Heterogenous Origins of Calcium Homeostasis Disorders Arising from Five Heterozygous Calcium-Sensing Receptor Variants

Abstract Context The human calcium-sensing receptor (CaSR) plays a key role in calcium homeostasis, and most identified CASR variants are associated with hypercalcemic and hypocalcemic disorders. Objective Here we characterized the pharmacological implications of 5 heterozygous CASR variants from individuals with familial hypocalciuric hypercalcemia 1 (FHH1: Y63C, I81T, Q459R, W818stop) or autosomal dominant hypocalcemia 1 (ADH1: R955stop). Methods Total and cell surface expression levels of wild-type (WT) and variant CaSRs expressed in human embryonic kidney 293T (HEK293T) cells were determined using enzyme-linked immunosorbent assay, and the pharmacological properties of the receptors were delineated in 2 functional assays. Results The Y63C and I81T variations in the extracellular domain (ECD) of CaSR yielded markedly reduced cell surface expression and Ca2+ responsiveness, while Q459R displayed WT-like expression and functional properties. Truncation of the 7-transmembrane domain (7TMD) in W818stop eliminated cell surface expression, whereas R955stop in the intracellular carboxy-terminal yielded modestly increased surface expression and Ca2+ potency compared with WT CaSR. Interestingly, the effectiveness of positive allosteric modulators (PAMs) at the variants varied. Ca2+-mediated signaling through Y63C and I81T was significantly augmented by 7TMD-binding PAMs (NPS R-568 and evocalcet) but not by ECD-binding PAMs (etelcalcetide and Nb4), whereas signaling through Q459R and R955stop were robustly potentiated by all four PAMs. Conclusion While the molecular phenotypes exhibited by the 5 CaSR variants concord with the clinical phenotypes in individuals harboring them, CASR variant–induced calcium homeostasis disorders clearly arise from diverse molecular origins, and the effectiveness of calcimimetics in these disorders could differ depending on the specific variants.