bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3494: Combining ATRA and GSK3 inhibition in AML
Authors: Gupta Kalpana , Sun Lillian , Agarwal Divya , Ramdeo Ritu , Wald David
Published: June 2012

Acute Myeloid Leukemia (AML) is the most common form of leukemia in adults. Unfortunately, the standard therapeutic agents used for this disease have high toxicities and poor efficacy. The one exception to these poor outcomes is the use of all-trans retinoic acid (ATRA) for a rare subtype of AML (APL, 5-10% of AML). The use of the differentiation agent, ATRA, in combination with low dose chemotherapy leads to the long term survival and presumed cure of 75-85% of patients. Unfortunately ATRA is not clinically useful for other subtypes of AML. Though the non-APL leukemic cells also express retinoic acid receptors (RAR), they require significantly higher concentrations of ATRA for effective differentiation. The requirement for high doses of ATRA for differentiation is at least partially due to the fact that ATRA initiates a negative feedback loop resulting in the degradation of RAR. This feedback loop limits the magnitude and duration of its effects. Despite the high clinical significance of inhibiting the degradation of RAR to enhance the efficacy of ATRA, little is known about the pathways leading to RAR degradation. Our data suggests that the kinase, GSK3, plays a role in RAR degradation. Utilizing genetic and biochemical approaches with AML cell lines and mouse xenograft studies, we have identified that inhibition of GSK3 in combination with ATRA is a promising strategy to induce AML differentiation and growth inhibition. For example, the combination of ATRA and GSK3 inhibitors including Lithium and SB415286 leads to a significant enhancement in anti-tumor activity in a mouse AML xenograft model system. Mechanistically, our data suggests that GSK3 enhances ATRA activity through modulating the expression of RAR. Utilizing HL-60 cells, we have found that not only can endogenous GSK3 and RAR physically interact, but GSK3 can directly phosphorylate RAR and modulate its stability. After the addition of GSK3 inhibitors, the interaction with RAR is disrupted and the GSK3-mediated phosphorylation of RAR is blocked. Overall, our studies suggest the clinical potential of ATRA and GSK3 inhibition for AML and provide a mechanistic framework to explain the promising activity of this combination regimen.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3494. doi:10.1158/1538-7445.AM2011-3494
Abstract 3495: Targeted therapies in melanoma – in vitro effects of single and combined drug treatment
Authors: Azimi Alireza , Egyházi Suzanne , Johansson Carolina , Hansson Johan
Published: June 2012

Malignant melanoma is one of the most rapidly increasing malignancies in Caucasian populations. So far, curative treatment is achieved only by surgical resection at an early stage, while the prognosis in cases with disseminated melanoma is poor, and we lack predictive tools to identify the few patients who respond to existing chemotherapy. Promising clinical trials of targeted therapies are ongoing, however, it is already clear that drug resistance will be a clinical reality also for these new therapies. Thus, novel predictive markers are urgently needed to identify responders to different types of therapy in melanoma.

The aim of the study is to unravel molecular mechanisms and pathways underlying resistance to new therapies in melanoma.

We are studying the effects of a set of MAPK and PI3K pathways inhibitors in melanoma cell lines that are wildtype or have different mutations/deletions in genes affecting these pathways (BRAF, NRAS, PTEN etc.). Cells are exposed to single drugs as well as combinations of inhibitors and/or chemotherapeutic agents.

Our results have shown that there is a wide range of sensitivity to the BRAF inhibitor PLX4720 among BRAF mutated cell lines, with IC50 values varying from 0.3 µM to 30 µM. We are investigating the underlying mechanisms causing this variation in sensitivity to the drug. Using gene expression profiling of a clinical material, we have identified several candidate genes that may predict response to DNA alkylating agents dacarbazine and temozolomide. The expression of some of these genes can be affected by PLX4720, and their potential relevance for response to different treatments is currently tested in vitro.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3495. doi:10.1158/1538-7445.AM2011-3495
Abstract 3508: High antitumor activity of cetuximab combined with taxotere-cisplatin-flourouracil (TPF) association on an orthotopic head and neck cancer model
Authors: Bozec Alexandre , Sudaka Anne , Etienne-Grimaldi Marie-Christine , Fischel Jean-Louis , Milano Gérard Alain
Published: June 2012

Background : Cetuximab (cetux), an anti-epidermal growth factor receptor (EGFR) monoclonal antibody is a standard of care in advanced head and neck (HN) cancer when combined with irradiation or cisplatin. The TPF association including taxotere (T), cisplatin (P) and fluorouracil (F) has become a chemotherapy of reference in the management of HN tumors. The aim of the study was to evaluate the antitumor activity of the association of cetux with TPF. In addition the potential benefit to add bevacizumab (beva), an anti-VEGF monoclonal antibody, was also examined.

Methods : investigations were performed on CAL33 human head and neck cancer cell line (high content of EGFR) injected as an orthotopic xenograft into the mouth floor of nude mice. The anti-tumor efficacy of cetux 2,5 mg/kg, beva 20 mg/kg and of TPF at the MTD (T : 20 mg/kg, P : 6 mg/kg, and F : 17mg/kg) administered alone or in combination at day 3 and 10 after tumor cell injection was assessed at day 12 (animal sacrifice for ethical reasons, 10 animals per treatment group). Tumor volume, tumor histology, tumor cell proliferation (Ki67 by immunohistochemistry) and apoptosis (Bcl2 by immunohistochemistry) were examined. Statistical analyses were performed according to non-parametric methods.

