The Use of Human Induced Pluripotent Stem Cells for Testing Neuroprotective Activity of Pharmacological Compounds

Abstract

Development of therapeutic preparations involves several steps, starting with the synthesis of chemical compounds and testing them in different models for selecting the most effective and safest ones to clinical trials and introduction into medical practice. Cultured animal cells (both primary and transformed) are commonly used as models for compound screening. However, cell models display a number of disadvantages, including insufficient standardization (primary cells) and disruption of cell genotypes (transformed cells). Generation of human induced pluripotent stem cells (IPSCs) offers new possibilities for the development of high-throughput test systems for screening potential therapeutic preparations with different activity spectra. Due to the capacity to differentiate into all cell types of an adult organism, IPSCs are a unique model that allows examining the activity and potential toxicity of tested compounds during the entire differentiation process in vitro. In this work, we demonstrated the efficiency of IPSCs and their neuronal derivatives for selecting substances with the neuroprotective activity using two classes of compounds — melanocortin family peptides and endocannabinoids. None of the tested compounds displayed cyto- or embryotoxicity. Both melanocortin peptides and endocannabinoids exerted neuroprotective effect in the neuronal precursors and IPSC-derived neurons subjected to hydrogen peroxide. The endo-cannabinoid N-docosahexaenoyl dopamine exhibited the highest neuroprotective effect (∼70%) in the differentiated cultures enriched with dopaminergic neurons; the effect of melanocortin Semax was ∼40%. The possibility of using other IPSC derivatives for selecting compounds with the neuroprotective activity is discussed.

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Abbreviations

alpha-MSH:

alpha-melanocyte-stimulating hormone

BDNF:

brain-derived neurotrophic factor

DA neuron:

dopaminergic neuron

DMSO:

dimethyl sulfoxide

EC:

endocannabinoid

G418:

geneticin

GDNF:

glial cell-derived neurotrophic factor

IPSC:

induced pluripotent stem cell

MC:

melanocortin

N-ADA:

N-arachidonoyl dopamine

N-DDA:

N-docosahexaenoyl dopamine

PGP:

melanocortin family peptide Pro-Gly-Pro

References

  1. 1.

    Grivennikov, I. A., Dolotov, O. V., Inozemtseva, L. S., Antonov, S. A., Kobylyanskii, A. G., and Myasoedov, N. F. (2011) The use of primary cultures of mammalian nerve and glial cells for selection of compounds with neuroprotective activity, Vestn. Biotekhnol. Fiz.-Khim. Biol. Yu. A. Ovchinnikova, 7, 24–31.

    Google Scholar 

  2. 2.

    Novosadova, E. V., Grivennikov, I. A., Bobrysheva, I. V., Grigorenko, A. P., Andreeva, L. A., Rogaeva, E. I., and Tarantul, V. Z. (2012) Semax positively affects viability of the transgenic pheochromocytoma line PC12 carrying human presenilin-1 mutant gene (hPS1), Vestn. Biotekhnol. Fiz.-Khim. Biol. Yu. A. Ovchinnikova, 8, 15–21.

    Google Scholar 

  3. 3.

    Shefer, K., Shpilmann, Kh., and Fetter, K. (2010) Drug Therapy during Pregnancy and Lactation [in Russian] (Romanov, B. K., ed.), Logosfera, Moscow.

  4. 4.

    Kalter, H. (2003) Teratology in the 20th century: environmental causes of congenital malformations in humans and how they were established, Neurotoxicol. Teratol., 25, 131–282.

    CAS  Article  Google Scholar 

  5. 5.

    Takahashi, K., Tanabe, K., Ohnuki, M., Narita, M., Ichisaka, T., Tomoda, K., and Yamanaka, S. (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors, Cell, 131, 861–872; doi: https://doi.org/10.1016/j.cell.2007.11.019.

    CAS  Article  Google Scholar 

  6. 6.

    Elitt, M. S., Barbar, L., and Tesar, P. J. (2018) Drug screening for human genetic diseases using iPSC models, Hum. Mol. Genet., 27, R89–R98; doi: https://doi.org/10.1093/hmg/ddy186.

