Sitokalasin B Tedavisi, Nrf2 Sinyal Yoluyla U87 Glioblastoma Hücrelerinde Anti-Proliferatif Etkiler Gösterdi

Öz

Giriş ve Amaç: Glioblastoma kötü prognozlu agrasif beyin tümörlerinden biridir ve glioblastoma için mevcut tedavi yöntemleri oldukça sınırlıdır. Sitokalasin B'nin (CB) kanser hücre hatları üzerinde inhibitör etki göstermektedir, ancak antikanser etkileri henüz tam olarak anlaşılamamıştır. Bu çalışmada, CB'nin U87 insan glioblastoma hücrelerinde nükleer faktör eritroid 2 ile ilişkili faktör (Nrf2) sinyal yolağı üzerinden oksidatif, antioksidan ve DNA hasar mekanizmaları üzerindeki etkisini araştırmayı hedefledik. Gereç ve Yöntemler: İlk olarak, CB'nin U87 hücrelerindeki sitotoksik konsantrasyonlarını MTT analizi ile belirledik. Ardından, CB'nin Nrf2 seviyeleri üzerindeki etkisini ve bununla bağlantılı olarak total oksidan kapasite (TOS), malondialdehit (MDA), süperoksit dismutaz (SOD) ve glutatyon peroksidaz (GPx) seviyeleri ölçüldü. Son olarak, CB ile tedavi edilen U87 hücrelerindeki DNA hasarını tespit edebilmek için 8-hidroksi-2'-deoksiguanozin (8-OHdG) seviyeleri ölçüldü. Bulgular: MTT analizine göre, CB'nin U87 hücrelerinde canlılığı konsantrasyona bağımlı bir şekilde azalttığını belirledik ve IC50 konsantrasyonunu 62,8 μM olarak tespit ettik. Ardından, 5,1, 33,6 ve 62,8 μM CB ile tedavi edilen U87 hücrelerinde TOS, MDA ve 8-OHdG seviyeleri konsantrasyon bağımlı bir şekilde artmıştır (p<0,05). Aksine, CB tedavisi U87 hücrelerindeki Nrf2, SOD ve GPx seviyeleri azalmıştır (p<0,05). CB, U87 glioblastoma hücre proliferasyonunu Nfr2 sinyal yolağı üzerinden inhibe etmiştir. Sonuç: CB tedavisi, U87 hücrelerinde oksidatif stresi ve DNA hasarını indüklemesinin yanı sıra antioksidan enzimlerin seviyelerini azaltmıştır, bu da CB'nin glioblastoma için potansiyel terapötik ajan olabileceğini düşündürdü. Ancak, CB'nin diğer kanser hücre hatlarında ve in vivo modellerdeki etkilerini araştıran daha ileri çalışmalara ihtiyaç vardır.

Anahtar Kelimeler

• Sitokalasin B
• U87 Glioblastoma Hücreler
• Oksidatif Stres
• Antioksidan
• DNA hasarı

Cytochalasin B Treatment Showed Anti-Proliferative Effects in U87 Glioblastoma Cells via Nrf2 Signaling Pathway

Öz

Objective: Glioblastoma is one of the aggressive brain tumors with a poor prognosis, and the available treatments for glioblastoma are quite limited. Cytochalasin B (CB) has an inhibitory effect on cancer cell lines, but its anticancer effects are not yet fully understood. In this study, we aimed to investigate the effect of CB on oxidative, antioxidant and DNA damage mechanisms via nuclear factor erythroid 2 associated factor (Nrf2) signaling pathway in U87 human glioblastoma cells. Materials and Methods: Firstly, we determined the cytotoxic concentrations of CB in U87 cells by MTT analysis. Then, the effect of CB on Nrf2 levels and related total oxidant capacity(TOS), malondialdehyde (MDA), superoxide dismutase(SOD) and glutathione peroxidase(GPx) levels were measured. Finally, 8-hydroxy-2'-deoxyguanosine(8-OHdG) levels were measured to detect DNA damage in U87 cells treated with CB. Results: According to MTT analysis, we determined that CB decreased viability in U87 cells in a concentration-dependent manner and determined the IC50 concentration to be 62.8μM. Subsequently, TOS, MDA, and 8-OHdG levels increased in a concentration-dependent manner in U87 cells treated with 5.1, 33.6, and 62.8μM CB (p<0.05). In contrast, Nrf2, SOD and GPx levels were decreased in CB-treated U87 cells (p<0.05). CB inhibited U87 glioblastoma cell proliferation via the Nfr2 signaling pathway. Conclusion: CB induced oxidative stress and DNA damage in U87 cells, as well as reduced antioxidant enzymes levels, suggesting that CB may be a potential therapeutic agent for glioblastoma. However, further studies investigating the effects of CB in other cancer cell lines and in vivo models are needed.

