bluuebunny/crossref_metadata_2025 huggingface dataset:
Wavelet based local fairing algorithm for surfaces with data compression
Authors: Wang Aizeng , Zhao Gang , Li Yong-Dong
Published: August 2015

By combining the T-spline and wavelet techniques together, this paper proposes a new local fairing algorithm for B-spline surfaces. There are two steps of operations in the new algorithm. The first is to decompose a local region into a scale part and a detail one by wavelet analysis. The second is to reconstruct the surface by removing the detail part and then replacing the local region by the scale one. Because the scale part constitutes a T-spline, the new algorithm can not only realize local fairing, but also simultaneously reduce the control points. The effectiveness of the new approach is verified by three examples, which also demonstrate that the major advantage of the new algorithm over the traditional ones lies in its local fairing capability.
A new signal characterization and signal-based Chou's PseAAC representation of protein sequences
Authors: Sanchez Victoria , Peinado Antonio M. , Pérez-Córdoba Jose L. , Gómez Angel M.
Published: August 2015

Most of the algorithms used for information extraction and for processing the amino acid chains that make up proteins treat them as symbolic chains. Fewer algorithms exploit signal processing techniques that require a numerical representation of amino acid chains. However, these algorithms are very powerful for extracting regularities that cannot be detected when working with a symbolic chain, which may be important for understanding the biological meaning of a sequence or in classification tasks. In this study, a new mathematical representation of amino acid chains is proposed, which is derived using a similarity measure based on the PAM250 amino acid substitution matrix and that generates 20 signals for each protein sequence. Using this representation 20 consensus spectra for a protein family are determined and the relevance of the frequency peaks is established, obtaining a group of significant frequency peaks that manifest common periodicities of the amino acid sequences that belong to a protein family. We also show that the proposed representation in 20 signals can be integrated into Chou's pseudo amino acid composition (PseAAC) and constitute a useful alternative to amino acid physicochemical properties in Chou's PseAAC.
On some branches of the Bruhat–Tits tree
Authors: Arenas-Carmona Luis , Saavedra Ignacio
Published: August 2015

We explicitly compute the largest subtree, in the local Bruhat–Tits tree for [Formula: see text], whose vertices correspond to maximal orders containing a fixed order generated by a pair of orthogonal pure quaternions. In other words, we compute the set of maximal integral valued lattices in a ternary quadratic space, whose discriminant is a unit, containing a pair of orthogonal vectors, extending thus previous computations by Schulze-Pilot. The maximal order setting makes these computations simpler. The method presented here can be applied to arbitrary sub-orders or sublattices. The shape of this subtree is described, when it is finite, by a set of two invariants. In a previous work, the first author showed that determining the shape of these local subtrees allows the computation of representation fields, a class field determining the set of isomorphism classes, in a genus of Eichler orders, containing an isomorphic copy of a given order. Some further applications are described here.
Rift magmatism on the Eurasia basin margin: U–Pb baddeleyite ages of alkaline dyke swarms in North Greenland
Authors: Thórarinsson Sigurjón B. , Söderlund Ulf , Døssing Arne , Holm Paul M. , Ernst Richard E. , Tegner Christian
Published: August 2015

The opening of the Arctic Ocean involved multiple stages of continental rifting and intrusion of extensive dyke swarms. To trace tectonomagmatic processes of the High Arctic, we present the first U–Pb ages for alkaline dyke swarms of North Greenland. Concordia ages of 80.8 ± 0.6 and 82.1 ± 1.5 Ma indicate that north–south and east–west dykes are coeval. The north–south dykes reflect initial east–west rifting that led to break-up along the Gakkel Ridge and formation of the Eurasia Basin. The east–west dykes reflect local variations in the stress field associated with reactivated Palaeozoic faults.

Supplementary materials:

U–Pb data are available at
http://www.geolsoc.org.uk/SUP18857
.
Silurian–Devonian magmatism, mineralization, regional exhumation and brittle strike-slip deformation along the Loch Shin Line, NW Scotland
Authors: Holdsworth R. E. , Dempsey E. , Selby D. , Darling J. R. , Feely M. , Costanzo A. , Strachan R. A. , Waters P. , Finlay A. J. , Porter S. J.
Published: August 2015

The Loch Shin Line is a geological–geophysical lineament associated with a zone of mantle-derived appinites, granites and strike-slip faulting that runs NW–SE across the Moine Nappe, northern Scotland. U–Pb zircon and Re–Os molybdenite dating of the Loch Shin and Grudie plutons, which lie immediately SW of the NW–SE Loch Shin–Strath Fleet fault system, yield
c
. 427–430 Ma ages that overlap within error. They also coincide with previously obtained U–Pb zircon ages for the Rogart pluton, which lies along-strike to the SE. Field and microstructural observations confirm the similarity and contemporaneous nature of the plutons and associated sulphide mineralization. Fluid inclusion analyses place further constraints on the
P
–
T
–
X
conditions during regional late Caledonian exhumation of the Moine Nappe. Synchronous to slightly younger brittle dextral strike-slip faulting along the WNW–ESE Loch Shin–Strath Fleet Fault System was probably antithetic to sinistral movements along the nearby Great Glen Fault Zone. Our findings support the hypothesis that the Loch Shin Line acted as a deep crustal channelway controlling the ascent and emplacement of Silurian magmas into the overlying Moine Nappe. We propose that this deep structure corresponds to the southeastern continuation of the Precambrian-age Laxford Front shear zone in the buried Lewisian autochthon.

