Development of a novel predictive in vitro non human primate proximal tubule model for drug transporter & nephrotoxicity studies

Nephrotoxicity is a major reason for drugs failing during clinical development. Currently there is no in vitro platform that enables crossspecies comparisons of drug transport or nephrotoxicity. Our innovative solution to this is to develop of an assay platform measuring both drug transport and a range of biomarkers associated with drug induced kidney injury using primary renal proximal tubule cells (PTCs) derived from key animal species.

To generate an Non human Primate (NHP) PTC platform, PTCs were isolated from kidney cortex using a collagenase digest and Percoll density gradient protocol. NHP PTCs were seeded onto Transwell filters and grown to confluency in a chemically defined media for 7 days. Gene expression was measured by qPCR. Digoxin flux was measured using [3H] digoxin. Biomarkers were assessed using an NHP specific clusterin ELISA assay. Monolayer integrity was measured as transepithelial electrical resistance (TEER) and cell viability using MTS. NHP PTCs grown on permeable filter supports formed confluent monolayers and expressed a range of key drug transporters including MDR1, MRP4, BCRP, OCT2, OATP4C1 and lower levels OAT1. Functionally, we could demonstrate a net digoxin secretion across NHP monolayers. To test the utility of NHP PTC monolayers as a predictive models of nephrotoxicity, cells were challenged with a range of well characterised nephrotoxins and the impact upon the biomarker clusterin, TEER and cell viability was assessed. In response to cisplatin challenge (10mM) for 72 hours, clusterin release in to the apical media was substantially increased. Cisplatin challenge was also accompanied by both a significant decrease in cell viability and of TEER value.

Taken together these results report for the first time the successful production of a primary NHP PTC monolayer which, similar to our human and rat aProximate™ PTC models may prove useful as a predictive in vitro PTC model of drug transport and drug toxicity.

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