Positive and Negative Allosteric Modulators Promote Biased Signaling at the Calcium-Sensing Receptor

Abstract

The calcium-sensing receptor (CaSR) is a G protein-coupled receptor whose function can be allosterically modulated in a positive or negative manner by calcimimetics or calcilytics, respectively. Indeed, the second-generation calcimimetic, cinacalcet, has proven clinically useful in the treatment of chronic kidney disease patients with secondary hyperparathyroidism but is not widely used in earlier stages of renal disease due to the potential to predispose such patients to hypocalcaemia and hyperphosphatemia. The development of a biased CaSR ligand that is more selective for specific signaling pathway(s) leading only to beneficial effects may overcome this limitation. The detection of such stimulus-bias at a G protein-coupled receptor requires investigation across multiple signaling pathways and the development of methods to quantify the effects of allosteric ligands on orthosteric ligand affinity and cooperativity at each pathway. In the current study, we determined the effects of the calcimimetics, NPS-R568 or cinacalcet, and the calcilytic, NPS-2143, on Ca o 2ϩ -mediated intracellular Ca 2ϩ mobilization, ERK1/2 phosphorylation, and plasma membrane ruffling in a stably transfected human embryonic kidney 293-TREx c-myc-CaSR cell line and applied a novel analytical model to quantify these modulator effects. We present quantitative evidence for the generation of stimulus bias by both positive and negative allosteric modulators of the CaSR, manifested as greater allosteric modulation of intracellular Ca 2ϩ mobilization relative to ERK1/2 phosphorylation, and a higher affinity of the modulators for the state of the CaSR mediating plasma membrane ruffling relative to the other two pathways. Our findings provide the first evidence that an allosteric modulator used in clinical practice exhibits stimulus bias. (Endocrinology 153: 1232-1241, 2012)

Biased agonism: a novel paradigm in G protein-coupled receptor signaling observed in acquired hypocalciuric hypercalcemia [Review]

“…LPI805 promotes destabilization of the NKA-NK2R complexes that are in a conformation that triggers cAMP production, whereas the access of NKA to the conformation that triggers intracellular calcium elevation is unchanged. In contrast, among family C GPCRs such as metabotropic glutamate receptors, taste receptors and the GABA B receptor, including a calcium sensing receptor, many positive and negative allosteric modulators have now been reported [28,29]. In contrast to the autoantibodies we described in our AHH patient, however, most of these factors are not biased modulators.…”

The Effective Application of Biased Signaling to New Drug Discovery

“…Because allosteric molecules are by nature permissive in that they may allow the interaction of the receptor with the natural agonist, there is a probability that the allosteric ligand will change the quality of the natural agonist signaling-that is, it will produce a bias for the natural agonist effect. Such bias has been noted for negative allosteric molecules in the selective blockade of NK2 receptors (Maillet et al, 2007), prostaglandin D2 receptors (Mathiesen et al, 2005), and calcium-sensing receptors (Davey et al, 2012;Cook et al, 2015) and positive allosteric modulators for glucagon-like peptide 1 (GLP-1) receptors (Koole et al, 2010) and mGlutamic acid 5 receptors (Bradley et al, 2011). The possibility of producing induced-bias in natural signaling can be viewed as a potential positive aspect of allosteric modulation of drug effect but also should be seen as an added consideration in the development of allosteric molecules.…”

Calcilytic Ameliorates Abnormalities of Mutant Calcium-Sensing Receptor (CaSR) Knock-In Mice Mimicking Autosomal Dominant Hypocalcemia (ADH)

“…(28) Several calcilytics have been reported to reduce the sensitivity of CaSR to extracellular Ca 2þ , and cause a rightward shift of the extracellular Ca 2þ -tointracellular Ca 2þ concentration curve. (29,30) JTT-305 (also known as MK-5442), a short-acting calcilytic, transiently elevates the endogenous PTH secretion, and increases bone mineral density (BMD) in rats. (31) Because activating mutations of CaSR cause ADH, and because calcilytics suppress the enhanced sensitivity of CaSR to extracellular Ca 2þ , there is a possibility that JTT-305/MK-5442 may become a new therapeutic agent to reverse most of the abnormalities in Ca metabolism in ADH patients caused by activating mutations of CaSR.…”