Anahtar Kelimeler

• Cytochalasin B
• Cytochalasin B
• Glioblastoma Cells
• Oxidative Stress
• Antioxidant
• DNA damage

Kaynakça

1. Ohgaki, H, Kleihues, P, Population-based studies on incidence, survival rates, and genetic alterations in astrocytic and oligodendroglial gliomas, Journal of neuropathology and experimental neurology, 2005, 64(6), 479–489.
2. Goodenberger, M.L, Jenkins, R.B, Genetics of adult glioma, Cancer genetics, 2012, 205(12), 613–621.
3. Molinaro, A.M, Taylor, J.W, Wiencke, J.K, Wrensch, M.R, Genetic and molecular epidemiology of adult diffuse glioma, Nature reviews, Neurology, 2019, 15(7), 405–417.
4. Jovcevska, I, Kočevar, N, Komel, R, Glioma and glioblastoma- how much do we (not) know? Molecular and clinical oncology, 2013, 1(6), 935–941.
5. Bahadur, S, Sahu, A.K, Baghel, P, Saha, S, Current promising treatment strategy for glioblastoma multiform: A review, Oncology reviews, 2019, 13(2), 417.
6. Haidle, A.M, Myers, A.G, An enantioselective, modular, and general route to the cytochalasins: synthesis of L-696,474 and cytochalasin B, Proceedings of the National Academy of Sciences of the United States of America, 2004, 101(33), 12048–12053.
7. Scherlach, K, Boettger, D, Remme, N, Hertweck, C, The chemistry and biology of cytochalasans, Natural product reports, 2010, 27(6), 869–886.
8. Hayot, C, Debeir, O, Van Ham, P, Van Damme, M, Kiss, R, Decaestecker, C, Characterization of the activities of actin-affecting drugs on tumor cell migration, Toxicology and applied pharmacology, 2006, 211(1), 30–40.