Supplementary Material:

Field photographs, photomicrographs and fluid inclusion information are available at
http://www.geolsoc.org.uk/SUP18859
.
The role of apolipoprotein A-IV in regulating glucagon-like peptide-1 secretion
Authors: Wang Fei , Yang Qing , Huesman Sarah , Xu Min , Li Xiaoming , Lou Danwen , Woods Stephen C. , Marziano Corina , Tso Patrick
Published: August 2015

Both glucagon-like peptide-1 (GLP-1) and apolipoprotein A-IV (apoA-IV) are produced from the gut and enhance postprandial insulin secretion. This study investigated whether apoA-IV regulates nutrient-induced GLP-1 secretion and whether apoA-IV knockout causes compensatory GLP-1 release. Using lymph-fistula-mice, we first determined lymphatic GLP-1 secretion by administering apoA-IV before an intraduodenal Ensure infusion. apoA-IV changed neither basal nor Ensure-induced GLP-1 secretion relative to saline administration. We then assessed GLP-1 in apoA-IV−/− and wild-type (WT) mice administered intraduodenal Ensure. apoA-IV−/− mice had comparable lymph flow, lymphatic triglyceride, glucose, and protein outputs as WT mice. Intriguingly, apoA-IV−/− mice had higher lymphatic GLP-1 concentration and output than WT mice 30 min after Ensure administration. Increased GLP-1 was also observed in plasma of apoA-IV−/− mice at 30 min. apoA-IV−/− mice had comparable total gut GLP-1 content relative to WT mice under fasting, but a lower GLP-1 content 30 min after Ensure administration, suggesting that more GLP-1 was secreted. Moreover, an injection of apoA-IV protein did not reverse the increased GLP-1 secretion in apoA-IV−/− mice. Finally, we assessed gene expression of GLUT-2 and the lipid receptors, including G protein-coupled receptor (GPR) 40, GPR119, and GPR120 in intestinal segments. GLUT-2, GPR40 and GPR120 mRNAs were unaltered by apoA-IV knockout. However, ileal GPR119 mRNA was significantly increased in apoA-IV−/− mice. GPR119 colocalizes with GLP-1 in ileum and stimulates GLP-1 secretion by sensing OEA, lysophosphatidylcholine, and 2-monoacylglycerols. We suggest that increased ileal GPR119 is a potential mechanism by which GLP-1 secretion is enhanced in apoA-IV−/− mice.
Short- and long-term regulation of intestinal Na+/H+exchange by Toll-like receptors TLR4 and TLR5
Authors: Cabral José Miguel , Grácio Daniela , Soares-da-Silva Patrício , Magro Fernando
Published: August 2015

Inappropriate activation of pattern recognition receptors has been described as a potential trigger in the development of inflammatory bowel disease (IBD). In this study, we evaluated the activity and expression of Na+/H+exchanger (NHE) subtypes in T84 intestinal epithelial cells during Toll-like receptor 4 (TLR4) activation by monophosphoryl lipid A and TLR5 by flagellin. NHE activity and intracellular pH were evaluated by spectrofluorescence. Additionally, kinase activities were evaluated by ELISA, and siRNA was used to specifically inhibit adenylyl cyclase (AC). Monophosphoryl lipid A (MPLA) (0.01–50.00 μg/ml) and flagellin (10–500 ng/ml) inhibited NHE1 activity in a concentration-dependent manner (MPLA short term −25.2 ± 5.0%, long term −31.9 ± 4.0%; flagellin short term −14.9 ± 2.0%, long term −19.1 ± 2.0%). Both ligands triggered AC3, PKA, PLC, and PKC signal molecules. Long-term exposure to flagellin and MPLA induced opposite changes on NHE3 activity; flagellin increased NHE3 activity (∼10%) with overexpression of membrane protein, whereas MPLA decreased NHE3 activity (−17.3 ± 3.0%). MPLA and flagellin simultaneously had synergistic effects on NHE activity. MPLA and flagellin impaired pHirecovery after intracellular acidification. The simultaneous exposure to MPLA and flagellin induced a substantial pHireduction (−0.55 ± 0.03 pH units). Activation of TLR4 and TLR5 exerts marked inhibition of NHE1 activity in intestinal epithelial cells. Transduction mechanisms set into motion during TLR4-mediated and long-term TLR5-mediated inhibition of NHE1 activity involve AC3, PKA, PLC, and PKC. However, short- and long-term TLR4 activation and TLR5 activation might use different signaling pathways. The physiological alterations on intestinal epithelial cells described here may be useful in the development of better IBD therapeutics.
Intestinal DMT1 is critical for iron absorption in the mouse but is not required for the absorption of copper or manganese
Authors: Shawki Ali , Anthony Sarah R. , Nose Yasuhiro , Engevik Melinda A. , Niespodzany Eric J. , Barrientos Tomasa , Öhrvik Helena , Worrell Roger T. , Thiele Dennis J. , Mackenzie Bryan
Published: August 2015

Divalent metal-ion transporter-1 (DMT1) is a widely expressed iron-preferring membrane-transport protein that serves a critical role in erythroid iron utilization. We have investigated its role in intestinal metal absorption by studying a mouse model lacking intestinal DMT1 (i.e., DMT1int/int). DMT1int/intmice exhibited a profound hypochromic-microcytic anemia, splenomegaly, and cardiomegaly. That the anemia was due to iron deficiency was demonstrated by the following observations in DMT1int/intmice: 1) blood iron and tissue nonheme-iron stores were depleted; 2) mRNA expression of liver hepcidin (Hamp1) was depressed; and 3) intraperitoneal iron injection corrected the anemia, and reversed the changes in blood iron, nonheme-iron stores, and hepcidin expression levels. We observed decreased total iron content in multiple tissues from DMT1int/intmice compared with DMT1+/+mice but no meaningful change in copper, manganese, or zinc. DMT1int/intmice absorbed64Cu and54Mn from an intragastric dose to the same extent as did DMT1+/+mice but the absorption of59Fe was virtually abolished in DMT1int/intmice. This study reveals a critical function for DMT1 in intestinal nonheme-iron absorption for normal growth and development. Further, this work demonstrates that intestinal DMT1 is not required for the intestinal transport of copper, manganese, or zinc.
Vascular endothelial cell membranes differentiate between stretch and shear stress through transitions in their lipid phases
Authors: Yamamoto Kimiko , Ando Joji
Published: August 2015

