Effect of vitamin E δ-tocotrienol and aspirin on Wnt signaling in human colon cancer stem cells and in adenoma development in APCmin/+ mice.

In this study, we evaluated the effects of vitamin E δ-tocotrienol (DT3) and aspirin on Wnt signaling in human colon cancer stem cells (CCSCs) and in the prevention of adenoma formation in APCmin/+ mice. We found that knockdown of the adenomatous polyposis coli (APC) gene led to subsequent activation of Wnt signaling in colon epithelial cells (NCM460-APCsiRNA) and induction of ß-catenin and its downstream target proteins c-MYC, cyclin D1, and survivin. When aspirin and DT3 were combined, cell growth and survival were inhibited and apoptosis was induced in colon epithelial cells and in CCSCs. However, DT3 and/or aspirin had little or no effect on control normal colon epithelial cells (NCM460-NCsiRNA). The induction of apoptosis was directly related to activation of caspase 8 and cleavage of BID to truncated BID. In addition, DT3 and/or aspirin-induced apoptosis was associated with cleaved PARP, elevated levels of cytosolic cytochrome c and BAX, and depletion of anti-apoptotic protein BCl-2 in CCSCs. The combination of aspirin and DT3 inhibited the self-renewal capacity, Wnt/ß-catenin receptor activity, and expression of ß-catenin and its downstream targets c-MYC, cyclin D1 and survivin in CCSCs. We also found that treatment with DT3 alone or combined with aspirin significantly inhibited intestinal adenoma formation and Wnt/ ß-catenin signaling and induced apoptosis, compared to vehicle, in APCmin/+ mice. Our study demonstrated a rationale for further investigation of the combination of DT3 and aspirin for colorectal cancer prevention and therapy.

A microRNA Signature Identifies Patients at Risk of Barrett Esophagus Progression to Dysplasia and Cancer.

BACKGROUND: Progression of Barrett esophagus (BE) to esophageal adenocarcinoma occurs among a minority of BE patients. To date, BE behavior cannot be predicted on the basis of histologic features. AIMS: We compared BE samples that did not develop dysplasia or carcinoma upon follow-up of ≥ 7 years (BE nonprogressed [BEN]) with BE samples that developed carcinoma upon follow-up of 3 to 4 years (BE progressed [BEP]). METHODS: The NanoString nCounter miRNA assay was used to profile 24 biopsy samples of BE, including 13 BENs and 11 BEPs. Fifteen samples were randomly selected for miRNA prediction model training; nine were randomly selected for miRNA validation. RESULTS: Unpaired t tests with Welch's correction were performed on 800 measured miRNAs to identify the most differentially expressed miRNAs for cases of BEN and BEP. The top 12 miRNAs (P < .003) were selected for principal component analyses: miR-1278, miR-1301, miR-1304-5p, miR-517b-3p, miR-584-5p, miR-599, miR-103a-3p, miR-1197, miR-1256, miR-509-3-5p, miR-544b, miR-802. The 12-miRNA signature was first self-validated on the training dataset, resulting in 7 out of the 7 BEP samples being classified as BEP (100% sensitivity) and 7 out of the 8 BEN samples being classified as BEN (87.5% specificity). Upon validation, 4 out of the 4 BEP samples were classified as BEP (100% sensitivity) and 4 out of the 5 BEN samples were classified as BEN (80% specificity). Twenty-four samples were evaluated, and 22 cases were correctly classified. Overall accuracy was 91.67%. CONCLUSION: Using miRNA profiling, we have identified a 12-miRNA signature able to reliably differentiate cases of BEN from BEP.

LncRNA PAINT is associated with aggressive prostate cancer and dysregulation of cancer hallmark genes.

Long noncoding RNAs (lncRNAs) play regulatory role in cellular processes and their aberrant expression may drive cancer progression. Here we report the function of a lncRNA PAINT (prostate cancer associated intergenic noncoding transcript) in promoting prostate cancer (PCa) progression. Upregulation of PAINT was noted in advanced stage and metastatic PCa. Inhibition of PAINT decreased cell proliferation, S-phase progression, increased expression of apoptotic markers, and improved sensitivity to docetaxel and Aurora kinase inhibitor VX-680. Inhibition of PAINT decreased cell migration and reduced expression of Slug and Vimentin. Ectopic expression of PAINT suppressed E-cadherin, increased S-phase progression and cell migration. PAINT expression in PCa cells induced larger colony formation, increased tumor growth and higher expression of mesenchymal markers. Transcriptome analysis followed by qRT-PCR validation showed differentially expressed genes involved in epithelial mesenchymal transition (EMT), apoptosis and drug resistance in PAINT-expressing cells. Our study establishes an oncogenic function of PAINT in PCa.

