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An Investigation of Anticancer Effects of Doxorubicin and Calcitriol Combination on MCF-7 Cells

Year 2018, Volume: 8 Issue: 2, 41 - 46, 12.11.2018

Abstract

DOI: 10.26650/experimed.2018.18001


Objectives: This study aimed to identify a
substance that both increases the efficiency and decreases the dose of
doxorubicin. Doxorubicin has considerable cardiac side effects at certain doses
when used treating breast cancer. Calcitriol, one of the vitamin D analogs
considered to have antiproliferative effects, was selected, and its cytotoxic
effects on the human breast cancer cell line MCF-7 were investigated in
combination with doxorubicin.



Materials and Methods: MCF-7 cell line was
treated with calcitriol in real time for 72 h in x-CELLigence. The
antiproliferative optimal dose of calcitriol was determined by time-dependent
cell index graph plotted using The xCELLigence Real-Time Cell Analysis (RTCA)
software program. The combination of different doses of doxorubicin and this
optimal dose of calcitriol was used to treat the MCF-7 cell line. Then,
Sulforhodamine-B (SRB) assay was conducted, and spectrophotometric measurements
were performed for cytotoxicity assay. The results of these spectrophotometric
measurements were analyzed by Student’s -test.



Results: The optimal antiproliferative
calcitriol dose detection of MCF-7 cells was performed using the time-dependent
cell index graph RTCA software program. Spectrophotometric measurements
obtained using the protein-staining sulforodamine B (SRB) assay for
cytotoxicity determination were statistically evaluated by the Student’s t-test
using the GraphPad Prism program. The optimal dose of calcitriol was determined
to be 250 nM. Different doses of doxorubicin (1.84-0.92 µM), calcitriol (250
nM), and calcitriol without the MCF-7 cell line were then used for detecting
the cytotoxic effect. The combination of 0.46 µM doxorubicin and the optimal
dose of calcitriol was found to be cytotoxic compared with other doses
(p=0.0087); however, it was not as effective as the dose reduction obtained
when using doxorubicin.



Conclusion: The combined use of doxorubicin
with calcitriol was found to have no significant effect in reducing the doses
presently being used. Hence, it is too early to state that a combination of
vitamin D and doxorubicin in breast cancer treatment will not have any
beneficial effects. Other vitamin D analogs might be potential candidates for
breast cancer treatment in further studies.

