bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3415: Therapeutic targeting of the ASPL-TFE3 gene fusion in ASPS-1, a novel cell line derived from metastatic alveolar soft part sarcoma (ASPS)
Authors: Vistica David T. , Kenney Susan , Burkett Sandra , Hollingshead Melinda , Borgel Suzanne , Butcher Donna , Schrump David S. , Shoemaker Robert H.
Published: February 2011

Alveolar soft part sarcoma (ASPS) is a rare soft tissue sarcoma primarily affecting young children and adolescents. ASPS is characterized by a nonreciprocal chromosomal translocation, der(17)t(X;17)(p11;q25), in which the N-terminal region of the alveolar soft part locus (ASPL) gene, located at 17q25, is fused to the C-terminal region of the transcription factor E3 (TFE3), located at Xp11. Two alternative fusion junctions have been observed resulting in the expression of 2 distinct fusion transcripts, ASPL-TFE3 type 1 and 2, and their respective proteins which function as hybrid transcription factors. Recently we have described the first xenograft model of ASPS, established from a lymph node metastasis derived from a female patient expressing the ASPL-TFE3 type 1 fusion, in NOD.SCID mice (J. Pediatr. Hematol. Oncol 31:561, 2009) and an ASPS cell line, ASPS-1, derived from it (Proc. Amer. Assoc, Cancer Res. 50:978, 2009). We report here our efforts to target the ASPL-TFE3 type 1 gene fusion (CGG GAG CGG CTG CCT GTG TCA) in ASPS-1 cells using siRNA directed against its nucleotide sequence and compared results with siRNA directed against the ASPL-TFE3 type 2 gene fusion (CGG ATT GAT GAT GTC ATT GAT). ASPL-TFE3 type 1 and 2 siRNA were evaluated in vitro in ASPS-1 cells, which express the ASPL-TFE3 type 1 gene fusion and protein, at concentrations ranging from 1-50 nM and treatment times ranging from 3-12 days. These results indicate that the ASPL-TFE3 type 1 siRNA, but not the ASPL-TFE3 type 2 siRNA or a control siRNA not known to target any human gene, was effective in reducing the ASPL-TFE3 type 1 fusion transcript, the ASPL-TFE3 type 1 fusion protein and retarding the growth of ASPS-1 cells. Importantly the ASPL-TFE3 type 1 siRNA down regulated several signature ASPS genes (BMC Cancer 9:22, 2009) including those involved in angiogenesis of this highly vascular tumor (ANGPTL2, MDK, c-MET, and TIMP-2). Additionally, PCSKI, a proprotein convertase responsible for processing of neuropeptides, growth factors and receptors, and several tissue restricted markers including ITGB1BP3, was strikingly down regulated. Together these results suggest that the ASPL-TFE3 fusion gene and its fusion protein play an important role in pathways involved in the growth of ASPS and present potentially exploitable targets for development of novel therapeutics.

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 3415.
Abstract 3422: Synthetic lethal siRNA screening reveals novel modifiers of insulin-like growth factor-1 receptor (IGF1R) inhibitor activity in childhood sarcomas
Authors: Potratz Jenny , Saunders Darren , Wai Daniel , Reynolds C Patrick , Buckley Jonathan D. , Arceci Robert J. , Reaman Gregory H. , Triche Timothy J. , Jürgens Heribert , Pollak Michael , Sorensen Poul HB
Published: February 2011

The insulin-like growth factor-1 receptor (IGF1R) is emerging as a therapeutic target in many cancers including Ewing family tumors and other childhood sarcomas, where IGF1R blocking antibodies show promising anti-tumor activity in some but not all patients. A key question remains as to what underlies IGF1R inhibitor sensitivity versus resistance. IGF1R mutations have to date not been found in these tumors. Instead, we hypothesize that compensatory signaling circuits by-passing targeted IGF1R inhibition are involved. We performed synthetic lethal siRNA screens in Ewing family tumor cell lines to study the IGF1R signaling axis and related protein tyrosine kinase networks in the presence and absence of sub-lethal doses of an IGF1R kinase inhibitor, with the goal of identifying siRNAs that restored full inhibitor activity. This strategy revealed (1) that Ewing sarcoma and rhabdomyosarcoma cells are exquisitely sensitive to loss of distal IGF1R signaling components such as the ribosomal protein S6 (RPS6); (2) IGF1R inhibitors fail to block RPS6 activation in resistant cell lines; and (3) that siRNA knock-down of certain protein tyrosine kinases such as the MET family receptor MST1R restores inhibitor efficacy, even in highly drug resistant cell lines. Moreover, we confirmed MST1R expression in childhood sarcomas and found that loss of MST1R blocks downstream RPS6 activation when combined with IGF1R inhibition in vitro. Taken together, we present an approach in principle applicable to any promising drug of interest. We demonstrate that key signaling effectors of the IGF1R axis in childhood sarcomas represent distal pathway proteins such as RPS6, and we identify the MST1R receptor tyrosine kinase as a potential therapeutic target whose knock-down markedly enhances sensitivity to IGF1R inhibition in high-risk sarcoma cell types, including resistant lines. These data underscore the importance of fully understanding protein tyrosine kinase networks for successful implementation of kinase inhibitor strategies.

