bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4124: Chronic estrogen and NNK stimulate small airway epithelial cells via modulation of nicotinic acetylcholine receptors
Authors: Al-Wadei Hussein AN , Al-Wadei Mohammed H. , Plummer Howard K. , Schuller Hildegard M.
Published: February 2011

Small airway epithelial cell-derived adenocarcinoma is the most common human lung cancer and is particularly prevalent in women. We have previously reported that the proliferation of immortalized human small airway epithelial cells HPL1D is stimulated by a single dose of the tobacco carcinogen NNK via cAMP signaling activated by binding of NNK to the beta-1-adrenergic receptor (β1-AR) and that estrogen enhances this response. In the current study we show that γ-aminobutyric acid (GABA) blocks this cooperative signaling of NNK and estrogen in HPL1D cells by inhibiting the activation of adenylyl cyclase. In addition, HPL1D cells produced the stress neurotransmitters noradrenaline and adrenaline in response to binding of a single dose of NNK to the α7 nicotinic acetylcholine receptor (α7nAChR). Since both stress neurotransmitters are beta-adrenergic agonists, this response further enhanced cAMP-dependent stimulatory signaling. On the other hand, GABA production by these cells was reduced by a single dose of NNK via desensitization of the α4β2 nAChR. Exposure to NNK, estrogen or the combination of both agents for 7 days upregulated and sensitized the α7nAChR, resulting in an enhanced noradrenergic response to agonist. At the same time, chronic NNK and estrogen suppressed the production of GABA by desensitizing its regulatory α4β2nAChR. The resulting predominance of stimulatory cAMP signaling over inhibitory GABA signaling may contribute to the development and progression of small airway-derived adenocarcinoma in women who smoke and offer a novel target for the marker-guided prevention of this cancer.

Supported by a grant from the National Lung Cancer Partnership and LUNGevity Foundation.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4124.
Abstract 4125: The effects of S100B on cell signaling in malignant melanoma
Authors: Gianni Kira , Wilder Paul T. , Vitolo Michele I. , Shapiro Paul , Weber David J.
Published: February 2011

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. The protein S100B is an effective prognostic marker for human malignant melanoma. Increased levels of serum S100B are 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. The initial hypothesis tested was that S100B inhibits the phosphorylation of ERK targets in a calcium-dependent manner. The S100B binding motif has been found to overlap with the ERK binding motif, suggesting that S100B binds the targets and physically prevents ERK from interacting with and subsequently phosphorylating them. ERK kinase assays revealed that the addition of S100B in the presence of calcium decreased ERK phosphorylation of RSK, but more dramatically Elk-1. Additionally, 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. Over-expression of normal human S100B, but not a mutant S100B incapable of binding calcium, results in decreased phosphorylated RSK at Thr573, yet increased phosphorylated ERK, interestingly. Since a rise in serum S100B levels is indicative of tumor progression, it was also of interest to see if the addition of extracellular S100B alone is sufficient to elicit the same signaling response. Preliminary data shows an increase in phosphorylated ERK after the addition of low concentrations of extracellular S100B, as well as effects on ERK targets. We demonstrate that the calcium-binding protein S100B affects the MAPK pathway by simultaneously increasing phosphorylated ERK and decreasing phosphorylated RSK and Elk-1, together which could have a significant impact on cancer progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4125.
Abstract 4130: Genomic profiling of ovarian tumor cell lines provides a predictive signature of sensitivity to PF-4691502, a dual PI3kinase and mTOR inhibitor
Authors: Sheppard Karen E. , Neilsen Amelia , Ng Gwyn , Chan Joanna , Tothill Richard W. , Ellul Jason , Hannan Ross D. , Johnstone Ricky W. , McArthur Grant , Phillips Wayne , Pearson Richard B.
Published: February 2011

The PI3kinase/AKT/mTOR pathway plays key roles in cell growth, proliferation and survival with mTOR providing a hub for coordinating signaling in response to growth factors, hormones and nutrients. This pathway is one of the most frequently mutated or abnormally activated pathways in ovarian cancer, thus making it an excellent therapeutic target for this disease.

The aims of this study were:

1. To investigate the potency of PF-4691502 (PF502) a dual PI3 kinase and mTOR inhibitor to inhibit ovarian tumor cell proliferation and induce cell death.

2. To profile exponentially growing ovarian tumor cells using gene expression, copy number variations and activation of the PI3 kinase pathway.

