The Isoquinolone Derived Prolyl Hydroxylase Inhibitor ICA Is a Potent Substrate of the Organic Anion Transporters 1 and 3

Abstract

Objective: Many cellular responses to hypoxia are mediated by the transcription factor complex hypoxia-inducible factor (HIF). HIF stability is governed by a family of dioxygenases called HIF prolyl hydroxylases (PHDs). Isoquinolone-derived PHD inhibitors, like 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate (ICA), which stabilize the intracellular HIF-α have been suggested as a potentially beneficial therapeutic strategy for the treatment of disorders associated with ischemia. To stabilize HIF-α, ICA has to be taken up into proximal tubule cells (PCTs) across the basolateral membrane by one of the organic anion transporters 1, 2 or 3 (OAT1, OAT2 or OAT3). The release into the urine across the luminal membrane may be mediated by OAT4. Method: To demonstrate interaction of ICA with human OAT1, OAT2, OAT3 and OAT4, ICA was tested on these transporters stably transfected in HEK293 cells by using p-aminohippurate (PAH), cGMP and estrone-3-sulfate (ES) as reference substrates, respectively. Results: Uptakes of PAH and ES in OAT1- and OAT3-transfected HEK293 cells were inhibited by ICA with half-maximal inhibition values of 0.29 ± 0.05 and 2.58 ± 0.16 µ<smlcap>M</smlcap>, respectively. OAT2 was less sensitive to ICA. Efflux experiments identified ICA as an OAT1 and OAT3 substrate. Preloading OAT4-transfected HEK293 cells with ICA stimulated ES uptake by 18.3 ± 3.8%. Conclusion: The uptake of ICA across the basolateral membrane of PCTs occurs mainly by OAT1 and the efflux into the tubular lumen by OAT4.

A Fluorescent Benzo[g]isoquinoline‐Based HIF Prolyl Hydroxylase Inhibitor for Cellular Imaging

“…[33] Thus, no such transporter for cellular uptake is needed, hence BCA uptake likely occurs via passive diffusiond ue to its lipophilicity.A ss hown in previous studies, lipophilic 2OG oxygenase inhibitors such as ICA do not rely on the uptake via at ransporter. [13,34] In contrast, hydrophilic inhibitors including 2,4-pyridinedicarboxylic acid (PDCA)a re transferred via the organic anion transporter (OATs). [13,34] BCA was injected into mice 2.5 hb efore kidney removal to test for HIF-1a stabilization in vivo.…”

“…[13,34] In contrast, hydrophilic inhibitors including 2,4-pyridinedicarboxylic acid (PDCA)a re transferred via the organic anion transporter (OATs). [13,34] BCA was injected into mice 2.5 hb efore kidney removal to test for HIF-1a stabilization in vivo. Strong nuclear HIF-1a signals were detectedi ne pithelial cells of all tubulars egments, while con-trol kidneys showedn uclear HIF-1a positivity only in isolated tubular cells of the distal nephron ( Figure 9).…”

Untersuchung der Transportvorgänge des Prolyl-Hydroxylase-Hemmers ICA an den Transportern OAT1, OAT2, OAT3 und OAT4 von proximalen Nierentubuluszellen

“…Bei Hemmung der PHD wird der Hypoxia Inducible Factor (HIF) hochreguliert, sodass in den Sauerstoffhaushalt des Körpers eingegriffen werden kann. Die Ergebnisse dieser Arbeit konnten veröffentlicht werden(Schulz et al 2015).Zunächst werden die grundlegendenFunktionsweisen der PHD und des HIF unter physiologischen Bedingungen im Körper im ersten Unterkapitel erläutert, um dann im zweiten Unterkapitel der Einleitung auf die Möglichkeiten einer Einflussnahme auf dieses System mit Hemmstoffen und im Speziellen mit ICA einzugehen, um zuletzt die OATs im Allgemeinen und die vier in dieser Arbeit untersuchten auf ihre möglichen Wechselwirkungen mit ICA zu diskutieren. Anschließend wird die Fragestellung dieser Arbeit formuliert.…”