bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2461: Gene deletion of mPGES-1 results in a significant reduction in tumor stage and burden in the BBN mouse model of bladder cancer
Authors: Nakanishi Masako , Choudhary Shilpa , Hegde Poornima , Pilbeam Carol , Rosenberg Daniel , Taylor John A.
Published: February 2011

Introduction and Objective: Prostaglandins (PG) play a critical role in the development of solid tumors. Inhibition of cyclooxygenase-2 (COX-2), the committed step in PG synthesis, has been shown to impact tumor development in the BBN mouse model of bladder cancer. However, clinical translation has been impeded by the well documented cardiovascular problems with COX-2 inhibitors. Inhibition of other synthases downstream of COX-2 in the PG cascade may prove useful for chemoprevention or chemotherapy. In the following study, we evaluated the impact of gene deletion of membrane-associated prostaglandin synthase-1 (mPGES-1), the terminal synthase in the generation of PGE2, in the BBN mouse model.

Methods: Two month old male C57Bl/6 mice with global homozygous deletion for Ptges (mPGES-1 KO) and wild-type (WT) were treated with and without BBN in drinking water (n=5/grp) for 23 weeks. Bladders were harvested, weighed, fixed, step sectioned and evaluated by H&E for pathologic stage and grade and percent involvement of tissue by tumor. Urine was collected prior to sacrifice and evaluated for PGE-2 metabolites (PGEM) by ELISA.

Results: All animals had high-grade cancer at the time of euthanasia. WT bladders were significantly heavier than KO bladders (mean 0.18 vs 0.04 g respectively; p=0.03). Four of 5 WT mice had pT3 and one pT2 disease. Three of 5 KO mice had pT1 and 2 had pT2 disease. % tissue involvement was significantly less in the KO animals as well (54 vs 87%, p=0.03). Pearson's correlation for bladder weight and percent tumor involvement was 0.73 (p=0.02). Pathologic stage was dependant on genotype by Wilcoxon Rank Sum Test (p=0.02). Urine metabolites were reduced 50% in the KO compared to WT animals (mean 18.9 vs 45.2 pg/ml respectively; p<0.01).

Conclusions: Genetic deletion of mPGES-1 resulted in significant decreases in PGE-2 as evidenced by lower urinary PGEM and in pathologic stage and % tissue involvement by tumor. Increased bladder weights in the WT mice compared to KO mice were directly related to the increased tumor burden. These results suggest that inhibitors of mPGES-1 may prove to be an alternative to COX inhibition and possibly prove useful in the management of high-grade bladder cancer.

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 2461.
Abstract 2468: Selective aldosterone blocker inhibits murine hepatocellular carcinoma development via angiogenesis suppression
Authors: Kaji Kosuke , Yoshiji Hitoshi , Ikenaka Yasuhide , Noguchi Ryuichi , Shirai Yusaku , Fukui Hiroshi
Published: February 2011

It is now widely recognized that angiogenesis is a crucial step in the development of tumors, including hepatocellular carcinoma (HCC). Therapies targeting the tumor vessels have proven successful for cancer treatment in experimental models and, in the clinical practice, several anti-angiogenic agents have already been employed. The renin-angiotensin-aldosterone system (RAAS) has also become known as a prerequisite for tumor angiogenesis, including HCC. We previously reported that angiotensin-II (AT-II) plays an important role in HCC development, and that suppression of AT-II by angiotensin-converting enzyme (ACE) inhibitor attenuated the tumor growth of HCC via suppression of intra-tumoral angiogenesis. Recent studies have also suggested the possible involvement of aldosterone (Ald) in neovascularization, although the actual role of Ald in tumor angiogenesis is still vague. The aim of our current study was to elucidate the effect of eplerenone (Epl), a clinically used selective Ald blocker, on HCC development, especially in conjunction with angiogenesis, using subcutaneous HCC model. To create an allograft model, we injected 1×106 of BNL-HCC cells into the flanks of BALB/c mice. The administration of Epl by daily gavage at a dose of 100 mg/kg/day was started on day 14, when the mean tumor volume was ∼ 200 mm3. The tumor volume was measured twice a week, and the mice were killed 35 days after tumor cell implantation. Administration of Epl at clinically comparable low dose could suppress HCC development as compared with the non-treated control. Epl treatment resulted in a marked increase of TUNEL-positive apoptosis in the tumor, whereas Ki67-positive tumor cell proliferation was not altered. Both CD31-immunopositive neo-vessels and VEGF expression in the tumor were markedly suppressed by this treatment in a magnitude similar to the inhibitory effect on the tumor growth. Our in-vitro study showed that Epl significantly suppressed the Ald-induced endothelial proliferation and tubular formation via inhibition of phosphorylation of the extracellular signal-regulated kinase 1/2. On the other hand, neither Ald nor Epl affected the proliferation of HCC cells. In conclusion, these results indicated that aldosterone plays a pivotal role in HCC development via VEGF-mediated tumor angiogenesis, and that Epl may be a potential new strategy for HCC therapy in the future.