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 3422.
Abstract 3437: GFP imaging of angiogenesis in lymph node metastasis
Authors: Aki Ryoichi , Amoh Yasuyuki , Kishimoto Hiroyuki , Kimura Hiroaki , Suetsugu Atsushi , Momiyama Masashi , Katsuoka Kensei , Hoffman Robert M.
Published: February 2011

In the present study, we developed a novel model to image angiogenesis of lymph node cancer metastasis. Transgenic nude mice carrying GFP under the control of the nestin promoter (ND-GFP mice), 6 to 8 weeks old, were used as hosts. ND-GFP mice express GFP in nascent blood vessels. B16F10-RFP cells were injected into the efferent lymph vessel of the inguinal lymph node of ND-GFP mice to establish experimental metastasis in the axillary lymph node. The axillary lymph nodes of the mice were imaged at days 0, 3, 7, 14, and 28 after injection of tumor cells. Three days after cancer-cell transplantation, nascent blood vessels were imaged by GFP fluorescence in the tumor-involved lymph nodes in the ND-GFP mice. Ten days after cancer-cell transplantation, ND-GFP expression demonstrated an angiogenesis network in the tumor-involved lymph node. Fourteen days after cancer-cell transplantation, ND-GFP expression in the lymph node tumor blood vessels began to be extinguished as the blood vessels matured. Twenty-eight days after cancer-cell transplantation, ND-GFP expression in blood vessels in the lymph node tumors was completely extinguished. This model is useful for direct visualization of nascent tumor angiogenesis in lymph nodes with metastatic cancer, a very critical site for early metastasis.

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 3437.
Abstract 3438: Expression of a novel tumor-derived axonal guidance molecule Sema7a in a breast cancer model
Authors: Garcia-Areas Ramon , Libreros Stephania , Iragavarapu-Charyulu Vijaya L.
Published: February 2011

The purpose of this study is to explore the expression of axonal guidance molecule Sema7a by breast cancer cells and the possible hijacking of macrophages by Sema7a to promote tumor growth. We have recently identified the novel expression and secretion of axonal guidance molecule Sema7a by two different mouse mammary tumor cells both in vitro and in vivo. No studies have yet elucidated the implications of Sema7a expression in cancer. The over-expression of Sema7a in murine breast cancer cell lines may parallel the expression of this axonal guidance molecule in human breast cancer. Furthermore, we have evidence that Sema7a is expressed at higher levels in the macrophages and activated T lymphocytes of tumor bearers compared to the normal mice. To determine the role of the tumor on Sema7a expression, Sema7a expression was assessed in macrophages and T cells from normal mice cultured with tumor-derived factors. Sema7a, originally discovered for its chemotactic activity, is a known immune modulator through α1β1integrins. Sema7a mediates chemotaxis of human peripheral blood monocytes and induces them to produce proinflammatory cytokines, including IL-1β, IL-6, TNF-α, and IL-8. Murine macrophages treated with Sema7a significantly increased their expression of CXCL2/MIP-2 an angiogenic chemokine that is a homologue of human IL-8. This proinflammatory chemokine is known to promote leukocyte-mediated angiogenesis and tumor growth. Monocytes recruited to the tumor adopt a macrophage phenotype that is of M2. M2 macrophages secrete angiogenic factors including VEGF, IL-8, and TNF-α, as well as TNF-α-dependent MMP-9 that degrades extracellular matrix to promote metastasis. It is not clear which tumor-derived factors drive the switch to M2 phenotype in the macrophages and their subsequent production of angiogenic chemokines and matrix metalloproteinases. A putative relationship between tumor-derived Sema7a and monocytes could prove valuable in establishing new research avenues towards unraveling important tumor-host immune interactions in breast cancer patients.

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 3438.
Abstract 3444: The role of p75ntr and TIMP-1 in U251 malignant glioma cells
Authors: Rojiani Mumtaz V. , Brem Steven , Murphy Susan , Obadia Marguerite , Rojiani Amyn M.
Published: February 2011

The neurotrophin receptor p75NTR and trk receptors play an important role in the diverse functions of neurotrophins in the CNS. These include survival, differentiation, neurite outgrowth and development in the nervous system. More recently, an emerging role for the p75NTR has been described in tumor biology, both within the nervous system as well as in non-nervous system neoplasms. Investigative studies in vitro and in vivo have identified both tumor promoting and tumor inhibitory functions of p75NTR, related to specific tumor types. p75NTR has been implicated as a tumor and metastasis suppressor gene and in promoting apoptosis, particularly within the prostate. It has been suggested that p75NTR interactions in the microenvironment result in decreasing MMP2 and MMP9, with a concomitant increase in TIMP1 expression. These have a negative effect on tumor kinetics. However, these tumor promoting and tumor inhibitory functions of p75NTR have been reported in context of specific tumor types. Previously, we have shown that over-expressing TIMP1 in H2009, a lung adenocarcinoma cell line resulted in decreased expression of p75NTR in the clone HB1 compared to H2009. This clone was found to be more aggressive in both in vitro and in vivo studies. Extending these studies to glial neoplasms we sought to define a role of TIMP1in the modulation of p75NTR activities. To this end, we over-expressed TIMP1 in U251 glioma cell line. The resulting stable clones expressed decreased levels of p75 RNA and again behaved more aggressively in vitro with increased tumorigenicity and angiogenesis as evidenced by enhanced colony formation in soft agar as well as increased capillary network formation on Matrigel when serum-free conditioned media was used from the clone compared to parental U251 cell line. We have utilized p75 siRNA to knockdown the levels of p75NTR in U251 glioma cell line. Stable clones expressing reduced levels of p75 showed an increase in the number as well as the size of colonies in soft agar assay compared to the parental cell line suggesting increased tumorigenicity. Collectively our studies appear to define a tumor suppressor function for p75NTR. Additional studies are ongoing to further define the role of p75NTR and TIMP1.

