bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3573: The novel FASN inhibitor UCMG028 overcomes resistance to trastuzumab and lapatinib
Authors: Puig Teresa , Gómez Daniel , Urruticoechea Ander , Cufí Sílvia , Turrado Carlos , Aguilar Helena , Campuzano Òscar , Brugada Ramon , López-Rodríguez María Luz , Colomer Ramon
Published: February 2011

The vast majority of Her-2 positive advanced breast cancer patients develop resistance to trastuzumab based therapies within the first year of treatment. We have shown that inhibition of Fatty Acid Synthase (FASN) activity, an enzyme overexpressed and hiperactivated in breast carcinoma, shows marked synergic anticancer activity combined with anti-HER drugs. To examine the molecular mechanisms of acquired resistance to HER2, we have generated clones of FASN+ and HER2+ cell lines (AU565 and SK-Br3) resistant to trastuzumab (TTZ) and lapatinib (LAP) for longer than 9 months, and have evaluated the anticancer effect of UCMG028. We further characterized the effect of UCM028 in a mice model of HER2+ xenograft (BT474). TTZ and LAP-resistant cells displayed an increased sensitivity to UCMG028 compared to previously known FASN inhibitors (C75 and EGCG). Gene expression profiling of LAP and TTZ-resistant cells showed a significant differential expression in different sets of genes involved on cell cycle pathways. Mice BT474 xenografts showed tumor regression after long-term exposure to UCMG028. Preliminary toxicity data showed no weight loss, and histological studies (H&E, Masson's Tricromic, Sudan Black B and Picrosirius Red Stains) showed no structural myocardial abnormalities in treated mice with UCMG028-treated mice. Thus, UCMG028 is an effective anticancer treatment for HER2 + breast cancer and its activity remains unchanged in TTZ and LAP resistant cells. Taken together, these data suggest that FASN activity inhibition by UCMG028 could be a new therapeutic strategy in HER2 + breast cancer with potential to overcome resistance to standard anti-HER2 therapies.

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 3573.
Abstract 3576: Pixantrone is chemically and mechanistically related to mitoxantrone (Mito) and doxorubicin (Dox) but lacks their iron binding properties and myocyte toxicity in vitro
Authors: Hacker Miles P. , Ata Tariq , Gesler Korie , Mazur Jakub , McKennon Marc , Singer Jack
Published: February 2011

The anthracyclines (Dox), anthracenediones (Mito) and anthrapyrazoles (losoxantrone) are all intercalating topoisomerase II inhibitors that have significant anti-tumor activity but have the potential to cause severe, treatment limiting cardiotoxicity. Each of these drugs contains at least one hydroxyquinone element capable of binding iron. Given the putative role of iron in free radical generation and drug induced cardiotoxicity, we synthesized Pixantrone (BBR 2778; Pix), an aza-anthracenedione heterocyclic compound devoid of hydroxyquinone moieties. Pix has recently successfully completed a Phase 3 clinical trial showing excellent anti-tumor activity. Results obtained in animal models demonstrated that Pix caused minimal cardiotoxicity in mice, even those pretreated with Dox. To determine whether Pix binds iron, we used a well established spectrophotometric method to assess drug: iron interactions. Adding increasing amounts of iron to a drug solution we noted the previously reported hypochromic and bathychromic shifts in drug absorption patterns for both Dox and Mito in a 3:1 and 2:1 drug: iron ratios, respectively. In contrast, no iron binding was noted when iron was added to Pix solution at iron: drug ratios ranging from 0.1 to 10.0. To assess the effect of pH on drug binding Pix was incubated with iron at pH of 5, 6, 7 and 8 and at no pH was iron binding detected. Further studies assessing the effect of incubation temperature (40C - 500C) and time of incubation (1 to120 min) demonstrated the lack of iron binding by Pix. Studies performed with the rat myocardial cell line H9C2 indicated that Pix (ID50>50 μg/ml) was far less toxic to confluent myocardial cells than doxorubicin (ID50∼1μg/ml). Further, Pix did not induce a significant increase in reactive oxygen species (ROS) compared to vehicle treated controls. In summary, our results demonstrate that the lack of iron binding and concomitant lack of SOD production may provide the mechanism for lack of cardiotoxicity in animals and an apparent decrease in cardiotoxic potential in humans for Pix. Our results also substantiate the idea that ROS are not necessary for tumor cell cytotoxicity.

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

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3576.
Abstract 3577: Sensitivity and resistance of non-small cell lung cancer to the telomerase inhibitor imetelstat
Authors: Frink Robin E. , Peyton Michael , Sullivan James P. , Wright Woodring E. , Schiller Joan , Shay Jerry W. , Minna John D.
Published: February 2011