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 2468.
Abstract 2469: Combination of branched-amino acids and ACE inhibitor attenuates the insulin resistance-based hepatocarcinogenesis via angiogenesis suppression
Authors: Shirai Yusaku , Yoshiji Hitoshi , Ikenaka Yasuhide , Noguchi Ryuichi , Kaji Kosuke , Fukui Hiroshi
Published: February 2011

A strong correlation between insulin resistance (IR) and hepatocellular carcinoma (HCC) has recently been identified, although little is known about the possible mechanisms involved. IR is a consistent finding in patients with type 2 diabetes mellitus (DM), and a close interaction between DM and HCC has been documented. Since neovascularization plays an important role in HCC including hepatocarcinogenesis, an angiostatic therapy would be a promising approach for chemoprevention against HCC. The aims of this study were to elucidate the role of angiogenesis in IR-based hepatocarcinogenesis, and to elucidate the therapeutic effect of combination of clinically used branched-chain amino acids (BCAA) and angiotensin-converting enzyme inhibitor (ACE-I), in conjunction with neovascularization. The combined effects on the development of liver enzyme-altered pre-neoplastic lesions, angiogenesis, and several indices were elucidated in obese diabetic rats. We also performed several sets of in-vitro experiments to examine the mechanisms involved. The IR status directly accelerated hepatocarcinogenesis, and treatment with both BCAA and ACE-I markedly inhibited the development of pre-neoplastic lesions under the condition of IR along with suppression of angiogenesis and VEGF expression in the liver. The combination treatment with BCAA and ACE-I exerted more potent inhibitory effects than single-agent treatments. Our in-vitro study demonstrated that BCAA and ACE-I inhibited the endothelial tubular formation almost in parallel with suppression of development of pre-neoplastic lesions. This combination regimen with BCAA and ACE-I showed a marked chemopreventive effect against hepatocarcinogenesis along with suppression of neovascularization and VEGF expression in obese diabetic rat as compared with either single-agent treatment. These inhibitory effects were achieved at clinically comparable low doses. Since both BCAA and ACE-I are widely used in the clinical practice, this combination therapy may represent a potential new strategy for chemoprevention against IR-based HCC in the future.

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 2469.
Abstract 248: Functional characterization of MED29 gene in pancreatic cancer
Authors: Kuuselo Riina , Savinainen Kimmo , Kallioniemi Anne
Published: February 2011

Pancreatic cancer is a highly aggressive malignancy characterized by rapid progression and an extremely poor prognosis. More knowledge is needed to better understand the mechanisms of the disease pathogenesis. Identification of new disease genes can improve our understanding about the biology of pancreatic cancer and might provide us tools for improving both diagnostics and treatment.

Previously, we performed a detailed characterization of the 19q13 locus, one the recurrently amplified regions in pancreatic cancer, and identified a set of putative amplification target genes that showed consistent overexpression in amplified cell lines versus non-amplified cell lines. Subsequent high-throughput RNAi screen targeting genes within the amplified region revealed a number of genes whose downregulation led to decreased cell viability in the amplified but not in the non-amplified cells. Of these, the mediator complex subunit 29 (MED29) was consistently overexpressed in all amplified cell lines and had the most dramatic effect on cell viability. By flow cytometry analysis, we showed that MED29 silencing resulted in G0/G1 cell cycle arrest and increased apoptosis in the amplified PANC-1 cells. MED29 is a subunit of a large multiprotein complex that is involved in transcriptional regulation and may function both as a transcriptional repressor and enhancer. To further define the function of MED29 we constructed a lentiviral vector for MED29 gene delivery. Mouse embryonic fibroblasts (NIH/3T3) and two pancreatic cancer cell lines with low basal MED29 expression (MIAPaCa2 and Hs700T) were transduced with a lentivirus vector containing MED29. Stable MED29 expression was confirmed in all three cell lines by quantitative RT-PCR and western blot analysis. MED29 expression led to statistically significant changes in proliferation and/or survival in both the NIH/3T3 cells and the pancreatic cancer cells. Finally, MED29 overexpressing human pancreatic cancer cell lines were subcutaneously inoculated into immunocompromized mice to follow tumor growth in vivo. Significant growth difference was observed between MED29 expressing cancer cells compared to control cells when injected into mice. To conclude, our results implicate MED29 as an important regulator of growth and survival in pancreatic cancer both in vivo and in vitro.

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 248.
Abstract 2487: Mechanism of action KX01 (KX2-391), a novel tubulin polymerization and Src signaling inhibitor proceeding to Phase II clinical trials
Authors: BU YAHAO , Smolinski Michael , Qu Jun , Kazim Latif , Gelman Irwin , Dyster Lyn , Hangauer David
Published: February 2011

