due, in part, through upregulated Wnt ligand expression and β-catenin activation, leading to paracrine and autocrine effects promoting tumorigenesis. To investigate additional drivers of DEK-induced tumorigenesis, we performed RNA-Sequencing on control and DEK over-expressing immortalized MCF10A cells. Gene ontology analysis identified several immune signaling processes. This is in alignment with the well-characterized role of DEK expression in the promotion of inflammatory diseases. We investigated the immune cell infiltration of tumors from MMTV-Ron/Dek+/+ and MMTV-Ron/Dek-/- mice. Interestingly, we discovered that Dek-expressing tumors demonstrated decreased tumor associated macrophage (TAM) infiltration, where TAMs were largely limited to the periphery. Furthermore, these TAMs demonstrated an iron-recycling phenotype (M2 polarization), whereas TAMs from Dek-knockout tumors retained iron (M1 polarization). M1 macrophages are typically associated with inflammatory responses whereas M2 macrophages have been linked to tissue remodeling, angiogenesis, and tumor promotion. To investigate this further, we compared Dek+/+ and Dek-/- bone marrow-derived macrophages (BMDM) and found no inherent difference in polarization. However, BMDM demonstrated different gene expression profiles when cultured in conditioned media from Dek-expressing murine breast cancer cell lines compared to isogenic Dek-/- cells. BMDM exposed to conditioned media from Dek-expressing cancer cells demonstrated the same iron recycling phenotype observed in vivo, likely caused by increased expression of ferroportin, as well as increased expression of M2 markers CXCR4, VEGF, and an elevated ARG1hi/NOSlo population. We thus used a combination of genetic models and small molecule inhibitors to identify the mechanism of M2 polarization of BMDM exposed to Dek-expressing cancer cells. NRF2 transcription activity was eliminated as a possible mechanism for ferroportin regulation. However, small molecule inhibitors and gene expression analyses implicated secreted Wnt ligands and S100A8/A9 alarmins from the cancer cells, and downstream C/EBP transcriptional activity in the macrophages, as possible mechanisms of Dek-induced M2 polarization. Combined, this suggests that the poor clinical outcome of high DEK expressing breast cancers may be due to not only cell intrinsic factors, but also cell extrinsic factors, such as the tumor-promoting M2 polarization of tumor associated macrophages. Citation Format: Privette Vinnedge LM, Pease NA, Cheek J. Expression of the DEK oncogene promotes M2 polarization and iron recycling in tumor associated macrophages [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr S4-05.

Critical role of DEK and its regulation in tumorigenesis and metastasis of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related mortality globally. Therefore, it is quite essential to identify novel HCC-related molecules for the discovery of new prognostic markers and therapeutic targets. As an oncogene, DEK plays an important role in cell processes and participates in a variety of cellular metabolic functions, and its altered expression is associated with several human malignancies. However, the functional significance of DEK and the involved complex biological events in HCC development and progression are poorly understood. Here, combing the results from clinical specimens and cultured cell lines, we uncover a critical oncogenic role of DEK, which is highly expressed in HCC cells. DEK protein encompasses two isoforms (isoforms 1 and 2) and isoform 1 is the most frequently expressed DEK isoform in HCC cells. DEK depletion by using shRNA inhibited the cell proliferation and migration in vitro and suppressed tumorigenesis and metastasis in mouse models. Consistently, DEK overexpression regardless of which isoform produced the opposite effects. Further studies showed that DEK induced cell proliferation through upregulating cell cycle related CDK signaling, and promoted cell migration and EMT, at least in part, through the repression of β-catenin/E-cadherin axis. Interestingly, isoform 1 induced cell proliferation more efficiently than isoform 2, however, no functional differences existed between these two isoforms in cell migration. Together, our study indicates that DEK expression is required for tumorigenesis and metastasis of HCC, providing molecular insights for DEK-related pathogenesis and a basis for developing new strategies against HCC.

Significance of DEK overexpression for the prognostic evaluation of non-small cell lung carcinoma.

