In Vitro Determination of Valproic Acid and Cisplatin Combination Antitumor Effect on Neuroblastoma Tumors Viability

Abstract

Neuroblastoma (SH-SY5Y) is one of the most common solid tumors in children, with aggressive and resistant features. Valproic acid (VPA) is a histone deacetylase inhibitor (HDAC) and is thought to have antitumoral activity. The aim of the current study is the evaluation of valproic acid and cisplatin (CSP) combination antitumor effects on SH-SY5Y. For this aim, the different doses of cisplatin (5, 10, and 15 μg/ml), VPA (5mM), and CSP (5, 10 and 15 μg/ml) + VPA (5 mM) were applied on SH-SY5Y tumor cell culture for 24 hours. For evaluation of cell viability, apoptosis, antioxidant and oxidant status, 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT), Anexin-V-FITC apoptosis, Total Antioxidant Capacity (TAC) and Total Oxidant Status (TOS) tests were done 24 hours after drug administration. According to our results, the combination of 15 μg/ml cisplatin and 5 mM VPA reduced cell viability and increased apoptosis status, compared with the negative control group (P<0.05). Our studies showed valproic acid increased the cisplatin effect and reduced the viability of tumor cells. The combination of VPA + CSP is more effective than only using cisplatin. In the light of these data, it is thought that the efficacy of VPA + CSP combined therapy should be investigated more comprehensively with in vivo studies.

Keywords

• Annexin V
• Cisplatin
• Neuroblastoma
• TAC
• TOS

Valproik Asit ve Cisplatin Kombinasyonunun Nöroblastom Tümörlerinde Canlılık Üzerine Antitümör Etkisinin İn Vitro Olarak Belirlenmesi

Öz

Nöroblastoma (SH-SY5Y), agresif ve dirençli özellikleri ile çocuklarda en sık görülen solid tümörlerden biridir. Valproik asit (VPA) bir histon deasetilaz inhibitörüdür (HDAC) ve antitümoral aktiviteye sahip olduğu düşünülmektedir. Bu çalışmanın amacı valproik asit ve sisplatin (CSP) kombinasyonun SH-SY5Y üzerindeki antitümör etkilerinin değerlendirilmesidir. Bu amaçla farklı dozlarda sisplatin (5, 10 ve 15 μg / ml), VPA (5mM) ve CSP (5, 10 ve 15 μg / ml) + VPA (5 mM) SH-SY5Y'ye hücre hattına 24 saat boyunca uygulanmıştır. Hücre canlılığı, apoptoz, antioksidan ve oksidan durumunun değerlendirilmesi için, 3- (4,5-Dimetiltiyazol-2-Yl) -2,5-Difeniltetrazolyum Bromür (MTT), Anexin-V-FITC apoptoz, Toplam Antioksidan Kapasite (TAC) ve Toplam Oksidan Durum (TOS) testleri ilaç uygulamasından 24 saat sonra yapılmıştır. Sonuçlarımıza göre, 15 μg / ml sisplatin ve 5 mM VPA kombinasyonu, negatif kontrol grubuna kıyasla hücre canlılığını azalttığı ve apoptoz durumunu arttırdığı tespit edilmiştir (P<0.05). Çalışmamızın sonuçları, valproik asidin cisplatin etkisini artırdığını ve kanser hücrelerinin canlılığını azalttığını ve VPA + CSP kombinasyonunun tek başına sispilatin uygulamasına kıyasla daha etkili olduğunu göstermektedir. Bu veriler ışığında VPA + CSP kombine tedavi etkinliği in vivo çalışmalar ile daha geniş kapsamlı araştırılması gerektiği düşünülmektedir.

Anahtar Kelimeler

• Annexin V
• Cisplatin
• Neuroblastoma
• TAC
• TOS

Kaynakça

1. 1. Matthay KK., Maris JM., Schleiermacher G., Nakagawara A., Mackall CL., Diller L., Weiss WA., 2016. Neuroblastoma. Nat Rev Dis Primers. Nov 10;2.
2. 2. Howman-Giles R., Shaw PJ., Uren RF., Chung DK., 2007. Neuroblastoma and other neuroendocrine tumors. Semin Nucl Med, 37, 286-302.
3. 3. Wagner LM., Danks MK., 2009. New Therapeutic targets for the treatment of high-risk neuroblastoma. J Cel Biochem, May 1, 46-57.
4. 4. Esmekaya MA., Kayhan H., Coskun A., Canseven AG., 2016. Effects of cisplatin electrochemotherapy on human neuroblastoma Cells. J Membrane Biol, 249, 601-610.
5. 5. Piskareva O., Harvey H., Nolan J., Conlon R., Alcock L., Buckley P., Dowling P., Henry M., O'Sullivan F., Bray I., Stallings RL., 2015. The development of cisplatin resistance in neuroblastoma is accompanied by epithelial to mesenchymal transition in vitro. Cancer Letters, 10; 142-155.
6. 6. Cruccetti A., Kiely EM., Spitz L., Drake DP., Pritchard J., Pierro A., 2000. Pelvic neuroblastoma: Low mortality and high morbidity. J Pediatr Surg. 35, 724-728.
7. 7. Matthay KK., Villablanca JG., Seeger RC., Stram DO., Harris RE., Ramsay NK., Swift P., Shimada H., Black CT., Brodeur GM., Gerbing RB, Reynolds CP., 1999. Treatment of high-risk neuroblastoma with intensive chemotherapy, radiotherapy, autologous bone marrow transplantation, and 13-cis-retinoic acid. Children's cancer group. The N Eng J Med, 14, 1165-1173.
8. 8. Whittle SB., Smith V., Doherty E., Zhao S., Mccarty S., Zage PE., 2017. Overview and recent advances in the treatment of neuroblastoma. Expert Rev Anticancer Ther, 17, 369-386.

