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Cinacalcet corrects hypercalcemia in mice with an inactivating G?11 mutation.

ABSTRACT: Loss-of-function mutations of GNA11, which encodes G-protein subunit ?11 (G?11), a signaling partner for the calcium-sensing receptor (CaSR), result in familial hypocalciuric hypercalcemia type 2 (FHH2). FHH2 is characterized by hypercalcemia, inappropriately normal or raised parathyroid hormone (PTH) concentrations, and normal or low urinary calcium excretion. A mouse model for FHH2 that would facilitate investigations of the in vivo role of G?11 and the evaluation of calcimimetic drugs, which are CaSR allosteric activators, is not available. We therefore screened DNA from > 10,000 mice treated with the chemical mutagen N-ethyl-N-nitrosourea (ENU) for GNA11 mutations and identified a G?11 variant, Asp195Gly (D195G), which downregulated CaSR-mediated intracellular calcium signaling in vitro, consistent with it being a loss-of-function mutation. Treatment with the calcimimetic cinacalcet rectified these signaling responses. In vivo studies showed mutant heterozygous (Gna11+/195G) and homozygous (Gna11195G/195G) mice to be hypercalcemic with normal or increased plasma PTH concentrations and normal urinary calcium excretion. Cinacalcet (30mg/kg orally) significantly reduced plasma albumin-adjusted calcium and PTH concentrations in Gna11+/195G and Gna11195G/195G mice. Thus, our studies have established a mouse model with a germline loss-of-function G?11 mutation that is representative for FHH2 in humans and demonstrated that cinacalcet can correct the associated abnormalities of plasma calcium and PTH.

SUBMITTER: Howles SA

PROVIDER: S-EPMC5846897 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Cinacalcet corrects hypercalcemia in mice with an inactivating Gα11 mutation.

Howles Sarah A SA Hannan Fadil M FM Gorvin Caroline M CM Piret Sian E SE Paudyal Anju A Stewart Michelle M Hough Tertius A TA Nesbit M Andrew MA Wells Sara S Brown Stephen Dm SD Cox Roger D RD Thakker Rajesh V RV

JCI insight 20171019 20

Loss-of-function mutations of GNA11, which encodes G-protein subunit α11 (Gα11), a signaling partner for the calcium-sensing receptor (CaSR), result in familial hypocalciuric hypercalcemia type 2 (FHH2). FHH2 is characterized by hypercalcemia, inappropriately normal or raised parathyroid hormone (PTH) concentrations, and normal or low urinary calcium excretion. A mouse model for FHH2 that would facilitate investigations of the in vivo role of Gα11 and the evaluation of calcimimetic drugs, which ...[more]

PMID: 29046478

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Project description:G-protein subunit ?-11 (G?11 ) couples the calcium-sensing receptor (CaSR) to phospholipase C (PLC)-mediated intracellular calcium (Ca2+i ) and mitogen-activated protein kinase (MAPK) signaling, which in the parathyroid glands and kidneys regulates parathyroid hormone release and urinary calcium excretion, respectively. Heterozygous germline loss-of-function G?11 mutations cause familial hypocalciuric hypercalcemia type 2 (FHH2), for which effective therapies are currently not available. Here, we report a novel heterozygous G?11 germline mutation, Phe220Ser, which was associated with hypercalcemia in a family with FHH2. Homology modeling showed the wild-type (WT) Phe220 nonpolar residue to form part of a cluster of hydrophobic residues within a highly conserved cleft region of G?11 , which binds to and activates PLC; and predicted that substitution of Phe220 with the mutant Ser220 polar hydrophilic residue would disrupt PLC-mediated signaling. In vitro studies involving transient transfection of WT and mutant G?11 proteins into HEK293 cells, which express the CaSR, showed the mutant Ser220 G?11 protein to impair CaSR-mediated Ca2+i and extracellular signal-regulated kinase 1/2 (ERK) MAPK signaling, consistent with diminished activation of PLC. Furthermore, engineered mutagenesis studies demonstrated that loss of hydrophobicity within the G?11 cleft region also impaired signaling by PLC. The loss-of-function associated with the Ser220 G?11 mutant was rectified by treatment of cells with cinacalcet, which is a CaSR-positive allosteric modulator. Furthermore, in vivo administration of cinacalcet to the proband harboring the Phe220Ser G?11 mutation, normalized serum ionized calcium concentrations. Thus, our studies, which report a novel G?11 germline mutation (Phe220Ser) in a family with FHH2, reveal the importance of the G?11 hydrophobic cleft region for CaSR-mediated activation of PLC, and show that allosteric CaSR modulation can rectify the loss-of-function Phe220Ser mutation and ameliorate the hypercalcemia associated with FHH2. © 2017 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc.

Dataset Information

Cinacalcet corrects hypercalcemia in mice with an inactivating G?11 mutation.

Publications

Cinacalcet corrects hypercalcemia in mice with an inactivating Gα11 mutation.

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