A cleavable cytolysin-neuropeptide Y bioconjugate enables specific drug delivery and demonstrates intracellular mode of action.
Myxobacterial tubulysins are promising chemotherapeutics inhibiting microtubule polymerization, however, high unspecific toxicity so far prevents their application in therapy. For selective cancer cell targeting, here the coupling of a synthetic cytolysin to the hY1-receptor preferring peptide [F(7),P(34)]-neuropeptide Y (NPY) using a labile disulfide linker is described. Since hY1-receptors are overexpressed in breast tumors and internalize rapidly, this system has high potential as peptide-drug shuttle system. Molecular characterization of the cytolysin-[F(7),P(34)]-NPY bioconjugate revealed potent receptor activation and receptor-selective internalization, while viability studies verified toxicity. Triple SILAC studies comparing free cytolysin with the bioconjugate demonstrated an intracellular mechanism of action regardless of the delivery pathway. Treatments resulted in a regulation of proteins implemented in cell cycle arrest confirming the tubulysin-like effect of the cytolysin. Thus, the cytolysin-peptide bioconjugate fused by a cleavable linker enables a receptor-specific delivery as well as a potent intracellular drug-release with high cytotoxic activity.
Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, Universität Leipzig, Brüderstraße 34, 04103 Leipzig, Germany.
Verena M
VM
Ahrens
Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, Universität Leipzig, Brüderstraße 34, 04103 Leipzig, Germany.
Katja B
KB
Kostelnik
OntoChem GmbH, Heinrich-Damerow-Straße 4, 06120 Halle (Saale), Germany.
Robert
R
Rennert
Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, Universität Leipzig, Brüderstraße 34, 04103 Leipzig, Germany.
David
D
Böhme
Department of Proteomics, Helmholtz Centre for Environmental Research - UFZ, Permoserstraße 15, 04318 Leipzig, Germany.
Stefan
S
Kalkhof
OntoChem GmbH, Heinrich-Damerow-Straße 4, 06120 Halle (Saale), Germany.
David
D
Kosel
OntoChem GmbH, Heinrich-Damerow-Straße 4, 06120 Halle (Saale), Germany.
Lutz
L
Weber
Department of Proteomics, Helmholtz Centre for Environmental Research - UFZ, Permoserstraße 15, 04318 Leipzig, Germany; Department of Metabolomics, Helmholtz Centre for Environmental Research - UFZ, Permoserstraße 15, 04318 Leipzig, Germany; Department of Biotechnology, Chemistry and Environmental Engineering, University of Aalborg, Denmark.
Martin
M
von Bergen
Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, Universität Leipzig, Brüderstraße 34, 04103 Leipzig, Germany. Electronic address: beck-sickinger@uni-leipzig.de.
Annette G
AG
Beck-Sickinger
D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002522:Chlorocebus aethiops; D004220:Disulfides / Q000737:chemistry; D016503:Drug Delivery Systems; D057809:HEK293 Cells; D006801:Humans; D009478:Neuropeptide Y / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology; D054353:Perforin / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology; D017476:Receptors, Neuropeptide Y / Q000235:genetics / Q000378:metabolism*
D004220:Disulfides; D009478:Neuropeptide Y; D017476:Receptors, Neuropeptide Y; D054353:Perforin
Breast cancer; Cytolysin; Neuropeptide Y; Peptide–drug conjugate; Quantitative proteomics; SILAC