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  • 1.
    Annie, Sernelin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Nora, Palm
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Julia, Axnér
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Sarina, Emad
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Johanna, Söderlund
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Capto Resin Integration Elevates the Phytip Portfolio: An Evaluation of Resins for Antibodies and Adeno-Associated Viruses with Recommendations for the Phytip Portfolio2024Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    The full text will be freely available from 2026-05-31 14:08
  • 2.
    Antonopoulou, Io
    et al.
    Luleå University of Technology, Department of Civil, Environmental and Natural Resources Engineering, Chemical Engineering.
    Sapountzaki, Eleftheria
    Luleå University of Technology, Department of Civil, Environmental and Natural Resources Engineering, Chemical Engineering.
    Rova, Ulrika
    Luleå University of Technology, Department of Civil, Environmental and Natural Resources Engineering, Chemical Engineering.
    Christakopoulos, Paul
    Luleå University of Technology, Department of Civil, Environmental and Natural Resources Engineering, Chemical Engineering.
    The Inhibitory Potential of Ferulic Acid Derivatives against the SARS-CoV-2 Main Protease: Molecular Docking, Molecular Dynamics, and ADMET Evaluation2022In: Biomedicines, E-ISSN 2227-9059, Vol. 10, no 8, article id 1787Article in journal (Refereed)
    Abstract [en]

    The main protease (Mpro) of SARS-CoV-2 is an appealing target for the development of antiviral compounds, due to its critical role in the viral life cycle and its high conservation among different coronaviruses and the continuously emerging mutants of SARS-CoV-2. Ferulic acid (FA) is a phytochemical with several health benefits that is abundant in plant biomass and has been used as a basis for the enzymatic or chemical synthesis of derivatives with improved properties, including antiviral activity against a range of viruses. This study tested 54 reported FA derivatives for their inhibitory potential against Mpro by in silico simulations. Molecular docking was performed using Autodock Vina, resulting in comparable or better binding affinities for 14 compounds compared to the known inhibitors N3 and GC376. ADMET analysis showed limited bioavailability but significantly improved the solubility for the enzymatically synthesized hits while better bioavailability and druglikeness properties but higher toxicity were observed for the chemically synthesized ones. MD simulations confirmed the stability of the complexes of the most promising compounds with Mpro, highlighting FA rutinoside and compound e27 as the best candidates from each derivative category.

  • 3.
    Aranzana-Climent, Vincent
    et al.
    Uppsala University, Sweden.
    Hughes, Diarmaid
    Uppsala University, Sweden.
    Cao, Sha
    Uppsala University, Sweden.
    Tomczak, Magdalena
    National Medicines Institute, Poland.
    Urbas, Malgorzata
    National Medicines Institute, Poland.
    Zabicka, Dorota
    Uppsala University, Sweden.
    Vingsbo Lundberg, Carina
    Statens Serum Institut, Denmark.
    Hansen, Jon
    Statens Serum Institut, Denmark.
    Lindberg, Johan
    RISE Research Institutes of Sweden, Bioeconomy and Health, Chemical and Pharmaceutical Toxicology.
    Hobbie, Sven N
    University of Zurich, Switzerland.
    Friberg, Lena E
    Uppsala University, Sweden.
    Translational in vitro and in vivo PKPD modelling for apramycin against Gram-negative lung pathogens to facilitate prediction of human efficacious dose in pneumonia2022In: Clinical Microbiology and Infection, ISSN 1198-743X, E-ISSN 1469-0691, Vol. 28, no 10, p. 1367-1374Article in journal (Refereed)
    Abstract [en]

    Objectives: New drugs and methods to efficiently fight carbapenem-resistant gram-negative pathogens are sorely needed. In this study, we characterized the preclinical pharmacokinetics (PK) and pharmacodynamics of the clinical stage drug candidate apramycin in time kill and mouse lung infection models. Based on in vitro and in vivo data, we developed a mathematical model to predict human efficacy. Methods: Three pneumonia-inducing gram-negative species Acinetobacter baumannii, Pseudomonas aeruginosa, and Klebsiella pneumoniae were studied. Bactericidal kinetics were evaluated with time-kill curves; in vivo PK were studied in healthy and infected mice, with sampling in plasma and epithelial lining fluid after subcutaneous administration; in vivo efficacy was measured in a neutropenic mouse pneumonia model. A pharmacokinetic-pharmacodynamic model, integrating all the data, was developed and simulations were performed. Results: Good lung penetration of apramycin in epithelial lining fluid (ELF) was shown (area under the curve (AUC)ELF/AUCplasma = 88%). Plasma clearance was 48% lower in lung infected mice compared to healthy mice. For two out of five strains studied, a delay in growth (∼5 h) was observed in vivo but not in vitro. The mathematical model enabled integration of lung PK to drive mouse PK and pharmacodynamics. Simulations predicted that 30 mg/kg of apramycin once daily would result in bacteriostasis in patients. Discussion: Apramycin is a candidate for treatment of carbapenem-resistant gram-negative pneumonia as demonstrated in an integrated modeling framework for three bacterial species. We show that mathematical modelling is a useful tool for simultaneous inclusion of multiple data sources, notably plasma and lung in vivo PK and simulation of expected scenarios in a clinical setting, notably lung infections. © 2022 The Author(s)

  • 4.
    Arkstål, Emil
    Linköping University, Department of Computer and Information Science, Human-Centered systems.
    Interactive Analytics and Visualization for Data Driven Calculation of Individualized COPD Risk2018Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Chronic obstructive pulmonary disease (COPD) is a high mortality disease, second to stroke and ischemic heart disease. This non-curable disease progressively exacerbates, leading to high personal and societal economic impact, reduced quality of life and often death. General treatment plans for COPD risk mistreating the individuals’ condition. To be effective, the treatment should be individualized following the practices of precision medicine. The aim of this thesis was to develop a data driven algorithm and system with visualization to assess individual COPD risk. With MRI body composition profile measurements, it is possible to accurately assess propensity of a multitude of metabolic conditions, such as coronary heart disease and type 2 diabetes.  The algorithm and system has been developed using Wolfram Language and R within the Wolfram Mathematica framework. The algorithm calculates individualized virtual control groups metabolically similar to the patient’s body composition and spirometric profile. Using UK Biobank data, our tool was used to assess patient COPD propensity using an individual-specific virtual control group with AUROC 0.778 (female) and 0.758 (men). Additionally, the tool was used to identify new body composition profiles related to COPD and associated comorbid conditions.

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    Interactive Visualization COPD Emil Arkstål
  • 5.
    Axelson, Linnéa
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Berg, Loova
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Blomkvist, Anna
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    El-Zein, Nora
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Grenholm, Elin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Hemmingsson, Nora
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Mosebach, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Exploring the Potential of Camelid Single-Domain Antibodies: Structure, Properties and Diverse Applications in Therapeutics and Diagnostics2023Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Download full text (pdf)
    fulltext
  • 6.
    Benevides, Kristina
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Broström, Oscar
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Elison Kalman, Grim
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Swenson, Hugo
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Vlassov, Andrei
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Ågren, Josefin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Stabil och antibiotikafri läkemedelsproduktion i rekombinant Escherichia coli2017Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [sv]

    Den här rapporten presenterar ett antibiotikafritt, stabilt och kromosombaserat expressionssystem för läkemedelsproduktion i Escherichia coli på beställning av företaget Affibody AB. E. coli-stammen BL21(DE3) valdes som värdorganism för expressionssystemet. Systemet består av en genkassett som innehåller en T7-promotor, en 5′-UTR från genen ompA och en terminatorsekvens från RNA-operonet rrnB. Fyra kopior av genkassetten ska integreras i pseudogenerna caiB, yjjM, hsdS och yjiV. En datormodell som modellerar det egentliga kopietalet i cellerna har skapats i mjukvaran MATLAB, vilket visar att det uppskattas vara maximalt 32 kopior av genkassetten per cell på grund av replikation av kromosomen. Ett högt pH i fermentorn; att använda fed-batch och blandade kolhydratkällor; och att använda stammen BL21(DE3) minskar acetatproduktionen i cellen. En lägre acetatproduktion kan leda till en högre produkthalt. En proteinutbytesmodell för mjukvaran MATLAB har konstruerats för att uppskatta koncentrationen av Affibody®-molekylen i en E. coli cell.

