bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5431: Histone deacetylase inhibitor (HDAC) panobinostat (LBH589) enhances the antiproliferative effect of a topoisomerase II (topo II) inhibitor in doxorubicin-resistant HL-60 cells, despite high MDR expression
Authors: Brady Anna K. , Advani Anjali S. , Grabowski Dale , Ganapathi Ram , Ganapathi Mahrukh K.
Published: February 2011

Acute myeloid leukemia (AML) is difficult to treat, particularly in the relapsed or refractory setting due to drug resistance. Topo II inhibitors are effectively used to treat AML, and histone deacetylase (HDAC) inhibitors have been used pre-clinically and clinically with some success. We previously reported that vorinostat (SAHA) was effective in combination with topo II inhibitors in AML cells (Proc. AACR, 2009, Abstract 4567). In the present study we evaluate LBH589, a novel class I/II HDAC inhibitor. Since resistance to HDAC inhibitors has not been well described, we hypothesized that LBH589 would be effective in combination with a topo II inhibitor in a doxorubicin-resistant AML cell line that expresses MDR and resistant by MDR-independent mechanism. A sensitive HL60 parent line (S) and the doxorubicin-resistant sub-line (R) were used. Early studies revealed that vorinostat (SAHA) had comparable anti-proliferative effects in S & R cells but the R cells were relatively more resistant to LBH589. This resistance was likely MDR-mediated. We then tested whether sub-lethal doses of SAHA or LBH589, used in combination with sub-lethal doses of etoposide (VP-16), would be effective. S and R cells were treated with VP-16 for 1 h, washed, and re-incubated with SAHA or LBH, or drug-free media, for a total of 144 hours. Cell viability and apoptosis were measured at 72 h and 144 h following treatment. At 72 h and at 144 h, the combination of VP-16 and LBH was significantly more effective (p<0.01) in reducing proliferation in HL60/S and HL60/R cells, compared to either agent alone. At both 72 h and 144 h the combination of VP-16 and LBH also led to significantly (p<0.05) greater apoptosis in both S & R cells. Western blotting for phospho-histone H2AX, a marker of DNA damage, showed increased DNA damage in the R cells treated with VP-16 and LBH589 compared to either drug alone. Overall, the effects of LBH589 in combination with VP-16 were significantly (p<0.05) more effective than SAHA in the S & R cells. In summary, our results demonstrate that MDR may mediate resistance to LBH589. Further, in spite of MDR expression, LBH589 is a useful adjunct to VP-16 treatment and enhances the anti-proliferative effect of the topo II inhibitor via increased apoptosis, even in the doxorubicin-resistant cells. Thus, post-treatment with LBH589 in combination with a topo II inhibitor may be useful treatment strategy in relapsed / refractory AML.

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 5431.
Abstract 5433: Prolonged histone hyperacetylation with a novel class of HDAC1/2 selective inhibitors
Authors: Methot Joey L. , Chenard Melissa , Joshua Close , Jonathan Cruz , Dahlberg William , Fleming Judith , Hamblett Chris , Julie Hamill , Harrington Paul , Harsch Andreas , Heidebrecht Richard , Hughes Bethany , Jung Joon , Kenific Candia , Kral Astrid , Middleton Richard , Ozerova Nicole , Sloman David , Stanton Matthew , Szewczak Alexander , Wilson Kevin , Witter David , Secrist Paul , Miller Thomas
Published: February 2011

The histone deacetylase (HDAC) metalloenzymes are intricately involved in gene expression through epigenetic regulation of histone acetylation. They also regulate the acetylation status of numerous non-histone proteins such as transcription factors p53, STAT1 and NF-κB as well as α-tubulin, Hsp90 and Ku70. Of the eleven zinc-dependent HDAC enzymes identified, HDACs 1 and 2 appear to be most critical in oncogenesis and tumor maintenance. They are overexpressed in many human cancers and RNAi knockdown leads to increased apoptosis.

We recently disclosed a family of novel HDAC1/HDAC2-selective biaryl inhibitors. In this presentation, we will describe unique features of these biaryl inhibitors that contribute to improved preclinical efficacy and tolerability. Desirable HDAC inhibitor properties identified from preclinical experience with Zolinza™ include subtype selectivity toward HDAC1/HDAC2 and prolonged target inhibition. Compelling in vitro and in vivo data indicates that solid tumor cell lines are most sensitive to HDAC inhibition under continuous exposure rather than intermittent exposure. These biaryl inhibitors exhibit extended target engagement in vivo, and are well tolerated in nude mice. Unlike other known HDAC inhibitors, these compounds exhibit a delay in and prolongation of histone hyperacetylation in nude mice bearing HCT116 tumors, extending beyond plasma clearance of the drug.

