Structural Modifications of the Quinolin-4-yloxy Core to Obtain New Staphylococcus aureus NorA Inhibitors

Tackling antimicrobial resistance (AMR) represents a social responsibility aimed at renewing the antimicrobial armamentarium and identifying novel therapeutical approaches. Among the possible strategies, efflux pumps inhibition offers the advantage to contrast the resistance against all drugs which can be extruded. Efflux pump inhibitors (EPIs) are molecules devoid of any antimicrobial activity, but synergizing with pumps-substrate antibiotics. Herein, we performed an in silico scaffold hopping approach starting from quinolin-4-yloxy-based Staphylococcus aureus NorA EPIs by using previously built pharmacophore models for NorA inhibition activity. Four scaffolds were identified, synthesized, and modified with appropriate substituents to obtain new compounds, that were evaluated for their ability to inhibit NorA and synergize with the fluoroquinolone ciprofloxacin against resistant S. aureus strains. The two quinoline-4-carboxamide derivatives 3a and 3b showed the best results being synergic (4-fold MIC reduction) with ciprofloxacin at concentrations as low as 3.13 and 1.56 µg/mL, respectively, which were nontoxic for human THP-1 and A549 cells. The NorA inhibition was confirmed by SA-1199B ethidium bromide efflux and checkerboard assays against the isogenic pair SA-K2378 (norA++)/SA-K1902 (norA-). These in vitro results indicate the two compounds as valuable structures for designing novel S. aureus NorA inhibitors to be used in association with fluoroquinolones.

Download Full-text

Antifungal Azoles as Tetracycline Resistance-Modifiers in Staphylococcus aureus

Applied and Environmental Microbiology ◽

10.1128/aem.00155-21 ◽

2021 ◽

Author(s):

Nisha Mahey ◽

Rushikesh Tambat ◽

Dipesh Kumar Verma ◽

Nishtha Chandal ◽

Krishan Gopal Thakur ◽

...

Keyword(s):

Staphylococcus Aureus ◽

In Silico ◽

Biofilm Formation ◽

Efflux Pump ◽

Efflux Pumps ◽

Major Facilitator Superfamily ◽

Efflux Pump Inhibitors ◽

Approved Drugs ◽

Major Facilitator

Staphylococcus aureus has developed resistance to antimicrobials since its first use. The S. aureus major facilitator superfamily (MFS) efflux pump Tet(K) contributes to resistance to tetracyclines. The efflux pump diminishes antibiotic accumulation, and biofilm hampers the diffusion of antibiotics. None of the currently known compounds have been approved as efflux pump inhibitors (EPIs) for clinical use. In the current study, we screened clinically approved drugs for possible Tet(K) efflux pump inhibition. In silico docking followed by in vitro checkerboard assays, we identified five azoles (the fungal ergosterol synthesis inhibitors) showing the putative EPI-like potential with a fractional inhibitory concentration index of ≤0.5, indicating synergism. The functionality of the azoles was confirmed using ethidium bromide (EtBr) accumulation and efflux inhibition assays. In time-kill kinetics, the combination treatment with butoconazole engendered a marked increase in the bactericidal capacity of tetracycline. When assessing the off-target effects of the azoles, we observed no disruption of bacterial membrane permeability and polarization. Finally, the combination of azoles with tetracycline led to a significant eradication of preformed mature biofilms. This study is the primary representation of azoles that can be repurposed as putative Tet(K) EPIs and to reduce biofilm formation at clinically relevant concentrations. IMPORTANCE Staphylococcus aureus use efflux pumps to transport antibiotics out of the cell and thus increase the dosage at which they endure antibiotics. Also, efflux pumps play a role in biofilm formation by the excretion of extracellular matrix molecules. One way to combat these pathogens may be to reduce the activity of efflux pumps and thereby increase pathogen sensitivity to existing antibiotics. We describe the in silico-based screen of clinically approved drugs that identified antifungal azoles inhibiting Tet(K); a pump belongs to the Major Facilitator Superfamily and shows that these compounds bind to and block the activity of the Tet(K) pump. Azoles enhanced the susceptibility of tetracycline against S. aureus and its methicillin-resistant strains. The combination of azoles with tetracycline led to a significant reduction in preformed biofilms. Repurposing of approved drugs may help solve the classical toxicity issues related to efflux pump inhibitors.

