PURPOSE: Breast cancer is one of the most common malignancies among females, and its prognosis is affected by a complex network of gene interactions. Weighted gene co-expression network analysis was used to construct free-scale gene co-expression networks and to identify potential biomarkers for breast cancer progression.
METHODS: The gene expression profiles of GSE42568 were downloaded from the Gene Expression Omnibus database. RNA-sequencing data and clinical information of breast cancer from TCGA were used for validation.
RESULTS: A total of ten modules were established by the average linkage hierarchical clustering. We identified 58 network hub genes in the significant module (R
CONCLUSIONS: AGO2, CDC20, CDCA5, MCM10, MYBL2, and TTK were identified as candidate biomarkers for further basic and clinical research on breast cancer based on co-expression analysis.

OBJECTIVE: Colorectal cancer (CRC) is an extremely common gastrointestinal malignancy. The present study aimed to identify microRNAs (miRNAs) and transcription factors (TFs) associated with tumor development.
METHODS: Three miRNA profile datasets were integrated and analyzed to elucidate the potential key candidate miRNAs in CRC. The starBase database was used to identify the potential targets of common differentially expressed miRNAs (DEMs). Transcriptional Regulatory Element Database and Transcriptional Regulatory Relationships Unraveled by Sentence-based Text databases were used to identify cancer-related TFs and the TF-regulated target genes. Functional and pathway enrichment analyses were performed using the Database for Annotation, Visualization and Integration Discovery (DAVID) database, and the miRNA-TF-gene networks were constructed by Cytoscape. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to detect the expression of genes and miRNAs.
RESULTS: In total, 14 DEMs were found in CRC. By bioinformatics analysis, 5 DEMs (miR-145, miR-497, miR-30a, miR-31, and miR-20a) and 8 TFs (ELK4 (ETS-family transcription factor), myeloblastosis proto-oncogene like (MYBL)1, MYBL2, CEBPA, PPARA, PPARD, PPARG, and endothelial PAS domain protein (EPAS1)) appeared to be associated with CRC and were therefore used to construct miRNA-TF-gene networks. From the networks, we found that miR-20a might play the most important role as an miRNA in the networks. By qRT-PCR, we demonstrated that miR-20a was significantly upregulated in CRC tissues. We also performed qRT-PCR to identify the expression of miR-20a-related TFs (PPARA, PPARD, PPARG, EPAS1). Three of them, PPARA, PPARG, and EPAS1, were downregulated in CRC tissues, with statistically significant differences, while the downregulation of PPARD in CRC tissues was not significantly different. Pathway enrichment analyses indicated that the phosphoinositide 3-kinase (PI3K)-Akt signaling pathway was the most significantly enriched pathway. Two main elements of the PI3K-Akt signaling pathway, phosphatase and tensin homolog deleted on chromosome 10 and B-cell lymphoma 2-associated agonist of cell death, were demonstrated to be downregulated in CRC.
CONCLUSION: The present study identified hub miRNAs and miRNA-related TF regulatory networks in CRC, which might be potential targets for the diagnosis and treatment of CRC.

Neuroblastoma derived from primitive sympathetic neural precursors is a common type of solid tumor in infants. MYCN proto‑oncogene bHLH transcription factor (MYCN) amplification and 1p36 deletion are important factors associated with the poor prognosis of neuroblastoma. Expression levels of MYCN and c‑MYB proto‑oncogene transcription factor (c‑myb) decline during the differentiation of neuroblastoma cells; E2F transcription factor 1 (E2F1) activates the MYCN promoter. However, the underlying mechanism of MYCN overexpression and amplification requires further investigation. In the present study, potential c‑Myb target genes, and the effect of c‑myb RNA interference (RNAi) on MYCN expression and amplification were investigated in MYCN‑amplified neuroblastoma cell lines. The mRNA expression levels and MYCN gene copy number in five neuroblastoma cell lines were determined by quantitative polymerase chain reaction. In addition, variations in potential target gene expression and MYCN gene copy number between pre‑ and post‑c‑myb RNAi treatment groups in MYCN‑amplified Kelly, IMR32, SIMA and MHH‑NB‑11 cell lines, normalized to those of non‑MYCN‑amplified SH‑SY5Y, were examined. To determine the associations between gene expression levels and chromosomal aberrations, MYCN amplification and 1p36 alterations in interphases/metaphases were analyzed using fluorescence in situ hybridization. Statistical analyses revealed correlations between 1p36 alterations and the expression of c‑myb, MYB proto‑oncogene like 2 (B‑myb) and cyclin dependent kinase inhibitor 1A (p21). Additionally, the results of the present study also demonstrated that c‑myb may be associated with E2F1 and L3MBTL1 histone methyl‑lysine binding protein (L3MBTL1) expression, and that E2F1 may contribute to MYCN, B‑myb, p21 and chromatin licensing and DNA replication factor 1 (hCdt1) expression, but to the repression of geminin (GMNN). On c‑myb RNAi treatment, L3MBTL1 expression was silenced, while GMNN was upregulated, indicating G2/M arrest. In addition, MYCN gene copy number increased following treatment with c‑myb RNAi. Notably, the present study also reported a 43.545% sequence identity between upstream of MYCN and Drosophila melanogaster amplification control element 3, suggesting that expression and/or amplification mechanisms of developmentally‑regulated genes may be evolutionarily conserved. In conclusion, c‑myb may be associated with regulating MYCN expression and amplification. c‑myb, B‑myb and p21 may also serve a role against chromosome 1p aberrations. Together, it was concluded that MYCN gene is amplified during S phase, potentially via a replication‑based mechanism.