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 5433.
Abstract 5434: Development and biological characterization of novel, potent and stable isoform-specific inhibitors of histone deacetylase-8 (HDAC8)
Authors: Balasubramanian Sriram , Sirisawad Mint , Verner Erik , Mani Chitra , Jejurkar Purvi , Buggy Joseph J.
Published: February 2011

Histone deacetylase (HDAC) inhibitors have shown promise for the treatment of hematological and solid tumors. The current clinical candidates all inhibit multiple isoforms of the 11-member zinc-dependent HDAC family. With a view to increasing disease specificity and decreasing toxicity, we have developed potent and selective inhibitors of one isoform, HDAC8. We have previously described one such molecule, PCI-34051, which inhibited HDAC8 with a Ki of 10 nM and greater than 200-fold selectivity over other HDAC isoforms. However, this compound displayed poor metabolic stability and was not suitable for in vivo testing in animal models. We have therefore developed a series of analogs to increase metabolic stability while maintaining potency and isoform selectivity. These compounds, which are based on different pharmacophores, have similar potency and selectivity to PCI-34051 but have increased cellular potency, inhibiting cell growth and inducing apoptosis with EC50 of ∼1-2 μM in Jurkat and Hut78 lymphoma cells. These compounds also showed increased metabolic stability in an in-vitro rat liver microsomal assay. Since increased HDAC8 expression has recently been shown to be correlated with poor survival in neuroblastoma, the biological activities of these novel HDAC8 inhibitors were tested in neuroblastoma cell lines. Interestingly, these compounds were found to inhibit cell growth and induce differentiation in neuroblastoma cells. However, incubation with higher concentrations of these compounds did induce caspase activation and apoptosis in these cells. In keeping with the HDAC8 selectivity, no tubulin or histone acetylation was observed at upto 25μM. The best of these inhibitors is currently being tested in mouse xenograft models of T-cell lymphoma as well as neuroblastoma and the results will be presented.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5434.
Abstract 5437: The histone deacetylase (HDAC) inhibitor SNDX-275 enhances the antitumor effect of interleukin 2 (IL-2) by suppressing regulatory T cell gene expression and function
Authors: Shen Li , Pili Roberto
Published: February 2011

In our previous studies we have reported that the combination of IL-2 and the HDAC inhibitor SNDX-275 had an in vivo synergistic antitumor effect in an immuno-competent murine model of renal cell carcinoma (RENCA). The antitumor effect was associated with a decreased number of T regulatory cells (Tregs). In this study we observed that SNDX-275 treatment affects Tregs function by modulating the transcriptional factor Foxp3, a key regulator of Treg function. We assessed the levels of CD4+Foxp3+ cells (Tregs) by fluorescence-activated cell sorting analysis (FACS) and evaluated mRNA level of Foxp3 by QRT-PCR. Both in vitro and in vivo treatment of SNDX-275 reduced Foxp3 protein level and mRNA level in Tregs. Moreover, in the RENCA tumor mouse model, by testing peripheral blood lymphocytes, we found that the antitumor activity of SNDX-275 was associated with reduced Foxp3 protein level in Treg cells. In addition, Tregs were purified from splenocytes of SNDX-275 treated or vehicle-treated mice. Regulatory T cell functional assay showed that Tregs from SNDX-275 (5mg/kg) treated mice had a significantly reduced inhibitory activity on proliferating T effector cells as compared to Tregs from vehicle-treated mice. While 20mg/kg dose of SNDX-275 had a dramatic inhibitory effect on T effector cells, lower dose SNDX-275 (5mg/kg) had either none or moderate negative effect on T effectors. Taken together, these results indicate that SNDX-275 inhibits Foxp3 gene and protein expression, which leads to inhibition of Tregs function, in a dose dependent fashion. Tregs-suppressing features of the HDAC inhibitor SNDX-275 provides the rationale to include this agent in combinational strategies involving immuno-modulatory therapies such as IL-2 to further improve antitumor immune response.

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 5437.
Abstract 5444: Polyamine-based HAT inhibitors as therapy sensitizers
Authors: Bandyopadhyay Keya , Banères Jean-Louis , Martin Aimée , Blonski Casimir , Parello Joseph , Gjerset Ruth A.
Published: February 2011

Histone acetylation plays an important role in regulating chromatin structure and is tightly controlled through the actions of histone acetyltransferases (HAT) and histone deacetylases (HDAC). Deregulation of either HAT or HDAC activity can affect chromatin remodeling needed to carry out its functions and contribute to cancer. While HATs are potential targets for anticancer therapeutics, the development of effective HAT inhibitors has been slow. We have examined the cellular effects of an S-substituted coenzyme A (CoA) inhibitor of histone acetylation, consisting of spermidine (Spd) linked to the S-terminus of CoA through a thioglycolic acid linkage (adduct abbreviated as Spd-CoA), as well as the effects of a truncated Spd-CoA derivative lacking the negatively charged portion of the CoA moiety. While exposure of cancer cells to Spd-CoA has little effect on cell viability, it causes a rapid inhibition of specific acetylated lysines, H3-K9 and H3-K56. That inhibition correlates with a transient arrest of DNA synthesis, a transient delay in S-phase progression, and an inhibition of nucleotide excision repair and DNA double strand break repair. Furthermore, Spd-CoA synergizes with a variety of commonly used DNA damaging chemotherapeutic agents, cisplatin (PlatinolTM) and 5-fluorouracil, camptothecin, as well as UV-C radiation to induce cancer cell killing. This suggests that a common mechanism, relevant to DNA damage, underlies the ability of histone acetylation inhibition to synergize with drugs and radiation. Normal human fibroblasts and epithelial cells are not sensitized to DNA damage by Spd-CoA due to a barrier to uptake, indicating that this differential uptake can be exploited to achieve cancer cell-specific sensitization. Furthermore, therapy sensitization occurs in both p53-null cancer cells and in cancer cells expressing wild-type p53, indicating that p53-mediated apoptosis is not required. The truncated Spd-CoA derivative displays similar but enhanced chemosensitization effects, suggesting that this class of inhibitors may be amenable to further refinement and have considerable clinical potential as a novel class of potent therapy sensitizers applicable to a broad range of conventional cancer treatments, particularly to reduce therapy toxicity and reverse therapy resistance.

