bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4444: Targeted delivery of siRNAs to tumor cells and the tumor microenvironment
Authors: da Silva Lígia Catarina Gomes , Ramalho José , Lima Maria Conceição Pedroso , Simões Sérgio , Moreira João Nuno
Published: June 2012

The limited effectiveness of conventional treatment strategies has generated considerable interest on the development of new types of anticancer agents, with improved molecular target specificity. In this context, gene silencing technology constitutes such new class of anticancer agents. The identification of activated oncogenes, as fundamental genetic differences relative to normal cells has made it possible to consider such genes as targets for antitumor therapy.

A tumor is an organ, not a single cell type, which encompasses multiple cell types, each one contributing to the overall tumor aggressiveness. It is now recognized that novel therapeutic strategies should envisage the simultaneous inhibition of multiple targets or pathways, at different cell levels within a tumor, that regulate processes promoting tumor development.

The main goal of this work was to design a novel ligand-mediated targeted lipid-based nanocarrier, containing a nucleic acid (siRNA), aiming at targeting, simultaneously, human breast cancer and endothelial cells from angiogenic vessels.

We have been able to develop a novel ligand-targeted sterically stabilized lipid-based nanoparticle with adequate features for systemic administration: it is characterized by high siRNA encapsulation efficiency, efficient protection of siRNA, average size around 200 nm, and charge close to neutrality.

Our results have shown that the covalent attachment of a specific ligand at the extremity of poly(ethylene glycol) chains, brings a major advantage as it significantly improves the internalization of the lipid-based nanoparticle by both human cancer cells (MDA-MB-435 and MDA-MB-231) and endothelial cells from angiogenic blood vessels (HMEC-1). Moreover, it was not observed a significant internalization by the non-transformed cell line, BJ fibroblasts, indicating the cellular specificity of the developed targeted nanoparticle.

In order to evaluate the potential of the developed targeted nanoparticle to silence a target gene, the green fluorescent protein (GFP)-overexpressing MDA-MB-435 cells were used. A specific downregulation of GFP, both at the protein and mRNA levels, was further observed with the targeted nanoparticle when compared with the non-targeted counterpart.

As the developed nanoparticle is adequate for the encapsulation and delivery of any siRNA sequence, studies with a siRNA targeting a validated molecular target are now ongoing.

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 4444. doi:10.1158/1538-7445.AM2011-4444
Abstract 4448: Rational design of a novel nanocarrier for doxorubicin delivery in cancer treatment
Authors: Luo Juntao , Xiao Kai , Lee Joyce S. , Li Yuanpei , Lam Kit S.
Published: June 2012

Doxorubicin is a very important anti-cancer drug and is currently being used in the treatment of many hematologic malignancies and solid tumors. There is a great need to develope a more effective nanoformulation of doxorubicin that can overcome cardiotoxicity of doxorubicin, as well as the problems associated with Doxil (PEGylated liposomal doxorubicin): large particle size, therefore unable to penetrate deep into the tumor, foot and hand syndrome, and myelotoxicity. We have recently developed a novel biocompatible telodendrimer (PEG block dendritic oligomer of cholic acids), which can self-assemble into stable micelles (∼50 nm) in water when mixed with a hydrophobic drug, such as paclitaxel, with high loading capacity. However, the doxorubicin delivery using this formulation suffers from lower drug loading capacity, less stability and series hemolytic properties. We have optimized the structure of the telodendrimer via introducing the specific building blocks, which has strong interactions with doxorubicin through TT- TT interaction and the hydrogen bonding. The engineered telodendrimer showed better doxorubicin drug loading capacity (4 mg/mL) and greater physical stability (18 nm, stable >3 months) compared with the original formulation. Nanocarrier is nontoxic in cell culture even at >1 mg/mL concentration. The doxorubicin-loaded formulation exhibited the similar potency as the free doxorubicin and Doxil in killing of cancer cells in the cell culture. This engineered formulation after loaded with doxorubicin can completely avoid the hemolysis in the in vitro red blood cell incubation. The in vivo biodistribution and the in vivo antitumor efficacy of this novel DOX-nanotherapeutics will be further studied in the animal 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 4448. doi:10.1158/1538-7445.AM2011-4448
Abstract 4450: Cisplatin intercalated zirconium phosphate nanoparticles are cytotoxic to MCF-7 breast cancer cells via apoptosis
Authors: Gonzalez Millie L. , Ortiz Mayra , Baez Adriana
Published: June 2012

Purpose: Nanoparticle-mediated drug delivery has great potential for anticancer therapy. The purpose of this study was to determine the effectiveness of zirconium phosphate (ZrP) nanoparticles as carrier of the cytotoxic drug cisplatin in MCF-7 breast cancer cells.

Methods: Cytotoxicity of ZrP, ZrP:cisplatin, and cisplatin were studied using the MTT Assay. Cell cycle analyses and apoptosis were analyzed using flow cytometry technology.