Diphenylbutylpiperidine Antipsychotic Drugs Inhibit Prolactin Receptor Signaling to Reduce Growth of Pancreatic Ductal Adenocarcinoma in Mice.

BACKGROUND & AIMS: Prolactin (PRL) signaling is up-regulated in hormone-responsive cancers. The PRL receptor (PRLR) is a class I cytokine receptor that signals via the Janus kinase (JAK)-signal transducer and activator of transcription and mitogen-activated protein kinase pathways to regulate cell proliferation, migration, stem cell features, and apoptosis. Patients with pancreatic ductal adenocarcinoma (PDAC) have high plasma levels of PRL. We investigated whether PRLR signaling contributes to the growth of pancreatic tumors in mice. METHODS: We used immunohistochemical analyses to compare levels of PRL and PRLR in multitumor tissue microarrays. We used structure-based virtual screening and fragment-based drug discovery to identify compounds likely to bind PRLR and interfere with its signaling. Human pancreatic cell lines (AsPC-1, BxPC-3, Panc-1, and MiaPaCa-2), with or without knockdown of PRLR (clustered regularly interspaced short palindromic repeats or small hairpin RNA), were incubated with PRL or penfluridol and analyzed in proliferation and spheroid formation. C57BL/6 mice were given injections of UNKC-6141 cells, with or without knockdown of PRLR, into pancreas, and tumor development was monitored for 4 weeks, with some mice receiving penfluridol treatment for 21 days. Human pancreatic tumor tissues were implanted into interscapular fat pads of NSG mice, and mice were given injections of penfluridol daily for 28 days. Nude mice were given injections of Panc-1 cells, xenograft tumors were grown for 2 weeks, and mice were then given intraperitoneal penfluridol for 35 days. Tumors were collected from mice and analyzed by histology, immunohistochemistry, and immunoblots. RESULTS: Levels of PRLR were increased in PDAC compared with nontumor pancreatic tissues. Incubation of pancreatic cell lines with PRL activated signaling via JAK2-signal transducer and activator of transcription 3 and extracellular signal-regulated kinase, as well as formation of pancospheres and cell migration; these activities were not observed in cells with PRLR knockdown. Pancreatic cancer cells with PRLR knockdown formed significantly smaller tumors in mice. We identified several diphenylbutylpiperidine-class antipsychotic drugs as agents that decreased PRL-induced JAK2 signaling; incubation of pancreatic cancer cells with these compounds reduced their proliferation and formation of panco spheres. Injections of 1 of these compounds, penfluridol, slowed the growth of xenograft tumors in the different mouse models, reducing proliferation and inducing autophagy of the tumor cells. CONCLUSIONS: Levels of PRLR are increased in PDAC, and exposure to PRL increases proliferation and migration of pancreatic cancer cells. Antipsychotic drugs, such as penfluridol, block PRL signaling in pancreatic cancer cells to reduce their proliferation, induce autophagy, and slow the growth of xenograft tumors in mice. These drugs might be tested in patients with PDAC.

CDX-2 Expression in Esophageal Biopsies Without Goblet Cell Intestinal Metaplasia May Be Predictive of Barrett's Esophagus.