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 3444.
Abstract 3446: Induction of colonic aberrant crypts in mice by an apparent N-nitroso compound fraction derived from hot dogs
Authors: Davis Michael E. , Lisowyj Michal P. , Wisecarver James L. , Gulizia James M. , Mirvish Sidney S.
Published: February 2011

Introduction: Nitrite-preserved meat products, e.g., hot dogs, may be a cause of colon cancer. This may occur because such products contain a possibly carcinogenic fraction called “total apparent N-nitroso compounds” (ANC). Feeding such products to humans and mice increased the fecal excretion of ANC. We tested here whether hot-dog-derived ANC could induce colonic aberrant crypts (ABC), a precursor lesion for colon cancer, in mice. Methods: We purified the ANC precursor (ANCP) fraction of commercial hot hogs by adsorption-desorption on silica gel and cation exchange resin, and treated the ANCP with nitrite to convert them to ANC. As a positive control group, adult female A/J mice were given 1 or 3 i.p. injections of 5 mg/kg of azoxymethane (AOM) [Experiment (Exp.) 1]. In Exps. 2 and 3, similar mice were continuously fed the purified hot dog ANC (initial dose, 100 nmol/g diet) in a high-fat or “high-fat stress” diet. In Exp. 2 the ANC dose initially dropped sharply, probably because the ANCP was unstable on thawing, but later in Exp 2 and throughout Exp 3 the ANCP level was stable and was maintained at 100 nmol/g diet. The mice were killed after 8 (for AOM tests) or 24-34 (for ANC tests) weeks of treatment. Feces of the ANC-treated mice contained up to 225 nmol ANC/g, indicating that the ANC reached the colon. The distal 2 cm of each colon was stained with methylne blue, the mucosal surfaces were scanned at 400 x magnification, and ABC were identified and counted. Results: Mean numbers of ANC/colon for 1 and 3 AOM injections, AOM controls, ANC (Exp 2), ANC (Exp. 3) and ANC controls were 28, 76, 14, 20, 14 and 5-8. Standard deviations were <50% of mean values. The effect of ANC was significant by the rank order test (P < 0.01) for both Exps. 2 and 3. Potency of ABC induction per mole ANC was 1.7% (Exp. 2) and 0.4% (Exp. 3) of that for AOM in Exp. 1. In Exp. 3 we noted that the ABC occurred as foci with 1-4 ABC/focus and a mean of 6.8 foci/colon. ANC treatment did not affect the number of foci. Conclusions: Hot-dog-derived ANC induced a significant number of ABC in the mouse colon, but the effect was relatively weak. These results provide a direct link between ANC in a nitrite-preserved meat product and effects in the colon. Partly supported by NIH grant RO1 CA-143460-01.

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 3446.
Abstract 3451: Cell proliferation and apoptosis during chloroform-induced hepatocarcinogenesis in male F-344/N rats
Authors: Carter Julia H. , Carter Harry W. , Richmond Raymond E. , Deddens James A. , Chiu Nancy , DeAngelo Anthony B.
Published: February 2011

The carcinogenic potential of chlorinated organics is of direct importance in human risk assessment. Most drinking water chlorinated organics are disinfection by products (DBPs) of water chlorination and many test positive in rodent bioassays. Trihalomethanes (THMs) are the most prevalent DBPs generated in chlorinated drinking water and chloroform (TCM) is the THM in highest concentration in finished drinking water. Human exposure to TCM occurs through ingestion of drinking water, inhalation and dermal exposure. TCM is carcinogenic to the liver and kidney of rodents including male and female B6C3F1 mice and male F-344/N and Osborne-Mendel rats. The carcinogenic mechanism of TCM in the rodent is not completely understood. Four mechanisms have been proposed: 1) mutagenicity; 2) reparative hyperplasia; 3) altered gene expression; and 4) secondary genotoxicity. Here, histopathology, immunohistochemistry, and quantitative image analysis were used to examine reparative cell proliferation and altered gene expression as potential mechanisms of TCM-induced hepatocellular carcinogenesis in male F-344/N rats. Animals were exposed to concentrations of 803 + 5 or 1592 + 21 mg/L in the drinking water for 78 or 100 weeks. Distilled water was the vehicle control. The high TCM dose increased the prevalence (% of animals with a lesion) of hepatocellular neoplasia (carcinoma and adenoma) 17.5% vs. 5.1% (p<0.05) and marginally enhanced the prevalence of combined preneoplastic and neoplastic tumors (20.5% vs 7.7%; 0.05<p<0.1). Cell proliferation (proliferating cell nuclear antigen), apoptosis (terminal transferase assay) and expression of tumor associated markers, transforming growth factor alpha (TGF-α) and the p21 Ras oncogene were evaluated in normal liver, altered foci (AF), large foci of cellular alteration (LFCA) and adenomas (AD). PCNA, TGF-α, and p21 Ras increased significantly during TCM-induced neoplastic progression (p = 0.0021, 0.0105, and 0.0167, respectively). A significant change in apoptosis was not found during neoplastic progression, however, a progressive imbalance between cell proliferation and death occurred (p = 0.0022). The phenotype of hepatocellular lesions in rats receiving TCM in their drinking water differed from that of the “spontaneous” lesions arising in rats given distilled water, suggesting a different mechanistic origin. Taken together the data are consistent with a carcinogenic mechanism for TCM that includes adaptation to a hepatotoxic chemical and selection of resistant cells. (This abstract does not reflect the views of the EPA).

