CYP3A5 is the primary CYP3A subfamily enzyme expressed in the human kidney and its aberrant expression may contribute to a broad spectrum of renal disorders. Pharmacogenetic studies have reported inconsistent linkages between CYP3A5 expression and hypertension, however, most investigators have considered CYP3A5*1 as active and CYP3A5*3 as an inactive allele. Observations of gender specific differences in CYP3A5*3/*3 protein expression suggest additional complexity in gene regulation that may underpin an environmentally responsive role for CYP3A5 in renal function. Reconciliation of the molecular mechanism driving conditional restoration of functional CYP3A5*3 expression from alternatively spliced transcripts, and validation of a morpholino-based approach for selectively suppressing renal CYP3A5 expression, is the focus of this work. Morpholinos targeting a cryptic splice acceptor created by the CYP3A5*3 mutation in intron 3 rescued functional CYP3A5 expression in vitro, and salt-sensitive cellular mechanisms regulating splicing and conditional expression of CYP3A5*3 transcripts are reported. The potential for a G-quadruplex (G4) in intron 3 to mediate restored splicing to exon 4 in CYP3A5*3 transcripts was also investigated. Finally, a proximal tubule microphysiological system (PT-MPS) was used to evaluate the safety profile of morpholinos in proximal tubule epithelial cells, highlighting their potential as a therapeutic platform for the treatment of renal disease.
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Date of publication : 25 February 2021; Scientific Reports volume 11, Article number: 4722 (2021)
Author Information: Kevin A. Lidberg (1), Andrew J. Annalora (2), Marija Jozic (2), Daniel J. Elson (2), Lu Wang (3), Theo K. Bammler (3), Susanne Ramm (4), Maria Beatriz Monteiro (5), Jonathan Himmelfarb (6), Craig B. Marcus (2), Patrick L. Iversen (2), Edward J. Kelly (1)
(1) Department of Pharmaceutics, University of Washington, Seattle, WA, USA
(2) Department of Environmental and Molecular Toxicology, Oregon State University, Corvallis, OR, USA
(3) Department of Environmental and Occupational Health Sciences, University of Washington, Seattle, WA, USA
(4) Victorian Centre for Functional Genomics, Peter MacCallum Cancer Centre, Melbourne, Australia
(5) Depto Clinica Medica, Faculdade de Medicina FMUSP, Universidade de Sao Paulo, São Paulo, Brazil
(6) Kidney Research Institute, University of Washington, Seattle, WA, USA