The oncogenic transcription factor B-Myb is an essential regulator of late cell cycle genes whose activation by phosphorylation is still poorly understood. We describe a stepwise phosphorylation mechanism of B-Myb, which involves sequential phosphorylations mediated by cyclin-dependent kinase (Cdk) and Polo-like kinase 1 (Plk1) and Pin1-facilitated peptidyl-prolyl cis/trans isomerization. Our data suggest a model in which initial Cdk-dependent phosphorylation of B-Myb enables subsequent Pin1 binding and Pin1-induced conformational changes of B-Myb. This, in turn, initiates further phosphorylation of Cdk-phosphosites, enabling Plk1 docking and subsequent Plk1-mediated phosphorylation of B-Myb to finally allow B-Myb to stimulate transcription of late cell cycle genes. Our observations reveal novel mechanistic hierarchies of B-Myb phosphorylation and activation and uncover regulatory principles that might also apply to other Myb family members. Strikingly, overexpression of B-Myb and of factors mediating its activation strongly correlates with adverse prognoses for tumor patients, emphasizing B-Myb's role in tumorigenesis.

The MuvB transcriptional regulatory complex, which controls cell-cycle-dependent gene expression, cooperates with B-Myb to activate genes required for the G2 and M phases of the cell cycle. We have identified the domain in B-Myb that is essential for the assembly of the Myb-MuvB (MMB) complex. We determined a crystal structure that reveals how this B-Myb domain binds MuvB through the adaptor protein LIN52 and the scaffold protein LIN9. The structure and biochemical analysis provide an understanding of how oncogenic B-Myb is recruited to regulate genes required for cell-cycle progression, and the MMB interface presents a potential therapeutic target to inhibit cancer cell proliferation.

Overexpression of the oncogene MYBL2 (B-Myb) is associated with increased cell proliferation and serves as a marker of poor prognosis in cancer. However, the mechanism by which B-Myb alters the cell cycle is not fully understood. In proliferating cells, B-Myb interacts with the MuvB core complex including LIN9, LIN37, LIN52, RBBP4, and LIN54, forming the MMB (Myb-MuvB) complex, and promotes transcription of genes required for mitosis. Alternatively, the MuvB core interacts with Rb-like protein p130 and E2F4-DP1 to form the DREAM complex that mediates global repression of cell cycle genes in G0/G1, including a subset of MMB target genes. Here, we show that overexpression of B-Myb disrupts the DREAM complex in human cells, and this activity depends on the intact MuvB-binding domain in B-Myb. Furthermore, we found that B-Myb regulates the protein expression levels of the MuvB core subunit LIN52, a key adapter for assembly of both the DREAM and MMB complexes, by a mechanism that requires S28 phosphorylation site in LIN52. Given that high expression of B-Myb correlates with global loss of repression of DREAM target genes in breast and ovarian cancer, our findings offer mechanistic insights for aggressiveness of cancers with MYBL2 amplification, and establish the rationale for targeting B-Myb to restore cell cycle control.

Genome-wide association studies of childhood acute lymphoblastic leukemia (ALL) have identified regions of association at PIP4K2A and upstream of BMI1 at chromosome 10p12.31-12.2. The contribution of both loci to ALL risk and underlying functional variants remain to be elucidated. We carried out single nucleotide polymorphism (SNP) imputation across chromosome 10p12.31-12.2 in Latino and non-Latino white ALL cases and controls from two independent California childhood leukemia studies, and additional Genetic Epidemiology Research on Aging study controls. Ethnicity-stratified association analyses were performed using logistic regression, with meta-analysis including 3,133 cases (1,949 Latino, 1,184 non-Latino white) and 12,135 controls (8,584 Latino, 3,551 non-Latino white). SNP associations were identified at both BMI1 and PIP4K2A. After adjusting for the lead PIP4K2A SNP, genome-wide significant associations remained at BMI1, and vice-versa (p

B-Myb has been shown to play an important oncogenic role in several types of human cancers, including non-small-cell lung cancer (NSCLC). We previously found that B-Myb is aberrantly upregulated in NSCLC, and overexpression of B-Myb can significantly promote NSCLC cell growth and motility. In the present study, we have further investigated the therapeutic potential of B-Myb in NSCLC. Kaplan⁻Meier and Cox proportional hazards analysis indicated that high expression of B-Myb is significantly associated with poor prognosis in NSCLC patients. A loss-of-function study demonstrated that depletion of B-Myb resulted in significant inhibition of cell growth and delayed cell cycle progression in NSCLC cells. Notably, B-Myb depletion also decreased NSCLC cell migration and invasion ability as well as colony-forming ability. Moreover, an in vivo study demonstrated that B-Myb depletion caused significant inhibition of tumor growth in a NSCLC xenograft nude mouse model. A molecular mechanistic study by RNA-seq analysis revealed that B-Myb depletion led to deregulation of various downstream genes, including insulin-like growth factor binding protein 3 (IGFBP3). Overexpression of IGFBP3 suppressed the B-Myb-induced proliferation and migration, whereas knockdown of IGFBP3 significantly rescued the inhibited cell proliferation and motility caused by B-Myb siRNA (small interfering RNA). Expression and luciferase reporter assays revealed that B-Myb could directly suppress the expression of IGFBP3. Taken together, our results suggest that B-Myb functions as a tumor-promoting gene via suppressing IGFBP3 and could serve as a novel therapeutic target in NSCLC.

PURPOSE: In this study, we aimed to assess the association between MYBL2 expression/transcription regulatory activity (TRA) and overall survival (OS) in patients with primary hepatocellular carcinoma (HCC) and to explore the factors related to B-Myb TRA.
MATERIALS AND METHODS: Bioinformatic analysis was performed based on data from the cancer genome atlas-liver hepatocellular carcinoma (TCGA-LIHC) and the human protein atlas (HPA).
RESULTS: The death group in TCGA-LIHC had significantly higher MYBL2 RNA and exon expression than the censor group. The high MYBL2 RNA and exon expression groups had significantly worse OS (P<0.01). Univariate and multivariate analysis confirmed that high MYBL2 expression was an independent prognostic factor of unfavourable OS (HR=1.591, 95%CI: 1.119-2.262, P=0.01). One hundred and fourteen out of 188 primary HCC cases in TCGA-LIHC had elevated transcription of B-Myb's downstream genes. High B-Myb TRA was associated with poor OS (P=0.013). Elevated expression of MYBL2, LIN9, LIN52 and FOXM1 were related to the higher TRA of B-Myb in HCC.
CONCLUSION: High MYBL2 expression/TRA are associated with inferior OS in patients with primary HCC. Increased expression of MYBL2, LIN9, LIN52 and FOXM1 are related to higher TRA of B-Myb in HCC.

