bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 103: Array-based DNA methylation profiling of oral squamous cell carcinoma with lymph node invasion and clinical implications
Authors: Huang Wei-Li , Chang Shu-Hao , Kao Shou-Yen , Su Pei-Fen
Published: June 2012

Background: Oral squamous cell carcinoma (OSCC) is the sixth common cancer and is highly mortal due to tumor recurrence and metastasis. Invasion of tumor cells into lymph nodes is one of the characters of regional metastasis. Here we assessed whether epigenetic modification of DNA methylation is correlated to OSCC regional metastasis and evaluated the potential clinical implications of DNA methylation-based biomarker.

Methods: A case-control study of OSCC patients with lymph node invasion (N=8), OSCC patients without lymph node invasion (N=6), and normal oral tissues (N=2) was performed. To characterize aberrant DNA methylation in OSCC tumors with regional metastasis at diagnosis, Illumina infinium beadsarrays containing 27,578 CpG loci which cover 14,475 genes were applied. The genomic DNA of test samples was individually bisulfite-converted and subjected to beadsarray analysis. β value, the indicator of methylation status of each tested CpG locus, was derived with proper normalization.

Results: Methylation β values allocated tested tissues into regional metastatic tumor, non-metastatic tumor, and control groups. A cluster of 111 CpG loci was selected based on the difference between the mean β value of OSCC tumors with and without regional metastasis. There were 69 loci and 42 loci that were hypermethylated (Δβ value ≫ 0.2) and hypomethylated (Δβ value ≪ −0.2), respectively, in OSCC with regional metastasis. Differentially methylated genes appeared to be mainly related to organ development, cell death, cell signaling, behavior, nucleic acid metabolism, and molecular transport. Several genes were further investigated for their methylation statuses by genomic bisulfite sequencing and DNA methyltransferase inhibitor reactivation in oral carcinoma cell lines with or without invasiveness. Characterization the association of hypermethylation of metastatic-associated genes and clinical pathological variables, we found combination of the hypermethylation status of a candidate gene and p16INK4A improved the prediction of lymph node invasiveness (p=0.003) and survival rate (p=0.010). Our results show the feasibility of this beadsarray analysis to identify aberrant methylation of metastatic-associated genes in OSCC as well as potential risk assessment application. The cohort for the risk assessment assay will be increased and the biological activities of candidate genes in carcinogenesis will be discussed.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 103. doi:10.1158/1538-7445.AM2011-103
Abstract 1034: A natural inhibitor of c-Jun N-terminal kinase 1
Authors: Yao Ke , Cho Yong-Yeon , Chen Hanyong , Song NuRy , Liu Kang-Dong , Wen Weihong , Bode Ann M. , Dong Zigang
Published: June 2012

The c-Jun N-terminal kinases (JNKs) pathway plays a important role in many physiological processes including cell proliferation, survival, death, DNA repair and metabolism induced by a variety of stress signals and pro-inflammatory stimuli. The JNK proteins are encoded by three genes, jnk1, jnk2, jnk3. The protein products of jnk1 and jnk2 are found in all cells and tissues, whereas the JNK3 protein is found primarily in the brain, but is also found in the heart and the testes. The unequal expression of the different isoforms of JNK is possibly determined by some unique characteristics. Each JNK appears to have a distinct function in cancer, asthma, diabetes, and Parkinson's disease. Recent studies demonstrate that JNK1 is a key contributor for the serine phosphorylation of the IRS-1. Several other studies also demonstrated the importance of JNK1, but not JNK2, in lung fibrosis and gastric cancer. We have identified a novel flavonoid isolated from natural sources, herein referred to Compound-A, that suppressed JNK1 activity, with no effect on JNK2 in vitro. At 10 μM, Compound-A specifically inhibited JNK1 by about 50%. We also found that compound-A inhibits UVB-induced c-Jun phosphorylation in a cell culture system. In contrast to SP600125, a non-selective JNKs inhibitor that binds to the ATP active site, compound-A appears to target the protein-protein interaction between JNK1 and its binding proteins, including the JNK-interacting protein-1(JIP1) rather than the ATP binding site. In a computer simulation model, compound-A complexed with JNK1, but not JNK2, and formed strong hydrogen bonding at Glu329. Glu329 is a key residue for the JNK complex with JIP1. This interaction induces a hinge motion between the JNK N- and C-terminal domains and decreases the affinity of the kinase and ATP. These results demonstrate that compound-A is a selective JNK1 inhibitor. In humans, JNK1 play a key role in obesity-induced type 2 diabetes mellitus and the absence of JNK1 results in decreased adiposity, significantly improved insulin sensitivity and an enhanced insulin receptor signaling capacity in mouse obesity models. Therefore, we suggest that because of JNK1's critical role in diabetes, compound-A might have therapeutic potential against this devastating disease.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1034. doi:10.1158/1538-7445.AM2011-1034
Abstract 1037: Effects of stimulary GTP binding protein on the expression of histone deacetylase 6 and 8
Authors: Lim Jeong Ah , Park Ji-Yeon , Juhnn Yong-Sung
Published: June 2012

Stimulatory heterotrimeric GTP-binding protein (Gαs protein) is one of the four families of α subunit in G proteins, and stimulates cAMP production. Gαs protein is involved in many cellular signaling pathways, and regulates various cellular metabolism and gene expression. Histone deacetylases (HDACs) are the enzymes that perform epigenetic regulation of gene expression by removing of acetyl groups from histones. The HDAC family is composed of 18 proteins, and belongs to two major families: the zinc-dependent HDACs (class I, II, and IV), and NAD-dependent (sirtuins-like class III). Human class I HDACs include HDAC1, 2, 3 and 8, and human class II HDACs include HDAC4, 5, 6, 7, 9 and 10.