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    fulltext
  • 7. Chaudhary, Himanshu
    et al.
    Meister, Sebastian
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Zetterberg, Henrik
    Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Mölndal, Sweden..
    Löfblom, John
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Engineering.
    Lendel, Christofer
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Chemistry, Applied Physical Chemistry.
    Dissecting the structural organization of multiprotein amyloid aggregates using a bottom-up approach2020In: ACS Chemical Neuroscience, E-ISSN 1948-7193, Vol. 11, no 10, p. 1447-1457Article in journal (Refereed)
    Abstract [en]

    Deposition of fibrillar amyloid β (Aβ) in senile plaques is a pathological signature of Alzheimer's disease. However, senile plaques also contain many other components, including a range of different proteins. Although the composition of the plaques can be analyzed in post mortem tissue, knowledge of the molecular details of these multiprotein inclusions and their assembly processes is limited, which impedes the progress in deciphering the biochemical mechanisms associated with Aβ pathology. We here describe a bottom-up approach to monitor how proteins from human cerebrospinal fluid associate with Aβ amyloid fibrils to form plaque particles. The method combines flow cytometry and mass spectrometry proteomics and allowed us to identify and quantify 128 components of the captured multiprotein aggregates. The results provide insights in the functional characteristics of the sequestered proteins and reveal distinct interactome responses for the two investigated Aβ variants, Aβ(1-40) and Aβ(1-42). Furthermore, the quantitative data is used to build models of the structural organization of the multiprotein aggregates, which suggests that Aβ is not the primary binding target for all the proteins; secondary interactions account for the majority of the assembled components. The study elucidates how different proteins are recruited into senile plaques and establishes a new model system for exploring the pathological mechanisms of Alzheimer's disease from a molecular perspective.

  • 8.
    Chotteau, Veronique
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Jiang, Yun
    Biovitrum/SOBI, Sweden.
    westin, Jeanette
    Biovitrum/SOBI, Sweden.
    Dahlenborg, K
    Biovitrum/SOBI, Sweden.
    Sjöblom-Hallén, A
    Biovitrum/SOBI, Sweden.
    Svensson, Erik
    Biovitrum/SOBI, Sweden.
    Öberg, Mikael
    Biovitrum/SOBI, Sweden.
    Development of a fed-batch process for the production of a recombinant protein X in CHO-GS system: Case study from the cell to reactor process ready for pilot scale cultivation2010In: Cells and Culture: Proceedings of the 20th ESACT / [ed] Noll T, Springer Science+Business Media B.V., 2010, p. 723-725Conference paper (Other academic)
    Abstract [en]

    A new cell line was created using CHO-GS system. The most promising clones were adapted to different base cultivation media leading to the selection of one medium. The fed-batch process development was performed in spinner, shake flask and bioreactor scale. It included the selection of a feed medium, the choice of the feed strategy and the optimisation of the glucose feeding. The process was then simplified by using a single feed including the feed medium and the glucose feed. Finally up-scaling parameters like aeration and CO2 stripping were studied in 3 L and 15 L bioreactors in preparation for pilot scale operation. This process proved to be robust, reproducible and suitable for large and commercial scale operation.

  • 9.
    Chotteau, Veronique
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Lindqvist, Anna
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Study of the effect of high pH and alkali addition in a cultivation of Chinese Hamster Ovary cell2012In: Proceedings of the 21st Annual Meeting of the European Society for Animal Cell Technology (ESACT) / [ed] Jenkins, Nigel; Barron, Niall; Alves, Paula, Springer Science+Business Media B.V., 2012, p. 323-326Conference paper (Refereed)
    Abstract [en]

    This work aimed at studying the impact of alkali addition in a Chinese Hamster Ovary cell culture. Two phenomena were studied, the kinetic rate of direct cell death in presence of high pH and the effect of transitory single contact of high pH on cell viability and growth. Contact with pH 11 or 10 did not provoke immediate cell lysis. The cells survived several minutes to such conditions. Contact with pH 11 during 2 minutes, with pH 10 during 5 minutes, with pH 9 during 5 minutes or 10 minutes did not affect the viability. In these conditions, the growth was not affected except after 5 minutes contact at pH 10 or 10 minutes contact at pH 9 for which the growth was slowed down the first day only. As expected, NaOH addition affected the cells more than Na2CO3 addition. This was due to a higher pH but could be even observed at the same pH (10).

  • 10.
    Chotteau, Veronique
    et al.
    KTH, School of Biotechnology (BIO).
    Wåhlgren, Caroline
    Biovitrum/SOBI, Sweden.
    Jiang, Yun
    Biovitrum/SOBI, Sweden.
    Svensson, Erik
    Biovitrum/SOBI, Sweden.
    Process for cultivating animal cells comprising the feeding of plant-derived peptones2005Patent (Other (popular science, discussion, etc.))
    Abstract [en]

    A process for cultivating animal cells producing complex proteins, wherein one plant-derived peptone or a combination of plant-derived peptones is fed to the cell culture, as well as a method for reducing the toxic effect of over-feeding amino acids during a fed-batch process for cultivating animal cells producing complex proteins.

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    attachment
  • 11.
    Chotteau, Veronique
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Wåhlgren, Caroline
    Biovitrum/SOBI, Sweden.
    Pettersson, Helena
    Biovitrum/SOBI, Sweden.
    Effect of Peptones and Study of Feeding Strategies in a CHO Based Fed-batch Process for the Production of a Human Monoclonal Antibody2007In: Cell Technology for Cell Products: Proceedings of the 19th ESACT Meeting, Harrogate, UK, June 5-8, 2005 / [ed] Smith R, Dordrecht, The Netherlands: Springer Netherlands, 2007, p. 371-374Conference paper (Other academic)
    Abstract [en]

    Eight commercial peptones, derived from plants, were studied for their ability of improving the cell growth and the productivity of a CHO cell line producing a human monoclonal antibody. They were also compared to yeast, lactalbumin and meat derived peptones. Seven plant peptones were selected and further studied in combination by Design of Experiment. The best three peptones were then tested in combinations in fed-batch cultivation. The fed-batch process was based on low concentrations of glucose and glutamine with feeding of amino acids, peptones and feed medium including vitamins, metal traces and biosynthesis precursors. This process was based on Biovitrum protein-free proprietary medium for the base medium and the feeding medium. Different feeding strategies, different peptone combinations and phosphate feeding were studied for their ability to improve the cell density, the cell specific productivity and the cultivation longevity

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    Chotteau 2007 Effect of Peptones and Study of Feeding Strategies in a CHO Based Fed-batch Process for the Production of a Human Monoclonal Antibody
  • 12.
    Clincke, Marie-Francoise
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Mölleryd, Carin
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Zhang, Ye
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Lindskog, Eva
    GE Healthcare, Uppsala, Sweden.
    Walsh, Kieron
    GE Healthcare, Westborough, MA, USA.
    Chotteau, Veronique
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Study of a recombinant CHO cell line producing a monoclonal antibody by ATF or TFF external filter perfusion in a WAVE Bioreactor™2011In: BMC Proceedings, 2011, Volume 5, Supplement 8, P105, BioMed Central, 2011, p. 105-Conference paper (Refereed)
    Abstract [en]

    Major advantages of perfusion are high cell numbers and high total production in a relatively small size bioreactor. Moreover, perfusion is optimal when the product of interest is unstable or if the product yield is low. On the other hand, disadvantages are for example technical challenges originating from non-robust cell separation devices as well as sterility concerns from the more complex set-up needed.

    In the present work, the use of a WAVE Bioreactor™ system 20/50 in perfusion mode with10 L disposable Cellbag™ bioreactors customized with two dip tubes in combination with disposable hollow fiber filters as external cell separating devices were investigated. A comparison between Alternating Tangential Flow (ATF) and Tangential Flow Filtration (TFF) was performed using a recombinant CHO cell line producing a monoclonal antibody (mAb) as a model system. 

  • 13.
    Dolfe, Lisa
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Process development for the control of solubility of Affibody® molecules2011Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    In this study the aim was to optimize the production of the Affibody fusion-protein Z03358-

    ABD094-(S4G)3-IL2 with regard to the amount of soluble protein produced. However,

    problems with reproducibility with this protein and the chosen expression system were

    encountered. Therefore, expression of the His-tagged Affibody His6-(Z05477)2 was

    evaluated using the same expression system as well as expression in another well

    characterized expression system.

    Both target proteins are of therapeutic interest. One of the proteins is an IL2 fusion

    protein (Z03358-ABD094-(S4G)3-IL2) that bind the platelet-derived growth factor receptor β

    (PDGFR-β). PDGF signaling is of interest in cancer treatment where, among other things, the

    effects of PDGF on tumor angiogenesis is researched. The His6-(Z05477)2 protein has a

    classified target but is developed as a therapeutic in the area of inflammation and autoimmune

    disease. Both model proteins are known to be difficult to purify due to low solubility.

    The two E. coli expression systems investigated and compared were BL21(DE3) and

    Lemo21(DE3). The fusion protein Z03358-ABD094-(S4G)3-IL2 was produced in

    BL21(DE3) in inclusion bodies with a yield of 4.95 g/l. An optimized process for the

    expression of His6-(Z05477)2 using BL21(DE3) was developed with a yield of 6.6 g/l soluble

    protein after expression at 30°C for 6 h.