Results : As compared to controls, TPF-cetux and TPF-cetux-beva had a significant impact on tumor volume and histological response. The highest efficacy was observed with the combination TPF-cetux. The effects of the association TPF-cetux-beva were not significantly different than those observed with TPF-cetux. Data were corroborated by a diminution in Ki67 labelling and Bcl2 expression.

Conclusion : the association of cetux with TPF could be an interesting clinical option to evaluate in advanced HN cancer patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3508. doi:10.1158/1538-7445.AM2011-3508
Abstract 3510: Fluvastatin enhances sorafenib cytotoxicity in melanoma cells via modulation of AKT and JNK signaling pathways
Authors: Doudican Nicole A. , Zhang Shali , Quay Ellinor , Orlow Seth J.
Published: June 2012

Background: Most metastatic melanomas are refractory to current available therapy, underscoring the need to identify new effective treatments. Sorafenib, a multikinase inhibitor of the RAF/RAS/MEK pathway, showed promise in earlier stages of clinical development, but ultimately failed to demonstrate efficacy as a single agent in the treatment of melanoma. In order to enhance the efficacy of sorafenib in the treatment of melanoma, we tested over 2,000 naturally occurring compounds and marketed drugs in the presence of a sorafenib in two chemoresistant human melanoma cell lines (M14 and SKM-173) resistant to sorafenib-induced apoptosis. Of the 9 compounds identified as sorafenib sensitizers, we prioritized the cholesterol lowering agent fluvastatin based on its favorable pharmacokinetics and safety profile.

Methodology: Growth inhibition was examined by MTT assay in a panel of 6 human melanoma cell lines. Drug interaction was analyzed by Calcusyn software which determines the effects of two drugs (i.e., synergy, antagonism, or additivity) through calculations of the combination index (CI). The effect of combination therapy on cellular signaling and induction of apoptotic pathways was evaluated by immunoblotting techniques.

Results: Our results demonstrate that fluvastatin at 1µM, a level which is safely achieved through oral administration, dramatically enhances the growth inhibitory activity of clinically achievable concentrations of sorafenib in M14 and SKM-173 melanoma cells with a 3.2 and 3.6-fold reduction in the sorafenib GI50, respectively, with similar effects observed in other melanoma cell lines. Combination indices analysis revealed a synergistic relationship between the two agents. Fluvastatin enhances sorafenib-mediated apoptosis as revealed through enhanced cleavage of PARP. In combination with sorafenib, fluvastatin treatment results in reduced levels of activated AKT along with enhanced levels of activated JNK. Sensitization to sorafenib is unique to fluvastatin as other statins (ie. pravastatin and simvastatin) do not enhance sorafenib-mediated growth inhibition.

Conclusions: These promising results warrant further investigation into the clinical applicability of fluvastatin as an agent for enhancing the efficacy of sorafenib in the treatment of melanoma. Fluvastatin sensitization to sorafenib will be further addressed through follow-up studies in mouse melanoma xenograft models.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3510. doi:10.1158/1538-7445.AM2011-3510
Abstract 3531: Functional characterization of genes that sensitize pancreatic cancer cells to gemcitabine
Authors: Bhattacharjee Vikram
Published: June 2012

Pancreatic cancer has been one of the most aggressive cancers to date. At the point of diagnosis, most patients’ life expectancies are measured in months. While several chemotherapeutic agents have been used to treat pancreatic cancer, none have been very successful in prolonging a patient's lifespan in a major way. Pancreatic adenocarcinoma, the major form of pancreatic cancer, has several risk factors including chronic pancreatitis, advanced age, smoking, obesity, diabetes, and genetic predispositions. Chronic pancreatitis has been thought to promote tumorigenesis ultimately resulting in pancreatic adenocarcinoma. Presently, several classes of drugs (anti-proliferative agents, platanating agents, fluoropyrimidines, and taxanes) are used in the clinic to treat pancreatic cancer, but patient response is very poor when compared to other cancers. Therefore, there is an urgent need to identify novel molecular targets that sensitize pancreatic cancer cells to chemotherapeutic agents.

Gemcitabine (an inhibitor of DNA replication) is of particular interest to us because it remains the drug of choice in the treatment of pancreatic cancer. We have completed a genome wide RNA interference (RNAi) screen and have identified numerous targets that greatly sensitize pancreatic cancer cells to gemcitabine. In the screen, every gene in the human genome was ablated individually via RNAi, so we can identify gene candidates that negatively affect cell survival following drug treatment. We have finalized our validated gene target list. We have utilized automated microscopy tools in addition to molecular biological approaches that are established in our lab in order to chronicle in real time how these pancreatic cancer cells are being sensitized to gemcitabine either via aberrant mitotic processes, direct induction of apoptosis, or other forms of cell death. Indeed, we have found two distinct stages of the cell cycle where death is prevalent. We have the capacity to observe the cells at every stage of the cell cycle before, during, and after the addition of drug. We have established pancreatic cancer cells lines that stably express fluorescent marker proteins that enable us to observe and get a handle on how these cancer cells are surviving drug treatment and how coupling siRNA treatment with chemotherapy sensitizes these cells. We have focused on two major pathways that are aiding cell survival in order to characterize major players that help pancreatic cancer cells survive drug treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3531. doi:10.1158/1538-7445.AM2011-3531
Abstract 3540: Characterization of a novel class of P70S6K in preclinical models
Authors: Araldi Gian Luca , Ronsheim Matthew , Diep Nhut , Guan Ruth , Kalyan Yuriy , Liolis Ira , Ronsheim Melanie , Tang Fuxing
Published: June 2012