Download Full-text

Virtual Screening for Novel Staphylococcus Aureus NorA Efflux Pump Inhibitors From Natural Products

Medicinal Chemistry ◽

10.2174/1573406410666140902110903 ◽

2015 ◽

Vol 11 (2) ◽

pp. 135-155 ◽

Cited By ~ 17

Author(s):

Khac-Minh Thai ◽

Trieu-Du Ngo ◽

Thien-Vy Phan ◽

Thanh-Dao Tran ◽

Ngoc-Vinh Nguyen ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Natural Products ◽

Virtual Screening ◽

Efflux Pump ◽

Efflux Pump Inhibitors

Download Full-text

From quinoline to quinazoline‐based S. aureus NorA efflux pump inhibitors by coupling focused scaffold hopping approach and pharmacophore search

ChemMedChem ◽

10.1002/cmdc.202100282 ◽

2021 ◽

Author(s):

Nicholas Cedraro ◽

Rolando Cannalire ◽

Andrea Astolfi ◽

Gianmarco Mangiaterra ◽

Tommaso Felicetti ◽

...

Keyword(s):

Efflux Pump ◽

Scaffold Hopping ◽

Efflux Pump Inhibitors

Download Full-text

Staphylococcus aureus Mutants Isolated via Exposure to Nonfluorinated Quinolones: Detection of Known and Unique Mutations

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.45.12.3422-3426.2001 ◽

2001 ◽

Vol 45 (12) ◽

pp. 3422-3426 ◽

Cited By ~ 10

Author(s):

Siddhartha Roychoudhury ◽

Tracy L. Twinem ◽

Kelly M. Makin ◽

Mark A. Nienaber ◽

Chuiying Li ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Sequence Analysis ◽

Efflux Pump ◽

Serial Passage ◽

Efflux Pump Inhibitor ◽

In Vitro Development ◽

Development Of Resistance ◽

High Level ◽

Vitro Development

ABSTRACT The in vitro development of resistance to the new nonfluorinated quinolones (NFQs; PGE 9262932, PGE 4175997, and PGE 9509924) was investigated in Staphylococcus aureus. At concentrations two times the MIC, step 1 mutants were isolated more frequently with ciprofloxacin and trovafloxacin (9.1 × 10−8 and 5.7 × 10−9, respectively) than with the NFQs, gatifloxacin, or clinafloxacin (<5.7 × 10−10). Step 2 and step 3 mutants were selected via exposure of a step 1 mutant (selected with trovafloxacin) to four times the MICs of trovafloxacin and PGE 9262932. The step 1 mutant contained the known Ser80-Phe mutation in GrlA, and the step 2 and step 3 mutants contained the known Ser80-Phe and Ser84-Leu mutations in GrlA and GyrA, respectively. Compared to ciprofloxacin, the NFQs were 8-fold more potent against the parent and 16- to 128-fold more potent against the step 3 mutants. Mutants with high-level NFQ resistance (MIC, 32 μg/ml) were isolated by the spiral plater-based serial passage technique. DNA sequence analysis of three such mutants revealed the following mutations: (i) Ser84-Leu in GyrA and Glu84-Lys and His103-Tyr in GrlA; (ii) Ser-84Leu in GyrA, Ser52-Arg in GrlA, and Glu472-Val in GrlB; and (iii) Ser84-Leu in GyrA, Glu477-Val in GyrB, and Glu84-Lys and His103-Tyr in GrlA. Addition of the efflux pump inhibitor reserpine (10 μg/ml) resulted in 4- to 16-fold increases in the potencies of the NFQs against these mutants, whereas it resulted in 2-fold increases in the potencies of the NFQs against the parent.

