bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2456: Mechanism of pancreatic cancer induction by smoking
Authors: Park ChangHwan , Lee In Seok , Abdulsattar Amro M. , Grippo Paul J. , Pandol Stephen J. , Gukovskaya Anna S. , Edderkaoui Mouad
Published: February 2011

Background & Aims: Cigarette smoking is a major risk factor for pancreatic cancer. However, the molecular signaling mechanisms of cigarette smoking remain poorly understood. We studied the effects of the cigarette smoking extract (CSE) and the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) on apoptosis, proliferation and autophagy and underlying signaling mechanisms both in vitro in human pancreatic ductal cells and in vivo in mouse pancreas. We also studied the effect of K-ras mutation on the smoking-induced cell survival and proliferation. Methods: Immortalized human ductal cells HPDE6 were cultured for up to 1 week in the presence or absence of CSE (0.04-4µg/ml) or NNK (1nM-1µM). Pancreas-specific K-ras transgenic mice EL-K-ras were used as a cancer model. Wild type and EL-K-ras mice were injected with NNK (100mg/Kg) once weekly for 4 weeks and animals sacrificed 72h after the last injection. Apoptosis was assessed by measuring DNA fragmentation with Cell death ELISA or TUNEL and proliferation by 3H-thymidine incorporation into DNA or BrdUrd labeling. Autophagy was measured with its marker LC3 using immunofluorescence staining. Molecular signaling mechanisms were evaluated with Western. Results: Exposure of human ductal cells to CSE and NNK for 24h to 1 week time-dependently decreased apoptosis and autophagy and increased proliferation. Exposure to CSE and NNK decreased AMPK phosphorylation and increased Akt phosphorylation. Akt activation occurred at the times of AMPK inhibition. Pharmacological Akt inactivation reversed the inhibition of AMPK phosphorylation by CSE and NNK and prevented the effects of CSE and NNK on apoptosis and autophagy, indicating a central role of Akt in the prosurvival effects of tobacco compounds. In K-ras mice model of pancreatic cancer NNK injections induced increase in Akt kinase and decrease in AMPK phosphorylation. NNK decreased autophagy and apoptosis and stimulated proliferation in the pancreas. The NNK inhibitory effect on apoptosis and autophagy and the stimulation of proliferation were more pronounced in K-ras mice compared to wild type mice. Conclusion: Our in vitro and in vivo data indicate that smoking compounds create conditions favorable for cancer promotion through activating survival pathways including activation of Akt kinase and inhibition of AMPK activities; this leads to a decrease in apoptosis and autophagy and stimulation of proliferation. Our data suggest that smoking works in concert with K-ras mutation to induce adenocarcinoma.

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 2456.
Abstract 2460: Epstein-Barr virus infection confer stress-resistant property in immortalized nasopharyngeal epithelial cells
Authors: Tsao George Sai-Wah , Tsang CM , Zhang GT , Deng Wen , Hau PM , Man CM , Kenzo Takada , Chen HL , Yip YL , Lo KW , Cao Ya , Cheung LM
Published: February 2011

Epstein-Barr virus (EBV) infection has been postulated to be an early event involved in the pathogenesis of NPC. The lack of representative premalignant nasopharyngeal epithelial cell system for EBV infection has hampered research investigation into the regulation and involvement of EBV infection in NPC pathogenesis. We have compared the efficiency of EBV infection in nasopharyngeal epithelial cells with different biological properties including immortalized, primary and cancerous nasopharyngeal epithelial cells. EBV infection could be achieved in all the nasopharyngeal epithelial cells examined with variable infection rate. TGF-β effectively enhanced EBV infection into nasopharyngeal epithelial cells both in the immortalized and primary nasopharyngeal epithelial cells. Stable infection of EBV was achieved in a telomerase-immortalized nasopharyngeal epithelial cell line, NP460hTert. The expression pattern of EBV-encoded genes and biological properties of this EBV infected cell line upon long term propagation were monitored. The EBV-infected nasopharyngeal epithelial cells acquired anchorage independent growth in soft-agar and exhibited invasive growth properties upon prolonged propagation. A distinguish feature of this EBV-infected nasopharyngeal epithelial cell model was its enhanced ability to survive under growth factor and nutrient starvation. This was evidenced by the suppressed activation of apoptotic markers and sustained activation of pAkt of EBV-infected cells compared to control cells under nutrient starvation. Examination of cytokine profiles of EBV-infected NP460hTert cells to nutrient and growth factor deprivation revealed upregulation expression of MCP-1 and GRO-α. The establishment of a stable EBV infection model of premalignant nasopharyngeal epithelial cells will facilitate research investigation into the pathogenic role of EBV in NPC development.

Acknowlegement: The authors wish to acknowlege the funding support from the Research Grant Council (Hong Kong) and the Area of Excellence on Nasopharyangeal Carcinoma Research Center (Hong Kong).

