Usnic Acid Triggers Apoptosis and Reduces Invasion by Activating Tumor Suppressor Genes in SH-SY5Y Neuroblastoma Cells.

Year 2024, , 497 - 502, 12.01.2025

Ceren Oy , Sema Serter Koçoğlu

https://doi.org/10.32708/uutfd.1587200

Abstract

Usnic acid is a dibenzofuran compound derived from Usnea diffracta Vain and has anti-inflammatory, antifungal and anticancer activities. However, the molecular mechanism of its antitumor effects has not been fully elucidated. This study aims to identify the apoptosis and autophagy-mediated mechanisms underlying the anticancer activity of usnic acid in neuroblastoma cells. The effects of usnic acid on SH-SY5Y neuroblastoma cell viability were evaluated by CCK-8 test, its effects on cell migration by wound healing assay, and its effects on invasion, apoptosis, autophagy and tumor suppressor genes by Western blot. Usnic acid was shown to reduce SH-SY5Y neuroblastoma cell viability in a dose and time dependent manner, and the IC50 dose was determined as 10 μM at 24 hours. Usnic acid has been shown to significantly reduce neuroblastoma cell migration and invasion. The control group wound area was determined to be 628.9 ± 35.59 cm2 on average at hour 0, while it was determined to be 234.8 ± 30.98 cm2 at 24 hours. Usnic acid wound area was 895.2 ± 30.07 cm2 at 0 hour and 801.9 ± 15.23 cm2 at 24 hours. It has been determined that usnic acid induces neuroblastoma cell apoptosis. Usnic acid caused a 4-fold increase in the expression of cleaved caspase-3, but did not cause any change in autophagy markers. Usnic acid application caused an increase in the expression of tumor suppressor genes in SH-SY5Y neuroblastoma cells. As a result, it has been shown that usnic acid can trigger apoptosis by activating tumor suppressor genes in neuroblastoma cells, thereby causing a decrease in invasion, migration and cell viability. The mechanisms underlying the anticancer activity of usnic acid in neuroblastoma cells need to be supported by further studies.

Keywords

usnic acid, neuroblastoma, apoptosis, autophagy, invasion

Usnik Asit SH-SY5Y Nöroblastom Hücrelerinde Tümör Baskılayıcı Genleri Aktive Ederek Apoptozu Tetikler ve İnvazyonu Azaltır

Abstract

Usnik asit, Usnea diffracta Vain'den kaynaklanan bir dibenzofuran bileşiğidir ve anti-inflamatuar, antifungal ve antikanser etkileri vardır. Ancak, antitümör etkilerinin moleküler mekanizması tam olarak açıklanmamıştır. Bu çalışma, usnik asidin nöroblastom hücrelerinde antikanser aktivitesinin altında yatan apoptoz ve otofaji aracılı mekanizmaları belirlemeyi amaçlamaktadır. Usnik asidin SH-SY5Y nöroblastom hücre canlılığı üzerine etkileri CCK-8 testi, hücre migrasyonu üzerine etkileri yara iyileşme deneyi, invazyon, apoptoz, otofaji ve tümör baskılayıcı genler üzerine etkileri ise Western blot ile değerlendirilmiştir. Usnik asidin SH-SY5Y nöroblastom hücre canlılığını doz ve zaman bağımlı olarak azalttığı gösterilmiş ve IC50 dozu 24 saatte 10 μM olarak belirlenmiştir. Usnik asidin nöroblastom hücre migrasyonu ve invazyonunu anlamlı olarak azalttığı gösterilmiştir. Kontrol grubu yara alanı 0. saatte ortalama 628,9 ± 35,59 cm2 iken 24 saatte 234,8 ± 30,98 cm2 olarak belirlenmiştir. Usnik asit yara alanı ise 0. saatte ortalama 895,2 ± 30,07 cm2 iken 24 saatte 801,9 ± 15,23 cm2’dir. Usnik asidin nöroblastom hücre apoptozunu indüklediği belirlenmiştir. Usnik asit kesilmiş kaspaz-3 ekspresyonunda 4 katlık bir artışa neden olmuş, otofaji markırlarında ise bir değişiklik oluşturmamıştır. Usnik asit uygulaması SH-SY5Y nöroblastom hücrelerinde tümör baskılayıcı genlerin ekspresyonunda artışa neden olmuştur. Sonuç olarak, usnik asidin nöroblastom hücrelerinde tümör baskılayıcı genleri aktive ederek apoptozu tetikleyebileceği, böylelikle invazyon, migrasyon ve hücre canlılığında azalışa neden olabileceği gösterilmiştir. Usnik asidin nöroblastom hücrelerinde antikanser aktivitesinin altında yatan mekanizmaların daha ileri çalışmalarla desteklenmesi gerekmektedir.

