Characterization of in Vivo Disulfide-Reduction Mediated Drug Release in Mouse Kidneys

Abstract

Due to the overexpression of a folate receptor (FR) on many malignant cells, folate-targeted drugs have been developed to improve the cancer specificity of chemotherapeutic agents. Therapeutic index is further enhanced with the use of self-immolative linkers that efficiently release the attached drug upon cellular internalization of the folate-drug conjugate. Because FR is also abundant in normal kidney proximal tubule (PT) cells, we sought to examine in real time the trafficking and release of folate-targeted drugs in the kidney in vivo. Thus, we conducted two-photon kidney imaging studies in mice utilizing a Förster resonance energy transfer (FRET) based folate conjugate that undergoes a color shift from red to green upon reduction of the disulfide bond linking folate to a surrogate drug molecule. Following infusion via intravenous injection, folate-FRET reached the kidney in its intact unreduced form. The folate-FRET conjugate was then filtered into the lumen of PT, where it was efficiently captured by FR. As FR transcytosed across PT, some disulfide reduction occurred, with reduced folate-FRET detectable in PT vesicles 30 min postinjection. Prolonged monitoring of folate-FRET in mice showed modest progression of reduction in PT cells over time. Moreover, inhibition of FR trafficking in PT cells by colchicine did not significantly affect the rate or extent of folate-FRET reduction. Finally, the lack of cytosolic accumulation of released drug surrogate in the PT suggests that drug release via disulfide bond reduction should cause little kidney toxicity.

Folate Receptor-β in Activated Macrophages: Ligand Binding and Receptor Recycling Kinetics

“…Except for the kidneys (known to express very high levels of FR-α), accumulation of folate conjugates in all other tissues was negligible (data not shown).…”

A bioinspired peptide scaffold with high antibiotic activity and low in vivo toxicity

“…Moreover, accumulation of PxB in kidneys could be correlated to its nephrotoxicity potential 33 34 . On the other hand, if this is the case with our novel PxB analogs, eventual uptake by renal cells would presumably reduce the disulfide and open up the peptide cycle due to the reducing intracellular environment (reduced glutathione and oxidorreductases) and thus facilitate the proteolysis of the linear sequence 35 36 37 . Hence, the disulfide link may provide PxB analogs with sufficient stability to reach the infectious target in vivo , whereas upon eventual accumulation and uptake by renal cells, reduction and subsequent decyclization would facilitate peptide proteolysis and potentially lower renal toxicity.…”

B cell–derived IL-4 acts on podocytes to induce proteinuria and foot process effacement

“…The preparation of kidney intravital 2-photon microscopy was adapted from previously described approaches (90)(91)(92). Briefly, 8-to 10-week-old mice previously given multimerized HEL were anesthetized with isofluorane and a small vertical incision was made in the flank of the mouse.…”