Gene Summary

Gene:SMAD4; SMAD family member 4
Aliases: JIP, DPC4, MADH4, MYHRS
Summary:This gene encodes a member of the Smad family of signal transduction proteins. Smad proteins are phosphorylated and activated by transmembrane serine-threonine receptor kinases in response to TGF-beta signaling. The product of this gene forms homomeric complexes and heteromeric complexes with other activated Smad proteins, which then accumulate in the nucleus and regulate the transcription of target genes. This protein binds to DNA and recognizes an 8-bp palindromic sequence (GTCTAGAC) called the Smad-binding element (SBE). The Smad proteins are subject to complex regulation by post-translational modifications. Mutations or deletions in this gene have been shown to result in pancreatic cancer, juvenile polyposis syndrome, and hereditary hemorrhagic telangiectasia syndrome. [provided by RefSeq, Oct 2009]
Databases:OMIM, VEGA, HGNC, Ensembl, GeneCard, Gene
Protein:mothers against decapentaplegic homolog 4
Source:NCBIAccessed: 16 March, 2015

Cancer Overview

Research Indicators

Publications Per Year (1990-2015)
Graph generated 16 March 2015 using data from PubMed using criteria.

Literature Analysis

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Tag cloud generated 16 March, 2015 using data from PubMed, MeSH and CancerIndex

Specific Cancers (8)

Latest Publications: SMAD4 (cancer-related)

Terris B, Cavard C
Diagnosis and molecular aspects of solid-pseudopapillary neoplasms of the pancreas.
Semin Diagn Pathol. 2014; 31(6):484-90 [PubMed] Related Publications
Solid-pseudopapillary neoplasm of the pancreas (SPN) is an uncommon low-grade malignant neoplasm occurring mostly in young women. In addition to its distinctive pathological appearance of pseudopapillae with poorly cohesive neoplastic cells, rare variants exist raising the differential diagnosis especially with neuroendocrine neoplasms. The overall prognosis for patients with SPNs is excellent after surgical resection. Nevertheless, 10% of cases may have malignant behavior characterized by tumor recurrence and/or metastasis. Despite numerous studies, the histogenesis of this neoplasm remains unclear. Distinctive molecular alterations such as the presence of CTNNB1 mutations are observed in nearly all cases, while mutations classically observed in ductal adenocarcinoma, such as KRAS, TP53, and SMAD4, are not observed in SPNs, reinforcing its distinct nature compared to all other pancreatic neoplasms. Recent transcriptional studies have shown that activation of the Wnt/beta-catenin pathway in these tumors is associated with the upregulation of genes belonging to Notch, Hedgehog, and androgen receptor signaling pathways.

Reid MD, Choi H, Balci S, et al.
Serous cystic neoplasms of the pancreas: clinicopathologic and molecular characteristics.
Semin Diagn Pathol. 2014; 31(6):475-83 [PubMed] Related Publications
We herein summarize the pathology and most recent advances in the molecular genetics of serous cystic neoplasms of the pancreas. They typically present as relatively large, well-demarcated tumors (mean size, 6cm), predominantly occurring in females. Pre-operative diagnosis remains challenging; imaging findings and cyst fluid analysis often prove non-specific and fine-needle aspiration often does not yield diagnostic cells. Pathologically, they are characterized by a distinctive cytology referred to as "serous." Although they have ductal differentiation, they distinctly lack the mucin production that characterizes most other pancreatic ductal tumors, including ductal adenocarcinoma and its variants, intraductal papillary mucinous neoplasm (IPMN) and mucinous cystic neoplasm (MCN). They instead produce abundant glycogen (glycogen-rich adenoma). Serous cystadenomas also lack the molecular alterations that characterize ductal neoplasms, such as mutation of KRAS (high prevalence in most mucinous ductal neoplasms), inactivation of SMAD4 (seen in ductal adenocarcinomas), and mutations in GNAS (seen in some IPMNs) and RNF43 (detected in MCNs and IPMNs). Instead, new molecular and immunohistochemical observations place serous pancreatic tumors closer to "clear cell neoplasms" seen in various other organs that are associated with the von Hippel-Lindau (VHL) pathway, such as clear cell renal cell carcinomas and capillary hemangioblastomas. Patients with VHL syndrome have an increased risk of developing serous pancreatic tumors and somatic mutations of the VHL gene are common in these tumors along with modification of its downstream effectors including hypoxia-inducible factor (HIF1), glucose uptake and transporter-1 (GLUT-1), a common factor in clear cell (glycogen-rich) tumors, as well as expression of vascular endothelial growth factor (VEGF), thought to be a factor in the striking capillarization of serous cystadenomas and other non-pancreatic clear cell tumors. VEGF may prove to be of significant diagnostic value since its elevation in cyst fluid has recently been found highly sensitive and specific for serous neoplasms. These molecular alterations establish serous tumors as prototypes of clear cell tumorigenesis and angiogenesis and may prove helpful both as diagnostic and non-surgical therapeutic targets.

Wood LD, Klimstra DS
Pathology and genetics of pancreatic neoplasms with acinar differentiation.
Semin Diagn Pathol. 2014; 31(6):491-7 [PubMed] Free Access to Full Article Related Publications
Pancreatic neoplasms with acinar differentiation, including acinar cell carcinoma, pancreatoblastoma, and carcinomas with mixed differentiation, are distinctive pancreatic neoplasms with a poor prognosis. These neoplasms are clinically, pathologically, and genetically unique when compared to other more common pancreatic neoplasms. Most occur in adults, although pancreatoblastomas usually affect children under 10 years old. All of these neoplasms exhibit characteristic histologic features including a solid or acinar growth pattern, dense neoplastic cellularity, uniform nuclei with prominent nucleoli, and granular eosinophilic cytoplasm. Exocrine enzymes are detectable by immunohistochemistry and, for carcinomas with mixed differentiation, neuroendocrine or ductal lineage markers are also expressed. The genetic alterations of this family of neoplasms largely differ from conventional ductal adenocarcinomas, with only rare mutations in TP53, KRAS, and p16, but no single gene or neoplastic pathway is consistently altered in acinar neoplasms. Instead, there is striking genomic instability, and a subset of cases has mutations in the APC/β-catenin pathway, mutations in SMAD4, RAF gene family fusions, or microsatellite instability. Therapeutically targetable mutations are often present. This review summarizes the clinical and pathologic features of acinar neoplasms and reviews the current molecular data on these uncommon tumors.

