bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5360: Leptin-induced SIRT1 expression in human colon cancer cells: a potential link between obesity and cancer
Authors: Song Na-Young , Surh Young-Joon
Published: February 2011

Recently, several epidemiological studies have shown that obesity is one of the prime factors in the etiology of some malignancies. The molecular mechanism underlying obesity-induced carcinogenesis, however, is not fully clarified yet. Leptin, a representative adipokine secreted from the white adipose tissue, is considered as a potential linker between obesity and cancer. Although leptin has been identified as an anorexic adipokine, obese subjects exhibit unexpectedly higher circulating levels of leptin than non-obese individuals. Increased serum levels of leptin may activate the pro-inflammatory signals and stimulate the tumor growth. In the present study, we have found that treatment with leptin (100 ng/ml) promotes migration and invasion of human colon cancer (HCT-116) cells, suggesting leptin as a molecular link between obesity and cancer. Interestingly, leptin induced expression as well as activity of SIRT1, a NAD+-dependent class III histone deacetylase, recognized as an important metabolic regulator. It has been reported that SIRT1 exerts an anti-aging effect. Paradoxically, SIRT1 gene is highly expressed in certain types of tumors, and is speculated to function as an oncogene. We found that leptin-induced HCT-116 cell migration and invasion were blocked by treatment with sirtinol (25 μM), an inhibitor of SIRT1. NF-E2-related factor 2 (Nrf2) is a redox-sensitive transcription factor responsible for regulating expression of a battery of stress-responsive genes. Considering that obesity is a state of chronic oxidative stress, Nrf2 might be involved in leptin-induced SIRT1 up-regulation. In line with this notion, several consensus antioxidant response elements (AREs) specified for Nrf2 binding are found within a promoter region of SIRT1. Treatment of HCT-116 cells with leptin provoked nuclear translocation and ARE binding of Nrf2. Moreover, siRNA knockdown of Nrf2 abrogated leptin-induced SIRT1 expression, lending further support to the notion that Nrf2 regulates SIRT1 expression in leptin-treated HCT-116 cells. We also found that treatment with leptin led to accumulation of reactive oxygene species (ROS) in HCT-116 cells. Pre-treatment with N-acetyl cysteine attenuated leptin-induced SIRT1 expression, indicating a possible involvement of ROS. In conclusion, leptin, a key adipokine in obesity, may mediate colon tumor progression via Nrf2-dependent SIRT1 expression.

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 5360.
Abstract 5366: NAD(P)H-Quinone oxidoreductase (NQO1): Potential role in the prevention of estrogen-induced breast cancer
Authors: Singh Bhupendra , Bhat Hari K.
Published: February 2011

The importance of estrogens in the etiology of breast cancer is widely recognized. However, the exact mechanisms underlying the initiation and progression of estrogen-related cancers are not clear. Literature evidence and our studies strongly support the role of estrogen metabolism-mediated oxidative stress in estrogen-induced breast carcinogenesis. The objective of the current study was to characterize the role of NQO1 in the prevention of estrogen-induced mammary tumorigenesis by antioxidants vitamin C or butylated hydroxyanisole (BHA). NQO1 converts 17β-estradiol (E2)-quinones back to E2-catechols, thus making E2-quinones unavailable for oxidant damage. Female ACI rats were treated with E2 (3 mg); vitamin C (1% in drinking water); E2 + vitamin C; BHA (0.7% in diet); and E2 + BHA for up to 8 months. The female ACI rat is an established animal model to study the mechanisms of E2-induced breast cancer. The protein isolated from breast tissue and breast tumor tissue of rats treated with E2 and antioxidants was analyzed for NQO1 expression. NQO1 was suppressed in E2-exposed mammary tissue after treatment of rats with E2 for 8 months and in mammary tumors. This suppression was overcome by co-treatment of rats with E2 and vitamin C or BHA. Co-treatment of rats with E2 and vitamin C resulted in a 4-fold induction of NQO1 compared to control mammary tissue. Similarly, co-treatment of rats with E2 and BHA induced NQO1 4.4-fold compared to control mammary tissue. There was no change in NQO1 expression during early time points of E2 exposure (7 and 15 days). The up-regulation of NQO1 by vitamin C or BHA was evident after 4 months of treatment. There was no effect of BHA or vitamin C treatment on NQO1 expression on other non-target organs like liver, lung and heart except kidney. These studies suggest a protective role of NQO1 in E2-induced breast carcinogenesis. NQO1 may thus have a great potential in the development of therapeutic strategies for the prevention of estrogen-induced neoplasia.

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 5366.
Abstract 5370: Potentiation of the novel topoisomerase I inhibitor indenoisoquinoline NSC743400 by Chk1/2 inhibitors in p53-deficient cells
Authors: Aris Sheena M. , Pommier Yves
Published: February 2011

Clincally available camptothecin derivatives, though highly effective against a broad range tumors, are limited by their chemical instability, solubility, ABCG2-mediated efflux and reversibility of Top1-DNA complexes. Indenoisoquinolines were identified as topoisomerase I (Top1) inhibitors by the COMPARE analysis of the NCI drug screen database. Although structurally similar to camptothecin, idenoisoquinolines are chemically stable, able to overcome ABCG2 efflux while forming more persistent Top1-DNA complexes and maintaining potent antitumor activity. The potential for indenoisoquinoline derivatives as possible clinical agents is further aided by the synergistic effect observed between camptothecin and Chk1/2 inhibitors. Chk1/2 kinases regulate cell cycle, DNA damage response via ATM, ATR and DNA-PK, which are activated by genomic instability. Previous studies have shown Chk1/2 inhibitors, such as UCN-01, can potentiate camptothecin-mediated cytotoxicity in p53 mutant cells by eliminating the DNA replication checkpoint.