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 5444.
Abstract 5449: Histone deacetylase inhibitor treatment in B-cell non-Hodgkin's lymphoma - effect of ex vivo treatment on primary patient samples
Authors: Nielsen Torsten H. , Petruccelli Luca A. , Dobocan Monica C. , Petterson Filippa , Haliotis Tina P. , Assouline Sarit , Mann Koren K. , Miller Wilson H.
Published: February 2011

The use of histone deacetylase inhibitors (HDACis) is approved in cutaneous T-cell lymphoma, but the efficacy in other lymphomas, particularly B-cell Non-Hodgkin's lymphoma (B-NHL), remains to be established. To investigate the hypothesis that HDACis have anti-neoplastic effects in B-NHL, we have tested a series of primary B-NHL patient samples for response to the two HDACis Vorinostat (SAHA) and Panobinostat (LBH589).

Lymph node biopsies from patients with B-NHL were obtained with patient consent. A single-cell suspension was prepared and mononuclear cells isolated by Ficoll gradient separation. Lymphocytes were treated with clinically achievable doses of SAHA (2.5 μM, 5 μM and 7 μM) and LBH589 (10 nM, 25 nM and 50 nM) ex vivo for 48 hours. Treatment effect was evaluated by assays of cell viability (trypan blue staining and Cell TiterGlo luminescence assay) and cell death (propidium iodide staining analyzed by flow cytometric quantification of the sub-G0 cell population). Expression of acetylated histone H4, Bcl-2 and BCL6 was evaluated by western blot.

To date, 16 patient samples, covering a wide variety of B-NHL, have been analyzed. Diagnoses include: follicular lymphoma (n=7), diffuse large B-cell lymphoma (n=5), mantle cell lymphoma (n=1), nodal marginal zone lymphoma (n=1), hairy cell leukemia (n=1) and small lymphocytic lymphoma/chronic lymphocytic leukemia (n=1). Fifteen out of 16 patient samples analyzed showed moderate to high sensitivity to one or both HDACis tested at the given concentrations. Overall, the effects of SAHA and LBH589 were comparable. Western blots of lymphocytes treated with both types of HDACi showed increased acetylation of histone H4 compared to untreated cells. No change in Bcl-2 expression was seen (n=2) and no expression of BCL6 was detected in the samples tested for this (n=2).

Based on the data outlined herein, B-NHL is sensitive to the growth inhibitory effects of HDACis at clinically achievable doses. Building upon this pilot study, we are developing cell line models of B-NHL in order to characterize the mechanism of action of HDACis in B-NHL, including combination therapy with currently used agents such as Rituximab. A phase 2 clinical trial of LBH589 in diffuse large B-cell lymphoma, including an arm evaluating combination therapy with Rituximab, is also 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 5449.
Abstract 5453: Inhibition of lactate dehydrogenase A induces oxidative stress and inhibits tumor progression
Authors: Le Anne , Cooper Charles R. , Gouw Arvin , Maitra Anirban , Dinavahi Ramani , Deck Lorraine , Royer Robert , Jagt David Vander , Semenza Gregg , Dang Chi Van
Published: February 2011

As the result of genetic alterations and tumor hypoxia, many cancer cells avidly take up glucose and generate lactate through lactate dehydrogenase A (LDHA), which is encoded by a target gene of c-Myc and HIF-1. Previous studies with reduction of LDHA expression indicate that LDHA is involved in tumor initiation, but its role in tumor maintenance and progression has not been established. Furthermore, how reduction of LDHA expression by interference or antisense RNA inhibits tumorigenesis is not well understood. Here, we report that reduction of LDHA by siRNA or its inhibition by a small molecule inhibitor (FX11) reduced ATP levels and induced significant oxidative stress and cell death that could be partially reversed by the antioxidant N-acetylcysteine. Furthermore, we document that FX11 inhibited the progression of sizable human lymphoma and pancreatic cancer xenografts. When used in combination with the NAD+ synthesis inhibitor FK866, FX11 induced lymphoma regression. Hence, inhibition of LDHA with FX11 is an achievable and tolerable treatment for LDHA-dependent tumors. Our studies document a therapeutic approach to the Warburg effect and demonstrate that oxidative stress and metabolic phenotyping of cancers are critical aspects of cancer biology to consider for the therapeutic targeting of cancer energy metabolism.

