Stratification of responders towards eculizumab using a structural epitope mapping strategy

The complement component 5 (C5)-binding antibody eculizumab is used to treat patients with paroxysmal nocturnal hemoglobinuria (PNH) and atypical haemolytic uremic syndrome (aHUS). As recently reported there is a need for a precise classification of eculizumab responsive patients to allow for a safe and cost-effective treatment. To allow for such stratification, knowledge of the precise binding site of the drug on its target is crucial. Using a structural epitope mapping strategy based on bacterial surface display, flow cytometric sorting and validation via haemolytic activity testing, we identified six residues essential for binding of eculizumab to C5. This epitope co-localizes with the contact area recently identified by crystallography and includes positions in C5 mutated in non-responders. The identified epitope also includes residue W917, which is unique for human C5 and explains the observed lack of cross-reactivity for eculizumab with other primates. We could demonstrate that Ornithodorus moubata complement inhibitor (OmCI), in contrast to eculizumab, maintained anti-haemolytic function for mutations in any of the six epitope residues, thus representing a possible alternative treatment for patients non-responsive to eculizumab. The method for stratification of patients described here allows for precision medicine and should be applicable to several other diseases and therapeutics

Critical role for hyperpolarization-activated cyclic nucleotide-gated channel 2 in the AIF-mediated apoptosis

This study establishes HCN2 channels as physiological relevant ‘calcium gates' that mediate apoptosis-inducing factor-dependent cell death in cancer and primary neuronal cells

Small molecule screening platform for assessment of cardiovascular toxicity on adult zebrafish heart

<p>Abstract</p> <p>Background</p> <p>Cardiovascular toxicity is a major limiting factor in drug development and requires multiple cost-effective models to perform toxicological evaluation. Zebrafish is an excellent model for many developmental, toxicological and regenerative studies. Using approaches like morpholino knockdown and electrocardiogram, researchers have demonstrated physiological and functional similarities between zebrafish heart and human heart. The close resemblance of the genetic cascade governing heart development in zebrafish to that of humans has propelled the zebrafish system as a cost-effective model to conduct various genetic and pharmacological screens on developing embryos and larvae. The current report describes a methodology for rapid isolation of adult zebrafish heart, maintenance <it>ex vivo</it>, and a setup to perform quick small molecule throughput screening, including an in-house implemented analysis script.</p> <p>Results</p> <p>Adult zebrafish were anesthetized and after rapid decapitation the hearts were isolated. The short time required for isolation of hearts allows dissection of multiple fishes, thereby obtaining a large sample size. The simple protocol for <it>ex vivo </it>culture allowed maintaining the beating heart for several days. The in-house developed script and spectral analyses allowed the readouts to be presented either in time domain or in frequency domain. Taken together, the current report offers an efficient platform for performing cardiac drug testing and pharmacological screens.</p> <p>Conclusion</p> <p>The new methodology presents a fast, cost-effective, sensitive and reliable method for performing small molecule screening. The variety of readouts that can be obtained along with the in-house developed analyses script offers a powerful setup for performing cardiac toxicity evaluation by researchers from both academics and industry.</p

A global view of protein expression in human cells, tissues, and organs

Defining the protein profiles of tissues and organs is critical to understanding the unique characteristics of the various cell types in the human body. In this study, we report on an anatomically comprehensive analysis of 4842 protein profiles in 48 human tissues and 45 human cell lines. A detailed analysis of over 2 million manually annotated, high-resolution, immunohistochemistry-based images showed a high fraction (>65%) of expressed proteins in most cells and tissues, with very few proteins (<2%) detected in any single cell type. Similarly, confocal microscopy in three human cell lines detected expression of more than 70% of the analyzed proteins. Despite this ubiquitous expression, hierarchical clustering analysis, based on global protein expression patterns, shows that the analyzed cells can be still subdivided into groups according to the current concepts of histology and cellular differentiation. This study suggests that tissue specificity is achieved by precise regulation of protein levels in space and time, and that different tissues in the body acquire their unique characteristics by controlling not which proteins are expressed but how much of each is produced

CDK‐mediated activation of the SCFFBXO28 ubiquitin ligase promotes MYC‐driven transcription and tumourigenesis and predicts poor survival in breast cancer

SCF (Skp1/Cul1/F‐box) ubiquitin ligases act as master regulators of cellular homeostasis by targeting key proteins for ubiquitylation. Here, we identified a hitherto uncharacterized F‐box protein, FBXO28 that controls MYC‐dependent transcription by non‐proteolytic ubiquitylation. SCFFBXO28 activity and stability are regulated during the cell cycle by CDK1/2‐mediated phosphorylation of FBXO28, which is required for its efficient ubiquitylation of MYC and downsteam enhancement of the MYC pathway. Depletion of FBXO28 or overexpression of an F‐box mutant unable to support MYC ubiquitylation results in an impairment of MYC‐driven transcription, transformation and tumourigenesis. Finally, in human breast cancer, high FBXO28 expression and phosphorylation are strong and independent predictors of poor outcome. In conclusion, our data suggest that SCFFBXO28 plays an important role in transmitting CDK activity to MYC function during the cell cycle, emphasizing the CDK‐FBXO28‐MYC axis as a potential molecular drug target in MYC‐driven cancers, including breast cancer

