Therapeutic Opportunities of Targeting Allosteric Binding Sites on the Calcium-Sensing Receptor

Abstract

The CaSR is a class C G protein-coupled receptor (GPCR) that acts as a multimodal chemosensor to maintain diverse homeostatic functions. The CaSR is a clinical therapeutic target in hyperparathyroidism and has emerged as a putative target in several other diseases. These include hyper- and hypocalcaemia caused either by mutations in the CASR gene or in genes that regulate CaSR signaling and expression, and more recently in asthma. The development of CaSR-targeting drugs is complicated by the fact that the CaSR possesses many different binding sites for endogenous and exogenous agonists and allosteric modulators. Binding sites for endogenous and exogenous ligands are located throughout the large CaSR protein and are interconnected in ways that we do not yet fully understand. This review summarizes our current understanding of CaSR physiology, signaling, and structure and how the many different binding sites of the CaSR may be targeted to treat disease.

Figure 1

Principal CaSR signaling pathways. CaSR activation of G_i/o inhibits adenylyl cyclase (AC) to reduce cAMP levels. CaSR coupling to G_q/11 activates phosphatidylinositol-specific phospholipase C (PI–PLC) to increase inositol triphosphate (IP₃) and diacyl glycerol (DAG) and trigger the release of Ca²⁺_i from stores. Ca²⁺_i activates phospholipase A₂ (PLA₂) and PKC. The CaSR also increases Ca²⁺_i via influx through L-type voltage-gated and transient receptor potential ion channels (IC), in part via PKC. The CaSR activates MAPK signaling cascades via G_q/11-mediated PKC, G_i/o-mediated activation of epidermal growth factor receptor (EGFR), and β arrestin.

Figure 2

CaSR model and predicted ligand binding sites. The published crystal structure of the CaSR ECD (PDB 5K5S) was superimposed onto a published model of the CaSR 7TM, ECLs, and ICLs based on homology with the mGlu₅ crystal structure (PDB 6N51). Numbers correspond to ligand binding sites predicted as follows: Ca²⁺ (sites 1–4) by anomalous scattering analysis and SO₄^2– (5–7), PO₄^3– (8–9), and l-Trp (10) by electron density distribution analysis of the crystallized ECD (PDBs 5K5S and 5K5T); TNCA (11), Mg²⁺ (12–13), and Gd³⁺ (13) by electron density distribution analysis of the crystallized VFT (PDBs 5FBK and 5FBH); etelcalcetide (15) from mutagenesis and mass spectrometry, where the yellow stick represents a putative disulfide bond as a rough depiction of where etelcalcetide is predicted to bind; quinazolinone-containing NAMs (16), arylalkylamine PAMs and NAMs (17), and AC265347 (18) from mutagenesis combined with homology modeling and computational docking.^,

Figure 3

CaSR agonists and PAMs and their structures.

Figure 4

An operational model of allosterism to quantify CaSR agonist, PAM, and NAM actions. Endogenous CaSR agonists such as Ca²⁺_o bind with a mean equilibrium dissociation constant, K_A, to multiple sites. When the receptor is occupied by an agonist, the agonist stimulates a response, depicted by an operational measure of efficacy, τ_A. Allosteric modulators may alter the equilibrium dissociation constant of the agonist via a cooperativity factor, α, or alter the efficacy of the orthosteric agonist via a scaling factor, β. Allosteric modulators may also have their own efficacy, τ_B.

Figure 5

CaSR NAMs and their structures.