Telomerase is expressed in the majority of lung cancers and functions to extend telomeres allowing for unlimited replication potential. Telomerase may also be essential in maintaining a cancer stem cell population making it an attractive therapeutic target. A new telomerase inhibitor, imetelstat (GRN163L), is currently in clinical trials for non-small cell lung cancer (NSCLC). Imetelstat, a lipidated 13-mer oligonucleotide N3′ P5′-thio-phosphoramidate, binds the functional RNA template of telomerase preventing telomerase from extending telomeres and thus potentially leading to telomere loss associated cancer cell death. In the present studies we treated multiple NSCLC cell lines (representing the full spectrum of oncogenotypes and chemotherapy drug response phenotypes) with imetelstat (1 μM, a concentration achieved in patients) continuously over several months and all tumor lines showed loss of telomerase activity and shortening of telomeres. NSCLC lines with initially short telomeres (NCI-H2073-initial telomere length (ITL) 2.7 Kb, Calu-3 (ITL 1.5 Kb), and NCI-H2087 (ITL 3 Kb)) either suffered massive apoptotic associated cell death (H2073) within one week, or developed very short telomeres and ceased proliferating at 2.5 weeks (Calu-3) or 20 weeks (H2087) of treatment, demonstrating primary sensitivity. However, a small subpopulation in H2073 continues to grow in culture in the presence of drug and is being tested for telomere length and sensitivity to higher concentrations of imetelstat. NSCLC lines with long telomeres (NCI-H1819 (ITL 15Kb) and NCI-H157 (ITL 14.5Kb)) treated with 1 µM imetelstat twice weekly for 32 weeks showed reduction of telomeres to 1.8 and 2 Kb respectively with no effect on mass culture proliferation but, in some cases, showed inhibition of colony formation. Removal of imetelstat resulted in return of telomerase activity and telomere re-growth. With continued 1 μM treatment, however, H1819 telomere length stabilized at 1.8Kb while H157 cells showed re-growth of telomeres to parental length and now requires >5 μM imetelstat to robustly inhibit telomerase. In summary, we have found a panel of NSCLC lines demonstrate several imetelstat response phenotypes: extreme initial sensitivity, maintenance of shortened telomeres with inhibition of colony formation, and acquired resistance to imetelstat therapy. We conclude that NSCLCs exhibit a diverse spectrum of responses to telomerase targeted therapy emphasizing the need to have tumor biomarker signatures for personalizing imetelstat therapy in NSCLC.

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 3577.
Abstract 3593: Genetic variants in NER pathway genes and ovarian cancer risk and outcomes
Authors: Gong Yilei , Lin Jie , Ye Yuanqing , Wu Lei , Pu Xia , Wu Xifeng , Liang Dong , Lu Karen
Published: February 2011

Nucleotide excision repair (NER) pathway is critical for protecting against damage from carcinogens. In this case-control study, we evaluated the associations between genetic variations in NER pathway and ovarian cancer risk in 339 Caucasian ovarian cancer cases and 349 Caucasian healthy controls. A comprehensive panel of 217 SNPs in 28 NER pathway genes was analyzed to assess their association with ovarian cancer risk. For the cases who received surgery and platinum-based chemotherapy, we also analyzed the association of these SNPs with overall survival and response to chemotherapy. Individually, 12 SNPs were significantly associated with the ovarian cancer risk. The most significant association was observed for SNP rs4150667 from GTF2H1 (OR = 0.64, 95% CI 0.47- 0.86). A strong gene-dosage effect was observed when these top 12 SNPs were combined for unfavorable genotype analysis. Compared to subjects carrying ≤ 2 unfavorable genotypes, individuals carrying 3 (OR = 2.11, 95% CI = 1.32−3.39), 4 (OR = 2.32, 95% CI = 1.47 − 3.66) or ≥ 5 (OR = 4.31, 95% CI = 2.75 − 6.78) unfavorable genotypes exhibited significantly increased risk for ovarian cancer (P for trend < 0.0001). Classification and regression tree (CART) analysis further revealed potential high order gene-gene interaction among the top 12 SNPs and identified 7 terminal nodes with the percentage of ovarian cancer cases in each terminal node range from 27% to 64%. In the clinical outcome study, there were 10 SNPs that exhibited significant association with overall survival. The unfavorable genotype analysis showed the median survival times for individuals carrying ≤3, 4∼7, and ≥ 8 unfavorable genotypes were 83.3, 40.0, and 24.7 months, respectively, indicating a strong gene-dosage effect (P for trend < 0.0001). We also observed significant gene-dosage effect in chemotherapy response. In summary, this pathway-based approach demonstrated that increasing numbers of unfavorable genotypes in the NER predispose susceptible individuals to increased risk of ovarian cancer and affect clinical outcomes in ovarian cancer patients. These findings warrant further replications in independent populations.

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 3593.
Abstract 3604: Significant effect of polymorphisms in CYP2D6 and DT-1 on clinical outcomes of adjuvant tamoxifen therapy for breast cancer patients
Authors: Zembutsu Hitoshi , Kiyotani Kazuma , Mushiroda Taisei , Imamura Chiyo K. , Hosono Naoya , Kubo Michiaki , Tanigawara Yusuke , Flockhart David A. , Skaar Todd C. , Desta Zeruesenay , Aki Fuminori , Hirata Koichi , Takatsuka Yuichi , Okazaki Minoru , Ohsumi Shozo , Yamakawa Takashi , Sasa Mitsunori , Nakamura Yusuke
Published: February 2011