    CAS  Article  Google Scholar 

  7. 7.

    Piccinno, M. S., Petrachi, T., Resca, E., Strusi, V., Bergamini, V., Mulas, G. A., Mari, G., Dominici, M., and Veronesi, E. (2018) Label-free toxicology screening of primary human mesenchymal cells and iPS-derived neurons, PLoS One, 13, e0201671; doi: https://doi.org/10.1371/journal.pone.0201671.

    Article  Google Scholar 

  8. 8.

    Vu, M., Li, R., Baskfield, A., Lu, B., Farkhondeh, A., Gorshkov, K., Motabar, O., Beers, J., Chen, G., Zou, J., Espejo-Mojica, A. J., Rodriguez-Lopez, A., Almeciga-Diaz, C. J., Barrera, L. A., Jiang, X., Ory, D. S., Marugan, J. J., and Zheng, W. (2018) Neural stem cells for disease modeling and evaluation of therapeutics for Tay-Sachs disease, Orphanet. J. Rare Dis., 13, 152; doi: https://doi.org/10.1186/s13023-018-0886-3.

    Article  Google Scholar 

  9. 9.

    Cota-Coronado, A., Ramirez-Rodriguez, P. B., Padilla-Camberos, E., Diaz, N. F., Flores-Fernandez, J. M., AvilaGonzalez, D., and Diaz-Martinez, N. E. (2018) Implications of human induced pluripotent stem cells in metabolic disorders: from drug discovery toward precision medicine, Drug Discov. Today, 24, 334–341; doi: https://doi.org/10.1016/j.drudis.2018.10.001.

    Article  Google Scholar 

  10. 10.

    Novosadova, E. V., Andreeva, L. A., Arsen’eva, E. L., Grivennikov, I. A., Illarioshkin, S. N., Lebedeva, O. S., Makarova, I. V., Manuilova, E. S., Myasoedov, N. F., and Tarantul, V. Z. (2016) Use of human induced pluripotent stem cells for testing of cyto- and embryotoxicity of pharmacological compounds, RF Patent 2599847 C1.

  11. 11.

    Novosadova, E. V., Andreeva, L. A., Arsen’eva, E. L., Grefenshtein, M. A., Grivennikov, I. A., Illarioshkin, S. N., Inozemtseva, L. S., Lebedeva, O. S., Manuilova, E. S., and Myasoedov, N. F. (2018) A method for testing neuroprotective activity of compounds in vitro, and the relevant testsystem, RF Patent 2646446 C1.

  12. 12.

    Novosadova, E. V., Nekrasov, E. D., Chestkov, I. V., Surdina, A. V., Vasina, E. M., Bogomazova, A. N., Manuilova, E. S., Arsenyeva, E. L., Simonova, V. V., Konovalova, E. V., Fedotova, E. Yu., Abramycheva, N. Yu., Khaspekov, L. G., Grivennikov, I. A., Tarantul, V. Z., Kiselev, S. L., and Illarioshkin, S. N. (2016) A platform for studying molecular and cellular mechanisms of Parkinson’s disease based on human induced pluripotent stem cells, Sovr. Tehnol. Med., 8, 155–164.

    Google Scholar 

  13. 13.

    Novosadova, E. V., Manuilova, E. S., Arsen’eva, E. L., Andreeva, L. A., Lebedeva, O. S., Grivennikov, I. A., and Myasoedov, N. F. (2016) Investigation of the effect of alpha-melanocyte-stimulating hormone on proliferation and early stages of differentiation of human induced pluripotent stem cells, Dokl. Biochem. Biophys., 467, 141–144; doi: https://doi.org/10.1134/S1607672916020174.

    CAS  Article  Google Scholar 

  14. 14.

