bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 676: Induction of apoptosis via control of cell adhesion by peroxisome proliferator activated receptor-gamma agonist in human uterine leiomyoma cells
Authors: Cho Chi-Heum , Jung Sun-Wook , Shin So-Jin , Kwon Sang-Hoon , Cha Soon-Do
Published: February 2011

Objective : Peroxisome Proliferator-Activated Receptor (PPAR)-gamma ligands constitute important insulin sensitizers that have already been approved for the treatment of human metabolic disorders. It has been previously demonstrated that growth inhibition in human uterine leiomyoma cells mediated by caspase-dependent pathway and death receptor. The present study was designed to further understand the mechanism after uterine leiomyoma and normal myometrial cells were exposed to PPAR gamma agonist, Ciglitizone.

Methods : In order to confirm the unknown cellular and biochemical responses to ciglitizone treatment with three uterine leiomyoma and normal myometrial cells, DNA chip analysis were performed. Highly up-regulated genes were confirmed to ascertain effect of PPAR gamma agonist by RT-PCR and Western-blot analysis. To analyze mechanism of invasion and metastasis -associating gene regulation by PPAR-γ ligands, wound healing assay and soft agar assay were performed.

Results : DNA chip analysis revealed genes are related with cell adhesion and metastasis genes including plasminogen activating urokinase (PLAU), claudin 1, matrix metallopeptidase 1,2, and 9. Highly down-regulated genes had defense system against cell metastasis in common and was related with adhesion. Treatment with ciglitizone made decrease expression level of above cell adhesion related genes in dose dependent manner by RT-PCR and Western-blot analysis. Furthermore, treatment with ciglitizone in uterine leiomyoma cells decrease the invasion ability of leiomyoma cells and the formation of anchorage-independent colonies in soft agar.

Conclusions : These results suggest that PPAR-gamma could be considered as a therapeutic target for diverse disease states in which excessive angiogenesis is implicated, including cancer and uterine leiomyoma. This phenomenon has not been previously reported for PPAR-gamma agonist in uterine leiomyoma. Defining the role of PPARγ in uterine leiomyoma cells will be paramount for a rational design of combinatorial therapy schemes that takes advantage of the potent and well-tolerated PPARγ agonists.

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 676.
Abstract 684: Trifluoperazine modulates DNA damage-induced cell death in human lung cancer cells through inhibition of DNA repair, cell cycle delays, and augmentation of pro-apoptotic signaling
Authors: Zong Dali , Hååg Petra , Yakymovych Ihor , Lewensohn Rolf , Viktorsson Kristina
Published: February 2011

Trifluoperazine modulates DNA damage-induced cell death in human lung cancer cells through inhibition of DNA repair, cell cycle delays, and augmentation of pro-apoptotic signaling

Resistance of human tumors to DNA damaging chemotherapy poses severe limitations to the efficacy of current available treatment. Small molecule compounds of the phenothiazine class, such as trifluoperazine (TFP), have documented chemosensitizing activities. We exposed human non-small cell lung cancer (NSCLC) cell lines to the DNA damaging drug bleomycin in the presence or absence of TFP, and studied potential differences with regard to DNA repair and cell death signaling. A colony formation assay showed that TFP significantly enhanced the cytotoxic potential of bleomycin, at doses achievable clinically. Bleomycin treatment resulted in a reversible arrest at the G2-M phase of the cell cycle. Notably, TFP delayed the repair of bleomycin-induced DNA double strand breaks (DSBs), causing higher residual levels of γH2AX 24 h after treatment. Despite the presence of unrepaired DNA DSBs, cells co-treated with bleomycin and TFP overcame the G2-M checkpoint arrest, as evidenced by the reappearance of the mitosis marker histone H3 phospho-Ser10 and reduction in CFSE fluorescence. This process, which is reminiscent of checkpoint adaptation, was associated with increased micronucleation and secondary arrest in G2-M phase during subsequent rounds of the cell cycle. Furthermore, the onset of secondary checkpoint arrest in cells co-treated with bleomycin and TFP coincided with the activation of pro-apoptotic Bak and Bax, and was followed by activation of caspase-3, which occurred preferentially in cells in the G2-M phase. In conclusion, we demonstrate here that the chemosensitizing effect of TFP in NSCLC cells involves interference with DNA DSBs repair, resulting in secondary G-M checkpoint arrest after initial checkpoint adaptation, and concomitant increases in pro-apoptotic signaling.

