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  • 1.
    Mateus, André
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci. Faculty of Pharmacy, University of Lisbon.
    Intracellular unbound drug concentrations: Methodology and application for understanding cellular drug exposure2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Most known drug targets and metabolizing enzymes are located inside cells. Interactions with these proteins are determined by intracellular unbound drug concentrations. Assessing intracellular drug exposure is technically challenging, but essential for predicting pharmacokinetic, pharmacological, and toxicological profiles of new drugs.

    This thesis aims at establishing and applying a straightforward methodology to measure intracellular unbound drug concentrations. This was achieved by separately measuring cellular drug binding (fu,cell), and total intracellular drug accumulation (Kp). This allowed the calculation of intracellular drug bioavailability (Fic), which represents the fraction of the concentration added to the cells that is unbound in the cell interior.

    The methodology was initially developed in HEK293 cells, where the Fic of 189 drug-like compounds was measured. Binding to HEK293 cells was governed by compound lipophilicity and was correlated with binding to more complex systems, such as hepatocytes and brain. Due to negligible expression of drug transporters, Fic in this cell line was consistent with pH-dependent subcellular sequestration of lipophilic cations in low pH compartments.

    The methodology was then applied to study the effects of drug transporters on Fic. The uptake transporter OATP1B1 increased the Fic of its substrates in a concentration-dependent manner. In contrast, the Fic of P-gp substrates was decreased when P-gp was present. In human hepatocytes, the methodology allowed the determination of Fic without prior knowledge of transporter mechanisms or metabolic activity.

    Finally, the methodology was applied to measure the impact of Fic on target binding and cellular drug response. Intracellular concentrations of active metabolites of pro-drugs targeting the intracellular target thymidylate synthase were in agreement with the level of binding to this target. Further, high Fic was generally required for kinase and protease inhibitors to be active in cellular assays.

    In conclusion, the methodology can be used to predict if new drug candidates reach their intracellular targets in sufficient amounts. Furthermore, the methodology can improve in vitro predictions of drug clearance and drug-drug interactions, by measuring the drug available for intracellular enzymes. Finally, this work can be expanded to other xenobiotics, e.g., to predict their intracellular toxicity.

    Delarbeten
    1. Rapid Measurement of Intracellular Unbound Drug Concentrations
    Öppna denna publikation i ny flik eller fönster >>Rapid Measurement of Intracellular Unbound Drug Concentrations
    2013 (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, E-ISSN 1543-8392, Vol. 10, nr 6, 2467-2478 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Intracellular unbound drug concentrations determine affinity to targets in the cell interior. However, due to difficulties in measuring them, they are often overlooked in pharmacology. Here we present a simple experimental technique for the determination of unbound intracellular drug concentrations in cultured cells that is based on parallel measurements of cellular drug binding and steady-state intracellular drug concentrations. Binding in HEK293 cells was highly correlated with binding in liver-derived systems, whereas binding in plasma did not compare well with cellular binding. Compound lipophilicity increased drug binding, while negative charge and aromatic functional groups decreased binding. Intracellular accumulation of unbound drug was consistent with pH dependent subcellular sequestration, as confirmed by modeling and by inhibition of subcellular pH gradients. The approach developed here can be used to measure intracellular unbound drug concentrations in more complex systems, for example, cell lines with controlled expression of transporters and enzymes or primary cells.

    Nyckelord
    intracellular unbound concentrations, drug binding, drug transport, drug accumulation, membrane partitioning
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-204292 (URN)10.1021/mp4000822 (DOI)000320015600037 ()
    Tillgänglig från: 2013-07-29 Skapad: 2013-07-29 Senast uppdaterad: 2016-04-04
    2. A High-Throughput Cell-Based Method to Predict the Unbound Drug Fraction in the Brain
    Öppna denna publikation i ny flik eller fönster >>A High-Throughput Cell-Based Method to Predict the Unbound Drug Fraction in the Brain
    2014 (Engelska)Ingår i: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 57, nr 7, 3005-3010 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Optimization of drug efficacy in the brain requires understanding of the local exposure to unbound drug at the site of action. This relies on measurements of the unbound drug fraction (f(u,brain)), which currently requires access to brain tissue. Here, we present a novel methodology using homogenates of cultured cells for rapid estimation of f(u,brain). In our setup, drug binding to human embryonic kidney cell (HEK293) homogenate was measured in a small-scale dialysis apparatus. To increase throughput, we combined drugs into cassettes for simultaneous measurement of multiple compounds. Our method estimated f(u,brain) with an average error of 1.9-fold. We propose that our simple method can be used as an inexpensive, easily available and high-throughput alternative to brain tissues excised from laboratory animals. Thereby, estimates of unbound drug exposure can now implemented at a much earlier stage of the drug discovery process, when molecular property changes are easier to make.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-224730 (URN)10.1021/jm401963n (DOI)000334572000017 ()
    Tillgänglig från: 2014-05-22 Skapad: 2014-05-19 Senast uppdaterad: 2016-04-04
    3. Impact of drug transporters on intracellular drug concentrations
    Öppna denna publikation i ny flik eller fönster >>Impact of drug transporters on intracellular drug concentrations
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    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-276093 (URN)
    Tillgänglig från: 2016-02-09 Skapad: 2016-02-09 Senast uppdaterad: 2016-04-04
    4. CETSA screening identifies known and novel thymidylate synthase inhibitors and slow intracellular activation of 5-fluorouracil
    Öppna denna publikation i ny flik eller fönster >>CETSA screening identifies known and novel thymidylate synthase inhibitors and slow intracellular activation of 5-fluorouracil
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    2016 (Engelska)Ingår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 7, 11040Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Target engagement is a critical factor for therapeutic efficacy. Assessment of compound binding to native target proteins in live cells is therefore highly desirable in all stages of drug discovery. We report here the first compound library screen based on biophysical measurements of intracellular target binding, exemplified by human thymidylate synthase (TS). The screen selected accurately for all the tested known drugs acting on TS. We also identified TS inhibitors with novel chemistry and marketed drugs that were not previously known to target TS, including the DNA methyltransferase inhibitor decitabine. By following the cellular uptake and enzymatic conversion of known drugs we correlated the appearance of active metabolites over time with intracellular target engagement. These data distinguished a much slower activation of 5-fluorouracil when compared with nucleoside-based drugs. The approach establishes efficient means to associate drug uptake and activation with target binding during drug discovery.

    Nationell ämneskategori
    Farmakologi och toxikologi
    Identifikatorer
    urn:nbn:se:uu:diva-276077 (URN)10.1038/ncomms11040 (DOI)000372887500001 ()27010513 (PubMedID)
    Forskningsfinansiär
    Karolinska Institutets ForskningsstiftelseVetenskapsrådetCancerfondenKnut och Alice Wallenbergs Stiftelse
    Tillgänglig från: 2016-02-09 Skapad: 2016-02-09 Senast uppdaterad: 2016-05-25Bibliografiskt granskad
    5. Impact of intracellular drug bioavailability on cellular drug response
    Öppna denna publikation i ny flik eller fönster >>Impact of intracellular drug bioavailability on cellular drug response
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    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nationell ämneskategori
    Farmakologi och toxikologi
    Identifikatorer
    urn:nbn:se:uu:diva-276089 (URN)
    Tillgänglig från: 2016-02-09 Skapad: 2016-02-09 Senast uppdaterad: 2016-04-04
  • 2.
    Hellrup, Joel
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Pharmaceutical Nanocomposites: Structure–Mobility–Functionality Relationships in the Amorphous State2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Amorphous materials are found in pharmaceutical formulations both as excipients and active ingredients. Indeed, these formulations are becoming an essential strategy for incorporating drugs into well-performing solid dosage forms. However, there is an unmet need of better understanding of the microstructure and component interactions in amorphous formulations to be able to design materials with improved functionalities. The aim of this thesis is to give deepened knowledge about structure-mobility-functionality relationships in amorphous for-mulations by studying composites produced from sugars and filler particles. The structure, the mobility, and physical stability of the composite materials were studied using calorimetry, X-ray diffraction, microscopy, spectroscopy, and molecular dynamics simulations. Further, the moisture sorption of the composites was determined with dynamic vapor sorption. The compression mechanics of the composites was evaluated with compression analysis.

    It was demonstrated that fillers change the overall properties of the amorphous material. Specifically, the physical stability of the composite was by far improved compared to the amorphous sugar alone. This effect was pronounced for formulations with 60 wt% filler content or more. Amorphous lactose that normally recrystallizes within a few minutes upon humidity exposure, could withstand recrystallization for several months at 60% RH in composites with 80 wt% cellulose nanocrystals (CNC) or sodium montmorillonite (Na-MMT). The increased physical stability of the amorphous sugars was related to intra-particle confinement in extra-particle voids formed by the fillers and to immobilization of the amorphous phase at the surface of the fillers. Also, the composite formation led to increased particle hardness for the lactose/CNC and the lactose/Na-MMT nanocomposites. The largest effect on particle hardness was seen with 40-60 wt% nanofiller and could be related to skeleton formation of the nanofillers within the composite particles. The hygroscopicity for the lactose/Na-MMT nanocomposites decreased as much as 47% compared to ideal simple mixtures of the neat components. The nanofillers did not influence the water sorption capacity in the amorphous domains; however, lactose (intercalated into Na-MMT) interacted with the sodium ions in the interlayer space which led to the lowered hygroscopicity of this phase.

    The thesis advanced the knowledge of the microstructure of amorphous pharmaceutical com-posites and its relationship with pharmaceutical functionalities. It also presented new approaches for stabilizing the amorphous state by using fillers. The concept illustrated here might be used to understand similar phenomena of stabilization of amorphous formulations.

    Delarbeten
    1. Pharmaceutical micro-particles give amorphous sucrose higher physical stability
    Öppna denna publikation i ny flik eller fönster >>Pharmaceutical micro-particles give amorphous sucrose higher physical stability
    2011 (Engelska)Ingår i: Internation Journal of Pharmaceutics, ISSN 0378-5173, E-ISSN 1873-3476, Vol. 409, nr 1-2, 96-103 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The aim of this study was to explore how pharmaceutical micro-sized filler particles affect the amorphous stability of sucrose in sucrose/filler particle composites produced by freeze-drying. Focus was put on the filler particles' properties crystallinity, hygroscopicity, hydrophobicity, and surface area, and their influence on physical stability of the amorphous phase. The micro-sized filler particles were examined with Blaine permeametry, gas adsorption, pycnometry, gravimetric vapour sorption, X-ray diffraction, and light microscopy before composites of sucrose and micro-sized filler particles were prepared by freeze-drying. The stability of the composites was examined with X-ray diffraction, differential scanning calorimetry (DSC), and microcalorimetry. All composites were amorphous and showed higher stability compared to pure amorphous sucrose, which was evident from a delay in heat and moisture-induced crystallization. However, calcium carbonate and oxazepam micro-sized filler particles lost their ability to stabilize the amorphous sucrose when exposed to humidity. The dry glass transition temperature (T-g) was higher for the composites, indicating the stabilization was mediated by a reduced molecular mobility of the amorphous phase.

    Nyckelord
    Amorphous, Sucrose, Physical stability, Freeze-drying, Crystallization, Micro-particles
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-153962 (URN)10.1016/j.ijpharm.2011.02.031 (DOI)000290135800012 ()21356288 (PubMedID)
    Tillgänglig från: 2011-05-23 Skapad: 2011-05-23 Senast uppdaterad: 2016-09-01Bibliografiskt granskad
    2. Inhibition of Recrystallization of Amorphous Lactose in Nanocomposites Formed by Spray-Drying
    Öppna denna publikation i ny flik eller fönster >>Inhibition of Recrystallization of Amorphous Lactose in Nanocomposites Formed by Spray-Drying
    2015 (Engelska)Ingår i: Journal of Pharmaceutical Sciences, ISSN 0022-3549, E-ISSN 1520-6017, Vol. 104, nr 11, 3760-3769 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    This study aims at investigating the recrystallization of amorphous lactose in nanocomposites. In particular, the focus is on the influence of the nano- to micrometer length scale nanofiller arrangement on the amorphous to crystalline transition. Further, the relative significance of formulation composition and manufacturing process parameters for the properties of the nanocomposite was investigated. Nanocomposites of amorphous lactose and fumed silica were produced by co-spray-drying. Solid-state transformation of the lactose was studied at 43%, 84%, and 94% relative humidity using X-ray powder diffraction and microcalorimetry. Design of experiments was used to analyze spray-drying process parameters and nanocomposite composition as factors influencing the time to 50% recrystallization. The spray-drying process parameters showed no significant influence. However, the recrystallization of the lactose in the nanocomposites was affected by the composition (fraction silica). The recrystallization rate constant decreased as a function of silica content. The lowered recrystallization rate of the lactose in the nanocomposites could be explained by three mechanisms: (1) separation of the amorphous lactose into discrete compartments on a micrometer length scale (compartmentalization), (2) lowered molecular mobility caused by molecular interactions between the lactose molecules and the surface of the silica (rigidification), and/or (3) intraparticle confinement of the amorphous lactose.

    Nyckelord
    amorphous, crystallization, glass transition, mobility, physical stability, solid state, stabilization, spray drying, factorial design
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-266687 (URN)10.1002/jps.24583 (DOI)000362984100013 ()26182904 (PubMedID)
    Tillgänglig från: 2015-11-12 Skapad: 2015-11-10 Senast uppdaterad: 2016-12-07
    3. Structure and mobility of lactose in lactose/sodium montmorillonite nanocomposites
    Öppna denna publikation i ny flik eller fönster >>Structure and mobility of lactose in lactose/sodium montmorillonite nanocomposites
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    2016 (Engelska)Ingår i: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 32, nr 49, 13214-13225 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    This study aims at investigating the molecular level organization and molecular mobility in montmorillonite nanocomposites with the uncharged organic low-molecular-weight compound lactose commonly used in pharmaceutical drug delivery, food technology, and flavoring. Nanocomposites were prepared under slow and fast drying conditions, attained by drying at ambient conditions and by spray-drying, respectively. A detailed structural investigation was performed with modulated differential scanning calorimetry, powder X-ray diffraction, solid-state nuclear magnetic resonance spectroscopy, scanning electron microscopy, microcalorimetry, and molecular dynamics simulations. The lactose was intercalated in the sodium montmorillonite interlayer space regardless of the clay content, drying rate, or humidity exposure. Although, the spray-drying resulted in higher proportion of intercalated lactose compared with the drying under ambient conditions, nonintercalated lactose was present at 20 wt % lactose content and above. This indicates limitations in maximum loading capacity of nonionic organic substances into the montmorillonite interlayer space. Furthermore, a fraction of the intercalated lactose in the co-spray-dried nanocomposites diffused out from the clay interlayer space upon humidity exposure. Also, the lactose in the nanocomposites demonstrated higher molecular mobility than that of neat amorphous lactose. This study provides a foundation for understanding functional properties of lactose/Na-MMT nanocomposites, such as loading capacity and physical stability.

    Nationell ämneskategori
    Materialkemi
    Identifikatorer
    urn:nbn:se:uu:diva-300158 (URN)10.1021/acs.langmuir.6b01967 (DOI)000389866300029 ()
    Tillgänglig från: 2016-08-05 Skapad: 2016-08-03 Senast uppdaterad: 2017-03-08
    4. Confinement of Amorphous Lactose in Pores formed upon Co-Spray-Drying with Nanoparticles
    Öppna denna publikation i ny flik eller fönster >>Confinement of Amorphous Lactose in Pores formed upon Co-Spray-Drying with Nanoparticles
    (Engelska)Ingår i: Journal of Pharmaceutical Sciences, ISSN 0022-3549, E-ISSN 1520-6017Artikel i tidskrift (Övrigt vetenskapligt) In press
    Abstract [en]

    This study aims at investigating factors influencing humidity induced recrystallization of amorphous lactose, produced by co-spray-drying with particles of cellulose nanocrystals (CNC) or sodium montmorillonite (Na-MMT). In particular, the focus is on how the nanoparticle shape and surface properties influence the nano- to micrometer length scale nanofiller arrangement in the nanocomposites and how the arrangements influence the mechanisms involved in the inhibition of the amorphous to crystalline transition. The nanocomposites were produced by co-spray-drying. Solid-state transformations were analyzed at 60-94% relative humidity using X-ray powder diffraction, microcalorimetry, and light microscopy. The recrystallization rate constant for the lactose/CNC and lactose/Na-MMT nanocomposites was lowered at nanofiller contents higher than 60% and were stable for months at 80% nanofiller. The most likely explanation to these results is spontaneous formations of mesoporous particle networks that the lactose is confined within upon co-spray-drying at high filler content. Compartmentalization and rigidification of the amorphous lactose proved to be less important mechanisms involved in the stabilization of lactose in the nanocomposites.

    Nationell ämneskategori
    Materialkemi
    Identifikatorer
    urn:nbn:se:uu:diva-300153 (URN)10.1016/j.xphs.2016.09.032 (DOI)
    Tillgänglig från: 2016-08-03 Skapad: 2016-08-03 Senast uppdaterad: 2016-12-07
    5. Humidity sorption of lactose/sodium montmorillonite nanocomposites
    Öppna denna publikation i ny flik eller fönster >>Humidity sorption of lactose/sodium montmorillonite nanocomposites
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    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Understanding of water sorption is of high importance in materials science as water may change materials properties promoting degradation, relaxations, or recrystallization. In this study, we investigated the humidity sorption in co-spray-dried lactose/sodium montmorillonite nanocomposites with varying lactose loading with the aim to increase the knowledge of the water sorption in this type of materials. It was demonstrated that the intercalation of lactose in the Na‑MMT clay decreased hygroscopicity of the composite despite high water affinity of both materials. As the cations in interlayer space of montmorillonite play an essential role in water sorption in the clay, we gained the molecular level understanding of Na+ interactions with lactose molecules and clay surface in the nanocomposites with molecular dynamic simulations and 23Na solid-state NMR. In conclusion, we demonstrated that the decreased hygroscopicity of the materials can be explained by interactions of lactose with the Na+ and the clay surfaces in the MMT interlayer space of lactose/Na-MMT nanocomposites.

    Nationell ämneskategori
    Materialkemi
    Identifikatorer
    urn:nbn:se:uu:diva-300154 (URN)
    Tillgänglig från: 2016-08-03 Skapad: 2016-08-03 Senast uppdaterad: 2016-09-01
    6. Powder compression mechanics of spray-dried lactose nanocomposites
    Öppna denna publikation i ny flik eller fönster >>Powder compression mechanics of spray-dried lactose nanocomposites
    (Engelska)Ingår i: International Journal of Pharmaceutics, ISSN 0378-5173, E-ISSN 1873-3476Artikel i tidskrift (Övrigt vetenskapligt) Submitted
    Abstract [en]

    The aim of this study was to investigate the structural impact of the nanofiller incorporation on the powder compression mechanics of spray-dried lactose. The lactose was co-spray-dried with three different nanofillers, that is, cellulose nanocrystals, sodium montmorillonite, and fumed silica, which led to lower micron sized nanocomposite particles with varying structure and morphology. The powder compression mechanics of the nanocomposites and physical mixtures of the neat spray-dried components were evaluated by a rational evaluation method with compression analysis as a tool using the Kawakita equation and the Shapiro-Konopicky-Heckel equation. Particle rearrangement dominated the initial compression profiles due to the small particle sizes of the materials. The strong contribution of particle rearrangement in the materials with fumed silica continued throughout the whole compression profile, which prohibited an in-depth material characterization. However, the lactose/cellulose nanocrystals and the lactose/sodium montmorillonite nanocomposites demonstrated increased yield pressure compared with the physical mixtures indicating increased particle hardness. This increase has likely to do with a reinforcement of the nanocomposite particles by skeleton formation of the nanoparticles. In summary, the rational evaluation applying compression analysis proved to be a valuable tool for mechanical evaluation for this type of materials unless they demonstrate particle rearrangement throughout the whole compression profile.