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    lisadolfe_examensarbete
  • 14.
    Einarsson, Ellen
    Linköping University, Department of Physics, Chemistry and Biology, Biotechnology.
    Evaluation of 5´- and 3´-UTR Translation Enhancing Sequences to Improve Translation of Proteins in CHO Cells2018Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    The purpose of this project was to identify and evaluate nucleotide sequences enhancing translation of proteins in Chinese hamster ovary (CHO) cells. Candidate sequences were placed in the 5´-untranslated region (UTR) or 3´ UTR respectively and evaluated in a CHO-based expression system with a fluorescent Fc-fusion protein as a model protein.Five plasmid vectors were constructed, two of which designed to have a randomized nucleotide library in their 5´ and 3´ UTR respectively, and three of which designed to hold varying repeats of a known enhancing translation (ET) sequence in their 5´ or 3´ UTR. The plasmid constructs were transfected into CHO cells and the protein expression was analyzed both by fluorescence intensity in single cells using flow cytometry and in bulk by monoclonal antibody titer analysis based on Protein A affinity.The main result is that both flow cytometry and titer analysis indicate that insertion of five repeats of the ET in the 5´UTR has a negative effect on protein expression as compared to the control which had no ET repeats. Results related to the insertion of three ETs in the 5´ UTR were ambiguous. The titer analysis indicated that it had a negative effect on the protein expression compared to the control which had no ET repeats, whereas the flow cytometry results suggest that the effect is negligible. Transfection of library plasmids was unsuccessful; hence no library expression analysis results were achieved. Due to the time constraints of the project, the reason for the unsuccessful transfection of library plasmids was not investigated, but the LTX transfection method is stated as a highly plausible cause.Based on the outcome of this study, two recommendations for future work are suggested. The first one is to continue the focus on UTR sequences in terms of library screening, and to improve the method of transfecting library plasmid constructs into CHO cells using lipofection. The second suggestion for further studies is to test different UTR sequence lengths without involving potential ETs, to rule out the effect and positions of the ETs and investigate the expressional effect of UTR length solely.

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    Master thesis Ellen Einarsson
  • 15.
    Eriksson, Sara
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Quality by Design (QbD) in Biochemical Applications: Possibilities and Limitations2013Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
  • 16.
    Garousi, Javad
    et al.
    Uppsala Univ, Dept Immunol Genet & Pathol, SE-75185 Uppsala, Sweden..
    Lindbo, Sarah
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Borin, Jesper
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    von Witting, Emma
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Vorobyeva, Anzhelika
    Uppsala Univ, Dept Immunol Genet & Pathol, SE-75185 Uppsala, Sweden..
    Oroujeni, Maryam
    Uppsala Univ, Dept Immunol Genet & Pathol, SE-75185 Uppsala, Sweden..
    Mitran, Bogdan
    Uppsala Univ, Dept Med Chem, Uppsala, Sweden..
    Orlova, Anna
    Uppsala Univ, Dept Med Chem, Uppsala, Sweden..
    Buijs, Jos
    Uppsala Univ, Dept Immunol Genet & Pathol, SE-75185 Uppsala, Sweden..
    Tolmachev, Vladimir
    Uppsala Univ, Dept Immunol Genet & Pathol, SE-75185 Uppsala, Sweden..
    Hober, Sophia
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Comparative evaluation of dimeric and monomeric forms of ADAPT scaffold protein for targeting of HER2-expressing tumours2019In: European journal of pharmaceutics and biopharmaceutics, ISSN 0939-6411, E-ISSN 1873-3441, Vol. 134, p. 37-48Article in journal (Refereed)
    Abstract [en]

    ADAPTs are small engineered non-immunoglobulin scaffold proteins, which have demonstrated very promising features as vectors for radionuclide tumour targeting. Radionuclide imaging of human epidermal growth factor 2 (HER2) expression in vivo might be used for stratification of patients for HER2-targeting therapies. ADAPT6, which specifically binds to HER2, has earlier been shown to have very promising features for in vivo targeting of HER2 expressing tumours. In this study we tested the hypothesis that dimerization of ADAPT6 would increase the apparent affinity to HER2 and accordingly improve tumour targeting. To find an optimal molecular design of dimers, a series of ADAPT dimers with different linkers, -SSSG- (DiADAPT6L1), -(SSSG)(2)- (DiADAPT6L2), and -(SSSG)(3)- (DiADAPT6L3) was evaluated. Dimers in combination with optimal linker lengths demonstrated increased apparent affinity to HER2. The best variants, DiADAPT6L2 and DiADAPT6L3 were site-specifically labelled with In-111 and I-125, and compared with a monomeric ADAPT6 in mice bearing HER2-expressing tumours. Despite higher affinity, both dimers had lower tumour uptake and lower tumour-to-organ ratios compared to the monomer. We conclude that improved affinity of a dimeric form of ADAPT does not compensate the disadvantage of increased size. Therefore, increase of affinity should be obtained by affinity maturation and not by dimerization.

  • 17.
    Haughey, Caitlin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Mesilaakso, Lauri
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Berner-Wik, Erik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Östlund, Emma
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Ulfsparre, Jonatan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Olin, Hampus
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Probes for ESBL: A Method for Production of Probe Targets in Antibiotic Resistant Genes2017Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    This project aimed to find a method for producing potential probe targets for identification of ESBL (Extended Spectrum Beta Lactamase) genes in bacteria. ESBLs are a type of enzymes responsible for antibiotic resistance in many bacteria. The result we developed was a semi-automated pipeline that utilises several Perl scripts to download gene sequences, identify sequence subgroups based on sequence similarity, find common target sequences among them and screen the target sequences against a background database. These target sequences should work with padlock probes and therefore had specific requirements regarding length and highest number of allowed mismatches. This report includes descriptions of the scripts and ideas for future improvements, as well as an ethical analysis about aspects relevant to research on antibiotic resistance.

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    fulltext
  • 18.
    Hendrikse, Natalie M.
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Fibre- and Polymer Technology, Coating Technology.
    Sandegren, Anna
    Andersson, Tommy
    Blomqvist, Jenny
    Makower, Åsa
    Possner, Dominik
    Su, Chao
    Thalén, Niklas
    Tjernberg, Agneta
    Westermark, Ulrica
    Svensson Gelius, Stefan
    Syrén, Per-Olof
    Nordling, Erik
    Ancestral lysosomal enzymes with increased activity harbor therapeutic potential for treatment of Hunter syndromeManuscript (preprint) (Other academic)
  • 19.
    Holm, Mikaela
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering.
    An exploratory study of the durability of biopharmaceuticals in AstraZeneca's Quality Control Operations2024Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    This master's thesis has been performed at the Quality Control department (QC) at Sweden Biomanufacturing Center (SBC), AstraZeneca. The main goal of the master's thesis was to examine if a monoclonal antibody-based (mAb) drug product (DP) at the QC laboratory is still representative of the batch after storage up to one month after opening, since today's practice states a longevity of 24h after opening. The purpose also includes investigating how the antibody is affected by different storage conditions. This could possibly lead to an extension of the durability, which would enable optimizing the usage of the biopharmaceuticals in QC operations, reducing waste and increasing the availability of DP. 

    The stability of biopharmaceuticals is monitored to ensure safe and highly effective drugs to patients. To investigate how the stability is affected by opening of the samples and different storage's, the stability of the biopharmaceutical has been evaluated by three methods: capillary electrophoresis (CE), high-performance size exclusion chromatography (HPSEC) and capillary isoelectric focusing (ciEF). These methods were performed each week during a month to monitor the progression of impurities e.g. aggregates and fragments in the DP. Results showed a slight decrease in purity for all samples. Samples in non-recommended storage condition showed a more progressed degradation than samples in recommended storage conditions. However, all samples stored in recommended storage condition passed the specifications and control limits set by AstraZeneca at all time points. Therefore, AstraZeneca is recommended to extend the durability of the DP, to two weeks after opening.

