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Combination of Emricasan with Ponatinib Synergistically Reduces Ischemia/Reperfusion Injury in Rat Brain Through Simultaneous Prevention of Apoptosis and Necroptosis

Abstract

Apoptosis and receptor-interacting protein kinase 1/3(RIPK1/3)-mediated necroptosis contribute to the cerebral ischemia/reperfusion (I/R) injury. Emricasan is an inhibitor of caspases in clinical trials for liver diseases while ponatinib could be a potential inhibitor for RIPK1/3. This study aims to investigate the effect of emricasan and/or ponatinib on cerebral I/R injury and the underlying mechanisms. Firstly, we evaluated the status of apoptosis and necroposis in a rat model of cerebral I/R under different conditions, which showed noticeable apoptosis and necroptosis under condition of 2-h ischemia and 24-h reperfusion; next, the preventive or therapeutic effect of emricasan or ponatinib on cerebral I/R injury was tested. Administration of emricasan or ponatinib either before or after ischemia could decrease the neurological deficit score and infarct volume; finally, the combined therapeutic effect of emricasan with ponatinib on I/R injury was examined. Combined application of emricasan and ponatinib could further decrease the I/R injury compared to single application. Emricasan decreased the activities of capase-8/-3 in the I/R-treated brain but not the protein levels of necroptosis-relevant proteins: RIPK1, RIPK3, and mixed lineage kinase domain-like (MLKL), whereas ponatinib suppressed the expressions of these proteins but not the activities of capase-8/-3. Combination of emricasan with ponatinib could suppress both capase-8/-3 and necroptosis-relevant proteins. Based on these observations, we conclude that combination of emricasan with ponatinib could synergistically reduce I/R injury in rat brain through simultaneous prevention of apoptosis and necroptosis. Our findings might lay a basis on extension of the clinical indications for emricasan and ponatinib in treating ischemic stroke.

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References

  1. 1.

    Catanese L, Tarsia J, Fisher M. Acute ischemic stroke therapy overview. Circ Res. 2017;120(3):541–58.

    Article  PubMed  CAS  Google Scholar 

  2. 2.

    Bai J, Lyden PD. Revisiting cerebral postischemic reperfusion injury: new insights in understanding reperfusion failure, hemorrhage, and edema. Int J Stroke. 2015;10(2):143–52.

    Article  PubMed  Google Scholar 

  3. 3.

    Khoshnam SE, Winlow W, Farzaneh M, Farbood Y, Moghaddam HF. Pathogenic mechanisms following ischemic stroke. Neurol Sci. 2017;38(7):1167–86.

    Article  PubMed  Google Scholar 

  4. 4.

    Galluzzi L, Lopez-Soto A, Kumar S, Kroemer G. Caspases connect cell-death signaling to organismal homeostasis. Immunity. 2016;44(2):221–31.

    Article  PubMed  CAS  Google Scholar 

  5. 5.

    Elmore S. Apoptosis: a review of programmed cell death. Toxicol Pathol. 2007;35(4):495–516.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. 6.

    Shabanzadeh AP, D'Onofrio PM, Monnier PP, Koeberle PD. Targeting caspase-6 and caspase-8 to promote neuronal survival following ischemic stroke. Cell Death Dis. 2015;6:e1967.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  7. 7.

    Harrison DC, Davis RP, Bond BC, Campbell CA, James MF, Parsons AA, et al. Caspase mRNA expression in a rat model of focal cerebral ischemia. Brain Res Mol Brain Res. 2001;89(1–2):133–46.

    Article  PubMed  CAS  Google Scholar 

  8. 8.

    Velier JJ, Ellison JA, Kikly KK, Spera PA, Barone FC, Feuerstein GZ. Caspase-8 and caspase-3 are expressed by different populations of cortical neurons undergoing delayed cell death after focal stroke in the rat. J Neurosci. 1999;19(14):5932–41.

    Article  PubMed  CAS  Google Scholar 

  9. 9.

    Li H, Deng CQ, Chen BY, Zhang SP, Liang Y, Luo XG. Total saponins of Panax notoginseng modulate the expression of caspases and attenuate apoptosis in rats following focal cerebral ischemia-reperfusion. J Ethnopharmacol. 2009;121(3):412–8.

    Article  PubMed  CAS  Google Scholar 

  10. 10.

    Inoue S, Davis DP, Drummond JC, Cole DJ, Patel PM. The combination of isoflurane and caspase 8 inhibition results in sustained neuroprotection in rats subject to focal cerebral ischemia. Anesth Analg. 2006;102(5):1548–55.

    Article  PubMed  CAS  Google Scholar 

  11. 11.

    Shiffman ML, Pockros P, McHutchison JG, Schiff ER, Morris M, Burgess G. Clinical trial: the efficacy and safety of oral PF-03491390, a pancaspase inhibitor—a randomized placebo-controlled study in patients with chronic hepatitis C. Aliment Pharmacol Ther. 2010;31(9):969–78.

    PubMed  CAS  Google Scholar 

  12. 12.

