Showing papers by "Novartis published in 2006"

Five-year follow-up of patients receiving imatinib for chronic myeloid leukemia

Abstract: BACKGROUND: The cause of chronic myeloid leukemia (CML) is a constitutively active BCR-ABL tyrosine kinase. Imatinib inhibits this kinase, and in a short-term study was superior to interferon alfa ...

mTOR Inhibition Induces Upstream Receptor Tyrosine Kinase Signaling and Activates Akt

Abstract: Stimulation of the insulin and insulin-like growth factor I (IGF-I) receptor activates the phosphoinositide-3-kinase/Akt/mTOR pathway causing pleiotropic cellular effects including an mTOR-dependent loss in insulin receptor substrate-1 expression leading to feedback down-regulation of signaling through the pathway. In model systems, tumors exhibiting mutational activation of phosphoinositide-3-kinase/Akt kinase, a common event in cancers, are hypersensitive to mTOR inhibitors, including rapamycin. Despite the activity in model systems, in patients, mTOR inhibitors exhibit more modest antitumor activity. We now show that mTOR inhibition induces insulin receptor substrate-1 expression and abrogates feedback inhibition of the pathway, resulting in Akt activation both in cancer cell lines and in patient tumors treated with the rapamycin derivative, RAD001. IGF-I receptor inhibition prevents rapamycin-induced Akt activation and sensitizes tumor cells to inhibition of mTOR. In contrast, IGF-I reverses the antiproliferative effects of rapamycin in serum-free medium. The data suggest that feedback down-regulation of receptor tyrosine kinase signaling is a frequent event in tumor cells with constitutive mTOR activation. Reversal of this feedback loop by rapamycin may attenuate its therapeutic effects, whereas combination therapy that ablates mTOR function and prevents Akt activation may have improved antitumor activity.

The MicroArray Quality Control (MAQC) project shows inter- and intraplatform reproducibility of gene expression measurements

Abstract: Over the last decade, the introduction of microarray technology has had a profound impact on gene expression research. The publication of studies with dissimilar or altogether contradictory results, obtained using different microarray platforms to analyze identical RNA samples, has raised concerns about the reliability of this technology. The MicroArray Quality Control (MAQC) project was initiated to address these concerns, as well as other performance and data analysis issues. Expression data on four titration pools from two distinct reference RNA samples were generated at multiple test sites using a variety of microarray-based and alternative technology platforms. Here we describe the experimental design and probe mapping efforts behind the MAQC project. We show intraplatform consistency across test sites as well as a high level of interplatform concordance in terms of genes identified as differentially expressed. This study provides a resource that represents an important first step toward establishing a framework for the use of microarrays in clinical and regulatory settings.

Nilotinib in imatinib-resistant CML and Philadelphia chromosome-positive ALL.

Abstract: Background Resistance to imatinib mesylate can occur in chronic myelogenous leukemia (CML) Preclinical in vitro studies have shown that nilotinib (AMN107), a new BCR-ABL tyrosine kinase inhibitor, is more potent than imatinib against CML cells by a factor of 20 to 50 Methods In a phase 1 dose-escalation study, we assigned 119 patients with imatinib-resistant CML or acute lymphoblastic leukemia (ALL) to receive nilotinib orally at doses of 50 mg, 100 mg, 200 mg, 400 mg, 600 mg, 800 mg, and 1200 mg once daily and at 400 mg and 600 mg twice daily Results Common adverse events were myelosuppression, transient indirect hyperbilirubinemia, and rashes Of 33 patients with the blastic phase of disease, 13 had a hematologic response and 9 had a cytogenetic response; of 46 patients with the accelerated phase, 33 had a hematologic response and 22 had a cytogenetic response; 11 of 12 patients with the chronic phase had a complete hematologic remission Conclusions Nilotinib has a relatively favorable safety profil