Vascular endothelial cells (ECs) respond to the hemodynamic forces stretch and shear stress by altering their morphology, functions, and gene expression. However, how they sense and differentiate between these two forces has remained unknown. Here we report that the plasma membrane itself differentiates between stretch and shear stress by undergoing transitions in its lipid phases. Uniaxial stretching and hypotonic swelling increased the lipid order of human pulmonary artery EC plasma membranes, thereby causing a transition from the liquid-disordered phase to the liquid-ordered phase in some areas, along with a decrease in membrane fluidity. In contrast, shear stress decreased the membrane lipid order and increased membrane fluidity. A similar increase in lipid order occurred when the artificial lipid bilayer membranes of giant unilamellar vesicles were stretched by hypotonic swelling, indicating that this is a physical phenomenon. The cholesterol content of EC plasma membranes significantly increased in response to stretch but clearly decreased in response to shear stress. Blocking these changes in the membrane lipid order by depleting membrane cholesterol with methyl-β-cyclodextrin or by adding cholesterol resulted in a marked inhibition of the EC response specific to stretch and shear stress, i.e., phosphorylation of PDGF receptors and phosphorylation of VEGF receptors, respectively. These findings indicate that EC plasma membranes differently respond to stretch and shear stress by changing their lipid order, fluidity, and cholesterol content in opposite directions and that these changes in membrane physical properties are involved in the mechanotransduction that activates membrane receptors specific to each force.
The cerebral venous system and hypoxia
Authors: Wilson Mark H. , Imray Christopher H. E.
Published: August 2015

Most hypobaric hypoxia studies have focused on oxygen delivery and therefore cerebral blood inflow. Few have studied venous outflow. However, the volume of blood entering and leaving the skull (∼700 ml/min) is considerably greater than cerebrospinal fluid production (0.35 ml/min) or edema formation rates and slight imbalances of in- and outflow have considerable effects on intracranial pressure. This dynamic phenomenon is not necessarily appreciated in the currently taught static “Monro-Kellie” doctrine, which forms the basis of the “Tight-Fit” hypothesis thought to underlie high altitude headache, acute mountain sickness, and high altitude cerebral edema. Investigating both sides of the cerebral circulation was an integral part of the 2007 Xtreme Everest Expedition. The results of the relevant studies performed as part of and subsequent to this expedition are reviewed here. The evidence from recent studies suggests a relative venous outflow insufficiency is an early step in the pathogenesis of high altitude headache. Translation of knowledge gained from high altitude studies is important. Many patients in a critical care environment develop hypoxemia akin to that of high altitude exposure. An inability to drain the hypoxemic induced increase in cerebral blood flow could be an underappreciated regulatory mechanism of intracranial pressure.
CD38-Expressing Myeloid-Derived Suppressor Cells Promote Tumor Growth in a Murine Model of Esophageal Cancer
Authors: Karakasheva Tatiana A. , Waldron Todd J. , Eruslanov Evgeniy , Kim Sang-Bae , Lee Ju-Seog , O'Brien Shaun , Hicks Philip D. , Basu Devraj , Singhal Sunil , Malavasi Fabio , Rustgi Anil K.
Published: August 2015

Myeloid-derived suppressor cells (MDSC) are an immunosuppressive population of immature myeloid cells found in advanced-stage cancer patients and mouse tumor models. Production of inducible nitric oxide synthase (iNOS) and arginase, as well as other suppressive mechanisms, allows MDSCs to suppress T-cell–mediated tumor clearance and foster tumor progression. Using an unbiased global gene expression approach in conditional p120-catenin knockout mice (L2-cre;p120ctnf/f), a model of oral–esophageal cancer, we have identified CD38 as playing a vital role in MDSC biology, previously unknown. CD38 belongs to the ADP-ribosyl cyclase family and possesses both ectoenzyme and receptor functions. It has been described to function in lymphoid and early myeloid cell differentiation, cell activation, and neutrophil chemotaxis. We find that CD38 expression in MDSCs is evident in other mouse tumor models of esophageal carcinogenesis, and CD38high MDSCs are more immature than MDSCs lacking CD38 expression, suggesting a potential role for CD38 in the maturation halt found in MDSC populations. CD38high MDSCs also possess a greater capacity to suppress activated T cells, and promote tumor growth to a greater degree than CD38low MDSCs, likely as a result of increased iNOS production. In addition, we have identified novel tumor–derived factors, specifically IL6, IGFBP3, and CXCL16, which induce CD38 expression by MDSCs ex vivo. Finally, we have detected an expansion of CD38+ MDSCs in peripheral blood of advanced-stage cancer patients and validated targeting CD38 in vivo as a novel approach to cancer therapy. Cancer Res; 75(19); 4074–85. ©2015 AACR.
Androgen-Regulated SPARCL1 in the Tumor Microenvironment Inhibits Metastatic Progression
Authors: Hurley Paula J. , Hughes Robert M. , Simons Brian W. , Huang Jessie , Miller Rebecca M. , Shinder Brian , Haffner Michael C. , Esopi David , Kimura Yasunori , Jabbari Javaneh , Ross Ashley E. , Erho Nicholas , Vergara Ismael A. , Faraj Sheila F. , Davicioni Elai , Netto George J. , Yegnasubramanian Srinivasan , An Steven S. , Schaeffer Edward M.
Published: August 2015

