Novel 1,2,3-Triazole-sulphadiazine-ZnO Hybrids as Potent Antimicrobial Agents against Carbapenem Resistant Bacteria

Abstract

Bacterial pneumonia is considered one of the most virulent diseases with high morbidity and mortality rates, especially in hospitalized patients. Moreover, bacterial resistance increased over the last decades which limited the therapy options to carbapenem antibiotics. Hence, the metallo-β-lactamase-producing bacteria were deliberated as the most deadly and ferocious infectious agents. Sulphadiazine-ZnO hybrids biological activity was explored in vitro and in vivo against metallo-β-lactamases (MBLs) producing Klebsiella pneumoniae. Docking studies against NDM-1 and IMP-1 MBLs revealed the superior activity of the 3a compound in inhibiting both MBLs enzymes in a valid reliable docking approach. The MBLs inhibition enzyme assay revealed the remarkable sulphadiazine-ZnO hybrids inhibitory effect against NDM-1 and IMP-1 MBLs. The tested compounds inhibited the enzymes both competitively and noncompetitively. Compound 3b-ZnO showed the highest antibacterial activity against the tested metallo-β-lactamase producers with an inhibition zone (IZ) diameter reaching 43 mm and a minimum inhibitory concentration (MIC) reaching 2 µg/mL. Sulphadiazine-ZnO hybrids were tested for their in vitro cytotoxicity in a normal lung cell line (BEAS-2Bs cell line). Higher cell viability was observed with 3b-ZnO. Biodistribution of the sulphadiazine-ZnO hybrids in the lungs of uninfected rats revealed that both [^124I]3a-ZnO and [^124I]3b-ZnO hybrids remained detectable within the rats' lungs after 24 h of endotracheal aerosolization. Moreover, the residence duration in the lungs of [^124I]3b-ZnO (t_1/2 4.91 h) was 85.3%. The histopathological investigations confirmed that compound 3b-ZnO has significant activity in controlling bacterial pneumonia infection in rats.

Keywords: IMP; NDM; docking; enzyme assay; in vivo; pneumonia; sulphadiazine-ZnO hybrids.

Figure 1

Certified drugs against bacterial pneumonia.

Scheme 1

Click synthesis of 1,2,3-triazoles with sulfa-drug tethers 3a–c.

Figure 2

XRD pattern of ZnO NPs.

Figure 3

FTIR spectra of the ZnO NPs.

Figure 4

Docking study of the synthesized compounds against IMP-1. (a) Two-dimensional binding mode of L-captopril, (b) 3D binding mode of L-captopril, (c) 2D binding mode of compound 3a, (d) 3D binding mode of compound 3a, (e) 2D binding mode of compound 3b, (f) 3D binding mode of compound 3b, (g) 2D binding mode of compound 3c, (h) 3D binding mode of compound 3c inside IMP-1 active site.

Figure 5

Three-dimensional binding mode of ZnO nanoparticles inside IMP-1 active site.

Figure 6

Docking study of the synthesized compounds against NDM-1. (a) Two-dimensional binding mode of L-captopril, (b) 3D binding mode of L-captopril, (c) 2D binding mode of compound 3a, (d) 3D binding mode of compound 3a, (e) 2D binding mode of compound 3b, (f) 3D binding mode of compound 3b, (g) 2D binding mode of compound 3c, (h) 3D binding mode of compound 3c inside NDM-1 active site.

Figure 7

Three-dimensional binding mode of ZnO nanoparticles inside NDM-1 active site.

Figure 8

Agarose gel electrophoresis showing: PCR detection for NDM (a) and IPM (b) genes. K. pneumoniae ATCC 13883 in lane 1 while K. pneumoniae tested strains were in lane from 2 to 10.

Figure 9

Antibacterial effect of the synthesized compounds and their hybrids against Kp5. (a) Disc diffusion method, (b) transmission electron microscopic study of Kp5 treated cell, and (c) time-kill curve.

Figure 10

Cytotoxic effect of the synthesized hybrids.

Figure 11

Endotracheal aerosolization in uninfected rats through PET-CT images of the biodistribution of radiolabeled compounds (a). Concentration of the radiolabeled compounds in the uninfected rats’ lungs versus time (b).

Figure 12

Bacterial load assessment in the lungs of pneumonic rats.

Figure 13

Photomicrograph of rat lung infected with Kp5 and given endotracheal aerosolization various preparations of 1,2,3-triazole-sulphadiazine-ZnO hybrids as treatments, 48 h post-infection. (A) Negative control group. (B) Positive control; (C) 3c-ZnO; (D) 3a-ZnO; and (E) 3b-ZnO. In which black arrows refer to focal accumulations of macrophages (in B); red arrows refer to normal small blood vessels for gas exchange; red stars (alveolar histiocytosis); green arrows refer to densely packed cluster of lymphocytes associated lymphoid tissue; TB refers to terminal bronchioles found to be alienated with normal histological structure in (D) and damaged ones observed in (C). H&E stain (magnification: 200×).

Figure 14

Photomicrograph of rat lung infected withKp5 and given endotracheal aerosolization various preparations of 1,2,3-triazole-sulphadiazine-ZnO hybrids as treatments, 96 h post-infection. (A) Negative control group. (B) Positive control; (C) 3c-ZnO; (D) 3a-ZnO; and (E) 3b-ZnO. Black arrows refer to increased macrophage focal accumulations in (B), with some remnants of macrophage accumulation only observed in (D); red arrows refer to the appearance of normal small blood vessels for gas exchange; green arrows refer to lymphatic-associated lymphoid tissue in (B,C) that is not symptoms of alveolar histiocytosis found in (C,D). Fewer signs of histiocytosis persisted in (E) with its elimination in (A,B). H&E stain (magnification: 200×).

Figure 15

Transmission electron microscope study of the first interval (a,c,e,g,i) and the second interval (b,d,f,h,j) of the experimental rat groups. Note: (a,b) and b represent the control group, (c,d) represent the positive control group, (e,f) represent group 3, (g,h) represent group 4, and finally (i,j) represent group 5. Nucleus (N); alveolar lumen (L); cellular debris (D); congestion (C); luminal micro villi (blue arrows); lamellar bodies (green arrows).