Antimicrobial resistance (AMR) is a major public health concern. In a recently published report, commissioned by the British government, economist Jim O’eill and co-authors estimate that by 2050 10 million lives a year will be at risk due to the rise of drug-resistant infections. The chronic misuse and overuse of our most potent antibiotics, together with the fact that conventional antibiotics impose immense selective pressure on bacteria has led to the generation of multi-resistant micro-organisms. Methicillin-resistant Staphylococcus aureus (MRSA) is one of these so-called ‘uperbugs’ To make matters worse, Staphylococcus aureus (S. aureus) is notorious for the formation of biofilms. The resistance of bacteria to antimicrobial products is much larger in biofilms than in planktonic form. While antibiotic resistance constitutes a major public health threat, the development of new antibiotics is extremely scarce and poses a problem. Mainly the enormous economic costs and the fear of developing drugs that are likely to become rapidly obsolete, create a situation in which pharmaceutical companies fail to cover the needs. Moreover, novel targets are needed to combat microbial infections. In Chapter I, a general overview of the problem of antimicrobial resistance is given, with a special focus on S. aureus. We also provide quite an elaborate outline of S. aureus pathogenicity as well as (future) treatment options and potential new platforms for controlling S. aureus infection. Potentiating the effect of existing antimicrobial agents and/or altering the virulence of pathogens may provide promising alternatives to conventional antibiotics. In this PhD work, compounds that are not antimicrobial in nature, but augment the susceptibility of existing antibiotics were developed. More specifically, we focused on inhibitors of quorum sensing in S. aureus. Quorum sensing (QS) is a bacterial intercellular communication system that controls gene expression. In S. aureus, QS is involved in biofilm formation and virulence in general. The recent discovery of the quorum sensing inhibitor (QSI) hamamelitannin (HAM), a natural substance isolated from the American witch hazel (Hamamelis virginiana), for drugresistant staphylococcal infections has provided an interesting starting point for further study in this important field of drug research. HAM increases the susceptibility of bacterial biofilms to antibiotics in vitro as well as in vivo. It is important to emphasize that HAM must not be considered as a classical antibiotic drug, since it doesn’ exert bactericidal nor bacteriostatic effects. HAM interferes with mechanisms that are responsible for the exceptional resistance in biofilms and thus can cause a potentiating effect in combination with antibiotics. However, from a medicinal chemistry perspective, the structure of HAM is not ideal. Therefore, we investigated structure-activity relationships (S.A.R.) around HAM, in order to find analogues with better drug-like properties to enhance metabolic stability and activity. The specific objectives of this work are described in Chapter II. In the form of a compilation of the peer-reviewed publications that have resulted from this project, Chapter III describes the scientific investigations that have been carried out over the course of the past four years. The first publication, in Angewandte Chemie International Edition (DOI: 10.1002/anie.201601973), describes how we gravitated from HAM towards more drug-like analogues. In this paper we focus on optimization of the 5-side (i.e. the “estern side” of the molecule. Our work resulted in the identification of a metabolically stable lead compound that shows promising potentiating properties in vitro as well as in two in vivo models of S. aureus infection. In the manuscript, we allude to the fact that ortho substitution of the 5-phenyl ring is most probably beneficial for activity. In our second publication, in Bioorganic & Medicinal Chemistry (DOI: 10.1016/j.bmc.2016.07.058), we further focus on the S.A.R. of the 5-position. The stereochemical requirements at C4 are investigated, alternative nitrogen-based linkers as well. The hypothesis mentioned above with regards to ortho substitution distorting the phenylamide moiety to a non-planar conformation, is corroborated. The third part of Chapter III constitutes our publication in ACS Medicinal Chemistry Letters (DOI: 10.1021/acsmedchemlett.6b00315). Here we attempt to shed light onto the structural requirements at the 2’-side (i.e. the “astern side” of the molecule. None of the bisbenzamides described herein proved exceptionally potent. Truncation of the 2’-acyl group, however, yielded several interesting potentiators. Furthermore, we built HAM analogues that combine favorable substituents of both sides in one single compound. This mix-andmatch endeavour yielded several highly potent hybrid molecules. A fourth manuscript, which was accepted for publication in European Journal of Medicinal Chemistry (DOI: 10.1016/j.ejmech.2016.10.056), forms the last part of Chapter III. In search for derivatives with improved synthetic accessibility, we describe several practical pathways towards the synthesis of HAM analogues with alternative central scaffolds. In this scaffold hopping approach, we were inspired by a ligand-based virtual screening procedure, and via the synthesis and biological evaluation of such compounds, we conclude that there is room for structural improvement in the central part of the molecule. A general conclusion is given in Chapter IV of this thesis. Chapter V puts the research conducted and the results obtained in this thesis in a broader international context. This part is dedicated to a discussion and critical evaluation of our hamamelitannin analogues as potentiators for existing antibiotics in the treatment of resistant S. aureus infections. Finally, future perspectives are outlined.