Ma C, Giardiello FM, Montgomery EA
Upper tract juvenile polyps in juvenile polyposis patients: dysplasia and malignancy are associated with foveolar, intestinal, and pyloric differentiation.
Am J Surg Pathol. 2014; 38(12):1618-26 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
Patients with juvenile polyposis syndrome (JPS), a hereditary autosomal dominant hamartomatous polyposis syndrome, are at increased risk for colorectal adenocarcinoma. The upper gastrointestinal tract is less often involved by JPS than the colorectum, and, consequently, upper tract juvenile polyps (JPs) are not well studied. We reviewed upper endoscopies and corresponding biopsies in JPS patients documented in our Polyposis Registry. A total of 199 upper gastrointestinal biopsies from 69 endoscopies were available in 22 of 41 (54%) JPS patients. Thirteen of the 22 patients (59%) had ≥1 gastric JP; 5 also had 6 small bowel JPs. Gastric JP was identified as early as age 7 in a patient with an SMAD4 gene mutation. Two patients (9%) had high-grade dysplasia in gastric JP. Invasive adenocarcinoma was diagnosed in the gastrectomy specimen of 1 patient. Five patients had a huge gastric polyp burden; 3 underwent total gastrectomy. Three patients died of complications associated with extensive upper JP. Histologically, 8 of the 56 (14%) gastric JPs identified had dysplasia. All of the 8 polyps demonstrated intestinalized and pyloric gland differentiation intermixed with foveolar epithelium. Dysplasia was seen arising in all 3 types of epithelium. The flat gastric mucosa in 11 patients was unremarkable without inflammation or intestinal metaplasia. The 6 small bowel JPs had no dysplasia. Our findings suggest that JPS patients are at increased risk for gastric adenocarcinoma. Detection of malignancy in syndromic gastric JP indicates that the current screening procedures are insufficient in removal of precursor lesions to prevent progression to carcinoma.

Aytac E, Sulu B, Heald B, et al.
Genotype-defined cancer risk in juvenile polyposis syndrome.
Br J Surg. 2015; 102(1):114-8 [PubMed] Related Publications
BACKGROUND: Germline mutations in SMAD4 and BMPR1A disrupt the transforming growth factor β signal transduction pathway, and are associated with juvenile polyposis syndrome. The effect of genotype on the pattern of disease in this syndrome is unknown. This study evaluated the differential impact of SMAD4 and BMPR1A gene mutations on cancer risk and oncological phenotype in patients with juvenile polyposis syndrome.
METHODS: Patients with juvenile polyposis syndrome and germline SMAD4 or BMPR1A mutations were identified from a prospectively maintained institutional registry. Medical records were reviewed and the clinical patterns of disease were analysed.
RESULTS: Thirty-five patients had germline mutations in either BMPR1A (8 patients) or SMAD4 (27). Median follow-up was 11 years. Colonic phenotype was similar between patients with SMAD4 and BMPR1A mutations, whereas SMAD4 mutations were associated with larger polyp numbers (number of patients with 50 or more gastric polyps: 14 versus 0 respectively). The numbers of patients with rectal polyps was comparable between BMPR1A and SMAD4 mutation carriers (5 versus 17). No patient was diagnosed with cancer in the BMPR1A group, whereas four men with a SMAD4 mutation developed gastrointestinal (3) or extraintestinal (1) cancer. The gastrointestinal cancer risk in patients with juvenile polyposis syndrome and a SMAD4 mutation was 11 per cent (3 of 27).
CONCLUSION: The SMAD4 genotype is associated with a more aggressive upper gastrointestinal malignancy risk in juvenile polyposis syndrome.

Nummela P, Saarinen L, Thiel A, et al.
Genomic profile of pseudomyxoma peritonei analyzed using next-generation sequencing and immunohistochemistry.
Int J Cancer. 2015; 136(5):E282-9 [PubMed] Related Publications
Pseudomyxoma peritonei (PMP) is a relatively rare clinical syndrome characterized by neoplastic epithelial cells growing in the peritoneal cavity and secreting mucinous ascites. Our aim was to explore the molecular events behind this fatal but under-investigated disease. We extracted DNA from 19 appendix-derived PMP tumors and nine corresponding normal tissues, and analyzed the mutational hotspot areas of 48 cancer-related genes by amplicon-based next-generation sequencing (NGS). Further, we analyzed the protein expression of V600E mutated BRAF, MLH1, MSH2, MSH6 and p53 from a larger set of PMP tumors (n = 74) using immunohistochemistry. With NGS, we detected activating somatic KRAS mutations in all of the tumors studied. GNAS was mutated in 63% of the tumors with no marked difference between low-grade and high-grade tumors. Only one (5.3%) tumor showed oncogenic PIK3CA mutation, one showed oncogenic AKT1 mutation, three (15.8%) showed SMAD4 mutations and none showed an APC mutation. P53 protein was aberrantly expressed in higher proportion of high-grade tumors as compared with low-grade ones (31.3 vs. 7.1%, respectively; p = 0.012) and aberrant expression was an independent factor for reduced overall survival (p = 0.002). BRAF V600E mutation was only found in one (1.4%) high-grade tumor by immunohistochemistry (n = 74). All the studied tumors expressed mismatch repair proteins MLH1, MSH2 and MSH6. Our results indicate that KRAS mutations are evident in all and GNAS mutations in most of the PMPs, but BRAF V600E, PIK3CA and APC mutations are rare. Aberrantly expressed p53 is associated with high-grade histology and reduced survival.

Wang Y, Wu J, Lin B, et al.
Galangin suppresses HepG2 cell proliferation by activating the TGF-β receptor/Smad pathway.
Toxicology. 2014; 326:9-17 [PubMed] Related Publications
Galangin can suppress hepatocellular carcinoma (HCC) cell proliferation. In this study, we demonstrated that galangin induced autophagy by activating the transforming growth factor (TGF)-β receptor/Smad pathway and increased TGF-β receptor I (RI), TGF-βRII, Smad1, Smad2, Smad3 and Smad4 levels but decreased Smad6 and Smad7 levels. Autophagy induced by galangin appears to depend on the TGF-β receptor/Smad signalling pathway because the down-regulation of Smad4 by siRNA or inhibition of TGF-β receptor activation by LY2109761 blocked galangin-induced autophagy. The down-regulation of Beclin1, autophagy-related gene (ATG) 16L, ATG12 and ATG3 restored HepG2 cell proliferation and prevented galangin-induced apoptosis. Our findings indicate a novel mechanism for galangin-induced autophagy via activation of the TGF-β receptor/Smad pathway. The induction of autophagy thus reflects the anti-proliferation effect of galangin on HCC cells.