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 2460.
Abstract 2463: Overexpression of galectin-7 in mouse skin inhibits tumorigenesis
Authors: Simper Melissa S. , Thomas Megan M. , Clifford John L. , Fischer Susan M.
Published: February 2011

Galectins are a family of proteins that contain conserved sequences in the carbohydrate-binding region that bind to β-galactosides. Galectin expression differs between tissues in addition to different stages of development and also has altered expression during tumor development. Galectin-7 (Gal-7) has been linked to both apoptosis and reduced cell growth in vitro, but conversely, upregulation of Gal-7 in skin has been reported in many transgenic mice that have endogenous skin tumor promoting activity in addition to being associated with an aggressive phenotype in lymphoma cells. To further investigate this conflicting evidence, the role of Gal-7 in skin carcinogenesis was examined. Gal-7 mRNA and protein levels were elevated in mouse skin after 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment, while Gal-7 stably-transfected cells showed a decrease in tumor growth when compared to vector stably-transfected cells after injection into nude mice, although no difference in cell growth in vitro was observed. Transgenic mice were then generated that overexpress a flag-tagged Gal-7 protein in the basal layer of the skin under the control of the bovine keratin 5 promoter (K5. Gal-7). These mice display no visible phenotype, but immunohistochemistry shows the transgene is present and expressed. Using a two-stage chemical carcinogenesis protocol, the K5. Gal-7 mice were shown to develop significantly fewer papillomas and squamous cell carcinomas than wild type (WT) mice, though no difference in proliferation or apoptosis was seen in response to TPA treatment. No difference in the inflammatory response to TPA was observed, with the exception of a spike in B cells 36 hours after TPA. However, untreated K5. Gal-7 mice showed a decrease in T and B cells compared to WT mice. This supports data from a reverse phase protein array that indicates annexin, which suppresses phospholipase A2 and inhibits inflammation, is upregulated in the K5. Gal-7 mice. These data suggest that overexpression of Gal-7 in the mouse skin inhibits tumorigenesis, possibly through the inhibition of inflammation. Supported by CA100140.

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 2463.
Abstract 2466: Tobacco carcinogen-enhanced lung tumorigenesis in Gprc5a knockout mice recapitulates human lung cancer development
Authors: Fujimoto Junya , Kadara Humam N. , Tao Qingguo , Men Taoyan , van Pelt Carolyn , Lotan Dafna , Lotan Reuben
Published: February 2011

Lung carcinogenesis occurs through a multi-step process, which involves the activation or over-expression of oncogenes and the inactivation or silencing of tumor-suppressor genes. Mouse models for human lung cancer have proven to be valuable tools for understanding the basic tumor biology as well as for the development and validation of new approaches to cancer prevention and therapy. Previously, we found that mice with a knockout (KO) of G-protein coupled receptor 5A (Gprc5a) develop lung tumors after a long latent period (12 to 24 months). To determine whether a tobacco carcinogen will enhance tumorigenesis in this model, we administered 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) i.p. to 2-months old Gprc5a-KO mice and sacrificed groups (n=5) of mice at 6, 9, 12, and 18 months later. Compared to control Gprc5a-KO mice, NNK-treated mice developed lung tumors at least 6 months earlier, exhibited 2- to 4-fold increased tumor incidence and multiplicity, and showed a dramatic increase in lesion size. A signature of differentially expressed genes derived by transcriptome analysis of epithelial cell lines from normal lungs of Gprc5a-KO mice and from NNK-induced adenocarcinoma was highly similar to differential expression patterns observed between normal and tumorigenic human lung cells. The signature also separated both mouse and human adenocarcinomas from normal lung tissues based on publicly available microarray datasets. A key feature of the signature, up-regulation of UBE2C, MCM2, and FEN-1, was validated in mouse normal lung and adenocarcinoma tissues and cells by immunohistochemistry and western blotting, respectively. Our findings support the following conclusions: 1) lung tumorigenesis in the Gprc5a-KO mouse model is augmented by NNK, 2) gene expression changes induced by tobacco carcinogen(s) are conserved between mouse and human lung epithelial cells, 3) the Gprc5a-KO mouse model recapitulates human lung adenocarcinoma. Thus, the Gprc5a-KO mouse model could be useful for cancer chemoprevention studies. Supported by the Samueal Waxman Cancer Research Foundation and the NCI Cancer Center Core Grant P30-CA16672.

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 2466.
Abstract 2476: Tumor development in transgenic mice constitutively expressing hypoxia-inducible factor-1α
Authors: Ide Masaru , Sueoka Eisaburo , Aragane Naoko , Sato Akemi , Nakamura Tomoko , Nakamura Hideaki , Hiyama Keiko , Kimura Shinya , Tanimoto Keiji
Published: February 2011