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 5453.
Abstract 5454: Novel STAT3:STAT3 small-molecule inhibitors as potential anticancer agents
Authors: Haque Mohammad R. , Zinzalla Giovanna , Kaye Samantha L. , Haider Shozeb , Basu B. Piku , Anderson John , Rahman Khondaker M. , Antonow Dyeison , Essex Samantha , Palmer Jonathan , Nkansah Edwin , Hasan Fyeza , Morgenstern Daniel , Wilderspin Andrew F. , Neidle Stephen , Thurston David E.
Published: February 2011

Transcription factors are important targets for cancer therapy. The inhibition of protein-protein interactions (PPIs) within signalling pathways known to be key regulators of transcriptional activity is a viable approach to novel chemotherapeutic strategies. Proof-of-concept studies in cell-culture and animal models have validated the potential of small-molecule inhibitors of STAT3 signalling in cancer therapy. In particular, the protein-protein interaction between two STAT3 monomers (i.e., the dimerisation event in the signalling cascade) has been identified as a valid target to inhibit DNA-binding and the resultant transcriptional activation. Of the approximately 20 small-molecule STAT3 inhibitors reported in the literature to date, only three are described as potential STAT3:STAT3 dimerisation inhibitors, and these have IC50 values in STAT3-expressing cell lines of between 10-90 μM.

Using in silico and medicinal chemistry-based approaches based on a published X-Ray structure of STAT3 (PDB: ID-1BG1) to identify “hit” inhibitors, a focussed library (approx. 50 members) was designed around one such “hit” and synthesized employing an efficient 4-step linear approach. Library members were then entered into a screening cascade involving initial evaluation of their ability to inhibit STAT3:STAT3 interaction in a Fluorescent Polarisation (FP)-based primary PPI binding assay. “Hit” molecules from this primary assay were then studied in two cell-based assays designed to test for STAT3 selectivity. The first was a comparative MTS assay between STAT3-expressing MDA MB231 breast cancer cells and STAT3-null colon A4 cells. The second was a luciferase reporter assay designed to measure transcription inhibition in STAT3-Luc-transformed Hela cells compared to SV40-Luc-transformed Hela control cells. A potential “lead” molecule, RH06, emerged from this screening cascade with potentially selective STAT3 inhibitory activity in the low micromolar (i.e., ∼1 µM) region. RH06 is currently being studied for its effect on STAT3 and pSTAT3 signal activation, and its potentially differential effect on upstream and downstream mediators (i.e., JAK2, Bcl-xl, Cyclin D1 and pSTAT1).

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 5454.
Abstract 5455: Antitumor activity of death receptor agonists for prostate cancer therapy
Authors: Shankar Sharmila , Marsh Luke , Jackman Chris , Srivastava Rakesh K.
Published: February 2011

Clinical trials on TRAIL and death receptor (TRAIL-R1 and TRAIL-R2) agonist antibodies are being carried out through out the world. Until today, there are no small organic molecules that bind to TRAIL-death receptors, and induce apoptosis mainly in cancer cells. Through molecular and biological approaches, we have identified novel death receptor agonists (small molecular weight organic compounds). Death receptor agonists termed as “DRAs” inhibited cell viability and colony formation, and induced apoptosis in androgen-dependent (LNCaP) and -independent (PC-3) human prostate cancer cell lines, but had no effect on human normal prostate epithelial cells and hepatocytes. The combinations of DRAs with TRAIL had additive effects on apoptosis in prostate cancer cells. Commonly used anticancer drugs further enhanced the apoptosis-inducing potential of DRAs. DRAs recruited FADD and activated caspase-8, and thus formed active TRAIL-DISC (death-inducing signaling complex) similar to that of TRAIL. DRAs inhibited the growth of PC-3 and LNCaP xenografts in nude mice, and prostate cancer growth in TRAMP mice. In vivo, DRAs inhibited tumor cells proliferation (Ki67 staining) and induced caspase-3 activity and apoptosis (TUNEL staining). Pharmacokinetics data demonstrate that DRA is stable, possesses a long terminal half-life, and meats the criteria of clinical trials. Our data suggest, for the first time, that DRAs are specific activators of death receptor pathway of apoptosis, and can be used to treat human prostate cancer. Further studies are underway to assess their clinical significance for the treatment of prostate cancer.

(Funding Support: Department of Defense, US Army)

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 5455.
Abstract 5457: High levels of fatty acid synthase expression in esophageal cancers represent a potential target for therapy
Authors: Orita Hajime , Coulter Jonathan , Tully Ellen , Montgomery Elizabeth , Abe Masaaki , Maekawa Hiroshi , Sato Kouichi , Tsurumaru Masahiko , Hino Okio , Gabrielson Edward
Published: February 2011

Fatty acid synthase (FAS) is overexpressed in many human cancers and is considered to be a promising target for therapy. To investigate the expression of this candidate target in esophageal cancer, we evaluated expression of FAS protein in - cases of esophageal squamous cancer, - cases of esophageal adenocarcinoma, and - cases of Barrett's esophagus with dysplasia - a lesion thought to represent a pre-invasive precursor to esophageal cancer. Using immunohistochemistry, we found significantly higher levels of FAS expression in -% of the squamous cancers and -% of the adenocarcinomas compared to normal esophageal epithelium. To evaluate the potential for inhibiting this enzyme as a treatment of esophageal cancer, we treated mice bearing xenografts of the Colo680N (squamous carcinoma) and Bic1 (adenocarcinoma) cell lines using C93, a rationally designed molecule that inhibits FAS activity. In these experiments, C93 significantly inhibited the growth of orthotopic xenograft tumors from without causing anorexia and weight loss in the treated animals. We conclude that, similar to several other common types of human cancer, FAS is expressed at very high levels in esophageal cancer, and growth of these cancers can be inhibited by pharmacological agents that target this enzyme. Moreover, this high expression of FAS is also seen in pre-invasive lesions of the esophagus, leading us to propose considering of FAS-inhibitors for purposes of esophageal cancer chemoprevention.