Results: ZrP nanoparticles were not cytotoxic to MCF-7 cells. However, cell viability was reduced to 70% when MCF-7 cells were grown with ZrP: cisplatin 100μM. Cisplatin was cytotoxic to MCF-7 cells with an IC50 of 10 μM. The cytotoxic effect of cisplatin: ZrP is mediated by induction of apoptosis. AnnexinV/PI treated MCF-7 cells showed the apoptotic events in cisplatin -induced apoptosis. ZrP nanoparticles alone did not cause apoptosis to MCF-7 cells or peripheral blood lymphocytes.

Conclusion: ZrP nanoparticles can significantly reduce side effects associated to systemic toxicity of anticancer agents and more studies are warranted.

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 4450. doi:10.1158/1538-7445.AM2011-4450
Abstract 446: Inhibition of peroxisome proliferator-activated receptor-gamma in myeloid lineage cells induces systemic inflammation, immunosuppression and tumorigenesis
Authors: Du Hong , Wu Lingyan , Yan Cong
Published: June 2012

Inflammation is a major mechanism to protect organisms from damage by responding to pathogen infection and tissue injury. However, failure to clear excessive amounts of inflammatory cells, or to control exuberant expression of pro-inflammatory molecules at the inflammatory sites can cause organ damage and cancer. Therefore, it is important to identify anti-inflammatory molecules and pathways that suppress overexpression of inflammatory molecules and eliminate excessive inflammatory cells to prevent chronic inflammation-induced cancer formation. At the cellular level, malformation and malfunction of myeloid cells contribute to inflammation and cancer formation. Peroxisome proliferator-activated receptor-γ (PPARγ) is an anti-inflammatory molecule. In order to study its biological function in myeloid cells, dominant negative PPARγ (dnPPARγ) was overexpressed in a myeloid-specific c-fms-rtTA/(TetO)7-CMV-dnPPARγ bitransgenic mouse model. In this bitransgenic system, overexpression of dnPPARγ-Flag fusion protein in myeloid lineage cells abnormally elevated frequencies and total numbers of LK, LSK, CMP and GMP progenitor populations in the bone marrow. DnPPARγ overexpression led to up-regulation of PPARγ downstream genes IL-1β, IL-6 and TNFα in the blood plasma, which are known to stimulate development and expansion of myeloid-derived suppressor cells (MDSCs). As a result, CD11b+Gr-1+ MDSCs were systemically increased in association with activation of oncogenic intracellular signaling molecules Stat3, NF-κB, Erk1/2 and p38 in doxycycline-treated bitransgenic mice. MDSCs from doxycycline-treated bitransgenic mice strongly inhibited proliferation and function of wild type CD4+ T cells in vitro. CD4+ T cells from doxycycline-treated bitransgenic mice showed much weaker activities of proliferation and lymphokine release function. As a result, both CD4+ and CD8+ T cell populations were decreased in doxycycline-treated bitransgenic mice. The bone marrow transplantation study indicated that the myeloid autonomous defect was responsible for MDSC expansion and immunosuppression. Multiple forms of carcinoma and sarcoma in the lung, liver, spleen and lymph nodes were observed in doxycycline-treated bitransgenic mice. These studies suggest that anti-inflammatory PPARγ in myeloid lineage cells plays a key role in controlling pro-inflammatory cytokine syntheses, MDSCs expansion, immunosuppression and cancer formation. Studies outlined here will have a significant impact on how to design a strategy to combat inflammation-induced cancer and pave the way for novel immunotherapy.

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 446. doi:10.1158/1538-7445.AM2011-446
Abstract 4466: Characterization of the pan class I PI3K inhibitor NVP-BKM120 mechanism of action across a broad range of concentrations
Authors: Brachmann Saskia M. , Kleylein-Sohn Julia , Gaulis Swann , Stauffer Frederic , Voshol Johannes , Guthy Daniel , Wilson Christopher , Huang Alan , Wartmann Markus , Bruemmendorf Thomas , Chene Patrick , Fritsch Christine , Hackl Wolfgang , Cosaert Jan , Schnell Christian , Hofmann Francesco , Maira Michel
Published: June 2012

NVP-BKM120 is a pan class I PI3K inhibitor that has recently entered phase II clinical trials. The compound was shown to inhibit cell proliferation and survival of cancer models displaying PI3K pathway dependency, in a dose-dependent manner, and proportionally to the extent of pathway inhibition. To further characterize NVP-BKM120, we have investigated its mechanism of action across a broad range of relevant models and concentrations of the molecule and compared it to other PI3K inhibitors (e.g. GDC0941 and ZSTK474). The effects observed on phenotypical read-outs were similar for all compounds, when tested up to concentrations necessary to achieve near complete pathway inhibition (IC90 for Akt-S473P). More profound effects were however observed with NVP-BKM120, at higher concentrations (>2 micromolar), in PI3K-independent models, suggesting that at these dose levels, NVP-BKM120 might display inhibitory activities other than PI3K.