    Novosadova, E. V., Arsenyeva, E. L., Manuilova, E. S., Khaspekov, L. G., Bobrov, M. Y., Bezuglov, V. V., Illarioshkin, S. N., and Grivennikov, I. A. (2017) Neuroprotective properties of endocannabinoids N-arachidonoyl dopamine and N-docosahexaenoyl dopamine examined in neuronal precursors derived from human pluripotent stem cells, Biochemistry (Moscow), 82, 1367–1372; doi: https://doi.org/10.1134/S0006297917110141.

    CAS  Article  Google Scholar 

  15. 15.

    Asmarin, I. P., Nezavibat’ko, V. N., Miasoedov, N. F., Kamenskii, A. A., Grivennikov, I. A., Ponomareva-Stepnaia, M. A., Andreeva, L. A., Kaplan, A. Ia., Koshelev, V. B., and Riasina, T V. (1997) A nootropic adrenocorticotropin analog 4-10-Semax (15-year experience in its design and study), Zh. Vyssh. Nerv. Deiat. Im. I. P. Pavlova, 47, 420–430.

    CAS  PubMed  Google Scholar 

  16. 16.

    Myasoedov, N. F., Skvortsova, V. I., Nasonov, E. L., Zhuravleva, E. Iu., Grivennikov, I. A., Arsenyeva, E. L., and Sukhanov, I. I. (1999) Investigation of mechanisms of neuroprotective effect of Semax in acute period of ischemic stroke, Zh. Nevrol. Psikhiatr. Im. S. S. Korsakova, 99, 15–19.

    Google Scholar 

  17. 17.

    Novosadova, E. V., and Grivennikov, I. A. (2014) Induced pluripotent stem cells: from derivation to application in biochemical and biomedical research, Biochemistry (Moscow), 79, 1425–1441; doi: https://doi.org/10.1134/S000629791413001X.

    CAS  Article  Google Scholar 

  18. 18.

    Cayo, M. A., Mallanna, S. K., and Di Furio, F. (2017) A drug screen using human iPSC-derived hepatocyte-like cells reveals cardiac glycosides as a potential treatment for hypercholesterolemia, Stem Cell, 20, 478–489; doi: https://doi.org/10.1016/j.stem.2017.01.011.

    CAS  Google Scholar 

  19. 19.

    Del’Alamo, J. C., Lemons, D., and Serrano, R. (2016) High throughput physiological screening of iPSC-derived cardiomyocytes for drug development, Biochim. Biophys. Acta, 1863, 1717–1727.

    Article  Google Scholar 

  20. 20.

    Malik, N., Efthymiou, A. G., Mather, K., Chester, N., Wang, X., Nath, A., Rao, M. S., and Steiner, J. P. (2014) Compounds with species and cell type specific toxicity identified in a 2000 compound drug screen of neural stem cells and rat mixed cortical neurons, Neurotoxicology, 45, 192–200; doi: https://doi.org/10.1016/j.neuro.2014.10.007.

    CAS  Article  Google Scholar 

  21. 21.

    Peng, J., Liu, Q., Rao, M. S., and Zeng, X. (2013) Using human pluripotent stem cell-derived dopaminergic neurons to evaluate candidate Parkinson’s disease therapeutic agents in MPP+ and rotenone models, J. Biomol. Screen, 18, 522–533; doi: https://doi.org/10.1177/1087057112474468.

    CAS  Article  Google Scholar 

  22. 22.

    Little, D., Ketteler, R., Gissen, P., and Devine, M. J. (2019) Using stem cell-derived neurons in drug screening for neurological diseases, Neurobiol. Aging, 78, 130–141; doi: https://doi.org/10.1016/j.neurobiolaging.2019.02.008.

    CAS  Article  Google Scholar 

  23. 23.

    Garcia-Leon, J. A., Vitorica, J., and Gutierrez, A. (2019) Use of human pluripotent stem cell-derived cells for neurodegenerative disease modeling and drug screening platform, Future Med. Chem., 11, 1305–1322; doi: https://doi.org/10.4155/fmc-2018-0520.

    CAS  Article  Google Scholar 

  24. 24.

    Desbaillets, I., Ziegler, U., Groscurth, P., and Gassmann, M. (2000) Embryoid bodies: an in vitro model of mouse embryogenesis, Exp. Physiol., 85, 645–651.