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 5457.
Abstract 5458: Clinical and biological significance of CCR9-CCL25 axis in prostate cancer
Authors: Sharma Praveen K. , Singh Rajesh , Grizzle William E. , Singh Shailesh
Published: February 2011

Bone metastasis is major cause of prostate cancer (PCa) associated deaths and handful treatment alternatives are available to treat this condition. Therefore, better understating of metastatic process and identification of molecular signature on metastatic cells are urgently needed to develop therapeutic for metastatic PCa. We have shown that CCR9, a CC chemokine receptor is highly expressed by the PCa tissue and cells and play an important role in regulating the metastatic process under the influence of its natural ligand CCL25. CCL25 is also known as a thymus-expressed chemokine (TECK) and primarily expressed by the thymus, which involutes with age. Interestingly, we have observed higher serum CCL25 in PCa patients in comparison to normal healthy donors. For the first time, we show secretion of CCL25 within the primary prostate cancer tissue in a paracrine manner. Interestingly, CCL25 was highly produced by the bone marrow cells isolated from tumor bearing mice in comparison to non-tumor bearing mice. Furthermore, inhibition of CCR9-CCL25 interaction in vivo using CCL25 neutralization antibodies, showed significant inhibition of bone metastasis after intra-cardiac injection of PC3 cells and tumor establishment after intra-tibial injection. These results provide clinical and biological significance of CCR9-CCL25 axis in prostate cancer progression and metastasis. We have also demonstrated that CCR9-CCL25 axis plays an important role in lowering the chemotherapeutic efficacy by supporting anti-apoptotic and survival signals in prostate cancer cells. Interestingly, significant inhibition in tumor burden was observed, when etoposide was given in combination CCL25 neutralization antibody in comparison to both as a single agent. Our data show strong clinical and biological significance of CCR9-CCL25 axis in prostate cancer. Furthermore, these results also show that activation of CCR9-CCL25 axis may be the possible cause behind low apoptosis rate in prostate cancer cells and could be a possible cause for unsatisfactory chemotherapeutic outcomes. Therefore, developing CCR9-CCL25 directed therapies, may improve the therapeutic outcome of prostate cancer either as a single agent or in combination with other therapeutic 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 5458.
Abstract 5460: Identification of c-Src as a potential therapeutic target for gastric cancer and of MET activation as a cause of resistance to c-Src inhibition
Authors: Okamoto Wataru , Okamoto Isamu , Yoshida Takeshi , Hatashita Erina , Yamada Yuki , Kuwata Kiyoko , Arao Tokuzo , Yanagihara Kazuyoshi , Nishio Kazuto , Fukuoka Masahiro , Nakagawa Kazuhiko
Published: February 2011

Therapeutic strategies that target c-Src hold promise for a wide variety of cancers. We have now investigated both the effects of dasatinib, which inhibits the activity of c-Src and several other kinases, on cell growth as well as the mechanism of dasatinib resistance in human gastric cancer cell lines. Immunoblot analysis revealed activation of c-Src at various levels in most gastric cancer cell lines examined. Dasatinib inhibited phosphorylation of extracellular signal-regulated kinase (ERK) and induced G1 arrest, as revealed by flow cytometry, in a subset of responsive cell lines. In other responsive cell lines, dasatinib inhibited both ERK and AKT phosphorylation and induced apoptosis, as revealed by an increase in caspase-3 activity and cleavage of poly(ADP-ribose) polymerase. Depletion of c-Src by RNA interference also induced G1 arrest or apoptosis in dasatinib-responsive cell lines, indicating that the antiproliferative effect of dasatinib is attributable to c-Src inhibition. Gastric cancer cell lines positive for activation of MET were resistant to dasatinib. Dasatinib had no effect on ERK or AKT signaling, whereas the MET inhibitor PHA-665752 induced apoptosis, in these cells. The subsets of gastric cancer cells defined by a response to c-Src or MET inhibitors were distinct and nonoverlapping. Our results suggest that c-Src is a promising target for treatment of gastric cancer, and that analysis of MET amplification might optimize patient selection for treatment with c-Src 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 5460.
Abstract 5468: JNK-1 inhibition leads to antitumor activity in ovarian cancer
Authors: Rodriguez-Aguayo Cristian , Vivas-Mejia Pablo , Benito Juliana Maria , Fernandez Ariel , Francois-Xavier Claret , Chavez-Reyes Arturo , Sood Anil K. , Lopez-Berestein Gabriel
Published: February 2011