3. To define a predictive signature or biomarker that defines ovarian tumor cell sensitivity to PF502.

A panel of 39 ovarian tumor cells were treated with PF502 (1nM-30µM) for 72h and the dose at which proliferation was inhibited by 50% (GI50) determined. Cells showed a 50-fold difference in PF502 sensitivity with GI50 values ranging from 15nM to 700nM, with the majority (85%) of cells having a GI50 of less than 200nM, these cells were classified as sensitive and the remaining 6 cell lines with a GI50 greater than 350nM were classified as resistant. PF502 also caused cell death in a subset of sensitive ovarian tumor cells. Increase in PIK3CA gene copy number was observed in 40% of the cell lines but this did not correlate with PF502 sensitivity nor did activating mutations in PIK3CA or inactivation of PTEN. Gene expression data was generated from exponentially growing cells using high-density oligonucleotide microarrays. A predictive model of response to PF502 was generated using the gene expression data. Exhaustive 10-fold cross validation of the model showed that tumour cell sensitivity could be predicted with high (96%) accuracy using approximately 20-30 genes. A differential gene expression analysis was performed using all data and 89 genes were found significantly different between sensitive and resistant cell lines. Pathway analysis revealed an over expression of inflammatory markers within the resistant class. Gene set enrichment analysis further supported an increased inflammatory signal within resistant cells, and revealed that both MYC and RAS signaling were also elevated providing a clear indication of potential targets for combination therapies in PF502 resistant disease.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4130.
Abstract 4132: Down Regulation of the CCK-B/C Receptor Inhibits Growth of Pancreatic Cancer
Authors: Fino Kristin K. , Matters Gail L. , McGovern Christopher O. , Kester Mark , Smith Jill P.
Published: February 2011

Background: Pancreatic cancer has a dismal prognosis with only 1% surviving 5 years. Reasons for this poor prognosis include the inability to diagnosis pancreatic cancer in early stages and its resistance to standard chemotherapy. The gastrointestinal peptide, gastrin, has been identified as a growth factor in pancreatic cancer. Gastrin stimulates growth of pancreatic cancer through the cholecystokinin (CCK)-B receptor and a splice variant of this receptor (CCK-C), which is expressed only in cancer cells.

Hypothesis: It is hypothesized that down regulation of the gastrin binding receptors, CCK-B/C, by small interfering RNAs (siRNAs) will inhibit growth of pancreatic cancer.

Methods: A series of human pancreatic cancer cells were screened by Real-Time RT-PCR for the presence and quantity of CCK-B/C receptor mRNA. Receptor binding was identified in pancreatic cancer cells with a CCK-B polyclonal antibody (1:10,000) by immunocytochemistry. PANC-1 cells were transfected with siRNAs that target a region of mRNA common to both the CCK-B and CCK-C receptor with either Lipofectamine 2000 or pegylated cationic nanoliposomes previously shown to be nontoxic after systemic intravenous administration. RNA was extracted and analyzed by TaqMan Real-Time RT-PCR for the receptor expression 3 days after treatment. Control cells were untreated, vehicle only treated (Lipofectamine 2000 or pegylated cationic nanoliposomes), or treated with a scrambled siRNA control. The effects of CCK-B/C knock-down on growth were evaluated by counting cells at various time points by trypan blue dye exclusion.

Results: The CCK-B/C receptor was detected in several human pancreatic cancer cell lines (PANC-1, MIA PaCa-2, AsPC-1, BxPC-3, and Capan-1) by Real-Time RT-PCR. The receptor was identified by immunofluorescence in the plasma membrane. CCK-B/C receptor expression was down regulated by CCK-B/C siRNA with both Lipofectamine 2000 and pegylated cationic nanoliposomes. PANC-1 cell growth was significantly decreased in CCK-B/C receptor targeted siRNA treated cells compared to untreated cells, vehicle only, and scrambled control on day 3 (p<0.0001, n=9). Control cells more than doubled in number while CCK-B/C siRNA treated cells increased by only 10%.

Conclusion: The CCK-B/C receptor is present in human pancreatic cancer cells and knock-down significantly decreases cell growth. Systemic administration of CCK-B/C siRNAs by pegylated cationic nanoliposomes may be a potential therapeutic modality for pancreatic cancer.

Supported by NIH CA117926.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4132.
Abstract 4134: Breast cancer antiestrogen resistance-3 promotes growth in the presence of tamoxifen through a mechanism involving p130cas and c-src
Authors: Guerrero Michael S. , Schuh Natasha R. , Bouton Amy H.
Published: February 2011

Tamoxifen, an antiestrogen and estrogen receptor (ER) ligand, has been successfully used to treat ER-positive breast tumors for almost 40 years. For women with ER-positive disease, tamoxifen treatment can decrease the rate of recurrence by 50% and reduce the risk of death by 25%. Despite such success, resistance occurs in up to one-third of patients. To date, many proteins have been identified that contribute to resistance. Breast Cancer Antiestrogen Resistance-3 (BCAR3) is one such molecule; its expression induces resistance to the antiestrogens tamoxifen and ICI 182, 780 (fulvestrant). To investigate the signaling pathway(s) driving BCAR3-mediated resistance, we generated clones of the ER-positive and antiestrogen-sensitive MCF7 cell line that overexpress BCAR3 (MCF7 B6). Utilizing these cells, we show here that BCAR3 overexpression confers a growth advantage under conditions of both hormone deprivation and tamoxifen treatment. We further employed this inducible cell line to examine the relationship between BCAR3 and two other molecules that have been implicated in both antiestrogen resistance and BCAR3 signaling: the scaffold/adaptor molecule p130Cas (Cas, also known as BCAR1) and the non-receptor tyrosine kinase c-Src (Src). Upon BCAR3 overexpression, we observed an alternative Src-Cas complex, increased Src kinase activity, and an increase in Src-dependent tyrosine phosphorylation of Cas. Interestingly, these BCAR3-induced effects are preserved following tamoxifen treatment. Conversely, upon knockdown of BCAR3 in the ER-negative BT549 breast cancer cell line, Src activity was reduced, as were Cas phosphorylation and Src-Cas interactions. We also examined the expression and subcellular localization of the cyclin-dependent kinase inhibitor and known Src substrate, p27. In MCF7 B6 cells overexpressing BCAR3, we observed a decrease in the nuclear accumulation of p27 under conditions of both hormone deprivation and tamoxifen treatment. Based on these data and the established role of Src-Cas signaling in the induction of antiestrogen resistance, we postulate that these factors play a significant role in the promotion and establishment of BCAR3-mediated tamoxifen resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4134.
Abstract 4136: Effects of magnolol on cell cycle, STAT 3, NF-kappa B and AKT signaling
Authors: Chilampalli Shivani , Kaushik Radhey S. , Chandrasekher Gudiseva , Fahmy Hesham , Dwivedi Chandradhar
Published: February 2011