    CAS  Article  Google Scholar 

  25. 25.

    Bobrov, M. Y., Lizhin, A. A., Andrianova, E. L., Gretskaya, N. M., Frumkina, L. E., Khaspekov, L. G., and Bezuglov, V. V. (2008) Antioxidant and neuroprotective properties of N-arachidonoyl dopamine, Neurosci. Lett., 431, 6–11.

    CAS  Article  Google Scholar 

  26. 26.

    Bobrov, M. Y., Lyzhin, A. A., Andrianova, E. L., Gretskaya, N. M., Zinchenko, G. N., Frumkina, L. E., Khaspekov, L. G., and Bezuglov, V. V. (2006) Antioxidant and neuroprotective properties of N-docosahexaenoyl dopamine, Bull. Exp. Biol. Med., 142, 425–427.

    CAS  Article  Google Scholar 

  27. 27.

    Niu, N., and Wang, L. (2015) In vitro human cell line models to predict clinical response to anticancer drugs, Pharmacogenomics, 16, 273–285; doi: https://doi.org/10.2217/pgs.14.170.

    CAS  Article  Google Scholar 

  28. 28.

    Bobrov, M. Yu., Bezuglov, V. V., Khaspekov, L. G., Illarioshkin, S. N., Novosadova, E. V., and Grivennikov, I. A. (2017) Expression of type I cannabinoid receptors at different stages of neuronal differentiation of human fibroblasts, Bull. Exp. Biol. Med., 163, 272–275; doi: https://doi.org/10.1007/s10517-017-3782-2.

    CAS  Article  Google Scholar 

  29. 29.

    Vendel, E., and de Lange, E. C. (2014) Functions of the CB1 and CB2 receptors in neuroprotection at the level of the blood-brain barrier, Neuromol. Med., 16, 620–642; doi: https://doi.org/10.1007/s12017-014-8314-x.

    CAS  Article  Google Scholar 

  30. 30.

    Hauser, D. N., and Cookson, M. R. (2011) Astrocytes in Parkinson’s disease and DJ-1, J. Neurochem., 117, 357–358.

    CAS  Article  Google Scholar 

  31. 31.

    Krencik, R., and Ullian, E. M. (2013) A cellular star atlas: using astrocytes from human pluripotent stem cells for disease studies, Front. Cell. Neurosci., 7, 25; doi: https://doi.org/10.3389/fncel.2013.00025.

    CAS  Article  Google Scholar 

  32. 32.

    Di Domenico, A., Carola, G., Calatayud, C., Pons-Espinal, M., Munoz, J. P., Richaud-Patin, Y., Fernandez-Carasa, I., Gut, M., Faella, A., Parameswaran, J., Soriano, J., Ferrer, I., Tolosa, E., Zorzano, A., Cuervo, A. M., Raya, A., and Consiglio, A. (2019) Patient-specific iPSC-derived astrocytes contribute to non-cell-autonomous neurodegeneration in Parkinson’s disease, Stem Cell Rep., 12, 213–229; doi: https://doi.org/10.1016/j.stemcr.2018.12.011.

    CAS  Article  Google Scholar 

  33. 33.

    Lancaster, M. A., Renner, M., and Martin, C.-A. (2013) Cerebral organoids model human brain development and microcephaly, Nature, 501, 373–379; doi: https://doi.org/10.1038/nature12517.

    CAS  Article  Google Scholar 

  34. 34.

    Amin, N. D., and Pas, S. P. (2018) Building models of brain disorders with three-dimensional organoids, Neuron, 100, 389–405; doi: https://doi.org/10.1016/j.neuron.2018.10.007.

    CAS  Article  Google Scholar 

  35. 35.

    Bordoni, M., Rey, F., Fantini, V., Pansarasa, O., Di Giulio, A. M., Carelli, S., and Cereda, C. (2018) From neuronal differentiation of iPSCs to 3D neuro-organoids: modelling and therapy of neurodegenerative diseases, Int. J. Mol. Sci., 19, 3972; doi: https://doi.org/10.3390/ijms19123972.

