Design and synthesis of new N-thioacylated ciprofloxacin derivatives as urease inhibitors with potential antibacterial activity

Abstract

A new series of N-thioacylated ciprofloxacin 3a-n were designed and synthesized based on Willgerodt-Kindler reaction. The results of in vitro urease inhibitory assay indicated that almost all the synthesized compounds 3a-n (IC₅₀ = 2.05 ± 0.03-32.49 ± 0.32 μM) were more potent than standard inhibitors, hydroxyurea (IC₅₀ = 100 ± 2.5 μM) and thiourea (IC₅₀ = 23 ± 0.84 μM). The study of antibacterial activity against Gram-positive species (S. aureus and S. epidermidis) revealed that the majority of compounds were more active than ciprofloxacin as the standard drug, and 3h derivative bearing 3-fluoro group had the same effect as ciprofloxacin against Gram-negative bacteria (P. aeruginosa and E. coli). Based on molecular dynamic simulations, compound 3n exhibited pronounced interactions with the critical residues of the urease active site and mobile flap pocket so that the quinolone ring coordinated toward the metal bi-nickel center and the essential residues at the flap site like His593, His594, and Arg609. These interactions caused blocking the active site and stabilized the movement of the mobile flap at the entrance of the active site channel, which significantly reduced the catalytic activity of urease. Noteworthy, 3n also exhibited IC₅₀ values of 5.59 ± 2.38 and 5.72 ± 1.312 µg/ml to inhibit urease enzyme against C. neoformans and P. vulgaris in the ureolytic assay.

Figure 1

Identified representative lead candidates.

Figure 2

Outline for the synthesis of N-thioacylated ciprofloxacin derivatives 3a–n. Reagents and conditions: (a) DMSO, 50 °C, 4–8 h.

Figure 3

A plausible mechanism for the preparation of 3a.

Figure 4

Representation of the compounds docking poses over the active site (a) close-up illustration of ciprofloxacin nucleuses relative to the binuclear center (b), the active site flap (colored in green color).

Figure 5

RMSD plot of the urease backbone in complexed with thiourea (in green) and compound 3n (in yellow) over 30 ns of the MD simulation time.

Figure 6

RMSF plot of the urease residue in complexed with thiourea (in green) and compound 3n (in yellow) (a), individual RMSF plot regards to ligand binding location over 30 ns MD simulation time (b). α-helical and ß-strand regions are highlighted in light pink and blue backgrounds, respectively.

Figure 7

The distance between Ala440 and Ile599 urease residues when complexed with thiourea (green), and compound 3n (yellow) during the whole MD simulation time. (a) Representative snapshots of MD simulations where the active site flap adopts the open (urease-thiourea complex) and closed (urease-compound 3n) conformations which depict in green and yellow color, respectively (b).

Figure 8

2D representation of ligand-residue interactions that occur at least 30% of simulation time at the equilibrated phase of MD simulation which include urease bound-state with thiourea (a) and compound 3n (b). Timeline rendering of interacting residues during the whole simulation time in urease complexed with thiourea (c) and compound 3n (d).

Figure 9

Two different configurations of 4-nitro phenyl thioamide moiety of compound 3n from the beginning of MD to the 16.56 ns (a) and from the time at 16.56 ns to the end of the MD simulation time (b). The RMSD of compound 3n during the whole simulation time (c).