BACKGROUND: CDX-2 is a nuclear homeobox transcription factor not normally expressed in esophageal and gastric epithelia, reported to highlight intestinal metaplasia (IM) in the esophagus. Pathological absence of goblet cells at initial screening via hematoxylin and eosin (HE) and alcian blue (AB) staining results in patient exclusion from surveillance programs. AIMS: This study aimed to determine whether non-goblet cell IM, as defined by CDX-2 positivity, can be considered to be a precursor to Barrett's esophagus (BE). METHODS: This study received IRB approval (17,284). Patients with gastroesophageal reflux disease (n = 181) who underwent upper-gastrointestinal endoscopy with biopsies of the distal esophagus to rule out BE using HE/AB staining and CDX-2 immunostaining were followed for 3 years. Initial and follow-up staining results were evaluated for age/sex. RESULTS: Differences between development of goblet cell IM in CDX-2-negative and CDX-2-positive groups were evaluated. A Kaplan-Meier curve showed that, out of the 134 patients initially positive for CDX-2, 25 (18.7%) had developed goblet cell IM after 2 years and 106 (79.1%) after 3 years. Conversely, of the 47 patients initially negative for CDX-2, 8 (17.9%) developed goblet cell IM after 24 months and only 11 (23.8%) after 40 to 45 months (P = .049; age-adjusted Cox proportional hazard regression model). CONCLUSION: In cases that are initially AB negative and CDX-2 positive, CDX-2 was demonstrated to have a potential prognostic utility for early detection of progression to BE. CDX-2 expression is significantly predictive for risk of goblet cell IM development 40 to 45 months after initial biopsy.

Clinicopathologic analysis of primary gastroenteropancreatic poorly differentiated neuroendocrine carcinoma; A ten year retrospective study of 68 cases at Moffit Cancer Center.

OBJECTIVE: To assess clinicopathological characteristics of primary gastro-entero-pancreatic poorly differentiated neuroendocrine carcinomas (GEP-PDNECAs) and evaluate overall survival in patients treated with systemic platinum and etoposide therapy. METHODS: A detailed retrospective review of clinico-pathologic data (1999-2009) on 68 consecutive adult patients with primary GEP-PDNECAs was carried out, from H Lee Moffit Cancer Center and Research Institute, Tampa, Florida; USA, based on electronic patient records, specialty consultation files, tumor registry, social security index and pathology archives. All available tumor slides were reviewed and subtyped by neuro-endocrine pathologists. Clinicopathologic data and patient survival were analyzed. RESULTS: Of 68 patients 41 were males and 27 females with a mean age of 42 years (range: 25-76 years). Regarding the site of origin, 39 patients were of the colorectal location, 19 from the pancreas, 04 from small intestines, 03 from stomach and 03 were multi-focal from colon, small intestine and pancreas. Sixty three of 68 (93%) patients presented with lymph node/distant metastases. Of 68 tumors 37 (54%) were classified as small cell carcinoma (SCCA), 16 (24%) large cell carcinoma (LCCA), 5 (7%) mixed small and large cell (MSLCCA) and 10 (15%) poorly differentiated carcinoma with neuroendocrine features (PDCA-NEF). Neuroendocrine differentiation was confirmed by positivity for chromogranin in 38/65 (55%), synaptophysin in 62/67 (92%) and CD56 in 17/21 (81%) cases. One neuroendocrine marker was positive in 22/68 (32%), 2 in 40/68 (59%) and all 3 were positive in 9/68 (13%) cases. Fifty-eight of 68 (85%) patients were treated with platinum and etoposide. Overall patient survival at 1, 3 5 and 10 years was 85%, 40%, 16% and 1.5% respectively. Patient survival was independent of age (r= 0.1022), sex (r= -0.909) and histologic tumor subtype (r=0.1028) (p= 0.128) but was related to distant metastases (r=0.306; p=0.0383). CONCLUSIONS: GEP-PDNECA occurred in many part of the GI tract, most commonly in the colorectal region. Positivity of neuroendocrine markers was variable, which helped to confirm neuro-endocrine differentiation and to avoid under-diagnosis of GEP-PDNECA, especially in metastatic setting. Overall prognosis of GEP-PDNECA patients following platinum and etoposide therapy in our series was relatively favorable but remained poor in the presence of distant metastases.

Vitamin E δ-tocotrienol sensitizes human pancreatic cancer cells to TRAIL-induced apoptosis through proteasome-mediated down-regulation of c-FLIPs.