KXO1 (KX2-391) has completed Phase I clinical trials in cancer patients and is now proceeding to Phase II. KXO1 was first characterized as a Src signaling inhibitor that exhibits potent anti-proliferative activity against a broad range of solid and liquid human cancers in vitro and in mouse models, including synergistic activity with other drugs. Using photo-affinity labeling technology, we have discovered that KXO1 also binds to tubulin at a novel binding site, and inhibits tubulin polymerization. KXO1 leads to cell cycle arrest at M phase with subsequent cell death mediated by canonical apoptosis pathways including phosphorylation of Bcl-2, loss of mitochondrial membrane permeability, and activation of caspases. Interestingly, inhibition of caspase activity by Z-VAD-FMK abrogates KXO1 induced cell death of HT29 and LNCaP cells but not in PC3-LN4 cells, suggesting the existence of caspase independent pathways capable of executing KXO1-induced cell death in certain cell types. In multi-drug resistant cancer cell lines, which overexpress p-glycoprotein, KXO1 still has equally potent anti-proliferative activity with GI50's in the low nanomolar range. KXO1 also exhibits strong anti-angiogenesis and vascular disrupting activities as revealed by an in vitro vasculature tube formation assay. KXO1 demonstrates robust activity in prostate cancer mouse xenograft models. The safety, tolerability and MTD were determined for KXO1 in a Phase 1 clinical study in patients with advanced malignancies refractory to conventional therapies. KXO1 was well tolerated at the MTD of 40 mg BID; only 1 out of 17 SAE's reported at or below the MTD was assessed as possibly KXO1-treatment related. Phase II clinical trial in prostate cancer patients will commence in early 2010.

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 2487.
Abstract 2502: p53 is critical for the Aurora B kinase inhibitor-mediated apoptosis in acute myelogenous leukemia cells
Authors: Ikezoe Takayuki , Yang Jing , Nishioka Chie , Yokoyama Akihito
Published: February 2011

We previously showed that AZD1152-HQPA, the inhibitor of Aurora B kinase potently induced growth arrest and apoptosis of various types of human leukemia cells including MV4-11 acute myelogenous leukemia (AML) cells, although the molecular mechanisms by which this class of kinase inhibitors induces apoptosis remain to be fully elucidated. We have recently established the MV4-11 subline, designated as MV4-11 TP53 R248W, which possesses transcriptionally inactive R248W mutation in the TP53 gene. MV4-11 TP53 R248W cells were relatively resistant to AZD1152-HQPA-mediated growth arrest, as measured by MTT and clonogenic assays. AZD1152-HQPA (10-100 nM, 48 hrs) strikingly induced apoptosis of MV4-11 cells, as assessed by Annexin V binding, loss of mitochondrial outer membrane potential, and activation of caspase cascade, in parallel with up-regulation of p53 and its target molecules Bax and Noxa. Notably, AZD1152-HQPA (10-100 nM, 48 hrs) induced polyploidy rather than apoptosis in MV4-11 TP53 R248W cells. The polyploid cells were eventually eliminated via apoptosis at later time period (72-120 hrs) in association with up-regulation of p73. Taken together, p53 plays an important role in AZD1152-HQPA induced growth arrest and early-onset of apoptosis in AML cells. P73 may mediate the late-onset of apoptosis to eliminate the polyploid cells caused by the inhibitor of Aurora B kinase.

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 2502.
Abstract 2521: The pan-RAF inhibitor PLX4032 inhibits ERK signaling and tumor cell proliferation in a V600E BRAF mutant-selective manner
Authors: Joseph Eric W. , Pratilas Christine A. , Poulikakos Poulikos I. , Tadi Madhavi , Wang WeiQing , Taylor Barry S. , Persaud Yogindra , Xing Feng , Halilovic Ensar , Viale Agnes , Chapman Paul , Tsai James , Bollag Gideon , Rosen Neal , Solit David B.
Published: February 2011

Activation of the RAS/RAF/MEK/ERK pathway occurs often in human cancer. Tumors with mutant BRAF and some with mutant RAS are dependent upon ERK signaling for proliferation and their growth is suppressed by selective inhibitors of MEK. In contrast, tumor cells with HER-kinase activation proliferate in a MEK-independent manner. These findings have led to the development of RAF and MEK inhibitors as anticancer agents. Like MEK inhibitors, the pan-RAF kinase inhibitor PLX4032 inhibits the proliferation of V600E BRAF mutant tumor cells, but not that of HER kinase dependent tumors. However, tumors with RAS mutation that are sensitive to MEK inhibition are insensitive to PLX4032. Whereas MEK inhibitors inhibit ERK phosphorylation in all normal and tumor cells, PLX4032 only inhibits ERK signaling in tumor cells with BRAF mutation. In contrast, the drug activates MEK and ERK phosphorylation in cells with wild-type BRAF. In mutant BRAF cells, the MEK and RAF inhibitors affect the expression of a common set of genes, whereas they have opposite effects on the expression of these genes in tumors with mutant RAS. Furthermore, PLX4032 inhibits ERK signaling output in mutant BRAF cells, but transiently activates it in tumor cells with wild-type RAF. Thus, PLX4032 inhibits ERK signaling output in a BRAF mutant-selective manner. These data suggest that PLX4032 may have a broader therapeutic index and help to explain the greater antitumor activity observed with this drug compared to MEK inhibitors, which inhibit ERK signaling in all cells.

