The Investigation of the Effects of Calcitriol on Human Ovarian Carcinoma Cells

Abstract

Purpose: Ovarian cancer is the fifth leading cause of cancer death in women, leading cause of death from gynecologic malignancies, and the second most commonly diagnosed gynecologic malignancy, however the underlying pathophysiology is not clearly understood. Calcitriol, the active form of vitamin D serves its activity by binding to the vitamin D receptor (VDR). Calcitriol regulates multiple signaling pathways such as proliferation, apoptosis, differentiation, inflammation, invasion, angiogenesis and metastasis. It has been found to have a broad effect on several cancer types such as colon, breast and prostate cancer. Therefore, the study aimed to investigate the effects of calcitriol on human ovarian cancer cells. Material and Methods: The human MDAH-2774 ovarian carcinoma cells were exposed to different dose ranges of calcitriol for 24 and 48 hours. Cultured cells were evaluated in terms of MTT assay and quantitative- Real time-PCR. Results: As evidenced by the MTT assay, calcitriol treatment resulted in the reduction of cell viability in human MDAH-2774 cells. The gene expressions of VDR and p53 were increased with the calcitriol treatment compared to control. Additionally the gene expression of proapoptotic marker Bax increased and the anti-apoptotic marker Bcl-2 decreased with the presence of the calcitriol. Conclusion: In conclusion calcitriol treatment decreased cell proliferation and induced apoptosis in ovarian cancer cells, therefore we can suggest that calcitriol, either by itself or in combination with chemotherapy drugs, may be effective in treating ovarian cancer.

Keywords

• apoptosis
• calcitriol
• mtt assay
• ovarian cancer
• qrt-pcr

Kalsitriol’ün İnsan Ovaryum Kanser Hücrelerine Etkisinin Araştırılması

Abstract

Amaç: Over kanseri jinekolojik malignitelerden ölümlerin önde gelen nedenlerindendir. Kadınlarda kansere bağlı ölümlerde beşinci sırada ye almaktadır. D Vitaminin aktif formu olan kalsitirol, vitamin D reseptörüne (VDR) bağlanarak fonksiyon göstermektedir. Kalsitriol, proliferasyon, apoptoz, diferansiasyon, inflamasyon, invazyon, anjiyogenez ve metastaz ile ilgili çoklu sinyal yollarını düzenleyerek kanser gelişimini ve büyümesini etkileme potansiyeline sahiptir. Kolon, meme ve prostat kanseri büyümesini sağlayan spesifik sinyal yollarının kalsitriol ile düzenlemesi incelendiğinde kalsitriolün birçok farklı kanser türünde kanser hücreleri üzerinde geniş bir etkiye sahip olduğu görülmüştür. Çalışmamızın amacı kalsitriolün ovaryum kanser hücrelerine karşı etkisini araştırmaktır. Araçlar ve Yöntem: Çalışmamızda MDAH-2774 insan ovaryum kanseri hücre hattı kullanılmıştır. Hücreler farklı dozlarda kalsitriole 24 ve 48 saat maruz bırakıldıktan sonra MTT testi ve kantitatif gerçek zamanlı -PCR yöntemi uygulanmıştır. Bulgular: MTT testi sonucunda, kalsitirolün ovaryum kanser hücrelerinin canlılığını azalttığı tespit edildi. Kalsitriol uygulanan grupta kontrol grubuna kıyasla VDR ve p53 gen ekspresyonlarında artış saptandı. Bunlara ek olarak, kalsitriol uygulamasının proapoptotik belirteç Bax'ın gen expresyonunda artışa ve anti-apoptotik Bcl-2 ekspresyonunda azalmaya neden olduğu tespit edildi. Sonuç: Sonuç olarak kalsitriol tedavisinin, ovaryum kanser hücrelerinin proliferasyonunu azalttığı ve apoptozu indüklediği saptanmış olup, kasitriolün tek başına veya kemoterapi ilaçlarıyla kombinasyon halinde kullanılmasının ovaryum kanser tedavilerinde potansiyel bir rolü olduğu düşünülmektedir.

