Sentezlenen Pirazol-Akridin Türevinin SH-SY5Y İnsan Nöroblastoma Hücrelerinde Antikanser Özelliklerinin Değerlendirilmesi

Abstract

Amaç: Bu çalışmanın amacı yeni bir pirazol akridin türevi (3-ACH) sentezlemek ve bu bileşiğin SH-SY5Y insan nöroblastoma hücreleri üzerindeki antikanser özelliklerini değerlendirmektir. Gereç ve Yöntemler: Pirazol-4-karbaldehit türevi, 3-ACH sentezlemek için dimedon ve p-nitroanilin ile siklize edildi. Bileşiğin karakterizasyonu FT-IR, NMR, HPLC-Q-TOF/MS ve element analizi kullanılarak yapıldı. 3-ACH bileşiğinin SH-SY5Y insan nöroblastoma hücreleri üzerindeki sitotoksik etkisi, doz (50, 100 ve 150 μg/mL) ve zaman (12, 24 ve 48 saat) bağımlı bir şekilde WST-1 testi kullanılarak değerlendirildi. 3-ACH'nin apoptoz üzerindeki etkisi, Kaspaz-3, -8, -9 ve BAX proteinleri için immün boyama yolu ile araştırıldı. Bulgular: 3-ACH, 100 μg/mL konsantrasyonda 24 saatlik tedaviden sonra SH-SY5Y hücre canlılığını %89,89±7,63 (p=0,002) oranında azaltmıştır. 24 saatte, daha yüksek (150 μg/mL) konsantrasyon benzer bir azalma gösterirken (%90,53±2,88, p=0,004), daha düşük (50 μg/mL) konsantrasyon hücre canlılığını korumuştur (%98,37±1,67, p=0,903). 50 μg/mL (%94,55±0,65), 100 μg/mL (%95,18±1,41) ve 150 μg/mL (%95,28±2,57) dozlarının tümü, 48 saatte hücre canlılık oranlarını önemli ölçüde azaltmıştır (tüm p<0,001). İmmün boyama, 24 saat boyunca 3-ACH (100 μg/mL) ile muamele edilen hücrelerde, BAX, Kaspaz-3, -8 ve -9 proteinlerinin sentezinde kontrol grubuna kıyasla önemli bir artış olduğunu ortaya koymuştur. Sonuç: Bu sonuçlar, 3-ACH'nin hem intrinsik hem de ekstrinsik yolakları aktive ederek SH-SY5Y hücrelerinde, sitotoksik etkilerine katkıda bulunabilecek olan, apoptozu indükleme potansiyeline sahip olduğunu göstermektedir. Bu çalışma, 3-ACH bileşiğinin antikanser terapötik ajan olarak potansiyelini destekleyen umut verici kanıtlar sunmaktadır.

Keywords

• Kimyasal sentez
• antineoplastik ajanlar
• nöroblastoma
• pirazoller
• akridinler
• sitotoksisite
• apoptoz

Assessment of Anticancer Properties of Synthesized Pyrazole-Acridine Derivative on SH-SY5Y Human Neuroblastoma Cells

Abstract

Aim: This study aimed to synthesize a novel pyrazole acridine derivative (3-ACH) and evaluate its anticancer properties on SH-SY5Y human neuroblastoma cells. Material and Methods: The pyrazole-4-carbaldehyde derivative was cyclized with dimedone and p-nitroaniline to synthesize the 3-ACH. Characterization of the compound was performed using FT-IR, NMR, HPLC-Q-TOF/MS, and elemental analysis. The cytotoxic impact of the 3-ACH compound on SH-SY5Y human neuroblastoma cells was evaluated using the WST-1 assay in a dose- (50, 100, and 150 μg/mL) and time-dependent (12, 24, and 48 hours) manner. The impact of 3-ACH on apoptosis was investigated through immunostaining for Caspase-3, -8, and -9, and BAX proteins. Results: 3-ACH reduced SH-SY5Y cell viability with the rate of 89.89±7.63% (p=0.002) at 100 μg/mL concentration after 24 hours of treatment. While a higher (150 μg/mL) concentration showed a similar reduction (90.53±2.88%, p=0.004), the lower (50 μg/mL) concentration maintained high cell viability (98.37±1.67%, p=0.903) at 24 hours. All doses of 50 μg/mL (94.55±0.65%), 100 μg/mL (95.18±1.41%), and 150 μg/mL (95.28±2.57%) significantly reduced cell viability rates, at 48 hours (all p<0.001). Immunostaining revealed a significant upregulation in the synthesis of BAX, Caspase-3, -8, and -9 proteins in cells treated with 3-ACH (100 μg/mL) for 24 hours compared to the control group. Conclusion: These results indicate that 3-ACH has the potential to induce apoptosis in SH-SY5Y cells by activating both the intrinsic and extrinsic pathways, which may contribute to its cytotoxic effects. This study provides promising evidence supporting the potential of the 3-ACH compound as an anticancer therapeutic agent.