References

  • 1. Haydaroğlu A, Dubova S, Özsaran Z. Ege Üniversitesinde Meme Kanserleri, 3897 Olgunun Değerlendirilmesi. Meme Sağlığı Dergisi 2005; 1.
  • 2. Türkiye’de Kanser Önleme ve Taramaları 2014 Kısa Raporu. Kanser Daire Başkanlığı.
  • 3. Aydıntuğ S. Meme Kanserinde Erken Tanı TTB STED 2004; 6: 228.
  • 4. Wang Z, Chen JQ, Liu JL, Qin XG, Serious neutropenia following neoadjuvant chemotherapy for locally advanced breast cancer: A case report. Oncol Lett 2016; 11: 1597-9.
  • 5. Norman AW. Sunlight, season, skin pigmentation, vitamin D and 25-hydroxy vitamin D: Integral components of the vitamin D endocrine system. Am J Clin Nutr 1998; 67: 1108-10.
  • 6. Harbeck N, Salem, M, Nitz U, Gluz O, Liedtke C. Personalized treatment of early-stage breast cancer: present concepts and future directions. Cancer Treat Rev 2010; 36: 584-94.
  • 7. Nabholtz JM. Docetaxel-anthracycline combinations in metastatic breast cancer. Breast Cancer Res Treat 2003; 79: 3-9.
  • 8. Bagheri F, Safarian S, Eslaminejad M, Sheibani N. Effect of adriamycin on DNA, RNA, and protein synthesis in cell-free systems and intact cells. Cancer Res 1976; 36: 2891-5.
  • 9. Gudkov AV, Zelnick CR, Kazarov AR, Thimmapaya R, Suttle DP, Beck WT, Roninson IB, Thimmapaya, R, Suttle DP, Beck WT, Roninson IB. Isolation of genetic suppressor elements, inducing resistance to topoisomerase IIinteractive cytotoxic drugs, from human topoisomerase II cDNA. Proc Natl Acad Sci USA 1993; 90: 3231-5.
  • 10. Thorn C, Oshiro C, Marsh S, Hernandez-Boussard, McLeod H, Klein T and Altman R. Doxorubicin pathways: pharmacodynamics and adverse effects. Pharmacogenet Genomics 2011; 21: 440-6.
  • 11. Dimitrakis P, Romay-Ogando MI, Timolati F, Suter TM, Zuppinger C. Effects of doxorubicin cancer therapy on autophagy and the ubiquitin-proteasome system in long-term cultured adult rat cardiomyocytes. Cell Tissue Res 2012; 350: 361-72.
  • 12. Rivankar S. An overview of doxorubicin formulations in cancer therapy. J Cancer Res Ther 2014; 10: 853-8.
  • 13. Ayla Ş, Oktar H, Tanrıverdi G, Cengiz M, Özkılıç AÇ, Böttjer N, et al. Doksorubisin ile Oluşturulmuş Deneysel Kardiyotoksisite Üzerine Nikotinamidin Etkisi. Cerrahpaşa Tıp Derg 2008; 39: 7-14.
  • 14. Barinaga M. From bench top to bedside. Science 1997; 278: 1036-9.
  • 15. Cross HS. Extrarenal vitamin D hydroxylase expression and activity in normal and malignant cells: Modification of expression by epigenetic mechanisms and dietary substances. Nutr Rev 2007; 65: S108-S112.
  • 16. Robsahm TE, Tretli S, Dahlback A, Moan J. Vitamin D3 from sunlight may improve the prognosis of breast, colon and prostate cancer (Norway). Cancer Causes Control 2004; 15: 149-58.
  • 17. Tuohimaa P. Vitamin D, aging, and cancer. Nutr Rev 2008; 66: S147-S152.
  • 18. Bernardi RJ, Johnson CS, Modzelewski RA, Trump DL. Antiproliferative effects of 1 alpha, 25- dihydroxy vitamin D3 and vitamin D analogs on tumor-derived endothelial cells. Endocrinology 2002; 143: 2508-14.
  • 19. Chen ST, Pan TL, Tsai YC and Huang CM. Proteomics reveals protein profile changes in doxorubicin-treated MCF-7 human breast cancer cells. Cancer Lett 2002; 181: 95-107.
  • 20. Hines SL, Jorn HKS, Thompson KM, Larson JM. Breast cancer survivors and vitamin D: A review. Nutrition 2010; 26: 255-62.
  • 21. Colston KW, Hansen CM. Mechanisms implicated in the growth regulatory effects of vitamin D in breast cancer. Endocr Relat Cancer 2001; 9: 45-59.
