Atorvastatin blocks Sonic Hedgehog medulloblastoma progression through apoptosis

R.E. Gordon^a,b*, Z.I. Abramova^a**

^aKazan Federal University, Kazan, 420008 Russia

^bFox Chase Cancer Center, Philadelphia, 19111 USA

E-mail: ^*renata.e.gordon@gmail.com, ^**ziabramova@mail.ru

Received July 25, 2017

Full text PDF

Abstract

Aberrant activation of Sonic Hedgehog signaling transduction leads to formation of numerous   malignancies, including medulloblastoma (MB) – the most common malignant pediatric tumor. Earlier, we have shown that endogenous cholesterol acts as a direct activator of the Sonic Hedgehog signaling and is a promising target for treating MB. Histochemical analysis of the effect of atorvastatin, which is widely used in clinical practice as a cholesterol biosynthesis inhibitor, revealed a high level of cell death in the tumor tissue via apoptosis. These data show the potential of using statins for the targeted therapy of medulloblastoma, as well as other Sonic Hedgehog-driven malignancies.

Keywords: medulloblastoma, Sonic Hedgehog, statins, atorvastatin, cholesterol, apoptosis

Acknowledgments. The study was supported by the Russian Government Program of Competitive Growth of Kazan Federal University.

Figure Captions

Fig. 1. The scheme of Shh signal transduction in the cell.

Fig. 2. The primary cell culture of mouse MB cells after 72-h incubation: a – control sample (DMSO), b – atorvastatin (10 nmol), c – atorvastatin (50 nmol). (dose-dependent decrease in the number of viable cells, identified by the low number of cells per visual field). Scale: 10 ?m.

Fig. 3. The assessment of the proliferation level (BrdU-positive cells) in the primary mouse MB cell culture after 72-h incubation using IF-analysis: a – control sample (DMSO), b – atorvastatin (10 nmol), c – atorvastatin (50 nmol). Scale: 10 ?m.

Fig. 4. The quantitative analysis of proliferation (BrdU-positive cells, %) in the primary mouse MB cell culture after 72-h incubation. * – p < 0.05, ** – p < 0.005.

Fig. 5. The analysis of Gli1 protein expression in the primary culture of Ptch1^+/– MB cell culture by Western blotting. GAPDH protein analysis was used as a control variant.

Fig. 6. The IF-analysis of apoptosis level (caspase3-positive cells) in the tissue of Ptch1^+/– MB mice after three weeks of atorvastatin administration: a – control sample (DMSO), b – atorvastatin (10 mg/kg/day), c – atorvastatin (40 mg/kg/day). Scale: 10 ?m.

Fig. 7. The quantitative analysis of apoptosis level (caspase3-positive cells, %) in mouse MB allograft tissues after three weeks of treatment with atorvastatin. * – p < 0.05, ns – p > 0.05.

References

1.     Klesse L.J., Bowers D.C. Childhood medulloblastoma: Current status of biology and treatment. CNS Drugs, 2010, vol. 24, no. 4, pp 285–301. doi: 10.2165/11530140-000000000-00000.

2.     Yang Z.J., Ellis T., Markant S.L., Read T.A., Kessler J.D., Bourboulas M., Schuller U., Machold R., Fishell G., Rowitch D.H., Wainwright B.J., Wechsler-Reya R.J. Medulloblastoma can be initiated by deletion of Patched in lineage-restricted progenitors or stem cells. Cancer Cell, 2008, vol. 14, no. 2, pp. 135–145. doi: 10.1016/j.ccr.2008.07.003.

3.     Packer R.J., Zhou T., Holmes E., Vezina G., Gajjar A. Survival and secondary tumors in children with medulloblastoma receiving radiotherapy and adjuvant chemotherapy: Results of Children's Oncology Group trial A9961. Neuro Oncol., 2013, vol. 15, no. 1, pp. 97–103. doi: 10.1093/neuonc/nos267.

4.     Roussel M.F., Robinson G. Medulloblastoma: Advances and challenges. F1000 Biol. Rep., 2011, vol. 3, no. 5, pp. 1–3. doi: 10.3410/B3-5.

5.     Taylor M.D., Northcott P.A., Korshunov A., Remke M., Cho Y.J., Clifford S.C., Eberhart C.G., Parsons D.W., Rutkowski S., Gajjar A., Ellison D.W., Lichter P., Gilbertson R.J., Pomeroy S.L., Kool M., Pfister S.M. Molecular subgroups of medulloblastoma: The current consensus. Acta Neuropathol., 2012, vol. 123, no. 4, pp. 465–472. doi: 10.1007/s00401-011-0922-z.

6.     Gajjar A.J., Robinson G.W. Medulloblastoma-translating discoveries from the bench to the bedside. Nat. Rev. Clin. Oncol., 2014, vol. 11, no. 12, pp. 714–722. doi: 10.1038/nrclinonc.2014.181.

7.     Marti E., Bovolenta P. Sonic hedgehog in CNS development: One signal, multiple outputs. Trends Neurosci., 2002, vol. 25, no. 2, pp. 89–96.

8.     Hui C.C., Angers S. Gli proteins in development and disease. Annu. Rev. Cell Dev. Biol., 2011, vol. 27, pp. 513–537. doi: 10.1146/annurev-cellbio-092910-154048.

9.     Fuse N., Maiti T., Wang B., Porter J.A., Hall T.M., Leahy D.J., Beachy P.A. Sonic hedgehog protein signals not as a hydrolytic enzyme but as an apparent ligand for patched. Proc. Natl. Acad. Sci. U. S. A., 1999, vol. 96, no. 20, pp. 10992–10999.