BACKGROUND: Vitamin E δ-tocotrienol (VEDT), a vitamin E compound isolated from sources such as palm fruit and annatto beans, has been reported to have cancer chemopreventive and therapeutic effects. METHODS: We report a novel function of VEDT in augmenting tumor necrosis factor-related apoptosis-inducing ligand- (TRAIL-) induced apoptosis in pancreatic cancer cells. The effects of VEDT were shown by its ability to trigger caspase-8-dependent apoptosis in pancreatic cancer cells. RESULTS: When combined with TRAIL, VEDT significantly augmented TRAIL-induced apoptosis of pancreatic cancer cells. VEDT decreased cellular FLICE inhibitory protein (c-FLIP) levels without consistently modulating the expression of decoy death receptors 1, 2, 3 or death receptors 4 and 5. Enforced expression of c-FLIP substantially attenuated VEDT/TRAIL-induced apoptosis. Thus, c-FLIP reduction plays an important part in mediating VEDT/TRAIL-induced apoptosis. Moreover, VEDT increased c-FLIP ubiquitination and degradation but did not affect its transcription, suggesting that VEDT decreases c-FLIP levels through promoting its degradation. Of note, degradation of c-FLIP and enhanced TRAIL-induced apoptosis in pancreatic cancer cells were observed only with the anticancer bioactive vitamin E compounds δ-, γ-, and ß-tocotrienol but not with the anticancer inactive vitamin E compounds α-tocotrienol and α-, ß-, γ-, and δ-tocopherol. CONCLUSIONS: c-FLIP degradation is a key event for death receptor-induced apoptosis by anticancer bioactive vitamin E compounds in pancreatic cancer cells. Moreover, VEDT augmented TRAIL inhibition of pancreatic tumor growth and induction of apoptosis in vivo. Combination therapy with TRAIL agonists and bioactive vitamin E compounds may offer a novel strategy for pancreatic cancer intervention.

Protein and glycomic plasma markers for early detection of adenoma and colon cancer.

OBJECTIVE: To discover and confirm blood-based colon cancer early-detection markers. DESIGN: We created a high-density antibody microarray to detect differences in protein levels in plasma from individuals diagnosed with colon cancer <3 years after blood was drawn (ie, prediagnostic) and cancer-free, matched controls. Potential markers were tested on plasma samples from people diagnosed with adenoma or cancer, compared with controls. Components of an optimal 5-marker panel were tested via immunoblotting using a third sample set, Luminex assay in a large fourth sample set and immunohistochemistry (IHC) on tissue microarrays. RESULTS: In the prediagnostic samples, we found 78 significantly (t-test) increased proteins, 32 of which were confirmed in the diagnostic samples. From these 32, optimal 4-marker panels of BAG family molecular chaperone regulator 4 (BAG4), interleukin-6 receptor subunit beta (IL6ST), von Willebrand factor (VWF) and CD44 or epidermal growth factor receptor (EGFR) were established. Each panel member and the panels also showed increases in the diagnostic adenoma and cancer samples in independent third and fourth sample sets via immunoblot and Luminex, respectively. IHC results showed increased levels of BAG4, IL6ST and CD44 in adenoma and cancer tissues. Inclusion of EGFR and CD44 sialyl Lewis-A and Lewis-X content increased the panel performance. The protein/glycoprotein panel was statistically significantly higher in colon cancer samples, characterised by a range of area under the curves from 0.90 (95% CI 0.82 to 0.98) to 0.86 (95% CI 0.83 to 0.88), for the larger second and fourth sets, respectively. CONCLUSIONS: A panel including BAG4, IL6ST, VWF, EGFR and CD44 protein/glycomics performed well for detection of early stages of colon cancer and should be further examined in larger studies.

Expression of FGD4 positively correlates with the aggressive phenotype of prostate cancer.

BACKGROUND: FGD4 (Frabin) is an F-actin binding protein with GTP/GDP exchange activity specific for CDC42. It is involved in reorganization of the actin cytoskeleton, which requires both actin binding and CDC42 activating function of FGD4. Expression of FGD4 is altered in patients with heterogeneous hereditary motor and sensory neuropathies as a result of demyelination of peripheral nerves. METHODS: In this study, we examined the expression of FGD4 in prostate cancer specimens using immunohistochemistry and studied the function of FGD4 in maintaining cell phenotype, behavior and drug sensitivity using overexpression and siRNA-based silencing approaches. We used Mann-Whitney test for comparative analysis of FGD4 expression. RESULTS: Our results show that the expression of FGD4 is upregulated in cancerous prostates compared to the luminal cells in benign prostatic hyperplasia, although the basal cells showed high staining intensities. We noted a gradual increase in the staining intensity of FGD4 with increasing aggressiveness of the disease. Inhibition of expression of FGD4 using siRNAs showed reduced proliferation and cell cycle arrest in G2/M phase of androgen dependent LNCaP-104S and androgen refractory PC-3 cells. Inhibition of FGD4 also resulted in reduced cell migration and CDC42 activities in PC-3 cells whereas, ectopic expression of FGD4 induced cell migration, altered expression of mesenchymal and epithelial markers and activation of CDC42/PAK signaling pathway. Reduced expression of FGD4 improved sensitivity of LNCaP-104S cells to the anti-androgen drug Casodex and PC-3 cells to the microtubule stabilizing drug docetaxel. CONCLUSIONS: Our data demonstrate a tumor promoting and a cell migratory function of FGD4 in prostate cancer cells and that inhibition of FGD4 expression enhances the response for both androgen-dependent and independent prostate cancer cells towards currently used prostate cancer drugs.