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  • 20.
    Hulme, Heather E.
    et al.
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Meikle, Lynsey M.
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Wessel, Hannah
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Strittmatter, Nicole
    AstraZeneca, Milton Sci Pk, Cambridge CB4 0WG, England..
    Swales, John
    AstraZeneca, Milton Sci Pk, Cambridge CB4 0WG, England..
    Thomson, Carolyn
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Nilsson, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Nibbs, Robert J. B.
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Milling, Simon
    Andrén, Per E.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Mackay, C. Logan
    Univ Edinburgh, Sch Chem, Edinburgh EH9 3FJ, Midlothian, Scotland..
    Dexter, Alex
    Natl Phys Lab, Teddington TW11 0LW, Middx, England..
    Bunch, Josephine
    Natl Phys Lab, Teddington TW11 0LW, Middx, England..
    Goodwin, Richard J. A.
    AstraZeneca, Milton Sci Pk, Cambridge CB4 0WG, England..
    Burchmore, Richard
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Wall, Daniel M.
    Univ Glasgow, Inst Infect Immun & Inflammat, Coll Med Vet & Life Sci, Glasgow G12 8QQ, Lanark, Scotland..
    Mass spectrometry imaging identifies palmitoylcarnitine as an immunological mediator during Salmonella Typhimurium infection2017In: Scientific Reports, E-ISSN 2045-2322, Vol. 7, article id 2786Article in journal (Refereed)
    Abstract [en]

    Salmonella Typhimurium causes a self-limiting gastroenteritis that may lead to systemic disease. Bacteria invade the small intestine, crossing the intestinal epithelium from where they are transported to the mesenteric lymph nodes (MLNs) within migrating immune cells. MLNs are an important site at which the innate and adaptive immune responses converge but their architecture and function is severely disrupted during S. Typhimurium infection. To further understand host-pathogen interactions at this site, we used mass spectrometry imaging (MSI) to analyse MLN tissue from a murine model of S. Typhimurium infection. A molecule, identified as palmitoylcarnitine (PalC), was of particular interest due to its high abundance at loci of S. Typhimurium infection and MLN disruption. High levels of PalC localised to sites within the MLNs where B and T cells were absent and where the perimeter of CD169(+) sub capsular sinus macrophages was disrupted. MLN cells cultured ex vivo and treated with PalC had reduced CD4(+) CD25(+) T cells and an increased number of B220(+) CD19(+) B cells. The reduction in CD4(+) CD25(+) T cells was likely due to apoptosis driven by increased caspase-3/7 activity. These data indicate that PalC significantly alters the host response in the MLNs, acting as a decisive factor in infection outcome.

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  • 21.
    Jiang, Yun
    et al.
    Biovitrum/SOBI, Sweden.
    Svensson, Erik
    Biovitrum/SOBI, Sweden.
    Chotteau, Veronique
    Improvement of a CHO Fed-Batch Process by Fortifying with Plant Peptones2010In: Cells and Culture: ESACT Proceedings, 2010, Volume 4, Part 3 / [ed] Noll T, Springer, 2010, p. 281-284Conference paper (Other academic)
    Abstract [en]

    A serum-free fed-batch process was developed for production of a human monoclonal antibody in Chinese hamster ovary (CHO) cells based on Biovitrum’s proprietary low protein serum-free medium without animal derived components (BVT4). The cells were fed with glucose, glutamine and Biovitrum’s proprietary low protein serum-free feed medium without animal derived components enriched with amino acids, vitamins, metal traces, peptones, and biosynthesis precursors. To improve the performance of the fed-batch process, we developed the use of plant peptones by studying the dose and timing of the peptone feeding. Different doses of peptone cocktail and amino acid cocktail, as well as different combinations of pep- tone and amino acid cocktails were first screened in 50 ml filter tubes on an AgCell shaker table. The best combinations were then assessed in spinner and 3 L bioreactor cultures. To reinforce our findings, the antibody-producing CHO cells were adapted to a disclosed serum-free medium DMEM/F12 and the beneficial effects of pep- tones were confirmed in a fed-batch process based on the DMEM/F12 serum-free medium.

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    Jiang 2010 Improvement of a CHO Fed-Batch Process by Fortifying with Plant Peptones
  • 22.
    Kandušer, Maša
    et al.
    University of Ljubljana, Slovenia.
    Ušaj, Marko
    University of Ljubljana, Slovenia.
    Cell electrofusion: past and future perspectives for antibody production and cancer cell vaccines2014In: Expert Opinion on Drug Delivery, ISSN 1742-5247, E-ISSN 1744-7593, Vol. 11, no 12, p. 1885-1898Article in journal (Refereed)
    Abstract [en]

    Introduction: In the past few decades, new methods for drug and gene delivery have been developed, among which electroporation and electrofusion have gained noticeable attention. Lately, advances in the field of immunotherapy have enabled new cancer therapies based on immune response, including monoclonal antibodies and cell vaccines. Efficient cell fusion is needed for both hybridoma production and cell vaccine preparation, and electrofusion is a promising method to achieve this goal.Areas covered: In the present review, we cover new strategies of cancer treatment related to antibody production and cell vaccines. In more detail, cell electroporation and electrofusion are addressed. We briefly describe principles of cell electroporation and focus on electrofusion and its influential factors, with special attention on the fusogenic state of the cell membrane, contact formation, the effect of electrofusion media and cell viability. We end the review with an overview of the very promising field of microfluidic devices for electrofusion.Expert opinion: In our opinion, electrofusion can be a very efficient method for hybridoma and cell vaccine production. Advances in the development of microfluidic devices and a better understanding of the underlying (biological) mechanisms will overcome the current limitations.

  • 23.
    Kanje, Sara
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Venskutonytė, Raminta
    Lund University.
    Scheffel, Julia
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Nilvebrant, Johan
    KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Engineering.
    Lindkvist-Petersson, Karin
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Centres, Centre for Bioprocess Technology, CBioPT. KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Protein engineering allows for mild affinity-based elution of therapeutic antibodies2018In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 430, no 18, p. 3427-3438Article in journal (Refereed)
    Abstract [en]

    Presented here is an engineered protein domain, based on Protein A, that displays a calcium-dependent binding to antibodies. This protein, ZCa, is shown to efficiently function as an affinity ligand for mild purification of antibodies through elution with ethylenediaminetetraacetic acid. Antibodies are commonly used tools in the area of biological sciences and as therapeutics, and the most commonly used approach for antibody purification is based on Protein A using acidic elution. Although this affinity-based method is robust and efficient, the requirement for low pH elution can be detrimental to the protein being purified. By introducing a calcium-binding loop in the Protein A-derived Z domain, it has been re-engineered to provide efficient antibody purification under mild conditions. Through comprehensive analyses of the domain as well as the ZCa–Fc complex, the features of this domain are well understood. This novel protein domain provides a very valuable tool for effective and gentle antibody and Fc-fusion protein purification

  • 24.
    Kennedy, Gordon T.
    et al.
    Beckman Laser Institute and Medical Clinic, USA.
    Lentsch, Griffin R.
    Beckman Laser Institute and Medical Clinic, USA.
    Trieu, Brandon
    Beckman Laser Institute and Medical Clinic, USA.
    Ponticorvo, Adrien
    Beckman Laser Institute and Medical Clinic, USA.
    Saager, Rolf B.
    Beckman Laser Institute and Medical Clinic, USA.
    Durkin, Anthony J.
    Beckman Laser Institute and Medical Clinic, USA.
    Design and fabrication of solid phantoms for NIR water fraction studies2017In: Proceedings Volume 10056, Design and Quality for Biomedical Technologies X; 100560A (2017) / [ed] Ramesh Raghavachari, Rongguang Liang, SPIE - International Society for Optical Engineering, 2017, Vol. 10056, article id 100560AConference paper (Refereed)
    Abstract [en]

    Tissue simulating phantoms provide a valuable platform for quantitative evaluation of the performance of diffuse optical devices. In this paper we report the development of a poly(dimethylsiloxane) (PDMS) tissue phantom that mimics the spectral characteristics of tissue water. We have developed these phantoms to mimic different water fractions in tissue for testing new devices within the context of clinical applications such as burn wound triage. Compared to liquid phantoms, PDMS phantoms are easier to transport and use, and have a longer usable life than gelatin based phantoms. The pthalocyanine dye 9606 was used to provide an absorption feature of in the vicinity of 970 nm. Scattering properties were independently determined by adding titanium dioxide powder to obtain reduced scattering coefficients similar to that of tissue in the near infrared. Phantom properties were characterized using the techniques of inverse adding doubling and spatial frequency domain imaging. Results presented here demonstrate that we can fabricate solid phantoms that can be used to simulate different water fractions.

  • 25.
    Khan, Anisha
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Molecular Biomimetics.
    Optimisation of Culture Conditions and Pilot Scale Production of a LALA-IAHA Fc-Mutated IgG1 Antibody2021Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Anaplastic thyroid cancer patients have a short life expectancy even with available treatments including radiation and invasive surgery. Over the past decades monoclonal antibodies have been growing in popularity on the pharmaceutical market. This study involves an IgG1 monoclonal antibody with LALA-IAHA double mutations developed for radioimmunotherapy of anaplastic thyroid cancer to prolong the life expectancy of these patients. Producing large amounts of pharmaceuticals is important in drug development for both preclinical and clinical studies. Pilot scale productionis important for generating material for preclinical studies and for acquiring process specific data to facilitate tech transfer to contract manufacturing organisations.