Further synthesis has led to the water-soluble indenoisoquinoline derivative, NSC743400 for clinical development. Current checkpoint kinase inhibitors, like PV1019 or AZD7762, offer a higher level of specificity than their predecessors that can be exploited by combination with indenoisoquinolines. Checkpoint inhibition showed synergistic antiproliferative activity with NSC743400 in human colon carcinoma HT-29 cells. Furthermore, AZD7762 was able to abrogate the late S-phase arrest induced by NSC743400 as early as 3h post treatment. Inhibition of checkpoint kinases also abrogated the DNA replication fork progression response to NSC743400. The effect of these inhibitors on the autophosphorylation of both Chk1 and Chk2 post NSC743400 was examined by Western blot. These results further substantiate the idea that checkpoint inhibition is a viable strategy in enhancing the antiproliferative effect of novel non-camptothecinn inhibitors such as the indenoisoquinolines.

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 5370.
Abstract 5371: Synergistic antitumor effect of HDAC inhibitor PXD101 in combination with irinotecan in colon cancer
Authors: Na Young-Soon , Jung Kyung-A , Kim Seung-Mi , Ryu Min-Hee , Hong Yong Sang , Kim Jin Cheon , Lee Jung Shin , Kim Tae Won
Published: February 2011

The histone deacetylase inhibitors (HDACi), such as PXD101 or SAHA, inhibit the proliferation and stimulate the apoptosis in tumors. The enhanced effect of HDACi combined with chemotherapy or radiotherapy has been reported in several cancers. In this study, we investigated the antitumor effect of HDACi PXD101 combined with irinotecan in colon cancer.

PXD101 and SN38, active form of irinotecan, had a dose-dependent anti-proliferative activity in HCT116 and HT29 colon cancer cells. PXD101 combined with SN38 had a synergistic effect in colon cancer cell lines, as shown by combination index. The combination of PXD101 with SN38 had an effect on XIAP and p21 protein expressions with time- and dose-dependent manner, more prominently in HCT116 than HT29 cells. These effects were further enhanced by the addition of PXD101 than irinotecan alone in both cell lines. In xenograft mice, PXD101 in combination with irinotecan dramatically inhibited tumor growth without additive toxicity. In addition, antiumor effect was more prominent in HCT116 than HT29 xenografts. Apoptotic effects of tumor in xenografts treated with these combinations were better than irinotecan alone. Combination of PXD101 and irinotecan resulted in 63 % of relative SUV mean in [18F]FLT-PET in mice with established HCT116 xenograft. Early response to these agents can be detected with [18F]FLT-PET. These data suggest that PXD101 increases the cytotoxic activity of iriontecan in colon cancer and these drug combinations should be explored in the treatment of colon 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 5371.
Abstract 5377: Disruption of akt/hif1 signaling can enhance the therapeutic efficacy of ischemic hypoxia and chemotherapy
Authors: Lau Chi-Keung , Yang Zhen Fan , Ho Wing Yuen , Ng Michael , Poon Tung Ping , Fan Sheung Tat
Published: February 2011

PURPOSE: This study investigates the possible molecular basis leading to failure in a treatment that is composed of hypoxia and chemotherapy in a rat orthotopic hepatoma model. EXPERIMENTAL DESIGNS: Cell viability after cisplatin treatments under normoxia and hypoxia would be measured by 3,[4,5-dimethylthiazol-2-yl] −2,5-diphenyl-tetrazolium bromide (MTT) assay and cytofluorometric annexin-V apoptotic assay would be used to compare the apoptotic cells induced by cisplatin under normoxia and hypoxia. Western blotting would be employed to determine the akt/hif1 signaling. In vivo hypoxic condition was induced by hepatic artery ligation, whereas chemotherapeutic effect was achieved by intraportal injection of cisplatin. Hif1 inhibitor was administered to blockade Hif-1α expression both in vitro and in vivo. RESULTS: Cell viability was higher under hypoxic than normoxic conditions shown by MTT assay and decreased apoptotic cells could be observed by apoptotic assay under hypoxia. Hif-1α and akt were regulated each other under hypoxic conditions to confer cisplatin resistance in vitro. In an rat orthotopic hepatoma model, combining blockade of Hif-1α activity with ischemic hypoxia significantly enhanced the efficacy of chemotherapy, leading to suppression of tumor growth and prolongation of animal survival. CONCLUSION: Combined Hif1 inhibitor with hypoxia and chemotherapy can be a new and promising therapeutic approach to the treatment of HCC.