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 3451.
Abstract 3468: Aldo-keto reductases protect lung epithelial cells from polycyclic aromatic hydrocarbons at the expense of increased oxidative stress
Authors: Abedin Zahidur , Sen Sushmita , Field Jeffrey
Published: February 2011

Tobacco exposure stimulates genes that are likely to be involved in the metabolism of tobacco and its combustion products. Four induced genes are aldo-keto reductases (AKR), a family of enzymes that reduce aldehydes and ketones to alcohols. AKRs act on a range of substrates including polycyclic aromatic hydrocarbons (PAH). AKRs convert PAH to form o-quinones. Alternatively, PAHs are metabolized to (+)-anti-benzo[α]pyrene diol epoxide ((+)-anti-BPDE)) by the combined action of P4501A1/1B1 and epoxide hydrolase. (+)-anti-BPDE forms DNA adducts directly, while PAH o-quinones cause oxidative stress through a futile redox cycle. To address the role of AKRs in PAH toxicity, we compared the toxicities of PAH metabolites and the effects of overexpressing AKR 1A1 in lung cells. We find that anti-BPDE and BP,-7,8-diol, an intermediate product, are toxic to A549 cells at concentrations with an IC50 of ∼2 µM. In addition, both anti-BPDE and BP 7,8-diol induce p53 expression in 6 hrs post-treatment at concentrations as low as 1 µM consistent with extensive DNA damage. In contrast, we found that the PAH o-quinone B[a]P-7,8-dione was about 10-fold less toxic to A549 cells with an IC 50 > 20 µM. B[a]P-7,8-dione treatment also did not induce p53 expression in 24 hrs at 20 µM. However, B[a]P-7,8-dione treatment generated reactive oxygen species (ROS) in A549 cells as previously reported. We also observed that overexpression of AKR 1A1 in H358 cells, which otherwise have low levels of AKR expression, protected cells 2-10 fold from the toxic effects of BP, 7,8-diol. These data suggest that overexpression of AKRs may protect lung cancer cells from the acute toxic effects of PAH at the expense of increasing the burden oxidative stress

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 3468.
Abstract 3469: Effect of age and mutagen on the enriched murine mammary epithelial stem and progenitor cell population
Authors: Bandyopadhyay Abhik M. , De Keya , Biswas Tanuka , Herzig Maryanne , Walter Christi A. , Sun LuZhe
Published: February 2011

Epidemiological studies indicated that aging is a significant risk factor for breast cancer. Yet, there is insufficient knowledge on the mechanisms affecting breast cancer initiation and promotion in older women. Recent studies indicated epithelial stem/progenitor cell population, responsible for lifelong tissue maintenance and regeneration, as a target of transformation. It has been reported earlier by serial transplantation of murine mammary tissues that functional mammary outgrowths indicating stem cell function is unaffected by the age of the donor. But it is unclear that whether spontaneous or induced mutagenesis in the aging stem/progenitor cell population has the potential to impair stem/progenitor functions and/or to initiate tumorigenesis. In this study, first we have isolated lineage negative (CD31−CD45−Ter119−) mammary epithelial cells (Lin−MEC) from the thoracic/inguinal mammary glands of young (6 months, n=7) and old (28 months, n=6) mice by serial enzymatic dissociation followed by magnetic nanoparticle separation method (StemCell Technology). These cells were utilized to form mammospheres, representing mammary stem/early progenitor cells. Although the frequency of mammosphere formation remained the same, the number of larger mammospheres (>60µm) from old mice were significantly lower than the young mice, probably indicating decreased stem cell activity in the older population. To study the effect of mutagen in vivo, we treated two young and two old mice with methyl nitrosourea (MNU, 25mg/kg body weight) once a week for three consecutive weeks. Animals were sacrificed one week after the last treatment. Both total and larger mammospheres (>60µm) from the isolated Lin−MEC of the treated older mice were observed in significantly higher number than the treated younger mice. In a recent clinical study larger mammospheres (>50µm) obtained from metastatic breast cancer cells has been correlated to their ability to induce tumors. These Lin−MEC cells were also analyzed and compared for DNA damage by alkaline comet assay and a higher percentage of cells (>10%) with irreversible DNA damage in the older population were observed compared to younger population (<2%) indicating probable lower DNA repair capacity of the former cells. Thus, our preliminary study indicated that enriched stem/progenitor cells in the old mice with decreased in vitro stem cell activity might have the potential to be vulnerable to DNA damage by mutagens and subsequent 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 3469.
Abstract 3475: PI3K inhibition by the βGBP cytokine in colon cancer cells. Combined activation of intrinsic and extrinsic apoptosis
Authors: Mallucci Livio , Shi Dong-yun , Lotti Lavinia , Mariani-Costantini Renato , Wells Valerie , Zicha Daniel
Published: February 2011