Toward early detection of Helicobacter pylori-associated gastric cancer.

BACKGROUND: Gastric cancer is typically diagnosed at a late stage, leading to poor prognoses. Helicobacter pylori is responsible for 70% of gastric cancers globally, and patients with this bacterial infection often present with early stages of the carcinogenic pathway such as inflammation or gastritis. Although many patients continue to progress to advanced-stage disease after antibacterial treatment, there are no follow-up screening protocols for patients with a history of H. pylori. METHODS: Several biomarkers (Lgr5, CD133, CD44) become upregulated during gastric carcinogenesis. A logistic regression model is developed using clinical data from 59 patients at different stages of the carcinogenic pathway to identify the likelihood of being at an advanced stage of disease for all combinations of age, sex, and marker positivity. Using these likelihood distributions and the observed rate of marker positivity increase, time to high likelihood (probability >0.8) of advanced disease for individual patients is predicted. RESULTS: A strong correlation between marker positivity and disease stage was found for all three markers. Disease stage was accurately classified by the respective regression models for more than 86% of retrospective patients. Highly patient-specific predictions of time to onset of dysplasia were made, allowing the classification of 17 patients initially diagnosed with intestinal metaplasia into high-, intermediate-, or low-risk categories. CONCLUSIONS: We present an approach designed to integrate pathology, mathematics, and statistics for detection of the earliest precancerous, treatable lesion. Given the simplicity and robustness of the framework, such technique has the potential to guide personalized screening schedules to minimize the risk of undetected malignant transformation.

Knockdown of CSE1L Gene in Colorectal Cancer Reduces Tumorigenesis in Vitro.

Human cellular apoptosis susceptibility (chromosomal segregation 1-like, CSE1L) gene plays a role in nuclear-to-cytoplasm transport and chromosome segregation during mitosis, cellular proliferation, and apoptosis. CSE1L is involved in colon carcinogenesis. CSE1L gene expression was assessed with three data sets using Affymetrix U133 + gene chips on normal human colonic mucosa (NR), adenomas (ADs), and colorectal carcinoma (CRC). CSE1L protein expression in CRC, AD, and NR from the same patients was measured by immunohistochemistry using a tissue microarray. We evaluated CSE1L expression in CRC cells (HCT116, SW480, and HT29) and its biological functions. CSE1L mRNA was significantly increased in all AD and CRC compared with NR (P < 0.001 and P = 0.02, respectivly). We observed a change in CSE1L staining intensity and cellular localization by immunohistochemistry. CSE1L was significantly increased during the transition from AD to CRC when compared with NR in a CRC tissue microarray (P = 0.01 and P < 0.001). HCT116, SW480, and HT29 cells also expressed CSE1L protein. CSE1L knockdown by shRNA inhibited protein, resulting in decreased cell proliferation, reduced colony formation in soft agar, and induction of apoptosis. CSE1L protein is expressed early and across all stages of CRC development. shRNA knockdown of CSE1L was associated with inhibition of tumorigenesis in CRC cells. CSE1L may represent a potential target for treatment of CRC.

The human, F-actin-based cytoskeleton as a mutagen sensor.