PURPOSE: The clinical efficacy of tamoxifen is suspected to be influenced by the activity of drug-metabolizing enzymes and transporters involved in the formation, metabolism and elimination of its active forms. We investigated relations of polymorphisms in transporter genes as well as CYP2D6 to clinical outcome of patients with tamoxifen treatment. PATIENTS AND METHODS: We enrolled 282 hormone receptor-positive, invasive breast cancer patients receiving tamoxifen monotherapy. We investigated the effects of allelic variants of CYP2D6 and tag-SNPs of DT-1, DT-2 and DT-3 on recurrence-free survivals using Kaplan-Meier and Cox regression analyses. Plasma concentrations of tamoxifen metabolites were measured in 98 patients taking 20 mg/day of tamoxifen. RESULTS: CYP2D6 variants were significantly associated with shorter recurrence-free survival (P = 3.6×10−5; hazard ratio (HR) 9.52 [95% CI 2.79-32.45] in patients with 2 variant alleles vs without variant allele). Among 51 tag-SNPs in transporter genes, a significant association was found at SNPY in DT-1(P = 1.7×10−4; HR 10.64 [95% CI 1.44-78.88] in patients with AA vs GG genotypes). The number of risk alleles of CYP2D6 and DT-1 showed cumulative effects on recurrence-free survivals (P = 5.5×10−8). The patients carrying 4 risk alleles had 45.25-times higher risk compared to those with ≤1 risk allele. CYP2D6 variants were associated with lower plasma levels of endoxifen and 4-hydroxytamoxifen (P = 4.3×10−6 and 5.2×10−4), whereas no significant difference was found among DT-1 genotype groups. CONCLUSION: Our results indicated that polymorphisms in CYP2D6 and DT-1 are important predictors for the prognosis of breast cancer patients treated with tamoxifen.

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 3604.
Abstract 3609: MGCD265, an orally active Met/VEGFR multitargeted kinase inhibitor in Phase II clinical development, potently inhibits clinically relevant Met mutants
Authors: Beaulieu Normand , Dupont Isabelle , Bonfils Claire , Dubay Marja , Ste-Croix Hélène , Isakovic Lubo , Claridge Stephen , Saavedra Oscar , Raeppel Franck , Raeppel Stephane , Mannion Michael , Vaisburg Arkadii , Besterman Jeffrey M. , Maroun Christiane R.
Published: February 2011

MGCD265 is a multitargeted receptor tyrosine kinase inhibitor in Phase II clinical development, which inhibits the activation of regulators of cancer development and progression by targeting Met, the three members of the VEGFR family, Tie-2 and Ron. Met is a key player in the epithelial-mesenchymal transition (EMT) and Met deregulation is described in several types of cancer. The mechanisms involved in Met activation include co-expression of Met with its ligand HGF, overexpression of the Met protein, Met and/or HGF gene amplification or activating mutations. Originally found in hereditary papillary renal carcinoma (HPRCC), activating missense mutations in the kinase domain provided the first evidence that Met activation was causative of this disease. Mutations outside of the tyrosine kinase domain, such as those in the juxtamembrane and extracellular sema domains have also been analyzed, and found in NSCLC, SCLC, melanoma, head and neck cancer, mesothelioma and pancreatic cancer. For the clinical development of MGCD265, we demonstrated the ability of MGCD265 to inhibit clinically relevant Met mutants described in a variety of cancers, including HPRCC and NSCLC. In cell-based assays, MGCD265 inhibits the activation of Met kinase domain mutants (e.g. Y1230C) as well as mutants within the juxtamembrane region (e.g. R988C), and the extracellular domain (e.g. N375S) with nanomolar potency. This is of particular interest, as several kinase domain mutations confer resistance of Met to Met-selective agents such as PF-4217903. MGCD265 also inhibits signaling pathways activated downstream from Met mutants. The phosphorylation of Gab1 multisubstrate docking protein following the activation of Met mutants is inhibited by MGCD265 as is the ensuing activation of the Erk pathway. Furthermore, MGCD265 inhibits the motility of renal carcinoma cells expressing an extracellular domain Met mutant, and blocks tumor growth in a NSCLC cell line expressing a juxtamembrane domain Met mutant. These results, together with our previous findings, demonstrate that MGCD265 is particularly efficient at inhibiting tumor growth in animal models in which wild-type Met or Met mutants are active, suggesting that MGCD265 may particularly benefit patients with activated Met.

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 3609.
Abstract 362: TR-FRET binding assays for PI3K family inhibitors
Authors: Hereley Sara B. , Wolken Jill K. , Frazee William J. , Eliason Hildegard C. , Riddle Steven M.
Published: February 2011

There is a great clinical need to develop selective, high affinity kinase inhibitors. While many high throughput kinase activity assays have become readily available, easy-to-use and cost-effective, activity-based assays have significant limitations in terms of both the extent of target coverage and the type of information they can provide about compounds. We have developed a competitive TR-FRET binding assay platform for characterization of protein kinase inhibitors. This assay platform has several advantages over traditional activity-based assays, in part due to simplicity of the assays as well as the ability to measure binding events in real time. To extend the breadth of target coverage of this platform, we have developed novel tracers for Class I PI3-kinases and the related protein kinase mTOR. We present a comparison of inhibitor potencies measured in binding assays relative to kinase activity assays for a diverse set of commonly used PI3 kinase family inhibitors. These novel assays enable simple, rapid measurements of IC50 values for PI3 kinases and mTOR.