Oral Fingolimod (FTY720) for Relapsing Multiple Sclerosis

Abstract: Background Fingolimod (FTY720) is a new oral immunomodulating agent under evaluation for the treatment of relapsing multiple sclerosis. Methods We randomly assigned 281 patients to receive oral fingolimod, at a dose of 1.25 mg or 5.0 mg, or a placebo once daily, and we followed these patients for 6 months with magnetic resonance imaging (MRI) and clinical evaluations (core study, months 0 to 6). The primary end point was the total number of gadolinium-enhanced lesions recorded on T1-weighted MRI at monthly intervals for 6 months. In an extension study in which the investigators and patients remained unaware of the dose assignments (months 7 to 12), patients who received placebo underwent randomization again to one of the fingolimod doses. Results A total of 255 patients completed the core study. The median total number of gadolinium-enhanced lesions on MRI was lower with 1.25 mg of fingolimod (1 lesion, P<0.001) and 5.0 mg of fingolimod (3 lesions, P=0.006) than with placebo (5 lesions). The annualized re...

Methods for evaluation of medication adherence and persistence using automated databases.

Abstract: Purpose Our aim was to perform a systematic review of the methods currently being used to assess adherence and persistence in pharmacoepidemiological and pharmacoeconomic studies using automated databases. Methods A MEDLINE search of English language literature was performed to identify studies published between January 1, 1980 and March 31, 2004 that evaluated adherence, compliance, persistence, switching, or discontinuations of medications using automated dispensing data (pharmacy records). Two study investigators independently reviewed the abstracts and articles to determine relevant studies according to specified criteria. Results A total of 136 articles met the criteria for evaluation. The types of measures of adherence and persistence commonly reported include the medication possession ratio and related measures of medication availability (77 studies), discontinuation/continuation (58 studies), switching (34 studies), medication gaps (13 studies), refill compliance (7 studies), and retentiveness/turbulence (4 studies). Specific issues considered include the assessment of exposed time to drug therapy and specification of the follow-up period. Conclusions The terminology, definitions, and methods to determine adherence and persistence differ greatly in the published literature. The appropriateness and choice of the specific measure employed should be determined by the overall goals of the study, as well as the relative advantages and limitations of the measures. Copyright © 2006 John Wiley & Sons, Ltd.

Exogenous induction of cerebral beta-amyloidogenesis is governed by agent and host.

Abstract: Protein aggregation is an established pathogenic mechanism in Alzheimer's disease, but little is known about the initiation of this process in vivo. Intracerebral injection of dilute, amyloid-beta (Abeta)-containing brain extracts from humans with Alzheimer's disease or beta-amyloid precursor protein (APP) transgenic mice induced cerebral beta-amyloidosis and associated pathology in APP transgenic mice in a time- and concentration-dependent manner. The seeding activity of brain extracts was reduced or abolished by Abeta immunodepletion, protein denaturation, or by Abeta immunization of the host. The phenotype of the exogenously induced amyloidosis depended on both the host and the source of the agent, suggesting the existence of polymorphic Abeta strains with varying biological activities reminiscent of prion strains.

A universal vaccine for serogroup B meningococcus

Abstract: Meningitis and sepsis caused by serogroup B meningococcus are two severe diseases that still cause significant mortality. To date there is no universal vaccine that prevents these diseases. In this work, five antigens discovered by reverse vaccinology were expressed in a form suitable for large-scale manufacturing and formulated with adjuvants suitable for human use. The vaccine adjuvanted by aluminum hydroxide induced bactericidal antibodies in mice against 78% of a panel of 85 meningococcal strains representative of the global population diversity. The strain coverage could be increased to 90% and above by the addition of CpG oligonucleotides or by using MF59 as adjuvant. The vaccine has the potential to conquer one of the most devastating diseases of childhood.

Trp ion channels and temperature sensation

Abstract: The abilities to sense environmental and internal temperatures are required for survival, both for maintenance of homeostasis and for avoidance of tissue-damaging noxious temperatures. Vertebrates can sense external physical stimuli via specialized classes of neurons in the peripheral nervous system that project to the skin. Temperaturesensitive neurons can be divided into two classes: innocuous thermosensors (warm or cool) and noxious thermonociceptors (hot or cold). ThermoTRPs, a subset of the transient receptor potential family of ion channels, which are expressed in sensory nerve endings and in skin, respond to distinct thermal thresholds. In this review, we examine the extent to which thermoTRPs are responsible for providing a molecular basis for thermal sensation.

