bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5354: The guilty bystander model for electrochemotherapy of cancer: Verification in vitro
Authors: Lambert W Clark , Tsongalis Gregory J. , Gagna Claude E. , Lambert Muriel W.
Published: August 2015

Electroporation (EPR) is the induction of pores in cellular membranes in cells by application of a short application of a high voltage, low amperage electric current. If the cells are immersed in a solution containing a large molecule which is normally impervious to this membrane, it may be introduced into the cells in this manner. Electrochemotherapy (ECT) is a clinical procedure in which a drug, such as bleomicin or cisplatin, is introduced into the blood of or intratumorally into an animal or human subject by intravenous injection followed by application of a short high dose, low amperage electric current into a neoplasm after a delay enabling the drug to reach the neoplasm through the bloodstream or by other means. ECT is normally used for palliative care in human subjects, however there are now a number of reports, numbering well over 10, of unexpected complete clearance of neoplasms following ECT. Many of these reports have come from university centers in Europe and elsewhere. Although ECT is not yet approved in the USA, we believe that these reports can no longer be ignored. We and others have used EPR in culture to introduce large molecules into cells for experimental purposes for decades. Even under the controlled experimental conditions we have employed, however, the proportion of cells into which these molecules have been introduced has been well below 100 per cent. To account for the clearance of some tumors by ECT, in which the proportion of cells in which drug is introduced would be expected to be lower than for EPR, we have proposed a “Guilty Bystander Model”, in which cells adjacent to neoplastic cells into which drug has been introduced are also cleared. To test this model in vitro, we optimized conditions for EPR but were unable to produce more than 88 per cent introduction of labeled large molecules into lymphoblastoid cells (GM 1989A, derived from a healthy 33 year old Caucasian male, and GM 3999, derived from a healthy 8 year old Caucasian female), even under conditions which reduced cell viability from 99 per cent to 79 per cent. Increasing voltage produced a marked loss of viability. We conclude that EPR does not produce 100 per cent introduction of large molecules into human cells, even under optimized conditions in vitro, and that there must remain neoplastic cells in tumors treated by ECT that are eliminated by means other than introduction of the drug.

Citation Format: W Clark Lambert, Gregory J. Tsongalis, Claude E. Gagna, Muriel W. Lambert. The guilty bystander model for electrochemotherapy of cancer: Verification in vitro. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5354. doi:10.1158/1538-7445.AM2015-5354
Abstract 5359: Antitumor efficacy of EC1456 in patient derived xenograft models of ovarian, endometrial, NSCLC and TNBC
Authors: Reddy Joseph A. , Bloomfield Alicia , Taylor Christina , Hargett Katherine , Nelson Melissa , Leamon Christopher
Published: August 2015

EC1456, a potent folate-targeted tubulysin conjugate, is currently being evaluated in a phase 1 dose escalation study in patients with advanced solid tumors. To assist in upcoming cancer indication selection decisions for advanced trials, we tested EC1456 against eight patient-derived xenograft (PDX) cancer models. Thus, EC1456 was evaluated as a single agent or in combination with standard of care (SOC) treatments against two folate receptor (FR)-positive models each of ovarian, endometrial, non-small cell lung (NSCLC) and triple negative breast (TNBC) tumor models. Nude mice engrafted subcutaneously with PDX tumors received 4 μmol/kg/week EC1456 for two weeks, following a once a week (SIW) or two times a week (BIW) schedules, either alone or in combination with 15 mg/kg, SIW x 2 Paclitaxel (ovarian and endometrial), 1 mg/kg SIW x 2 eribulin mesylate (TNBC) or 15 mg/kg, SIW docetaxel (NSCLC). EC1456 alone demonstrated tumor growth suppression or stasis in most of the tumor models and significantly greater anti-tumor activity (including cures) with no detectable increase in toxicity when tested in combination with the SOC agents. Overall, EC1456 significantly augmented the growth inhibitory effects of SOC treatments against all four PDX cancer indications examined, thus lending support to future clinical development of this novel FR-targeted agent.

Citation Format: Joseph A. Reddy, Alicia Bloomfield, Christina Taylor, Katherine Hargett, Melissa Nelson, Christopher Leamon. Antitumor efficacy of EC1456 in patient derived xenograft models of ovarian, endometrial, NSCLC and TNBC. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5359. doi:10.1158/1538-7445.AM2015-5359
Abstract 5361: In vitro results and electric fields simulations suggest Tumor Treating Fields (TTFields) to be an effective treatment against Mesothelioma
Authors: Giladi Moshe , Munster Mijal , Blat Roni , Schneiderman Rosa , Porat Yaara , Bomzon Zeev , Urman Noa , Voloshin Tali , Kirson Eilon David , Weinberg Uri , Palti Yoram
Published: August 2015

Mesothelioma is an aggressive cancer affecting the membrane lining of the lungs and abdomen, and is associated with poor prognosis. Current available treatment options for Mesothelioma, including surgery, radiation therapy and chemotherapy, offer only modest improvement in survival, therefore emphasizing the desperate need for new treatment options. Tumor Treating Fields (TTFields) is a treatment modality based on non-invasive application of low intensity intermediate frequency alternating electric fields that disrupt mitosis.

The goal of the present work was to determine whether TTFields in combination with standard of care drugs, could serve as an effective treatment against Mesothelioma.

