# Mechanism of Resistance Avoidance in Synthetically Evolved Antibacterial Peptides

> **NIH NIH R21** · TULANE UNIVERSITY OF LOUISIANA · 2022 · $228,000

## Abstract

Summary
In the ongoing fight against drug resistant bacterial infections it will be critical to identify novel antibiotic
chemotypes that are less likely to induce resistance. We have used synthetic molecular evolution (SME), iterative
library design and screening, to identify novel antimicrobial peptides that have potent broad-spectrum sterilizing
activity against all ESKAPE pathogens tested, including drug-resistant and biofilm-forming pathogens, in vitro
and in vivo. These peptides are highly soluble, protease resistant, have low cytotoxicity, and are active in the
presence of concentrated host cells and/or serum proteins. Most importantly, they do not induce resistance in P.
aeruginosa under conditions that rapidly enable development of resistance to conventional antibiotics and some
AMPs. Resistance to some other AMPs has been reported, and it has been experimentally selected. However,
avoidance or delay in resistance to AMPs have also been reported. Here we seek to understand the mechanism
of resistance-avoidance. The most commonly observed AMP resistance mechanism, by far, is the modification
of Lipid A phosphates with cationic moieties, which dramatically reduces the potential for AMP interactions with
the outer membrane(OM). We have previously shown that some AMPs bind strongly and accumulate
significantly on OM LPS. In this work we will test the hypothesis that peptides requiring large scale accumulation
on the outer membrane LPS, including Lipid A, before they can cross it will be able to drive selection for
resistance using this mechanism. AMPs that can bypass the OM to reach the inner membrane target without
requiring large scale accumulation on the OM will be slower or unable to elicit resistance. To test this hypothesis,
we have identified two related AMPs, one that rapidly invokes resistance in Pseudomonas aeruginosa, and one
that does not invoke any resistance at all in multiple passages under the same conditions. We will use these two
peptides, and others, to study resistance and resistance-avoidance mechanisms in P. aeruginosa and other
Gram-negative bacteria using i) direct measurements of AMP-bacteria binding, ii) genetic assessment of
resistance pathways, iii) direct assessment of Lipid A modifications. We will also test for resistance avoidance in
a panel of other Gram-negative pathogens to test for the generality of the mechanism.

## Key facts

- **NIH application ID:** 10412134
- **Project number:** 5R21AI154284-02
- **Recipient organization:** TULANE UNIVERSITY OF LOUISIANA
- **Principal Investigator:** WILLIAM C WIMLEY
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $228,000
- **Award type:** 5
- **Project period:** 2021-06-01 → 2024-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10412134

## Citation

> US National Institutes of Health, RePORTER application 10412134, Mechanism of Resistance Avoidance in Synthetically Evolved Antibacterial Peptides (5R21AI154284-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10412134. Licensed CC0.

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