βGBP (monomeric β-galactoside binding protein) is a newly discovered antiproliferative cytokine produced by CD4+ and CD8+-activated T cells and endogenously released by somatic cells. In normal cells βGBP negatively regulates the cell cycle; in cancer cells βGBP induces apoptosis through pathways which involve inhibition of mitogenic signaling and inhibition of survival signaling. Mechanisms of action initiate with downregulation of class 1A and class 1B phosphatidylinositol 3-OH kinase (PI3K) via functional inhibition of their p110 catalytic subunit.

The study has the purpose of defining signaling routes to apoptosis downstream of PI3K in cells from colorectal carcinomas, which epitomise cancers made aggressive and refractory to therapies by accumulated genetic changes. We examined cancer cells of primary (SW480) and metastatic derivation (SW620, LoVo). Pathways investigated consequent to PI3K downregulation are those relating to Ras-ERK signaling and cell cycle parameters, to Akt signaling and to Rac mediated signaling. Apoptosis was assessed by changes in mitochondrial membrane potential, redistribution of phosphatidylserine, caspase 3 activation and DNA fragmentation.

We found colon cancer cells to be sensitive to minimal doses of βGBP which, while unable to inhibit Ras-ERK activity and Akt expression consequent to inhibition of PI3K activity, led to apoptosis through unprecedented outstanding events: a dramatic rearrangemant of the cytoskeletal network and the concurrent activation of both intrinsic and extrinsic apoptosis. Mechanisms involved related to two set of events. One event was characterised by overexpression and stabilisation of cyclin E, inhibition of DNA synthesis, Chk2 activation and overexpression of deregulated E2F1, a condition for the activation of intrinsic apoptosis; the other related to changes in macromolecular mobility in the plane of the membrane and CD95/Fas clustering, a condition for the activation of extrinsic apoptosis.

In vivo experiments in immunodeficient mice show that βGBP inhibits/prevents tumor development with no harm to the mouse.

The ability of βGBP to induce death in cancer cells notoriously resistant to apoptosis, and to induce death by combined routes is a unique property with important therapeutic implications.

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 3475.
Abstract 3488: In vitro activity of the IGF-1R inhibitor CP-751,871, as a single agent and in combination with paclitaxel, in lung and colon cell lines
Authors: Agullo-Ortuño M. Teresa , Díaz-García C. Vanessa , Rodríguez Analía , Cortijo Ana , López-Martín José A. , García-Carbonero Rocío , López-Ríos Fernando , Cortés-Funes Hernán , Paz-Ares Luis
Published: February 2011

Background: Signaling through the insulin-like growth factor 1 receptor (IGF-1R) has been implicated in the resistance to a number of clinically relevant anti-cancer agents.

Objective: To assess the direct anti-tumour effects of the IGF-1R monoclonal antibody CP-751,871, as a single agent and in combination with paclitaxel, in lung and colon cancer cell lines.

Methods: Four lung (A549, HCC78, H1299 and H460) and four colon (LS180, Colo205, HT29 and DLD1) cancer cell lines were selected. The experiments were designed to test the effects of simultaneous or sequential exposure to with CP-751,871 and/or paclitaxel. In the sequential arm, cells were pretreated with progressive doses of paclitaxel (from 0 to1000 nM) for 24 hours, drug removed by washing the cells twice, followed by the addition of cell culture medium plus CP-751,871 (50 μg/mL). After 48 hours of continuous presence of CP-751,871, cell growth was evaluated by WST-1 cell survival assays. In simultaneous treatment, cells were treated with both Paclitaxel (0-1000 nM) and CP-751,871 (50 μg/mL) for 48 hours. Flow cytometric analysis was used to estimate the effects on cell cycle and apoptosis. IGF-IR mRNA expression was determined by quantitative real-time PCR. Relative gene expression values were calculated by the ΔΔCt method.

Results: No correlation between basal IGF-IR mRNA expression and response to CP-751,871 was observed. Flow cytometric analysis demonstrated that CP-751,871 enhanced cell cycle arrest at the G1/G0 checkpoint with minimal effects on apoptosis. Combined exposure to both CP-751,871 and Paclitaxel showed improved response in cell growth inhibition on HCC78, H1299, Colo205 and HT29 cell lines, and was statistically superior to Paclitaxel alone (p< 0.001) and CP-751,871 alone (p< 0.001). The results were even better after a sequential exposure. On the contrary, in H460, LS180 and DLD-1 cell lines, concomitant, but no sequential treatment resulted in antagonistic interactions. Conclusions: CP-751,871 showed a modest anti-tumour activity (about 30% growth inhibition in lung cancer cells; 10% growth inhibition in DLD1 and HT29 cell lines; and bigger inhibition in Colo205 (IC50=27.6 μg/mL) and LS180 (IC50=2.33 μg/mL) cell lines), predominantly cytostatic, that is not related to the mRNA expression. The effects of CP-751,871 in combination with paclitaxel depend on the cell line model and on the sequence of administration.