    Nationell ämneskategori
    Materialkemi
    Identifikatorer
    urn:nbn:se:uu:diva-300156 (URN)
    Tillgänglig från: 2016-08-05 Skapad: 2016-08-03 Senast uppdaterad: 2016-12-07
  • 3.
    Singh, Shalini
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Amphiphilic Peptide Interactions with Complex Biological Membranes: Effect of peptide properties on antimicrobial and anti-inflammatory effects2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    With increasing problem of resistance development in bacteria against conventional antibiotics, as well as problems associated with diseases either triggered or enhanced by infection, there is an urgent need to identify new types of effective therapeutics for the treatment of infectious diseases and its consequences. Antimicrobial and anti-inflammatory peptides have attracted considerable interest as potential new antibiotics in this context. While antimicrobial function of such peptides is being increasingly understood demonstrated to be due to bacterial membrane disruption, the mechanisms of their anti-inflammatory function are poorly understood. Since bacterial membrane component lipopolysaccharide triggers inflammation, this thesis aims at clarifying importance of lipopolysaccharide (LPS)-peptide interactions while investigating possible modes of action of peptides exhibiting anti-inflammatory effect. Furthermore, effect of poly(ethylene)glycol (PEG)-conjugation was investigated to increase performance of such peptides.

    Results presented in this thesis demonstrate that peptide-induced LPS- and lipid A binding/scavenging is necessary but not sufficient criterium for anti-inflammatory effects of peptides. Furthermore, preferential binding to LPS over lipid membrane, as well as higher binding affinity to the lipid A moiety within LPS, are seen for these peptides. In addition, results demonstrate that apart from direct LPS scavenging, membrane-localized peptide-induced LPS scavenging seem to contribute partially to anti-inflammatory effect. Furthermore, fragmentation and densification of LPS aggregates, in turn dependent on the peptide secondary structure on LPS binding, as well as aromatic packing interactions, correlate to the anti-inflammatory effect, thus promoting peptide-induced packing transition in LPS aggregates as key for anti-inflammatory functionality. Thus, peptide-induced LPS aggregate disruption together with reduction of the negative charge of LPS suggests the importance of phagocytosis as an alternative to the inflammatory pathway, which needs to be further investigated. Furthermore, PEG conjugation of peptide results in strongly reduced toxicity at a cost of reduced antimicrobial activity but markedly retained anti-inflammatory effect.

    Taken together, the results obtained in this work have demonstrated several key issues which need to be taken into consideration in the development of effective and selective anti-inflammatory peptide therapeutics for the treatment of severe Gram-negative bacterial infections.

    Delarbeten
    1. Membrane and lipopolysaccharide interactions of C-terminal peptides from S1 peptidases
    Öppna denna publikation i ny flik eller fönster >>Membrane and lipopolysaccharide interactions of C-terminal peptides from S1 peptidases
    2012 (Engelska)Ingår i: Biochimica et Biophysica Acta - Biomembranes, ISSN 0005-2736, E-ISSN 1879-2642, Vol. 1818, nr 9, 2244-2251 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The mechanisms underlying antimicrobial and anti-endotoxic effects were investigated for a series of structurally related peptides derived from the C-terminal region of S1 peptidases. For this purpose, results on bacterial killing were compared to those on peptide-induced liposome leakage, and to ellipsometry and dual polarization interferometry results on peptide binding to, and disordering of, supported lipid bilayers. Furthermore, the ability of these peptides to block endotoxic effects caused by bacterial lipopolysaccharide (LPS), monitored through NO production in macrophages, was compared to the binding of these peptides to LPS, and to secondary structure formation in the peptide/LPS complex. Bacteria killing, occurring through peptide-induced membrane lysis, was found to correlate with liposome rupture, and with the extent of peptide binding to the lipid membrane, no adsorption threshold for peptide insertion being observed. Membrane and LPS binding was found to depend on peptide net charge, illustrated by LPS binding increasing with increasing peptide charge, and peptides with net negative charge being unable to lyse membranes, kill bacteria, and block LPS-induced endotoxic effect. These effects were, however, also influenced by peptide hydrophobicity. LPS binding was furthermore demonstrated to be necessary, but not sufficient, for anti-endotoxic effect of these peptides. Circular dichroism spectroscopy showed that pronounced helix formation occurs in peptide/LPS complexes for all peptides displaying anti-endotoxic effect, hence potentially linked to this functionality. Similarly, ordered secondary structure formation was correlated to membrane binding, lysis, and antimicrobial activity of these peptides. Finally, preferential binding of these peptides to LPS over the lipid membrane was demonstrated.

    Nyckelord
    Antimicrobial peptide, Dual polarization interferometry, Ellipsometry, Lipopolysaccharide, Liposome, Membrane
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-180268 (URN)10.1016/j.bbamem.2012.03.017 (DOI)000306882600019 ()
    Tillgänglig från: 2012-09-03 Skapad: 2012-09-03 Senast uppdaterad: 2016-06-01Bibliografiskt granskad
    2. Lipopolysaccharide Interactions of C-Terminal Peptides from Human Thrombin
    Öppna denna publikation i ny flik eller fönster >>Lipopolysaccharide Interactions of C-Terminal Peptides from Human Thrombin
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    2013 (Engelska)Ingår i: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 14, nr 5, 1482-1492 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Interactions with bacterial lipopolysaccharide (LPS), both in aqueous solution and in lipid membranes, were investigated for a series of amphiphilic peptides derived from the C-terminal region of human thrombin, using ellipsometry, dual polarization interferometry, fluorescence spectroscopy, circular dichroism (CD), dynamic light scattering, and z-potential measurements. The ability of these peptides to block endotoxic effects caused by LPS, monitored through NO production in macrophages, was compared to peptide binding to LPS and its endotoxic component lipid A, and to size, charge, and secondary structure of peptide/LPS complexes. While the antiendotoxic peptide GKY25 (GKYGFYTHVFRL-KKWIQKVIDQFGE) displayed significant binding to both LPS and lipid A, so did two control peptides with either selected D-amino acid substitutions or with maintained composition but scrambled sequence, both displaying strongly attenuated antiendotoxic effects. Hence, the extent of LPS or lipid A binding is not the sole discriminant for the antiendotoxic effect of these peptides. In contrast, helix formation in peptide/LPS complexes correlates to the antiendotoxic effect of these peptides and is potentially linked to this functionality. Preferential binding to LPS over lipid membrane was furthermore demonstrated for these peptides and preferential binding to the lipid A moiety within LPS inferred.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-202380 (URN)10.1021/bm400150c (DOI)000319034600027 ()
    Tillgänglig från: 2013-06-24 Skapad: 2013-06-24 Senast uppdaterad: 2016-06-01Bibliografiskt granskad
    3. Effects of linear amphiphilicity on membrane interactions of C-terminal thrombin peptides
    Öppna denna publikation i ny flik eller fönster >>Effects of linear amphiphilicity on membrane interactions of C-terminal thrombin peptides
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    2014 (Engelska)Ingår i: RSC Advances, ISSN 2046-2069, Vol. 4, nr 71, 37582-37591 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Effects of linear amphiphilicity on membrane interactions of antimicrobial peptides were investigated by ellipsometry, dual polarization interferometry, fluorescence spectroscopy, light scattering, and circular dichroism. In doing so, the thrombin-derived GKY25 (GKYGFYTHVFRLKKWIQKVIDQFGE) was compared to WFF25 (WFFFYYLIIGGGVVTHQQRKKKKDE) of identical composition, but with amino acids sorted according to hydrophobicity, the latter peptide thus displaying pronounced linear amphiphilicity. In addition, GKY25d (GKYG(f) YTH(v) FRL(k) KWI(q) KVI(d) QFGE; with an identical sequence but with selected D-amino acid substitutions) was included as a control peptide, for which conformationally induced (helix-related) amphiphilicity was suppressed. Through its pronounced linear amphiphilicity, WFF25, but not the less amphiphilic GKY25 and GKY25d, forms aggregates in solution. Through its terminal W/F stretch, WFF25 also displays pronounced selectivity, with higher membrane binding and liposome rupture than GKY25 and GKY25d for anionic membranes, but suppressed peptide insertion and lytic effects for zwitterionic ones. In addition, WFF25 binds extensively to anionic polyelectrolyte components in bacterial membranes, i.e., lipopolysaccharide and lipoteichoic acid, resulting in reduced antimicrobial effects through peptide scavenging, not seen for the less amphiphilic GKY25 and GKY25d peptides. Taken together, the results thus demonstrate a series of striking effects for highly amphiphilic peptides, which need to be recognized in the development of such compounds as potential peptide therapeutics.

    Nationell ämneskategori
    Klinisk medicin Kemi
    Identifikatorer
    urn:nbn:se:uu:diva-233621 (URN)10.1039/c4ra05420b (DOI)000341454600018 ()
    Tillgänglig från: 2014-10-07 Skapad: 2014-10-07 Senast uppdaterad: 2016-06-01Bibliografiskt granskad
    4. Importance of lipopolysaccharide aggregate disruption for the anti-endotoxic effects of heparin cofactor II peptides
    Öppna denna publikation i ny flik eller fönster >>Importance of lipopolysaccharide aggregate disruption for the anti-endotoxic effects of heparin cofactor II peptides
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    2013 (Engelska)Ingår i: Biochimica et Biophysica Acta - Biomembranes, ISSN 0005-2736, E-ISSN 1879-2642, Vol. 1828, nr 11, 2709-2719 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Lipid membrane and lipopolysaccharide (LPS) interactions were investigated for a series of amphiphilic and cationic peptides derived from human heparin cofactor II (HCII), using dual polarization interferometry, ellipsometry, circular dichroism (CD), cryoTEM, and z-potential measurements. Antimicrobial effects of these peptides were compared to their ability to disorder bacterial lipid membranes, while their capacity to block endotoxic effects of LPS was correlated to the binding of these peptides to LPS and its lipid A moiety, and to charge, secondary structure, and morphology of peptide/LPS complexes. While the peptide KYE28 (KYEITTIHNLERKLTHRLFRRNEGYTLR) displayed potent antimicrobial and anti-endotoxic effects, its truncated variants KYE21 (KYEITTIHNLFRKLTHRLFRR) and NLF20 (NLFRKLTHRLFRRNFGYTLR) provide some clues on structure-activity relations, since KYE21 retains both the antimicrobial and anti-endotoxic effects of KYE28 (although both attenuated), while NLF20 retains the antimicrobial but only a fraction of the anti-endotoxic effect, hence locating the anti-endotoxic effects of KYE28 to its N-terminus. The antimicrobial effect, on the other hand, is primarily located at the C-terminus of KYE28. While displaying quite different endotoxic effects, these peptides bind to a similar extent to both LPS and lipid A, and also induce comparable LPS scavenging on model eukaryotic membranes. In contrast, fragmentation and densification of LPS aggregates, in turn dependent on the secondary structure in the peptide/LPS aggregates, correlate to the anti-endotoxic effect of these peptides, thus identifying peptide-induced packing transitions in LPS aggregates as key for anti-endotoxic functionality. This aspect therefore needs to be taken into account in the development of novel anti-endotoxic peptide therapeutics. 

    Nyckelord
    Antimicrobial peptide, Dual polarization interferometry, Ellipsometry, Lipopolysaccharide, Liposome, Membrane
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-211434 (URN)10.1016/j.bbamem.2013.06.015 (DOI)000326143200038 ()
    Tillgänglig från: 2013-11-27 Skapad: 2013-11-25 Senast uppdaterad: 2016-06-01Bibliografiskt granskad
    5. Effects of PEGylation on Membrane and Lipopolysaccharide Interactions of Host Defense Peptides
    Öppna denna publikation i ny flik eller fönster >>Effects of PEGylation on Membrane and Lipopolysaccharide Interactions of Host Defense Peptides
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    2014 (Engelska)Ingår i: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 15, nr 4, 1337-1345 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Effects of poly(ethylene glycol) (PEG) conjugation on peptide interactions with lipid membranes and lipopolysaccharide (LPS) were investigated for KYE28 (KYEITTIHNLFRKLTHRLFRRNFGYTLR), an antimicrobial and anti-inflammatory peptide derived from human heparin cofactor II. In particular, effects of PEG length and localization was investigated by ellipsometry, circular dichroism, nanoparticle tracking analysis, and fluorescence/electron microscopy. PEGylation of KYE28 reduces peptide binding to lipid membranes, an effect accentuated at increasing PEG length, but less sensitive to conjugation site. The reduced binding causes suppressed liposome leakage induction, as well as bacterial lysis. As a result of this, the antimicrobial effects of KYE28 is partially lost with increasing PEG length, but hemolysis also strongly suppressed and selecticity improved. Through this, conditions can be found, at which the PEGylated peptide displays simultaneously efficient antimicrobial affects and low hemolysis in blood. Importantly, PEGylation does not markedly affect the anti-inflammatory effects of KYE28. The combination of reduced toxicity, increased selectivity, and retained anti-inflammatory effect after PEGylation, as well as reduced scavenging by serum proteins, thus shows that PEG conjugation may offer opportunities in the development of effective and selective anti-inflammatory peptides.

    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-225069 (URN)10.1021/bm401884e (DOI)000334571600026 ()
    Tillgänglig från: 2014-06-23 Skapad: 2014-05-27 Senast uppdaterad: 2016-06-01Bibliografiskt granskad
    6. Role of Aromatic Amino Acids in Lipopolysaccharide and Membrane Interactions of Antimicrobial Peptides for use in Plant Disease Control
    Öppna denna publikation i ny flik eller fönster >>Role of Aromatic Amino Acids in Lipopolysaccharide and Membrane Interactions of Antimicrobial Peptides for use in Plant Disease Control
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    2016 (Engelska)Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 291, nr 25, 13301-13317 s.Artikel i tidskrift, Meeting abstract (Refereegranskat) Published
    Abstract [en]

    KYE28(KYEITTIHNLFRKLTHRLFRRNFGYTLR), the representative sequence  of helix D of heparin co-factor II, was demonstrated to be potent against agronomically important Gram-negative plant pathogens X. vesicatoria and X. oryzae,capable of inhibiting disease symptoms in detached tomato leaves. NMR studies in presence of lipopolysaccharide provided structural insights into the mechanisms underlying this, notably in relation to outer membrane permeabilisation. The three-dimensional solution structure of KYE28 in LPS is characterised by a N-ter helical segment, an intermediate loop and an extended C-ter. The two termini are in close proximity to each other via aromatic packing interactions, while the positively charged residues formed an exterior polar shell. To further demonstrate the importance of the aromatic residues for this, a mutant peptide KYE28A, with Ala substitutions at F11, F19, F23 and Y25 showed attenuated antimicrobial activity at high salt concentrations, as well as lower membrane disruption and LPS binding abilities compared to KYE28. In contrast to KYE28, KYE28A adopted an opened out helical structure in LPS with extended N- and C-ter and a small break in between the helical segments. Aromatic packing interactions were completely lost, although hydrophobic interaction between the side chains of hydrophobic residues were still partly retained, imparting an amphipathic character and explaining its residual antimicrobial activity and LPS binding as observed from ellipsometry and ITC. We thus present important structural aspects of KYE28, constituting an aromatic zipper, of potential importance, for the development of novel plant protection agents and therapeutic agents.

    Nyckelord
    LPS, Antimicrobial
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Forskningsämne
    Farmaceutisk fysikalisk kemi
    Identifikatorer
    urn:nbn:se:uu:diva-282778 (URN)10.1074/jbc.M116.719575 (DOI)000379770500033 ()27137928 (PubMedID)
    Tillgänglig från: 2016-04-06 Skapad: 2016-04-06 Senast uppdaterad: 2016-08-18Bibliografiskt granskad
  • 4.
    Heidarian Höckerfelt, Mina
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    On the chemical and processing stability of pharmaceutical solids: Solid form dependent water presenting capacity and process induced solid form transformation2015Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    There is a need for improving our knowledge and understanding about formation mechanisms and nature of amorphous state in order to prevent the unintended presence of disorder in solid pharmaceutical products and reduce the related stability issues. The suggested theory that water binding capacity of amorphous cellulose affects the chemical stability of hydrolysis sensitive drugs in formulations with cellulose based excipients needs a clarification and water-cellulose interaction profiles need to be examined.  This thesis has addressed these questions.

    Chemical, mechanical and thermal methods have been used to create partially or predominantly amorphous solids. Mechanisms and the pathways for transformation to amorphous phase and the characteristic qualities of this phase is studied in order to give some tools to predict, to control or prevent the creation of disorder in a crystalline structure. The water interaction with amorphous pharmaceutical materials has been studied to improve stability of hydrolysis sensitive drugs. 

     

    The transition to amorphous state during handling of pharmaceutical material, referred to as mechanical activation in processes like blending, mixing and compression is substantially a consequence of vitrification. The process is described as creation of hot spots where friction caused by particle sliding raise the temperature above the melting point of the material. The fast cooling process promotes creation of a local disordered molecular arrangement. It is possible to decrease the degree of amorphisation and undesired stability problems by reducing the friction and inhibit the creation of crystal defects during processing.

     

    The glass-forming propensity is an inherent material characteristic related to molecular size and structure and is not process dependent. Molecules with a couple of aromatic rings are often poor glass-formers. Less symmetrical, branched molecular structures with presence of electronegative atoms are more readily transformed to and exist in amorphous state when handled and stored at temperatures below their glass transition temperature.

     

    The interaction profile of cellulose with water is strongly dependent on solid state structure of cellulose. Crystallinity is the key parameter in water presenting capacity of cellulose. Amorphous regions have a capacity to bind the water and decrease water mobility and in that way reduce cellulose water presenting capacity despite higher moisture content in partially amorphous cellulose compared to crystalline cellulose. The fact that higher amorphous content decreases cellulose water presenting capacity is a promising lead to improve stability of hydrolysis sensitive drugs in compositions with cellulose. This knowledge could be applicable to other pharmaceutical materials as the differences between crystalline and amorphous states of material are generally the same for different kind of materials.

