Experimental Cell Line Models for Nephrotoxicity Screening

The aim of the study was to review literature data on cell models for experimental assessment of drug nephrotoxicity in vitro. Because of nephrotoxicity, 2% of new investigational medicinal products are discarded at the stage of preclinical in vivo studies in laboratory animals, and 19%—after phase 3 clinical trials. Prediction of toxicity in cell models could make drug development more cost-effective and help to reduce/avoid animal testing. At present, there are no official international guidelines for assessment of nephrotoxicity in vitro, but there is a lot of research underway. The main toxicity target in kidneys is renal proximal tubule epithelial cells, therefore the main research is focused on the development of renal proximal tubule epithelial cell lines with stable functional characteristics. Another important aspect in nephrotoxicity modeling is the choice of relevant test methods and end points which would reflect potential toxicity mechanisms. The paper reviews existing human renal proximal tubule epithelial cell lines and current test methods for assessing cytotoxicity. Promising areas for future development of cell models for nephrotoxicity assessment— are optimisation and standardisation of in vitro systems that would help to make preclinical predictions of drug nephrotoxicity in vivo.  

1. Redfern WS, Ewart L, Hammond TG, Bialecki R, Kinter L, Lindgren S, et al. Impact and frequency of different toxicities throughout the pharmaceutical life cycle. Toxicologist. 2010;114(S1):1081.

2. Zou L, Stecula A, Gupta A, Prasad B, Chien HC, Yee SW, et al. Molecular mechanisms for species differences in organic anion transporter 1, OAT1: implications for renal drug toxicity. Mol Pharmacol. 2018;94(1):689–99. https://doi.org/10.1124/mol.117.111153

3. Steimberg N, Bertero A, Chiono V, Dell’Era P, Di Angelantonio S, Hartung T, et al. iPS, organoids and 3D models as advanced tools for in vitro toxicology. ALTEX. 2020;37(1):136–40. https://doi.org/10.14573/altex.1911071

4. Nigam SK, Wu W, Bush KT, Hoenig MP, Blantz RC, Bhatnagar V. Handling of drugs, metabolites, and uremic toxins by kidney proximal tubule drug transporters. Clin J Am Soc Nephrol. 2015:10(11):2039–49. https://doi.org/10.2215/CJN.02440314

5. Giacomini KM, Huang SM, Tweedie DJ, Benet LZ, Brouwer KL, Chu X, et al. Membrane transporters in drug development. Nat Rev Drug Discov. 2010;9(3):215–36. https://doi.org/10.1038/nrd3028

6. Eshbach ML, Weisz OA. Receptor-mediated endocytosis in the proximal tubule. Annu Rev Physiol. 2017;79(1):425–48. https://doi.org/10.1146/annurev-physiol-022516-034234

7. Bhargava P, Schnellmann RG. Mitochondrial energetics in the kidney. Nat Rev Nephrol. 2017;13(10):629–46. https://doi.org/10.1038/nrneph.2017.107

8. Linkermann A, Chen G, Dong G, Kunzendorf U, Krautwald S, Dong Z. Regulated cell death in AKI. J Am Soc Nephrol. 2014;25(12):2689–701. https://doi.org/10.1681/ASN.2014030262

9. Li Q, Guo D, Dong Z, Zhang W, Zhang L, Huang SM, et al. Ondansetron can enhance cisplatin-induced nephrotoxicity via inhibition of multiple toxin and extrusion proteins (MATEs). Toxicol Appl Pharmacol. 2013;273(1):100–9. https://doi.org/10.1016/j.taap.2013.08.024

10. Zhou Y, Yang Y, Wang P, Wei M, Ma Y, Wu X. Adefovir accumulation and nephrotoxicity in renal interstitium: Role of organic anion transporters of kidney. Life Sci. 2019;224:41–50. https://doi.org/10.1016/j.lfs.2019.03.042

