Showing papers by "University of Hamburg published in 2006"

The amphibian tree of life

Abstract: The evidentiary basis of the currently accepted classification of living amphibians is discussed and shown not to warrant the degree of authority conferred on it by use and tradition. A new taxonomy of living amphibians is proposed to correct the deficiencies of the old one. This new taxonomy is based on the largest phylogenetic analysis of living Amphibia so far accomplished. We combined the comparative anatomical character evidence of Haas (2003) with DNA sequences from the mitochondrial transcription unit H1 (12S and 16S ribosomal RNA and tRNAValine genes, ≈ 2,400 bp of mitochondrial sequences) and the nuclear genes histone H3, rhodopsin, tyrosinase, and seven in absentia, and the large ribosomal subunit 28S (≈ 2,300 bp of nuclear sequences; ca. 1.8 million base pairs; x = 3.7 kb/terminal). The dataset includes 532 terminals sampled from 522 species representative of the global diversity of amphibians as well as seven of the closest living relatives of amphibians for outgroup comparisons. The...

Reconstructing the early evolution of Fungi using a six-gene phylogeny

Abstract: The ancestors of fungi are believed to be simple aquatic forms with flagellated spores, similar to members of the extant phylum Chytridiomycota (chytrids). Current classifications assume that chytrids form an early-diverging clade within the kingdom Fungi and imply a single loss of the spore flagellum, leading to the diversification of terrestrial fungi. Here we develop phylogenetic hypotheses for Fungi using data from six gene regions and nearly 200 species. Our results indicate that there may have been at least four independent losses of the flagellum in the kingdom Fungi. These losses of swimming spores coincided with the evolution of new mechanisms of spore dispersal, such as aerial dispersal in mycelial groups and polar tube eversion in the microsporidia (unicellular forms that lack mitochondria). The enigmatic microsporidia seem to be derived from an endoparasitic chytrid ancestor similar to Rozella allomycis, on the earliest diverging branch of the fungal phylogenetic tree.

Femtosecond diffractive imaging with a soft-X-ray free-electron laser

Abstract: Theory predicts that with an ultrashort and extremely bright coherent X-ray pulse, a single diffraction pattern may be recorded from a large macromolecule, a virus, or a cell before the sample explodes and turns into a plasma. Here we report the first experimental demonstration of this principle using the FLASH soft X-ray free-electron laser. An intense 25 fs, 4 x 10{sup 13} W/cm{sup 2} pulse, containing 10{sup 12} photons at 32 nm wavelength, produced a coherent diffraction pattern from a nano-structured non-periodic object, before destroying it at 60,000 K. A novel X-ray camera assured single photon detection sensitivity by filtering out parasitic scattering and plasma radiation. The reconstructed image, obtained directly from the coherent pattern by phase retrieval through oversampling, shows no measurable damage, and extends to diffraction-limited resolution. A three-dimensional data set may be assembled from such images when copies of a reproducible sample are exposed to the beam one by one.

Engineered heart tissue grafts improve systolic and diastolic function in infarcted rat hearts

Abstract: The concept of regenerating diseased myocardium by implantation of tissue-engineered heart muscle is intriguing, but convincing evidence is lacking that heart tissues can be generated at a size and with contractile properties that would lend considerable support to failing hearts. Here we created large (thickness/diameter, 1-4 mm/15 mm), force-generating engineered heart tissue from neonatal rat heart cells. Engineered heart tissue formed thick cardiac muscle layers when implanted on myocardial infarcts in immune-suppressed rats. When evaluated 28 d later, engineered heart tissue showed undelayed electrical coupling to the native myocardium without evidence of arrhythmia induction. Moreover, engineered heart tissue prevented further dilation, induced systolic wall thickening of infarcted myocardial segments and improved fractional area shortening of infarcted hearts compared to controls (sham operation and noncontractile constructs). Thus, our study provides evidence that large contractile cardiac tissue grafts can be constructed in vitro, can survive after implantation and can support contractile function of infarcted hearts.