    In this report feeding regimen and temperature were optimised for high antibody titre of a stable CHO cell line in shake flasks in fed-batch mode. The optimised culture conditions were daily feed starting culture day three of 3 % Cell Boost 7a and 0.3 % Cell Boost 7b of start culture volume and glucose maintained at 4 g/L (if < 2 g/L in the culture) with a temperature shift from 37 °C to 32 °C. The fed-batch process established in shake flask culturewas further tested and proven to work in a bioreactor which shows process scalability. However, the antibody titre was not as high as expected. There are still parameters to improve such as agitation speed and feeding of the bioreactor in order to decrease toxic lactate and ammonia levels. The collected data is still useful for future process development and production of the antibody with this cell line.

    The full text will be freely available from 2024-10-05 12:20
  • 26.
    Kronander, Björn
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Quantification of alpha-synuclein in cerebrospinal fluid2012Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    To date there is no accepted clinical diagnostic test for Parkinson's disease (PD) based on biochemical analyses of blood or cerebrospinal uid. Currently, diagnosis, measurement of disease progression and response to therapeutic intervention are based on clinical observation, but the rst neuronal dysfunction precede the earliest recognition of symptom by at least 5 - 10 years. A potential diagnostic biomarker is oligomeric alpha-synuclein which in recent papers have reported a signicant quantitative dierence between PD and controls. In this master thesis, a method for measuring oligomeric levels of alpha-synuclein is presented together with a monomeric measuring commercial kit used to measure alpha-synuclein in a preclinical model of PD. A signicant dierence of monomeric levels could be detected between two weeks and four weeks post injection of a vector containing the gene for human alpha-synuclein, no signicant dierence between four and eight weeks was found.

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    Quantification of alpha-synuclein in cererospinal fluid
  • 27.
    Källgren, Joanna
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Strukturella och funktionella studier av fyra enzymer involverade i cellväggsbiosyntes hos Mycobacterium tuberculosis2015Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    The pathogenic bacterium Mycobacterium tuberculosis (Mt) is the causative agent of tuberculosis, a widespread and fatal infectious disease. Today, treatment against tuberculosis involves a combination of drugs, which need to be taken for at least six months and which often causes severe side effects. Therefore, new drugs that are more effective and that give fewer side effects are needed. A characteristic feature of the Mt bacterium is its very complex and thick cell wall, which prevents many potential drug molecules from penetrating it. Inhibiting any one of the enzymes that are involved in its biosynthesis would therefore seem to be a good strategy for eliminating the Mt bacteria. The aim of this study was to characterize four enzymes involved in Mt cell wall biosynthesis. In order to do that, they were produced recombinantly in E. coli and purified. Crystallization experiments were set up in order to produce diffracting crystals, with the aim of structure determination and drug design.

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  • 28.
    Langer, Krzysztof
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology.
    Jernström, Sandra
    Mikkonen, Piia
    University of Helsinki, Helsinki, Finland.
    Östling, Päiivi
    Seashore-Ludlow, Brinton A
    Jönsson, Håkan
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Engineering.
    A conversational robotic lab assistant for automated microfluidic 3d microtissue production2019In: 23rd International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2019, Chemical and Biological Microsystems Society , 2019, p. 888-889Conference paper (Refereed)
    Abstract [en]

    The future of life science is linked to automation and microfluidics. Here we present a robotic lab assistant, a general automation platform for droplet microfluidics including a conversational mobile interface. We demonstrate the automated production of human cancer microtissues in droplets at a throughput of 85000 spheroids per microfluidic circuit per hour. The capability of automated spheroid generation is directly applicable to precision medicine and drug screening. Multiple cell lines were successfully tested, including cancer cell lines, co-cultures, and primary cells. The 3D-microtissues/spheroids were automatically assembled-incubated-retrieved with high viability for further drug screening analysis - the platform interfaces with standard labware.

  • 29.
    Langer, Krzysztof
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Jönsson, Håkan
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Rapid production and recovery of cell spheroids by automated droplet microfluidics2019Manuscript (preprint) (Other academic)
    Abstract [en]

    Droplet microfluidics enables high throughput cell processing, analysis and screening by miniaturizing the reaction vessels to nano- or pico-liter water-in oil droplets, but like many other microfluidic formats, droplet microfluidics have not been interfaced with or automated by laboratory robotics. Here we demonstrate automation of droplet microfluidics based on an inexpensive liquid handling robot for the automated production of human scaffold-free cell spheroids, using pipette actuation and interfacing the pipetting tip with a droplet generating microfluidic chip. In this chip we produce highly mono-disperse 290μm droplets with diameter CV of 1.7%. By encapsulating cells in these droplets, we produce cell spheroids in droplets and recover them to standard formats at a throughput of 85000 spheroids per microfluidic circuit per hour. The viability of the cells in spheroids remains high after recovery only decreased by 4% starting from 96% after 16 hours incubation in nanoliter droplets. Scaffold-free cell spheroids and 3D tissue constructs recapitulate many aspects of functional human tissue more accurately than 2D or single cell cultures, but assembly methods for spheroids, e.g. hanging drop micro-plates, has had limited throughput. The increased throughput and decreased cost of our method enables spheroid production at the scale needed for lead discovery drug screening and approaches the cost where these micro tissues could be used as building blocks for organ scale regenerative medicine.

  • 30.
    Langer, Krzysztof
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology.
    Jönsson, Håkan
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology. KTH, Novo Nordisk Fdn Ctr Biosustainabil, Stockholm, Sweden.
    Rapid Production and Recovery of Cell Spheroids by Automated Droplet Microfluidics2020In: SLAS TECHNOLOGY, ISSN 2472-6303, Vol. 25, no 2, p. 111-122Article in journal (Refereed)
    Abstract [en]

    The future of the life sciences is linked to automation and microfluidics. As robots start working side by side with scientists, robotic automation of microfluidics in general, and droplet microfluidics in particular, will significantly extend and accelerate the life sciences. Here, we demonstrate the automation of droplet microfluidics using an inexpensive liquid-handling robot to produce human scaffold-free cell spheroids at high throughput. We use pipette actuation and interface the pipetting tip with a droplet-generating microfluidic device. In this device, we produce highly monodisperse droplets with a diameter coefficient of variation (CV) lower than 2%. By encapsulating cells in these droplets, we produce cell spheroids in droplets and recover them to standard labware containers at a throughput of 85,000 spheroids per microfluidic circuit per hour. The viability of the cells in spheroids remains high throughout the process and decreases by >10% (depending on the cell line used) after a 16 h incubation period in nanoliter droplets and automated recovery. Scaffold-free cell spheroids and 3D tissue constructs recapitulate many aspects of functional human tissue more accurately than 2D or single-cell cultures, but assembly methods for spheroids (e.g., hanging drop microplates) have limited throughput. The increased throughput and decreased cost of our method enable spheroid production at the scale needed for lead discovery drug screening, and approach the cost at which these microtissues could be used as building blocks for organ-scale regenerative medicine.

  • 31.
    Lehto, Tõnis
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Guidelines for developing safe and efficient acylated cell-penetrating peptides for nanoparticle mediated non-covalent nucleic acid delivery in vivoManuscript (preprint) (Other academic)
    Abstract [en]

    We have previously described how the in vitro therapeutic window of cell-penetrating peptides (CPPs) in non-covalent oligonucleotide delivery can be widened by increasing the N-terminal acyl chain length and by reducing the free fraction of peptide in the peptide/nucleic acid complexes. Here we show on the same peptide library how varying the acyl chain length from 2-22 carbons influences the complexation and plasmid delivery both in cell culture and systemically in vivo.

    For that we first show by DLS and electron microscopy that for efficient complexation of plasmid DNA into stable and condensed nanoparticles hydrophobic interactions play the key role. Moreover, these cationic nanoparticles maintain their size in serum containing cell culture media, but not in serum free DMEM. When the transfection ability of these peptide/pDNA complexes was assessed in cell cultures, the more stable analogs (C14-22) showed similar functional activity. On the contrary, when these complexes were intravenously administered to mice, the most hydrophobic analog (C22) showed superior gene induction in liver and lungs over other tested analogs. Moreover, at optimal peptide/pDNA charge ratio (CR2) the complexes with intermediate carbon chain analogs (C8-C14) caused acute death in around 50% of the animals, whereas, with shorter and longer analogs survival was not affected. To emphasize the discrepancies between gene induction in cell culture and in vivo even further, we demonstrate by changing the size of the complexes and centrifugation that in cell culture the transfection efficiency is in part dependent on sedimentation, which can be misleading when translating these formulations to in vivo. Collectively these findings provide guidelines for how to design safe and efficient next generation CPPs for nanoparticle-mediated intravenous nucleic acid delivery.