TTFields were applied to Mesothelioma cell cultures using the inovitro system. Cytotoxicity and effect on clonogenicity were determined following 72 hours of TTFields treatment. Combination index (CI) was calculated for the treatment of TTFields with each of the following drugs: Paclitaxel, Cisplatin, Pemetrexed and Vinorelbine. Finite Element Mesh (FEM) simulations were performed using the Sim4life software package (ZMT, Zurich, Switzerland) for the calculations of the electric fields intensities in the mesothelium of a realistic human computational phantom.

Frequency titration experiments revealed 150 kHz to be the optimal TTFields frequency for Mesothelioma cell cultures, leading to 69% reduction in the number of cells (p<0.001) and 78% reduction in clonogenicity (p<0.05) as compared to control. The combined treatment of TTFields with Paclitaxel, Cisplatin, Pemetrexed and Vinorelbine resulted in combination index of 0.99, 0.88, 1.29 and 1.13, respectively. FEM simulations demonstrated that electric field intensities within the mesothelium are approximately 2 V/cm RMS which is above the minimal threshold required for TTFields effect.

Taken together, these results suggest that combining TTFields therapy with systemic chemotherapy may be an approach that offers enhanced treatment efficacy for Mesothelioma. A clinical study testing the combined efficacy of TTFields Cisplatin and Pemetrexed is under development.

Citation Format: Moshe Giladi, Mijal Munster, Roni Blat, Rosa Schneiderman, Yaara Porat, Zeev Bomzon, Noa Urman, Tali Voloshin, Eilon David Kirson, Uri Weinberg, Yoram Palti. In vitro results and electric fields simulations suggest Tumor Treating Fields (TTFields) to be an effective treatment against Mesothelioma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5361. doi:10.1158/1538-7445.AM2015-5361
Abstract 5366: Propentofylline inhibits TROY/TNFRSF19 signaling to enhance therapeutic efficacy in invasive glioblastoma cells
Authors: Dhruv Harshil D. , Tuncali Serdar , Roos Alison , Tomboc Patrick , Jameson Nathan , Chavez Ashley , Loftus Joseph , Berens Michael E. , Tran Nhan L.
Published: August 2015

Glioblastoma (GBM) is the most common primary tumor of the CNS and carries a dismal prognosis. The aggressive invasion of GBM cells into the surrounding normal brain makes complete resection impossible, significantly increases resistance to the standard therapy regimen, and virtually assures tumor recurrence. Median survival for newly diagnosed GBM is 14.6 months and declines to 8 months for patients with recurrent GBM. New therapeutic strategies that target the molecular drivers of invasion are required for improved clinical outcome. We have demonstrated that TROY (TNFRSF19), a member of the TNFR super-family, plays an important role in GBM invasion and resistance. TROY expression increases with glial tumor grade and inversely correlates with patient survival. TROY stimulates GBM cell invasion and increases resistance to temozolomide (TMZ) and radiation treatment. Conversely, knockdown of TROY expression inhibits GBM cell invasion, increases sensitivity to temozolomide, and prolongs survival in an intracranial xenograft model. Propentofylline (PPF), an atypical synthetic methylxanthine compound, has been extensively studied in Phase II and Phase III clinical trials for Alzheimer's disease and vascular dementia where it has demonstrated blood-brain permeability and minimal adverse side effects. In this study, we demonstrated that PPF decreases TROY protein expression in glioma cells, and subsequently suppress activation downstream signaling effectors including AKT, NF-κB, and Rac1. PPF treatment of glioma cells also suppressed glioma cell migration and invasion, demonstrated by transwell migration assay and reduced membrane ruffling. Finally, PPF treatment increased vulnerability of glioma cells to Temozolomide (TMZ) and radiation. In summary, this study demonstrates that PPF provides a pharmacologic approach to target TROY to inhibit glioma cell invasion and reduce therapeutic resistance to TMZ and radiation.

Citation Format: Harshil D. Dhruv, Serdar Tuncali, Alison Roos, Patrick Tomboc, Nathan Jameson, Ashley Chavez, Joseph Loftus, Michael E. Berens, Nhan L. Tran. Propentofylline inhibits TROY/TNFRSF19 signaling to enhance therapeutic efficacy in invasive glioblastoma cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5366. doi:10.1158/1538-7445.AM2015-5366
Abstract 5368: Metarrestin effectively disassembles PNCs and inhibits metastasis
Authors: Wang Chen , Frankowski Kevin , Yaroslav Teper , Patnaik Samarjit , Schoenen Frank , Southall Noel , Sun Wei , Titus Steve , Griner Lesley , Dextras Christopher , Sultan Jamey , Kandela Irawati , Lewandowska Marzena , Wen Yi-Ping , Norton John , Kang Jin Sol , Mazar Andrew , Zhang Wei , Aubé Jeffrey , Ferrer Marc , Rudloff Udo , Marugan Juan Jose , Huang Sui
Published: August 2015

The perinucleolar compartment (PNC) is a nuclear body that forms specifically in highly malignant cancer cells. PNC prevalence is shown to reflect metastatic potential of cancer cells from solid tissue origins. A high-content screen was performed to identify small molecules that reduce PNC prevalence. Medicinal chemistry optimizations of confirmed hits yield metarrestin, having 95% of oral bioavailability and a PNC disassembly IC50 between 100 to 400 nM across multiple cancer cell lines. Metarrestin inhibits soft agar growth and the invasive capabilities of cancer cells in vitro. In vivo mouse xenograft metastatic studies showed that metarrestin effectively inhibits lung and/or liver metastasis of prostate cancer (PC3M), pancreatic cancer (organotypic spheroids of PANC1), and a breast cancer patient xenograft (PDX) model derived from metastatic cells isolated from a pleural effusion. Treated animals tolerate metarrestin well with no discernable impact on animal well-being including weight compared with control animals. These data supports the idea that the pharmacological disassembly of PNC produces the inhibition of malignant behavior of cancer cells both in vitro and in vivo. Cellular and molecular analyses of the mechanisms of action of metarrestin show that this compound induces significant and reversible changes in the nucleolar structure, accompanied by a selective reduction of pol I transcription. These studies suggest that PNC structural integrity directly associates with pol I function and nucleolar structure. Experiments are currently underway to identify the cellular factors and pathways by which metarrestin reduces PNCs, disrupts nucleolar structure, regulates transcriptions, and blocks malignant behaviors.

