Selectively Antibacterial Silver-Gold Alloy Nanoparticles Conjugated with Target Specific Aptamer Sequences

• Applicants: Professor Dr.-Ing. Stephan Barcikowski; Professorin Dr. Meike Stiesch
• Subject Area: Biomaterials ; Synthesis and Properties of Functional Materials
• Term: from 2018 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 356685838

• Final Report Year: 2024

Final Report Abstract

Implant-associated infections are a challenge in medicine and arise from bacteria that organize themselves into resistant biofilms. Limitations of current therapeutic approaches are that their effect on bacteria is also associated with a risk to human cells. Therefore, there is a need for antibacterial strategies with an extended therapeutic window. For this purpose, this project developed silver-gold alloy nanoparticle (AgAu NP)-aptamer conjugates designed to I) release antimicrobial silver ions, and II) bind to pathogenic bacteria in biofilm via a specific aptamer. A size-controlled synthesis of AgAu NP could be achieved by laser ablation in liquids. Alloy particle surface composition was characterized in-depth by XPS and cyclic voltammetry. A protocol for the conjugation of thiolated aptamers to AgAu NP and in vitro functional assays for the conjugates was developed. The maximum functionality was hardly affected by the Ag:Au ratio in the particle, but selective binding of the conjugates to the target protein A, predicted in the literature, could not be reproduced. The study of ion release in gel phantoms showed a significant reduction in the presence of the aptamer ligands. Collaborative single-particle hyperspectroscopy studies with Mercator Fellow Stephan Link established trends for composition-dependent particle dissolution and derived a two-step model for the dissolution kinetics of AgAu NP. Biological analysis of AgAu NP and corresponding conjugates showed antibacterial effects against Staphylococcus aureus and Porphyromonas gingivalis. Detailed studies demonstrated a dependence of the effect on nanoparticle concentration and bacterial growth phase, especially biofilm formation. By applying activity assays, membrane staining, and gene expression analysis, bacterial metabolism was identified as the major target of AgAu NP. The nanoparticles also exhibited cytotoxic effects against human fibroblasts and keratinocytes, which, however, were attenuated when cells and bacteria were cocultured. Overall, this project provided fundamental chemical and biological insights into AgAu NP as antibiotic alternatives and identified key approaches for further development toward clinical application.

Publications

• How does nano-silver get inside bacteria? Mechanistic studies using AgAu alloy nanoparticles. Abstracts of Papers of the American Chemical Society 2019, 257. Conference Talk
  Rehbock, C.; Streich, C.; Jakobi, J.; Grade, S.; Kuhnel, M.; Migunov, V.; Knura, T.; Sures, B.; Stiesch, M. & Barcikowski, S.
  
• Selective Aerobic Oxidation of 5‐(Hydroxymethyl)furfural over Heterogeneous Silver‐Gold Nanoparticle Catalysts. Advanced Synthesis & Catalysis, 362(24), 5681-5696.
  Schade, Oliver R.; Stein, Frederic; Reichenberger, Sven; Gaur, Abhijeet; Saraҫi, Erisa; Barcikowski, Stephan & Grunwaldt, Jan‐Dierk
  
• Antibacterial properties of AgAu-nanoparticles produced by laser ablation. Annual Meeting of the German Society for Biomaterials, 2021. Conference Poster
  Heine N., Stein F., Doll K., Winkel A., Rehbock C., Barcikowski S. & Stiesch M.
  
• Influence of Gold/Silver Ratio in Ablative Nanoparticles on Their Interaction with Aptamers and Functionality of the Obtained Conjugates. Bioconjugate Chemistry, 32(11), 2439-2446.
  Stein, Frederic; Schielke, Andreas; Barcikowski, Stephan & Rehbock, Christoph
  
• Single-Particle Hyperspectral Imaging Reveals Kinetics of Silver Ion Leaching from Alloy Nanoparticles. ACS Nano, 15(5), 8363-8375.
  Al-Zubeidi, Alexander; Stein, Frederic; Flatebo, Charlotte; Rehbock, Christoph; Hosseini, Jebeli Seyyed Ali; Landes, Christy F.; Barcikowski, Stephan & Link, Stephan
  
• Antibacterial and cytotoxic properties of AgAu-nanoparticles produced by laser ablation in a co-culture model. Annual Meeting of the German Society for Biomaterials, 2022. Conference Poster and Talk
  Heine N., Stein F., Kheirmand-Parizi M., Doll K., Winkel A., Rehbock C., Barcikowski S. & Stiesch M.
  
• Anti-Biofilm Properties of Laser-Synthesized, Ultrapure Silver-Gold-Alloy Nanoparticles against Staphylococcus aureus. Research Square Platform LLC.
  Heine, Nils; Doll-Nikutta, Katharina; Stein, Frederic; Jakobi, Jurij; Ingendoh-Tsakmakidis, Alexandra; Rehbock, Christoph; Winkel, Andreas; Barcikowski, Stephan & Stiesch, Meike
  
• Bacteria‐Epithelial Cell Interaction Influences Cytotoxicity and Antibacterial Effect of Silver‐Gold Alloy Nanoparticles on a Species‐Specific Level. ChemNanoMat, 10(2).
  Doll‐Nikutta, Katharina; Heine, Nils; Kheirmand‐Parizi, Marjan; Stein, Frederic; Ulrich, Jennifer; Rehbock, Christoph; Winkel, Andreas; Barcikowski, Stephan & Stiesch, Meike
  
• Disproportional surface segregation in ligand-free gold–silver alloy solid solution nanoparticles, and its implication for catalysis and biomedicine. Faraday Discussions, 242(2023), 301-325.
  Stein, Frederic; Kohsakowski, Sebastian; Martinez-Hincapie, Ricardo; Reichenberger, Sven; Rehbock, Christoph; Colic, Viktor; Guay, Daniel & Barcikowski, Stephan
  
• The Origin of the Intracellular Silver in Bacteria: A Comprehensive Study using Targeting Gold–Silver Alloy Nanoparticles. Advanced Healthcare Materials, 12(30).
  Streich, Carmen; Stein, Frederic; Jakobi, Jurij; Ingendoh‐Tsakmakidis, Alexandra; Heine, Nils; Rehbock, Christoph; Winkel, Andreas; Grade, Sebastian; Kühnel, Mark; Migunov, Vadim; Kovács, András; Knura, Thomas; Stiesch, Meike; Sures, Bernd & Barcikowski, Stephan