  • 32.
    Li, Jing
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Chemistry, Glycoscience.
    Wang, Damao
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Chemistry, Glycoscience.
    Xing, Xiaohui
    Adelaide Glycomics, School of Agriculture, Food and Wine, University of Adelaide, Waite Campus, Urrbrae, SA 5064, Australia.
    Cheng, Ting-Jen Rachel
    Genomics Research Centre, Academia Sinica, Sec. 2, 128 Academia Road, Nankang, Taipei 115, Taiwan.
    Liang, Pi-Hui
    School of Pharmacy, College of Medicine, National Taiwan University, Taipei 100, Taiwan.
    Bulone, Vincent
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Chemistry, Glycoscience. Adelaide Glycomics, School of Agriculture, Food and Wine, University of Adelaide, Waite Campus, Urrbrae, SA 5064, Australia.
    Park, Jeong Hill
    College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul, 08826, Republic of Korea.
    Hsieh, Yves S. Y.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Chemistry, Glycoscience. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Centres, Wallenberg Wood Science Center.
    Structural analysis and biological activity of cell wall polysaccharides extracted from Panax ginseng marc2019In: International Journal of Biological Macromolecules, ISSN 0141-8130, E-ISSN 1879-0003, Vol. 135, p. 29-37Article in journal (Refereed)
    Abstract [en]

    Ginseng marc is a major by-product of the ginseng industry currently used as animal feed or fertilizer. This fibrous, insoluble waste stream is rich in cell wall polysaccharides and therefore a potential source of ingredients for functional food with health-promoting properties. However, the extraction of these polysaccharides has proved problematic and their exact composition remains unknown. Here we have analysed the composition, structure and biological activity of polysaccharides from ginseng root, stem and leaf marc fractionated using a chelator and alkali solutions. The pectic fraction has been extracted from root marc in high abundance and can activate the production of interleukine-1α and the hematopoietic growth factor by RAW 264.7 murine macrophage cells, which are important immune regulators of T-cells during inflammatory responses and infection processes. Our study reveals the potential to increase the value of ginseng marc by generating carbohydrate-based products with a higher value than animal feed.

  • 33. Li, Yanping
    et al.
    Zheng, Yuting
    Zhang, Ye
    Yang, Yuanyuan
    Wang, Peiyao
    Imre, Balázs
    Wong, Ann C. Y.
    Hsieh, Yves S. Y.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Chemistry, Glycoscience. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Centres, Wallenberg Wood Science Center.
    Wang, Damao
    Brown Algae Carbohydrates: Structures, Pharmaceutical Properties, and Research Challenges2021In: Marine Drugs, E-ISSN 1660-3397, Vol. 19, article id 620Article, review/survey (Refereed)
    Abstract [en]

    Brown algae (Phaeophyceae) have been consumed by humans for hundreds of years. Current studies have shown that brown algae are rich sources of bioactive compounds with excellent nutritional value, and are considered functional foods with health benefits. Polysaccharides are the main constituents of brown algae; their diverse structures allow many unique physical and chemical properties that help to moderate a wide range of biological activities, including immunomodulation, antibacterial, antioxidant, prebiotic, antihypertensive, antidiabetic, antitumor, and anticoagulant activities. In this review, we focus on the major polysaccharide components in brown algae: the alginate, laminarin, and fucoidan. We explore how their structure leads to their health benefits, and their application prospects in functional foods and pharmaceuticals. Finally, we summarize the latest developments in applied research on brown algae polysaccharides.

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  • 34.
    Lindbo, Sarah
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Generation and engineering of ABD-derived affinity proteins for clinical applications2018Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Proteins that specifically recognize and bind to other molecules or structures are important tools in industrial and medical applications. Binding proteins engineered from small stable scaffold proteins have been utilized for several purposes due to their favorable biophysical properties, tolerance to mutagenesis, efficient tissue penetration and ease of production. The 46 amino acid long albumin-binding domain (ABD) derived from the bacterial receptor Protein G is a promising scaffold that has been explored in this thesis. The scaffold was subjected to combinatorial protein engineering for generation of ABD-derived binding proteins with novel specificities. Furthermore, the medical potential of engineered ABD- derived affinity proteins (ADAPTs) was evaluated in a series of pre-clinical studies.

    In the first studies, ADAPTs suitability as tracers for radionuclide molecular imaging was evaluated. Factors influencing biodistribution and tumor targeting properties were assessed in mice models bearing HER2 positive xenografts. All tested ADAPT constructs demonstrated high and specific targeting of HER2-expressing tumor cells as well as fast clearance from circulation. The results also showed that the size and character of the N- terminus affected the biodistribution profile of ADAPTs. Moreover, the targeting properties of ADAPTs proved to be highly influenced by the residualizing properties of the attached radionuclide label. Taken together, the results provided the first evidence that tumor imaging can be performed using ADAPTs and the favorable pharmacokinetic profiles in the studied mice models suggest that the scaffold is a promising candidate for clinical applications.

    In the last study, a platform for generation of stable ABD-derived affinity proteins with novel binding specificities was established using a multi-step approach combining directed evolution and rational protein design. A broad combinatorial protein library with 20 randomized positions in ABD was designed and binders against three distinct targets were selected using phage display. Characterization of the selected binders provided information regarding optimal positions to randomize in a final library. In addition, the isolated binders were subjected to mutagenesis in certain surface exposed positions and mutations that provided increased stability were introduced into the original scaffold. Finally, a more focused combinatorial protein library consisting of 11 randomized positions was designed and constructed. The library was validated by selections against the same set of targets as for the first, broad library. The isolation of highly stable affinity ligands confirms that the library can be used for generation of diverse and stable affinity molecules.

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    Sarah Lindbo 2018
  • 35.
    Matheis, Sebastian
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Preparation of a Class A certification in the field of pharmaceutical packaging through mapping and optimization of business processes and implementation of the Oliver Wight Class A Behaviors for Business Excellence.2014Independent thesis Advanced level (degree of Master (Two Years)), 30 credits / 45 HE creditsStudent thesis
    Abstract [en]

    On the road to success, companies need to fulfil their stakeholders’ expectations. On the road to business excellence, companies need to exceed these expectations. Oliver Wight Inc. has established a certification called Class A Business, which shows that a company is exceeding stakeholders’ expectations and that it performs in the upper quartile in its respective industry. The Class A Business certification is awarded, once a company fulfils a certification checklist with Class A Business criteria. To get to this point, a company can design their road to business excellence by following a specific set of nine Class A Behaviors.

    This study focuses on four of these behaviors, divided in three parts, and how they are implemented at a pharmaceutical packaging department at Roche in Kaiseraugst, Switzerland. In addition, through employee feedback potential areas of improvement are identified.

    For a company to understand how it is running, it has to understand its underlying processes. Once the processes are in place, a process-oriented way of thinking can change a company to make decisions based on process’ needs rather than on individual preferences. Business processes and their potential for continuous improvement were the first part of the study. The second part of the study investigated the communication of different functions in the packaging process and how the flow of information could be improved. In the third part, the usage of operational metrics in the packaging department is researched by a user feedback survey.

    An innovative way to visualize meeting conversations was developed in this study to make meetings more tangible for the reader. This is a newly developed and never before described method for business research colorfully showing interactions in meetings.

    The results are very intriguing. Simple thought business elements seem to pose larger hurdles than would be expected sometimes.

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    Master Thesis of Sebastian Matheis
  • 36.
    Meister, Sebastian
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Hjelm, Linnea C.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Dannemeyer, Melanie
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Tegel, Hanna
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Lindberg, Hanna
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Ståhl, Stefan
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Löfblom, John
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Engineering.
    An Affibody Molecule Is Actively Transported into the Cerebrospinal Fluid via Binding to the Transferrin Receptor2020In: International Journal of Molecular Sciences, ISSN 1661-6596, E-ISSN 1422-0067, Vol. 21, no 8, p. 2999-Article in journal (Refereed)
    Abstract [en]

    Theuseofbiotherapeuticsforthetreatmentofdiseasesofthecentralnervoussystem(CNS) is typically impeded by insufficient transport across the blood–brain barrier. Here, we investigate a strategy to potentially increase the uptake into the CNS of an affibody molecule (ZSYM73) via binding to the transferrin receptor (TfR). ZSYM73 binds monomeric amyloid beta, a peptide involved in Alzheimer’s disease pathogenesis, with subnanomolar affinity. We generated a tri-specific fusion protein by genetically linking a single-chain variable fragment of the TfR-binding antibody 8D3 and an albumin-binding domain to the affibody molecule ZSYM73. Simultaneous tri-specific target engagementwasconfirmedinabiosensorexperimentandtheaffinityformurineTfRwasdetermined to 5 nM. Blockable binding to TfR on endothelial cells was demonstrated using flow cytometry and in a preclinical study we observed increased uptake of the tri-specific fusion protein into the cerebrospinal fluid 24 h after injection.