In the present study, we explored the role of DEK expression for the prognostic evaluation of non-small cell lung carcinoma (NSCLC). DEK protein and mRNA expression levels were detected in NSCLC cells and fresh tissue samples of NSCLC paired with adjacent non-tumor tissues, respectively. NSCLC cases (n=196) meeting strict follow-up criteria were selected for immunohistochemical staining of DEK protein. Correlations between DEK expression and clinicopathological features of the NSCLC cases were evaluated using Chi-square tests. Survival rates were calculated using the Kaplan-Meier method, and the relationship between prognostic factors and patient overall survival was analyzed using Cox proportional hazard analysis. Based on the results, the levels of DEK protein and mRNA were significantly upregulated in 6 fresh tissue samples of NSCLC. Immunohistochemical analysis showed that the DEK expression rate was significantly higher in the NSCLC samples compared with either the adjacent non-tumor tissues or normal lung tissues. DEK expression was correlated with poor differentiation and late pathological stage of NSCLC. DEK expression was also correlated with low disease-free survival and overall survival rates. In the early-stage group, disease-free and overall survival rates of patients with DEK expression were significantly lower than those of patients without DEK expression. Further analysis using a Cox proportional hazard regression model revealed that DEK expression emerged as a significant independent hazard factor for the overall survival rate of patients with NSCLC. Consequently, DEK plays an important role in the progression of NSCLC. DEK may potentially be used as an independent biomarker for the prognostic evaluation of NSCLC.

Overexpression of DEK in human gastric carcinoma and its clinicopathological significance.

75 Background: Gastric cancer (GAC) is the second most common cancer worldwide and the fifth leading cause of cancer-related death in Taiwan. To improve the survival of gastric cancer patients, biomarkers for early detection and effective anticancer therapy are required. Methods: Dysregulated proteins in GAC tumor cells were captured and identified by laser capture microdissection (LCM) and an isobaric tags for relative and absolute quantitation (iTRAQ)-labeling following basic-RP HPLC coupled to LTQ-orbitrap MS analysis. DEK, one of the most highly-expressed proteins in both intestinal and diffuse gastric cancers in iTRAQ analysis, was selected for further study. DEK proto-oncogene is originally identified as a fusion with the CAN nucleoporin protein in a subset of acute myeloid leukemia (AML) patients carrying the t(6;9) translocation, and DEK has been implicated in acute myeloid leukemia, melanoma, glioblastoma, hepatocellular carcinoma, and bladder cancer. The overexpression of DEK was confirmed using real-time quantitative-reverse transcription-polymerase chain reaction (Q-RT–PCR), Western blot, and immunohistochemical (IHC) analysis. Results: The data revealed that the expression of DEK mRNA was up-regulated in tumor tissues (compared with adjacent non-tumor tissues) of about 55.1% gastric patients. The expression of DEK protein was also up-regulated in tumor tissues identified by Western blot and IHC. To further understand the role of DEKgene in the human gastric cancer cells, the DEK knockdown stable clones were prepared in AZ521. The cells migration ability was reduced in DEK depletion stable clones, and the antitumor drug sensitivity and anoikis were increased in DEK depletion cells. Conclusions: These results indicate that DEK overexpression may contribute the poor prognosis of patients.

DEK is a potential marker for aggressive phenotype and irinotecan-based therapy response in metastatic colorectal cancer.

BackgroundDEK is a transcription factor involved in stabilization of heterochromatin and cruciform structures. It plays an important role in development and progression of different types of cancer. This study aims to analyze the role of DEK in metastatic colorectal cancer.MethodsBaseline DEK expression was firstly quantified in 9 colorectal cell lines and normal mucosa by WB. SiRNA-mediated DEK inhibition was carried out for transient DEK silencing in DLD1 and SW620 to dissect its role in colorectal cancer aggressiveness. Irinotecan response assays were performed with SN38 over 24 hours and apoptosis was quantified by flow cytometry. Ex-vivo assay was carried out with 3 fresh tumour tissues taken from surgical resection and treated with SN38 for 24 hours. DEK expression was determined by immunohistochemistry in 67 formalin-fixed paraffin-embedded tumour samples from metastatic colorectal cancer patients treated with irinotecan-based therapy as first-line treatment.ResultsThe DEK oncogene is overexpressed in all colorectal cancer cell lines. Knock-down of DEK on DLD1 and SW620 cell lines decreased cell migration and increased irinotecan-induced apoptosis. In addition, low DEK expression level predicted irinotecan-based chemotherapy response in metastatic colorectal cancer patients with KRAS wild-type.ConclusionsThese data suggest DEK overexpression as a crucial event for the emergence of an aggressive phenotype in colorectal cancer and its potential role as biomarker for irinotecan response in those patients with KRAS wild-type status.