11. Lin Z, Will Y. Evaluation of drugs with specific organ toxicities in organ-specific cell lines. Toxicol Sci. 2012;126(1):114–27. https://doi.org/10.1093/toxsci/kfr339

12. Li Y, Kandasamy K, Chuah JK, Lam YN, Toh WS, Oo ZY, Zink D. Identification of nephrotoxic compounds with embryonic stem-cell-derived human renal proximal tubular-like cells. Mol Pharm. 2014;11(7):1982–90. https://doi.org/10.1021/mp400637s

13. Narayanan K; Schumacher K, Tasnim F, Kandasamy K, Schumacher A, Ni M et al. Human embryonic stem cells differentiate into functional renal proximal tubular-like cells. Kidney Int. 2013;83(4):593−603. https://doi.org/10.1038/ki.2012.442

14. Shaw G, Morse S, Ararat M, Graham FL. Preferential transformation of human neuronal cells by human adenoviruses and the origin of HEK 293 cells. FASEB J. 2002;16(8):869−71. https://doi.org/10.1096/fj.01-0995fje

15. Ryan MJ, Johnson G, Kirk J, Fuerstenberg SM, Zager RA, Torok-Storb B. HK-2: an immortalized proximal tubule epithelial cell line from normal adult human kidney. Kidney Int. 1994;45(1):48–57. https://doi.org/10.1038/ki.1994.6

16. Jenkinson SE, Chung GW, van Loon E, Bakar NS, Dalzell AM, Brown CDA. The limitations of renal epithelial cell line HK-2 as a model of drug transporter expression and function in the proximal tubule. Pflugers Arch. 2012;464(6):601–11. https://doi.org/10.1007/s00424-012-1163-2

17. Wu Y, Connors D, Barber L, Jayachandra S, Hanumegowda UM, Adams SP. Multiplexed assay panel of cytotoxicity in HK-2 cells for detection of renal proximal tubule injury potential of compounds. Toxicol In Vitro. 2009;23(6):1170–8. https://doi.org/10.1016/j.tiv.2009.06.003

18. Li Y, Oo ZY, Chang SY, Huang P, Eng KG, Zeng JL, et al. An in vitro method for the prediction of renal proximal tubular toxicity in humans. Toxicol Res. 2013;2(5):352–65. https://doi.org/10.1039/c3tx50042j

19. Lash LH, Putt DA, Cai H. Drug metabolism enzyme expression and activity in primary cultures of human proximal tubular cells. Toxicology. 2008;244(1):56–65. https://doi.org/10.1016/j.tox.2007.10.022

20. Wieser M, Stadler G, Jennings P, Streubel B, Pfaller W, Ambros P, et al. hTERT alone immortalizes epithelial cells of renal proximal tubules without changing their functional characteristics. Am J Physiol Renal Physiol. 2008;295(5):F1365–75. https://doi.org/10.1152/ajprenal.90405.2008

21. Wilmer MJ, Saleem MA, Masereeuw R, Ni L, van der Velden TJ, Russel FG, et al. Novel conditionally immortalized human proximal tubule cell line expressing functional influx and efflux transporters. Cell Tissue Res. 2010;339(2):449–57. https://doi.org/10.1007/s00441-009-0882-y

22. Nieskens T, Peters J, Schreurs M, Smits N, Woestenenk R, Jansen K, et al. A human renal proximal tubule cell line with stable organic anion transporter 1 and 3 expression predictive for antiviral-induced toxicity. AAPS J. 2016;18(2):465–75. https://doi.org/10.1208/s12248-016-9871-8

23. Li S, Zhao J, Huang R, Steiner T, Bourner M, Mitchell M, et al. Development and application of human renal proximal tubule epithelial cells for assessment of compound toxicity. Curr Chem Genom Transl Med. 2017;11:19–30. https://doi.org/10.2174/2213988501711010019

