Development and Application of Human Renal Proximal Tubule Epithelial Cells for Assessment of Compound Toxicity

Abstract

Kidney toxicity is a major problem both in drug development and clinical settings. It is difficult to predict nephrotoxicity in part because of the lack of appropriate in vitro cell models, limited endpoints, and the observation that the activity of membrane transporters which plays important roles in nephrotoxicity by affecting the pharmacokinetic profile of drugs is often not taken into account. We developed a new cell model using pseudo-immortalized human primary renal proximal tubule epithelial cells. This cell line (SA7K) was characterized by the presence of proximal tubule cell markers as well as several functional properties, including transporter activity and response to a few well-characterized nephrotoxicants. We subsequently evaluated a group of potential nephrotoxic compounds in SA7K cells and compared them to a commonly used human immortalized kidney cell line (HK-2). Cells were treated with test compounds and three endpoints were analyzed, including cell viability, apoptosis and mitochondrial membrane potential. The results showed that most of the known nephrotoxic compounds could be detected in one or more of these endpoints. There were sensitivity differences in response to several of the chemicals between HK-2 and SA7K cells, which may relate to differences in expressions of key transporters or other components of nephrotoxicity pathways. Our data suggest that SA7K cells appear as promising for the early detection of renal toxicants.

Implementation of a Human Renal Proximal Tubule on a Chip for Nephrotoxicity and Drug Interaction Studies

“…This explains the lack of sensitivity of RPTEC towards tenofovir, as this substance requires the OAT1 transporter for cellular uptake. 18,24 Epithelial barrier function is also a key functional parameter, as a leaky epithelium is often an indicator of impaired kidney function, previously demonstrated in this model. 15 Regarding biochemical parameters, cell viability, LDH release and gene expression of toxicity markers were determined on both PTEC lines to assess the effect of the compounds.…”

“…22 In this study, we implemented wellcharacterized PTEC lines, considered relevant for nephrotoxicity and drug-transporter interaction studies, as potential alternatives to animal experimentation. 14e18, 23,24 Supply and reproducibility (low batch-to-batch variability) of these commercially available cells are guaranteed. The PTECs were cultured in the OrganoPlate, generating a proximal tubule-on-a-chip consisting of 40 chips on a 384-well microtiter plate format.…”

“…Membrane drug transporters are an important characteristic of PTECs and expression and activity of transporters have been demonstrated previously. 6,17,18,24 Differences in response towards toxicant exposures between ciPTEC-OAT1 and RPTEC could be explained by different expression levels of drug transporters. For instance, ciPTEC-OAT1, but not RPTEC, functionally express OAT1.…”

Review of high-content screening applications in toxicology

“…Acute renal failure can result from exposure to a variety of drugs, natural products, and industrial and environmental chemicals. Several in vitro cell models have been used for nephrotoxicity evaluations including human embryonic kidney 293 (HEK293), porcine kidney (LLC-PK1), human kidney-2 (HK-2), hTERT immortalized human renal proximal tubular epithelial cell line (RPTEC/TERT1), modified human primary renal proximal tubule epithelial cell (SA7K), and human iPSC-derived renal cells [90][91][92][93][94]. The renal cell types differentiated from pluripotent stem cells, and microfluidic and 3D culture systems are more physiologically relevant, improving the ability to identify and characterize the nephrotoxicants [95].…”

Emerging Kidney Models to Investigate Metabolism, Transport, and Toxicity of Drugs and Xenobiotics

“…In 1994, human renal cortex-derived cells were transfected with the human papilloma virus 16 E6/E7 genes to create the immortalized human kidney (HK-2) cell line (Ryan et al, 1994). HK-2 cells have since been used to show that P-gp is suppressed by vancomycin (Im et al, 2017), to elucidate the biotransformation and toxicity of acyclovir (Gunness et al, 2011), to assess in vitro biomarkers of cisplatin nephrotoxicity (Sohn et al, 2013), and to evaluate general nephrotoxicity of compounds (Wu et al, 2009;Li et al, 2017). P-gp (Tramonti et al, 2001) and monocarboxylate transporter (MCT) (Wang et al, 2006) have been fully characterized in HK-2 cells; however, an extensive genetic analysis of HK-2 cells revealed that OAT1, OAT2, OAT3, OCT2, MRP2, and BCRP were not expressed by HK-2 cells, bringing into question their overall utility in assessing renal transporter function or transporter-related toxicities (Jenkinson et al, 2012).…”