The Unc93b1 mutation 3d disrupts exogenous antigen presentation and signaling via Toll-like receptors 3, 7 and 9.

Abstract: Here we have identified 'triple D' (3d), a recessive N-ethyl-N-nitrosourea-induced mutation and phenotype in which no signaling occurs via the intracellular Toll-like receptors 3, 7 and 9 (sensors for double-stranded RNA, single-stranded RNA and unmethylated DNA, respectively). The 3d mutation also prevented cross-presentation and diminished major histocompatibility complex class II presentation of exogenous antigen; it also caused hypersusceptibility to infection by mouse cytomegalovirus and other microbes. By positional identification, we found 3d to be a missense allele of Unc93b1, which encodes the 12-membrane-spanning protein UNC-93B, a highly conserved molecule found in the endoplasmic reticulum with multiple paralogs in mammals. Innate responses to nucleic acids and exogenous antigen presentation, which both initiate in endosomes, thus seem to depend on an endoplasmic reticulum–resident protein, which suggests communication between these organellar systems.

DNA methyltransferases control telomere length and telomere recombination in mammalian cells.

Abstract: Here, we describe a role for mammalian DNA methyltransferases (DNMTs) in telomere length control. Mouse embryonic stem (ES) cells genetically deficient for DNMT1, or both DNMT3a and DNMT3b have dramatically elongated telomeres compared with wild-type controls. Mammalian telomere repeats (TTAGGG) lack the canonical CpG methylation site. However, we demonstrate that mouse subtelomeric regions are heavily methylated, and that this modification is decreased in DNMT-deficient cells. We show that other heterochromatic marks, such as histone 3 Lys 9 (H3K9) and histone 4 Lys 20 (H4K20) trimethylation, remain at both subtelomeric and telomeric regions in these cells. Lack of DNMTs also resulted in increased telomeric recombination as indicated by sister-chromatid exchanges involving telomeric sequences, and by the presence of 'alternative lengthening of telomeres' (ALT)-associated promyelocytic leukaemia (PML) bodies (APBs). This increased telomeric recombination may lead to telomere-length changes, although our results do not exclude a potential involvement of telomerase and telomere-binding proteins in the aberrant telomere elongation observed in DNMT-deficient cells. Together, these results demonstrate a previously unappreciated role for DNA methylation in maintaining telomere integrity.

Maintenance of self-renewal ability of mouse embryonic stem cells in the absence of DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b.

Abstract: DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b cooperatively regulate cytosine methylation in CpG dinucleotides in mammalian genomes, providing an epigenetic basis for gene silencing and maintenance of genome integrity. Proper CpG methylation is required for the normal growth of various somatic cell types, indicating its essential role in the basic cellular function of mammalian cells. Previous studies using Dnmt1 ‐/‐ or Dnmt3a ‐/‐ Dnmt3b ‐/‐ ES cells, however, have shown that undifferentiated embryonic stem (ES) cells can tolerate hypomethylation for their proliferation. In an attempt to investigate the effects of the complete loss of CpG DNA methyltransferase function, we established mouse ES cells lacking all three of these enzymes by gene targeting. Despite the absence of CpG methylation, as demonstrated by genome-wide methylation analysis, these triple knockout (TKO) ES cells grew robustly and maintained their undifferentiated characteristics. TKO ES cells retained pericentromeric heterochromatin domains marked with methylation at Lys9 of histone H3 and heterochromatin protein-1, and maintained their normal chromosome numbers. Our results indicate that ES cells can maintain stem cell properties and chromosomal stability in the absence of CpG methylation and CpG DNA methyltransferases.

Prolonged diabetes reversal after intraportal xenotransplantation of wild-type porcine islets in immunosuppressed nonhuman primates.

