New Human CYP17A1 Fluorescent Ligands

Using spectrophotometric titration approach several fluorescent and profluorescent compounds were tested against human CYP17A1. It was found that 20αand 20β-NBD (7-nitrobenzoxadiazole) derivatives of pregnenolone bind in the active site of human CYP17A1 in a substrate-like manner with dissociation constants K_d = 21.5 ± 3.1 mcМ (20a-NBD-Preg) and K_d = 19.8 ± 1.8 mcМ (20b-NBD-Preg), respectively. It was found that the compounds are not metabolized by the enzyme due to the non-optimal positioning of the ligands in the active site caused by the presence of the bulky NBD group. The formation of ligand complexes with CYP17A1 leads to fluorescence quenching, which is stronger for 20a isomer, possibly due to the formation of a hydrogen bond with Asn202, as it was found using molecular docking simulation approach. The identified ligands are promising lead compounds for the development of novel molecular tools that can be used to study the functioning mechanism of human CYP17A1 and to search for high-affinity modulators of enzyme activity — novel effective drugs.

1. P. F. Hall. J. Steroid Biochem. Mol. Biol., 40, N 4-6 (1991) 527—532

2. R. Yadav, E. M. Petrunak, D. F. Estrada, E. E. Scott. Mol. Cell Endocrinol., 441 (2017) 68—75

3. W. L. Miller. J. Steroid Biochem., 27, N 4-6 (1987) 759—66

4. D. Porubek. Current Top. Med. Chem., 13 (2013) 1364—1384

5. F. P. Guengerich. Trends Pharm. Sci., 6 (1992) 625—640

6. N. M. DeVore, E. E. Scott. Nature, 482, N 7383 (2012) 116—119

7. S. Dharia, A. Slane, M. Jian, M. Conner, A. J. Conley, C. R. Parker Jr. Biol. Reprod., 71, N 1 (2004) 83—88

8. A. Slominski, G. Ermak, M. Mihm. J. Clin. Endocrinol. Metab., 81, N 7 (1996) 2746—2749

9. V. Pezzi, J. M. Mathis, W.E. Rainey, B. R. Carr. J. Steroid Biochem. Mol. Biol., 87, N 2-3 (2003) 181—189

10. M. I. New. Mol. Cell Endocrinol., 211, N 1-2 (2003) 75—83

11. E. M. Petrunak, N. M. DeVore, P. R. Porubsky, E. E. Scott. J. Biol. Chem., 289 (2014) 32952—32964

12. G. Attard, A. H. Reid, R. J. Auchus, B. A Hughes, A. M. Cassidy, E. Thompson, N. B. Oommen, E. Folkerd, M. Dowsett, W. Arlt, J. S de Bono. J. Clin. Endocrinol. Metab., 97 (2012) 507—516

13. J. S de Bono, C. J. Logothetis, A. Molina, K. Fizazi, S. North, L. Chu, K. N. Chi, R. J. Jones, O. B. Goodman Jr, F. Saad, J. N. Staffurth, P. Mainwaring, S. Harland, T. W. Flaig, T. E. Hutson, T. Cheng, H. Patterson, J. D. Hainsworth, C. J. Ryan, C. N. Sternberg, S. L. Ellard, A. Fléchon, M. Saleh, M. Scholz, E. Efstathiou, A. Zivi, D. Bianchini, Y. Loriot, N. Chieffo, T. Kheoh, C. M. Haqq, H. I. Scher. N. Engl. J. Med., 134 (2011) 563—574

14. E. A. Mostaghel, B. T. Marck, S. Plymate, R. Vessella, S. Balk, A. M. Matsumoto, P. S. Nelson, B. Montgomery. Clin. Cancer Res., 17 (2011) 5913—5925

15. E. M. Petrunak, S. A. Rogers, J. Aube, E. E. Scott. Drug Metab. Dispos., 45, N 6 (2017) 635—645

16. E. Baston, F. R. Leroux. Recent Pat, Anticancer, Drug Discov., 2, N 1 (2007) 31—58

17. Q. Hu, M. Negri, S. Olgen, R. W. Hartmann. Chem. Med. Chem., 5 (2010) 899—910

18. S. S. Sandhu, H. Shukla, R. P. Aharwal, S. Kumar, S. Shukla. Natural Products J., 4, N 2 (2014) 140—152

19. N. Almassi, C. Reichard, J. Li, C. Russell, J. Perry, C. J Ryan, T. Friedlander, N. Sharifi. JAMA Oncol., 4 (2018) 554—557