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 5377.
Abstract 5382: Etoposide enhances TRAIL cytotoxicity by increase of TRAIL-R2 expression via a NF-κB pathway in neuroblastoma cells
Authors: Lee Myoung Woo , Kim Hye Ryung , Kim Dae Seong , Noh Yoo Hun , Kim Hye Jin , Lee Soo Hyun , Cheuh Hee Won , Yoo Keon Hee , Sung Ki Woong , Koo Hong Hoe , Jung Hye Lim
Published: February 2011

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family, which induces apoptosis in tumor cells while not in normal cells. TRAIL binds with two types of receptors which are apoptosis-inducing death receptors (TRAIL-R1 and TRAIL-R2) and non-apoptosis-inducing decoy receptors (TRAIL-R3 and TRAIL-R4). TRAIL has been considered as a promising agent for selective cancer therapy, although some cancer cells show resistance to apoptosis induction by this ligand. In this study, we examined whether combination treatment of chemotherapeutic drugs with TRAIL induces synergistic cell death in neuroblastoma cells. We showed that pre-treatment with sublethal concentrations of etoposide followed by exposure to TRAIL synergistically enhanced apoptotic cell death compared to treatment with etoposide or TRAIL alone in SK-N-MC cells (28 ± 2% versus 12 ± 3% or 2 ± 2% apoptotic cell death in 48 h), while not in IMR-32 cells (26 ± 4% versus 26 ± 7% or 0.5 ± 0.6% apoptotic cell death in 48 h). These synergistic cell death was inhibited by addition of fusion chimera protein (recombinant human TRAIL-R2: Fc), which can block TRAIL binding to TRAIL-R1 or TRAIL-R2. In addition, pre-treatment with sublethal concentrations of etoposide in SK-N-MC cells induced degradation of IκBα, resulting in NF-κB activation and up-regulation of TRAIL-R2 expression. The increase of TRAIL-R2 expression resulted in TRAIL-induced synergistic cell deaths, which were effectively inhibited by exposure to NF-κB inhibitor. Moreover, we showed that combination treatment significantly decreased the Mcl-1 expression and increased caspase-3 activation compared to treatment with etoposide or TRAIL alone. Our data suggest that pre-treated etoposide activates NF-κB by down-regulation of IκBα, and then NF-κB up-regulates TRAIL-R2 expression enhancing TRAIL-induced apoptosis in neuroblastoma 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 5382.
Abstract 5384: Tanshinone IIA synergizes with arsenic trioxide to promote human NB4 promyelocytic leukemia cell differentiation and apoptosis
Authors: Wang Zhibo , Johnson Daniel E. , Cui Naiqiang , Wu Xianzhong , Herberman Ronald , Ferrone Soldano , Wang Wenjun
Published: February 2011

Background: Tanshinone IIA (TIIA), a natural compound isolated from Salvia Miltiorrhiza, has been used for hematological malignancies as an adjunctive therapy in combination with arsenic in other countries. However, the molecular mechanism of TIIA as an adjunctive therapy has not been well documented. The goal of this study is to develop novel strategies to treat human promyelocytic leukemia.

Methods: Purified TIIA was used for this pilot study; and purified arsenic trioxide (ATO) was purchased from Sigma. Western blotting and flow cytometry were employed to evaluate biomarker expression in human NB4 promyelocytic leukemia cells prior to treatment and 2-5 days after treatment.

Results: TIIA induced NB4 cell apoptosis at the concentrations above 0.75 µM, a concentration that is below plasma concentrations that can be achieved in vivo. TIIA was not toxic to peripheral blood mononuclear cells (PBMC) from healthy individuals at 3 µM, the highest concentration tested. TIIA also promoted NB4 differentiation at and above 0.75 µM. TIIA up-regulated the percentage of CD11b+ NB4 cells, as well as increasing the percentage of NBT+ cells. In addition, There were not significant differences between TIIA (1µM) and ATRA (1 µM), in terms of their impact on NB4 cell differentiation, as assessed by NBT and CD11b assays. TIIA also inhibited NB4 cell colony formation in CFU-L assays. Western blotting demonstrated that TIIA down-regulated c-Myc expression in NB4 cells. The combination of ATO and TIIA increased NB4 cell death when compared with single drug treatment (TIIA or ATO). In addition, the combination of TIIA and ATO had stronger impact on the induction of NB4 cell differentiation, as assessed by CD11b expression.

Conclusion: TIIA is a potential novel adjunctive agent to be considered in combination with ATO for the treatment of human promyelocytic leukemia.

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 5384.
Abstract 539: Endosialin (TEM1) is a tumor stroma marker and receptor for the metastasis-related Mac-2 binding protein
Authors: Wieland Matthias , Becker Renate , Vijayshankar Sivanandam , Mogler Carolin , Besemfelder Eva , Christian Sven , Augustin Hellmut G.
Published: February 2011

Endosialin is a heavily sialylated C-type lectin. It is a single transmembrane molecule that was identified by two independent studies as an angiogenic endothelial specific marker. However, recent high resolution confocal expression studies by our and other laboratories have revealed that Endosialin is not expressed by endothelial cells but instead by angiogenic blood vessel associated pericytes. Detailed tissue array-based expression profiling validated Endosialin as an oncofetal gene product of the mesenchymal lineage that is hardly detectable in normal tissues but abundantly expressed by tumor vessel-associated pericytes and tumor stromal fibroblasts (myofibroblasts). As such, Endosialin was not just identified as a marker of activated pericytes but based on its exclusive tumor-associated expression also as a novel therapeutic tumor stroma target. Cytokine induction experiments identified Endosialin as a major downstream target of TGFß stimulation. Cellular siRNA-based loss-of-function experiments unravelled a role of Endosialin in the regulation of mesenchymal cell adhesion, migration and proliferation. These findings prompted experiments aimed at identifying the extracellular ligand of Endosialin. Using Endosialin-Fc as an affinity probe, the metastasis associated molecule Mac-2BP/90K was identified as high affinity extracellular Endosialin ligand. As Endosialin, Mac-2BP/90K displays an exclusive tumor progression associated expression pattern. However, in contrast to the stromal expression of Endosialin, Mac-2BP/90K is strictly expressed by the tumor cell compartment. The mutually exclusive expression pattern suggests a repulsive interaction between Endosialin and Mac-2BP/90K which was confirmed in cellular experiments. Ongoing experiments are aimed at mechanistically studying the role of the Endosialin - Mac-2BP/90K interaction in controlling the crosstalk between the tumor cell and the stromal cell compartments during tumor progression and metastasis.