In order to determine this potential off-target activity, a gene expression profiling study was performed in a PI3K insensitive model, comparing effects of GDC0941 and NVP-BKM120 at equipotent concentrations. Gene-Set Enrichment Analysis (GSEA) revealed that NVP-BKM120 at the highest dose only (3.6 micromolar, 2 fold above the S473-Akt IC90 of PI3K sensitive models), led to increased expression of genes involved in G2 and mitotic (M) phases. Subsequent FACS analysis showed that in contrast to the other pan-PI3K inhibitors, NVP-BKM120 was indeed able to induce a strong G2/M arrest in several PI3K non addicted cell lines when used at concentrations higher than 2 micromolar. DAPI and tubulin immuno-histochemistry studies showed that the NVP-BKM120 induced block was phenotypically similar to that of Nocodazole, suggesting effects on spindle dynamics in prometaphase. Indeed, in cellular or in in vitro purified systems, NVP-BKM120 greatly reduced microtubule polymerization.Based on analysis the antitumor activity observed in vivo in PI3K-dependent animal models, it appears that efficacy is solely due to pure PI3K inhibition, as these off-target activities are generally observed at concentrations (corrected for free fraction) that could not be achieved in animals. Based on modeling of human PK data, a similar conclusion can be reached for patients, as the exposure currently observed in plasma does predict sole coverage of PI3K inhibitory activities.

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 4466. doi:10.1158/1538-7445.AM2011-4466
Abstract 447: A mechanism of hypoxia-induced immune escape in cancer cells
Authors: Hamilton Thomas K. , Barsoum Ivraym , Li Xin , Miles Ellen A. , Elliott Janet , Siemens D. Robert , Graham Charles H.
Published: June 2012

We previously showed that hypoxia induces resistance to natural killer (NK) cell-mediated lysis in tumor cells through a mechanism that involves the shedding of NK cell-activating ligands (MICA) from the cell surface. Furthermore, we were able to block this hypoxia-induced shedding of MICA, as well as resistance to NK cell-mediated lysis, by activating NO signaling in the cancer cells. The objective of the present study was to fully elucidate this mechanism of hypoxia-induced immune escape. We used siRNA knock-down to determine the role of hypoxia-inducible factor-1 (HIF-1), a transcriptional activator of hypoxia-inducible genes, and ADAM10, a metalloproteinase known to be involved in the shedding of MICA, in the hypoxia-induced immune escape in DU145 prostate cancer cells and MDA-MB-231 breast cancer cells. Our findings revealed that in vitro exposure to hypoxia increases ADAM10 expression in a HIF-1-dependent manner, and that knock-down of ADAM10 abolishes the hypoxia-induced MICA release and resistance to NK cell-mediated lysis. Treatment of the tumor cells with low concentrations of nitroglycerin inhibited the hypoxia-induced accumulation of HIF-1α, ADAM10 up-regulation, the shedding of the NK-activating ligands, and resistance to NK cell-mediated lysis. More importantly, continuous transdermal administration of nitroglycerin (1.8 µg/h) to nude mice attenuated the growth of transplanted human DU145 prostate tumors; however, this effect of nitroglycerin was lost in mice made deficient in NK cells by treatment with anti-asialo GM1 antibody. This latter finding indicates that nitroglycerin attenuates tumor growth by sensitizing tumor cells to innate immunity. These findings are important because they indicate that nitric oxide mimetics could potentially be used as immunosensitizers in the treatment and/or prevention of 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 447. doi:10.1158/1538-7445.AM2011-447
Abstract 4473: Sorafenib suppresses postsurgical recurrence and metastasis of hepatocellular carcinoma in an orthotopic mouse model
Authors: Feng Yu-Xiong , Zhao Jiang-Sha , Li Jing-Jing , Xie Dong
Published: June 2012

Surgical resection is the first-line treatment for hepatocellular carcinoma (HCC) patients with well-preserved liver function. Nevertheless, postoperative recurrences occur up to 70% at 5 years, gravely jeopardizing the therapeutic outcome. Clearly, new approaches are needed for preventing relapse of this deadly disease. Taking advantage of a luciferase-labeled orthotopic xenograft model of HCC, we examined the role of Sorafenib, the first approved systemic drug for advanced HCC patients, in prevention of HCC recurrence. We found that Sorafenib suppressed the development of postsurgical intrahepatic recurrences, as well as abdominal metastasis, consequently leading to a prolonged postoperative survival of mice in this model. Furthermore, hyperactivity of ERK signaling caused by elevated levels of growth factors associated with postoperative liver regeneration enhanced the sensitivity of HCC cells to Sorafenib, providing a plausible explanation to the observation that recurrent tumors are more responsive to growth inhibition by Sorafenib. Our results strongly suggest a potential application of Sorafenib in early stage HCC patients who have received hepatectomy with curative intention, by effectively reducing postoperative recurrences.

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 4473. doi:10.1158/1538-7445.AM2011-4473
Abstract 4474: Cellular effects of fluorescent quinolinium drugs on normal lymphoblast and A431 carcinoma cells
Authors: Velez Christian , Molina Denisse , Rosado Carlos , Cox Osvaldo , Zayas Beatriz
Published: June 2012