Citation Format: Chen Wang, Kevin Frankowski, Teper Yaroslav, Samarjit Patnaik, Frank Schoenen, Noel Southall, Wei Sun, Steve Titus, Lesley Griner, Christopher Dextras, Jamey Sultan, Irawati Kandela, Marzena Lewandowska, Yi-Ping Wen, John Norton, Jin Sol Kang, Andrew Mazar, Wei Zhang, Jeffrey Aubé, Marc Ferrer, Udo Rudloff, Juan Jose Marugan, Sui Huang. Metarrestin effectively disassembles PNCs and inhibits metastasis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5368. doi:10.1158/1538-7445.AM2015-5368
Abstract 5376: Identification of the first small molecule PRMT6 inhibitor tool compound
Authors: Drew Allison , Mitchell Lorna , Rioux Nathalie , Swinger Kerren , Ribich Scott , Jacques-O'hagan Suzanne , Lingaraj Trupti , Wigle Tim , Riera Tom , Chesworth Richard , Smith Jesse
Published: August 2015

PRMT6 is a nuclear-localized arginine methyl transferase (RMT) capable of adding omega-N(G)-monomethylarginine and asymmetric omega-N(G),N(G)-dimethylarginine derivatives to histone and other protein substrates containing a GAR motif. PRMT6 can methylate both histone and non-histone substrates and is the only RMT known to methylate the H3R2 mark. This mark can act in opposition to the activating H3K4me3 mark, effectively acting as a transcriptional repressor.

PRMT6 overexpression has been reported in several cancer types including melanoma and bladder, lung, and prostate carcinoma suggesting that inhibition of PRMT6 activity might have therapeutic utility. However, until now no small molecule PRMT6 inhibitor has been available to carry out in vitro and in vivo studies.

Here we report identification of an aryl pyrazole as the first small molecule PRMT6 inhibitor. This tool compound has an IC50 of 10nM in a PRMT6 biochemical assay and the binding mode has been elucidated through crystallography. The compound inhibited methylation of H3R2 in a dose dependent manner in an engineered model in which PRMT6 was transiently expressed in the A375 cell line. The tool compound showed a half-life (t½) of 8.54 ± 1.43 h after a 1 mg/kg intravenous administration to rats and bioavailability of 65.6 ± 4.3% for a 5 mg/kg subcutaneous dose, enabling its use in further in vitro and in vivo target validation.

Citation Format: Allison Drew, Lorna Mitchell, Nathalie Rioux, Kerren Swinger, Scott Ribich, Suzanne Jacques-O'hagan, Trupti Lingaraj, Tim Wigle, Tom Riera, Richard Chesworth, Jesse Smith. Identification of the first small molecule PRMT6 inhibitor tool compound. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5376. doi:10.1158/1538-7445.AM2015-5376
Abstract 5388: Methionine S-adenosyltransferase 2A (MAT2A) inhibitors for cancer treatment
Authors: Zhang Wen , Sviripa Vitaliy , Watt David , Liu Chunming
Published: August 2015

Methionine S-adenosyltransferase 2A (MAT2A) is the catalytic subunit for synthesis of S-adenosylmethionine (SAM), the principal methyl donor in many biological processes. MAT2A is up-regulated in many cancers, including liver cancer and colorectal cancer (CRC) and is a potentially important drug target. In normal liver, methionine S-adenosyltransferase 1 (MAT1), but not MAT2A is expressed. MAT2A is induced in liver cancer. We developed a family of fluorinated N,N-dialkylaminostilbene agents, called FIDAS agents, that inhibit the proliferation of CRC cells and liver cells in vitro and in vivo. Using a biotinylated FIDAS analog, we identified the catalytic subunit of MAT2A as the direct and exclusive binding target of these FIDAS agents. FIDAS agents do not bind MAT1A. MAT2B, an associated regulatory subunit of MAT2A, binds indirectly to FIDAS agents through its association with MAT2A. FIDAS agents inhibited MAT2A activity in SAM synthesis, and depletion of MAT2A by shRNAs inhibited CRC and liver cancer cell growth. FIDAS agents also inhibited specific histone methylation and repressed transcription of c-myc and other oncogenes. FIDAS agents delivered orally repressed CRC xenografts in athymic nude mice. Based on SAR and docking studies, new generations of FIDAS agents were synthesized and tested. An active in vivo metabolite of FIDAS was identified. PK/PD studies were performed. Our findings suggest that FIDAS analogs targeting MAT2A represent a family of novel and potentially useful agents for cancer treatment.