  • 22. Jensen S, Madsen MW, Lukas J, Binderup L, Bartek J. Inhibitory effects of 1,25-dihydroxyvitamin D3 on the G1-S phase controlling machinery. Mol Endocrinol 2001; 15: 1370-80.
  • 23. Mehta RG, Mehta RR. Vitamin D and cancer. J Nutr Biochem 2002; 13: 252-64.
  • 24. Welsh JE, Wietzke JA, Zinser GM, Smyczek S, Romu S, Tribble E, et al. Impact ofthe vitamin D receptor on growth regulatory pathways in mammary gland and breast cancer. J Steroid Biochem Mol Biol 2002; 83: 85-92.
  • 25. Fu GK, Lin D, Zhang MY, Bikle DD, Shackleton CH, Miller WL, et al. Cloning of human 25-hydroxyvitamin D-1-hydroxylase and mutations causing vitamin D dependent rickets type I. Mol Endocrinol 1997; 11: 1961-70.
  • 26. Takeyama K, Kitanaka S, Sato T, Kobori M, Yanagisawa J, Kato S. 25-Hydroxyvitamin D3-1-hydroxylase and vitamin D synthesis. Science 1997; 277: 1827-30.
  • 27. Friedrich M, Reichrath J, Chen T, et al. Expression of 25- hydroxyvitamin D3 1- hydroxylase in breast tissue. In: Anthony W. Norman Jürgen Roth Lelio Orci, eds. Vitamin D endocrine system: structural, biological, genetic and clinical aspects. University of Riverside Press, 2000; 189-91.
  • 28. Bonadonna G, Brusamolino E, Valagussa P, Rossi A, Brugnatelli L, Brambilla C, et al. Combination chemotherapy as an adjuvant treatment in operable breast cancer. N Engl J Med 1976; 294: 405-10.
  • 29. Carter SK. Single and combination nonhormonal chemotherapy in breast cancer. Cancer 1972; 30: 1543-55.
  • 30. Jones SE, Durie BG, Salmon SE. Combination chemotherapy with adriamycin and cyclophosphamide for advanced breast cancer. Cancer 1975; 36: 90-7.
  • 31. Crown J, O’Leary M, Ooi WS. Docetaxel and paclitaxel in the treatment of breast cancer: a review of clinical experience. Oncologist 2004; 9: 24-32.
  • 32. Dieras V. Docetaxel in combination with doxorubicin: a phase I dose-finding study. Oncology (Williston Park) 1997; 11: 17-20.
  • 33. Kars MD, Iseri OD, Gündüz U, Ural AU, Arpaci F, Molnár J. Development of rational in vitro models for drug resistance in breast cancer and modulation of MDR by selected compounds. Anticancer Res 2006; 26: 4559-68.
  • 34. Li J, Xu LZ, He KL, Guo WJ, Zheng YH, Xia P, et al. Reversal effects of nomegestrol acetate on multidrug resistance in adriamycin-resistant MCF-7 breast cancer cell line. Breast Cancer Res 2001; 3: 253-63.
  • 35. Martin M. Docetaxel, doxorubicin and cyclophosphamide (the TAC regimen): an effective adjuvant treatment for operable breast cancer. Womens Health (Lond) 2006; 2: 527-37.
  • 36. Prados J, Melguizo C, Rama AR, Ortiz R, Segura A, Boulaiz H, et al. GEF gene therapy enhances the therapeutic efficacy of doxorubicin to combat growth of MCF-7 breast cancer cells. Cancer Chemother Pharmacol 2010; 66: 69-78.
  • 37. Bemd GJCM, Chang GTG. Vitamin D and Vitamin D Analogs in Cancer Treatment. Current Drug Targets 2002; 3: 85-94.
  • 38. Gissel T, Rejnmark L, Mosekilde L,Vestergaard P. Intake of vitamin D and risk of breast cancer; A meta-analysis. J Steroid Biochem Mol Biol 2008; 111: 195-9.
  • 39. Reichrath J, Friedrich M, Vogt T. Vitamin D and its analogs in cancer prevention and therapy. Anticancer Res 2012; 32: 209-10.
  • 40. Holick MF. Vitamin D: Its Role in cancer prevention and treatment. Prog Biophys Mol Biol 2006; 92: 49-59.
  • 41. Trump DL, Hershberger PA, Bernardi RJ, Ahmed S, Muindi J, Fakih M, et al. Anti Tumor activity of calcitriol. J Steroid Biochem Mol Biol 2004; 89-90: 519-26.

