Pharmacological Characterization of [³H]VUF11211, a Novel Radiolabeled Small-Molecule Inverse Agonist for the Chemokine Receptor CXCR3

Abstract

Chemokine receptor CXCR3 has attracted much attention, as it is thought to be associated with a wide range of immunerelated diseases. As such, several small molecules with different chemical structures targeting CXCR3 have been discovered. Despite limited clinical success so far, these compounds serve as interesting tools for investigating receptor activation and antagonism. Accumulating evidence suggests that many of these compounds are allosteric modulators for CXCR3. One feature of allosteric ligands is that the magnitude of the mediated allosteric effect is dependent on the orthosteric probe that is used. Consequently, there is a risk for incorrect assessment of affinity for allosteric modulators with orthosteric radioligands, which has so far been the most applied approach for chemokine receptors.

Combined CXCR3/CXCR4 measurements are of high prognostic value in chronic lymphocytic leukemia due to negative co-operativity of the receptors

“…Performing chemotaxis assays as previously described, 8 we found that CLL cells pre-stimulated with either CXCR3 ligand CXCL9, CXCL10, or CXCL11 exhibited significantly reduced directed cell migration towards CXCL12 ( Figure 2Ai). Pre-stimulation with a small-molecule high-affinity CXCR3 agonist (VUF11418) 9 confirmed this observation and an antagonist (VUF11211) 9 did not interfere with chemotactic ability (Figure 2Aii).…”

Metabolic profiling of ligands for the chemokine receptor CXCR3 by liquid chromatography-mass spectrometry coupled to bioaffinity assessment

“…G418 was purchased from Duchefa (Haarlem, The Netherlands). 125 I- CXCL10, [ 3 H]-VUF11211 (38.4 Ci/mmol) [ 31 ], and Whatman GF/C filter plates were purchased from Perkin Elmer (Groningen, The Netherlands).…”

“…The radioligand binding assay was performed in a similar fashion as described previously by Scholten et al [ 31 ]. Shortly, the [ 3 H]-VUF11211 radioligand was dissolved in binding buffer (50 mM Tris-HCl buffer pH 7.4 at room temperature, 100 mM NaCl, 0.1 % Tween 80, and 0.1 % BSA) and added to wells containing ligand or collected high-resolution fractions of the ligands.…”

“…Metabolites were separated using reversed phase liquid chromatography (LC) followed, in parallel, by high-resolution nanofractionation onto 96-well plates to assess the bioaffinity of parent compounds and metabolites and accurate mass MS and MS 2 analysis for structural identification of formed metabolites. The bioaffinity was assessed using a radioligand binding assay employing the recently described small molecule radioligand [ 3 H]-VUF11211 [ 31 ]. In total, ten metabolites of NBI-74330 and eight metabolites of VUF11211 have been identified and their structures were partially or fully elucidated.…”

Cholesterol impacts chemokine CCR5 receptor ligand‐binding activity

“…In that sense, the receptor is the target of ligands that through receptor‐conformational changes aim at blocking different stages in the viral fusion process. One such ligand is maraviroc, the first CCR5 antagonist approved by both the European Medicines Agency and the US Food and Drug Administration (FDA) for treatment of HIV‐infected patients experiencing virological failure due to resistance to other classes of antiretroviral drugs . It represents a novel class of antiretroviral drugs, as it is the first therapeutic targeting a cellular rather than a viral protein.…”