Oligonucleotides encapsulation

Ropert, C., Malvy, C., Couvreur, P. (1993). Inhibition of the Friend retrovirus by antisense oligonucleotides encapsulated in liposomes mechanism of action. Pharm. Res., 10, 1427-1433. 

Fenske DB, Maurer N, Cullis PR. Encapsulation of weakly-basic drugs, anti-sense oligonucleotides and plasmid DNA within large unilamellar vesicles for drug delivery applications. In Torchilin VP, Weissig V, eds. Liposomes A Practical Approach. 2nd ed. Oxford Oxford University Press, 2002, Chapter 6. 

Both approaches are simple and allow efficient encapsulation of nucleic acid-based molecules such as oligonucleotides (9,10) and pDNA (8,10,12) in liposomes that are small in size (about lOOnm diameter) and stable in circulation, protecting the cargo from degradation. In the sections to follow, we will provide a brief overview of these methods. 

Encapsulation efficiencies are listed as a function of ethanol concentration for distearoyl-phosphatidyl-choline/choles-terol/l,2-dioleoyl-3-dimethylammoniumpropane large unilamellar vesicle (LUVs). The initial oligonucleotide-to-lipid ratio was 0.034 mol/mol (0.3mg/mg). The LUVs used for these experiments were 99 22 nm in size. The encapsulation values are given as mean SD. 

Figure 2 Encapsulation as a function of ethanol concentration. Oligonucleotides were added to distearoyl-phosphatidyl-choline/cholesterol/l-0-(2 -(co-methoxy-poly-ethylene-glycol)succinoyl)-2-iV-myristoyl-sphingosine /1,2-dioleoyl-3-dimethylam-monium propane liposomes in varying concentrations of ethanol at an initial oligonucleotide-to-lipid ratio of 0.24mg/mg. Abbreviations AS, antisense oligonucleotide %EtOH(v/v), percentage of ethanol in volume/volume.

Semple SC, Klimuk SK, Harasym TO, et al. Efficient encapsulation of antisense oligonucleotides in lipid vesicles using ionizable aminolipids formation of novel small multilamellar vesicle structures. Biochim Biophys Acta 2001 1510 152. 

D Chen, DL Cole, GS Srivatsa. Determination of free and encapsulated oligonucleotides in liposome formulated drug product. J Pharm Biomed Anal 22 791-801, 2000. 

Poly(ethylene oxide)-b-poly (L-lysine) (PEO-h-PLL) Block copolymer In vitro transfection, systemic administration of plasmids, encapsulation of oligonucleotides (Kataoka et al., 1996 Wolfert et al., 1996)... 

Yu RZ, Geary RS, Leeds JM, Watanabe T, Fitchett JR, Matson JE, Mehta R, Hardee GR, Templin MV, Huang K, Newman MS, Quinn Y, Uster P, Zhu G, Working PK, Homer M, Nelson J, Levin AA (1999) Pharmacokinetics and tissue disposition in monkeys of an antisense oligonucleotide inhibitor of Ha-ras encapsulated in stealth liposomes. Pharm Res 16(8) 1309—1315... 

B. Calabretta, G. Zupi, and G. Zon. 1999. Enhanced anti-tumor effects with micro-encapsulated c-myc antisense oligonucleotide. Antisense Nucleic Acid Drug Dev. 9 451-458. 

By encapsulating oligonucleotides in liposomes, it is possible to reduce the acute effects of siRNA compounds. This was demonstrated for a PS ODN in a stealth liposome formulation [50], These types of delivery systems are being increasingly employed with siRNA. Oligonucleotide formulations can reduce the propensity to produce the typical PS ODN effects, but it will be necessary to characterize their acute effects. 

FIGURE 12 Survival of NB-bearing nude mice after injection of oligonucleotides (free or encapsulated within liposomal formulations). Nude mice were injected intravenously with 3.5 x 106 HTLA-230 neuroblastoma cells. After 4h each mouse received 50 ig of oligonucleotides either free or encapsulated in targeted or nontargeted liposomes. Control mice received HEPES-buffered saline. (Reprinted from ref. 385 with permission of Elsevier.)... 

---