Breast cancer remains a public health issue on a global scale. The present study aimed to explore the functional role of MYB proto-oncogene like 2 (MYBL2) in breast cancer, as well as underlying mechanisms. The regulatory relationship between miR-143-3p and MYBL2 was analyzed, and the effects of dysregulation of miR-143-3p and MYBL2 on cell proliferation and apoptosis were investigated. The results showed that MYBL2 and miR-143-3p were inversely expressed in breast cancer tissues and cells: MYBL2 was highly expressed, whereas miR-143-3p was lowly expressed. MYBL2 was confirmed as a target gene of miR-143-3p. Suppression of MYBL2 inhibited proliferation and induced apoptosis of breast cancer cells, which was similar to the effects of overexpression of miR-143-3p. Our findings reveal that MYBL2 is targeted by miR-143-3p and regulates breast cancer cell proliferation and apoptosis.

Ginkgetin has been reported to display antitumor activity. However, the relevant pathway integrating cell cycle regulation and signaling pathways involved in growth inhibition in CRC cells remains to be identified. In this study, ginkgetin-treated HCT116 CRC cells exhibited significant dose-dependent growth inhibition with a GI50 value of 4.0 µM for 48-h treatment, together with apoptosis, via G2-phase cell cycle arrest. When HCT116 cells were treated with 10 µM ginkgetin for 48 h, the percentage of cells in G2/M phase increased by 2.2-fold (43.25%) versus the untreated control (19.69%). Ginkgetin regulated the expression of genes that are critically involved in G2 phase arrest cells, such as b‑Myb, CDC2 and cyclin B1. Furthermore, we found that the suppression of b‑Myb expression by ginkgetin was rescued ~5.1-fold by treatment with a miR-34a inhibitor (500 nM) and b‑Myb was downregulated by >80% by 100 nM miR‑34a mimic. These data suggest that the miRNA34a/b‑Myb/cyclin B1 cascade plays a critical role in ginkgetin-induced G2 cell cycle arrest, as well as in the inhibition of HCT116 cell proliferation. Moreover, the administration of ginkgetin (10 mg/kg) reduced tumor volumes by 36.5% and tumor weight by 37.6% in the mice xenografted with HCT116 cells relative to their vehicle-treated counterparts. Therefore, ginkgetin is the first compound shown to regulate b‑Myb by modulating miR-34a, and we suggest the use of ginkgetin as an inducer of G2 arrest for the treatment of CRC.

The chaperone protein and guanine nucleotide exchange factor SmgGDS (RAP1GDS1) is a key promoter of cancer cell proliferation and tumorigenesis. SmgGDS undergoes nucleocytoplasmic shuttling, suggesting that it has both cytoplasmic and nuclear functions that promote cancer. Previous studies indicate that SmgGDS binds cytoplasmic small GTPases and promotes their trafficking to the plasma membrane. In contrast, little is known about the functions of SmgGDS in the nucleus, or how these nuclear functions might benefit cancer cells. Here we show unique nuclear localization and regulation of gene transcription pathways by SmgGDS. Strikingly, SmgGDS depletion significantly reduces expression of over 600 gene products that are targets of the DREAM complex, which is a transcription factor complex that regulates expression of proteins controlling the cell cycle. The cell cycle regulators E2F1, MYC, MYBL2 (B-Myb) and FOXM1 are among the DREAM targets that are diminished by SmgGDS depletion. E2F1 is well known to promote G1 cell cycle progression, and the loss of E2F1 in SmgGDS-depleted cells provides an explanation for previous reports that SmgGDS depletion characteristically causes a G1 cell cycle arrest. We show that SmgGDS localizes in nucleoli, and that RNAi-mediated depletion of SmgGDS in cancer cells disrupts nucleolar morphology, signifying nucleolar stress. We show that nucleolar SmgGDS interacts with the RNA polymerase I transcription factor upstream binding factor (UBF). The RNAi-mediated depletion of UBF diminishes nucleolar localization of SmgGDS and promotes proteasome-mediated degradation of SmgGDS, indicating that nucleolar sequestration of SmgGDS by UBF stabilizes SmgGDS protein. The ability of SmgGDS to interact with UBF and localize in the nucleolus is diminished by expressing DiRas1 or DiRas2, which are small GTPases that bind SmgGDS and act as tumor suppressors. Taken together, our results support a novel nuclear role for SmgGDS in protecting malignant cells from nucleolar stress, thus promoting cell cycle progression and tumorigenesis.

The precise timing of cell cycle gene expression is critical for the control of cell proliferation; de-regulation of this timing promotes the formation of cancer and leads to defects during differentiation and development. Entry into and progression through S phase requires expression of genes coding for proteins that function in DNA replication. Expression of a distinct set of genes is essential to pass through mitosis and cytokinesis. Expression of these groups of cell cycle-dependent genes is regulated by the RB pocket protein family, the E2F transcription factor family, and MuvB complexes together with B-MYB and FOXM1. Distinct combinations of these transcription factors promote the transcription of the two major groups of cell cycle genes that are maximally expressed either in S phase (G1/S) or in mitosis (G2/M). In this review, we discuss recent work that has started to uncover the molecular mechanisms controlling the precisely timed expression of these genes at specific cell cycle phases, as well as the repression of the genes when a cell exits the cell cycle.