A Biologic Definition of Burkitt's Lymphoma from Transcriptional and Genomic Profiling

Abstract: Background The distinction between Burkitt’s lymphoma and diffuse large-B-cell lymphoma is unclear. We used transcriptional and genomic profiling to define Burkitt’s lymphoma more precisely and to distinguish subgroups in other types of mature aggressive B-cell lymphomas. Methods We performed gene-expression profiling using Affymetrix U133A GeneChips with RNA from 220 mature aggressive B-cell lymphomas, including a core group of 8 Burkitt’s lymphomas that met all World Health Organization (WHO) criteria. A molecular signature for Burkitt’s lymphoma was generated, and chromosomal abnormalities were detected with interphase fluorescence in situ hybridization and array-based comparative genomic hybridization. Results We used the molecular signature for Burkitt’s lymphoma to identify 44 cases: 11 had the morphologic features of diffuse large-B-cell lymphomas, 4 were unclassifiable mature aggressive B-cell lymphomas, and 29 had a classic or atypical Burkitt’s morphologic appearance. Also, five did not have a detectable IG-myc Burkitt’s translocation, whereas the others contained an IG-myc fusion, mostly in simple karyotypes. Of the 176 lymphomas without the molecular signature for Burkitt’s lymphoma, 155 were diffuse large-B-cell lymphomas. Of these 155 cases, 21 percent had a chromosomal breakpoint at the myc locus associated with complex chromosomal changes and an unfavorable clinical course. Conclusions Our molecular definition of Burkitt’s lymphoma clarifies and extends the spectrum of the WHO criteria for Burkitt’s lymphoma. In mature aggressive B-cell lymphomas without a gene signature for Burkitt’s lymphoma, chromosomal breakpoints at the myc locus were associated with an adverse clinical outcome.

In Praise of Convergent Thinking

Abstract: Free production of variability through unfettered divergent thinking holds out the seductive promise of effortless creativity but runs the risk of generating only quasicreativity or pseudocreativity if it is not adapted to reality. Therefore, creative thinking seems to involve 2 components: generation of novelty (via divergent thinking) and evaluation of the novelty (via convergent thinking). In the area of convergent thinking, knowledge is of particular importance: It is a source of ideas, suggests pathways to solutions, and provides criteria of effectiveness and novelty. The way in which the 2 kinds of thinking work together can be understood in terms of thinking styles or of phases in the generation of creative products. In practical situations, divergent thinking without convergent thinking can cause a variety of problems including reckless change. Nonetheless, care must be exercised by those who sing the praises of convergent thinking: Both too little and too much is bad for creativity.

Non-invasive brain stimulation: a new strategy to improve neurorehabilitation after stroke?

Abstract: Summary Background Motor impairment resulting from chronic stroke can have extensive physical, psychological, financial, and social implications despite available neurorehabilitative treatments. Recent studies in animals showed that direct epidural stimulation of the primary motor cortex surrounding a small infarct in the lesioned hemisphere (M1 lesioned hemisphere ) elicits improvements in motor function. Recent developments In human beings, proof of principle studies from different laboratories showed that non-invasive transcranial magnetic stimulation and direct current stimulation that upregulate excitability within M1 lesioned hemisphere or downregulate excitability in the intact hemisphere (M1 intact hemisphere ) results in improvement in motor function in patients with stroke. Possible mechanisms mediating these effects can include the correction of abnormally persistent interhemispheric inhibitory drive from M1 intact hemisphere to M1 lesioned hemisphere in the process of generation of voluntary movements by the paretic hand, a disorder correlated with the magnitude of impairment. In this paper we review these mechanistically oriented interventional approaches. What next? These findings suggest that transcranial magnetic stimulation and transcranial direct current stimulation could develop into useful adjuvant strategies in neurorehabilitation but have to be further assessed in multicentre clinical trials.

Buried alive: how osteoblasts become osteocytes.