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 2521.
Abstract 2526: CP690550 inhibits interleukin-2 (IL-2), IL-9 and IL-15 mediated JAK3 activation and proliferation of T cells obtained from patients with HTLV-1-associated adult T cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP)
Authors: Ju Wei , Zhang Meili , Bryant Bonita R. , Chen Jing , Morris John C. , Janik John E. , Waldmann Thomas A. , Jiang Jian-kang , Thomas Craig J. , Jacobson Steven , Oh Unsong
Published: February 2011

The retrovirus, human T cell lymphotrophic virus-1 (HTLV-1) is the etiologic agent of ATL and the neurological disorder, HAM/TSP. Infection with HTLV-1 results in viral tax protein mediated constitutive activation of the JAK3/STAT5 pathway in patients' T-lymphocytes that is believed to result in malignant transformation or autoimmunity. JAK3 plays a key role in cytokine receptor-mediated signal transduction including the receptors for IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21 that use the common γ- chain. This suggests that inhibitors of JAK3 may be of benefit in HTLV-1-associated diseases. We evaluated the effect of a novel small molecule JAK3 inhibitor, CP690550 on the cytokine-dependent cell line, NK92. CP690550 at 50 or 100 nM effectively inhibited the proliferation of IL-2-stimulated NK92 cells, but not NK92 that were stimulated with IL-12 or the combination of IL-6 and its receptor (IL-6R) that do not utilize the common γ-chain and JAK3 for signaling and proliferation. We also demonstrated the relative specificity of this inhibitor on another cytokine-dependent cell line, 32Dβ where at 50 or 100 nM CP690550 inhibited IL-2 and JAK3 mediated proliferation but did not affect murine IL-3-mediated cell proliferation. The activation mediated phosphorylation of STAT5 in NK92 cells in response to IL-2 stimulation was inhibited by CP690550. We evaluated the action of CP690550 on the ex vivo proliferation of peripheral blood mononuclear cells (PBMC) from selected patients with the smoldering or chronic subtypes of ATL, or HAM/TSP whose PBMC are associated with the production of cytokines, in particular, IL-2, IL-9 and IL-15 that use the common γ-chain and JAK3. CP690550 at 50 nM inhibited the ex vivo spontaneous proliferation of PBMC from 11 patients with ATL and 9 patients with HAM/TSP at 6-day by means of 66% and 86.4%, respectively. CP690550 also inhibited the proliferation of NK92 cells stimulated with the 6-day ex vivo PBMC culture supernatants from ATL and HAM/TSP patients by a mean of 83.9% and 82%, respectively. We also demonstrated that CP690550 effectively inhibited the activation of the JAK3/STAT5 pathway in isolated T-cells from ATL patients. On the basis of this study, the JAK3 inhibitor CP690550 deserve further study as a potential therapeutic agent for select patients with HTLV-1-associated ATL and HAM/TSP.

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 2526.
Abstract 2537: The novel sphingosine kinase inhibitor ABC294640 blocks survival and tumorigenesis in chemo- And endocrine resistant breast cancer
Authors: Antoon James W. , White Martin D. , Gestaut Matthew M. , Slaughter Evelyn M. , Elliott Steven , Driver Jennifer L. , Khalili Hafez S. , Smith Charles D. , Burow Matthew E. , Beckman Barbara S.
Published: February 2011

Resistance to chemotherapy is a major cause of breast cancer treatment failure today. The sphingolipid metabolizing enzyme sphingosine kinase converts pro-apoptotic ceramide into pro-survival sphingosine-1-phosphate (S1P). Sphingosine kinase has been shown to be involved in breast cancer endocrine- and chemo-resistance and is proposed to be a key regulator of proliferation. Here, we studied the effects of the novel sphingosine kinase inhibitor, ABC294640 (3-(4-chlorophenyl)-adamantane-1-carboxylic acid (pyridin-4-ylmethyl) amide), on human breast cancer and compared them to its parent compound, SKI-II (4-[[4-(4-Chlorophenyl)-2-thiazolyl]amino]phenol). Treatment with ABC294640 increased levels of endogenous long chain ceramides while simultaneously reducing levels of S1P in vitro. ABC294640 blocked both viability and survival at low micromolar IC50 concentrations in the endocrine therapy resistant - MDA-MB-231, MDA-MB-468 and MDA-MB-361 cells, as well as the chemotherapy resistant MCF-7TN-R cell line. Furthermore, treatment with the inhibitor significantly reduced proliferation, as seen in immunofluorescence staining of Ki-67 in vitro. Interestingly, ABC294640 can induce apoptosis and release caspase-9, possibly through modulation of activity of the transcription factor NF-kappaB. Treatment with 10μM of the inhibitor blocked transcription of genes known to be mediated by the NF-kappaB, such as at IAP1, IAP2, and SOD1. Finally, utilizing xenograft immunocompromised mouse models, ABC294640 significantly reduced breast cancer tumorigenesis of MDA-MB-468 and MCF-7TN-R in vivo. Treatment with 50 mg/kg of the inhibitor for 14 days decreased MDA-MB-468 tumor volume by 37.3% (n=10) compared to vector control. Similarly, there was a 67.4% decrease in tumor volume following 18 days of treatment in the chemoresistant MCF-7TN-R xenograft mice (n=10). These results indicate that pharmacological inhibition of sphingosine kinase with an orally bioavailable selective inhibitor, ABC294640, has therapeutic potential in the treatment of chemo- and endocrine therapy- resistant breast cancer.