Citation Format: Wen Zhang, Vitaliy Sviripa, David Watt, Chunming Liu. Methionine S-adenosyltransferase 2A (MAT2A) inhibitors for cancer treatment. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5388. doi:10.1158/1538-7445.AM2015-5388
Abstract 5394: First-in-class dual PIM/FLT3 kinase inhibitor SEL24-B489 for the treatment of hematological malignancies
Authors: Brzózka Krzysztof , Czardybon Wojciech , Gołas Aniela , Windak Renata , Gałęzowski Michał , Gabor-Worwa Ewelina , Winnik Bożena , Przybyłowicz Agnieszka , Szydłowski Maciej , Białopiotrowicz Emilia , Sewastianik Tomasz , Mądro Elżbieta , Lech-Marańda Ewa , Warzocha Krzysztof , Juszczyński Przemysław
Published: August 2015

Despite huge effort spent on understanding acute myeloid leukemia (AML), current standards of care are still based on the same chemotherapy agents as two decades ago - typically based on cytarabine chemotherapy with an anthracycline. Although patients aged <60 years may achieve remission to induction therapy, most patients will relapse. One of primary reasons for that is high heterogeneity of the disease. Recently one of the most extensively investigated approaches to the treatment of AML, are targeted therapies, especially inhibition of certain kinases. Probably mostly investigated up to date are inhibitors of FLT3 with most advanced clinical compound - AC220 (Quizartinib). FLT3 inhibitors are very effective in killing cancer lines with a major drawback of the therapy being fast development of resistance.

PIM kinases are expressed in various cancers including AML but also other liquid tumors as well as in some solid tumors. Inhibition of PIM kinases which are downstream in the FLT3 signaling cascade have already shown influence on cancer cell survival. In addition there are great hopes related to the fact that PIM kinases were shown to contribute in resistance to FLT3 inhibitors. This leads to the conclusion that combined inhibition of PIM and FLT3 may be a rational strategy.

Selvita has developed series of inhibitors combining activity against PIM and FLT3 kinases. Selected clinical candidate SEL24-B489 - has shown excellent anticancer efficacy. It is active against broad panel of AML cell lines and primary blasts, but also against other hematological cancers: DLBCL, CLL, HL - both in vitro on cell lines, patient samples and in vivo. Head to head comparison of SEL24-B489 with PIM (AZD1208) and FLT3 (AC220) inhibitors currently in clinical development will be presented. Comparison shows strong activity of SEL24-B489 against broader panel of cell lines in vitro and in vivo (in xenograft models) than AZD1208 or AC220. SEL24-B489 is currently in preclinical development and details of toxicology profile will also be discussed.

Citation Format: Krzysztof Brzózka, Wojciech Czardybon, Aniela Gołas, Renata Windak, Michał Gałęzowski, Ewelina Gabor-Worwa, Bożena Winnik, Agnieszka Przybyłowicz, Maciej Szydłowski, Emilia Białopiotrowicz, Tomasz Sewastianik, Elżbieta Mądro, Ewa Lech-Marańda, Krzysztof Warzocha, Przemysław Juszczyński. First-in-class dual PIM/FLT3 kinase inhibitor SEL24-B489 for the treatment of hematological malignancies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5394. doi:10.1158/1538-7445.AM2015-5394
Abstract 5398: In vivo development of pan-Pim kinase small molecule inhibitors
Authors: Mattson Bethany , Sastri Christine. E. , Guerrero Nadia , Hickman Dean , Chen Jie , Wu Tian , Wang Hui-Ling , Taskar Andrew , Lanman Brian , Reed Anthony B. , Canon Jude , Lipford J. Russell , Rex Karen
Published: August 2015

Pim-1,-2, and -3 are constitutively active serine-threonine kinases which are partially redundant and regulate multiple pathways important for tumor growth and survival. One or more of the human Pims are over-expressed in multiple hematological tumor types (e.g. multiple myeloma (MM), NHL and AML) and in some solid tumors (e.g. prostate and SCLC). Pim over-expression correlates with malignancy and poor prognosis in several indications. Our goal was to generate a pan Pim kinase inhibitor with acceptable physical chemical properties and in vivo anti-tumor efficacy. Here we present data on two ATP-competitive, orally bioavailable pan Pim inhibitors, Compound I and Compound II. These inhibitors have potent enzymatic and cellular activity, acceptable pharmacokinetic properties (PK) and robust in vivo efficacy. In a kinase enzyme assay Compound I inhibits Pim-1 and Pim-2 activity with 0.4 nM and 0.7 nM IC50s, respectively, while Compound II is even more potent with Pim-1 and Pim-2 IC50s of 0.1 nM/0.1 nM. In a cellular assay which measures inhibition of the Pim downstream substrate phospho-BAD (p-BAD), compounds I and II demonstrate IC50s of 56 and 16 nM, respectively. In an in vivo pharmacodynamic assay (PD) to demonstrate on-target Pim activity, compounds I and II significantly inhibited p-BAD in KMS-12-BM multiple myeloma tumors for 16 hours post dose. Treatment of KMS-12-BM tumor xenografts with Compound I demonstrated robust in vivo anti-tumor efficacy resulting in 23% tumor regression at 50 mg/kg BID and tumor stasis at 100 mg/kg QD. Compound II demonstrated improved PK properties leading to greater anti-tumor efficacy of 33% tumor regression at 100 mg/kg QD and tumor stasis at 50 mg/kg QD. Compound II showed efficacy in an orthotopic model of multiple myeloma and in models of AML and DLBCL. Combination treatment of Compound II and the standard of care Dexamethasone in the multiple myeloma RPMI-8226 xenograft model demonstrated enhanced tumor growth inhibition compared to either single agent activity. In summary, Compound I and II are potent and selective inhibitors of Pim kinases with excellent in vivo properties. Pim kinase inhibitors, either as monotherapy or in combination with dexamethasone, may be effective clinical strategies for certain cancer patients.