Hypoxia-inducible factor-1α (HIF-1α) has an essential transcription factor for regulation of various genes associated with hypoxic condition in tissues. Elevated expression of HIF-1α has been reported in many cancers associated with tumor progression, invasion and metastasis. To investigate the role of HIF-1α in tumor development and metastasis, we established transgenic mice constitutively expressing HIF-1α gene under regulation of CMV promoter. The transgenic mice showed no gross morphological abnormality until 3 months after birth, but the mice developed tumors in lymphoid, lung and breast tissues. The incidence of tumor bearing in transgenic mice was up to 90% until 18 months after birth. Expression of HIF-1α mRNA and protein was varied among tissues, but the amount of HIF-1α gene was gradually increased in an age-dependent manner. To clarify the molecular mechanism of lymphomagenesis in the transgenic mice, subpopulation of the lymphocytes, mitogenic responsiveness and cytokine production on lymphocyte from transgenic mice were analyzed. We next analyzed the expression of HIF-1α in human lymphoma tissues and found apparent expression of the gene in angioimmunoblastic T cell lymphoma (AILT). Since VEGF expression and marked increase of small vessels in tumor tissues are common features in AILT, HIF-1α may play an important role in pathogenesis of the lymphoma. We present the results concerning the role of HIF-1α in lymphoma occurrence, and will discuss the contribution of the transcription factor in human cancer 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 2476.
Abstract 2478: Early and enhanced development of N-nitrosobis(2-oxopropyl)amine-induced hamster pancreatic adenocarcinomas by high-fat diet
Authors: Takahashi Mami , Hori Mika , Kitahashi Tsukasa , Imai Toshio , Ishigamori Rikako , Takasu Shinji , Mutoh Michihiro , Sugimura Takashi , Wakabayashi Keiji
Published: February 2011

Epidemiologically, obesity is associated with pancreatic cancer risk, but the mechanisms of obesity-associated carcinogenesis have not yet been clearly understood. Hypertrophic white adipose tissues secrete many adipokines and inflammatory factors, which are expected to enhance cancer development. Syrian golden hamsters, which are used in pancreatic carcinogenesis studies, are in a hyperlipidemic state even under normal diet conditions. We previously have shown that pioglitazone, a PPAR γ ligand, improves hyperlipidemia and suppresses N-nitrosobis(2-oxopropyl)amine (BOP)-induced pancreatic ductal carcinoma development. In the present study, effects of a high-fat diet (HFD) on hyperlipidemia and pancreatic carcinogenesis in BOP-treated hamsters were examined. Six-week-old female Syrian golden hamsters were subcutaneously injected with BOP (10 mg/kg body weight) four times in a week and from 8 weeks of age, the animals in the HFD groups were fed Quick Fat Diet (CLEA Japan, crude fat 14.3 %) for 6 or 17 weeks. At 14 weeks of age, serum levels of triglycerides, total cholesterol, free fatty acids and leptin were significantly elevated in hamsters fed the HFD, and pancreatic ductal adenocarcinomas were already observed in the BOP + HFD group. Pancreatic fatty infiltration was observed to increase in BOP-treated and HFD-fed hamsters. At 25 weeks of age, serum levels of total cholesterol and free fatty acids were further increased in each group and pancreatic ductal adenocarcinomas developed both in the BOP + standard diet group and BOP + HFD group. The numbers of dysplasia and adenocarcinomas in the pancreas were significantly higher in the BOP + HFD group than in the BOP + standard diet group. Apparent fatty infiltration was observed in relatively large adenocarcinomas in the BOP + HFD group. In conclusion, the HFD aggravated the hyperlipidemia, shortened the latency periods of cancer and increased numbers of BOP-induced pancreatic precancerous lesions and adenocarcinomas. Fatty infiltration was apparent in the pancreas of the HFD group and may be associated with pancreatic cancer development in hamsters.

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 2478.
Abstract 2490: The flavonoid quercetin interferes with the activity of anti-microtubule drugs
Authors: Samuel Temesgen , Fadlalla Khalda , Turner Timothy , Yehualaeshet Teshome
Published: February 2011

Dietary compounds with anticancer properties such as polyphenols are intensively studied for their chemopreventive or therapeutic potentials. Quercetin is a polyphenolic flavonoid with anticancer properties and is abundantly available in fruits and green vegetables. In this study, we examined the effects of quercetin on the cell cycle, viability and proliferation of colorectal and prostate cancer cells, either singly or in combination with the microtubule targeting drugs taxol or nocodazole. Quercetin inhibited cell proliferation in a dose dependent manner in both HCT116 colorectal and PPC1 prostate cancer cells. However, unexpectedly, combination treatment of the cells with 50µM or lower quercetin and taxol or nocodazole neutralized the ability of both drugs to induce G2/M arrest and to decrease cell viability. BrdU incorporation was inhibited by quercetin or quercetin-drug combination treatment of cells, but the microtubule targeting of taxol or nocodazole was not inhibited. Moreover, colony formation by both wild type and p53-null HCT116 colorectal cancer cells was inhibited by the same dose of quercetin that neutralized G2/M arrest by Taxol and nocodazole, suggesting a p53-independent effect. We conclude that while quercetin by itself may reduce cell proliferation and survival, in the short-term, the interference of quercetin with cell cycle progression diminishes the ability of taxol and nocodazole to arrest cells at G2/M. Our study adds microtubule targeting drugs to the list of anticancer drugs the activity of which might be antagonized by flavonoids such as quercetin. Further studies are needed to examine the mechanisms of interactions between anticancer drugs and polyphenolic dietary components or supplements, and how these interactions may influence outcomes of cancer therapy, survivorship or recurrence.

