Biomol Ther  
Designing Tyrosinase siRNAs by Multiple Prediction Algorithms and Evaluation of Their Anti-Melanogenic Effects
Ok-Seon Kwon1,†, Soo-Jung Kwon1,†, Jin Sang Kim2, Gunbong Lee2, Han-Joo Maeng3, Jeongmi Lee4, Gwi Seo Hwang5, Hyuk-Jin Cha1,* and Kwang-Hoon Chun3,*
1Department of Life Sciences, Sogang University, Seoul 04107,
2Leaders Cosmetics Co., Ltd., Anseong 17599,
3Gachon Institute of Pharmaceutical Sciences, College of Pharmacy, Gachon University, Incheon 21936,
4School of Pharmacy, Sungkyunkwan University, Suwon 16419,
5Laboratory of Cell Differentiation Research, College of Korean Medicine, Gachon University, Seongnam 13120, Republic of Korea
E-mail: khchun@gachon.ac.kr (Chun KH), hjcha@sogang.ac.kr (Cha HJ)
Tel: +82-32-820-4951 (Chun KH), +82-2-705-4761 (Cha HJ)
Fax: +82-32-820-4823 (Chun KH), +82-2-704-3601 (Cha HJ)
The first two authors contributed equally to this work.
Received: May 30, 2017; Revised: August 3, 2017; Accepted: August 7, 2017; Published online: December 8, 2017.
© The Korean Society of Applied Pharmacology. All rights reserved.

Abstract
Melanin is a pigment produced from tyrosine in melanocytes. Although melanin has a protective role against UVB radiation-induced damage, it is also associated with the development of melanoma and darker skin tone. Tyrosinase is a key enzyme in melanin synthesis, which regulates the rate-limiting step during conversion of tyrosine into DOPA and dopaquinone. To develop effective RNA interference therapeutics, we designed a melanin siRNA pool by applying multiple prediction programs to reduce human tyrosinase levels. First, 272 siRNAs passed the target accessibility evaluation using the RNAxs program. Then we selected 34 siRNA sequences with ΔG ≥-34.6 kcal/mol, i-Score value ≥65, and siRNA scales score ≤30. siRNAs were designed as 19-bp RNA duplexes with an asymmetric 3’ overhang at the 3’ end of the antisense strand. We tested if these siRNAs effectively reduced tyrosinase gene expression using qRT-PCR and found that 17 siRNA sequences were more effective than commercially available siRNA. Three siRNAs further tested showed an effective visual color change in MNT-1 human cells without cytotoxic effects, indicating these sequences are anti-melanogenic. Our study revealed that human tyrosinase siRNAs could be efficiently designed using multiple prediction algorithms.
Keywords: Tyrosinase, Melanin, siRNA, Melanocytes, Whitening


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