Thus we analyzed the effect of Gαs on the expression of HDAC isoforms and its underlying molecular mechanism in H1299 human lung cancer cells. We found the expression of HDAC6 and HDAC8 isoforms were increased by the overexpression of the constitutively active Gαs proteins (GαsQL). The levels of HDAC6 and HDAC8 mRNA were increased following GαsQL expression. Treatment with forskolin also increased HDAC6 and HDAC8 levels, and treatment with H89 inhibited the increase in HDAC6 and HDAC8 levels induced by GαsQL. Furthermore, prostaglandin E2, a Gαs activating signal, also increased in HDAC8 levels. The increased expression of HDAC6 through this signaling pathway caused a decrease in acetylation level of α-tubulin. From these results, it is concluded that Gαs-cAMP signaling pathway augments HDAC6 and HDAC8 expression in H1299 lung cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1037. doi:10.1158/1538-7445.AM2011-1037
Abstract 1038: Regulation of cyclin A by FoxO3a drives proliferation in anaplastic thyroid carcinoma
Authors: Marlow Laura , von Roemeling Christina , Zhang Yilin , Cooper Simon , Rohl Stephen , Tun Han , Storz Peter , Smallridge Robert , Copland John A.
Published: June 2012

The Forkhead transcription factor, FoxO3a, is a known suppressor of primary tumor growth and is negatively regulated by growth factors via phosphorylation and mislocalization from the nucleus to the cytoplasm. Here we show for the first time that FoxO3a is nuclear in anaplastic thyroid carcinoma (ATC) and it transcriptionally upregulates cyclin A thereby promoting tumor growth in human ATC cells. Silencing FoxO3a in ATC cell lines leads to down-regulation of cyclin A mRNA and protein while over-expression of FoxO3a leads to upregulation of cyclin A protein. Transient co-transfection of cyclin A promoter linked to luciferase with FoxO3a leads to enhanced luciferase activity, which can be ablated when mutating the FoxO3a site on the cyclin A promoter. This combined data implicates an entirely novel function for FoxO3a in modulating cancer progression by promoting cell cycle progression and tumor growth. ATC is a poorly differentiated carcinoma with average survival upon diagnosis of 3–4 months with 99% lethality, notably the most aggressive and deadliest cancers known. This data also suggest that any therapies under development to inactivate FOXO3a may be effective at blocking tumor expansion.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1038. doi:10.1158/1538-7445.AM2011-1038
Abstract 1041: Alterations of INPP4B, PIK3CA and pAkt of the PI3K pathway are associated with squamous cell carcinoma of the lung
Authors: Magnussen Annika , Karlsson Christina , Söderkvist Peter , Karlsson Mats G. , Thunell Lena K.
Published: June 2012

The aim of the study was to investigate how alterations in the PI3K pathway correlate with the non-small cell lung cancer subtypes squamous cell carcinoma (SSC) and adenocarcinoma (ADCA). We analyzed copy number variation (CNV) and protein expression of the newly identified phosphatase INPP4B, protein expression of pAkt, PDPK1 and PTEN as well as mutational status of PIK3CA and PTEN in 180 lung cancer cases. Nineteen % of all cases displayed loss of INPP4B copy whereas 47% lacked expression. Both CNV and expression analyses showed a correlation between loss of INPP4B and the squamous subtype (RR = 1.44, p = 0.06; RR = 1.86, p = 0.00003). Elevated level of pAkt expression was seen in 63% of all cases and also correlated specifically to SCC (RR = 1.71, p = 0.001). PDPK1 was expressed in 70% of all cases, more in male than female patients, but no difference between the tumor subtypes. Regarding PTEN, 50% of all cases displayed loss of expression, of which seven cases were identified with mutations in the phosphatase or C2 domains. We detected nine cases (5%) of PIK3CA mutations, all identified as the E545K hot spot mutation in the helical domain, all except one in SCC (RR = 1.69, p = 0.05). When analyzing all PI3K pathway components together, we show that patients with at least one alteration in the PI3K pathway are twice as likely to have SCC, than ADCA (RR = 2.01, p = 0.02). Interestingly, we also found a strong correlation between high pAkt expression and PTEN expression, which we speculate can be explained by a feedback-loop via EGR1 and ARF recently proposed by Yu et al, 2009. As comparison, we also analyzed MAPK pathway genes, where we identified fifteen KRAS mutations (8%) and one BRAF mutation (1%), significantly associated to ADCA. The intermediate insulin receptor substrate 1 (IRS-1) is involved in activation of both the PI3K and MAPK pathways and we were interested in studying the Gly972Arg polymorphism previously shown to be associated with prostate, colon and breast cancer. However, we could not detect any associations to any subgroup. In conclusion, we show here that several components of the PI3K pathway, alone and in combination, are correlated to development of squamous cell carcinoma of the lung.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1041. doi:10.1158/1538-7445.AM2011-1041
Abstract 1042: Balance between several phosphatases is crucial for statin- and ATP-induced depletion of nuclear pAkt in prostatic cancer cells
Authors: Ghalali Aram , Högberg Johan , Stenius Ulla
Published: June 2012

