الفهرس 
Antisense oligonucleotide in dermatologyOnly 14 pages are availabe for public view
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Abstract
Antisense oligonucleotides (ASOs) are synthetic single-stranded DNA fragments that bind to messenger RNA to stop translation of the mRNA and hence protein synthesis expressed by the targeted gene. The sequence complementarity of mRNA is referred to as the “sense” strand because it is “read” directly by ribosomes in the assembly of amino acids. Oligonucleotides designed to hybridize with this “sense” nucleic acid necessarily have the opposite complementarity, analogous to the double- stranded pairing in nuclear DNA and therefore, are referred to as “antisense”.
The first step in antisense therapeutic development is the identification of a protein as a potential therapeutic target in a disease process. The ability of an ASO to form a hybrid depends on its binding affinity and sequence specificity.
Mechanism of action of antisense oligonucleotide includes formation of the ASO–mRNA heteroduplex either triggers RNase H activity, leading to mRNA degradation, induces translational arrest by steric hindrance of ribosomal activity, interferes with mRNA maturation by inhibiting splicing or destabilizes pre-mRNA in the nucleus, resulting in down-regulation of target protein expression.
Factors affecting mechanism of action of antisense oligonucleotides include stability against nucleases, the ability to be taken up by cells, a strong affinity and specificity for the target mRNA and the ability to elicit RNase H for RNA cleavage. ASOs can also display properties that have an adverse effect on their efficiency. These include the poly-anionic nature of some ASOs and the ability to stimulate the immune system.
Synthetic antisense oligonucleotide should activate RNase H–degradation pathways ,Be easy to make and inexpensive ,Not be physiologically toxic ,Not be easily degraded ,Not disrupt normal Watson–Crick base-pairing ,Not induce any unanticipated sequence-independent biologic effects.
ASOs are natural phosphodiester compounds. However, critical drawbacks such as their poor stability versus nuclease activity in vitro and in vivo and their low intracellular penetration and low bioavailability have limited their use in therapeutics. As a result, clinical applications of antisense oligonucleotides have required chemical modifications with the aim of retaining their capacity to knock down protein expression while increasing stability and cellular penetration.
First generation antisense oligonucleotides are those containing a Phosphorothioate (PS)-modified backbone, in which one of the non-bridging oxygen atoms in the phosphodiester bond is replaced by a sulphur atom. Phosphorothioate modification confers higher resistance against nuclease degradation leading to higher bioavailability of the oligonucleotide and promote RNase H-mediated cleavage of target mRNA.
To further enhance nuclease resistance and increase binding affinity for target mRNA, second-generation ASOs with 2´-alkyl modifications of the ribose were developed. 2´-O-Methyl (2´-OMe) and 2´-O-Methoxyethyl (2´-MOE) modifications of PS- modified ASOs are the two most widely studied second-generation ASOs but they do not support RNase H-mediated cleavage of target mRNA which dampens the efficacy of the ASO to circumvent this shortcoming a chimeric ASO was developed.
To further enhance target affinity, nuclease resistance, biostability and pharmacokinetics, a third generation of ASO was developed mainly by chemical modifications of the furanose ring of the nucleotide. Peptide nucleic acid (PNA), locked nucleic acid (LNA) and phosphoroamidate morpholino oligomer (PMO) are the three most studied third-generation ASOs.
It is very important to meet all the critical parameters corresponding to an optimal activity: improved stability, increased blood half-life, tissue and cellular targeting, improve cellular penetration, release the nucleic acids in the right intracellular compartment.
The stratum corneum acts as the major barrier to penetration of molecules across the skin. Large negatively charged molecules such as antisense oligonucleotides do not easily penetrate the stratum corneum as they have low lipid solubility.
In order to down-regulate gene expression, ASOs must penetrate into the targeted cells and reach the cytoplasm. Uptake occurs through active transport.
Polymeric carriers or the use of alternative administration route such as oral for obtaining systemic absorption or to produce a local effect (e.g. ocular and skin) are used to improve their delivery and to circumvent the hurdles for their clinical applications.
Advantages of the skin as an antisense target, antisense treatment of diseases that require systemic delivery of the oligonucleotides has been complicated in the past by generalised toxicity, particularly of phosphorothioate oligonucleotides causing thrombocytopaenia, lymphoid hyperplasia and renal tubular degeneration. The systemic absorption of a topically delivered oligonucleotide is likely to be significantly less than for intravenous administration, due to the relatively low lipophilicity of these molecules. The attraction of antisense therapy for skin diseases therefore, is great .It is suggested that the use of antisense oligonucleotides as cutaneous drugs will similarly provide major advantages over intravenous antisense oligonucleotides. Cutaneous delivery will allow direct access to target cells in the skin and will minimize systemic toxicity.
The potential applications of antisense oligonucleotides in the skin are in viral skin diseases, Wound healing, Vitiligo, Diseases of skin pigmentation Non-malignant hyperplasias of the skin (Psoraisis), Atopic dermatitis, Androgenetic alopecia, Malignant melanoma, Non-melanoma skin cancers, Merkel cell carcinoma, and Kaposi’ sarcoma