    Delarbeten
    1. Dry mixing transformed micro-particles of a drug from a highly crystalline to a highly amorphous state
    Öppna denna publikation i ny flik eller fönster >>Dry mixing transformed micro-particles of a drug from a highly crystalline to a highly amorphous state
    2009 (Engelska)Ingår i: Pharmaceutical development and technology (Print), ISSN 1083-7450, E-ISSN 1097-9867, Vol. 14, nr 3, 233-239 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    In this paper, the degree of mechanical activation of particles due to mechanical straining without subsequent breakage has been studied. Griseofulvin micro-particles of about 2 mum in size were mixed with glass beads (proportion 1:99) in a tumbling mixer. After a series of mixing times, ranging from 2-96 hours, samples were withdrawn and the particle size and the degree of crystallinity were assessed. The mixing process gave no detectable change in particle size. The degree of disorder of the drug particles increased with mixing time and highly amorphous particles were obtained after about 24 h of mixing. The results thus indicate that particles can be completely activated by mechanical treatment without a parallel size reduction of the particles. It is suggested that the activation is caused by repeated deformation of the particles, gradually transforming the crystalline state into an amorphous state.

    Nyckelord
    Mixing, activation, crystallinity, solubility
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-109431 (URN)10.1080/10837450802585252 (DOI)000268243600001 ()19519178 (PubMedID)
    Tillgänglig från: 2009-10-15 Skapad: 2009-10-15 Senast uppdaterad: 2015-11-26Bibliografiskt granskad
    2. Toward In Silico Prediction of Glass-Forming Ability from Molecular Structure Alone: A Screening Tool in Early Drug Development
    Öppna denna publikation i ny flik eller fönster >>Toward In Silico Prediction of Glass-Forming Ability from Molecular Structure Alone: A Screening Tool in Early Drug Development
    2011 (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, Vol. 8, nr 2, 498-506 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    We present a novel computational tool which predicts the glass-forming ability of drug compounds solely from their molecular structure. Compounds which show solid-state limited aqueous solubility were selected, and their glass-forming ability was determined upon spray-drying, melt-quenching and mechanical activation. The solids produced were analyzed by differential scanning calorimetry (DSC) and powder X-ray diffraction. Compounds becoming at least partially amorphous on processing were classified as glass-formers, whereas those remaining crystalline regardless of the process method were classified as non-glass-forming compounds. A predictive model of the glass-forming ability, designed to separate between these two classes, was developed through the use of partial least-squares projection to latent structure discriminant analysis (PLS-DA) and calculated molecular descriptors. In total, ten of the 16 compounds were determined experimentally to be good glass-formers and the PLS-DA model correctly sorted 15 of the compounds using four molecular descriptors only. An external test set was predicted with an accuracy of 75%, and, hence, the PLS-DA model developed was shown to be applicable for the identification of compounds that have the potential to be designed as amorphous formulations. The model suggests that larger molecules with a low number of benzene rings, low level of molecular symmetry, branched carbon skeletons and electronegative atoms have the ability to form a glass. To conclude, we have developed a predictive, transparent and interpretable computational model for the identification of drug molecules capable of being glass-formers. The model allows an assessment of amorphization as a formulation strategy in the early drug development process, and can be applied before compound synthesis.

    Nyckelord
    glass-forming ability, prediction, in silico models, molecular descriptors, amorphous
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-151964 (URN)10.1021/mp100339c (DOI)000289008600019 ()21344945 (PubMedID)
    Tillgänglig från: 2011-04-29 Skapad: 2011-04-20 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    3. Mechanism of Amorphisation of Micro-Particles of Griseofulvin During Powder Flow in a Mixer
    Öppna denna publikation i ny flik eller fönster >>Mechanism of Amorphisation of Micro-Particles of Griseofulvin During Powder Flow in a Mixer
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    2013 (Engelska)Ingår i: Journal of Pharmaceutical Sciences, ISSN 0022-3549, E-ISSN 1520-6017, Vol. 102, nr 11, 4036-4045 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The purpose of the research was to investigate the degree of solid-state amorphisation during powder flow and to propose a mechanism for this transformation. Micro-particles of griseofulvin (about 2m in diameter) were mixed in a shear mixer under different conditions to influence the inter-particulate collisions during flow, and the degree of amorphisation was determined by micro-calorimeter. The amorphisation of griseofulvin particles (GPs)during repeated compaction was also determined. The GPs generally became disordered during mixing in a range from about 6% to about 86%. The degree of amorphisation increased with increased mixing time and increased batch size of the mixer, whereas the addition of a lubricant to the blend reduced the degree of amorphisation. Repeated compaction using the press with ejection mode gave limited amorphisation, whereas repeated compaction without an ejection process gave minute amorphisation. It is concluded that during powder flow, the most important inter-particulate contact process that cause the transformation of a crystalline solid into an amorphous state is sliding. On the molecular scale, this amorphisation is proposed to be caused by vitrification, that is the melting of a solid because of the generation of heat during sliding followed by solidification into an amorphous phase.

    Nyckelord
    powder technology, powder flow, compaction, mixing, amorphisation, mechanical activation, vitrification, crystal defect, friction, sliding
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-210559 (URN)10.1002/jps.23713 (DOI)000325550400019 ()
    Tillgänglig från: 2013-11-13 Skapad: 2013-11-11 Senast uppdaterad: 2017-03-19Bibliografiskt granskad
    4. Influence of water-cellulose binding energy on stability of acetylsalicylic acid
    Öppna denna publikation i ny flik eller fönster >>Influence of water-cellulose binding energy on stability of acetylsalicylic acid
    2006 (Engelska)Ingår i: International Journal of Pharmaceutics, ISSN 0378-5173, E-ISSN 1873-3476, Vol. 323, nr 1-2, 139-145 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The aim of the present study was to investigate how the energies of water binding in cellulose tabletting excipients influence the availability of moisture to induce hydrolysis of acetylsalisylic acid (ASA). Cellulose powders of varying degree of order, denoted as low-crystallinity cellulose (LCC) and high-crystallinity cellulose (HCC), were produced by treating ordinary microcrystalline cellulose (MCC) in ZnCl2 solutions of varying concentrations. Microcrystalline cellulose (MCC) and lactose monohydrate were used as reference excipients. The samples were then studied by X-ray diffraction, scanning electron microscopy, and differential scanning calorimetry (DSC). Different ratios of each excipient mixed with ASA were stored at 40% RH and 50 degrees C for 35 days to investigate the hydrolytic stability of the mixtures. Stability studies indicated that as concentration of HCC and MCC in binary mixtures with ASA was raised from 1 to 50% (w/w), ASA became increasingly unstable with respect to hydrolysis. Although LCC contained more moisture than the other celluloses, no such trend was observed in the LCC and lactose samples. DSC analysis revealed that each water molecule on the average was bound by more than three hydrogen bonds in the LCC and lactose structures and therefore remained predominantly unavailable to induce hydrolysis. The current study elucidates the necessity of evaluating the energy of water bindings in a pharmaceutical excipient when predicting the excipient's performance in mixtures comprising moisture-sensitive drugs.

    Nyckelord
    microcrystalline cellulose, lactose, crystallinity, moisture content, water interactions, stability of moisture-sensitive drug
    Nationell ämneskategori
    Farmaceutisk vetenskap Teknik och teknologier
    Identifikatorer
    urn:nbn:se:uu:diva-83335 (URN)10.1016/j.ijpharm.2006.05.058 (DOI)000241346400018 ()16854539 (PubMedID)
    Tillgänglig från: 2007-01-17 Skapad: 2007-01-17 Senast uppdaterad: 2016-11-30Bibliografiskt granskad
    5. The crystallinity of cellulose controls the physical distribution of sorbed water and the capacity to present water for chemical degradation of a solid drug
    Öppna denna publikation i ny flik eller fönster >>The crystallinity of cellulose controls the physical distribution of sorbed water and the capacity to present water for chemical degradation of a solid drug
    2014 (Engelska)Ingår i: International Journal of Pharmaceutics, ISSN 0378-5173, E-ISSN 1873-3476, Vol. 477, nr 1-2, 326-333 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The purpose of the research was to investigate the effect of moisture content of cellulose on the degradation of a drug in binary mixtures with cellulose. Physical mixtures of acetylsalicylic acid and two forms of cellulose, either microcrystalline cellulose or low crystalline cellulose, in the proportion 1:1 were stored at 50°C at a series of relative humidities (0-90%) for up to 175 days. The degradation rate constant of the drug increased with increased cellulose moisture content in a bi-regional fashion, with a low and a high degradation rate region. The shift from region 1 to 2 occurred at higher moisture content for the low crystalline cellulose. The relationships between rate constant and the temperature of maximum endothermic value overlapped for the two celluloses. It is proposed that the amount of water available for degradation of a solid drug is controlled by the water presenting capacity of cellulose which is dependent of the mechanism of sorption of water in cellulose. The water sorption of water can for cellulose satisfactorily be described by a two-site residence model with cellulose crystallinity as the structural correlate to the distribution between the two residence sites.

    Nationell ämneskategori
    Farmakologi och toxikologi
    Identifikatorer
    urn:nbn:se:uu:diva-246090 (URN)10.1016/j.ijpharm.2014.10.034 (DOI)000347623800036 ()25455777 (PubMedID)
    Tillgänglig från: 2015-03-02 Skapad: 2015-03-02 Senast uppdaterad: 2015-11-26Bibliografiskt granskad
  • 5.
    Lilienberg, Elsa
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Biopharmaceutical Evaluation of Intra-arterial Drug-Delivery Systems for Liver Cancer: Investigations in healthy pigs and liver cancer patients2015Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    There are currently two types of intra-arterial drug-delivery system (DDS) in clinical use in the palliative treatment of primary liver cancer. The chemotherapeutic drug doxorubicin (DOX) can be formulated into a drug-in-lipiodol emulsion (LIPDOX) or a microparticulate drug-eluting bead system (DEBDOX). To facilitate development of future DDSs, we need to understand the release and local distribution of drug from these DDSs into the complex, in vivo, pathological environment.

    The overall aim of this project was to assess and improve understanding of the in vivo release of DOX from LIPDOX and DEBDOX and its local disposition in the liver. These processes were investigated in detail in a multisampling-site, healthy pig model and in human patients with liver cancer. The mechanisms involved in DOX disposition were studied by examining potential interactions between DOX and lipiodol and/or cyclosporine A (CsA) in pigs.  

    In this project, the main elimination pathway for DOX and its primary metabolite doxorubicinol (DOXol) was via bile; their extensive canalicular carrier-mediated transport (e.g. ATP-binding cassette transporters ABCB1, ABCC1, ABCC2 and ABCG2) was inhibited by CsA. CsA had no effect on the carbonyl and aldo-keto reductases responsible for the metabolism of DOX into DOXol. LIPDOX released DOX more rapidly and to a greater extent into the circulation than DEBDOX, which had only released 15% of the dose in patients after 24 hrs. The systemic exposure to DOX was lower for DEBDOX than for LIPDOX. Greater fractions of DOXol were formed in blood and bile with LIPDOX than with DEBDOX. This may have been because DOX was more widely distributed into regions with increased metabolic capacity or because of increased intracellular uptake when DOX was delivered in LIPDOX. The excipient lipiodol in the LIPDOX formulation did not interact with transporters, enzymes or membranes that would explain the increased cellular uptake of DOX.

    In conclusion, the release of DOX from DEBDOX is more controlled in vivo than that from LIPDOX, indicating that DEBDOX is a more robust pharmaceutical product. The formulations for future optimized DDSs should therefore be more similar to DEBDOX than to LIPDOX. 

    Delarbeten
    1. Investigation of Hepatobiliary Disposition of Doxorubicin Following Intrahepatic Delivery of Different Dosage Forms
    Öppna denna publikation i ny flik eller fönster >>Investigation of Hepatobiliary Disposition of Doxorubicin Following Intrahepatic Delivery of Different Dosage Forms
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    2014 (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, Vol. 11, nr 1, 131-144 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Unresectable, intermediate stage hepatocellular carcinoma (HCC) is often treated palliatively in humans by doxorubicin (DOX). The drug is administered either as a drug-emulsified-in-Lipiodol (DLIP) or as drug loaded into drug eluting beads (DEB), and both formulations are administered intrahepatically. However, several aspects of their in vivo performance in the liver are still not well-understood. In this study, DLIP and DEB were investigated regarding the local and systemic pharmacokinetics (PK) of DOX and its primary metabolite doxorubicinol (DOXol). An advanced PK-multisampling site acute in vivo pig model was used for simultaneous sampling in the portal, hepatic, and femoral veins and the bile duct. The study had a randomized, parallel design with four treatment groups (TI–TIV). TI (n = 4) was used as control and received an intravenous (i.v.) infusion of DOX as a solution. TII and TIII were given a local injection in the hepatic artery with DLIP (n = 4) or DEB (n = 4), respectively. TIV (n = 2) received local injections of DLIP in the hepatic artery and bile duct simultaneously. All samples were analyzed for concentrations of DOX and DOXol with UPLC-MS/MS. Compared to DLIP, the systemic exposure for DOX with DEB was reduced (p < 0.05), in agreement with a slower in vivo release. The approximated intracellular bioavailability of DOX during 6 h appeared to be lower for DEB than DLIP. Following i.v. infusion (55 min), DOX had a liver extraction of 41 (28–53)%, and the fraction of the dose eliminated in bile of DOX and DOXol was 20 (15–22)% and 4.2 (3.2–5.2)%, respectively. The AUCbile/AUCVP for DOX and DOXol was 640 (580–660) and 5000 (3900–5400), respectively. In conclusion, DLIP might initially deliver a higher hepatocellular concentration of DOX than DEB as a consequence of its higher in vivo release rate. Thus, DLIP delivery results in higher intracellular peak concentrations that might correlate with better anticancer effects, but also higher systemic drug exposure and safety issues.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-210381 (URN)10.1021/mp4002574 (DOI)000329529700012 ()24171458 (PubMedID)
    Tillgänglig från: 2013-11-06 Skapad: 2013-11-06 Senast uppdaterad: 2016-01-13Bibliografiskt granskad
    2. The Effects of Lipiodol and Cyclosporin A on the Hepatobiliary Disposition of Doxorubicin in Pigs
    Öppna denna publikation i ny flik eller fönster >>The Effects of Lipiodol and Cyclosporin A on the Hepatobiliary Disposition of Doxorubicin in Pigs
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    2014 (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, Vol. 11, nr 4, 1301-1313 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Doxorubicin (DOX) emulsified in Lipiodol (LIP) is used as local palliative treatment for unresectable intermediate stage hepatocellular carcinoma. The objective of this study was to examine the poorly understood effects of the main excipient in the drug delivery system, LIP, alone or together with cyclosporin A (CsA), on the in vivo liver disposition of DOX. The advanced, multi-sampling-site, acute pig model was used; samples were collected from three blood vessels (v. portae, v. hepatica and v. femoralis), bile and urine. The four treatment groups (TI-TIV) all received two intravenous 5 min infusions of DOX into an ear vein: at 0 and 200 min. Before the second dose, the pigs received a portal vein infusion of saline (TI), LIP (TII), CsA (TIII) or LIP and CsA (TIV). Concentrations of DOX and its active metabolite doxorubicinol (DOXol) were analyzed using UPLC-MS/MS. A multi-compartment model was developed to describe the distribution of DOX and DOXol in plasma, bile and urine. LIP did not affect the pharmacokinetics of DOX or DOXol. CsA (TIII and TIV) had no effect on the plasma pharmacokinetics of DOX, but a 2-fold increase in exposure to DOXol and a significant decrease in hepatobiliary clearance of DOX and DOXol was observed. Model simulations supported that CsA inhibits 99% of canalicular biliary secretion of both DOX and DOXol, but does not affect the metabolism of DOX to DOXol. In conclusion, LIP did not interact with transporters, enzymes and/or biological membranes important for the hepatobiliary disposition of DOX.

    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-222282 (URN)10.1021/mp4007612 (DOI)000334092700022 ()24558959 (PubMedID)
    Tillgänglig från: 2014-04-09 Skapad: 2014-04-09 Senast uppdaterad: 2016-01-13Bibliografiskt granskad
    3. Lipiodol does not affect the tissue distribution of intravenous doxorubicin infusion in pigs
    Öppna denna publikation i ny flik eller fönster >>Lipiodol does not affect the tissue distribution of intravenous doxorubicin infusion in pigs
    (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, E-ISSN 1543-8392Artikel i tidskrift (Övrigt vetenskapligt) Submitted
    Ort, förlag, år, upplaga, sidor
    Uppsala:
    Nyckelord
    doxorubicin, doxorubicinol, lipiodol, tissue distribution, Kp, cyclosporine A
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-267119 (URN)
    Tillgänglig från: 2015-11-21 Skapad: 2015-11-18 Senast uppdaterad: 2016-01-13
    4. Comparison of drug release, pharmacokinetics and pharmacodynamics after lipiodol-based emulsion or drug-eluting bead delivery to patients with hepatocellular carcinoma
    Öppna denna publikation i ny flik eller fönster >>Comparison of drug release, pharmacokinetics and pharmacodynamics after lipiodol-based emulsion or drug-eluting bead delivery to patients with hepatocellular carcinoma
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    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nyckelord
    Doxorubicin, doxorubicinol, drug eluting beads, focal delivery, focal therapy, hepatocellular carcinoma, liver cancer, lipiodol, transarterial chemoembolization, transarterial infusion chemotherapy
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-267395 (URN)
    Forskningsfinansiär
    Vetenskapsrådet, 521-2011-373
    Tillgänglig från: 2015-11-21 Skapad: 2015-11-21 Senast uppdaterad: 2016-01-13
  • 6.
    Widenbring, Ronja
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Microgel Interactions with Peptides and Proteins: Consequence of Peptide and Microgel Properties2015Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Microgels are lightly cross-linked hydrogel particles in the sub-micrometer to micrometer size range with a capacity to drastically change their volume in response to changes in the external environment. Microgels have an ability to bind and store substances such as biomacromolecular drugs, notably proteins and peptides, and release them upon stimuli, making them potential candidates as drug delivery vehicles and functional biomaterials. This thesis aims at clarifying important factors affecting peptide-microgel interactions. These interactions were studied by micromanipulator-assisted light and fluorescence microscopy focusing on microgel deswelling in response to peptide binding, as well as re-swelling in response to peptide release or enzymatic degradation. To evaluate peptide uptake in microgels, solution depletion measurements were used whereas the peptide secondary structure was investigated by circular dichroism. In addition, the peptide and enzyme distribution within microgels was analyzed with confocal microscopy.

    Results presented in this thesis demonstrate that peptide incorporation into microgels, as well as peptide-induced microgel deswelling, increases with peptide length and charge density. In addition, results demonstrate that the peptide charge (length) rather than peptide charge density determines microgels deswelling. End-to-end cyclization is shown to not noticeably influence peptide-microgel interactions, suggesting that peptide cyclization can be used in combination with oppositely charged microgel carriers to improve the proteolytic and chemical stability of the peptide compared to the corresponding linear variant. Peptide secondary structure is found to drastically affect peptide incorporation into, and release from, oppositely charged microgels. Furthermore, it is shown that microgel charge density, peptide molecular weight, and enzyme concentration all greatly influence microgel bound peptide degradation. Of importance for applications, protective effects of microgels against proteolytic peptide degradation are observed only at sufficiently high microgel charge densities. Enzyme-mediated microgel degradation is shown to increase with increasing enzyme concentration, while an increased peptide loading in microgels causes a concentration-dependent decrease in microgel degradation.