BACKGROUND: MYB-related protein B (B-MYB/MYBL2), a member of the myeloblastosis family of transcription factors, has been reported for its role in the genesis and progression of tumors. Forkhead box M1 (FoxM1), another transcriptional factor, is considered to be an independent predictor of poor survival in many solid cancers. The aim of the present study was to investigate the clinical significance of the correlation between MYBL2 and FoxM1 in glioma and the possible mechanism of FoxM1and MYBL2 expression.
METHODS: MYBL2 and FoxM1expression in cancerous tissues and cell lines were determined by reverse transcription-PCR (RT-PCR), Western blotting and immunostaining. The co-expression of MYBL2 and FoxM1 was analyzed in low-grade glioma (LGG) and glioblastoma (HGG) cohorts of TCGA using cBioportal and UCSC Xena. And, the role of MYBL2 and FoxM1 in glioma cell progression and the underlying mechanisms were studied by using small interfering RNA (si-RNA) and pcDNA3.1 + HAvectors. Furthermore, the effects of MYBL2 and FoxM1 in cell proliferation, cell cycle progression, apoptosis, migration, invasion, and adhesion were determined by cell proliferation assays, flow cytometry analysis, transwell migration and cell adhesion assay.
RESULTS: MYBL2 and FoxM1 expression are significantly associated with clinical stages and overall survival of glioma patients. In cohorts of TCGA, patients with high MYBL2 but without radio-chemotherapy had the highest hazard ratio (adjusted HR = 5.29, 95% CI = 1.475-18.969, P < 0.05). Meanwhile, MYBL2 closely related to the FoxM1 expression in 79 glioma tissues (r = 0.742, p < 0.05) and LGG (r = 0.83) and HGG (r = 0.74) cohorts of TCGA. Down regulation of FoxM1 and MYBL2 by siRNAs induced the cell cycle arrest, apoptosis and EMT of glioma cells. Furthermore, inactivations of Akt/FoxM1 signaling by Akt inhibitor and siRNA-FoxM1 reduce the expression of MYBL2 in glioma cells.
CONCLUSIONS: MYBL2 is a key downstream factor of Akt/FoxM1 signaling to promote progression of human glioma, and could be a new candidate gene for molecular targeting therapy and biomarker for radiotherapy of glioma.
TRIAL REGISTRATION: CTXY-1300041-3-2. ChiCTR-COC-15006186 . Registered date: 13 September 2013.

BACKGROUND: An absence of reliable molecular markers has hampered individualised breast cancer treatments, and a major limitation for translational research is the lack of fresh tissue. There are, however, abundant banks of formalin-fixed paraffin-embedded (FFPE) tissue. This study evaluated two platforms available for the analysis of DNA copy number and gene expression using FFPE samples.
METHODS: The cDNA-mediated annealing, selection, extension, and ligation assay (DASL™) has been developed for gene expression analysis and the Molecular Inversion Probes assay (Oncoscan™), were used for copy number analysis using FFPE tissues. Gene expression and copy number were evaluated in core-biopsy samples from patients with breast cancer undergoing neoadjuvant chemotherapy (NAC).
RESULTS: Forty-three core-biopsies were evaluated and characteristic copy number changes in breast cancers, gains in 1q, 8q, 11q, 17q and 20q and losses in 6q, 8p, 13q and 16q, were confirmed. Regions that frequently exhibited gains in tumours showing a pathological complete response (pCR) to NAC were 1q (55%), 8q (40%) and 17q (40%), whereas 11q11 (37%) gain was the most frequent change in non-pCR tumours. Gains associated with poor survival were 11q13 (62%), 8q24 (54%) and 20q (47%). Gene expression assessed by DASL correlated with immunohistochemistry (IHC) analysis for oestrogen receptor (ER) [area under the curve (AUC) = 0.95], progesterone receptor (PR)(AUC = 0.90) and human epidermal growth factor type-2 receptor (HER-2) (AUC = 0.96). Differential expression analysis between ER+ and ER- cancers identified over-expression of TTF1, LAF-4 and C-MYB (p ≤ 0.05), and between pCR vs non-pCRs, over-expression of CXCL9, AREG, B-MYB and under-expression of ABCG2.
CONCLUSION: This study was an integrative analysis of copy number and gene expression using FFPE core biopsies and showed that molecular marker data from FFPE tissues were consistent with those in previous studies using fresh-frozen samples. FFPE tissue can provide reliable information and will be a useful tool in molecular marker studies.
TRIAL REGISTRATION: Trial registration number ISRCTN09184069 and registered retrospectively on 02/06/2010.

AIMS: Adenoid cystic carcinoma (AdCC) is one of the most common salivary gland malignancies and the long-term prognosis is poor. In this study, we examined alterations of AdCC-associated genes, MYB, MYBL1, MYBL2 and NFIB, and their target molecules, including MYC. The results were correlated to clinicopathological profile of the patients.
METHODS AND RESULTS: Using paraffin tumour sections from 33 cases of salivary gland AdCC, we performed a detailed fluorescence in-situ hybridization (FISH) analysis for gene splits and fusions of MYB, MYBL1, MYBL2 and NFIB. We found that 29 of 33 (88%) AdCC cases showed gene splits in either MYB, MYBL1 or NFIB. None of the cases showed an MYBL2 gene alteration. AdCCs were divided genetically into six gene groups, MYB-NFIB (n = 16), MYB-X (n = 4), MYBL1-NFIB (n = 2), MYBL1-X (n = 1), NFIB-X (n = 6) and gene-split-negative (n = 4). AdCC patients showing the MYB or MYBL1 gene splits were associated with microscopically positive surgical margins (P = 0.0148) and overexpression of MYC (P = 0.0164). MYC expression was detected in both ductal and myoepithelial tumour cells, and MYC overexpression was associated with shorter disease-free survival of the patients (P = 0.0268).
CONCLUSIONS: The present study suggests that (1) nearly 90% of AdCCs may have gene alterations of either MYB, MYBL1 or NFIB, suggesting the diagnostic utility of the FISH assay, (2) MYB or MYBL1 gene splits may be associated with local aggressiveness of the tumours and overexpression of MYC, which is one of the oncogenic MYB/MYBL1 targets and (3) MYC overexpression may be a risk factor for disease-free survival in AdCC.

