Rhamnolipid-Based Liposomes as Promising Nano-Carriers for Enhancing the Antibacterial Activity of Peptides Derived from Bacterial Toxin-Antitoxin Systems

Beatriz Cristina Pecoraro Sanches,1 Camila Aguiar Rocha,1 Jose Gregorio Martin Bedoya,1 Vinicius Luiz da Silva,2 Patrícia Bento da Silva,3 Ana Marisa Fusco-Almeida,4 Marlus Chorilli,3 Jonas Contiero,2 Edson Crusca,1 Reinaldo Marchetto1 1São Paulo State University (UNESP), Institute of Chemistry, Department of Biochemistry and Organic Chemistry, Araraquara, SP, Brazil; 2São Paulo State University (UNESP), Institute of Biosciences, Department of General and Applied Biology, Rio Claro, SP, Brazil; 3São Paulo State University (UNESP), School of Pharmaceutical Sciences, Department of Drugs and Medicines, Araraquara, SP, Brazil; 4São Paulo State University (UNESP), School of Pharmaceutical Sciences, Department of Clinical Analysis, Araraquara, SP, BrazilCorrespondence: Reinaldo MarchettoUNESP Institute of Chemistry, Rua Prof. Francisco Degni, 55, Araraquara, 14.800-060, SP, BrazilTel +55 16 3301 9670Fax +55 16 3322 2308Email reinaldo.marchetto@unesp.brBackground: Antimicrobial resistance poses substantial risks to human health. Thus, there is an urgent need for novel antimicrobial agents, including alternative compounds, such as peptides derived from bacterial toxin-antitoxin (TA) systems. ParELC3 is a synthetic peptide derived from the ParE toxin reported to be a good inhibitor of bacterial topoisomerases and is therefore a potential antibacterial agent. However, ParELC3 is inactive against bacteria due to its inability to cross the bacterial membranes. To circumvent this limitation we prepared and used rhamnolipid-based liposomes to carry and facilitate the passage of ParELC3 through the bacterial membrane to reach its intracellular target - the topoisomerases.Methods and Results: Small unilamellar liposome vesicles were prepared by sonication from three formulations that included 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine and cholesterol. ParELC3 was loaded with high efficiency into the liposomes. Characterization by DLS and TEM revealed the appropriate size, zeta potential, polydispersity index, and morphology. In vitro microbiological experiments showed that ParELC3 loaded-liposomes are more efficient (29 to 11 μmol·L− 1) compared to the free peptide (> 100 μmol·L− 1) at inhibiting the growth of standard E. coli and S. aureus strains. RL liposomes showed high hemolytic activity but when prepared with POPC and Chol this activity had a significant reduction. Independently of the formulation, the vesicles had no detectable cytotoxicity to HepG2 cells, even at the highest concentrations tested (1.3 mmol·L− 1 and 50 μmol·L− 1 for rhamnolipid and ParELC3, respectively).Conclusion: The present findings suggest the potential use of rhamnolipid-based liposomes as nanocarrier systems to enhance the bioactivity of peptides.Keywords: rhamnolipid liposomes, biosurfactants, bioactive peptides, antimicrobial activity