Dendrimer oligonucleotide

The surface of each layer has two types of single stranded arms (e.g. one 3 — 5 and one 5 — 3 strand) which can bind to other monomers to render 1st-and 2nd-generation dendrimers 25 and 26. Therefore the molecular scaffold grows exponentially with each sequential layer of hybridization. If an oligonucleotide contains a sequence complementary to those at the surface of these networks it should be hybridized. The remaining free sequences from the other type of arms then bind in a standard nucleic acid blot (after they are bound to... 

Dendrimers bearing certain fluorescent molecules attached on their periphery have been shown to be environmentally sensitive probes for the presence of certain metal ions or to changes in pH (Balzani et al., 2000 Paola et al., 2005). In addition, PAMAM dendrimers modified with the relatively hydrophobic dye Oregon Green 488 were shown to be a more effective transfection agent for anti-sense oligonucleotides than the dendrimer alone-plus the complex could be tracked within the cell due to the fluorescence of the dye (Yoo and Juliano, 2000). 

Yoo, H., and Juliano, R.L. (2000) Enhanced delivery of antisense oligonucleotides with fluorophore-con-jugated PAMAM dendrimers. Nucleic Acids Res. 28, 4225-4231. 

Particles from cationic lipids may also be useful for antisense therapy of skin disease — a nontoxic increase in the oligonucleotide uptake by cultivated keratinocytes and a sebocyte cell line has been reported [66]. Moreover, cationic dendri-mers also efficiently transfer reporter gene DNA to human keratinocytes cultivated in vitro. In the skin of hairless mice, in vivo transfection was possible with complexes, yet reporter gene expression was localized to perifollicular areas. Transfection, however, failed with the naked plasmid. For prolonged contact, biodegradable membranes coated with dendrimer/DNA complexes were used [67]. This hints at a follicular uptake of these complexes and indicates that gene transfection also may be possible with human skin, which has a thicker stratum comeum compared with mouse skin (eight to ten vs. two to three layers [58]). 

One particular dendrimer system was reported to have increased the loading capacity 10-fold over direct coupling of the oligonucleotide to the surface. Unfortunately, the presumed improvement in hybridization efficiency was not discussed in quantitative terms. Reusability of the array was a primary objective of the work. Again, quantitative information was not provided. As the authors note, the array withstood >7 actual hybridization stripping cycles and 30 simulated (lacking target) cycles. 

Poly(ethylene oxide)-b-poly (L-lysine) dendrimer (PEO-b-dendr PLL) Block copolymer, dendrimer PLL segment Encapsulation of plasmids and oligonucleotides (Choi et al, 1999)... 

Bielinska, A., Kukowska-Latallo, J.F., Johnson, J., Tomalia, D.A. and Baker, Jr., J.R. (1996) Regulation of in vitro gene expression using antisense oligonucleotides or antisense expression plasmids transfected using starburst PAMAM dendrimers. Nucleic Acids Res., 24, 2176-2182. 

Access to nucleic acid dendrimers is initiated by a zip-fastener like dissociation of the DNA double strand by heating. The double strand separates into the two individual strands by thermal motion (denaturation). Subsequent association, hybridisation of complementary sequences, is followed by stepwise cross-linking to form DNA dendrimers, which can contain up to two million oligonucleotide-end group strands (Fig. 8.19). The latter can be labelled with fluorescence or radioactive markers. 

Hybridisation can thus be used to produce probes, for example, for the detection of oligonucleotides in chick embryo tissue or of Epstein-Barr viruses (glandular fever) in transplant recipients - by detection of specific RNA strands. DNA dendrimers and corresponding detection devices have meanwhile become commercially available (Genisphere company). Fluorescence-labelled polynucleotide dendrimers have also been used for signal intensification in DNA microchip technology [59]. 

Fig. 24. Various oligonucleotide branch cell reagents [ A), (B), (C), (D)] suitable for constructing DNA/RNA dendrons and dendrimers...

Parekh, H. S., et al. (2006), Synthesis of a library of polycationic lipid core dendrimers and their evaluation in the delivery of an oligonucleotide with hVEGF inhibition, Bioorg. Med. Chem., 14(14), 4775 1780. 

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