Biomolecules & Therapeutics

The Korean Society of Applied Pharmacology (한국응용약물학회)
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Designing Tyrosinase siRNAs by Multiple Prediction Algorithms and Evaluation of Their Anti-Melanogenic Effects

  • Kwon, Ok-Seon (Department of Life Sciences, Sogang University) ;
  • Kwon, Soo-Jung (Department of Life Sciences, Sogang University) ;
  • Kim, Jin Sang (Leaders Cosmetics Co., Ltd.) ;
  • Lee, Gunbong (Leaders Cosmetics Co., Ltd.) ;
  • Maeng, Han-Joo (Gachon Institute of Pharmaceutical Sciences, College of Pharmacy, Gachon University) ;
  • Lee, Jeongmi (School of Pharmacy, Sungkyunkwan University) ;
  • Hwang, Gwi Seo (Laboratory of Cell Differentiation Research, College of Korean Medicine, Gachon University) ;
  • Cha, Hyuk-Jin (Department of Life Sciences, Sogang University) ;
  • Chun, Kwang-Hoon (Gachon Institute of Pharmaceutical Sciences, College of Pharmacy, Gachon University)
  • Received : 2017.05.30
  • Accepted : 2017.08.07
  • Published : 2018.05.01
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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 ${\Delta}G{\geq}-34.6kcal/mol$, i-Score value ${\geq}65$, and siRNA scales score ${\leq}30$. 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.

