Druggability of G-quadruplexes, promising modulators for antimicrobial resistance

Project Information

The G-Q-reat ESKAPE

Grant agreement ID: 101106871

DOI

10.3030/101106871

Project closed

EC signature date 7 July 2023

Start date 1 September 2023

End date 31 August 2025

Funded under

• Marie Skłodowska-Curie Actions (MSCA)

Total cost

No data

EU contribution

€ 172 750,08

Coordinated by

UNIVERSITA DEGLI STUDI DI SIENA
Italy

Periodic Reporting for period 1 - The G-Q-reat ESKAPE (Druggability of G-quadruplexes, promising modulators for antimicrobial resistance)

Reporting period: 2023-09-01 to 2025-08-31

Summary of the context and overall objectives of the project

Antimicrobial resistance (AMR) is one of today’s most pressing global health threats, responsible for millions of deaths every year. "The G-Q-reat ESKAPE" project (Druggability of G-quadruplexes, promising modulators for antimicrobial resistance) explored an entirely new class of bacterial drug targets, namely bacterial G-quadruplexes (G4s). These are four-stranded DNA structures that can regulate key genes involved in replication, virulence and antibiotic resistance. "The G-Q-reat ESKAPE" project combined computational chemistry, molecular dynamics (MD) simulations and biophysical experiments to identify, model and validate bG4s as potential antimicrobial targets, focusing on six priority pathogens of the WHO “ESKAPE” list: Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Staphylococcus aureus and Streptococcus pneumoniae.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

During the two-year fellowship (2023 - 2025), at least one G4 structure was modeled and analyzed for each target bacterium. Advanced MD simulations were used to elucidate loop dynamics, cation dependence and topology, while circular dichroism (CD) and UV-Vis spectroscopy confirmed the folding and stability of selected bacterial G4s.

• For K. pneumoniae, the study revealed how structural features influence G4 topology, thermal profiles and ligand binding.
• In P. aeruginosa, CD and thermal melting profiles led to the identification of a natural compound able to bind and stabilize pivotal G4s.
• In A. baumannii, new G4s were discovered within genes essential for nutrient uptake, confirmed experimentally to adopt hybrid or antiparallel folds.
• In parallel, a new mixed-solvent MD pipeline was created to discover G4-binding molecules. This innovative workflow bridges virtual screening and experimental validation and will be openly shared with the research community.

A comprehensive open-access review, “The Rise of Bacterial G-Quadruplexes in Current Antimicrobial Discovery” (ACS Omega 2024), already disseminates the conceptual framework of "The G-Q-reat ESKAPE" project. Three manuscripts are in preparation on (i) K. pneumoniae G4 structures, (ii) the mixed-solvent MD pipeline, and (iii) ligand binding to P. aeruginosa G4s.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

"The G-Q-reat ESKAPE" project established, for the first time, a systematic approach to treat bacterial G4s as druggable antimicrobial targets, shifting the paradigm from protein- to nucleic-acid-centered drug discovery. The outcomes include:

• 3D structural models of bG4s across six pathogens;
• a validated in vitro/in silico workflow for identifying G4 ligands;
• new hit compounds that stabilize P. aeruginosa G4s;
• a computational pipeline for rapid target validation in emerging pathogens.

These advances open new avenues to design next-generation antimicrobials acting on conserved, mutation-resistant genomic regions. By enabling faster response to outbreaks and reducing dependence on classical antibiotics, "The G-Q-reat ESKAPE" project directly supports EU priorities in health, research innovation and antimicrobial stewardship.

Summary and pipeline of the "The G-Q-reat ESKAPE" project