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.

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