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 539.
Abstract 5397: Crocetin, a carotenoid compound derived from Saffron, enhances antitumor effects of paclitaxol and cisplatin in pancreatic cancer
Authors: Dhar Animesh , Wiegmann Peter S. , Reed Greg , Gutheil William , Veldhuizen Peter Van
Published: February 2011

Pancreatic cancer is the fourth leading cause of cancer deaths in the United States and no significant treatment is at present available. Although there are an increasing number of therapeutic options available for patients with advanced disease, their efficacy is time limited and non-curative. Presently approximately 50-60% of cancer patients in the United States utilize therapies derived from plants, herbs, flowers, or nutrients (complementary and alternative medicine [CAM]), exclusively or concurrently with their traditional therapies such as chemotherapy or radiation therapy. One such CAM therapy is “crocetin”, a carotenoid compound isolated from the saffron plant. Recent studies demonstrated that crocetin has a significant antitumorigenic effect in both in vitro and in vivo on pancreatic cancer. Two conventional chemotherapeutic agents, paclitaxol (a microtubule-targeted agent) and cisplatin (a platinum based anticancer drug binds to DNA), have been demonstrated significant reduction of growth and proliferation in pancreatic cancer. Therefore, the aim of the series of experiments was to determine whether crocetin enhances paclitaxol or cisplatin induced proliferation in pancreatic cancer. MIA-PaCa-2 and BxPC3 cells were treated with crocetin and paclitaxol or cisplatin together at different doses and a proliferation assay was utilized using Click-it Edu flurorometric assay. Paclitaxol or cisplatin alone at higher doses (50-100nM) inhibited proliferation in both MIA PaCa2 and BxPC3 cells but crocetin (200μM) in combination of paclitaxol or cisplatin showed significant additive effect even at lower doses (10 and 25nM) of paclitaxol or cisplatin. The combinational effect is more pronounced in BxPC3 cells. Paclitaxol or cisplatin alone at higher doses (50-100nM) stimulated apoptosis using Deadend flurorometric TUNEL apoptosis assay in MIA PaCa2 and BxPC3 cells and apoptosis is more pronounced at lower doses in combination with crocetin. This study indicated that crocetin in combination with lower doses of paclitaxol or cisplatin inhibited proliferation and stimulated apoptosis in pancreatic cancer. The antitumorigenic effect is more pronounced in BxPC3 cells and paclitaxol is more effective than cisplatin at lower doses in combination with crocetin.

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 5397.
Abstract 5400: Targeting the PI3K pathway improves the effect of PARP-1 inhibition in BRCA1-null breast cancer cells
Authors: Kimbung Siker S. , Berglund Pontus , Gruvberger-Saal Sofia , Hedenfalk Ingrid A.
Published: February 2011

The development of targeted therapies has greatly improved the prognosis for many women affected by breast cancer. Unfortunately, tumors with a triple-negative, ‘basal-like’ phenotype do not express the targets of these agents and remain a treatment challenge. New approaches to specifically target these groups of breast tumors are in need. The majority of BRCA1 mutated tumors display a triple-negative phenotype and are characterized by homologous recombination defects. They frequently harbor mutations in the PTEN gene leading to activation of the PI3K pathway. The absence of BRCA1 and PTEN expression both confer sensitivity to poly-(ADP-ribose) polymerase 1 (PARP-1) inhibitors whilst the activated PI3K signaling most likely promotes cell survival and resistance to apoptotic cues. The therapeutic effects of PARP-1 inhibitors in BRCA1 deficient cells and tumors have been investigated. Even though pre-clinical and clinical studies show promising results, further ways to improve and broaden the scope of application of this therapeutic approach is warranted. We have investigated the possibility of improving the response of BRCA1 mutated cells to PARP-1 inhibition by co-targeting the PI3K pathway. The BRCA1 and PTEN mutated human breast cancer cell line MDA-MB-436 was treated with the PARP-1 inhibitor AG14361 (1-(4-dimethylaminomethyl-phenyl)-8-9-dihydro-7H-2,7,9a-benzo[cd]azulen-6-one) and the PI3K inhibitor LY294002 (2-(4-morpholinyl)-8-phenyl-chromone), both as single agents and in combination and the effects on cell growth, cell cycle phase distribution and DNA integrity were analyzed. Our results confirm that PARP-1 inhibition alone induces DNA damage conferring a G2/M arrest and a profound decrease in cell growth. LY294002, at the low concentrations used, had less pronounced effects on both cell cycle phase distribution and cell growth. Importantly, combining AG14361 and LY294002 resulted in reduced G2/M accumulation concomitant with an even greater growth inhibition compared to AG14361 alone (p<0.01). These observations could indicate a combination specific upregulation of apoptosis. Our results suggest that the combination of PARP-1 and P13K inhibitors is superior over PARP-1 inhibition alone and may represent an improved selective targeted treatment strategy for triple-negative breast cancers with DNA repair defects and activated PI3K signaling.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5400.
Abstract 5403: Akt inhibitor MK-2206 combined with Dasatinib synergistically inhibits the growth of sarcoma cells
Authors: Cubitt Christopher L. , Menth Jillaina L. , Dawson Jana L. , Sullivan Daniel M.
Published: February 2011