Abstract: Cell-based diabetes therapy requires an abundant cell source. Here, we report reversal of diabetes for more than 100 d in cynomolgus macaques after intraportal transplantation of cultured islets from genetically unmodified pigs without Gal-specific antibody manipulation. Immunotherapy with CD25-specific and CD154-specific monoclonal antibodies, FTY720 (or tacrolimus), everolimus and leflunomide suppressed indirect activation of T cells, elicitation of non-Gal pig-specific IgG antibody, intragraft expression of proinflammatory cytokines and invasion of infiltrating mononuclear cells into islets.

Hypoxia facilitates Alzheimer's disease pathogenesis by up-regulating BACE1 gene expression

Abstract: The molecular mechanism underlying the pathogenesis of the majority of cases of sporadic Alzheimer's disease (AD) is unknown. A history of stroke was found to be associated with development of some AD cases, especially in the presence of vascular risk factors. Reduced cerebral perfusion is a common vascular component among AD risk factors, and hypoxia is a direct consequence of hypoperfusion. Previously we showed that expression of the β-site β-amyloid precursor protein (APP) cleavage enzyme 1 (BACE1) gene BACE1 is tightly controlled at both the transcriptional and translational levels and that increased BACE1 maturation contributes to the AD pathogenesis in Down's syndrome. Here we have identified a functional hypoxia-responsive element in the BACE1 gene promoter. Hypoxia up-regulated β-secretase cleavage of APP and amyloid-β protein (Aβ) production by increasing BACE1 gene transcription and expression both in vitro and in vivo. Hypoxia treatment markedly increased Aβ deposition and neuritic plaque formation and potentiated the memory deficit in Swedish mutant APP transgenic mice. Taken together, our results clearly demonstrate that hypoxia can facilitate AD pathogenesis, and they provide a molecular mechanism linking vascular factors to AD. Our study suggests that interventions to improve cerebral perfusion may benefit AD patients.

Structural basis for the activation of flaviviral NS3 proteases from dengue and West Nile virus.

Abstract: The replication of flaviviruses requires the correct processing of their polyprotein by the viral NS3 protease (NS3pro). Essential for the activation of NS3pro is a 47-residue region of NS2B. Here we report the crystal structures of a dengue NS2B-NS3pro complex and a West Nile virus NS2B-NS3pro complex with a substrate-based inhibitor. These structures identify key residues for NS3pro substrate recognition and clarify the mechanism of NS3pro activation.

Minimizing the risk of reporting false positives in large-scale RNAi screens

Abstract: Large-scale RNA interference (RNAi)-based analyses, very much as other ‘omic’ approaches, have inherent rates of false positives and negatives. The variability in the standards of care applied to validate results from these studies, if left unchecked, could eventually begin to undermine the credibility of RNAi as a powerful functional approach. This Commentary is an invitation to an open discussion started among various users of RNAi to set forth accepted standards that would insure the quality and accuracy of information in the large datasets coming out of genome-scale screens.

Ion formation mechanisms in UV-MALDI

Abstract: Matrix Assisted Laser Desorption/Ionization (MALDI) is a very widely used analytical method, but has been developed in a highly empirical manner. Deeper understanding of ionization mechanisms could help to design better methods and improve interpretation of mass spectra. This review summarizes current mechanistic thinking, with emphasis on the most common MALDI variant using ultraviolet laser excitation. A two-step framework is gaining acceptance as a useful model for many MALDI experiments. The steps are primary ionization during or shortly after the laser pulse, followed by secondary reactions in the expanding plume of desorbed material. Primary ionization in UV-MALDI remains somewhat controversial, the two main approaches are the cluster and pooling/photoionization models. Secondary events are less contentious, ion–molecule reaction thermodynamics and kinetics are often invoked, but details differ. To the extent that local thermal equilibrium is approached in the plume, the mass spectra may be straightforwardly interpreted in terms of charge transfer thermodynamics.

AMN107 (nilotinib): a novel and selective inhibitor of BCR-ABL.

