title: Combined MUC1-specific nanobody-tagged PEG-polyethylenimine polyplex targeting and transcriptional targeting of tBid transgene for directed killing of MUC1 over-expressing tumour cells.
abstract: We provide evidence for combining a single domain antibody (nanobody)-based targeting approach with transcriptional targeting as a safe way to deliver lethal transgenes to MUC1 over-expressing cancer cells. From a nanobody immune library, we have isolated an anti-DF3/Mucin1 (MUC1) nanobody with high specificity for the MUC1 antigen, which is an aberrantly glycosylated glycoprotein over-expressed in tumours of epithelial origin. The anti-MUC1 nanobody was covalently linked to the distal end of poly(ethylene glycol)(3500) (PEG(3500)) in PEG(3500)-25kDa polyethylenimine (PEI) conjugates and the resultant macromolecular entity successfully condensed plasmids coding a transcriptionally targeted truncated-Bid (tBid) killer gene under the control of the cancer-specific MUC1 promoter. The engineered polyplexes exhibited favourable physicochemical characteristics for transfection and dramatically elevated the level of Bid/tBid expression in both MUC1 over-expressing caspase 3-deficient (MCF7 cells) and caspase 3-positive (T47D and SKBR3) tumour cell lines and, concomitantly, induced considerable cell death. Neither transgene expression nor cell death occurred when the MUC1 promoter was replaced with the CNS-specific synapsin I promoter. Since PEGylated PEI was only responsible for DNA compaction and played no significant role in direct transfection and cell killing, our attempts overcome previously reported PEI-mediated apoptotic and necrotic cell death, which is advantageous for future in vivo transcriptional targeting as this will minimize (or eliminate) non-targeted cell damage.
authors.affiliation: Department of Medical Biotechnology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: Elham
authors.initials: E
authors.lastname: Sadeqzadeh
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Rahbarizadeh
authors.forename: Davoud
authors.initials: D
authors.lastname: Ahmadvand
authors.forename: Mohammad J
authors.initials: MJ
authors.lastname: Rasaee
authors.forename: Ladan
authors.initials: L
authors.lastname: Parhamifar
authors.forename: S Moein
authors.initials: SM
authors.lastname: Moghimi
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000008:administration & dosage* / Q000235:genetics; D051023:BH3 Interacting Domain Death Agonist Protein / Q000235:genetics*; D001943:Breast Neoplasms / Q000628:therapy; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D004337:Drug Carriers / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D018390:Gene Targeting; D041722:Genes, Transgenic, Suicide; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D051379:Mice; D018396:Mucin-1 / Q000235:genetics*; D053758:Nanoparticles / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000737:chemistry*; D011094:Polyethyleneimine / Q000031:analogs & derivatives* / Q000737:chemistry; D014162:Transfection
chemical_list: D000911:Antibodies, Monoclonal; D051023:BH3 Interacting Domain Death Agonist Protein; D004337:Drug Carriers; C060192:MUC1 protein, human; D018396:Mucin-1; C544016:poly(ethylene glycol)-co-poly(ethyleneimine); D011092:Polyethylene Glycols; D011094:Polyethyleneimine; D053148:Caspase 3