Yamaguchi J, Nagayama S, Chino A, et al.
Identification of coding exon 3 duplication in the BMPR1A gene in a patient with juvenile polyposis syndrome.
Jpn J Clin Oncol. 2014; 44(10):1004-8 [PubMed] Related Publications
Juvenile polyposis syndrome is an autosomal dominant inherited disorder characterized by multiple juvenile polyps arising in the gastrointestinal tract and an increased risk of gastrointestinal cancers, specifically colon cancer. BMPR1A and SMAD4 germline mutations have been found in patients with juvenile polyposis syndrome. We identified a BMPR1A mutation, which involves a duplication of coding exon 3 (c.230+452_333+441dup1995), on multiple ligation dependent probe amplification in a patient with juvenile polyposis syndrome. The mutation causes a frameshift, producing a truncated protein (p.D112NfsX2). Therefore, the mutation is believed to be pathogenic. We also identified a duplication breakpoint in which Alu sequences are located. These results suggest that the duplication event resulted from recombination between Alu sequences. To our knowledge, partial duplication in the BMPR1A gene has not been reported previously. This is the first case report to document coding exon 3 duplication in the BMPR1A gene in a patient with juvenile polyposis syndrome.

Liszka L
Ductal adenocarcinoma of the pancreas usually retained SMAD4 and p53 protein status as well as expression of epithelial-to-mesenchymal transition markers and cell cycle regulators at the stage of liver metastasis.
Pol J Pathol. 2014; 65(2):100-12 [PubMed] Related Publications
There are limited data on the biology of metastatic pancreatic ductal adenocarcinoma (PDAC). The aim of the present study was to compare the expression of immunohistochemical markers that may be involved in the development of metastatic disease in primary PDAC and in synchronous liver metastatic tissues. Thirty-two stains (corresponding to proteins encoded by 31 genes: SMAD4, TP53, ACTA2, CDH1, CDKN1A, CLDN1, CLDN4, CLDN7, CTNNB1, EGFR, ERBB2, FN1, KRT19, MAPK1/MAPK3, MAPK14, MKI67, MMP2, MMP9, MUC1 (3 antibodies), MUC5AC, MUC6, MTOR, MYC, NES, PTGS2, RPS6, RPS6KB1, TGFB1, TGFBR1, VIM) were evaluated using tissue microarray of 26 pairs of primary PDACs and their liver metastases. There were no significant differences in expression levels of examined proteins between primary and secondary lesions. In particular, metastatic PDAC retained the primary tumour's SMAD4 protein status in all and p53 protein status in all but one case. This surprising homogeneity also involved expression levels of markers of epithelial-to-mesenchymal transition as well as cell cycle regulators studied. In conclusion, the biological profiles of primary PDACs and their liver metastases seemed to be similar. Molecular alterations of PDAC related to a set of immunohistochemical markers examined in the present study were already present at the stage of localized disease.

Weaver JM, Ross-Innes CS, Shannon N, et al.
Ordering of mutations in preinvasive disease stages of esophageal carcinogenesis.
Nat Genet. 2014; 46(8):837-43 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
Cancer genome sequencing studies have identified numerous driver genes, but the relative timing of mutations in carcinogenesis remains unclear. The gradual progression from premalignant Barrett's esophagus to esophageal adenocarcinoma (EAC) provides an ideal model to study the ordering of somatic mutations. We identified recurrently mutated genes and assessed clonal structure using whole-genome sequencing and amplicon resequencing of 112 EACs. We next screened a cohort of 109 biopsies from 2 key transition points in the development of malignancy: benign metaplastic never-dysplastic Barrett's esophagus (NDBE; n=66) and high-grade dysplasia (HGD; n=43). Unexpectedly, the majority of recurrently mutated genes in EAC were also mutated in NDBE. Only TP53 and SMAD4 mutations occurred in a stage-specific manner, confined to HGD and EAC, respectively. Finally, we applied this knowledge to identify high-risk Barrett's esophagus in a new non-endoscopic test. In conclusion, mutations in EAC driver genes generally occur exceptionally early in disease development with profound implications for diagnostic and therapeutic strategies.

Zhang R, Zhao C, Xiong Z, Zhou X
Pathway bridge based multiobjective optimization approach for lurking pathway prediction.
Biomed Res Int. 2014; 2014:351095 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
Ovarian carcinoma immunoreactive antigen-like protein 2 (OCIAD2) is a protein with unknown function. Frequently methylated or downregulated, OCIAD2 has been observed in kinds of tumors, and TGFβ signaling has been proved to induce the expression of OCIAD2. However, current pathway analysis tools do not cover the genes without reported interactions like OCIAD2 and also miss some significant genes with relatively lower expression. To investigate potential biological milieu of OCIAD2, especially in cancer microenvironment, a nova approach pbMOO was created to find the potential pathways from TGFβ to OCIAD2 by searching on the pathway bridge, which consisted of cancer enriched looping patterns from the complicated entire protein interactions network. The pbMOO approach was further applied to study the modulator of ligand TGFβ1, receptor TGFβR1, intermediate transfer proteins, transcription factor, and signature OCIAD2. Verified by literature and public database, the pathway TGFβ1-TGFβR1-SMAD2/3-SMAD4/AR-OCIAD2 was detected, which concealed the androgen receptor (AR) which was the possible transcription factor of OCIAD2 in TGFβsignal, and it well explained the mechanism of TGFβ induced OCIAD2 expression in cancer microenvironment, therefore providing an important clue for the future functional analysis of OCIAD2 in tumor pathogenesis.

Shen G, Lin Y, Yang X, et al.
MicroRNA-26b inhibits epithelial-mesenchymal transition in hepatocellular carcinoma by targeting USP9X.
BMC Cancer. 2014; 14:393 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
BACKGROUND: Metastasis is responsible for the rapid recurrence and poor survival of malignancies. Epithelial-mesenchymal transition (EMT) has a critical role in metastasis. Increasing evidence indicates that EMT can be regulated by microRNAs (miRNAs). The aim of this study was to investigate the role of miR-26b in modulating epithelial-mesenchymal transition (EMT) in hepatocellular carcinoma (HCC), as well as to identify its underlying mechanism of action.
METHODS: The expression level of miR-26b was assessed in multiple HCC cell lines (HepG2, MHCC97H, Hep3B, MHCC97L, HCCC9810, BEL-7402, Huh7 and QGY-7703), as well as in liver tissue from patients with HCC. Follow-up studies examined the effects of a miR-26b mimic (increased expression) and a miR-26b inhibitor (decreased expression) on markers of EMT, wound healing and cell migration. The molecular target of miR-26b was also identified using a computer algorithm and confirmed experimentally.
RESULTS: MiR-26b expression was decreased in HCC cell lines and was inversely correlated with the grade of HCC. Increased expression of miR-26b inhibited the migration and invasiveness of HCC cell lines, which was accompanied by decreased expression of the epithelial marker E-cadherin and increased expression of the mesenchymal marker vimentin, at both the mRNA and protein expression levels. A binding site for miR-26b was theoretically identified in the 3'UTR of USP9X. Further studies revealed that overexpression of miR-26b repressed the endogenous level of USP9X protein expression. Overexpression of miR-26b also repressed Smad4 expression, whereas its inhibition elevated Smad4 expression.
CONCLUSIONS: Taken together, our results indicate that miR-26b were inhibited in HCC. In HCC cell lines, miR-26b targeted the 3'UTR of USP9X, which in turn affects EMT through Smad4 and the TGF-β signaling pathway. Our analysis of clinical HCC samples verifies that miR-26b also targets USP9X expression to inhibit the EMT of hepatocytes. Thus, miR-26b may have some effects on the EMT of HCC cells.

