Synthetic mRNA delivered to human cells leads to expression of Cpl-1 bacteriophage-endolysin with activity against Streptococcus pneumoniae

Moritz K Jansson; Dat Tien Nguyen; Stefan Mikkat; Carolin Warnke; Marc Benjamin Janssen; Philipp Warnke; Bernd Kreikemeyer; Nadja Patenge

doi:10.1016/j.omtn.2024.102145

Synthetic mRNA delivered to human cells leads to expression of Cpl-1 bacteriophage-endolysin with activity against Streptococcus pneumoniae

Mol Ther Nucleic Acids. 2024 Feb 8;35(1):102145. doi: 10.1016/j.omtn.2024.102145. eCollection 2024 Mar 12.

Authors

Moritz K Jansson¹, Dat Tien Nguyen¹, Stefan Mikkat², Carolin Warnke¹, Marc Benjamin Janssen¹, Philipp Warnke¹, Bernd Kreikemeyer¹, Nadja Patenge¹

Affiliations

¹ Institute of Medical Microbiology, Virology and Hygiene, University Medicine Rostock, Rostock, Germany.
² Core Facility Proteome Analysis, University Medicine Rostock, Rostock, Germany.

Abstract

Endolysins are bacteriophage-encoded hydrolases that show high antibacterial activity and a narrow substrate spectrum. We hypothesize that an mRNA-based approach to endolysin therapy can overcome some challenges of conventional endolysin therapy, namely organ targeting and bioavailability. We show that synthetic mRNA applied to three human cell lines (HEK293T, A549, HepG2 cells) leads to expression and cytosolic accumulation of the Cpl-1 endolysin with activity against Streptococcus pneumoniae. Addition of a human lysozyme signal peptide sequence translocates the Cpl-1 to the endoplasmic reticulum leading to secretion (hlySP-sCpl-1). The pneumococcal killing effect of hlySP-sCpl-1 was enhanced by introduction of a point mutation to avoid N-linked-glycosylation. hlySP-sCpl-1N215D, collected from the culture supernatant of A549 cells 6 h post-transfection showed a significant killing effect and was active against nine pneumococcal strains. mRNA-based cytosolic Cpl-1 and secretory hlySP-sCpl-1N215D show potential for innovative treatment strategies against pneumococcal disease and, to our best knowledge, represent the first approach to mRNA-based endolysin therapy. We assume that many other bacterial pathogens could be targeted with this novel approach.

Keywords: Cpl-1 endolysin; MT: Delivery Strategies; Streptococcus pneumoniae; bacterial infection; bacteriophage endolysins; infectious diseases; mRNA therapy; mRNA-based antibacterial therapy; mRNA-based endolysins; mRNA-encoded endolysins.