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References

  1. Amarzguioui, M. and Prydz, H. (2004) An algorithm for selection of functional siRNA sequences. Biochem. Biophys. Res. Commun. 316, 1050-1058. https://doi.org/10.1016/j.bbrc.2004.02.157
  2. Chang, C. I., Hong, S. W., Kim, S. and Lee, D. K. (2007) A structureactivity relationship study of siRNAs with structural variations. Biochem. Biophys. Res. Commun. 359, 997-1003. https://doi.org/10.1016/j.bbrc.2007.06.004
  3. Chu, C. Y. and Rana, T. M. (2008) Potent RNAi by short RNA triggers. RNA 14, 1714-1719. https://doi.org/10.1261/rna.1161908
  4. Czauderna, F., Fechtner, M., Dames, S., Aygun, H., Klippel, A., Pronk, G. J., Giese, K. and Kaufmann, J. (2003) Structural variations and stabilising modifications of synthetic siRNAs in mammalian cells. Nucleic Acids Res. 31, 2705-2716. https://doi.org/10.1093/nar/gkg393
  5. Elbashir, S. M., Harborth, J., Lendeckel, W., Yalcin, A., Weber, K. and Tuschl, T. (2001a) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411, 494-498. https://doi.org/10.1038/35078107
  6. Elbashir, S. M., Harborth, J., Weber, K. and Tuschl, T. (2002) Analysis of gene function in somatic mammalian cells using small interfering RNAs. Methods 26, 199-213. https://doi.org/10.1016/S1046-2023(02)00023-3
  7. Elbashir, S. M., Martinez, J., Patkaniowska, A., Lendeckel, W. and Tuschl, T. (2001b) Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate. EMBO J. 20, 6877-6888. https://doi.org/10.1093/emboj/20.23.6877
  8. Engels, J. W. (2013) Gene silencing by chemically modified siRNAs. N. Biotechnol. 30, 302-307. https://doi.org/10.1016/j.nbt.2012.07.002
  9. Fire, A., Xu, S., Montgomery, M. K., Kostas, S. A., Driver, S. E. and Mello, C. C. (1998) Potent and specific genetic interference by double- stranded RNA in Caenorhabditis elegans. Nature 391, 806-811. https://doi.org/10.1038/35888
  10. Gillbro, J. M. and Olsson, M. J. (2011) The melanogenesis and mechanisms of skin-lightening agents--existing and new approaches. Int. J. Cosmet. Sci. 33, 210-221. https://doi.org/10.1111/j.1468-2494.2010.00616.x
  11. Hearing, V. J. and Tsukamoto, K. (1991) Enzymatic control of pigmentation in mammals. FASEB J. 5, 2902-2909. https://doi.org/10.1096/fasebj.5.14.1752358
  12. Hosoi, J., Abe, E., Suda, T. and Kuroki, T. (1985) Regulation of melanin synthesis of B16 mouse melanoma cells by 1 alpha, 25-dihydroxyvitamin D3 and retinoic acid. Cancer Res. 45, 1474-1478.
  13. Huesken, D., Lange, J., Mickanin, C., Weiler, J., Asselbergs, F., Warner, J., Meloon, B., Engel, S., Rosenberg, A., Cohen, D., Labow, M., Reinhardt, M., Natt, F. and Hall, J. (2005) Design of a genomewide siRNA library using an artificial neural network. Nat. Biotechnol. 23, 995-1001. https://doi.org/10.1038/nbt1118
  14. Ichihara, M., Murakumo, Y., Masuda, A., Matsuura, T., Asai, N., Jijiwa, M., Ishida, M., Shinmi, J., Yatsuya, H., Qiao, S., Takahashi, M. and Ohno, K. (2007) Thermodynamic instability of siRNA duplex is a prerequisite for dependable prediction of siRNA activities. Nucleic Acids Res. 35, e123. https://doi.org/10.1093/nar/gkm699
  15. Kim, D. H., Behlke, M. A., Rose, S. D., Chang, M. S., Choi, S. and Rossi, J. J. (2005) Synthetic dsRNA Dicer substrates enhance RNAi potency and efficacy. Nat. Biotechnol. 23, 222-226. https://doi.org/10.1038/nbt1051
  16. Ladunga, I. (2007) More complete gene silencing by fewer siRNAs: transparent optimized design and biophysical signature. Nucleic Acids Res. 35, 433-440. https://doi.org/10.1093/nar/gkm352
  17. Lerner, A. B. and Fitzpatrick, T. B. (1950) Biochemistry of melanin formation. Physiol. Rev. 30, 91-126. https://doi.org/10.1152/physrev.1950.30.1.91
  18. Matveeva, O., Nechipurenko, Y., Rossi, L., Moore, B., Saetrom, P., Ogurtsov, A. Y., Atkins, J. F. and Shabalina, S. A. (2007) Comparison of approaches for rational siRNA design leading to a new efficient and transparent method. Nucleic Acids Res. 35, e63. https://doi.org/10.1093/nar/gkm088
  19. Muckstein, U., Tafer, H., Hackermuller, J., Bernhart, S. H., Stadler, P. F. and Hofacker, I. L. (2006) Thermodynamics of RNA-RNA binding. Bioinformatics 22, 1177-1182. https://doi.org/10.1093/bioinformatics/btl024
  20. Pancoska, P., Moravek, Z. and Moll, U. M. (2004) Efficient RNA interference depends on global context of the target sequence: quantitative analysis of silencing efficiency using Eulerian graph representation of siRNA. Nucleic Acids Res. 32, 1469-1479. https://doi.org/10.1093/nar/gkh314
  21. Reynolds, A., Leake, D., Boese, Q., Scaringe, S., Marshall, W. S. and Khvorova, A. (2004) Rational siRNA design for RNA interference. Nat. Biotechnol. 22, 326-330. https://doi.org/10.1038/nbt936
  22. Rose, S. D., Kim, D. H., Amarzguioui, M., Heidel, J. D., Collingwood, M. A., Davis, M. E., Rossi, J. J. and Behlke, M. A. (2005) Functional polarity is introduced by Dicer processing of short substrate RNAs. Nucleic Acids Res. 33, 4140-4156. https://doi.org/10.1093/nar/gki732
  23. Sano, M., Sierant, M., Miyagishi, M., Nakanishi, M., Takagi, Y. and Sutou, S. (2008) Effect of asymmetric terminal structures of short RNA duplexes on the RNA interference activity and strand selection. Nucleic Acids Res. 36, 5812-5821. https://doi.org/10.1093/nar/gkn584
  24. Schwarz, D. S., Hutvagner, G., Du, T., Xu, Z., Aronin, N. and Zamore, P. D. (2003) Asymmetry in the assembly of the RNAi enzyme complex. Cell 115, 199-208. https://doi.org/10.1016/S0092-8674(03)00759-1
  25. Shabalina, S. A., Spiridonov, A. N. and Ogurtsov, A. Y. (2006) Computational models with thermodynamic and composition features improve siRNA design. BMC Bioinformatics 7, 65. https://doi.org/10.1186/1471-2105-7-65
  26. Sheth, V. M. and Pandya, A. G. (2011) Melasma: a comprehensive update: part II. J. Am. Acad. Dermatol. 65, 699-714. https://doi.org/10.1016/j.jaad.2011.06.001
  27. Slominski, A., Tobin, D. J., Shibahara, S. and Wortsman, J. (2004) Melanin pigmentation in mammalian skin and its hormonal regulation. Physiol. Rev. 84, 1155-1228. https://doi.org/10.1152/physrev.00044.2003
  28. Tafer, H., Ameres, S. L., Obernosterer, G., Gebeshuber, C. A., Schroeder, R., Martinez, J. and Hofacker, I. L. (2008) The impact of target site accessibility on the design of effective siRNAs. Nat. Biotechnol. 26, 578-583. https://doi.org/10.1038/nbt1404
  29. Ui-Tei, K., Naito, Y., Takahashi, F., Haraguchi, T., Ohki-Hamazaki, H., Juni, A., Ueda, R. and Saigo, K. (2004) Guidelines for the selection of highly effective siRNA sequences for mammalian and chick RNA interference. Nucleic Acids Res. 32, 936-948. https://doi.org/10.1093/nar/gkh247
  30. Vert, J. P., Foveau, N., Lajaunie, C. and Vandenbrouck, Y. (2006) An accurate and interpretable model for siRNA efficacy prediction. BMC Bioinformatics 7, 520. https://doi.org/10.1186/1471-2105-7-520

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