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 2490.
Abstract 2494: A non-covalent binding platinum drug, TriplatinNC, induces a G1 phase cell cycle arrest and p53-independent mechanisms of apoptosis
Authors: Kabolizadeh Peyman , Kipping Ralph , Menon Vijay , Ryan John , Peterson Erica , Povirk Lawrence , Farrell Nicholas
Published: February 2011

TriplatinNC, is a novel trinuclear derivative of the Phase II clinical drug, BBR3464. The binding to DNA is through non-covalent (hydrogen bonding and electrostatic) interactions. The absence of substation-labile leaving groups results in insensitivity to glutathione concentration in vitro while the high (8+) charge results in enhanced cellular accumulation compared to BBR3464 and especially cisplatin. The antitumor activity of TriplatinNC was examined in vivo showing higher activity against 2008 ovarian carcinoma cells than c-DDP at the doses administered. Studies using HCT116 colon carcinoma cells have shown that TriplatinNC is cytotoxic in the presence and absence of p53. Cell cycle studies show that TriplatinNC differs from BBR3464 in inducing a G1 arrest rather than a G2 arrest in HCT116 cells. A p53-independent activation of caspase 8 and caspase 9 leading to the cleavage of caspase 3 and its downstream substrate, PARP-1, may indicate that TriplatinNC induces both intrinsic and extrinsic pathways of cell death. Our study illustrates fundamental differences in the mechanisms of action between non-covalent binding drugs like TriplatinNC and related covalent binding drugs such as c-DDP, oxaliplatin, and BBR3464.

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 2494.
Abstract 2498: BNC105 is a tubulin polymerization inhibitor that exhibits vascular disruptive activity in renal cancer and causes upregulation in HIF1alpha, HIF2alpha and mTOR
Authors: Leske Annabell F. , Beaumont Donna M. , Lavranos Tina C. , Gasic Jelena , Coles Carolyn J. , Matthews Clayton A. , Phillips Renee J. , Kremmidiotis Gabriel
Published: February 2011

BNC105 is a tubulin polymerization inhibitor. This agent selectively disrupts the vasculature in tumors, and in addition acts as a direct cytotoxic against cancer cells. BNC105 has demonstrated single agent efficacy in a number of cell lines and tumor models, including breast, colon, prostate, brain and lung cancer. We have investigated the activity of BNC105 in renal cancer cell lines and renal tumor xenografts. BNC105 exhibits cytotoxic activity with EC50 values in the range of 1-20nM. A number of other chemotherapeutic and targeted agents exhibit activity that is comparable or considerably lower than BNC105. Furthermore, we investigated the activity of BNC105 in disrupting the vasculature of tumors arising from human renal cancer cell lines. Our investigations included the renal cell lines Caki-1 and A498. Caki-1 cells express the wild type of the VHL gene, whereas A498 cells expresses a non-functional mutant form of VHL. In addition A498 cells do not express HIF1alpha. Somatic mutation of the VHL gene is the most frequent genetic alteration observed in renal cell carcinoma leading to inappropriate accumulation of HIF1alpha and HIF2alpha. Single dose BNC105 treatment of mice bearing tumor xenografts arising from these cell lines resulted in substantial tumor vascular disruption giving rise to large areas of tumor necrosis. Immunohistochemistry analysis revealed that expression of HIF1alpha was dramatically increased following BNC105 treatment in tumors arising from the renal cell line Caki-1. HIF1alpha expression in A498 tumors was restricted to endothelial cells and did not change following BNC105 treatment. HIF2alpha expression was found to be upregulated in cells surrounding areas of BNC105 induced necrosis in both Caki-1 and A498 tumors. These findings suggest that vascular disruption and necrosis caused by BNC105 lead to an angiogenic rebound effect that may contribute to tumor recovery from the vascular disruption insult. Given the key role of mTOR signaling in upregulation of HIF we examined the expression of phosphorylated mTOR in renal tumor xenografts following treatment with BNC105. Expression of phosphorylated mTOR was distinctly upregulated in viable cell zones surrounding tumor VDA-induced necrotic areas in both Caki-1 and A498 tumors. These findings suggest that tumor recovery from the necrotic insult of BNC105 is mediated through upregulation of mTOR activity. The potential benefit of combining BNC105 with mTOR inhibitors on the growth of renal tumors is currently being investigated.

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 2498.
Abstract 2501: Rapamycin inhibits epidermal cell proliferation in a xenograft model of tuberous sclerosis skin tumors
Authors: Li Shaowei , Thangapazham Rajesh , Wang Ji-an , Rajesh Sangeetha , Moss Joel , Darling Thomas N.
Published: February 2011