BACKGROUND: Forty years ago the actin cytoskeleton was determined to be disrupted in fibroblasts from persons with DNA repair-defective, hereditary colon cancer, with no clear connection between the cytoskeleton and DNA repair defects at that time. Recently, the large number of sequenced genomes has indicated that mammalian mutagenesis has a large stochastic component. As a result, large coding regions are large mutagen targets. Cytoskeletal protein-related coding regions (CPCRs), including extra-cellular matrix proteins, are among the largest coding regions in the genome and are indeed very commonly mutated in cancer. METHODS: To determine whether mutagen sensitivity of the actin cytoskeleton could be assessed experimentally, we treated tissue culture cells with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and quantified overall cytoskeleton integrity with rhodamine-phalloidin stains for F-actin. RESULTS: The above approach indicated cytoskeletal degradation with increasing mutagen exposure, consistent with increased mutagenesis of CPCRs in TCGA, smoker samples, where overall mutation rates correlate with CPCR mutation rates (R2 = 0.8694; p < 0.00001). In addition, mutagen exposure correlated with a decreasing cell perimeter to area ratio, raising questions about potential decreasing, intracellular diffusion and concentrations of chemotherapy drugs, with increasing mutagenesis and decreasing cytoskeleton integrity. CONCLUSION: Determination of cytoskeletal integrity may provide the opportunity to assess mutation burdens in nonclonal cell populations, such as in intact tissues, where DNA sequencing for heterogeneous mutation burdens can be challenging.

TGF-ß-inducible microRNA-183 silences tumor-associated natural killer cells.

Transforming growth factor ß1 (TGF-ß), enriched in the tumor microenvironment and broadly immunosuppressive, inhibits natural killer (NK) cell function by yet-unknown mechanisms. Here we show that TGF-ß-treated human NK cells exhibit reduced tumor cytolysis and abrogated perforin polarization to the immune synapse. This result was accompanied by loss of surface expression of activating killer Ig-like receptor 2DS4 and NKp44, despite intact cytoplasmic stores of these receptors. Instead, TGF-ß depleted DNAX activating protein 12 kDa (DAP12), which is critical for surface NK receptor stabilization and downstream signal transduction. Mechanistic analysis revealed that TGF-ß induced microRNA (miR)-183 to repress DAP12 transcription/translation. This pathway was confirmed with luciferase reporter constructs bearing the DAP12 3' untranslated region as well as in human NK cells by use of sense and antisense miR-183. Moreover, we documented reduced DAP12 expression in tumor-associated NK cells in lung cancer patients, illustrating this pathway to be consistently perturbed in the human tumor microenvironment.

Histone deacetylase 10 regulates DNA mismatch repair and may involve the deacetylation of MutS homolog 2.

MutS homolog 2 (MSH2) is an essential DNA mismatch repair (MMR) protein. It interacts with MSH6 or MSH3 to form the MutSα or MutSß complex, respectively, which recognize base-base mispairs and insertions/deletions and initiate the repair process. Mutation or dysregulation of MSH2 causes genomic instability that can lead to cancer. MSH2 is acetylated at its C terminus, and histone deacetylase (HDAC6) deacetylates MSH2. However, whether other regions of MSH2 can be acetylated and whether other histone deacetylases (HDACs) and histone acetyltransferases (HATs) are involved in MSH2 deacetylation/acetylation is unknown. Here, we report that MSH2 can be acetylated at Lys-73 near the N terminus. Lys-73 is highly conserved across many species. Although several Class I and II HDACs interact with MSH2, HDAC10 is the major enzyme that deacetylates MSH2 at Lys-73. Histone acetyltransferase HBO1 might acetylate this residue. HDAC10 overexpression in HeLa cells stimulates cellular DNA MMR activity, whereas HDAC10 knockdown decreases DNA MMR activity. Thus, our study identifies an HDAC10-mediated regulatory mechanism controlling the DNA mismatch repair function of MSH2.

Long non-coding RNAs (LncRNA) regulated by transforming growth factor (TGF) ß: LncRNA-hit-mediated TGFß-induced epithelial to mesenchymal transition in mammary epithelia.