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 684.
Abstract 687: Structure-based design, discovery, and pharmacological profiling of human AP endonuclease (APE1) inhibitors for cancer therapy
Authors: Mohammed Mohammed Z. , Vyjayanti Vaddadi N. , Laughton Charles , Dekker Lodewijk , Fischer Peter M. , Wilson David M. , Madhusudan Srinivasan
Published: February 2011

Potentially cytotoxic AP site is an obligatory intermediate in base excision repair that is repaired by APE1. In the previous study we used fluorescence-based high through-put screening of a chemical library, biochemical and cellular investigations to identify compounds that selectively inhibited the DNA repair domain of APE1. To identify drug-like leads, we recently adopted an industry-standard high through-put virtual screening strategy. Sybyl8.0 molecular modelling software suite was used to build inhibitor templates based on first generation inhibitors. Similarity searching strategies, using ROCS2.3, were then applied to extract pharmacophorically related subsets of compounds from a chemically diverse database of 2546389 compounds resulting in 2535 potential hits. These 2535 compounds were then 3D-encoded, and subjected to docking against the active site of the APE1 model, using algorithm-based programme GOLD2.7. Predicted ligand poses were ranked on the basis of two scoring functions (GOLD score and CHEMO score). The top hits were examined for favourable pharmaceutical properties and underwent detailed biochemical analyses. Several novel inhibitors of APE1 have been identified. Further modelling and biochemical studies in polymorphic variants of APE1, showed that many APE1 inhibitors preferentially bound to the D148E polymorph compared to the wild type. APE1 inhibitors potentiated the cytoxicity of chemotherapeutic agents in melanoma, glioma, breast cancer and pancreatic cancer cell line models implying therapeutic applications in these tumour types.

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 687.
Abstract 69: Functional metabolomic profiling of human melanocyte and melanoma cells during hypoxic adaptation
Authors: Richardson Adam D. , Scott David A. , Filipp Fabian V. , Knutzen Christine A. , Chiang Gary G. , Ronai Ze'ev A. , Osterman Andrei L. , Smith Jeffrey W.
Published: February 2011

Adaptation to hypoxia is an important part of the cellular progression from normal melanocyte to malignant melanoma. This adaptation can be comprised of short-term remodeling of the cellular metabolic network and/or the permanent, gene-driven development of a new metabolic program. In either case, low oxygen conditions require a unique metabolic program for continued survival and proliferation. However, beyond a general shift from respiration to glycolysis, the details of this permissive metabolic program in melanoma are generally unknown. In order to better understand the changes to both the metabolome and metabolic flux in this system, we applied functional metabolomic profiling to seven human melanoma cell lines and to two human melanocyte cell lines under both hypoxic and normoxic conditions. The melanoma panel consists of four cell lines driven by B-raf mutations (WM35, UACC903, WM793 and LU1205), two containing N-ras mutations (MEL501 and WM1346) and one p53 mutant (MeWo). These cells, along with the two melanocyte lines, were grown in media containing a 1:1 mixture of natural and [U-13C]-labeled glucose for 24 hours in either 1% oxygen or atmospheric conditions. We then employed GCMS, NMR and biochemical experiments to map flux through five metabolic hubs and quantify the abundance of 28 key metabolites for each cell line and condition. These maps of central carbon metabolism encompass glycolysis, the pentose phosphate pathway, fatty acid biosynthesis, the glutamate-glutamine-proline network, and glycine/serine de novo synthesis, along with a number of other important metabolites. Analyzing these data with unbiased hierarchical clustering and Ingenuity Pathway Analysis, we discovered a metabolite-derived signature for hypoxic adaptation in melanocytes. Furthermore, we found that certain hypoxic responses are unique to either the B-raf or N-ras of the cell line subset. Interestingly, the hypoxic response of the mutant p53 cell line is more like that of the B-raf mutant cell lines than the B-raf and N-ras sets are to each other. The information gained from this functional metabolomic profiling advances our ability to identify hypoxia-adapted melanomas and develop metabolism-based therapeutic interventions.

This work was supported by P01 CA128814-01 from the National Cancer Institute.

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 69.
Abstract 690: Estrogen receptor beta (ERβ) enhances the chemosensitivity of breast cancer cells by altering DNA damage response signaling
Authors: Thomas Christoforos , Lindberg Karolina , Strom Anders , Damdimopoulos Tassos , Gustafsson Jan-Ake
Published: February 2011

Estrogen Receptor beta (ERβ) is a mediator of estrogen signaling that regulates growth in mammary gland and is involved in breast cancer. In response to DNA damage most tumor cells due to disruptions in the p53 pathway and consequent loss of G1 cell cycle arrest are entirely dependent in S and G2/M checkpoints to maintain genomic integrity. Inhibition of these checkpoints following DNA damage induces selective killing of p53-defective tumor cells. We show that expression of ERβ caused abrogation of Cdc25C-mediated G2/M checkpoint following cisplatin and doxorubicin exposure selectively in p53-deficient cells, resulting in mitotic catastrophe and decreased cancer cell survival. The loss of G2/M checkpoint observed was due to ERβ-mediated inhibition of the DNA damage-induced expression of brca1. ERβ by modulating DNA damage response signaling seems to selectively enhance the chemosensitivity of tumor cells. This suggests that co-operation of compounds that specifically induce the ERβ signaling with DNA damaging chemotherapeutic drugs may represent an attractive new cancer therapeutic strategy.

