DPD-Inspired Discovery of Novel LsrK Kinase Inhibitors: An Opportunity To Fight Antimicrobial Resistance

Abstract

Antibiotic resistance is posing a continuous threat to global public health and represents a huge burden for society as a whole. In the last decade, the interference with bacterial Quorum Sensing (QS) (i.e., cell-cell communication) mechanisms has extensively been investigated as a valid therapeutic approach in the pursuit of a next generation of antimicrobials. (S)-4,5-dihydroxy-2,3pentanedione, commonly known as (S)-DPD, a small signaling molecule that modulates QS in both Gram-negative and Gram-positive bacteria, is phosphorylated by LsrK and the resulting phospho-DPD activates QS. We designed and prepared a small library of DPD derivatives, characterized by five different scaffolds and evaluated their LsrK inhibition in the context of QS interference. SAR studies highlighted the pyrazole moiety as an essential structural element for LsrK inhibition. Particularly, four compounds were found to be micromolar LsrK inhibitors (IC50 ranging between 100 µM and 500 µM) encouraging further exploration of novel analogues as potential new antimicrobials. imination reagent and then deprotected (Scheme 1). Briefly, alkyne 1 was dissolved in DMF and then Pd(OAc)2, PPh3, KOAc and the appropriate o-bromoaldehyde were added. The reaction mixtures were irradiated at 80 °C for 1-2 hours, ammonium acetate was added and the resulting mixtures were irradiated again at 150 °C, thus affording the desired protected products 3a-g (Scheme 1). Heteroaromatic o-bromoaldehydes (i.e., 3-bromofuran-2-carbaldehyde 2e, 3-bromothiophene-2carbaldehyde 2f and 5-bromo-2-methyl-4-thiazolecarboxaldehyde 2g) led to isolation of the corresponding furopyridine, thienopyridine and thiazolopyridine derivatives 3e-f. Final acidic treatment yielded the monosubstituted annulated pyridines-Het-DPD derivatives 4a-g (Scheme 1). Scheme 1: Synthesis of monosubstituted annulated pyridines-Het-DPD derivatives 4a-g. Reagents and conditions: (a) 2a-g (0.9 eq), Pd(OAc)2 (1.8% mol), PPh3 (3.6% mol), KOAc (1.8 eq), DMF, mw, 80 °C, 1h-2h; (a') NH4OAc (1.8 eq), MW, 150 °C, 2h-3h; (b) 12M HCl (cat.), 1,4-dioxane, 0 °C to rt, 1h-3h Sonogashira coupling 43 of terminal alkyne 1 with acyl chlorides, followed by addition of amidinium salts to the corresponding ynones and final acidic removal of the acetal protecting group provided rapid access to 2,4,6-trisubstituted pyrimidines-Het-DPD derivatives (Figure 2, series B). Benzoyl chloride 5a was selected to screen three different conditions for the synthesis of ynone 6a. Surprisingly, copper-, ligand-and solvent-free acylation 51 as well as the use of a mixture of PdCl2(CH3CN)2, Sphos and Cs2CO3 26 did not furnish the desired product. Good results were obtained with the copper-and palladium-catalyzed system proposed by Karpov et al. 50 which allowed us to obtain compound 6a with a yield of 87%. 52 Three different ynones (6a-c) were reacted with six different amidinium salts and the resulting products 7a-f treated under acidic conditions (i.e., Scheme 2, conditions c) to afford the 2,4,6trisubstituted pyrimidines-Het-DPD derivat...

Resistance of Gram-Negative Bacteria to Current Antibacterial Agents and Approaches to Resolve It

“…2-Substituted aminobenzoic acids are other compounds structurally related to DPD that were identified as LsrK inhibitors by virtual screening. Working on a hit-to-lead process to increase LsrK inhibition by changing in the heterocyclic core can be used as QS-interfering compounds in the treatment of bacterial infections [82]. Modulation or inhibition of QS has emerged as a potential therapeutic that can control several bacterial virulence factors such as biofilm formation and reduce the bad effect of bacterial infections.…”

Quorum-Sensing Regulation of Antimicrobial Resistance in Bacteria

“…This strategy may be useful in mixed infections because LsrK can phosphorylate DPD as a precursor molecule of AI-2 [130]. In addition, it may be effective regardless of the structure and transport/sensor mechanism of AI-2 for different bacterial QS systems [131]. It has been reported that exogenous imidazole is a furan ring analog of AI-2, which reduces the resistance of E. coli to β-lactam antibiotics by inhibiting the function of AI-2 [113].…”

Regulatory Mechanisms of the LuxS/AI-2 System and Bacterial Resistance

“…This strategy might be particularly effective in mixed infections because LsrK can phosphorylate DPD (the precursor molecule of AI-2) (91). Moreover, it may be effective regardless of the AI-2 structure and transport/sensor mechanism used by different bacterial QS systems (92). Yu et al (93) and others reported that exogenous imidazole, a furan carbocyclic analogue of AI-2, reduced the antibiotic resistance of clinical E. coli strains to ␤-lactam antibiotics by inhibiting the function of AI-2.…”