Gα11 mutation in mice causes hypocalcemia rectifiable by calcilytic therapy
Caroline M. Gorvin, Fadil M. Hannan, Sarah A. Howles, Valerie N. Babinsky, Sian E. Piret, Angela Rogers, Andrew J. Freidin, Michelle Stewart, Anju Paudyal, Tertius A. Hough, M. Andrew Nesbit, Sara Wells, Tonia L. Vincent, Stephen D.M. Brown, Roger D. Cox, Rajesh V. Thakker
Caroline M. Gorvin, Fadil M. Hannan, Sarah A. Howles, Valerie N. Babinsky, Sian E. Piret, Angela Rogers, Andrew J. Freidin, Michelle Stewart, Anju Paudyal, Tertius A. Hough, M. Andrew Nesbit, Sara Wells, Tonia L. Vincent, Stephen D.M. Brown, Roger D. Cox, Rajesh V. Thakker
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Research Article Endocrinology Metabolism

Gα11 mutation in mice causes hypocalcemia rectifiable by calcilytic therapy

Abstract

Heterozygous germline gain-of-function mutations of G-protein subunit α11 (Gα11), a signaling partner for the calcium-sensing receptor (CaSR), result in autosomal dominant hypocalcemia type 2 (ADH2). ADH2 may cause symptomatic hypocalcemia with low circulating parathyroid hormone (PTH) concentrations. Effective therapies for ADH2 are currently not available, and a mouse model for ADH2 would help in assessment of potential therapies. We hypothesized that a previously reported dark skin mouse mutant (Dsk7) — which has a germline hypermorphic Gα11 mutation, Ile62Val — may be a model for ADH2 and allow evaluation of calcilytics, which are CaSR negative allosteric modulators, as a targeted therapy for this disorder. Mutant Dsk7/+ and Dsk7/Dsk7 mice were shown to have hypocalcemia and reduced plasma PTH concentrations, similar to ADH2 patients. In vitro studies showed the mutant Val62 Gα11 to upregulate CaSR-mediated intracellular calcium and MAPK signaling, consistent with a gain of function. Treatment with NPS-2143, a calcilytic compound, normalized these signaling responses. In vivo, NPS-2143 induced a rapid and marked rise in plasma PTH and calcium concentrations in Dsk7/Dsk7 and Dsk7/+ mice, which became normocalcemic. Thus, these studies have established Dsk7 mice, which harbor a germline gain-of-function Gα11 mutation, as a model for ADH2 and have demonstrated calcilytics as a potential targeted therapy.

Authors

Caroline M. Gorvin, Fadil M. Hannan, Sarah A. Howles, Valerie N. Babinsky, Sian E. Piret, Angela Rogers, Andrew J. Freidin, Michelle Stewart, Anju Paudyal, Tertius A. Hough, M. Andrew Nesbit, Sara Wells, Tonia L. Vincent, Stephen D.M. Brown, Roger D. Cox, Rajesh V. Thakker

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Figure 4

MAPK responses of the Val62 Gα11 mutant and effect of NPS-2143 treatment.

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MAPK responses of the Val62 Gα11 mutant and effect of NPS-2143 treatment...
(A) Western blot analysis of lysates from HEK-CaSR cells used for phosphorylated ERK (pERK) experiments. (B) pERK fold-change responses to changes in [Ca2+]o of cells transfected with WT (solid line) or Val62 Gα11 mutant (dashed line). (C) Western blot analysis of lysates from cells used to assess effect of NPS-2143 (2143) on pERK responses. (D) Effect of NPS-2143 on pERK responses of Val62 Gα11 mutant. (E) Western blot analysis of lysates from HEK-CaSR cells used for serum response element (SRE) reporter experiments. (F) SRE reporter fold-change responses to changes in [Ca2+]o of cells transfected with WT (solid line) or Val62 Gα11 mutant (dashed line). (G) Western blot analysis of lysates from cells used to assess effect of NPS-2143 on SRE reporter responses. (H) Effect of NPS-2143 on the SRE reporter responses of the Val62 Gα11 mutant. The Val62 Gα11 mutant led to significantly increased pERK and SRE fold-change responses following stimulation with Ca2+o. In the absence of Ca2+o, the pERK and SRE responses of the Val62 Gα11 mutant were not significantly different from WT, thereby indicating the Ile62Val Gα11 mutation to be nonconstitutively activating. The addition of 20 nM NPS-2143 decreased the pERK and SRE responses of cells expressing the Val62 Gα11 mutant (patterned bars) compared with untreated cells (solid bars), so that these were not significantly different from WT (open bars). The fold-change responses are shown as the mean ± SEM of 4–8 independent transfections. *P < 0.05, **P < 0.01. Mann-Whitney U test for B, D, F, and H.