Keywords

• Chemical synthesis
• antineoplastic agents
• neuroblastoma
• pyrazoles
• acridines
• cytotoxicity
• apoptosis

References

1. Nandurkar D, Danao K, Rokde V, Shivhare R, Mahajan U. Pyrazole scaffold: Strategies toward the synthesis and their applications. In: Kumari P, Patel AB, editors. Strategies for the synthesis of heterocycles and their applications. London, UK: IntechOpen; 2022. p15-38.
2. Kumar V, Kaur K, Gupta GK, Sharma AK. Pyrazole containing natural products: Synthetic preview and biological significance. Eur J Med Chem. 2013;69:735-53.
3. Karrouchi K, Radi S, Ramli Y, Taoufik J, Mabkhot YN, Al-Aizari FA, et al. Synthesis and pharmacological activities of pyrazole derivatives: A review. Molecules. 2018;23(1):134.
4. Nitulescu GM. Quantitative and qualitative analysis of the anti-proliferative potential of the pyrazole scaffold in the design of anticancer agents. Molecules. 2022;27(10):3300.
5. Costa RF, Turones LC, Cavalcante KVN, Rosa Júnior IA, Xavier CH, Rosseto LP, et al. Heterocyclic compounds: Pharmacology of pyrazole analogs from rational structural considerations. Front Pharmacol. 2021;12:666725.
6. Alfei S, Brullo C, Caviglia D, Piatti G, Zorzoli A, Marimpietri D, et al. Pyrazole-based water-soluble dendrimer nanoparticles as a potential new agent against staphylococci. Biomedicines. 2021;10(1):17.
7. Kornis GI. Pyrazoles, pyrazolines, and pyrazolones. In: Kirk-Othmer encyclopedia of chemical technology. 3rd ed. New York: John Wiley & Sons; 2000. p.436-53.
8. National Center for Biotechnology Information. PubChem compound summary for CID 9215, Acridine. 2023. [Cited: 2024 Jul 23]. Available from: https://pubchem.ncbi.nlm.nih.gov/compound/Acridine

