CIT8, a machine learning-optimized antimicrobial peptide, exhibits potent activity against carbapenem-resistant Acinetobacter baumannii
6 affiliations
- 1Department of Medicine, Houston Methodist Hospital
- 2Laboratory of Medicinal Chemistry, Section of Pharmaceutical Chemistry, Department of Pharmacy, National and Kapodistrian University of Athens
- 3Department of Medicine, Weill Cornell Medical College
- 4School of Pharmaceutical Sciences, Rajiv Gandhi Technological University
- 5School of Biomolecular Engineering and Biotechnology, Rajiv Gandhi Technological University
- 6Department of Electrical and Computer Engineering, National Technical University of Athens
Abstract
As the burden of antimicrobial resistance continues to rise worldwide, the need for novel antimicrobial therapeutics has become increasingly urgent. Machine learning (ML) has emerged as a promising strategy for accelerating antimicrobial peptide discovery; however, experimental characterization remains essential to define the biological properties and broader applicability of ML-optimized peptides. CIT8 is a 13-residue, citropin-based peptide previously developed through ML-optimization with potent antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA). Here, we demonstrate that its activity extends to multidrug-resistant Gram-negative pathogens. Among these, CIT8 displayed the most potent activity against Acinetobacter baumannii , maintaining an MIC of 4 μg/mL against 15 of 16 A. baumannii strains tested, including multiple carbapenem-resistant (CRAB) clinical isolates. At 20 μg/mL (5 × MIC), CIT8 reduced an initial population of approximately 5 × 10 ⁷ CFU/mL of exponential-phase A. baumannii to below the limit of detection within 15 min. Consistent with its rapid bactericidal activity, CIT8 induced rapid, dose-dependent membrane depolarization and permeabilization, accompanied by leakage of intracellular ATP. In addition, CIT8 and, even more notably, its D-enantiomer CIT8-D exhibited strong antibiofilm activity while maintaining relatively low mammalian cytotoxicity, with therapeutic indices of >32 for CIT8 in both cell lines and 16.2 and 18.5 for CIT8-D in HaCaT and HKC-8 cells, respectively. Moreover, CIT8 potentiated the bactericidal activity of meropenem against CRAB, with the combination exhibiting bactericidal synergy by time-kill assay. These findings highlight CIT8 as a promising scaffold for further optimization and preclinical evaluation against multidrug-resistant A. baumannii .
Related
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BibTeX
@article{dellis2026cit8,
title = {{CIT8, a machine learning-optimized antimicrobial peptide, exhibits potent activity against carbapenem-resistant Acinetobacter baumannii}},
author = {Charilaos Dellis and Biswajit Mishra and Liyang Zhang and Anindya Basu and Lewis Oscar Felix and Fadi Shehadeh and Narchonai Ganesan and Antonios Kolocouris and Eleftherios Mylonakis},
journal = {Frontiers in Microbiology},
year = {2026},
month = oct,
doi = {10.3389/fmicb.2026.1967486},
url = {https://www.frontiersin.org/articles/10.3389/fmicb.2026.1967486/full},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - CIT8, a machine learning-optimized antimicrobial peptide, exhibits potent activity against carbapenem-resistant Acinetobacter baumannii
AU - Charilaos Dellis
AU - Biswajit Mishra
AU - Liyang Zhang
AU - Anindya Basu
AU - Lewis Oscar Felix
AU - Fadi Shehadeh
AU - Narchonai Ganesan
AU - Antonios Kolocouris
AU - Eleftherios Mylonakis
PY - 2026
DA - 2026/10/19
JO - Frontiers in Microbiology
DO - 10.3389/fmicb.2026.1967486
UR - https://www.frontiersin.org/articles/10.3389/fmicb.2026.1967486/full
AB - As the burden of antimicrobial resistance continues to rise worldwide, the need for novel antimicrobial therapeutics has become increasingly urgent. Machine learning (ML) has emerged as a promising strategy for accelerating antimicrobial peptide discovery; however, experimental characterization remains essential to define the biological properties and broader applicability of ML-optimized peptides. CIT8 is a 13-residue, citropin-based peptide previously developed through ML-optimization with potent antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA). Here, we demonstrate that its activity extends to multidrug-resistant Gram-negative pathogens. Among these, CIT8 displayed the most potent activity against Acinetobacter baumannii , maintaining an MIC of 4 μg/mL against 15 of 16 A. baumannii strains tested, including multiple carbapenem-resistant (CRAB) clinical isolates. At 20 μg/mL (5 × MIC), CIT8 reduced an initial population of approximately 5 × 10 ⁷ CFU/mL of exponential-phase A. baumannii to below the limit of detection within 15 min. Consistent with its rapid bactericidal activity, CIT8 induced rapid, dose-dependent membrane depolarization and permeabilization, accompanied by leakage of intracellular ATP. In addition, CIT8 and, even more notably, its D-enantiomer CIT8-D exhibited strong antibiofilm activity while maintaining relatively low mammalian cytotoxicity, with therapeutic indices of >32 for CIT8 in both cell lines and 16.2 and 18.5 for CIT8-D in HaCaT and HKC-8 cells, respectively. Moreover, CIT8 potentiated the bactericidal activity of meropenem against CRAB, with the combination exhibiting bactericidal synergy by time-kill assay. These findings highlight CIT8 as a promising scaffold for further optimization and preclinical evaluation against multidrug-resistant A. baumannii .
SN - 1664-302X
ER -