    Taken together, results obtained in this work have provided some insight into factors of importance for rational use of microgels as delivery systems for protein or peptide drugs, but also in a host of other biomedical applications using weakly cross-linked polymer systems.

    Delarbeten
    1. Effects of Peptide Secondary Structure on the Interaction with Oppositely Charged Microgels
    Öppna denna publikation i ny flik eller fönster >>Effects of Peptide Secondary Structure on the Interaction with Oppositely Charged Microgels
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    2011 (Engelska)Ingår i: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 12, nr 2, 419-424 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The importance of peptide secondary structure on the interaction between antimicrobial peptides and oppositely charged poly(acrylic acid-co-acrylamide) microgels of various charge density was investigated for EFKRIVQRIKDFLRNLV (EFK17). Through D-enantiomer (EFK17-d/a; E(dF)KR(dI)VQR(dI)KD(dF)LRNLV) or tryptophan (EFK17-W/a; EWKRWVQRWKDFLRNLV) substitutions, both conformation-dependent and -independent amphiphilicity of this peptide could be precisely controlled. Peptide secondary structure was investigated by circular dichroism, whereas microgel deswelling and reswelling in response to peptide binding and release were studied by micromanipulator-assisted light and fluorescence microscopy, and peptide uptake in the microgels was determined from solution depletion measurements. Results show that peptide binding to the microgel is highly influenced by peptide secondary structure. EFK17-a, characterized by an idealized helix with all polar/charged amino acids located at one side of the helix, and all nonpolar/hydrophobic residues on the other, displays pronounced alpha-helix induction on peptide binding to the microgels. EFK17-d/a, on the other hand, displays no such amphiphilic helix induction. Mirroring this, EFK17-a displays substantially higher binding to the microgels than EFK17-d/a as well as much larger peptide-induced microgel deswelling. For EFK17-W/a, both conformation-dependent and -independent amphiphilicity effects were demonstrated. Overall, the results show that peptide conformational aspects need to be considered in peptide/microgel interactions, for example, in the design of microgel carrier systems for peptide drugs.

    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-149054 (URN)10.1021/bm101165e (DOI)000287175700017 ()21182237 (PubMedID)
    Tillgänglig från: 2011-03-15 Skapad: 2011-03-15 Senast uppdaterad: 2015-03-11Bibliografiskt granskad
    2. Effects of peptide cyclization on the interaction with oppositely charged microgels
    Öppna denna publikation i ny flik eller fönster >>Effects of peptide cyclization on the interaction with oppositely charged microgels
    2011 (Engelska)Ingår i: Colloids and Surfaces A: Physicochemical and Engineering Aspects, ISSN 0927-7757, Vol. 391, nr 1-3, 62-68 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The effect of peptide cyclization on the interaction between antimicrobial peptides and oppositely charged poly(acrylic acid-co-acrylamide) microgels of various charge density was investigated for linear and cyclic variants of peptide oligomers (C(ARKKAAKA)nC) (n = 1, 1.5, 2, 3). Through this, peptide length could be varied without substantially affecting peptide charge density and mean hydrophobicity. Furthermore, the peptides were demonstrated to display random coil conformation both in aqueous solution and when bound to oppositely charged microgels, allowing effects of cyclization to be monitored without interference from conformational transitions. With increasing peptide length, both cyclic and linear peptide variants displayed increased binding affinity to oppositely charged microgels. For all peptide lengths, however, the difference between cyclic and linear peptide variants was marginal at most, hence cyclization had little or no influence in peptide incorporation to oppositely charged microgels. In parallel, microgel deswelling increased with peptide length for both linear and cyclic peptide variants, while linear and cyclic peptide variants of the same length displayed very similar peptide-induced deswelling. Also electrolyte-induced peptide desorption from the microgels was similar for linear and cyclic peptide variants. Taken together, these findings demonstrate that end-to-end cyclization does not markedly affect peptide incorporation into, and release from, oppositely charged microgels. This opens up opportunities for the use of microgels as carriers for peptides which have been cyclized in order to improve their proteolytic and chemical stability, or in order to achieve other therapeutic advantages compared to the corresponding linear peptide variant.

    Nyckelord
    Cyclization, Microgel, Peptide
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-168831 (URN)10.1016/j.colsurfa.2011.01.029 (DOI)000299068100009 ()
    Anmärkning
    18th International Symposium on Surfactants in Solution (SIS), Melbourne Australia, 14-19 November 2010Tillgänglig från: 2012-02-16 Skapad: 2012-02-16 Senast uppdaterad: 2015-03-11Bibliografiskt granskad
    3. Peptide-Microgel Interactions in the Strong Coupling Regime
    Öppna denna publikation i ny flik eller fönster >>Peptide-Microgel Interactions in the Strong Coupling Regime
    2012 (Engelska)Ingår i: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 116, nr 35, 10964-10975 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The interaction between lightly cross-linked poly(acrylic acid) microgels and oppositely charged peptides was investigated as a function of peptide length, charge density, pH, and salt concentration, with emphasis on the strong coupling regime at high charge contrast. By micromanipulator-assisted light microscopy, the equilibrium volume response of single microgel particles upon oligolysine and oligo(lysine/alanine) absorption could be monitored in a controlled fashion. Results show that microgel deswelling, caused by peptide binding and network neutralization, increases with peptide length (3 < 5 < 10) and charge density (30% < 50% < 100%). Furthermore, oligomer-induced microgel deswelling was more pronounced at pH 5 than at pH 8, reflecting the lower network charge density in the former case (pK(a) for the isolated acrylic acid approximate to 4.7). In order to describe these highly coupled systems, a model was developed, in which counterion/peptide-mediated electrostatic attraction between the network chains is described using an exponential force law, and the network elasticity by the inverse Langevin theory. The model was used to calculate the composition of microgels in contact with reservoir solutions of peptides and simple electrolytes. At high electrostatic coupling, the calculated swelling curves were found to display first-order phase transition behavior. The model was demonstrated to capture pH- and electrolyte-dependent microgel swelling, as well as effects of peptide length and charge density on microgel deswelling. The analysis demonstrated that the peptide charge (length), rather than the peptide charge density, determines microgel deswelling. Furthermore, a transition between continuous and discrete network collapse was identified, consistent with experimental results in the present investigations, as well as with results from the literature on microgel deswelling caused by multivalent cations.

    Nationell ämneskategori
    Kemi
    Identifikatorer
    urn:nbn:se:uu:diva-182754 (URN)10.1021/jp306121h (DOI)000308339400060 ()
    Tillgänglig från: 2012-10-18 Skapad: 2012-10-15 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    4. Factors Affecting Enzymatic Degradation of Microgel-Bound Peptides
    Öppna denna publikation i ny flik eller fönster >>Factors Affecting Enzymatic Degradation of Microgel-Bound Peptides
    2013 (Engelska)Ingår i: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 14, nr 7, 2317-2325 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Proteolytic degradation and release of microgel-bound peptides was investigated for trypsin, poly(acrylic acid-co-acrylamide) microgels (70-90 mu m in diameter), and oppositely charged polylysine, using a method combination of confocal microscopy and micromanipulator-assisted light microscopy. Results show that trypsin-induced release of polylysine increased with increasing trypsin concentration, decreasing microgel charge density and decreasing peptide molecular weight. While the microgel offered good protection against enzymatic degradation at high microgel charge density, it was also observed that the cationic peptide enabled trypsin to bind throughout the peptide-loaded microgels, even when it did not bind to the peptide-void ones. With the exception of highly charged microgels, proteolytic degradation throughout the peptide-loaded microgel resulted in the generation of short and non-adsorbing peptide stretches, giving rise to the concentration and peptide length dependence observed. A simple random scission model was able to qualitatively capture these experimental findings. collectively, the results demonstrate that microgel charge density, peptide molecular weight, and enzyme concentration greatly influence degradation/release of microgel-bound peptides and need to be considered in the use of microgels, e.g., as carriers for protein and peptide drugs.

    Nationell ämneskategori
    Naturvetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-204977 (URN)10.1021/bm400431f (DOI)000321793700021 ()
    Tillgänglig från: 2013-08-16 Skapad: 2013-08-13 Senast uppdaterad: 2015-03-11Bibliografiskt granskad
    5. Chain and Pore-Blocking Effects on Matrix Degradation in Protein-Loaded Microgels
    Öppna denna publikation i ny flik eller fönster >>Chain and Pore-Blocking Effects on Matrix Degradation in Protein-Loaded Microgels
    2014 (Engelska)Ingår i: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 15, nr 10, 3671-3678 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Factors affecting matrix degradation in protein-loaded microgels were investigated for dextran-based microgels, the sugar-binding protein Concanavalin A (ConA), and the dextran-degrading enzyme Dextranase. For this system, effects of enzyme, protein, and glucose concentrations, as well as pH, were considered. Microgel network degradation was monitored by micromanipulator-assisted light microscopy, whereas enzyme and protein distributions were monitored by confocal microscopy. Results show that Dextranase-mediated microgel degradation increased with increasing enzyme concentration, whereas an increased ConA loading in the dextran microgels caused a concentration-dependent decrease in microgel degradation. In the presence of glucose, competitive release of microgel-bound ConA restored the microgel degradation observed in the absence of ConA. To clarify effects of mass transport limitations, microgel degradation was compared to that of non-cross-linked dextran, demonstrating that ConA limits enzyme substrate access in dextran microgels primarily through pore blocking and induction of pore shrinkage. The experimentally observed effects were qualitatively captured by a modified Michaelis-Menten approach for spherical symmetry, in which network blocking by ConA was included. Taken together, the results demonstrate that matrix degradation of protein-loaded microgels depends sensitively on a number of factors, which need to be considered in the use of microgels in biomedical applications.

    Nationell ämneskategori
    Biokemi och molekylärbiologi
    Identifikatorer
    urn:nbn:se:uu:diva-236549 (URN)10.1021/bm5009525 (DOI)000343026600022 ()25144139 (PubMedID)
    Tillgänglig från: 2014-11-26 Skapad: 2014-11-19 Senast uppdaterad: 2015-03-11Bibliografiskt granskad
  • 7.
    Vildhede, Anna
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    In vitro and in silico Predictions of Hepatic Transporter-Mediated Drug Clearance and Drug-Drug Interactions in vivo2015Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The liver is the major detoxifying organ, clearing the blood from drugs and other xenobiotics. The extent of hepatic clearance (CL) determines drug exposure and hence, the efficacy and toxicity associated with exposure. Drug-drug interactions (DDIs) that alter the hepatic CL may cause more or less severe outcomes, such as adverse drug reactions. Accurate predictions of drug CL and DDI risk from in vitro data are therefore crucial in drug development.

    Liver CL depends on several factors including the activities of transporters involved in the hepatic uptake and efflux. The work in this thesis aimed at developing new in vitro and in silico methods to predict hepatic transporter-mediated CL and DDIs in vivo. Particular emphasis was placed on interactions involving the hepatic uptake transporters OATP1B1, OATP1B3, and OATP2B1. These transporters regulate the plasma concentration-time profiles of many drugs including statins.

    Inhibition of OATP-mediated transport by 225 structurally diverse drugs was investigated in vitro. Several novel inhibitors were identified. The data was used to develop in silico models that could predict OATP inhibitors from molecular structure. Models were developed for static and dynamic predictions of in vivo transporter-mediated drug CL and DDIs. These models rely on a combination of in vitro studies of transport function and mass spectrometry-based quantification of protein expression in the in vitro models and liver tissue. By providing estimations of transporter contributions to the overall hepatic uptake/efflux, the method is expected to improve predictions of transporter-mediated DDIs. Furthermore, proteins of importance for hepatic CL were quantified in liver tissue and isolated hepatocytes. The isolation of hepatocytes from liver tissue was found to be associated with oxidative stress and degradation of transporters and other proteins expressed in the plasma membrane. This has implications for the use of primary hepatocytes as an in vitro model of the liver. Nevertheless, by taking the altered transporter abundance into account using the method developed herein, transport function in hepatocyte experiments can be scaled to the in vivo situation. The concept of protein expression-dependent in vitro-in vivo extrapolations was illustrated using atorvastatin and pitavastatin as model drugs.

    Delarbeten
    1. Classification of Inhibitors of Hepatic Organic Anion Transporting Polypeptides (OATPs): Influence of Protein Expression on Drug - Drug Interactions
    Öppna denna publikation i ny flik eller fönster >>Classification of Inhibitors of Hepatic Organic Anion Transporting Polypeptides (OATPs): Influence of Protein Expression on Drug - Drug Interactions
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    2012 (Engelska)Ingår i: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 55, nr 10, 4740-4763 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The hepatic organic anion transporting polypeptides (OATPs) influence the pharmacokinetics of several drug classes and are involved in many clinical drug-drug interactions. Predicting potential interactions with OATPs is, therefore, of value. Here, we developed in vitro and in silico models for identification and prediction of specific and general inhibitors of OATP1B1, OATP1B3, and OATP2B1, The maximal transport activity (MTA) of each OATP in human liver was predicted from transport kinetics and protein quantification. We then used MTA to predict the effects of a subset of inhibitors on atorvastatin uptake in vivo. Using a data set of 225 drug-like compounds, 91 OATP inhibitors were identified. In silico models indicated that lipophilicity and polar surface area are key molecular features of OATP inhibition. MTA predictions identified OATP1B1 and OATP1B3 as major determinants of atorvastatin uptake in vivo. The relative contributions to overall hepatic uptake varied with isoform specificities of the inhibitors.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-176227 (URN)10.1021/jm300212s (DOI)000304338800017 ()
    Tillgänglig från: 2012-06-19 Skapad: 2012-06-18 Senast uppdaterad: 2015-03-09Bibliografiskt granskad
    2. Hepatic Uptake of Atorvastatin: Influence of Variability in Transporter Expression on Uptake Clearance and Drug-Drug Interactions
    Öppna denna publikation i ny flik eller fönster >>Hepatic Uptake of Atorvastatin: Influence of Variability in Transporter Expression on Uptake Clearance and Drug-Drug Interactions
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    2014 (Engelska)Ingår i: Drug Metabolism And Disposition, ISSN 0090-9556, E-ISSN 1521-009X, Vol. 42, nr 7, 1210-1218 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Differences in the expression and function of the organic anion transporting polypeptide (OATP) transporters contribute to interindividual variability in atorvastatin clearance. However, the importance of the bile acid transporter sodium taurocholate cotransporting polypeptide (NTCP, SLC10A1) in atorvastatin uptake clearance (CLupt) is not yet clarified. To elucidate this issue, we investigated the relative contribution of NTCP, OATP1B1, OATP1B3, and OATP2B1 to atorvastatin CLupt in 12 human liver samples. The impact of inhibition on atorvastatin CLupt was also studied, using inhibitors of different isoform specificities. Expression levels of the four transport proteins were quantified by liquid chromatography tandem mass spectrometry. These data, together with atorvastatin in vitro kinetics, were used to predict the maximal transport activity (MTA) and interindividual differences in CLupt of each transporter in vivo. Subsequently, hepatic uptake impairment on coadministration of five clinically interacting drugs was predicted using in vitro inhibitory potencies. NTCP and OATP protein expression varied 3.7- to 32-fold among the 12 sample donors. The rank order in expression was OATP1B1 > OATP1B3 approximate to NTCP approximate to OATP2B1. NTCP was found to be of minor importance in atorvastatin disposition. Instead, OATP1B1 and OATP1B3 were confirmed as the major atorvastatin uptake transporters. The average contribution to atorvastatin uptake was OATP1B1 > OATP1B3 >> OATP2B1 > NTCP, although this rank order varied among individuals. The interindividual differences in transporter expression and CLupt resulted in marked differences in drug-drug interactions due to isoform-specific inhibition. We conclude that this variation should be considered in in vitro to in vivo extrapolations.

    Nationell ämneskategori
    Farmakologi och toxikologi Medicinska grundvetenskaper
    Identifikatorer
    urn:nbn:se:uu:diva-229430 (URN)10.1124/dmd.113.056309 (DOI)000338341300016 ()
    Tillgänglig från: 2014-08-08 Skapad: 2014-08-07 Senast uppdaterad: 2015-03-09Bibliografiskt granskad
    3. Comparative proteomics of human liver tissue and hepatocytes reveal systematic differences in proteins determining hepatic drug exposure
    Öppna denna publikation i ny flik eller fönster >>Comparative proteomics of human liver tissue and hepatocytes reveal systematic differences in proteins determining hepatic drug exposure
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    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nyckelord
    Quantitative proteomics, Oxidative stress, plasma membrane protein, drug transport and metabolism, pitavastatin, Total Protein Approach, ADME, OATP, CYP, UGT
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-241374 (URN)
    Tillgänglig från: 2015-01-12 Skapad: 2015-01-12 Senast uppdaterad: 2015-03-09
    4. Mechanistic Modeling of Pitavastatin Disposition in Sandwich-Cultured Human Hepatocytes: A Proteomics-Informed Bottom-Up Approach
    Öppna denna publikation i ny flik eller fönster >>Mechanistic Modeling of Pitavastatin Disposition in Sandwich-Cultured Human Hepatocytes: A Proteomics-Informed Bottom-Up Approach
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    2016 (Engelska)Ingår i: Drug Metabolism And Disposition, ISSN 0090-9556, E-ISSN 1521-009X, Vol. 44, nr 4, 505-516 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Isolated human hepatocytes are commonly used to predict transporter-mediated clearance in vivo. Such predictions, however, do not provide estimations of transporter contributions and consequently do not allow predictions of the outcome resulting from a change in the activity of a certain transporter, e.g., by inhibition or a genetic variant with reduced function. Pitavastatin is a drug that is heavily dependent on hepatic transporters for its elimination and it is mainly excreted as unchanged drug in the bile. For this reason, pitavastatin was used as a model drug to demonstrate the applicability of a bottom-up approach to predict transporter-mediated disposition in sandwich-cultured human hepatocytes (SCHH), allowing for the estimation of transporter contributions. Transport experiments in transfected HEK293 cells and inverted membrane vesicles overexpressing each of the relevant transport proteins were used to generate parameter estimates for the mechanistic model. By adjusting for differences in transporter abundance between the in vitro systems and individual SCHH batches, the model successfully predicted time profiles of medium and intracellular accumulation. Our predictions of pitavastatin bile accumulation could, however, not be confirmed due to a very low biliary excretion of pitavastatin in relation to the hepatic uptake in our SCHH. This study is, to our knowledge, the first to successfully simulate transporter-mediated processes in a complex system such as SCHH at the level of individual transport proteins using a bottom-up approach.