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 690.
Abstract 691: In vitro and in vivo characterization of selective orally available Parp-1 inhibitors with demonstrated antitumor efficacy in BRCA negative cancer models
Authors: Papeo Gianluca , Rainoldi Sonia , Gianellini Laura , Ciavolella Antonella , Leone Antonella , Lupi Rosita , Sola Francesco , Caprera Francesco , Paolucci Mauro , Thieffine Sandrine , Perego Rita , Albanese Clara , Casale Elena , Forte Barbara , Guanci Marco , Orsini Paolo , Posteri Helena , Scolaro Alessandra , Riccardi-Sirtori Federico , Zuccotto Fabio , Donati Daniele , Moll Jurgen , Montagnoli Alessia
Published: February 2011

Poly (ADP-ribose) polymerase 1 and 2 (Parp-1 and Parp-2) are nuclear enzymes responsible for signaling the presence of DNA damages by catalyzing the addition of ADP-ribose units to DNA, histones and various DNA repair enzymes, thus facilitating DNA repair. Parp-1 has been gaining increasing interest as a therapeutic target for cancer in combination with DNA damaging agents but also as single agent in particular tumor settings, such as BRCA mutated tumors. While Parp-1 knock out mice are viable and display only defects in DNA repair, double Parp-1 and Parp-2 knock-out are embryonic lethal indicating that the absence of Parp-1 and Parp-2 is less tolerated in normal cells. We confirmed this concept in vitro by RNAi of Parp-1 and Parp-2, showing that the inhibition of Parp-1 alone is enough to kill tumor cells but spares normal cells. Hence a selective Parp-1 inhibitor might have a better safety profile, particularly in view of a long treatment. Here we report the discovery and the in vitro and in vivo characterization of a novel class of potent, orally available Parp-1 selective small molecule inhibitors with no activity on other Parp isoforms such as Parp-2, −3 and −5a (tankyrase), which differentiates it from current clinical Parp inhibitors. The most potent compounds were further evaluated in vitro and in vivo. These compounds are highly potent in inhibiting DNA damage-induced Parp-1 activity in cells and exhibit selective anti-proliferative and pro-apoptotic activity in tumor cell lines harbouring defects in DNA repair. In vivo, the compounds show an excellent pharmacokinetic profile with almost complete oral bioavailability and demonstrate efficacy as a single agent in xenograft tumor models with DNA repair deficiencies superior to reference Parp inhibitors. These compounds are very well tolerated with no overt toxicity even after prolonged exposure and their mechanism of action is confirmed both in tumors and peripheral blood cells of treated mice.

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 691.
Abstract 699: Uterine serous papillary carcinomas overexpress human trophoblast-cell-surface marker (Trop-2) and are highly sensitive to immunotherapy with hRS7, a humanized monoclonal anti-Trop-2 antibody
Authors: Santin Alessandro D. , Cocco Emiliano , Varughese Joyce , Richter Christine E. , Casagrande Francesca , Bellone Stefania , El-Sahwi Karim , Bellone Marta , Todeschini Paola , Silasi Dan-Arin , Azodi Masoud , Schwartz Peter E. , Rutherford Thomas J. , Pecorelli Sergio
Published: February 2011

Purpose:

To evaluate the expression of human trophoblast cell-surface marker (Trop-2) and the potential of hRS7, a humanized monoclonal anti-Trop-2 antibody, as a novel therapeutic strategy against Uterine Serous Papillary Carcinoma (USPC), a biologically aggressive and chemotherapy resistant variant of endometrial cancer.

Methods:

Trop-2 expression was evaluated by real time-PCR and flow cytometry in six primary uterine serous papillary carcinoma cell lines recently characterized in our laboratory. Sensitivity to hRS7 antibody-dependent-cellular-cytotoxicity (ADCC) was tested in standard 5-hrs 51Cr release-assays against primary USPC cell lines expressing different levels of Trop-2. Finally, to investigate the effect of interleukin-2 (IL-2) on hRS7-mediated ADCC, 5-hrs 51Cr assays were also conducted in the presence of low doses of IL-2 (i.e., 50-100 IU/ml).

Results:

Trop-2 mRNA transcript was significantly overexpressed in 3 out of 6 USPC primary cell lines when compared to normal human endometrial-cells (NEC) [p=0.005]. Consistent with RT-PCR data, high surface expression of Trop-2 was detected by flow cytometry in Trop-2 overexpressing cell lines [i.e., percentage of positive cells = 100% in all 3 positive cell lines, median (minimum-maximum) MFI (mean fluorescence intensity) of 184.2 (62.5-276.2)]. Importantly, while these USPC cell lines were found highly resistant to natural killer dependent cytotoxicity in the absence of the hRS7 antibody (range of killing 1.1% to 12.4%), they showed high sensitivity to hRS7-mediated ADCC in vitro (range of killing 28.2% to 64.4%) (P< 0.001). Incubation with IL-2 in addition to hRS7 further increased the cytotoxic activity against USPC cell lines overexpressing Trop-2 (p= 0.01).

Conclusion:

Primary USPC cell lines may overexpress Trop-2 at mRNA and protein level and are highly sensitive to hRS7-mediated cytotoxicity in vitro. hRS7 may represent a novel, potentially highly effective therapeutic strategy in patients harbouring advanced, recurrent or metastatic USPC refractory to standard treatment modalities.