Automatic Filtering and Substantiation of Drug Safety Signals

Drug safety issues pose serious health threats to the population and constitute a major cause of mortality worldwide. Due to the prominent implications to both public health and the pharmaceutical industry, it is of great importance to unravel the molecular mechanisms by which an adverse drug reaction can be potentially elicited. These mechanisms can be investigated by placing the pharmaco-epidemiologically detected adverse drug reaction in an information-rich context and by exploiting all currently available biomedical knowledge to substantiate it. We present a computational framework for the biological annotation of potential adverse drug reactions. First, the proposed framework investigates previous evidences on the drug-event association in the context of biomedical literature (signal filtering). Then, it seeks to provide a biological explanation (signal substantiation) by exploring mechanistic connections that might explain why a drug produces a specific adverse reaction. The mechanistic connections include the activity of the drug, related compounds and drug metabolites on protein targets, the association of protein targets to clinical events, and the annotation of proteins (both protein targets and proteins associated with clinical events) to biological pathways. Hence, the workflows for signal filtering and substantiation integrate modules for literature and database mining, in silico drug-target profiling, and analyses based on gene-disease networks and biological pathways. Application examples of these workflows carried out on selected cases of drug safety signals are discussed. The methodology and workflows presented offer a novel approach to explore the molecular mechanisms underlying adverse drug reactions

Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network

Using the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of transcription start sites (TSSs) in several species. Strikingly, ~72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probe these unassigned TSSs and show that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we develop Cap Trap RNA-seq, a technology which combines cap trapping and long read MinION sequencing. We train sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveil the importance of STR surrounding sequences not only to distinguish STR classes, but also to predict the level of transcription initiation. Importantly, genetic variants linked to human diseases are preferentially found at STRs with high transcription initiation level, supporting the biological and clinical relevance of transcription initiation at STRs. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism

Resursstöd vid internationalisering : Fallstudie av S.B.E. Svensk Brikettenergi AB

I teorier kring företagens internationaliseringsprocess går det att identifiera ett behov av resursstöd från moderbolaget till dotterbolaget under etableringen. Det är svårt att finna svar på vilka resurser som avses med detta stöd med hjälp av den forskning som finns inom de berörda områdena. Syftet med denna studie är att identifiera vilka resurser som moderbolaget stöder dotterbolaget med vid en utlandsetablering. Detta görs genom att studera företaget S.B.E. Svensk Brikettenergi AB:s etablering i Lettland. Studien begränsar sig till att undersöka resursstöd ur moderbolagets perspektiv. För att genomföra undersökningen har studien använt sig av en kvalitativ undersökningsmetod med resursbaserade teorier som grund. Resursstöd undersöks i de fem resurskategorierna finansiella, fysiska, humana, organisatoriska och teknologiska. Resursstödet har delats upp i företagets fem funktionsområden forskning och utveckling, tillverkning, marknadsföring, finans och ledning. I studien har det identifierats resursstöd inom alla resurskategorier. De finansiella resurserna var svåra att relatera till ett specifikt funktionsområde då överföringarna där var mer generella och gick till hela verksamheten. De fysiska resurserna var den kategori där vi kunde identifiera resursstöd till minst funktionsområden och det som skett var mestadels på grund av det dåliga utbudet på den lettiska marknaden. Inom övriga områden, humana, organisatoriska och teknologiska resurser, kunde vi identifiera ett brett stöd där de olika resurskategorierna ofta var kopplade till varandra

Stöd eller börda? : Fallstudie av Svensk kod för bolagsstyrning i BioGaia

Ett antal bolagsskandaler i början av 2000-talet i exempelvis Enron och Parmalat försämrade allmänhetens förtroende för styrelser och bolagsledningar. I och med oegentligheterna i bland annat Skandia påverkades även de svenska börsbolagen negativt. I ett försök att stärka bolagens förtroende tillsatte regeringen en förtroendekommission med uppgift att se över vilka åtgärder som kunde vara lämpliga. Som ett resultat av detta togs Svensk kod för bolagsstyrning (Koden) fram. Koden utgör självreglering inom det svenska näringslivet och bygger på principen följ eller förklara. Det innebär att man kan anses följa Koden som helhet trots att man avviker från enstaka regler i den, förutsatt att man förklarar varför man avviker från regeln. Inledningsvis har Koden tillämpats av svenska noterade företag med ett börsvärde överstigande tre miljarder kronor. Förslaget är nu att Koden ska tillämpas av samtliga företag noterade på den nordiska börsen samt NGM Equity. Med anledning av detta har vi undersökt vilken effekt Koden skulle få för BioGaia, ett bolag noterat på den nordiska börsen för mindre bolag. Uppsatsen är en fallstudie och syftet är att jämföra dagens rutiner i bolaget med vad Koden skulle ställa för krav. Vi har även försökt få svar på vad bolaget anser om Koden samt undersökt om det är något särskilt avsnitt i Koden som bör revideras inför Kodens implementering för denna grupp företag. Vi har kommit fram till att det i BioGaia finns en medvetenhet om koden och att viss frivillig anpassning redan skett. Trots detta så har vi kommit fram till att Koden skulle få relativt stor effekt på bolaget och att bolaget troligen kommer att välja att avvika från ett antal regler i Koden. Avvikelserna motiveras oftast av bolagets storlek eller av det koncentrerade ägandet. I bolaget finns en viss oro för att Koden kommer att innebära stora kostnader och en massmedial debatt där bolag som avviker mer än ett visst antal gånger klassificeras som oseriösa företag. Vi har även kommit fram till att det i några avseenden vore önskvärt med en viss revidering av Koden inför implementeringen hos ytterligare bolag från den 1 juli 2008