Kalsitriol ve Doksorubisin Kombinasyonunun MCF-7 Üzerine Olası Anti Kanser Etkilerinin Araştırılması

Year 2018, Volume: 8 Issue: 2, 41 - 46, 12.11.2018

Abstract

DOI: 10.26650/experimed.2018.18001


Amaç: Bu çalışmada amacımız, belirli bir
dozun üzerinde kardiyak yan etkileri oldukça fazla olan ve meme kanseri
tedavisinde kullanılan kemoterapötiklerden doksorubisinin, tedavide kullanım
miktarlarını azaltmayı ve etkinliğini arttırmayı sağlayan bir maddeyi
saptamaktı. Etken madde olarak anti proliferatif etkisi olduğu düşünülen
vitamin D analoglarından biri olan kalsitriol seçilmiş ve doksorubisin ile
kombine tedavisinin, insan meme kanseri hücre hattı MCF-7 üzerine sitotoksik
etkisinin saptanması amaçlanmıştır.

Gereç ve Yöntem: MCF-7 insan meme kanseri
hücre hattı kalsitriol ile muamele edilerek, gerçek zamanlı olarak, x-CELLigence
cihazında 72 saat inkübasyona bırakıldı ve kalsitriolün anti-proliferatif
optimum doz tespiti zamana bağlı hücre indeksi grafiği The xCELLigence
Real-Time Cell Analysis (RTCA) software programı kullanılarak yapıldı.
Kalsitriol optimum dozu ve doksorubisinin farklı dozlarının kombinasyonu MCF-7
hücre kültürü ile muamele edilerek sitotoksisite tayini için Sulforhodamine-B
(SRB) uygulaması ve spektrofotometrik ölçüm uygulandı. Spektrofotometrik ölçüm
sonuçları anlamlılık sonuçları için Student’s t-Testi ile değerlendirildi.

Bulgular: MCF-7 hücrelerinin
antiproliferative optimum kalsitriol doz tespiti zamana bağlı hücre indeksi
grafiği RTCA software programı kullanılarak yapılmıştır. Sitotoksisite tayini
için uygulanan SRB yöntemi sonucu elde edilen spektrofotometrik ölçüm sonuçları
GraphPad Prism programı kullanılarak Student’s t-testi ile istatistiksel olarak
değerlendirilmiştir. Kalsitriol optimum dozu 250 nM tespit edilmiştir.
Doksorubisinin farklı dozları (1,84 µM-0,92 µM), kalsitriol (250 nM) ve
kalsitriol olmaksızın MCF-7 hücre hattı ile sitotoksik etki saptaması için
muamele edilmiştir. 0,46 µM doksorubisin ve optimum kalsitriol kombinasyonunun
sitotoksik açıdan diğer dozlara göre anlamlı olduğu (p=0,0087) fakat
doksorubisin kullanımında doz azaltımını sağlayacak kadar etkin olmadığı
saptanmıştır.







Sonuç: Doksorubisinin, kalsitriol ile
kombine kullanımının günümüzde kullanılan dozlar üzerinde azaltıcı yönde
anlamlı bir etkisinin olmadığını saptanmıştır. Vitamin D ve doksorubisin
birlikte kullanımının meme kanserinde fayda sağlamayacağını söylemek için ise
henüz erkendir. İlerleyen çalışmalarda vitamin D’nin farklı analogları ile
çeşitli çalışmalar yapılabilir.