We have previously shown that cholesterol-lowering drugs, statins, or extracellular ATP, induce a complex and coordinated response in insulin-stimulated non-small cell lung cancer cells (A549), leading to depletion of nuclear phosphorylated Akt (pAkt). The response involved activation of protein/lipid phosphatases PTEN, PHLPP1 and −2, PP2A and calcineurin (J Biol Chem, 285, 27900, 2010). As Akt might be critical for prostatic cancer (PC) development and statins may prevent aggressive PC the effects of statins and ATP on Akt signaling in PC cells was studied. It was found that atorvastatin in micromolar concentrations decreased constitutive levels of pAkt in DU145 and 22RV1 cells. The effect was associated with a decreased phosphorylation of downstream targets of pAkt. Furthermore, we found that insulin-induced pAkt was inhibited by atorvastatin. Our previous data indicated that PTEN and PHLPP are essential for depletion of nuclear pAkt. In line with this no down-regulation of nuclear Akt and its downstream targets was induced by statins in PTEN deficient LNCaP cells. However, the statin-induced down-regulation of nuclear Akt was restored when cells were transfected for PTEN. We also transfected PTEN deficient PC3 cells with PTEN. However, in this cell line PTEN transfection did not restore the capacity of statins to deplete nuclear pAkt. Instead, we found that in PC3 cells the PTEN transfection lowered levels of PHLPP2 and PHLPP1. When the cells were transfected for both PTEN and PHLPP2 the capacity of statins to deplete nuclear pAkt was restored in PC3 cells. The crosstalk between PHLPP and PTEN was further studied in LNCap, PC3, 22RV1 cells and it was found that increasing PTEN levels depressed the levels of PHLPP 1 and 2 and vice versa. The similar cross talk was also observed between PHLPP1 and 2. These data indicate that a balance between different phosphatases is essential for statin-induced depletion of nuclear Akt. This might be due to the fact that formation of complexes between Akt and different phosphatases is needed for the depletion of nuclear pAkt. Our data suggest that a well-controlled balance between essential phosphatases is needed for nuclear pAkt depletion. This balance seems disturbed in many prostate cancer cell lines.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1042. doi:10.1158/1538-7445.AM2011-1042
Abstract 1053: Mouse Kremen 1 negatively regulates Wnt/β-catenin signaling in human breast cancer MDA-MB-231 cells
Authors: King Taj , Li Yonghe
Published: June 2012

The canonical Wnt/β-catenin signaling pathway is a major developmental pathway that regulates embryonic development, cell motility, polarity, growth, and progression. Wnts are secreted glycoproteins that bind to the low-density lipoprotein receptor-related protein 5/6 (LRP5/6) and Frizzled, a seven pass transmembrane receptor protein, which increase cytosolic β-catenin levels and subsequent β-catenin transcriptional activity via the inhibition of glycogen synthase kinase-3β. Kremen receptors are single-pass transmembrane proteins that consist of two isoforms (Kremen 1 and 2) in mammals. Dickkopf (Dkk) proteins are secreted glycoproteins that inhibit Wnt signaling. Kremen antagonizes Wnt signaling by forming a ternary complex with Dkk1 and LRP5/6, while Kremen alone is able to promote LRP6 cell-surface localization and stimulate LRP6 signaling. Previous studies have shown that aberrant Wnt/β-catenin signaling promotes tumorigenesis in breast cancer and that LRP6 is an important mediator in mammary gland development and in breast cancer tumorigenesis. In the present studies, we found that breast cancer MDA-MB-231 cells express LRP6 and Dkk1, but not Kremen 1 and 2. Therefore, MDA-MB-231 breast cancer cells were used to make stable cell lines that conditionally express mouse Kremen 1 in the presence of doxycycline. We found that doxycycline-induced Kremen 1 expression resulted in the down-regulation of endogenous LRP6 expression and inhibition of Wnt/β-catenin signaling in MDA-MB-231 cells. Furthermore, we demonstrated that Kremen 1 expression blocked Wnt-3a-indced activation of Wnt/β-catenin signaling in MDA-MB-231 cells. Future studies will focus on the effects of Kremen 1 on tumor progression in vitro and in vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1053. doi:10.1158/1538-7445.AM2011-1053
Abstract 1054: S100B affects cell signaling in malignant melanoma
Authors: Gianni Kira , Vitolo Michele I. , Wilder Paul T. , Martin Stuart S. , Weber David J.
Published: June 2012