Magnolol is plant lignan isolated from bark and seed cones of Magnolia glaucan has been investigated for its anticancer effects. Previous studies from our laboratory have shown magnolol to be chemopreventive against UVB-caused skin carcinogenesis by inducing apoptosis. The objective of present study is to investigate other mechanisms involved for its chemopreventive effects. For this investigation, A431 human epidermoid carcinoma cells were used. Effects of magnolol on cell cycle were determined by flow cytometry. Expressions of various proteins involved in cell cycle were determined by Western blotting using appropriate antibodies. A431 human epidermoid carcinoma cells were incubated with different concentrations at different time periods. We further investigated the effects of magnolol on signal transducer and activator of transcription (STAT 3), nuclear factor-kappa B (NF-kappa B/p65) and AKT signaling pathways that play an important role in cell proliferation and survival. Magnolol at 100 μM and 125 µM caused G0/G1 cell cycle arrest by decreasing cyclin D1, cyclin D2, and CDK 4 and by increasing CDK inhibitors p21 and p27 at 24 and 48 h. We further found that magnolol inhibited expressions of STAT 3, NF-kappa B and AKT by inhibiting their phosphorylation at 24 and 48 h. These results indicate that chemopreventive effects of magnolol are mediated through multiple mechanisms.

This study is supported by Translational Cancer Research Center funded as one of the 2010 Research Initiative Center by the State of South Dakota.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4136.
Abstract 4138: Resveratrol induces COX-2-dependent apoptosis in human ovarian cancer cells
Authors: Lin Cassie , Sebuyira Annette , Davis Faith B. , Davis Paul J. , Lin Hung-Yun
Published: February 2011

Resveratrol is a naturally-occurring trihydroyxl-diphenylethylene compound that has been shown experimentally to have beneficial effects in the treatment of cancer and cardiovascular disease. We have studied the molecular basis of the anti-cancer actions of resveratrol, using human ovarian carcinoma (OVCAR-3) cells. Resveratrol induces programmed cell death (apoptosis) in these cells and activates important signal transducing proteins including extracellular signal-regulated kinases 1 and 2 (ERK1/2) in cancer cells. Resveratrol also causes nuclear accumulation of the enzyme cyclooxygenase-2 (COX-2) and of the oncogene suppressor protein, p53. Expressed constitutively, cytoplasmic COX-2 is a marker of tumor cell aggressiveness, but there is a resveratrol-inducible pool of nuclear COX-2 that is important to activation (Ser-15 phosphorylation) of the oncogene suppressor protein, p53. Our findings include the following: (1) Nuclear accumulation of COX-2 in resveratrol-treated cells is blocked by the ERK1/2 inhibitor, PD98059. (2) An inhibitor of COX-2 activity, NS398, prevents accumulation in the nucleus of ERK1/2, COX-2, activated p53 and SUMO-1. (3) Apoptosis, quantitated by nucleosome ELISA and the nuclear abundance of the pro-apoptotic protein, Bcl-xs, were inhibited by NS398. This finding implicates nuclear COX-2 in p53-mediated apoptosis induced by resveratrol. Sumoylation is important to stabilization of p53 and a COX-2/SUMO-1 interaction suggests sumoylation of COX-2 in resveratrol-treated cells. (4) Chromatin immunoprecipitation (ChIP) studies showed binding of induced nuclear COX-2 to the promoter region of p21, a pro-apoptotic gene target of transcriptionally active p53. In summary, nuclear accumulation of activated ERK1/2 and COX-2 are essential to resveratrol-induced, pSer-15-p53-mediated apoptosis in human ovarian cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4138.
Abstract 4140: Inhibition of mTOR signaling by psoralidin in breast cancer
Authors: Damodaran Chendil , Kumar Raj , Rohr Jurgen
Published: February 2011

