Biased Agonist Pharmacochaperones of the AVP V2 Receptor May Treat Congenital Nephrogenic Diabetes Insipidus

Abstract

X-linked congenital nephrogenic diabetes insipidus (cNDI) results from inactivating mutations of the human arginine vasopressin (AVP) V2 receptor (hV 2 R). Most of these mutations lead to intracellular retention of the hV 2 R, preventing its interaction with AVP and thereby limiting water reabsorption and concentration of urine. Because the majority of cNDI-hV 2 Rs exhibit protein misfolding, molecular chaperones hold promise as therapeutic agents; therefore, we sought to identify pharmacochaperones for hV 2 R that also acted as agonists. Here, we describe high-affinity nonpeptide compounds that promoted maturation and membrane rescue of L44P, A294P, and R337X cNDI mutants and restored a functional AVP-dependent cAMP signal. Contrary to pharmacochaperone antagonists, these compounds directly activated a cAMP signal upon binding to several cNDI mutants. In addition, these molecules displayed original functionally selective properties (biased agonism) toward the hV 2 R, being unable to recruit arrestin, trigger receptor internalization, or stimulate mitogen-activated protein kinases. These characteristics make these hV 2 R agonist pharmacochaperones promising therapeutic candidates for cNDI. The antidiuretic hormone arginine-vasopressin (AVP) is crucial for osmoregulation, cardiovascular control, and water homeostasis. The human AVP V 2 receptor (hV 2 R), localized in the principal cells of the kidney collecting duct, mediates AVP antidiuretic effect and therefore helps in maintaining physiologic plasma osmolality, blood volume, and arterial pressure. Binding of AVP to hV 2 R first triggers a cAMP signal through activation of the G protein ␣ s (Gs) subunit and adenylyl cyclase (AC). Then, the cAMP-activated protein kinase A phosphorylates aquaporin 2 water channels, resulting in their insertion into the luminal membrane of principal cells and finally to water reabsorption. 1 AVP binding to hV 2 R also induces arrestin recruitment, receptor internalization, 2 and mitogen-activated protein kinase (MAPK) activation. 3 Mutations in the hV 2 R gene lead to the X-linked congenital nephrogenic diabetes insipidus (cNDI), a rare disease characterized by the kidney's inability to concentrate urine despite normal or elevated plasma concentrations of AVP. 4 More than 200 different mutations have been described and are responsible for polyuria, a main consequence of the disease. Most of the mutant receptors (cNDIhV 2 Rs), trapped in the endoplasmic reticulum

Structural insights into biased G protein-coupled receptor signaling revealed by fluorescence spectroscopy

“…These results demonstrate that NAPol-reconstituted V2R interacts with and efficiently activates Gs protein in a ligand-dependent manner. Importantly, the efficacy profiles of the ligands correlated well with those observed in living cells (18,21).…”

“…To determine the functionality of NAPol-reconstituted V2R, we first measured ligand-induced incorporation of [ 35 S]GTPγS to purified Gαs proteins (1:2.5 receptor/G protein molar ratio). This assay was done using two full agonists for the Gs pathway, the endogenous hormone AVP and the biased ligand MCF14 (18), as well as with a Gs inverse agonist (SR121463) (21) (Fig. 2A).…”

“…V2R represents an interesting model to study molecular bases of functional selectivity for two reasons: (i) its pharmacology has been well-characterized using a large panel of ligands with different efficacies toward the Gs signaling pathway (17), and (ii) several V2R ligands are biased agonists. For instance, although the unbiased natural hormone AVP is a full agonist toward Gs protein and β-arrestin (Gs agonist/Arr agonist), two nonpeptide-biased synthetic ligands, MCF14 (Gs agonist/Arr antagonist) and SR121463 (Gs inverse agonist/Arr partial agonist), have been described (12,18).…”

Chaperons pharmacologiques

“…La découverte très récente d'agonistes biaisés du V2R (MCF14, MCF18, MCF57) offre une nouvelle opportunité thérapeutique dans le DINc [13]. Un agoniste biaisé est une molécule qui ne possède qu'une partie des propriétés pharmacologiques du ligand activateur endogène.…”

“…Ils offrent donc un avantage certain par rapport aux agonistes classiques et aux antagonistes, tout en ayant une affinité forte vis-à-vis du V2R. L'association de ces caractéristiques semble idéale pour traiter les patients atteints de DINc puisque ces molécules induisent effectivement le ciblage de mutants du V2R à la surface cellulaire [13]. Ces composés devraient permettre a priori une activation soutenue et durable dans le temps des V2R mutés puisque ceux-ci ne sont pas internalisés.…”