    Nyckelord
    pitavastatin, SCHH, hepatocytes, drug transport, OATP, NTCP, hepatic uptake, Pgp, BCRP, MRP2, MRP3
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-241375 (URN)10.1124/dmd.115.066746 (DOI)000372880600005 ()26842596 (PubMedID)
    Forskningsfinansiär
    Vetenskapsrådet, 2822Stiftelsen Lars Hiertas Minne
    Tillgänglig från: 2015-01-12 Skapad: 2015-01-12 Senast uppdaterad: 2016-05-18Bibliografiskt granskad
  • 8.
    Fagerberg, Jonas H.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Experimental and Computational Predictions of Drug Solubility in Human Gastrointestinal Fluids2014Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The aqueous solubility of a drug is viewed as a pivotal property for its oral absorption since only dissolved molecules can permeate the gut wall and reach the systemic circulation. The fluids in the intestine, however, do not only consist of water and therefore poor water solubility may not necessarily imply a poor solubility in the intestinal fluids and resulting low bioavailability. This thesis addresses the determination of drug solubility and dissolution rates in biorelevant dissolution media (BDM) with the aim of applying these methods to the early stages of drug discovery, where there is a need to reduce the volume of the medium and the amount of solid drug used in testing. The thesis also addresses the need for computational methods for predicting solubility in intestinal fluids and, hence, allowing in silico screening of drugs yet to be synthesized. The apparent solubility and dissolution behavior of large series of lipophilic and other diverse compounds in BDM were studied using a miniaturized method developed herein. The media used in the experimental design provided an opportunity to assess the effects of charge, solubilization in mixed lipid aggregates, and ethanol in BDM. Highly lipophilic and uncharged drugs were efficiently solubilized by aggregates in the BDM while solubilization was decreased with charge. The decrease was more pronounced for negatively charged drugs. The solubility of anionic and neutral drugs was significantly increased by the addition of ethanol to the medium and absorption simulations showed that intake of alcohol could lead to increased plasma concentrations of neutral compounds. Statistical models based on calculated molecular descriptors that accurately predicted the apparent solubility in fasted-state simulated intestinal fluid and in aspirated human intestinal fluid were also developed. In summary, the work undertaken in this thesis has resulted in new experimental and computational models for assessment of the dissolution and solubility of poorly water-soluble compounds in BDM. The models are applicable in the early discovery and development phases for predicting physiologically relevant solubility and the effects thereof on drug absorption.  

    Delarbeten
    1. Dissolution Rate and Apparent Solubility of Poorly Soluble Drugs in Biorelevant Dissolution Media
    Öppna denna publikation i ny flik eller fönster >>Dissolution Rate and Apparent Solubility of Poorly Soluble Drugs in Biorelevant Dissolution Media
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    2010 (Engelska)Ingår i: Molecular pharmaceutics, ISSN 1543-8384, Vol. 7, nr 5, 1419-1430 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    A series of poorly soluble BCS class II compounds with "grease ball" characteristics were assessed for solubility and dissolution rate in biorelevant dissolution media (BDM) with the purpose of investigating which molecular structures gain most in solubility when dissolved under physiologically relevant conditions. The compounds were studied in four media (simulated intestinal fluid in fasted (FaSSIF pH 6.5) and fed state (FeSSIF pH 5.0), and their corresponding blank buffers (FaSSIF(blk) and FeSSIFblk)) at a temperature of 37 degrees C. The experimental results were used to analyze which molecular characteristics are of importance for the solubility in BDM and for in silico modeling using multivariate data analysis. It was revealed that a majority of the compounds exhibited a higher dissolution rate and higher solubility in the FaSSIF and FeSSIF than in their corresponding blank buffers. Compounds which were neutral or carried a positive charge were more soluble in FeSSIF than FaSSIF. The acidic compounds displayed clear pH dependency, although the higher concentration of solubilizing agents in FeSSIF than FaSSIF also improved the solubility. Five of the ten compounds were upgraded to BCS class I when dissolved in FaSSIF or FeSSIF, i.e., the maximum dose of these compounds given orally was soluble in 250 mL of these BDMs. Lipophilicity as described by the log D-oct value was identified as a good predictor of the solubilization ratio (R-2 = 0.74), and computed molecular descriptors were also shown to successfully predict the solubilities in BDM for this data set. To conclude, the physiological solubility of "grease ball" molecules may be largely underestimated in in vitro solubility assays unless BDM is used. Moreover, the results herein indicate that the improvement obtained in BDM may be possible to predict from chemical features alone.

    Nyckelord
    Poor solubility, dissolution rate, biorelevant dissolution media, FaSSIF, FeSSIF, apparent BCS, physicochemical properties, molecular features, solvation limited
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-134347 (URN)10.1021/mp100049m (DOI)000282304200006 ()
    Tillgänglig från: 2010-11-25 Skapad: 2010-11-24 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    2. Ethanol Effects on Apparent Solubility of Poorly Soluble Drugs in Simulated Intestinal Fluid
    Öppna denna publikation i ny flik eller fönster >>Ethanol Effects on Apparent Solubility of Poorly Soluble Drugs in Simulated Intestinal Fluid
    2012 (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, Vol. 9, nr 7, 1942-1952 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Ethanol intake can lead to an unexpected and possibly problematic increase in the bioavailability of druglike compounds. In this work we investigated the effect of ethanol on the apparent solubility and dissolution rate of poorly soluble compounds in simulated intestinal fluid representing a preprandial state. A series of 22 structurally diverse, poorly soluble compounds were measured for apparent solubility and intrinsic dissolution rate (37 degrees C) in phosphate buffer pH 6.5 (PhB6.5) and fasted state simulated intestinal fluid (FaSSIF, pH 6.5) with and without ethanol at 5% v/v or 20% v/v. The obtained data were used to understand for which molecules ethanol results in an increased apparent solubility and, therefore, may increase the amount of drug absorbed. In FaSSIF(20%ethanol) 59% of the compounds displayed >3-fold higher apparent solubility than in pure FaSSIF, whereas the effects of 5% ethanol on solubility, in most cases, were negligible. Acidic and neutral compounds were more solubilized by the addition of ethanol than by lecithin/taurocholate aggregates, whereas bases showed a more substance-specific response to the additives in the buffer. The stronger solubilizing capacity of ethanol as compared to the mixed lipid aggregates in FaSSIF was further identified through Spearman rank analyses, which showed a stronger relationship between FaSSIF(20%ethanol) and PhB6.5,20%ethanol (r(S) of 0.97) than FaSSIF(20%ethanol) and FaSSIF (r(S) of 0.86). No relationships were found between solubility changes in media containing ethanol and single physicochemical properties, but multivariate data analysis showed that inclusion of ethanol significantly reduced the negative effect of compound lipophilicity on solubility. For this data set the higher concentration of ethanol gave a dose number (Do) <1 for 30% of the compounds that showed incomplete dissolution in FaSSIF. Significant differences were shown in the melting point, lipophilicity, and dose profiles between the compounds having a Do < 1 and Do > 1, with the latter having higher absolute values in all three parameters. In conclusion, this study showed that significant effects of ethanol on apparent solubility in the preprandial state can be expected for lipophilic compounds. The results herein indicate that acidic and neutral compounds are more sensitive to the addition of ethanol than to the mixed lipid aggregates present in the fasted intestine.

    Nyckelord
    apparent solubility, dissolution rate, ethanol, biorelevant dissolution medium, poorly soluble compounds, molecular properties, dose number
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-178097 (URN)10.1021/mp2006467 (DOI)000305917600009 ()
    Tillgänglig från: 2012-07-30 Skapad: 2012-07-27 Senast uppdaterad: 2014-04-29Bibliografiskt granskad
    3. Concomitant intake of alcohol may increase the absorption of poorly soluble drugs
    Öppna denna publikation i ny flik eller fönster >>Concomitant intake of alcohol may increase the absorption of poorly soluble drugs
    2015 (Engelska)Ingår i: European Journal of Pharmaceutical Sciences, ISSN 0928-0987, E-ISSN 1879-0720, Vol. 67, 12-20 s.Artikel i tidskrift (Refereegranskat) Published
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-220869 (URN)10.1016/j.ejps.2014.10.017 (DOI)000348257500002 ()25444841 (PubMedID)
    Anmärkning

    Title in Thesis list of papers: Concomitant Intake of Alcohol Increases the Absorption of Poorly Soluble Drugs Administered as Immediate Release Formulations

    Tillgänglig från: 2014-03-27 Skapad: 2014-03-21 Senast uppdaterad: 2016-04-12Bibliografiskt granskad
    4. Computational Prediction of Drug Solubility in Fasted Simulated and Aspirated Human Intestinal Fluid
    Öppna denna publikation i ny flik eller fönster >>Computational Prediction of Drug Solubility in Fasted Simulated and Aspirated Human Intestinal Fluid
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    2015 (Engelska)Ingår i: Pharmaceutical research, ISSN 0724-8741, E-ISSN 1573-904X, Vol. 32, nr 2, 578-589 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Purpose

    To develop predictive models of apparent solubility (Sapp) of lipophilic drugs in fasted state simulated intestinal fluid (FaSSIF) and aspirated human intestinal fluid (HIF).

    Methods

    Measured Sapp values in FaSSIF, HIF and phosphate buffer pH 6.5 (PhBpH6.5) for 86 lipophilic drugs were compiled and divided into training (Tr) and test (Te) sets. Projection to latent structure (PLS) models were developed through variable selection of calculated molecular descriptors. Experimentally determined properties were included to investigate their contribution to the predictions.

    Results

    Modest relationships between Sapp in PhBpH6.5 and FaSSIF (R2 = 0.61) or HIF (R2 = 0.62) were found. As expected, there was a stronger correlation obtained between FaSSIF and HIF (R2 = 0.78). Computational models were developed using calculated descriptors alone (FaSSIF, R2 = 0.69 and RMSEte of 0.77; HIF, R2 = 0.84 and RMSEte of 0.81). Accuracy improved when solubility in PhBpH6.5 was added as a descriptor (FaSSIF, R2 = 0.76 and RMSETe of 0.65; HIF, R2 = 0.86 and RMSETe of 0.69), whereas no improvement was seen when melting point (Tm) or logDpH 6.5 were included in the models.

    Conclusion

    Computational models were developed, that reliably predicted Sapp of lipophilic compounds in intestinal fluid, from molecular structures alone. If experimentally determined pH-dependent solubility values were available, this further improved the accuracy of the predictions.

    Ort, förlag, år, upplaga, sidor
    Springer, 2015
    Nyckelord
    biorelevant solubility, human intestinal fluid, simulated intestinal fluid, in silico, prediction
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Farmaceutisk fysikalisk kemi
    Identifikatorer
    urn:nbn:se:uu:diva-220867 (URN)10.1007/s11095-014-1487-z (DOI)000349357700019 ()25186438 (PubMedID)
    Forskningsfinansiär
    Vetenskapsrådet, 621-2008-3777Vetenskapsrådet, 621-2011-2445
    Tillgänglig från: 2014-03-27 Skapad: 2014-03-21 Senast uppdaterad: 2015-03-19Bibliografiskt granskad
  • 9.
    Persson, Ann-Sofie
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Flow and Compression of Granulated Powders: The Accuracy of Discrete Element Simulations and Assessment of Tablet Microstructure2013Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Simulations are powerful and important tools for gaining insight into powder processes. Ultimately, simulations have the potential to replace experiments. Thus, accurate models and insight into the essential factors for descriptions of powder behaviour are required. In this thesis, discrete element method (DEM) simulations of granule flow and compression were evaluated to deduce parameters and potential models essential for the experimental and numerical correspondence. In addition, the evolution in tablet microstructure during compression was studied using mercury porosimetry.

    Granule flow was measured using angle of repose, discharge rate, and shear. The granular flow depended primarily on particle shape and surface texture due to the mutual influence of these two parameters on the inter-particle forces. Rolling friction stabilised both the heap formation and promoted shear in the elastic quasi-static flow regime. Thus, rolling friction was established to be an essential simulation parameter for the correspondence to experiments.

    Current compression models often neglect the elastic compact deformation during particle loading. In this thesis, two fundamentally different models were evaluated with focus of including the elastic deformation. The first model comprised a maximal particle overlap, where elastic deformation commences. The second model accounted for the contact dependence and impingement at high relative densities. This model was based on a truncated-sphere followed by a Voronoi extension. The validity of the models was demonstrated by the elastic qualitative correspondence to experimental compressions for ductile materials.

    In tablets, the void (inter-granular pore) diameter was dependent on the degree of compression. Thus, the degree of compression provides an indication of the tablet microstructure. The microstructure was subsequently observed to be related to the tablet tensile strength as inferred from a percolation threshold required for formation of coherent tablets.

    In summary, this thesis has shed light onto the potential of simulating flow and compression of granulated pharmaceutical powders using DEM. Continuous work in the area are required to further improve the models to increase the experimental and numerical correspondence.

    Delarbeten
    1. Flowability of surface modified pharmaceutical granules: A comparative experimental and numerical study
    Öppna denna publikation i ny flik eller fönster >>Flowability of surface modified pharmaceutical granules: A comparative experimental and numerical study
    2011 (Engelska)Ingår i: European Journal of Pharmaceutical Sciences, ISSN 0928-0987, E-ISSN 1879-0720, Vol. 42, nr 3, 199-209 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Flowability - as measured by hopper discharge rate, angle of repose and Carr's index (CI) - of surface modified microcrystalline cellulose granules was investigated experimentally. Three-dimensional simulations of the granule flow were performed, using the discrete element method (DEM), including either sliding and rolling friction or sliding friction and cohesion in the model. Granule surface modification with polymer coating and lubrication was found to have a significant effect on the sliding friction coefficient. This effect was also reflected in the ensuing flow behaviour, as quantified by the experimental discharge rate and angle of repose, whereas the results for the Cl were inconclusive. The numerical results demonstrated that granular flow was qualitatively different for non-cohesive and cohesive granules, occurring in the form of individual particles for the former and in larger clusters for the latter. Rolling friction and cohesion nevertheless affected the simulated discharge rate in a similar manner, producing results comparable to those observed experimentally and calculated with the Beverloo equation. The numerical results for the cohesive granules demonstrated that cohesion alone was sufficient to produce stable heaps. However, the agreement with experimental data was satisfactory only for the non-cohesive granules, demonstrating the importance of rolling friction.

    Nyckelord
    Discharge rate, Angle of repose, Discrete element method, Rolling and sliding friction, Cohesion
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-149033 (URN)10.1016/j.ejps.2010.11.011 (DOI)000287616300004 ()21112389 (PubMedID)
    Tillgänglig från: 2011-03-15 Skapad: 2011-03-15 Senast uppdaterad: 2014-01-23Bibliografiskt granskad
    2. The influence of rolling friction on the shear behaviour of non-cohesive pharmaceutical granules: An experimental and numerical investigation
    Öppna denna publikation i ny flik eller fönster >>The influence of rolling friction on the shear behaviour of non-cohesive pharmaceutical granules: An experimental and numerical investigation
    2013 (Engelska)Ingår i: European Journal of Pharmaceutical Sciences, ISSN 0928-0987, E-ISSN 1879-0720, Vol. 49, nr 2, 241-250 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Granule shear behaviour was investigated experimentally and numerically to evaluate the reliability of the numerical model. Additionally, parameters affecting the ensuing flow regimes - elastic quasi-static and inertial non-collisional - were highlighted. Furthermore, the influence of using the Lees-Edwards periodic boundary conditions or the standard boundary conditions was studied. Experiments were performed with microcrystalline cellulose granules of three size distributions using the FT4 powder rheometer. The numerical parameters, particle size, effective density, and particle stiffness were selected to match the experimental conditions. Experimentally, an unexpected particle size effect was evident where the resistance to shear increased with particle size. Numerically, combining rolling friction. and increased shear rate enabled a transition from the inertial non-collisional to the elastic quasi-static regime at a reduced sliding friction coefficient. Presumably, this is an effect of increased particle overlap creating stronger contacts and facilitating force chain formation. Both boundary conditions provided comparable results provided a correction of system size was made, where larger systems were required for the standard boundary conditions. A satisfactory qualitative agreement between the experimentally and numerically determined yield loci emphasised the predictive capacity of the DEM. Rolling friction was in addition concluded to be an essential model parameter for obtaining an improved quantitative agreement.

    Nyckelord
    Shear, Discrete element method, Flow regimes, Rolling friction, Shear rate
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-204131 (URN)10.1016/j.ejps.2013.02.022 (DOI)000319639800017 ()
    Tillgänglig från: 2013-07-22 Skapad: 2013-07-22 Senast uppdaterad: 2014-04-15Bibliografiskt granskad
    3. An experimental evaluation of the accuracy to simulate granule bed compression using the discrete element method
    Öppna denna publikation i ny flik eller fönster >>An experimental evaluation of the accuracy to simulate granule bed compression using the discrete element method
    2012 (Engelska)Ingår i: Powder Technology, ISSN 0032-5910, E-ISSN 1873-328X, nr 219, 249-256 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    In this work, granule compression is studied both experimentally and numerically with the overall objective of investigating the ability of the discrete element method (DEM) to accurately simulate confined granule bed compression. In the experiments, granules of microcrystalline cellulose (MCC) in the size range 200-710 mu m were used as model material. Unconfined uniaxial compression of single granules was performed to determine granule properties such as the yield pressure and elastic modulus and compression profiles of the MCC granules were obtained from granule bed compression experiments. By utilizing the truncated Hertzian contact model for elastic-perfectly plastic materials, the rearrangement and plastic deformation stages of the force displacement curve were found to be in reasonable agreement with experiments. In an attempt to account for the final compression stage, elastic deformation of the compact, a simple modification of the contact model was proposed. This modification amounted to the introduction of a maximal plastic overlap, beyond which elastic deformation was the only deformation mode possible. Our results suggest that the proposed model provides an improved, although not perfect, description of granule bed compression at high relative densities and hence may be used as a basis for future improvements.

    Nyckelord
    Compression, Discrete element method, Contact model, Plastic overlap, Elastic deformation
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-172045 (URN)10.1016/j.powtec.2011.12.054 (DOI)000301310400033 ()
    Tillgänglig från: 2012-04-02 Skapad: 2012-04-01 Senast uppdaterad: 2014-05-05Bibliografiskt granskad
    4. On the role of bulk modulus and granule hardness on the simulation accuracy of confined bulk granule compression
    Öppna denna publikation i ny flik eller fönster >>On the role of bulk modulus and granule hardness on the simulation accuracy of confined bulk granule compression
    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-208807 (URN)
    Tillgänglig från: 2013-10-08 Skapad: 2013-10-08 Senast uppdaterad: 2014-01-23
    5. The degree of compression of spherical granular solids controls the evolution of microstructure and bond probability during compaction
    Öppna denna publikation i ny flik eller fönster >>The degree of compression of spherical granular solids controls the evolution of microstructure and bond probability during compaction
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    2013 (Engelska)Ingår i: International Journal of Pharmaceutics, ISSN 0378-5173, E-ISSN 1873-3476, Vol. 442, nr 1-2, 3-12 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The effect of degree of compression on the evolution of tablet microstructure and bond probability during compression of granular solids has been studied. Microcrystalline cellulose pellets of low (about 11%) and of high (about 32%) porosity were used. Tablets were compacted at 50, 100 and 150 MPa applied pressures and the degree of compression and the tensile strength of the tablets determined. The tablets were subjected to mercury intrusion measurements and from the pore size distributions, a void diameter and the porosities of the voids and the intra-granular pores were calculated. The pore size distributions of the tablets had peaks associated with the voids and the intra-granular pores. The void and intra-granular porosities of the tablets were dependent on the original pellet porosity while the total tablet porosity was independent. The separation distance between pellets was generally lower for tablets formed from high porosity pellets and the void size related linearly to the degree of compression. Tensile strength of tablets was higher for tablets of high porosity pellets and a scaled tablet tensile strength related linearly to the degree of compression above a percolation threshold. In conclusion, the degree of compression controlled the separation distance and the probability of forming bonds between pellets in the tablet. 

