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Polymer Conjugated Cyclic Peptide Nanotubes as Next Generation Antibiotics

Principal Supervisor: Prof Sébastien Perrier - Department of Chemistry

Co-supervisor: Prof Chris Dowson - SLS

PhD project title: Polymer Conjugated Cyclic Peptide Nanotubes as Next Generation Antibiotics

University of Registration: University of Warwick

Project outline:

Antimicrobial peptides (AMP) are effective antibacterial agents which target the phospholipid membranes of bacteria. The key of their activity resides in an optimal amphiphilic balance, that enables them to insert into the negatively charged bacterial membrane, which ultimately leads to pore formation, membrane disruption and bacterial cell death. Unfortunately, bacteria have developed resistance to AMPs by modifying their membrane chemistry (charge, hydrophobicity, fluidity, membrane proteins), or by increasing the secretion of proteases that degrade AMP, thus lowering their effect and limiting their applications.1

Cylic peptide nanotubes (CPNT) are a special class of AMPs which have shown great promises as anti-bacterial agents.2 CPNT have the potential to overcome bacterial resistances, as they can easily be modified to address changes in the physical properties of the bacterial membrane, and are less sensitive to proteolytic cleavage due to their cyclic alternating D-/L-structure. However, their high tendency to aggregate and low selectivity between bacterial and mammalian cells has hampered their use as antibiotics.

Recently, we have shown that by attaching a hydrophilic polymer shell to CPNT, aggregation can be prevented to form water soluble tubes, compatible with biological systems, which do not interfere with bacterial membranes.3 Therefore, by introducing a cleavable linker between the polymeric shell and CPNT it is possible to prevent the interaction of the peptide with bacterial membrane until the polymer is released, and the peptide activity is switched on, thus achieving controlled toxicity. Non-aggregating polymer-CPNT conjugates should show low overall biological activity, while cleavage of the polymeric shell should result in biologically active systems. Furthermore, by mixing various types of peptides polymer conjugates it is possible to easily generate libraries of CPNT with a range of physical properties, thus increasing the difficulty for bacteria to develop resistance.

We aim to test the effect of CPNTs in different bacterial strains, and achieve trigger controlled toxicity upon cleavage of the polymer shell. A small library of CPNT varying the amphiphilic balance will be synthesized, based on previously reported antibacterial sequences.4 Biocompatible polymers showing stealth behavior (Poly(2-ethyl-2-oxazoline) will be attached using stimuli responsive linkers (i.e. disulfide bonds). Both, non-conjugated CP (pure and mixed) as well as polymer conjugates are to be tested regarding their toxicity towards mammalian cells and different bacteria strains. Their membrane activity will be assessed using bacterial membrane models (phospholipid vesicles).

The project should allow us to demonstrate the activation of polymer-CPNT systems upon cleavage of the polymer shell, and to explore libraries of CPNT formed in situ as a strategy to overcome drug resistance. The successful outcome of the project will lead to the design of selective systems that are effective against specific strains of resistant bacteria; i.e. by generating membrane active CPNT upon activation by proteases secreted by bacteria like pseudomonas aeruginosa (pseudolysin).

References:

  1. K.A. Brogden, Nature Rev. Microbiol. 2005, 3, 238-250.
  2. Ghadiri et al., Nature 2001, 412, 452-455.
  3. B. S. Perrier et al. Chem. Eur. J. 2014, 20, 12745-12749.
  4. Ghadiri et al., Chem. Eur. J. 2007, 13, 4008-4013.

BBSRC Strategic Research Priority: Molecules, cells and systems

Techniques that will be undertaken during the project:

The project will combine materials, chemical and biological techniques:

  • Polymer and nanoparticle synthesis
  • Solid-phase peptide synthesis
  • Labelling of nanoparticles
  • Bioconjugation techniques
  • Polymer / nanoparticles characterization techniques: GPC, SLS and DLS, SAXS, SANS, TEM, SEM, DCS.
  • Small molecules characterization techniques: NMR, IR, UV, Fluorescence.
  • Maintenance of several human cancer cell lines
  • MIC and Haemolysis
  • Study of materials interaction with bacteria

Contact: Prof Sébastien Perrier, University of Warwick