Germline de novo variant F747S extends the phenotypic spectrum of CACNA1D Ca2+ channelopathies.

Germline gain-of-function missense variants in the pore-forming Cav1.3 α1-subunit (CACNA1D gene) confer high risk for a severe neurodevelopmental disorder with or without endocrine symptoms. Here, we report a 4-week-old new-born with the novel de novo missense variant F747S with a so far not described prominent jittering phenotype in addition to symptoms previously reported for CACNA1D mutations including developmental delay, elevated aldosterone level and transient hypoglycemia. We confirmed the pathogenicity of this variant in whole-cell patch-clamp experiments with wild-type and F747S mutant channels heterologously expressed together with α2δ1 and cytosolic ß3 or membrane-bound ß2a subunits. Mutation F747S caused the quantitatively largest shift in the voltage dependence of activation (-28 mV) reported so far for CACNA1D germline mutations. It also shifted inactivation to more negative voltages, slowed the time course of current inactivation and slowed current deactivation upon repolarization with both co-expressed ß-subunits. In silico modelling and molecular docking, simulations revealed that this gain-of-function phenotype can be explained by formation of a novel inter-domain hydrogen bond between mutant residues S747 (IIS6) with N1145 (IIIS6) stabilizing selectively the activated open channel state. F747S displayed 2-6-fold increased sensitivity for the L-type Ca2+ channel blocker isradipine compared to wild type. Our data confirm the pathogenicity of the F747S variant with very strong gain-of-function gating changes, which may contribute to the novel jittering phenotype. Increased sensitivity for isradipine suggests this drug for potential symptomatic off-label treatment for carriers of this mutation.

Cav 1.3-selective inhibitors of voltage-gated L-type Ca2+ channels: Fact or (still) fiction?

Voltage-gated L-type Ca2+ -channels (LTCCs) are the target of Ca2+ -channel blockers (CCBs), which are in clinical use for the evidence-based treatment of hypertension and angina. Their cardiovascular effects are largely mediated by the Cav 1.2-subtype. However, based on our current understanding of their physiological and pathophysiological roles, Cav 1.3 LTCCs also appear as attractive drug targets for the therapy of various diseases, including treatment-resistant hypertension, spasticity after spinal cord injury and neuroprotection in Parkinson's disease. Since CCBs inhibit both Cav 1.2 and Cav 1.3, Cav 1.3-selective inhibitors would be valuable tools to validate the therapeutic potential of Cav 1.3 channel inhibition in preclinical models. Despite a number of publications reporting the discovery of Cav 1.3-selective blockers, their selectivity remains controversial. We conclude that at present no pharmacological tools exist that are suitable to confirm or refute a role of Cav 1.3 channels in cellular responses. We also suggest essential criteria for a small molecule to be considered Cav 1.3-selective.