Establishing the mechanism by which S100B effects ERK and its downstream proteins will provide valuable insight into the role of S100B in the progression of melanoma and potentially aid in developing new pharmacological drugs. S100B is a 21.5 kDa symmetric homodimer that is highly conserved and expressed in a number of tissues and cell lines, including melanocytes. Generally, low levels of S100B have trophic effects, while higher levels are problematic, as is the case in human malignant melanoma. S100B is an effective and widely used prognostic marker for malignant melanoma, with increased levels of serum S100B being predictive of disease stage, increased recurrence, and low overall survival. It has previously been found that S100B directly interacts with the tumor suppressor protein p53, down-regulating its protein levels, thereby allowing damaged cells to proliferate and mutations to propagate. S100B has also been implicated in several other signaling pathways, including the MAPK pathway (BRAF-MEK-ERK). The initial hypothesis tested was that S100B inhibits the phosphorylation of ERK targets in a calcium-dependent manner. To evaluate the significance of S100B in melanoma cell lines, stable S100B knock-down clones were created using lentiviral shRNA particles. The extent of the knock-down varies among the clones and shows a correlation between the level of remaining S100B and the level of phosphorylated ERK, where the clone with the least amount of S100B also has the lowest level of phosphorylated ERK. The clones also display a decreased rate of proliferation measured with both an MTT based assay, as well as with electrical impedance. Over-expression studies were also conducted where normal human S100B, but not a mutant S100B incapable of binding calcium, results in increased phosphorylated ERK, but decreased phosphorylated RSK at Thr573. To determine if S100B was interacting with RSK directly, pull-down assays were performed, and RSK was detected in S100B pull-downs in the presence of calcium; however, it was not detected in the presence of the calcium chelator EDTA. In conclusion, we demonstrate that the calcium-binding protein S100B affects the MAPK pathway by simultaneously increasing phosphorylated ERK and decreasing phosphorylated RSK, together which are consistent with the increased growth rates and cancer progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1054. doi:10.1158/1538-7445.AM2011-1054
Abstract 106: Epigenetic regulation of snoRNAs in cancer
Authors: Ferreira Humberto , Heyn Holger , Balint Balazs , Martin-Subero Jose Ignacio , Esteller Manel
Published: June 2012

Cancer has been associated with epigenetic abnormalities leading to deregulated expression of both coding and non-coding RNAs (ncRNAs) involved in oncogenesis. Among these ncRNAs, microRNAs (miRNAs) were identified acting as oncogenes or tumor suppressors by repressing the expression of cancer related genes. In addition, other studies suggest that other ncRNAs are similarly involved in tumorigenesis, including long non-coding RNAs (lncRNAs) and transcribed Ultraconserved Regions (t-UCRs) by different mechanisms.

Recent studies have shown the impact of small nucleolar RNAs (snoRNAs) in the onset or progression of cancer. SnoRNAs encoded by the ncRNA GAS5 that is repressed in breast cancer or the snoRNA U50, which is mutated in breast and prostate cancer, are examples of the connection between snoRNAs and cancer. It is also known that some snoRNAs are differentially expressed in cancer in comparison with the correspondent matched tissue, indicating their possible role in this disease.

This study focuses in the analysis of the DNA methylation profile associated with the differentially expression of snoRNAs in cancer considering a host gene dependent or independent regulation. To select snoRNAs regulated by DNA methylation we used two different epigenetic approaches: snoRNA expression arrays upon DNA methylation inhibition and global genomic DNA methylation arrays. Herein, we were able to indentify snoRNAs with different DNA methylation that might be involved in cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 106. doi:10.1158/1538-7445.AM2011-106
Abstract 1062: Identification of flotilin-1 as an H-Ras-interacting lipid raft protein: Its implications in breast epithelial cell invasion
Authors: Yong Hae-Young , Son Hwajin , Kim Eun-Sook , Hwang Jin-Sun , Kim Myeong-Ok , Choi Wahn Soo , Noh Dong-Young , Hwang Ki-Tae , Kim Hyeong-Reh Choi , Kim Ki-Bum , Choi Yoomi , Kim Eun Joo , Park So Yeon , Moon Aree
Published: June 2012

Metastatic spread is the major cause of death from breast cancer. We previously showed that H-Ras, but not N-Ras, induces invasive/migratory phenotypes of MCF10A human breast epithelial cells, while both H-Ras and N-Ras induce proliferation/transformation. Here, we identified flotillin-1 as an H-Ras-induced lipid raft protein through comparative proteome profiling of lipid raft proteins. Flotillin-1 interacts with H-Ras in lipid raft at a higher affinity than with N-Ras, elucidating a molecular nature of the crucial link between lipid rafts and the function of H-Ras. Small interfering RNA (siRNA)-mediated flotillin-1 knockdown significantly reduced H-Ras activation and H-Ras-mediated motility/invasion in MCF10A cells engineered to express H-Ras and in highly invasive human breast carcinoma Hs578T cells that express endogenous H-Ras. These results suggest a positive signal amplification loop between flotillin-1 and H-Ras for the invasive signaling program in breast epithelial cells. Importantly, tissue microarrays of 289 patients with invasive breast cancer revealed that flotillin-1 expression positively correlates with lymph node metastases (p=0.027), HER2/neu oncogene expression (p<0.001), and high histologic grade (p=0.014). Although infrequent, the membranous flotillin-1 expression is significantly associated with poor disease-free survival of patients (p=0.005), suggesting the clinical importance of predicting the characteristics of a small subpopulation of malignant breast cancer. Taken together, our findings provide new insight into the molecular basis of the Ras isoform-specific signaling mechanism that depends on the lipid-based sorting platforms. [This work was supported by the Korea government (Nos.ROA-2008-000-20070-0)]