    Article  Google Scholar 

  36. 36.

    Eremeev, A. V., Volovikov, E. A., Shuvalova, L. D., Davidenko, A. V., Khomyakova, E. A., Bogomiakova, M. E., Lebedeva, O. S., Zubkova, O. A., and Lagarkova, M. A. (2019) Necessity is the mother of invention or inexpensive, reliable, and reproducible protocol for generating organoids, Biochemistry (Moscow), 84, 321–328; doi: https://doi.org/10.1134/S0006297919030143.

    CAS  Article  Google Scholar 

  37. 37.

    McArdle, P., Engberg, S., Bennett, N., Blackett, C., and Wigglesworth, M. (2017) Enabling 1536-well high-throughput cell-based screening through the application of novel centrifugal plate washing, SLAS Discov., 22, 732–742; doi: https://doi.org/10.1177/2472555216683650.

    PubMed  Google Scholar 

  38. 38.

    Sherman, S. P., and Bang, A. G. (2018) High-throughput screen for compounds that modulate neurite growth of human induced pluripotent stem cell-derived neurons, Dis. Model Mech., 11, No. 2, dmm031906; doi: https://doi.org/10.1242/dmm.031906.

    Article  Google Scholar 

  39. 39.

    Han, Y., Miller, A., Mangada, J., Liu, Y., and Swistowski, A. (2009) Identification by automated screening of a small molecule that selectively eliminates neural stem cells derived from hESCs but not dopamine neurons, PLoS One, 4, e7155; doi: https://doi.org/10.1371/journal.pone.0007155.

    Article  Google Scholar 

  40. 40.

    Wang, C., Ward, M. E., Chen, R., Liu, K., Tracy, T. E., Chen, X., Xie, M., Sohn, P. D., Ludwig, C., Meyer-Franke, A., Karch, C. M., Ding, S., and Li, G. (2017) Scalable production of iPSC-derived human neurons to identify taulowering compounds by high-content screening, Stem Cell Rep., 9, 1221–1233; doi: https://doi.org/10.1016/j.stemcr.2017.08.019.

    CAS  Article  Google Scholar 

  41. 41.

    Traub, S., and Heilker, R. (2019) hiPS cell-derived neurons for high-throughput screening, Methods Mol. Biol., 1994, 243–263; doi: https://doi.org/10.1007/978-1-4939-9477-9_23.

    Article  Google Scholar 

  42. 42.

    Little, D., Luft, C., Pezzini-Picart, O., Mosaku, O., Ketteler, R., Devine, M. J., and Gissen, P. (2019) Seeding induced pluripotent stem cell-derived neurons onto 384-well plates, Methods Mol. Biol., 1994, 159–164; doi: https://doi.org/10.1007/978-1-4939-9477-9_14.

    Article  Google Scholar 

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Acknowledgements

The authors express gratitude to V. V. Bezuglov (Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences) for kindly providing endocannabinoids for this study.

Funding

This work was supported by the Russian Foundation for Basic Research (project 17-04-01661a) and the Program of the Presidium of the Russian Academy of Sciences “Fundamental studies for biomedical technologies”. The equipment used in the study was provided by the Center of Collective Use of the Institute of Molecular Genetics, Russian Academy of Sciences (Center for Cell and Gene Technologies).

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Correspondence to E. V. Novosadova or I. A. Grivennikov.

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This article does not contain any studies with human participants or animals performed by any of the authors.

Russian Text © The Author(s), 2019, published in Biokhimiya, 2019, Vol. 84, No. 11, pp. 1610–1621.

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Novosadova, E.V., Arsenyeva, E.L., Antonov, S.A. et al. The Use of Human Induced Pluripotent Stem Cells for Testing Neuroprotective Activity of Pharmacological Compounds. Biochemistry Moscow 84, 1296–1305 (2019). https://doi.org/10.1134/S0006297919110075

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Keywords

  • test-system
  • embryotoxicity
  • neuroprotection
  • induced pluripotent stem cells
  • oxidative stress