# Genetically-encoded fluorescent RNA sensors for measuring transport of antibiotics into the cytoplasm of Gram-negative pathogens and development of efflux pump inhibitors

> **NIH NIH R01** · HARVARD MEDICAL SCHOOL · 2020 · $703,931

## Abstract

Project Summary
Complications from infections caused by difficult to treat Gram-negative pathogens resistant to
multiple antibiotics (the MDR phenotype) has become a major public health threat. The basis of
resistance in the most important members of this group, including Pseudomonas aeruginosa, is
the poor penetration of the antibiotics through a relatively impermeable bacterial cell envelope
and their expulsion (efflux) by various pumps expressed by the microorganisms. Consequently,
the intracellular concentrations of antibiotic are kept below bactericidal levels. This proposal
aims at creating new classes of diagnostic tools and therapeutic agents to combat efflux-based
resistance by a project organized into two phases. First, we will develop genetically encoded
fluorescent sensors activated by binding of antibiotics following their entry into the bacterial
cytoplasm. This tool will enable us to measure antibiotic flux under different environmental
conditions and on a single cell basis. During the second phase of the project, working with our
industrial partner, we will implement a screening program, using a library of several hundred
billion small molecules, to identify inhibitors targeting different components of efflux pumps.
These inhibitors should activate the fluorescent detector by causing enhanced intracellular
accumulation of the antibiotics resulting from inhibition of efflux. Functional studies are proposed
to assess the ability of the active compounds to promote killing of P. aeruginosa by antibiotics
that are normally expelled by the efflux pumps. Crystal structures of inhibitors bound to target
proteins will be determined and these will guide medicinal chemistry efforts with the goal of
creating additional derivatives with increased potencies and expanded spectrum, capable of
inhibiting efflux pumps of other important Gram-negative pathogens.

## Key facts

- **NIH application ID:** 9841903
- **Project number:** 5R01AI136789-03
- **Recipient organization:** HARVARD MEDICAL SCHOOL
- **Principal Investigator:** STEPHEN LORY
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $703,931
- **Award type:** 5
- **Project period:** 2018-02-13 → 2023-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9841903

## Citation

> US National Institutes of Health, RePORTER application 9841903, Genetically-encoded fluorescent RNA sensors for measuring transport of antibiotics into the cytoplasm of Gram-negative pathogens and development of efflux pump inhibitors (5R01AI136789-03). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/9841903. Licensed CC0.

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