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  • 1. Geh, Katharina J.
    et al.
    Hubert, Madlen
    Department of Pharmacy, Pharmaceutical Technology and Biopharmaceutics, Ludwig-Maximilians Universität München, Munich, Germany.
    Winter, Gerhard
    Optimisation of one-step desolvation and scale-up of gelatine nanoparticle production2016Ingår i: Journal of Microencapsulation, ISSN 0265-2048, E-ISSN 1464-5246, Vol. 33, nr 7, s. 595-604Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Gelatine nanoparticles (GNPs) are biodegradable and biocompatible drug delivery systems with excellent clinical performances. A two-step desolvation is commonly used for their preparation, although this methodology has several shortcomings: lack of reproducibility, small scales and low yields. A straightforward and more consistent GNP preparation approach is presented here focusing on the development of a one-step desolvation with the use of a commercially available gelatine type. Controlled stirring conditions and ultrafiltration are used to achieve large-scale production of nanoparticles of up to 2.6g per batch. Particle size distributions are conserved and comparable to those determined for two-step desolvation on small scale. Additionally, a range of cross-linking agents is examined for their effectiveness in stabilising GNPs as an alternative to glutaraldehyde. Glyceraldehyde demonstrated outstanding properties, which led to high colloidal stability. This approach optimises the manufacturing process and the scale-up of the production capacity, providing a clear potential for future applications.

  • 2. Geh, Katharina J.
    et al.
    Hubert, Madlen
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Winter, Gerhard
    Progress in formulation development and sterilisation of freeze-dried oligodeoxynucleotide-loaded gelatine nanoparticles2018Ingår i: European journal of pharmaceutics and biopharmaceutics, ISSN 0939-6411, E-ISSN 1873-3441, Vol. 129, s. 10-20Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Oligodeoxynucleotide (ODN)-loaded gelatine nanoparticles (GNPs) have proven their outstanding potential in the treatment of allergic diseases such as equine asthma and canine atopic dermatitis, which are appropriate models for the corresponding human diseases. To encourage the development of a marketable product, long term stability and sterility needs to be ensured. In this work, we aimed to advance freeze-drying options to stabilise ODN-loaded GNPs. Matrix-assisted laser desorption/ionisation mass spectrometry time-of-flight was implemented as a versatile tool to assess ODN stability. With this method long-term storage stability of lyophilised ODN-loaded GNPs formulated in sucrose or trehalose was achieved. Controlled nucleation was further introduced to optimise the lyophilisation approach. This allowed shortening of the process in comparison to standard freeze-drying procedures. Particle sizes, polydispersity indices, ODN stability, residual moisture and glass transition temperature were maintained upon storage. Excipient portfolio was enlarged by novel amino acid containing formulations for lyophilisates. His emerged as an excellent excipient in stabilising lyophilised ODN-loaded GNPs, whereas addition of Arg and Gly revealed to be inadequate at accelerated conditions. Lastly, gamma irradiation was evaluated as a suitable sterilisation method of ODN-loaded GNPs.

  • 3.
    Holst, Mikkel Roland
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Vidal-Quadras, Maite
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Larsson, Elin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Song, Jie
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Hubert, Madlen
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Blomberg, Jeanette
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lundborg, Magnus
    Landström, Maréne
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB). Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Clathrin-Independent Endocytosis Suppresses Cancer Cell Blebbing and Invasion2017Ingår i: Cell reports, ISSN 2211-1247, E-ISSN 2211-1247, Vol. 20, nr 8, s. 1893-1905Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Cellular blebbing, caused by local alterations in cellsurface tension, has been shown to increase the invasiveness of cancer cells. However, the regulatory mechanisms balancing cell-surface dynamics and bleb formation remain elusive. Here, we show that an acute reduction in cell volume activates clathrinindependent endocytosis. Hence, a decrease in surface tension is buffered by the internalization of the plasma membrane (PM) lipid bilayer. Membrane invagination and endocytosis are driven by the tension- mediated recruitment of the membrane sculpting and GTPase-activating protein GRAF1 (GTPase regulator associated with focal adhesion kinase-1) to the PM. Disruption of this regulation by depleting cells of GRAF1 or mutating key phosphatidylinositol- interacting amino acids in the protein results in increased cellular blebbing and promotes the 3D motility of cancer cells. Our data support a role for clathrin-independent endocytic machinery in balancing membrane tension, which clarifies the previously reported role of GRAF1 as a tumor suppressor.