Long noncoding RNAs (lncRNAs) are emerging as key regulators in various biological processes. Epithelial-to-mesenchymal transition (EMT) is a developmental process hijacked by tumor cells to depart from the primary tumor site, invade surrounding tissue, and establish distant metastases. Transforming growth factor ß (TGFß) signaling has been shown to be a major inducer of EMT and to facilitate breast cancer metastasis. However, the role of lncRNAs in this process remains largely unknown. Here we report a genome-wide lncRNA profile in mouse mammary epithelial NMuMG cells upon TGFß induction of EMT. Among 10,802 lncRNAs profiled, over 600 were up-regulated and down-regulated during the EMT, respectively. Furthermore, we identify that lncRNA-HIT (HOXA transcript induced by TGFß) mediates TGFß function, i.e. depletion of lncRNA-HIT inhibits TGFß-induced migration, invasion, and EMT in NMuMG. LncRNA-HIT is also significantly elevated in the highly metastatic 4T1 cells. Knockdown of lncRNA-HIT in 4T1 results in decrease of cell migration, invasion, tumor growth, and metastasis. E-cadherin was identified as a major target of lncRNA-HIT. Moreover, lncRNA-HIT is conserved in humans and elevated expression associates with more invasive human primary breast carcinoma. Collectively, these data suggest that a subset of lncRNAs such as lncRNA-HIT play a significant role in regulation of EMT and breast cancer invasion and metastasis, and could be potential therapeutic targets in breast cancers.

YAP1 Regulates OCT4 Activity and SOX2 Expression to Facilitate Self-Renewal and Vascular Mimicry of Stem-Like Cells.

Non-small cell lung cancer (NSCLC) is highly correlated with smoking and has very low survival rates. Multiple studies have shown that stem-like cells contribute to the genesis and progression of NSCLC. Our results show that the transcriptional coactivator yes-associated protein 1 (YAP1), which is the oncogenic component of the Hippo signaling pathway, is elevated in the stem-like cells from NSCLC and contributes to their self-renewal and ability to form angiogenic tubules. Inhibition of YAP1 by a small molecule or depletion of YAP1 by siRNAs suppressed self-renewal and vascular mimicry of stem-like cells. These effects of YAP1 were mediated through the embryonic stem cell transcription factor, Sox2. YAP1 could transcriptionally induce Sox2 through a physical interaction with Oct4; Sox2 induction occurred independent of TEAD2 transcription factor, which is the predominant mediator of YAP1 functions. The binding of Oct4 to YAP1 could be detected in cell lines as well as tumor tissues; the interaction was elevated in NSCLC samples compared to normal tissue as seen by proximity ligation assays. YAP1 bound to Oct4 through the WW domain, and a peptide corresponding to this region could disrupt the interaction. Delivery of the WW domain peptide to stem-like cells disrupted the interaction and abrogated Sox2 expression, self-renewal, and vascular mimicry. Depleting YAP1 reduced the expression of multiple epithelial-mesenchymal transition genes and prevented the growth and metastasis of tumor xenografts in mice; overexpression of Sox2 in YAP1 null cells rescued these functions. These results demonstrate a novel regulation of stem-like functions by YAP1, through the modulation of Sox2 expression.

Pathology Report: Presacral Noncommunicating Enteric Duplication Cyst.

BACKGROUND: Gastrointestinal (GI) tract duplication cysts or enteric duplication cysts are rare congenital malformations sometimes found on the mesenteric aspect of segments of the alimentary tract. Enteric duplication cysts are lined by normal GI epithelium and may be classified as foregut, mid-gut, and hindgut cysts. Except in very rare cases of retroperitoneal enteric duplication cysts, these cysts communicate with the GI tract and share a common blood supply. Concurrent congenital malformations are not uncommon and malignant transformation within enteric duplication cysts has also been reported. METHODS: We describe a case of a noncommunicating enteric duplication cyst in a 52-year-old woman. RESULTS: The patient presented with a presacral cystic mass requiring frequent drainage procedures that was primarily believed to be of neural origin. Upon resection, the lesion contained heterotopic tissue, including ciliated bronchial epithelium, squamous and transitional epithelia, and pancreatic and gastric tissue. Focal, low-grade intestinal adenoma was present, but malignancy was not detected in this case. CONCLUSION: To our knowledge, this is the sixth reported case of a noncommunicating enteric duplication cyst in the English medical literature.

Systematic Review and Case Series Report of Acinar Cell Carcinoma of the Pancreas.