Prostate cancer is a leading cause of cancer death in men due to the subset of cancers that progress to metastasis. Prostate cancers are thought to be hardwired to androgen receptor (AR) signaling, but AR-regulated changes in the prostate that facilitate metastasis remain poorly understood. We previously noted a marked reduction in secreted protein, acidic and rich in cysteine-like 1 (SPARCL1) expression during invasive phases of androgen-induced prostate growth, suggesting that this may be a novel invasive program governed by AR. Herein, we show that SPARCL1 loss occurs concurrently with AR amplification or overexpression in patient-based data. Mechanistically, we demonstrate that SPARCL1 expression is directly suppressed by androgen-induced AR activation and binding at the SPARCL1 locus via an epigenetic mechanism, and these events can be pharmacologically attenuated with either AR antagonists or HDAC inhibitors. We establish using the Hi-Myc model of prostate cancer that in Hi-Myc/Sparcl1−/− mice, SPARCL1 functions to suppress cancer formation. Moreover, metastatic progression of Myc-CaP orthotopic allografts is restricted by SPARCL1 in the tumor microenvironment. Specifically, we show that SPARCL1 both tethers to collagen in the extracellular matrix (ECM) and binds to the cell's cytoskeleton. SPARCL1 directly inhibits the assembly of focal adhesions, thereby constraining the transmission of cell traction forces. Our findings establish a new insight into AR-regulated prostate epithelial movement and provide a novel framework whereby SPARCL1 in the ECM microenvironment restricts tumor progression by regulating the initiation of the network of physical forces that may be required for metastatic invasion of prostate cancer. Cancer Res; 75(20); 4322–34. ©2015 AACR.
The Neuronal Pentraxin-2 Pathway Is an Unrecognized Target in Human Neuroblastoma, Which Also Offers Prognostic Value in Patients
Authors: Bartolini Alice , Di Paolo Daniela , Noghero Alessio , Murgia Daniele , Sementa Angela R. , Cilli Michele , Pasqualini Renata , Arap Wadih , Bussolino Federico , Ponzoni Mirco , Pastorino Fabio , Marchiò Serena
Published: August 2015

Neuronal pentraxins (NPTX) and their corresponding receptors (NPTXR) have been studied as synapse-associated proteins in the nervous system, but their role in cancer is largely unknown. By applying a multidisciplinary, high-throughput proteomic approach, we have recently identified a peptide ligand motif for targeted drug delivery to neuroblastoma. Here, we report the sequence similarity between this peptide and a conserved portion of the pentraxin domain that is involved in the homo- and hetero-oligomerization of NPTX2 and NPTXR. We show that, in comparison with normal tissues, NPTX2 and NPTXR are overexpressed in vivo in mouse models, as well as in human Schwannian stroma-poor, stage IV neuroblastoma. Both proteins are concentrated in the vicinity of tumor blood vessels, with NPTXR also present on neuroblastic tumor cells. In vivo targeting of NPTX2 and NPTXR with the selected peptide or with specific antibodies reduces tumor burden in orthotopic mouse models of human neuroblastoma. In vitro interference with this ligand/receptor system inhibits the organization of neuroblastoma cells in tumor-like masses in close contact with vascular cells, as well as their adhesion to normal microenvironment-derived cells, suggesting a role in the cross-talk between tumor and normal cells in the early steps of neuroblastoma development. Finally, we show that NPTX2 is a marker of poor prognosis for neuroblastoma patients. Cancer Res; 75(20); 4265–71. ©2015 AACR.
Akt Kinase-Interacting Protein 1 Signals through CREB to Drive Diffuse Malignant Mesothelioma
Authors: Yamada Tadaaki , Amann Joseph M. , Fukuda Koji , Takeuchi Shinji , Fujita Naoya , Uehara Hisanori , Iwakiri Shotaro , Itoi Kazumi , Shilo Konstantin , Yano Seiji , Carbone David P.
Published: August 2015

Diffuse malignant mesothelioma (DMM) is a tumor of serosal membranes with propensity for progressive local disease. Because current treatment options are largely ineffective, novel therapeutic strategies based on molecular mechanisms and the disease characteristics are needed to improve the outcomes of patients with this disease. Akt kinase interacting protein 1 (Aki1; Freud-1/CC2D1A) is a scaffold protein for the PI3K–PDK1–Akt signaling module that helps determine receptor signal selectivity for EGFR. Aki1 has been suggested as a therapeutic target, but its potential has yet to be evaluated in a tumor setting. Here, we report evidence supporting its definition as a therapeutic target in DMM. In cell-based assays, Aki1 silencing decreased cell viability and caused cell-cycle arrest of multiple DMM cell lines via effects on the PKA–CREB1 signaling pathway. Blocking CREB activity phenocopied Aki1 silencing. Clinically, Aki1 was expressed in most human DMM specimens where its expression correlated with phosphorylated CREB1. Notably, Aki1 siRNA potently blocked tumor growth in an orthotopic implantation model of DMM when administered directly into the pleural cavity of tumor-bearing mice. Our findings suggest an important role for the Aki1–CREB axis in DMM pathogenesis and provide a preclinical rationale to target Aki1 by intrathoracic therapy in locally advanced tumors. Cancer Res; 75(19); 4188–97. ©2015 AACR.
Integrating Image-Based High-Content Screening with Mouse Models Identifies 5-Hydroxydecanoate as a Neuroprotective Drug for Paclitaxel-Induced Neuropathy
Authors: Chen Li-Hsien , Sun Yuan-Ting , Chen Yih-Fung , Lee Mei-Yi , Chang Lian-Yun , Chang Jang-Yang , Shen Meng-Ru
Published: August 2015

