Evaluation Of the Potential Cytotoxic, Antimetastatic, and Antioxidant Abilities Of Chrysin and Astaxanthin İn Triple-Negative Breast Cancer Cells

Öz

Aim: Triple-negative breast cancer (TNBC) has worst overall survival of all breast cancers. The aim of this study was to investigate the effects of chrysin and astaxanthin on cell viability/cytotoxicity, metastasis, and oxidative stress in MDA-MB-231 cells. Material and Methods: The effects of chrysin (5, 10, 15, 20, 25, 40, 50, 75, 90, 100 µg/ml) and astaxanthin (5, 10, 15, 20, 40, 50, 75, 90, 100 µg/ml) on cell viability/cytotoxicity in TNBC (MDA-MB-231) cells were determined by WST-1. The efficacy of chrysin and astaxanthin on cell migration and metastasis was determined by scratch assay. In addition, the effect of chrysin and astaxanthin on the level of reactive oxygen species (ROS) in MDA-MB-231 cells was determined by DCF-DA analysis. Results: Astaxanthin did not suppress cell proliferation in MDA-MB-231 cells according to our WST-1 data. However, cell viability of the MDA-MB-231 cell line at higher chrysin doses decreased to %70 at all-time intervals. After 48 hours of exposure to chrysin (40 µg/ml) and astaxanthin (25 µg/ml), the scratch in the MDA-MB-231 cells was closed. Astaxanthin at a dose of 25 µg/ml was found not to cause oxidative stress at 24 hours after exposure, but a high fluorescence intensity was detected at 48 hours. On the other hand, after the administration of 40 µg/ml chrysin, more fluorescence intensity was detected at both 24 and 48 hours. Conclusion: Chrysin and astaxanthin may have effects on cell migration and intracellular ROS accumulation, however, they did not inhibit cell proliferation in MDA-MB-231 cells.

Anahtar Kelimeler

• Triple Negative Breast Cancer (TNBC)
• MDA-MB-231
• cell culture
• chrysin
• astaxanthin

Üçlü negatif meme kanseri hücrelerinde chrysin ve astaxanthin'in potansiyel sitotoksik, antimetastatik ve antioksidan etkilerinin değerlendirilmesi

Öz

Giriş ve Amaç: Üçlü negatif meme kanseri (TNBC), tüm meme kanserleri arasında en kötü genel sağkalım oranına sahiptir. Bu çalışmanın amacı, MDA-MB-231 hücrelerinde chrysin ve astaxanthin 'in hücre canlılığı/sitotoksisitesi, metastaz ve oksidatif stres üzerindeki etkilerini araştırmaktır. Gereç ve Yöntem: Chrysin (5, 10, 15, 20, 25, 40, 50, 75, 90, 100 µg/ml) ve astaxanthin’in (5, 10, 15, 20, 40, 50, 75, 90, 100 µg/ml) TNBC (MDA-MB-231) hücrelerinde hücre canlılığı/sitotoksisitesi üzerindeki etkisi WST-1 ile belirlendi. Chrysin ve astaxanthin’in hücre göçü ve metastaz üzerindeki etkinliği çizik analizi ile belirlendi. Ayrıca chrysin ve astaxanthin’in MDA-MB-231 hücrelerindeki reaktif oksijen türleri (ROS) seviyesi üzerindeki etkisi DCF-DA analizi ile belirlendi. Bulgular: Astaxanthin, WST-1 verilerimize göre MDA-MB-231 hücrelerinde hücre proliferasyonu üzerine etki göstermedi. Ancak yüksek chrysin dozları MDA-MB-231 hücre hattında hücre canlılığını tüm zaman aralıklarında %70'e kadar düşürdü. Ayrıca chrysin (40 µg/ml) ve astaksantin’e (25 µg/ml) 48 saat maruz kaldıktan sonra MDA-MB-231 hücrelerindeki çizik kapandı. 25 µg/ml dozunda astaxanthin maruziyetinden 24 saat sonra oksidatif strese neden olmadığı ancak 48. saatte yüksek floresans şiddeti tespit edildi. Öte yandan 40 µg/ml krisin uygulamasından sonra hem 24. hem de 48. saatte daha fazla floresans yoğunluğu tespit edildi. Sonuç: Chrysin ve astaksantin’in MDA-MB-231 hücrelerinde hücre göçü ve hücre içi ROS birikimi üzerine etkileri olabilir ancak hücre çoğalması üzerine etkisi saptanmadı.