9. Prasher P, Sharma M. Medicinal chemistry of acridine and its analogues. Medchemcomm. 2018;9(10):1589-618.
10. Varakumar P, Rajagopal K, Aparna B, Raman K, Byran G, Gonçalves Lima CM, et al. Acridine as an anti-tumour agent: A critical review. Molecules. 2022;28(1):193.
11. Gackowski M, Szewczyk-Golec K, Pluskota R, Koba M, Mądra-Gackowska K, Woźniak A. Application of multivariate adaptive regression splines (MARSplines) for predicting antitumor activity of anthrapyrazole derivatives. Int J Mol Sci. 2022;23(9):5132.
12. Adjei AA. Current status of pyrazoloacridine as an anticancer agent. Invest New Drugs. 1999;17(1):43-8.
13. Elmusa M, Elmusa S, Mert S, Kasımoğulları R, Türkan F, Atalar MN, et al. One-pot three-component synthesis of novel pyrazolo-acridine derivatives and assessment of their acetylcholinesterase inhibitory properties: An in vitro and in silico study. J Mol Struct. 2023;1274(1):134553.
14. Küçükbağrıaçık Y, Dastouri MR, Elmusa M, Elmusa F, Yılmaz H, Kasımoğulları R. Evaluation of the potential anticancer activity of pyrazole-acridine derivative synthesis on SKBR-3 human breast cancer cell line. KÜ Tıp Fak Derg. 2024;26(1):76-85. Turkish.
15. Kucukbagriacik Y, Dastouri M, Yilmaz H, Altuntas EG. The apoptotic effect of the Lycopodium clavatum extracts on MCF-7 human breast cancer cells. Med Oncol. 2023;40(10):289.
16. Nandiyanto ABD, Oktiani R, Ragadhita R. How to read and interpret FTIR spectroscope of organic material. Indonesian J Sci Technol. 2019;4(1):97-118.
17. Gunawan R, Nandiyanto ABD. How to read and interpret 1H-NMR and 13C-NMR spectrums. Indonesian J Sci Technol. 2021;6(2):267-98.
18. Zhai H, Zhang S, Ampomah-Wireko M, Wang H, Cao Y, Yang P, et al. Pyrazole: An important core in many marketed and clinical drugs. Russ J Bioorg Chem. 2022;48(6):1175-89.
19. Alam MJ, Alam O, Naim MJ, Nawaz F, Manaithiya A, Imran M, et al. Recent advancement in drug design and discovery of pyrazole biomolecules as cancer and inflammation therapeutics. Molecules. 2022;27(24):8708.
20. Di Nicola C, Marchetti F, Pettinari C, Pettinari R, Brisdelli F, Crucianelli M, et al. Synthesis and characterization of a new alkyne functionalized bis(pyrazolyl)methane ligand and of its Pd(II) complexes: Evaluation of their in vitro cytotoxic activity. Inorgonica Chim Acta. 2017;455(2):677-82.
21. Mohan G, Sridhar G, Laxminarayana E, Chary MT. Synthesis and biological evaluation of 1,2,4-oxadiazole incorporated 1,2,3-triazole-pyrazole derivatives as anticancer agents. Chem Data Coll. 2021;34:100735.
22. Akhtar W, Marella A, Alam MM, Khan MF, Akhtar M, Anwer T, et al. Design and synthesis of pyrazole-pyrazoline hybrids as cancer-associated selective COX-2 inhibitors. Arch Pharm (Weinheim). 2021;354(1):e2000116.
23. Varakumar P, Rajagopal K, Aparna B, Raman K, Byran G, Gonçalves Lima CM, et al. Acridine as an anti-tumour agent: A critical review. Molecules. 2022;28(1):193.
24. Rupar JS, Dobričić VD, Aleksić MM, Brborić JS, Čudina OA. A review of published data on acridine derivatives with different biological activities. Kragujevac J Sci. 2018;40:83-101.
25. Joubert JP, Smit FJ, du Plessis L, Smith PJ, N’Da DD. Synthesis and in vitro biological evaluation of aminoacridines and artemisinin-acridine hybrids. Eur J Pharm Sci. 2014;56:16-27.
26. Hu Y, Krishan A, Nie W, Sridhar KS, Mayer LD, Bally M. Synergistic cytotoxicity of pyrazoloacridine with doxorubicin, etoposide, and topotecan in drug-resistant tumor cells. Clin Cancer Res. 2004;10(3):1160-9.
27. Plaxe SC, Blessing JA, Bookman MA, Creasman WT. Phase II trial of pyrazoloacridine in recurrent platinum-sensitive ovarian cancer: a Gynecologic Oncology Group study. Gynecol Oncol. 2002;84(1):32-5.
28. Ramaswamy B, Mrozek E, Kuebler JP, Bekaii-Saab T, Kraut EH. Phase II trial of pyrazoloacridine (NSC#366140) in patients with metastatic breast cancer. Invest New Drugs. 2011;29(2):347-51.
29. Sugaya T, Mimura Y, Shida Y, Osawa Y, Matsukuma I, Ikeda S, et al. 6H-pyrazolo[4,5,1-de]acridin-6-ones as a novel class of antitumor agents. Synthesis and biological activity. J Med Chem. 1994;37(7):1028-32.
30. Jan R, Chaudhry GE. Understanding apoptosis and apoptotic pathways targeted cancer therapeutics. Adv Pharm Bull. 2019;9(2):205-18.