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 362.
Abstract 3628: JNJ-38877605: A selective oral Met inhibitor for the treatment of cancer
Authors: King Peter , Janssens Boud , Verhulst Tinne , Argent Richard , Kumari Rajendra , Watson Susan , Andries Luc , Mevellec Laurence , Page Martin , Perera Tim
Published: February 2011

JNJ-38877605 is a potent and selective Met receptor tyrosine kinase inhibitor that has substantial pre-clinical antitumor activity. We recently described JNJ-38877605 as an ATP-competitive, inhibitor of the catalytic activity of Met kinase (IC50 4 nM) with a unique binding mode that leads to selectivity, exhibiting a 600-fold selectivity (vs cFMS IC50 2.6μM; the next potently inhibited kinase) for Met compared with a panel of 250 diverse tyrosine and serine-threonine kinases. Previously, JNJ-38877605 was shown to inhibit Met phosphorylation in tumor xenografts up to 16 h following a single oral dose, and that inhibition of receptor phosphorylation was associated with dose-dependent tumor growth inhibition. Herein we describe the activity of JNJ-38877605, on a broader range of preclinical solid tumor xenograft models, at doses below MTD and with different schedules of administration. In addition, we now also show data with primary patient-tumor derived xenografts where we have dosed JNJ-39977605 at the MTD using gastric, colorectal and liver metastatic (derived from primary CRC) samples.

JNJ-38877605 was observed to induce significant tumor regression in large well established MET gene amplified gastric cancer models and in Met pathway activated (autocrine or paracrine) models. Significant growth inhibition was achieved when dosing JNJ-38877605 at MTD (and lower doses) with T/C < 42% in tumors of MKN-45, GTL-16, SNU-5 and Kato II met gene amplified models. Significant inhibition was also observed in K-ras mt (dependent) NCI-H441 NSCLC xenografts. Regression of U87 MG PTEN negative glioblastoma tumors (autocrine loop), grown either as subcutaneous or orthotopic models was also observed.

Using primary patient derived xenograft models, we have shown that differential expression of the c-Met/HGF axis and epithelial-mesenchymal transition (EMT)-related gene markers between primary colorectal carcinomas and liver metastases in primary tissue, play a role in the activity of this compound in vivo.

The strong preclinical properties of the JNJ-38877605 compound has resulted in the selection of this potent and uniquely selective Met inhibitor for clinical evaluation. JNJ-38877605 is currently in Phase I 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 3628.
Abstract 363: Therapeutic targeting of insulin receptor-A with a dual IGF-1R/IR inhibitor: Overcoming a potential escape mechanism from IGF-1R-specific signaling inhibition in insulin receptor-A transfected cancer cells
Authors: Hillerman Stephen M. , Greer Ann F. , Hurlburt Warren , Huang Fei , Reeves Karen , Ryseck Rolf , Finckenstein Friedrich Graf , Dhar Arindam , Kramer Robert A. , Gottardis Marco M. , Carboni Joan M.
Published: February 2011

Type I Insulin-like Growth Factor Receptor (IGF-1R) is a tyrosine kinase receptor that is activated by binding to IGF-1 and IGF-II ligands; IGF-1R has been shown to play a role in cancer development and progression and therapies targeting IGF-1R have resulted in clinical benefit in cancer patients. Insulin Receptor (IR) which is closely related to IGF-1R, is expressed in normal tissues and tumors, and is expressed as two isoforms, IR-A and IR-B. IR-A exhibits mitogenic activity and is preferentially found in fetal tissue and cancer cells, whereas IR-B regulates glucose metabolism in response to insulin. Both IR isoforms bind insulin, and IR-A also binds IGF-II. Several cancer cell types that express IR-A also overexpress IGF-II, suggesting a possible autocrine loop that enhances tumor survival. Since IGF-1R antibodies bind specifically to IGF-1R and not to IR, IR signaling could represent a potential escape mechanism to antibody treatment through the IR-A pathway. BMS-754807, a small molecule inhibitor with dual activity against IGF-1R and IR, recently entered clinical development. It is anticipated that BMS-754807 can prevent or overcome therapeutic resistance resulting from increased IR-activated signaling. The goal of the presented studies was to demonstrate whether dual IGF-1R/IR inhibition by BMS-754807 may prevent IR-mediated resistance to IGF-1R inhibition.

To test this hypothesis, a human rhabdomyosarcoma cell line, Rh41 expressing IGF-1R, but little IR, was engineered to express either IR-A (Rh41-IR-A) or IR-B (Rh41-IR-B). IR expression was confirmed by PCR analysis of specific IR isoforms. Rh41-IR-A cells showed phosphorylation of IR after stimulation with IGF-II and/or insulin. MAB391, a neutralizing monoclonal antibody to IGF-1R, inhibited the proliferation of parental Rh41 and Rh41-IR-B cells in vitro, but Rh41-IR-A cells exhibited no sensitivity to the IGF-1R mAb. Combination treatment with both IGF-1R and IR antibodies in Rh41-IR-A cells were strongly synergistic, suggesting dual receptor blocking is required for enhanced anti-tumor efficacy. Inhibition of IGF-1R and IR with BMS-754807 showed the same inhibitory activity in Rh41-IR-A cells as was observed in the parental Rh41 cells. Therefore, dual IGF-1R/IR inhibition by BMS-754807 treatment may overcome the ability of cancer cells to utilize the IR-A pathway and provide an advantage over IGF-1R specific antibodies in the treatment of cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 363.
Abstract 3631: AN019, a potent anticancer agent, suppresses tumors, ErbB-2, and angiogenesis in pancreatic cancer cells
Authors: Gondi Christopher S. , Gorantla Bharathi , Rao A.K.S. Bhujanga , Amala K. , Naidu M.U.R. , Jogi K.V. , Ramana G.V. , Chowdary M. Praveen , Rao Jasti S.
Published: February 2011