References

  • 1. Haydaroğlu A, Dubova S, Özsaran Z. Ege Üniversitesinde Meme Kanserleri, 3897 Olgunun Değerlendirilmesi. Meme Sağlığı Dergisi 2005; 1.
  • 2. Türkiye’de Kanser Önleme ve Taramaları 2014 Kısa Raporu. Kanser Daire Başkanlığı.
  • 3. Aydıntuğ S. Meme Kanserinde Erken Tanı TTB STED 2004; 6: 228.
  • 4. Wang Z, Chen JQ, Liu JL, Qin XG, Serious neutropenia following neoadjuvant chemotherapy for locally advanced breast cancer: A case report. Oncol Lett 2016; 11: 1597-9.
  • 5. Norman AW. Sunlight, season, skin pigmentation, vitamin D and 25-hydroxy vitamin D: Integral components of the vitamin D endocrine system. Am J Clin Nutr 1998; 67: 1108-10.
  • 6. Harbeck N, Salem, M, Nitz U, Gluz O, Liedtke C. Personalized treatment of early-stage breast cancer: present concepts and future directions. Cancer Treat Rev 2010; 36: 584-94.
  • 7. Nabholtz JM. Docetaxel-anthracycline combinations in metastatic breast cancer. Breast Cancer Res Treat 2003; 79: 3-9.
  • 8. Bagheri F, Safarian S, Eslaminejad M, Sheibani N. Effect of adriamycin on DNA, RNA, and protein synthesis in cell-free systems and intact cells. Cancer Res 1976; 36: 2891-5.
  • 9. Gudkov AV, Zelnick CR, Kazarov AR, Thimmapaya R, Suttle DP, Beck WT, Roninson IB, Thimmapaya, R, Suttle DP, Beck WT, Roninson IB. Isolation of genetic suppressor elements, inducing resistance to topoisomerase IIinteractive cytotoxic drugs, from human topoisomerase II cDNA. Proc Natl Acad Sci USA 1993; 90: 3231-5.
  • 10. Thorn C, Oshiro C, Marsh S, Hernandez-Boussard, McLeod H, Klein T and Altman R. Doxorubicin pathways: pharmacodynamics and adverse effects. Pharmacogenet Genomics 2011; 21: 440-6.
  • 11. Dimitrakis P, Romay-Ogando MI, Timolati F, Suter TM, Zuppinger C. Effects of doxorubicin cancer therapy on autophagy and the ubiquitin-proteasome system in long-term cultured adult rat cardiomyocytes. Cell Tissue Res 2012; 350: 361-72.
  • 12. Rivankar S. An overview of doxorubicin formulations in cancer therapy. J Cancer Res Ther 2014; 10: 853-8.
  • 13. Ayla Ş, Oktar H, Tanrıverdi G, Cengiz M, Özkılıç AÇ, Böttjer N, et al. Doksorubisin ile Oluşturulmuş Deneysel Kardiyotoksisite Üzerine Nikotinamidin Etkisi. Cerrahpaşa Tıp Derg 2008; 39: 7-14.
  • 14. Barinaga M. From bench top to bedside. Science 1997; 278: 1036-9.
  • 15. Cross HS. Extrarenal vitamin D hydroxylase expression and activity in normal and malignant cells: Modification of expression by epigenetic mechanisms and dietary substances. Nutr Rev 2007; 65: S108-S112.
  • 16. Robsahm TE, Tretli S, Dahlback A, Moan J. Vitamin D3 from sunlight may improve the prognosis of breast, colon and prostate cancer (Norway). Cancer Causes Control 2004; 15: 149-58.
  • 17. Tuohimaa P. Vitamin D, aging, and cancer. Nutr Rev 2008; 66: S147-S152.
  • 18. Bernardi RJ, Johnson CS, Modzelewski RA, Trump DL. Antiproliferative effects of 1 alpha, 25- dihydroxy vitamin D3 and vitamin D analogs on tumor-derived endothelial cells. Endocrinology 2002; 143: 2508-14.
  • 19. Chen ST, Pan TL, Tsai YC and Huang CM. Proteomics reveals protein profile changes in doxorubicin-treated MCF-7 human breast cancer cells. Cancer Lett 2002; 181: 95-107.
  • 20. Hines SL, Jorn HKS, Thompson KM, Larson JM. Breast cancer survivors and vitamin D: A review. Nutrition 2010; 26: 255-62.
  • 21. Colston KW, Hansen CM. Mechanisms implicated in the growth regulatory effects of vitamin D in breast cancer. Endocr Relat Cancer 2001; 9: 45-59.
  • 22. Jensen S, Madsen MW, Lukas J, Binderup L, Bartek J. Inhibitory effects of 1,25-dihydroxyvitamin D3 on the G1-S phase controlling machinery. Mol Endocrinol 2001; 15: 1370-80.
  • 23. Mehta RG, Mehta RR. Vitamin D and cancer. J Nutr Biochem 2002; 13: 252-64.
