title: Native chemical ligation for conversion of sequence-defined oligomers into targeted pDNA and siRNA carriers.
abstract: Native chemical ligation (NCL) was established for the conversion of sequence-defined oligomers of different topologies into targeted and PEG shielded pDNA and siRNA carriers. From an existing library of non-targeted oligoethanamino amides, six oligomers containing N-terminal cysteines were selected as cationic cores, to which monodisperse polyethylene glycol (PEG) containing terminal folic acid as targeting ligand (or terminal alanine as targeting negative control ligand) were attached by NCL. Ligated conjugates plus controls (in sum 18 oligomers) were evaluated for pDNA or siRNA gene delivery. Biophysical characteristics including nucleic acid binding in the absence or presence of serum, as well as biological activities in cellular uptake and gene transfer (or gene silencing, respectively) were determined. In most cases, the folic acid-PEG-ligated oligomers displayed a strongly improved cellular binding, uptake and gene transfer into receptor-positive KB cells as compared to the alanine-PEG controls. Changing the topological structures by increasing the number of cationic arms, adding tyrosine trimers as polyplex stabilizing domains, or histidines facilitating endosomal escape resulted in beneficial gene transfer characteristics. The screen revealed different requirements for pDNA and siRNA delivery. A folate-PEG ligated histidinylated four-arm oligomer was most effective for pDNA delivery but inactive for siRNA, whereas a folate-PEG-ligated three-arm oligomer with tyrosine trimer modifications was most effective in siRNA mediated gene silencing. The results demonstrate the site-selective NCL reaction as powerful method to modify existing oligomers. Thus multifunctional targeted carriers can be obtained with ease and used to identify lead structures for subsequent in vivo delivery.
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany; School of Chemistry and Chemical Engineering, South China University of Technology, 510640 Guangzhou, People's Republic of China.
authors.forename: Can Yang
authors.initials: CY
authors.lastname: Zhang
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany; Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany.
authors.forename: Petra
authors.initials: P
authors.lastname: Kos
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany; Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Müller
authors.affiliation: Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany; Physical Chemistry, Department of Chemistry, Munich Center for Integrated Protein Science (CiPSM), Germany.
authors.forename: Waldemar
authors.initials: W
authors.lastname: Schrimpf
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Troiber
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany; Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Lächelt
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany; Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Scholz
authors.affiliation: Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany; Physical Chemistry, Department of Chemistry, Munich Center for Integrated Protein Science (CiPSM), Germany.
authors.forename: Don C
authors.initials: DC
authors.lastname: Lamb
authors.affiliation: Pharmaceutical Biotechnology, Center for NanoScience (CeNS), Ludwig-Maximilians-University (LMU) Munich, Butenandtstrasse 5-13, D-81377 Munich, Germany; Nanosystems Initiative Munich, Schellingstraße 4, D-80799 Munich, Germany. Electronic address: ernst.wagner@cup.uni-muenchen.de.
authors.forename: Ernst
authors.initials: E
authors.lastname: Wagner
mesh_terms: D000577:Amides / Q000737:chemistry; D002460:Cell Line; D003545:Cysteine / Q000031:analogs & derivatives; D004247:DNA / Q000008:administration & dosage* / Q000235:genetics; D004337:Drug Carriers / Q000737:chemistry*; D005492:Folic Acid / Q000031:analogs & derivatives; D018014:Gene Transfer Techniques; D006801:Humans; D010957:Plasmids / Q000008:administration & dosage* / Q000235:genetics; D011092:Polyethylene Glycols / Q000737:chemistry; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics
chemical_list: D000577:Amides; D004337:Drug Carriers; D034741:RNA, Small Interfering; D011092:Polyethylene Glycols; D004247:DNA; D005492:Folic Acid; D003545:Cysteine
keywords: Folate; Gene delivery; Library screening; Native chemical ligation; pDNA; siRNA