Citation Format: Bethany Mattson, Christine. E. Sastri, Nadia Guerrero, Dean Hickman, Jie Chen, Tian Wu, Hui-Ling Wang, Andrew Taskar, Brian Lanman, Anthony B. Reed, Jude Canon, J. Russell Lipford, Karen Rex. In vivo development of pan-Pim kinase small molecule inhibitors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5398. doi:10.1158/1538-7445.AM2015-5398
Abstract 54: Ezrin enhances signaling and nuclear translocation of the epidermal growth factor receptor in non-small cell lung cancer cells
Authors: Saygideger Kont Yasemin , Celik Haydar , Erkizan Hayriye V. , Minas Tsion , Han Jenny , Toretsky Jeffrey , Uren Aykut
Published: August 2015

The cytoskeletal cross linker protein ezrin is a member of the ezrin-radixin-moesin (ERM) family and plays important roles not only in cell motility, cell adhesion, and apoptosis, but also in various cell-signaling pathways. Ezrin interacts with EGFR in the cell membrane and involves in cell motility events, but little is known about the effects of this interaction on the EGFR signaling pathway. We investigated the role of Ezrin in EGFR signaling and nuclear trafficking in non-small cell lung cancer (NSCLC) cell lines. The ligand induced interaction between Ezrin and EGFR was evaluated by immunoprecipitation (IP) and immunofluorescence (IF) in H292 and A549 cells. Ezrin levels were reduced using siRNA in these two cell lines. Downstream signaling protein phosphorylation and nuclear localization of EGFR were detected after EGF treatment. Expressions of nuclear EGFR target genes were evaluated by qPCR. Endogenous Ezrin was found in a complex with EGFR in IP and IF. When Ezrin protein expression was inhibited, phosphorylation levels of EGFR at Y1068, Y1101 and Y845 were reduced as well as phosphorylation levels of downstream signaling pathway proteins ERK and STAT3. Cell fractionation revealed that EGFR nuclear translocation after EGF treatment significantly reduced in Ezrin-knockdown cells. Further, mRNA levels of EGFR target genes AuroraK-A, COX2, Cyclin D1 and iNOS were decreased in Ezrin-knockdown A549 cells. Small molecule ezrin inhibitors showed strong synergy with EGFR inhibitors in cytotoxicity assays. These results suggest that Ezrin has a role as an enhancer in the EGFR pathway and targeting ezrin may potentiate anti-EGFR based therapies in NSCLC.

Citation Format: Yasemin Saygideger Kont, Haydar Celik, Hayriye V. Erkizan, Tsion Minas, Jenny Han, Jeffrey Toretsky, Aykut Uren. Ezrin enhances signaling and nuclear translocation of the epidermal growth factor receptor in non-small cell lung cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 54. doi:10.1158/1538-7445.AM2015-54
Abstract 540: A MALDI imaging mass spectrometry approach using tissue microarrays to identify an N-glycan biomarker panel for pancreatic cancers
Authors: Drake Richard R. , Powers Thomas W. , Neely Benjamin A.
Published: August 2015

We have recently developed a MALDI imaging mass spectrometry (MALDI-IMS) method to spatially profile N-linked glycans in frozen and formalin-fixed paraffin-embedded (FFPE) tissue sections and tissue microarrays (TMAs). Tissues are incubated with peptide N-glycosidase, and released N-glycans are detected directly using MALDI-FTICR, linked directly with tissue histopathology. Other methods to detect the localization of glycans in tissues rely on detection of broader glycan structural motifs (i.e., lectins or carbohydrate antigen antibodies), whereas our method is able to simultaneously identify and distinguish 40 or more components of the N-glycome on a single slide. To demonstrate the ability of MALDI-IMS to generate biomarker panels, pancreatic cancer tissue blocks and six TMAs containing matched tumor and non-tumor regions from over 70 patients were profiled. Aberrant glycosylation, such as elevated CA-19-9, is well documented in pancreatic cancer, making this an ideal sample set. To best analyze this complex data set, we developed an in-house analysis script that can extract spectra from tissue cores, and return non-biased statistics for observed glycan ions. Panels were generated using a training set of data and tested on external validation data set. The most accurate panel of 12 glycans achieved an overall sensitivity of 92.9% and specificity of 86.7%. Structural identification of N-glycans has been confirmed by techniques such as on-tissue CID, ethylation analysis, sequential glycosidase digestions, and comparison to glycan structural databases. Furthermore, imaging of entire FFPE sections matched with histological analysis revealed a vast diversity of N-glycan localizations, which could distinguish not only tumor from normal tissue, but also regions of pancreatitis and pancreatic intraepithelial neoplasia. The generated glycan tissue maps will also be used to determine specific glycoprotein carriers of biomarker candidate glycans in tissue and biofluid samples.

Citation Format: Richard R. Drake, Thomas W. Powers, Benjamin A. Neely. A MALDI imaging mass spectrometry approach using tissue microarrays to identify an N-glycan biomarker panel for pancreatic cancers. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 540. doi:10.1158/1538-7445.AM2015-540
Abstract 5401: TGF-β signaling inhibition as a potential approach to target suboptimally debulked ovarian tumors
Authors: Wei Wei , Birrer Michael J.
Published: August 2015

The current standard of care for all patients with high-grade, advanced-stage serous ovarian cancer is surgery followed by adjuvant chemotherapy. Resection of all visible residual disease to less than 1 cm is associated with a favorable prognosis, but this optimal debulking can only be achieved in ∼70% of patients. Patients with suboptimal cytoreduction are presumed to have lower response rates to subsequent chemotherapy and are known to have a poor prognosis.