Chemotherapy-induced neurotoxicity is a common adverse effect of cancer treatment. No medication has been shown to be effective in the prevention or treatment of chemotherapy-induced neurotoxicity. This study aimed to discover potential neuroprotective drugs for paclitaxel-induced neurotoxicity. An image-based high-content platform was first developed to screen for potential neuroprotective drugs. The screening system comprised of automated image acquisition and multiparameter analysis, including neuronal viability, neurite outgrowth, and synaptogenesis. By this platform, we obtained a candidate list from compound libraries. In the drug screening from compound libraries of ion channel ligands, REDOX and GABAergic ligands, 5-hydroxydecanoate (5-HD) exhibited the most significant neuroprotective effects against paclitaxel-induced neurotoxicity in both cortical and dorsal root ganglion (DRG) neurons. In mouse behavioral tests, 5-HD restored the thermal sensitivity and alleviated mechanical allodynia induced by paclitaxel. Electron micrographs of sciatic nerve revealed that 5-HD reduced the damages caused by paclitaxel in the nonmyelinated and smaller myelinated fibers. The mechanistic study on DRG neurons suggested that 5-HD rescued the dysregulation of intracellular calcium homeostasis provoked by paclitaxel. Importantly, 5-HD did not jeopardize the antitumor effect of paclitaxel in tumor xenograft models. In conclusion, we established an imaged-based high-content screening platform and a protocol for verifying the neuroprotective effect in vivo, by which 5-HD was identified and validated as a potential neuroprotective drug for paclitaxel-induced neuropathy. Mol Cancer Ther; 14(10); 2206–14. ©2015 AACR.
A Novel Inhibitor of AKT1–PDPK1 Interaction Efficiently Suppresses the Activity of AKT Pathway and Restricts Tumor Growth In Vivo
Authors: Mäemets-Allas Kristina , Viil Janeli , Jaks Viljar
Published: August 2015

The serine/threonine kinase AKT/PKB has a critical role in the regulation of cell proliferation. Because AKT signaling is deregulated in numerous human malignancies, it has become an attractive anticancer drug target. A number of small molecule AKT kinase inhibitors have been developed; however, severe side effects have prevented their use in clinical trials. To find inhibitors of AKT1 signaling with principally novel mechanism of action, we carried out a live cell-based screen for small molecule inhibitors of physical interaction between AKT1 and its primary activator PDPK1. The screen revealed one molecule—NSC156529, which downregulated AKT1 signaling, efficiently decreased the proliferation of human cancer cells in vitro, and substantially inhibited the growth of prostate tumor xenografts in vivo. Interestingly, the treated tumor xenografts exhibited higher expression level of normal prostate differentiation markers but did not show augmented cell death, suggesting that the identified compound primarily enhances the differentiation of malignant cells toward normal prostate epithelium and thus poses as an attractive lead compound for developing novel antitumor agents with less cytotoxic side effects. Mol Cancer Ther; 14(11); 2486–96. ©2015 AACR.
Antibody Format and Drug Release Rate Determine the Therapeutic Activity of Noninternalizing Antibody–Drug Conjugates
Authors: Gébleux Rémy , Wulhfard Sarah , Casi Giulio , Neri Dario
Published: August 2015

The development of antibody–drug conjugates (ADC), a promising class of anticancer agents, has traditionally relied on the use of antibodies capable of selective internalization in tumor cells. We have recently shown that also noninternalizing antibodies, coupled to cytotoxic drugs by means of disulfide linkers that can be cleaved in the tumor extracellular environment, can display a potent therapeutic activity. Here, we have compared the tumor-targeting properties, drug release rates, and therapeutic performance of two ADCs, based on the maytansinoid DM1 thiol drug and on the F8 antibody, directed against the alternatively spliced Extra Domain A (EDA) domain of fibronectin. The antibody was used in IgG or in small immune protein (SIP) format. In both cases, DM1 was coupled to unpaired cysteine residues, resulting in a drug-antibody ratio of 2. In biodistribution studies, SIP(F8)-SS-DM1 accumulated in the tumor and cleared from circulation more rapidly than IgG(F8)-SS-DM1. However, the ADC based on the IgG format exhibited a higher tumor uptake at later time points (e.g., 33%IA/g against 8%IA/g at 24 hours after intravenous administration). In mouse plasma, surprisingly, the ADC products in IgG format were substantially more stable compared with the SIP format (half-lives >48 hours and <3 hours at 37°C, respectively), revealing a novel mechanism for the control of disulfide-based drug release rates. Therapy experiments in immunocompetent mice bearing murine F9 tumors revealed that SIP(F8)-SS-DM1 was more efficacious than IgG(F8)-SS-DM1 when the two products were compared either in an equimolar basis or at equal milligram doses. Mol Cancer Ther; 14(11); 2606–12. ©2015 AACR.
Abstract A04: The tumor microenvironment is the main source of IL-6 for plasmacytoma development in mice.
Authors: Rosean Timothy R. , Tompkins Van S. , Sompallae Ramakrishna , Norian Lyse A. , Morse Herbert C. , Waldschmidt Thomas J. , Janz Siegfried
Published: August 2015

The development of targeted interventions for hematopoietic neoplasms of the B-lymphocyte lineage, such as multiple myeloma (MM), may benefit from preclinical experimental model systems in which complex interactions of neoplastic and nonmalignant bystander cells in the tumor microenvironment (TME) can be genetically dissected. Here we employ peritoneal plasmacytomagenesis in mice, which is strictly dependent on the pro-inflammatory cytokine, interleukine 6 (IL-6), to demonstrate that the nonmalignant bystanders are the main source of IL-6 for tumor development. B cell derived IL-6 is dispensable. The finding that lack of IL-6 in the TME significantly delays tumor onset in mice supports translational research efforts to target IL-6 production in bone marrow stroma cells for myeloma treatment and prevention.