20. F. K. Yoshimoto, R. J. Auch. J. Steroid Biochem. Mol. Biol., 151 (2015) 52—65

21. L. Gomez, J. R. Kovac, D. J. Lamb. Steroids, 95 (2015) 80—87

22. A. S. Latysheva, A. Yu. Misharin. Biomed. Chem. Res. Methods, 1, N 2 (2018) e00020

23. L. Yin, Q. Hu. Nat. Rev. Urol., 11 (2014) 32—42

24. A. S. Latysheva, V. A. Zolottsev, V. S. Pokrovsky, I. I. Khan, A. Yu. Misharin. Current Med. Chem., 28, N 40 (2021) 8416—8432

25. Я. В. Панада, Я. В. Фалетров, Н. С. Фролова, В. М. Шкуматов. Биомедицинская химия, 65, № 4 (2019) 324—330

26. A. Luthra, I. G. Denisov, S. G. Sligar. Arch. Biochem. Biophys., 507, N 1 (2011) 26—35

27. S. Leavitt, E. Freire. Current Opin. Struct. Biol., 11, N 5 (2001) 560—566

28. Folta-Stogniew. Methods Mol. Biol., 328 (2006) 97—112

29. E. Bozkurt, H. I. Gul. J. Photochem. Photobiol. A: Chemistry, 383, N 1 (2019) 111996

30. Дж. Лакович. Основы флуоресцентной спектроскопии, Москва, Мир (1986)

31. M. R. Eftink. Fluorescence Quenching: Theory and Applications, Topics in Fluorescence Spectroscopy, 2, Boston, Springer (2002) 53—126

32. M. Weerta, L. Stellab. J. Mol. Struct., 998, N 1-3 (2011) 144—150

33. K. V. Tugaeva, Y. V. Faletrov, E. S. Allakhverdiev, V. M. Shkumatov, E. G. Maksimov, N. N. Sluchanko. Biochem. Biophys. Res. Commun., 497 (2018) 58—64

34. D. S. Auld, H. Veith, J. J. Cali. In: Cytochrome P450 Protocols, Methods in Molecular Biology, 987, New York (2013) 1—9

35. D. M. Stresser, S. D. Turner, A. P. Blanchard, V. P. Miller, C. L. Crespi. Drug Metab. Dispos., 30 (2002) 845—852

36. I. N. White. Anal. Biochem., 172 (1988) 304—310

37. M. T. Donato, N. Jiménez, J. V. Castell, M. J. Gómez-Lechón. Drug Metab. Dispos., 32 (2004) 699—706

38. Компания ThermoFisher Scientific Inc. Инструкция к набору Vivid® CYP450 Screening Kits. [Электронный ресурс]. Режим доступа: http://tools.thermofisher.com/content/sfs/manuals/Vivid_CYP450_Screening_Kits _man.pdf.

39. Y. Dzichenka, M. Shapira, A. Yantsevich, T. Cherkesova, L. Grbovic, M. Savic, S. Usanov, Jovanovic-Santa. J. Steroid Biochem. Mol. Biology, 205 (2021) 105777

40. С. V. Locuson, J. M. Hutzler, T. S. Tracy. Drug Metab. Dispos., 35, N 4 (2007) 614—622

41. Г. Н. Некрасова, Я. В. Фалетров, Я. В. Диченко. Свиридовские чтения: сб. ст., 18 (2022) 109—116

42. B. N. Tyler, K. Chichak, R. B. Neil. Tetrahedron, 58, N 4 (2002) 639—651

43. J. Hidalgo, A. Sánchez-Coronilla, M. A. Muñoz, C. Carmona, M. Balón. J. Lumin., 127, N 2 (2007) 671—677

44. M. H. Mostafa, F. C. Khalil, D. Schryver, H. Keller, J.-M. Lehn. Supramolecular Sci., 2, N 3-4 (1995) 175—182