Togashi Y, Sakamoto H, Hayashi H, et al.
Homozygous deletion of the activin A receptor, type IB gene is associated with an aggressive cancer phenotype in pancreatic cancer.
Mol Cancer. 2014; 13:126 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
BACKGROUND: Transforming growth factor, beta (TGFB) signal is considered to be a tumor suppressive pathway based on the frequent genomic deletion of the SMAD4 gene in pancreatic cancer (PC); however; the role of the activin signal, which also belongs to the TGFB superfamily, remains largely unclear.
METHODS AND RESULTS: We found a homozygous deletion of the activin A receptor, type IB (ACVR1B) gene in 2 out of 8 PC cell lines using array-comparative genomic hybridization, and the absence of ACVR1B mRNA and protein expression was confirmed in these 2 cell lines. Activin A stimulation inhibited cellular growth and increased the phosphorylation level of SMAD2 and the expression level of p21CIP1/WAF1 in the Sui66 cell line (wild-type ACVR1B and SMAD4 genes) but not in the Sui68 cell line (homozygous deletion of ACVR1B gene). Stable ACVR1B-knockdown using short hairpin RNA cancelled the effects of activin A on the cellular growth of the PC cell lines. In addition, ACVR1B-knockdown significantly enhanced the cellular growth and colony formation abilities, compared with controls. In a xenograft study, ACVR1B-knockdown resulted in a significantly elevated level of tumorigenesis and a larger tumor volume, compared with the control. Furthermore, in clinical samples, 6 of the 29 PC samples (20.7%) carried a deletion of the ACVR1B gene, while 10 of the 29 samples (34.5%) carried a deletion of the SMAD4 gene. Of note, 5 of the 6 samples with a deletion of the ACVR1B gene also had a deletion of the SMAD4 gene.
CONCLUSION: We identified a homozygous deletion of the ACVR1B gene in PC cell lines and clinical samples and proposed that the deletion of the ACVR1B gene may mediate an aggressive cancer phenotype in PC. Our findings provide novel insight into the role of the activin signal in PC.

Li X, Nadauld L, Ootani A, et al.
Oncogenic transformation of diverse gastrointestinal tissues in primary organoid culture.
Nat Med. 2014; 20(7):769-77 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
The application of primary organoid cultures containing epithelial and mesenchymal elements to cancer modeling holds promise for combining the accurate multilineage differentiation and physiology of in vivo systems with the facile in vitro manipulation of transformed cell lines. Here we used a single air-liquid interface culture method without modification to engineer oncogenic mutations into primary epithelial and mesenchymal organoids from mouse colon, stomach and pancreas. Pancreatic and gastric organoids exhibited dysplasia as a result of expression of Kras carrying the G12D mutation (Kras(G12D)), p53 loss or both and readily generated adenocarcinoma after in vivo transplantation. In contrast, primary colon organoids required combinatorial Apc, p53, Kras(G12D) and Smad4 mutations for progressive transformation to invasive adenocarcinoma-like histology in vitro and tumorigenicity in vivo, recapitulating multi-hit models of colorectal cancer (CRC), as compared to the more promiscuous transformation of small intestinal organoids. Colon organoid culture functionally validated the microRNA miR-483 as a dominant driver oncogene at the IGF2 (insulin-like growth factor-2) 11p15.5 CRC amplicon, inducing dysplasia in vitro and tumorigenicity in vivo. These studies demonstrate the general utility of a highly tractable primary organoid system for cancer modeling and driver oncogene validation in diverse gastrointestinal tissues.

Li L, Li Z, Kong X, et al.
Down-regulation of microRNA-494 via loss of SMAD4 increases FOXM1 and β-catenin signaling in pancreatic ductal adenocarcinoma cells.
Gastroenterology. 2014; 147(2):485-97.e18 [PubMed] Related Publications
BACKGROUND & AIMS: Dysregulation of β-catenin and the transcriptional activator FOXM1 mediate oncogenesis, but it is not clear how these proteins become dysregulated in tumors that do not typically carry mutations in adenomatous polyposis coli (APC) or β-catenin, such as pancreatic ductal adenocarcinomas (PDACs). We searched for microRNAs that regulate levels of FOXM1 in PDAC cells and samples from patients.
METHODS: We identified microRNAs that affect levels of FOXM1 in PDACs using bioinformatic, genetic, and pharmacologic approaches. We altered expression of the microRNA-494 (miR-494) in PDAC cell lines (AsPC-1 and PANC-1) and examined the effects on FOXM1 and β-catenin signaling and cell proliferation and colony formation. The cells were injected into immunocompromised mice and growth of xenograft tumors and liver metastases were measured. We performed immunohistochemical analyses of 10 paired PDAC and nontumor pancreatic tissue samples collected from untreated patients during surgery.
RESULTS: We identified miR-494 as a negative regulator of FOXM1 levels in PDAC cells, and found that levels of this microRNA were reduced in PDAC specimens, compared with nontumor tissues. Loss of response of PDAC cells to transforming growth factor β, owing to SMAD4 deficiency, reduced expression of miR-494. Transgenic expression of miR-494 in PDAC cells produced the same effects as reducing expression of FOXM1 or blocking nuclear translocation of β-catenin, reducing cell proliferation, migration, and invasion, and increasing their sensitivity to gemcitabine. Reduced expression of miR-494 correlated with PDAC metastasis and reduced survival times of patients.
CONCLUSIONS: Loss of SMAD4 in PDAC cells leads to reduced levels of miR-494, increased levels of FOXM1, and nuclear localization of β-catenin. miR-494 might be developed as a prognostic marker for patients with PDAC or a therapeutic target.