Citation Format: Timothy R. Rosean, Van S. Tompkins, Ramakrishna Sompallae, Lyse A. Norian, Herbert C. Morse, III, Thomas J. Waldschmidt, Siegfried Janz. The tumor microenvironment is the main source of IL-6 for plasmacytoma development in mice. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A04.
Abstract A13: Anti-CD38-attenukine: A myeloma-targeting immunocytokine containing an engineered IFNα that provides >10,000-fold enhanced tumor-specific activity compared to native IFNα.
Authors: Pogue Sarah , Taura Tetsuya , Bi Mingying , Mikesell Glen , Yun Yong , Sho Angela , Sanchez Eric , Chen Haiming , Berenson James , Behrens Collette , Stevens Maxwell , Domagala Teresa , Sokolovsky Maya , Hallak Hussein , Rosenstock Moti , Doyle Anthony , Wilson David
Published: August 2015

Multiple myeloma (MM) remains an incurable disease despite recent approvals of novel therapies. Treatments which utilize alkylating agents, corticosteroids, proteasome inhibitors and immunomodulatory drugs have provided significant survival benefits; however, tumor relapse eventually occurs with the use of these agents. The cytokine, interferon-α (IFNα), has been used clinically to treat MM for over 30 years. IFNα exerts good anti-myeloma tumor activity but durable responses are rarely achieved due to dose limiting toxicities. Tumor specificity can be modestly increased by attaching IFNα to an anti-CD38 antibody, thus targeting the cytokine to MM cells. However, by introducing an attenuating mutation to the IFNα portion of this fusion protein, the activity of the cytokine is dramatically reduced on CD38 negative cells while exhibiting a similar potency to native IFNα on CD38+ MM cells. As a result, the anti-CD38-attenuated IFNα, referred to as anti-CD38-attenukine™, has over 10,000-fold greater myeloma-specificity than native IFNα in vitro. In vivo, even though the off-target activity is decreased, anti-CD38-attenukine™ has profound anti-tumor effects in murine MM and NHL xenograft models. Treatment with this fusion protein can cure mice with large (>700mm3) tumors—an effect not observed with any other MM agents tested. Here we report that in a Velcade (bortezomib) refractory, IFNα-insensitive primary MM xenograft model, the combination of Velcade with anti-CD38-attenukine™ has potent anti-tumor activity, resulting in tumour elimination and curing of mice. Our findings indicate that anti-CD38-attenukine™ may be a well tolerated, potent anti-MM treatment that may be utilized alone or in combination with Velcade.

Citation Format: Sarah Pogue, Tetsuya Taura, Mingying Bi, Glen Mikesell, Yong Yun, Angela Sho, Eric Sanchez, Haiming Chen, James Berenson, Collette Behrens, Maxwell Stevens, Teresa Domagala, Maya Sokolovsky, Hussein Hallak, Moti Rosenstock, Anthony Doyle, David Wilson. Anti-CD38-attenukine: A myeloma-targeting immunocytokine containing an engineered IFNα that provides >10,000-fold enhanced tumor-specific activity compared to native IFNα. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A13.
Abstract A17: Non-RGD-based strategies to target the thyroid hormone receptor-integrin αvβ3: Lessons from myeloma cells.
Authors: Cohen Keren , Ellis Martin , Davis Paul J. , Hercbergs Aleck , Ashur-Fabian Osnat
Published: August 2015

Introduction. Integrin αvβ3 is a plasma membrane structural protein expressed/activated in tumor cells and dividing blood vessel cells. The extracellular domain of αvβ3 has an Arg-Gly-Asp (RGD) recognition site that enables the integrin to bind specific extracellular matrix (ECM) proteins that contain the RGD sequence and to transduce the binding into discrete intracellular signals. Such ECM proteins have pivotal roles in solid tumors and the RGD site has been tested as a chemotherapeutic target. Recently, a receptor for thyroid hormone (L-thyroxine, T4; 3, 5, 3'-triiodo-L-thyronine, T3) has been described on αvβ3 that is distinct from the RGD recognition site and has been shown to mediate proliferative effects of T4 and T3 on solid tumor cells via the MAPK pathway. We have recently shown that similar T4- and T3-induced proliferation occurs in multiple myeloma (MM) cells. Actions of T4 and T3 at the non-RGD site can be blocked by tetraiodothyroacetic acid (tetrac), a naturally-occurring deaminated T4 analog. In addition, tetrac has anti-tumor effects that are independent of blockade of actions of T4 and T3. In this report, we define mechanisms of tetrac action in MM cells.

Methods. MM cell lines (CAG, RPMI 8226, ARK, ARP-1, U266) and primary cultures of marrow cells from MM patients were grown with T4/T3 and with/without tetrac (100 nM-50 μM). Harvested cells were subjected to the following analyses: cell counts (FACS, CyQuant), viability (WST-1), cell cycle (FACS, PI), cell death (Annexin-PI ± pan-caspase inhibitor, ZVAD, FACS; cleaved caspase-3, cleaved PARP, western blots), DNA damage response (pATM and PARP, western blots) and transcription of apoptosis-relevant genes (RT-PCR).