P70S6K is a serine/threonine protein kinase responsible for the phosphorylation/ regulation of several important targets like S6, BAD, and IRS1/2. It acts downstream of the mammalian target of Rapamycin (mTOR) and regulates cell growth, proliferation, survival and migration via control of protein translation and a number of important downstream effectors. The over-expression of P70S6K confers clinical aggressiveness in solid tumors, and several findings using indirect inhibition (mTOR inhibitors) or biological tools (RNAi) suggest a critical role for p70S6K signaling in the regulation of invasion and cell motility, making selective inhibitors of this kinase potential anti-metastatic agents. A novel class of phthalazine derivatives were developed that are potent and selective inhibitors of P70S6K, with IC50s for P70S6K in the low nM to sub μμ ρανγε. We have used these inhibitors to further demonstrate and confirm the usefulness of this target for the treatment of metastatic cancers. The effect of the P70S6K inhibitors in regulating cell invasion and motility were determined in several breast (MDA-MB-231, MDA-MB-468), prostate (DU-145, PC-3, and RWPE-2), and melanoma (A-375) cancer cell lines using the modified Boyden chamber assay. All the compounds tested showed dose-dependent inhibitory activity against tumor cell motility and invasion. The activity of these compounds is broad against all the cancer cell lines tested. The activity of our p70S6K inhibitors is comparable to Rapamycin and MEK inhibitors (PD325901), supporting the hypothesis that p70S6K may be a key component for both pathways. Besides the potent in vitro activity, our Lead compound was also effective in the systematic B16-F10 murine melanoma tumor model.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3540. doi:10.1158/1538-7445.AM2011-3540
Abstract 3548: Selective targeting of pancreatic tumors lacking methylthioadenosine phosphorylase (MTAP)
Authors: Bertino Joseph R. , Lee Kuo-Chi , Johnson-Farley Nadine , Lubin Martin
Published: June 2012

The pancreatic cancer cell line, Mia PACA, lacks MTAP, and as a result is unable to salvage adenine and methionine, products of methylthioadenosine (MTA) metabolism by MTAP, and is more sensitive to de novo purine synthesis inhibitors (thiopurines, methotrexate, azaserine and L-alanosine). In addition, MTAP deficiency creates a condition where normal, but not tumor tissues, can be protected from 5-fluorouracil (5-FU) toxicity by MTA. In normal tissue, MTA is converted to adenine, consuming 5-phosphoribose-1-pyrophosphate (PRPP) and competitively inhibiting phosphoribosylation of 5-FU (and subsequent cellular retention and inhibition of thymidylate synthase and incorporation into RNA). In MTAP deficient tumors, MTA can not generate adenine; the relative lack of adenine, and abundance of PRPP favors phosphoribosylation of 5-FU to nucleotides. The MTAP gene is linked to the tumor suppressor p16INK4 locus on chromosome 9p21. This locus is frequently deleted in human cancer. As a result, relative or complete MTAP deficiency is common in several human cancers, including pancreatic cancer (33%). We have transfected the Mia PACA cell line with MTAP cDNA and generated three sublines that express MTAP. The parenteral cell line lacking MTAP expression is more sensitive to 5-FU (4 h treatment), and toxicity is not ameliorated by pretreatment with MTA. In contrast, the three sublines expressing MTAP are a log less sensitive to 5FU, when pretreated with MTA.

As pancreatic cancer is known to be somewhat responsive to treatment with 5-FU, but doses are limited by toxicity, the protection of the host from toxicity by MTA pretreatment may allow higher doses of 5-FU to be safely given and allow gemcitabine, another antimetabolite used to treat patients with pancreatic cancer, to be added in full doses.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3548. doi:10.1158/1538-7445.AM2011-3548
Abstract 3553: Scutellaria barbata inhibits angiogenesis through downregulation of HIF-1 α expression in lung tumor
Authors: Lee Che-Hsin , Shen Yu-Ting , Wu Chao-Liang , Chow N-H , Shiau Ai-Li
Published: June 2012