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1062. doi:10.1158/1538-7445.AM2011-1062
Abstract 1066: ERK/RSK and JNK signaling cooperatively mediate oncogenic Ras-induced death receptor 5 expression through enhancing CHOP, Elk1 and c-Jun-dependent transcription
Authors: Oh You-Take
Published: June 2012

Oncogenic mutation of Ras frequently occurs in many types of cancers and plays a critical role in cell transformation and oncogenesis. Interestingly, the oncogenic Ras has been documented to induce the expression of death receptor 5 (DR5), a pro-apoptotic death domain-containing cell surface receptor for TRAIL, through currently undefined mechanisms. The current study attempted to address the mechanisms by which the oncogenic Ras induces DR5 expression. Enforced expression of the oncogenic H-Ras or K-Ras (e.g., RasV12) increased DR5 promoter activity and DR5 expression. This was also accompanied by activation of both ERK/RSK and JNK signaling pathways as evidenced by increased levels of p-ERK, p-RSK, p-JNK and p-c-Jun as well as their downstream proteins, CHOP, Elk1 and c-Jun. Gene silencing-mediated inhibition of either ERK, RSK or JNK activation blocked RasV12-induced DR5 expression. Moreover, knockdown of CHOP, Elk1 or c-Jun abrogated RasV12-induced DR5 expression as well. These data collectively indicate that both ERK/RSK and JNK signaling pathways are required for RasV12-induced DR5 expression. Triple gene (CHOP, Elk1 and c-Jun) co-expressed resulted in the highest DR5 promoter transactivation and protein expression in comparison to single or double gene expression. Consistently, oligonucleotide pull-down assay could detect Elk1, CHOP and c-Jun proteins bound to the DR5 promoter in cells co-transfected with CHOP, Elk1 and c-Jun genes or transfected with Ras-V12 gene, indicating that these proteins together bind to the DR5 promoter region. Thus we suggest that these three proteins cooperatively participate in RasV12-induced DR5 expression by enhancing its transcription. Importantly, we found that RasV12-transformed human ovarian cells were more sensitive than their non-transformed counterparts to undergo TRAIL-induced apoptosis. In summary, this study demonstrated that oncogenic Ras induces DR5 expression through activation of both ERK/RSK and JNK signaling pathways and subsequent CHOP, Elk1 and c-Jun-mediated gene transactivation. Cancers with Ras mutation may be suitable for TRAIL-based cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1066. doi:10.1158/1538-7445.AM2011-1066
Abstract 1067: SPROUTY-2 represses E-cadherin expression and vitamin D action in colorectal cancer
Authors: Barbachano Antonio , Ordonez-Moran Paloma , Pereira Fabio , Sanchez Agustin , Rojas Jose Maria , Munoz Alberto
Published: June 2012

Introduction: Colorectal cancer (CRC) is one of the highest in prevalence and mortality worldwide. Epidemiological and preclinical studies indicate that 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3), the most active metabolite of vitamin D, and several analogs have protective effects against CRC. Previously we have reported that 1,25(OH)2D3 inhibits the proliferation and promotes the differentiation of human colon cancer cells at least in part by the induction of the adhesion molecule E-cadherin and the antagonism of the Wnt/β-catenin pathway (Pálmer et al., J. Cell Biol., 2001; Aguilera et al., Carcinogenesis, 2007). A transcriptomic analysis revealed a decrease in the RNA level of SPROUTY-2 (SPRY2), a gene that modulates signaling mediated by several tyrosine kinase receptors following treatment of human SW480-ADH cells with 1,25(OH)2D3 (5.6-fold at 4 h post-treatment; Pálmer et al., Cancer Res., 2003).

Methods: Protein expression in cell lines were done by Western blot analysis. RNA expression were performed by quantitative RT-PCR. To knock-down gene expression, cells were infected with lentiviral particles containing a U6 promoter driving an shRNA targeting the respective RNA. Protein expression in human colon tumors were performed by immunofluorescence. Cell proliferation was measure by MTT assay.

Results: In this study, we show that the expression of SPRY2 is inhibited by 1,25(OH)2D3 in colon cancer cells by E-cadherin-dependent and -independent mechanisms. In turn, ectopic expression of SPRY2 in SW480-ADH cells inhibits both basal and 1,25(OH)2D3-induced E-cadherin expression. SPRY2 induces ZEB-1 RNA and protein, a transcriptional repressor of CDH1/E-cadherin, and prevents the induction of an adhesive phenotype by 1,25(OH)2D3. A study of the mutual repression between SPRY2 and E-cadherin proteins revealed that their expression levels are inversely correlated in colon cancer cell lines and in xenografted tumors. In colon cancer patients, SPRY2 expression correlates also inversely with that of E-cadherin and is upregulated in invasive areas and high grade tumors. In addition, the ectopic expression of SPRY2 precludes other 1,25(OH)2D3 actions such as the antagonism of Wnt/beta-catenin signaling and the inhibition of cell proliferation. Conclusions: Our data indicate that SPRY2 is a negative regulator of E-cadherin and 1,25(OH)2D3 action and has a putative tumorigenic action in colorectal cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1067. doi:10.1158/1538-7445.AM2011-1067
Abstract 1069: Clinicopathological significance of CARD10 alterations in Middle Eastern colorectal cancer
Authors: Abubaker Jehad , Bavi Prashant , Jehan Zeenath , Ahmed Maqbool , Al-Haqawi Wael , Sultana Mehar , Prabhakaran Sarita , Al-Sanea Nasser , Abduljabbar Alaa , Ashari Luai H. , Alhomoud Samar , Al-Dayel Fouad , Uddin Shahab , Al-Kuraya Khawla S.
Published: June 2012