9. 9. Vetter I., Mozar CA., Durek T., Wingerd JS., Alewood PF., Christie MJ., Lewis RJ., 2012. Characterisation of na(V) types endogenously expressed in human SH-SY5Y neuroblastoma cells. Biochem Pharmacol, 1, 1562-1571.
10. 10. Eckschlager T., Plch J., Stiborova M., Hrabeta J., 2017. Histone deacetylase inhibitors as anticancer drugs. Int J Mol Sci, 1, 18.
11. 11. Halaweish I., Bambakidis T., Chang Z., Wei H., Liu B., Li Y., Bonthrone T., Srinivasan A., Bonham T., Chtraklin K., Alam HB., 2015. Addition af low-dose valproic acid to saline resuscitation provides neuroprotection and improves long-term outcomes in a large animal model of combined traumatic brain injury and hemorrhagic shock. J Trauma Acute Care Surg, 79, 911-919.
12. 12. Hwabejire JO., Lu J., Liu B., Li Y., Halaweish I., Alam HB., 2014. Valproic acid for the treatment of hemorrhagic shock: a dose-optimization study. J Surg Res, 186, 363-370.
13. 13. Stockhausen MT., Sjolund J., Manetopoulos C., Axelson H., 2005. Effects of the histone deacetylase inhibitor valproic acid on notch signalling in human neuroblastoma cells. British J Cancer, 28, 751-759.
14. 14. Groh T., Hrabeta J,. Khalil MA., Doktorova H., Eckschlager T., Stiborova M., 2015. The synergistic effects of dna-damaging drugs cisplatin and etoposide with a histone deacetylase inhibitor valproate in high-risk neuroblastoma cells. Int J Oncology, 47, 343-352.
15. 15. Beinert T., Masuhr F., Mwela E., Schweigert M., Flath B., Harder H., B inder D., Oehm C., Behse F., Possinger K., 2000. Neuropathy under chemotherapy. European J Med Res, 30, 415-423.
16. 16. Rabik CA., Dolan ME., 2007. Molecular mechanisms of resistance and toxicity associated with platinating agents. Canc Treatment Rev, 33, 9-23.
17. 17. Mascaro-Cordeiro B., Oliveira ID., Tesser-Gamba F., Pavon LF., Saba-Silva N., Cavalheiro S., Dastoli P., Toledo SRC., 2018. Valproic acid treatment response in vitro is determined by tp53 status in medulloblastoma. Child's Nerv Sys, 34, 1497-1509.
18. 18. Sajadpoor Z., Amini-Farsani Z., Teimori H., Shamsara M., Sangtarash MH., Ghasemi-Dehkordi P., Yadollahi F., 2018. Valproic acid promotes apoptosis and cisplatin sensitivity through downregulation of h19 noncoding rna in ovarian a2780 cells. Appl Biochem Biotech, 185, 1132-1144.
19. 19. Taghizadehghalehjoughi A., Hacimuftuoglu A., Cetin M., Ugur AB., Galateanu B., Mezhuev Y., Okkay U., Taspinar N., Taspinar M., Uyanik A., Gundogdu B., Mohammadzadeh M., Nalci KA., Stivaktakis P., Tsatsakis A, Jung TW., Jeong JH., El-Aty AM., 2018. Effect of metformin/irinotecan-loaded poly-lactic-co-glycolic acid nanoparticles on glioblastoma: in vitro and in vivo studies. Nanomedicine, 13, 1595-1606.
20. 20. Kamalak H., Kamalak A., Taghizadehghalehjoughi A., Hacimuftuoglu A., Nalci KA., 2018. Cytotoxic and biological effects of bulk fill composites on rat cortical neuron cells. Odontology, 106, 377-388.
21. 21. Taghizadehghalehjoughi A., Yeni Y., Hacimuftuoglu A., 2018. Is Na+ channel blocker increase vincristine antitumor effect on neuroblastoma? J Clin Anal Med.
22. 22. Stiborova M., Eckschlager T., Poljakova J., Hrabeta J., Adam V., Kizek R., Frei E., 2012. The synergistic effects of DNA-targeted chemotherapeutics and histone deacetylase inhibitors as therapeutic strategies for cancer treatment. Current Med Chem. 19, 4218-4238.
23. 23. Tang R., Faussat AM., Majdak P., Perrot JY., Chaoui D., Legrand O., Marie JP., 2004. Valproic acid inhibits proliferation and induces apoptosis in acute myeloid leukemia cells expressing P-Gp and MRP1. Leukemia, 18, 1246-1251.
24. 24. Cerna T., Hrabeta J., Eckschlager T., Frei E., Schmeiser HH., Arlt VM., Stiborova M., 2018. The histone deacetylase inhibitor valproic acid exerts a synergistic cytotoxicity with the DNA-damaging drug ellipticine in neuroblastoma Cells. Int J Mol Sci, 5, 19.
25. 25. Blaheta RA., Michaelis M., Natsheh I., Hasenberg C., Weich E., Relja B., Jonas D., Doerr HW., Cinatl Jr J., 2007. Valproic acid inhibits adhesion of vincristine- and cisplatin-resistant neuroblastoma tumour cells to endothelium. Br J Cancer, 4, 1699-1706.
26. 26. Schuchmann M., Schulze-Bergkamen H., Fleischer B., Schattenberg JM., Siebler J., Weinmann A., Teufel A, Wörns M, Fischer T., Strand S., Lohse AW., Galle PR., 2006. Histone deacetylase inhibition by valproic acid down-regulates C-FLIP/CASH and sensitizes hepatoma cells towards CD95- and trail receptor-mediated apoptosis and chemotherapy. Oncology Rep, 15, 227-230.