Aortic Binding of AZD5248: Mechanistic Insight and Reactivity Assays To Support Lead Optimzation

Abstract

The oral dipeptidyl peptidase 1 (DPP1) inhibitor AZD5248 showed aortic binding in a rat quantitative whole-body autoradiography (QWBA) study, and its development was terminated prior to human dosing. A mechanistic hypothesis for this finding was established invoking reactivity with aldehydes involved in the cross-linking of elastin, a major component of aortic tissue. This was tested by developing a simple aldehyde chemical reactivity assay and a novel in vitro competitive covalent binding assay. Results obtained with AZD5248, literature compounds, and close analogues of AZD5248 support the mechanistic hypothesis and provide validation for the use of these assays in a two tier screening approach to support lead optimization. The strengths and limitations of these assays are discussed.

Synthesis of 2 C‐14 labeled cathepsin C inhibitors: The use of a cyanide to displace a Benzotriazole

“…Building on our past experience in labeling AZD5248 with C‐14, we proposed to conduct the synthesis is an analogous manner . Nitrile 3 had previously been prepared from 4‐iodobromobenzene via a CuI catalyzed reaction with K 14 CN to give a 9:1 mixture of 4‐bromobenzo‐[ 14 C]‐nitrile and 4‐iodobenzo‐[ 14 C]‐nitrile .…”

“…Building on our past experience in labeling AZD5248 with C-14, we proposed to conduct the synthesis is an analogous manner. 2,6,7 Nitrile 3 had previously been prepared from 4-iodobromobenzene via a CuI catalyzed reaction with K 14 CN to give a 9:1 mixture of 4-bromobenzo-[ 14 C]-nitrile and 4-iodobenzo-[ 14 C]-nitrile. 6 This mixture was reacted with boronate 4 to give amide 5 in 80% radiochemical yield and amide 5 was dehydrated with Burgess reagent 8,9 to give the penultimate compound, 6, in 95% yield (Scheme 2).…”

“…If possible, the identity of the compounds was determined by co‐elution on HPLC and by LCMS (Pos ESI, PenTOF 200‐1500 Da, 4 scans/s, Poroshell EC‐C18, 1 × 50 mm 2.7 um, 50 uL/min, 5 to 95% MeCN‐H 2 O in 4 minutes or Shimadzu LCMS2020 with Shimadzu LC20 series liquid handling system and method B). The specific activities of the products were determined by LC/MS using Isopat to deconvolute the MS signals . Liquid scintillation counting was performed with a Beckman LS 6500 scintillation counter.…”

“…During the safety evaluation of AZD5248, aortic binding was observed in the quantitative whole‐body autoradiography (QWBA) and binding to elastin was suspected to be the cause . Elastin can be found in relatively high abundance in the aorta and has been demonstrated to interact with rofecoxib .…”

“…During the safety evaluation of AZD5248, 1 aortic binding was observed in the quantitative whole-body autoradiography (QWBA) and binding to elastin was suspected to be the cause. 2 Elastin can be found in relatively high abundance in the aorta and has been demonstrated to interact with rofecoxib. 3,4 A hypothesis was advanced to explain the mechanism of the interaction in which an allysine residue of elastin reacts with the 4-aminotetrahydro-2H-pyran-4-carboxamide of the drug molecule to form a bicyclic ring system (Scheme 1).…”