Abstract: During osteogenesis, osteoblasts lay down osteoid and transform into osteocytes embedded in mineralized bone matrix. Despite the fact that osteocytes are the most abundant cellular component of bone, little is known about the process of osteoblast-to-osteocyte transformation. What is known is that osteoblasts undergo a number of changes during this transformation, yet retain their connections to preosteoblasts and osteocytes. This review explores the osteoblast-to-osteocyte transformation during intramembranous ossification from both morphological and molecular perspectives. We investigate how these data support five schemes that describe how an osteoblast could become entrapped in the bone matrix (in mammals) and suggest one of the five scenarios that best fits as a model. Those osteoblasts on the bone surface that are destined for burial and destined to become osteocytes slow down matrix production compared to neighbouring osteoblasts, which continue to produce bone matrix. That is, cells that continue to produce matrix actively bury cells producing less or no new bone matrix (passive burial). We summarize which morphological and molecular changes could be used as characters (or markers) to follow the transformation process.

EFNS guideline on the drug treatment of migraine--revised report of an EFNS task force.

Abstract: Migraine is one of the most frequent disabling neurological conditions with a major impact on the patients' quality of life. To give evidence-based or expert recommendations for the different drug treatment procedures of the different migraine syndromes based on a literature search and an consensus in an expert panel. All available medical reference systems were screened for all kinds of clinical studies on migraine with and without aura and on migraine-like syndromes. The findings in these studies were evaluated according to the recommendations of the EFNS resulting in level A,B, or C recommendations and good practice points. For the acute treatment of migraine attacks, oral non-steroidal anti-inflammatory drugs (NSAIDs) and triptans are recommended. The administration should follow the concept of stratified treatment. Before intake of NSAIDs and triptans, oral metoclopramide or domperidon is recommended. In very severe attacks, intravenous acetylsalicylic acid or subcutaneous sumatriptan are drugs of first choice. A status migrainosus can probably be treated by steroids. For the prophylaxis of migraine, betablockers (propranolol and metoprolol), flunarizine, valproic acid, and topiramate are drugs of first choice. Drugs of second choice for migraine prophylaxis are amitriptyline, naproxen, petasites, and bisoprolol.

Observations of the Crab nebula with HESS

Abstract: The Crab nebula was observed with the H.E.S.S. stereoscopic Cherenkov-telescope array between October 2003 and January 2005 for a total of 22.9 hours (after data quality selection). Observations were made with three operational telescopes in late 2003 and with the complete 4 telescope array in January - February 2004 and October 2004 - January 2005. The observations are discussed and used as an example to detail the flux and spectral analysis procedures of H.E.S.S., and to evaluate the systematic uncertainties in H.E.S.S. flux measurements. The flux and spectrum of gamma-rays from the source are calculated on run-by-run and monthly time-scales, and a correction is applied for long-term variations in the detector sensitivity. Comparisons of the measured flux and spectrum over the observation period, along with the results from a number of different analysis procedures are used to estimate systematic uncertainties in the measurements. The energy spectrum is found to follow a power law with an exponential cutoff, with photon index $\Gamma = 2.39 \pm 0.03\stat$ and cutoff energy $E_{c} = (14.3 \pm 2.1\stat) \textrm{TeV}$ between 440 GeV and 40 TeV. The observed integral flux above 1 TeV is $(2.26 \pm 0.08\stat) \times 10^{-11} cm^{-2} s^{-1}$. The estimated systematic error on the flux measurement is estimated to be 20%, while the estimated systematic error on the spectral slope is 0.1.

A classification of assembly line balancing problems

Abstract: Assembly lines are special flow-line production systems which are of great importance in the indus-trial production of high quantity standardized commodities. Recently, assembly lines even gained importance in low volume production of customized products (mass-customization). Due to high capital requirements when installing or redesigning a line, its configuration planning is of great rele-vance for practitioners. Accordingly, this attracted attention of plenty researchers, who tried to sup-port real-world configuration planning by suited optimization models (assembly line balancing prob-lems). In spite of the enormous academic effort in assembly line balancing, there remains a consider-able gap between requirements of real configuration problems and the status of research. To ease communication between researchers and practitioners, we provide a classification scheme of assem-bly line balancing. This is a valuable step in identifying remaining research challenges which might contribute to closing the gap.