    Nyckelord
    Tablets, Pore structure, Microstructure, Degree of compression, Tensile strength, Percolation theory
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-196531 (URN)10.1016/j.ijpharm.2012.08.011 (DOI)000314690200002 ()
    Tillgänglig från: 2013-03-13 Skapad: 2013-03-11 Senast uppdaterad: 2014-04-17Bibliografiskt granskad
  • 10.
    Pedersen, Jenny M.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    ATP-Binding-Cassette Transporters in Biliary Efflux and Drug-Induced Liver Injury2013Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Membrane transport proteins are known to influence the absorption, distribution, metabolism, excretion and toxicity (ADMET) of drugs. At the onset of this thesis work, only a few structure-activity models, in general describing P-glycoprotein (Pgp/ABCB1) interactions, were developed using small datasets with little structural diversity. In this thesis, drug-transport protein interactions were explored using large, diverse datasets representing the chemical space of orally administered registered drugs. Focus was set on the ATP-binding cassette (ABC) transport proteins expressed in the canalicular membrane of human hepatocytes.

    The inhibition of the ABC transport proteins multidrug-resistance associated protein 2 (MRP2/ABCC2) and bile salt export pump (BSEP/ABCB11) was experimentally investigated using membrane vesicles from cells overexpressing the investigated proteins and sandwich cultured human hepatocytes (SCHH). Several previously unknown inhibitors were identified for both of the proteins and predictive in silico models were developed. Furthermore, a clear association between BSEP inhibition and clinically reported drug induced liver injuries (DILI) was identified. For the first time, an in silico model that described combined inhibition of Pgp, MRP2 and breast cancer resistance protein (BCRP/ABCG2) was developed using a large, structurally diverse dataset. Lipophilic weak bases were more often found to be general ABC inhibitors in comparison to other drugs. In early drug discovery, in silico models can be used as predictive filters in the drug candidate selection process and membrane vesicles as a first experimental screening tool to investigate protein interactions.

    In summary, the present work has led to an increased understanding of molecular properties important in ABC inhibition as well as the potential influence of ABC proteins in adverse drug reactions. A number of previously unknown ABC inhibitors were identified and predictive computational models were developed.

    Delarbeten
    1. Prediction and identification of drug interactions with the human ATP-binding cassette transporter multidrug-resistance associated protein 2 (MRP2; ABCC2)
    Öppna denna publikation i ny flik eller fönster >>Prediction and identification of drug interactions with the human ATP-binding cassette transporter multidrug-resistance associated protein 2 (MRP2; ABCC2)
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    2008 (Engelska)Ingår i: Journal of Medicinal Chemistry, ISSN 0022-2623, Vol. 51, nr 11, 3275-3287 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The chemical space of registered oral drugs was explored for inhibitors of the human multidrug-resistance associated protein 2 (MRP2, ABCC2), using a data set of 191 structurally diverse drugs and drug-like compounds. The data set included a new reference set of 75 compounds, for studies of hepatic drug interactions with transport proteins, CYP enzymes, and compounds associated with liver toxicity. The inhibition of MRP2-mediated transport of estradiol-17 beta-D-glucuronide was studied in inverted membrane vesicles from Sf9 cells overexpressing human MRP2. A total of 27 previously unknown MRP2 inhibitors were identified, and the results indicate an overlapping but narrower inhibitor space for MRP2 compared with the two other major ABC efflux transporters P-gp (ABCB1) and BCRP (ABCG2). In addition, 13 compounds were shown to stimulate the transport of cstradiol-17 beta-D-glucuronide. The experimental results were used to develop a computational model able to discriminate inhibitors from noninhibitors according to their molecular structure, resulting in a predictive power of 86% for the training set and 72% for the test set. The inhibitors were in general larger and more lipophilic and presented a higher aromaticity than the noninhibitors. The developed computational model is applicable in an early stage of the drug discovery process and is proposed as a tool for prediction of MRP2-mediated hepatic drug interactions and toxicity.

    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-96583 (URN)10.1021/jm7015683 (DOI)000256504800025 ()18457386 (PubMedID)
    Tillgänglig från: 2007-12-20 Skapad: 2007-12-20 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    2. Identification of novel specific and general inhibitors of the three major human ATP-binding cassette transporters P-gp, BCRP and MRP2 among registered drugs
    Öppna denna publikation i ny flik eller fönster >>Identification of novel specific and general inhibitors of the three major human ATP-binding cassette transporters P-gp, BCRP and MRP2 among registered drugs
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    2009 (Engelska)Ingår i: Pharmaceutical research, ISSN 0724-8741, E-ISSN 1573-904X, Vol. 26, nr 8, 1816-1831 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    PURPOSE: To study the inhibition patterns of the three major human ABC transporters P-gp (ABCB1), BCRP (ABCG2) and MRP2 (ABCC2), using a dataset of 122 structurally diverse drugs. METHODS: Inhibition was investigated in cellular and vesicular systems over-expressing single transporters. Computational models discriminating either single or general inhibitors from non-inhibitors were developed using multivariate statistics. RESULTS: Specific (n = 23) and overlapping (n = 19) inhibitors of the three ABC transporters were identified. GF120918 and Ko143 were verified to specifically inhibit P-gp/BCRP and BCRP in defined concentration intervals, whereas the MRP inhibitor MK571 was revealed to inhibit all three transporters within one log unit of concentration. Virtual docking experiments showed that MK571 binds to the ATP catalytic site, which could contribute to its multi-specific inhibition profile. A computational model predicting general ABC inhibition correctly classified 80% of both ABC transporter inhibitors and non-inhibitors in an external test set. CONCLUSIONS: The inhibitor specificities of P-gp, BCRP and MRP2 were shown to be highly overlapping. General ABC inhibitors were more lipophilic and aromatic than specific inhibitors and non-inhibitors. The identified specific inhibitors can be used to delineate transport processes in complex experimental systems, whereas the multi-specific inhibitors are useful in primary ABC transporter screening in drug discovery settings.

    Nyckelord
    ABC transporters, drug transport, inhibition, structure-activity relationships, transport proteins
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-109429 (URN)10.1007/s11095-009-9896-0 (DOI)000267685800003 ()19421845 (PubMedID)
    Tillgänglig från: 2009-10-15 Skapad: 2009-10-15 Senast uppdaterad: 2013-08-19Bibliografiskt granskad
    3. Biliary Drug Excretion and Drug-drug Interactions in Sandwich Cultured Human Hepatocytes Predicted from Inverted Membrane Vesicles and Targeted Proteomics
    Öppna denna publikation i ny flik eller fönster >>Biliary Drug Excretion and Drug-drug Interactions in Sandwich Cultured Human Hepatocytes Predicted from Inverted Membrane Vesicles and Targeted Proteomics
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    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    The ABC multidrug resistance proteins in the canalicular membrane are important contributors to the pharmacokinetic properties of drugs and endogenous compounds. In this study, a new approach for determining the individual contribution from three ABC transporters to the biliary clearance was investigated. First, the inhibition of P-glycoprotein (Pgp/ABCB1), Breast Cancer Resistance Protein (BCRP/ABCG2) and Multidrug-Resistance Associated Protein 2 (MRP2/ABCC2) was investigated in inverted membrane vesicles from HEK293 cells overexpressing each of the ABC proteins.  Inhibition profiles of prototypic substrate transport were obtained for 24 compounds and compared with results obtained in alternative expression systems. A common substrate for the three transporters was found in estradiol 17β-glucuronide (E17G). The contribution of each transporter to the E17G biliary efflux was investigated in HEK vesicles and in sandwich cultured human hepatocytes (SCHH), using nine inhibitors with different specificity towards each of the ABC-proteins. After quantification of transport kinetics and protein expression, the maximal transport activity (MTA) of each of the transporters was calculated and their contribution to the biliary clearance of E17G was predicted. The results show a good correlation between HEK-MRP2 and Sf9-MRP2 vesicle data. However, up to 40-fold lower IC50-values were obtained for Pgp and BCRP in the HEK vesicles compared to cellular expression systems. The physiologically more relevant SCHH were found to identify ABC inhibitors observed in the vesicle system that lost their ABC inhibition in this more complex model. SCHH therefore contribute with important additional information impossible to assess in vesicular systems. Finally, the MTA and subsequent of biliary clearance determinations were used to predict the absolute biliary excretion of E17G in SCHH, which was predicted with a prediction accuracy of 90%.

    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-204199 (URN)
    Tillgänglig från: 2013-07-23 Skapad: 2013-07-23 Senast uppdaterad: 2013-08-19
    4. Early Identification of Clinically Relevant Drug Interactions with the Human Bile Salt Export Pump (BSEP; ABCB11)
    Öppna denna publikation i ny flik eller fönster >>Early Identification of Clinically Relevant Drug Interactions with the Human Bile Salt Export Pump (BSEP; ABCB11)
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    2013 (Engelska)Ingår i: Toxicological Sciences, ISSN 1096-6080, E-ISSN 1096-0929, Vol. 136, nr 2, 328-343 s.Artikel i tidskrift (Övrigt vetenskapligt) Published
    Abstract [en]

    A comprehensive analysis was performed to investigate how inhibition of the human bile salt export pump (BSEP/ABCB11) relates to clinically observed drug induced liver injury (DILI). Inhibition of taurocholate (TA) transport was investigated in BSEP membrane vesicles for a dataset of 250 compounds, and 86 BSEP inhibitors were identified. Structure-activity modeling identified BSEP inhibition to correlate strongly with compound lipophilicity, while positive molecular charge was associated with a lack of inhibition. All approved drugs in the dataset (n=182) were categorized according to DILI warnings in drug labels issued by the FDA and a strong correlation between BSEP inhibition and DILI was identified. As many as 38 of the 61 identified BSEP inhibitors were associated with severe DILI, including nine drugs not previously linked to BSEP inhibition. Further, among the tested compounds, every second drug associated with severe DILI was a BSEP inhibitor. Finally, sandwich cultured human hepatocytes (SCHH) were used to investigate the relationship between BSEP inhibition, TA transport and clinically observed DILI in detail. BSEP inhibitors associated with severe DILI greatly reduced the TA canalicular efflux while BSEP inhibitors with less severe or no DILI resulted in weak or no reduction of TA efflux in SCHH. This distinction illustrates the usefulness of SCHH in refined analysis of BSEP inhibition. In conclusion, BSEP inhibition in membrane vesicles was found to correlate to DILI severity, and altered disposition of TA in SCHH  was shown to separate BSEP inhibitors associated with severe DILI from those with no or mild DILI. 

    Nyckelord
    Bile Salt Export Pump, ABCB11, BSEP, ABC transporters, drug transport, inhibition, structure-activity relationships, transport proteins, Drug Induced Liver Injury, DILI
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-204168 (URN)10.1093/toxsci/kft197 (DOI)000328695500006 ()
    Tillgänglig från: 2013-07-23 Skapad: 2013-07-23 Senast uppdaterad: 2015-02-19Bibliografiskt granskad
  • 11.
    Gernandt, Jonas
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    On the phase behaviour of hydrogels: A theory of macroion-induced core/shell equilibrium2013Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Colloidal macroions are known to interact very strongly with oppositely charged polyionic hydrogels. Sometimes this results in a non-uniform distribution of the macroions within the gel, a phenomenon that is not fully understood. This thesis is a summary of four papers on the development of a theory of the thermodynamics of macroions interacting with hydrogels, aimed at explaining the phenomenon of core/shell separation in spherical gels. It is the first theory of such interactions to use a rigorous approach to whole-gel mechanics, in which the elastic interplay between different parts of the gel is treated explicitly.

    The thesis shows that conventional theories of elasticity, earlier used on gels in pure solvent, can be generalised to apply also to gels in complex fluids, and that the general features of the phase behaviour are the same if mapped to corresponding system variables. It is found that the emergence of shells is due to attractions between macroions in the gel, mediated by polyions. Since the shell state is unfavourable from the perspective of the shell itself, being deformed from its preferred state, there will be a hysteresis between the uptake and the release of the macroion, like already known to occur with the uptake and release of pure solvent.

    Due to the elastic interplay, growth of the shell makes further growth progressively more favourable. Thus, unless there is a limited amount of macroions available the system will not reach equilibrium until complete phase transition has taken place. If the amount is limited the core/shell separation can be in equilibrium, so the volume of the solution that the gel is in contact with plays a very important part in determining the thermodynamic resting point of the system. The ability of a macroion/hydrogel to phase separate thus depends on the molecular properties whereas the ultimate fate of such a separation depends on the proportions in number between the ingoing components.

    Delarbeten
    1. A model describing the internal structure of core/shell hydrogels
    Öppna denna publikation i ny flik eller fönster >>A model describing the internal structure of core/shell hydrogels
    2011 (Engelska)Ingår i: Soft Matter, ISSN 1744-683X, Vol. 7, nr 21, 10327-10338 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    We apply a theory to the constrained swelling of gel particles, explicitly accounting for the propagation of elastic forces through the particle. This approach, together with conventional thermodynamics of gel swelling, allows modelling of the equilibrium state of gels with properties that are spatially inhomogeneous. In our case we consider both a discrete inhomogeneity in the form of assigning different water solubilities to the core and shell domains of the particle, and a continuous inhomogeneity in allowing the density of chemical cross-links to vary gradually through the network. The model is used to understand the behaviour of temperature-sensitive poly(N-isopropyl acrylamide) core/poly(N-isopropyl methacrylamide) shell microgels investigated in an earlier experimental study. How the swelling of the core and shell is affected by the presence of each other at different temperatures is investigated and explained from a mechanical and thermodynamic perspective.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-161771 (URN)10.1039/c1sm05694h (DOI)000296026700065 ()
    Tillgänglig från: 2011-11-21 Skapad: 2011-11-17 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    2. Core-shell separation of a hydrogel in a large solution of proteins
    Öppna denna publikation i ny flik eller fönster >>Core-shell separation of a hydrogel in a large solution of proteins
    2012 (Engelska)Ingår i: Soft Matter, ISSN 1744-683X, Vol. 8, nr 42, 10905-10913 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Upon absorption of large oppositely charged electrolytes such as proteins, polyionic hydrogels are frequently observed to separate into dense shell–swollen core states. We have developed a theory that in a detailed way takes into account the inhomogeneous swelling and distribution of the protein within such a core–shell separated gel. With this we investigated whether the core–shell separation can be an equilibrium state or if it must be understood as a dynamical phenomenon. Restricting ourselves to spherical gels with an unlimited supply of protein, we found that as an intermediate between a swollen and a collapsed gel the core–shell state can indeed be the one of lowest free energy but this state is not stable. In such cases where formation of a shell could occur spontaneously there was no further thermodynamic barrier to complete collapse of the gel (but possibly dynamical ones). The core–shell separation was favourable in systems of high charge and low ionic strength and was explained, within our theory, by the fact that the energy gain in packing proteins and polyions closely together outweighs the entropy loss of the uneven distribution.

    Ort, förlag, år, upplaga, sidor
    Royal Society of Chemistry, 2012
    Nationell ämneskategori
    Fysikalisk kemi
    Identifikatorer
    urn:nbn:se:uu:diva-184518 (URN)10.1039/C2SM26227D (DOI)000310829400014 ()
    Tillgänglig från: 2012-11-08 Skapad: 2012-11-08 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    3. Hysteresis in the surfactant-induced volume transition of hydrogels
    Öppna denna publikation i ny flik eller fönster >>Hysteresis in the surfactant-induced volume transition of hydrogels
    2015 (Engelska)Ingår i: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 119, nr 4, 1717-1725 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The discontinuous uptake and release of surfactants by hydrogels and the accompanying discontinuous volume transition is known to occur with a hysteresis. We have performed a theoretical analysis in order to find the mechanistic origin of this phenomenon. Using a mean-field model, we have quantitatively reproduced the experimental behavior by considering the cost of elastically deforming the gel material to allow phase coexistence. The major part of the hysteresis is due to the high phase coexistence cost of the swelling transition, since in this direction the coexistence cost depends not only on the elasticity of the network (being a weak force in comparison) but also on the entropy of the monovalent nonsurfactant electrolyte present in the system.

    Nationell ämneskategori
    Fysikalisk kemi
    Identifikatorer
    urn:nbn:se:uu:diva-188149 (URN)10.1021/jp5087416 (DOI)000348753600047 ()25567724 (PubMedID)
    Tillgänglig från: 2012-12-12 Skapad: 2012-12-12 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    4. Stable core/shell separation in hydrogels induced by surfactants
    Öppna denna publikation i ny flik eller fönster >>Stable core/shell separation in hydrogels induced by surfactants
    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nationell ämneskategori
    Fysikalisk kemi
    Identifikatorer
    urn:nbn:se:uu:diva-188150 (URN)
    Tillgänglig från: 2012-12-12 Skapad: 2012-12-12 Senast uppdaterad: 2013-02-11
  • 12.
    Mahmoodi, Foad
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Compression Mechanics of Powders and Granular Materials Probed by Force Distributions and a Micromechanically Based Compaction Equation2012Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The internal dynamics of powder systems under compression are as of yet not fully understood, and thus there is a necessity for approaches that can help in further clarifying and enhancing the level of understanding on this subject. To this end, the internal dynamics of powder systems under compression were probed by means of force distributions and a novel compaction equation.

    The determination of force distributions hinged on the use of carbon paper as a force sensor, where the imprints transferred from it onto white paper where converted through calibration into forces. Through analysis of these imprints, it was found that the absence of friction and bonding capacity between the particles composing the powder bed had no effect on how the applied load was transferred through the system. Additionally, it was found that pellet strength had a role to play in the homogeneity of force distributions, where, upon the occurrence of fracture, force distributions became less homogenous.

    A novel compaction equation was derived and tested on a series of systems composed of pellets with differing mechanical properties. The main value of the equation lay in its ability to predict compression behavior from single particle properties, and the agreement was especially good when a compact of zero porosity was formed.

    The utility of the equation was tested in two further studies, using a series of pharmaceutically relevant powder materials. It was established that the A parameter of the equation was a measure of the deformability of the powder material, much like the Heckel 1/K parameter, and can be used as a means to rank powders according to deformability, i.e. to establish plasticity scale. The equation also provided insights into the dominating compression mechanisms through an invariance that could be exploited to determine the point, at which the powder system became constrained, i.e. the end of rearrangement. Additionally, the robustness of the equation was demonstrated through fruitful analysis of a set of diverse materials.