Mechanisms underlying cancer growth and apoptosis by DEK overexpression in colorectal cancer.

Our previous study indicated that DEK protein was overexpressed in colorectal carcinoma (CRC) compared with the normal colorectal mucosa. DEK was also significantly correlated with the prognostic characteristics of patients with CRC, demonstrating that DEK played an important role in CRC progression. In this work, we evaluate the effects of DEK on biological behaviors in CRC and explore the related molecular mechanisms. The results showed that DEK was overexpressed in human CRC tissues, and was correlated with the Ki-67 index and the apoptotic index. DEK depletion by RNAi in SW-620 and HCT116 cells significantly decreased cell proliferation, but increased cell apoptosis. Upregulation of DEK was involved in the p53/MDM, Bcl-2 family, and caspase pathways. Our study demonstrates that DEK promotes the growth of CRC, and could be a therapeutic target in CRC.

208P - Clinical Significance of Dek Overexpression in Oral Squamous Cell Carcinoma

ABSTRACT Aim: Human DEK gene on chromosome 6p encodes a 43kD nuclear phospoprotein that was originally identified as part of a fusion protein found in a subset of acute myeloid leukemia carrying a t(6;9) translocation. According to several in vitro studies, DEK overexpression may play a role in tumor development through suppressing cellular senescence, apoptosis, and differentiation, which result in promoting cell growth and survival. Although DEK upregulation has been described in a number of human malignancies and was significantly associated with high histologic grade, lymph node metastasis and/or advanced clinical stage, no previous report has evaluated the expression of DEK protein and its clinical significance in oral squamous cell carcinoma (OSCC). Our aims were to determine DEK expression in tissue samples of normal oral mucosa and OSCC by immunohistochemistry, to analyze the correlation between DEK expression and clinicopathologic parameters, and to evaluate the value of DEK as a predictive marker for clinical outcome in OSCC. Methods: Ten normal oral mucosa samples and 60 OSCC samples were studied by immunohistochemistry. The immunostaining for DEK was scored as 0 (no or 50% positive cells). Cases with a score ≥ 2 were defined as high expression. Correlation between DEK expression and clinicopathological parameters were analyzed by k2 test and Fisher's exact tests. The survival rates were calculated using the Kaplan-Meier method and analyzed using the log-rank test. P Results: High expression of DEK protein (score ≥ 2) was found in 68.3% of OSCC cases. Statistical analysis revealed that DEK overexpression in OSCC was positively correlated with high histologic grade (p = 0.001), lymph node metastasis (p = 0.003), and advanced clinical stage (p = 0.039). In the Kaplan-Meier survival analysis, DEK overexpression was significantly associated with decreased overall survival in patients with OSCC (p = 0.019). Conclusions: We demonstrated the clinical significance of DEK overexpression in OSCC tissue samples. DEK overexpression correlated with worse clinical outcome of patients with OSCC. Therefore, our results suggest that DEK overexpression may be a reliable marker to predict the clinical outcome in OSCC. Disclosure: All authors have declared no conflicts of interest.

The DEK oncogene promotes cellular proliferation through paracrine Wnt signaling in Ron receptor-positive breast cancers.

