Development of a New Narrow Spectrum Antibacterial

Agent against Gram-Positive MDR Pathogens

Rashedul Islam1, Charlotte Hind2, Mathew Wand2, J Mark Sutton2 and
K Miraz Rahman1
1

Institute of Pharmaceutical Science, Faculty of Life Sciences &

Medicine, Kings College London, London SE1 1DB United Kingdom
2

Public Health England, Porton Down, Salisbury SP4 0JG, United

Kingdom
Background: Antibiotic discovery is considered as one of the
greatest achievements of modern times, but the evolution of
multidrug resistance (MDR) in clinically significant bacteria has put
that achievement at risk. Most of the frontline antibiotics are
gradually becoming resistant and no new classes of antibiotics have
been discovered in the last 30 years. Due to the rapid emergence of
multiple drug resistant strains, there is an urgent need to develop
novel agents that are active against MDR pathogens. A new
chemical scaffold has been designed by modifying the structure of a
naturally occurring alkaloid to facilitate the introduction of halogen
atoms. The antibacterial activity of the synthesized compounds was
tested against a panel of MDR Gram-positive and Gram-negative
bacterial strains.
Methods: The compounds were synthesized using a linear
synthetic scheme starting with commercially available anthranilic
acid. All compounds were purified using standard liquid column
chromatography and fully characterized using Nuclear Magnetic
Resonance (NMR), Liquid Chromatography-Mass Spectroscopy (LCMS) and High-Resolution Mass Spectroscopy (HRMS) before
biological evaluation. The compounds were screened against MDR
clinical strains of the ESKAPE panel using the broth dilution method
to determine their minimum inhibitory concentrations (MIC). Lead
compounds were further evaluated against several MDR isolates to
determine their minimum bactericidal concentration (MBC) and
Time-Kill kinetics assays were carried out to study their mode of
action and potential for emergence of resistance.
Results: To date, 32 compounds have been synthesized and tested
for their antibacterial activity. From the initial screening, the
compound RI-63.65 with fluorine at specific positions in ring A (C 1)
and ring D (C8) emerged as the lead compound with excellent
activity against a large panel of MDR Gram-positive clinical isolates

including methicillin resistant Staphylococcus aureus (EMRSA-15,

EMRSA-16) and vancomycin resistant Enterococci (NCTC12201,
NCTC12204). RI-63.65 showed MICs in the range of 0.5-16g/mL
with identical MBC values. Time-kill kinetics study demonstrated a
bactericidal mode of action for this compound with 7-log kill within
30 minutes of treatment. No persistent population was observed
during the study which suggests the emergence of resistance is
unlikely. The compounds work by inhibiting DNA Gyrase, a finding
that was further supported by advanced computational chemistry
experiments. RI-63-65 didnt show any eukaryotic toxicity at the
highest concentration tested.
Conclusion:
A new class of antibacterial agent with rapid
bactericidal activity against MDR Gram-positive bacterial strains
have been identified. A detailed structure activity relationship (SAR)
profile is currently being established by synthesizing compounds
that cover the patent chemical space. Further microbiological
profiling of RI-63-65 will involve antibacterial efficacy testing in a
non-animal Galleria model at Public Health England an in vivo
infection-imaging model at the University of Nottingham.