The number of effective drugs available for the treatment of soft tissue and bone sarcomas is limited to a small group of antineoplastic agents. Previously, we have identified synergistic anti-tumor activities of the Akt inhibitor, Triciribine, when combined with the Src family inhibitor Dasatinib. The goal of this study was to determine the effectiveness of MK-2206, a newly developed inhibitor of Akt, when combined with Dasatinib in a panel of nine sarcoma cell lines. CT-Blue cell viability experiments were performed to test dilutions of MK-2206 alone and in combination with Dasatinib at a 1:1 constant molar ratio. IC50s for MK-2206 alone ranged from 3 to 12.5 µM in the nine cell lines (Table). Using CI analyses (effective dose levels of 0.7, 0.9 & 0.95), a synergistic or strongly synergistic reduction in cell viability (CI <0.7 or <0.3 respectively) was observed in all cell lines. Synergy was observed over a wide dose range and remained synergistic when as little as 0.8 µM of each drug was combined.

Table. IC50s and Chou and Talalay Combination Index (CI) Synergy Analysis. Inhibitory concentration of MK-2206 required for 50% reduction in viable cells after 72 hour treatment (IC50). CI values represent the level of synergy resulting from concurrent MK-2206 plus Dasatinib treatment. +/− symbols indicate calculated CI range (+++, synergism; ++++, strong synergism).Cell lineSarcoma typeIC50 (M)Combination Index (CI)meanSEMnmeanSEMnSynergyU2-OSosteo12.5E-6809.5E-9120.400.103+++MNNG HOSosteo9.3E-61.2E-6110.210.202++++A-673Ewing9.6E-6963.4E-9120.440.173+++RD-ESEwing8.5E-61.0E-6100.180.032++++SK-ES-1Ewing3.0E-6371.3E-9120.230.084++++SK-LMS-1leiomyo8.2E-6936.4E-9110.320.033+++SK-UT-1leiomyo6.6E-6889.7E-9110.260.153++++SW-872lipo9.0E-61.1E-6120.480.205+++HT-1080fibro11.7E-6694.9E-9110.540.203+++

In Caspase 3/7 activation assays, MK-2206 induced apoptosis in most cell lines at low µM concentrations. Concurrent MK-2206 plus Dasatinib treatment synergistically enhanced apoptosis during the first 24 hrs of treatment in 4 of 9 cell lines. Ongoing experiments are underway to identify the effects of drug sequencing on cell viability and find mechanisms that can account for the synergistic interactions. However, results of current experiments indicate that a MK-2206, Dasatinib combination may be an effective treatment regimen for sarcoma patients. MK-2206 was provided by Merck & Co. Inc. Funded by the Pediatric Cancer Foundation.

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 5403.
Abstract 5404: Inhibition of AKT sensitizes HeLa-S3 cells to arsenite-induced mitotic cell apoptosis
Authors: Yih Ling-Huei , Hsu Nai-Chi , Wu Yi-Chen , Yen Wen-Yen , Tang Yu-Hsuan
Published: February 2011

Arsenic trioxide (ATO) was proved effective in the treatment of promyelocytic leukemia by inducing apoptosis. However, the use of ATO as a single agent in clinical trials against solid tumors was found to be mostly not effective or too toxic. Induction of mitotic arrest was recently shown to be one of the major mechanisms for arsenite-induced apoptosis in cancer cells. Reagents that could effectively enhance ATO-induced mitotic arrest might help improve the therapeutic potential ATO. Cotreatement of HeLa-S3 cells with ATO and LY294002 or a specific AKT inhibitor significantly enhanced mitotic cell accumulation and suppressed the viability of cells at concentrations at which ATO alone was relatively ineffective. The suppression of cell viability was associated with enhanced induction of ATO-induced mitotic abnormalities and was due to induction of mitotic cell apoptosis. Ectopic expression of the constitutively active AKT1 had no effect on ATO-induced mitotic abnormalities, but accelerated mitosis exit, prevented mitotic cell apoptosis, and enhanced the formation of micro- or multi-nuclei after cell division. In addition, AKT1 activation upregulated the expression of aurora kinase B (AURKB) and survivin, and depletion of AURKB and survivin reversed the resistance of AKT1-activated cells to arsenite-induced apoptosis. These results indicate that AKT1 activation suppresses arsenite-induced mitotic cell apoptosis, despite the presence of numerous spindle abnormalities, by upregulation of AURKB and survivin and that inhibition of AKT effectively sensitizes cancer cells to ATO by enhancing mitotic cell apoptosis.