Anahtar Kelimeler

• Üçlü negatif meme kanseri (TNBC)
• MDA-MB-231
• hücre kültürü
• chrysin
• astaxanthin

Kaynakça

1. 1. Bertucci F, Finetti P, Simeone I, Hendrickx W, Wang E, Marincola FM, et al. The immunologic constant of rejection classification refines the prognostic value of conventional prognostic signatures in breast cancer. Br J Cancer. 2018;119(11):1383-91.
2. 2. Garrido-Castro AC, Lin NU, Polyak K. Insights into Molecular Classifications of Triple-Negative Breast Cancer: Improving Patient Selection for Treatment. Cancer Discov. 2019;9(2):176-98.
3. 3. Foukakis T, Fornander T, Lekberg T, Hellborg H, Adolfsson J, Bergh J. Age-specific trends of survival in metastatic breast cancer: 26 years longitudinal data from a population-based cancer registry in Stockholm, Sweden. Breast Cancer Res Treat. 2011;130(2):553-60.
4. 4. Shahbaz M, Naeem H, Imran M, Ul Hassan H, Alsagaby SA, Al Abdulmonem W, et al. Chrysin a promising anticancer agent: recent perspectives. Int J Food Prop. 2023;26(1):2294-337.
5. 5. Zhu ZY, Wang WX, Wang ZQ, Chen LJ, Zhang JY, Liu XC, et al. Synthesis and antitumor activity evaluation of chrysin derivatives. Eur J Med Chem. 2014;75:297-300.
6. 6. Khoo BY, Chua SL, Balaram P. Apoptotic effects of chrysin in human cancer cell lines. Int J Mol Sci. 2010;11(5):2188-99.
7. 7. Salari N, Faraji F, Jafarpour S, Faraji F, Rasoulpoor S, Dokaneheifard S, et al. Anti-cancer Activity of Chrysin in Cancer Therapy: a Systematic Review. Indian J Surg Oncol. 2022;13(4):681-90.
8. 8. Gao S, Siddiqui N, Etim I, Du T, Zhang Y, Liang D. Developing nutritional component chrysin as a therapeutic agent: Bioavailability and pharmacokinetics consideration, and ADME mechanisms. Biomed Pharmacother. 2021;142:112080.