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 3488.
Abstract 3492: A novel PI3K/mTOR dual inhibitor provides correlations of pharmacokinetic pharmacodynamic and tumor growth inhibition in a prostate PC3 xenograft model for application in clinical trials
Authors: Carley William W. , Bagrodia Shubha , Yuan Jing , Ye Jingjing , Cheng Hengmiao , Sun Shaoxian , Cullnane Carleen , McArthur Grant
Published: February 2011

PF-04691502 is a dual inhibitor of both PI3K and mTOR, inhibits PI3K signaling in cancer cell lines, and exhibits in vitro and in vivo anti-proliferative activity in PI3K-pathway driven cell lines. It is a nanomolar inhibitor of all 4 isoforms of the catalytic subunit of PI3K and of both TORC1 and TORC2. Anti-cancer activity of the inhibitor is hypothesized to be through inhibition of survival, proliferative, and anti-apoptotic processes. Its predicted human bioavalibility is roughly 60% and it will soon be entering phase 1 studies.

To provide an improved predictability of this compound and its PD markers in the clinic we correlated PK, PD (phosphoproteins and FDG-PET imaging) and tumor growth inhibition in a single model. Since this inhibitor exhibits an IC50 of 37 nM in the PC3 prostate cell line (PTEN null) in vitro we chose this cell line for a xenograft model. Three groups (Vehicle, 5 and 7.5 mg/kg.) were dosed p.o. daily and subsequently evaluated. Tissues were harvested on day 7 and 13 at 4.5 hr post-dose. FDG-PET imaging was completed on days 1, 3, 7 and 13 at 3 hr post-dose.

First, negative correlations were established between blood levels of PF-04691502 and the following tumor phosphoprotein epitopes: pS6, pAKTS473 and pAKTT308. The R2 values for these proteins were −0.3, −0.4 and −0.4, respectively. P-values of the correlations are all significant for all three proteins. Second, all PD markers were positively correlated with TGI. When correlating FDG-PET and TGI, day 7 has the best correlation and an R2 of 0.3 while days 3 and 13 have poorer correlations (R2 of 0.04 and 0.13, respectively). Although the R2 isn't high for day 7, the positive trend between image and TGI is statistically significant. When correlating PD markers and TGI, R2 values for pS6, pAKTS473 and pAKTT308 were 0.6, 0.8 and 0.6, respectively. In conclusion, the data from this single model indicates PD phosphoprotein markers are more reflective of TGI than FDG-PET imaging. These results will be applied to estimating the size of the cohort recruited for mechanistic evaluation in phase 1 trials of PF-04691502.

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 3492.
Abstract 3493: ON 01910. Na, a clinical trial stage multi-kinase inhibitor, induces apoptosis in chronic lymphocytic leukemia (CLL) cells through inhibition of PI3K/AKT and activation of the JNK pathway resulting in NOXA and BIM upregulation
Authors: Perez-Galan Patricia , Chapman Colby M. , Sun Xiameng , Gibellini Federica , Liu Poching , Raghavachari Nalini , Wiestner Adrian
Published: February 2011

Chronic Lymphocytic leukemia (CLL), the most common leukemia in Western countries, is a clinically heterogeneous disease characterized by the accumulation of mature B lymphocytes, where disease progression is driven by survival and proliferation signals provided by the tumor microenvironment. ON 01910. Na (Onconova Therapeutics) belongs to the family of styryl benzyl sulfones, a novel family of non-ATP competitive kinase inhibitors. ON 01910. Na is under clinical development in hematologic and solid tumors. Here we show that ON 01910. Na induced apoptosis in CLL samples (n=28) in vitro, without showing significant toxicity against T-cells or normal B-cells. Twenty four CLL samples were highly sensitive to ON 01910. Na (mean IC50=0.89 μM) a concentration readily achieved in phase I clinical trials, three showed moderate sensitivity (mean IC50=4.26 μM) and one was resistant (IC50 >8 μM). There were no significant differences in ON01910. Na cytotoxicity against CLL cells expressing mutated (mean IC50=1.33 μM) or unmutated (mean IC50 =1.22μM) IgVH sequences. ON 01910. Na was similarly effective against tumor cells carrying 17p or 11q deletions. ON 01910. Na activated Bax and Bak, leading to mitochondrial depolarization. To delineate the upstream pathways underlying ON 01910. Na induced apoptosis, we performed Gene Expression Profiling (GEP) in CLL cells treated in vitro for 4 and 10 hours with ON 01910. Na. Gene Set Enrichment Analysis (GSEA) identified gene sets indicating BCR and PI3K inhibition (FDR<0.2). In keeping with PI3K inhibition, we found that ON 19010. Na inhibited phosphorylation of Akt and Foxo3a, and FOXO3 target genes, including Bim, were upregulated. In addition, GSEA identified induction of AP-1 gene sets (FDR<0.01) suggesting the generation of reactive oxygen species (ROS) by ON 01910. Na. Consistently, we found a rapid increase in ROS in cells treated with ON 01910. Na and ROS scavengers protected cells from its cytotoxic effect. ROS upregulated the proapoptotic BH3-only protein Noxa, and knockdown of Noxa by shRNA decreased the sensitivity to the drug by 60%. Our results identify ON 01910. Na as a promising agent in the treatment of CLL with an interesting dual mechanism of action: inhibition of the PI3K/AKT survival pathway with Foxo mediated upregulation of BIM and the induction of oxidative stress resulting in Noxa upregulation.