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 5404.
Abstract 541: TGF-β signaling in the tumor microenvironment promotes tumor formation and tumor progression
Authors: Moore Lakisha D. , Isayeva Tatyana , Gurley Jessica , Ponnazhagan Selvarangan
Published: February 2011

Recent studies indicated that the tumor microenvironment plays an important role in tumor growth. Both tumor cells and stromal cells produce TGF-β1, which contributes to autocrine and paracrine functions in the tumor microenvironment. During cell transformation, TGF-β induces angiogenesis, produces reactive stroma, and tumor invasiveness suggesting that crosstalk between the tumor and the stroma is very important in the progression of metastatic disease. The present study examined the effects of TGF-β1 crosstalk in the tumor microenvironment and its role in mediating tumor formation and tumor development by targeted abrogation of TGF-β expression in the tumor cells in situ. Athymic female mice were injected subcutaneously with either normal or tumor derived fibroblast, unmodified MDA-MB 435 breast cancer cells, MDA-MB 435 clones containing TGF-β siRNA either alone or in combination with fibroblasts. Implantation of the cells subcutaneously showed significant difference in tumor formation between the groups. Control animals injected with fibroblast only developed no tumors. In animals injected with TGF-β-silenced MDA-MB 435 clones there was 12.5% tumor incidence. When these same cells were injected in combination with fibroblast the tumor incidence increased to 57%. Unmodified MDA-MB 435 cells showed a tumor incidence of 85% when injected alone and 100% in the presence of fibroblasts. This data suggests that when TGF-β is inhibited in the tumor cells the surrounding stroma can provide growth factors that continue to support tumor formation. In addition to measuring the tumor incidence, tumor growth was assessed every two days over the course of 6 weeks or until tumor was removed due to necrosis. Control animals injected with fibroblasts alone showed no tumor growth. The tumors formed from TGF-β-silenced MDA-MB 435 clones display an average tumor volume of 11.2 mm3. When these cells were co-injected with fibroblasts, the average tumor volume increased to 76 mm3. Unmodified MDA-MB 435 cells produced an average tumor volume of 164.3 mm3 but when injected with fibroblast the volume increased to 229.1 mm3. This data suggests that signaling from the tumor microenvironment can promote tumor progression through TGF-β signaling pathway. Thus, understanding the role of TGF-β1 expression in the tumor microenvironment will help to develop targeted therapies for 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 541.
Abstract 5410: Simultaneously tackling NFκB and JNK pathways to sensitise breast cancer cell lines to conventional anticancer drugs
Authors: Yip* Nga C. , Fombon* Ivo S. , Liu Peng , Brown Sarah , Armesilla Angel L. , Cassidy James , Darling John L. , Wang Weiguang
Published: February 2011

Many anticancer drugs induce reactive oxygen species (ROS) in cancer cells. High ROS will activate JNK pathway and induce cancer cell apoptosis. Meanwhile, ROS also strongly induces NFκB activity which triggers expression of many antiapoptotic factors and inhibits ROS/JNK pathway therefore antagonises anticancer drug-induced programmed cell death. The fate of anticancer agent-treated cancer cells is highly determined by the cross-talk between NFκB and JNK pathways. In this study, we demonstrated that a clinically used anti-alcoholism drug, disulfiram (DS), simultaneously activated ROS-JNK pathway and inhibited NFκB. DS was cytotoxic to breast cancer cell lines in vitro. As a divalent metal ion chelator, the cytotoxic effect of DS on breast cancer cell lines was highly copper-dependent. In the medium containing physiological concentration of copper (Cu, 1µM) the IC50 concentrations of DS to breast cancer cell lines were 200 - 500nM. The chemosensitizing effect of DS on 3 anti-breast-cancer agents, gemecitabine (dFdC), doxorubicine (DOX) and paclitaxel (PAC), was determined in 3 breast cancer cell lines (MCF-7, MDA-MB-231 and T47D). In combination with DS/Cu, the cytotoxicity of dFdC, DOX and PAC was significantly enhanced (Table 1. DOX: 8 - 11-fold; dFdC: 1.2 - 4-fold; PAC: 4 - 10-fold). DS also reversed dFdC resistance in acquired dFdC resistant cell lines. CI-isobologram analysis demonstrated synergistic effect between DS and anticancer drugs. Flow cytometric analysis showed DS enhanced anticancer drug-induced apoptosis. DS enhanced anticancer drug-induced ROS activity. The phosphorylated c-Jun and JNK protein in breast cancer cell lines was significantly induced by exposure of cancer cells to DS/Cu complex. Transfection of NFκB p50 and p65 induced dFdC resistance in MCF7 cells. DS inhibited drug-induced IκBα degradation and NFκB activation. Our data suggested that DS may be developed as a chemosensitizer for BC chemotherapy.