Epidermal growth factor receptor (EGFR), a member of the HER family of receptor tyrosine kinases, has been identified as a therapeutic target for pancreatic cancer. Excessive ErbB-2 signaling is associated with the development of a wide variety of solid malignancies. In this study, we used the pancreatic cancer cell lines MIA PaCa-2 and PANC-1 to demonstrate the suppressive, anti-angiogenic, and anti-proliferative effects of a novel compound known as AN019. Here, we show that AN019 inhibits the invasiveness of pancreatic cancer cells in a dose-dependent manner and is superior to treatment with Gemcitabine alone. Angiogenic assays demonstrated that AN019 treatment retarded the invasive potential of pancreatic cancer cells and provided greater angiogenic inhibition than Gemcitabine. In addition, western blot analysis demonstrated that AN019 inhibits expression of ErbB-2 in a dose-dependent manner in MIA PaCa-2 and PANC-1 cells. In contrast, Gemcitabine lacked a similar dose response. Finally, the proliferation assay showed that while AN019 retarded proliferation in a dose-dependent manner, Gemcitabine did not. Our in vivo studies showed that expression of ErbB-2 and VEGF decreased in a dose-dependent manner in AN019-treated animals with subcutaneously implanted MIA PaCa-2 tumors. A treatment combination of AN019 and Gemcitabine had an effect similar to that of treatment with AN019 alone, thereby indicating no specific contribution from Gemcitabine. Our results demonstrate AN019 is superior to Gemcitabine, and AN019 is a specific suppressor of ErBB-2 in vitro and in vivo. In addition, AN019 has anti-proliferative and anti-angiogenic properties. Taken together, these results clearly indicate the potential therapeutic use of AN019 as an anti-cancer agent.

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 3631.
Abstract 3633: eEF2 in acquired lapatinib resistance in HER2 positive breast cancer cells
Authors: McDermott Martina , Browne Brigid , Crown John , O'Brien Neil , Slamon Dennis , Henry Michael , Meleady Paula , Clynes Martin , Dowling Paul , O'Donovan Norma
Published: February 2011

Approximately 25% of breast cancers overexpress HER2. Although HER2 targeted therapies, trastuzumab and more recently lapatinib, have improved prognosis for HER2 breast cancer patients not all HER2 positive tumors respond to HER2 targeted therapies. Using phosphoproteomic profiling, we examined mechanisms of resistance in an in vitro model of acquired lapatinib resistance.

SKBR3 cells were conditioned in lapatinib containing media (200 nM) for six months, after which time the resulting SKBR3-L cells exhibited significantly reduced growth inhibition (27.7 ± 0.1 %) when treated with lapatinib (1 µM) treatment compared to parental SKBR3 (90.7 ± 0.2 %) (p<0.01). Using phosphoproteomic profiling, eukaryotic elongation factor 2 (eEF2) was identified as a phosphoprotein which was significantly altered in the SKBR3-L cells compared to the parental SKBR3 cells. One form of phospho-eEF2 was up-regulated and 6 forms were down-regulated in SKBR3-L compared to SKBR3. eEF2 is a phosphoprotein with an essential role in protein synthesis. Phosphorylation of eEF2 at Thr56 and Thr58 by eEF2 kinase inhibits eEF2 activity. eEF2 kinase activity is regulated by the mTOR pathway, via phosphorylation by p70S6 kinase.

To examine the role of eEF2 in lapatinib resistance, levels of phospho-eEF2, eEF2, phospho-p70S6K and p70S6K, were examined in a panel of HER2 positive breast cancer cell lines following 24 hour treatment with lapatinib (1 µM). Treatment with lapatinib resulted in a decrease in phospho-p70S6K and an increase in phospho-eEF2, with no change in the levels of total protein, in lapatinib sensitive cell lines.

In order to further examine the role the mTOR pathway and eEF2, plays in acquired resistance to lapatinib, the effect of rapamycin, alone and in combination with lapatinib, was investigated in SKBR3 par and SKBR3-L cells. Combined treatment with lapatinib (200 nM) and rapamycin (2 nM) results in similar inhibition of growth (64.5 ± 3.3 %) in SKBR3-L cells as lapatinib alone in SKBR3 parental cells (62.5 ± 0.6 %).

In conclusion, response to lapatinib is associated with decreased eEF2 activity in HER2 positive cell lines. Furthermore, inhibition of mTOR signaling, which causes inhibition of eEF2 activity, appears to restore sensitivity to lapatinib in our model of acquired lapatinib resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3633.
Abstract 363A: PAX2 is an oncogene in type I ovarian cancer
Authors: Song Huijuan , Tung Celestine S. , Tsang Yvonne TM , Mok Samuel C. , Gershenson David M. , Wong Kwong-Kwok
Published: February 2011

Ovarian cancer is the second most common gynecologic malignancy, but the most lethal gynecologic cancer. Ovarian cancer is divided into Type I and Type II subgroups. Type I tumors include low-grade micropapillary serous carcinoma, mucinous, endometrioid, and clear cell carcinomas and are characterized by high frequency of KRAS, BRAF, PTEN, or beta-catenin mutations. Type II tumors include high-grade serous carcinoma, malignant mixed mesodermal tumors (carcinosarcomas) and undifferentiated carcinomas and are characterized by high genetic instability and high frequency of TP53 mutation. Using gene expression profiling, we found that Pax2, a transcription factor of the Pax gene family, is highly expressed in type I ovarian tumors but is less expressed in Type II tumors. Pax2 is one of the nine Pax genes which have a conserved DNA sequence motif called the paired box, a 128 amino acid domain in the amino-terminal portion of the protein. Pax2 regulates tissue development and cellular differentiation in embryos and Pax2 expression is attenuated after development. Unattenuated Pax2 expression has been found in several cancer types, but the function of Pax2 in ovarian cancer is not clearly understood.