Liu X, Mody K, de Abreu FB, et al.
Molecular profiling of appendiceal epithelial tumors using massively parallel sequencing to identify somatic mutations.
Clin Chem. 2014; 60(7):1004-11 [PubMed] Related Publications
BACKGROUND: Some epithelial neoplasms of the appendix, including low-grade appendiceal mucinous neoplasm and adenocarcinoma, can result in pseudomyxoma peritonei (PMP). Little is known about the mutational spectra of these tumor types and whether mutations may be of clinical significance with respect to therapeutic selection. In this study, we identified somatic mutations using the Ion Torrent AmpliSeq Cancer Hotspot Panel v2.
METHODS: Specimens consisted of 3 nonneoplastic retention cysts/mucocele, 15 low-grade mucinous neoplasms (LAMNs), 8 low-grade/well-differentiated mucinous adenocarcinomas with pseudomyxoma peritonei, and 12 adenocarcinomas with/without goblet cell/signet ring cell features. Barcoded libraries were prepared from up to 10 ng of extracted DNA and multiplexed on single 318 chips for sequencing. Data analysis was performed using Golden Helix SVS. Variants that remained after the analysis pipeline were individually interrogated using the Integrative Genomics Viewer.
RESULTS: A single Janus kinase 3 (JAK3) mutation was detected in the mucocele group. Eight mutations were identified in the V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) and GNAS complex locus (GNAS) genes among LAMN samples. Additional gene mutations were identified in the AKT1 (v-akt murine thymoma viral oncogene homolog 1), APC (adenomatous polyposis coli), JAK3, MET (met proto-oncogene), phosphatidylinositol-4,5-bisphosphate 3-kinase (PIK3CA), RB1 (retinoblastoma 1), STK11 (serine/threonine kinase 11), and tumor protein p53 (TP53) genes. Among the PMPs, 6 mutations were detected in the KRAS gene and also in the GNAS, TP53, and RB1 genes. Appendiceal cancers showed mutations in the APC, ATM (ataxia telangiectasia mutated), KRAS, IDH1 [isocitrate dehydrogenase 1 (NADP+)], NRAS [neuroblastoma RAS viral (v-ras) oncogene homolog], PIK3CA, SMAD4 (SMAD family member 4), and TP53 genes.
CONCLUSIONS: Our results suggest molecular heterogeneity among epithelial tumors of the appendix. Next generation sequencing efforts have identified mutational spectra in several subtypes of these tumors that may suggest a phenotypic heterogeneity showing mutations that are relevant for targeted therapies.

Ke TW, Hsu HL, Wu YH, et al.
MicroRNA-224 suppresses colorectal cancer cell migration by targeting Cdc42.
Dis Markers. 2014; 2014:617150 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
The metastatic spread of tumor cells is the major risk factor affecting the clinical prognosis of colorectal cancer (CRC) patients. The metastatic phenotype can be modulated by dysregulating the synthesis of different structural and functional proteins of tumor cells. Micro(mi)RNAs are noncoding RNAs that recognize their cognate messenger (m)RNA targets by sequence-specific interactions with the 3' untranslated region and are involved in the multistep process of CRC development. The objective of this study was to investigate the expression and biological roles of miR-224 in CRC. The miR-224 expression level was assessed by a quantitative real-time PCR in 79 CRC and 18 nontumor tissues. Expression levels of miR-224 in CRC tissues were significantly lower than those in nontumor tissues. Its expression level was associated with the mutation status of the APC gene. Ectopic expression of miR-224 suppressed the migratory ability of CRC cell line, but cell proliferation was less affected. Increased miR-224 diminished Cdc42 and SMAD4 expressions at both the protein and mRNA levels and inhibited the formation of actin filaments. Overall, this study indicated a role of miR-224 in negatively regulating CRC cell migration. The expression level of miR-224 may be a useful predictive biomarker for CRC progression.

Jin X, Chen Z, Xiang L, et al.
Colorectal polyp model established by transplacental BMP4 RNAi.
Mol Med Rep. 2014; 10(1):33-8 [PubMed] Article available free on PMC after 01/12/2015 Related Publications
Previous studies have shown that disruption of the bone morphogenetic protein (BMP) signaling pathway is an important cause of intestinal cancer in human and animal models. Thus, the purpose of this study was to construct a Balb/C model of colorectal polyps. Pregnant mice at 9.5 days gestation were injected via the tail vein with the pSES-Si BMP4 plasmid bearing a fluorochrome (DsRed) reporter, in order to silence the BMP4 gene in the first generation (F1); this group of mice was named the pSES-BMP4 group Intestinal fluorescence was detected at 1-, 4- and 8-week‑old F1 mice, and reverse transcription-polymerase chain reaction (RT-PCR) and western-blotting assays were used to determine changes in the expression of BMP4. A dissecting microscope and hematoxylin and eosin (H&E) staining were used to observe the cell morphology and appearance of the polyps. DsRed fluorescence was observed in the intestines of 1-week-old F1 mice of the pSES-BMP4 group. BMP4 expression at the mRNA and protein level was reduced in 1-, 4- and 8-week-old F1 mice (P<0.05). However, the level of Smad4 mRNA was only reduced in 8-week-old F1 mice (P<0.05). Multiple hyperplasic polyps emerged in the colon and rectum of the intestines of 4-week-old F1 mice in the pSES-BMP4 group. The size of colorectal polyps increased at 8 weeks, when vessels and polyp pedicles became apparent. In conclusion, silencing of the BMP4 gene using transplacental RNAi injection can induce formation of colorectal polyps in mice.

Guo W, Zhang M, Shen S, et al.
Aberrant methylation and decreased expression of the TGF-β/Smad target gene FBXO32 in esophageal squamous cell carcinoma.
Cancer. 2014; 120(16):2412-23 [PubMed] Related Publications
BACKGROUND: F-box protein 32 (FBXO32) (also known as atrogin-1), a member of the F-box protein family, has recently been identified as a transforming growth factor beta (TGF-β)/Smad target gene involved in regulating cell survival, and it may be transcriptionally silenced by epigenetic mechanisms in some kinds of carcinomas, yet its role in esophageal squamous cell carcinoma (ESCC) has not been defined.
METHODS: The role of FBXO32 in ESCC and the correlation of FBXO32 methylation with a series of pathologic parameters were studied in a large cohort of patients with ESCC.
RESULTS: Decreased messenger RNA (mRNA) expression and protein expression of FBXO32 were observed in esophageal cancer cell lines, and the silencing of FBXO32 could be reversed by treatment with 5-aza-2'-deoxycytidine or trichostatin A in the TE13 cell line. In addition, aberrant methylation of FBXO32 and histone deacetylation was capable of suppressing FBXO32 mRNA and protein expression in TE13 cells. Decreased mRNA and protein expression of FBXO32 was observed in ESCC tumor tissues and was associated with FBXO32 promoter methylation status. A positive correlation between FBXO32 and phosphorylated SMAD family members 2 and 3 expression and Smad4 protein expression also was observed in clinical specimens. FBXO32 methylation status and protein expression were independently associated with survival in patients with ESCC.
CONCLUSIONS: FBXO32 may be a functional tumor suppressor. Its inactivation through promoter methylation could play an important role in ESCC carcinogenesis, and reactivation of the FBXO32 gene may have therapeutic potential and might be used as a prognostic marker for patients with ESCC.