This study aims to evaluate the cellular effects of amino benzazolo [3,2- a] quinolinium drugs on normal lymphoblast cells and compare them to effects on A431 epitheloid carcinoma cells. The drugs under study are amino containing heterocyclic compounds possessing a positive charge that could facilitate their interaction with cell organelles specially mitochondria. The fluorescent properties could also facilitate observation of their cellular binding location. These compounds have also been demonstrated to have mutagenic and DNA binding capacity on A431 cancer cells. A431 and TK6 cells in culture where exposed for 48 hours to determine the IC50 dose, mitochondrial membrane permeability, ROS generation and fluorescent microscopy analysis of intracellular binding. IC50 was determined by trypan blue exclusion and mitochondrial membrane permeability applying the Nucleo counter 3000, JC-1 assay. For ROS determination the fluorescent dye 2,7-dichlorofluorescein diacetate (DCFH-DA) was applied. Determination of intracellular binding was performed applying confocal microscopy. In the TK6 normal lymphoblast cells the observed IC50 concentrations were as follow: ABQ38=310µM, and ABQ95=270µM. On A431 IC50s were as follows: ABQ38 = 32 µM and ABQ95 = 36 µM. Overall, normal TK6 demonstrated higher tolerance to the tested ABQS than A 431. Preliminary results on ROS generation indicated a stronger ROS release on A431 than on TK6 lymphoblast. Analysis of the effects of ABQS on the mitochondrial permeability where also consistent with the cytotoxicity observed, demonstrating higher levels of membrane permeability on treated tumor cells than on normal. Microscopy images of ABQS treated cells revealed location of these fluorescent compounds in organelles such as mitochondria and DNA. DNA fragmentation analyses are being implemented. This study presents preliminary evidence of an overall higher tolerance of normal TK6 lymphoblast to the tested fluorescent drugs in comparison to treated A431.

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 4474. doi:10.1158/1538-7445.AM2011-4474
Abstract 4476: The effect of electrochemotherapy on metastatic potential of human melanoma cells SK-Mel28
Authors: Cemazar Maja , Todorovic Vesna , Mlakar Vid , Glavac Damjan , Sersa Gregor
Published: June 2012

Electrochemotherapy is a local treatment, combining chemotherapy and application of electric pulses to the tumour. Electrochemotherapy with bleomycin or cisplatin has demonstrated its effectiveness in controlling local tumour growth in the treatment of malignant melanoma and is proposed as one of the treatment options for in-transit melanoma metastases in Europe. An important aspect of successful treatment is prevention of metastasis, however, it is known that metastasis can be induced by different treatment modalities. Alterations in tumour microenvironment caused by surgery or radiation therapy, can lead to increased formation of metastasis. Furthermore, formation of metastases can be directly linked to migratory potential of the cells. Although electrochemotherapy is now routinely used for treatment of various cancers in many cancer centres, the effect of electrochemotherapy on the metastatic potential of tumour cells is not known. Therefore, the aim of this study was to evaluate the effect of electrochemotherapy with bleomycin and cisplatin on the metastatic potential of human malignant melanoma cells.

Human malignant melanoma cells SK-MEL 28 were treated by electrochemotherapy with a range of bleomycin concentrations from 0.01 nM to 1μM and cisplatin concentrations from 2 to 80 μg/mL. SK-MEL 28 cells that were viable 48 h after electrochemotherapy were tested for their metastatic potential by ability to migrate and invade through Matrigel coated porous membrane. In addition, cell adhesion and proliferation using MTS assay were also tested. Furthemore, RNA was isolated from the cells 16 h after each treatment and differentially expressed genes were investigated by microarray analysis to evaluate the effect of electrochemotherapy with bleomycin or cisplatin on expression of genes involved in the development of cancer. The Gene Ontology Tree Machine program was used for gene enrichment analysis.

Migration and invasion of melanoma cells that survived 48h after electrochemotherapy with bleomycin or cisplatin were not changed compared to control untreated cells. Also, there were no changes observed in cell adhesion on Matrigel. Gene expression analysis demonstrated that a very low number of genes were differentially expressed after electrochemotherapy with either bleomycin or cisplatin. Specifically, the expression of metastasis promoting genes was not increased after electrochemotherapy.

Our data suggest that electrochemotherapy with bleomycin or cisplatin does not increase metastatic behaviour of human melanoma cells, having important clinical implication for safe use in treatment of melanoma metastases.

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 4476. doi:10.1158/1538-7445.AM2011-4476
Abstract 4477: Characterization of AZD5363, an orally bioavailable, potent ATP-competitive inhibitor of AKT kinases with pharmacodynamic and antitumor activity in preclinical models
Authors: Davies Barry R. , Luke Richard , Dudley Phillippa , Greenwood Hannah , Crafter Claire , Cosulich Sabina , Page Ken , Yates James , Lane Clare , Yu Derek , Zhang Jingchuan , Pass Martin
Published: June 2012

