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Title: Design, Synthesis and Evaluation of Aminoacyl-tRNA Synthetase Inhibitors Based on Microcin C and Albomycin
Other Titles: Design, synthese en evaluatie van aminoacyl-tRNA synthetase inhibitoren gebaseerd op microcin C en albomycin
Authors: Vondenhoff, Gaston; S0021311
Issue Date: 2-Apr-2012
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UnhideWhenUsed="false" Name="Colorful Grid Accent 6"/> UnhideWhenUsed="false" QFormat="true" Name="Subtle Emphasis"/> UnhideWhenUsed="false" QFormat="true" Name="Intense Emphasis"/> UnhideWhenUsed="false" QFormat="true" Name="Subtle Reference"/> UnhideWhenUsed="false" QFormat="true"Name="Intense Reference"/> UnhideWhenUsed="false" QFormat="true" Name="Book Title"/> /* Style Definitions */ table.MsoNormalTable {mso-style-name:Standaardtabel; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}Summary mso-ansi-language:EN-US" lang="EN-US"> justify;line-height:150%">mso-ansi-language:EN-GB;mso-fareast-language:NL" lang="EN-GB">This dissertation developedfrom an initial interest in extending the McC concept to a series of potentantimicrobial compounds that could target a variety of aaRSs (Ch.2). Along theway, several new side projects wereinvestigated that evolved in response tofurther optimize drug-likeness (Ch. 3) and selectivity for the bacterial enzyme(Ch. 5). We alsoundertook several attempts to prevent possible resistancemechanismsto potential antibiotics that might result from this work (Ch. 4). justify;line-height:150%">mso-ansi-language:EN-GB" lang="EN-GB">Increasing resistance to antibiotics is a majorproblem worldwide and provides the stimulus for development of new bacterialinhibitors with preferably different modes of action. In search for new leads,several new bacterial targets are being exploited beside the use oftraditionalscreening methods. Hereto, inhibition of bacterial protein synthesis is along-standing validated target. Aminoacyl-tRNA synthetases (aaRSs) play anindispensable role in protein synthesis and their structures proved quiteconserved in prokaryotes and eukaryotes.However, some divergence has occurredallowing the development of selective aaRS inhibitors. Following an outline onthe action mechanismof aaRSs, an overview has been given of already existingaaRS inhibitors, which are largely based on mimics of the aminoacyl-adenylates,the natural reaction intermediates. This is followed by a discussion on morerecent developments in the field and the bioavailability problemand theso-called Trojan horse inhibitors, as based on microcin C (McC) and thesideromycins. justify;line-height:150%">mso-ansi-language:EN-GB" lang="EN-GB">Microcin C (21 )is a potent antibacterial compound produced by some normal">E. coli strains. McC functions through a Trojan horse mechanism: itis actively taken up inside a sensitive cell through the function of theYejABEF transporter and then processed by cellular aminopeptidases. ProcessedMcC (22 ) is a non-hydrolyzableaspartyl-adenylate analogue that inhibits aspartyl-tRNA synthetase (AspRS). Anew synthesis is described that allows for the production of a wide variety ofMcC analogues in acceptable amounts. Using this synthesis a number of diversecompounds was synthesized with altered target specificity. It was shown thatthese compounds exhibit potent whole-cell activity. In addition, furthercharacteristics of the YejABEF transporter were determined using thesecompounds, showing a preference for the more polar aminoacyl residues at the C -terminal position. justify;line-height:150%">mso-ansi-language:EN-GB" lang="EN-GB">McC analogues with variable length of the peptidemoiety were synthesized and evaluated in order to characterize substratepreferences of the YejABEF transporter. It was shown that a minimal peptide-chainlength of six amino acids and the presence for N -terminalformyl-methionyl-arginyl sequence are required for transport. justify;line-height:150%">N -methylatedaaSAs were synthesized to investigate their potential as aaRS inhibitors and toestablish if these would escape acetylation of the alpha amine (withconcomitant inactivation of the inhibitor), which is a self-protectivemechanism in the bacterial cell. It was shown however that these compounds arenot able anymore to effectively inhibit their respective aaRSs. In addition weshowed that ( font-family:"Arial","sans-serif";mso-ansi-language:EN-GB" lang="EN-GB">D )D-SA(i.e. Asp-SA with a mso-ansi-language:EN-GB" lang="EN-GB">D mso-ansi-language:EN-GB" lang="EN-GB">-configuration of the Asp), is a potent inhibitor ofAspRS. However, we also showed that the inhibitory effect of ( mso-ansi-language:EN-GB" lang="EN-GB">D mso-ansi-language:EN-GB" lang="EN-GB">)D-SA is relatively short-lasting. This was shown tobe attributable to a large extent to acetylation by RimL, although it was alsoshown that other unknown factors also play a role in the inactivation of thiscompound. justify;line-height:150%">mso-ansi-language:EN-GB" lang="EN-GB">McC analogues with mso-ansi-language:EN-GB" lang="EN-GB">D mso-ansi-language:EN-GB" lang="EN-GB">-amino acids at positions two to six were shown to beresistant against metabolization by the different peptidases and couldtherefore not liberate the active moiety. This observation could not bereversed by the incorporation of L-amino acids at position six, showing that noneof the peptidases could exhibit endopeptidase activity. justify;line-height:150%">mso-ansi-language:EN-GB" lang="EN-GB">The substitution of the adenine by aryl-tetrazolemoieties linked via a two-carbonspacer in aaSA proved unsuccessful. Although all synthesized compounds, both inMcC as in SDC form, showed nice potential when evaluated innormal"> in vitro aminoacylation experiments, the whole-cell activity wasvirtually abolished. Onlythe activity of compound normal">7 (CB432) against S. aureus couldbe confirmed. It was shown that lack of activity in whole-cell assays was dueto an inability of these compounds to pass the cell membrane. Nevertheless, itwas also shown that the synthesized SDCs could readily be metabolized by thepeptidases PepA, PepB and PepN. 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Publication status: published
KU Leuven publication type: TH
Appears in Collections:Medicinal Chemistry (Rega Institute)

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