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 2537.
Abstract 254: A HDAC1-binding domain within FATS bridges p21 turnover to radiation-induced tumorigenesis
Authors: Li Zheng , Zhang Qian , Mao Jian-Hua , Weise Anja , Mrasek Kristin , Fan Xueyu , Zhang Xifeng , Liehr Thomas , Lu Karen H. , Balmain Allan , Cai Wei-Wen
Published: February 2011

There is a gap between the initial formation of cells carrying radiation-induced genetic damage and their contribution to cancer development. Herein we reveal a previously uncharacterized gene FATS through a genome-wide approach and demonstrate its essential role in regulating the abundance of p21 in surveillance of genome integrity. A large exon coding the NH2-terminal domain of FATS, deleted in spontaneous mouse lymphomas, is much more frequently deleted in radiation-induced mouse lymphomas. Its human counterpart is a fragile site gene at a previously identified loss of heterozygosity (LOH) site. FATS is essential for maintaining steady-state level of p21 protein and sustaining DNA damage checkpoint. Furthermore, the NH2-terminal FATS physically interacts with HDAC1 to enhance the acetylation of endogenous p21, leading to the stabilization of p21. Our results reveal a molecular linkage between p21 abundance and radiation-induced carcinogenesis.

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 254.
Abstract 2545: Regulation of NF-YB by miR-485-3p affects TopoisomeraseIIα expression
Authors: Chen Cheng-Fen , Beck William T. , He Xiaolong
Published: February 2011

DNA topoisomerase IIα (TopoIIα) is an essential nuclear enzyme. Our teniposide-resistant human leukemic lymphoblastic cells (CEM/VM-1-5) express reduced TopoIIα protein. To determine the role of TopoIIα in drug resistance, previously we knocked down TopoIIα in parental CEM cells by RNAi. Compared to control cells, CEM-siTopoIIα cells are more resistant to etoposide. Our previous work suggested that the transcription factor NF-YB is a negative regulator of TopoIIα, working through the TopoIIα promoter (Morgan and Beck, Mol. Pharmacol 59:203,2001). We have found that NF-YB protein expression is increased in CEM/VM-1-5 cells compared to CEM cells. This suggests that increased NF-YB may be the cause of reduced TopoIIα in CEM/VM-1-5 cells. We asked what causes the up-regulation of NF-YB in CEM/VM-1-5 cells. MicroRNAs are a recently identified group of non-protein coding RNAs that regulate posttranscriptional gene expression. In our previous study, we found by microRNA profiling that hsa-miR-485-3p, is lower in CEM/VM-1-5 cells compared to CEM cells. MicroRNA target prediction programs also reveal that the 3′ UTR of NF-YB harbors a putative miR-485-3p binding site. We hypothesized that hsa-miR-485-3p regulates drug resistance by decreasing NF-YB expression, which in turn negatively regulates TopoIIα expression. We overexpressed miR-485-3p in CEM/VM-1-5 cells and this led to reduced expression of NF-YB and a corresponding upregulation of TopoIIα. To validate the binding of miR-485-3p to NF-YB 3′-UTR, present experiments with HEK293Tcells using luciferase reporters carrying the 3′-UTR of the NF-YB gene are being done to clarify the role of miR-485-3p in this phenomenon. (Supported in part by CA40570 from NCI and in part by UIC)

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 2545.
Abstract 2580: Single-chain antibody based immunotoxins targeting Her2/neu: design optimization and impact of affinity on antitumor effects
Authors: Cao Yu , Marks James D. , Hittelman Walter N. , Adams Gregory P. , Simmons Heidi H. , Cheung Lawrence H. , Rosenblum Michael G.
Published: February 2011

Recombinant immunotoxins, consisting of single-chain antibodies (scFv) genetically fused to polypeptide toxins represent potentially effective candidates for cancer therapeutics. Antibody affinity, internalization rates and intracellular fate of molecular constructs play critical roles in the eventual cytotoxic activities of these constructs although these events related to the design characteristics of fusion constructs have not been comprehensively explored. We evaluated the relationship between affinity of various anti-Her2 scFv fused to recombinant gelonin (rGel) and its effect on cytotoxic activity. A series of rGel-based immunotoxins were created from the human anti-Her2/neu scFv C6.5 and various affinity mutants (designated ML3.9, MH3B1 and B1D3-in increasing affinity order). The affinities of the scFv ranged from 10−8 to 10−11 M and the rGel fusion toxins retained the affinities of the scFv from which they were derived. Against BT474M1 human breast tumor cells, the immunotoxins demonstrated IC50 values of 18.9, 10.9, 3.9 and 1.2nM for the C6.5, ML3.9, MH3B1 and B1D3 based immunotoxins respectively. Targeting indices were highest for the B1D3/rGel construct with the highest affinity. The internalization rate of the fusion constructs into target cells directly correlated with the cytotoxic effects. Competition studies showed that addition of soluble Her2/neu extracellular domain (ECD) as the competitive factor significantly reduced the cytotoxic effects of the highest-affinity fusion construct (B1D3/rGel) while the lower-affinity constructs showed lesser impact of Her2/neu ECD addition on the cytotoxic effect. Animal model efficacy studies against several Her2/neu overexpressing tumors are ongoing but appear to suggest that the construct containing the highest affinity antibody was comparatively less effective in vivo than constructs containing antibodies of intermediate affinity. These studies suggest that human MH3B1/rGel fusion construct with an apparent affinity of 10−10 M displayed an overall optimal cell-killing activity to several Her2/neu positive cancer cells especially in the presence of free Her2/neu ECD. These findings demonstrate the necessity to consider antibody affinity in design considerations for targeted therapeutics. In addition, the potential impact of soluble target or truncated receptor molecules in the environment need to be considered when evaluating the efficacy of targeted therapeutics. Research Conducted, in part, by the Clayton Foundation for Research.