Breast carcinoma is the most common malignancy in women and is the second leading cause of cancer-related deaths in the United States. Mammalian target of rapamycin (mTOR) is a major effector of the PI3Kinase/Akt pro-survival signaling pathway and has received significant attention as a therapeutic target for the treatment of various malignancies. Rapamycin and its analogs are specific mTOR antagonists; however their toxicity hampers wide use in the clinical setting for the treatment of breast cancer. This compels development of new strategies for the treatment of advanced, triple and double negative breast cancers. Psoralidin is one of the major active ingredients in Psoralea corylifolia, herbal plant that is extensively used in Asian and African traditional medicines. In our study we tested the effects of psoralidin on the mTOR pathway in estrogen-responsive (MCF-7) and -refractory (MDA 231) breast cancer cells. Our results suggest that psoralidin inhibits phoshporlyation of mTOR resulting in the inhibition of the phosphorylation of mTOR effectors (p70S6K and 4EBP1) in both MCF-7 and MDA 231 cells. Interestingly, there was no significant down regulation of pAkt expression in either cell lines following treatment with psoralidin. While dissecting the effect of psoralidin on epithelial-mesenchymal transition (EMT), we found that psoralidin inhibits expression of β-catenin with a concomitant increase in the expression of E-cadherin in both estrogen-responsive and -refractory breast cancers cells. In addition, treatment with psoralidin effectively inhibits the growth of both estrogen-responsive (MCF-7) and -refractory (MDA 231) breast cancer cells and induces apoptosis without causing significant toxicity to normal breast epithelial cells (MCF-10A). Currently we are pursuing studies using xenograft models to determine the effect of psoralidin in estrogen-responsive and -refractory breast cancers animal models. The outcome of this study may provide a mechanistic validation of psoralidin as a potent compound that may be used for the treatment of breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4140.
Abstract 4144: The treatment with a derivative of Vitamin A (ATRA) modulates the expression and subcellular localization of Protein Kinase C (PKC) delta and induces growth inhibition in normal and tumor derived mammary cells
Authors: Berardi Damian E. , Bessone María I. Díaz , de Kier Joffé Elisa D. Bal , Todaro Laura B. , Urtreger Alejandro J.
Published: February 2011

Retinoids may exert some of their effects on cell differentiation and malignant phenotype reversion through the interaction with different PKC isoforms. In the present work we addressed the following issues: A)- the role of ATRA on the in vitro growth potential of LM3, a murine mammary tumor-derived cell line; NMuMG, a normal murine mammary cell line and MDA-MB231, a human breast cancer-derived cell line. B)- the effect of retinoid treatment on the expression and subcellular distribution of PKCδ and C)- the importance of PKCδ in the activity of retinoid receptors.

ATRA treatment (1μM, 72h) induced a significant growth inhibition in the analyzed murine cell lines. This phenomenon was associated with the reduction of pERK1 levels (80±9% and 40±5% in LM3 and NMuMG cells respectively) and the increase of the cell cycle inhibitor p27 in the nuclear fraction without altering the expression of Cyclin D1 in both murine cell lines. None of these modulations were observed in the ATRA unresponsive MDA-MB241 cell line.

By Western blot we could determine that 72h treatment with ATRA induced an increase of PKCδ protein levels both in the nuclear fraction of LM3 cells (4-fold) and in the cytoplasm (2- and 3-fold for LM3 and NMuMG cells respectively). On the contrary, in MDA-MB241 cells PKCδ expression was reduced after ATRA exposure.

Interestingly, pharmacological silencing of PKCδ in LM3 cells prevented the activation of retinoid receptors by ATRA, as evidenced by a reporter gene assay (RARE-Luciferase).

Our results suggest that, in ATRA-responsive cells, the treatment with this vitamin A derivate inhibits in vitro growth, by reducing mitogenic signaling, by increasing the expression of a cell-cycle inhibitory protein as well as by increasing PKCδ an isoform usually associated with growth inhibition and apoptosis induction. The subcellular localization was also altered by this treatment in LM3 cells. Moreover, the effect of ATRA exerted through the activation of its specific nuclear receptors would depend on the nuclear translocation of this PKC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4144.
Abstract 4148: Rapamycin potentiates the effects of paclitaxel in uterine papillary serous carcinoma (UPSC) cells through inhibition of cell proliferation and induction of apoptosis
Authors: Mendivil Albert , Zhou Chunxiao , Cantrell Leigh , Gehrig Paola , Malloy Kim , Bae-Jump Victoria L.
Published: February 2011

Objectives: Treatment of women with uterine papillary serous carcinoma (UPSC) using cytotoxic chemotherapeutic agents has been met with limited success. This has led to the search for an additional agent that could be used in combination with more traditional therapies to dramatically increase therapeutic efficacy while not increasing toxicities. Among the most promising of these for endometrial cancer are inhibitors of the mammalian target of rapamycin (mTOR). Rapamycin and paclitaxel have been found to be synergistic in type I endometrial cancer cell lines. Thus, our objective was to examine the effects of combined therapy with rapamycin and paclitaxel in a UPSC cell line.

Methods: The UPSC cell line, SPEC-2 was used. Cell proliferation was assessed after exposure to rapamycin or paclitaxel alone or in combination, and viable cell densities were determined by measuring conversion of the metabolic dye, MTT. Apoptosis was assessed by flow cytometric analysis of annexin-V expression. hTERT expression, as a marker of telomerase activity, was determined by real-time RT-PCR. Western immunoblotting was performed to determine the expression of the downstream targets of the mTOR pathway, including phosphorylated S6 and 4E-BP-1.