  • 37.
    Meister, Sebastian
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Parks, Luke
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Kolmar, Leonie
    Ståhl, Stefan
    KTH, Superseded Departments (pre-2005), Biotechnology. KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova.
    Löfblom, John
    KTH, School of Engineering Sciences (SCI). KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Engineering.
    Engineering the substrate specificity of TEV protease towards an Aβ-cleaving enzymeManuscript (preprint) (Other academic)
    Abstract [en]

    Due to their ability to catalytically cleave proteins and peptides, proteases present unique opportunities for the use in industrial, biotechnological, and therapeutic applications. The possibility to engineer proteases with redesigned substrate specificities has the potential to expand the scope of practical applications of this enzyme class. We here apply a combinatorial protease engineering screening method that links proteolytic activity to the solubility and correct folding of a fluorescent reporter protein to redesign the substrate specificity of Tobacco Etch Virus (TEV) protease. The target substrate EKLVFQA differs at three of seven positions from the TEV consensus substrate sequence and exhibits high sequence similarity to the aggregation-inducing hydrophobic core region of the amyloid beta (Aβ) peptide. Flow cytometric sorting of a semi-rational TEV protease library led to the enrichment of a set of protease variants that recognize and cleave the novel target substrate.

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  • 38. Melle, Giovanni
    et al.
    Bruno, Giulia
    Maccaferri, Nicolò
    Department of Physics and Materials Science, University of Luxembourg, Luxembourg, Luxembourg.
    Iachetta, Giuseppina
    Colistra, Nicolò
    Barbaglia, Andrea
    Dipalo, Michele
    De Angelis, Francesco
    Intracellular Recording of Human Cardiac Action Potentials on Market-Available Multielectrode Array Platforms2020In: Frontiers in Bioengineering and Biotechnology, E-ISSN 2296-4185, Vol. 8, article id 66Article in journal (Refereed)
    Abstract [en]

    High quality attenuated intracellular action potentials from large cell networks can be recorded on multi-electrode arrays by means of 3D vertical nanopillars using electrical pulses. However, most of the techniques require complex 3D nanostructures that prevent the straightforward translation into marketable products and the wide adoption in the scientific community. Moreover, 3D nanostructures are often delicate objects that cannot sustain several harsh use/cleaning cycles. On the contrary, laser optoacoustic poration allows the recording of action potentials on planar nanoporous electrodes made of noble metals. However, these constraints of the electrode material and morphology may also hinder the full exploitation of this methodology. Here, we show that optoacoustic poration is also very effective for porating cells on a large family of MEA electrode configurations, including robust electrodes made of nanoporous titanium nitride or disordered fractal-like gold nanostructures. This enables the recording of high quality cardiac action potentials in combination with optoacoustic poration, providing thus attenuated intracellular recordings on various already commercial devices used by a significant part of the research and industrial communities.

  • 39.
    Ohlander, Anna
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology. Fraunhofer EMFT, Hansastraße 27d, Munich, 80686, Germany.
    Bose, Indranil
    Fraunhofer EMFT, Hansastraße 27d, Munich, 80686, Germany. nstitute of Physics, Universität der Bundeswehr München, 85577, Germany.
    Njenda, Duncan T.
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Huddinge, Stockholm, Sweden.
    Neogi, U.
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Huddinge, Stockholm, Swede.
    Kutter, Christoph
    Fraunhofer EMFT, Hansastraße 27d, Munich, 80686, Germany. nstitute of Physics, Universität der Bundeswehr München, 85577, Germany.
    Russom, Aman
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology.
    Lab-on-foil based portable μPCR for POC nucleic acid testing of HIV-12020In: 21st International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2017, Chemical and Biological Microsystems Society , 2020, p. 1246-1247Conference paper (Refereed)
    Abstract [en]

    We present a low-cost μPCR platform using a thin film microheater on foil, fabricated in only one metallization step using roll-to-roll fabrication. The μPCR module was successfully applied for amplification of HIV-1 viral load in three clinical samples of which one was negative control. The μPCR module was able to amplify the target in the two HIV positive samples whereas the COBAS TaqMan HIV-1 from Roche and an in-house developed qPCR assay using TaqMan probes could not. These results indicate that the μPCR module has the potential to be very sensitive. The low cost μPCR system opens up for the possibility to perform molecular diagnostics at the POC - ideally suited at low-resource settings.

  • 40.
    Parakkal Sreenivasan, Akshai
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Deep learning prediction of Quantmap clusters2021Independent thesis Advanced level (degree of Master (Two Years)), 30 credits / 45 HE creditsStudent thesis
    Abstract [en]

    The hypothesis that similar chemicals exert similar biological activities has been widely adopted in the field of drug discovery and development. Quantitative Structure-Activity Relationship (QSAR) models have been used ubiquitously in drug discovery to understand the function of chemicals in biological systems. A common QSAR modeling method calculates similarity scores between chemicals to assess their biological function. However, due to the fact that some chemicals can be similar and yet have different biological activities, or conversely can be structurally different yet have similar biological functions, various methods have instead been developed to quantify chemical similarity at the functional level. Quantmap is one such method, which utilizes biological databases to quantify the biological similarity between chemicals. Quantmap uses quantitative molecular network topology analysis to cluster chemical substances based on their bioactivities. This method by itself, unfortunately, cannot assign new chemicals (those which may not yet have biological data) to the derived clusters. Owing to the fact that there is a lack of biological data for many chemicals, deep learning models were explored in this project with respect to their ability to correctly assign unknown chemicals to Quantmap clusters. The deep learning methods explored included both convolutional and recurrent neural networks. Transfer learning/pretraining based approaches and data augmentation methods were also investigated. The best performing model, among those considered, was the Seq2seq model (a recurrent neural network containing two joint networks, a perceiver and an interpreter network) without pretraining, but including data augmentation.

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  • 41.
    Possnert, Oliver
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Schön, Adam
    A case study research of asymmetrical relationshipsbetween service providers and emerging companieswithin the healthcare industry2018Independent thesis Basic level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    This master thesis report aims to highlight the importance ofinterorganizational relationships between experienced serviceproviders and emerging biopharmaceutical (EBP) companies within theSwedish healthcare industry. A shift in innovation strategiesregarding new pharmaceutical- and medical device products hasprompted a paradigm shift within a complex industry wherecollaborations between organisations has become increasinglycrucial. With a better understanding of how these companiesoperates, increased collaboration efforts could result in a fasterand more precise product development with new products reaching themarket improving the health for people around the world. In order toallow experienced service providers to enhance services towards EBPcompanies, a fundamental understanding of how decision makers withinthese EBP companies prefer to conduct relationships is needed. Wehave examined relationship preferences of EBP companies byconducting a qualitative case study through 14 interviews withdecision makers combined with a quantitative conjoint analysis.Eight factors was identified as important for when EBP companiesdecide to engage with a service provider: cost behavior,professional competence, adaptability, communication, personalrelationship, stability, EBP insight and size. The factorsadaptability, personal relationship, cost and size were used in theconjoint analysis to determine their relative importance which showthat adaptability and cost behavior was of the largest importance.With descriptions of each factor, we have provided a meaningfulguide to action of how to address these factors as a serviceprovider. The relationships is largely investigated as relationshipsbetween contract research organizations (as service providers) andEBP companies, but we have created a framework applicable forservice providers within the healthcare industry in general.

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  • 42.
    Randek, Judit
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Mandenius, Carl-Fredrik
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    In situ scanning capacitance sensor with spectral analysis reveals morphological states in cultures for production of biopharmaceuticals2020In: Sensors and actuators. B, Chemical, ISSN 0925-4005, E-ISSN 1873-3077, Vol. 313, article id 128052Article in journal (Refereed)
    Abstract [en]

    In situ capacitance sensing is shown to be capable of monitoring critical morphological changes in industrial cultures by analyzing the sensor’s frequency spectrum. Scanning the frequency of an alternating current between the electrodes of a capacitance sensor, placed in a cell culture, allowed detection of the size change of microbial cells from shifts in the spectra. The frequency was scanned between 0.1–15 MHz and cell size was measured from 1 to 20 μm. The analysis of the spectra was verified with two recombinant strains, one producing human insulin and another Green Fluorescence Protein (GFP). Both the insulin and GFP cultivations were carried out in 6 L fed-batch bioreactors using typical industrial procedures. The spectral analysis provided critical information about the changes in the size of the cells. It is suggested that this information may have high relevance for a better assessment of the state of cultivations producing proteins, for optimization and for improving the economy of large-scale biopharmaceutical production.

  • 43.
    Rockberg, Johan
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.
    Split-GFP and droplet microfluidics allows high-throughput screening of mammalian cell factories2016In: New Biotechnology, ISSN 1871-6784, E-ISSN 1876-4347, Vol. 33, p. S51-S51Article in journal (Other academic)
  • 44.
    Shamsudin Khan, Yasmin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational Biology and Bioinformatics.
    Non-Steroidal Anti-Inflammatory Drugs in Cyclooxygenases 1 and 2: Binding modes and mechanisms from computational methods and free energy calculations2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Non-steroidal anti-inflammatory drugs (NSAIDs) are one of the most commonly used classes of drugs. They target the cyclooxygenases (COX) 1 and 2 to reduce the physiological responses of pain, fever, and inflammation. Due to their role in inducing angiogenesis, COX proteins have also been identified as targets in cancer therapies.