Hypoxia, a hallmark of many solid tumors, is associated with angiogenesis and tumor progression. Hypoxia-inducible factor-1 (HIF-1) plays a significant role in tumor angiogenesis. In this study, we constructed a selective platform to screen the traditional Chinese medicine as anti-angiogenic agent. We examined the molecular mechanism by which Scutellaria barbata (S. barbata) regulates HIF-1-dependent expression of vascular endothelial growth factor (VEGF), which is an important angiogenic factor. Hypoxia stimulated with the HIF-1 expression promotes angiogenesis by increasing VEGF expression and secretion. Herein, the expression of VEGF was decreased by treatment with S. barbata in tumor cells. Meanwhile, S. barbata reduced the migration and proliferation of endothelial cells under hypoxic condition. S. barbata inhibited the expression of HIF-1α, as well as phosphorylated their upstream signal mediators AKT. S. barbata significant retarded the tumor growth by 56% in vivo and immunohistochemical studies in the tumors revealed decreased intratumoral microvessel density. These results suggest that the traditional Chinese medicine therapy using S. barbata, which exerts anti-angiogenic activities, represents a promising strategy for the treatment of tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3553. doi:10.1158/1538-7445.AM2011-3553
Abstract 3559: A new selective ALK inhibitor CH5424802 shows potent efficacy against ALK-positive cancers including the gatekeeper mutant-driven tumors
Authors: Sakamoto Hiroshi , Tsukaguchi Toshiyuki , Hiroshima Sayuri , Kodama Tatsushi , Kobayashi Takamitsu , Fukami Takaaki A. , Oikawa Nobuhiro , Tsukuda Takuo , Ishii Nobuya , Aoki Yuko
Published: June 2012

The recent development of targeted protein kinase inhibitors has provided new opportunity in cancer treatment. However, certain factors limit the efficacy of cancer therapies, such as a narrow therapeutic index caused by inhibition of multiple kinases, and the emergence of resistant mutants. Thus, kinase inhibitors with more potent and selective properties and effectiveness against resistant mutants need to be developed. Anaplastic lymphoma kinase (ALK) is a tyrosine kinase that is constitutively activated in certain cancers following gene alterations such as chromosomal translocation, amplification, or point mutation. Here, we identified CH5424802, a potent, selective, and orally available ALK inhibitor with a new chemical scaffold, showing preferential antitumor activity against cancers with gene alterations of ALK, such as non-small cell lung cancer (NSCLC) cells expressing EML4-ALK fusion and anaplastic large-cell lymphoma (ALCL) cells expressing NPM-ALK fusion in vitro and in vivo. CH5424802 inhibited ALK L1196M, which corresponds to the gatekeeper mutation conferring common resistance to kinase inhibitors, and blocked EML4-ALK L1196M-driven tumors. Our results support the potential for the clinical evaluation of CH54242802 for long-term treatment of patients with ALK-driven tumors. CH5424802 is currently being investigated in Phase I/II clinical trials for patients with ALK-positive NSCLC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3559. doi:10.1158/1538-7445.AM2011-3559
Abstract 3561: Pazopanib inhibits a subpopulation of PDGFR+ astrocytes in the neuro-inflammatory microenvironment of breast cancer brain metastases
Authors: Gril Brunilde , Vidal-Vanaclocha Fernando , Qian Yong , Liewehr David J. , Steinberg Seth M. , Palmieri Diane , Steeg Patricia S.
Published: June 2012

Brain metastases occur in approximately 35% of metastatic breast cancer patients whose tumors overexpress HER2. Using a quantitative model system to study breast cancer brain metastasis, HER2 overexpression increased the brain colonization of a brain seeking subline of MDA-MB-231 cells (231-BR-HER2). Since HER2 has been shown to induce overexpression of VEGF, we hypothesized that the anti-angiogenic drug pazopanib would reduce the metastatic outgrowth of breast cancer cells in the brain. Pazopanib is a multispecific tyrosine kinase inhibitor targeting VEGFR-1, -2, and -3, PDGFR alpha and beta, and c-kit. We previously showed that pazopanib directly targets the tumor cell through the disruption of B-Raf pathway. Pazopanib prevented the outgrowth of 231-BR-HER2 large brain metastasis by 73% (p<0.0001). Metastasis has been described as an interaction of tumor cells and their environment. Here we evaluated the effect of pazopanib on the brain neuro-inflammatory microenvironment.

Previously, activated microglia and activated GFAP-expression astrocytes were observed around and within experimental brain metastases as well as from resected human brain metastases of breast cancer. In our experiments, the 231-BR-HER2 cell line was injected into the left cardiac ventricle of immunocompromised mice; mice were treated with vehicle, 30mg/kg or 100mg/kg pazopanib twice a day beginning on day 3 post-injection. Brain metastases were counted using H&E staining and brain slides were immunostained with markers for brain micro-environment components.

In the brain metastases, using an antibody to the phosphorylation site tyrosine 751 of PDGFR beta, a subpopulation of GFAP+ astrocytes was observed, previously undescribed. P-PDGFR beta + astrocytes were only detected around the brain metastases, suggesting a specific activation of astrocytes by tumor cells. Pazopanib treatment resulted in a trend, 76% (p=0.13) and 80% (p=0.070) decrease of the pY751 PDGFR beta + astrocyte population, at 30 and 100mg/kg respectively. In contrast, Pazopanib did not have any effect on NG2+pericyte, collagen IV or microglia expression. The data suggest a specific effect of pazopanib on the brain microenvironment. Collectively, the data identify an effect of pazopanib on the brain microenvironment, as well as tumor cells, which makes this drug an interesting candidate for brain metastasis prevention.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3561. doi:10.1158/1538-7445.AM2011-3561
Abstract 3563: A highly selective Erk1/2 Inhibitor with in-vivo anti tumor potency
Authors: Seipelt Irene , Guenther Eckhard , Blumenstein Lars , Mueller Gilbert , Schmidt Peter , Aicher Babette , Teifel Michael , Gerlach Matthias
Published: June 2012

Mutations in receptor tyrosine kinases, Ras or B-Raf frequently manifest constitutive activation of the Ras/Raf/Mek/Erk pathway in a variety of human cancers. The mitogen activated protein kinase Erk1/2 is an important regulator of cell proliferation and was shown to be over-activated by these dysregulated or mutated upstream signals.