    In summary, this thesis has provided insights and tools that can be translated into more efficient development and manufacturing of medicines in the form of tablets.

    Delarbeten
    1. An effective-medium analysis of confined compression of granular materials
    Öppna denna publikation i ny flik eller fönster >>An effective-medium analysis of confined compression of granular materials
    2009 (Engelska)Ingår i: Powder Technology, ISSN 0032-5910, E-ISSN 1873-328X, Vol. 194, nr 3, 228-232 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    A simplified model for confined compression of granular materials is considered, which idealizes the collection of particles as a (central) force network. Applying an effective-medium procedure, an equation with micromechanically well-defined parameters is derived, which relates the applied pressure to the engineering strain of the powder during uniaxial compression. Despite the simplicity of the model, comparison with experimental data for mm-sized spherical granules indicates that this equation is able to satisfactorily predict the overall compression profile from single-particle data.

    Nyckelord
    Granular materials, Compression, Effective-medium theory, Pharmaceuticals, Mathematical modelling, Powder technology
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-123115 (URN)10.1016/j.powtec.2009.04.012 (DOI)000267842700009 ()
    Tillgänglig från: 2010-04-23 Skapad: 2010-04-23 Senast uppdaterad: 2016-05-13Bibliografiskt granskad
    2. Effect of lubrication on the distribution of force between spherical agglomerates during compression
    Öppna denna publikation i ny flik eller fönster >>Effect of lubrication on the distribution of force between spherical agglomerates during compression
    2010 (Engelska)Ingår i: Powder Technology, ISSN 0032-5910, E-ISSN 1873-328X, Vol. 198, nr 1, 69-74 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    We employ the carbon paper technique to aid the understanding of in die force and spatial distributions, upon compression of approximately I mm sized spherical agglomerates (pellets) of microcrystalline cellulose (MCC). The aim in this study was to test for the effect of lubricant film on force and spatial distributions. Pellets of MCC were formed via granulation and extrusion/spheronisation. Investigation of pellet bed compression was performed on a materials tester. Prior to compression studies the pellets were characterised for bulk density. size and deformability. Two pellet types were investigated; MCC and MCC lubricated with magnesium stearate. The carbon paper technique relies upon carbon paper as the medium for transferring imprints from compressed pellets onto photo quality paper. The digitised images of these imprints form the basis of analysis through the use of image processing software. Using the carbon paper technique within the range of 10-30 MPa indicates that lubrication does not have a significant effect on the distribution of forces between spherical agglomerates during uniaxial compression. Spatial analysis of the imprints revealed that the lubricated pellets exhibited a higher packing order than the unlubricated ones at low applied pressures (10 and 20 MPa), a difference that could not be observed at 30 MPa. Hence interparticle friction and/or cohesion appear to influence the initial particle rearrangement, whereas confinement is suggested to dominate at higher pressures.

    Nyckelord
    Granular materials, Compression, Force distributions, Carbon paper technique, Lubrication
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-137888 (URN)10.1016/j.powtec.2009.10.016 (DOI)000274281400010 ()
    Tillgänglig från: 2010-12-16 Skapad: 2010-12-16 Senast uppdaterad: 2013-10-31Bibliografiskt granskad
    3. Effect of spherical-agglomerate strength on the distribution of force during uniaxial compression
    Öppna denna publikation i ny flik eller fönster >>Effect of spherical-agglomerate strength on the distribution of force during uniaxial compression
    2011 (Engelska)Ingår i: Powder Technology, ISSN 0032-5910, E-ISSN 1873-328X, Vol. 206, nr 3, 283-290 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    We employ the carbon paper technique with the aim of investigating the effect of spherical-agglomerate (pellet) strength on force distributions, through confined compression of approximately 1 mm sized pellets formed from microcrystalline cellulose and polyethylene glycol. The carbon paper technique relies on the transference of imprints from compressed pellets onto white photo quality paper, which are digitised and processed via image processing software. The investigated pellets can both deform plastically and develop localised cracks in response to an applied stress, while remaining largely intact during confined compression. Our results indicate that such crack formation - henceforth referred to as fracture - has a decisive influence on force distributions. Previous work on non-fracturing systems has found that the distribution of normalized forces tends to narrow with increasing particle deformation. No narrowing is observed after the point of fracture in this study and the width of the distributions - as quantified by the standard deviation of non-normalized forces - is found to increase with the difference between non-normalized mean force and fracture force. Additional corroborative results show that spatial force-force correlations typically exhibit a marked change once the fracture force is exceeded.

    Nyckelord
    Granular materials, Compression, Force distributions, Carbon paper technique, Particle failure
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-147764 (URN)10.1016/j.powtec.2010.09.031 (DOI)000286299200011 ()
    Tillgänglig från: 2011-03-01 Skapad: 2011-02-28 Senast uppdaterad: 2013-11-06Bibliografiskt granskad
    4.
    Posten kunde inte hittas. Det kan bero på att posten inte längre är tillgänglig eller att du har råkat ange ett felaktigt id i adressfältet.
    5. On the practical utility of an effective medium based compaction equation: application to a diverse set of pharmaceutically relevant materials
    Öppna denna publikation i ny flik eller fönster >>On the practical utility of an effective medium based compaction equation: application to a diverse set of pharmaceutically relevant materials
    Visa övriga...
    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-171873 (URN)
    Tillgänglig från: 2012-03-28 Skapad: 2012-03-28 Senast uppdaterad: 2012-08-01
  • 13.
    Carlert, Sara
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Investigation and Prediction of Small Intestinal Precipitation of Poorly Soluble Drugs: a Study Involving in silico, in vitro and in vivo Assessment2012Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The main objectives of the present project were to increase the understanding of small intestinal precipitation of poorly soluble pharmaceutical drugs, investigate occurrence of crystalline small intestinal precipitation and effects of precipitation on absorption. The aim was to create and evaluate methods of predicting crystalline small intestinal drug precipitation using in vivo, in vitro and in silico models.

    In vivo small intestinal precipitation from highly supersaturated solutions of two weakly basic model drugs, AZD0865 and mebendazole, was investigated in humans and canine models. Potential precipitation of AZD0865 was investigated by examining dose dependent increases in human maximum plasma concentration and total exposure, which turned out to be dose linear over the range investigated, indicating no significant in vivo precipitation. The small intestinal precipitation of mebendazole was investigated from drug concentrations and amount of solid drug present in dog jejunum as well as through the bioavailability after direct duodenal administration in dogs. It was concluded that mebendazole small intestinal precipitation was limited, and that intestinal supersaturation was measurable for up to 90 minutes.

    In vitro precipitation methods utilizing simulated or real fasted gastric and intestinal fluids were developed in order to simulate the in vivo precipitation rate. The methods overpredicted in vivo precipitation when absorption of drug was not simulated. An in vitro-in silico approach was therefore developed, where the in vitro method was used for determining the interfacial tension (γ), necessary for describing crystallization in Classical Nucleation Theory (CNT). CNT was evaluated using a third model drug, bicalutamide, and could successfully describe different parts of the crystallization process of the drug. CNT was then integrated into an in silico absorption model. The in vivo precipitation results of AZD0865 and mebendazole were well predicted by the model, but only by allowing the fundamental constant γ to vary with concentration. Thus, the in vitro-in silico approach could be used for small intestinal precipitation prediction if the in vitro concentration closely matched in vivo small intestinal concentrations.

    Delarbeten
    1. Nucleation and crystal growth in supersaturated solutions of a model drug
    Öppna denna publikation i ny flik eller fönster >>Nucleation and crystal growth in supersaturated solutions of a model drug
    2008 (Engelska)Ingår i: Journal of Colloid and Interface Science, ISSN 0021-9797, E-ISSN 1095-7103, Vol. 325, nr 2, 404-413 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The crystallization process in aqueous solutions of the drug bicalutamide and the effect of the polymer polyvinylpyrrolidone (PVP) have been studied. Results show that PVP decreased the crystallization rate significantly in a system with PVP concentrations as low as 0.01% (w/w), without changing the polymorph formed. The crystal habit was changed already at PVP concentrations as low as 0.001% (w/w). Measurements made with self-diffusion NMR indicated that the decrease in crystallization rate was not because of a reduced supersaturation due to bicalutamide binding to PVP in solution. Furthermore, in experiments designed to specifically study crystal nucleation, the same nucleation rate was found in the absence and presence of PVP. Instead, PVP adsorbes to the crystals formed in solution and by doing so, the crystal growth rate is reduced. This was confirmed in separate experiments using bicalutamide nanocrystals. By combining theories describing classical nucleation and crystal growth, with some modifications, a consistent description of several independent experiments performed in polymer-free systems was obtained. From these experiments a crystal-water interfacial tension of 22.1 mN/m was extracted. We also analyze the interfacial tension of other crystalline organic solids and find that it varies approximately as the logarithm of the solubility. This finding is discussed within the framework of the Bragg-Williams regular solution theory where we also compare with the tension of liquid alkanes.

    Nyckelord
    Crystal nucleation, Crystal growth, Poorly soluble drugs, Classical nucleation theory
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-178050 (URN)10.1016/j.jcis.2008.05.034 (DOI)
    Tillgänglig från: 2012-07-25 Skapad: 2012-07-25 Senast uppdaterad: 2013-01-22Bibliografiskt granskad
    2. Predicting intestinal precipitation: a case example for a basic BCS class II drug
    Öppna denna publikation i ny flik eller fönster >>Predicting intestinal precipitation: a case example for a basic BCS class II drug
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    2010 (Engelska)Ingår i: Pharmaceutical research, ISSN 0724-8741, E-ISSN 1573-904X, Vol. 27, nr 10, 2119-2130 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    PURPOSE: To investigate the prediction accuracy of in vitro and in vitro/in silico methods for in vivo intestinal precipitation of basic BCS class II drugs in humans. METHODS: Precipitation rate of a model drug substance, AZD0865 (pKa = 6.1; log K(D) = 4.2), was investigated in vitro using simulated intestinal media, and calculations of the crystallization rates were made with a theoretical model. Human intestinal precipitation was estimated by analysis of pharmacokinetic data from clinical studies at different doses. RESULTS: All in vitro models predicted rapid drug precipitation, where the intestinal concentration of dissolved AZD0865 at the highest dose tested was expected to decrease to half after less than 20 min. However, there was no indication of precipitation in vivo in humans as there was a dose proportional increase in drug plasma exposure. The theoretical model predicted no significant precipitation within the range of expected in vivo intestinal concentrations. CONCLUSIONS: This study indicated that simple in vitro methods of in vivo precipitation of orally administered bases overpredict the intestinal crystalline precipitation in vivo in humans. Hydrodynamic conditions were identified as one important factor that needs to be better addressed in future in vivo predictive methods.

    Nyckelord
    absorption-biopharmaceutics classification system . gastrointestinal . in vitro/in vivo correlations (IVIVC) . precipitation
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-132389 (URN)10.1007/s11095-010-0213-8 (DOI)000281860900010 ()20717839 (PubMedID)
    Tillgänglig från: 2010-10-19 Skapad: 2010-10-19 Senast uppdaterad: 2013-01-22Bibliografiskt granskad
    3.
    Posten kunde inte hittas. Det kan bero på att posten inte längre är tillgänglig eller att du har råkat ange ett felaktigt id i adressfältet.
    4. Evaluation of the use of Classical Nucleation Theory for predicting intestinal crystallization of two weakly basic BCS class II drugs
    Öppna denna publikation i ny flik eller fönster >>Evaluation of the use of Classical Nucleation Theory for predicting intestinal crystallization of two weakly basic BCS class II drugs
    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    The aim of this work was to evaluate an in vitro-in silico approach for prediction of small intestinal crystallization of two weakly basic model BCS class II drugs, AZD0865 and mebendazole, and the effect crystallization would have on the absorption prediction of the drug. The crystallization rates were investigated in an in vitro method using simulated gastric and intestinal media, and the result was modeled by using Classical Nucleation Theory (CNT). The effect of varying in vitro parameters (initial drug concentration, rate of mixing gastric and intestinal fluid, stirring and filtration) on the interfacial tension g, being a key parameter in CNT, was investigated. The initial drug concentration had the most significant effect on g for both substances tested, although g is a fundamental parameter independent of concentration according to CNT. In the subsequent in silico prediction of drug absorption an empirical approach was used where g was predicted at expected in vivo small intestinal concentrations. The results showed that lack of crystallization effects on absorption in man of the model drug AZD0865 up to doses of 4 mg/kg could be predicted. Mebendazole intestinal precipitation in canines was also well described by the model, where mean predicted amount precipitated was 111% (range 41-166%) of measured solid amount, and mean predicted supersaturation was 106% (range 73-118%) of measured supersaturation. The plasma concentration of mebendazole after duodenal administration of a solution could not be predicted by the model with the same precision in the absence of measured intestinal drug concentrations as basis for estimating the g value. In conclusion, the in vitro-in silico approach can be used for predictions of absorption effects of crystallization, but the model could benefit from further development work on the theoretical crystallization model and in vitro experimental design.

    Nyckelord
    precipitation, in silico prediction, absorption, biopharmaceutics classification system, crystallization, classical nucleation theory, in vitro-in vivo correlations
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-178052 (URN)
    Tillgänglig från: 2012-07-25 Skapad: 2012-07-25 Senast uppdaterad: 2013-01-22
  • 14.
    Fagerlind, Hanna
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Patient-Physician Communication in Oncology Care: The character of, barriers against, and ways to evaluate patient-physician communication, with focus on the psychosocial dimensions2012Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The overall aim of this thesis was to characterize patient-physician communication in oncology care with focus on the content and quality of the consultations from the perspectives of patients, oncologists and observer. Further, the aim was to explore oncologists’ perceived barriers against psychosocial communication in out-patient consultations. Finally, the aim was to evaluate different methods for evaluating communication in this setting.

    Routine oncology out-patient consultations from two different hospitals were audio-recorded. After the consultations, patients and oncologists perceptions of the content and quality of the communication were assessed using a self-report questionnaire. A nation-wide survey was performed to assess oncologists’ perceived barriers against psychosocial communication. Finally, the audio-recorded consultations were used for evaluating inter-rater reliability and feasibility of two different communication analysis instruments.

    Patient-physician consultations in oncology care are focused on the physical aspects of disease and treatment, both in terms of how often these issues were discussed and in terms of the amount of time spent on discussing them. Psychosocial issues, such as the disease’s effects on patients’ emotional or social functioning, are not always discussed during consultations, and the time spent on such discussions is limited. When psychosocial issues are discussed during the medical consultations, they are most often patient-initiated. Reasons for why psychosocial aspects are seldom discussed during the medical consultations can be the barriers concerning this kind of communication perceived by a large majority (93%) of the oncologists. Barriers against psychosocial communication were identified at organizational levels (including guidelines, routines, and resources) and individual levels (including physicians’ knowledge and attitudes).

    Furthermore, this thesis shows that there are methods with high feasibility and reliability for evaluating the content of patient-physician communication, in large study samples in oncology care. The method (observation/self-report) and perspective (patient, physician, and observer) used when evaluating communication affects the results. This needs to be considered when choosing evaluation methods in intervention studies.

    There are reasons to continue to evaluate, promote and implement promising ways of achieving better communication in clinical practice. Research should focus on how to overcome barriers against psychosocial communication.

    Delarbeten
    1. Patient-physician communication during oncology consultations
    Öppna denna publikation i ny flik eller fönster >>Patient-physician communication during oncology consultations
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    2008 (Engelska)Ingår i: Psycho-Oncology, ISSN 1057-9249, E-ISSN 1099-1611, Vol. 17, nr 10, 975-985 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    OBJECTIVE: The aim of this study was to characterize the content of patient-physician communication in standard oncology care. METHODS: The sample consisted of 19 patients with gastrointestinal cancer. The consultations were audio-recorded, transcribed verbatim, and analyzed according to qualitative content analysis. RESULTS: The analysis resulted in seven main categories: Disease and treatment, Healthcare planning, Everyday living, Psychological well-being, Coping with disease, Expressions of concerns and feelings, and Other aspects of communication. The main focus during the consultations was on disease and treatment. Physicians tended to concentrate on response to treatment and types and severity of side effects and how to treat them. More patient-centered subjects of psychosocial character like coping and psychological well-being were discussed only briefly, if at all. CONCLUSIONS: This study adds to the information given by the existing communication analysis systems, and hence we suggest a development of the psychosocial content categories of those systems to make them more valid.

    Nyckelord
    cancer, oncology, patient-physician communication, consultation, qualitative
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-86840 (URN)10.1002/pon.1410 (DOI)000260351000003 ()18677715 (PubMedID)
    Tillgänglig från: 2008-12-08 Skapad: 2008-12-08 Senast uppdaterad: 2013-02-11Bibliografiskt granskad
    2. Communication analysis in oncology care: Performance of a combination of a content analysis system and a global scale
    Öppna denna publikation i ny flik eller fönster >>Communication analysis in oncology care: Performance of a combination of a content analysis system and a global scale
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    2011 (Engelska)Ingår i: Psycho-Oncology, ISSN 1057-9249, E-ISSN 1099-1611, Vol. 20, nr 9, 992-1000 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Objective: The aim was to assess the feasibility and reliability of Velikova's Content Analysis System (VCAS) and the Medical Interaction Process System (MIPS) global scale for evaluation of communication in oncology care.

    Methods: Seventy routine physician consultations with gastro-intestinal (GI) cancer patients were audio-recorded. Two coders applied VCAS and MIPS global scale to the consultations. VCAS captures aspects of communication like symptoms, side effects, functional issues (e.g. emotional, social, physical), health-related quality of life and medical decision making. MIPS global scale measures the total impression of the consultation, e.g. patient centredness and psychosocial focus.

    Results: In total, 61 of 70 consultations were coded. The coding took twice the consultations' actual durations in minutes for VCAS. The time for coding MIPS global scale equalled the consultations length. However, the coder had then listened to the consultation twice before, coding for VCAS. Cohen's kappa for all aspects measured by VCAS varied between 0.20 and 1, mean 0.80. One category (Info on test) had a kappa of 0.20, the other categories were all above 0.60. Weighted Kappa for MIPS global scale varied between 0.25 and 0.73, mean 0.42.

    Conclusions: VCAS and MIPS global scale is a feasible combination of tools for evaluating patient-physician communication regarding content, medical decision making and global aspects of communication. VCAS showed high reliability. The MIPS global scale showed lower reliability, due to its sensitivity to the individual coders' unique values, common for all global scales. Further development of the combination of content and global instruments would be valuable.

