A Novel Gain-of-Function CACNA1C Variant (Gly856Asp) Associated With QT Prolongation and Sudden Death

Abstract

The investigation of sudden death is challenging, but of utmost importance to guide screening in potentially at-risk relatives. We present a case of a 14-year-old boy of Central Asian ancestry with palpitations and recent sudden death of his sister. Autopsy revealed ischemic injury of uncertain etiology. Family evaluation revealed mild QT prolongation in relatives cosegregating with a novel CACNA1C variant (Gly856Asp). Functional in vitro analyses show slower inactivation kinetics (gain of function) as shown in the long QT syndrome type 8. The case highlights the importance of a holistic approach to the investigation of sudden death and screening of at-risk relatives.

Graphical Abstract

History of Presentation

A 14-year-old boy of Central Asian ancestry was seen in the pediatric arrhythmia clinic for palpitations and family history of sudden cardiac death (SCD). He was the oldest living child of a family of 5 siblings (II-2 in pedigree) (Figure 1). He was physically active and never had syncope. His physical examination was normal.

Learning Objectives

• •To recognize nonsyndromic long QT syndrome type 8 as a rare potential cause of SCD.
• •To apply a holistic interdisciplinary approach when investigating complex cases of SCD.

Figure 1.

Pedigree

Circles represent female family members. Squares represent male family members. Pedigree identifiers are shown in bold characters below circles and squares, along with the corrected QT interval (QTc) (in ms) measured using the threshold method and Bazett correction (see ECGs in Figure 2 and Supplemental Figures 1 to 5). The percentile of the QTc in the control population is shown in parentheses. Heterozygous carriership status of the CACNA1C:p.Gly856Asp variant is shown as + (carrier) and – (noncarrier). The oldest daughter (II-1) had sudden cardiac death (SCD) at age 14 years.

Past Medical History

The patient had no prior medical history. His older sister (II-1) had SCD at 14 years of age a few months before his evaluation. SCD occurred during sleep and the coroner’s investigation was ongoing at the time of assessment.

Differential Diagnosis

Palpitations are most often benign. In the context of family history of SCD, exclusion of inherited structural or electrical heart disease was warranted.

Investigations

A resting electrocardiogram (ECG) (Supplemental Figure 1) showed sinus rhythm with a mildly increased corrected QT interval (QTc) (472 and 448 ms using Bazett [QTc-B] and Fridericia [QTc-F] corrections, respectively, and averaging measures over 5 beats using the threshold method¹). Holter monitoring and exercise testing were normal. Cardiac echocardiography was normal. Characterization of the rhythm at the time of palpitations was not possible.

In the context of family history of SCD, all living siblings were evaluated. His 8-year-old brother (II-4) was asymptomatic. His QT interval was also mildly prolonged (QTc-B = 483 ms and QTc-F = 469 ms) (Figure 2). The 2 sisters were asymptomatic; the older one (10 years of age; II-3) had a resting QTc-B of 482 ms and QTc-F of 458 ms (Supplemental Figure 2) and the younger one (4 years of age; II-5) had a resting QTc-B of 434 ms and QTc-F of 378 ms (Supplemental Figure 3). The mother had a normal ECG at rest (Supplemental Figure 4) (QTc-B = 415 ms, QTc-F = 384 ms). The father’s ECG revealed a QTc interval at the upper limit of normal (QTc-B = 436 ms; QTc-F = 430 ms) (Supplemental Figure 5). Given the mildly prolonged QT intervals in several family members (Supplemental Table 1) and family history of SCD, long QT syndrome (LQTS) was suspected. A LQTS genetic panel including sequencing of CACNA1C, CALM1, CALM2, KCNE1, KCNE2, KCNH2, KCNJ2, KCNQ1, and SCN5A was performed for the sibling (II-4) with the longest QTc at rest (Figure 2). The test revealed a heterozygous variant in CACNA1C (NM_000719.6:c.2567G>A; p.Gly856Asp) that was initially classified as being of uncertain significance. Cosegregation analysis revealed that all 4 living siblings inherited the CACNA1C variant from the father (Figure 1).

Figure 2.

Resting Electrocardiogram of Youngest Brother (II-4) of the SCD Victim Carrying the CACNA1C:Gly856Asp Variant

ECG recorded at the age of 8 years showing a prolonged corrected QT interval (QTc). QTc was averaged over 5 beats using the threshold method (end of T) and calculated as 483 and 469 ms using the Bazett and Fridericia methods, respectively. Such measures are beyond the 99th percentile of QTc in the general population as determined by Vink et al.^1,6

The coroner’s investigation concluded a natural death of undetermined etiology. The cardiac examination performed by a cardiac pathologist revealed acute transmural ischemic injury of the right ventricle and atrium (Supplemental Figure 6), with normal coronary anatomy. There was a hemopericardium and extensive interstitial hemorrhage within the ischemic right atrium, possibly secondary to vigorous resuscitation maneuvers on the ischemic myocardium. The heart was otherwise normal. Neuropathology showed cerebral edema compatible with SCD. Postmortem genetic testing was performed (including arrhythmia and cardiomyopathy genes listed in the Supplemental Methods). The deceased sister also carried the familial CACNA1C variant. No other genetic variant was reported.