Results. Early (4-8 h) and late (24 h) effects of tetrac were observed. In the early phase, tetrac reduced cell proliferation (p <<0.05) and induced a DNA damage response (ATM and PARP) and apoptosis (cleaved caspase-3 and cleaved PARP). After 24 h, tetrac significantly (p < 0.05) reduced cell number and viability and induced apoptotic cell death, with a parallel increase in pro-apoptotic gene expression.

Conclusion. The thyroid hormone-binding (non-RGD) site on integrin αvβ3 in myeloma cells appears to be a novel target for chemotherapy with tetrac.

Citation Format: Keren Cohen, Martin Ellis, Paul J. Davis, Aleck Hercbergs, Osnat Ashur-Fabian. Non-RGD-based strategies to target the thyroid hormone receptor-integrin αvβ3: Lessons from myeloma cells. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A17.
Abstract A21: Inhibition of USP7 for the treatment of multiple myeloma and other malignancies.
Authors: Nicholson Benjamin , Kodrasov Matthew P. , LaRocque James P. , Eddins Michael J. , Kumar Suresh , Chauhan Dharminder , Anderson Kenneth C. , Mattern Michael R.
Published: August 2015

The conjugation and deconjugation of ubiquitin controls the stability, localization and activity of many cellular proteins. Deconjugation of ubiquitin is performed by a family of ~80 deubiquitylating enzymes (DUBs) including USP7 which specifically removes ubiquitin from HDM2, Foxp3, claspin and other therapeutically relevant substrates. Importantly, USP7 is overexpressed in human multiple myeloma cells and the expression level of USP7 inversely correlates with prognosis in multiple myeloma patients. Previously, we reported the discovery and characterization of P0005091 a USP7 inhibitor which exhibited anti-tumor activity in in vitro and in vivo models of multiple myeloma. Additionally, we reported that P0005091 inhibits USP47 a closely related DUB which is an emerging oncology target in its own right. Furthermore, a structural analog of P0005091, P0022077 has recently been reported by another research group to inhibit neuroblastoma growth in an orthotopic mouse model suggesting that in addition to hematological tumors, solid tumors may be treatable via inhibition of USP7. To obtain additional starting points for a USP7 or a USP7/USP47 specific drug discovery program we recently performed a new screening campaign. By utilizing a biophysical assay platform we were able to exclude compounds that bind to the active site of USP7 thus increasing the possibility of identifying allosteric modifiers of USP7 activity. Hits from this screen are currently the subject of hit to lead optimization and will be evaluated in biophysical and biochemical assays as well as in vitro and in vivo models of hematological malignancies. Data will be presented describing these results.

Citation Format: Benjamin Nicholson, Matthew P. Kodrasov, James P. LaRocque, Michael J. Eddins, Suresh Kumar, Dharminder Chauhan, Kenneth C. Anderson, Michael R. Mattern. Inhibition of USP7 for the treatment of multiple myeloma and other malignancies. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A21.
Abstract A26: Cancer-associated mutations impair the functional association of ETO/MTG family members with E proteins.
Authors: Acharya Pankaj , Hiebert Scott W.
Published: August 2015

MTG8 and MTG16 are disrupted by the t(8;21) and t(16;21) chromosomal translocations respectively, yielding a fusion protein containing the N-terminal DNA-binding domain of RUNX1 fused to almost all of the MTG8 or MTG16. The t(8;21) is responsible for up to 15% of de novo AML cases, however, t(16;21) is relatively rare and has been liked to therapy-related AML. MTG16 is also disrupted by the inv(16)(p13.3q24.3) in 27% of non-Downs syndrome acute megakaryocytic leukemia (AMKL). This translocation fuses the first 3/4th of MTG16 to GLIS2. That two different ETO/MTG family members are disrupted by frequent translocations highlights the key regulatory role of this gene family, but we are just beginning to understand how ETO/MTG8 and MTG16/ETO2 act in transcriptional control and tumorigenesis. Proteomic studies suggest that these proteins link DNA binding proteins such as the “E proteins” E2A and HEB to the transcriptional machinery that controls RNA polymerase II transcriptional initiation and elongation. E-proteins also play a critical role in B- and T-cell development. Here, we have tested mutations uncovered by The Cancer Genome Atlas (TCGA) project in solid cancers that we predicted would affect the association between MTGs and E proteins. We found that the majority of these changes are silent in our assays but selected alterations impaired MTG-dependent interference with E2A-dependent transcriptional activation. This was also true in complementation assays in vivo. Furthermore, the mutations also affected E-protein-MTG interactions. These results suggest that inactivating mutations in MTG family members may contribute to solid cancers in specific instances, and also deregulate normal B- and T-cell development.