Keywords

Usnic asit, nöroblastom, apoptoz, otofaji, invazyon

References

• 1. Brodeur GM. Neuroblastoma: biological insights into a clinical enigma. Nat Rev Cancer. 2003;3(3):203-216. doi:10.1038/nrc1014
• 2. Maris JM. Recent advances in neuroblastoma. N Engl J Med. 2010;362(23):2202-2211. doi:10.1056/NEJMra0804577
• 3. Radic-Sarikas B, Halasz M, Huber KVM, vd. Lapatinib potentiates cytotoxicity of YM155 in neuroblastoma via inhibition of the ABCB1 efflux transporter. Sci Rep. 2017;7(1):1-8. doi:10.1038/s41598-017-03129-6
• 4. Vella S, Penna I, Longo L, vd. Perhexiline maleate enhances antitumor efficacy of cisplatin in neuroblastoma by inducing over-expression of NDM29 ncRNA. Sci Rep. 2015;5(November):1-13. doi:10.1038/srep18144
• 5. Alexander F. Neuroblastoma. Urol Clin North Am.2000;27(3):383-392, vii. doi:10.1016/s0094-0143(05)70087-2
• 6. DuBois SG, Kalika Y, Lukens JN, vd. Metastatic sites in stage IV and IVS neuroblastoma correlate with age, tumor biology,and survival. J Pediatr Hematol Oncol. 1999;21(3):181-189. doi:10.1097/00043426-199905000-00005
• 7. Ishola TA, Chung DH. Neuroblastoma. Surg Oncol. 2007;16(3):149-156. doi:10.1016/j.suronc.2007.09.005
• 8. Gimła M, Herman-Antosiewicz A. Multifaceted Properties ofUsnic Acid in Disrupting Cancer Hallmarks. Biomedicines. 2024;12(10). doi:10.3390/BIOMEDICINES12102199
• 9. Huneck S, Yoshimura I. Identification of Lichen Substances. Identification of Lichen Substances. Published online 1996:11-123. doi:10.1007/978-3-642-85243-5_2
• 10. Sun TX, Li MY, Zhang ZH, vd. Usnic acid suppresses cervical cancer cell proliferation by inhibiting PD-L1 expression and enhancing T-lymphocyte tumor-killing activity. Phytother Res.2021;35(7):3916-3935. doi:10.1002/PTR.7103
• 11. Serter Kocoglu S, Oy C, Secme M, Sunay FB. Investigation ofthe anticancer mechanism of monensin via apoptosis-related factors in SH-SY5Y neuroblastoma cells. Clin Transl Sci. 2023;16(9):1725-1735. doi:10.1111/CTS.13593
• 12. Kocoglu SS, Seçme M, Sunay FB. Erianin as a Promising Novel Agent in the Treatment of Neuroblastoma: The Anticancer Effects and Underlying Molecular Mechanisms. Anticancer Agents Med Chem. 2023;23(10):1204-1210. doi:10.2174/1871520623666230228095429
• 13. Serter Kocoglu S, Secme M, Oy C, Korkusuz G, Elmas L.Monensin, an Antibiotic Isolated from StreptomycesCinnamonensis, Regulates Human Neuroblastoma Cell Proliferation via the PI3K/AKT Signaling Pathway and ActsSynergistically with Rapamycin. Antibiotics (Basel).2023;12(3). doi:10.3390/ANTIBIOTICS12030546
• 14. Kumar K, Mishra JPN, Singh RP. Usnic acid induces apoptosis in human gastric cancer cells through ROS generation andDNA damage and causes up-regulation of DNA-PKcs and γ-H2A.X phosphorylation. Chem Biol Interact. 2020;315. doi:10.1016/J.CBI.2019.108898
• 15. Elmore S. Apoptosis: a review of programmed cell death. Toxicol Pathol. 2007;35(4):495-516. doi:10.1080/01926230701320337
• 16. Boice A, Bouchier-Hayes L. Targeting apoptotic caspases in cancer. Biochim Biophys Acta Mol Cell Res. 2020;1867(6). doi:10.1016/J.BBAMCR.2020.118688
• 17. Eskandari E, Eaves CJ. Paradoxical roles of caspase-3 in regulating cell survival, proliferation, and tumorigenesis. J Cell Biol. 2022;221(6):e202201159. doi:10.1083/JCB.202201159
• 18. Geng X, Zhang X, Zhou B, vd. Usnic Acid Induces CycleArrest, Apoptosis, and Autophagy in Gastric Cancer Cells InVitro and In Vivo. Med Sci Monit. 2018;24:556-566. doi:10.12659/MSM.908568
• 19. Özben R, Cansaran-Duman D. The expression profiles of apoptosis-related genes induced usnic acid in SK-BR-3 breast cancer cell. Hum Exp Toxicol. 2020;39(11):1497-1506. doi:10.1177/0960327120930257
• 20. Liu N, Yang C, Yang L, vd. Matrine induces autophagy in human neuroblastoma cells via blocking the AKT-mTOR pathway. Med Oncol. 2022;39(11). doi:10.1007/S12032-022-01762-4
• 21. Gimła M, Pyrczak-Felczykowska A, Malinowska M, vd. The pyrazole derivative of usnic acid inhibits the proliferation of pancreatic cancer cells in vitro and in vivo. Cancer Cell Int.2023;23(1):1-13. doi:10.1186/S12935-023-03054-X/FIGURES/6
• 22. Galanty A, Koczurkiewicz P, Wnuk D, vd. Usnic acid and atranorin exert selective cytostatic and anti-invasive effects on human prostate and melanoma cancer cells. Toxicol In Vitro.2017;40:161-169. doi:10.1016/J.TIV.2017.01.008
• 23. Justus CR, Leffler N, Ruiz-Echevarria M, Yang L V. In vitro cell migration and invasion assays. J Vis Exp. 2014;(88). doi:10.3791/51046
• 24. Lee JM, Bernstein A. Apoptosis, cancer and the p53 tumour suppressor gene. Cancer and Metastasis Reviews. 1995;14(2):149-161. doi:10.1007/BF00665797