AKT is a key node in the most frequently de-regulated signaling pathway in human cancer, and has been shown to mediate resistance to a range of cytotoxic, anti-hormonal and targeted therapies. AZD5363 inhibits all AKT isoforms with a potency of <10 nM, and inhibits phosphorylation of the AKT substrate PRAS40 in BT474c cells with a potency of 0.31 μM. AZD5363 monotherapy inhibits the proliferation of 22/150 solid and hematological tumour cell lines with a potency of < 1μM. Cell lines derived from breast cancers show the highest frequency of response. Pharmacodynamics have been studied in BT474c xenografts growing in nude mice following oral dosing. A dose of 300 mg/kg AZD5363 significantly inhibits pPRAS40, pGSK3β and pS6 for at least 24 h; pPRAS40 was inhibited by 70-90% for 24 h following a 300 mg/kg dose. These biomarkers were also significantly inhibited for at least 8 h following a 100 mg/kg dose. A good correlation was observed between plasma exposure of AZD5363 and inhibition of PRAS40 phosphorylation (EC50 ∼ 0.1 μM). Chronic dosing of AZD5363 causes dose-dependent inhibition of BT474c xenografts; schedules of 300 mg/kg qd and 150 mg/kg bd resulted in >90% inhibition of BT474c xenograft growth. Chronic monotherapy dosing resulted in profound growth inhibition or regression of multiple other xenografts derived from a range of solid tumour types. AZD5363 also greatly enhanced the anti-tumor activity of cytotoxic and novel targeted therapies in xenograft models. It is concluded that AZD5363 is a potent inhibitor of AKT with pharmacodynamic and anti-tumor activity in vivo, and has potential to treat a range of solid and hematological malignancies. AZD5363 is currently in phase 1 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 4477. doi:10.1158/1538-7445.AM2011-4477
Abstract 448: Mechanism of action differences of transmembrane versus secretory tumor necrosis factor-α on tumor growth; role of tumor-associated myeloid cells
Authors: Ardestani Shidrokh , Li Bin , Deskins Desirae , Young Pampee
Published: June 2012

Tumor Necrosis Factor alpha (TNF-α) is a major inflammatory cytokine, produced by a wide variety of cell types including macrophages, T lymphocytes endothelial cells and tumor cells. TNF-α is expressed as a membrane-tethered protein (tmTNF-α), cleaved by tumor necrosis factor-α-converting enzyme (TACE) and subsequently released into extracellular space as a soluble TNF-α (sTNF-α). Recent studies from our laboratory and others have shown that low level expression of TNF-α by tumor cells substantially increases tumor growth, vascularity and decreases tumor necrosis, in part by increasing tumor content of a proangiogenic myeloid subtype (Li et al., Cancer Res. 2009, 1;69(1):338-48). To better understand the mechanism by which TNF-α modulates the myeloid phenotype, we generated murine Lewis lung carcinoma (LLC) and B16F10 melanoma tumor cells transduced with retrovirus expression vectors directing modest overexpression of a membrane tethered moiety, tmTNF-α. Heterotopic implantation of tmTNF-α expressing lines resulted in significantly decreased growth than control tumors in both tumor models. Our studies showed that this was not due to effects on the tumor cells themselves but likely due to effects on tumor stroma as the effect of tmTNF-α was lost in TNF-α receptor deficient mice. Further evaluation showed an absence of tumor associated myeloid cells within the tumor, which suggested apoptosis of migrated myeloid cells or inhibition of myeloid recruitment into the tumor environment. There is a significant gap in our knowledge on the mechanism of transmembrane isoform of TNF-α and its potential anti-tumorogenic role. In this study our goal is to identify the mechanism in which tmTNF-α promotes tumor regression and to understand how it modulates host bone marrow derived myeloid cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 448. doi:10.1158/1538-7445.AM2011-448
Abstract 4480: Combination of a PI3K/mTOR inhibitor and a MEK inhibitor at suboptimal dose significantly inhibits growth of lung adenocarcinoma tumors in KrasG12D-LSL mice
Authors: Shojaei Farbod , Simmons Brett H. , Lee Joseph H. , Wong Anthony , Los Gerrit , Kan Julie , Christensen James G.
Published: June 2012

The KrasG12D-LSL conditional GEMMs (genetically engineered mouse models) are a valuable tool in translational research to study tumor progression and to investigate efficacy of anti-tumor compounds in a variety of cancer types. Here, intranasal delivery of adeno-Cre resulted in the expression of mutant Kras leading to development of tumor lesions in the lungs including adenomatous hyperplasia, large adenoma and adenocarcinoma. Treatment of tumor bearing mice at adenocarcinoma stage (at 14 weeks post adeno-cre inhalation) with suboptimal dose of PF-04691502 (PI3Ka/mTOR inhibitor; 7.5 mg/kg; SID) and PD-0325901 (MEK inhibitor, 1.5 mg/kg; SID) reduced tumor lesions in mice treated with single agents and significantly inhibited tumor growth in mice treated with combination of both compounds. In addition, immunohistochemistry showed inhibition of pS6 in the treated animals particularly in the combination group providing a proof of mechanism for the tumor growth inhibition. Similar response was observed when mice bearing H460 (carry Kras and PIK3CA mutations) orthotopic lung tumors were treated with the combination regimen. These observations provide some clue to further pursue efficacy of PF-04691502 plus PD-0325901 in the clinical settings and also in more invasive models such as KrasG12D-LSLp53flox/flox mice.

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 4480. doi:10.1158/1538-7445.AM2011-4480
Abstract 4483: SR13668, a selective 12-lipoxygenase inhibitor, suppresses tumor growth by modulating the Akt signaling pathway
Authors: Jong Ling , Chao Wan-Ru , Green Carol , Kapetanovic Izet , Sambucetti Lidia , Collins Nathan
Published: June 2012

SR13668, an orally active cancer therapeutic agent, exhibits potent antitumor activity against various human cancer xenografts (e.g., breast, prostate, lung and ovarian cancer) without significant toxicity. An exploratory IND study of SR13668 has been completed with a two-stage design to compare the bioavailability of five SR13668 formulations, and the preferred formulation will be subjected to investigation in subsequent phase I testing.