Laforest A, Aparicio T, Zaanan A, et al.
ERBB2 gene as a potential therapeutic target in small bowel adenocarcinoma.
Eur J Cancer. 2014; 50(10):1740-6 [PubMed] Related Publications
AIM OF THE STUDY: Small bowel adenocarcinoma (SBA) is a rare and aggressive tumour with poor outcomes. Because of its low incidence, the number prospective studies remains insufficient leading to poor knowledge and absence of standard of care. Aiming to better understand small bowel carcinogenesis we investigated the frequency of somatic mutations in a large data set of patients in more than 740 mutational hotspots among 46 genes.
METHODS: In total, 83 SBA cases were selected from two European databases. The sequencing was performed using the Ion 316 Chip. Additionally we looked into ERBB2 expression and microsatellite instability (MSI) status.
RESULTS: The tumours most frequently were duodenal (47%) and stage ⩾3 (63%). Eight genes were mutated with a frequency >5%: KRAS, TP53, APC, SMAD4, PIK3CA, ERBB2, BRAF and FBXW7. ERBB2 alterations are present in 10 patients (12%) through mutations (7 cases) or amplifications (3 cases). ERBB2 mutations were significantly associated with duodenal tumour location (P=0.04). In this group, there was a positive association with dMMR status (P=0.006) and APC mutation (P=0.02) but negative association with p53 mutations (P=0.038).
CONCLUSIONS: This study describes the first large screening of somatic mutations in SBA using next generation sequencing. The ERBB2 mutation was revealed to be one of the most frequent alterations in SBA with a distribution dependent on tumour location. In most cases ERBB2 mutation was identical (p.L755S). In clinical practice, this may suggest that more than 10% of the patients with SBA could be treated using an anti-ERBB2-targeted agent.

Bai W, Chen Y, Yang J, et al.
Aberrant miRNA profiles associated with chronic benzene poisoning.
Exp Mol Pathol. 2014; 96(3):426-30 [PubMed] Related Publications
Chronic occupational benzene exposure is associated with an increased risk of hematological malignancies. To gain an insight into the new biomarkers and molecular mechanisms of chronic benzene poisoning, miRNA profiles and mRNA expression pattern from the peripheral blood mononuclear cells of chronic benzene poisoning patients and health controls matched age and gender without benzene exposure were performed using the Exiqon miRNA PCR ARRAY and Gene Chip Human Gene 2.0ST Arrays, respectively. Totally, 6 up-regulated miRNAs (miR-34a, miR-205, miR-10b, let-7d, miR-185 and miR-423-5p-2) and 7 down-regulated miRNAs (miR-133a, miR-543, hsa-miR-130a, miR-27b,miR-223, miR-142-5p and miR-320b) were found in chronic benzene poisoning group compared to health controls (P ≤ 0.05). By integrating miRNA and mRNA expression data, these differential miRNAs were mainly involved in regulation of transcription from RNA polymerase II promoter, axon guidance, regulation of transcription, DNA-dependent, nervous system development, and regulation of actin cytoskeleton organization. Further, pathway analysis indicated that SMAD4, PLCB1, NFAT5, GNAI2, PTEN, VEGFA, BCL2, CTNNB1 and CCND1 were key target genes of differential miRNAs which were implicated in Adherens junction, TGF-beta signaling pathway, Wnt signaling pathway, tight junction and Pathways in cancer. In conclusion, the aberrant miRNAs might be a potential biomarker of chronic benzene poisoning.

Fotouhi O, Adel Fahmideh M, Kjellman M, et al.
Global hypomethylation and promoter methylation in small intestinal neuroendocrine tumors: an in vivo and in vitro study.
Epigenetics. 2014; 9(7):987-97 [PubMed] Article available free on PMC after 01/07/2015 Related Publications
Aberrant DNA methylation is a feature of human cancer affecting gene expression and tumor phenotype. Here, we quantified promoter methylation of candidate genes and global methylation in 44 small intestinal-neuroendocrine tumors (SI-NETs) from 33 patients by pyrosequencing. Findings were compared with gene expression, patient outcome and known tumor copy number alterations. Promoter methylation was observed for WIF1, RASSF1A, CTNNB1, CXCL14, NKX2-3, P16, LAMA1, and CDH1. By contrast APC, CDH3, HIC1, P14, SMAD2, and SMAD4 only had low levels of methylation. WIF1 methylation was significantly increased (P = 0.001) and WIF1 expression was reduced in SI-NETs vs. normal references (P = 0.003). WIF1, NKX2-3, and CXCL14 expression was reduced in metastases vs. primary tumors (P<0.02). Low expression of RASSF1A and P16 were associated with poor overall survival (P = 0.045 and P = 0.011, respectively). Global methylation determined by pyrosequencing of LINE1 repeats was reduced in tumors vs. normal references, and was associated with loss in chromosome 18. The tumors fell into three clusters with enrichment of WIF1 methylation and LINE1 hypomethylation in Cluster I and RASSF1A and CTNNB1 methylation and loss in 16q in Cluster II. In Cluster III, these alterations were low-abundant and NKX2-3 methylation was low. Similar analyses in the SI-NET cell lines HC45 and CNDT2 showed methylation for CDH1 and WIF1 and/or P16, CXCL14, NKX2-3, LAMA1, and CTNNB1. Treatment with the demethylating agent 5-azacytidine reduced DNA methylation and increased expression of these genes in vitro. In conclusion, promoter methylation of tumor suppressor genes is associated with suppressed gene expression and DNA copy number alterations in SI-NETs, and may be restored in vitro.

Nye MD, Almada LL, Fernandez-Barrena MG, et al.
The transcription factor GLI1 interacts with SMAD proteins to modulate transforming growth factor β-induced gene expression in a p300/CREB-binding protein-associated factor (PCAF)-dependent manner.
J Biol Chem. 2014; 289(22):15495-506 [PubMed] Article available free on PMC after 30/05/2015 Related Publications
The biological role of the transcription factor GLI1 in the regulation of tumor growth is well established; however, the molecular events modulating this phenomenon remain elusive. Here, we demonstrate a novel mechanism underlying the role of GLI1 as an effector of TGFβ signaling in the regulation of gene expression in cancer cells. TGFβ stimulates GLI1 activity in cancer cells and requires its transcriptional activity to induce BCL2 expression. Analysis of the mechanism regulating this interplay identified a new transcriptional complex including GLI1 and the TGFβ-regulated transcription factor, SMAD4. We demonstrate that SMAD4 physically interacts with GLI1 for concerted regulation of gene expression and cellular survival. Activation of the TGFβ pathway induces GLI1-SMAD4 complex binding to the BCL2 promoter whereas disruption of the complex through SMAD4 RNAi depletion impairs GLI1-mediated transcription of BCL2 and cellular survival. Further characterization demonstrated that SMAD2 and the histone acetyltransferase, PCAF, participate in this regulatory mechanism. Both proteins bind to the BCL2 promoter and are required for TGFβ- and GLI1-stimulated gene expression. Moreover, SMAD2/4 RNAi experiments showed that these factors are required for the recruitment of GLI1 to the BCL2 promoter. Finally, we determined whether this novel GLI1 transcriptional pathway could regulate other TGFβ targets. We found that two additional TGFβ-stimulated genes, INTERLEUKIN-7 and CYCLIN D1, are dependent upon the intact GLI1-SMAD-PCAF complex for transcriptional activation. Collectively, these results define a novel epigenetic mechanism that uses the transcription factor GLI1 and its associated complex as a central effector to regulate gene expression in cancer cells.