In this study, we further validated the expression of Pax2 in Type I ovarian cancer by immunohistochemistry, western blot analysis and quantitative real-time PCR. Furthermore, the expression of Pax2 in two Type I ovarian cancer cell lines (RMUGL and TOV21G) was knockdown by Pax2 specific shRNA. By comparing to the control non-target shRNA cancer cell lines and the parental cell lines, Pax2 knockdown cell lines have significant reduction in growth rate as measured by WST cell proliferation assay. From the gene expression profiles of three independent Pax2 knockdown cell lines, we identified 196 genes that were significantly affected by the down-regulation of Pax2. Many of these genes are involved in cell cycle control, cellular growth and proliferation, and the TP53 pathway such as BBC3, CHEK1, GADD45A and GADD45B and JUN genes. This suggests that Pax2 may play an oncogenic role in the pathogenesis of ovarian cancer through a TP53-dependent pathway. Since immunotherapy targeting Pax2 is currently being developed, Pax2 could be a therapeutic target for Type I ovarian cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 363A.
Abstract 3642: AS1411 causes a specific increase in levels of cell surface nucleolin in responsive cell lines
Authors: Teng Yun , Jones David , McLaughlin Fiona , Bates Paula J.
Published: February 2011

AS1411 is a 26-mer guanine-rich oligodeoxynucleotide that has antiproliferative activity against many types of cancer cells, but not against normal cells. It is currently being tested as an anticancer agent in Phase II clinical trials. Previously, we reported that this agent has a novel mechanism of action that involves formation of a stable G-quadruplex structure that can bind as an aptamer to nucleolin, a multifunctional protein expressed at high levels by cancer cells. Nucleolin is a predominantly nuclear protein, but can also be present in the cytoplasm and on the surface of cells, where it has been reported to mediate the endocytosis of several ligands. Cell surface nucleolin is believed to play an important role in AS1411 activity, although the precise details have yet to be elucidated. In the present study, we have evaluated the levels of cell surface nucleolin in human cell lines that are responsive or resistant to AS1411, before and after treatment with AS1411. Levels were examined by flow cytometry, with gating to exclude non-viable permeable (PI-positive) cells, and by western blotting of cell surface proteins, isolated using a cell-impermeable biotinylating agent. The amount of cell surface nucleolin appeared to be similar in untreated cells that were either sensitive to AS1411 (DU145 prostate cancer cells and MV-4-11 leukemia cells) or resistant to AS1411 (Hs27 non-malignant skin fibroblasts). However, in the sensitive cell lines, we observed a dramatic and time-dependent increase in levels of cell surface nucleolin in response to AS1411 treatment, whereas there was no significant change in the resistant Hs27 cells. We determined that this AS1411-induced increase in cell surface nucleolin in the cancer cells was not simply a reflection of apoptosis because it was not observed when apoptosis was induced by UV irradiation. We did not observe any changes in the levels of nucleolin in nuclear, cytoplasmic, or membrane extracts, or in total cell lysates, following treatment with AS1411, suggesting a specific re-localization of nucleolin rather than an increase in overall levels. These new results do not support our previous hypothesis that baseline levels of cell surface nucleolin determine responsiveness to AS1411. However, they do clearly demonstrate that “externalization” of nucleolin is a specific response to AS1411 and thereby confirm the important role of cell surface nucleolin in the mechanism of action of AS1411.

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 3642.
Abstract 3654: Clofarabine: factors influencing cytotoxicity and resistance in hematologic and solid malignancies
Authors: Wang Xin , Mirzaee Saeedeh , Lotfi Kourosh , Eriksson Staffan , Albertioni Freidoun
Published: February 2011