Disease progression and recurrence are major barriers to surviving breast cancer. Understanding the etiology of recurrent or metastatic breast cancer and underlying mechanisms is critical for the development of new treatments and improved survival. Here, we report that two commonly over-expressed breast cancer oncogenes, Ron and DEK, cooperate to promote advanced disease through multi-pronged effects on β-catenin signaling. The Ron receptor is commonly activated in breast cancers, and Ron over-expression in human disease stimulates β-catenin nuclear translocation and is an independent predictor of metastatic dissemination. Dek is a chromatin-associated oncogene whose expression has been linked to cancer through multiple mechanisms, including β-catenin activity. We demonstrate here that Dek is a downstream target of Ron receptor activation in murine and human models. The absence of Dek in the MMTV-Ron mouse model led to a significant delay in tumor development, characterized by decreased cell proliferation, diminished metastasis, and fewer cells expressing cancer stem cell markers. Dek complementation of cell lines established from this model was sufficient to promote cellular growth and invasion in vitro and in vivo. Mechanistically, Dek expression stimulated the production and secretion of Wnt ligands to sustain an autocrine/paracrine canonical β-catenin signaling loop. Finally, we show that Dek over-expression promotes tumorigenic phenotypes in immortalized human mammary epithelial MCF10A cells and, in the context of Ron receptor activation, correlates with disease recurrence and metastasis in patients. Overall, our studies demonstrate that DEK over-expression, due in part to Ron receptor activation, drives breast cancer progression through the induction of Wnt/β-catenin signaling.

High expression of DEK predicts poor prognosis of gastric adenocarcinoma

BackgroundDEK, as an oncoprotein, plays an important role in cancer development and progression. This study aimed to investigate the clinicopathological significance of DEK overexpression in patients with gastric cancer.Materials and methodsThe expression of DEK protein was evaluated by immunohistochemical (IHC) staining of 172 gastric cancer samples with complete clinicopathological features, and the correlation between DEK expression and clinicopathological features was examined. Survival rates were also calculated using the Kaplan-Meier method in gastric cancer patients with complete survival data.ResultsDEK protein showed a strictly nuclear staining pattern in gastric cancers with IHC and immunofluorescence. The strongly positive rate of DEK protein was 60.5% (104/172) in gastric cancers, which was significantly higher than that in either gastric dysplasia (19.4%, 7/36) or adjacent normal mucosa (0%, 0/27). DEK expression in gastric cancer correlated to tumor size, differentiation, clinical stage, disease-free survival, and overall survival rates. Further analysis showed that patients with early-stage gastric cancer and high DEK expression had shorter disease-free survival and overall survival duration than those with low DEK expression.ConclusionHigh level of DEK protein expression predicts the poor prognosis of patients with gastric cancer. DEK expression might be potentially used as an independent effective biomarker for prognostic evaluation of gastric cancers.Virtual slidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/5050145571193097

DEK promotes HPV-positive and -negative head and neck cancer cell proliferation.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common malignancy worldwide, and patient outcomes using current treatments remains poor. Tumor development is etiologically associated with tobacco or alcohol use and/or HPV infection. HPV positive HNSCCs, which frequently harbor wild-type p53, carry a more favorable prognosis and are a biologically distinct subgroup when compared to their HPV negative counterparts. HPV E7 induces expression of the human DEK gene, both in vitro and in vivo. In keratinocytes, DEK overexpression is sufficient for causing oncogenic phenotypes in the absence of E7. Conversely, DEK loss results in cell death in HPV positive cervical cancer cells at least in part through p53 activation, and Dek knockout mice are relatively resistant to the development of chemically induced skin papillomas. Despite the established oncogenic role of DEK in HPV associated cervical cancer cell lines and keratinocytes, a functional role of DEK has not yet been explored in HNSCC. Using an established transgenic mouse model of HPV16 E7 induced HNSCC, we demonstrate that Dek is required for optimal proliferation of E7-transgenic epidermal cells and for the growth of HNSCC tumors. Importantly, these studies also demonstrate that DEK protein is universally up-regulated in both HPV positive and negative human HNSCC tumors relative to adjacent normal tissue. Furthermore, DEK knockdown inhibited the proliferation of HPV positive and negative HNSCC cells, establishing a functional role for DEK in human disease. Mechanistic studies reveal that attenuated HNSCC cell growth in response to DEK loss was associated with reduced expression of the oncogenic p53 family member, ΔNp63. Exogenous ΔNp63 expression rescued the proliferative defect in the absence of DEK, thereby establishing a functional DEK-ΔNp63 oncogenic pathway that promotes HNSCC. Taken together, our data demonstrate that DEK stimulates HNSCC cellular growth and identify ΔNp63 as a novel DEK effector.

DEK and WT1 Affect Alternative Splicing of Genes Involved in Hematopoietic Cell Lineage and Resistance to Chemotherapy in Acute Myeloid Leukemia Cells.