AEZS-131 selectively inhibits Erk1/2 with an IC50 of 4 nM, blocks cellular Rsk-1 phosphorylation, modulates downstream cellular substrate activation, arrests tumor cells in G1 and inhibits the growth of multiple human tumor cell lines in the nanomolar range. We have studied the response of human cancer cells (Hct116) in in-vivo mouse xenograft experiments. AEZS-131 significantly inhibited tumor growth at daily doses of 30mg/kg. Currently, AEZS-131 is undergoing in-vivo combination experiments with PI3K inhibitors.

Since pharmacological inhibition of Erk1/2 reverses Ras and Raf activation also in cells demonstrating resistancy to common Raf inhibitors like GDC-0879 and PLX4720, it appears likely to put more attention on the downstream kinase Erk1/2 as therapeutic target. Furthermore, in instances of coincident activation of the Raf and PI3K pathways, combinations of AEZS-131 and PI3K inhibitors may prove efficacious.

Here we present the in-vitro and in-vivo characterisation of the first in-class Erk1/2 inhibitor showing in-vivo efficacy as single therapy agent.

Early development of the Erk1/2 inhibitor AEZS-131 by Æterna Zentaris is integral part of our in-house kinase research program comprising the investigation of different compounds for single Erk1/2 inhibition, single PI3K inhibition and dual kinase inhibitions. All compounds are exclusively synthesized by our Drug Discovery Department.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3563. doi:10.1158/1538-7445.AM2011-3563
Abstract 3569: Changes in signaling pathways induced by vandetanib in a human medullary thyroid carcinoma model as detected by reverse phase protein arrays
Authors: Broutin Sophie , Commo Frédéric , De Koning Leane , Hego Caroline , Marty Bérengère , Talbot Monique , Caillou Bernard , Dupuy Corinne , Ryan Anderson Joseph , Schlumberger Martin , Bidart Jean-Michel
Published: June 2012

Introduction: Medullary thyroid carcinomas (MTC) frequently involve activating mutations in the proto-oncogene tyrosine-kinase receptor RET. Vandetanib (ZD6474, ZACTIMATM) is a potent inhibitor of RET and VEGFR-2 tyrosine-kinase activities which is currently in phase III clinical trial in MTC. The aim of this study was to investigate in vitro and in vivo molecular effects of vandetanib on MTC signaling pathways using a proteomic approach based on reverse-phase protein arrays (RPPA).

Methods: The human MTC TT cell line, bearing a RETC634W activating mutation, was cultured in the absence or the presence of vandetanib (100, 500, 1000 nM) during 4, 24 and 72 hours. MTC TT xenografted athymic nu/nu mice received vandetanib (50 mg/kg/day p.o.) or placebo and xenografts were removed after 1 and 4 days of treatment. Twenty-eight protein extracts from in vitro (n=24) and in vivo (n=4) experiments were profiled by RPPA analysis. Equal amounts of lysates were printed in serial dilutions onto nitrocellulose-coated glass slides. Each slide was then probed with each of a 20-antibodies set, specific for oncogenic pathways (PI3K/AKT/mTOR, MEK/p44-42, STAT3, NFκB) and cell cycle/apoptotic proteins (CDK1, PARP, cleaved-PARP).

Results: In in vitro experiments, vandetanib treatment induced an increase of cleaved-PARP (p<10-5) with a 3-fold rise after 72 hours at 100 nM. Phosphorylation of NFKB was not significantly modified when compared to actin. In contrast, phosphorylation of STAT3 significantly decreased after 72 hours of vandetanib treatment. Overall, the MAPK signaling pathway was dramatically inhibited by vandetanib, as substantiated by the decrease of MEK and p44-42 phosphorylation when compared to their total protein forms (p<10-3). Inhibition of p44/42 and STAT3 phosphorylation were also observed in MTC TT xenografted mice treated by vandetanib. Several proteins were re-analyzed by western-blot and/or immunohistochemistry analyses and the results were in agreement with those observed in RPPA.