9. 9. Simat V, Rathod NB, Cagalj M, Hamed I, Generalic Mekinic I. Astaxanthin from Crustaceans and Their Byproducts: A Bioactive Metabolite Candidate for Therapeutic Application. Mar Drugs. 2022;20(3).
10. 10. Nishida Y, Nawaz A, Hecht K, Tobe K. Astaxanthin as a Novel Mitochondrial Regulator: A New Aspect of Carotenoids, beyond Antioxidants. Nutrients. 2021;14(1).
11. 11. Chung BY, Park SH, Yun SY, Yu DS, Lee YB. Astaxanthin Protects Ultraviolet B-Induced Oxidative Stress and Apoptosis in Human Keratinocytes via Intrinsic Apoptotic Pathway. Ann Dermatol. 2022;34(2):125-31.
12. 12. Faraone I, Sinisgalli C, Ostuni A, Armentano MF, Carmosino M, Milella L, et al. Astaxanthin anticancer effects are mediated through multiple molecular mechanisms: A systematic review. Pharmacol Res. 2020;155:104689.
13. 13. Kim MS, Ahn YT, Lee CW, Kim H, An WG. Astaxanthin Modulates Apoptotic Molecules to Induce Death of SKBR3 Breast Cancer Cells. Mar Drugs. 2020;18(5).
14. 14. Karimian A, Mir Mohammadrezaei F, Hajizadeh Moghadam A, Bahadori MH, Ghorbani-Anarkooli M, Asadi A, et al. Effect of astaxanthin and melatonin on cell viability and DNA damage in human breast cancer cell lines. Acta Histochem. 2022;124(1):151832.
15. 15. Seyhan MF, Yilmaz E, Timirci-Kahraman O, Saygili N, Kisakesen HI, Gazioglu S, et al. Different propolis samples, phenolic content, and breast cancer cell lines: Variable cytotoxicity ranging from ineffective to potent. IUBMB Life. 2019;71(5):619-31.
16. 16. Esen M, Guven Y, Seyhan MF, Ersev H, Tuna-Ince EB. Evaluation of the genotoxicity, cytotoxicity, and bioactivity of calcium silicate-based cements. BMC Oral Health. 2024;24(1):119.
17. 17. Ryu S, Lim W, Bazer FW, Song G. Chrysin induces death of prostate cancer cells by inducing ROS and ER stress. J Cell Physiol. 2017;232(12):3786-97.
18. 18. Lee JH, Yoo ES, Han SH, Jung GH, Han EJ, Choi EY, et al. Chrysin Induces Apoptosis and Autophagy in Human Melanoma Cells via the mTOR/S6K Pathway. Biomedicines. 2022;10(7).
19. 19. Zhang J, Peng CA. Enhanced proliferation and differentiation of mesenchymal stem cells by astaxanthin-encapsulated polymeric micelles. PLoS One. 2019;14(5):e0216755.
20. 20. Kim JH, Park JJ, Lee BJ, Joo MK, Chun HJ, Lee SW, et al. Astaxanthin Inhibits Proliferation of Human Gastric Cancer Cell Lines by Interrupting Cell Cycle Progression. Gut Liver. 2016;10(3):369-74.
21. 21. Siangcham T, Vivithanaporn P, Sangpairoj K. Anti-Migration and Invasion Effects of Astaxanthin against A172 Human Glioblastoma Cell Line. Asian Pac J Cancer Prev. 2020;21(7):2029-33.
22. 22. Jiang L, Wang Y, Liu G, Liu H, Zhu F, Ji H, et al. C-Phycocyanin exerts anti-cancer effects via the MAPK signaling pathway in MDA-MB-231 cells. Cancer Cell Int. 2018;18:12.
23. 23. Xia M, Yu H, Gu S, Xu Y, Su J, Li H, et al. p62/SQSTM1 is involved in cisplatin resistance in human ovarian cancer cells via the Keap1-Nrf2-ARE system. Int J Oncol. 2014;45(6):2341-8.
24. 24. Agus HH, Sengoz CO, Yilmaz S. Oxidative stress-mediated apoptotic cell death induced by camphor in sod1-deficient Schizosaccharomyces pombe. Toxicol Res (Camb). 2019;8(2):216-26.
25. 25. Orrantia-Borunda E, Anchondo-Nunez P, Acuna-Aguilar LE, Gomez-Valles FO, Ramirez-Valdespino CA. Subtypes of Breast Cancer. In: Mayrovitz HN, editor. Breast Cancer. Brisbane (AU)2022.
26. 26. Zhu S, Wu Y, Song B, Yi M, Yan Y, Mei Q, et al. Recent advances in targeted strategies for triple-negative breast cancer. J Hematol Oncol. 2023;16(1):100.
27. 27. Deshmukh PK, Mutha RE, Surana SJ. Electrostatic deposition assisted preparation, characterization and evaluation of chrysin liposomes for breast cancer treatment. Drug Dev Ind Pharm. 2021;47(5):809-19.
28. 28. Mohammadinejad S, Jafari-Gharabaghlou D, Zarghami N. Development of PEGylated PLGA Nanoparticles Co-Loaded with Bioactive Compounds: Potential Anticancer Effect on Breast Cancer Cell Lines. Asian Pac J Cancer Prev. 2022;23(12):4063-72.
29. 29. Liu X, Zhang X, Shao Z, Zhong X, Ding X, Wu L, et al. Pyrotinib and chrysin synergistically potentiate autophagy in HER2-positive breast cancer. Signal Transduct Target Ther. 2023;8(1):463.