Recent findings suggest that JNK mediates oncogenic functions in several cancer types including head and neck, gastric, and breast cancers, and melanoma, thus it may be an attractive therapeutic target. Specifically, JNK1 but not JNK2 has been shown to be important in promoting cell survival by controlling cell cycle arrest and apoptosis. Little is known about the role of JNK1 in ovarian cancer growth. Therefore, we examined the potential of JNK1 as a therapeutic target in the current study. We utilized a novel small molecule inhibitor, WBZ_4, which was designed, synthesized, and characterized by our team to curb the potential for side effects of the original compound, imatinib. WBZ_4 was designed to target both c-Kit and JNK1 while avoiding the Abl-kinase. The redesign process was based on comparing the residence time of water molecules that solvate the interfacial aligned residues across known targets of imatinib. Non-conserved sites with low residence time that had a higher propensity to dehydrate (de-wetting hot spots) were used to build blueprints for each target. By adding a methyl group to the original imatinib, we were able to attack a de-wetting hot spot present in C-Kit and JNK1 but absent from BCR-Abl, thereby increasing the specificity of the drug towards the desired targets. Here, we report on the biological effects of JNK-1 inhibition, both in vitro and in vivo using experimental models of ovarian cancer. We found a significant association of pJNK with progression free survival in the 116 epithelial ovarian cancers obtained at primary debulking therapy. WBZ_4 led to cell growth inhibition and increased apoptosis in a dose dependent fashion in four ovarian cancer cell lines. In vivo, while imatinib had no effect on tumor growth, WBZ_4 inhibited tumor growth in orthotopic murine models of ovarian cancer. The anti-tumor effect was further increased in combination with docetaxel. Silencing of JNK-1 with systemically administered siRNA led to significantly reduced tumor weights as compared to non-silencing siRNA controls, indicating that indeed the antitumor effects observed were due to JNK-1 inhibition. These studies identify JNK-1 as an attractive therapeutic target in ovarian carcinoma and that the re-designed WBZ_4 compound should be considered for further clinical development.

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 5468.
Abstract 547: Changes in proteomic and epigenetic expression patterns in tumour associated fibroblasts (TAF) by interaction with urinary bladder carcinoma cells
Authors: Enkelmann Astrid , Heinzelmann Joana , Escher Niko , Walter Martina , Weidig Michaela , Wunderlich Heiko , Junker Kerstin
Published: February 2011

Background: Tumour development and progression is strongly affected by interaction of tumour cells and tumour stroma. For different tumour models (e.g. breast cancer) a supportive effect of TAF on the tumour genesis was demonstrated. Purpose of the present work is the isolation and characterisation of TAF from primary urinary bladder tumour specimen. A further part of this study will deal with the influence of urinary bladder carcinoma cell lines on protein expression of TAF.

Material and Methods: TAF were isolated from cultured urinary bladder tumour specimen. Therefore, primary tumour material was treated with EDTA followed by differential trypsinisation. Non-tumour fibroblasts were isolated from foreskin and normal urinary bladder tissue. Analyses of protein patterns were carried out on cultivated fibroblasts by SELDI-TOF-MS. TAF and foreskin fibroblasts were co-cultivated with urinary bladder cancer cell lines in separated cell culture compartments followed by SELDI-TOF-MS analyses. Furthermore total RNA was isolated from TAF and non-tumour fibroblasts to analyse the miRNA expression profile by miRNA Microarray.

Results: By optimizing cell culture routines it was possible to isolate and subsequently cultivate TAF from primary tumour material of the urinary bladder. SELDI-TOF-MS measurements reveal differences in the proteomic patterns of TAF and non-tumour fibroblasts. Microarray analyses indicate different expression levels of several miRNAs in TAF and non-tumour fibroblasts. Co-cultivation of urinary bladder carcinoma cells and TAF or non-tumour fibroblasts induces modified protein patterns in the different cell types.

Conclusion: TAF can be isolated and cultivated separately from primary tumour material. TAF are characterised by the expression of a specific protein pattern and miRNA profile in comparison to non-tumour fibroblasts. Co-cultivation with tumour cells revealed the induction of a modified expression profile in fibroblasts and vice versa. The present results will provide a more detailed understanding of the function of TAF in the tumour development of urinary bladder carcinoma.

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 547.
Abstract 5470: Safe and targeted anticancer therapeutic efficacy of a novel antioxidant-conjugated difluoro-diarylidenyl-piperidone (HO-3867) against ovarian cancer
Authors: Kuppusamy Periannan , Selvendiran K. , Tong L. , Bratasz A. , Kuppusamy ML. , Trigg NJ. , Rivera BK. , Kálai T. , Hideg K.
Published: February 2011

