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Microgels as carriers of antimicrobial peptides – effects of peptide PEGylation
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.ORCID iD: 0000-0001-5626-3959
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.ORCID iD: 0000-0001-5236-9107
Bose Insitute, Kolkata, India.
Indian Insitute of Science, Bangalore, India.
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2019 (English)In: Colloids and Surfaces A: Physicochemical and Engineering Aspects, ISSN 0927-7757, E-ISSN 1873-4359, Vol. 565, p. 8-15Article in journal (Refereed) Published
Abstract [en]

Delivery systems are likely to be central for the translation of antimicrobial peptides (AMPs) towards therapeutics. Addressing AMP interactions with microgel carriers, we here investigate how poly(ethylene glycol) conjugation ('PEGylation') of AMPs affect their loading and release to/from microgels, combining structural studies using nuclear magnetic resonance (NMR) with ellipsometry, circular dichroism spectroscopy (CD), and light scattering. Such studies demonstrate that poly(ethyl acrylate-co-methacrylic acid) microgels bind considerable amounts of the positively charged AMP KYE28 (KYEITTIHNLFRKLTHRLFRRNFGYTLR) and its PEGylated variants KYE28-PEG48, PEG48-KYE28, and PEG24-KYE28-PEG24. Z-potential measurements indicate that KYE28 resides primarily inside the microgel core, and that localization of the PEGylated peptides is shifted towards the microgel corona. Furthermore, while all peptides are disordered in solution, CD measurements report on helix induction on microgel binding, particularly so for the PEGylated peptides. Addressing such conformational changes in more detail, NMR structural studies showed that peptide-microgel interactions are facilitated by a hydrophobic domain formed by the peptide after microgel binding, and with modulating electrostatic/salt bridge interaction between the positively charged peptide residues and negative microgel charges. As the microgels remain negatively charged also at high peptide load, membrane disruption and antimicrobial effects necessitates peptide release, demonstrated to be promoted by PEGylation and high ionic strength. Importantly, microgel loading, as well as peptide localization, conformation, and release, did not depend significantly on PEG conjugation site, but instead seems to be dictated by the PEG content of the peptide conjugates.

Place, publisher, year, edition, pages
2019. Vol. 565, p. 8-15
Keywords [en]
Antimicrobial peptide, Drug delivery, Microgel, NMR, PEGylation
National Category
Pharmaceutical Sciences
Identifiers
URN: urn:nbn:se:uu:diva-361401DOI: 10.1016/j.colsurfa.2018.12.049ISI: 000457062300002OAI: oai:DiVA.org:uu-361401DiVA, id: diva2:1250426
Funder
Swedish Research Council, 2012-1842Swedish Research Council, 2015-06720Available from: 2018-09-24 Created: 2018-09-24 Last updated: 2019-10-12Bibliographically approved
In thesis
1. Microgels as Carriers for Antimicrobial Peptides: Surface-bound microgels, and factors affecting peptide interactions
Open this publication in new window or tab >>Microgels as Carriers for Antimicrobial Peptides: Surface-bound microgels, and factors affecting peptide interactions
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

With a growing number of multi-resistant bacteria against conventional antibiotics, there is an urgent need to identify new antimicrobial therapeutics. One example that has gained considerable interest is antimicrobial peptides (AMPs). For AMPs to reach their full potential as therapeutics, as well as for other peptide and protein drugs, the right drug delivery system may overcome reported shortcomings, such as fast clearance in the bloodstream and proteolytic degradation. Microgels are weakly cross-linked polymer colloids, which can be made responsive to various stimuli. In the context of drug delivery, microgels are of particular interest as carriers for biomacromolecular drugs, such as peptides and proteins, as their water-rich environment offers both protection against enzymatic degradation and triggered release possibilities. Combining these, the aim of this thesis was to investigate electrostatically triggered surface-bound microgels as a delivery system for AMPs, as well as evaluate such systems as an antimicrobial and anti-inflammatory coating for biomaterials.