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 2580.
Abstract 2582: Branched neurotensin peptides for the selective targeting of human colon and pancreas carcinoma
Authors: Bracci Luisa , Falciani Chiara , Lelli Barbara , Brunetti Jlenia , Lozzi Luisa , Pini Alessandro , Pagliuca Chiara , Ravenni Niccolò , Bencini Lapo , Menichetti Stefano , Moretti Renato
Published: February 2011

The aim of this study was to validate oligo-branched peptides as selective targeting agents that might be effective either for spotlighting tumor cells that over-express peptide receptors, or for killing them, simply by exchanging the functional moiety coupled to the conserved receptor-targeting core. Tetra-branched peptides containing neurotensin (NT) sequence are described here for selective targeting of human colon, pancreas and prostate cancer. Fluorophore-conjugated peptides were used to measure tumor versus healthy tissue binding in human surgical samples, resulting in validation of neurotensin receptors as highly promising tumor-biomarkers. Drug-armed branched peptides were synthesized with different conjugation methods, resulting in uncleavable adducts or drug-releasing molecules. Cytotoxicity on human cell lines from colon (HT-29), pancreas (PANC-1) or prostate (PC-3) carcinoma indicated branched NT conjugated with MTX and 5-FdU as the most active agents on PANC-1 (EC50 4.4e-007 M) and HT-29 (1.1e-007 M), respectively. Tetra-branched NT armed with 5-FdU was used for in vivo experiments in HT-29-xenografted mice and produced a 50% reduction in tumor growth with respect to animals treated with the same amount of free drug. An unrelated branched peptide carrying the same drug was completely ineffective. In vitro and in vivo results indicated that branched peptides are valuable new tools for tumor selective targeting.

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 2582.
Abstract 2589: Robust in vitro anti-tumor activity of Trastuzamab antibody drug conjugates in HER2 amplified cell lines
Authors: Mohan Sankar , O'Brien Carol , Savage Heidi , Phillips Gail , Lackner Mark , Pandita Ajay
Published: February 2011

Breast cancer is a heterogeneous disease with molecular subtypes characterized by distinct gene expression signatures and histopathology. The defining characteristic of the HER2-amplified subtype is high-level amplification of the HER2 oncogene and overexpression of HER2 protein. Assays for protein overexpression and chromosomal amplification are currently used as diagnostic markers to select patients for HER2-targeting therapies. In the current study, we compared the sensitivity of several HER2-targeting therapies across a panel of 31 breast cancer cell lines and correlated the sensitivity of the cell lines with HER2 positivity detected by IHC, qPCR and FISH. The agents used in screening were trastuzamab, two antibody-drug conjugates (ADCs) consisting of trastuzamab conjugated to an anti-mitotic agent, and a small molecule inhibitor of HER2 kinase activity. HER2 testing revealed 32% of the cell lines to be positive for HER2 by qPCR and FISH, and 90% of these demonstrated a high-level amplification as demonstrated by HER2 amplifications in the form of clusters by FISH and more than 10 copies by qPCR. Several of the cell lines showed polysomy for chromosome 17 and extra copies of HER2 by FISH assay but did not meet the criteria for HER2 amplification. The HER2 IHC assay showed 10 cell lines to be positive (2+, 3+) by IHC, with 8 illustrating high-level expression (3+). The agents tested were more efficacious in the HER2-amplified cell lines, with the broadest activity observed in response to the ADCs. Cell lines exhibiting polysomy for HER2 and 1+ expression by IHC were less responsive to any of the agents tested. Consistent with previous reports, many of the cell lines resistant to trastuzamab or the small molecule HER2 inhibitor demonstrated deregulation of the PI3K pathway illustrated by mutations in the PIK3CA gene or loss of PTEN protein expression. Interestingly, the majority of these resistant cell lines were sensitive to ADCs. In summary, our study illustrates a high sensitivity of ADCs in breast cancer cell lines harboring a high-level amplification of the HER2 gene and suggests these agents may overcome in vitro resistance provided by activation of the PI3K pathway.

