Safety evaluation of conditionally immortalized cells for renal replacement therapy
Metrics: PDF 596 views | Full Text 1000 views | ?
Milos Mihajlovic1, Sam Hariri1, Koen C.G. Westphal1, Manoe J. Janssen1, Miriam J. Oost1, Laura Bongiovanni2, Lambertus P. van den Heuvel3, Alain de Bruin2, Luuk B. Hilbrands4 and Rosalinde Masereeuw1
1 Division of Pharmacology, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands
2 Dutch Molecular Pathology Centre, Department of Pathobiology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands
3 Department of Pediatric Nephrology, Radboud University Medical Center, Nijmegen, The Netherlands
4 Department of Nephrology, Radboud University Medical Center, Nijmegen, The Netherlands
Keywords: bioartificial kidney; cell therapy safety; conditionally immortalized proximal tubule epithelial cells; tumorigenicity; viral integration
Received: June 26, 2019 Accepted: July 29, 2019 Published: September 03, 2019
End-stage kidney disease represents irreversible kidney failure. Dialysis and transplantation, two main treatment options currently available, present various drawbacks and complications. Innovative cell-based therapies, such as a bioartificial kidney, have not reached the clinic yet, mostly due to safety and/or functional issues. Here, we assessed the safety of conditionally immortalized proximal tubule epithelial cells (ciPTECs) for bioartificial kidney application, by using in vitro assays and athymic nude rats. We demonstrate that these cells do not possess key properties of oncogenically transformed cells, including anchorage-independent growth, lack of contact inhibition and apoptosis-resistance. In late-passage cells we did observe complex chromosomal abnormalities favoring near-tetraploidy, indicating chromosomal instability. However, time-lapse imaging of ciPTEC-OAT1, confined to a 3D extracellular matrix (ECM)-based environment, revealed that the cells were largely non-invasive. Furthermore, we determined the viral integration sites of SV40 Large T antigen (SV40T), human telomerase (hTERT) and OAT1 (SLC22A6), the transgenes used for immortalization and cell function enhancement. All integrations sites were found to be located in the intronic regions of endogenous genes. Among these genes, early endosome antigen 1 (EEA1) involved in endocytosis, and BCL2 Like 1 (BCL2L1) known for its role in regulating apoptosis, were identified. Nevertheless, both gene products appeared to be functionally intact. Finally, after subcutaneous injection in athymic nude rats we show that ciPTEC-OAT1 lack tumorigenic and oncogenic effects in vivo, confirming the in vitro findings. Taken together, this study lays an important foundation towards bioartificial kidney (BAK) development by confirming the safety of the cell line intended for incorporation.
All site content, except where otherwise noted, is licensed under a Creative Commons Attribution 3.0 License.