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

Abstract

Activating mutations of calcium-sensing receptor (CaSR) cause autosomal dominant hypocalcemia (ADH). ADH patients develop hypocalcemia, hyperphosphatemia, and hypercalciuria, similar to the clinical features of hypoparathyroidism. The current treatment of ADH is similar to the other forms of hypoparathyroidism, using active vitamin D 3 or parathyroid hormone (PTH). However, these treatments aggravate hypercalciuria and renal calcification. Thus, new therapeutic strategies for ADH are needed. Calcilytics are allosteric antagonists of CaSR, and may be effective for the treatment of ADH caused by activating mutations of CaSR. In order to examine the effect of calcilytic JTT-305/MK-5442 on CaSR harboring activating mutations in the extracellular and transmembrane domains in vitro, we first transfected a mutated CaSR gene into HEK cells. JTT-305/MK-5442 suppressed the hypersensitivity to extracellular Ca 2þ of HEK cells transfected with the CaSR gene with activating mutations in the extracellular and transmembrane domains. We then selected two activating mutations locating in the extracellular (C129S) and transmembrane (A843E) domains, and generated two strains of CaSR knock-in mice to build an ADH mouse model. Both mutant mice mimicked almost all the clinical features of human ADH. JTT-305/MK-5442 treatment in vivo increased urinary cAMP excretion, improved serum and urinary calcium and phosphate levels by stimulating endogenous PTH secretion, and prevented renal calcification. In contrast, PTH(1-34) treatment normalized serum calcium and phosphate but could not reduce hypercalciuria or renal calcification. CaSR knock-in mice exhibited low bone turnover due to the deficiency of PTH, and JTT-305/MK-5442 as well as PTH(1-34) increased bone turnover and bone mineral density (BMD) in these mice. These results demonstrate that calcilytics can reverse almost all the phenotypes of ADH including hypercalciuria and renal calcification, and suggest that calcilytics can become a novel therapeutic agent for ADH.

Knockin mouse with mutant Gα11 mimics human inherited hypocalcemia and is rescued by pharmacologic inhibitors

“…This finding is notably different from the increased BMD typically present in patients with other forms of hypoparathyroidism (29,30). It is also different from the findings of increased BMD in knockin mice expressing an activated CASR mutant (31). By contrast, transgenic mice expressing an oncogenic variant of the closely related Gα q (Gα q Q207L ) in osteoblasts through the col. 1 promoter have osteoporosis (32), and related studies in vitro show that activation of Gα q signaling inhibits osteoblast differentiation (32).…”

GENETICS IN ENDOCRINOLOGY: Gain and loss of function mutations of the calcium-sensing receptor and associated proteins: current treatment concepts

“…Soon it was, therefore, proposed that these compounds could correct the molecular defect caused by inactivating or activating CASR mutations and thus might be useful in the treatment of associated diseases (41,42,55,56).…”

“…Relative hypercalciuria is another feature, which is present in almost all ADH patients (21, 100) and poses them at risk for nephrocalcinosis, nephrolithiasis and renal failure. Besides chronic hypocalcemia and relative hypercalciuria, low or even undetectable PTH levels may also contribute to some of the pathophysiological consequences of ADH (56). In principle, one would like to correct the underlying pathophysiological mechanism to prevent or ameliorate these risks even in patients who are seemingly asymptomatic.…”

“…These are exactly the effects which are desired in patients with activating CASR mutations. Recent studies using knock-in mouse models of ADH and BS type 5 showed, that calcilytics could increase plasma calcium and PTH, reverse hypercalciuria and renal calcification and were more effective than PTH (56,114). The most common adverse effects in humans reported were mild neurological and gastrointestinal symptoms like fatigue, headache, constipation, diarrhea, nausea, and dyspepsia.…”

Novel calcium-sensing receptor cytoplasmic tail deletion mutation causing autosomal dominant hypocalcemia: molecular and clinical study

“…In a few cases of ADH due to CASR mutations, the long-term use of human PTH has been shown to be of benefit (37,38). In addition, the potential for using calcilytics, compounds that desensitize the CASR, has recently been demonstrated in mouse models of ADH (39,40).…”