MYBL2 (B-MYB), a member of the MYB family of transcription factor genes, regulates the expression of genes in the process of tumorigenesis. Many studies have shown that MYBL2 is high expresssion in several human malignancies including pancreatic ductal adenocarcinoma (PDAC). However, its role in PDAC is still unclear. The present study is designed to investigate MYBL2 expression levels and prognostic significance in PDAC patients. We assessed MYBL2 expression level by immunohistochemistry in tumor tissues from 93 PDAC patients undergoing curative resection. The association of MYBL2 expression with clinicopathological parameters was evaluated by Pearson's chi-square (χ2) test, Fisher's exact test, and Spearman's rank. Kaplan-Meier survival analysis and Cox proportional hazards models were used to estimate the effect of MYBL2 expression on survival. The expression of MYBL2 was significantly higher in PDAC cells compared with adjacent non-cancerous tissues (P=0.000). The overexpression of MYBL2 in the tumor tissues was significantly correlated with a higher T classification (p=0.002), peri-neural invasion (PNI) (p=0.013) and vital status (p=0.045). Kaplan-Meier analysis indicated that high MYBL2 expression was significantly associated with shorter overall survival times in PDAC patients. Moreover, univariate and multivariate analysis confirmed MYBL2 expression (P=0.010), histological grade (P=0.001) as independent prognostic factors in PDAC. These results suggested that overexpression of MYBL2 might serve as a novel prognostic biomarker in PDAC patients.

B-Myb is a transcription factor that is overexpressed and plays an oncogenic role in several types of human cancers. However, its potential implication in lung cancer remains elusive. In the present study, we have for the first time investigated the expression profile of B-Myb and its functional impact in lung cancer. Expression analysis by quantificational real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry demonstrated that B-Myb expression is aberrantly overexpressed in non-small cell lung cancer (NSCLC), and positively correlated with pathologic grade and clinical stage of NSCLC. A gain-of-function study revealed that overexpression of B-Myb significantly increases lung cancer cell growth, colony formation, migration, and invasion. Conversely, a loss-of-function study showed that knockdown of B-Myb decreases cell growth, migration, and invasion. B-Myb overexpression also promoted tumor growth in vivo in a NSCLC xenograft nude mouse model. A molecular mechanistic study by RNA-sequencing (RNA-seq) analysis showed that B-Myb overexpression causes up-regulation of various downstream genes (e.g.,

In this study, we investigated whether there is any association between the expression of FSCN1 and SNAI2 and the possible underlying mechanisms in head and neck squamous cell carcinoma (HNSC). In addition, we also investigated whether FSCN1 modulates epithelial-to-mesenchymal transition (EMT) in HNSC cells. Microarray data of dysregulated genes in HNSC were searched in GEO datasets. The association between FSCN1 expression and the 5-year/10-year overall survival (OS), as well as the correlation between the expression of FSCN1 and SOX2, MYBL2, SNAI2, STAT1, and SOX4, was analyzed based on data in TCGA HNSC cohort (TCGA-HNSC). The binding site of SNAI2 in FSCN1 promoter was verified using luciferase reporter assay. SCC9 and SCC15 cells were transfected with pCMV-SNAI2 or pCMV-FSCN1 expression vector or the empty control. Alteration of E-cadherin, Claudin 1, Vimentin, and N-cadherin was then quantified. Our results showed that FSCN1 is significantly upregulated in HNSC tissues compared with the normal control tissues. High FSCN1 expression is associated with worse 5-year and 10-year OS among the HNSC patients. Bioinformatic prediction showed a highly possible SNAI2 binding site in FSCN1 promoter and following luciferase reporter assay verified this site. SNAI2 overexpression significantly increased FSCN1 expression at both mRNA and protein level. FSCN1 overexpression reduced the expression of E-cadherin and Claudin 1, but increased the expression of Vimentin and N-cadherin in SCC9 and SCC-15 cells. Therefore, we infer that FSCN1 is a downstream effector of SNAI2 in promoting EMT in HNSC cells.

Selective serotonin reuptake inhibitors (SSRI) are common antidepressants which cytotoxicity has been assessed in cancers notably colorectal carcinomas and glioma cell lines. We assessed and compared the cytotoxicity of 2 SSRI, citalopram and escitalopram, on neuroblastoma cell lines. The study was performed on 2 non-MYCN amplified cell lines (rat B104 and human SH-SY5Y) and 2 human MYCN amplified cell lines (IMR32 and Kelly). Citalopram and escitalopram showed concentration-dependent cytotoxicity on all cell lines. Citalopram was more cytotoxic than escitalopram. IMR32 was the most sensitive cell line. The absence of toxicity on human primary Schwann cells demonstrated the safety of both molecules for myelin. The mechanisms of cytotoxicity were explored using gene-expression profiles and quantitative real-time PCR (qPCR). Citalopram modulated 1 502 genes and escitalopram 1 164 genes with a fold change ≥ 2. 1 021 genes were modulated by both citalopram and escitalopram; 481 genes were regulated only by citalopram while 143 genes were regulated only by escitalopram. Citalopram modulated 69 pathways (KEGG) and escitalopram 42. Ten pathways were differently modulated by citalopram and escitalopram. Citalopram drastically decreased the expression of MYBL2, BIRC5 and BARD1 poor prognosis factors of neuroblastoma with fold-changes of -107 (p<2.26 10-7), -24.1 (p<5.6 10-9) and -17.7 (p<1.2 10-7). CCNE1, AURKA, IGF2, MYCN and ERBB2 were more moderately down-regulated by both molecules. Glioma markers E2F1, DAPK1 and CCND1 were down-regulated. Citalopram displayed more powerful action with broader and distinct spectrum of action than escitalopram.

