Date: Sunday, May 3, 2015
Session Time: 5:30pm-6:30pm
Presentation Time: 5:30pm-6:30pm
Location: Exhibit Hall E
Background: Multiple studies have demonstrated the importance of genetic variation in CYP3A5 and ABCB1 for calcineurin inhibitor (CNI) disposition and CNI-related nephrotoxicity. Renal CNI metabolism occurs in the proximal tubule cells (PTC). The present study was designed to study the functional implications of variation in CYP3A5 and ABCB1 in human PTC exposed to CNIs .
Methods: A technique was developed to obtain immortalized PTC clones from a routine renal allograft biopsy. Cells were characterized for CYP3A5(rs776746) /ABCB1(rs1045642) genotype. We selected specific sets of PTCs according to different combinations of aforementioned genetic variants. Quantitative RT-PCR, WB and immunohistochemistry were performed for gene expression. CYP3A5 activity was assessed by differential midazolam (MDZ) hydroxylation and Pgp (ABCB1 product) activity by calcein efflux assay. Drug metabolism was assessed by tacrolimus (Tac) disappearance. Cytokine production by RT-PCR.
Results: 11 PTC clones from different donor-recipient pairs were selected. CYP3A5*1 carriers had increased 1-OH/4-OH MDZ formation vs. *3/*3 carriers ((mean: 2,36 (95%CI: 1,11-3,40) vs. 0,88 (95%CI:0,48-1,27); p<0,05). Pgp activity was confirmed by calcein accumulation (mean 38,6%; 95% CI: 32,8- 44,4%), and was higher in cell lines with the ABCB1 3435TT than in the CC/CT genotype (46,2% versus 35,5%; 95% CI: 28,7-42,2%). Tac disappearance was circa 2-fold higher in A5*1 allele carriers and was highest in the combined A5*1/ 3435TT. Increasing "physiological" concentrations of Tac caused increasing cytokines expression (hypoxia, ox. stress, EMT, fibrosis), in particular for A5*1/ 3435TT.
Conclusion: Immortalized human PTC cell lines demonstrate functional expression of genes involved in CNI metabolism. Differences in protein function, Tac metabolism and cytokine expression were detected according to genetic variants in CYP3A5 and ABCB1. This in vitro model allows studying the role of human pharmacogenetic variations in renal drug disposition and toxicity.
To cite this abstract in AMA style:Knops N, Heuvel Lvanden, Masereeuw R, Loor Hde, Levtchenko E, Kuypers D. Pharmacogenetic Variation in Renal Calcineurin Inhibitor (CNI) Metabolism and the Response of Allograft-Derived Proximal Tubule Cells [abstract]. Am J Transplant. 2015; 15 (suppl 3). https://atcmeetingabstracts.com/abstract/pharmacogenetic-variation-in-renal-calcineurin-inhibitor-cni-metabolism-and-the-response-of-allograft-derived-proximal-tubule-cells/. Accessed August 14, 2020.
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