    In this thesis, I describe computational protocols of molecular docking, molecular dynamics simulations and free energy calculations. These methods were used in this thesis to determine structure-activity relationships of a diverse set of NSAIDs in binding to their target proteins COX-1 and 2. Binding affinities were calculated and used to predict the binding modes. Based on combinations of molecular dynamics simulations and free energy calculations, binding mechanisms of sub-classes of NSAIDs were also proposed. Two stable conformations of COX were probed to understand how they affect inhibitor affinities. Finally, a brief discussion on selectivity towards either COX isoform is discussed. These results will be useful in future de novo design and testing of third-generation NSAIDs.

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  • 45.
    Smedsrud, Sabina
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Industrial Engineering & Management.
    Ekdahl, Simon
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Industrial Engineering & Management.
    Näslund, Emil
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Industrial Engineering & Management.
    Optimising a launch: Important factors affecting a new pharmaceutical launch in Sweden2019Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    This master thesis explores the launch process of new pharmaceuticals in Sweden. The path of a pharmaceutical from idea to innovation is a long and arduous process with only few new products actually reaching the patients in the end. Seeing as the drug development is also an expensive process, it is of importance that the products that get approval meet their expected revenue. New pharmaceuticals can also be life changing for the patient, and thus it is important that once approval is received the patients gain access to the new treatments. This study focuses on the post regulatory approval processes in Sweden, as well as activities carried out by the companies that affect the adoption of a new product.

    By utilizing a qualitative study, this thesis aims to describe the internal and external factors that affect the pharmaceutical launch process in Sweden. As well as exploring what future initiatives and possible changes that might affect it. Ten interviews with different company representatives as well as six interviews with governmental and regional stakeholders were analysed using grounded theory to answer what factors affect the adoption of new pharmaceuticals. Factors that were found to be important were: Utilisation of cross-functional teams, clear and simple strategy that includes the whole organisation, communicating with national and regional authorities, and get feedback from these, communicate with patient representatives and organisations as well as developing utility services for the product. From a couple of these factors a trend towards the servicification of the pharmaceutical industry was discovered.

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  • 46.
    Strömme, Maria
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Physics.
    Niklasson, Gunnar A.
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Physics.
    Ek, Ragnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
    Densification-induced conductivity percolation in high-porosity pharmaceutical microcrystalline cellulose compacts2003In: Applied Physics Letters, ISSN 0003-6951, E-ISSN 1077-3118, Vol. 82, no 4, p. 648-650Article in journal (Refereed)
    Abstract [en]

    The percolation theory is established as a useful tool in the field of pharmaceutical materials science.It is shown that percolation theory, developed for analyzing insulator–conductor transitions, can beapplied to describe imperfect dc conduction in pharmaceutical microcrystalline cellulose duringdensification. The system, in fact, exactly reproduces the values of the percolation threshold andexponent estimated for a three-dimensional random continuum. Our data clearly show a crossoverfrom a power-law percolation theory region to a linear effective medium theory region at a celluloseporosity of ;0.7.

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  • 47.
    Svahn, Carl Fabian
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Khan, Anisha
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Wahlsten, Amanda
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Larsson, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Koivula, Therese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Andersson, Thomas
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Drugs of the Future - Bispecific Antibodies: An investigaion of future development needs2019Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    This report reviews the field of bispecific antibodies, artificially engineered antibodies thathave the ability to bind two or more different antigen simultaneously. Historical as well asrecently developed techniques are demonstrated, together with formats in preclinical andclinical development. We studied the field with the future needs of the developers in mind,when it comes to the processes and tools that can be offered by GE Healthcare BiosciencesAB.

    The development of bispecific antibodies gave rise to new challenges and product-relatedimpurities, which are handled by various methods. We argue for, based on the formats inclinical and preclinical development, that the methods already used to purify monospecificantibodies remain the most successful methods for the purification of bispecific antibodies.This, together with the design strategies that resolve the initial bottle-necks, ensures that theneeds of the developers are met to the same extent as for monoclonal antibodies. The methodsand formats demonstrated here do not represent all that are available or under trial.

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  • 48.
    Vadi Dris, Sam
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
    Development of ultra-sensitive immunoassay on Gyrolab microfluidic platform using Binding Oligo Ladder Detection: Enhancing Gyrolab biomarker assays using Exazym®2024Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Immunoassays are widely used for detection of antigens in a wide range of applications including assays in pharmaceutical development. Immunoassays are continuously improved in many aspects including automatization, miniaturization and extending the dynamic range. The need to measure low abundance molecules are challenging and the need to improve the sensitivity is desired. The Gyrolab technology is a miniaturized immunoassay performed in an automated system covering a broad concentration range. In order to  extend the sensitivity, the technology is combined with Binding Oligo Ladder Detection (BOLD) amplification. The technology behind BOLD or Exazym

    ® utilizes a DNA primer, a polymerase, and a template (RNA) to generate a ladder-like modified DNA strand. Antibodies with affinity for the polymerized DNA:RNA hybrid strand (duplex) conjugated with reporter molecules are introduced to the system, resulting in an increased number of signal-generating molecules associated with each bound analyte molecule. In this thesis, the development of an ultra-sensitive immunoassay is pursued by applying Exazym

    ® add-on reagents to the Gyrolab platform, comparing performance with the standard Gyrolab sandwich assay and other commercially available high-performing TNF-α assays. The work includes characterization of a wide range of reaction variables involved in the BOLD signal amplification process including hybridization, polymerization, and detection of a synthetic oligonucleotide. The breakthrough involves the introduction of Allophycocyanin (APC) as a fluorescent conjugate, significantly improving sensitivity and signal-to-noise ratios. The BOLD amplified sensitivity for the TNF-α assay approaches levels seen in ultra-sensitive biomarker assays like Erenna

    ® and Simoa®. Exazym® technology on the Gyrolab platform allows highly sensitive biomarker assays with minimal sample volume and a 1–2-hour run-time. The study marks substantial progress in achieving ultra-sensitive biomarker assays on the Gyrolab platform through BOLD signal amplification. The use of APC-conjugated detection reagents holds promise for future optimization studies.

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  • 49.
    Volk, Anna-Luisa
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Cell line and protein engineering tools for production and characterization of biologics2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Our increasing understanding of disease mechanisms coupled with technological advances has facilitated the generation of pharmaceutical proteins, which are able to address yet unmet medical needs. Diseases that were fatal in the past can now be treated with novel biological medications improving and prolonging life for many patients. Pharmaceutical protein production is, however, a complex undertaking, which is by no means problem-free. The demand for more complex proteins and the realization of the importance of post-translational modifications have led to an increasing use of mammalian cells for protein expression. Despite improvements in design and production, the costs required for the development of pharmaceutical proteins still are far greater than those for conventional, small molecule drugs. To render such treatments affordable for healthcare suppliers and assist in the implementation of precision medicine, further progress is needed. In five papers this thesis describes strategies and methods that can help to advance the development and manufacturing of pharmaceutical proteins. Two platforms for antibody engineering have been developed and evaluated, one of which allows for efficient screening of antibody libraries whilst the second enables the straightforward generation of bispecific antibodies. Moreover, a method for epitope mapping has been devised and applied to map the therapeutic antibody eculizumab’s epitope on its target protein. In a second step it was shown how this epitope information can be used to stratify patients and, thus, contribute to the realization of precision medicine. The fourth project focuses on the cell line development process during pharmaceutical protein production. A platform is described combining split-GFP and fluorescence-activated droplet sorting, which allows for the efficient selection of highly secreting cells from a heterogeneous cell pool. In an accompanying study, the split-GFP probe was improved to enable shorter assay times and increased sensitivity, desirable characteristics for high-throughput screening of cell pools. In summary, this thesis provides tools to improve design, development and production of future pharmaceutical proteins and as a result, it makes a contribution to the goal of implementing precision medicine through the generation of more cost-effective biopharmaceuticals for well-characterized patient groups.

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  • 50.
    Volk, Anna-Luisa
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Hansen, Henning G.
    Lundqvist, Magnus
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Hammar, Petter
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Bai, Yunpeng
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Kol, Stefan
    Kildegaard, Helene F.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab. Technical University of Denmark, Denmark.
    Joensson, Haakan N.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Rockberg, Johan
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Droplet microfluidics and split-GFP complementation enable selection of Chinese hamster ovary cells with high specific productivity of therapeutic glycoproteinsManuscript (preprint) (Other academic)
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