    Barreyro FJ, Holod S, Finocchietto PV, Camino AM, Aquino JB, Avagnina A, et al. The pan-caspase inhibitor emricasan (IDN-6556) decreases liver injury and fibrosis in a murine model of non-alcoholic steatohepatitis. Liver Int. 2015;35(3):953–66.

    Article  PubMed  CAS  Google Scholar 

  13. 13.

    Rotman Y, Sanyal AJ. Current and upcoming pharmacotherapy for non-alcoholic fatty liver disease. Gut. 2017;66(1):180–90.

    Article  PubMed  CAS  Google Scholar 

  14. 14.

    Brumatti G, Ma C, Lalaoui N, Nguyen NY, Navarro M, Tanzer MC, et al. The caspase-8 inhibitor emricasan combines with the SMAC mimetic birinapant to induce necroptosis and treat acute myeloid leukemia. Sci Transl Med. 2016;8(339):339ra69.

    Article  PubMed  CAS  Google Scholar 

  15. 15.

    Baskin-Bey ES, Washburn K, Feng S, Oltersdorf T, Shapiro D, Huyghe M, et al. Clinical trial of the pan-caspase inhibitor, IDN-6556, in human liver preservation injury. Am J Transplant. 2007;7(1):218–25.

    Article  PubMed  CAS  Google Scholar 

  16. 16.

    Conrad M, Angeli JP, Vandenabeele P, Stockwell BR. Regulated necrosis: disease relevance and therapeutic opportunities. Nat Rev Drug Discov. 2016;15(5):348–66.

    Article  PubMed  CAS  Google Scholar 

  17. 17.

    Vanden Berghe T, Linkermann A, Jouan-Lanhouet S, Walczak H, Vandenabeele P. Regulated necrosis: the expanding network of non-apoptotic cell death pathways. Nat Rev Mol Cell Biol. 2014;15(2):135–47.

    Article  CAS  Google Scholar 

  18. 18.

    Moreno-Gonzalez G, Vandenabeele P, Krysko DV. Necroptosis: a novel cell death modality and its potential relevance for critical care medicine. Am J Respir Crit Care Med. 2016;194(4):415–28.

    Article  PubMed  CAS  Google Scholar 

  19. 19.

    Zille M, Karuppagounder SS, Chen Y, Gough PJ, Bertin J, Finger J, et al. Neuronal death after hemorrhagic stroke in vitro and in vivo shares features of ferroptosis and necroptosis. Stroke. 2017;48(4):1033–43.

    Article  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Yang XS, Yi TL, Zhang S, ZW X, ZQ Y, Sun HT, et al. Hypoxia-inducible factor-1 alpha is involved in RIP-induced necroptosis caused by in vitro and in vivo ischemic brain injury. Sci Rep. 2017;7(1):5818.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  21. 21.

    Zhang T, Zhang Y, Cui M, Jin L, Wang Y, Lv F, et al. CaMKII is a RIP3 substrate mediating ischemia- and oxidative stress-induced myocardial necroptosis. Nat Med. 2016;22(2):175–82.

    Article  PubMed  CAS  Google Scholar 

  22. 22.

    Fauster A, Rebsamen M, Huber KV, Bigenzahn JW, Stukalov A, Lardeau CH, et al. A cellular screen identifies ponatinib and pazopanib as inhibitors of necroptosis. Cell Death Dis. 2015;6:e1767.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  23. 23.

    SH F, Zhang HF, Yang ZB, Li TB, Liu B, Lou Z, et al. Alda-1 reduces cerebral ischemia/reperfusion injury in rat through clearance of reactive aldehydes. Naunyn Schmiedeberg's Arch Pharmacol. 2014;387(1):87–94.

    Article  CAS  Google Scholar 

  24. 24.

    He Z, Yamawaki T, Yang S, Day AL, Simpkins JW, Naritomi H. Experimental model of small deep infarcts involving the hypothalamus in rats: changes in body temperature and postural reflex. Stroke. 1999;30(12):2743–51. discussion 51

    Article  PubMed  CAS  Google Scholar 

  25. 25.

    Li F, Liu KF, Silva MD, Omae T, Sotak CH, Fenstermacher JD, et al. Transient and permanent resolution of ischemic lesions on diffusion-weighted imaging after brief periods of focal ischemia in rats: correlation with histopathology. Stroke. 2000;31(4):946–54.

    Article  PubMed  CAS  Google Scholar 

  26. 26.

    Luo CX, Zhu XJ, Zhou QG, Wang B, Wang W, Cai HH, et al. Reduced neuronal nitric oxide synthase is involved in ischemia-induced hippocampal neurogenesis by up-regulating inducible nitric oxide synthase expression. J Neurochem. 2007;103(5):1872–82.

    Article  PubMed  CAS  Google Scholar 

  27. 27.

    HY W, Tang Y, Gao LY, Sun WX, Hua Y, Yang SB, et al. The synergetic effect of edaravone and borneol in the rat model of ischemic stroke. Eur J Pharmacol. 2014;740:522–31.

    Article  CAS  Google Scholar 

  28. 28.

    Yousuf S, Atif F, Sayeed I, Tang H, Stein DG. Progesterone in transient ischemic stroke: a dose-response study. Psychopharmacology. 2014;231(17):3313–23.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  29. 29.