9. Van Goietsenoven, G, Mathieu, V, Andolfi, A, Cimmino, A, Lefranc, F, Kiss, R, Evidente, A, In vitro growth inhibitory effects of cytochalasins and derivatives in cancer cells, Planta medica, 2011, 77(7), 711–717.
10. Hwang, J, Yi, M, Zhang, X, Xu, Y, Jung, J.H, Kim, D.K, Cytochalasin B induces apoptosis through the mitochondrial apoptotic pathway in HeLa human cervical carcinoma cells, Oncology reports, 2013, 30(4), 1929–1935.
11. Ma, Q, Role of nrf2 in oxidative stress and toxicity, Annual review of pharmacology and toxicology, 2013, 53, 401–426.
12. Bellezza, I, Giambanco, I, Minelli, A, Donato, R, Nrf2-Keap1 signaling in oxidative and reductive stress, Biochimica et biophysica acta Molecular cell research, 2018, 1865(5), 721–733.
13. Hacioglu, C, Kar, F, Kacar, S, Sahinturk, V, Kanbak, G, Bexarotene inhibits cell proliferation by inducing oxidative stress, DNA damage and apoptosis via PPARγ/ NF-κB signaling pathway in C6 glioma cells, Medical oncology (Northwood, London, England), 2021, 38(3), 31.
14. Lowry, O.H, Rosebrough, N.J, Farr, A.L, Randall, R.J, Protein measurement with the Folin phenol reagent, The Journal of biological chemistry, 1951, 193(1), 265–275.
15. Tong, Z.G, Liu, N, Song, H.S, Li, J.Q, Jiang, J, Zhu, J.Y, Qi, J.P, Cytochalasin B inhibits the proliferation of human glioma U251 cells through cell cycle arrest and apoptosis, Genetics and molecular research: GMR, 2014, 13(4), 10811–10822.
16. Heidarzadeh, S, Motalleb, G.H, Zorriehzahra, M.J, Evaluation of Tumor Regulatory Genes and Apoptotic Pathways in The Cytotoxic Effect of Cytochalasin H on Malignant Human Glioma Cell Line (U87MG), Cell journal, 2019, 21(1), 62–69.
17. Li, J, Gu, B, Chen, G, Ma, B, Xu, J, Zhang, G, et al., Cytochalasin E, a potential agent for anti-glioma therapy, efficiently induces U87 human glioblastoma cell death, Latin American Journal of Pharmacy, 2012, 31(1), 147-151.
18. Hwang, J, Yi, M, Zhang, X, Xu, Y, Jung, JH, Kim, DK, Cytochalasin B induces apoptosis through the mitochondrial apoptotic pathway in HeLa human cervical carcinoma cells, Oncology reports, 2013, 30(4), 1929–1935.
19. Rojo de la Vega, M, Chapman, E, Zhang, D.D, NRF2 and the Hallmarks of Cancer, Cancer cell, 2018, 34(1), 21–43.
20. Murakami, S, Motohashi, H, Roles of Nrf2 in cell proliferation and differentiation, Free radical biology & medicine, 2015, 88(Pt B), 168–178.
21. Wu, S, Lu, H, Bai, Y, Nrf2 in cancers: A double-edged sword, Cancer medicine, 2019, 8(5), 2252–2267.
22. Solis, L.M, Behrens, C, Dong, W, Suraokar, M, Ozburn, N.C, Moran, C.A, et al., II, Nrf2 and Keap1 abnormalities in non-small cell lung carcinoma and association with clinicopathologic features, Clinical cancer research: An official journal of the American Association for Cancer Research, 2010, 16(14), 3743–3753.
23. Sajadimajd, S, Khazaei, M, Oxidative Stress and Cancer: The Role of Nrf2, Current cancer drug targets, 2018, 18(6), 538–557.
24. Zimta, A.A, Cenariu, D, Irimie, A, Magdo, L, Nabavi, S.M, Atanasov, AG, Berindan-Neagoe, I, The Role of Nrf2 Activity in Cancer Development and Progression, Cancers, 2019, 11(11), 1755.
25. Basak, P, Sadhukhan, P, Sarkar, P, Sil, P.C, Perspectives of the Nrf-2 signaling pathway in cancer progression and therapy, Toxicology reports, 2017, 4, 306–318.
26. Ma, X, Zhang, J, Liu, S, Huang, Y, Chen, B, Wang, D, Nrf2 knockdown by shRNA inhibits tumor growth and increases efficacy of chemotherapy in cervical cancer, Cancer chemotherapy and pharmacology, 2012, 69(2), 485–494.
27. Lister, A, Nedjadi, T, Kitteringham, N.R, Campbell, F, Costello, E, Lloyd, B, et al., Nrf2 is overexpressed in pancreatic cancer: implications for cell proliferation and therapy, Molecular cancer, 2011, 10, 37.
28. DeNicola, G.M, Karreth, F.A, Humpton, T.J, Gopinathan, A, Wei, C, Frese, K, et al., Oncogene-induced Nrf2 transcription promotes ROS detoxification and tumorigenesis, Nature, 2011, 475(7354), 106–109.
29. Galadari, S, Rahman, A, Pallichankandy, S, Thayyullathil, F, Reactive oxygen species and cancer paradox: To promote or to suppress? Free radical biology & medicine, 2017, 104, 144–164.
30. Kim, J, Kim, J, Bae, JS, ROS homeostasis and metabolism: a critical liaison for cancer therapy, Experimental & molecular medicine, 2016, 48(11), e269.
31. Kalyanaraman, B, Cheng, G, Hardy, M, Ouari, O, Bennett, B, Zielonka, J, Teaching the basics of reactive oxygen species and their relevance to cancer biology: Mitochondrial reactive oxygen species detection, redox signaling, and targeted therapies, Redox biology, 2018, 15, 347–362.
32. Nguyen, C, Pandey, S, Exploiting mitochondrial vulnerabilities to trigger apoptosis selectively in cancer cells, Cancers, 2019, 11(7), 916.
33. Castaldo, S.A, Freitas, J.R, Conchinha, N.V, Madureira, P.A, The tumorigenic roles of the cellular REDOX regulatory systems, Oxidative medicine and cellular longevity, 2016, 8413032.
34. Hayes, J.D, McMahon, M, NRF2 and KEAP1 mutations: permanent activation of an adaptive response in cancer, Trends in biochemical sciences, 2009, 34(4), 176–188.
35. Rushworth, S.A, Bowles, K.M, MacEwan, D.J, High basal nuclear levels of Nrf2 in acute myeloid leukemia reduces sensitivity to proteasome inhibitors, Cancer research, 2011, 71(5), 1999–2009.
36. Takaishi, K, Kinoshita, H, Feng, G.G, Azma, T, Kawahito, S, Kitahata, H, Cytoskeleton-disrupting agent cytochalasin B reduces oxidative stress caused by high glucose in the human arterial smooth muscle, Journal of pharmacological sciences, 2020, 144(4), 197–203.