10.   Ruiz i Altaba A., Sanchez P., Dahmane N. Gli and hedgehog in cancer: tumours, embryos and stem cells. Nat. Rev. Cancer, 2002, vol. 2, no. 5, pp. 361–372. doi: 10.1038/nrc796.

11.   Orth M., Bellosta S. Cholesterol: Its regulation and role in central nervous system disorders. Cholesterol, 2012, vol. 2012, art. 292598, pp. 1–19. doi: 10.1155/2012/292598.

12.   Riobo N.A. Cholesterol and its derivatives in Sonic Hedgehog signaling and cancer. Curr. Opin. Pharmacol., 2012, vol. 12, no. 6, pp. 736–741. doi: 10.1016/j.coph.2012.07.002.

13.   Ryan K.E., Chiang C. Hedgehog secretion and signal transduction in vertebrates. J. Biol. Chem., 2012, vol. 287, no. 22, pp. 17905–17913. doi: 10.1074/jbc.R112.356006.

14.   Byrne E.F.X., Sircar R., Miller P.S., Hedger G., Luchetti G., Nachtergaele S., Tully M.D., Mydock-McGrane L., Covey D.F., Rambo R.P., Sansom M.S.P, Newstead S., Rohatgi R., Siebold C. Structural basis of Smoothened regulation by its extracellular domains. Nature, 2016, vol. 535, no. 7613, pp. 517–522. doi: 10.1038/nature18934.

15.   Huang P., Nedelcu D., Watanabe M., Jao C., Kim Y., Liu J., Salic A. Cellular cholesterol directly activates Smoothened in Hedgehog signaling. Cell, 2016, vol. 166, no. 5, pp. 1176–1187. doi: 10.1016/j.cell.2016.08.003.

16.   Luchetti G., Sircar R., Kong J.H., Nachtergaele S., Sagner A., Byrne E.F., Covey D.F., Siebold C., Rohatgi R. Cholesterol activates the G-protein coupled receptor Smoothened to promote Hedgehog signaling. eLife, 2016, vol. 5, art. e20304, pp. 1–22. doi: 10.7554/eLife.20304.

17.   Zhang J., Liu Q. Cholesterol metabolism and homeostasis in the brain. Protein Cell, 2015, vol. 6, no. 4, pp. 254–264. doi: 10.1007/s13238-014-0131-3.

18.   Dietschy J.M., Turley S.D. Cholesterol metabolism in the brain. Curr. Opin. Lipidol., 2001, vol. 12, no. 2, pp. 105–112.

19.   Goldstein J.L., Brown M.S. Regulation of the mevalonate pathway. Nature, 1990, vol. 343, no. 6257, pp. 425–430. doi: 10.1038/343425a0.

20.   Collins R., Reith C., Emberson J., Armitage J., Baigent C., Blackwell L., Blumenthal R., Danesh J., Smith G.D., DeMets D., Evans S., Law M., MacMahon S., Martin S., Neal B., Poulter N., Preiss D., Ridker P., Roberts I., Rodgers A., Sandercock P., Schulz K., Sever P., Simes J., Smeeth L., Wald N., Yusuf S., Peto R. Interpretation of the evidence for the efficacy and safety of statin therapy. Lancet, 2016, vol. 388, no. 10059, pp. 2532–2561. doi: 10.1016/S0140-6736(16)31357-5.

21.   Dimitroulakos J., Ye L.Y., Benzaquen M., Moore M.J., Kamel-Reid S., Freedman M.H., Yeger H., Penn L.Z. Differential sensitivity of various pediatric cancers and squamous cell carcinomas to lovastatin-induced apoptosis: Therapeutic implications. Clin. Cancer Res., 2001, vol. 7, no. 1, pp. 158–167.

22.   Macaulay R.J., Wang W., Dimitroulakos J., Becker L.E., Yeger H. Lovastatin-induced apoptosis of human medulloblastoma cell lines in vitro. J. Neurooncol., 1999, vol. 42, no. 1, pp. 1–11.

23.   Wang W., Macaulay R.J. Mevalonate prevents lovastatin-induced apoptosis in medulloblastoma cell lines. Can. J. Neurol. Sci., 1999, vol. 26, no. 4, pp. 305–310.

24.   Sasai K., Romer J.T., Lee Y., Finkelstein D., Fuller C., McKinnon P.J., Curran T. Shh pathway activity is down-regulated in cultured medulloblastoma cells: Implications for preclinical studies. Cancer Res., 2006, vol. 66, no. 8, pp. 4215–4222. doi: 10.1158/0008-5472.CAN-05-4505.

25.   Gage G.J., Kipke D.R., Shain W. Whole animal perfusion fixation for rodents. J. Visualized Exp., 2012, vol.65, art. e3564, pp. 1–9. doi: 10.3791/3564.

26.   Porter A.G., Janicke R.U. Emerging roles of caspase-3 in apoptosis. Cell Death Differ., 1999, vol. 6, no. 2, pp. 99–104. doi: 10.1038/sj.cdd.4400476.

For citation: Gordon R.E., Abramova Z.I. Atorvastatin blocks Sonic Hedgehog medulloblastoma progression through apoptosis. Uchenye Zapiski Kazanskogo Universiteta. Seriya Estestvennye Nauki, 2017, vol. 159, no. 4, pp. 550–563. (In Russian)

The content is available under the license Creative Commons Attribution 4.0 License.