Familial and acquired hemophagocytic lymphohistiocytosis

Abstract: Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening condition of severe hyperinflammation caused by the uncontrolled proliferation of activated lymphocytes and histiocytes secreting high amounts of inflammatory cytokines. Cardinal signs and symptoms are prolonged fever, hepatosplenomegaly and pancytopenia. Characteristic biochemical markers include elevated triglycerides, ferritin and low fibrinogen. HLH occurs on the basis of various inherited or acquired immune deficiencies. Impaired function of natural killer (NK) cells and cytotoxic T-cells (CTL) is shared by all forms of HLH. Genetic HLH occurs in familial forms (FHLH) in which HLH is the primary and only manifestation, and in association with the immune deficiencies Chediak-Higashi syndrome 1 (CHS 1), Griscelli syndrome 2 (GS 2) and x-linked lymphoproliferative syndrome (XLP), in which HLH is a sporadic event. Most patients with acquired HLH have no known underlying immune deficiency. Both acquired and genetic forms are triggered by infections, mostly viral, or other stimuli. HLH also occurs as a complication of rheumatic diseases (macrophage activation syndrome) and of malignancies. Several genetic defects causing FHLH have recently been discovered and have elucidated the pathophysiology of HLH. The immediate aim of therapy in genetic and acquired HLH is suppression of the severe hyperinflammation, which can be achieved with immunosuppressive/immunomodulatary agents and cytostatic drugs. Patients with genetic forms have to undergo stem cell transplantation to exchange the defective immune system with normally functioning immune effector cells. In conclusion, awareness of the clinical symptoms and of the diagnostic criteria of HLH is crucial in order not to overlook HLH and to start life-saving therapy in time.

A low level of extragalactic background light as revealed by gamma-rays from blazars.

Abstract: The diffuse Extragalactic Background Light (EBL) contains unique information about the epochs of formation and the history of evolution of galaxies. Unfortunately, direct measurements are subject to large systematic uncertainties due to the difficulties in the accurate model-based subtraction of the bright foregrounds. An alternative approach is based on the detection and identification of EBL absorption features in high-energy spectra of objects of known redshift. Here we exploit this method on the blazars H 2356-309 (z=0.165) and 1ES 1101-232 (z=0.186), newly discovered at TeV energies by the H.E.S.S. Collaboration. They are the most distant sources with measured spectra known so far at these energies. Their hard spectra provide the most stringent upper limit to date on the EBL in the Opt--NIR band, which appears significantly lower than expected from the current "direct" estimates and very close to the absolute lower limit represented by the integrated light of resolved galaxies. In addition to important cosmological implications, this result shows that the intergalactic space is more transparent to gamma-rays than previously thought, expanding the horizon of the TeV Universe.

Control of Electron Localization in Molecular Dissociation

Abstract: We demonstrated how the subcycle evolution of the electric field of light can be used to control the motion of bound electrons. Results are presented for the dissociative ionization of deuterium molecules (D2 ⇒ D+ + D), where asymmetric ejection of the ionic fragment reveals that light-driven intramolecular electronic motion before dissociation localizes the electron on one of the two D+ ions in a controlled way. The results extend subfemtosecond electron control to molecules and provide evidence of its usefulness in controlling reaction dynamics.

Drivers of Brand Extension Success

Abstract: The research presented in this article addresses the issue of the significance and relative importance of the determinants of extension success by simultaneously investigating ten success factors. The empirical analysis considers the direct relationships between success factors and extension success, the structural relationships among investigated factors, and moderating effects. The authors find that fit between the parent brand and an extension product is the most important driver of brand extension success, followed by marketing support, parent-brand conviction, retailer acceptance, and parent-brand experience. The authors also find several important structural relationships among the investigated success factors (e.g., marketing support → fit → retailer acceptance → extension success). Finally, the interaction terms of fit with the quality of the parent brand and with parent-brand conviction are statistically significant, albeit of relatively low importance.