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 2589.
Abstract 259: Functional analysis of the β1 subunit of AMP-activated protein kinase in ovarian cancer
Authors: Li Cuilan , Liu Vincent W. S. , Chan David W. , Ngan Hextan Y. S.
Published: February 2011

AMP-activated protein kinase (AMPK) is a heterotrimer composed of a catalytic subunit (α) and two regulatory subunits (β and γ). Each subunit has different isoforms, namely, α1, α2, β1, β2, γ1, γ2 and γ3 encoded by distinct genes, which enable the yielding of 12 possible heterotrimeric complexes. Originally known as a cellular energy-balancing sensor, AMPK was recently found to have strong tumour suppression function. Roles of AMPK have recently been extensively studied in metabolic diseases and human cancers. However, the functional aspects of individual subunits of AMPK are barely investigated. Using quantitative RT-PCR, we found that the AMPK-β1 subunit was over-expressed in ovarian cancer as compared with normal ovary. Over-expression of the AMPK-β1 was further confirmed by immunohistochemical staining (IHC) of the paraffin-embedded ovarian tissues. Clinicopathological data analysis indicates that over-expression of AMPK-β1 correlates with low-grade tumour. Furthermore, from the IHC, the geographical location of the AMPK-β1 in ovarian cancer tissues was also known. In addition to its appearance in the cytoplasm, the AMPK-β1 is also presence in the cell membrane. Since the active form of AMPK (phospho-AMPK) only locates at the cytoplasm, we postulate that AMPK-β1 has addition functions other than just component of the AMPK molecule. To determine the role of AMPK-β1 in ovarian cancer, we have carried out functional analysis of the expression of AMPK-β1 in ovarian cancer. Interestingly, AMPK-β1 was found to inhibit cell growth when forced expressed in ovarian cancer cells. Enforced expression of AMPK-β1 can activate AMPK or down-regulate the level of phospho-AKT in ovarian cancer cells. Additional assays are being studied to further determine the role of AMPK-β1 in ovarian cancer. Understanding the functional role of AMPK-β1 may facilitate the design and development of a new strategy for therapeutic intervention.

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 259.
Abstract 2590: Preclinical evaluation of ASG-5ME, a novel antibody-drug conjugate for pancreatic cancer
Authors: Smith Leia M. , Gudas Jean M. , Lewis Tim , Duniho Steve , An Zili , Morrison Kendall , Jia Xia C. , Raitano Art B. , Moser Ruth , Lyon Robert P. , Hayes Dawn , Senter Peter D. , Jakobovits Aya , Benjamin Dennis R.
Published: February 2011

Antibody-drug conjugates (ADCs) have demonstrated promising activity and tolerability profiles in oncology clinical trials, including for Hodgkin lymphoma (brentuzumab vedotin) and breast cancer (trastuzumab-DM1), motivating the development of new ADCs targeting antigens in other prevalent cancers. An immunohistochemical survey of expression of the novel tumor antigen AGS-5 revealed strong staining in 90% of pancreatic cancer specimens tested. Importantly, this staining was largely uniform, with essentially all transformed cells demonstrating membranous staining. This observation along with the observation of equally abundant expression of AGS-5 in prostate and other cancers motivated the development of ASG-5ME, an ADC targeting AGS-5. The ADC is comprised of a fully human IgG2 monoclonal antibody conjugated to the potent tubulin-binding drug monomethylauristatin E, conjugated with an average of 3.7 drug molecules per antibody. Conjugation and analysis of this IgG2 ADC will be discussed. The resulting conjugate demonstrated antigen-dependent internalization and in vitro cytotoxicity at sub-saturating ADC concentrations, as well as potent in vivo antitumor activity in patient-derived xenograft models of pancreatic cancer. ASG-5ME has a long (12 day) T1/2 in mice. Taken together, these results support the clinical evaluation of ASG-5ME for treatment of pancreatic cancer

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 2590.
Abstract 2591: Differential oncolysis by vesicular stomatitis virus in prostate cancers arising in transgenic mice
Authors: Moussavi Maryam , Fazli Ladan , Tearle Howard , Cox Michael E. , Bell John C. , Ong Christopher , Jia William , Rennie Paul S.
Published: February 2011

Prostate cancer (PCa) is the most prevalent malignancy and the second leading cause of cancer related mortality in North American men. Survival prospects for patients with locally advanced or metastatic prostate cancer are low and the side effects associated with current therapies seriously compromise quality of life in PCa patients. Oncolytic viral therapy provides an opportunity to efficiently target and kill primary and metastatic cancer cells while sparing normal cells. Vesicular Stomatitis Virus (VSV) is an oncolytic virus which can infect and kill cells that have defects in their cellular anti-viral immunity such as the interferon (IFN) response pathway. In this study VSV (AV3) strain with enhanced IFN sensitivity was injected intra-prostatically into two different transgenic mouse models (prostate specific PTEN null mice and transgenic adenocarcinoma mice [TRAMP]) and monitored for tissue bioluminescence over 96 h. While the virus readily dispersed throughout the body of each group after only 3 h, it persisted >72 h in prostates of tumor bearing mice compared to low levels ∼48 h in control mice. Plaque assay provided a similar pattern, with much higher concentrations of replicating virus in tumor bearing transgenic mice than in control. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) analysis of paraffin embedded tissues demonstrated that VSV(AV3) is capable of selectively infecting and killing malignant prostate cells while sparing normal cells, specifically at the 48h time point. This cell death is due to disrupted IFN response in the PTEN null prostates. Moreover, > 60 % of TRAMP mice treated with VSV(AV3) did not present any metastasis while approximately 70% of TRAMP mice treated with inactivated virus presented metastatic lesions either in lymph nodes or lungs. In summary, our data demonstrates that VSV(AV3) may be used as a potential oncolytic viral therapy to target prostate cancer.