The incidence of Colorectal Carcinoma (CRC) in Saudi Arabia is rising and CRC is among the top 3 Saudi cancers in both men and women. NF-κB gene plays a role in tumorgenesis through the transcriptional activation of genes associated with cell proliferation, angiogenesis, metastasis, tumor promotion, inflammation and suppression of apoptosis. NF-κB protein activation, defined as nuclear expression of NF-kB by immunohistochemistry was seen in 43.2% (164/380) CRC and was an independent prognostic marker for poor survival. CARD10 and A20 genes are known positive and negative regulators of the NF-kB activation pathway and can activate the NF-kB activity. CARD10 is an enzyme that is encoded by the CARD10 gene and it associates with IKK complex resulting in NF-κB activation. CARD11 which shares a high degree of structure, domain and functional homology to CARD10 is oncogenic in diffuse large B cell lymphoma and results in activation of NF-κB. Considering the potential therapeutic utility of targeting NF-kB and its key modulators, we studied CARD10 genetic alterations (mutations and amplifications) and immunohistochemical expression of CARD10 in over 300 CRC cases and 13 colon cell lines. We detected missense mutations in 5.2 % (15/288) and 2 colon cell lines. Our results showed that CARD10 protein is widely expressed in CRC (67.7%) and expression levels of CARD10 progressively increase from normal colorectal mucosa to pre-malignant colorectal adenomas to CRC. CARD10 protein expression associated significantly with NF-KB activation (P = 0.0313). Interestingly, there was a mutual exclusivity between CARD10 protein overexpression and A20 loss of expression which might suggest that NFKB can be activated by alteration in one of these genes in CRC. Furthermore, CARD10 amplification (15.3%) was significantly associated with CARD10 protein expression and presence of distance metastasis (p=0.0487) Interestingly CARD10 mutations were detected in 21.4% of MSI CRC and only 78.6% of MSS CRC (p=0.0251), suggesting that CARD10 mutations play a role in both subsets of CRC but might represent more important oncogenic event for the development/progression of MSI CRC than MSS CRC. Our study highlights for the first time the role of CARD10 protein overexpression as a novel mechanism for NF-kB activation and demonstrates clinico-pathological significance of CARD10 alterations in Middle Eastern CRC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1069. doi:10.1158/1538-7445.AM2011-1069
Abstract 1075: STAT3 signaling in thyroid cancer
Authors: Silva Joana , Soares Paula , Knauf Jeffrey , Fagin James , Sobrinho-Simoes Manuel , Ghossein Ronald , Bromberg Jacqueline F.
Published: June 2012

Here, we examined the contribution of STAT3 activation in papillary thyroid carcinoma (PTC), which constitutes 80% of all thyroid malignancies. We initially examined STAT3 activation (i.e. Y705 phosphorylation) by IHC in 80 primary PTCs. Approximately 50% of the tumors expressed nuclear pSTAT3. In general, positive cancer cells were found on the edge of the tumor and in association with the stroma (also strongly pSTAT3 positive). The inner part of the tumor was generally negative, as well as all solid PTCs. Additionally, we examined human thyroid cancer derived cell lines and determined that STAT3 was phosphorylated in ∼50% and was regulated by the IL-6/gp130/JAK pathway, similarly to what has been described in breast and lung cancers.

To assess STAT3's role in thyroid cancer, we evaluated the effects of STAT3 under-expression in two models: thyroid cancer derived cell lines where STAT3 levels were reduced using a short hairpin and a transgenic model of PTC lacking STAT3. We observed no significant effects on in vitro growth, migration and invasion as a consequence of reduced STAT3 levels in cancer cell lines. However, in vivo (xenografts in nude mice) the tumors with low STAT3 were significantly larger compared to control (STAT3 expressing) tumors. Similarly, in a transgenic model of PTC lacking STAT3, tumors were larger, more cellular with evidence of increased proliferation compared to the STAT3 expressing cancers. RNA expression analysis of the human xenografts suggested that tumors deficient in STAT3 have downregulated death-promoting genes. In conclusion, our data suggests that in thyroid cancer STAT3 can function as a tumor suppressor, contrary to the current dogma where STAT3 is considered to be oncogenic. We are currently determining whether STAT3's effect on tumorigenesis is dependent on “activated” or total STAT3 and hope to identify the specific molecular pathways involved.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1075. doi:10.1158/1538-7445.AM2011-1075
Abstract 108: FGF-2 regulates cell proliferation, migration and angiogenesis through a novel NDY1/KDM2B-miR101-EZH2 pathway
Authors: Kottakis Filippos , Polytarchou Christos , Kampranis Sotirios , Tsichlis Philip N.
Published: June 2012