Clofarabine is a new-generation nucleoside analog that was synthesized to combine the most favorable pharmacological properties of its congener fludarabine and cladribine. It acts as a strong inhibitor of DNA polymerase, DNA synthesis and ribonucletide reductase (R1/R2). Like other nucleoside analogues, clofarabine must be activated within cells to the 5′-triphodphate form initially by deoxycytidine kinase (dCK). In this study, the anticancer effect of clofarabine was investigated in three solid tumor cell lines, HCT116 (a colon carcinoma cell line), U1810 (a human non-small cell lung carcinoma cell line), and MCF7 (a mammary carcinoma cell line) and compared to four leukemic cell lines, CCRF-CEM, HL60, K562 and MOLT4. Clofarabine was investigated to enter into HCT116, U1810, and MCF7 in a clearly detectable manner by LigandTracer® White. At 10 µM a high signal is achieved and approaches equilibrium after 1∼2 hrs. Pre-incubation with 10 µM clofarabine resulted in rapid inhibition of thymidine incorporation and DNA synthesis. Induction of apoptosis by clofarabine was cell line dependent, but the average apoptotic level was higher in leukemic cell lines than solid tumor cell lines; however it seemed the sensitivity for apoptosis of cell line is correlated with the activity of dCK and thymidine kinase 1 (TK1) in the solid tumor cell lines. Upon treatment with 10 μM clofarabine for 24 hrs, apoptosis induction increased compared to untreated cells 3.1-, 2.9- and 2.1-fold; the activity of dCK 7.3-, 3.7- and 3.1-fold and the activity of TK1 4.2-, 1.4- and 1.2-fold in U1810, HCT116 and MCF7 cells, respectively. In general, the expression of R1, dCK and TK1 in leukemic cells was higher than in solid tumor cells, but the expression of R2, p53R2 and p53 varied among the cell lines. However, there was an inverse correlation between induction of apoptosis at 100nM clofarabine (upon 24 hours incubation) and the expression of p53R2 (spearman rank test −0.67, p=0.1). In addition, clofarabine treatment causes retardation in S-phase, which seems to be related to the activity of TK1 and the protein expression of R2 in U1810, HCT116 and MCF7 cells. This study is a first step to extend the application of clofarabine toward the solid tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3654.
Abstract 3655: Metronomic chemotherapy with cyclophosphamide (Cy) and celecoxib (Cel) in murine mammary adenocarcinoma tumor models: Mechanism of action
Authors: Mainetti Leandro E. , Rozados Viviana R. , Rossa Ana , Roggero Eduardo , Rico María J. , Gervasoni Silvia I. , Giordano Ricardo , Scharovsky O. Graciela
Published: February 2011

Metronomic chemotherapy (MCT) refers to the chronic, equally spaced, delivery of low doses of different chemotherapeutic drugs, without extended interruptions. CY, administered in low dose, is able to modulate the immune response. It can also reduce tumor angiogenesis, like the COX-2 inhibitor CEL does. We have already demonstrated that the administration, as an intervention strategy, of metronomic CY + CEL in M-406 and M-234p mammary adenocarcinomas-bearing mice inhibited tumor growth, increasing the survival rate. Our present aim was to study the mechanism by which that therapeutic effect was achieved. CBi and Balb/c inbred mice were s.c. challenged on day 0 with M-406 or M-234p mammary tumors, respectively, and treated from day 8 (M-406) and 5 (M-234p) on with: I) no further treatment (controls); II) CY in the drinking water (25-30 mg/kg body weight/day); III) CEL (30 mg/kg p.o.), five times/week; IV) Treated as II + III. Blood samples were taken on days 0, 20 (M-406) and 30 (M-324p) to evaluate VEGF serum level determined by ELISA and the % of circulating regulatory T cells (Tregs) by flow cytometry. The same tumor challenges and therapeutic schedules were performed in nude mice. Tumors were excised on days 20 and 30 for M-406 and M-234p, respectively, fixed and paraffin embedded for detection by immunohistochemistry of CD34 and Ki67 molecules. The VEGF serum level of M-406-bearing control animals (group I) was, on day 20, significantly higher than that measured on day 0 (p<0,001), while groups II, III y IV showed, on day 20, lower values than group I (p<0.01) in the same day, and similar than those on day 0. The % of CD4+ CD25+ Foxp3+ Treg cells in serum showed no differences between days 0 and 20 in all the experimental groups. The antitumoral effect of MCT was the same in euthymic and athymic mice. Immunohistochemistry for CD34 showed slight differences between control and treated animals that were not significant. Also, the staining for the proliferation marker Ki67 did not evinced differences among the experimental groups. The results obtained with M-234p tumor model were similar to those of M-406. In conclusion, the antitumor effect of MCT with CY plus CEL in mammary tumor models would be achieved, at least in part, by the inhibition of tumor angiogenesis, without modulation of the antitumor immune response.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3655.
Abstract 3657: 6-(Het)aryl-7-Deazapurine Ribonucleosides as Potent Novel Cytostatic Agents that Interfere with RNA Synthesis
Authors: Dzubak Petr , Nauŝ Petr , Pohl Radek , Votruba Ivan , Ameral Ria , Birkus Gabriel , Wang Ting , Mackman Richard , Ray Adrian , Cihlar Tomas , Hocek Michal , Hajdúch Marián
Published: February 2011

A series of 33 7-deazapurine ribonucleosides bearing an alkyl, aryl or hetaryl group in position 6 and H, F or Cl atom in position 7 was successfully synthesized. Significant cytostatic effects against both hematological and solid tumor cell lines were found for most of these compounds under in vitro conditions. The most potent were 7-H and 7-F derivatives of 6-furyl-/ 6-thienyl-7-deazapurine ribosides displaying nanomolar GIC50 values (comparable to gemcitabine or clofarabine) in several cancer cell lines. Intracellular phosphorylation to mono- and tri-phosphates and consequent inhibition of RNA synthesis, but not DNA synthesis, were demonstrated in preliminary studies of metabolism and mechanism of action. One of the most active compounds (6-thienyl-7-F-7-deazapurine riboside) was tested for in vivo toxicology in mice, and dose limiting toxicities were weight lost and leucopenia. However, the compound demonstrated highly significant antitumor activity at doses equivalent to MTD and ½ MTD, and afforded the extension of survival in treated mice bearing P388D1 leukemia (p<0.001 compared to untreated animals). Antitumor effect was comparable to doxorubicin in P388D1 model. Interestingly, the intraperitoneal administration of the compound resulted in good local control of intraperitoneally transplanted tumors. Supported by grants awarded by the Czech Ministry of Education (MSM 6198959216 and LC07017).