Conclusions: Together these observations demonstrated that, at the protein level, vandetanib induces apoptosis and mainly inhibits MAPK and AKT pathways in MTC. Furthermore, our experimental approach confirmed the interest of this method of protein arrays for analyzing changes in signaling pathways induced by tyrosine-kinase inhibitors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3569. doi:10.1158/1538-7445.AM2011-3569
Abstract 3581: Characterization of human breast cancer cells with acquired resistance to the EGFR/ErbB-2 tyrosine kinase inhibitor lapatinib
Authors: De Luca Antonella , Faraso Stefania , D'Alessio Amelia , Gallo Marianna , Lamura Luana , Maiello Monica R. , Normanno Nicola
Published: June 2012

The epidermal growth factor receptor (EGFR) and the ErbB-2 receptor are co-expressed in a subset of breast carcinomas that show a more aggressive phenotype. The dual EGFR/ErbB-2 tyrosine kinase inhibitor lapatinib has been recently approved for the treatment of metastatic breast cancer patients whose tumors overexpress ErbB-2. We investigated the mechanisms of acquired resistance to lapatinib in breast cancer cells. For this purpose, we used as model the SK-Br-3 human breast carcinoma cell line that express high levels of ErbB-2, and moderate levels of EGFR and ErbB-3. We selected lapatinib-resistant SK-Br-3 cells by stepwise dose escalation of lapatinib. After six months of selection, we isolated SK-Br-3 cells that were able to grow in 1 μM lapatinib. The IC50 of lapatinib for parental SK-Br-3 cells was 140 nM, whereas the IC50 for the lapatinib-resistant SK-Br-3 derived cell line (Lap-R) was > 4 μM. Treatment with lapatinib significantly inhibited the activation of EGFR, ErbB-2 and ErbB-3 in both parental and Lap-R SK-Br-3 cells. However, following treatment with lapatinib, Lap-R cells showed a persistent activation of AKT and MAPK, which in contrast were completely inhibited in parental SK-Br-3 cells, as measured by both western blotting and a Luminex-based phosphoprotein array. In addition, treatment with lapatinib produced a slight activation of c-Src in parental cells, whereas significant levels of c-Src activation were detected in Lap-R cells cultured in presence of lapatinib. Lap-R cells also showed higher levels of expression of the chemokine receptor CXCR-4 than parental cells. In agreement with these findings, Lap-R cells showed a significantly higher ability to invade a matrigel-coated membrane as compared with parental SK-Br-3 cells, as measured with a Boyden-chamber based colorimetric invasion assay. Preliminary findings demonstrate that inhibition of c-Src with the selective src kinase inhibitor saracatinib produces a more significant inhibition of Lap-R cell invasion as compared with parental SK-Br-3 cells. Taken together, these findings suggest that activation of c-Src signalling might play a role in acquired resistance to lapatinib in breast cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3581. doi:10.1158/1538-7445.AM2011-3581
Abstract 3585: Antitumor activities of JTP-74057 (GSK1120212), a novel MEK1/2 inhibitor, on colorectal cancer cell lines in vitro and in vivo
Authors: Yamaguchi Takayuki , Kakefuda Reina , Tajima Nobuyuki , Sowa Yoshihiro , Sakai Toshiyuki
Published: June 2012

Purpose: MAPK-pathway is one of the most important pathways for a new anticancer drug development. We performed a high-throughput screening for compounds that induce expressions of p15INK4b, and finally identified a novel MEK1/2 inhibitor JTP-74057 (GSK1120212) being evaluated in clinical trials. We characterized its antitumor activities in vitro and in vivo.

Experimental design: The MEK inhibitory activity was evaluated using recombinant proteins, and its specificity was confirmed against multiple kinases. Several colorectal cancer cell lines were used for proliferation assay, apoptosis assay, and xenograft model.

Results: JTP-74057 strongly inhibited MEK1/2 kinase activities, but did not inhibit other 98 kinase activities. Treatment with JTP-74057 resulted in growth inhibition accompanied with upregulation of p15INK4b and/or p27KIP1 in most colorectal cancer cell lines, which we tested. Daily oral administration of JTP-74057 for 14 days suppressed tumor growth both of HT-29 and COLO205 xenografts in nude mice. Interestingly, the tumor regression was observed only in COLO205 xenografts, and COLO205 was also much more sensitive to JTP-74057 for inducing apoptosis than HT-29 in vitro. Treatment with an Akt inhibitor enhanced the JTP-74057-induced apoptosis in HT-29 cells. Finally, JTP-74057 exhibited an additive or a synergistic effect in combination with the standard-of-care agents, 5-fluorouracil, oxaliplatin or SN-38.

Conclusions: JTP-74057, a highly specific and potent MEK1/2 inhibitor, exerts favorable antitumor activities in vitro and in vivo. Sensitivity to JTP-74057-induced apoptosis might be an important factor for the estimation of in vivo efficacy, and the treatment with an Akt inhibitor enhanced the induction of apoptosis. These results suggest the usefulness of JTP-74057 in therapeutic applications for colorectal cancer patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3585. doi:10.1158/1538-7445.AM2011-3585
Abstract 3587: XL184: c-Met inhibition is effective in a mouse xenograft model of metastatic uveal melanoma
Authors: Yeh Iwei , Griewank Klaus , Ding Vivianne W. , Bastian Boris C.
Published: June 2012

Oncogenic mutation of GNAQ is an early event in uveal melanoma development. Identification of critical downstream effectors of GNAQ could allow for targeted therapy of this deadly form of melanoma. We identified c-Met upregulation as a downstream effect of GNAQ Q209L mutation by analyzing expression changes in immortalized mouse melanocytes transduced with GNAQ Q209L. By Western blot, we established that c-Met protein expression is increased in human uveal melanoma cell lines that harbor a GNAQ activating mutation compared to lines with wild-type GNAQ.