  • 4.
    Hubert, Madlen
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Larsson, Elin
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Caveolae dynamics is strongly influenced by the lipid composition of the plasma membrane2017Ingår i: European Biophysics Journal, ISSN 0175-7571, E-ISSN 1432-1017, Vol. 46, s. S121-S121Artikel i tidskrift (Övrigt vetenskapligt)
  • 5.
    Mohan, Jagan
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Hubert, Madlen
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Larsson, Elin
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Schweiger, C
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Structural arrangement of membrane-bound cavinManuskript (preprint) (Övrigt vetenskapligt)
  • 6.
    Moodie, Lindon W. K.
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hubert, Madlen
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Zhou, Xin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Albers, Michael Franz
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Wanrooij, Sjoerd
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Hedberg, Christian
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Photoactivated Colibactin Probes Induce Cellular DNA Damage2019Ingår i: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 58, nr 5, s. 1417-1421Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Colibactin is a small molecule produced by certain bacterial species of the human microbiota that harbour the pks genomic island. Pks(+) bacteria induce a genotoxic phenotype in eukaryotic cells and have been linked with colorectal cancer progression. Colibactin is produced in a benign, prodrug form which, prior to export, is enzymatically matured by the producing bacteria to its active form. Although the complete structure of colibactin has not been determined, key structural features have been described including an electrophilic cyclopropane motif, which is believed to alkylate DNA. To investigate the influence of the putative "warhead" and the prodrug strategy on genotoxicity, a series of photolabile colibactin probes were prepared that upon irradiation induced a pks(+) like phenotype in HeLa cells. Furthermore, results from DNA cross-linking and imaging studies of clickable analogues enforce the hypothesis that colibactin effects its genotoxicity by directly targeting DNA.

  • 7. Rodrigues, Leticia
    et al.
    Kyriakos, Konstantinos
    Schneider, Fabian
    Dietz, Hendrik
    Winter, Gerhard
    Papadakis, Christine M.
    Hubert, Madlen
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB). Ludwig Maximilians Univ Munchen, Dept Pharm Pharmaceut Technol & Biopharm, Munich, Germany.
    Characterization of Lipid-Based Hexosomes as Versatile Vaccine Carriers2016Ingår i: Molecular Pharmaceutics, ISSN 1543-8384, E-ISSN 1543-8392, Vol. 13, nr 11, s. 3945-3954Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Subunit vaccines typically show insufficient immunogenicity. To address this issue, we developed a novel self-adjuvanting particulate carrier system based upon the lipids phytantriol (Phy) and mannide monooleate (MaMo). Phy is a lipid known to form nonlamellar phases in fully hydrated systems, whereas MaMo has been found to promote immune responses in emulsion form. A bulk phase composition of Phy/MaMo (14 wt %) showed hexagonal (HII) phase behavior over a practical temperature range (including room and body temperature), and was therefore used for particle development. Hexosomes stabilized with different concentrations of either poloxamer 407, Myrj 59, or Pluronic F108 were successfully prepared. To demonstrate the versatile nature of these systems, the particles were further modified with either positively or negatively charged lipids and loaded with model antigens, while maintaining the HII structure. These hexosomes are structurally robust and amenable to customization, rendering them suitable as antigen delivery carriers.

  • 8. Rodrigues, Leticia
    et al.
    Schneider, Fabian
    Zhang, Xiaohan
    Larsson, Elin
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Moodie, Lindon W. K.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Dietz, Hendrik
    Papadakis, Christine M.
    Winter, Gerhard
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Hubert, Madlen
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Cellular uptake of self-assembled phytantriol-based hexosomes is independent of major endocytic machineries2019Ingår i: Journal of Colloid and Interface Science, ISSN 0021-9797, E-ISSN 1095-7103, Vol. 553, s. 820-833Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Despite increasing interests in non-lamellar liquid crystalline dispersions, such as hexosomes, for drug delivery, little is known about their interactions with cells and mechanism of cell entry. Here we examine the cellular uptake of hexosomes based on phytantriol and mannide monooleate by HeLa cells using live cell microscopy in comparison to conventional liposomes. To investigate the importance of specific endocytosis pathways upon particle internalization, we silenced regulatory proteins of major endocytosis pathways using short interfering RNA. While endocytosis plays a significant role in liposome internalization, hexosomes are not taken up via endocytosis but through a mechanism that is dependent on cell membrane tension. Biophysical studies using biomembrane models highlighted that hexosomes have a high affinity for membranes and an ability to disrupt lipid layers. Our data suggest that direct biomechanical interactions of hexosomes with membrane lipids play a crucial role and that the unique morphology of hexosomes is vital for their membrane activity. Based on these results, we propose a mechanism, where hexosomes destabilize the bilayer, allowing them to "phase through" the membrane. Understanding parameters that influence the uptake of hexosomes is critical to establish them as carrier systems that can potentially deliver therapeutics efficiently to intracellular sites of action.

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