Drug delivery of antisense oligonucleotides or peptides to tissuesin vivousing an avidin–biotin system

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Abstract
The pharmacologic efficacy of antisense oligonucleotide- or peptide-based therapeutics in intact organisms is limited by the poor transport of these highly charged molecules across cellular membranes. Since recent studies have shown that avidin, a cationic protein, is taken up by cells via absorptive-mediated endocytosis, the present studies examine whether the avidin-biotin system may be used to develop soluble transport vectors that mediate the organ uptake in vivo of monobiotinylated antisense oligonucleotides or peptides. The model antisense oligonucleotide used is a 21-mer complementary to the bovine GLUT1 glucose transporter mRNA, and the model peptide used is [desamino-Cys1, d-Lys8]lysine vasopressin (DDLVP). The antisense oligonucleotide was bound to avidin with a noncleavable biotin linker using N-hydroxysuccinimidobiotin, and the DDLVP was coupled to the avidin vector with a cleavable biotin linker using sulfosuccinimidyl - 2 - (biotinamido) ethyl -1, 3 - dithiopropionate. Pharmacokinetic studies showed that avidin was cleared biexponentially from the circulation with half-times of 22 ± 13 s and 53 ± 5 min. Conversely, streptavidin, a slightly acidic protein, was cleared monoexponentially with a half-time of 2.4 ± 0.2 h. The volume of distribution (Vd) of [3H]biotin in liver and kidney was enriched nearly 1000-fold 2 h after a single injection when the biotin was co-injected with avidin compared to when it was co-injected with streptavidin. The hepatic clearance of the vasopressin analogue was in-creased manyfold by co-injection with avidin, and the increased organ uptake was blocked by pretreatment of the conjugate with sulfhydryl reducing agents. The clearance of the oligonucleotide in liver and kidney was greatly increased by avidin co-injection but not by streptavidin co-injection. In conclusion, these studies demonstrate that the avidin–biotin system provides a simple and efficient approach to drug delivery of biotinylated antisense oligonucleotide- or peptide-based therapeutics to tissues in vivo.

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