24. Birdsall H, Hammond T. Role of shear stress on renal proximal tubular cells for nephrotoxicity assays. J Toxicol. 2021;2021:6643324. https://doi.org/10.1155/2021/6643324

25. Secker PF, Luks L, Schlichenmaier N, Dietrich DR. RPTEC/TERT1 cells form highly differentiated tubules when cultured in a 3D matrix. ALTEX. 2018;35(2):223–34. https://doi.org/10.14573/altex.1710181

26. Bernal-Barquero CE, Vazquez-Zapien GJ, Mata-Miranda MM. Revisión de las alteraciones en la expresión génica y vías apoptóticas provocadas en la nefrotoxicidad inducida por cisplatino. Nefrologia. 2019;39:362–71. https://doi.org/10.1016/j.nefro.2018.11.012

27. Stoddart MJ. Cell viability assays: introduction. Methods Mol Biol. 2011;740:1–6. https://doi.org/10.1007/978-1-61779-108-6_1

28. Trusov GA, Chaplenko AA, Semenova IS, Melnikova EV, Olefir YuV. Use of flow cytometry for quality evaluation of biomedical cell products. BIOpreparations. Prevention, Diagnosis, Treatment. 2018;18(1):16–24 (In Russ). https://doi.org/10.30895/2221-996X-2018-18-1-16-24

29. Camaño S, Lazaro A, Moreno-Gordaliza E, Torre AM, de Lucas C, Humanes B, et al. Cilastatin attenuates cisplatin-induced proximal tubular cell damage. J Pharmacol Exp Ther. 2010;334(2):419–29. https://doi.org/10.1124/jpet.110.165779

30. Chen S, Einspanier R, Schoen J. Transepithelial electrical resistance (TEER): a functional parameter to monitor the quality of oviduct epithelial cells cultured on filter supports. Histochem Cell Biol. 2015;144(5):509–15. https://doi.org/10.1007/s00418-015-1351-1

31. Ke N, Wang X, Xu X, Abassi YA. The xCELLigence system for real-time and label-free monitoring of cell viability. Methods Mol Biol. 2011;740:33–43. https://doi.org/10.1007/978-1-61779-108-6_6

32. Sakamuru S, Attene-Ramos MS, Xia M. Mitochondrial membrane potential assay. Methods Mol Biol. 2016;1473:17–22. https://doi.org/10.1007/978-1-4939-6346-1_2

33. Limonciel A, Aschauer L, Wilmes A, Prajczer S, Leonard MO, Pfaller W, et al. Lactate is an ideal non-invasive marker for evaluating temporal alterations in cell stress and toxicity in repeat dose testing regimes. Toxicol In Vitro. 2011;25(8):1855–62. https://doi.org/10.1016/j.tiv.2011.05.018

34. Sies H., Jones D.P. Reactive oxygen species (ROS) as pleiotropic physiological signalling agents. Nat Rev Mol Cell Biol 2020;21(7):363–83. https://doi.org/10.1038/s41580-020-0230-3

35. Aschauer L, Limonciel A, Wilmes A, Stanzel S, Kopp-Schneider A, Hewitt P, et al. Application of RPTEC/TERT1 cells for investigation of repeat dose nephrotoxicity: A transcriptomic study. Toxicol In Vitro. 2015;30(1 Pt A):106–16. https://doi.org/10.1016/j.tiv.2014.10.005

36. Dieterle F, Sistare F, Goodsaid F, Papaluca M, Ozer JS, Webb CP, et al. Renal biomarker qualification submission: a dialog between the FDA-EMEA and Predictive Safety Testing Consortium. Nat Biotechnol. 2010;28(5):455–62. https://doi.org/10.1038/nbt.1625

37. Qiu X, Miao Y, Geng X, Zhou X, Li B. Evaluation of biomarkers for in vitro prediction of drug-induced nephrotoxicity in RPTEC/TERT1 cells. Toxicol Res (Camb). 2020;9(2):91–100. https://doi.org/10.1093/toxres/tfaa005