*Equal contributionTable 1.DS/Cu sensitised cytotoxicity of anticancer drugs to breast cancer cell linesTreatmentsDrug: DS/CuMCF7MDA-MB-231T47DDOX (nM) 440.5 (16.2)178.5 (11.9)160.0 (5.5)DOX + DS/Cu1:540.3 (9.3)22.0 (2.0)21.5 (3.1)dFdC (nM) 22.1 (3.4)12.3 (2.2)35.0 (4.3)dFdC + DS/Cu1:104.9 (0.5)11.3 (1.2)12.4 (0.7)PAC (nM) 4.3 (1.4)9.3 (0.7)2.6 (0.3)PAC + DS/Cu1:62.50.4 (0.1)0.6 (0.02)0.7 (0.1)

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 5410.
Abstract 5415: Histone deacetylase inhibitors and 15-deoxy-Δ12,14-prostaglandin J2 synergistically induce apoptosis
Authors: Koyama Makoto , Izutani Yasuyuki , Goda Ahmed E. , Matsui Taka-aki , Horinaka Mano , Tomosugi Mitsuhiro , Fujiwara Jun , Nakamura Yoshitaka , Wakada Miki , Yogosawa Shingo , Sowa Yoshihiro , Sakai Toshiyuki
Published: February 2011

The histone deacetylase inhibitors (HDIs) valproic acid (VPA) and suberoylanilide hydroxamic acid (SAHA, vorinostat) are used clinically for the treatment of epilepsy and cutaneous T-cell lymphoma, respectively. HDIs have been shown to be more effectively utilized in combination with other agents despite the promising anti-tumor effect of HDIs alone. Therefore, additional mechanism-based therapeutic strategies for use of HDIs in combination with other agents should be investigated to overcome resistance to HDIs or to increase the efficacy of HDIs. The natural endogenous ligand of peroxisome proliferator-activated receptor γ (PPARγ) 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) is a member of cyclopentenone prostaglandins and known to be a potent anti-neoplastic agent. Thus, we examined whether VPA or SAHA in combination with 15d-PGJ2 could show synergistic anti-tumor activitiy in colon cancer DLD-1 cells. Cell viability was determined using a Cell Counting Kit-8 assay. Apoptosis and reactive oxygen species (ROS) generation were determined using flow cytometry analysis. Western blotting and real-time RT-PCR analysis were carried out to investigate the expressions of apoptosis-related molecules. Small interfering RNA or transient transfection with plasmids was used for gene silencing, gene overexpression or luciferase assays. Mice bearing DLD-1 xenograft were divided into 4 groups (n=5) and injected everyday (i.p.) with diluent, VPA (100 mg/kg), 15d-PGJ2 (5 mg/kg) or a combination for 25 days. Cotreatment with VPA or SAHA at clinically achievable concentrations and 15d-PGJ2 synergistically induced cell death and drastically caused caspase-dependent apoptosis in DLD-1 cells. The apoptosis induced by VPA/15d-PGJ2 cotreatment was due to alterations of apoptosis-related genes such as an antiapoptotic Bcl-2 family member Bcl-XL, X-chromosome-linked inhibitor of apoptosis (XIAP), C/EBP homologous protein (CHOP) and death receptor 5 (DR5) via ROS generation, subsequent endoplasmic reticulum stress and/or histone deacetylase inhibition. Moreover, VPA/15d-PGJ2 cotreatment induced ROS-dependent apoptosis in other malignant tumor cells and was more effective than a VPA or 15d-PGJ2 monotherapy in vivo. These results suggest that cotreatment with the clinically relevant HDIs and 15d-PGJ2 may warrant clinical investigation as a rational combination therapy against a broad spectrum of malignant tumors.

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 5415.
Abstract 5420: Multi-targeted approach for inhibiting pancreatic tumor growth in vivo consistent with in vitro results
Authors: Ali Shadan , Banerjee Sanjeev , Schaffert Jacqueline M. , El-Rayes Bassel F. , Philip Philip A. , Sarkar Fazlul H.
Published: February 2011

Inactivation of survival pathways such as NF-κB, cyclooxygenase (COX-2) or epidermal growth factor receptor (EGFR) signaling individually may not be sufficient for the treatment of advanced pancreatic cancer (PC). Since these pathways are well known in the pathogenesis and aggressiveness of PC, we hypothesized that a multitargeted approach for the inhibition of NF-κB, COX-2 and EGFR may therefore be useful to improve the outcome of anti-EGFR therapies and will also potentiate the anti-tumor effects of gemcitabine. In this study, we determined the cell viability in seven PC cell lines with 3,3′-diindolylmethane (B-DIM), erlotinib or gemcitabine alone or in combination using MTT assay. Significant inhibition in cell viability and induction in apoptosis was observed in PC cells expressing high levels of COX-2, EGFR, and NF-κB proteins. A significant down-regulation in the expression of COX-2, NF-κB and EGFR in BxPC-3, COLO-357 and HPAC cells was observed, suggesting that multi-targeting of EGFR, NF-κB and COX-2 is more effective than targeting either signaling pathway separately. Most importantly, our in vivo results using animal model showed that B-DIM in combination with erlotinib and gemcitabine was significantly more effective than individual agents and these results were consistent with in vitro findings. Based on our pre-clinical in vitro and in vivo results, we conclude that this multi-targeted combination could be developed for the treatment of PC patients whose tumors express high levels of COX-2, EGFR and NF-κB.

