An Unexpected Switch in the Modulation of AI-2-Based Quorum Sensing Discovered through Synthetic 4,5-Dihydroxy-2,3-pentanedione Analogues

Abstract

Bacteria secrete and respond to small chemical signals, or autoinducers, in a cell densitydependent process known as quorum sensing (QS). 1 As the number of cells, and consequently autoinducer concentration, increases, bacterial populations coordinate their gene expression to behave as a unified group. This coordinated effort of bacteria might result in deleterious effects for humans, as certain bacteria use QS to regulate the formation of biofilms and the secretion of virulence factors. As a result, the modulation of QS has emerged as a therapeutic target of considerable interest. 2,3One class of autoinducers is AI-2, derived from the precursor (S)-4,5-dihydroxy-2,3-pentanedione (DPD), and the gene encoding the DPD synthase, LuxS, has been identified in over 55 bacterial species. 4 QS systems based on this autoinducer have been shown to regulate bioluminescence in Vibrio harveyi, virulence and biofilm formation in Vibrio cholera, AI-2 transport and virulence expression in Salmonella typhimurium, and mixed-species biofilm development in oral pathogens. 5,6 Illustrated in this final example is the fact that AI-2 QS, in contrast to other QS systems that are used for communication between members of the same species, serves as a mechanism of interspecies communication. 4 Thus, the development of agonists or antagonists for this system would have implications for broad range QS modulation.Despite the wide distribution of the AI-2 synthase LuxS, the discrete structures of DPD-based autoinducers and their respective receptor proteins have only been identified in two species: V. harveyi and S. typhimurium. These two signals are distinct despite the fact that both are derived from DPD and rapidly interconvert in solution: S. typhimurium responds to the Rtetrahydroxytetrahydrofuran (R-THMF) form of DPD, whereas V. harveyi responds to the S-THMF borate diester form of DPD (Supporting Information, Figure S1). 4 This lack of structural information has been detrimental to the development of agonists and antagonists of AI-2-based QS, and reports of modulators of this system remain limited. Several reports have identified weak and partial agonists, 7-9 while reports of antagonists remain largely limited to one class of natural products. 10,11 Because of this dearth of information regarding the modulation of AI-2 QS, there is no solid rationale for the design of new ligands. Herein, we report the discovery of a class of synergistic compounds toward the QS of V. harveyi, as well as a remarkable switch in the biological transmission of AI-2-based QS in S. typhimurium stemming from the addition of methylene groups to the C1 position of DPD. E-mail: kdjanda@scripps.edu. Supporting Information Available: Experimental procedures, spectral data, and biological protocols. This material is available free of charge via the Internet at http://pubs.acs.org. As a first step toward the discovery of new modulators of AI-2-based QS, we created a panel of C1-substituted DPD analogues according to Scheme 1. The synthesis is founded upon previous...

Exploiting Quorum Sensing To Confuse Bacterial Pathogens

“…First, as anticipated, numerous DPD derivatives were found to be AI-2 QS antagonists in one or multiple bacterial species (209)(210)(211)(212)(213). Second, some of the analogues were found to be synergistic agonists despite having no agonist activity on their own (209)(210)(211)213).…”

“…Likewise, cyclic DPD analogues generally inhibited AI-2 QS in E. coli, whereas they either had no activity or functioned as synergistic agonists in S. Typhimurium (209). Additionally, the activity of a given analogue in a given strain may be context dependent, as multiple C1-alkyl DPD derivatives were found to function as either antagonists or synergistic agonists in V. harveyi under different assay conditions (210)(211)(212)(213).…”

“…The best inhibitor identified from these screens was pyrogallol (Table 3), with an IC 50 of 2 M. More recently, there have been multiple reports of the generation and screening of C1-and C2-alkyl DPD derivatives. C1-alkyl derivatives have included linear, branched, cyclic, and aromatic DPD analogues (209)(210)(211)(212)(213), whereas the C2 derivatives tested have thus far been limited to linear substitutions of various alkyl chain lengths (214). None of these C1-or C2-alkyl analogues were more potent inhibitors than the previously identified pyrogallol, but some did have AI-2-modulating activity.…”

“…Also, given the species specificity exhibited by many of the analogues tested to date, disruption of AI-2 signaling in multispecies environments will likely require multiple AI-2 analogues used in combination that can inhibit AI-2 QS in various species simultaneously (209,212) or in combination with inhibitors of other QS pathways that contribute to virulence alongside the AI-2 pathway. Finally, although AI-2 derivatives do not appear to be cytotoxic toward mammalian cells (211), work will need to be done to assess the stability and efficacy of AI-2 analogues in vivo. One concern for in vivo efficacy is the general instability of AI-2 molecules.…”

Innovative Strategies for Combating Biofilm-Based Infections

“…6.3 ), which activate the AI-2 pathway more potently than DPD, exhibiting submicromolar EC 50 values in a V. harveyi reporter strain (Tsuchikama et al 2012 ). Several C1-substituted DPD analogues act as antagonists of AI-2-based QS, with propyl-DPD 19 and butyl-DPD 20 exhibiting IC 50 values ten- fold below that of DPD (Lowery et al 2008 ). Another C1-substituted analogue, isobutyl-DPD 21 signifi cantly inhibits maturation of Escherichia coli biofi lms and also brings about near complete clearance of pre-formed E. coli biofi lms when administered in combination with gentamicin (Roy et al 2013 ).…”