    Rupadevi M, Parasuraman S, Raveendran R. Protocol for middle cerebral artery occlusion by an intraluminal suture method. J Pharmacol Pharmacother. 2011;2(1):36–9.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  30. 30.

    Ye SY, Gao WY. Hydroxysafflor yellow A protects neuron against hypoxia injury and suppresses inflammatory responses following focal ischemia reperfusion in rats. Arch Pharm Res. 2008;31(8):1010–5.

    Article  PubMed  CAS  Google Scholar 

  31. 31.

    Saleem S, Ahmad M, Ahmad AS, Yousuf S, Ansari MA, Khan MB, et al. Behavioral and histologic neuroprotection of aqueous garlic extract after reversible focal cerebral ischemia. J Med Food. 2006;9(4):537–44.

    Article  PubMed  CAS  Google Scholar 

  32. 32.

    Schabitz WR, Schade H, Heiland S, Kollmar R, Bardutzky J, Henninger N, et al. Neuroprotection by hyperbaric oxygenation after experimental focal cerebral ischemia monitored by MRI. Stroke. 2004;35(5):1175–9.

    Article  PubMed  CAS  Google Scholar 

  33. 33.

    Salvesen GS, Riedl SJ. Caspase mechanisms. Adv Exp Med Biol. 2008;615:13–23.

    Article  PubMed  CAS  Google Scholar 

  34. 34.

    Mishra R, Das MK, Singh S, Sharma RS, Sharma S, Mishra V. Articulatin-D induces apoptosis via activation of caspase-8 in acute T-cell leukemia cell line. Mol Cell Biochem. 2017;426(1–2):87–99.

    Article  PubMed  CAS  Google Scholar 

  35. 35.

    Beisner DR, Ch'en IL, Kolla RV, Hoffmann A, Hedrick SM. Cutting edge: innate immunity conferred by B cells is regulated by caspase-8. J Immunol. 2005;175(6):3469–73.

    Article  PubMed  CAS  Google Scholar 

  36. 36.

    Tonnus W, Linkermann A. The in vivo evidence for regulated necrosis. Immunol Rev. 2017;277(1):128–49.

    Article  PubMed  CAS  Google Scholar 

  37. 37.

    Zhang S, Tang MB, Luo HY, Shi CH, Xu YM. Necroptosis in neurodegenerative diseases: a potential therapeutic target. Cell Death Dis. 2017;8(6):e2905.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  38. 38.

    Degterev A, Huang Z, Boyce M, Li Y, Jagtap P, Mizushima N, et al. Chemical inhibitor of nonapoptotic cell death with therapeutic potential for ischemic brain injury. Nat Chem Biol. 2005;1(2):112–9.

    Article  PubMed  CAS  Google Scholar 

  39. 39.

    Yang R, Hu K, Chen J, Zhu S, Li L, Lu H, et al. Necrostatin-1 protects hippocampal neurons against ischemia/reperfusion injury via the RIP3/DAXX signaling pathway in rats. Neurosci Lett. 2017;651:207–15.

    Article  PubMed  CAS  Google Scholar 

  40. 40.

    Sorel N, Cayssials E, Brizard F, Chomel JC. Treatment and molecular monitoring update in chronic myeloid leukemia management. Ann Biol Clin (Paris). 2017;75(2):129–45.

    Google Scholar 

  41. 41.

    Filozof C, Goldstein BJ, Williams RN, Sanyal A. Non-alcoholic steatohepatitis: limited available treatment options but promising drugs in development and recent progress towards a regulatory approval pathway. Drugs. 2015;75(12):1373–92.

    Article  PubMed  PubMed Central  Google Scholar 

  42. 42.

    Zhang J, Salminen A, Yang X, Luo Y, Wu Q, White M, et al. Effects of 31 FDA approved small-molecule kinase inhibitors on isolated rat liver mitochondria. Arch Toxicol. 2017;91(8):2921–38.

    Article  PubMed  CAS  Google Scholar 

  43. 43.

    Miller GD, Bruno BJ, Lim CS. Resistant mutations in CML and Ph(+)ALL—role of ponatinib. Biologics. 2014;8:243–54.

    PubMed  PubMed Central  CAS  Google Scholar 

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Funding

This work was supported by the National Natural Science Foundation of China, China (No. 81573430 to Xiu-Ju Luo; No. 81373409 to Jun Peng) and the Natural Science Foundation of Hunan Province, China (No. 2015JJ2156 to Xiu-Ju Luo).

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Correspondence to Xiu-Ju Luo or Jun Peng.

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Tian, J., Guo, S., Chen, H. et al. Combination of Emricasan with Ponatinib Synergistically Reduces Ischemia/Reperfusion Injury in Rat Brain Through Simultaneous Prevention of Apoptosis and Necroptosis. Transl. Stroke Res. 9, 382–392 (2018). https://doi.org/10.1007/s12975-017-0581-z

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Keywords

  • Emricasan
  • Ponatinib
  • Ischemia/reperfusion
  • Apoptosis
  • Necroptosis
  • Brain