AbstractAbstract 2392In humans, the majority of all protein-coding transcripts contain introns that are removed by mRNA splicing carried out by spliceosomes. Mutations in the spliceosome machinery have recently been identified using whole-exome/genome technologies in myelodysplastic syndromes (MDS) and in acute myeloid leukemia (AML). In MDS the frequency of somatic spliceosomal mutations (SSM) range from 1–3% for U2AF1 in RARS/RCMD-RS to more than 70% for SF3B1 in ARSI. These values are significantly lower in AML whereas AML cells cumulate numerous splicing defects. Beside SSMs, one can propose that alternative splicing (AS) might be disturbed by other processes such as abnormal protein-protein interactions. DEK and WT1 are 2 oncogenes overexpressed in most patients with AML. They physiologically influence AS through physical interactions with the heterodimer U2AF1/U2AF2 involved in the recognition of splice acceptor site by the splicing machinery. It is therefore possible that the leukemogenic overexpression of DEK or WT1 might deregulate AS in AML cells, even in the absence of SSM. Here we show that DEK and WT1 affect AS in AML cells. Exon expression profiling was performed in triplicate with MOLM13, KASUMI and KG1 AML cells stably knocked down or not for DEK and WT1 through shRNA. The efficiency of shRNA-mediated silencing was confirmed by western blot and total RNA was analyzed using the Exon microarray platform GeneChip Human Exon 1.0 ST (Affymetrix). Microarray data were cross-compared between cell lines and only statistically significant modifications (p<0.05) shared by MOLM13, KASUMI and KG1 cells were selected. DEK and WT1 knock-down induced the transcriptional deregulation of 1613 (813 up) and 3280 (998 up) genes in AML cell lines, respectively. AS events were selected and annotated with fasterDB database (http://fasterdb.com/faster/home.pl) for genes displaying either no or low (<2) differential transcription. With this approach, differential expression of DEK coincided with changes in 1049 AS events over 934 genes. Those were distributed in 4 alternative acceptor sites (ACC), 222 first exons (AFE), 257 last exons (ALE), 539 spliced exon (ASE), 6 deletions (DEL), 21 donor sites (DON). Differential expression of WT1 led to modifying 1371 alternative splicing entities over 1198 genes. Those were distributed in 6 ACC, 385 AFE, 343 ALE, 590 ASE, 13 DEL, and 34 DON. Genes with AS events were then sorted based on gene function with DAVID bioinformatics resources version 6.7 (http://david.abcc.ncifcrf.gov/). The results indicated that, in both DEK- and WT1-dependent assays, a large subset of genes were related to hematopoietic cell lineage followed by other functional categories such as calcium signaling, ATP-binding cassette (ABC) transporters, and focal adhesion pathways that have been previously reported to be affected in AML cells and involved in resistance to chemotherapy. Differential expression of WT1 modified AS of CD3E, CD9, CSF1R, CSF3R, CR2, GP1BA, ITGA1, ITGA3, ITGB3, IL1A and IL6 while that of DEK led to modulate AS of CD19, CD1d, CD36, CD3G, CSF1R, CR2, ITGA1, ITGA2B, ITGA4, ITGA6, IL7, HLA-DRB5, and MME. Microarray data were validated by exon specific RT-PCR. Exon expression profiling of fresh AML bone marrow samples with or without U2AFs mutations and various levels of DEK or WT1 expression is currently in progress and will be presented. In conclusion in AML cells, DEK and WT1 overexpression affects AS of numerous key genes involved in hematologic differentiation, leukemogenesis and resistance to chemotherapy. These posttranscriptional effects occur in the absence of transcriptional change and therefore highlight hitherto unknown phenotypic alterations having putative diagnostic, prognostic and therapeutic interests. DEK and WT1 are overexpressed in more than 80% of AML and target AS via U2AFs that are mutated in only 0–8% of cases. Accordingly present results strongly suggest that other factors than SSM divert AS in AML. Disclosures:No relevant conflicts of interest to declare.