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 699.
Abstract 70: Mitochondrial localization of adiponectin receptor 1 (AdipoR1) induces apoptosis in cancer cells
Authors: Lu JianPing , Hou ZheFeng , Beatty Laura K. , Pinthus Jehonathan H.
Published: February 2011

INTRODUCTION

Previous studies by us and others demonstrated tumor suppressive effects of adiponectin, an adipocyte derived hormone. Adiponectin signaling occurs through two distinct membrane receptors; AdipoR1 and AdipoR2. There is paucity of data how their activation leads to tumor suppression. Here we present data indicating localization of AdipoR1 to the mitochondria and its role in apoptosis of cancer cells.

MATERIALS AND METHODS

Human kidney (RCC1932) and cervical (HeLa CCL-2) cancer cells as well as human embryonic kidney cells (HEK 293) were used as models. Full length AdipoR1 and its N-terminal (ligand binding) and C-terminal (intracellular moiety) truncated mutants (mutAdipoR1) ORFs were subcloned into YFP expressing vectors (pEYFP- AdipoR1, pEYFPmutAdipoR1). Cells were seeded onto glass cover slips in multi-well culture plates and transfected with vectors through Lipofectamine-2000 (Invitrogen) method 24 h after seeding. The effect of adiponectin (BioVendor) supplementation was also tested. Immunocytochemistry staining were with MitoTracker labeling or anti-MnSOD (mitochondria), anti-Stearoyl-CoA desaturase (SCD-1, ER), anti-Reb5, anti-manose-6-phosphatase receptor (M6P, endosome); or anti-Catalase (peroxisome). Images of the cells were collected with DM IRB Deconvolution Microscope (Leica) or Confocal microscope LSM510 (Carl Zeiss). Apoptosis was determined by sub-diploid population detection and or annexin binding using flow cytometry.

RESULTS

Over-expression of AdipoR1 in the cells induced apoptosis which was further augmented by adiponectin treatment. This was evident only in cells expressing full length AdipoR1. Furthermore, cells expressing mutAdipoR1 had different intracellular distributions with loss of mitochondrial localization of the receptor and pro-apoptotic activity.

CONCLUSIONS

This is the first report showing that AdipoR1 can be localized in the mitochondria. Adiponectin induce apoptosis in cancer cells through mitochondrial localization related mechanism that requires both ligand and signaling domains of AdipoR1.

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 70.
Abstract 705: Crosstalk with stromal fibroblasts contributes to EGFR inhibitor resistance in colorectal cancer cells
Authors: Liska David , Chen Chin-Tung , Ding Pei-Rong , Weiser Martin R.
Published: February 2011

The mechanism for resistance to EGFR targeted therapies in colorectal cancer (CRC) with intact KRAS is still unknown. Host stromal cells contribute to tumorigenicity; however their effect on the sensitivity to EGFR targeted therapies has still to be determined. We hypothesized that crosstalk between stromal fibroblasts and CRC cancer cells contributes to cetuximab resistance via activation of the HGF/MET axis.

Primary CRC associated fibroblasts (CAFs) were isolated and expanded from five surgical specimens. Fibroblasts were enriched by magnetic activated cell sorting (MACS®) and activated fibroblast lineage was confirmed by alpha smooth muscle actin (SMA) immunofluorescence. Supernatant HGF content and cellular mRNA expression levels in primary CRC CAFs, an embryonic lung fibroblast cell line (MRC-5), and a normal colon fibroblast cell line (CCD18) were determined by ELISA and quantitative RT-PCR. The effect of fibroblasts or HGF (75ng/ml) on the CRC cell lines LIM1215 and DiFi treated with cetuximab (5nM) for 72 hours was determined by co-culture using Transwells and WST-1 cell proliferation assays. Alterations in proliferation were compared by ANOVA.

Mean fibroblast HGF secretion (8.3 ng/ml) and relative mRNA expression (2.0) could be augmented by co-culture with conditioned media (CM) from the CRC cell LIM1215 (14.1 ng/ml and 4.1, respectively) (Table). Further, co-culture of CRC cells with HGF, CCD18 cells, or primary CAFs reversed cetuximab-induced growth inhibition. LIM1215 proliferation was inhibited by cetuximab (55.3 ± 14.4%) which could be reversed by co-culture with either HGF (93.1 ± 10.2%) or CCD18 cells (107.2 ± 10.8%) (p<0.01 for both).