The H.E.S.S. Survey of the Inner Galaxy in Very High Energy Gamma Rays

Abstract: We report on a survey of the inner part of the Galactic plane in very high energy gamma rays with the H.E.S.S. Cerenkov telescope system. The Galactic plane between +/-30° in longitude and +/-3° in latitude relative to the Galactic center was observed in 500 pointings for a total of 230 hr, reaching an average flux sensitivity of 2% of the Crab Nebula at energies above 200 GeV. Fourteen previously unknown sources were detected at a significance level greater than 4 σ after accounting for all trials involved in the search. Initial results on the eight most significant of these sources were already reported elsewhere (Aharonian and coworkers). Here we present detailed spectral and morphological information for all the new sources, along with a discussion on possible counterparts in other wavelength bands. The distribution in Galactic latitude of the detected sources appears to be consistent with a scale height in the Galactic disk for the parent population smaller than 100 pc, consistent with expectations for supernova remnants and/or pulsar wind nebulae.

Manipulation of Host Hepatocytes by the Malaria Parasite for Delivery into Liver Sinusoids

Abstract: The merozoite stage of the malaria parasite that infects erythrocytes and causes the symptoms of the disease is initially formed inside host hepatocytes. However, the mechanism by which hepatic merozoites reach blood vessels (sinusoids) in the liver and escape the host immune system before invading erythrocytes remains unknown. Here, we show that parasites induce the death and the detachment of their host hepatocytes, followed by the budding of parasite-filled vesicles (merosomes) into the sinusoid lumen. Parasites simultaneously inhibit the exposure of phosphatidylserine on the outer leaflet of host plasma membranes, which act as "eat me" signals to phagocytes. Thus, the hepatocyte-derived merosomes appear to ensure both the migration of parasites into the bloodstream and their protection from host immunity.

Mechanisms of placebo analgesia: rACC recruitment of a subcortical antinociceptive network.

Abstract: Placebo analgesia is one of the most striking examples of the cognitive modulation of pain perception and the underlying mechanisms are finally beginning to be understood. According to pharmacological studies, the endogenous opioid system is essential for placebo analgesia. Recent functional imaging data provides evidence that the rostral anterior cingulate cortex (rACC) represents a crucial cortical area for this type of endogenous pain control. We therefore hypothesized that placebo analgesia recruits other brain areas outside the rACC and that interactions of the rACC with these brain areas mediate opioid-dependent endogenous antinociception as part of a top-down mechanism. Nineteen healthy subjects received and rated painful laser stimuli to the dorsum of both hands, one of them treated with a fake analgesic cream (placebo). Painful stimulation was preceded by an auditory cue, indicating the side of the next laser stimulation. BOLD-responses to the painful laser-stimulation during the placebo and no-placebo condition were assessed using event-related fMRI. After having confirmed placebo related activity in the rACC, a connectivity analysis identified placebo dependent contributions of rACC activity with bilateral amygdalae and the periaqueductal gray (PAG). This finding supports the view that placebo analgesia depends on the enhanced functional connectivity of the rACC with subcortical brain structures that are crucial for conditioned learning and descending inhibition of nociception.

Dissociable Systems for Gain- and Loss-Related Value Predictions and Errors of Prediction in the Human Brain

Abstract: Midbrain dopaminergic neurons projecting to the ventral striatum code for reward magnitude and probability during reward anticipation and then indicate the difference between actual and predicted outcome. It has been questioned whether such a common system for the prediction and evaluation of reward exists in humans. Using functional magnetic resonance imaging and a guessing task in two large cohorts, we are able to confirm ventral striatal responses coding both reward probability and magnitude during anticipation, permitting the local computation of expected value (EV). However, the ventral striatum only represented the gain-related part of EV (EV+). At reward delivery, the same area shows a reward probability and magnitude-dependent prediction error signal, best modeled as the difference between actual outcome and EV+. In contrast, loss-related expected value (EV−) and the associated prediction error was represented in the amygdala. Thus, the ventral striatum and the amygdala distinctively process the value of a prediction and subsequently compute a prediction error for gains and losses, respectively. Therefore, a homeostatic balance of both systems might be important for generating adequate expectations under uncertainty. Prevalence of either part might render expectations more positive or negative, which could contribute to the pathophysiology of mood disorders like major depression.