SR13668 inhibits growth factor-stimulated phosphorylation of Akt and its downstream target GSK3β in PC-3 prostate and high pAkt expressing MDA-MB-468 breast cancer cells in a dose-dependent manner. SR13668 was also shown to block FGF- and IGF-stimulated Akt activation. Screening a broad selection of kinase targets, including Akt(1,2,3), PI3K, and PDK1, indicated that SR13668 is not a kinase inhibitor. Preliminary mechanistic studies showed that SR13668 selectively inhibits 12-lipoxygenase (12-LOX), but does not affect other lipoxygenases (5-LOX and 15-LOX), or cyclooxygenases (COX-1 and COX-2).

12-LOX plays a major role in promoting cancer progression, invasion and metastasis as well as tumor angiogenesis. 12-LOX has been regarded as a novel target for cancer therapy, since it is generally absent in normal epithelia and is often constitutively expressed in various epithelial cancers. Unlike direct inhibitors of PI3K or Akt, SR13668 has no adverse effects on fasting glucose levels or body weights after 14 days of oral treatment with SR13668 at 500 mg/kg/day, a dose more than 10 times higher than that needed for antitumor activity.

Pharmacologic intervention studies with lipoxygenase inhibitors for the prevention or inhibition of cancers have been difficult to interpret, as there are very few isoform-selective LOX inhibitors available. SR13668 is a highly specific 12-LOX inhibitor. No off-target activities were observed, when screened against a broad collection of 40 diverse enzymes and 80 transmembrane and soluble receptors, ion channels and monoamine transporters. The Phase 0 clinical study showed that SR13668 has long half-life (11.2 ± 3.1 hours), and patients did not experience significant adverse events related to SR13668 treatment; thus SR13668 is a promising candidate for further clinical investigation.

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 4483. doi:10.1158/1538-7445.AM2011-4483
Abstract 4490: Tocotrienols inhibit PI3/Akt and ERK pathways to induce growth arrest in pancreatic cancer cell lines by downregulation of Her-2/ErbB2 receptors
Authors: Chakraborty Kanishka , Ramsauer Victoria , Kang S Shin , Lightner Janet , Campbell S , Stone WL , Krishnan Koyamangalath
Published: June 2012

Abstract: Pancreatic cancer is the fifth leading cause of cancer death in the United States. Alternate strategies to treat and prevent this cancer are urgently required. In this regard, novel therapeutic agents that can inhibit signaling pathways implicated in the proliferation and survival of pancreatic cancer cells are of immense interest. Tocotrienols are members of the Vitamin E family but, unlike tocopherols, possess an unsaturated isoprenoid side-chain that confers superior anti-cancer properties. The ability of tocotrienols to selectively inhibit the HMG CoA reductase pathway through post-translational degradation of HMG CoA reductase and to suppress the activity of transcription factor NFκβ could be the basis for some of these properties. Data: Our studies indicate that γ- and Δ-tocotrienol have potent anti-proliferative activity in pancreatic cancer cells (Panc-28, MIA-PACA-2 and BXPC-3). Indeed both tocotrienols induced cell death (> 50%) by the MTT cell viability assay in all three pancreatic cancer cell lines. We also examined the effects of the tocotrienols on the Akt and the Ras/Raf/MEK/ERK signaling pathways by Western blotting analysis. γ- and Δ-Tocotrienol treatment of cells reduced the activation of ERK MAP kinase and that of its downstream mediator RSK (ribosomal protein S6k) in addition to suppressing the activation of protein kinase, Akt. Suppression of activation of Akt by gamma-tocotrienol led to downstream upregulation of Fox-03 and GSK-3b. These effects were mediated by the downregulation of Her2/ErbB2 at the messenger level. Tocotrienols but not tocopherols were able to induce the observed effects. Conclusion: Our results suggest that the tocotrienol isoforms of vitamin E can induce apoptosis in pancreatic cancer cells through the suppression of vital cell survival and proliferative signaling pathways such as those mediated by the PI3/Akt and ERK/MAP kinases via downregulation of Her2/ErbB2 expression. The molecular components for this mechanism are not completely elucidated and needs further investigation.

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 4490. doi:10.1158/1538-7445.AM2011-4490
Abstract 4496: Preclinical efficacy of the novel dual PI3K/mTOR inhibitor, BEZ235 compared with RAD001 in HER2+ breast tumors
Authors: De Pradip K.R , Sun Yuliang , Dey Nandini , Leyland-Jones Brian
Published: June 2012

Background: HER2 overexpression occurs in 20-25% of patients with breast cancer and is associated with aggressive disease and decreased survival. HER2 activates multiple cellular signaling pathways including the PI3K-AKT-mTOR pathway. The sustained activation of the PI3K-AKT-mTOR pathway or the activating mutation of PIK3CA (p110α) is one of the major causes of drug resistance including trastuzumab.