XL184 is a small-molecule tyrosine kinase inhibitor that has activity against c-Met. In vitro assays of XL184 demonstrated preferential activity against human uveal melanoma cell lines (OMM1.3 and Mel202) with GNAQ activating mutations. Intravenous injection of 1×106 OMM1.3 cells into immunocompromised NSG mice results in the development of multiple liver tumor nodules after 6-8 weeks, recapitulating liver tropism (over 80% of patients with metastatic uveal melanoma have liver metastases). We treated mice starting four weeks after OMM1.3 intravenous injection with XL184. After four weeks of treatment, the treated mice demonstrated decreased liver tumor size with central tumor necrosis on histologic examination. The activity of XL184 is likely through its inhibition of c-Met signaling as we found little expression of the other targets of XL184 in OMM1.3. In conclusion, XL184 demonstrates activity in a mouse xenograft model of metastatic uveal melanoma, and has potential to be active against hepatic metastases of uveal melanoma, the major cause of mortality in this rare cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3587. doi:10.1158/1538-7445.AM2011-3587
Abstract 3589: Dovitinib (TKI258), a multikinase inhibitor of FGFR, PDGFR, and VEGFR tyrosine kinases, induces growth inhibition in endometrial carcinoma cells
Authors: Konecny Gottfried E. , Winterhoff Boris , Guorong Ellen Yang , Qi Jingwei , Le Julie , Shi Michael , Dugan Margaret , Linnartz Ronald , Finn Richard S. , Slamon Dennis J.
Published: June 2012

Background: Endometrial carcinoma is the most common gynecological malignancy in the western world. Activating mutations of the fibroblast growth factor receptor 2 (FGFR2) have been described in 12-16% of endometrial cancers. Moreover, vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), and platelet-derived growth factor (PDGF) and their receptors are involved in the neovascularization, invasiveness, and metastatic potential of endometrial cancer. Therefore, suppression of FGFR/VEGFR/PDGFR signaling by TKI258 may represent a novel approach for treatment of endometrial cancer.

Experimental Design: The aims of this study were 1.) to assess the effect of TKI258 on tumor cell growth in two-dimensional (2D) and three-dimensional (3D) culture assays using a panel of 20 human endometrial cancer cell lines, 2.) to identify candidate molecular markers predicting sensitivity using baseline gene expression profiling and mutational analyses, and 3.) to determine the in vivo antineoplastic activity of TKI258 in endometrial cancer xenograft models. In addition, we screened 200 fresh frozen endometrial cancer specimens to comprehensively assess the distribution pattern of PI3K, PTEN, and FGFR mutations in endometrial cancer.

Results: Concentration-dependent anti-proliferative effects of TKI285 using 2D assays were seen in all endometrial cancer cell lines tested, but varied significantly between individual cell lines (IC50 range: 0.42µM – 3.06 µM). The three most sensitive endometrial cancer cell lines demonstrated activating FGFR2 mutations (MFE296: N549K, IC50 0.42µM; AN3CA: N549K and K310R, IC50 0.50µM; MFE280: S252W, IC50 0.66µM). Assessment of TKI258 responses in 3D assays demonstrated similar results to those observed in 2D culture assays in that the 3 cell lines harboring FGFR2 mutations were the most sensitive to TKI258 achieving 100% growth inhibition at a concentration of 1µM. AN3CA and MFE296 endometrial cancer cells formed xenografts in nude mice and antitumor activity was studied in both models. Inhibition of p-FGFR, p-PDGFR, p-VEGFR-2, pAKT and p-ERK1/2 were observed. Comprehensive data on the pattern of PI3K, PTEN and FGFR mutations in endometrial cancer will be provided.

Conclusion: TKI258 demonstrates significant antitumor activity in endometrial cancer cells with FGFR2 mutations. This study provides a strong rationale for the clinical evaluation of TKI258 in patients with endometrial cancer, specifically in those harboring FGFR2 mutations.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3589. doi:10.1158/1538-7445.AM2011-3589
Abstract 3601: CP-868,596, a highly potent and selective PDGFR TKI inhibits growth of PDGFRα-driven lung cancer cells
Authors: Peyton Michael , Chaudhary Preeti , Ramachandran Abhijit , Minna John
Published: June 2012

Background: Platelet-derived growth factor receptor alpha (PDGFRα) has been identified as an oncogenic kinase in lung cancer tumors and cell lines. CP-868,596 is an orally bioavailable, highly potent and selective PDGFR tyrosine kinase inhibitor (TKI). CP-868,596 is a benzimidazole compound that has been designed to block phosphorylation of PDGFRα and PDGFRβ at 1-2 nM concentrations. The 50% inhibition concentration (IC50) of CP-868,596 for PDGFRα and PDGFRβ is 0.9 nM and 1.8 nM, respectively. A phase I trial of CP-868,596 showed good oral bioavailability and a favorable safety profile. Additionally, serum concentrations as high as 2000 nM were achieved. One bronchoalveolar carcinoma patient showed clinical improvement and remained on the trial for more than 170 days. We have tested the activity of CP-868,596 in ∼20 selected NSCLC cell lines, including H1703, which has been identified as a model system to study PDGFRα-driven lung cancer. Each cell line tested was well-characterized with respect to DNA copy number gain, mRNA and protein expression level.