Patients with tuberous sclerosis complex (TSC) develop hamartomatous tumors of the skin that have increased dermal fibroblast-like cells and a thickened epidermis. Our previous studies indicate that dermal cells are neoplastic, showing loss of TSC1/TSC2 function and activation of signaling through the mTORC1 pathway, and that overlying epidermal keratinocytes are reactive. The goals of this study were to determine whether TSC skin tumor cells induce epidermal changes and to assess the potential utility and mechanism of action of rapamycin, an mTORC1 inhibitor. TSC2-null skin tumor cells or fibroblasts from patient normal-appearing skin were incorporated into collagen gels overlaid with normal human neonatal keratinocytes, and these constructs were grafted to the backs of nude mice. Mice grafted with TSC tumor cells (n=27) or TSC normal fibroblasts (n=23) received rapamycin (2 mg/kg) (n=24) or an equal volume of vehicle (n=26) by intraperitoneal injection on alternate days for 12 weeks beginning 5 weeks after grafting. Mice were sacrificed 24 hours after the last injection for analysis of the grafts by immunohistochemistry. Grafts containing TSC tumor cells showed greater activation of the mTORC1 signaling pathway than grafts with normal fibroblasts, as shown by greater numbers of dermal cells staining for phosphorylated ribosomal protein S6 (p-S6). Xenografted epidermis overlying TSC tumor cells also showed stronger p-S6 staining than epidermis overlying TSC fibroblasts, replicating what we had observed in TSC skin tumors. Rapamycin treatment of tumor xenografts dramatically inhibited p-S6 staining in both epidermal and dermal cells. Tumor xenografts showed greater epidermal cell proliferation (as determined by staining with anti-Ki-67 specific antibody) than normal xenografts (p=0.009). Rapamycin treatment decreased the number of Ki-67 positive epidermal cells in grafts containing TSC tumor cells (p=0.046) but not in grafts with TSC normal fibroblasts. These results indicate that TSC tumor cells stimulate mTORC1 signaling and proliferation of overlying epidermal cells, and suggest that one of rapamycin effects on TSC skin tumors is through decreasing epidermal cell proliferation.

This work was supported by an Idea Development Award (TS080064) from the Congressionally Directed Medical Research Program to Dr. Darling. Dr. Moss was supported by the Intramural Research Program, NIH, NHLBI.

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 2501.
Abstract 2505: Sorafenib inhibits non-small cell lung cancer cell growth by targeting B-RAF in KRAS wild-type cells and C-RAF in KRAS mutant cells
Authors: Takezawa Ken , Okamoto Isamu , Yonesaka Kimio , Yamada Yuki , Hatashita Erina , Fukuoka Masahiro , Nakagawa Kazuhiko
Published: February 2011

Sorafenib is a multikinase inhibitor whose targets include B-RAF and C-RAF, both of which function in the extracellular signal-regulated kinase (ERK) signaling pathway but which also have distinct downstream targets. The relative effects of sorafenib on B-RAF and C-RAF signaling in tumor cells remain unclear, however. We have now examined the effects of sorafenib as well as of B-RAF or C-RAF depletion by RNA interference on cell growth and ERK signaling in non-small cell lung cancer (NSCLC) cell lines with or without KRAS mutations. Sorafenib inhibited ERK phosphorylation in cells with wild-type KRAS but not in those with mutant KRAS. Despite this difference, sorafenib inhibited cell growth and induced G1 arrest in both cell types. Depletion of B-RAF, but not that of C-RAF, inhibited ERK phosphorylation as well as suppressed cell growth and induced G1 arrest in cells with wild-type KRAS. In contrast, depletion of C-RAF inhibited cell growth and induced G1 arrest, without affecting ERK phosphorylation, in cells with mutant KRAS; depletion of B-RAF did not induce G1 arrest in these cells. These data suggest that B-RAF-ERK signaling and C-RAF signaling play the dominant roles in regulation of cell growth in NSCLC cells with wild-type or mutant KRAS, respectively. The G1 arrest induced by either C-RAF depletion or sorafenib in cells with mutant KRAS was associated with down-regulation of cyclin E. Our results thus suggest that sorafenib inhibits NSCLC cell growth by targeting B-RAF in cells with wild-type KRAS and C-RAF in those with mutant KRAS.

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 2505.
Abstract 2506: Palomid 529, a PI3K/Akt/mTOR dual TORC1/2 inhibitor, is a radiosensitizer with effect in both subcutaneous and orthotopic U251 glioblastoma tumor xenograft models
Authors: Cerna David , Carter Donna , Flaherty Siobhan , Cao Liang , Sherris David , Yoo Stephen S.
Published: February 2011

Palomid 529 (P529) is a small molecule drug created through three generations of computational design. P529 is an anti-tumor agent able to target and inhibit the PI3K/Akt/mTOR signal transduction pathway, specifically as an allosteric dual TORC1/TORC2 inhibitor causing the dissociation of the TORC complexes. Furthermore, P529 has been shown to broadly inhibit cell lines of the NCI 60 cell screen, inhibit HIF-1α, and show tumor growth delay in murine tumor xenograft models. P529 is able to cross the blood-brain barrier and show little or no effect of two transporters (P-gp and Bcrp1) in brain uptake of P529. Here we show that P529 has a more than additive radiation sensitizing activity inhibiting tumor growth in U251 glioblastoma tumor xenograft models both subcutaneous and orthotopic. P529 was administered in a dose-dependent manner at 25, 50 and 95 mg/kg oral dosing every day for 5 days with 4 Gy radiation given on day three. Tumor xenografts in both subcutaneous and orthotopic models were evaluated daily for tumor size or survival respectively up to 40 days post tumor implantation. P529 treatment alone showed marked decrease in tumor growth (subcutaneous) and increase in survival (orthotopic). With addition of radiation, there was a synergistic increase in activity of P529 on the inhibition of U251 tumor growth and survival. All in all, work described here provides evidence that P529 has activity as a radiosensitizer in murine models of glioblastoma and the possibility of efficacious activity in treatment of human patients under a novel mechanism of inhibition of the PI3K/Akt/mTOR pathway.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2506.
Abstract 2509: CDDO-Im induces DNA damage, G2/M arrest and apoptosis in BRCA1-mutated breast cancer cells
Authors: Kim Eun-Hee , Sporn Michael B. , Liby Karen T.
Published: February 2011