  • 24. Welsh JE, Wietzke JA, Zinser GM, Smyczek S, Romu S, Tribble E, et al. Impact ofthe vitamin D receptor on growth regulatory pathways in mammary gland and breast cancer. J Steroid Biochem Mol Biol 2002; 83: 85-92.
  • 25. Fu GK, Lin D, Zhang MY, Bikle DD, Shackleton CH, Miller WL, et al. Cloning of human 25-hydroxyvitamin D-1-hydroxylase and mutations causing vitamin D dependent rickets type I. Mol Endocrinol 1997; 11: 1961-70.
  • 26. Takeyama K, Kitanaka S, Sato T, Kobori M, Yanagisawa J, Kato S. 25-Hydroxyvitamin D3-1-hydroxylase and vitamin D synthesis. Science 1997; 277: 1827-30.
  • 27. Friedrich M, Reichrath J, Chen T, et al. Expression of 25- hydroxyvitamin D3 1- hydroxylase in breast tissue. In: Anthony W. Norman Jürgen Roth Lelio Orci, eds. Vitamin D endocrine system: structural, biological, genetic and clinical aspects. University of Riverside Press, 2000; 189-91.
  • 28. Bonadonna G, Brusamolino E, Valagussa P, Rossi A, Brugnatelli L, Brambilla C, et al. Combination chemotherapy as an adjuvant treatment in operable breast cancer. N Engl J Med 1976; 294: 405-10.
  • 29. Carter SK. Single and combination nonhormonal chemotherapy in breast cancer. Cancer 1972; 30: 1543-55.
  • 30. Jones SE, Durie BG, Salmon SE. Combination chemotherapy with adriamycin and cyclophosphamide for advanced breast cancer. Cancer 1975; 36: 90-7.
  • 31. Crown J, O’Leary M, Ooi WS. Docetaxel and paclitaxel in the treatment of breast cancer: a review of clinical experience. Oncologist 2004; 9: 24-32.
  • 32. Dieras V. Docetaxel in combination with doxorubicin: a phase I dose-finding study. Oncology (Williston Park) 1997; 11: 17-20.
  • 33. Kars MD, Iseri OD, Gündüz U, Ural AU, Arpaci F, Molnár J. Development of rational in vitro models for drug resistance in breast cancer and modulation of MDR by selected compounds. Anticancer Res 2006; 26: 4559-68.
  • 34. Li J, Xu LZ, He KL, Guo WJ, Zheng YH, Xia P, et al. Reversal effects of nomegestrol acetate on multidrug resistance in adriamycin-resistant MCF-7 breast cancer cell line. Breast Cancer Res 2001; 3: 253-63.
  • 35. Martin M. Docetaxel, doxorubicin and cyclophosphamide (the TAC regimen): an effective adjuvant treatment for operable breast cancer. Womens Health (Lond) 2006; 2: 527-37.
  • 36. Prados J, Melguizo C, Rama AR, Ortiz R, Segura A, Boulaiz H, et al. GEF gene therapy enhances the therapeutic efficacy of doxorubicin to combat growth of MCF-7 breast cancer cells. Cancer Chemother Pharmacol 2010; 66: 69-78.
  • 37. Bemd GJCM, Chang GTG. Vitamin D and Vitamin D Analogs in Cancer Treatment. Current Drug Targets 2002; 3: 85-94.
  • 38. Gissel T, Rejnmark L, Mosekilde L,Vestergaard P. Intake of vitamin D and risk of breast cancer; A meta-analysis. J Steroid Biochem Mol Biol 2008; 111: 195-9.
  • 39. Reichrath J, Friedrich M, Vogt T. Vitamin D and its analogs in cancer prevention and therapy. Anticancer Res 2012; 32: 209-10.
  • 40. Holick MF. Vitamin D: Its Role in cancer prevention and treatment. Prog Biophys Mol Biol 2006; 92: 49-59.
  • 41. Trump DL, Hershberger PA, Bernardi RJ, Ahmed S, Muindi J, Fakih M, et al. Anti Tumor activity of calcitriol. J Steroid Biochem Mol Biol 2004; 89-90: 519-26.
There are 41 citations in total.

Details

Primary Language English
Journal Section Research Article
Authors

Özge Bildiren This is me

Buse Cevatemre This is me

Ebru Nur Ay This is me

Güneş Özen This is me

Ceylan Hepokur This is me

Merve Erkısa This is me

Özlem Küçükhüseyin This is me

Engin Ulukaya This is me

Serap Kuruca This is me

İlhan Yaylım

Publication Date November 12, 2018
Submission Date July 5, 2018
Published in Issue Year 2018 Volume: 8 Issue: 2

Cite

Vancouver Bildiren Ö, Cevatemre B, Ay EN, Özen G, Hepokur C, Erkısa M, Küçükhüseyin Ö, Ulukaya E, Kuruca S, Yaylım İ. An Investigation of Anticancer Effects of Doxorubicin and Calcitriol Combination on MCF-7 Cells. Experimed. 2018;8(2):41-6.