Stromal-tumor interactions may play an important role in CRC resistance to cetuximab via the HGF/MET axis. Inhibition of both EGFR and MET may be necessary to realize the full potential of EGFR targeted therapies in CRC.Supernatant HGF levels and relative mRNA expression in fibroblasts treated with LIM1215 CM LIM1215 (Control)MRC5CCD18CAF3CAF4CAF5CAF6HGF (ng/ml)       Ctrl<0.11.217.310.97.24.08.9LIM1215-CM<0.10.840.015.08.45.015.5        HGF mRNA       Ctrl00.81.82.63.01.6 LIM1215-CM01.07.65.44.91.8 

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 705.
Abstract 708: A novel hedgehog signaling inhibitor blocks hedgehog-mediated carcinogenesis
Authors: Xie Jingwu , Fan Eric Qipeng , Schelgel Chelsea , Zhang Xiaoli , Garrisson Massound , He Miao
Published: February 2011

The link of hedgehog (Hh) signaling activation to human cancer and synthesis of a variety of Hh signaling inhibitors raise great expectation that inhibiting Hh signaling may be effective in human cancer treatment. We have developed a novel hedgehog signaling inhibitor Cyc-T, which has better solubility and higher efficacy in inhibiting hedgehog signaling. In analyses of motor neuron differentiation and osteoblast differentiation, we found that Cyc-T is very effective in preventing hedgehog-mediated functions. Using a mouse model based on keratin 14 promoter-driven expression of an activated form of SMO, SmoM2, we found that a cyclopamine derivative cyc-T are effective in shrinkage of Smo-mediated tumors. K14cre/R26SMOm2yfp mice develop skin dysplasia on postnatal day 3 and BCC-like tumors by day 15. We performed topical application of cyclopamine or its derivatives CPA006, Cyc-T on the skin of K14cre/R26SMOm2yfp mice daily for seven days with the solvent ethanol as the control. The effects of these reagents on the tumors were examined by histological examination of skin biopsies and expression of Hh target genes. In comparison with the control group, we found that 6 out of 9 mice treated with cyclopamine reduced tumor mass and decreased expression of Hh target genes. The mice with no effects by cyclopamine and its derivatives had no changes in Hh target gene expression. In an orthotopic mouse model of pancreatic cancer, we measured the tumor progression following drug treatment and obtained promising results. Taken together, we discovered that Cyc-T are an effective inhibitor for Hh signaling-mediated carcinogenesis.

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 708.
Abstract 716: CXCR4 antagonist, CTCE-9908, inhibits PC-3 cell invasiveness and metastasis
Authors: Chinni Sreenivasa R. , Kandagatla Pridvi , Wong Donald , Korz Walter
Published: February 2011

Chemokines and their receptors function in the migration and homing of cells to target tissues. Current evidence suggests that cancer cells activate chemokines and their receptors during dissemination and metastasis formation at secondary sites. Previously, in prostate cancer cells, we showed that binding of the chemokine CXCL12 to its receptor CXCR4 induced signaling events leading to MMP-9 expression, migration and invasion. CXCL12/CXCR4 transactivation of epidermal growth factor family receptors in lipid raft membrane microdomains on cell surface mediates these invasive signaling events and subsequent expansion of the skeletal metastatic deposits. Herein, we tested the efficacy of CXCR4 antagonist CTCE-9908 in prostate cancer cell growth, invasion, and metastasis. Methods: We used in vitro growth and invasion assays and in vivo orthotopic model system to asses the tumor growth and metastasis. Results: We found that (a) CTCE-9908 treatment resulted in no significant change in the growth of PC-3 cells; (b) CTCE-9908 compound at 50 μg/ml inhibited CXCL12 mediated invasion of PC-3 cells; (c) in vivo CTCE-9908 compound did not significantly altered the growth of primary tumors as measured by calipers and (d) the total tumor burden in the animal including the growth of prostate and soft tissue metastases to lymph node and distant organ tissues were significantly decreased upon CTCE-9908 administration. Histological analysis showed that CTCE-9908 treatment resulted in tumor necrosis in primary prostate tumors. Conclusions: These data suggest that CXCR4 inhibition by CTCE-9908 compound decreased the invasive signaling and metastatic spread of tumor cells without apparently affecting the primary tumor growth in animal model. Hence, CTCE-9908 is an efficacious antagonist for CXCL12/CXCR4 signaling pathway mediating invasive spread of tumor cells in secondary metastasis formation. Studies of CTCE-9908 in phase II are planned.

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 716.
Abstract 717: The diterpenoid lactone andrographolide (andro) induces apoptosis in all- trans-retinoic-acid (ATRA) resistant acute promyelocytic leukemia (APL) cell lines: Potential therapeutic implications
Authors: Yang Shuo , Evens Andrew M. , Prachand Sheila , Singh Amareshwar T.K. , Bhalla Savita , Platanias Leonidas , Gordon Leo I.
Published: February 2011