Keywords

• apoptoz
• kalsitriol
• mtt analizi
• ovaryum kanseri
• qrt-pcr

References

1. 1. Marczak A, Denel M. Trabectedin as a single agent and in combination with pegylated liposomal doxorubicin – activity against ovarian cancer cells. Contemp Oncol. 2014;18(3):149-152.
2. 2. Mersch J, Jackson MA, Park M, et al. Cancers associated with BRCA1 and BRCA2 mutations other than breast and ovarian. Cancer.2015;121(2):269-275.
3. 3. Liotta LA, Steeg PS, Stetler-Stevenson WG. Cancer metastasis and angiogenesis: an imbalance of positive and negative regulation. Cell.1991;64(2):327-336.
4. 4. Harter P, Bois A, Hahmann M, et al. Surgery in recurrent ovarian cancer. Ann Sur Oncol. 2006;13(12): 1702-1710.
5. 5. Elangovan H, Chahal S, Gunton JE. Vitamin D in Liver Disease: Current Evidence and Potential Directions. Biochim Biophys Acta Mol Basis Dis. 2017;1863(4):907-916.
6. 6. Wacker M, Holick MF. Vitamin D Effects on skeletal and extraskeletal health and the need for supplementation. Nutrients. 2013;5(1):111-148.
7. 7. Christakos S, Dhawan P, Verstuyf A, et al.Vitamin D: Metabolism, Molecular Mechanism of Action, and Pleiotropic Effects Vitamin D Analogs. Physiol Rev. 2016; 96(1):365-408.
8. 8. Feldman D, Krishnan AV, Swami S, et al.The role of vitamin D in reducing cancer risk and progression. Nat Rev Cancer. 2014;14(5):342-357.