The breast cancer-associated gene 1 (BRCA1) is the most frequently mutated tumor suppressor gene in familial breast cancers. BRCA1 mutations also predispose to other types of cancers, pointing to a fundamental role of this pathway in tumor suppression. BRCA1 plays important roles in DNA damage and repair, homologous recombination, cell-cycle regulation, and apoptosis. The synthetic triterpenoid 1-[2-cyano-3,12-dioxooleana-1, 9(11)-dien-28-oyl]imidazole (CDDO-Im) is a promising anticancer agent with potent anti-proliferative and apoptotic activities against a wide variety of cancer types. However the mechanisms responsible for the selective apoptotic effects of CDDO-Im in cancer cells remain elusive. In the present work, treatment of BRCA1-mutated mammary tumor cells with CCDO-Im (0.3-1 μM) led to apoptosis as characterized by decreased thymidine incorporation, increased annexin V-binding, and cleavage of PARP and caspases-3. In addition, 0.3 μM CDDO-Im treatment depleted the G1 population of cells, which was accompanied by an accumulation of cells at G2 phase. CDDO-Im also activated the DNA damage response by inducing the phosphorylation of Chk1/2 and the up-regulation of p21, prior to the induction of apoptosis in BRCA1-mutated cancer cells. The inhibition of Chk1 by UCN-01, a Chk1 inhibitor abrogated G2/M arrest induced by CDDO-Im. To investigate how CDDO-Im activates the DNA damage response in BRCA1-mutated breast cancer cells, we performed a comet assay and Western Blot for γH2AX, a marker of DNA damage. CDDO-Im induced DNA damage and the phosphorylation of H2AX in BRCA1-mutated mammary tumor cells. Pretreatment with 0.3 μM CDDO-Im sensitized BRCA1-mutated cancer cells to carboplatin, a DNA-damaging agent, suggesting that CDDO-Im potentiates the effect of this drug. Moreover, CDDO-Im also induced reactive oxygen species (ROS) generation in these tumor cells, which is associated with the induction of DNA damage. The inhibition of ROS generation by uric acid prevented CDDO-Im-induced DNA damage. Furthermore, treatment with CDDO-Im was not cytotoxic to normal mouse 3T3 fibroblasts, highlighting a selective therapeutic potential of CDDO-Im for BRCA1-associated breast cancer. Taken together, our results demonstrate that CDDO-Im induces ROS and subsequent DNA damage, thereby facilitating the activation of the DNA damage checkpoint, G2/M arrest and finally apoptosis in Brca1-mutated cancer cells. These findings are being explored in vivo to determine whether CDDO-Im could be used therapeutically for BRCA1-associated breast cancer.

This study was supported by Breast Cancer Research 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 2509.
Abstract 2516: Antitumor effects of TAK-733, a novel MEK1/2 inhibitor, against mesothelioma cells
Authors: Miyake Hiroshi , Tsuchiya Shuntarou , Hoshino Takashi , Hori Akira , Vincent Patrick W.
Published: February 2011

Malignant mesothelioma (MM) is a highly aggressive tumor characterized by local invasion of the pleura and metastasis, and is regarded as an almost incurable cancer with increasing incidence worldwide. Various cytokines and growth factors are reported to play important roles in the physiology of MM, possibly through MEK/ERK cascade. Thus, antitumor effects of TAK-733, a highly potent, selective, and non-ATP competitive inhibitor of mitogen-activated protein kinase kinase 1/2 (MEK1/2), were investigated against MM cell lines, NCI-H226, MSTO-211H, NCI-H2052, NCI-H2452, and NCI-H28 harboring wild-type KRAS and BRAF and display moderate levels of basal ERK phosphorylation.

TAK-733 completely inhibited the ERK phosphorylation of 5 mesothelioma cell lines at 1-10 nM. TAK-733 also inhibited the proliferation of these cell lines in a dose dependent manner (IC50 = 4.9-240 nM). Treatment with TAK-733 clearly induced G1 cell cycle arrest with cyclin D1 down regulation and without up regulation of cleaved PARP in NCI-H226 and MSTO-211H cells. Once daily, oral administration of 1, 3, and 10 mg/kg TAK-733 potently inhibited the tumor growth of MSTO-211H xenograft in nude mice with T/C values of 31, 8, and −4% (T/C: treated per control on tumor volume growth over treatment period), respectively, with good tolerability. TAK-733 also showed potent antitumor activities against NCI-H226 and NCI-H2052 xenografts. Additionally, in an intrapleural MSTO-211H xenograft nude mouse model, daily oral administration of 3 and 10 mg/kg TAK-733 for 14 days significantly increased survival. These results demonstrate that TAK-733 is useful for the treatment of malignant mesothelioma with an activate MEK-ERK pathway.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2516.
Abstract 2520: TAK-733, a novel MEK1/2 inhibitor, inhibits tumor angiogenesis without affecting vascular permeability
Authors: Akira Hori , Tsuchiya Shuntarou , Kakoi Yuichi , Mizutani Akio , Vincent Patrick
Published: February 2011