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 2591.
Abstract 2626: Phospho-Akt is the molecular determinant of bortezomib sensitivity to head and neck squamous cells
Authors: Yu-Chin Lin , Chen Kuen-Feng , Chen Kui-Chiu , Chen Ching-Chow , Cheng Ann-Lii
Published: February 2011

Introduction: Head and neck squamous cell carcinoma (HNSCC) is a worldwide prevalent cancer with aggressive clinical courses and dismal outcomes. Incorporation of cetuximab, an antibody against epidermal growth factor receptor, had led to advance of treatment, but the efficacy is modest. Therefore, new treatment for HNSCC is needed. Bortezomib, a proteasome inhibitor approved for treatment of myeloma and lymphoma, demonstrated activities for various solid tumors. Here, we report the differential sensitivity of bortezomib for HNSCC cells and identify the major molecular determinant responsible for activity. Material and method: HNSCC cells, including Ca9-22, SAS, SCC-25 and FaDu, were treated with bortezomib and evaluated for viability, apoptosis, and signal transduction. MTT assay was used for viability. Flow cytometry and Western blotting were performed for apoptosis and signal transduction analysis. Gene silencing was done by small interference RNA (siRNA). Results: Four HNSCC cells showed differential sensitivities to bortezomib. Thorough comparison of molecular change before and after treatment of bortezomib, we found that Akt played a significant role in mediating bortezomib sensitivity to HNSCC cells. Our data demonstrated that bortezomib down-regulated phospho-Akt (Ser473) in dose- and time-dependent manner, and significantly correlated with apoptosis. Knocking down Akt1 by small interference RNA enhanced sensitivity of bortezomib whereas ectopic expression of Akt conferred resistance to bortezomib, suggesting that Akt is indispensible in mediating bortezomib's effect on HNSCC cells. Conclusion: Down-regulation of phospho-Akt represents a molecular determinant of bortezomib-induced apoptosis in HNSCC cells and may be a biomarker for predicting clinical response to HNSCC treatment. Targeting Akt signaling enhances sensitivity to bortezomib in HNSCC cells, which might provide a new approach for the combinational therapy of HNSCC.

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 2626.
Abstract 2628: BU32: A novel proteasome inhibitor for treatment of multiple myeloma and other types of cancer
Authors: ROY SUDIPA SAHA , KIRMA NAMEER , NAIR HAREESH , TEKMAL RAJESHWAR RAO , AGYIN JOSEPH
Published: February 2011

Background: Multiple myeloma (MM) is an incurable neoplasm characterized by devastating and progressive bone destruction. Standard chemotherapeutic agents have not been effective at significantly prolonging the survival of MM patients and these agents are typically associated with often severe, dose-limiting side effects. Proteasome inhibition provides an attractive approach to cancer therapy and may have application in the treatment of MM and also other types of cancers. However, results of recent clinical trials to evaluate the effect of the proteasome inhibitor bortezomib (Velcade®, also called PS-341) in MM patients have shown limited activity when used as a single agent. This underscores the need to find new and more efficacious proteasome inhibitors for treatment of MM. We have recently synthesized a novel proteasome inhibitor BU-32 and tested its growth inhibitory effects in different MM cells including RPMI8226 (human) and 5TGM1 (mouse) and other types of cancer cells.

Methods and Results: In this study, we evaluate the efficacy of the novel proteasome inhibitor BU-32 (NSC D750499) using in vitro and in vivo MM models. BU-32 exhibits strong cytotoxicity in a panel of MM cell lines - RPMI8226 and 5TGM1. Furthermore, proteasome inhibition assay reveals that BU-32 potently inhibits the chymotryptic- and caspase-like activities of the 26S proteasome. Results from our studies using real-time PCR array analyses show that BU-32 effectively down-regulates an array of angiogenesis and inflammatory markers. We show from Annexin V FITC binding studies that BU-32, like bortezomib, induces apoptosis in different MM cell lines but the effect is more prominent in case of BU-32 treated cells. Studies using an in vivo MM model show significant effect and marked reduction in osteolytic bone lesions in radiography with 0.3 mg/kg body weight dose of BU-32. Further testing of BU-32 on pancreatic cancer and various hormone regulated cancers like ovarian and prostate are currently under way. Preliminary cell viability studies on these cancer cell lines indicate BU-32 is highly effective against other cancer cells also. Annexin V FITC binding analyses show that BU-32 drug induces apoptosis to these cell lines. These studies elucidate that BU-32 is a most promising candidate drug for clinical trial as a novel treatment modality for MM and other types of cancer.

Conclusion: Our results suggest that BU-32 might be a potential chemotherapeutic agent with promising antitumor activity for the treatment of MM and other types of cancer.

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 2628.