Novel Activating Mutations of the Calcium-Sensing Receptor: The Calcilytic NPS-2143 Mitigates Excessive Signal Transduction of Mutant Receptors

Abstract

Context and Objective: Activating mutations in the calcium-sensing receptor (CaSR) gene cause autosomal dominant hypocalcemia (ADH). The aims of the present study were the functional characterization of novel mutations of the CaSR found in patients, the comparison of in vitro receptor function with clinical parameters, and the effect of the allosteric calcilytic NPS-2143 on the signaling of mutant receptors as a potential new treatment for ADH patients. Methods: Wild-type and mutant CaSR (T151R, P221L, E767Q, G830S, and A844T) were expressed in human embryonic kidney cells (HEK 293T). Receptor signaling was studied by measuring intracellular free calcium in response to different concentrations of extracellular calcium ([Ca2+]o) in the presence or absence of NPS-2143. Results: All ADH patients had lowered serum calcium ranging from 1.7 to 2.0 mm and inadequate immunoreactive PTH and urinary calcium excretion. In vitro testing of CaSR mutations from these patients revealed exaggerated [Ca2+]o-induced cytosolic Ca2+ responses with EC50 values for [Ca2+]o ranging from 1.56 to 3.15 mm, which was lower than for the wild-type receptor (4.27 mm). The calcilytic NPS-2143 diminished the responsiveness to [Ca2+]o in the CaSR mutants T151R, E767Q, G830S, and A844T. The mutant P221L, however, was only responsive when coexpressed with the wild-type CaSR. Conclusion: Calcilytics might offer medical treatment for patients with autosomal dominant hypocalcemia caused by calcilytic-sensitive CaSR mutants.

Activating mutations in the calcium-sensing receptor: genetic and clinical spectrum in 25 patients with autosomal dominant hypocalcaemia - a German survey

“…Eight mutations have been described and characterized in the literature (F128T, P221L, Q245R, E556K and E830S), 6,[14][15][16] including three by our group (T151R, E767Q and A844T). 11,17 Here, we describe one novel variant, M734T, in a single case with asymptomatic hypocalcaemia, inappropriately low PTH level and normal calcium excretion. These biochemical findings argue for the diagnosis of ADH.…”

“…6 Hypocalciuric substances like PTH and thiazides are used to overcome these problems, while calcilytics are being examined. [9][10][11] It is important to recognize ADH and define these patients by genetic testing because they can exhibit unusual sensitivity to treatments for hypocalcaemia, leading to toxic renal effects. Here, we describe a large series of 25 patients carrying activating mutations in the CASR gene, their clinical presentation and long-term outcomes under low-dose calcitriol treatment.…”

The calcium-sensing receptor in bone metabolism: from bench to bedside and back

“…Calcilytics may indeed be employed in conditions characterized by hypocalcemia and inappropriately low PTH levels, such as ADH, caused by activating mutation of the CaSR, since they revert the abnormal set point for systemic calcium homeostasis [103,104]. In this setting, CaSR antagonists mainly act through preserving calcium eliciting paracellular calcium transport through the thick Fig.…”

Chemotactic and proangiogenic role of calcium sensing receptor is linked to secretion of multiple cytokines and growth factors in breast cancer MDA-MB-231 cells

“…Whereas 10 μg/ml of 7F8, a control monoclonal antibody that recognizes CaSR but does not significantly alter its function, did not interfere with the secretion of chemotactic factors by MDA-MB-231 cells. In addition, we investigated the effect of NPS-2143, a calcilytic known as a selective antagonist of CaSR [53][54][55][56][57]. As shown in Fig.…”