Next Generation Antibiotics: Progress Toward the Synthesis of an
Enzyme-Activated Nitric Oxide-Releasing Fluorescent Prodrug
Hailey A. J. Hibbard, Melissa M. Reynolds
Department of Chemistry, Colorado State University
Abstract
Bacterial resistance to antibiotics is spreading at an alarming rate, and without the
development of new antibiotics, common infections will become deadly. The goal of this project is to synthesize an enzyme-activated antibiotic prodrug that detects and kills
bacteria. The antibiotic will incorporate nitric oxide, a known antibacterial agent, and a fluorescent compound to visualize bacterial presence. A synthesis procedure was
developed to synthesize a fluorescent compound attached to a small signaling compound. A nitric oxide donor will be added in the future. In the presence of bacteria, the antibiotic prodrug is designed to simultaneously fluoresce and release nitric oxide.
Esterase
Anti-leukemia activity
Cytochrome P450 enzymes Protects against liver toxicity
AcOM-PYRRO/NO V-PYRRO/NO
N-diazeniumdiolates
NO donor group
Project Idea
Create a small molecule that detects and kills bacteria
• Enzymatically-activated NO prodrug senses bacteria, releases NO to kill bacteria
Model compound Non-fluorescent compound
Bacterial studies performed by Bella Neufeld of the Reynolds group
Fluorescent color of synthesized compounds changes as expected in the presence of Pseudomonas aeruginosa
Add diazeniumdiolate group
Special thanks to Dr. Melissa Reynolds for her help and guidance. Thanks to Bella Neufeld of the Reynolds group for performing the bacterial studies, and Lei Yang of the Schoenfisch group for performing the NO addition reactions.
Introduction
Multiple mechanisms of action against bacteria
Nitric Oxide (NO)
Inactive drug metabolism Active drug Prodrugs
Cellier, et. al. PLOS ONE, 2016, 11.
amino acid NO sensor enzyme NO spontaneous amino acid NO sensor sensor Fluorescent compound
Protease Sensor Synthesis
Fluorescent Sensor Synthesis
Model Compound Synthesis
Non-Fluorescent Compound Synthesis
NO addition reactions run by Lei Yang of the Schoenfisch group at UNC Chapel Hill
A B C
λmax A: 263, 413 nm λmax B: 258 nm λmax C: 250 nm
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 230 260 290 320 350 380 410 440 470 500 Abs orbanc e (a.u .) Wavelength (nm)
Compounds A-C in 0.01 M NaOH
A B C 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 0 5 10 15 20 25 30 Abs orbanc e (a.u .) Time (hours)
Water Decomposition Experiment
B C
Bacterial Studies
Positive control Boc-protected fluorescent sensor Boc-deprotected fluorescent sensor No new antibiotics Deadly infectionsUV-Vis Product Analysis
Conclusions
References and Acknowledgements
Reynolds group
Future Directions
• Synthesized fluorescent sensor, attached amino acid, fluoresces in presence of P.
aeruginosa
• Attached amino acid to model compound
• Attached amino acid to non-fluorescent compound
• Analyzed samples reacted with NO, diazeniumdiolate group on non-fluorescent compound
Measure NO release
Measure antimicrobial activity against bacteria
Antibiotic Resistance Threats in the United States, 2013. CDC.
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