The CACNA1C gene codes for the main subunit of the cardiac voltage-gated calcium channel (Ca_V1.2), which underlies the inward L-type calcium current. Variants that disrupt inactivation of Ca_V1.2 cause LQTS type 8 by extending calcium entry into cardiomyocytes and therefore prolonging the action potential. LQTS type 8 classically presents with syndromic extracardiac features (known as Timothy syndrome) including syndactyly, and is caused by the recurrent Gly406Arg variant in CACNA1C or other less common variants (including Gly402Arg and Gly419Arg).2, 3, 4 LQTS type 8 can also present in isolation with no extracardiac features (ie, nonsyndromic LQTS type 8).³ Several genetic variants have been reported in nonsyndromic LQTS type 8 with a cluster in the cytoplasmic loop linking domains 2 and 3, more specifically in amino acid residues 857 to 860.⁵ The variant identified in the reported family (Gly856Asp) is mapped to this same mutational hotspot and was therefore considered likely causal of QT prolongation and SCD in the family. Nonetheless, considering the limited QT prolongation, we sought to explore the functional impact of the variant.

The functional properties of the variant were characterized following recombinant expression of the Ca_V1.2 Gly856Asp variant in HEK293T cells.⁴ As compared with the wild-type (WT) channel, the inactivation kinetics of the clinical variant were significantly slower and comparable with the archetypical Timothy Syndrome variant Ca_V1.2 Gly406Arg (Figure 3). Other biophysical properties of Ca_V1.2 Gly856Asp were unaffected in contrast to the gain-of-function in the activation voltage (negative shift) measured for Ca_V1.2 Gly406Arg under the same experimental conditions (Supplemental Table 2). The slower inactivation kinetics of Ca_V1.2 Gly856Asp could thus be a putative molecular mechanism accounting for the increased repolarization interval. Expanded methods regarding the site-directed mutagenesis, cell culture and transfection, electrophysiological measurements, and data analysis associated with in vitro functional analysis are available in the Supplemental Appendix.

Figure 3.

Functional Analysis of Ca_V1.2 Channels

(A) Whole-cell representative currents of cardiac voltage-gated calcium channel (Ca_V1.2) wildtype (WT), Ca_V1.2 Gly856Asp (G856D, variant identified in this presented family), and Ca_V1.2 Gly406Arg (G406R, a known Timothy syndrome variant) recombinant channels. Ca²⁺ current traces were activated by applying 450-ms pulses between −60 and +60 mV in 5-mV increments from a holding potential of −100 mV (see protocol in inset). (B) Averaged current-voltage relationship for Ca_V1.2 channels. Peak inward currents were normalized to cell capacitance for Ca_V1.2 current density and plotted against the pulse potentials. (C) Superimposed traces are shown to compare the inactivation kinetics. Current traces were induced from a holding potential of −100 mV to a testing potential of +5 mV for Ca_V1.2 WT (black), Ca_V1.2 G856D (red), or −5 mV for Ca_V1.2 G406R (blue) and normalized to their maximal amplitudes.

Management

The functional studies showing slowed inactivation kinetics support the CACNA1C:p.Gly856Asp variant as a cause for mild QT prolongation in this family and as a likely cause of SCD. Carriers of the CACNA1C:p.Gly856Asp variant were instructed to avoid use of QT-prolonging drugs and to seek urgent care in case of syncope. All 4 siblings of the sister with SCD were treated with beta-blockers, as recommended for LQTS more broadly. It remains unclear whether beta-blocker therapy is as effective in LQTS type 8 as in other LQTS types.

Discussion

This family exemplifies the importance of interdisciplinary investigation of SCD cases, including expertise in clinical cardiology, cardiogenetics, cardiac pathology, and basic electrophysiology. While the coroner’s investigation of SCD in the 14-year-old girl was ongoing, the family sought medical attention for nonspecific symptoms (palpitations). The QTc intervals in relatives carrying the CACNA1C variant ranged from 434 to 482 ms. Although such QTc values are certainly not diagnostic of LQTS in isolation, they should raise suspicion for LQTS in the context of family history of unexplained SCD. The diagnosis of LQTS is challenging because of dynamic QTc changes and the overlap between QTc distributions in the normal population and the population of patients with LQTS, with a majority of relatives with LQTS having a QTc within the normal range. Vink et al¹ showed that the mean QTc in a cohort of patients with LQTS was 459 ms for type 1 LQTS, 464 ms for type 2, and 447 ms for type 3. The authors suggest to diagnose LQTS using QTc thresholds adjusted for age, sex, QT measurement method (threshold vs tangent), and heart rate correction method. They provide an online tool⁶ to calculate the QTc percentile using a reference control population, recommending that a 99th percentile be used as a threshold to conservatively diagnose LQTS. In the present family, 3 of 5 carriers of the CACNA1C variant had a QTc beyond such a 99th percentile threshold.

The sister’s SCD is supportive of a diagnosis of LQTS. Her cardiac autopsy showed ischemic changes in the right atrium and ventricle. We suspect that such changes likely represent a combination of ischemia secondary to cardiac arrest in the setting of lethal arrhythmia and vigorous manual resuscitation. This highlights the importance of molecular genetic testing in cases where the etiology of SCD remains unclear even in presence of macroscopic or microscopic changes.⁷

Follow-up

Given the supporting functional and co-segregation data, the CACNA1C variant initially classified as a variant of uncertain significance has been upgraded to being likely pathogenic by the diagnostic laboratory. No other arrhythmic event occurred in the family during the first 3 years of follow-up.

Conclusions

We report a family of Central Asian ancestry with SCD and mild QTc prolongation caused by a novel gain-of-function genetic variant in CACNA1C, compatible with nonsyndromic LQTS type 8. The case highlights the value of interdisciplinary expertise when investigating complex SCD cases.

Funding Support and Author Disclosures

Dr Tadros is supported by the Philippa and Marvin Carsley Chair in Cardiology and holds the Canada Research Chair in Translational Cardiovascular Genetics. This work was completed in part with the operating grant 159556 from the Canadian Institutes of Health Research to Dr Parent. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose.