bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3720: Encouraging activity of bicalutamide and everolimus in castration-resistant prostate cancer (CRPC): Early results from a phase I/II clinical trial
Authors: Zhang Hongyong , Wang Yu , Ghosh Paramita , Lara Primo N. , Pasquinelli Patricia , Beckett Laurel , deVere White Ralph W. , Pan Chong-Xian
Published: February 2011

Background: Multiple signaling pathways are involved in the development of CRPC. We previously showed that the mTOR pathway is activated in CRPC cell lines while inhibition of this pathway results in upregulation of androgen receptor (AR) signaling (Wang et al, Oncogene. 2008). Simultaneous blockade of the mTOR and AR pathways synergize in inducing PCa cell death and delaying tumor formation in mouse models. We hypothesize that simultaneous blockade of the AR and mTOR pathways in CRPC patients with bicalutamide and everolimus will result in improved efficacy compared to bicalutamide alone.

Methods: A phase II clinical trial with a lead-in safety phase was designed to determine the efficacy and tolerability of the bicalutamide and everolimus combination in CRPC patients compared with bicalutamide alone. Patients must have histologically confirmed disease and demonstrated disease progression (either by PSA or radiographically) while on androgen deprivation therapy. At the lead-in phase, all patients receive both agents. At the phase II stage, patients are randomized to bicalutamide +/− everolimus. The primary endpoint is PSA response. The second endpoints include progression-free survival, time-to-treatment failure, overall survival and toxicity. Here, we report the results of the lead-in phase.

Results: To-date, 13 patients have been screened and 8 have been enrolled with the age ranging XX-XX, and performance status of ECOG 0-1. In the lead-in phase, bicalutamide + everolimus has been well tolerated with no unexpected toxicities. One patient developed grade 3 interstitial pneumonitis (attributed to everolimus) that improved with dose reduction and steroid treatment. Seven of 8 patients have had either stable PSA or a PSA response after at least 8 weeks of therapy. Tumor and blood specimens are being collected for molecular correlative studies of mTOR pathway markers. Conclusions: Thus far, the rational combination of bicalutamide and everolimus appears to have promising anti-tumor activity and an acceptable toxicity profile. The randomized phase of the clinical trial is currently ongoing and will be reported. (Supported by Novartis)

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 3720.
Abstract 373: Circulating endothelial cell measurement allows early prediction of outcome in patients with metastatic colorectal cancer treated with first-line bevacizumab and chemotherapy. Findings from the FNCLCC ACCORD 13/0503 randomized phase II trial
Authors: Boige Valerie , Mendiboure Jean , Adenis Antoine , Boucher Evelyne , Pierga Jean-Yves , Francois Eric , Conroy Thierry , Pignon Jean-Pierre , Ducreux Michel , Farace Françoise , Malka David
Published: February 2011

Purpose - There is no validated biomarker to predict the efficacy of bevacizumab. We tested the hypothesis that circulating endothelial cells (CEC) levels could predict clinical outcome of metastatic colorectal cancer (mCRC) patients starting treatment with bevacizumab and chemotherapy.

Patients and Methods - Among the 145 mCRC patients treated with first-line bevacizumab and irinotecan-based chemotherapy (either FOLFIRI or XELIRI regimen) in the FNCLCC ACCORD 13/0503 randomized phase II trial, 99 agreed to blood sampling at day [D]1 (before treatment) and D15 or 22 (at the end of cycle 1). CEC (CD45-CD31+CD146+7-amino-actinomycin- cells) were quantified by four-color flow cytometry according to a previously validated method. We assessed whether D1 or D15/22 CEC levels, or changes in CEC levels between D1 and D15/22, correlated with patient characteristics, 6-month objective response rate (ORR), and 6-month progression-free survival (PFS) rate (the trial primary endpoint).

Results - CEC levels were higher at D1 in patients with altered PS (p=0.02). CEC levels were higher at D15/22 in the absence of objective response (p=0.04), and at D1 (p=0.01) and D15/22 (p=0.03) in patients whose disease progressed within 6 months of treatment. The best outcome (6-month PFS rate, 96%) was observed in patients with low CEC levels at D1 and D15/22 (cutoff, baseline median=16/mL) compared to patients with high CEC levels at either D1 or D15/22 (79%) and patients with high CEC levels at D1 and D15/22 (68%; trend test, p=0.01).

Conclusions - CEC levels may help in early prediction of efficacy of first-line bevacizumab and chemotherapy in mCRC patients.

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 373.
Abstract 3730: Akt activity is suppressed by doxorubicin through downregulation of ErbB receptors and its enhanced activity is associated with higher sensitivity to doxorubicin in breast cancer
Authors: Harada Amane , Suminaka Chihiro , Oyama Tomoko , Gohda Keigo , Ishihara Hideki , Kim Seung Jin , Noguchi Shinzaburo
Published: February 2011

Anthracyclines are the backbone of breast cancer therapy and benefit patients in both the adjuvant and metastatic setting whereas they are effective for 50-60% of breast cancer patients and cause substantial morbidity and occasional life-threatening toxic effects. For these reasons, it has been eagerly anticipated to develop diagnostic markers which can predict efficacy of the treatment. HER2 amplification or overexpression of its protein (HER2 positivity) has been shown to be associated with relative benefit from anthracycline-based chemotherapy (Gennari A. et al., J. Natl. Cancer Inst, 100, 14-20, 2008). However, HER2 is positive in 15-25% of breast tumor tissues and no clear biological mechanism between HER2 positivity and action of anthracyclines has been reported.

In this study, we have shown that doxorubicin exerts its anti-tumor activity through inhibiting ErbBs-Akt signal pathway. Promoted activity of Akt was associated with not only HER2 but EGFR overexpression and responsiveness to doxorubicin in human breast cancer cell lines. As well as doxorubicin, PI3 kinase inhibitor caused growth inhibition and apoptosis in cells with promoted Akt activity more significantly than cells with weakly activated Akt. Doxorubicin-induced cytotoxic stress induced internalization and degradation of ErbBs followed by inactivation of Akt. This was confirmed with the observation that doxorubicin caused decrease in the amount of immunocomplex formed between HER3 and p85, PI3 kinase regulatory subunit. These data indicate that cancer cells with ErbBs overexpression and promoted Akt activity are dependent on activities of these factors for their growth and survival, and doxorubicin inhibits the ErbBs-Akt signal pathway through inducing downregulation of ErbB receptors. Furthermore, we found that highly phosphorylated Akt in tumors corresponds to improved recurrence-free survival in breast cancer patients treated with anthracycline-based adjuvant chemotherapy relative to weakly phosphorylated Akt. Taken together, we provide mechanistic insight into molecular basis of correlation between HER2 positivity and sensitivity to anthracyclines, and propose a new possibility that Akt activity is a useful predictor of response to anthracycline-based chemotherapy.

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 3730.
Abstract 3733: Heregulin as a novel resistant factor for cetuximab
Authors: Yonesaka Kimio , Okamoto Isamu , Satoh Taroh , Takeda Kohji , Takada Minoru , Nishio Kazuto , Fukuoka Masahiro , Janne Pasi A. , Nakagawa Kazuhiko
Published: February 2011

Background

Epidermal growth factor receptor (EGFR) is a therapeutic target in colorectal cancer (CRC). Patients with CRC with wild type KRAS have significant tumor regressions with cetuximab. However cancers ultimately acquire resistance. The mechanisms of these are still unknown. We investigated mechanisms of acquired resistance to cetuximab.

Methods

The cetuximab-resistance cell line, A431ER, was established from cetuximab-sensitive parent A431 by exposing cetuximab. In vitro sensitivity was analyzed using MTS assay. Expression of ErbBs and heregulin (HRG) was analyzed with RTK array, immunoblotting (IB) and ELISA. Plasma samples were studied using ELISA among 44 pts with CRC, treated with cetuximab.

Result

A431 was so sensitive to cetuximab that the growth inhibition following 10μg/ml cetuximab (GI10) was about 50% in MTS. In contrast, A431ER was completely resistance (GI10 <10%). The RTK array showed that A431ER but not A431 had markedly phosphorylated ErbB2 and 3. Although ErbB2 and 3 were highly phosphorylated in A431ER, total protein level of those was equivalent to A431 in IB. Finally heregulin, a ligand for ErbB3, was detected significantly higher in A431 than A431ER using IB and also ELISA in culture mediums (355 vs 1025pg/ml). HRG inhibition with siRNA could restore the sensitivity to cetuximab in A431ER. And exogenous HRG induced A431 parent cell line resistant to cetuximab. The exogenous HRG also phosphorylated both Erb2 and 3 in A431, which means hetero-dimerization of those. So we did MTS assay with cetuximab and pertuzumab, which inhibited hetero-dimerization between ErbB2 and 3. Cetuximab alone or pertuzumab alone minimally effected in A431ER (GI10 was about 10%). But the combination of two drugs abundantly effected (GI10 was about 60%). About clinical samples, pts with progressive disease (PD, n=23) following cetuximab had higher expression of HRG in plasma than pts with non-PD (n=15) (median 1188 vs 4854pg/ml, p=0.0008). And low HRG group (<2000pg/ml, n=22) had longer progression free survival (HR 0.2371 (95%CI 0.095-0.593), p=0.0021) and overall survival (HR 0.2274 (0.075-0.693), p=0.0092) than high HRG group (>2000pg/ml, n=22). Finally, post- treatment samples had significantly higher HRG than pre-treatment in 6 pts, who acquired resistance after 5 months treatment (mean 904 vs 1890pg/ml, p=0.0161).

Conclusion

Our findings suggest that HRG could lead both de novo and acquired resistance to cetuximab. HRG-ErbB3-ErbB2 complex is a warranted target for overcoming resistance to cetuximab.

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 3733.
Abstract 374: VEGFR2/KDR polymorphisms and gene copy gain with outcome in non-small cell lung cancer
Authors: Yang Fei , Tang Ximing , Corvalan Alejandro , Behrens Carmen , Lin Heather , Raso Maria Gabriela , Lee J. Jack , Minna John D. , Wistuba Ignacio Ivan
Published: February 2011

Background. The VEGF receptor 2 (VEGFR2) is the predominant mediator of VEGF-stimulated endothelial cell function. Recently, VEGFR2/KDR copy number gain and mutation, and VEGF copy number gain, have been described in lung adenocarcinoma tumor specimens. To better characterize the molecular changes of these two genes in non-small cell lung carcinoma (NSCLC), we investigated their abnormalities in NSCLC tumor tissue specimens and correlated with patients clinico-pathological characteristics.

Methods. We extracted DNA from microdissected tissue obtained from 200 surgically resected NSCLCs. KDR single nucleotide polymorphism (SNP) 889G/A (rs2305948), SNP 1416A/T (rs1870377) and SNP −37A/G (rs2219471) were genotyped by PCR-based sequencing. KDR and VEGF copy number were examined by quantitative (q)-PCR. Protein expressions of VEGF and VEGFR2, and CD34 for microvascular density (MVD) analysis were studied by immunohistochemistry. SNP genotypes, genes copy number and protein expression were correlated with NSCLC clinico-pathological features, including overall survival (OS) and recurrence-free survival (RFS).

Results. KDR 1416 AT/TT genotypes had significantly improved OS (HR = 0.56, 95% CI 0.33 to 0.96, P = 0.035) compared with KDR 1416 AA in all NSCLC patients in the multivariate analysis with adjustment of histology and tumor stage. In lung adenocarcinomas, KDR 1416 AT/TT genotypes and KDR −37AG/GG were associated with a favorable OS (HR = 0.43, 95% CI 0.20 to 0.92, P = 0.029; HR = 0.47, 95% CI 0.23 to 0.96, P = 0.039, respectively). Strikingly, KDR −37 AG/GG genotypes predicted a superior OS compared with KDR −37 AA genotype in the NSCLC patients treated with adjuvant therapy (HR = 2.45, 95% CI 1.06 to 5.66, P = 0.036). No genotype in KDR SNPs was associated with RFS in NSCLC patients. Gene copy gain of KDR and VEGF were detected respectively in 34/91 (37.4%) and 2/91 (2.2%) NSCLC tumors. Gene copy gain of KDR was associated with a poor overall survival in NSCLC patients (HR= 2.96, 95% CI 1.41 to 6.24, P = 0.004). Furthermore, tumors with gene copy gain of KDR showed significantly higher cytoplasmic (P = 0.013) and membrane (P = 0.009) VEGFR2 protein expression, lower cytoplasmic VEGF expression (P = 0.044), and higher MVD (P = 0.018) and larger vessel areas (P = 0.033) compared with tumors lacking KDR gene copy gain.

Conclusion. Our findings suggest an association between KDR SNP genotypes and survival in NSCLC patients receiving adjuvant therapy. KDR copy number gain was frequently identified in NSCLC and was associated with worse survival in NSCLC patients. (Supported by grant US DoD W81XWH-07-1-0306).

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 374.
Abstract 3741: Adjuvant active specific immunotherapy improves survival of patients with micro-satellite stable colorectal tumors, but not with micro-satellite instable tumors
Authors: de Weger Vincent A. , Euler Zelda , van Tinteren Harm , Bril Herman , Meijer Gerrit A. , Hooijberg Erik
Published: February 2011

Purpose: It has been documented that Active Specific Immunotherapy (ASI), given as adjuvant treatment, improves survival of patients with stage II and III colorectal cancer (CRC). Here we report on the effects of ASI on Micro-Satellite Stable (MSS) versus Micro-Satellite Instable (MSI) tumors.

Materials and Methods: From 196 patients with stage II or stage III CRC who previously participated in the ASI trial published previously (Vermorken et al. Lancet), we collected frozen tumor samples and/or paraffin embedded tumor material. From this material DNA was extracted and microsatellite stability status was assessed. Microsatellite status was associated with clinical outcome in both the placebo and ASI treatment groups using Kaplan Meier analysis.

Results: From 196 patients participating in the trial we retrieved good quality archival tumor DNA for further analysis of microsatellite stability (MS). We subsequently identified 162 (83%) MSS tumors and 34 MSI (17%) tumors. Irrespective of MS status a significant survival benefit for patients receiving ASI treatment (P=0.012) was observed, confirming previous results. New to this is that irrespective of treatment, patients with MSI tumors showed significantly less recurrences than patients with MSS tumors (P = 0.03). Taking both MS status and treatment into account we found no significant difference in survival between ASI treated versus placebo receiving patients with MSI tumors. In contrast, patients with MSS tumors benefited from ASI treatment showing a significantly improved recurrence free survival (P=<0,05).

Conclusion: It can be concluded that only patients with MSS tumors will benefit from adjuvant ASI since only they showed significant improved recurrence free survival on ASI therapy. For the MSI group there is most likely no benefit of ASI therapy.

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 3741.
Abstract 3753: Quantitation of EGFR and phosphoEGFR in FFPE tissue
Authors: Hembrough Todd , Krizman David , Heidbrink-Thompson Jenny , Thyparambil Sheeno , Burrows Jon , Darfler Marlene , Taylor Paul , Tong Jiefie , Shi Warren , Tsao Ming , Moran Michael
Published: February 2011

The epidermal growth factor receptor (EGFR) is a drug target for both small molecule and antibody therapeutic approaches for several cancers; however, current methods of selecting patients that will most likely respond to anti-EGR therapy are not effective. Better methods for patient stratification are needed. To this end we have developed an approach which can determine both absolute EGFR levels and the phosphorylation status of EGFR directly in formalin-fixed paraffin-embedded (FFPE) patient tissue. This approach is based on the Liquid Tissue®-SRM technology platform, a combination of tissue microdissection, Liquid Tissue® processing which turns dissected tissue to a complete solubilized tryptic digest, and mass spectrometry-based selected reaction monitoring (SRM). This approach was used to measure the EGFR protein and its phosphorylation status in formalin fixed tissue culture cells, xenograft tumors, and patient tumor tissue. For assay development, 3 distinct tryptic peptides were assessed for absolute protein quantitation and multiple peptides where specific residues are known to become phosphorylated (pT693 and pY1197) were assessed for assaying the phosphorylation status of the EGFR protein. We demonstrate the ability to detect and quantify the EGFR protein and to monitor its phosphorylation status directly in patient tumor tissue. This approach offers a dynamic range and quantification which is superior to traditional IHC methods, and that could be used to identify and stratify patients most likely to benefit from anti-EGFR therapies.

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 3753.
Abstract 3764: Promoter methylation in exfoliated breast cells isolated from breast milk of women who have had a breast biopsy
Authors: Browne Eva P. , Punska Elizabeth C. , Anderton Douglas L. , Arcaro Kathleen F.
Published: February 2011

Analysis of promoter methylation patterns in genes frequently downregulated in breast cancer may serve as an early indicator of breast cancer risk. To date, studies of methylation patterns in breast epithelial cells from asymptomatic women have been limited to cells obtained from fine nipple aspirate and ductal lavage. Analysis of promoter hypermethylation in exfoliated epithelial cells isolated from breast milk provides a non-invasive opportunity to assess breast cancer risk. Our preliminary data indicate that we can detect a cancer methylation phenotype in the milk of women at high risk. We propose that epithelial cells isolated from breast milk of lactating women scheduled to have a breast biopsy provide a resource for assessing early changes in DNA related to increased breast cancer risk.

Two hundred and fifty breast milk samples from women who are scheduled to have, or have had a breast biopsy will be collected. Of these women, we expect that roughly ten percent will be diagnosed with breast cancer. Breast milk will be collected from both the biopsied and contralateral breast of each participant. To date, 153 participants have donated breast milk, yielding 300 breast milk samples with an average size of 50 ml (±30 ml) per breast. After collection, the epithelial enriched and epithelial depleted cells from each breast were isolated. Subsequent DNA isolation of both the epithelial enriched and depleted fraction of cells yielded an average of 372 ng (range 12-5000 ng) of high quality DNA.

Using the DNA extracted from epithelial enriched and depleted fractions of cells isolated from breast milk, the promoter methylation level of 9 genes known to be methylated in breast cancer will be examined using pyrosequencing technology. We will assess the promoter methylation levels of: CDH1, DAP-K, ER-α, GSTP1, HIN1, P16, RAR-ß, RASSF1A, and SFRP1 in the epithelial enriched and depleted bisulfite treated DNA of each breast milk sample. Preliminary pyrosequencing assays conducted in our lab have shown that there is a difference in RASSF1A methylation levels between the epithelial enriched DNA from the cancerous breast compared to the epithelial depleted DNA from the non-cancerous breast of the same participant. Further methylation analysis may prove that breast milk can be used as non-invasive alternative to ductal lavage or fine needle aspirate, which may yield accurate breast cancer risk assessment.

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 3764.
Abstract 3779: Combination chemoprevention of hepatocellular carcinoma using acyclic retinoid
Authors: Shimizu Masahito , Moriwaki Hisataka
Published: February 2011

Background and aims: A malfunction of retinoid X receptor-α (RXRα) due to phosphorylation by the Ras/MAPK signaling pathway is associated with the development of hepatocellular carcinoma (HCC). Acyclic retinoid (ACR: the same material as NIK-333), which has been clinically shown to reduce the incidence of a post-therapeutic recurrence of HCC, can inhibit both the Ras activity and phosphorylation of the ERK and RXRα proteins. The effects of a combination of ACR plus other agents on the inhibition of cell growth in human HCC cells were herein examined to develop a more effective strategy for the prevention of HCC.

Methods and results: The combination of ACR plus vitamin K2, which inhibits the phosphorylation of the RXRα protein through the inhibition of Ras/MAPK activation, synergistically induced apoptosis and inhibited the growth of human HCC cells without affecting the growth of normal human hepatocytes. ACR and trastuzumab, the humanized anti-HER2 monoclonal antibody, also synergistically inhibited the activation of HER2, a member of receptor tyrosine kinase (RTK), and its downstream signaling pathways, thus subsequently inhibiting the phosphorylation of RXRα and the growth of HCC cells. ACR in combination with valproic acid, a histone deacetylase inhibitor, also synergistically inhibited the growth of human HCC cells and this was associated with the inhibition of phosphorylation of the RXRα and Akt proteins by valproic acid.

Conclusion: The combination of ACR plus a specific agent that targets RTK and its downstream signaling pathways, including the Ras/MAPK and PI3K/Akt pathways, may be able to inhibit the phosphorylation of RXRα and it may therefore be a promising strategy to prevent the development of HCC. ACR, which targets phosphorylated RXRα, may thus play a critical role in preventing the development of HCC when it is used either alone or in combination with other agents.

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 3779.
Abstract 3782: Benzyl isothiocyanate-induced apoptosis in human breast cancer cells is mediated by downregulation of X-linked inhibitor of apoptosis protein
Authors: Kim Su-Hyeong , Singh Shivendra V.
Published: February 2011

Benzyl isothiocyanate (BITC) is a highly promising cancer chemopreventive constituent of edible cruciferous vegetables such as gardencress. We have shown previously that BITC treatment inhibits viability of MDA-MB-231 (an estrogen-independent cell line expressing mutant p53) and MCF-7 (an estrogen-responsive cell line with wild-type p53) by causing reactive oxygen species-mediated apoptosis. In addition, oral administration of BITC inhibits growth of MDA-MB-231 human breast cancer cells subcutaneously or orthotopically implanted in female athymic mice without causing weight loss or any other side effects. More recent studies from our laboratory have indicated that dietary feeding of BITC offers protection against mammary hyperplasia and carcinoma development in MMTV-neu transgenic mice. The present study extends these exciting observations and determines role of p53 and inhibitor of apoptosis (IAP) family proteins in regulation of BITC-induced apoptosis. Exposure of MCF-7 and MDA-MB-231 cells to apoptosis inducing concentrations of BITC (2.5 and 5.0 μM) resulted in up-regulation of p53 in MCF-7 cell line and down-regulation of X-linked inhibitor of apoptosis (XIAP) in both cell lines. The BITC-mediated down-regulation of XIAP was evident as early as 8 h post-treatment. BITC-treated cell also exhibited induction of survivin especially in the MCF-7 cell line. The level of cIAP2 protein was only modestly increased upon treatment of MCF-7 and MDA-MB-231 cells with BITC. The BITC-induced apoptosis was only marginally affected by knockdown of p53 protein level by RNA interference. On the otehr hand, forced expression of XIAP protein conferred significant protection against BITC-mediated cytoplasmic histone-associated apoptotic DNA fragmentation in both cell lines. The BITC-induced cytoplasmic histone-associated apoptotic DNA fragmentation was significantly increased by knockdown of survivin protein in the MCF-7 cell line but not in the MDA-MB-231. These results indicate that down-regulation of XIAP contributes to the BITC-induced apoptosis in breast cancer cells. In addition, induction of survivin confers resistance against BITC-mediated cell death at least in the MCF-7 cell line. This study was supported by the National Cancer Institute grant CA129347-03.

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 3782.
Abstract 3786: Activation of KLF4 by 3,3′-Diindolylmethane contributes to the cell differentiation and grow inhibition of pancreatic cancer cells
Authors: Wei Daoyan , Xie Keping
Published: February 2011

3,3′-Diindolylmethane (DIM), the major bioactive metabolite of the phytochemical indole-3-carbinol present in cruciferous vegetables, is a promising cancer preventative and anti-tumor agent for various tumor types, including pancreatic cancer. However the underlying molecular mechanisms by which DIM exerts its antitumor activity remain poorly understood. Our previous studies indicate that KLF4 has a tumor suppressive function in pancreatic cancer by up-regulation of p27 expression and induction of cell cycle arrest, which has also been suggested to be the mechanistic action of DIM in pancreatic cancer cells. However, whether KLF4 is involved in DIM's antitumor activity and if so, what are the underlying molecular mechanisms in pancreatic cancer have not been defined. In the present study, we found that DIM treatment dose dependently induced the expressions of KLF4 and its downstream target gene p27 and p21 in several, and cytokeratin 19 in Panc-28 and PANC-1 pancreatic cancer cells. Consistently, induction of KLF4 expression correlated with significant inhibition of cell cycle progression and cell growth as determined by Flow cytometry and MTT assay, while knockdown or conditional knockout of KLF4 expression significantly reduced the sensitivity of pancreatic cancer cells to DIM treatment determined by IC50 assay. These results indicate that KLF4 plays a critical role in mediating the antitumor activity of DIM in pancreatic cancer cells. To further investigate the molecular mechanism underlying DIM's induction of KLF4 expression, KLF4 promoter deletion mutation analysis identified a DIM responsible region (from nucleotide −780 to −346) in the KLF4 promoter, within which a candidate Egr-1 cis-acting element was found by computer-based motive analysis. Functionally, significant induction of Egr-1 protein expression was observed as early as 1 hr after DIM treatment in time-course experiments, which was prior to the induced expression of KLF4 in FG cells. Consistently, forced or knocking down Egr-1 expression increased or reduced the response of BxPC3 cells to DIM treatment, which was correlated with increased or decreased KLF4 expression. In addition, chromatin immunoprecipitation assay showed that Egr-1 protein physically interacted with and DIM treatment increased the binding of Egr-1 to the DIM responsive region of KLF4 promoter in FG cells in vivo. Collectively, our results suggest that Egr-1/KLF4 signaling pathways play an important role in DIM's antitumor activity in pancreatic cancer cells.

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 3786.
Abstract 3789: Elevated inorganic phosphate stimulates immediate early gene expression and requires FGF receptor signaling
Authors: Beck George R. , Camalier Corinne E. , Hood Brian , Conrads Thomas P.
Published: February 2011

Dietary inorganic phosphate represents a novel chemoprevention target. Recent results from two different mouse models of tumorigenesis; the two-stage skin carcinogenesis and KrasLA1 lung tumor models suggest that a high phosphate diet increases tumor/papilloma number by approximately 50% relative to a low phosphate diet. Additionally, a low phosphate diet resulted in delayed incidence and growth of papillomas. These results suggest that increased consumption of dietary inorganic phosphate may have long-term consequences on overall health and particularly to tumorigenesis and consequently reducing dietary phosphate intake may be a novel chemoprevention strategy. Our in vitro studies have revealed an active role of inorganic phosphate in altering cell function in various cell types and suggest that the amount of available inorganic phosphate may have significant cell autonomous consequences on cell behavior including activation of ras and ERK1/2 signaling, and increased expression of transformation and metastasis associated genes such as osteopontin (spp1). However, the mechanism by which cells sense and respond to changes in extracellular phosphate resulting in increased proliferation and transformation is not fully understood. We have performed an analysis of the temporal changes in gene expression in response to elevated phosphate. Results identified the strong stimulation of immediate early genes within 15 minutes of exposure to elevated phosphate. Reporter and DNA binding assays revealed the requirement of serum response and AP-1 transcriptional elements and taken together characterize phosphate as a novel mitogen. Furthermore, we have identified the requirement of FGF receptor signaling as one of the earliest events in the phosphate signaling cascade. This finding provides a novel link between growth factor signaling, nutrient sensing and cell growth and tumorigenesis.

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 3789.
Abstract 38: Saturated free fatty acids activate a c-Src-JNK1 axis within lipid rafts: Implications for the development of insulin resistance, obesity, and obesity-associated cancer
Authors: Holzer Ryan , Tran Helen , Salinas Giovanni , Karin Michael
Published: February 2011

Obesity is a risk factor for the development of insulin resistance/type 2 diabetes and several types of cancer, including cancers of the colon, breast, liver, endometrium and esophagus. Obese individuals have increased levels of free fatty acids (FFAs) in their serum, and so we are investigating signaling pathways that are activated by FFAs in vitro to gain insight into pathways that may trigger insulin resistance and the activation of oncogenes in obese individuals in vivo. Obesity is associated with chronic activation of c-Jun NH2-terminal kinase (JNK)1 in liver, fat and muscle tissue, and studies using JNK1-deficient mice demonstrate that JNK activation is important for adiposity, peripheral insulin resistance and the production of the proinflammatory cytokines, but the mechanism of JNK1 activation in obesity is unclear. Our data demonstrate that JNK1 is activated by FFAs in vitro in a c-Src-dependent manner, and that activation of c-Src occurs within lipid rafts. Src-deficient murine embryonic fibroblasts (MEFs) and SYF MEFs, which are deficient in c-Src family kinases c-Src, c-Yes and c-Fyn, do not activate JNK1 after exposure to FFAs in vitro, while NIH 3T3 MEFs and SYF MEFs reconstituted with c-Src activate JNK1 normally. In contrast, SYF MEFs reconstituted with c-Yes and c-Fyn do not activate JNK1. Knockdown of c-Src with shRNA in HEK 293T cells also inhibits FFA-induced activation of JNK1, and inhibition of c-Src with pharmacologic inhibitors has the same effect in J774 mouse macrophages. Pharmacologic inhibition of c-Src also prevents the FFA-induced upregulation of tumor necrosis factor (TNF)-α mRNA in macrophages. Importantly, c-Src is activated within lipid rafts after FFA treatment, as seen by both detergent-dependent and detergent-independent methods of raft isolation. Interestingly, omega-3 polyunsaturated fatty acids (PUFAs), which can protect against the onset of obesity, will also block JNK activation following treatment with saturated FFAs, and we suspect that this effect may be due to the ability of PUFAs to exclude c-Src from lipid rafts. Finally, preliminary data suggests that c-Src may be aberrantly activated in liver tissue from obese mice that were fed a high-fat diet. Experiments are currently underway to verify the role of c-src in the activation of JNK1 in vivo by acutely infusing mice with a free fatty acid emulsion in the presence of a pharmacologic inhibitor of c-src.

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 38.
Abstract 3801: p53 acetylation by green tea polyphenols leads to upregulation of GSTP1 activity
Authors: Thakur Vijay S. , Gupta Karishma , Gupta Sanjay
Published: February 2011

Epigenetic modifications including DNA methylation and post-translational changes in histone proteins cause alteration in gene regulation. Expression of glutathione-S-transferase Pi (GSTP1), an enzyme of phase II cellular metabolism involved in detoxification of carcinogens and other cellular functions found to be silenced in 80-90% prostate cancers due to hypermethylation of its promoter. We have recently demonstrated that green tea polyphenols (GTPs) cause re-expression of GSTP1 in human prostate cancer LNCaP cells [Int J Cancer. 2009; PMID: 19856314]. However, the mechanism of GSTP1 re-expression is not fully elucidated. Similar to GSTP1 gene, p53 tumor suppressor is also involved in a number of important cellular events including cell cycle, DNA repair, and apoptosis which is reported to be mutated or has defective signaling in most human cancers including prostate cancer. Human GSTP1 gene has a p53-binding motif located in intron 4 and is highly homologous to consensus p53-binding motifs in other p53-responsive genes having the ability to transcriptionally activate GSTP1. Whether p53 participates in GTP-mediated re-expression of GSTP1 is unknown. In the present study we attempted to elucidate the role of tumor suppressor p53 in GSTP1 re-expression after treatment with GTPs. Human prostate cancer LNCaP cells which possess epigenetic signatures of GSTP1 silencing and functional p53 were treated with physiologically achievable doses of GTPs (5μg/mL) or its major constituent epigallocatechin-3-gallate (20μM) up to 7 days. Exposure of LNCaP cells to green tea polyphenols caused acetylation of p53 at its C-terminal amino acid positions K373 and K382, other than K305, which correlated with increased GSTP1 expression. Discontinuation of treatment with GTPs resulted in the loss of p53 acetylation with progressive decrease in GSTP1 expression in these cells. Furthermore, GSTP1 levels were significantly reduced after GTP treatment in cells with the knockdown of p53 expression achieved using shRNA in lentiviral vector. Taken together, our results demonstrate that expression of GSTP1, in part, could be enhanced through p53 acetylation after treatment with green tea polyphenols and thus protect the cells from undergoing epigenetic alterations which may contribute to prostate tumorigenesis.

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 3801.
Abstract 3808: Impact of MIC expression and shedding on prostate tumor development and progression in double transgenic TRAMP-MIC mouse models
Authors: Wu Jennifer D. , Liu Gang , Wang Xuanjun , Plymate Stephen , Greenberg Norm M.
Published: February 2011

Shedding of the NKG2D ligand MIC has been shown to be associated with poor disease prognosis in cancer patients, in particular in cancers of epithelial origins. However, the relationship of MIC expression or MIC shedding with cancer development and progression has never been demonstrated experimentally. In order to evaluate the impact of MIC expression and MIC shedding on cancer development and progression, we generated two double transgenic mouse models for prostate carcinomas, the TRAMP-MICB and TRAMP-MICB. A2 mouse models. In these models, the natural cleavable MICB or the mutant membrane-bound non-cleavable MICB. A2 was expressed in the prostate epithelium of the TRAMP (transgenic adenocarcinoma of the mouse prostate) mouse driven by the rat probasin promoter. Both MICB and MICB. A2 can be recognized by mouse NKG2D and activate mouse NK cells. Our results showed that the TRAMP-MICB. A2 mice only develop PIN (prostate intraepithelial neoplasia) lesions but do not progress to carcinomas. NK cell function in these mice is comparable to wild type mice. On the contrary, more than 60% of the TRAMP-MICB mice develop poorly differentiated (PD) prostate carcinomas and associated metastasis by 24 weeks of age whereas development of PD carcinomas in simple TRAMP mice of the same age is rare. In TRAMP-MICB mice, ELISA showed that mice bearing PD tumors had significantly highly levels of serum sMICB than those only developed well-differentiated (WD) carcinomas. Concomitantly, NKG2D-dependent splenic NK cell function was significantly impaired in TRAMP-MICB mice that developed PD tumors. Immunohistochemistry staining of MIC expression in TRAMP-MICB tumors revealed loss of surface MIC expression in PD carcinomas; in stead, strong MIC immune reactivity was seen in the interstitial associated with infiltration of lymphocytes. Also, in simple TRAMP mice, the development of PD tumor has been shown to be associated with NKG2D-induced tumor immunoediting through down-regulation of endogenous mouse NKG2D ligand expression. However, in TRAMP-MICB mice, PD carcinomas retained high levels of endogenous mouse NKG2D ligand expression, presumably due to sMIC-induced down-regulation of NKG2D function. Our findings suggest: 1) sustained expression of membrane-bound MIC can stimulate NKG2D tumor immunity whereas persistent presence of sMIC impairs NKG2D function; 2) MIC shedding promotes prostate tumor development and progression; 3) the MIC-NKG2D interaction in TRAMP-MICB mouse model closely mimics which in clinical cancers and thus the TRAMP-MICB mouse model can be used as a pre-clinical model for validation of reagents to inhibit MIC shedding for cancer therapy.