Download Full-text

Development of an accumulation assay and evaluation of the effects of efflux pump inhibitors on the retention of chlorhexidine digluconate in Pseudomonas aeruginosa and Staphylococcus aureus

BMC Research Notes ◽

10.1186/s13104-017-2637-2 ◽

2017 ◽

Vol 10 (1) ◽

Cited By ~ 5

Author(s):

Molly Mombeshora ◽

Stanley Mukanganyama

Keyword(s):

Staphylococcus Aureus ◽

Pseudomonas Aeruginosa ◽

Efflux Pump ◽

Chlorhexidine Digluconate ◽

Efflux Pump Inhibitors ◽

And Staphylococcus Aureus

Download Full-text

Influence of P-glycoprotein and MRP efflux pump inhibitors on the intracellular activity of azithromycin and ciprofloxacin in macrophages infected by Listeria monocytogenes or Staphylococcus aureus

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkg223 ◽

2003 ◽

Vol 51 (5) ◽

pp. 1167-1173 ◽

Cited By ~ 77

Author(s):

C. Seral

Keyword(s):

Staphylococcus Aureus ◽

Listeria Monocytogenes ◽

Efflux Pump ◽

Intracellular Activity ◽

P Glycoprotein ◽

Efflux Pump Inhibitors

Download Full-text

Quinazolinone Fungal Efflux Pump Inhibitors. Part 2. In vitro Structure?Activity Relationships of (N-Methyl-piperazinyl)-Containing Derivatives.

ChemInform ◽

10.1002/chin.200503155 ◽

2005 ◽

Vol 36 (3) ◽

Author(s):

William J. Watkins ◽

et al. et al.

Keyword(s):

Efflux Pump ◽

Structure Activity Relationships ◽

Structure Activity ◽

Efflux Pump Inhibitors

Download Full-text

5-Arylideneimidazolones with Amine at Position 3 as Potential Antibiotic Adjuvants against Multidrug Resistant Bacteria

Molecules ◽

10.3390/molecules24030438 ◽

2019 ◽

Vol 24 (3) ◽

pp. 438 ◽

Cited By ~ 3

Author(s):

Aneta Kaczor ◽

Karolina Witek ◽

Sabina Podlewska ◽

Joanna Czekajewska ◽

Annamaria Lubelska ◽

...

Keyword(s):

Efflux Pump ◽

Enterobacter Aerogenes ◽

Multidrug Resistant ◽

Resistant Bacteria ◽

Aromatic Rings ◽

Efflux Pump Inhibitor ◽

Fold Reduction ◽

Dual Action ◽

New Compounds

Searching for new chemosensitizers of bacterial multidrug resistance (MDR), chemical modifications of (Z)-5-(4-chlorobenzylidene)-2-(4-methylpiperazin-1-yl)-3H-imidazol-4(5H)-one (6) were performed. New compounds (7–17), with fused aromatic rings at position 5, were designed and synthesized. Crystallographic X-ray analysis proved that the final compounds (7–17) were substituted with tertiary amine-propyl moiety at position 3 and primary amine group at 2 due to intramolecular Dimroth rearrangement. New compounds were evaluated on their antibiotic adjuvant properties in either Gram-positive or Gram-negative bacteria. Efflux pump inhibitor (EPI) properties towards the AcrAB-TolC pump in Enterobacter aerogenes (EA289) were investigated in the real-time efflux (RTE) assay. Docking and molecular dynamics were applied to estimate an interaction of compounds 6–17 with penicillin binding protein (PBP2a). In vitro ADME-Tox properties were evaluated for compound 9. Most of the tested compounds reduced significantly (4-32-fold) oxacillin MIC in highly resistant MRSA HEMSA 5 strain. The anthracene-morpholine derivative (16) was the most potent (32-fold reduction). The tested compounds displayed significant EPI properties during RTE assay (37–97%). The naphthyl-methylpiperazine derivative 9 showed the most potent “dual action” of both oxacillin adjuvant (MRSA) and EPI (E. aerogenes). Molecular modeling results suggested the allosteric mechanism of action of the imidazolones, which improved binding of oxacillin in the PBP2a active site in MRSA.

Download Full-text