Chemical and Biomolecular Engineering - Theses
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ItemEnzyme-responsive nanomaterials for the delivery of antimicrobial peptides( 2023-10)The rate of resistance to antibiotics that are commonly used in the clinic is escalating rapidly, surpassing the introduction of new antimicrobial drugs. To address this problem, alternative strategies are being explored, such as the re-evaluation of antibiotics, that have not yet gained widespread clinical application. Antimicrobial peptides (AMPs) represent one of those antibiotics, offering remarkable antimicrobial efficacy against various pathogens. However, in clinical settings, AMPs are typically considered a last-resort option due to their off-target effects and poor stability in-vivo resulting from their cationic and amphiphilic peptide nature. Therefore, most of current strategies addressing these limitations focus primarily on the control and shielding of the cationic charge and the amphiphilic nature of AMPs. These can potentially be achieved through encapsulation of AMPs inside stimuli-responsive polyelectrolyte complexes (PECs) by combining the cationic drug with anionic polyelectrolytes. Stimuli-responsive polymers can be employed as encapsulation materials in PECs to design systems that activate drug release in response to specific changes encountered during microbial infection, such as variations in pH, enzyme activity, or temperature. The overarching aim of this Thesis was to explore the creation of PECs capable of encapsulating the clinically approved antimicrobial peptide, Polymyxin B and its subsequent enzyme-induced release. In Chapters 2 and 3, the aims were: Firstly, to synthesize anionic and helical polymers incorporating enzyme-degradable peptide side chains (Aim 1.1), followed by evaluation of the degradation properties of these polymers in response to the enzyme released by gram-negative bacterium Pseudomonas aeruginosa (Aim 1.2). In Chapter 4, the first objective (Aim 1.3) was to assemble Polymyxin B and the anionic enzyme-degradable polymers into PECs. The next objective (Aim 1.4 ) involved investigating the P. aeruginosa-induced drug release from these PECs, while Aim 1.5 focused on assessing the antimicrobial activity of the developed PECs against P. aeruginosa strains. Chapter 1 provides a review of the current developments in the field of the stimuli- responsive delivery of AMPs using polyelectrolyte complexes. Chapters 2 and 3 discuss the synthesis of polymers with poly(methacrylamide) and poly(acetylene) backbones respectively coupled to enzyme-degradable peptide side chains. Of the synthesized materials, two poly(methacrylamide) polymers from Chapter 2 were found to be the most effective in terms of degradation by the enzyme released by P.aeruginosa, while none of the synthesized acetylene-containing peptides from Chapter 3 polymerized. Subsequently, the poly(methacrylamide) polymer with the highest multivalency was used to form PECs with Polymyxin B in Chapter 4. Eight different formulations of PECs were created, with one being the most optimized in terms of encapsulation efficacy and physiological stability. The stability of the particles was further improved by the addition of Tannic Acid, which acted as a protective coating and a cross-linker. The Thesis then evaluated the ability of the PECs to release Polymyxin B under enzymatic degradation. Finally, the preliminary evaluation of the antimicrobial activity of the PECs against various P.aeruginosa strains were presented.
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ItemStructurally nanoengineered peptide polymers for combating multidrug-resistant bacteria( 2016)Antimicrobial resistance has been named as one of the clinical ‘super-challenges’ of the 21st century. With the rise and prevalence of multi-drug resistant (MDR) ‘superbugs’, such as methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), and more recently, the ‘ESKAPE’ pathogens, a return to the pre-antibiotic era is rapidly becoming a reality in many parts of the world. Infections caused by MDR pathogens are a major burden to modern healthcare, as the available treatment options are drastically reduced, leading to increased treatment costs, and high morbidity and mortality rates. However, this growing epidemic of infections caused by MDR pathogens has not been accompanied by an increase in the discovery of novel antimicrobials. In fact, it has been reported that aside from a few narrow spectrum drugs and teixobactin, no new chemical class of antibiotics has appeared in the last 40 years. The challenge remains to develop antimicrobial agents with new mechanisms of action that can overcome acquired resistance without contributing to resistance development. Over the past few years, our work in drug and gene delivery has demonstrated the potential of star peptide polymers as therapeutic agents, with significant advantages over linear peptides. Building on our prior knowledge, this thesis explores the possibility of using macromolecular engineering techniques to design and develop star peptide polymers that could function as novel antimicrobial agents capable of combating antibiotic-resistant bacteria. Inspired by antimicrobial peptides (AMPs) that form part of the innate immune response in multicellular organisms, 16-and 32-arm star peptide polymers were synthesized, with arms composed of cationic and hydrophobic amino acid moieties co-polymerized in a random fashion. The star polymers were found to demonstrate superior efficacy against clinically-relevant Gram-negative bacteria, including MDR species, compared to their linear ‘one arm’ equivalent and several well-known AMPs, while possessing high therapeutic indices. The lack of any observable bacterial resistance development against the star peptide polymers was attributed to their unique, multi-modal antimicrobial mechanism, which differs from that of antibiotics and AMPs. Based on these attributes, the star peptide polymers were classified as a new class of antimicrobial agents, referred to as ‘Structurally Nanoengineered Antimicrobial Peptide Polymers’ (SNAPPs). The subsequent part of this thesis focused on developing further understanding on the structural design and bio-nano interactions of SNAPPs. Through a structure-activity relationship study, the effects of the star arm (co)polymer structure and overall macromolecular architecture on antimicrobial activity and biocompatibility were investigated. Further, the behavior of SNAPPs in physiologically-relevant settings, such as in a bloodstream-mimicking environment, was probed in terms of their antimicrobial efficacy and mode of action. Lastly, this thesis also demonstrated the ability of SNAPPs to synergize with different classes of antibiotics, potentially offering a method to ‘revive’ antibiotics that have already been deemed ineffective. Collectively, the discovery and development of SNAPPs, as presented in this thesis, represents a breakthrough in the fight against infections caused by antibiotic-resistant bacteria and, hence, a significant advancement in the field of antimicrobial research. This thesis also provides fundamental understanding on the properties and performance of SNAPPs, which will be useful for the optimization of next-generation SNAPPs with enhanced antimicrobial performance and minimal toxic side-effects in vivo. It is thus with hope that this thesis will not only serve as a platform for the development of SNAPPs for actual clinical use, but also act as a stimulus for other researchers in the pursuit of innovative and more effective treatment methods against ‘superbugs’.