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 3493.
Abstract 3494: Experimentally validated modeling for optimizing therapeutic combinations of oncolytic adenoviruses and MEK inhibitor
Authors: Shiina Marisa , Bagheri Neda , Lauffenburger Douglas , Korn Wolfgang Michael
Published: February 2011

Oncolytic adenoviruses are therapeutic anti-cancer agents that selectively replicate in and lyse cancer cells motivating their use in targeted gene therapy. Successful entry of adenoviruses into target cells depends on attachment of the adenovirus fiber-knob protein to the Coxsackie-and Adenovirus Receptor (CAR), which is often down-regulated in highly malignant cells. In a previous study, we demonstrate that MEK inhibition may counter this effect by up-regulating CAR expression. In this study we optimize infection by investigating the combinatorial effect of oncolytic adenovirus infection and MEK inhibition through data-driven models and model-driven experiments. To motivate and test our hypotheses, we use HCT116 colon cancer cell line treated with MEK inhibitor, CI1040, and infected with oncolytic adenoviruses, ONYX-015, Delta-24 and Delta-24RGD. We developed a nonlinear ordinary differential equation model that characterizes the dynamics of these cancer cells subject to ONYX-015 infection modulated by CI1040. Resulting simulations offer predictions that may help guide experimental design and inform optimal timing of CI1040 treatment and ONYX-015 infection to minimize cancer growth. Experimental results demonstrate that HCT116 cells treated with CI1040 for 2 days and infected with adenoviruses have stronger cytopathic effects, greater virus uptake, and greater production when compared to the DMSO control. We also found that G1 cell cycle arrest, caused by CI1040 treatment, is associated with increased virus production. To assess the role of release from cell cycle arrest as a mechanism leading to enhanced production of oncolytic adenoviruses, we synchronize cells by density arrest for 48 hours, release and infect with adenoviruses. The results showed a more potent cell killing effect and virus production on cells arrested in G1 and released. We expanded our analysis to a panel of 21 pancreatic cell lines, as this tumor type is a possible target for adenoviral therapies. Enhanced virus replication and cell killing were observed in different pancreatic cell lines, suggesting that combination therapy of CI1040 and oncolytic adenoviruses have potential implications for the design of future clinical trials and improved oncolytic adenoviruses 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 3494.
Abstract 3498: The cytotoxicity and genotoxicity of etoposide and other phenolic and non-phenolic agents in human myeloid leukemia HL-60 cells: role of myeloperoxidase (MPO)
Authors: Yalowich Jack C. , Hornbaker Marisa , Bodnar Andrew J. , Jin ShunQian , Zhang Yuxun , Day Billy W. , Fuchs James R. , Li Pui-Kai
Published: February 2011

The clinical efficacy of the anticancer agent etoposide (VP-16) is compromised by its ability to cause acute myeloid leukemias (t-AML) associated with MLL gene rearrangements. We proposed previously that myeloperoxidase (MPO) found in myeloid precursors converts the phenolic drug VP-16 to its phenoxyl radical (VP-16-O[[Unable to Display Character: ∙]]), which redox cycles, leading to the generation of reactive oxygen species, oxidative DNA damage linked to DNA topoisomerase II (topo II)-mediated strand cleavage, and resultant recombination events causal for t-AML. After knockdown with an shRNA for MPO in myeloid leukemia HL-60 cells, MPO activity was reduced to 13% of shRNA controls. In these MPO knockdown cells, VP-16-O[[Unable to Display Character: ∙]] formation and VP-16-induced topo II/DNA covalent complexes were reduced compared to shRNA controls demonstrating that VP-16 activity was, in part, dependent on MPO. MPO can also oxidize VP-16 to its ortho-quinone metabolite which is subsequently inactivated by conjugation with glutathione. The cytotoxicities of VP-16, curcumin (a phenolic agent), and a curcumin analog (FLLL-10) were increased in MPO knockdown cells compared to shRNA controls. Podophyllotoxin, the non-phenolic VP-16 parental compound, was equally cytotoxic in MPO knockdown and control cells. Additionally, the cytotoxicity of hydrogen peroxide, an MPO co-substrate, was unaffected by MPO levels. Results indicate that MPO converts VP-16 to a pro-oxidant, which leads to genotoxicity, while simultaneously protecting cells from drug-induced cytotoxicity likely by glutathione conjugation to VP-16 ortho-quinone. The combined results suggest a mechanism by which MPO-containing myeloid progenitor cells can be relatively protected against VP-16-induced apoptosis/cytotoxicity while suffering increased genotoxic and recombinogenic insults responsible for leukemogenesis. Support: NIH CA090787.