Materials and Methods: Lung cancer cell lines including H1703 (PDGFRA, PDGFC amplification; PDGFRα overexpression), NCI-H1693, NCI-H23 and NCI-H661 (PDGFRA amplification), H1770 (PDGFRA mutation), H1975 (EGFR T790M mutation), EBC1 (MET amplification), H3122 (EML4-ALK translocation), etc. were cultured in HyQ RPMI-1640 medium (HyClone) supplemented with 5% FBS (HyClone) in 96 well plates. Cells were incubated overnight to allow attachment and drug was added ∼24 hours later as a gradient of concentrations to establish the dose-response curves. 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assays were performed 96 hours after drug addition to determine the IC50 of CP-868,596.

Results: Lung cancer cell lines were clustered into 2 categories based on their degree of sensitivity to CP-868,596: highly sensitive and resistant. H1703, the only example of a highly sensitive cell line, is reported to have amplification of both PDGFRA (4q12) and PDGFC (4q32) genes on chromosome 4. In H1703, CP-868,596 has an IC50 of ∼80 nM which is about 2 fold lower than the IC50 for sorafenib, a VEGFR/RAF TKI also known to inhibit PDGFR. The remaining cell lines, including the PDGFRA-amplified lines had IC50 values between 2-8 µM.

Conclusions: CP-868,596 potently inhibits the growth of the cell line, H1703, suggesting that the drug may have single agent activity in PDGFR-driven NSCLC tumors. We plan to identify biomarkers for sensitivity to CP-868,596 in NSCLC to help enrich patients for a potential clinical trial.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3601. doi:10.1158/1538-7445.AM2011-3601
Abstract 3604: Preclinical antitumor study of famitinib, an orally available multi-targeted kinase inhibitor of VEGFR/PDGFR/c-Kit in phase I clinical trials
Authors: Lou Liguang , Mi Yi , Xu Yongping , Xie Chengying , Zhao Hongbing
Published: June 2012

Angiogenesis plays a key role in tumor progression and anti-angiogenic agents including sunitinib and sorafenib that target VEGF/VEGFR signaling pathway have been proved to an effective therapy for cancer in the clinic. However, severe side effects such as hypertension and bone marrow toxicity limit their clinical application. Thus, development of novel anti-angiogenic agents with fewer side effects is still an unmet challenge. In this study, we characterized the in vitro and in vivo antitumor activity of famitinib, an orally active multi-targeted kinase inhibitor. Famitinib inhibited the activity of c-kit, VEGFR-2, PDGFRα and PDGFRβ with IC50 values of 2.3 nM, 4.7 nM and 6.6 nM, respectively. In addition, Famitinib inhibited the VEGF-induced proliferation, migration and tubule formation of human umbilical vein endothelial cells, and micro-vessel spouting from matrigel-embedded rat aortic rings. In vivo, famitinib exhibited broad and potent anti-tumor activity, leading to regression or growth arrest of various established xenografts derived from human tumor cell lines. Moreover, famitinib significantly enhanced the efficacy of oxaliplatin or 5-fluorouracil when they were combined. In summary, famitinib has potent preclinical antitumor activity which supports its further evaluation in clinic. Famitinib is currently in phase I clinical trials in China.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3604. doi:10.1158/1538-7445.AM2011-3604
Abstract 3612: A novel and selective c-Met inhibitor against subcutaneous xenograft and othotopic brain tumor models
Authors: Cui Yumin , Dai Guangxiu , Ren Yongxin , Zhou Feng , Fan Shiming , Sai Yang , Gu Yi , Yan James , Li Jia , Qing Weiguo , Su Weiguo
Published: June 2012

HM5016504 is a novel, highly potent and selective c-Met inhibitor under development by HMPL. In preclinical studies, it demonstrated strong in vitro activity against c-Met and its downstream signaling targets, thus blocking the related cellular functions, including proliferation, migration, invasion, scattering and the secretion of VEGF that plays a pivotal role in tumor angiogenesis. The cross-talk between c-Met and EGFR was investigated, and their contribution to downstream signaling cascade was identified in certain tumor cell lines with dysregulation of both c-Met and EGFR. HM5016504 could effectively inhibit the in vivo growth of a number of human tumor xenografts. Particularly, the tumor cells or xengorafts with high fold of c-Met gene amplification showed high sensitivity to HM5016504, such as gastric cancer SNU-5, Hs746T and lung cancer H1993 and EBC-1 cells or their respective xenografts. The compound not only showed activity in subcutaneous xenograft models, but also demonstrated efficacy in an orthotopic brain tumor model induced by U87MG, indicating it could pass blood brain barrier. In vitro and in vivo activities of HM5016504 were found to be closely correlated with the c-Met expression level and activation status. In addition, HM5016504 exhibited a reasonable pharmacokinetic profile and safety window based on the toxicology study results in rats, dogs, and monkeys. The pre-clinical study results suggest that HM5016504 could be a promising c-Met targeting anti-cancer agent.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3612. doi:10.1158/1538-7445.AM2011-3612