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 3808.
Abstract 3811: Robust anti-tumor activity through targeting integrin α5β1 and eliciting ADCC with a dual functional monoclonal antibody PF-04605412
Authors: Hu-Lowe Dana D. , Li Gary , Chen Enhong , Wang Jianying , Zhang Lianglin , Chen Jeffrey H. , Amundson Karin , James Zobel , Kraynov Eugenia , Sharp Leslie , Wickman Grant , Jiang Xin , Bender Steve , Casperson Gerald
Published: February 2011

Integrin α5β1 and its ligand fibronectin are significantly and coordinately over-expressed in tumor associated blood vessels and cancer cells. These molecules have been reported to mediate angiogenesis, tumor survival and metastasis. α5β1 has also been implicated as a poor prognostic marker in a number of human tumor types including NSCLC and ovarian carcinoma. Thus, α5β1 is a potential target for cancer therapy. Accumulating evidence suggests an important role of innate immune effector cells in tumor killing via antibody dependent cellular cytotoxicity (ADCC). Taking advantage of both of these mechanisms, we generated an α5 integrin-specific neutralizing monoclonal antibody (mAb, PF-04605412) with high affinity for Fcγ receptors. In vitro, PF-04605412 potently inhibited α5β1-mediated intracellular signaling, cell adhesion, migration, endothelial cell tubulogenesis, and induced apoptosis of endothelial and tumor cells. Compared to a wild type IgG1 of identical antigen specificity (wt IgG1), PF-04605412 induced a significantly greater tumor and endothelial cell cytotoxicity. ADCC activity correlated with the abundance of NK cells within PBMCs of human donors, but was independent of donor FcγRIIIa polymorphism. In in vivo studies using human xenograft, immuno-xenograft and syngeneic tumor models, PF-04605412 (and an anti-murine α5 surrogate mAb) displayed robust and dose-dependent anti-angiogenesis and anti-tumor efficacy including tumor growth delay, regression and metastasis inhibition, all of which were superior to effects observed with an IgG2 or wt IgG1 of identical antigen specificity. Western blotting and immunohistochemistry analysis of tumor samples showed that anti-tumor efficacy was associated with reduction of total and phosphorylated focal adhesion kinase and positively correlated with α5 expression, infiltrating macrophages, markers of effector cell activity, and apoptosis. Furthermore, depletion of host macrophages abrogated anti-tumor efficacy, indicating a critical contribution of ADCC/phagocytosis to the anti-tumor activity of PF-04605412. The PK-PD relationship, efficacy of PF-04605412 in combination with anti-angiogenic and tumor targeting agents will also be reported. The distinct dual mechanism of action and robust anti-tumor efficacy of PF-04605412 supports the clinical development of this agent for the treatment of human cancers.

Short Title: Dual functional mAb PF-04605412

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 3811.
Abstract 3815: Murine tumors demonstrate variable responses to ischemia-reperfusion injury
Authors: Sheu Eric G. , Wakatsuki Kohei , Oakes Sean M. , Ahmadi-Yazdi Cyrus , Moore Francis D.
Published: February 2011

Introduction: Acute ischemia/reperfusion (I/R) of normal tissues causes an inflammatory reaction initiated by binding of auto-reactive IgM antibody to an ischemia-induced self-antigen. Whether this inflammatory cascade also affects cancers - for example, after acute I/R during surgical resection or in conditions of chronic ischemia - is unclear. We examined the effect of I/R injury in three murine tumor models: B16F10 melanoma, Lewis lung carcinoma (LLC), and small intestinal adenomas of the APCmin/+ mouse.

Methods: LLC and B16F10 cell lines were implanted into the hindlimbs of C57BL/6 mice. Ischemia was induced by external tourniquet ligation of the femoral vessels. Small intestinal tumors in APC min/+ mice were subjected to ischemia by vascular clip occlusion of the superior mesenteric artery. Control animals underwent hindlimb ischemia to the non-tumor bearing hindlimb or laparotomy alone. Following graded times of ischemia and reperfusion, tumors and surrounding normal tissue were harvested for histological examination. Immunohistochemistry was performed for IgM and activated caspase-3. For hindlimb injury, mice were recovered and followed for LLC and B16F10 tumor growth.

Results: Ischemia-reperfusion causes injury of LLC and B16F10 tumors. In contrast, intestinal adenomas of APCmin/+ mice showed no signs of injury following ischemia, despite induction of necrosis and apoptosis of adjacent normal intestinal epithelium. Tumor susceptibility to ischemic injury correlated with immuno-histochemical deposition of IgM antibody. Ischemic injury of B16F10 significantly reduced tumor growth (36% volume reduction compared to control injury, p <0.05). In contrast, LLC tumor growth was significantly increased after ischemia (35% increased compared to control injury, p < 0.01)

Conclusions: The three murine tumor models examined are each affected differently by an acute I/R injury, in contrast to the conserved inflammatory cascade induced in normal tissues following ischemia. APCmin/+ adenomas evade ischemia-induced IgM binding and injury. While both LLC and B16F10 are acutely injured, the long term effects on growth are divergent. Targeted manipulation of the host inflammatory response to ischemia-reperfusion may be a method to influence tumor progression and growth.

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 3815.