The homogeneity and heterogeneity in somatic mutations, copy number alterations and methylation across different cancer types have been extensively explored. However, the related exploration based on transcriptome data is lacking. In this study we explored gene expression profiles across 33 human cancer types using The Cancer Genome Atlas (TCGA) data. We identified consistently upregulated genes (such as E2F1, EZH2, FOXM1, MYBL2, PLK1, TTK, AURKA/B and BUB1) and consistently downregulated genes (such as SCARA5, MYOM1, NKAPL, PEG3, USP2, SLC5A7 and HMGCLL1) across various cancers. The dysregulation of these genes is likely to be associated with poor clinical outcomes in cancer. The dysregulated pathways commonly in cancers include cell cycle, DNA replication, repair, and recombination, Notch signaling, p53 signaling, Wnt signaling, TGFβ signaling, immune response etc. We also identified genes consistently upregulated or downregulated in highly-advanced cancers compared to lowly-advanced cancers. The highly (low) expressed genes in highly-advanced cancers are likely to have higher (lower) expression levels in cancers than in normal tissue, indicating that common gene expression perturbations drive cancer initiation and cancer progression. In addition, we identified a substantial number of genes exclusively dysregulated in a single cancer type or inconsistently dysregulated in different cancer types, demonstrating the intertumor heterogeneity. More importantly, we found a number of genes commonly dysregulated in various cancers such as PLP1, MYOM1, NKAPL and USP2 which were investigated in few cancer related studies, and thus represent our novel findings. Our study provides comprehensive portraits of transcriptional landscape of human cancers.

BACKGROUND: The purpose of this study was to test whether immunohistochemical (IHC) Ki67 levels after short-term preoperative hormone therapy (post-Ki67) predict similar numbers of patients with favorable prognoses as genomic markers.
RESULTS: Thirty paired cases (60 samples) were enrolled in this study. Post-Ki67 levels were significantly lower than pre-treatment Ki67 levels (P < 0.001). Post-Ki67 predicted more low-risk cases (83.3%, 25/30) than pre-genomic surrogate signature(GSS) (66.7%: 20/30), but the difference in predictive power was not significant (P = 0.233). Proliferation (MKI67, STK15, Survivin, CCNB1, and MYBL2) and estrogen (ER, PGR, BCL2, and SCUBE2) related signatures were significantly downregulated after therapy (P < 0.001 and 0.041, respectively).
MATERIALS AND METHODS: Core needle biopsy specimens of primary breast cancer were collected at Okayama University Hospital from hormone receptor-positive and human epidermal growth factor 2-negative patients that subsequently received two weeks of neoadjuvant hormone therapy. Paired post-treatment specimens from surgical samples were also collected. IHC Ki67 levels and GSS were compared between pre- and post-hormone treatment samples. Changes of gene expression pattern in short-term hormone therapy were also assessed.
CONCLUSIONS: IHC based post-Ki67 levels may have distinct predictive power compared with the naïve IHC Ki67. Future studies with larger cohorts and longer follow-up periods may be needed to validate our results.

BACKGROUND: Lung adenocarcinoma is one of most threatening disease to human health. Although many efforts have been devoted to its genetic study, few researches have been focused on the transcription factors which regulate tumor initiation and progression by affecting multiple downstream gene transcription. It is proved that proper transcription factors may mediate the direct reprogramming of cancer cells, and reverse the tumorigenesis on the epigenetic and transcription levels.
METHODS: In this paper, a computational method is proposed to identify the core transcription factors that can regulate as many as possible lung adenocarcinoma associated genes with as little as possible redundancy. A greedy strategy is applied to find the smallest collection of transcription factors that can cover the differentially expressed genes by its downstream targets. The optimal subset which is mostly enriched in the differentially expressed genes is then selected.
RESULTS: Seven core transcription factors (MCM4, VWF, ECT2, RBMS3, LIMCH1, MYBL2 and FBXL7) are detected, and have been reported to contribute to tumorigenesis of lung adenocarcinoma. The identification of the transcription factors provides a new insight into its oncogenic role in tumor initiation and progression, and benefits the discovery of functional core set that may reverse malignant transformation and reprogram cancer cells.

The key signature of cancer genomes is the accumulation of DNA mutations, the most abundant of which is the cytosine-to-thymine (C-to-T) transition that results from cytosine deamination. Analysis of The Cancer Genome Atlas (TCGA) database has demonstrated that this transition is caused mainly by upregulation of the cytosine deaminase APOBEC3B (A3B), but the mechanism has not been completely characterized. We found that B-Myb (encoded by MYBL2) binds the A3B promoter, causing transactivation, and this is responsible for the C-to-T transitions and DNA hypermutation in breast cancer cells. Analysis of TCGA database yielded similar results, supporting that MYBL2 and A3B are upregulated and putatively promote C-to-T transitions in multiple cancer types. Moreover, blockade of EGF receptor with afatinib attenuated B-Myb-A3B signaling, suggesting a clinically relevant means of suppressing mutagenesis. Our results suggest that B-Myb-A3B contributes to DNA damage and could be targeted by inhibiting EGF receptor.