Recently, we performed a meta-analysis of 1,525 publicly available expression profiling assays and identified a 200-gene expression signature of primary ovarian tumors to predict the outcome of debulking surgery. The signature was validated using two independent sets of tumors. This debulking signature consists of transcripts positively regulated by TGF-β, indicating that hyperactivation of TGF-β signaling may be responsible of the aggressive biological and clinical behavior of suboptimally debulked tumors.

In this study, we evaluated the potential anti-tumor effect of TGF-β signaling blockade in ovarian cancer cells in vitro and in vivo. Integrated TGF-β signaling was observed in 8 of 10 ovarian cancer cell lines tested by a SMAD-binding element based luciferase assay. The potential anti-tumor effect of a small molecular TGFBR1 kinase inhibitor LY2157299 was further investigated in SKOV3 cell line, which presents both endogenous and inducible TGF-β signaling and is highly disseminative when inoculated into immunodeficient mice. Although imposing minimum effect on in vitro cellular proliferation, LY2157299 significantly inhibited tumor-dissemination related biological processes in SKOV3 cells including migration, invasion and mesothelial cell adhesion. In an interperitoneal SKOV3 xenograft model, administration of LY2157299 significantly increased cisplatin related cytotoxicity and tumor burden reduction. Taken together, our study has provided evidence for the potential clinical use of LY2157299 to improve the treatment and survival of women with suboptimal surgery.

Citation Format: Wei Wei, Michael J. Birrer. TGF-β signaling inhibition as a potential approach to target suboptimally debulked ovarian tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5401. doi:10.1158/1538-7445.AM2015-5401
Abstract 5415: Therapeutic effects of TOPK inhibitor on triple-negative breast cancer
Authors: Park Jae-Hyun , Miyamoto Takashi , Matsuo Yo , Nakamura Yusuke
Published: August 2015

TOPK (T-LAK cell-originated protein kinase) is a novel mitotic kinase and frequently up-regulated in multiple types of cancer, including triple-negative breast cancer (TNBC), but not expressed in normal organs except testis. We previously showed that knockdown of TOPK using siRNAs induced cytokinesis defect in dividing cancer cells with elongated intercellular bridge. In addition, we recently reported that a potent TOPK inhibitor (OTS964) caused the cytokinesis defect and subsequently induced apoptosis of cancer cells in vitro as well as in in vivo xenograft models of human lung cancer. More importantly, liposomal formulation of OTS964 led to complete regression of transplanted tumors without any detectable adverse reactions in mice. Since the TOPK gene is up-regulated in the great majority of TNBCs, we examined therapeutic effect of OTS964 on TNBC cell lines. Western blot analysis revealed that TOPK was highly expressed in all of five TNBC cell lines examined. OTS964 was effective to MDA-MB-231 cells with the IC50 value of 73 nM, and caused cytokinesis defect and apoptosis. Moreover, in MDA-MB-231 xenograft model, oral administration of 75 mg/kg OTS964 for 2 weeks achieved 65% tumor growth inhibitory (TGI) effect at day 15, without any body weight loss. Collectively, our findings imply that targeting TOPK is a very promising therapeutic option for the treatment of TNBC patients who still have very limited treatment modalities.

Citation Format: Jae-Hyun Park, Takashi Miyamoto, Yo Matsuo, Yusuke Nakamura. Therapeutic effects of TOPK inhibitor on triple-negative breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5415. doi:10.1158/1538-7445.AM2015-5415
Abstract 5417: The identification of BMS-595, an orally active imidazo[1,2-b]pyridazine CK2 inhibitor with in vivo anti-tumor activity
Authors: Tarby Christine M. , He Liqi , Fink Brian E. , Nation Andrew , Zhao Yufen , Kim Soong-Hoon , Chen Libing , Tokarski John S. , Yu Chiang , Pabalan Jonathan G. , Roongta Urvashi V. , Lippy Jonathan , Obermeier Mary , Elzinga Paul A. , Fura Aberra , Henley Benjamin , Fargnoli Joseph J. , Foster William R. , Gavai Ashvinikumar V. , Wong Tai W. , Hunt John T. , Vite Gregory D. , Purandare Ashok V. , Rupnow Brent A.
Published: August 2015

CK2 is a highly conserved, and constitutively active family of serine/threonine kinases abnormally elevated in a wide variety of cancers and linked to poor prognosis and disease progression. The enzymes form as hetero-tetrameric complexes comprised of two highly related catalytic subunits (α or α´) with two regulatory β subunits in various combinations and distributions, depending on cell type. While CK2 plays a role in normal growth and development, deregulation of the enzymes has been shown to promote and maintain a malignant phenotype through mechanisms in both the anti-apoptotic and the pro-proliferative signaling pathways. CK2 has been reported to modulate the activity of several oncogenic transcription factors including CREB, Myc, Jun and Fos. Studies with RNAi and small molecule compounds have demonstrated tumor cell dependence on CK2.

We sought to identify potent CK2 inhibitors to probe the function of CK2 in cancer-linked pathways and for evaluation in CK2 dependent tumor xenograft models. Herein we report SAR studies in the imidazo[1,2-b]pyridazine chemotype leading to the discovery of BMS-595, a highly potent and selective ATP-competitive CK2 inhibitor with a commensurate level of cellular potency. BMS-595 demonstrates strong PK/PD correlations and robust, oral anti-tumor efficacy in CK2-driven xenograft models at tolerated doses.