Results: Paclitaxel inhibited proliferation in a dose-dependent manner in the SPEC-2 cell line, with an IC50 value of 1-10 nM. Simultaneous exposure of cells to various doses of paclitaxel in combination with rapamycin (1 nM) resulted in a significant synergistic anti-proliferative effect (p=0.0032-0.0059). In addition, rapamycin increased paclitaxel-induced apoptosis over that of paclitaxel alone. Rapamycin decreased hTERT mRNA expression while paclitaxel alone had no effect on telomerase activity. Rapamycin decreased phosphorylation of the S6 and 4E-BP-1 proteins after 24 hrs of exposure. In contrast, paclitaxel had little effect on 4E-BP-1 phosphorylation but did induce phosphorylation of S6 at higher doses (10 nM). Treatment with paclitaxel and rapamycin resulted in overall decreased phosphorylation of both S6 and 4E-BP-1.

Conclusions: We demonstrate that rapamycin potentiates the effects of paclitaxel in UPSC cells through inhibition of cell proliferation and induction of apoptosis. This suggests that the combination of rapamycin and paclitaxel may be a promising effective targeted therapy for UPSC, an aggressive and often lethal subtype of endometrial cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4148.
Abstract 4149: A newly developed GSK-3 inhibitor, CG9, inhibits breast cancer cell proliferation through estrogen receptor α degradation
Authors: Kim Hyun Mi , Kim Choung-Soo , Lee Je-Hwan , Cho Joong Myung , Hwang Jung Jin , Jang Sejin , Ro Seonggu , Hyun Young-Lan , Choi Jene
Published: February 2011

GSK-3 (glycogen synthase kinase-3) has a major role in Wnt and Hedgehog signaling pathways and regulates the cell cycle, stem cell renewal and differentiation, apoptosis, and insulin action. The pharmacological inhibitors of GSK-3 could be used to treat several diseases, including Alzheimer's disease, other neurodegenerative diseases and diabetes. In this study, we investigated the cellular and molecular effects of a newly developed GSK-3 inhibitor, CG9, in seven breast cancer cell lines of the NCI-60 cell line panel. CG9 treatment increased inhibitory phosphorylation of GSK-3α/β (Ser21/9), whereas decreased activating phosphorylation of GSK-3β (Tyr216), together with the overall reduction of the level of GSK-3 proteins. CG9 also induced growth inhibition and stimulated a caspase-dependent apoptotic pathway regardless of estrogen receptor (ER) positivity with IC50 of 12 μM in T47D to ≤ 0.1 μM in MCF7 cells. The GSK inhibitor down-regulated the cellular levels of ERα and cyclin D in ERα-positive MCF7 cells. Immunoprecipitation experiments showed that CG9 treatment inhibits the interaction of Hsp90 with ERα in cytoplasm. Taken together these results indicate that CG9 may be an effective anti-cancer agent in human breast cancers. Currently, we are investigating the efficiency of CG9 compared with that of antiestrogen tamoxifen (Tam) and a possibility of combined treatment with CG9 and Tam.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4149.
Abstract 4153: Short term opioid receptor blockade inhibits cancer cell proliferation independent of immune function by upregulation of the OGF-OGFr axis: A tissue culture model of low dose naltrexone (LDN)
Authors: Donahue Renee N. , McLaughlin Patricia J. , Zagon Ian S.
Published: February 2011

The opioid growth factor (OGF) is an autocrine produced and constitutively expressed peptide that interacts with the OGF receptor (OGFr) and subsequently targets the cyclin dependent kinase inhibitory (CKI) pathways to inhibit proliferation at the G1/S phase of the cell cycle. Naltrexone (NTX) is a nonselective opioid receptor antagonist that blocks this interaction. In vivo administration of a low dosage of NTX (LDN, 0.1 mg/kg) blocks opioid receptors for 4-6 h, resulting in a compensatory upregulation of OGF and OGFr. Following clearance of NTX, the elevated levels of OGF and OGFr interact to decrease cell proliferation. To investigate whether LDN inhibits cancer cell proliferation independent of immune function, a tissue culture model using short term NTX in human ovarian cancer cell lines (SKOV-3, OVCAR-3) was established. Cell replication and cell number were determined over 96 h in cultures treated with a single 6 h application of NTX (10−6 M). In SKOV-3 and OVCAR-3 cultures, cell number was decreased by 21-28% and 27-32%, respectively, between 48 and 96 h; DNA synthesis was inhibited ∼ 33% at 72 h. Peptide specificity studies using a variety of classical μ, δ, or κ opioid receptor agonists confirmed that only OGF inhibited cell number. Antibody neutralization of OGF in cultures revealed that endogenous OGF invoked the inhibitory growth effects of short term NTX. To evaluate which opioid receptor mediated the inhibitory action of short term NTX on growth, cultures were treated with siRNAs against the μ, δ, κ, and OGFr receptors; only OGFr was associated with short term NTX treatment. Immunocytochemical studies and semiquantitative analyses documented an upregulation of OGF and OGFr expression following short term NTX exposure in cancer cells; western blot analysis and receptor binding for OGFr revealed 2-fold increases of receptors 48 h after a brief exposure to NTX. NTX's inhibitory action was not related to apoptosis (TUNEL) or necrosis (trypan blue staining). To investigate whether short term NTX's action on cell proliferation was transduced by the p16 and/or p21 CKI pathways, siRNAs for p16 and p21 were utilized in combination with short term NTX. Silencing of p16 and/or p21 blocked the inhibitory action of short term NTX, indicating a mechanism directed toward the CKI pathways. Independent of immune function, short term opioid receptor antagonism has been demonstrated to inhibit cancer cell proliferation by upregulation of the OGF-OGFr axis. Upregulation of the OGF-OGFr axis by a brief opioid receptor blockade may provide a novel target for biotherapy of cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4153.
Abstract 4162: Gal4/UAS-mediated pancreatic tumorigenesis in zebrafish
Authors: Liu Shu , Parsons Michael J. , Davison Jon M. , Leach Steven D.
Published: February 2011