DEK overexpression is correlated with the clinical features of breast cancer

To investigate the clinicopathological significance of DEK overexpression in breast cancers, a total of 196 cases, including 20 of normal tissues, 12 of intraductal hyperplasia, 31 of ductal carcinoma in situ (DCIS) and 133 of invasive ductal carcinoma of the breast, were selected from the Department of Pathology, Yanbian Tumor Hospital for immunohistochemical staining of DEK, estrogen (ER), progesterone (PR) and Ki-67 proteins. In results, DEK protein had higher positivity in DCIS, compared with the adjacent normal breast tissues. Also, DEK protein was strongly positive in invasive ductal carcinoma of the breast on immunohistochemistry, which was significantly higher than normal breast tissues. However, only two (2/12) cases of intraductal hyperplasia of the breast showed positive staining for DEK protein. Additionally, DEK overexpression was significantly correlated with the increased proliferating index of Ki-67. For the histological grade, DEK positive rate was only 39.6% in G1 breast cancers, but significantly higher in G2 (92.3%) and G3 (97.0%) cases (P<0.05). Also, a strongly positive rate of DEK was lower in Stage-0 (21.4%) and Stage-I (40.9%) compared with Stage-IIa (87.5%), Stage-IIb (89.7%) and Stage-IIIa (92.3%) (P<0.05). And DEK protein showed higher expression level in < 3 years disease free survival breast cancers than it did in ≥ 3 years disease free survival cases (P<0.05). However, no statistically difference was found among DEK expression, lymph node metastasis, and ER and PR expressions. In conclusion, DEK overexpression appears to be associated with breast cancer progression and DEK may potentially be used as a breast cancer biomarker for the early diagnosis, prognostic evaluation and therapeutic target for breast cancer.

The human DEK oncogene stimulates β-catenin signaling, invasion and mammosphere formation in breast cancer

Breast cancer is a major cause of cancer-related deaths in American women; therefore, the identification of novel breast-cancer related molecules for the discovery of new markers and drug targets remains essential. The human DEK gene, which encodes a chromatin-binding protein and DNA topology regulator, is up-regulated in many types of cancer. DEK has been implicated as an oncogene in breast cancer based on mRNA expression studies, but its functional significance in breast cancer growth and progression has not yet been tested directly. We demonstrate that DEK is highly expressed in breast cancer cells compared to normal tissue, and functionally important for cellular growth, invasion and mammosphere formation. DEK over-expression in non-tumorigenic MCF10A cells resulted in increased growth and motility with a concomitant down-regulation of E-cadherin. Conversely, DEK knockdown in MCF7 and MDA-MB-468 breast cancer cells resulted in decreased growth and motility with up-regulation of E-cadherin. The use of DEK-proficient and -deficient breast cancer cells in orthotopic xenografts provided further in vivo evidence that DEK contributes to tumor growth. Activation of the β-catenin signaling pathway is important for normal and cancer stem cell character, growth and metastasis. We show that DEK expression stimulated and DEK knockdown repressed β-catenin nuclear translocation and activity. Importantly, the expression of constitutively active β-catenin rescued breast cancer invasion defects of DEK knockdown cells. Together, our data indicate that DEK expression stimulates the growth, stem cell character, and motility of breast cancer cells, and that DEK-dependent cellular invasion occurs at least in part via β-catenin activation.

DEK expression in melanocytic lesions

The diagnosis of malignant melanoma presents a clinical challenge and relies principally on histopathological evaluation. Previous studies have indicated that increased expression of the DEK oncogene, a chromatin-bound factor, could contribute to the development of melanoma and may be a frequent event in melanoma progression. Here, we investigated DEK expression by immunohistochemistry in a total of 147 melanocytic lesions, including ordinary nevi, dysplastic nevi, Spitz nevi, melanoma in situ, primary invasive melanomas, and metastatic melanomas. Most benign nevi (ordinary, dysplastic, and Spitz nevi) were negative or exhibited weak staining for DEK, with only 4 of 49 cases showing strong staining. Similar to benign nevi, melanoma in situ also demonstrated low levels of DEK expression. In contrast, the expression of DEK in primary invasive melanomas was significantly higher than benign nevi (P < .0001). Moreover, DEK expression was significantly increased in deep melanomas (Breslow depth >1 mm) and metastatic melanomas as compared with superficial melanomas (Breslow depth ≤1 mm) (P < .05). Our findings indicate that DEK overexpression may be a frequent event in invasive melanomas, and further augmentation of DEK expression may be associated with the acquisition of ominous features such as deep dermal invasion and metastasis. These data suggest a role of DEK in melanoma progression.

