The calcilytic agent NPS2143 rectifies hypocalcaemia in a mouse model, Nuf, that is due to an activating calcium-sensing-receptor (CaSR) mutation: relevance to autosomal dominant hypocalcaemia with hypercalciuria
Fadil Hannan1, Gerard Walls1, Eniko Kallay1, M Andrew Nesbit1, Tertius Hough2, Roger Cox2, Jianxin Hu3, Allen Spiegel4 & Rajesh Thakker1
The G-protein coupled calcium-sensing-receptor (CaSR) regulates calcium homeostasis and inactivating mutations result in familial hypocalciuric hypercalcaemia (FHH), whilst activating mutations result in autosomal dominant hypocalcaemia with hypercalciuria (ADHH). Allosteric CaSR modulators consist of: calcimimetics, which activate the CaSR e.g. Cinacalcet, that is used to treat the hypercalcaemia of chronic renal failure and metastatic parathyroid carcinoma; and calcilytics e.g. NPS2143 that antagonise the CaSR. We have assessed NPS2143 for use in ADHH by in vitro and in vivo studies utilising HEK293 cells transiently transfected with wild-type or mutant CaSRs, and a hypocalcaemic mouse model, Nuf, that harbours an activating CaSR mutation, Leu723Gln. Mice were used in accordance with UK Home Office welfare guidelines and project license restrictions. In vitro studies using HEK293 cells demonstrated that 20 nM NPS2143 restored the significantly lower EC50 (1.94 mM±0.07, P< 0.01) of the mutant CaSR (Gln723) to a value (2.79 mM ±0.19) that was indistinguishable from that of the wild-type (Leu723) (EC50=2.53 mM±0.14, P=0.33). Moreover, 40 and 80 nM NPS2143 significantly raised the EC50 (3.97±0.1 and 4.56±0.01 respectively, P< 0.001) of the mutant Gln723 CaSR. In vivo studies, using heterozygous Nuf and wild-type mice that were given 1.1mg of NPS2143 intraperitoneally, demonstrated that the hypocalcaemia of Nuf mice was significantly improved from 1.74±0.02 to 2.15 mmol/l±0.05 (P<0.0001) at 1 h, and 2.30 mmol/l±0.08 (P<0.0001) at 24 h, which approached the values observed in wild-type mice (2.46 mmol/l±0.03). Thus, NPS2143 has a significant effect on the EC50, and hence set-point, of the Nuf mutant CaSR and NPS2143 rectifies, in vivo, the hypocalcaemia of Nuf mice. Our findings indicate that calcilytics may be a useful treatment for ADHH.