Andro is a diterpenoid lactone isolated from Andrographis paniculata, an important herbal medicine used in Asia to treat a range of diseases, (respiratory infection, fever, diarrhea). Andro is the major bioactive component extracted from Andrographis paniculata. The three hydroxyls at C-3, C-19 and C-14 are responsible for its biological activities. Recently, the anti-cancer properties of Andro have been recognized, and we found that Andro results in mitochondrial-mediated apoptosis in lymphoma cell lines and fresh malignant cells from patients with lymphoma. Based on the mechanism of action of this novel diterepenoid lactone and a prior report (Manikam et al Journal of Pharmacy and Pharmacol 2009, 61:69), we hypothesized that Andro may have activity in APL. The cell differentiation inducing agent all-trans retinoic acid (ATRA) combined with chemotherapy has become standard first-line treatment for APL through a mechanism that involves cell differentiation, but recent data suggest that ATRA alone or ATRA and arsenic trioxide, which induces apoptosis by degradation of the chimeric fusion protein PML-RARα, may replace ATRA/chemotherapy as initial treatment. Because Andro appears to work through a different mechanism than ATRA, we utilized ATRA-sensitive NB4, ATRA-resistant NB4 (NB4-007/6 and NB4-306) cell lines to test the in-vitro anti-leukemia effect of Andro on APL cells by measuring cell viability (MTT assay), apoptosis (Annexin V-FITC/PI, FACS), mitochondrial membrane potential (MMP) (TMRE/ FACS), cell differentiation (CD11B staining/FACS) and signaling pathways (Western blot). We found that Andro reduced APL cell viability, MMP and increased apoptosis in a time & dose-dependent manner. Moreover ATRA-resistant NB4-007/6 or NB4-306 cell lines were more sensitive to Andro compared with the ATRA-sensitive NB4 cell line. We found that the mechanism of Andro activity is not related to cell differentiation as measured by CD11b. In order to interrogate signaling pathways involved, we studied p27kip1 and FasL expression and found that Andro resulted in up-regulation of both. One of the forkhead box O (FOXO) proteins, FOXO1, a transcription factor, also demonstrated increased phosphorylation at Thr24 at 1h, 3h, and 24h Andro, but reduced phosphorylation at Thr24 at 6h and 16h. Taken together, these data suggest that Andro may have activity in patients with ATRA-resistant APL by a mechanism of action that is distinct from ATRA. Thus, further studies of diterpenoid lactones in APL are warranted.

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 717.
Abstract 726: Targeting the extracellular and intracellular interfaces of CXCR4
Authors: Meier Jason B. , Jala Venkatakrishna R. , Bodduluri Haribabu , Trent John O.
Published: February 2011

CXCR4 is a G protein coupled receptor that has multiple critical functions in normal and pathologic physiology that include regulation of the metastatic behavior of mammary carcinoma, and utilization as a coreceptor for infection by T-tropic strains of HIV-1. We have targeted CXCR4 via an interdisciplinary approach including molecular dynamics simulations of partial and fully constitutively active, and inactive forms of CXCR4 performed in a solvated lipid bilayer to reproduce the microenvironment of this integral membrane protein. We have discovered that it is the partially activated form that can interact with the G-protein complex. We have used in silico virtual screening to select possible CXCR4 inhibitors. Two sites were chosen to disrupt CXCR4 function. The extracellular site was targeted to interfere with the natural ligand for CXCR4, SDF-1. The intracellular site was targeted to interfere with the G-protein: CXCR4 interface. The ranked lists of compounds were tested for their ability to alter chemotaxis, an in vitro model for metastasis, and calcium flux, a measure of CXCR4 signaling capability. Several small molecules were found to inhibit these functions of CXCR4. The presented approach combines computational studies with molecular biological data and provides a powerful tool in the search for new inhibitors.

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 726.
Abstract 727: Highly potent first examples of dual aromatase-steroid sulfatase inhibitors based on a diaryl template
Authors: Woo L. W. Lawrence , Jackson Toby , Putey Aurelien , Cozier Gyles , Leonard Philip , Acharya K. Ravi , Chander Surinder K. , Purohit Atul , Reed (Deceased) Michael J. , Potter Barry V. l.
Published: February 2011

Designing single agents that act against multiple biological targets are of increasing interest and prominence. In recent years, an increasing volume of work has been published exemplifying the successful use of this strategy. Estrogen deprivation has been an effective therapeutic intervention for hormone-dependent breast cancer (HDBC). One established and clinically proven approach involves the inhibition of the aromatase enzyme, although the inhibition of steroid sulfatase (STS) is an emerging new strategy as demonstrated by the promising results of STX64 (BN83495),1 the first STS inhibitor that entered clinical trials. While both aromatase and STS are targets for treating HDBC, it is reasoned that estrogen deprivation may be more comprehensively achieved by dual inhibition of both enzymes. To this end, the reversible aromatase inhibitory pharmacophore, which is principally a heme-ligating nitrogen-containing heterocycle, was thus introduced into a known sulfamate-based STS inhibitor to give a series of dual aromatase-sulfatase inhibitors (DASIs). Several compounds were found to be good aromatase or STS inhibitors individually. However, two compounds show exceptional dual in vitro potency in JEG-3 cells, inhibiting aromatase with IC50 values of 0.5 and 2.0 nM, and STS with IC50 values of 5.5 and 35 nM. Both compounds share the common structural feature of having a sulfamate group on one aryl ring, and a heterocycle and a H-bond acceptor on the other. At 1 mg/kg p.o., both DASIs reduced plasma estradiol levels strongly and inhibited liver STS activity potently in vivo. One of the compounds was found to be non-estrogenic and potently inhibit human carbonic anhydrase II (hCAII IC50 = 86 nM). A complex of hCAII with the inhibitor was crystallized and solved by X-ray crystallography. This class of DASI should encourage further development towards multi-targeted therapeutic intervention in HDBC.