The MuvB multiprotein complex, together with B-MYB and FOXM1 (MMB-FOXM1), plays an essential role in cell cycle progression by regulating the transcription of genes required for mitosis and cytokinesis. In many tumors, B-MYB and FOXM1 are overexpressed as part of the proliferation signature. However, the transcriptional targets that are important for oncogenesis have not been identified. Given that mitotic kinesins are highly expressed in cancer cells and that selected kinesins have been reported as target genes of MMB-FOXM1, we sought to determine which mitotic kinesins are directly regulated by MMB-FOXM1. We demonstrate that six mitotic kinesins and two microtubule-associated non-motor proteins (MAPs) CEP55 and PRC1 are direct transcriptional targets of MuvB, B-MYB and FOXM1 in breast cancer cells. Suppression of KIF23 and PRC1 strongly suppressed proliferation of MDA-MB-231 cells. The set of MMB-FOXM1 regulated kinesins genes and 4 additional kinesins which we referred to as the mitotic kinesin signature (MKS) is linked to poor outcome in breast cancer patients. Thus, mitotic kinesins could be used as prognostic biomarker and could be potential therapeutic targets for the treatment of breast cancer.

How lncRNA MALAT1 is regulated by miRNAs at posttranscriptional level in pancreatic cancer and their regulative effects on the cancer cells remain largely unknown. By retrieving previous miRNA array data and performing primary qRT-PCR, we observed a significant negative correlation between miR-216a and MALAT1 in pancreatic ductal adenocarcinoma (PDAC) tissues and adjacent normal tissues. The following in vitro cell assay further confirmed a direct binding between miR-216a and MALAT1 and the suppressive effect of miR-216a on MALAT1 expression. MiR-216a overexpression had similar effects as MALAT1 siRNA on restoring p21 and p27 expression and inhibiting B-MYB, RAF1 and PCNA1 expression in both PANC-1 and BxPC3 cells. MiR-216a overexpression and MALAT1 knockdown induced cell cycle arrest at G2/M phase. MiR-216a overexpression not only significantly induced cell apoptosis, but also reduced cell viability and increased cell apoptosis in response to gemcitabine in the cancer cells. Based on these findings, we infer that miR-216a induces apoptosis both in the presence and absence of gemcitabine in pancreatic cancer cells by silencing MALAT1 expression.

Epithelial ovarian cancer (EOC) generates multicellular aggregates called spheroids that detach from the primary tumor and disseminate through ascites. Spheroids possess a number of characteristics of tumor dormancy including withdrawal from the cell cycle and resistance to chemotherapeutics. This report systematically analyzes the effects of RNAi depletion of 21 genes that are known to contribute to negative regulation of the cell cycle in 10 ovarian cancer cell lines. Interestingly, spheroid cell viability was compromised by loss of some cyclin-dependent kinase inhibitors such as p57

PURPOSE: Neoadjuvant systemic therapy (NAC) is currently used in the treatment of stage II/III breast cancer. Pathological complete response as a surrogate endpoint for clinical outcomes is not completely validated for all subgroups of breast cancers. Therefore, there is a need for reliable predictive tests of the most effective treatment.
METHODS: We used a combination of predictive clinical, pathological, and gene expression-based markers of response to NAC in a prospective phase II multicentre randomized clinical trial in breast cancer patients, with a long follow-up (8 years). This study concerned the subpopulation of 188 patients with similar levels of pathological response rates to sequential epirubicin/cyclophosphamide and docetaxel to determine predictive marker of pCR and DFS. We used a set of 45 genes selected from high throughput analysis and a standardized RT-qPCR. We analyzed the predictive markers of pathological complete response (pCR) and DFS in the overall population and DFS the subpopulation of 159 patients with no pCR.
RESULTS: In the overall population, combining both clinical and genomic variables, large tumor size, low TFF1, and MYBL2 overexpression were significantly associated with pCR. T4 Stage, lymphovascular invasion, negative PR status, histological type, and high values of CCNB1 were associated with DFS. In the no pCR population, only lymphovascular invasion and high values of BIRC5 were associated with DFS.
CONCLUSIONS: We confirm the importance of ER-related and proliferation genes in the prediction of pCR in NAC-treated breast cancer patients. Furthermore, we identified BIRC5 (survivin) as a main pejorative prognostic factor in patients with breast cancers with no pCR. These results also open perspective for predictive markers of new targeted therapies.

BACKGROUND: African American (AA) colon cancer patients have a worse prognosis than Caucasian (CA) colon cancer patients, however, reasons for this disparity are not well understood. To determine if tumor biology might contribute to differential prognosis, we measured recurrence risk and gene expression using the Oncotype DX® Colon Cancer Assay (12-gene assay) and compared the Recurrence Score results and gene expression profiles between AA patients and CA patients with stage II colon cancer.
METHODS: We retrieved demographic, clinical, and archived tumor tissues from stage II colon cancer patients at four institutions. The 12-gene assay and mismatch repair (MMR) status were performed by Genomic Health (Redwood City, California). Student's t-test and the Wilcoxon rank sum test were used to compare Recurrence Score data and gene expression data from AA and CA patients (SAS Enterprise Guide 5.1).
RESULTS: Samples from 122 AA and 122 CA patients were analyzed. There were 118 women (63 AA, 55 CA) and 126 men (59 AA, 67 CA). Median age was 66 years for AA patients and 68 for CA patients. Age, gender, year of surgery, pathologic T-stage, tumor location, the number of lymph nodes examined, lymphovascular invasion, and MMR status were not significantly different between groups (p = 0.93). The mean Recurrence Score result for AA patients (27.9 ± 12.8) and CA patients (28.1 ± 11.8) was not significantly different and the proportions of patients with high Recurrence Score values (≥41) were similar between the groups (17/122 AA; 15/122 CA). None of the gene expression variables, either single genes or gene groups (cell cycle group, stromal group, BGN1, FAP, INHBA1, Ki67, MYBL2, cMYC and GADD45B), was significantly different between the racial groups. After controlling for clinical and pathologic covariates, the means and distributions of Recurrence Score results and gene expression profiles showed no statistically significant difference between patient groups.
CONCLUSION: The distribution of Recurrence Score results and gene expression data was similar in a cohort of AA and CA patients with stage II colon cancer and similar clinical characteristics, suggesting that tumor biology, as represented by the 12-gene assay, did not differ between patient groups.