TAK-733 is a highly potent, selective, and non-ATP competitive inhibitor of mitogen-activated protein kinase kinase 1/2 (MEK1/2). In preclinical studies, TAK-733 exhibits potent antitumor activity against a variety of human tumor xenografts, but does not affect vascular permeability. The effects of TAK-733 on tumor angiogenesis were investigated in this preclinical study. TAK-733 completely inhibited the vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF)-induced ERK phosphorylation in human umbilical vein endothelial cells (HUVECs) at 100 nM. TAK-733 also inhibited VEGF- and FGF-induced HUVEC proliferation (IC50 = 1.3 and 18 nM, respectively) and HUVEC tube formation cocultured with fibroblasts (IC50 = 0.49 and 0.34 nM, respectively). The anti-angiogenic effect of TAK-733 in vivo was evaluated in a Matrigel plug assay following daily oral administration of TAK-733 at 3 and 10 mg/kg for 7 days. TAK-733 significantly inhibited the bFGF-induced hemoglobin accumulation in Matrigel plugs. TAK-733 significantly suppressed micro vessel density, examined by immunostaining with an anti-CD31 (PECAM) antibody, in human lung carcinoma A549 xenografts 4 and 7 days after initial treatment of 3 and 10 mg/kg. This was associated with significant antitumor effects in these xenograft models. These results indicate that potent anti-angiogenic activity of TAK-733 contributes to the anti-tumor activities. Angiogenesis inhibitors that modulate VEGF or VEGFR have all displayed hypertensive toxicity. Lack of nitric oxide production as a result of VEGF inhibition produces vasoconstriction and decreased vascular permeability leading to hypertensive effects (Fukumura et. al, PNAS 98, p2604, 2001). The effects of TAK-733 and VEGFR tyrosine kinase inhibitor on tumor interstitial fluid pressure (IFP) were examined by using wick-in-needle method in A549 xenograft model. TAK-733 did not affect the tumor IFP 6 hours after single administration of 30 mg/kg, while the VEGFR tyrosine kinase inhibitor significantly decreased it at 60 mg/kg. Consistent with the effects on tumor IFP, TAK-733 did show change of the vascular permeability unlike the VEGFR tyrosine kinase inhibitor by means of the Evans blue extravasation technique in A549 xenograft model. Although TAK-733 shows potent antiangiogenic effect, these profiles of TAK-733 on vascular permeability would not be expected to induce the typical toxicity of VEGF inhibitor.

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 2520.
Abstract 2523: BRAF is not the only predictor of sensitivity in vitro and in vivo for TAK-733, a selective potent inhibitor of MEK1/2
Authors: O'Connell Shawn M. , Tsuchiya Shuntarou , Shi Lihong , Fabrey Robyn , Zhang Lilly , Dong Qing , Hori Akira , Vincent Patrick
Published: February 2011

MEK has emerged as an attractive target for pharmacologic intervention in cancer as the RAS/RAF/MEK/ERK cascade has proven to play a central role in the signaling required for cellular transformation and proliferation. Inhibition of MEK impacts a diverse array of additional cellular events, including differentiation, apoptosis, and angiogenesis. Identification of an orally active inhibitor of MEK which shows significant efficacy as a mono-therapy for the treatment of cancer would be ideal. TAK-733 is a potent non-ATP competitive allosteric inhibitor of MEK1/2 with IC50 values for the inhibition of MEK1/2 signaling of 2-5 nM at high (400 μM) and low (10 μM) ATP concentrations. TAK-733 was selective for mammalian MEK-1/2 in a panel of 71 enzyme assays. These included phosphodiesterases, 17 kinases (IC50 values >25 µM), cyclases, and 14 other enzymes (IC50 values >10 µM). Using an MTS (a tetrazolium salt) viability assay, TAK-733 was found to inhibit proliferation of numerous cancer cell lines expressing BRAF, KRAS, NRAS, PI3K, MEK1, EGFR mutations, cMET amplification, and wild type MAPK signaling with EC50 values ranging from 2 to 90 nM. A quantitative PK/PD relationship was established in the HT-29 human colorectal carcinoma murine xenograft model. This model showed a range of PD activity (ie, from no effect to maximum effect) which correlated well with tumor and plasma TAK-733 concentrations. The relationship between TAK-733 plasma concentration (PK) and target modulation (PD) was evaluated by modeling percent inhibition of tumor ERK phosphorylation and plasma TAK-733 concentrations at 4 hours postdose, using an inhibitory sigmoidal concentration response model. The plasma EC50 was calculated to be 45 ng/mL. TAK-733 exhibited broad-spectrum in vivo antitumor activity against a panel of human tumor xenografts with and without mutations in the MAPK pathway. These xenografts were of colon (COLO 205/ BRAF V600E and HT-29/ BRAF V600E/PI3K P449T), NSCLC (A549/ KRAS G12S, PC-9/ EGFRΔE746-A750, NCI-H1975/ EGFR L858R T790M), AML (HL-60/ NRAS), cholangio-carcinoma (EGI-1/ KRAS G12D) and melanoma (A375/ BRAF V600E) origins. A high incidence of tumor regressions was observed in this collection of cancer models. Collectively, preclinical pharmacology studies indicate TAK-733 is a specific inhibitor of MEK with the potential to significantly impair growth of a large number of cancers relying on the MAPK signaling cascade for growth and survival. and provides a strong rationale for study of this agent in clinical trials.