Citation Format: Christine M. Tarby, Liqi He, Brian E. Fink, Andrew Nation, Yufen Zhao, Soong-Hoon Kim, Libing Chen, John S. Tokarski, Chiang Yu, Jonathan G. Pabalan, Urvashi V. Roongta, Jonathan Lippy, Mary Obermeier, Paul A. Elzinga, Aberra Fura, Benjamin Henley, Joseph J. Fargnoli, William R. Foster, Ashvinikumar V. Gavai, Tai W. Wong, John T. Hunt, Gregory D. Vite, Ashok V. Purandare, Brent A. Rupnow. The identification of BMS-595, an orally active imidazo[1,2-b]pyridazine CK2 inhibitor with in vivo anti-tumor activity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5417. doi:10.1158/1538-7445.AM2015-5417
Abstract 5424: Therapeutic targeting of KRAS mutation-driven tumor progression in colorectal cancer
Authors: Wu Christina , Lin Peng-Chan , Chu Po-Chen , Chuang Hsiao-Ching , Kulp Samuel , Bekaii-Saab Tanios , Chen Ching-Shih
Published: August 2015

KRAS is the most frequently altered gene in colorectal cancer (CRC), with mutations occurring in 30-40% of colorectal cancer. Although KRAS mutation is associated with poor prognosis and resistance to anti-epidermal growth factor receptor therapy, the mechanism by which it promotes tumor metastasis remains undefined. Our study explored a new hypothesis that targeting the Y-box binding protein-1 (YB-1)-insulin-like growth factor-I receptor (IGF-IR) signaling axis which is downstream of KRAS, with a novel integrin-linked kinase inhibitor, T315, represents a therapeutically relevant strategy to block KRAS mutation-driven tumor progression. In liver metastases from CRC patients, we observed that there was a preponderance (79/108) of over-expression in both YB-1 and IGF-1R by immunohistochemistry. In colon cancer cell lines HCT-116 and SW480, knockdown and over-expression of KRAS affected the protein and mRNA levels of IGF-1R in a dose-dependent manner. But KRAS only affected the protein level and not mRNA of YB-1, thus suggesting that KRAS regulated YB-1 expression at the post-transcriptional level. Manipulation of YB-1 via plasmid overexpression and siRNA affected IGF-1R protein and mRNA levels in a dose-dependent manner, and YB-1 and IGF-1R promoter binding was confirmed via ChIP assay. There was a dose-dependent decrease in YB-1 and IGF-1R protein levels with MEK162, a MEK inhibitor, but not with LY294002m a PI3K inhibitor. This demonstrated that the KRAS regulation of the YB-1- IGFR-1R signaling axis to be via the MEK signaling pathway and not PI3K/Akt. Our novel drug T315 targets YB-1, and was shown to inhibit cell viability and cell migration in a dose-dependent manner. T315 treatment of the colon cancer cells also decreased protein levels of YB-1 and IGF-1R and affected epidermal mesenchymal transition markers such as E-cadherin, vimentin, and snail. These results suggest that a combination of a MEK inhibitor and T315 may be an effective therapeutic strategy to target KRAS mutation driven CRC.

Citation Format: Christina Wu, Peng-Chan Lin, Po-Chen Chu, Hsiao-Ching Chuang, Samuel Kulp, Tanios Bekaii-Saab, Ching-Shih Chen. Therapeutic targeting of KRAS mutation-driven tumor progression in colorectal cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5424. doi:10.1158/1538-7445.AM2015-5424
Abstract 5432: Discovery and characterization of USP22 inhibitors as novel anti-cancer agents
Authors: Wang Feng , Stanek Timothy R. , Biswas Leelabati , Kodrasov Matthew , LaRocque James , Wu Jian , Sterner David , Weinstock Joseph , Mattern Michael , McMahon Steven B. , Kumar Suresh
Published: August 2015

The degradation of most cellular proteins is regulated by coordinated addition and removal of ubiquitin by families of ubiquitin E3 ligases and deubiquitylating enzymes (DUBs) respectively. DUBs proteolytically cleave ubiquitin molecules from proteins resulting in modifications of protein activity, localization and function. Several DUBs are aberrantly regulated in cancer, including the best studied, USP7, selective inhibitors of which are active in cancer models. USP22, is another validated anticancer target, being one of 11 genes in the death-from-cancer gene signature, a component of the human SAGA transcriptional cofactor complex regulating myc transcription, and a regulator of the expression of p21, the histone deacetylase Sirt 1, and p53 activity. USP22 is overexpressed in oral squamous cell carcinoma, breast, non-small cell lung, colorectal, and other cancers and its expression is inversely correlated with survival. Unlike most other DUBs, USP22 exhibits robust activity only as a component of a multi-subunit complex. Initial studies reported activity solely as a member of the 2MDa SAGA complex, but more recently it has been demonstrated that USP22 exhibits similar activity in a four-protein DUB module derived from the SAGA complex. USP22 inhibitors are expected to have broad anti-cancer activities. Progenra's UbiProTM discovery platform was utilized to screen ∼200K member diversity based library of small molecules for identifying novel USP22 inhibitors. Data will be presented describing these results.