Citation Format: Pankaj Acharya, Scott W. Hiebert. Cancer-associated mutations impair the functional association of ETO/MTG family members with E proteins. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A26.
Abstract A43: Opposing roles of the 19S regulatory- and 20S core-proteasomal subunits in controlling sensitivity of multiple myeloma cells to proteasome inhibition.
Authors: Acosta-Alvear Diego , Kampmann Martin , Cho Min Y. , Wild Thomas , Choudhary Chunaram , Shuman Marc A. , Weissman Jonathan S. , Walter Peter
Published: August 2015

Multiple myeloma (MM) remains a devastating incurable disease. Because of their secretory nature, MM cells heavily rely on proteostasis networks to maintain homeostasis. Indeed, paraprotein production by MM cells imposes a protein-folding burden that provides an opportunity for therapeutic intervention: because protein production in MM cells can exceed the buffering capacity of the cell for protein degradation, they are exquisitely sensitive to proteasome inhibitors. While successful, standard-of-care proteasome inhibitors, like bortezomib, remain not curative, as patients ultimately become refractory, suggesting the selection of genetic escape routes. Employing an ultra-complex shRNA library, we performed pooled shRNA screens in RPMI-8226, U-266, and JJN-3 MM cells, and in K-562 leukemia cells, and identified genes whose diminished function impact the cell's response to proteasome inhibition. Unexpectedly, we found that, contrary to knockdown of subunits in the 20S proteasome core-particle, the genetic ablation of subunits within the 19S regulatory-particle led to partial desensitization towards bortezomib. Proteome-wide analyses of in vivo ubiquitylated sites in K562 cells, in which we constitutively ablated a component of the 19S regulatory-particle, revealed potential candidate substrates that were differentially degraded in knockdown cells. We are investigating the mechanistic basis for this differential degradation to pinpoint key substrates mediating proteasome inhibitor resistance. Our results reveal potential escape routes in MM tumor cells to proteasome inhibitors, suggesting new promising targets to pre-empt resistance.

Citation Format: Diego Acosta-Alvear, Martin Kampmann, Min Y. Cho, Thomas Wild, Chunaram Choudhary, Marc A. Shuman, Jonathan S. Weissman, Peter Walter. Opposing roles of the 19S regulatory- and 20S core-proteasomal subunits in controlling sensitivity of multiple myeloma cells to proteasome inhibition. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A43.
Abstract A44: p53 protects against MG132-induced apoptosis in a cell state-dependent manner.
Authors: Legesse-Miller Aster , Cheng Jacky Ho Wa
Published: August 2015

Proteasome inhibition is an important strategy for treating many types of cancer. While proteasome inhibitors induce apoptosis in certain rapidly proliferating cancer cells, some cells enters quiescence and survives. The mechanism by which quiescent cells become resistant to proteasome inhibitors is unknown. The tumor suppressor protein p53 plays an essential role in establishing and maintaining quiescence in human cells. It also regulates several cell survival mechanisms, such as cell cycle arrest, autophagy, and activation of detoxifying enzymes. Here, we investigate the role of p53 in establishing resistance against proteasome inhibitors in quiescent cells. We found that MG132 (a known proteasome inhibitor) treatment induces the accumulation of p53 and its downstream target p21 more in quiescent than in proliferating primary human fibroblasts. shRNA-mediated down-regulation of p53 sensitizes both proliferating and quiescent fibroblasts to MG132-mediated apoptosis and cell death, suggesting that p53 plays a protective role in all cell states. Additional experiments demonstrated that proliferating fibroblasts are susceptible to inhibition of cytoplasmic p53 activity, whereas quiescent fibroblasts are sensitive to inhibition of either nuclear or cytoplasmic p53 activity. These findings suggest that proliferating and quiescent cells depend on different p53-induced survival mechanisms to be protected from proteasome inhibition. Autophagy, an alternative degradation pathway regulated by p53, is also induced in both cell states upon MG132 treatment. Identification of p53-mediated regulatory mechanisms that are activated in quiescent cells in response to proteasome inhibition may help to suggest strategies for improving the effectiveness of proteasome inhibitors in treating cancer.

Citation Format: Aster Legesse-Miller, Jacky Ho Wa Cheng. p53 protects against MG132-induced apoptosis in a cell state-dependent manner. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A44.
Abstract A46: Real time niche tracking of cancer stem cell cycle kinetics using a novel bi-cistronic lentiviral reporter.
Authors: Pineda Gabriel , Lennon Kathleen M. , Lambert-Fliszar Florence , Riso Gennarina L. , Jamieson Catriona
Published: August 2015

Human leukemia stem cells (LSC) are quiescent self-renewing cells that drive leukemic transformation of myeloproliferative neoplasms and myelodysplastic syndromes. Currently, there are few methods to quantitatively visualize cell cycle progression in (LSC) and to interrogate stem cell cycle gene regulatory networks in live cells. Human LSC are characterized as quiescent self-renewing cells and current methods require sequential transduction and clonal selection. These methods and conditions are not feasible for studying primary patient samples and LSC, which are generally limited in cell numbers. To alleviate this challenge, we generated a lentiviral biscistronic vector encoding FUCCI2 (fluorescent ubiquitination-based cell-cycle indicator) probes. We characterized the fidelity of the cell cycle by confirming normal cell division and distinct nuclear staining of either green or red fluorescence in both transiently and stably transduced 293 cells. Further, analysis by flow cytometry of 293 cell confirmed that S/G2/M and G1/G0 phases of cell cycle correlate with expression of fluorescent reporters expressed from the bicistronic lentiviral vector and recapitulate FACS based analysis using traditional Ki67 and 7AAD staining. Recently, using this lentiviral biscistronic reporter we succeeded in tracking cell cycle progression in CD34+ cells from CML primary patient samples co-cultured with SL/M2 cells. Time-lapse confocal imaging of these samples reveals distinct differences within the population of CML cells. In summary using this new Dual FUCCI2 lentiviral reporter makes it possible to track real time cell cycle kinetics in cancer stem cells. Future studies will include measuring LSC, cell cycle gene regulatory networks in live cells at different stages of the cell cycle.

Citation Format: Gabriel Pineda, Kathleen M. Lennon, Florence Lambert-Fliszar, Gennarina L. Riso, Catriona Jamieson. Real time niche tracking of cancer stem cell cycle kinetics using a novel bi-cistronic lentiviral reporter. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr A46.