BACKGROUND: Acinar cell carcinoma of the pancreas is a rare malignancy representing less than 1% of all pancreatic malignancies. METHODS: We report on a case series of 21 patients with acinar cell carcinoma of the pancreas treated at a high-volume quaternary center. A systematic review of the medical literature was performed that described typical therapeutic management approaches for acinar cell carcinoma of the pancreas and reported on disease control and survival rates. Data for the case series were obtained from a prospective database. RESULTS: In our systematic review of 6 articles, study patients had a median age of 61 years, 66% were male, 52% had stage I/II disease, and 55% of lesions were located in the pancreatic head. The rates of median survival were approximately 47 months after resection with adjuvant therapy, 38 months for nonmetastatic, locally unresectable disease, and 17 months for metastatic disease treated with chemotherapy. Combination fluoropyrimidine-based chemotherapy regimens had better rates of disease control than other therapies. Our case series included 21 study patients, 14 of whom required resection and 7 who had metastatic disease. The rates of median survival were 40.2 ± 31.9 months in those who underwent surgery and were treated with adjuvant therapy and 13.8 ± 11.3 months for patients with metastatic disease. CONCLUSIONS: Multidisciplinary treatment for acinar cell carcinoma of the pancreas should be considered due to the rarity of the disease and its lack of high-level therapeutic data. Progress in the molecular analysis of this tumor may improve outcomes through the use of personalized therapy based on underlying tumor mutations.

Construction and Validation of a Multi-Institutional Tissue Microarray of Invasive Ductal Carcinoma From Racially and Ethnically Diverse Populations.

BACKGROUND: The scarcity of tissues from racial and ethnic minorities at biobanks poses a scientific constraint to research addressing health disparities in minority populations. METHODS: To address this gap, the Minority Biospecimen/Biobanking Geographic Management Program for region 3 (BMaP-3) established a working infrastructure for a "biobanking" hub in the southeastern United States and Puerto Rico. Herein we describe the steps taken to build this infrastructure, evaluate the feasibility of collecting formalin-fixed, paraffin-embedded tissue blocks and associated data from a single cancer type (breast), and create a web-based database and tissue microarrays (TMAs). RESULTS: Cancer registry data from 6 partner institutions were collected, representing 12,408 entries from 8,279 unique patients with breast cancer (years 2001-2011). Data were harmonized and merged, and deidentified information was made available online. A TMA was constructed from formalin-fixed, paraffin-embedded samples of invasive ductal carcinoma (IDC) representing 427 patients with breast cancer (147 African Americans, 168 Hispanics, and 112 non-Hispanic whites) and was annotated according to biomarker status and race/ethnicity. Biomarker analysis of the TMA was consistent with the literature. CONCLUSIONS: Contributions from participating institutions have facilitated a robust research tool. TMAs of IDC have now been released for 5 projects at 5 different institutions.

Tumor regression grade in gastric cancer: Predictors and impact on outcome.

BACKGROUND: The clinical value and prognostic implications of histologic response to neoadjuvant chemotherapy in gastric cancer is unknown. METHODS: Tumor regression grade (TRG) was recorded in 58 gastric cancer patients identified from two institutional surgical databases. TRG 1a/b represented histologic responders (<10% viable tumor), while TRG 2/3 represented non-responders (>10% viable tumor). RESULTS: TRG 1a/b was recorded in 10 patients (17%), while 48 patients (83%) had a TRG 2/3 response. Larger tumor size (OR 0.24; 95%CI 0.09, 0.64; P = 0.004) and clinical downstaging (OR 30.0; 95%CI 3.26, 276; P = 0.003) were the only factors predictive of histologic response. TRG 1a/b responders had 3-year survival of 70.0% and an estimated overall survival of >69.8 months compared to 38.2% and 22.8 months in non-responders; however, this trend was not statistically significant (P = 0.535). While TRG could not predict survival (OR 2.40; 95%CI 0.46, 12.57; P = 0.300), patient age (OR 1.06; 95%CI 1.00, 1.11; P = 0.035), and the number of positive lymph nodes (≥7; OR 0.05; 95%CI 0.07, 0.27; P < 0.001) were independent predictors of survival. CONCLUSIONS: Few gastric cancers demonstrate histologic response to neoadjuvant chemotherapy. While TRG may be a valid marker for treatment response, its predictive value and clinical application in gastric cancer remains unclear. J. Surg. Oncol. 2016;114:434-439. © 2016 Wiley Periodicals, Inc.