    Nyckelord
    oncology, cancer, patient-physician communication, content analysis systems, clinical practice
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-160203 (URN)10.1002/pon.1808 (DOI)000295122700011 ()
    Tillgänglig från: 2011-10-18 Skapad: 2011-10-18 Senast uppdaterad: 2013-02-11Bibliografiskt granskad
    3. Different perspectives on communication quality and emotional functioning during routine oncology consultations
    Öppna denna publikation i ny flik eller fönster >>Different perspectives on communication quality and emotional functioning during routine oncology consultations
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    2012 (Engelska)Ingår i: Patient Education and Counseling, ISSN 0738-3991, E-ISSN 1873-5134, Vol. 88, nr 1, 16-22 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Objective: To determine quality of communication in routine oncology consultations from patient, physician, and observer perspectives, and to determine agreement of emotional function content in consultations from these three perspectives.

    Methods: In total, 69 consultations were included. Perceived quality of communication and whether or not emotional functioning had been discussed was evaluated with patient- and physician-reported questionnaires. Observer perspective was evaluated by content analysis of audio records of the consultations. Agreement between perspectives was analyzed and means compared using linear mixed models.

    Results: The patients' ratings of communication quality differed significantly from those of both the physician and observer. Observer and physician scores did not differ significantly. Physicians rated emotional functioning as discussed more often than was reported from patient and observer perspectives.

    Conclusion: The patients' view of the quality of communication differed from that of the physician and observer. Whether emotional functioning was discussed or not was also perceived differently by patients, physicians, and observer.

    Practice implications: The underpinnings and implications of these results need to be further explored regarding how to move toward a higher degree of shared understanding, where different perspectives are more in alignment, and how to develop more valid methods for evaluating communication.

    Nyckelord
    Oncology, Communication, Content analysis, Emotional functioning, Communication analysis, Clinical practice
    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-179580 (URN)10.1016/j.pec.2011.12.002 (DOI)000306541300004 ()
    Tillgänglig från: 2012-08-20 Skapad: 2012-08-20 Senast uppdaterad: 2013-02-11Bibliografiskt granskad
    4. Barriers against psychosocial communication: Oncologists' perceptions
    Öppna denna publikation i ny flik eller fönster >>Barriers against psychosocial communication: Oncologists' perceptions
    2013 (Engelska)Ingår i: Journal of Clinical Oncology, ISSN 0732-183X, E-ISSN 1527-7755, Vol. 31, nr 30Artikel i tidskrift (Refereegranskat) Published
    Nyckelord
    oncology, communication, clinical practice, barriers, oncologists
    Nationell ämneskategori
    Folkhälsovetenskap, global hälsa, socialmedicin och epidemiologi Farmaceutisk vetenskap
    Forskningsämne
    Samhällsfarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-183660 (URN)10.1200/JCO.2012.45.1609 (DOI)000330540400019 ()
    Tillgänglig från: 2012-10-31 Skapad: 2012-10-31 Senast uppdaterad: 2014-03-20Bibliografiskt granskad
  • 15.
    Thörn, Helena Anna
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    First-pass Intestinal Metabolism of Drugs: Experiences from in vitro, in vivo and simulation studies2012Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The bioavailability of a drug can be described as the fraction of an orally administered dose that reaches the systemic circulation and is often limited by first-pass metabolism in the gut and the liver. It is important to have knowledge about these processes since the systemic blood drug concentration is tightly connected to the effect of the drug.

    The general aim of this project was to quantitatively examine the role of the intestine in relation to the liver in first-pass metabolism of orally administered drugs. The first-pass metabolism of verapamil and raloxifene was investigated in detail with in vivo, in vitro and simulation studies, using the pig as an experimental model.

    The intestine contributed to the same extent as the liver to first-pass metabolism of R/S-verapamil in vivo in pigs. The S-isomer of verapamil was found in lower plasma concentrations compared to the R-isomer after oral dosing. The in vitro metabolism of verapamil in pig and human liver showed interspecies similarity and indicated equal intrinsic clearance for R- and S-verapamil. Through physiologically based pharmacokinetic modeling the stereoselectivity was explained by a combination of several processes, including enantioselective plasma protein binding, blood-to-plasma partition, and gut and liver tissue distribution. For raloxifene the intestine was the dominating organ in first-pass glucuronidation in vivo in pigs. Furthermore, the raloxifene concentration entering the intestine or the dose administered in the gut did not influence the plasma PK of raloxifene and indicated that the intestinal metabolism was not saturable with clinical relevant doses. For both verapamil and raloxifene, a time-dependent hepatic metabolism was noted with major consequences to the pharmacokinetic of the drugs.

    This project has pointed out the importance of intestinal metabolism in the overall first-pass extraction of drugs and indicates that intestinal metabolism should be considered and evaluated early in drug development.

    Delarbeten
    1. Different effects of ketoconazole on the stereoselective first-pass metabolism of R/S-verapamil in the intestine and the liver: important for the mechanistic understanding of first-pass drug-drug interactions
    Öppna denna publikation i ny flik eller fönster >>Different effects of ketoconazole on the stereoselective first-pass metabolism of R/S-verapamil in the intestine and the liver: important for the mechanistic understanding of first-pass drug-drug interactions
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    2009 (Engelska)Ingår i: Drug Metabolism And Disposition, ISSN 0090-9556, E-ISSN 1521-009X, Vol. 37, nr 11, 2186-2196 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    In this acute study a pig jejunal intestinal perfusion model with multiple plasma sampling sites and three different administration routes was used to investigate the quantitative contribution of the intestine versus liver to the first-pass extraction of each enantiomer of verapamil (VER). A subclinical dose of ketoconazole (8 mg) was coadministered in the perfusion solution to selectively inhibit gut wall CYP3A. Both enantiomers of VER and its main metabolite norverapamil (NOR) as well as the inhibitor ketoconazole were quantified in all plasma compartments by liquid chromatography-tandem mass spectrometry. The overall first-pass metabolic extraction of VER and the metabolite NOR was shown to be stereoselective with the S-isomer being more extensively extracted. For VER the ratio of R- enantiomer to S-enantiomer was greater in the hepatic vein than in the portal vein (approximately 2.2 versus 1.4), indicating that the stereoselective metabolism of VER in pigs mainly occurs on the first pass through the liver and not in the intestine. Ketoconazole increased the area under the curve from time 0 to 6 h and C(max) of R- and S-VER at least 3-fold in the portal vein, most likely explained by inhibition of gut wall metabolism. Conversely, hepatic extraction was increased because the effect of gut wall metabolism was not observed at the peripheral sampling sites. In conclusion, this study provided novel and more direct information on the contribution of the intestine and the liver, respectively, to the overall first-pass extraction of racemic VER.

    Nyckelord
    verapamil, ketoconazole, liver metabolism, intestinal metabolism, pig metabolism, first-pass metabolism
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-110886 (URN)10.1124/dmd.109.028027 (DOI)000270876600010 ()19687151 (PubMedID)
    Tillgänglig från: 2009-11-30 Skapad: 2009-11-30 Senast uppdaterad: 2012-05-08Bibliografiskt granskad
    2. Drug metabolism of CYP3A4, CYP2C9 and CYP2D6 substrates in pigs and humans
    Öppna denna publikation i ny flik eller fönster >>Drug metabolism of CYP3A4, CYP2C9 and CYP2D6 substrates in pigs and humans
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    2011 (Engelska)Ingår i: European Journal of Pharmaceutical Sciences, ISSN 0928-0987, E-ISSN 1879-0720, Vol. 43, nr 3, 89-98 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Pigs are becoming increasingly used as a test animal both in pharmacological and toxicological assessment of new drug compounds. For interspecies comparisons and predictions it is important to characterize the expression and function of membrane transport and enzymatic proteins in pigs, particularly at a mechanistic level which will make extrapolation of observation between pig and man to be made with more confidence. The major objective of this report was to increase the integrative knowledge of drug metabolism in pigs and to compare with corresponding data from human liver microsomes. This was done by using human substrates of CYP3A4 (verapamil and testosterone), CYP2C9 (diclofenac) and CYP2D6 (dextromethorphan). In addition, the mRNA expression of important drug metabolizing enzymes and carrier-mediated transporters were assessed in intestine and liver tissues from pigs. It was shown that CYP3A4 activity is quantitatively comparable between the two species but data suggest that qualitative differences may exist. Verapamil showed similar metabolism pattern as in humans and the CYP3A4 inhibitor ketoconazole was able to inhibit the depletion of both R- and S-verapamil. A correlation between individual pig CYP3A mRNA expression and in vivo hepatic extraction ratio was established which indicates that CYP3A is the major determinant factor in both pigs and humans. However, investigations of the metabolism of testosterone resulted in qualitative different metabolite pattern between pigs and humans. The metabolism of diclofenac was very low in pig liver microsomes and did not correlate to corresponding activity in human liver microsomes. In contrast dextromethorphan exhibited a very extensive and rapid metabolism in pig liver microsomes compared to human data. Together with previously determined gene expression data it confirms that CYP2D6 substrates will be very rapidly metabolized in pigs. The mRNA data increased the knowledge of the interindividual variability and the relative expression of different enzymes and transporters in pig intestine and liver. In conclusion, this study has increased the understanding of similarities and differences between pig and human biotransformation of drugs by providing new data for four different model compounds.

    Nyckelord
    Verapamil, Testosterone, Pig pharmacokinetics, Dextromethorphan, In vitro metabolism, Pig mRNA expressison
    Nationell ämneskategori
    Farmaceutisk vetenskap Cell- och molekylärbiologi
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-155943 (URN)10.1016/j.ejps.2011.03.008 (DOI)000291906500001 ()
    Tillgänglig från: 2011-07-04 Skapad: 2011-07-04 Senast uppdaterad: 2012-05-08Bibliografiskt granskad
    3. Binding processes determine the stereoselective intestinal and hepatic extraction of verapamil in vivo
    Öppna denna publikation i ny flik eller fönster >>Binding processes determine the stereoselective intestinal and hepatic extraction of verapamil in vivo
    2012 (Engelska)Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, Vol. 9, nr 11, 3034-3045 s.Artikel i tidskrift (Övrigt vetenskapligt) Published
    Abstract [en]

    The aim of this study was to investigate the mechanisms that might explain the observed route-dependent stereoselective pharmacokinetics (PK) of R/S-verapamil (R/S-VER) following oral and intravenous (iv) administration, by using a novel pig-specific physiologically based pharmacokinetic (PBPK) model suitable for investigations of first-pass extraction in the gut (EG) and the liver (EH). The PBPK model consisted of eight tissue compartments and was designed to simultaneously model the plasma concentration–time (PCT) profiles from three sampling sites after intrajejunal (ij) or iv administration of VER. The PBPK model successfully described the observed PCT profiles and EH over time for R- and S-VER. Extensive tissue binding to gut mucosa, liver, and lungs was an important determinant of the observed PK data. The stereoselective PK of VER was explained by a combination of several processes, including enantioselective plasma protein binding, blood-to-plasma partition, and gut mucosa and liver tissue distribution. The absence of stereoselectivity after iv dosing indicates that the first-pass tissue binding effect is an important factor in determining the steroselective PK of R/S-VER after oral administration. Additionally a combination of extensive liver tissue binding and a metabolite inhibition mechanism explained the time-dependent EH for both R- and S-VER. An in vitroin vivocorrelation of absorption needs to consider these processes because tissue binding may confound analysis of a drug’s biopharmaceutical properties when using classical deconvolution or convolution techniques. In conclusion, a combination of PK data from multiple plasma sampling sites and a PBPK modeling approach provided a mechanistic understanding of processes involved in the intestinal absorption and first-pass extraction ofR- and S-VER.

    Nyckelord
    verapamil, PBPK, hepatic extraction, gut wall extraction, first-pass metabolism
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-165509 (URN)10.1021/mp3000875 (DOI)000313769200008 ()
    Tillgänglig från: 2012-01-09 Skapad: 2012-01-09 Senast uppdaterad: 2013-02-26Bibliografiskt granskad
    4. Extensive intestinal glucuronidation of raloxifene in vivo in pigs and impact for oral drug delivery
    Öppna denna publikation i ny flik eller fönster >>Extensive intestinal glucuronidation of raloxifene in vivo in pigs and impact for oral drug delivery
    2012 (Engelska)Ingår i: Xenobiotica, ISSN 0049-8254, E-ISSN 1366-5928, Vol. 42, nr 9, 917-928 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    1. In this study an advanced multisampling site pig model, with simultaneous venous blood sampling pre- and post liver, was applied to quantify the role of the intestine in relation to the liver in first-pass glucuronidation of raloxifene in vivo. The pharmacokinetic of raloxifene (a BCS/BDDCS class II compound) in humans is characterized by extensive metabolism (>90%) and the major metabolite is the 4'-beta-glucuronide (R-4-G).

    2. Following intra-jejunal (i.j.) single dose administration in pigs raloxifene was metabolized in the gut (E G) during first-pass to more than 70% and a high concentration (AUC(0-6 h) ratio R-4-G/raloxifene >100) of R-4-G was reached in the portal vein. The hepatic extraction (E-H) of raloxifene was similar to 50% and as in humans the bioavailability become low (similar to 7%) in pigs. Interestingly the E-H of raloxifene and R-4-G was time-dependent after i.j. administration.

    3. It is clear that the gut was the dominating organ in first-pass extraction of raloxifene in vivo in pigs. The quantification in this study support earlier human data and emphasize that intestinal glucuronidation should be considered early in the pharmaceutical development.

    Nyckelord
    Raloxifene, glucuronidation, pigs, intestinal metabolism, first-pass metabolism
    Nationell ämneskategori
    Farmaceutisk vetenskap
    Forskningsämne
    Biofarmaci
    Identifikatorer
    urn:nbn:se:uu:diva-165511 (URN)10.3109/00498254.2012.683497 (DOI)000307301900011 ()
    Tillgänglig från: 2012-01-09 Skapad: 2012-01-09 Senast uppdaterad: 2012-09-28Bibliografiskt granskad
  • 16.
    Andersson, Martin
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaci.
    Phase Phenomena in Polymer Networks: Empirical Studies on the Influence of Hydrophobicity, Charge Density and Crosslinks on Macroion-Induced Phase Transitions in Polyelectrolyte Gels2011Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The thesis concerns polyelectrolyte gels in contact with oppositely charged proteins and surfactant micelles, and includes of four papers (I-IV). In paper I confocal Raman spectroscopy was introduced as a method to trace micelles and investigate the structure of gel-surfactant complexes, in phase separated gel spheres. In paper II, the binding of surfactants to microspheres (~50-100 µm) was investigated by means of a micromanipulator-assisted microscopy method. The two surfactants were found to display qualitative difference respect to degree of swelling, surfactant distribution in the gels, and the difference is discussed in terms of absence/presence of hydrophobic attraction to the polyelectrolyte gel network. Kinetics of volume change in gels were analyzed. Aggregation numbers of micelles in polystyrenesulfonate (PSS) solutions, obtained from fluorescence quenching measurements, are presented. In paper III, phase behaviour, protein assembly and diffusion, was studied in PSS gel microspheres. Interpretation of results was aided by measurements of osmotic swelling of individual gel networks, and by combining the results with studies of protein diffusion in macroscopic (cm-sized) gel spheres. Complexes formed were further analyzed with small angle x-ray spectroscopy. In paper IV phase behaviour of mixed ionic/nonionic surfactant micelles is investigated in cm-sized gel spheres. The coexistence of three phases, the formation of dense shells in the bulk of the gels and other phenomena are described for the first time, and the results are presented along with discussion on the charge-density of spherical micelles and of  network induced hysteresis effects in gels. The composition and microstructure of phases are investigated by confocal Raman spectroscopy and small-angle x-ray scattering respectively. The results are interpreted with aid of highly detailed theoretical model calculations.

    Delarbeten
    1. Differences in binding of a cationic surfactant to cross-linked sodium poly(acrylate) and sodium poly(styrene sulfonate) studied by Raman spectroscopy
    Öppna denna publikation i ny flik eller fönster >>Differences in binding of a cationic surfactant to cross-linked sodium poly(acrylate) and sodium poly(styrene sulfonate) studied by Raman spectroscopy
    2005 (Engelska)Ingår i: Langmuir, ISSN 0743-7463, Vol. 21, nr 7, 2761-2765 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Raman spectroscopy has been used to investigate the structure of gel-surfactant complexes. Cross-linked sodium poly(acrylate) and sodium poly(styrene sulfonate) were immersed in solutions of the cationic surfactant dodecyl trimethylammonium bromide. During the deswelling process, two distinct regions could be observed for both types of gels. Looking at the Raman spectra, however, for the poly(styrene sulfonate), the surfactant could be found throughout the gel particle, whereas for poly(acrylate), essentially all the surfactant was bound in a surface layer.

    Nationell ämneskategori
    Oorganisk kemi
    Forskningsämne
    oorganisk kemi
    Identifikatorer
    urn:nbn:se:uu:diva-70227 (URN)10.1021/la0468693 (DOI)15779946 (PubMedID)
    Tillgänglig från: 2008-12-16 Skapad: 2008-12-16 Senast uppdaterad: 2011-02-24
    2. Single microgel particle studies demonstrate the influence of hydrophobic interactions between charged micelles and oppositely charged polyions.
    Öppna denna publikation i ny flik eller fönster >>Single microgel particle studies demonstrate the influence of hydrophobic interactions between charged micelles and oppositely charged polyions.
    2005 (Engelska)Ingår i: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 21, nr 9, 3773-3781 s.Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The binding of two cationic surfactants, dodecyltrimethylammonium bromide (DoTAB) and N-(1,1,2,2-tetrahydroperfluorodecanyl)pyridinium bromide (HFDePB), to covalently cross-linked sodium poly(styrenesulfonate) (PSS) microgels has been investigated by means of micromanipulator-assisted time-resolved light microscopy on single gels. It is demonstrated that repeated measurements on the same microgel under conditions of controlled liquid flow give highly reproducible results. The two surfactants are found to behave very differently with respect to degree of swelling, surfactant distribution in the gels, both during shrinking and at equilibrium, and kinetics of volume changes induced by them. The main difference is attributed to the presence of a hydrophobic interaction between PSS and the DoTAB micelles, absent in the case of HFDePB. Kinetic shrinking curves are recorded and analyzed using a model for steady-state transport of surfactant between the solution and the gels. Aggregation numbers for DoTAB in PSS solutions obtained from fluorescence quenching measurements are presented. A strong dependence on the surfactant-to-polyion concentration ratio is observed. Relations between surfactant binding isotherms, phase diagrams for linear polyelectrolyte/surfactant/water systems, and the binding to gels are discussed.

    Nationell ämneskategori
    Medicin och hälsovetenskap
    Identifikatorer
    urn:nbn:se:uu:diva-92344 (URN)10.1021/la047316v (DOI)15835936 (PubMedID)
    Tillgänglig från: 2004-10-29 Skapad: 2004-10-29 Senast uppdaterad: 2016-04-27Bibliografiskt granskad
    3. Lysozyme Incorporation in Poly(styrenesulfonate) Microspheres: Dynamics of Core/Shell formation
    Öppna denna publikation i ny flik eller fönster >>Lysozyme Incorporation in Poly(styrenesulfonate) Microspheres: Dynamics of Core/Shell formation
    (Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
    Identifikatorer
    urn:nbn:se:uu:diva-145378 (URN)