30. 30. Javan Maasomi Z, Pilehvar Soltanahmadi Y, Dadashpour M, Alipour S, Abolhasani S, Zarghami N. Synergistic Anticancer Effects of Silibinin and Chrysin in T47D Breast Cancer Cells. Asian Pac J Cancer Prev. 2017;18(5):1283-7.
31. 31. Eatemadi A, Daraee H, Aiyelabegan HT, Negahdari B, Rajeian B, Zarghami N. Synthesis and Characterization of Chrysin-loaded PCL-PEG-PCL nanoparticle and its effect on breast cancer cell line. Biomed Pharmacother. 2016;84:1915-22.
32. 32. Mohammadinejad S, Akbarzadeh A, Rahmati-Yamchi M, Hatam S, Kachalaki S, Zohreh S, et al. Preparation and Evaluation of Chrysin Encapsulated in PLGA- PEG Nanoparticles in the T47-D Breast Cancer Cell Line. Asian Pac J Cancer Prev. 2015;16(9):3753-8.
33. 33. Samarghandian S, Azimi-Nezhad M, Borji A, Hasanzadeh M, Jabbari F, Farkhondeh T, et al. Inhibitory and Cytotoxic Activities of Chrysin on Human Breast Adenocarcinoma Cells by Induction of Apoptosis. Pharmacogn Mag. 2016;12(Suppl 4):S436-S40.
34. 34. Geng A, Xu S, Yao Y, Qian Z, Wang X, Sun J, et al. Chrysin impairs genomic stability by suppressing DNA double-strand break repair in breast cancer cells. Cell Cycle. 2022;21(4):379-91.
35. 35. Javan N, Khadem Ansari MH, Dadashpour M, Khojastehfard M, Bastami M, Rahmati-Yamchi M, et al. Synergistic Antiproliferative Effects of Co-nanoencapsulated Curcumin and Chrysin on MDA-MB-231 Breast Cancer Cells Through Upregulating miR-132 and miR-502c. Nutr Cancer. 2019;71(7):1201-13.
36. 36. Li Y, Zhang Q, He J, Yu W, Xiao J, Guo Y, et al. Synthesis and biological evaluation of amino acid derivatives containing chrysin that induce apoptosis. Nat Prod Res. 2021;35(4):529-38.
37. 37. Kim KM, Jung J. Upregulation of G Protein-Coupled Estrogen Receptor by Chrysin-Nanoparticles Inhibits Tumor Proliferation and Metastasis in Triple Negative Breast Cancer Xenograft Model. Front Endocrinol (Lausanne). 2020;11:560605.
38. 38. Hong TB, Rahumatullah A, Yogarajah T, Ahmad M, Yin KB. Potential effects of chrysin on MDA-MB-231 cells. Int J Mol Sci. 2010;11(3):1057-69.
39. 39. Yang B, Huang J, Xiang T, Yin X, Luo X, Huang J, et al. Chrysin inhibits metastatic potential of human triple-negative breast cancer cells by modulating matrix metalloproteinase-10, epithelial to mesenchymal transition, and PI3K/Akt signaling pathway. J Appl Toxicol. 2014;34(1):105-12.
40. 40. Huang C, Chen YJ, Chen WJ, Lin CL, Wei YX, Huang HC. Combined treatment with chrysin and 1,2,3,4,6-penta-O-galloyl-beta-D-glucose synergistically inhibits LRP6 and Skp2 activation in triple-negative breast cancer and xenografts. Mol Carcinog. 2015;54(12):1613-25.
41. 41. Jafari S, Dabiri S, Mehdizadeh Aghdam E, Fathi E, Saeedi N, Montazersaheb S, et al. Synergistic effect of chrysin and radiotherapy against triple-negative breast cancer (TNBC) cell lines. Clin Transl Oncol. 2023;25(8):2559-68.
42. 42. Kim K, Cho HR, Son Y. Astaxanthin Induces Apoptosis in MCF-7 Cells through a p53-Dependent Pathway. Int J Mol Sci. 2024;25(13).
43. 43. McCall B, McPartland CK, Moore R, Frank-Kamenetskii A, Booth BW. Effects of Astaxanthin on the Proliferation and Migration of Breast Cancer Cells In Vitro. Antioxidants (Basel). 2018;7(10).
44. 44. Atalay PB, Kuku G, Tuna BG. Effects of carbendazim and astaxanthin co-treatment on the proliferation of MCF-7 breast cancer cells. In Vitro Cell Dev Biol Anim. 2019;55(2):113-9.
45. 45. Ahn YT, Kim MS, Kim YS, An WG. Astaxanthin Reduces Stemness Markers in BT20 and T47D Breast Cancer Stem Cells by Inhibiting Expression of Pontin and Mutant p53. Mar Drugs. 2020;18(11).
46. 46. Malhao F, Ramos AA, Macedo AC, Rocha E. Cytotoxicity of Seaweed Compounds, Alone or Combined to Reference Drugs, against Breast Cell Lines Cultured in 2D and 3D. Toxics. 2021;9(2).
47. 47. Bharathiraja S, Manivasagan P, Quang Bui N, Oh YO, Lim IG, Park S, et al. Cytotoxic Induction and Photoacoustic Imaging of Breast Cancer Cells Using Astaxanthin-Reduced Gold Nanoparticles. Nanomaterials (Basel). 2016;6(4).