Abstract: Chronic myelogenous leukaemia (CML) and Philadelphia chromosome positive (Ph+) acute lymphoblastic leukaemia (ALL) are caused by the BCR-ABL oncogene. Imatinib inhibits the tyrosine kinase activity of the BCR-ABL protein and is an effective, frontline therapy for chronic-phase CML. However, accelerated or blast-crisis phase CML patients and Ph+ ALL patients often relapse due to drug resistance resulting from the emergence of imatinib-resistant point mutations within the BCR-ABL tyrosine kinase domain. This has stimulated the development of new kinase inhibitors that are able to over-ride resistance to imatinib. The novel, selective BCR-ABL inhibitor, AMN107, was designed to fit into the ATP-binding site of the BCR-ABL protein with higher affinity than imatinib. In addition to being more potent than imatinib (IC50<30 nM) against wild-type BCR-ABL, AMN107 is also significantly active against 32/33 imatinib-resistant BCR-ABL mutants. In preclinical studies, AMN107 demonstrated activity in vitro and in vivo against wild-type and imatinib-resistant BCR-ABL-expressing cells. In phase I/II clinical trials, AMN107 has produced haematological and cytogenetic responses in CML patients, who either did not initially respond to imatinib or developed imatinib resistance. Dasatinib (BMS-354825), which inhibits Abl and Src family kinases, is another promising new clinical candidate for CML that has shown good efficacy in CML patients. In this review, the early characterisation and development of AMN107 is discussed, as is the current status of AMN107 in clinical trials for imatinib-resistant CML and Ph+ ALL. Future trends investigating prediction of mechanisms of resistance to AMN107, and how and where AMN107 is expected to fit into the overall picture for treatment of early-phase CML and imatinib-refractory and late-stage disease are discussed.

Pili in Gram-positive pathogens

Abstract: Most bacterial pathogens have long filamentous structures known as pili or fimbriae extending from their surface. These structures are often involved in the initial adhesion of the bacteria to host tissues during colonization. In Gram-negative bacteria, pili are typically formed by non-covalent interactions between pilin subunits. By contrast, the recently discovered pili in Gram-positive pathogens are formed by covalent polymerization of adhesive pilin subunits. Evidence from studies of pili in the three principal streptococcal pathogens of humans indicates that the genes that encode the pilin subunits and the enzymes that are required for the assembly of these subunits into pili have been acquired en bloc by the horizontal transfer of a pathogenicity island.

Phase I/II Study of Imatinib Mesylate for Recurrent Malignant Gliomas: North American Brain Tumor Consortium Study 99-08

Abstract: Purpose: Phase I: To determine the maximum tolerated doses, toxicities, and pharmacokinetics of imatinib mesylate (Gleevec) in patients with malignant gliomas taking enzyme-inducing antiepileptic drugs (EIAED) or not taking EIAED. Phase II: To determine the therapeutic efficacy of imatinib. Experimental Design: Phase I component used an interpatient dose escalation scheme. End points of the phase II component were 6-month progression-free survival and response. Results: Fifty patients enrolled in the phase I component (27 EIAED and 23 non-EIAED). The maximum tolerated dose for non-EIAED patients was 800 mg/d. Dose-limiting toxicities were neutropenia, rash, and elevated alanine aminotransferase. EIAED patients received up to 1,200 mg/d imatinib without developing dose-limiting toxicity. Plasma exposure of imatinib was reduced by ∼68% in EIAED patients compared with non-EIAED patients. Fifty-five non-EIAED patients (34 glioblastoma multiforme and 21 anaplastic glioma) enrolled in the phase II component. Patients initially received 800 mg/d imatinib; 15 anaplastic glioma patients received 600 mg/d after hemorrhages were observed. There were 2 partial response and 6 stable disease among glioblastoma multiforme patients and 0 partial response and 5 stable disease among anaplastic glioma patients. Six-month progression-free survival was 3% for glioblastoma multiforme and 10% for anaplastic glioma patients. Five phase II patients developed intratumoral hemorrhages. Conclusions: Single-agent imatinib has minimal activity in malignant gliomas. CYP3A4 inducers, such as EIAEDs, substantially decreased plasma exposure of imatinib and should be avoided in patients receiving imatinib for chronic myelogenous leukemia and gastrointestinal stromal tumors. The evaluation of the activity of combination regimens incorporating imatinib is under way in phase II trials.