Zhang GJ, Li Y, Zhou H, et al.
miR‑20a is an independent prognostic factor in colorectal cancer and is involved in cell metastasis.
Mol Med Rep. 2014; 10(1):283-91 [PubMed] Related Publications
Accumulating evidence indicates that dysregulated microRNAs (miRNAs) are involved in cancer development, progression and metastasis. miR‑20a was found to be involved in invasion and epithelial‑mesenchymal transition (EMT) programs, with its aberrant expression having been observed in a variety of malignant tumors. However, the molecular mechanisms underlying the role of miR‑20a in colorectal cancer (CRC) development remain to be fully elucidated. In the present study, the expression of miR‑20a was compared between CRC tissue samples and the normal adjacent mucosa using quantitative polymerase chain reaction. The association of miR‑20a expression with clinicopathological characteristics was assessed using appropriate statistical analysis. The migration and invasion of SW480 cells was examined following transfection of the cells with either miR‑20a precursor or a negative control miRNA precursor. The effect of miR‑20a on the EMT in CRC cells in vitro was also analyzed. The regulatory effect of miR‑20a on SMAD family member 4 (SMAD4) was evaluated using a dual‑luciferase reporter assay. Relative expression levels of miR‑20a were significantly higher in CRC tissue than those in the normal adjacent mucosa, and high expression of miR‑20a correlated with lymph node metastases and distant metastases. Kaplan‑Meier analysis indicated that patients with increased miR‑20a levels exhibited unfavorable overall survival. Furthermore, multivariate analysis showed that miR‑20a was an independent prognostic factor. The transfection of SW480 CRC cells with miR‑20a promoted migration and invasion in vitro, and the upregulation of miR‑20a induced EMT in CRC cells. An inverse correlation between the levels of miR‑20a and SMAD4 was observed in patients with CRC. Overexpression of miR‑20a in CRC cells decreased SMAD4 expression and decreased SMAD4‑driven luciferase reporter activity. The present study revealed that miR‑20a was an independent prognostic factor in CRC. Furthermore, miR‑20a induced EMT and regulated migration and invasion of SW480 cells, at least in part via suppression of SMAD4 expression. The present study suggests that miR‑20a may serve as a novel prognostic marker and therapeutic target for CRC.

Abdel-Rahman WM, Nieminen TT, Shoman S, et al.
Loss of p15INK⁴b expression in colorectal cancer is linked to ethnic origin.
Asian Pac J Cancer Prev. 2014; 15(5):2083-7 [PubMed] Related Publications
Colorectal cancers remain to be a common cause of cancer-related death. Early-onset cases as well as those of various ethnic origins have aggressive clinical features, the basis of which requires further exploration. The aim of this work was to examine the expression patterns of p15INK4b and SMAD4 in colorectal carcinoma of different ethnic origins. Fifty-five sporadic colorectal carcinoma of Egyptian origin, 25 of which were early onset, and 54 cancers of Finnish origin were immunohistochemically stained with antibodies against p15INK4b and SMAD4 proteins. Data were compared to the methylation status of the p15INK4b gene promotor. p15INK4b was totally lost or deficient (lost in ≥ 50% of tumor cell) in 47/55 (85%) tumors of Egyptian origin as compared to 6/50 (12%) tumors of Finnish origin (p=7e-15). In the Egyptian cases with p15INK4b loss and available p15INK4b promotor methylation status, 89% of cases which lost p15INK4b expression were associated with p15INK4b gene promotor hypermethylation. SMAD4 was lost or deficient in 25/54 (46%) tumors of Egyptian origin and 28/48 (58%) tumors of Finnish origin. 22/54 (41%) Egyptian tumors showed combined loss/deficiency of both p15INK4b and SMAD4, while p15INK4b was selectively lost/deficient with positive SMAD4 expression in 24/54 (44%) tumors. Loss of p15INK4b was associated with older age at presentation (>50 years) in the Egyptian tumors (p=0.04). These data show for the first time that p15INK4b loss of expression marks a subset of colorectal cancers and ethnic origin may play a role in this selection. In a substantial number of cases, the loss was independent of SMAD4 but rather associated with p15INK4b gene promotor hypermethylation and old age which could be related to different environmental exposures.

Yamazaki K, Masugi Y, Effendi K, et al.
Upregulated SMAD3 promotes epithelial-mesenchymal transition and predicts poor prognosis in pancreatic ductal adenocarcinoma.
Lab Invest. 2014; 94(6):683-91 [PubMed] Related Publications
In pancreatic ductal adenocarcinoma (PDAC), features of epithelial-mesenchymal transition (EMT) are often seen in tumor tissue, and such features correlate with poor prognosis. Solitary infiltration of tumor cells represents a morphological phenotype of EMT, and we previously reported that a high degree of solitary cell infiltration correlates with EMT-like features, including reduced E-cadherin and elevated vimentin levels. Using solitary cell infiltration to evaluate the degree of EMT, gene-expression profiling of 12 PDAC xenografts was performed, and SMAD3 was identified as an EMT-related gene. Immunohistochemistry using clinical specimens (n=113) showed that SMAD3 accumulated in the nuclei of tumor cells, but was not detected in most epithelial cells in the pancreatic duct. Moreover, SMAD3 upregulation correlated with malignant characteristics, such as higher tumor grade and lymph node metastasis, as well as with EMT-like features. SMAD4, which plays a key role in transforming growth factor-β (TGF-β) signaling, is inactivated in approximately half of PDAC cases. In this study, the nuclear accumulation of SMAD3 was immunohistochemically detected even in SMAD4-negative cases. SMAD3 knockdown resulted in upregulated E-cadherin, downregulated vimentin, and reduced cell motility in pancreatic cancer cells regardless of SMAD4 status. In addition, TGF-β-treatment resulted in EMT induction in cells carrying wild-type SMAD4, and EMT was suppressed by SMAD3 knockdown. Patients with upregulated SMAD3 and a high degree of solitary cell infiltration had shorter times to recurrence and shorter survival times after surgery, and multivariate analysis showed that both factors were independent prognostic factors linked to unfavorable outcomes. These findings suggest that SMAD3 in PDAC is involved in the promotion of malignant potential through EMT induction in tumor cells regardless of SMAD4 status and serves as a potential biomarker of poor prognosis.

