AIM: Clinical studies have demonstrated the importance of genetic variation for tacrolimus disposition and suggested a role in the development of renal fibrosis associated with long-term tacrolimus treatment. Our aim was to explore the implications of tacrolimus exposure in a model of human proximal tubule cells incorporating genetic variation in CYP3A5 and ABCB1 on the expression of profibrotic cytokine CTGF and correlate these findings with CTGF expression in kidney allograft biopsies.

METHODS: We selected 8 clones of human conditional immortalized PTC (ciPTC) with 4 different combinations of CYP3A5 (rs776746) and ABCB1 (rs1045642) genotypes. Cells were incubated with vehicle, 50 ng/ml and 300 ng/ml tacrolimus (=tissue concentration range in allografts). Quantitative RT-PCR and western blot were performed to study CTGF expression. In addition, CTGF staining was performed on protocol biopsies with a known pharmacogenetic background derived from 17 allograft recipients over a period of 2 years.

RESULTS: CTGF mRNA and protein expression increased with tacrolimus concentration (CTGF vs. β-actin vs. vehicle at 50ng/ml: + 34.1% and at 300 ng/ml: +45.0%; p<0.001). Subgroup analysis demonstrated 46% higher CTGF protein expression in CYP3A5 *3/*3 allele carriers vs. *1 allele carriers (p=0.047) and more than 2-fold higher CTGF expression in ABCB1 3435 TTs, while in the genetic CC/CT counterparts CTGF expression decreased (p=0.01). Immunohistochemical studies of protocol biopsies demonstrated a 38.3% increase in tubular cell CTGF staining between 3 to 24 months in kidneys from 3435 TT genotype donors, while in CC/CT donor grafts the percentage of CTGF positive tubuli remained stable (p=0.046).

CONCLUSIONS: Tacrolimus exposure in human PTCs results in a concentration-dependent increase in CTGF expression. Tacrolimus exposure for 72 hours resulted in increased CTGF expression in PTC derived from CYP3A5 *3/*3 allele carriers, and in particular with the ABCB1 3435TT genotype. Immunohistochemical studies on protocol biopsies confirm increasing CTGF expression over time in donor kidneys with the ABCB1 3435TT genotype.

CITATION INFORMATION: Knops N., Ramazani Y., Van den Heuvel B., Goldschmeding R., Nguyen T., Levtchenko E., Kuypers D. Tacrolimus Exposure in Human Proximal Tubule Cells Results in Differentially Increased CTGF Expression in Relation to Pharmacogenetic Variants of CYP3A5 and ABCB1 Am J Transplant. 2017;17 (suppl 3).

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