Discovery of very-high-energy |[gamma]|-rays from the Galactic Centre ridge

Abstract: Events at the centre of our Galaxy are key to our understanding of high-energy processes in the Universe, since it contains examples of virtually every type of exotic object known to astronomers. The very-high-energy γ-ray emission from the Galactic Centre region has now been measured using HESS, the High Energy Stereoscopic System recently constructed in Namibia, South West Africa. HESS operates at energies above the regime accessible to satellite-based detectors, taking γ-ray astronomy into new territory. The results show that these clouds are glowing in very high energy γ-rays. The glow is caused by constant bombardment of the clouds by cosmic rays — probably protons and nuclei — produced close to the central black hole or in the expanding blast waves of supernova explosions. The source of Galactic cosmic rays (with energies up to 1015 eV) remains unclear, although it is widely believed that they originate in the shock waves of expanding supernova remnants1,2. At present the best way to investigate their acceleration and propagation is by observing the γ-rays produced when cosmic rays interact with interstellar gas3. Here we report observations of an extended region of very-high-energy (> 1011 eV) γ-ray emission correlated spatially with a complex of giant molecular clouds in the central 200 parsecs of the Milky Way. The hardness of the γ-ray spectrum and the conditions in those molecular clouds indicate that the cosmic rays giving rise to the γ-rays are likely to be protons and nuclei rather than electrons. The energy associated with the cosmic rays could have come from a single supernova explosion around 104 years ago.

Genetic analysis of cavefish reveals molecular convergence in the evolution of albinism

Abstract: The genetic basis of vertebrate morphological evolution has traditionally been very difficult to examine in naturally occurring populations. Here we describe the generation of a genome-wide linkage map to allow quantitative trait analysis of evolutionarily derived morphologies in the Mexican cave tetra, a species that has, in a series of independent caves, repeatedly evolved specialized characteristics adapted to a unique and well-studied ecological environment. We focused on the trait of albinism and discovered that it is linked to Oca2, a known pigmentation gene, in two cave populations. We found different deletions in Oca2 in each population and, using a cell-based assay, showed that both cause loss of function of the corresponding protein, OCA2. Thus, the two cave populations evolved albinism independently, through similar mutational events.

Polypeptides and 100 years of chemistry of alpha-amino acid N-carboxyanhydrides.

Abstract: Syntheses and polymerizations of alpha-amino acid N-carboxyanhydrides (NCAs) were reported for the first time by Hermann Leuchs in 1906. Since that time, these cyclic and highly reactive amino acid derivatives were used for stepwise peptide syntheses but mainly for the formation of polypeptides by ring-opening polymerizations. This review summarizes the literature after 1985 and reports on new aspects of the polymerization processes, such as the formation of cyclic polypeptides or novel organometal catalysts. Polypeptides with various architectures, such as diblock, triblock, and multiblock sequences, and star-shaped or dendritic structures are also mentioned. Furthermore, lyotropic and thermotropic liquid-crystalline polypeptides will be discussed and the role of polypeptides as drugs or drug carriers are reviewed. Finally, the hypothetical role of NCAs in molecular evolution on the prebiotic Earth is discussed.

Search for neutral MSSM Higgs bosons at LEP

Abstract: The four LEP collaborations, ALEPH, DELPHI, L3 and OPAL, have searched for the neutral Higgs bosons which are predicted by the Minimal Supersymmetric standard model (MSSM). The data of the four collaborations are statistically combined and examined for their consistency with the background hypothesis and with a possible Higgs boson signal. The combined LEP data show no significant excess of events which would indicate the production of Higgs bosons. The search results are used to set upper bounds on the cross-sections of various Higgs-like event topologies. The results are interpreted within the MSSM in a number of “benchmark” models, including CP-conserving and CP-violating scenarios. These interpretations lead in all cases to large exclusions in the MSSM parameter space. Absolute limits are set on the parameter cosβ and, in some scenarios, on the masses of neutral Higgs bosons.

Vascular wall resident progenitor cells: a source for postnatal vasculogenesis.