Pancreatic cancer remains the 4th deadliest cancer in the United States in 2009. The 5-year survival rate for all stages remains at 5%, the lowest rate among major malignancies. Lack of methods for early detection, as well as ineffective treatments, contribute to the poor outcome of the disease. Over 90% of the pancreatic cancers are pancreatic ductal adenocarcinomas (PDACs). Mutations in the KRAS gene were found in majority of the PDAC lesions and played critical role in the initiation of tumorigenesis as demonstrated in transgenic mouse models. Recent pancreatic cancer genome sequencing results have highlighted the major pathways that can be altered in PDACs. The study also produced a list of novel oncogene candidates that may play parts in the rise of pancreatic cancer. To facilitate the evaluation of these candidate oncogenes’ function in pancreatic cancer in vivo, we developed zebrafish models that can overexpress transgenes specifically in pancreas, alone or in the presence of a human KRAS mutant transgene. The transgenes were introduced into zebrafish via Tol2 transposon elements. The Gal4/UAS transcriptional regulatory system was utilized to ensure targeted expressions of multiple transgenes in the pancreas, controlled by the regulatory element of the Ptf1a gene, a transcription factor critical for pancreas development. Pancreas-specific expression of the mutant KRAS transgene alone in zebrafish can give rise to pancreatic cancers similar to those observed in humans. We now have the opportunity to functionally assess a wide variety of genetic lesions for their ability to modify pancreatic cancer initiation and/or progression, achieving a level of throughput not technically feasible in the mouse.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4162.
Abstract 4163: JNK2 reduction increases intestinal tumor susceptibility in Apc+/− mice
Authors: Bi Xiuli , Hu Dong , Pohl Nicole M. , Yin Zhinan , Yang Wancai
Published: February 2011

Previous studies have shown distinct functions between c-jun NH2 terminal kinases (JNK) 1 and 2. Indeed, loss of JNK1 causes spontaneous intestinal tumor formation in mice (Tong et al, Am. J. Pathol, 2007; Hu et al, Carcinogenesis, 2008), but loss of JNK2 does not have any phenotype in intestine, although in vitro studies have shown that JNK2, like JNK1, interacts with and inhibits β-catenin signaling, in which GSK3β and proteasome pathway play a critical role (Hu et al, LoS One, 2009). To determine the role of genetic interaction between JNK2 and beta-catenin in intestinal tumorigenesis and to elucidate JNK2-mediated intestinal carcinogenesis, we crossed the JNK2−/−mouse with Apc+/− mice that carry inactivated Apc allele and develops intestinal tumor due to beta-catenin activation and nuclear accumulation. We found that the introduction of mutant JNK2 into Apc+/− did not increase intestinal tumorigenesis when the mice were fed a defined AIN-76A control diet. However, JNK2 reduction significantly increased animal body weight in the Apc+/−/JNK2+/− and Apc+/−/JNK2−/− mice. Interestingly, JNK2 reduction was synergistic with a Western-style high-risk diet (high fat and phosphate, low calcium and vitamin D)(WD) to accelerate intestinal tumorigenesis. Tumor number was significantly increased from 1.89 (AIN-76A diet) to 3.56 (WD) in the Apc+/−/JNK2+/− mice (P<0.01) and to 4.14 (WD) in the Apc+/−/JNK2−/− mice from 1.92 (AIN-76A diet) (P<0.01), even though there was a slight increase of tumor formation in JNK2 wild-type mice (Apc+/−/JNK2+/+) by such a diet (1.55 v.s. 2.0, p=0.4). Therefore, we concluded that JNK2 might have function in controlling fat metabolism, and that loss of JNK2 increases the risk of obesity, the latter synergists with high-fat diet to increase intestinal tumor susceptibility. This data strongly suggests the important role of JNK2 in intestinal carcinogenesis and the essential of dietary manipulation for cancer prevention in the population if their JNK2 is inactivated. The underlying mechanisms are under investigation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4163.
Abstract 4164: HBx induced IGF-II expression is essential for hepatic dysplasia formation in mice
Authors: Ha Hye-lin , Shin Hye–Jun , Yu Dae-Yuel
Published: February 2011

HCC is one of the most common cancers in the world. Chronic infection with HBV considered as a major risk factor. Among the four proteins from the HBV genome, HBx is a multifunctional regulatory protein which has been reported to be linked with hepatocellular carcinogenesis. We reported HBx induces liver cancer in transgenic mice.