Purpose: Inhibitors of mTOR1 have been shown to be active in patients with solid tumors. As the phosphatidylinositol 3-Kianse (PI3K) pathway activates numerous other kinases, transcription factors, and proteins associated with cell growth/survival and apoptosis besides mammalian target of rapamycin (mTOR), suppression of this pathway upstream of mTOR may be more effective than inhibition of mTOR1 in HER2+ breast cancer models.

Experimental Design: To examine this possibility, the dual pan-PI3K/mTOR inhibitor, BEZ235 was compared with mTOR1 inhibitor, RAD001 in trastuzumab-sensitive HER2+ (BT474 and SKBR3), trastuzumab-resistant (BT474HR), and PIK3CA mutated/HER2+ breast cancer cells (HCC1954 and U893)

Results: Treatment of HER2+ breast cancer cell lines with BEZ235 or RAD001 in vitro showed 1) RAD001 mediated AKT activation was blocked when cells were treated with BEZ235, 2) BEZ235 dose- and time-dependently blocked AKT phosphorylation (both at Ser473 and Thr308), 3) both drugs completely blocked the activation P70S6K, but BEZ235 more efficiently blocked 4EBP1 phosphorylation than RAD001, 4) ERK activation was significantly attenuated by BEZ235, while inhibition of mTOR1 by RAD001 lead to activated ERK, 5) BEZ235 was significantly more effective than RAD001 in reducing HIF1α expression following heregulin stimulation, 6) activation of caspase3 (from the expression level of cleaved caspase3) was also greater following treatment with BEZ235 than RAD001, and 7) integrin-dependent breast tumor cell migration was significantly abrogated with BEZ235 in association with inhibition of RAC1-GTP, but this was not seenwith RAD001.

Conclusion: Our preclinical in vitro data demonstrate that concurrent inhibition of PI3K and mTOR is likely to be a more effective strategy in the treatment of both sensitive and resistant HER2+ breast cancers than the inhibition of mTOR1 alone.

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 4496. doi:10.1158/1538-7445.AM2011-4496
Abstract 4499: HM5016699, a novel and selective PI3K/mTOR dual inhibitor
Authors: Ren Yongxin , Zhang Weihan , Cui Yumin , Guo Mingchuan , Long Jingwen , Sai Yang , Wang Jian , Yan James , Li Jia , Qing Weiguo , Su Weiguo
Published: June 2012

PI3KCA gene amplification and mutations have been identified in many types of tumors, suggesting the dysregulation of PI3K/AKT/mTOR axis plays pivotal roles in tumorigenesis and development. Reported here is the discovery of HM5016699 as a potent, reversible ATP-competitive PI3K/mTOR dual inhibitor for treatment of cancer. HM5016699 showed potent kinase inhibition with IC50 of 0.01, 0.206, 0.052, 0.051 and 0.059 µM on PI3Kα, β, γ, Δ and mTOR, respectively. It demonstrated high selectivity on PI3K family and mTOR over a panel of 274 kinases. Of the 67 tumor cell lines screened from lung, colon, breast, ovarian, and other tumor types, we found that the tumor cells with HER2 gene amplification or PTEN loss tended to be sensitive to HM5016699, while the tumor cells harboring PI3KCA mutations and wild type Ras/Raf were mostly sensitive to the compound. HM5016699 could also show p-Erk inhibition and apoptotic induction at higher concentrations in the tumor cells harboring Ras/Raf mutations, suggesting it might bring benefits to patients carrying Ras/Raf mutations in the clinic. Consistent with in vitro studies, HM5016699 exhibited dose dependent tumor growth inhibition on multiple human xenografts models. It was found that the tumors with PIK3CA mutation or PTEN deletion were highly sensitive to HM5016699 with ED50s ranging from 0.6 to 1.8 mg/kg in U87MG (pTEN null), SKOV3 (H1047) and H1975 (G118D), whereas in tumors with Ras/Raf mutations, such as DLD-1(G13D), A549 (G12S) and HT-29(V600E), its ED50s were from 3.0 to 15.0 mg/kg. The decreased phosphorylation of AKT and S6 was observed in the tumor xenograft tissues treated with HM5016699, which was well-correlated with compound exposure. Therefore, we concluded that HM5016699 demonstrated both in vitro and in vivo activity through acting on PI3K/mTOR pathway. HM5016699 exhibited a favorable pharmacokinetic profile and acceptable safety window in rats. The pre-clinical study results suggest that HM5016699 could be a promising anti-cancer agent.

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 4499. doi:10.1158/1538-7445.AM2011-4499
Abstract 45: Accurate total and allele-specific copy number measurements in mosaic tumors
Authors: Curry Bo , Sampas Nick , Anderson Paige , Peter Brian
Published: June 2012

For the characterization of genomic copy number changes that occur in the development and progression of cancer, oligonucleotide array comparative genomic hybridization (aCGH) offers high-resolution and precise determination of chromosomal copy number and genome-wide aberrations. We recently reported a new method for making simultaneous measurements of single nucleotide polymorphisms (SNPs) and copy number alterations in the same microarray assay. The combined assay can detect copy-number neutral events, such as acquired loss of heterozygosity (LOH), as well as allelic imbalance in and around amplified regions.