Abstract: Here, we report the existence of endothelial precursor (EPC) and stem cells in a distinct zone of the vascular wall that are capable to differentiate into mature endothelial cells, hematopoietic and local immune cells, such as macrophages. This zone has been identified to be localized between smooth muscle and adventitial layer of human adult vascular wall. It predominantly contains CD34-positive (+) but CD31-negative (-) cells, which also express VEGFR2 and TIE2. Only few cells in this zone of the vascular wall are positive for CD45. In a ring assay using the fragments of human internal thoracic artery (HITA), we show here that the CD34+ cells of the HITA-wall form capillary sprouts ex vivo and are apparently recruited for capillary formation by tumor cells. New vessels formed by these vascular wall resident EPCs express markers for angiogenically activated endothelial cells, such as CEACAM1, and also for mature endothelial cells, such as VE-cadherin or occludin. Vascular wall areas containing EPCs are found in large and middle sized arteries and veins of all organs studied here. These data suggest the existence of a ;vasculogenic zone' in the wall of adult human blood vessels, which may serve as a source for progenitor cells for postnatal vasculogenesis, contributing to tumor vascularization and local immune response.

A combined computational and microarray-based approach identifies novel microRNAs encoded by human gamma-herpesviruses.

Abstract: We have developed an approach to identify microRNAs (miRNAs) that is based on bioinformatics and array-based technologies, without the use of cDNA cloning. The approach, designed for use on genomes of small size (<2 Mb), was tested on cells infected by either of two lymphotropic herpesviruses, KSHV and EBV. The viral genomes were scanned computationally for pre-miRNAs using an algorithm (VMir) we have developed. Candidate hairpins suggested by this analysis were then synthesized as oligonucleotides on microarrays, and the arrays were hybridized with small RNAs from infected cells. Candidate miRNAs that scored positive on the arrays were then subjected to confirmatory Northern blot analysis. Using this approach, 10 of the known KSHV pre-miRNAs were identified, as well as a novel pre-miRNA that had earlier escaped detection. This method also led to the identification of seven new EBV-encoded pre-miRNAs; by using additional computational approaches, we identified a total of 18 new EBV pre-miRNAs that produce 22 mature miRNA molecules, thereby more than quadrupling the total number of hitherto known EBV miRNAs. The advantages and limitations of the approach are discussed.

Dasatinib (BMS-354825), a dual SRC/ABL kinase inhibitor, inhibits the kinase activity of wild-type, juxtamembrane, and activation loop mutant KIT isoforms associated with human malignancies.

Abstract: Activating mutations of the activation loop of KIT are associated with certain human neoplasms, including the majority of patients with systemic mast cell disorders, as well as cases of seminoma, acute myelogenous leukemia (AML), and gastrointestinal stromal tumors (GISTs) The small-molecule tyrosine kinase inhibitor imatinib mesylate is a potent inhibitor of wild-type (WT) KIT and certain mutant KIT isoforms and has become the standard of care for treating patients with metastatic GIST However, KIT activation loop mutations involving codon D816 that are typically found in AML, systemic mastocytosis, and seminoma are insensitive to imatinib mesylate (IC50 > 5-10 micromol/L), and acquired KIT activation loop mutations can be associated with imatinib mesylate resistance in GIST Dasatinib (formerly BMS-354825) is a small-molecule, ATP-competitive inhibitor of SRC and ABL tyrosine kinases with potency in the low nanomolar range Some small-molecule SRC/ABL inhibitors also have potency against WT KIT kinase Therefore, we hypothesized that dasatinib might inhibit the kinase activity of both WT and mutant KIT isoforms We report herein that dasatinib potently inhibits WT KIT and juxtamembrane domain mutant KIT autophosphorylation and KIT-dependent activation of downstream pathways important for cell viability and cell survival, such as Ras/mitogen-activated protein kinase, phosphoinositide 3-kinase/Akt, and Janus-activated kinase/signal transducers and activators of transcription Furthermore, dasatinib is a potent inhibitor of imatinib-resistant KIT activation loop mutants and induces apoptosis in mast cell and leukemic cell lines expressing these mutations (potency against KIT D816Y >> D816F > D816V) Our studies suggest that dasatinib may have clinical efficacy against human neoplasms that are associated with gain-of-function KIT mutations