SMALL RNA AS ANTIAGING COSMECEUTICALS 457 microRNAs (miRNA) and small interference RNAs (siRNA) are classes of regulating small RNA molecules, ranging from 18 to 24 nucleotides in length. They function by altering the stability or translational effi ciency of messenger RNAs (mRNAs) with which they share sequence complementarity, as shown in Figure 1, and are predicted to affect up to one-third of all human genes (16). RNAi-based therapeutics has recently emerged for treatment of cancer, infectious dis- eases, and other diseases associated with specifi c gene disorders. Since the discovery of RNAi, there have been more than 30 clinical trials involving 21 different biological drugs known as siRNA or miRNA (18). In principle, the RNAi mechanism elicits a posttranscriptional gene silencing phenomenon capable of inhibiting specifi c gene func- tion with high potency at a few nanomolar dosages. The molecular mechanisms underly- ing some aspects of aging are now being unraveled, certain aging processes are associated with personal gene activities (8,19–21). Therefore, small RNA agents could potentially be used to develop new cosmetic designs and products by suppressing those unwanted gene activities, in particular for skin care such as aging prevention (22) (Table I). Figure 1. General mechanism for miRNA and siRNA (from Ref. 17).The nascent pri-microRNA (pri-miRNA) transcripts are fi rst processed into ~70-nucleotide pre-miRNAs by Drosha inside the nucleus. Pre-miRNAs are transported to the cytoplasm by Exportin 5 and are processed into miRNA: miRNA* duplexes by Dicer. Dicer also processes long dsRNA molecules into small interfering RNA (siRNA) du- plexes. Only one strand of the miRNA:miRNA* duplex or the siRNA duplex is preferentially assembled into the RNA-induced silencing complex (RISC), which subsequently acts on its target by translational re- pression or mRNA cleavage, depending, at least in part, on the level of complementarity between the small RNA and its target. ORF, open reading frame.

JOURNAL OF COSMETIC SCIENCE 458 SMALL RNA IN SKIN WHITENING COSMECEUTICALS There are many pigmentary disorders that pose cosmetic problems in humans. Melasma, freckles, and aging spots are among the most common ones (34). The number and amount of melanocytes, as well as the type and distribution of melanin in skin, are main factors affecting the color of skin. Synthesis of melanin pigment is completed by a series of oxida- tive reactions, along which tyrosinase (TYR) and microphthalmia-associated tran- scription factors (MITF) are the key regulators (35–37). TYR converts tyrosine to dihydroxyphenylalanine and further to dopaquinone dopaquinone is subsequently auto- oxidized to dopachrome and, fi nally, to dihydroxyindole or dihydroxyindole-2-carboxylic acid to form eumelanin (38). Other major regulator in melanocyte development, func- tion, and survival is MITF. MITF is thought to mediate signifi cant effects of α-melanocyte stimulating hormone on differentiation by transcriptionally regulating enzymes that are essential to melanin production, i.e., TYR, tyrosinase-related protein 1 (TYRP1) and TYR-related protein-2 (TYRP2)/dopachrome-tautomerase (DCT) (39,40). Inhibition of TYR activity is still the most reported approach as a means of skin lightening. Other methods used include MITF inhibition, downregulation of melanocortin 1 receptor (MC1R) activity, interference with melanosomal transfer, and melanocyte loss (41). Using miRNAs to target and reduce the expression of TYR presents a novel and feasible approach for achieving skin whitening. Chen et al. developed an artifi cial miRNA expres- sion system which synthesizes miRNAs that downregulate TYR expression by binding and degrading TYR mRNAs. This anti-tyrosinase miRNA expression system success- fully demonstrated the feasibility of miRNA-mediated skin whitening in mice and hu- mans (23). Alternatively, novel siRNAs have been developed that specifi cally inhibited TYR expression, in which selected siRNAs exhibited such high activity that the siRNAs could be applied at a dose of less than 1 nM. Furthermore, the association of these TYR- specifi c siRNAs with cationic particles less than 1 μm in size makes it possible to sig- nifi cantly improve their penetration into target cells in a three-dimensional model such as skin (24). In one clinical study, 31 patients with pigmented facial lesions were treated using a MITF siRNA (MITF-siR) cream that contained a highly effi cient peptide-based transdermal vehicle (25,26). It demonstrated that MITF-siR silenced MITF gene expression effectively Table I Small RNA in Antiaging Cosmeceuticals Type of small RNA Gene target Application References miRNA Tyrosinase Whitening 23,24 siRNA MITF Whitening 25,26 siRNA P53 Whitening 7 miRNA inhibitor miR-29 Antiwrinkle 27,28 miRNA Hyaluronidase Moisturizing 19 siRNA Androgen receptors Hair care 29,30 siRNA Tbx21 Hair care 31 miRNA inhibitor miR-31 Hair care 32 miRNA LSD1/2, DNMT1, MECP1/2 Antiaging 33