A "dock and lock" approach to preparation of targeted liposomes.
We developed a strategy for covalent coupling of targeting proteins to liposomes decorated with a standard adapter protein. This strategy is based on "dock and lock" the interactions between two mutated fragments of human RNase I, a 1-15-aa fragment with the R4C amino acid substitution, (Cys-tag), and a 21-127-aa fragment with the V118C substitution, (Ad-C). Upon binding to each other, Cys-tag and Ad-C spontaneously form a disulfide bond between the complimentary 4C and 118C residues. Therefore, any targeting protein expressed with Cys-tag can be easily coupled to liposomes decorated with Ad-C. Here, we describe the preparation of Ad-liposomes followed by coupling them to two Cys-tagged targeted proteins, human vascular endothelial growth factor expressed with N-terminal Cys-tag, and a 254-aa long N-terminal fragment of anthrax lethal factor carrying C-terminal Cys-tag. Both proteins retain functional activity after coupling to Ad-C-decorated drug-loaded liposomes. We expect that our "dock and lock" strategy will open new opportunities for development of targeted therapeutic liposomes for research and clinical use.
Backer
Marina V
MV
SibTech, Inc., Brookfield, CT, USA.
Backer
Joseph M
JM
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001427:Bacterial Toxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008081:Liposomes / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D008081:Liposomes; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; D042461:Vascular Endothelial Growth Factor A; C030325:anthrax toxin; D012259:Ribonuclease, Pancreatic; D003545:Cysteine