Ref 1: a) Stanway, S. J. et al. Clin. Cancer Res. 2006, 12, 1585-1592. b) Stanway, S. J. et al. Oncologist 2007, 12, 370-374.

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 727.
Abstract 73: Mitochondrial 10-formyltetrahydrofolate dehydrogenase: A novel enzyme in folate metabolism
Authors: Dubard Marianne E. , Strickland Kyle C. , Oleinik Natalia V. , Krupenko Natalia I. , Krupenko Sergey A.
Published: February 2011

Cytosolic 10-formyltetrahydrofolate dehydrogenase (FDH, Aldh1L1) is an important regulator of intracellular folate pools, which displays antiproliferative effects in cancer cells. We have identified a gene at the chromosome locus 12q24.11 in the human genome, the product of which has 87% sequence similarity with cytosolic FDH. This protein has an extra amino-terminal sequence of 22 amino acid residues enriched in arginines, which is predicted to be a mitochondrial translocation signal. The mitochondrial targeting function of the leader has been confirmed in Cos7 cells: green fluorescent protein (GFP)-tagged at the amino-terminus with the leader, localizes to mitochondria. Transfection of Cos7 or A549 cell lines with a construct, in which GFP has been introduced between the leader sequence and the rest of the putative mitochondrial FDH (mtFDH), has also shown mitochondrial localization, suggesting that the identified gene encodes a mitochondrial enzyme. To evaluate the abundance of mtFDH, we have measured its mRNA levels in a wide array of human tissues by real-time PCR, and compared them to the levels of mRNA that encode cytosolic FDH. While cytosolic FDH mRNA is highest in liver, kidney and pancreas, mtFDH mRNA is most highly expressed in pancreas, heart and brain, but not in liver or kidney. In contrast to the cytosolic enzyme, which is non detectable in human cancer cell lines, the presence of mtFDH mRNA was demonstrated in A549 and PC3 cells by conventional and real-time PCR. The presence of the endogenous enzyme in mitochondria of A549 cells has been further confirmed using specific polyclonal antibody generated against purified recombinant mtFDH. We have also shown that recombinant mtFDH, similar to the cytosolic enzyme, catalyzes NADP+-dependent oxidation of the 10-formyltetrahydrofolate to tetrahydrofolate and CO2. Thus, the enzyme is a likely source of CO2 production in mitochondria and we propose that it plays an essential role in distribution of one-carbon groups between cytosolic and mitochondrial compartments of the cell.

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 73.
Abstract 730: Targeting colchicine binding site in tubulin for the treatment of advanced melanoma
Authors: Li Wei , Wang Zhao , Chen Jianjun , Lu Yan , Li Chien-Ming , Dalton James T. , Miller Duane
Published: February 2011

Inhibition of tubulin dynamics has been a fruitful drug target for cancer treatment. Out the three ligand binding sites in tubulin, drugs targeting taxane binding site (e.g. taxol and epithilones) and vinca alkaloids binding site (e.g. vinblastine) are clinically very successful drugs. However, no FDA-approved cancer drugs targeting the colchicine binding site in tubulin currently exist. In our ongoing efforts to search for effective drugs for the treatment of advanced melanoma that are highly resistant to all existing cancer treatment, we have discovered a new class of compounds that are not substrates of P-glycoprotein (Pgp) and are highly efficacious against human melanoma tumors in A375 xenograft models. At relatively low doses (5∼15 mg/kg), these compounds inhibit melanoma tumor growth more effectively than DTIC (the gold standard drug for advanced melanoma) at 60 mg/kg. Extensive biological and molecular modeling studies suggest that these compounds work by competitively binding to the colchicine site in tubulin. Further optimization of these compounds hold great promise in providing an effective agent for the treatment of advanced melanoma.

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 730.
Abstract 736: Novel, potent MEK inhibitor CIP-1374 for the treatment of cancer
Authors: Chordia Mahendra , Khire Uday , Bhargava Ajay
Published: February 2011

Mitogen-activated protein kinases (MAPKs) are clinically validated, well-established targets for seeking anti-cancer therapy. MAPKs specifically phosphorylate serine/threonine residues of proteins, which are activated by variety of external stimuli (for example mitogens and growth factors) to manifest its actions inside the cell. The activation of MAPKs regulates many functions of the cells with profound physiological implications such as cell growth, survival, apoptosis, differentiation, proliferation and gene expression. MEK 1 and 2 are two human kinases presented in the middle of the classical MAPK-cascade involving upstream RAS-RAF and downstream ERKs. This signal transduction cascade ending in phophorylation of ERKs is extensively studied in cancer pathology. The phosphorylated-ERK upon its translocation to nucleus activates several transcription factors to induce the expression of many genes required for cell survival and proliferation. Because of very high selectivity conferred on MEKs to phophorylate only ERK1 and ERK2, targeting its inhibition offers a very attractive strategy for anti-cancer drug discovery. In addition, the mechanism of action of the known MEK inhibitors such as PD98059 and U0126 is non ATP-competitive (binding to allosteric site) and thus may have least side effects in clinical utility sets in high priority to discover new inhibitors. Currently there are few MEK inhibitors undergoing clinical studies, these includes ARRY-162 (Array Biopharma), RDEA-119 (Ardea biosciences) in combination with sorafenib displaying significant response in sorafenib resistant hepatoma cells and XL-518 (Exelixis) for solid tumors. We present here the discovery of a potent novel MEK inhibitor CIP-1374 for the treatment of cancer. The preliminary in vitro data including inhibition of ERK phophorylation, and whole cell anti-proliferation assays in variety of cancer cell lines will be presented. CIP-1374 exhibited MEK-1 IC50 21.0 nM. In the presence of 1.0 mM ATP, CIP-1374 retained inhibitory activity indicating allosteric mode of binding in ATP non-competitive fashion. The selectivity for MEK inhibition leading to non-proliferation will be presented from the cells without RAS mutation.

