bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 639: Inhibition of HER2 signaling using targeted therapeutics in breast cancer cell lines
Authors: Korkola James , Bayani Nora , Cooper Brian , Heiser Laura M. , Spellman Paul T. , Feiler Heidi S. , Wooster Richard , Gray Joe W.
Published: June 2012

ERBB2/HER2 amplification occurs in 20-30% of all breast cancers and is associated with an aggressive clinical behavior. Therapeutics that target HER2 offer a modest survival benefit when given as monotherapy. Recent studies indicate that in the case of the HER2/EGFR inhibitor lapatinib, there is an initial down-regulation of the PI3K-AKT signaling pathway followed by reactivation of the pathway through p-HER3. Inhibition with much higher doses of lapatinib in vitro prevents the reactivation of the pathway, implying that more effective inhibition of HER2 could lead to more durable responses. To explore this phenomenon more fully, we examined response to lapatinib in a panel of 22 HER2 amplified breast cancer cell lines. Of the 22, there were 5 that were resistant to lapatinib, failing to reach 50% growth inhibition (GI50) even at doses as high as 5 uM. These lines have also been reported to be resistant to trastuzumab. However, all the cell lines in the panel were sensitive to BIBW-2992, which is an irreversible EGFR/ERBB2 inhibitor, with GI50 values in the nanomolar range. Initial western blot analysis indicated that in response to BIBW-2992, there was down-regulation of p-AKT and p-HER3, but that there was recovery of both over time to levels that were comparable to untreated cells. Finally, we observed strong synergistic interactions between BIBW-2992 in combination with a PI3K inhibitor in almost every HER2+cell line tested. These results suggest that even in the setting of resistance to lapatinib and trastuzumab, all HER2 amplified cell lines in our panel are still addicted to HER2 signaling, and that more efficient inhibition of HER2 can result in significant inhibition of growth and loss of viability in HER2 positive breast cancer cell lines. BIBW-2992 was effective against HER2 amplified cells even though inhibition of p-AKT and p-HER3 was not complete. However, addition of PI3K pathway inhibitors resulted in strong synergism in combination with BIBW-2992. These studies set the stage for clinical investigations into the efficacy of such irreversible HER2 inhibitors in patients with HER2 positive breast 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 639. doi:10.1158/1538-7445.AM2011-639
Abstract 643: Targeting cancer with a selective ATP-mimetic inhibitor of polo like kinase-2
Authors: Jatiani Shashidhar S. , Cosenza Stephen C. , Billa Vinay K. , Reddy M.V. Ramana , Reddy E Premkumar
Published: June 2012

The Polo like kinases play key roles in mitosis. While the upregulation of PLK1 in cancer is well documented and PLK3 has been demonstrated to be a tumor suppressor, little is known about the oncogenic significance of PLK2. PLK2 kinase activity is essential for centriolar duplication and is also believed to play a regulatory role in the survival pathway by physically stabilizing the TSC1/2 complex in tumor cells under hypoxic conditions. Protein tyrosine hyper-phosphorylation and the upregulation of serine/threonine kinases associated with cell cycle progression have been linked to the etiology of cancer. In our attempt to develop ATP-mimetic compounds that are cytotoxic against a panel of cancer cell lines, we identified several sulfonyl pyridopyrimidines that possess nanomolar activity. The most potent of these compounds, ON1231320, was found to be a specific PLK2 inhibitor when profiled against a panel of 288 wild-type, 55 mutant and 12 special kinases. Further in vitro testing revealed that ON1231320 is a selective inhibitor of PLK2 with no inhibitory activity against PLK1, PLK3 and PLK4. Confirmation of this interaction and its selectivity was obtained from pull down studies using lysates of tumor cells and a biotinylated analog of ON1231320. The cytotoxic effect of the drug is mediated by apoptosis as evidenced by the induction of Caspase 3/7 activity and by the cleavage of PARP in a dose dependent manner. ON1231320 affects cell cycle progression by blocking tumor cells in the G2/M phase however it does not affect normal human fibroblasts. ON123120 exhibits an excellent safety profile with no overt signs of toxicity, no loss of body weight and 100% survival in mice given a single peritoneal dose of 200 mg/kg. Our ongoing efforts include efficacy studies in nude mouse models, identification of the structural determinants of the interaction between ON1231320 and PLK2 by computational and crystallographic methods and the identification of novel PLK2 substrates to elucidate its role in cancer biology.

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 643. doi:10.1158/1538-7445.AM2011-643
Abstract 649: Mirk/dyrk1B kinase inactivation increases gemcitabine efficacy towards quiescent pancreatic cancer cells
Authors: Ewton Daina Z. , Vilenchik Maria , Polonskaia Ann , Hoffman Ann , Zipf Karen , Boylan John F. , Heimbrook Dave , Hu Jing , Friedman Eileen A.
Published: June 2012

A major problem in the treatment of cancer is the presence of residual quiescent cancer cells that are relatively insensitive to most chemotherapeutic drugs and radiation. Such cancer cells are believed to cause tumor recurrence when they re-enter the cell cycle under favorable conditions of the microenvironment. Gemcitabine is the drug most often used for treatment of pancreatic cancer, but the response rate is less than 20%. Since stratagems to increase delivery of gemcitabine by inhibiting stromal hedgehog signaling have shown some success, identifying methods to increase gemcitabine efficacy is timely. The serine/threonine kinase Mirk/dyrk1B is expressed at very low levels in most normal tissues, but is overexpressed or amplified in most pancreatic cancers. Mirk maintains pancreatic cancer cells in a viable quiescent state through its transcriptional co-activator functions for a set of antioxidant proteins that decrease intracellular levels of reactive oxygen species (Cancer Res 2009;69:3317-24). We now show that Panc1 and SU86.86 pancreatic cancer cells undergo a reversible quiescent arrest in G0 through activation of Chk2 and autophagy to reduce ribosomal RNA levels, with little apoptosis or senescence. Depletion of Mirk/dyrk1B by either induced shRNA or transfected RNAi duplexes or inhibition of Mirk's kinase activity by a small molecule inhibitor increased DNA damage in quiescent pancreatic cancer cells, either grown attached to petri dishes or in small clumps in suspension as a model for ascites. DNA damage was detected by increased phosphorylation of the histone protein H2AX and by an increase in S phase checkpoints after the quiescent cells re-entered cycle in response to serum mitogens. Pharmacological inhibition of Mirk kinase induced a dose-dependent cleavage of the apoptotic marker proteins PARP and caspase 3, reduced Akt activation, and led to cleavage of the autophagic marker protein LC3. Pretreatment with the Mirk kinase inhibitor increased tumor cell kill 5-fold by short exposures to marginally toxic gemcitabine concentrations. In prior studies Mirk depletion had no detectable effect on the viability of normal diploid fibroblasts (Cancer Res 2007;67:7247-55), while other investigators have shown that embryonic knockout was not lethal (Leder et al, Biochem J 2003;372:881-8). Thus the small molecule Mirk/dyrk1B kinase inhibitor sensitized pancreatic cancer cells to low gemcitabine concentrations, while knockout or depletion of Mirk/dyrk1B had no detectable effect on normal tissue.

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 649. doi:10.1158/1538-7445.AM2011-649
Abstract 662: Evaluation of the anti-cancer effects of the tumor selective vascular disruption agent BNC105 in preclinical renal cancer models
Authors: Lavranos Tina C. , Leske Annabell F. , Beaumont Donna M. , Gasic Jelena , Kremmidiotis Gabriel
Published: June 2012

The disruption of blood vessels that feed tumors represents one of the most promising therapeutic strategies for treating cancer. BNC105 is a tubulin targeting dual acting vascular disruption agent with cytotoxic ability in solid tumors. A prodrug formulation, BNC105P, is under evaluation in phase II clinical trials for malignant mesothelioma and metastatic renal cell carcinoma as a 10min IV infusion. We have conducted a number of preclinical evaluations that provide a strong rationale for combining BNC105P treatment with agents that target mTOR signalling. BNC105 activity operates through the selective disruption of tumor blood vessels. Over 95% of blood flow is disrupted in tumors grown in xenograft animal models using human cancer cell lines representing a number of different cancer types, including breast, lung, colon, brain and mesothelioma. In the renal cancer setting we evaluated the anti-cancer effects of BNC105 in a Caki-1 xenograft model and a syngeneic murine renal cancer orthotopic model. Animals carrying Caki-1 solid tumors were treated with a single dose of BNC105P. Disruption of blood flow within the tumors was observed as early as 3 hr post-treatment. Similarly, blood flow disruption was seen in mice carrying solid tumors orthotopically inoculated in the kidney capsule using the RENCA mouse renal cancer cell line. Interestingly, BNC105P treatment also caused blood flow disruption in lung metastatic lesions seen in a number of the animals orthotopically inoculated with RENCA. The vasculature in non-cancerous portions of the kidney and all normal tissues examined, remained intact. Tumor re-vascularisation following BNC105P treatment was observed 2 days following administration. Immunohistochemical and Western blot analyses in BNC105P treated tumors revealed that a number of proteins involved in the mTOR signalling pathway exhibit expression changes consistent with activation of this pathway possibly a result of BNC105P induced tumor hypoxia. Up-regulation in phosphorylated mTOR, Hif1a and down-regulation of phosphorylated 4EBP1 were observed. The consequences of targeting mTOR in combination with BNC105P are currently under investigation in these preclinical models.

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 662. doi:10.1158/1538-7445.AM2011-662
Abstract 67: High-throughput cellular assays for interrogating epigenetic histone modifications
Authors: Hallis Tina , Bi Kun , Machleidt Thomas , Hermanson Spencer , Wilkinson Jennifer , Wetter Justin
Published: June 2012

Epigenetic histone modifications such as, methylation, acetylation, and phosphorylation play important roles in regulating the structure and functions of histones which in turn regulate essential gene expression. Dysregulation of the enzymes responsible for these modifications has been linked to cancer and many other diseases. In particular, histone deacetylases (HDACs) and histone methyl transferase (HMTs) have been a recent focus for drug discovery. However, detection of these enzymatic activities in an HTS cell-based format has remained challenging, especially since many of these enzymes may require the formation of protein complexes for relevant activity. To help overcome these challenges, we have developed and validated high-throughput compatible LanthaScreen® cellular assays for the analysis of histone methylation, acetylation, and phoshorylation, in cell backgrounds of interest. The acetylation, methylation and phosphorylation of H3 at specific residues can be detected with terbium-labeled antibodies against the specific site and modification. The expression of GFP-H3 substrate allows for detection of antibody binding in cell lysates via TR-FRET between the terbium and GFP. Since the actual modification takes place in the intact cell, native protein complexes that may be important for enzyme function are preserved. Together, these assays enable interrogation of multiple enzymatic activities responsible for epigenetic modifications of challenging histone targets

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 67. doi:10.1158/1538-7445.AM2011-67
Abstract 675: Effects of PBN and OKN007 in rodent glioma models assessed by 1H MR spectroscopy
Authors: He Ting , Doblas Sabrina , Saunders Debra , Casteel Rebba , Floyd Robert A. , Towner Rheal A.
Published: June 2012

Gliomas are the most common and lethal primary brain tumors in the adult, with a median survival time of 12 to 15 months for grade IV gliomas, glioblastoma multiforme. PBN and OKN007 are nitrones which have demonstrated beneficial effects in many aging diseases. In this study, we evaluated the anti-tumor effects of PBN or OKN007 in several rodent glioma models (C6, RG2, and GL261) by assessing metabolite alterations with magnetic resonance spectroscopy (MRS). PBN or OKN007 was administered in drinking water prior to or post tumor formation. MRI and short echo time, single-voxel point-resolved spectroscopy (PRESS) were obtained to assess tumor morphology and metabolites, respectively, following therapy. Major metabolite ratios (Choline, N-acetyl aspartate (NAA), Lipid (methylene), and Lipid (methyl), all compared to Creatine) were assessed. Nitrones induced tumor metabolism changes that resulted in restoring major metabolite ratios close to their normal levels, in the glioma regression phase. Nitrones treatment decreased the Lipid (methylene) to Creatine ratio significantly, which can be a useful marker for evaluation of the efficacy of the treatments, and was found to be related with the reduction of necrosis. OKN007 was more effective than PBN when administered post-tumor formation in the C6 glioma model. OKN007 was able to inhibit angiogenic factors (VEGF, bFGF and HGF) and induce apoptosis (indicated by an increase of the apoptotic marker m30) in C6 gliomas, which might be potential mechanisms that are involved in the anti-glioma effects. In conclusion, OKN007 and PBN are effective in inhibiting the growth of several experimental gliomas and may be considered as potential therapeutics for a clinical trial in human gliomas.

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 675. doi:10.1158/1538-7445.AM2011-675
Abstract 681: PTX-008, a novel galectin-1 targeting agent, improves the effects of cytotoxic drugs in colon cancer cells
Authors: Astorgues-Xerri Lucile , Serova Maria , Riveiro Maria E. , Cvitkovic Esteban , Noel Kay , Raymond Eric , Faivre Sandrine
Published: June 2012

Background: Addition of targeted therapies has been shown to potentiate the activity of cytotoxic drugs. Compounds with innovative mechanism of action such as PTX-008, a novel galectin-1 targeting agent, might represent an interesting candidate for cytotoxic-based combinations. We explored the effect of PTX-008 in combination with several drugs commonly used in cancer chemotherapy such as cisplatin, oxaliplatin, 5-FU, 5’-DFUR, taxotere and gemcitabine in colon cancer cell lines.

Methods: Antiproliferative effects were evaluated in two human colon (Colo205 and HT29) cancer cell lines by MTT assay. Effects of different schedules of exposure of PTX-008-based combinations were determined by median effect plot analysis (Chou & Talalay).

Results: PTX-008 induced significant growth inhibition at concentrations below 10 µM in HT29 and Colo205 cancer cell lines. Combination of PTX-008 with cisplatin and oxaliplatin was additive or synergistic in both Colo205 and HT29 cells for all administration schedules. Combination of PTX-008 with antimetabolites 5-FU and 5’-DFUR was synergistic for sequential administration schedule, especially when PTX-008 was given prior to 5-FU and 5’-DFUR in colon cancer cells. Simultaneous exposure to these drugs induced additive effects. Combination with taxotere was at least additive using all schedules of administration. PTX-008/gemcitabine combination were highly synergistic in Colo205 and HT29 cells for sequential schedules and additive/synergistic for simultaneous administration.

Conclusions: Combination studies showed that PTX-008 displays at least additive, often synergistic effects with a wide range of cytotoxic drugs (including cisplatin, oxaliplatin, 5-FU, gemcitabine and taxotere). Those data provide preliminary evidences that inhibition of galectin-1 using PTX-008 may be an attractive approach for combinations with anticancer drugs in clinical trials.

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 681. doi:10.1158/1538-7445.AM2011-681
Abstract 687: Enhanced expression of multidrug resistance (MDR) protein in malignant pleural mesothelioma (MPM) patients is possibly achieved via Osteoponin, CD44 variants and p-AKT expression
Authors: Bahnassy Abeer A. , Zekri Abdel-Rahman N. , Eid Salem , Abdel-Rahman Abdel-Rahman M. , Mahmoud Moustafa R. , Gaafar Rabab M.
Published: June 2012

Malignant pleural mesothelioma (MPM) is an aggressive disease, which is discovered late. It is resistant to chemotherapy and thus shows a dismal prognosis. Recently, in vitro studies demonstrated that osteoponin (is involved in the development of MPM via induction of multidrug resistance through unidentified mechanism(s).

Tissue and serum samples from 55 Egyptian cases of MPM were assessed for OPN, MDR, p-AKT and CD44 proteins and RNA by ELISA, immunohistochemistry (IHC) and RT-PCR. Normal pleural samples (20) were used as a control. Results were correlated with standard prognostic factors and survival.

In tissues, OPN, CD44, MDR and p-AKT protein overexpression was detected in 39, 26, 37 and 25 cases; respectively whereas RNA expression was reported in 31, 24, 30, 24 cases; respectively. In sera, OPN, CD44, MDR and p-AKT proteins were found in 36, 20, 40, 28 cases; respectively. There was a significant correlation between OPN expression and other markers, at the protein and RNA levels (p<0.05). The concordance between OPN and MDR expression was the highest (98%). Control samples were negative for the studied markers except for 4 cases that showed faint focal expression of p-AKT and CD44 proteins. OS was significantly associated with performance status, increased expression of OPN, MDR and CD44 in univariate and multivariate analysis (p<0.01). A borderline significance was reported between poor response to treatment and increased expression of OPN, CD44, MDR either singly (p=0.063) or combined (p=0.045)

In conclusion, OPN, CD44, and p-AKT could be used as poor prognostic markers in MPM being associated with reduced OS rates and poor response to treatment. This could be achieved via enhanced MDR expression with subsequent resistance to chemotherapy.

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 687. doi:10.1158/1538-7445.AM2011-687
Abstract 698: β-arrestin-1 as nuclear signalling element essential for endothelin A receptor-induced epithelial to mesenchymal transition and chemoresistance
Authors: Rosanò Laura , Cianfrocca Roberta , Castro Valeriana Di , Spinella Francesca , Nicotra Maria Rita , Natali Pier Giorgio , Bagnato Anna
Published: June 2012

The onset of chemoresistance is the major challenge in epithelial ovarian cancer (EOC). Emerging evidence suggests molecular and phenotypic association between chemoresistance and epithelial-mesenchymal transition (EMT) in cancer. We recently demonstrated that β-arrestin-1, scaffold and signaling protein of G-protein-coupled receptors, is recruited to endothelin A receptor (ETAR) to form molecular signaling complexes which concur to activate β-catenin transcriptional activity, EMT and metastasis. Here, by using biochemical and imaging techniques, we provide evidence that in EOC cells, ETAR activation by endothelin-1 (ET-1) induces nuclear translocation of β-arrestin-1 and its association with β-catenin. ChIP assays demonstrated that following this translocation, β-arrestin-1 was selectively enriched at specific promoters of β-catenin target genes, such as ET-1, MMP-2, cyclin D1. Further experiments showed that ET-1 promotes the nuclear association between histone acetyltransferase p300 and β-arrestin and the recruitment of p300 on these promoters, resulting in H3 and H4 histone acetylation and enhanced gene transcription. In chemoresistant EOC cells, ET-1/ETAR axis is overexpressed associated with EMT marker expression. Most importantly, the transcriptional activity of β-catenin was upregulated in chemoresistant EOC cells, associated with enhanced ETAR and β-arrestin-1 expression, providing molecular evidence that the ETAR/β-arrestin-1-mediated EMT signaling in EOC can represent a “salvage pathway” occurring during chemoresistance development. Interestingly, ETAR blockade with zibotentan, a specific ETAR antagonist, or its silencing with siRNA, abrogates both the engagement of β-arrestin-1 in the nucleus and the interplay between ET-1 and the β-catenin in controlling gene transcription. In vivo, zibotentan inhibited tumor growth of sensitive and resistant EOC xenografts, associated with reduction of EMT markers, and sensitized to chemotherapy. Analysis of EOC human tissues revealed that ETAR is overexpressed in resistant tumors and is associated with EMT phenotype. Our data provide the first evidence that blockade of ETAR/β-arrestin-1-driven EMT can overcome chemoresistance and inhibit tumor progression, improving the outcome of EOC patients’ treatment. Supported by AIRC

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 698. doi:10.1158/1538-7445.AM2011-698
Abstract 699: HSP70 specifically binds to 15 bp deletion mutant EGFR and modulates sensitivity to EGFR-TKI
Authors: Kadofuku Tsuyoki , Kanome Tomoko , Ichihashi Yoko , Yamaoka Toshimitsu , Hirose Takashi , Adachi Mitsuru , Saijo Nagahiro , Ohmori Tohru
Published: June 2012

[Purpose] Non small lung cancer (NSCLC) cells that expressed mutant EGFR are more sensitive to EGFR-tyrosine kinase inhibitors (EGFR-TKIs) than that expressed wild type EGFR. Recent clinical trials revealed that more than 70% NSCLC patients that expressed mutant EGFR showed sensitivity to EGFR-TKIs, such as gefitinib and erlotinib. To elucidate the mechanism of this hypersensitivity, we explored the difference of EGFR-binding proteins between wild type EGFR and 15 bp deletion mutant EGFR using respective stable transfectants.

[Methods] Wild type EGFR and a 15 bp deletion mutant EGFR plasmids were transfected into HEK293 cells. The stable transfectant cells were established, and were designated 293_pEGFR and 293_pΔ15, respectively. Cells were incubated in a medium with/without 10 ng/ml of TGFα for 1 h and lysed. Cell lysate was precleared by centrifugation at 15,000 rpm for 15 min. The supernatant was mixed with a polyclonal EGFR antibody for 1 h and EGFR was immunoprecipitated by Protein A-Sepharose. After adequate washing, coprecipitated proteins that bound to EGFR were eluted and separated by 2D-PAGE (Immobiline DryStrip (pH3-10 NL, 7 cm) and 10% SDS-PAGE). Proteins were visualized by silver staining and identified by LC-MS/MS. HSP70 siRNA was transfected into the cells by lipofection method. Sensitivity to gefitinib was measured by MTT assay. EGFR binding affinity to gefitinib was measured using [14C] gefitinib.

[Results and Discussion] We detected several EGFR binding proteins. Among these proteins, one candidate (lesser binding to wild type EGFR than the mutant EGFR) was identified as HSP 70 by LC-MS/MS. The total amount of HSP70 protein was not different between 293_pEGFR and293_pΔ15 cells. A specific binding of this protein to the mutant EGFR was confirmed by Western blotting analysis. This binding was not modulated by EGFR activation. Knockdown of HSP 70 protein by a specific siRNA enhanced gefitinib sensitivity in the mutant EGFR transfectant.

These results suggest that HSP 70 may decrease sensitivity to EGFR-TKIs in the cells that expressed 15 bp deletion mutant EGFR. There is a possibility that HSP70 overexpression might be a newly resistant mechanism to EGFR-TKIs in NSCLC patients that expressed mutant EGFR.

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 699. doi:10.1158/1538-7445.AM2011-699
Abstract 70: A role for the chromatin remodeling protein CHD3 in ovarian cancer therapy response
Authors: Cutter Noelle , Banerjee Nila , Vigliotti Michele , Janevski Angel , Levine Douglas , Dimitrova Nevenka , Lucito Robert
Published: June 2012

Carboplatin and cisplatin are chemotherapeutic agents that are used extensively for treating epithelial ovarian cancer. These drugs can be highly effective, yet tumors are frequently refractory to treatment or become resistant upon tumor relapse. Epigenetic silencing, particularly at promoter regions of genes regulates important cell function and has been associated with all stages of tumor formation and progression and may contribute to therapy response. We analyzed the epigenome of 50 primary ovarian tumors and 12 normal ovarian samples using an array based method previously developed in our lab and associated Affymetrix U133 expression data. We then identified gene candidates that segregate patients based on platinum sensitivity and patient survival. These candidates were then pooled into a genome-wide RNAi-based screen where we validated a gene encoding a chromatin remodeling protein, CHD3, a member of the Mi-2 NuRD complex, and show that it is linked to chemoresistance. CHD3 is silenced through an epigenetic mechanism in both ovarian cancer cell lines and primary ovarian tumors. When ovarian cancer cell lines that are transcriptionally silenced for CHD3 are challenged with carboplatin they display a striking slow growth phenotype as well as increased resistance to the chemotherapy drugs carboplatin and cisplatin. Taken together, we provide the first evidence for a role for CHD3 as an important mediator of chemoresistance in ovarian cancer. Furthermore, CHD3 might represent a response predictor and potential therapeutic target for predicting chemoresistance in this 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 70. doi:10.1158/1538-7445.AM2011-70
Abstract 707: Acquisition of chemoresistance and epithelial to mesenchymal phenotype is linked with activation of the endothelin A receptor pathway in ovarian carcinoma cells
Authors: Cianfrocca Roberta , Rosanò Laura , Castro Valeriana Di , Spinella Francesca , Nicotra Maria Rita , Lucidi Alessandro , Ferrandina Gabriella , Natali Pier Giorgio , Bagnato Anna
Published: June 2012

Understanding the molecular mechanisms underlying chemoresistance is of utmost importance for improving the treatment of epithelial ovarian cancer (EOC). Emerging evidence suggests molecular and phenotypic association between chemoresistance and epithelial-mesenchymal transition (EMT) in cancer cells. Endothelin-1 (ET-1) and the endothelin A receptor (ETAR) axis is implicated in the pathobiology of EOC by driving different tumor-promoting effects, including EMT. By employing two cisplatin- and taxol-resistant ovarian cancer cell lines, we demonstrated that in the resistant EOC cells ET-1 and ETAR are upregulated, at both mRNA and protein level, paralleled by enhanced MAPK and AKT phosphorylation and cell proliferation. To assess whether the chemoresistance in A2780 cells was associated with EMT molecular changes, we examined the expression of epithelial and mesenchymal markers. Enhanced Snail, Slug, Twist, N-cadherin and vimentin mRNA expression was observed in resistant cells, associated with a concomitant decrease in E-cadherin expression and promoter activity. Moreover, ET-1 induced a significant induction of Snail promoter activity in resistant cells, and the E-cadherin promoter sequences were detected bound to Snail in a time-dependent manner, as demonstrated by chromatin immunoprecipitation assays, suggesting that the ET-1-dependent and sustained binding of Snail in the E-cadherin promoter might account for EMT and chemoresistant phenotype of these cells. Moreover, the invasiveness and proteinase activity of chemoresistant cells was greater, compared with sensitive cells. Interestingly, ETAR blockade with the specific ETAR antagonist, zibotentan, or its silencing with siRNA, reverted EMT, restored drug sensitivity to cytotoxic-induced apoptosis, and inhibited the invasive potential of resistant cells. In vivo, zibotentan inhibited tumor growth of sensitive and resistant EOC xenografts, and sensitized to chemotherapy. Finally, analysis of EOC tissues, with different responses to chemotherapy, revealed that ETAR is overexpressed in resistant tumors and is associated with EMT marker expression. All these data indicate that blockade of ETAR-driven EMT can overcome chemoresistance in EOC, improving the outcome of EOC patients’ treatment. Supported by AIRC

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 707. doi:10.1158/1538-7445.AM2011-707
Abstract 709: Development of resistance towards artesunate in MDA-MB-231 human breast cancer cells
Authors: Bachmeier Beatrice E. , Killian Peter H. , Kronski Emanuel , Efferth Thomas
Published: June 2012

Breast cancer is the most common cancer and the second leading cause of cancer death in industrialized countries. Systemic treatment of breast cancer is effective at the beginning of therapy. However, after a variable period of time, progression occurs due to therapy resistance. Artesunate (ART), clinically used as anti-malarial agent, has recently revealed remarkable anti-tumor activity offering a role as novel candidate for cancer chemotherapy. We analyzed the anti-tumor effects of ART in metastasizing breast carcinoma in vitro and in vivo. Unlike as expected, ART induced resistance in highly metastatic human breast cancer cells MDA-MB-231. Likewise acquired resistance led to abolishment of apoptosis and cytotoxicity in pre-treated MDA-MB-231 cells. In contrast, ART was more cytotoxic towards the less tumorigenic MDA-MB-468 cells without showing resistance. Unraveling the underlying molecular mechanisms, we found that resistance was induced due to activation of the tumor progression related transcription factors NFκB and AP 1. Thereby transcription, expression and activity of the matrix-degrading enzyme MMP-1, whose function is correlated with increased invasion and metastasis, was up-regulated upon acquisition of resistance. Additionally, activation of the apoptosis-related factor NFκB lead to increased expression of ant-apoptotic bcl2 and reduced expression of pro-apoptotic bax. Application of ART in vivo in a model of xenografted breast cancer showed, that tumors growth was not efficiently abolished as compared to the control drug doxorubicin. Taken together our in vitro and in vivo results correlate well showing for the first time that ART induces resistance in highly metastatic breast 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 709. doi:10.1158/1538-7445.AM2011-709
Abstract 721: Putative molecular mechanism contributing AZD6244 resistance in lung cancer
Authors: Yamaguchi Hirohito , Hsu Jennifer , Ko How-Wen , Wang Ying-Nai , Chang Shih-Shin , Hung Mien-Chie
Published: June 2012

The K-ras proto-oncogene is mutated in about 10-30% of non-small cell lung cancer, and these mutations are responsible for constitutive activation of the mitogen-activated protein kinase (MAPK) signaling pathway, Raf/MEK/ERK. Therefore, several MEK inhibitors are currently under clinical trials to target K-Ras mutant lung cancer. AZD6244 (ARRY-142886) is an orally active and selective inhibitor that specifically targets MEK. We investigated the effect of AZD6244 on NSCLC cell lines to further understand the detailed molecular mechanism contributing to the sensitivity to this drug. Interestingly, two NSCLC cell lines, which have previously been reported as AZD6244-sensitive, exhibited a differential response to MEK inhibition by AZD6244. Specifically, long-term treatment by this drug eventually induced resistance in one of the two cell lines. We analyzed protein expression profile of the Bcl-2 family members which we hypothesized might play a role in the resistance to AZD6244 treatment. We found that while Bim appears to be stabilized in both apoptosis-resistant and -sensitive lung cancer cells, MCl-1 expression is increased in apoptosis-resistant lung cancer cells, suggesting that the increase in anti-apoptotic MCl-1 expression likely contributes to resistance under long-term AZD6244 treatment.

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 721. doi:10.1158/1538-7445.AM2011-721
Abstract 728: Molecular mechanisms of resistance to anti-IGF-1R therapies in Ewing's sarcoma
Authors: Garofalo Cecilia , Manara Maria Cristina , Marino Maria Teresa , Lollini Pier Luigi , Picci Piero , Scotlandi Katia
Published: June 2012

IGF-1R represents a promising therapeutic target, particularly in sarcomas. Identification of patient selection criteria and understanding the potential mechanisms involved in development of resistance will be crucial for an appropriate and successful design of clinical trials.

Here we evaluated efficacy of the anti-IGF-IR monoclonal antibody (HAb) AVE1642, in a panel of sarcoma cell lines and we developed a model of resistance to the anti-IGF-1R human antibody AVE1642 or to the small tyrosine kinase inhibitor (TKI) NVP-AEW541 starting from two highly sensitive Ewing's sarcoma cell lines to identify indicators of intrinsic and adaptive resistance to anti-IGF1R therapies.

Cells made resistant to AVE1642 showed same growth ability either in monolayer and in anchorage-independent condition respect to sensitive cells. Notably, resistant cells retained the same level of sensitivity to doxorubicin, vincristine, and ifosfamide as the parental cell lines. Annotation analysis highlighted that the insulin pathway seems to be significant in resistant cells. In fact, these cells showed higher proliferative response to insulin, in keeping with a switch towards insulin pathways sustaining proliferation and malignancy, rather than metabolism. Molecular characterization demonstrate that enhanced IR-A formation and IGF-II production are heavily involved in native and acquired resistance to IGF targeted therapies. This finding was extended to cells resistant to the small tyrosine kinase inhibitor NVP-AEW541.

We also show that tumors with low IGF-1R:IR-A ratio are unlikely to greatly benefit from anti-IGF-1R therapies and that the efficacy of anti-IGF-1R therapies should be evaluated in relationship to the IR-A:IGF-1R ratio in cancer cells.

Our findings demonstrate a role for IR-A in eliciting intrinsic and adaptive resistance to anti-IGF1R therapies and provide evidences supporting IR-A as an important target in sarcoma therapy. Future trials with these highly selective drugs should involve preselection of patients with tumors that express low levels of IR-A and IGF-2.

Studies are supported by grants from the Italian Association for Cancer Research, the Italian Ministry of Health, the Italian Ministry of Research and Instruction and the EU project Eurobonet.

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 728. doi:10.1158/1538-7445.AM2011-728
Abstract 733: A therapeutic opportunity in melanoma: Targeting CXCR1/2-dependent signaling attenuates therapy resistance
Authors: Wu Sheng , Varney Michelle L. , Singh Seema , Singh Rakesh K.
Published: June 2012

Advanced melanoma is one of the most intractable tumors, due to its insensitivity to all current treatment. Systemic chemotherapy is still considered the mainstay of treatment for advanced melanoma with limited survival impact. Failure of most agents is attributed to development of therapy resistance. Our previous findings demonstrate that CXCL-8 is a critical mediator in tumor growth, invasion, and angiogenesis. Two high affinity receptors for CXCL-8, CXCR1 and CXCR2, are differentially expressed on melanoma and endothelial cells. Based on these observations, we hypothesize that targeting CXCR1/2-dependent signaling will attenuate chemotherapy resistance and will synergistically enhance chemotherapeutic response for malignant melanoma. We used human melanoma cell lines A375P (low metastatic) and A375SM (highly metastatic), which express low and high levels of CXCL-8 respectively. First, we evaluated the role of CXCR1/2 ligands in chemo-resistance. CXCL-1 and -8 mRNA and protein expression was analyzed by quantitative realtime PCR and enzyme-linked immunosorbant assay, following treatment of dacarbazine and temozolomide. A significant increase in the CXCL-1 and -8 levels was observed in A375P and A375SM cells which was dependent on concentration and duration of chemotherapeutic treatment. In order to test whether blocking CXCR1/2 expression using genetic knock-down modulates chemotherapeutic responses, we used A375SM cells expressing different levels of CXCR1 and CXCR2. Our data suggest that knockdown of CXCR1 significantly enhanced the cytotoxic effect of dacarbazine in vitro. However, we did not observe a similar response in melanoma cells knocked down for CXCR2. Taken together, our preliminary data demonstrate that melanoma cells that survive chemotherapy express higher levels of CXCR1/2 ligands and targeting the CXCR1/2-dependent pathway might be a potential therapeutic approach to be combined with chemotherapy in advanced melanoma.

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 733. doi:10.1158/1538-7445.AM2011-733
Abstract 750: Identification of tumor antigens in transgenic mouse model as vaccine targets for the prevention of human breast cancer
Authors: Mao Jianning , Wu Mei , Gad Ekram , Lu Hailing , Disis Mary
Published: June 2012

The development of a vaccine to prevent breast cancer would be facilitated if tumor antigens were identified which were associated with the initiation of the malignant transformation. Transgenic mouse models have been proven to demonstrate a similar antigenic repertoire as that seen in breast cancer patients, therefore, we propose to use these models to identify tumor antigens as vaccine targets for breast cancer prevention. We have chosen two mouse models: TgMMTV-neu and TgC3(1)-Tag for study. TgMMTV-neu mouse has a genotype similar to luminal breast cancer and the TgC3(1)-Tag model reflects a basal phenotype of breast cancer. We identified potential vaccine candidate antigens in one of two ways; (1) determining which proteins expressed by the cancer are immunogenic very early in the malignant process by screening pre-diagnostic sera for immunogenicity against tumor cDNA libraries, and (2) injecting parental animals with a syngeneic tumor cell line resulting in “tumor rejection” which we have shown previously is mediated by T cells. To date, we have identified nine pre-diagnostic tumor antigens in TgMMTV-neu mouse; Rpl5, TNFaip3/A20, Pdhx, Otud6b, Stk39, Zfp238, Dnajc10, Lgals8 and Vps35. Moreover, we have identified four rejection antigens in TgC3(1)-Tag mouse; Ddx21, Sfrs11, Edh1 and Tdg. We have initiated vaccination experiments to examine the anti-tumor effect of vaccination with plasmids encoding these antigens. We found that vaccination targeting Ddx21, Dnajc10, and Pdhx mediated tumor growth inhibition in implant models compared with a control group (p<0.01). Our preliminary studies also demonstrate that several of the proteins are immunogenic in humans and we have identified putative promiscuous class II epitopes suitable for screening human PBMC for T cells specific to these proteins.

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 750. doi:10.1158/1538-7445.AM2011-750
Abstract 771: Therapy of spontaneous primary breast tumors in neu-N transgenic mice by adoptive transfer of vaccine-primed LN T cells also induces a humoral response
Authors: Wang Li-Xin , Plautz Gregory E.
Published: June 2012

To investigate factors which regulate T-cell mediated therapy of primary breast tumors in situ, we utilized neu-N transgenic mice. These neu-tolerant mice develop spontaneous breast tumors, several of which we have established as aggressive transplantable tumors and selected stem-like progenitor cells in vitro. We vaccinated neu-N mice with irradiated neupos breast cancer progenitor cells to prime T cells in draining LNs. We purified a minor subset (5% to 10%) of LN cells with a CD62Llow CD4+ or CD8+ phenotype representing recently activated effector cells. CD4 and CD8 subsets were separately activated in vitro with anti-CD3/IL-2/IL-7 for two stimulation cycles to induce 200-fold numerical expansion. Hosts bearing spontaneous primary breast tumors average 80 mm2 were treated with 35 × 106 of each CD4+ and CD8+ subsets. Even though the neu-tolerant hosts did not receive lymphodepletion prior to T cell transfer, the T cells significantly delayed tumor growth compared with untreated control mice (n=4 per group, p=0.037). This extends our previous work showing effective T cell therapy of inoculated lung metastases to primary in situ neu-N breast tumors. Neu is a transmembrane tumor antigen with shared expression between vaccine cells and spontaneous tumors that could contribute to the anti-tumor response. We also searched for evidence of response against other cross-reactive antigens. Interestingly, treated mice developed a humoral response and post-treatment serum IgG bound to neuneg variant tumor cells as well as neupos cell lines whereas pre-treatment serum was non-reactive. The development of an IgG serologic response following effective T cell-mediated therapy (in the absence of co-transferred B cells) indicates that transferred CD4+ T cells provided cognate helper function and immunoglobulin class switch signals to B cells and reversed tolerance to neu as well as other shared tumor antigens.

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 771. doi:10.1158/1538-7445.AM2011-771
Abstract 776: Therapeutic immunity against melanoma by IL-12 cDNA electroporation: a possible therapeutic mechanism
Authors: Sin Jeong-Im , Ryoo Hun Mo
Published: June 2012

We observed that large-established melanoma was eradicated by intratumoral (IT)-electroporation (EP) of IL-12 cDNA in an animal model. In this study, we explored the mechanism(s) as to how this tumor eradication is mediated in vivo and evaluated if long-term memory responses to parental tumor cell re-challenge can be induced. Melanoma (B16) cells were subcutaneously inoculated. When tumor reached 6-8 mm in mean diameter, 50 μg of IL-12 cDNA was intratumorally injected and electroporated using Cellectra® of VGXI at 0.2A for 4 sec. To further determine the antitumor mechanism(s), IT-EP was performed in tumor-bearing wild type, IFN-γ knockout (KO), and perforin KO animals. Furthermore, immune cell infiltration was measured by histological analysis. In vitro and in vivo immune cell subset depletion assay was also performed to see an involvement of T and NK cell populations in tumor Ag-specific IFN-γ production and tumor control. IT-EP of IL-12 cDNA resulted in eradicating tumors established on the skin without any significant levels of long-term antitumor memory immunity. This effect required mainly CD8+ and to a lesser degree CD4+ T cells, but not NK cells. Both IFN-γ KO animal studies and IT-EP assay of IFN-γ cDNA confirmed that IFN-γ was indirectly involved in melanoma control. Furthermore, perforin was found to be essential for the tumor-eradicating activity. However, B16 melanoma cells expressed Fas, but failed to exhibit Fas-mediated cell lysis, suggesting that perforin/granzyme B-mediated killing pathway is associated with CD8+ T cell-mediated melanoma control. Taken together, these studies show that IT-EP of IL-12 cDNA is an important approach to control melanoma on the skin and this antitumor effect is mediated mainly by CD8+ T cells (as a killer T cell) through perforin/granzyme B-mediated killing pathway. This study also suggests that additional approaches are needed to induce long-term antitumor memory responses.

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 776. doi:10.1158/1538-7445.AM2011-776