Citation Format: Feng Wang, Timothy R. Stanek, Leelabati Biswas, Matthew Kodrasov, James LaRocque, Jian Wu, David Sterner, Joseph Weinstock, Michael Mattern, Steven B. McMahon, Suresh Kumar. Discovery and characterization of USP22 inhibitors as novel anti-cancer agents. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5432. doi:10.1158/1538-7445.AM2015-5432
Abstract 5433: Discovery and development of novel USP2 inhibitors for cancer therapy
Authors: Kumar Suresh , LaRocque James P. , Marblestone Jeffrey G. , Wu Jian , Weinstock Joseph , Sterner David E. , Mattern Michael R.
Published: August 2015

Ubiquitylation and deubiquitylation control the stability, localization and activity of many cellular proteins. Deubiquitylation is performed by a family of ∼80 enzymes (DUBs) and many of these DUBs have been implicated in a wide range of diseases including cancer. In particular, USP2 removes ubiquitin selectively from Cyclin D1, Cyclin A1, Aurora A, MDM2 and FAS which are all implicated in oncogenic processes. Importantly, USP2 is overexpressed in prostate and ovarian cancers and inhibition of USP2 has been shown to sensitize prostate carcinoma cells to apoptosis. Therefore USP2 is an attractive therapeutic target. Progenra's unique UbiProTM discovery platform was utilized to screen ∼200K member diversity based library of small molecules for identifying novel USP2 inhibitors. Hits from the screen were subjected to orthogonal selectivity assays to identify USP2 specific inhibitors. Here we report discovery and hit to lead optimization of novel USP2 inhibitors showing cellular efficacy and anti-cancer activities against relevant cancer cell models. The most promising compounds are being further developed as novel anti-cancer agents.

Citation Format: Suresh Kumar, James P. LaRocque, Jeffrey G. Marblestone, Jian Wu, Joseph Weinstock, David E. Sterner, Michael R. Mattern. Discovery and development of novel USP2 inhibitors for cancer therapy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5433. doi:10.1158/1538-7445.AM2015-5433
Abstract 5434: A high through-put bioluminescent assay to monitor the deamidation of asparagine and isomerization of aspartate residues in therapeutic proteins and antibodies
Authors: Goueli Said A. , Hsiao Kevin , Patel Rushikesh , Nashine Vishal
Published: August 2015

Spontaneous degradation reactions such as oxidation, glycation, deamidation, Isomerization, and racemization are non-enzymatic modifications and produce functionally damaged species reflecting aging effect at the molecular level. L-asparagine (Asn) and L-aspartate (Asp) are among the most unstable residues in proteins- linked to deamidation, isomerization and racemization Rx. Under physiological conditions, succinimide-linked deamidation of Asn occurs with t1/2as short as 6 hrs while that for isoaspartate are generally 10-fold longer. A number of biological peptides and proteins possess labile Asn and Asp residues and the formation of IsoAsp at these sites adversely affect their function. The degradation of Asn and Asp sites via succinimide pathway has significant effect on protein function. Loss of function associated with isoAsp formation are found in many biologically critical proteins and is a major source of antibody instability and micro heterogeneity, Thus it is critical to accurately determine both the levels and the site of Asn deamidation in therapeutic antibodies and proteins. Although mass Spectroscopic platforms have been at the forefront of technologies to quantitate and identify sites of isoAsp formation, it fails in accurately determining Asn level and has problems with Asp isomerization due to the various steps required for sample preparation such as denaturation, reduction, alkylation, and enzyme digestion of the Ab before analysis resulting in problems with Asp isomerization and Asn determination artifacts. Although peptide mapping is the most powerful tool in isoAsp analyses, it is very time consuming and may itself actually cause sample degradation during preparation and analysis and is not always practical when large number of samples are needed for analysis. To improve on and complement existing technologies, we have developed a bioluminescent, homogenous, robust, sensitive and easy to use assay to accurately monitor the deamidation of Asn and isomerization of aspartate. The assay is based on the use of protein isoaspartate methyltransferase to catalyze the methylation of isoaspartate using S-adenosylmethionine (SAM) and resulting in the formation of S-adenosylhomocysteine and methylated isoaspartate. The assay is very sensitive, it can detect as little as 100 fmol of IsoAsp in as little as 5-10 pmol of proteins.

Citation Format: Said A. Goueli, Kevin Hsiao, Rushikesh Patel, Vishal Nashine. A high through-put bioluminescent assay to monitor the deamidation of asparagine and isomerization of aspartate residues in therapeutic proteins and antibodies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5434. doi:10.1158/1538-7445.AM2015-5434
Abstract 5435: A bioluminescent assay for measuring glucose uptake
Authors: Valley Michael P. , Sobol Mary , Vidugiriene Jolanta , Cali James J.
Published: August 2015

Glucose management is a key biological process. In adipose tissue and skeletal muscle, decreased rates of glucose uptake commonly associated with diabetes mellitus lead to chronic hyperglycemia and a wide array of physiological damage. In contrast, the overexpression of glucose transporters in cancer cells results in increased rates of glucose uptake to support their high rates of proliferation. Thus, activators and inhibitors of glucose uptake are useful for both diabetes management and anticancer therapies. The standard method of assaying glucose uptake involves addition of a radioactive glucose analog (2-deoxyglucose) and measurement of the accumulation of the stable and impermeable phosphorylated derivative, 2-deoxyglucose-6-phosphate (2DG6P). However, radioactive assays are laborious, costly, and require special handling of radioactive materials and waste. A simpler assay can be made by measuring the production of NADPH through the oxidation of 2DG6P by glucose-6-phosphate dehydrogenase. We have developed a bioluminescent glucose uptake assay that is both rapid and convenient and exhibits a larger signal window than comparable fluorescent or colorimetric approaches. The utility of this assay will be demonstrated with both cancer cells and insulin-sensitive 3T3L1 adipocytes and L6 myotubes.

Citation Format: Michael P. Valley, Mary Sobol, Jolanta Vidugiriene, James J. Cali. A bioluminescent assay for measuring glucose uptake. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5435. doi:10.1158/1538-7445.AM2015-5435