Polymyxins are clinically used antibiotics, discovered in mid-20th century. Once abandoned due to excessive nephrotoxicity, they are now used increasingly to address infections caused by multi-drug... Show morePolymyxins are clinically used antibiotics, discovered in mid-20th century. Once abandoned due to excessive nephrotoxicity, they are now used increasingly to address infections caused by multi-drug resistant Gram-negative bacteria.In this thesis, we describe the development and synthesis of analogues of polymyxin, aimed at reducing its associated nephrotoxicity. Analogues were made by semisynthesis, with modifications introduced mostly in the exocyclic portion of the molecule. Especially the introduction of a disulfide bond within the linked lipid helped in reducing the toxicity of the molecules, as evidenced by testing on proximal tubule epithelial cells. For most potent analogues, the antimicrobial activity was completely retained.In addition, this thesis describes studies on the mechanism of action of polymyxin, mostly based on the full stereoisomer of polymyxin B4. This analogue lacks antimicrobial activity, indicating its original stereochemistry to be of utmost importance for its use as an antibiotic.Hybrids based on polymyxin B derivatives are described, addressing non-conventional targets. A hybrid with vancomycin (typically active on Gram-positive bacteria only) shows activity on Gram-negative bacteria. A polymyxin-based hybrid coupled to a peptide with a beta-hairpin motif addresses Gram-negative bacteria, presumably by binding to outer membrane protein BamA. Show less
Global healthcare is on the verge of an antibiotic availability crisis as bacteria have evolved resistance to nearly all known antibacterials. Identifying new antibiotics that operate via novel... Show moreGlobal healthcare is on the verge of an antibiotic availability crisis as bacteria have evolved resistance to nearly all known antibacterials. Identifying new antibiotics that operate via novel modes-of-action is therefore of high priority.This thesis contains two drug discovery projects, originating from a antibacterial screen of a compound library. In both projects chemical hits are first structurally optimized, after which their mode-of-action is determined.The first project entails optimizing a hit with potency against MRSA into a submicromolar active antibiotic. By using a chemical proteomics approach, the targets of this compound were elucidated, along with the targets that are most important in its antibacterial activity.The second project concerns Gram-negative bacteria, where a hit molecule is optimized into the conformationally restricted LEI-800. The target of LEI-800 is found to be DNA gyrase, a common antibiotic target. However, it is that LEI-800 inhibits DNA gyrase differently, and more potently, than the status quo. Show less
Extending our current arsenal of antibiotics is key to staying ahead in the arms race between humans and resistant bacteria. Classes of antibiotics otherwise limited to the treatment of Gram... Show moreExtending our current arsenal of antibiotics is key to staying ahead in the arms race between humans and resistant bacteria. Classes of antibiotics otherwise limited to the treatment of Gram-positive pathogens may be potentiated against Gram-negative bacteria by disruption of their outer membrane. The work described in this thesis focuses on the development of novel synergists designed to selectively disrupt the outer membrane and in doing potentiate the activity of antibiotics that are otherwise inactive against Gram-negative bacteria. Show less
The research described in this thesis focussed on the role of apolipoproteins in lipid metabolism, inflammation and bacterial sepsis, with specific emphasis on apoCI. From studies in human APOC1_... Show moreThe research described in this thesis focussed on the role of apolipoproteins in lipid metabolism, inflammation and bacterial sepsis, with specific emphasis on apoCI. From studies in human APOC1_-transgenic and apoc1-/- mice, we were able to identify apoCI as a potent inhibitor of triglyceride hydrolysis by inhibiting lipoprotein lipase. Since APOC1 mice have thus increased VLDL levels, and VLDL protects against bacterial infection, we studied whether apoCI could play a role in inflammation and infection. We found that apoCI was able to bind lipopolysaccharide (LPS), the main toxic component of Gram-negative bacteria. Interestingly, although other apolipoproteins which have been studied have anti-inflammatory properties, we found that apoCI is a pro-inflammatory protein. By enhancing the biological response towards LPS and Gram-negative bacteria, apoCI dose-dependently improved the anti-bacterial attack, and protected against intrapulmonal Klebsiella pneumoniae-induced sepsis. Consistent with these experimental findings we also found that subjects with high plasma apoCI levels were less prone to infection-related mortality during follow-up, independent of plasma lipid levels. Likewise, survivors of severe sepsis showed higher plasma apoCI levels as compared to non-survivors, again independent of lipid levels. Taken together, our findings indicate that apoCI is an important determinant of the inflammatory response in mice and humans. Show less