Anti-miRNA oligonucleotides (AMOs): ammunition to target miRNAs implicated in human disease?

Abstract: MicroRNAs (miRNAs) are endogenous 19-25 nucleotide RNAs that have recently emerged as a novel class of important gene-regulatory molecules involved in many critical developmental and cellular functions. miRNAs have been implicated in the pathogenesis of several human diseases, such as neurodegenerative disorders, cancer, and more recently in viral and metabolic diseases. Unraveling the roles of miRNAs in cellular processes linked to human diseases will lead to novel opportunities for the regulation of protein function and will help to evaluate their potential for therapeutic intervention. Approaches to interfere with miRNA function in vitro and in vivo based on synthetic anti-miRNA oligonucleotides (AMOs) are discussed in this review.

The 7 TM G-protein-coupled receptor target family.

Abstract: Chemical biology approaches have a long history in the exploration of the G-protein-coupled receptor (GPCR) family, which represents the largest and most important group of targets for therapeutics. The analysis of the human genome revealed a significant number of new members with unknown physiological function which are today the focus of many reverse pharmacology drug-discovery programs. As the seven hydrophobic transmembrane segments are a defining common structural feature of these receptors, and as signaling through heterotrimeric G proteins is not demonstrated in all cases, these proteins are also referred to as seven transmembrane (7 TM) or serpentine receptors. This review summarizes important historic milestones of GPCR research, from the beginning, when pharmacology was mainly descriptive, to the age of modern molecular biology, with the cloning of the first receptor and now the availability of the entire human GPCR repertoire at the sequence and protein level. It shows how GPCR-directed drug discovery was initially based on the careful testing of a few specifically made chemical compounds and is today pursued with modern drug-discovery approaches, including combinatorial library design, structural biology, molecular informatics, and advanced screening technologies for the identification of new compounds that activate or inhibit GPCRs specifically. Such compounds, in conjunction with other new technologies, allow us to study the role of receptors in physiology and medicine, and will hopefully result in novel therapies. We also outline how basic research on the signaling and regulatory mechanisms of GPCRs is advancing, leading to the discovery of new GPCR-interacting proteins and thus opening new perspectives for drug development. Practical examples from GPCR expression studies, HTS (high-throughput screening), and the design of monoamine-related GPCR-focused combinatorial libraries illustrate ongoing GPCR chemical biology research. Finally, we outline future progress that may relate today's discoveries to the development of new medicines.

Therapeutic approaches to Alzheimer's disease.

Abstract: Alzheimer's disease is an age-related progressive neurodegenerative disorder with an enormous unmet medical need. It is the most common form of dementia affecting ∼5% of adults over 65 years. In view of our ageing society the number of patients, as well as the economical and social impact, is expected to grow dramatically in the future. Currently available medications appear to be able to produce moderate symptomatic benefits but not to stop disease progression. The search for novel therapeutic approaches targeting the presumed underlying pathogenic mechanisms has been a major focus of research and it is expected that novel medications with disease-modifying properties will emerge from these efforts in the future. In this review, currently available drugs as well as novel therapeutic strategies, in particular those targeting amyloid and tau pathologies, are discussed.

Allosteric inhibitors of Bcr-abl–dependent cell proliferation

Abstract: Chronic myelogenous leukemia (CML) is a myeloproliferative disorder characterized at the molecular level by the expression of Bcr-abl, a 210-kDa fusion protein with deregulated tyrosine kinase activity. Encouraged by the clinical validation of Bcr-abl as the target for the treatment of CML by imatinib, we sought to identify pharmacological agents that could target this kinase by a distinct mechanism. We report the discovery of a new class of Bcr-abl inhibitors using an unbiased differential cytotoxicity screen of a combinatorial kinase-directed heterocycle library. Compounds in this class (exemplified by GNF-2) show exclusive antiproliferative activity toward Bcr-abl-transformed cells, with potencies similar to imatinib, while showing no inhibition of the kinase activity of full-length or catalytic domain of c-abl. We propose that this new class of compounds inhibits Bcr-abl kinase activity through an allosteric non-ATP competitive mechanism.