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 3657.
Abstract 3662: Melatonin inhibits cell proliferation and induces apoptosis in human acute leukemia T and lymphoma B cells via a receptor independent mechanism
Authors: Sanchez-Hidalgo Marina , Lee Melanie , de la Lastra Catalina Alarcón , Guerrero Juan M. , Packham Graham
Published: February 2011

Loss of melatonin (MLT) production due to age contributes to immunosenescence onset and age-related diseases such as cancer. MLT reduces proliferation in many different cancer cell lines. However, MLT effects on subjects with immune cell cancer are controversial, and little is known about its action mechanisms in human malignant lymphoid cells. In the current study, we examined the effect of different doses of MLT (0.5, 1 and 2 mM) administration on human acute leukemia T cells (Jurkat), Burkitt (Ramos), diffuse large B cell lymphoma (Sudhl4) and follicular B no-Hodgkin lymphoma (Dohh2) cells analyzing cell cycle arrest, apoptosis, caspase cascade activation pathway and the role of melatonin transmembrane receptors. The data showed that high concentrations (millimolar) of melatonin significantly inhibited progression from G1 to S phase of the cell cycle and increased apoptotic cell death via caspase cascade activation and poly-ADP-ribose polymerase (PARP) and caspase-3 and −9 cleavages in all cell lines studied. There was also evidence of caspase-8 activation in Ramos and Sudhl4 cells. Reduced levels of procaspase-8 were detected in Dohh2 cells after melatonin treatment, thus evidence of activation even though the cleaved fragment was not detected. Interestingly, downregulation of Bcl-2 and Bcl-XL expression was also observed in melatonin treatment in Dohh2 cells whereas Mcl-1 antiapoptotic protein expression remained unchanged in each malignant cell line studied. A significant decrease in mitochondrial membrane potential (ΔΨm) was also detected in Ramos cells. Finally, using luzindole (LZD), as a MT1 and MT2 melatonin membrane receptor antagonist, we observed that MLT inhibited cell proliferation and apoptotic cell death by means of a melatonin membrane receptor independent pathway. This novel finding shows that a nontoxic natural indoleamine may be a potential therapy for some types of human lymphoid malignancies.

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 3662.
Abstract 367: Identification of tumor cell-derived factors associated with resistance to anti-VEGF therapy in non-small cell lung cancer
Authors: Sullivan Laura A. , Carbon Juliet G. , Toombs Jason , Xie Yang , Minna John D. , Brekken Rolf A.
Published: February 2011

Vascular endothelial growth factor-A (VEGF) functions during physiological and pathological angiogenesis. Strategies that inhibit VEGF are now an established anti-cancer paradigm. We have developed a human monoclonal antibody (mAb), r84 that recognizes mouse and human VEGF and specifically blocks VEGF binding and activation of VEGFR2 but does not interfere with VEGF:VEGFR1 interaction. Therefore, r84 allows for the functional evaluation of tumor cell and stromal cell derived VEGF-induced activation of VEGFR2 signaling in tumor progression and provides a useful comparison to other anti-VEGF strategies in tumor xenograft models. Bevacizumab (bev) is a humanized mAb that binds human VEGF and prevents VEGF-induced activation of VEGFR1 and VEGFR2. The FDA has approved bev for the treatment of several tumor types, including non-squamous non-small cell lung cancer (NSCLC). Given the cost, toxicities and great variation in patient response to bev, it would be desirable to have biomarkers that predict response and allow for “personalized” treatment. The variation of NSCLC response to anti-VEGF therapy suggests a complex mechanism of drug action within tumors and indicates that some tumors may be intrinsically resistant to bev. Therefore, successful use of anti-VEGF therapy may rely on a better understanding of genes associated with responsiveness or resistance. To evaluate intrinsic resistance to anti-VEGF therapy we have performed an in vivo screen of a panel of NSCLC xenografts (n=11) for response to anti-VEGF therapy (bev or r84). We identified NSCLC xenografts that are very sensitive, moderately sensitive or display intrinsic resistance to bev or r84 therapy with final tumor weight treatment/control (T/C) ratios ranging from 0.1 to 0.65. Sensitive tumors had low T/C ratios and relevant vascular changes including decreased tumor microvessel density, lymphatic vessel density, and VEGFR2 expression. Microarray analysis of sensitive and resistant cell lines have been used to generate a preliminary gene signature that may mark relevant tumor cell derived genes associated with response to anti-VEGF treatment. Validation of this signature is currently underway. We conclude that NSCLC xenografts can exhibit dramatically different response phenotypes to anti-VEGF mAbs. A better understanding of the mechanisms of response to anti-VEGF therapy may lead to the development of more effective therapies and could ultimately assist in the identification of patients that would benefit the most from anti-VEGF strategies.

Supported by the Randall Bridwell Lung Cancer Research Grant from Uniting Against Lung Cancer and Lung Cancer SPORE P50CA70907.

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 367.