9. 9. Wang Y, Zhu J,De Luca HF. Where is the vitamin D receptor? Arch Biochem Biophys. 2012;523(1):123-133.
10. 10. Sintov AC, Rarmolinsky L, Dahan A, et al. Pharmacological effects of vitamin D and its analogs: recent developments. Drug Discov Today. 2014;19(11):1769-1774.
11. 11. Koeffle HF, Hirji K, Itri L. 1,25-Dihydroxyvitamin D3: in vivo and in vitro effects on human preleukemic and leukemic cells. Cancer Treat Rep. 1985;69(2): 1399-1407.
12. 12. Hershberger PA, Modzelewski RA, Shurin ZR, et al. 1,25-Dihydroxycholecalciferol (1,25-D3) inhibits the growth of squamous cell carcinoma and down-modulates p21(Waf1/Cip1) in vitro and in vivo. Cancer Res. 1999;59(11):2644-2649.
13. 13. Lokeshwar BL, Schwartz GG, Selzer MG,et al. Inhibition of prostate cancer metastasis in vivo: a comparison of 1,23-dihydroxyvitamin D (calcitriol) and EB1089. Cancer Epidemiol Biomarkers Prev. 1999;8(3):241-248.
14. 14. Welsh J. Vitamin D and breast cancer: insights from animal models. Am J Clin Nutr. 2004;80(6):1721-1724.
15. 15. Diaz GD, Paraskeva C,Thomas MG,et al. Apoptosis is induced by the active metabolite of vitamin D3 and its analogue EB1089 in colorectal adenoma and carcinoma cells: possible implications for prevention and therapy. Cancer Res. 2000;60(8):2304-2312.
16. 16. Fatsa T, Hoşbul T, Elçi MP, et al. Antiproliferative, anti-inflammatory, antitumoral and proapoptotic effects of calcitriol on MCF-7 and MCF-10A cell lines. Indian J. Exp. Biol. 2023;61(5):320-328.
17. 17. Ricciardi CJ, Bae J, Esposito D, et al. 1,25-dihydroxyvitamin D3/vitamin D receptor suppresses brown adipocyte differentiation and mitochondrial respiration. Eur J Nutr. 2015;54(6):1001-1012.
18. 18. Gesmundo I, Silvagno F, Banfi D, et al. Calcitriol Inhibits Viability and Proliferation in Human Malignant Pleural Mesothelioma Cells. Front. Endocrinol. 2020;11:559-586.
19. 19. Feldman D, Krishnan AV, Swami S,et al. The role of vitamin D in reducing cancer risk and progression. Nat Rev Cancer. 2014;14(5):342-357.
20. 20. Bandera Merchan B, Morcillo S, Martin-Nunez G, et al. The role of vitamin D and VDR in carcinogenesis: through epidemiology and basic sciences. J Steroid Biochem Mol Biol. 2017;167:203-220. 21. Carlberg C, Munoz A. An update on vitamin D signaling and cancer. Semin Cancer Biol. 2020;79:217-230.
21. 22. Yin L, Grandi N, Raum E, Haug U, Arndt V, Brenner H. Meta-analysis: circulating vitamin D and ovarian cancer risk. Gynecol Oncol. 2011;121(2):369-375.
22. 23. Webb PM, de Fazio A, Protani MM, et al. Circulating 25-hydroxyvitamin D and survival in women with ovarian cancer. Am J Clin Nutr. 2015;102(1):109-114.
23. 24. Wong G, Lim WH, Lewis J, et al. Vitamin D and cancer mortality in elderly women. BMC Cancer. 2015;15:106. 25. Andraž D,Nina Fokter D. Vitamin D and Ovarian Cancer: Systematic Review of the Literature with a Focus on Molecular Mechanisms. Cells. 2020;9:335.
24. 26. Sajo E, Okunade K, Olorunfemi G, Rabiu K, Anorlu R. Serum Vıtamın D Defıcıency And Rısk Of Epıthelıal Ovarıan Cancer In Lagos, Nigeria. ecancer. 2020;14:1078.
25. 27. Ewa P, Agnieszka D, Zenon M, et.al. Calcitriol and Calcidiol Can Sensitize Melanoma Cells to Low–LET Proton Beam Irradiation. Int J Mol Sci. 2018;19(8): 2236.
26. 28. Consiglio M, Destefanis M, Morena D, et al.The vitamin D receptor inhibits the respiratory chain, contributing to the metabolic switch that is essential for cancer cell proliferation. PLoS ONE. 2014;9(12):e115816. 29. Ricca C , Aillon A, Bergandi L, et al. Vitamin D receptor is necessary for mitochondrial function and cell health. Int J Mol Sci. 2018;19(6):1672.
27. 30. Slominski AT, Brożyna AA, Zmijewski MA, et al. Vitamin D signaling and melanoma: Role of vitamin D and its receptors in melanoma progression and management. Lab Investig. 2017;97(6):706-724.
28. 31. Consiglio M, Viano W, Casarin S, et al. Mitochondrial and lipogenic effects of vitamin D on differentiating and proliferating human keratinocytes. Exp Dermatol. 2015;24(10):748-753.
29. 32. Argiris A, Cohen E, Karrison T, et al. A phase II trial of perifosine, an oral alkylphospholipid, in recurrent or metastatic head and neck cancer. Cancer Biol Ther. 2006;5(7):766-770.
30. 33. Bao A, Li Y, Tong Y. 1,25-Dihydroxyvitamin D3 and cisplatin syner¬gistically induce apoptosis and cell cycle arrest in gastric cancer cells. Int J Mol Med. 2014;33(5):1177-1184.
31. 34. Chaitanya GV, Alexander JS, Babu PP. PARP-1 cleavage fragments: Signatures of cell-death proteases in neurodegeneration. Cell Commun Signal. 2010;8:31.
32. 35. Ohnishi T,Takahashi A, Ohnishi K. Studies about space radiation promote new fields in radiation biology. J Radiat Res. 2002;43:7‐12.
33. 36. Campbell MJ, Trump DL. Vitamin D receptor signaling and cancer. Endocrinol Metab Clin North Am. 2017; 46:1009-1038.