I. Purpose Dyschromia in skin is considered aesthetically undesirable. A variety of blemishes exist in skin, some of which are due to an overexpression of melanin, most commonly seen in freckles. By inhibiting the genes that express these traits with a nontoxic substance, skin could be whitened to achieve a more uniform skin color. II. Definitions Tyrosinase: An enzyme that catalyzes the production of melanin through oxidation (i.e. exposure to air) DOPA oxidase: Transforms L-DOPA to DOPAchrome in the pathway to melanin synthesis. hydroxylation of tyrosine to L-DOPA (L-dihydroxyphenylalanine) and dehydroxylation of L-DOPA to DOPAquinone and then to DOPAchrome, then melanin synthesis TRP-1: Tyrosinase-related Protein 1, forms a complex with tyrosinase TRP-2: Tryosinase-related Protein 2, protects the carboxylic acids of melanin PKC-ßI: protein kinase C-ßI Antisense Oligonucleotide: A short sequence of nucleotides that are synthesized to modify gene expression Melanogenesis: Production of melanin; occurs in melanocytes. Cytotoxic: The quality of being toxic to cells Chromameter: A device that calculates an Individual Typological Angle (ITA) from a light variable and the chromaticity of skin color based on a blue-yellow axis Mexameter: A device that obtains a melanin index from skin Formazan: Artificial substance that absorbs at 450 nm III. Background information Current depigmenting agents are unstable/cytotoxic/skin irritants/possible carcinogens/otherwise ineffective Well-known example is hydroquinone, a tyrosinase-inhibiting compound that has been banned in some countries for being a possible carcinogen, despite its effectiveness in skin bleaching. IV. Materials and Methods Cultured human melanocytes were used to monitor the activity of tyrosinase, PKC-ßI and the antisense oligonucleotides that would modulate them. Cell viability was monitored to ensure that the treatment conditions were not cytotoxic. XTT, a tetrazolium salt was added. The XTT salt is reduced to a formazan; measuring this colored product's intensity with a spectrophotometer, we can determine the number of living cells 30 Asian women with pigmented spots on both hands volunteered to evaluate the product containing antisense nucleotides. The test product was applied twice daily to specific spots, at least 5mm in diameter, on each hand. Melanin index on the pigmented spots were measured before treatment, 4 weeks into treament, and 8 weeks into treatment using the chromameter and mexamater. Additionally, the same method was used on non-pigmented spots for comparison (also done before treatment, 4 weeks into, and 8 weeks into.) This data was analyzed by numerous tests, including the t-test, with significance at 5%. V. Results The cultured melanocytes were treated daily over 5 days with 250 and 500 nM concentrations of the oligonucleotide in order to determine the concentration of PKC-ßI targeting nucleotide necessary to obtain maximum inhibition of DOPA oxidase activity. Fig2 A similar test was performed using an oligonucleotide to target the TRP-1 sequence; additionally, human melanocytes were treated with kojic acid (a chemical used to lighten skin) at the maximum non-cytotoxic concentration. Fig3 A third test using both oligonucleotides to target the TRP-1 and PKC-ßI in sequence was found to be the most effective, with a 36% inhibition rate of DOPA-oxidase activity. VI. Conclusions This was probably the first known test of using antisense oligonucleotides The in vitro and in vivo tests showed that the antisense oligonucleotide method of lightening skin is effective and safe VII. Personal analysis This article was complex and I had difficulty even finding definitions for some of the terms used in it, but overall it was worth understanding the research that took place. The idea of controlling skin pigmentation has fascinated me for years, and this looks to be the first step of many in battling dyschromia and allowing you to get rid of a sunspot as easily as a pimple or wart. VIII. Applications/Directions for Future Research Using oligonucleotides to inhibit melanin expression appears to be effective, and hopefully more efficient methods of genetic modulation will be discovered in the future.