Previously reported algorithms for analyzing Agilent CGH+SNP data were able to genotype primarily diploid samples and to detect regions of constitutional LOH. However, tumor heterogeneity, aneuploidy and variable sample quality create significant challenges for both solid and liquid tumors. Our earlier methods were often unable to cope with the high levels of aneuploidy sometimes found in solid tumors, or with admixtures of normal cells and aberrant cells. We now describe new computational methods which can determine total copy number, aneuploid fraction, and allele-specific copy number in many aneuploid tumor samples, even when mixed with up to 80% normal tissue.

We report results from genomic DNA isolated from a cancer cell line, a blood sample, and a solid tumor. Each of these sample types presents novel challenges. The cell line we studied, though largely monoclonal, is highly aneuploid. The blood DNA is of high quality, but the fraction of tumor cells in the sample is low. The solid tumor DNA is of relatively low quality, and composed of multiple aberrant clones. We were able to determine the aneuploid fraction of the tumors, and to measure copy number variation in samples with as little as 20% tumor cell content. In addition to detecting copy-neutral LOH regions, we also measured allele-specific copy number, both of the aberrant clone and of admixed normal cells.

The new analysis methods allow the extension of the extra allelic information available from the CGH+SNP assay to cancer samples.

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 45. doi:10.1158/1538-7445.AM2011-45
Abstract 4505: Development of small molecules to target ezrin as anti-metastatic agents
Authors: Bulut Gülay , Hong Sung-Hyeok , Chen Kevin , Rahim Said , Kosturko George W. , Beauchamp Elspeth M. , Glasgow Eric , Toretsky Jeffrey A. , Khanna Chand , Uren Aykut
Published: June 2012

The most common cause for mortality in osteosarcoma (OS) is respiratory failure due to metastasis to lungs. Thus targeting underlying molecular events that lead to metastasis can provide a significant benefit to patients with fatal metastatic disease. Accumulating evidence from experimental animal models and human cases suggests that ezrin is a key factor in the metastasis of OS cells. Ezrin is a multifunctional protein that connects the actin cytoskeleton to extracellular matrix through transmembrane proteins. We screened small molecule libraries for compounds that directly interact with ezrin protein. Two small molecules that directly bind to ezrin with low micromolar affinity were selected based on inhibiting ezrin function in multiple assays. They inhibited ezrin phosphorylation, ezrin-actin interaction, and ezrin mediated motility of OS cells in culture. These compounds were unique in their mechanism of action such that they inhibited in vitro phosphorylation of ezrin by protein kinase C (PKC) by binding to the substrate not by inhibiting the kinase activity of the enzyme. PKC was able to phosphorylate other substrates in the presence of these novel ezrin inhibitors. They also mimicked the ezrin morpholino phenotype and caused a unique developmental defect consistent with reduced cell motility in zebrafish. Following tail vein injection of osteosarcoma cells to mice, both molecules inhibited lung metastasis of ezrin-sensitive cells, but not ezrin-resistant cells. These two molecules demonstrate a novel targeted therapy that directly inhibits ezrin protein as an approach to prevent tumor metastasis in osteosarcoma and other tumors with elevated ezrin in their metastatic subclones.

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 4505. doi:10.1158/1538-7445.AM2011-4505
Abstract 451: Macrophages are critical for promotion of urethane-induced lung carcinogenesis during early stages
Authors: Zaynagetdinov Rinat , Sherrill Taylor P. , Polosukhin Vasiliy V. , Han Wei , McMahon Frank B. , Gleaves Linda A. , Ausborn Jamie A. , McLoed Allyson G. , Yull Fiona E. , Blackwell Timothy S.
Published: June 2012

Among the multiple factors initiating and supporting tumor growth, inflammation plays one of the most important roles. Macrophages, which are the primary immune cells in the lungs, have established roles in tumor growth and metastasis; however, information about the role of macrophages in early carcinogenesis is limited. The purpose of this study was to evaluate the role of macrophages in modulation of tumor formation during different stages of urethane-induced lung carcinogenesis. To assess the role of macrophages in our lung tumor model, we developed a method of non-invasive long-term lung macrophage depletion by repetitive intratracheal injections of liposomal clodronate. Clodronate encapsulated liposomes are well known to cause selective apoptosis of macrophages. Depletion of macrophages during the entire experiment (4 Months) or during the late promotion and progression stages (Months 2-4), when macrophages primarily exhibited a M2 phenotype, significantly reduced tumor vasculogenesis and lung tumor multiplicity. Interestingly, depletion of alveolar macrophages during lung tumor initiation and early promotion stages (Week 1-2), when macrophages were polarized toward an M1 phenotype, caused similar attenuation of tumor vasculogenesis and lung carcinogenesis as observed after depletion of cells during the entire experiment or late promotion/progression stages. Moreover, early macrophage depletion, but not late depletion, significantly reduced formation of atypical adenomatous hyperplasia (AAH) lesions in the lungs. Our results demonstrate that macrophages are critical for promotion of lung carcinogenesis during early stages and suggest that the M1/M2 paradigm may not be an important concept for describing the impact of macrophages on tumor promotion.

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 451. doi:10.1158/1538-7445.AM2011-451