The anticancer efficacy of a novel difluorodiarylidenyl piperidone (H-4073) and its N-hydroxypyrroline modification (HO-3867) against ovarian cancer is reported. Studies were performed using established human ovarian cancer cell lines (A2870, A2780cDDP, OV-4, SKOV3, PA-1 and OVCAR3, as well as in a murine xenograft tumor (A2780) model. Both compounds were comparably and significantly cytotoxic to A2780 cells. However, HO-3867 demonstrated a preferential toxicity towards ovarian cancer cells, while sparing healthy cells. HO-3867 induced G2/M cell-cycle arrest in A2780 cells by modulating cell-cycle regulatory molecules p53, p21, p27, cdk2 and cyclin, and promoted apoptosis by caspase-8 and caspase-3 activation. It also caused an increase in the expression of functional Fas/CD95 and decreases in STAT3 (Tyr705) and JAK1 phosphorylation. There was a significant reduction in STAT3 downstream target protein levels including Bcl-xL, Bcl-2, survivin, and vascular endothelial growth factor (VEGF), suggesting that HO-3867 exposure disrupted the JAK/STAT3 signaling pathway. In addition, HO-3867 significantly inhibited the growth of the ovarian tumors in a dosage-dependent manner without any apparent toxicity. We further analyzed of xenografted tumor tissues showed that HO-3867 inhibits pSTAT3 (Tyr705 and Ser727) and Jak1 and increased apoptotic markers of cleaved caspase-3 and PARP. The results clearly showed that HO-3867 exhibited significant cytotoxicity towards ovarian cancer cells by inhibiting the JAK/STAT3-signaling pathway. The study suggested that HO-3867 may be useful as a safe and effective anticancer agent for ovarian cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5470.
Abstract 5472: Class III beta-tubulin in castration resistant human prostate cancer
Authors: Terry Stephane , Ploussard Guillaume , Maillé Pascale , Allory Yves , Nicolaiew Nathalie , Soyeux Pascale , Kheuang Laurence , Culine Stephane , Buttyan Ralph , de la Taille Alexandre , Vacherot Francis
Published: February 2011

BACKGROUND: Class III β-tubulin (βIII-tubulin) is expressed in tissues of neuronal lineage but it is also expressed in several human malignancies including non small cell lung carcinoma, breast and ovarian cancer. Overexpression of βIII-tubulin in these tumors is associated with an unfavorable outcome and resistance to taxane-based therapies. βIII-tubulin deregulation however is still poorly documented in prostate cancer. Previously, we reported that βIII-tubulin expression is increased in prostate cancer (PCa) cells by androgen ablation. Here we sought to better characterize the contribution of βIII-tubulin to castration resistant PCa, the stage of the tumor most responsible for mortality from PCa. METHODS: βIII-tubulin expression was assayed by immunohistochemistry in prostate tumors obtained from patients with castration resistant PCa. Protein expression was scored as null (0), weak (1), moderate (2) and strong (3) and considered significant when the score was 2 or more in more than 10% of cancer cells. Consecutive sections were also immunostained for neuroendocrine biomarkers, neuron specific enolase (NSE) and chromogranin A (CgA). An androgen independent (AI) variant of LNCaP cells (AI-LNCaP) was assessed for the effects of docetaxel treatment on βIII-tubulin expression. Finally, DU145 cells stably overexpressing βIII-tubulin were tested for their sensitivity to docetaxel relative to control vector-transfected cells. RESULTS: 24/40 specimens of castration resistant tumors were found to express significant levels of βIII-tubulin. In 6 out of 22 cases analyzed, regions of the tumor positively co-stained for both βIII tubulin and NSE, however none of 8 evaluated cases showed co-staining for βIII-tubulin and CgA. AI-LNCaP cells treated with 5 to 10 nM docetaxel showed a time dependent increase in βIII-tubulin expression. DU145 cells stably overexpressing βIII-tubulin were found to be more resistant to docetaxel than the parental counterpart with a 2-fold increase in the IC50. CONCLUSION: Our data indicate that βIII-tubulin is expressed in the majority of castration resistant prostate tumors. Although βIII-tubulin immunostaining of prostate tumor cells sometimes coincides with staining for neuroendocrine markers, most βIII-tubulin positive tumor cells examined did not show coincidental staining so increased βIII-tubulin expression may occur independently of NE differentiation. Regardless, our in vitro studies support a role for βIII-tubulin in the emergence of doxetaxel resistance by androgen independent human PCa 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 5472.
Abstract 5475: Dynamin inhibition: A new therapeutic approach for the treatment of cancer
Authors: Joshi Sanket , Gaddipatti Swetha , Gilbert Jayne , Smith Charlotte M. , Gordon Christopher P. , Sakoff Jennette , McCluskey Adam , Robinson Phillip J. , Braithwaite Antony W. , Chircop Megan
Published: February 2011

Cytokinesis is the final stage of cell division and involves ingression of the plasma membrane between segregating chromosomes followed by membrane abscission, resulting in the production of two daughter cells. Abscission involves the formation of a central γ-tubulin midbody ring within the intracellular bridge prior to final separation. The endocytic protein dynamin II (dynII) is required for this abscission step. We propose that dynII inhibition represents a new pharmacological intervention for the treatment of cancer via the induction of cytokinesis failure. Our pilot data in Hela cells shows that inhibition of dynII with myristyl trimethylammonium bromide (MiTMAB)(IC50 = 3.1 ± 0.2uM) and octadecyl trimethylammonium bromide (OcTMAB)(IC50 = 1.9 ± 0.2uM) induces cytokinesis failure. Using immunoflourescence, cell cycle analysis and live cell imaging we have shown that cells exposed to these compounds progress through mitosis with normal timing and complete furrow ingression successfully. However, abscission fails causing the membrane to regress generating binucleated cells. These cells initially accumulate in the G2+M phase of the cell cycle but ultimately undergo apoptotic cell death. In further support of our findings we have shown Chlorpromazine, a registered anti-psychotic known to induce cytokinesis failure, to also inhibit dynII (IC50 = 1.3 ± 0.4uM). Using the MTT growth inhibition assay in a panel of 9 cancer cell lines we have shown MiTMAB, OcTMAB and Chlorpromazine to be potent inhibitors of cell growth (IC50 = 0.6 - 3.9uM, 0.5 - 3.5 uM, and 13 - 32uM, respectively). Multinucleation and cytokinesis failure was also induced in Hela cells depleted of dynII using small interfering RNA (siRNA). These findings highlight dynamin II inhibition as a novel therapy for the treatment 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 5475.
Abstract 5476: gp130 knockdown by short hairpin RNA synergistic effect of bortezomib induced apoptosis in multiple myeloma cells
Authors: Park Juwon , Lee Chansu , Oh Jeong In , Kim Byung-Su , Kwon Jung Hye , Kim Byoung Kook , Ahn Kwang-Sung , Yoon Sung-Soo
Published: February 2011