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 5420.
Abstract 5428: Histone deacetylase (HDAC) enzyme inhibition potentiates taxanes and doxorubicin activity in thyroid cancer cell lines
Authors: Owonikoko Taofeek K. , Zhang Guojing , Ramalingam Suresh S. , Khuri Fadlo R. , Sun Shi-Yong
Published: February 2011

Thyroid cancer is the fastest growing cancer in the US. Doxorubicin, the only approved medical therapy, has limited efficacy; therefore alternative treatment options are warranted. We previously showed increased cytotoxicity when HDAC inhibitor was combined with taxanes and carboplatin in lung cancer cell lines. We now studied the combination of HDAC inhibitor and cytotoxic agents in thyroid cancer cell lines. Exponentially growing cells (Cal 62, PTC1, and BCPAP) were treated by continuous exposure for 72 hrs to vehicle or increasing concentrations of doxorubicin, paclitaxel or docetaxel ± fixed concentration of LBH589. Surviving cell fraction was estimated by sulforhodamine B (SRB) colorimetric assay. IC50 concentration was determined by non-linear regression modeling of OD measurements. In a separate experiment, Western blot assay for markers of apoptosis (caspase and PARP cleavage), cell cycle regulation (p21, p27) and tubulin acetylation was performed using cell lysates from cells treated for 48 hrs with IC50 concentration of chemotherapy drugs ± LBH589. Similarly treated cells were analyzed for changes in cell cycle distribution using propidium iodide staining and flow cytometry. Doxorubicin, docetaxel and paclitaxel showed potent nanomolar activity with up to 80% reduction in the IC50 concentrations when combined with LBH589 (paclitaxel: 0% - 70%; docetaxel: 60% - 84%; doxorubicin: 2% - 65%). Flow cytometry showed qualitative evidence for increased cell death with a 1.5 to 6-fold increase in the subG1 apoptotic cell fraction with combined doxorubicin and LBH589 versus doxorubicin alone. The increased cytotoxicity correlated with increased PARP cleavage and increased p21 protein expression; p27 induction by doxorubicin was not augmented by co-treatment with LBH589. HDAC inhibition augments in vitro cytotoxicity of microtubule and DNA-damaging agents in thyroid cancer cell lines. This combination deserves further evaluation in this disease.DrugIC50 Concentrations in Thyroid Cancer Cell Lines (nM)  BCPAPCAL 62PTC1  − L+L− L+L− L+L Paclitaxel3.60.641.310.538.6631.365Docetaxel6.9852.1450.710.282.252.25Doxorubicin34.2512.142278.6778.7188.542LBH569 16.84 11.94 22.71Flow Cytometry Analysis Following Propidium Iodide Staining BCPAPCAL 62PTC1TreatmentVDoxDox + LVDoxDox + LVDoxDox + L%Sub G145.5456.362.286.825.6766.775.456.914.21V: Vehicle;Dox: Doxorubicin;L: LBH589Full details of Western blot assay and complete flow analysis to be presented at the meeting.Acknowledgement: Taofeek Owonikoko, Suresh Ramalingam, Fadlo Khuri and Shi-Yong Sun are all supported by the Georgia Cancer Coalition as Distinguished Cancer Scholars.

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 5428.
Abstract 5429: Inhibition of LSD1 and HDACs causes synergistic cell death in glioblastoma cells
Authors: Singh Melissa , Manton Christa , Tsai Wen-Wei , Barton Michelle , Chandra Joya
Published: February 2011

Glioblastoma multiforme is a particularly aggressive brain tumor and remains a clinically devastating disease. More than 90% of children diagnosed with glioblastoma die within 2 years making it among the leading causes of cancer related morbidity and mortality. Despite innovative therapies for the treatment of glioblastoma, there has not been a significant increase in patient survival over the past decade. Enzymes that control epigenetic alterations have become popular targets for cancer therapy due to their ability to control cellular processes that lead to oncogenesis. Several inhibitors of histone deacetylases (HDACi) have been developed and are in clinical use for cancer therapy. Histone methylation can be modulated by HDACi and here we propose to examine the effect of dually inhibiting deacetylases and demethylases. Specifically, we focused on LSD1, lysine specific demethylase 1, a demethylase known to be in complex with HDAC1/2 and implicated in several high risk cancers. We evaluated apoptosis induction upon knockdown of LSD1 in combination with HDACi in glioblastoma cell lines. Our results demonstrate a greater than two-fold increase in apoptosis in LSD1 knockdown cells treated with increasing doses (1-5 μM) of HDACi for 48 hours as compared to control as measured by DNA fragmentation by propidium iodide staining and analysis by flow cytometry. Furthermore, pretreatment of LSD1 knockdown cells with the general caspase inhibitor zVAD-fmk prior to treatment with vorinostat only partially protects against DNA fragmentation suggesting that cell death proceeds through both caspase-dependent and -independent mechanisms. Similar results were obtained by pharmacologically inhibiting LSD1 with the monoamine oxidase inhibitor tranylcypromine. Treatment of LN-18 cells with combinations of tranylcypromine (100-1000 μM) and vorinostat (1-5 μM) for 72 hours demonstrate that these two agents synergize to promote cell death with 1000 μM tranylcypromine and 5 μM vorinostat showing the most synergistic effect (CI 0.313). Surprisingly, the increase in apoptosis observed upon LSD1 inhibition, by knockdown and pharmacological inhibitors, and HDACi also correlates with an increase in intracellular superoxide as measured by hydroethidum staining and analysis by flow cytometry. These data suggest that reactive oxygen species may play a role in the apoptosis induced by the combination of LSD1 and HDAC inhibition. Further studies aimed at understanding the mechanism by which LSD1 and HDAC inhibition leads to enhanced tumor cell death will substantially increase our knowledge of how epigenetic modifiers interact to control cell survival and may suggest novel options for 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 5429.