Among the multistep progress of liver carcinogenesis, enlargement (dysplasia) of the liver considered most critical point of this progression. HBx mice showed dysplasia within 4 weeks, dysplastic nodules and HCA after 6 months and HCC after 9 months of age. In this point we found out different expression pattern of IGF-II in HBx liver compare to wild type. In general this protein hormone predominantly active in fetal liver is strictly down-regulated shortly after birth. HBx mice have an increased liver/body weight ratio as well as cyclinD1 expression compared to wild type mice after 2 weeks. At that point, highly expressed IGF-II in wild mice started to decrease but in HBx mice the IGF-II expression still remained high level. The HBx-HepG2 cell line also has a higher expressed IGF-II level than the Mock-HepG2 cell line. Like in vivo data, the HBx cell line showed fast growth and increased cyclinD1 expression levels. Furthermore we found that abnormal cell growth was inhibited by neutralizing secreted IGF-II in vitro system with treatment of IGF-II antibody. In conclusion, we suggest IGF-II leads to liver dysfunction via over-proliferation of an HBx damaged hepatocyte.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4164.
Abstract 4166: Targeted expression of the Pleomorphic adenoma gene 1 in the pancreas of mice leads to islet hyperplasia
Authors: Declercq Jeroen , Kumar Anujith , Diepen Janna Van , Vroegrijk Irene , Gysemans Conny , Pietro Caterina Di , Voshol Peter , Mathieu Chantal , Ectors Nadine , Van de Ven Wim , Vefaillie Catherine
Published: February 2011

Pleomorphic adenoma gene 1 (PLAG1) encodes a transcription factor that is involved in several human neoplasias such as in pleomorphic adenomas of the salivary glands, lipoblastoma, hepatoblastoma and AML. The oncogenic capacity of PLAG1 has been further demonstrated in vivo in PLAG1 transgenic mice, in which the activation of the PLAG1 expression is Cre-mediated. Targeted expression of PLAG1 in the salivary gland or in the mammary gland leads to pleomorphic adenomas of the salivary glands and adenomyoepitheliomatous lesions of the mammary glands, respectively. Here, we investigated the impact of targeted PLAG1 expression in the pancreas. Pdx1-Cre-mediated targeted expression of PLAG1 in the whole pancreas, leads to islet hyperplasia. Although there was initially a balanced increase in the insulin, somatostatin and glucagon content of the pancreas, in older P1-Pdx1Cre mice the production of insulin and somatostatin far exceeded that of glucagon (82%, 275% and 9% respectively in 1 year old P1-Pdx1Cre mice as compared to littermate Pdx1-Cre mice). This was associated with hyperinsulinemia and an increased ability to dispose glucose in P1-Pdx1Cre mice, and progressive development of severe hypoglycaemia. Only 1/10 one year old mice developed a pancreatic tumor. We also investigated the consequence of overexpression of PLAG1 only in β-cells by using RIP-Cre mice. Similar to P1-Pdx1Cre mice, P1-RIPCre mice displayed islet hyperplasia, hyperinsulinemia and an increased ability to dispose glucose. Nevertheless, in contrast to P1-Pdx1Cre mice, P1-RIPCre mice stay normoglycemic and compensate for the hyperinsulinemia by the development of hepatic insulin resistance. In conclusion our data demonstrates that targeted expression of PLAG1 in the pancreas leads to islet hyperplasia rather than to pancreatic tumor formation.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4166.
Abstract 4178: N-cadherin haploinsufficiency prolongs survival in a genetically engineered mouse model of pancreatic cancer
Authors: Su Yanrong , Li Jifen , Griffith Leeanne , Radice Glenn
Published: February 2011

Cadherin switching (E-cadherin to N-cadherin) associated with epithelial-mesenchymal transition is implicated in the transition from benign tumors to invasive, malignant cancer and the subsequent metastatic dissemination of tumor cells. Although N-cadherin upregulation has been observed in many cancer types including pancreatic ductal adenocarcinoma (PDA), the importance of this cell adhesion receptor in disease progression is not well understood. The concomitant expression of oncogenic KrasG12D and a mutant Trp53R172H allele in the developing murine pancreas results in metastatic PDA that recapitulates the cognate features of human pancreatic cancer providing an excellent animal model to identify genetic changes required for tumor progression. Here we determine the consequences of genetically blocking the switch to N-cadherin in a mouse model of PDA. Two independent N-cadherin mutant alleles were used in these genetic studies. Remarkably, pancreatic cancer mice heterozygous for N-cadherin (n=17) survived 25% longer (177 vs. 142 days, p <0.05) than N-cadherin wild-type mice (n=16). The survival benefit may be due, at least in part, to increased cell death in the N-cadherin heterozygous tumors. Furthermore, cell death was also increased in tumor cell lines isolated from the N-cadherin heterozygous tumors demonstrating a cell-autonomous apoptotic phenotype. Interestingly, tumors lacking N-cadherin (i.e. null) did not show a survival benefit possibly due to compensation by an alternative metastatic promoting molecule(s). In conclusion, our genetic studies indicate that interfering with N-cadherin-mediated adhesion/signaling delays tumor progression, at least in part, by promoting tumor cell death, thus providing further evidence that N-cadherin antagonists may have therapeutic potential for the treatment of pancreatic cancer in humans.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4178.
Abstract 4179: Quercetin inhibits growth of pancreatic cancer in vitro and in vivo
Authors: Angst Eliane , Park Jenny L. , Moro Aune , Lu Qing-Yi , King Jonathan C. , Go Vay Liang W. , Reber Howard A. , Eibl Guido , Hines O. Joe
Published: February 2011