Results presented in this thesis demonstrate effects of microgel charge density, pH, and ionic strength on microgel volume transitions at solid interfaces, surface-induced microgel deformation and nanomechanical properties. In addition, effects of both microgel properties (charge density) and peptide properties (molecular weight, charge density, and posttranslational modifications) on peptide loading and release from surface-bound microgels were investigated. The presented thesis also reports in vitro studies of AMP-loaded microgels in dispersion and surface-bound, as either mono- or multilayers. Notably, the interplay between surface- and release-related effects for the antimicrobial properties of AMP-loaded microgels are investigated. In addition, anti-inflammatory properties of AMP-loaded microgels are also reported.

Taken together, microgels prove an interesting and versatile drug delivery system for AMPs. Results obtained in this thesis have demonstrated that several key factors need to be taken into consideration in the development of surface-bound microgels as a carrier for AMPs, and that small changes in microgel and peptide properties can alter peptide loading and release profiles.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2018. p. 66
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 259
Keywords
Antimicrobial peptides, Biomaterial coating, Drug delivery, Host defence peptides, Microgels, pH-responsive, Surface-bound
National Category
Pharmaceutical Sciences Biomaterials Science Physical Chemistry
Research subject
Pharmaceutical Physical Chemistry
Identifiers
urn:nbn:se:uu:diva-360241 (URN)978-91-513-0473-1 (ISBN)
Public defence
2018-11-30, A1:111a, BMC, Husargatan 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2018-11-09 Created: 2018-10-12 Last updated: 2018-11-19
2. Polymeric Nanoparticles as Carriers for Antimicrobial Peptides: Factors Affecting Peptide and Membrane Interactions
Open this publication in new window or tab >>Polymeric Nanoparticles as Carriers for Antimicrobial Peptides: Factors Affecting Peptide and Membrane Interactions
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

As resistance towards conventional antibiotics is becoming more pronounced, cationic antimicrobial peptides (AMPs) have received considerable attention as possible therapeutic alternatives. Thousands of potent AMPs occur in humans, animals, plants and fungi as a natural part of the immune system. However, there are several challenges with AMP therapeutics related to formulation and delivery. Examples include proteolytic sensitivity and serum protein binding, resulting in quick degradation, loss of activity and clearance. Therefore, it is important to find a suitable drug delivery system to meet these protection and delivery challenges. Micro-/nanogels are loosely crosslinked polymer colloids with high water content that can be made to trigger at a wide range of stimuli. They have shown promise as delivery systems for AMPs, as the aqueous environment they create allows the peptides to maintain their natural conformation, while their gel networks offer protection and triggered release. This thesis aims towards expanding the knowledge about degradable and non-degradable pH-responsive micro-/nanogels as carriers for AMPs.

The results in this thesis show that factors relating to the drug delivery system (degradability, charge and crosslinker density), the surrounding media (pH and ionic strength) and the peptide properties (length, charge, PEGylation) all affect the peptide loading to, protection, release from and effect of AMP-loaded gels. Studies of the interaction of AMP-loaded microgels with bacteria-modelling liposomes and lipid bilayers have verified peptide effect after gel incorporation, as further demonstrated by in vitro studies on several bacterial strains. Neutron reflectometry provided detailed mechanistic information on the interaction between AMP-loaded gels and bacteria-modelling lipid bilayers, showing that the antimicrobial unit is the released peptide. All gels showed low, promising hemolysis and some gels could offer protection against proteolytic degradation of AMPs.

In summary, non-degradable and degradable micro-/nanogels are versatile and interesting candidates as AMP carriers. Small changes in the gel composition or the AMP used can dramatically change the peptide loading, release and effect. It is therefore necessary to carefully consider and evaluate the optimal carrier for every AMP and the application at hand.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2019. p. 74
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 280
Keywords
antimicrobial peptide, microgel, degradable, nanogel, drug delivery, PEGylation, secondary structure, model membrane, lipid bilayer, neutron reflectometry, ellipsometry
National Category
Pharmaceutical Sciences Physical Chemistry
Research subject
Pharmaceutical Physical Chemistry
Identifiers
urn:nbn:se:uu:diva-383639 (URN)978-91-513-0778-7 (ISBN)
Public defence
2019-11-29, Room A1:107a, BMC, Husargatan 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2019-11-07 Created: 2019-10-12 Last updated: 2019-11-27

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Nordström, RandiNyström, LinaMalmsten, Martin

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