Although bortezomib, one of targeted anti-cancer drugs, effectively suppresses the growth of multiple myeloma (MM) cells via inactivation of NF-κB, activation of IL-6 mediated cell signaling could be suppressed bortezomib mediated apoptosis. Previously we found that the expression levels of IL-6 and sIL-6R in MM patient's bone marrow serum was higher than control. A set of reports suggested that those levels affect the progression of MM. In addition, bortezomib mediated apoptosis directly associated with the activities of NF-κB in MM cells. However, at present, their precise roles, especially IL-6 mediated cell signaling associating with NF-κB activation, in the progression of MM and therapeutic response to anti-cancer treatment still remain largely unknown. In this study, U266 cells treated with combination of IL-6 and sIL-6R showed the prolongation of the phosphorylation of ERK and STAT3 when compared with IL-6 alone. Transient transfection of gp130 shRNA in MM cells (MM/gp130 shRNA) showed inhibition of IL-6/sIL-6R induced ERK and STAT3 phosphorylation. Next, the effect of IL-6 and sIL-6R in an anti-myeloma agent, bortezomib, was investigated. In cell proliferation assay, U266 and RPMI8226 cells exposed to IL-6 and sIL-6R were decreased induction of apoptosis by bortezomib. In addition, MM/gp130 shRNA cells treated with bortezomib showed significantly inhibited c-Myc and cyclinD1 and increased apoptosis by borteozomib compared with parental cells. In this study indicated that the synergic activation of ERK and STAT3 phosphorylation as the result of IL-6 with sIL-6R could diminished by knockdown of gp130 and its effect could be potentiated the apoptotic efficacy of bortezomib in MM 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 5476.
Abstract 548: Interaction between mammary adenocarcinoma cells and macrophages in a three-dimensional model of tumor spheroids
Authors: Roldan Julieta S. , Najenson Ana C. , Roca Fernanda , Marino Lina , Jasnis Maria A. , Fiszman Gabriel L.
Published: February 2011

Solid tumors contain heterogeneous cellular subpopulation due to a limit supply of oxygen, nutrients and wastes. This microenvironment affects local cell growth, survival and overall therapeutic efficacy. Three-dimensional (3D) cell model reflect this situation and is a good model to study of the interaction with the immune system.

Spheroids of the murine LM3 and human MCF-7 mammary adenocarcinoma cell lines were cultured with RAW 264.7 macrophages and human monocytes isolated from peripheral blood from normal volunteers respectively Spheroids were maintained in culture dishes pre-treated with 1.5% agar. In both tumor models, macrophages/monocytes were added after 10-13 days spheroids growth. Control cultures were maintained as monolayers in normoxic and hypoxic conditions (hypoxic chamber 1% O2 saturated during 4-6h). A growth curve of the spheroids were performed, reaching a peak of >300um (+/-) at day 15. In MCF-7 model, cell viability, monocyte distribution inside spheroids and protein expression were determined by hematoxiline-eosine staining and immunohitochemistry with specific antibodies. Results showed that 3D growth followed a concentric organization of proliferant (Ki67+), quiescent (Ki67-/Bcl-2+) and dead cells (CAS+) from periphery towards the hypoxic core (HIF-1+). In LM3 spheroids an enhanced arginase activity (140,64±2,05 vs. 18,17±1,22, p<0,05) and nitric oxide (NO) levels (10,21±1,15 vs 0,55±0,50, p<0,05) were also detected compared to LM3 cells growing as monolayer. It was also detected a decrease NO levels during spheroids growth (4 days: 38,7±8,3 vs. 13 days: 10,21±1,15). On the other hand, arginase activity and NO levels of spheroids cocultured with macrophages were lower than the spheroids alone (96,41±3,63 vs.317,7048±8,82 and 4,1±1,56 vs. 10,22±1,15; p<0,05 respectively). These results show that arginine metabolic pathways, either by NOS or arginase, are both more activated in spheroids than in tumor cell monolayer and coculture spheroids. In MCF7 and LM3 models, HIF-1alpha and iNOS expression was detected by western blot using hypoxic monolayers as a control.

Culture system in 3D appears to be an ideal model to analyze the role of hypoxia and the mechanisms that modulate interactions between tumor and immune 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 548.