Background and Aim: Polyphenols are common constituents of many fruits and vegetables, and have recently gained interest as potential therapeutic agents for various human malignancies. The most abundant of these polyphenols are flavonoids - quercetin being the most ubiquitous. The aim of this study was to assess the growth-inhibitory and anti-angiogenic properties of quercetin on pancreatic cancer (PaCa) cells in vitro and in vivo. Methods and Results: Using cell count and BrdU assay, we measured a dose-dependent inhibition of cell proliferation by quercetin (0-75µM) for MIA PaCa-2 (MP) and BxPC-3 (Bx) cells (p<0.001 at 30µM). Quercetin (10 µM) increased apoptosis by 22% and 8% in MP and Bx, respectively (p<0.05). Further, quercetin (10 µM) inhibited mRNA and protein expression of the pro-angiogenic chemokine CXCL8 (p<0.05) in both cell lines as measured by ELISA and real-time PCR. Conditioned media from PaCa cells treated with 50 µM quercetin reduced endothelial cell tube formation by ∼50% compared to control (p<0.05). A feeding study was conducted utilizing an orthotopic xenograft model of PaCa in nude mice. Luciferase-expressing MP (3×106) cells were injected subcutaneously into the flank of donor mice. After four weeks, 1 mm3 donor tumor sections were transplanted into the pancreatic tail of recipient mice. These animals were randomly allocated to control diet or 1% quercetin diet groups utilizing the AIN93G-based diet (n=12 per group). In addition, 6 mice from each group were injected weekly with gemcitabine (120 mg/kg mouse, i.p.). Animals were imaged weekly after tumor implantation until sacrifice at 6 weeks. Light emission was reduced in animals receiving quercetin, gemcitabine, and both treatments by 12%, 8%, and 16%, respectively at day 24. Furthermore, CXCL8 serum levels were significantly reduced in the quercetin/gemcitabine group (p<0.001). Using HPLC analysis, quercetin and its metabolite, isorhamnetin, were detected in tumor tissues (2.6 ± 2.7 and 2.1 ± 0.9 nmol/gram, respectively). Conclusion: Quercetin significantly inhibits the growth of PaCa in vitro and in an orthotopic murine model. PaCa growth was inhibited utilizing a bioavailable quercetin preparation in the diet. Quercetin may, in part, be acting through a reduction of angiogenesis. From this data we surmise that quercetin may be useful as an adjunct to current treatment for pancreatic cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4179.
Abstract 418: Introducing different genetic elements into human mesenchymal stem cells to transform into osteosarcoma
Authors: Austin-Page Lukas , Thein Sajida , Lam Matthrew , Park Amy , Li Nan , Yang Rui , Gorlick Richard
Published: February 2011

Despite Osteosarcoma (OS) being the most common primary malignant bone tumor in children and young adults, little clarity exists in where the cell of origin exists in the pathway of differentiation between a human mesenchymal stem cell (hMSC) and a mature osteoblast. It is similarly unclear the primary genetic alterations which initiate oteosarcoma formation in patients. Since high grade OS usually demonstrates histologic variability, and the potential for multilineage differentiation, along with some direct experimental observations some consider hMSC as the cell of origin of OS whereas others believe the pre-osteoblast or osteoblast to be the mot likely cell of origin. Our previous studies have shown that serially introducing 3 different viral constructs, hTERT, SV40TAg, and H-RAS, transforms hMSC into 2 distinct sarcomas cell lines, but not OS. This may be attributable to the fact that oncogenic H-Ras does not seem to play a direct role in the pathogenesis of OS. Therefore, the MSC-TS cell line, hMSC transfected with hTERT and SV40TAg can be used as a platform for further analysis of the impact of genetic transformation of hMSCs. The MSC-TS cells were transformed with lentivirus containing human β-catenin (β), and IGF-1R (I)

, separately. Both β-catenin and IGF-1R have been suggested to be involved in the pathogenesis of OS due to their proliferative and/or anti-apoptotic effects. Drug resistant colonies were picked up 21 days after selection to obtain stably transfected cell lines, MSC-TS-β, and MSC-TS-I, respectively. Western blots were carried out to detect the protein of β-catenin and IGF-1R, respectively. Clones, which have highest, lowest, and intermediate protein levels were selected, and comparison between the lines and characterizations are completed and underway. Further characterization includes: 1) Soft Agar Assays , 2)Subcutaneous/Orthotopic Tumorigenicity Assays,3) Routine histological examination and also staining with immunohistochemical markers, 4) Microarray Analysis, 5) Spectral Karyotyping / Comparative Genomic Hybridization, 6) Motility and Migration Assays, and 7) Differentiation Assays.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 418.