DEK oncoprotein regulates transcriptional modifiers and sustains tumor initiation activity in high-grade neuroendocrine carcinoma of the lung

Lung cancer shows diverse histological subtypes. Large-cell neuroendocrine cell carcinoma and small-cell lung carcinoma show similar histological features and clinical behaviors, and can be classified as high-grade neuroendocrine carcinoma (HGNEC) of the lung. Here we elucidated the molecular classification of pulmonary endocrine tumors by copy-number profiling. We compared alterations of copy number with the clinical outcome of HGNEC and identified a chromosomal gain of the DEK oncogene locus (6p22.3) that was significantly associated with poor prognosis. We further confirmed that DEK overexpression was associated with poor prognosis in a larger set of HGNEC. Downregulation of DEK by small hairpin RNA led to a marked reduction of in vitro colony formation, in vivo tumorigenicity and chemo-resistance, and was associated with loss of lung cancer stem cell markers. Gene expression profiling revealed that DEK downregulation was associated with altered expression of transcriptional regulators, which specifically include known targets of interchromosomal translocations in hematopoietic tumors, and knockdown of these epigenetic modifiers affected colony formation activity. Our study showed that DEK overexpression, partly through an increase in its gene dose, mediates the activity of global transcriptional regulators and is associated with tumor initiation activity and poor prognosis in HGNEC.

Clinicopathological significance of DEK overexpression in serous ovarian tumors

To investigate the significance of DEK protein expression in ovarian lesions, a total of 113 ovarian serous tumors, including 62 serous cystadenocarcinomas and 19 serous borderline tumors, were studied on immunohistochemistry. For comparison, 32 benign serous tumors, including 12 serous papillary cystadenomas, 10 serous cystadenomas, and 10 serous surface papillomas, were also included. DEK was positive in 93.5% of serous cystadenocarcinomas (58/62), 63.2% of serous borderline tumors (12/19), and weakly positive in 15.6% of benign serous tumors (5/32). The strong positive signal was detected only in serous adenocarcinomas (80.6%, 50/62) and borderline tumors (21.1%, 4/19), but no serous benign tumors were strongly positive (0%, 0/32). Meanwhile, the strong positivity rate of DEK protein was significantly higher in grade 2 and grade 3 than in grade 1 ovarian cancers (P < 0.05), but there was no significant association between DEK protein expression level and International Federation of Gynecology and Obstetrics (FIGO) stage of serous ovarian adenocarcinoma (P > 0.05). In summary, DEK plays an important role in the progression of ovarian serous cancers. The detection of DEK protein expression should be useful for the diagnosis and prognosis of ovarian serous cancers, and DEK might be a useful molecular target for ovarian cancer therapy.

DEK Proto-Oncogene Expression Interferes with the Normal Epithelial Differentiation Program

Overexpression of the DEK gene is associated with multiple human cancers, but its specific roles as a putative oncogene are not well defined. DEK transcription was previously shown to be induced by the high-risk human papillomavirus (HPV) E7 oncogene via E2F and Rb pathways. Transient DEK overexpression was able to inhibit both senescence and apoptosis in cultured cells. In at least the latter case, this mechanism involved the destabilization of p53 and the decreased expression of p53 target genes. We show here that DEK overexpression disrupts the normal differentiation program in a manner that is independent of either p53 or cell death. DEK expression was distinctly repressed upon the differentiation of cultured primary human keratinocytes, and stable DEK overexpression caused epidermal thickening in an organotypic raft model system. The observed hyperplasia involved a delay in keratinocyte differentiation toward a more undifferentiated state, and expansion of the basal cell compartment was due to increased proliferation, but not apoptosis. These phenotypes were accompanied by elevated p63 expression in the absence of p53 destabilization. In further support of bona fide oncogenic DEK activities, we report here up-regulated DEK protein levels in both human papilloma virus-positive hyperplastic murine skin and a subset of human squamous cell carcinomas. We suggest that DEK up-regulation may contribute to carcinoma development at least in part through increased proliferation and retardation of differentiation.

