Nanotechnology in Medicine Krešimir Pavelić, Marijeta Kralj and Damir Kralj Division of Molecular Medicine and Division of Material Chemistry Ruđer Bošković Institute Zagreb, Croatia

Nanotechnology Antisense therapy gene therapy conventional drug delivery

Antisense Therapy The aim is to interface with gene expression by preventing the translation of proteins from mRNA. Mechanisms of mRNA interactions: sterical blocking of mRNA by antisense binding and destruction antisense mRNA hybrids by RnaseH enzyme formation of triple helix between genomic double-stranded DNA and oligonucleotides the cleavage of target RNA by ribozymes.

Antisense Therapy and Nanoparticles Problems: poor stability of antisense oligonucleotides versus nuclease activity in vitro and in vivo, and their low intracellular penetration have limited their use in therapeutics. Solutions: to increase antisense stability, to improve cell penetration and also to avoid non-specific aptameric effects (leading to non-specific binding of antisense oligonucleotides) the use of particulate carriers such as nanoparticles, has been considered. (The size of nanocapsules - 350 + 100 nm).

Gene Therapy Method for treatment or prevention of genetic disorders based on delivery of repaired or the replacement of incorrect genes. Aimed at treating or eliminating the cause of disease.

Gene Therapy and Nanoparticles Problems: gene delivery, vector immunogenicity, stability. Solutions: nanotechnology in gene therapy would be able to replace the currently used viral vectors by potentially less immunogenic nanosize gene carriers. Vector based on nanoparticles (50 to 500 nm in size) were developed to transport plazmid DNA.

Conventional Drugs and Nanoparticles Problems: delivery by oral route can not be used with most proteins due to both the degradation of these molecules within the intestine and their poor uptake across the intestinal wall. Solutions: pharmaceuticals can be incorporated within biodegradable nanoparticles. This has advantages of protecting the pharmaceuticals from proteolysis within the intestine, or amplifying the uptake capacity of the oral delivery system.

Soluble Inorganic Vector Layered double hydroxides (LDHs) cationic brucite-like layers exchangeable interlayer anions

Hypothesis The unique anion exchange capability of LDHs meet the requirement of inorganic matrices for encapsulating functional biomolecules with negative charge in aqueous media. Such biomolecules can be incorporated between hydroxide layers by a simple ion-exchange reaction to form bio-LDH nanohybrids.

Soluble Inorganic Vector The negatively charged biomolecules intercalated in the gallery spaces would gain extra stabilization energy due to the electrostatic interaction between cationic brucite layers and anionic DNA molecule.

Soluble Inorganic Vector Hydroxide layers can play role of a reservoir to protect intercalated DNA from DNase degradation If desired, the hydroxide layers can be intentionally removed by dissolving in an acidic media, which offers a way of recovering the encapsulated biomolecules.

LDH Interlayer ion Ion exchange Nanohybridization DNA-LDH hybrid Recognition and uptake Schematic illustration of the hybridization and transfer mechanism of the DNA-LDH hybrid into a cell

Conclusions Inorganic supramolecules, such as nanoscale LDHs, can act as biomolecule reservoirs and gene/drug carriers LDH itself is nontoxic Cellular uptake experiments reveal that the FITC-LDH hybrid is effectively transferred into NIH3T3 cells. Antisense myc-LDH hybrid cause strong antitumor effect in vitro. LDHs can act as new inorganic carrier, completely different from existing nonviral vectors in terms of its chemical nature.