Huhn S, Bevier M, Pardini B, et al.
Colorectal cancer risk and patients' survival: influence of polymorphisms in genes somatically mutated in colorectal tumors.
Cancer Causes Control. 2014; 25(6):759-69 [PubMed] Related Publications
PURPOSE: The first two studies aiming for the high-throughput identification of the somatic mutation spectrum of colorectal cancer (CRC) tumors were published in 2006 and 2007. Using exome sequencing, they described 69 and 140 candidate cancer genes (CAN genes), respectively. We hypothesized that germline variants in these genes may influence CRC risk, similar to APC, which is causing CRC through germline and somatic mutations.
METHODS: After excluding the well-established CRC genes APC, KRAS, TP53, and ABCA1, we analyzed 35 potentially functional single-nucleotide polymorphisms (SNPs) in 10 CAN genes (OBSCN, MLL3, PKHD1, SYNE1, ERCC6, FBXW7, EPHB6/TRPV6, ELAC1/SMAD4, EPHA3, and ADAMTSL3) using KBiosciences Competitive Allele-Specific PCR™ genotyping assays. In addition to CRC risk (1,399 CRC cases, 838 controls), we also considered the influence of the SNPs on patients' survival (406 cases).
RESULTS: In spite of the fact that our in silico analyses suggested functional relevance for the studied genes and SNPs, our data did not support a strong influence of the studied germline variants on CRC risk and survival. The strongest association with CRC risk and survival was found for MLL3 (rs6464211, OR 1.50, p = 0.002, dominant model; HR 2.12, p = 0.020, recessive model). Two SNPs in EPHB6/TRPV6 (dominant model) showed marginal associations with survival (rs4987622 HR 0.58 p = 0.028 and rs6947538 HR 0.64, p = 0.036, respectively).
CONCLUSION: Although somatic mutations in the CAN genes have been related to the development and progression of various types of cancers in several next-generation sequencing or expression analyses, our study suggests that the studied potentially functional germline variants are not likely to affect CRC risk or survival.

Zizi-Sermpetzoglou A, Myoteri D, Arkoumani E, et al.
A study of Smad4 and Smad7 expression in surgically resected samples of gastric adenocarcinoma and their correlation with clinicopathological parameters and patient survival.
J BUON. 2014 Jan-Mar; 19(1):221-7 [PubMed] Related Publications
PURPOSE: The canonical signaling pathway for the transforming growth factor-beta (TGF-β) family is through the Smad proteins which are pivotal intracellular mediators of TGF-β family members. Recently, disruption of the TGF-β pathway in cancer has been demonstrated at the level of the Smad signal transducers. In this study, we examined Smad4 and Smad7 expression in gastric carcinomas to elucidate their role in tumor progression.
METHODS: The immunohistochemical expression of Smad4 and Smad7 was evaluated in 151 surgically resected samples of gastric adenocarcinoma in order to examine their correlation with clinicopathologic findings and patients' survival.
RESULTS: Smad4 and Smad7 expression (low, moderate or strong) was observed in 86.7% (131/151) and 33.1% (50/151) of gastric adenocarcinoma tumor samples, respectively. Our results revealed that the loss of Smad4 expression correlated significantly with the intestinal type, male sex, depth of tumor and poor survival. Smad7 expression was significantly more frequent in intestinal type and well differentiated gastric adenocarcinomas and significantly correlated with the duration of disease-free survival.
CONCLUSION: Smad signal transducers are considered as important molecules in tumor development and progression and the evaluation of their expression in human gastric cancer could be useful in selecting stage I patients who should be considered as candidates for adjuvant chemotherapy.

Uitdehaag JC, de Roos JA, van Doornmalen AM, et al.
Comparison of the cancer gene targeting and biochemical selectivities of all targeted kinase inhibitors approved for clinical use.
PLoS One. 2014; 9(3):e92146 [PubMed] Article available free on PMC after 30/05/2015 Related Publications
The anti-proliferative activities of all twenty-five targeted kinase inhibitor drugs that are in clinical use were measured in two large assay panels: (1) a panel of proliferation assays of forty-four human cancer cell lines from diverse tumour tissue origins; and (2) a panel of more than 300 kinase enzyme activity assays. This study provides a head-on comparison of all kinase inhibitor drugs in use (status Nov. 2013), and for six of these drugs, the first kinome profiling data in the public domain. Correlation of drug activities with cancer gene mutations revealed novel drug sensitivity markers, suggesting that cancers dependent on mutant CTNNB1 will respond to trametinib and other MEK inhibitors, and cancers dependent on SMAD4 to small molecule EGFR inhibitor drugs. Comparison of cellular targeting efficacies reveals the most targeted inhibitors for EGFR, ABL1 and BRAF(V600E)-driven cell growth, and demonstrates that the best targeted agents combine high biochemical potency with good selectivity. For ABL1 inhibitors, we computationally deduce optimized kinase profiles for use in a next generation of drugs. Our study shows the power of combining biochemical and cellular profiling data in the evaluation of kinase inhibitor drug action.

Layfield LJ, Ehya H, Filie AC, et al.
Utilization of ancillary studies in the cytologic diagnosis of biliary and pancreatic lesions: the Papanicolaou Society of Cytopathology guidelines for pancreatobiliary cytology.
Diagn Cytopathol. 2014; 42(4):351-62 [PubMed] Article available free on PMC after 30/05/2015 Related Publications
The Papanicolaou Society of Cytopathology has developed a set of guidelines for pancreatobiliary cytology including indications for endoscopic ultrasound-guided fine-needle aspiration, terminology and nomenclature of pancreatobiliary disease, ancillary testing, and post-biopsy management. All documents are based on the expertise of the authors, a review of the literature, discussions of the draft document at several national and international meetings, and synthesis of selected online comments of the draft document. This document presents the results of these discussions regarding the use of ancillary testing in the cytologic diagnosis of biliary and pancreatic lesions. Currently, fluorescence in situ hybridization (FISH) appears to be the most clinically relevant ancillary technique for cytology of bile duct strictures. The addition of FISH analysis to routine cytologic evaluation appears to yield the highest sensitivity without loss in specificity. Loss of immunohistochemical staining for the protein product of the SMAD4 gene and positive staining for mesothelin support a diagnosis of ductal adenocarcinoma. Immunohistochemical markers for endocrine and exocrine differentiation are sufficient for a diagnosis of endocrine and acinar tumors. Nuclear staining for beta-catenin supports a diagnosis of solid-pseudopapilary neoplasm. Cyst fluid analysis for amylase and carcinoembryonic antigen aids in the preoperative classification of pancreatic cysts. Many gene mutations (KRAS, GNAS, VHL, RNF43, and CTNNB1) may be of aid in the diagnosis of cystic neoplasms. Other ancillary techniques do not appear to improve diagnostic sensitivity sufficiently to justify their increased costs.

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