DEK overexpression in uterine cervical cancers

The purpose of the present paper was to investigate the significance of DEK protein expression in uterine cervical lesions and its relationship with HPV infection status. DEK protein expression was studied in 253 cervical lesions, including 30 non-neoplastic cervix with or without squamous metaplasia, 64 cervical intra-epithelial neoplasias (CIN; CIN-1, n = 28; CIN-2, n = 17; CIN-3, n = 19), 102 squamous cell carcinomas (SCC), 51 adenocarcinomas, and six adenosquamous cell carcinomas (adenoSCC) on immunohistochemistry. For comparison, HPV-positive and -negative cervical cancer cell lines were also included. The HPV screening was performed using TaKaRa polymerase chain reaction. On immunohistochemistry DEK was found to be negative in all 30 non-neoplastic cervical epithelia, but it was positive in 96.1% of SCC (98/102), 92.2% of adenocarcinomas (47/51), 100% of adenoSCC (6/6), 85.7% of CIN-1 (24/28), 94.1% of CIN-2 (16/17), and 89.5% of CIN-3 (17/19). There was no significant difference between HPV-positive and -negative cervical lesions. Also, strongly positive staining was observed in all aforementioned cervical cancer cell lines regardless of HPV infection, according to immunocytochemistry. In summary, DEK plays an important role in the carcinogenesis of cervical cancers, and can be helpful for early diagnosis, and is a potential therapeutic target.

Caspase‐dependent apoptosis induction by targeted expression of DEK in drosophila involves histone acetylation inhibition

As a nuclear phosphoprotein, proto-oncogene protein DEK is capable to changing chromatin structure. DEK was recently identified as an inhibitor of histone acetylation mediated by p300 and PCAF and to facilitate transcriptional repression. To elucidate the biological functions of DEK in vivo, we have constructed transgenic flies that overexpress the human DEK in the developing eye. Transgenic flies developed a severe rough eye phenotype, which is indicative of ectopically induced apoptosis. Genetic and biochemical analyses, including the rescue of the apoptotic phenotype by pan-caspase inhibitor protein p35 and caspase activity analyses, suggested that DEK induces apoptotic cell death through a caspases-9 and -3 dependent pathway. Using extracts from larval salivary glands, we have determined that the global histone acetylation levels of histone H3 Lys9 and H4 Lys5 were decreased upon DEK overexpression. Using chromatin immunoprecipitation assays, we have demonstrated that overexpression of DEK induced the histone H3 and H4 hypoacetylation of promoter of the antiapoptotic gene bcl-2. Co-expression of bcl-2 also rescued apoptosis and the reduced expression of bcl-2 gene was analyzed by real-time PCR. Our results indicate that acidic domain containing protein DEK might have a role in modulating both transcriptional regulation and apoptosis through HAT inhibitory activity.

The Human DEK Proto-Oncogene Is a Senescence Inhibitor and an Upregulated Target of High-Risk Human Papillomavirus E7

The human DEK proto-oncogene is a nucleic acid binding protein with suspected roles in human carcinogenesis, autoimmune disease, and viral infection. Intracellular DEK functions, however, are poorly understood. In papillomavirus-positive cervical cancer cells, downregulation of viral E6/E7 oncogene expression results in cellular senescence. We report here the specific repression of DEK message and protein levels in senescing human papillomavirus type 16- (HPV16-) and HPV18-positive cancer cell lines as well as in primary cells undergoing replicative senescence. Cervical cancer cell senescence was partially overcome by DEK overexpression, and DEK overexpression was sufficient for extending the life span of primary keratinocytes, supporting critical roles for this molecule as a senescence regulator. In order to determine whether DEK is a bona fide HPV oncogene target in primary cells, DEK expression was monitored in human keratinocytes transduced with HPV E6 and/or E7. The results identify high-risk HPV E7 as a positive DEK regulator, an activity that is not shared by low-risk HPV E7 protein. Experiments in mouse embryo fibroblasts recapitulated the observed E7-mediated DEK induction and demonstrated that both basal and E7-induced regulation of DEK expression are controlled by the retinoblastoma protein family. Taken together, our results suggest that DEK upregulation may be a common event in human carcinogenesis and may reflect its senescence inhibitory function.