The histone H3K27 methyltransferase EZH2 plays an important role in oncogenesis, by mechanisms that are incompletely understood. Here we show that NDY1/KDM2B, a JmjC domain-containing histone H3K36me2, H3K36me1 and H3K4me3 demethylase synergizes with EZH2 to silence the EZH2 inhibitor miR-101. NDY1/KDM2B and EZH2 repress miR-101 by binding its promoter in concert via a process that is activated by upregulation of NDY1/KDM2B. EZH2 does not bind the miR-101 promoter when overexpressed in the absence of NDY1/KDM2B, while NDY1/KDM2B binds the miR-101 promoter when overexpressed in the absence of EZH2, but fails to repress transcription. The concerted action of NDY1/KDM2B and EZH2, triggered by the upregulation of NDY1/KDM2B, promotes the initial upregulation of EZH2 and initiates a feedforward loop that results to the gradual upregulation of EZH2 and other miR-101 targets. NDY1/KDM2B expression was induced by FGF-2 and to a lesser degree by VEGF, via CREB phosphorylation and activation, downstream of the DYRK1A kinase. The FGF-2-NDY1-miR-101-EZH2 axis was documented in primary fibroblasts in culture, and in a set of human tumor cell lines, and it was shown to be responsible for the FGF-2 and EZH2 effects on cell proliferation, migration and angiogenesis. The axis appeared to also be active in a set of transitional cell bladder carcinomas, in which NDY1/KDM2B and FGF-2 were overexpressed in concert and their upregulation correlated with the repression of miR-101 and the upregulation of EZH2. These data delineate a novel pathway that links FGF-2 signaling with the functional and transcriptional regulation of EZH2 via NDY1/KDM2B and miR-101, and contributes to oncogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 108. doi:10.1158/1538-7445.AM2011-108
Abstract 1080: Protein phosphatase 2A regulates Chk2 phosphorylation at constitutive level in human ovarian cancer cells
Authors: Liang Xiaobing , Guo Yi , Yu Jing Jie
Published: June 2012

Protein phosphatase 2A (PP2A) comprises a diverse family of phosphoserine- and phosphothreonine-specific enzymes ubiquitously expressed in eukaryotic cells. This principal Ser/Thr phosphatase plays an integral role in the regulation of a number of major signaling pathways, the deregulation of which can contribute to multiple human cancers, Alzheimer's disease and increased susceptibility to pathogen infections. The checkpoint kinase Chk2 is an important multifunctional regulator of the cellular DNA damage repair response. Chk2 activation is triggered by the phosphorylation of Thr-68 following DNA damage and protects genome integrity by promoting cell cycle arrest or apoptosis. We have previously reported that ovarian cancer A2780 cells show an interaction between PP2A and Chk2 after cisplatin treatment, and PP2A negatively regulates Chk2 phosphorylation at Thr-68. In this study we report that Chk2 and PP2A are associated to maintain the cell normal function. This conclusion is based on the following observations: 1) Co-immunoprecipatation of the holoenzyme PP2A (subunits A, B and C) with p-Chk2 shown immune complexes; 2) Inhibition of PP2A activity by okadaic acid, an inhibitor of PP2A, and by siRNA to PP2A, resulted in enhanced Chk2 phosphorylation at Thr-68 in studied cell model. These findings demonstrate that PP2A has a role in controlling the constitutive Chk2 phosphorylation at Thr-68 and that Chk2 and PP2A are responsible for maintenance of cell metabolic equilibrium.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1080. doi:10.1158/1538-7445.AM2011-1080
Abstract 1082: The amplification and overexpression of the calcium activated chloride channel TMEM16A (aka DOG1) has a prognostic role in patients with head and neck squamous cell carcinoma
Authors: Ruiz Christian , Zlobec Inti , Rudin Florian , Schneider Sandra , Rufle Alex , Fischer Claude , Tornillo Luigi , Terracciano Luigi M. , Kunzelmann Karl , Bubendorf Lukas
Published: June 2012

Introduction: The calcium activated chloride channel TMEM16A, which is also known as DOG1, is used as a sensitive marker for the diagnosis of gastrointestinal tumors (GIST). Its expression has also been described in head and neck squamous cell carcinomas (HNSCC) and in esophageal SCC. Surprisingly, until recently (2008) its role as chloride channel was not known. The TMEM16A gene is localized very close to the Cyclin D1 gene (CCND1), which is a putative target gene of the 11q13 amplicon. Aim of this study was the analysis of the genomic and expression status of TMEM16A in HNSCC.

Methods: We applied fluorescence in-situ hybridization (FISH) and immunohistochemistry to a tissue microarray (TMA) containing tissue specimens from 216 primary tumors from the oral cavity, 40 local recurrences and 76 neck dissections. We validated our results on a second cohort consisting of additional 158 HNSCCs. In order to analyze the prognostic role of TMEM16A, a subset of 365 primary tumors with complete follow-up data was selected.

