Characterization of quinazolinone calcilytic therapy for autosomal dominant hypocalcemia type 1 (ADH1)

Gain-of-function mutations of the calcium-sensing receptor (CaSR) result in autosomal dominant hypocalcemia type 1 (ADH1), which may cause symptomatic hypocalcemia with low parathyroid hormone concentrations. Negative allosteric CaSR modulators, known as calcilytics, have potential as a targeted ADH1 therapy and comprise two main classes, which are the amino alcohols and the quinazolinones. Amino alcohol calcilytics have been assessed as ADH1 therapies but may not be effective for all ADH1-causing mutations. We therefore evaluated quinazolinone calcilytics (ATF936 and AXT914) as an alternate ADH1 treatment. Calcilytic docking studies were performed using reported cryo-EM CaSR structures. In vitro dose-response studies were performed using CaSR-expressing HEK293 cells and in vivo studies undertaken in mice with a gain-of-function CaSR mutation, Leu723Gln, known as Nuf. ATF936 and AXT914, as well as the amino alcohol calcilytics, NPS 2143 and NPSP795, were shown to bind at a common region within the CaSR transmembrane domain, which is also an ADH1 mutational hotspot. Treatment of cells expressing the Nuf mutant (Gln723) CaSR with 1 to 20 nM AXT914 caused dose-dependent decreases in CaSR-mediated intracellular calcium responses with 10 nM AXT914 normalizing the gain of function. Oral administration of 10 mg/kg AXT914 to Nuf mice increased parathyroid hormone to 104 ± 29 pmol/l compared with 23 ± 4 pmol/l for vehicle-treated mice, p < 0.05; and increased plasma albumin-adjusted calcium to 2.03 ± 0.02 mmol/l compared with 1.84 ± 0.02 mmol/l for vehicle-treated mice, p < 0.001. These studies indicate that quinazolinone calcilytics may have potential for treating ADH1.