Cell cycle (CC) and TP53 regulatory networks are frequently deregulated in cancer. While numerous genome-wide studies of TP53 and CC-regulated genes have been performed, significant variation between studies has made it difficult to assess regulation of any given gene of interest. To overcome the limitation of individual studies, we developed a meta-analysis approach to identify high confidence target genes that reflect their frequency of identification in independent datasets. Gene regulatory networks were generated by comparing differential expression of TP53 and CC-regulated genes with chromatin immunoprecipitation studies for TP53, RB1, E2F, DREAM, B-MYB, FOXM1 and MuvB. RNA-seq data from p21-null cells revealed that gene downregulation by TP53 generally requires p21 (CDKN1A). Genes downregulated by TP53 were also identified as CC genes bound by the DREAM complex. The transcription factors RB, E2F1 and E2F7 bind to a subset of DREAM target genes that function in G1/S of the CC while B-MYB, FOXM1 and MuvB control G2/M gene expression. Our approach yields high confidence ranked target gene maps for TP53, DREAM, MMB-FOXM1 and RB-E2F and enables prediction and distinction of CC regulation. A web-based atlas at www.targetgenereg.org enables assessing the regulation of any human gene of interest.

The expression of genes required for progression through the cell cycle is highly modulated through a regulatory axis containing the E2F transcription factor and retinoblastoma tumour suppressor protein families. One of the genes regulated through this mechanism encodes the B-Myb transcription factor, which has been shown to be critically required for early embryonal development in the mouse. Transcriptional activity of B-Myb is substantially enhanced in S phase through modification by cyclin A/cdk2, and the evidence points squarely to the major role being played by B-Myb during this phase of the cell cycle. We discuss in this review recent findings suggesting that B-Myb is a multifunctional protein that has, in addition to its transcriptional properties, the ability to interact directly with other regulators of the cell cycle.

A defining feature of basal-like breast cancer, a breast cancer subtype with poor clinical prognosis, is the high expression of 'proliferation signature' genes. We identified B-Myb, a MYB family transcription factor that is often amplified and overexpressed in many tumor types, as being highly expressed in the proliferation signature. However, the roles of B-Myb in disease progression, and its mammary-specific transcriptional targets, are poorly understood. Here, we showed that B-Myb expression is a significant predictor of survival and pathological complete response to neoadjuvant chemotherapy in breast cancer patients. We also identified a significant association between the G/G genotype of a nonsynonymous B-Myb germline variant (rs2070235, S427G) and an increased risk of basal-like breast cancer [OR 2.0, 95% CI (1.1-3.8)]. In immortalized, human mammary epithelial cell lines, but not in basal-like tumor lines, cells ectopically expressing wild-type B-Myb or the S427G variant showed increased sensitivity to two DNA topoisomerase IIalpha inhibitors, but not to other chemotherapeutics. In addition, microarray analyses identified many G2/M genes as being induced in B-Myb overexpressing cells. These results confirm that B-Myb is involved in cell cycle control, and that its dysregulation may contribute to increased sensitivity to a specific class of chemotherapeutic agents. These data provide insight into the influence of B-Myb in human breast cancer, which is of potential clinical importance for determining disease risk and for guiding treatment.

The B-MYB proto-oncogene is a transcription factor belonging to the MYB family that is frequently overexpressed or amplified in different types of human malignancies. While it is suspected that B-MYB plays a role in human cancer, there is still no direct evidence of its causative role. Looking for mutations of the B-MYB gene in human cell lines and primary cancer samples, we frequently isolated two nonsynonymous B-MYB polymorphic variants (rs2070235 and rs11556379). Compared to the wild-type protein, the B-MYB isoforms display altered conformation, impaired regulation of target genes and decreased antiapoptotic activity, suggesting that they are hypomorphic variants of the major allele. Importantly, the B-MYB polymorphisms are common; rs2070235 and rs11556379 are found, depending on the ethnic background, in 10-50% of human subjects. We postulated that, if B-MYB activity is important for transformation, the presence of common, hypomorphic variants might modify cancer risk. Indeed, the B-MYB polymorphisms are underrepresented in 419 cancer patients compared to 230 controls (odds ratio 0.53; (95%) confidence interval 0.385-0.755; P=0.001). This data imply that a large fraction of the human population is carrier of B-MYB alleles that might be associated with a reduced risk of developing neoplastic disease.

The transcription factors of the Myb family are expressed in several tissues and play an important role in cell proliferation, differentiation, and survival In this study, the expression of A-myb, B-myb, and c-myb was investigated in a group of 64 neuroblastomas at different dinical stages by a sensitive reverse transcription-PCR tchnique and correlated with patients' survival. All of the myb genes were frequently expressed in neuroblastoma tumors. Interestingly, the expression of B-myb, which was detected in 33 cases, was associated with an increased risk of death (P = 0.027 in a univariate analysis), whereas there was no correlation with A-myb and c-myb expression. In addition, in a multivariate Cox regression analysis that included myb gene expression, MYCN status, age at diagnosis, and tumor staging, MYCN amplification and B-myb expression were independently associated to an increased risk (P < 0.01 and P = 0.015, respectively). In overall survival curves obtained by stratifying the neuroblastoma cases on the basis of MYCN status and B-myb expression, the group of patients without MYCN amplification and positive for B-myb expression had worse survival probability than that without MYCN amplification and nonexpressing B-myb (P < 0.01). In summary, these findings provide the first demonstration that B-myb expression can be a useful prognostic marker in human neuroblastoma. Moreover, B-myb expression has a prognostic value complementary to MYCN amplification and can identify a group of high-risk patients that would not be predicted on the basis of the MYCN status only.