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 2523.
Abstract 253: Identification of novel EWSR1-CREB3L1 fusion transcript in small cell osteosarcoma
Authors: Debelenko Larisa V. , Shivakumar Bangalore R. , McGregor Lisa , Dorfman Howard D. , Raimondi Susana
Published: February 2011

Small cell osteosarcoma (SCOS) is an aggressive variant of osteogenic sarcoma that has some morphologic similarities to Ewing sarcoma. Little is known about the molecular biology of SCOS and no specific molecular abnormality has been reported in this tumor subtype to date. We performed a molecular biology study of a SCOS that was positive for the Ewing sarcoma breakpoint region 1 (EWSR1) gene rearrangement by interphase fluorescent in situ hybridization and negative for EWSR1-FLI1, EWSR1-ERG and EWSR1-WT1 fusion transcripts by reverse transcriptase polymerase chain reaction. Rapid amplification of cDNA ends with the EWSR1 gene-specific forward primers revealed the exon 6 of the cAMP responsive element binding protein 3-like 1 gene (CREB3L1) fused in-frame to the EWSR1 exon 11, consistent with the EWSR1-CREB2L1 fusion transcript expressed in tumor tissue. The corresponding chimeric gene in the tumor DNA was confirmed by amplification and subsequent sequencing of the genetic breakpoint located in introns 11 and 5 of the EWSR1 and CREB3L1, respectively. No reciprocal fusion transcript or gene could be amplified in the tumor. An approximately 68 kDa product detected in tumor tissue lysate by immunoblotting with the CREB3L1 carboxyterminal antibody was consistent with a 656 aa predicted chimeric protein. This is the first report of a fusion transcript in osteosarcoma that demonstrates the relation of molecular mechanism of SCOS subtype to that of Ewing sarcoma. The 3'end fusion partner and the inferred structure of the new EWSR1-CREB3L1 chimera, however, are different from those underlying Ewing sarcoma, predicting different functions of the novel fusion product.

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 253.
Abstract 2532: In vitro cytotoxicity, and pharmacokinetics, tissue distribution, and metabolism of the protein kinase D inhibitors, kb-NB142-70 and kb-NB184-43, in mice bearing human cancer xenografts
Authors: Jianxia Guo , Parise Robert A. , Beumer Jan H. , Clausen Dana M. , Egorin Merrill J. , Lazo John S. , Wipf Peter , Wang Q. Jane , Eiseman Julie L.
Published: February 2011

Introduction: Protein kinase D (PKD) contributes to prostate and pancreatic cancer cell growth and survival. Therefore, PKD inhibitors are potential anticancer agents. kb-NB142-70 (NB142) and its methoxy-analogue, kb-NB184-43 (NB184), inhibit PKD in vitro.

Methods: The in vitro inhibition of growth of Pc 3 prostate cells and Panc-1 pancreatic cells by NB142 and NB184 was evaluated by MTT assay. We evaluated the pharmacokinetics of NB142 in SCID mice bearing Pc 3 human prostate cancer xenografts and of NB184 in SCID mice bearing Panc-1 cancer xenografts. NB142 or NB184 was administered i.v. at 25 mg/kg. Three mice per time point were euthanized at 5, 15, 30, 60, 120, 240, 360, 960 and 1440 min after dosing. Plasma, tissues and urine were collected, and concentrations of NB142 and NB184 were quantitated by HPLC-UV. Metabolites in plasma and urine were characterized by LC-MS/MS.

Results: The IC50s of NB142 and NB184 were 21.0 µM and 24.3 µM, respectively, against Pc 3; and 33.7 µM and 27.4 µM, respectively, against Panc-1. After dosing NB142, the plasma NB142 Cmax (at 5 min) was 36.9 µM, and the NB142 Pc 3 tumor Cmax was 11.8 µM. NB142 was not detected in plasma, liver, kidney, spleen, or heart beyond 30 min. After dosing NB184, the plasma, liver, and kidney NB184 Cmax, (at 5 min) were 61.9 µM, 42.6 µM, and 94.8 µM, respectively. The NB184 Cmax in Panc-1 tumors (at 15 min) was 8.0 µM. Areas under the concentration vs time curves of NB142 and NB184 in plasma were 409 and 782 nmol·min/ml, respectively. The plasma half-lives of NB142 and NB184 were 6 and 14 min, respectively. The major metabolite of NB142 was a glucuronide. Between 0-6 h, urinary excretion of NB142 and metabolites accounted for 2.5 % and 7.6%, respectively, of the administered dose. NB184 underwent oxidation and glucuronidation, and more than 10 metabolites were observed. NB142 was not detected in plasma after administration of NB184. Between 0-16 h, urinary excretion of NB184 and metabolites accounted for 2.1 % and 5.3%, respectively, of the administered dose.

Conclusions: Although the inclusion of a methoxy group in NB184 increased its plasma half-life approximately 3-fold compared to NB142, metabolic and urinary elimination of both compounds was relatively rapid. Peak tumor concentrations of NB142 and NB184 were of the same order of magnitude as the concentrations required in vitro for cytotoxicity.

(Support: P30-CA47904 and P01-CA078039)

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 2532.