Attaining the full therapeutic utility of antisense and siRNA oligonucleotides will require understanding of the biological barriers that stand between initial administration of these drugs and their final actions within cells. This article examines some of the key barriers that affect the biodistribution of oligonucleotides both in molecular form and when they are associated with nanocarriers. An understanding of the biological processes underlying these barriers will aid in the design of more effective delivery systems.
We describe the synthesis and characterization of a 5′ conjugate between a 2′-O-Me phosphorothioate antisense oligonucleotide and a bivalent RGD (arginine–glycine–aspartic acid) peptide that is a high-affinity ligand for the αvβ3 integrin. We used αvβ3-positive melanoma cells transfected with a reporter comprised of the firefly luciferase gene interrupted by an abnormally spliced intron. Intranuclear delivery of a specific antisense oligonucleotide (termed 623) corrects splicing and allows luciferase expression in these cells. The RGD–623 conjugate or a cationic lipid-623 complex produced significant increases in luciferase expression, while ‘free’ 623 did not. However, the kinetics of luciferase expression was distinct; the RGD–623 conjugate produced a gradual increase followed by a gradual decline, while the cationic lipid-623 complex caused a rapid increase followed by a monotonic decline. The subcellular distribution of the oligonucleotide delivered using cationic lipids included both cytoplasmic vesicles and the nucleus, while the RGD–623 conjugate was primarily found in cytoplasmic vesicles that partially co-localized with a marker for caveolae. Both the cellular uptake and the biological effect of the RGD–623 conjugate were blocked by excess RGD peptide. These observations suggest that the bivalent RGD peptide–oligonucleotide conjugate enters cells via a process of receptor-mediated endocytosis mediated by the αvβ3 integrin.
Dendrimer-oligonucleotide complexes were moderately effective for delivery of antisense and only poorly effective for delivery of siRNA. Conjugation of the dendrimer with the Tat cell penetrating peptide failed to further enhance the effectiveness of the dendrimer.
In this contribution we discuss the sintering of an inkjet-printed copper nanoparticle ink based on electrical performance and microstructure analysis. Laser and intense pulsed light (IPL) sintering are employed in order to compare the different techniques and their feasibility for electronics manufacturing. A conductivity of more than 20% of that of bulk copper material has been obtained with both sintering methods. Laser and IPL sintering techniques are considered to be complementary techniques and are highly suitable in different application fields.
We demonstrate intense pulsed light (IPL) sintering of inkjet-printed CuO layers on a primer-coated porous PET substrate to convert the electrically insulating CuO into conductive Cu. With this approach, conductive layers are obtained in less than 1 s after the printing process. The IPL sintering was performed for high productivity with minimum duration and repetition of IPL irradiation to evaluate the effect of pulse number and energy output on the conductivity and morphology of the sintered Cu layers. Depending on the energy output, sheet resistances were measured as 0.355, 0.131, and 0.121 Ω·□(-1) by exposure energy of 5.48 (single pulse), 7.03 (double pulse), and 7.48 J·cm(-2) (triple pulse), respectively. In contrast, an excessive energy with relatively short pulse duration causes a delamination of the Cu layer. The lowest resistivity of about 55.4 nΩ·m (corresponds to about 30% conductivity of bulk Cu) was obtained by an IPL sintering process of 0.26 s after the printing, which was composed of 2 ms triple pulses with 10 Hz frequency.
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