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 3498.
Abstract 35: Metabolomic analysis of MCF-7 and MCF-10 cells after thiaminase I exposure
Authors: Richardson Adam D. , Liu Shuqian , Moscow Jeffrey A.
Published: February 2011

We have recently shown that recombinant thiaminase I enzyme has cytotoxic activity against breast cancer cell lines (Liu, S., Monks, N.R., Hanes, J.W., Begley, T.P., Yu, H. and Moscow, J.A. Sensitivity of breast cancer cell lines to recombinant thiaminase I. Cancer Chemotherapy and Pharmacology, 2009). Because thiamine is a cofactor in several key metabolic enzymes, including pyruvate dehydrogenase, 2-oxoglutarate dehydrogenase and transketolase, information regarding the metabolic consequences of thiaminase I treatment may increase our understanding of the cytotoxic mechanism-of-action of thiaminase I. We analyzed the metabolomes of the MCF-7 breast cancer cell line and the immortalized but non-malignant breast cell line MCF-10A with or without incubation for 4 days in medium containing thiaminase I at the predetermined cytotoxic IC50 concentrations (0.125 U/ml for MCF-7 and 0.25 U/ml for MCF-10A), with the addition of [U-13C]-labeled glucose in the final 2 days. By tracking the metabolism of [U-13C]-glucose using GCMS and NMR we were able to quantify intracellular metabolite pools and calculate both relative and absolute flux rates through several key pathways. Thiaminase I treatment decreased the amount of protein per MCF-7 cell by 30% (163 pg/cell to 115 pg/cell) while the total protein remained essentially unchanged in the MCF-10A cells (141 pg/cell vs 152 pg/cell). While both cell lines displayed increased glucose consumption and lactate production during thiaminase I treatment, these responses were greater in the MCF-7 cell line. Metabolite analysis revealed significant alterations to glycolysis and the TCA cycle in both cell lines, with glycolysis-linked metabolites being depleted in the MCF-7 cell line and changes in the equilibrium of TCA cycle intermediates in the MCF-10A cell line. The 13C- labeling patterns suggest that the MCF-10A cell line responds to thiaminase I treatment by increasing the catabolism of alternate, non-glucose carbon sources through the activation of specific metabolic pathways. In contrast, the MCF-7 cell line appears to have a less adaptable metabolic program and must down-regulate its overall metabolic throughput. Therefore, the possibility is raised that differential metabolic effects of thiaminase I could be exploited therapeutically.

Supported by the Susan G. Komen For the Cure Foundation (JAM) and the California Breast Cancer Research Program (ADR).

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 35.
Abstract 3502: Understanding the pathways involved in the repair of CNDAC induced DNA damage
Authors: Frame Sheelagh , Armour Sian , Munro Clare , Hogben Morag , Jones Ruth , Blake David G. , MacCallum David E. , Green Simon R.
Published: February 2011

CNDAC is the active metabolite of sapacitabine, which is currently being evaluated in Phase II clinical trials for acute myeloid leukaemia, myelodysplastic syndrome and non small cell lung cancer. CNDAC (2′-C-Cyano-2′-deoxy-β-D-arabino-pentafuranosylcytosine) was designed as a novel cytosine analogue with a unique mechanism of action. The presence of the cyano-group within the ribose moiety of the molecule causes the formation of single-stranded DNA strand breaks, following incorporation of CNDAC into an extending DNA chain. These breaks are difficult to repair and are processed into double-strand DNA breaks that activate the dsDNA damage checkpoint. As a consequence of this unique mechanism of action, CNDAC arrests cells in the G2/M phase of the cell cycle in contrast to other nucleoside agents such as cytarabine and gemcitabine which cause an S-phase arrest. As such CNDAC may have unique therapeutic applications as a single agent in certain tumour types as well as in combination with other agents compared with standard nucleoside analogues.

In order to identify options for maximising that activity of CNDAC, two approaches were taken to evaluate the repair mechanisms involved for CNDAC induced DNA damage. First a small scale siRNA screen was used to identify genes that were synthetically lethal with CNDAC. Ten targets were initially selected; prioritising genes known to be involved in DNA repair and including BRCA, ATM, CHK and ERCC1. The most dramatic increase in CNDAC sensitivity was seen when BRCA2 was targeted by siRNA, indicating that the homologous recombination DNA repair pathway is involved in the repair of CNDAC induced DNA damage. The second approach involved a cytotoxicity screen evaluating synergy with commercially available agents that either target DNA repair or induce DNA damage themselves. The most promising combinations were then followed up with flow cytometry analysis to examine the induction of cell death. Using this approach the best synergy was detected between CNDAC and either the ATM inhibitor, KU55933 or the CHK inhibitor PF477736. Again these data indicate that the homologous recombination pathway is involved in the repair of CNDAC induced DNA damage.

These data suggest that it would be feasible to explore the clinical use of sapacitabine in diseases where the homologous recombination DNA repair pathway is compromised, such as BRCA deficient tumours. Data from these ongoing studies will be presented.

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 3502.