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 736.
Abstract 74: Differential regulation of Erk signaling by mitochondrial superoxide controls the metastatic potential of bladder cancer cells
Authors: Ye Hanqing , Hempel Nadine , Moore Erin , Mian Badar , Melendez Juan A.
Published: February 2011

Microarray profiling and analysis of in vivo-selected metastatic bladder cancer has revealed a strong association between manganese superoxide dismutase (Sod2) expression and invasiveness. However, whether increased Sod2 expression contributes to progression from transitional cell carcinoma (TCC) to invasive bladder cancer has not been established. To test this hypothesis we evaluated the impact of Sod2 overexpression on invasive and migratory properties of two distinct TCC cell lines (UC3 and UC6). Sod2 overexpression increased wound healing and matrigel transwell migration of UC6 while inhibiting these activities in the UC3 cell line. Increases or decreases in invasiveness were accompanied by parallel shifts in the levels of matrix metalloproteinases and VEGF. The opposing effects of Sod2 on TCC invasion and the secretory phenotype were associated with differential Erk activation. Sod2-dependent suppression of Erk activation and MMP-1 expression was reversed by EGF activation in UC3 cells while Erk inhibition blocked the Sod2-mediated elevation in ERK activity, MMP-1 and VEGF levels. Activation or inhibition of Erk by Sod2 in the UC6 and UC3 cell lines, respectively, was driven by superoxide (O2−.) and not H2O2. Increasing mitochondrial superoxide by treatments with complex I or III inhibitors reversed the effects of Sod2 overexpression in both cell lines. Sod2-dependent inhibition of invasion in vitro was associated with impairment in growth of orthotopically implanted tumors. These findings unveil distinct redox regulated attributes of TCC that participate in controlling their invasiveness both in vitro and in vivo. In addition, we have identified the ERK signaling network as a primary target for controlling divergent tumorigenic responses to Sod2 overexpression.

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 74.
Abstract 740: Oncodesign translational drug discovery process
Authors: Mirjolet Jean-Francois , Bichat Francis , Tizon Xavier , Serin Guillaume , Koob Zina , Duchamp Olivier , Blom Petra , Berthet Cyril , Genne Philippe , Hoflack Jan
Published: February 2011

Despite major advances in translational methods, oncology drug discovery remains a largely sequential process, in most cases starting from a molecular target with some initial but often limited relevance to the disease. It takes multiple years and investments in the order of 5-10 million US$ before a compound is validated in advanced efficacy and safety models. In addition, due to the remoteness of many of the models to real disease pathophysiology, attrition rates in clinical phases remain very high.

We have developed and validated a flexible and iterative “Translational Drug Discovery Process”, in which multiple translational research platforms create a tight link between molecule development and cancer pathophysiology.

Our “Translational Drug Discovery process” is based on multiple research platforms that are used as flexible building blocks to design an optimal process to advance compounds and position them towards precise oncology applications. These platforms include cellular models, in vivo tumourgrafts, phenotypic models and a multi-modality imaging platform. Recent additions that are based on fresh patient-derived tumour tissue include ex vivo 3D models and low passage tumourgrafts. We have also initiated a program to discover 18F-labeled kinase inhibitor based biomarkers/imaging tracers for use in PET applications. Our research base was recently completed with the addition of a diverse collection of potent and selective kinase inhibitors and the associated proprietary chemical technology.

We will describe how we have validated the ex vivo 3D model, based on tumour tissue collected directly in patients or from low passage tumourgrafts in mice. After mincing in small fragments and culturing in poly-HEMA coated plates, the tumour fragments are treated with drug for 24h. Tissue-microarrays are created by fixing and embedding the fragments in paraffin. Different classes of reference compounds were analysed by immunohistochemistry of Ki67 (proliferation) and M30 (apoptosis) using automated quantification. Following this validation, the model is used to select promising new compounds from our internal collection for further optimization or to position advanced compounds from our clients.

We will finally demonstrate how our “Translational Drug Discovery process” has allowed the identification of attractive compounds of high disease relevance at a fraction of the typical drug discovery cost within reduced timelines.

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 740.