Results: We found genomic amplification of the TMEM16A gene in 13% of the primary head and neck squamous cell carcinomas and in 12% of the local recurrences. Although the prevalence of TMEM16A protein expression was lower (8%), a strong correlation between TMEM16A amplification and overexpression was found (p<0.0001). FISH analysis of Cyclin D1 (CCND1) revealed co-amplification of these two genes (p<0.0001). Interestingly, both, genomic amplification and expression of TMEM16A identified a subgroup of HNSCC patients with poor survival (p<0.05).

Conclusions: TMEM16A positivity characterizes a small subgroup of HNSCC with poor survival. The strong correlation between genomic amplification and expression suggests that TMEM16A is one of the driving genes of the 11q13 amplicon in HNSCC. Additional studies may show how this chloride channel can be used as a therapeutic target in patients with TMEM16A positive tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1082. doi:10.1158/1538-7445.AM2011-1082
Abstract 1086: RHA plays an important role in EGFR-mediated gene transcription in cancer cells
Authors: Huo Longfei , Wang Ying-Nai , Xia Weiya , Hsu Sheng-Chieh , Lai Chein-Chen , Li Long-Yuan , Chang Wei-Chao , Wang Yan , Hsu Ming-Chuan , Yu Yung-Luen , Huang Tzu-Hsuan , Ding Qingqing , Chen Chung-Hsuan , Tsai Chang-Hai , Hung Mien-Chie
Published: June 2012

EGF treatment induces the translocation of its receptor (i.e. EGFR) from the cell surface into the nucleus. Nuclear expression of EGFR is negatively correlated with overall survival of cancer patients and EGFR nuclear translocation is important for cancer cell's resistance to the treatment with Cetuximab, cisplatin and radiation. Moreover, EGFR in the nucleus can associates with other proteins including STAT3, STAT5A, DNA-PK and PCNA to regulate cell transformation, proliferation, and DNA repair and replication. It has been demonstrated that nuclear EGFR regulates gene transcription through its binding to an AT-rich sequence (ATRS) of target gene promoter. However, EGFR does not have a DNA-binding domain and there is no evidence to support the direct binding of EGFR to the specific DNA sequence. Thus, identification of a DNA-binding partner of nuclear EGFR is crucial for us to understand how EGFR regulates gene transcription in the nucleus. Using a non-biased approach, we identified RNA helicase A (i.e. RHA), which is a highly conserved, multiple functional transcriptional activator, could interact with nuclear EGFR upon EGF stimulation in several cancer cells. The EGFR/RHA complex then associated with gene promoter through binding of RHA to the AT-rich sequence of the promoter to activate its transcription. Knockdown of RHA expression was found to abrogate the binding of EGFR to its target gene promoter, thereby reducing EGF/EGFR-induced gene expression. Moreover, interruption of EGFR-RHA interaction decreased EGFR-induced promoter activity. Interestingly, we found that the EGFR/RHA-regulated gene promoter activity is dependent on the EGFR tyrosine kinase activity but independent of RHA helicase activity. Finally, we observed a positive correlation among nuclear expression of EGFR, RHA and cyclin D1 in human breast cancer tissue samples. These results, taken together, indicate that RHA is an important mediator for EGFR-induced gene transactivation in cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1086. doi:10.1158/1538-7445.AM2011-1086
Abstract 1087: The role of fibroblast growth factor 9 in the recurrence after radical prostatectomy
Authors: Teishima Jun , Hayashi Tetsutaro , Shoji Koichi , Matsubara Akio
Published: June 2012

Introduction and Objectives: The aim of this study was to investigate the role of fibroblast growth factor 9 (FGF9) in the recurrence after radical prostatectomy. Methods: Immunohistochemical staining using anti-FGF9 antibody was performed for tissues derived from radical prostatectomy in 133 male patients. Cell viability, apoptosis and invasion of LNCaP and androgen-independent LNCaP (AI-LNCaP) cells were assessed with the MTT assay, Apopercentage apoptosis assay and Matrigel invasion assay, respectively, in the presence or absence of treatment with recombinant FGF9 or anti-FGF9 neutralizing antibody. Results: In immunohistochemical staining, FGF9-positive cells were detected in 20 samples. In cases with a Gleason score of 8 or higher, the incidence of cases with FGF9-positive cells was 34.2%, which was significantly higher than that for those with Gleason scores of 7 or lower (7.3%, p=0.0003), respectively. The 3-year biochemical relapse-free survival rate in cases with FGF9-positive cells was 17.5%, which was significantly lower than that in cases in which FGF9-positive cells were not detectable (76.6%, p<0.0001). In androgen-depleted medium, the cell viability of LNCaP was significantly lower than that of AI-LNCaP, and was enhanced to the same level by treatment with recombinant FGF9. Invasion was significantly enhanced and apoptosis by androgen deprivation was significantly suppressed by treatment of LNCaP cells with recombinant FGF9. Furthermore, these effects were significantly suppressed by treatment with anti-FGF9 neutralizing antibody. Conclusions: These results indicate that FGF9 contributes to postoperative recurrence of prostate cancer through stimulation of cell proliferation and invasion, and induction of an anti-apoptotic effect.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1087. doi:10.1158/1538-7445.AM2011-1087