University of Groningen Phospholamban p.Arg14del cardiomyopathy te Rijdt, Wouter

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University of Groningen

Phospholamban p.Arg14del cardiomyopathy

te Rijdt, Wouter

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Publication date: 2019

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te Rijdt, W. (2019). Phospholamban p.Arg14del cardiomyopathy: Clinical and morphological aspects supporting the concept of arrhythmogenic cardiomyopathy. Rijksuniversiteit Groningen.

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Introduction and outline of the thesis

CHAPTER 1

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1.1 General introduction

Cardiomyopathies are a major cause of heart disease worldwide, aff ecting around 7.9 million people in all age groups.1,2_{In the Netherlands, the prevalence of cardiomyopathy is approximately}

40,000 (0.25%).3_{Cardiomyopathies comprise a group of heart disorders where a primary defect in}

the cardiac myocytes leads to abnormal structure and/or function of the myocardium. Therefore, other possible secondary causes (e.g. coronary artery disease, hypertension and valvular heart disease) must fi rst be excluded in order to make the diagnosis. The most common forms, classifi ed according to their phenotypic expression, are hypertrophic cardiomyopathy (HCM), dilated cardiomyopathy (DCM), and arrhythmogenic cardiomyopathy (ACM), but overlapping phenotypes do exist and are well recognized nowadays.4_{They can be further classifi ed into}

inherited forms and non-inherited forms (e.g. DCM due to alcohol abuse or HCM due to amyloidosis etc.). In inherited forms the inheritance pattern is most often autosomal dominant, but autosomal recessive, X-linked and mitochondrial inheritance patterns have been observed. Inherited cardiomyopathies are typically late-onset diseases: usual start of symptoms is in adult life although (severe) phenotypical expression in childhood has been described. In general, symptomatology in individuals carrying a cardiomyopathy-related mutation steadily increases until advanced age but some carriers will remain unaff ected, i.e. incomplete and age-related penetrance.

Since the discovery of the fi rst causative mutations for cardiomyopathies in the early 1990s, the importance of identifying the underlying genetic cause has been increasingly recognized.5,6_{Moreover, in the past decade the development and implementation of}

next-generation sequencing (NGS) techniques have allowed identifi cation of a still increasing number of pathogenic gene mutations related to human cardiomyopathies. These discoveries have led to a better understanding of disease pathogenesis and introduced genetic evaluation into clinical practice for aff ected individuals and their relatives via genetic cascade screening.6,7_{This allows}

early identifi cation of asymptomatic mutation carriers who may be at increased risk for sudden cardiac death even before the onset of any symptoms.

This thesis focuses on ACM caused by the non-desmosomal c.40_42delAGA (p.Arg14del) mutation in the phospholamban (PLN) gene.

1.2 Arrhythmogenic cardiomyopathy

ACM is a heritable myocardial disorder characterized by fi bro-fatty replacement of the myocardium that predisposes patients to ventricular arrhythmias and to slowly progressive ventricular dysfunction.8_{Sudden cardiac death may be the presenting symptom in up to 50% of index cases.}9

ACM encompasses a broad spectrum of disease that includes the classical right-dominant forms (ARVC, arrhythmogenic right ventricular cardiomyopathy), predominant left-sided involvement (also referred to as left-dominant arrhythmogenic cardiomyopathy (LDAC) and biventricular subtypes. The diagnosis is based on International Task Force Criteria (table1), which were modifi ed in 2010 but are in its current form still focused on the right-dominant form of ACM.10

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12 Chapter 1

The pathology, genetics, pathogenesis, translational aspects and the clinical utility of genetic testing in ACM will be illustrated and discussed extensively in chapter 2 and 3. Mutations in genes encoding proteins of the cardiac desmosome are found in up to 60% of ACM-cases.11,12

However, more recently, non-desmosomal genes have also been identified. One of these is the

PLN gene.

Table 1. 2010 revised Task Force criteria for the diagnosis of ARVCTable 1. 2010 revised Task Force criteria for the diagnosis of ARVC

Revised Task Force criteria I. Global or regional dysfunction and structural alterations*

Major By 2D echo: _{ Regional RV akinesia, dyskinesia, or aneurysm}

 and 1 of the following (end diastole):

- PLAX RVOT ≥32 mm (corrected for body size [PLAX/BSA] ≥19 mm/m2₎

- PSAX RVOT ≥36 mm (corrected for body size [PSAX/BSA] ≥21 mm/m2₎

- or fractional area change ≤33 percent

By MRI:

 Regional RV akinesia or dyskinesia or dyssynchronous RV contraction  and 1 of the following:

- Ratio of RV end-diastolic volume to BSA ≥110 mL/m2_{(male) or ≥100 mL/m}2_(female)

- or RV ejection fraction ≤40 percent

By RV angiography:

 Regional RV akinesia, dyskinesia, or aneurysm

Minor By 2D echo: _{ Regional RV akinesia or dyskinesia}

 and 1 of the following (end diastole):

- PLAX RVOT ≥29 to <32 mm (corrected for body size [PLAX/BSA] ≥16 to <19 mm/m2₎

- PSAX RVOT ≥32 to <36 mm (corrected for body size [PSAX/BSA] ≥18 to <21 mm/m2) - or fractional area change >33 percent to ≤40 percent

By MRI:

 Regional RV akinesia or dyskinesia or dyssynchronous RV contraction  and 1 of the following:

- Ratio of RV end-diastolic volume to BSA ≥100 to <110 mL/m2_{(male) or ≥90 to <100 mL/m}2

(female)

- or RV ejection fraction >40 percent to ≤45 percent

II. Tissue characterization of wall

Major  Residual myocytes <60 percent by morphometric analysis (or <50 percent if estimated), with fibrous _{replacement of the RV free wall myocardium in ≥1 sample, with or without fatty replacement of tissue}

on endomyocardial biopsy

Minor  Residual myocytes 60 percent to 75 percent by morphometric analysis (or 50 percent to 65 percent if _{estimated), with fibrous replacement of the RV free wall myocardium in ≥1 sample, with or without}

fatty replacement of tissue on endomyocardial biopsy

III. Repolarization abnormalities

Major  Inverted T waves in right precordial leads (V_{the absence of complete right bundle-branch block QRS ≥120 ms)}1, V2, and V3) or beyond in individuals >14 years of age (in Minor  Inverted T waves in leads V1 and V2 in individuals >14 years of age (in the absence of complete right

bundle-branch block) or in V4, V5, or V6

 Inverted T waves in leads V1, V2, V3, and V4 in individuals >14 years of age in the presence of complete

right bundle-branch block

IV. Depolarization/conduction abnormalities

Major  Epsilon wave (reproducible low-amplitude signals between end of QRS complex to onset of the T wave) _{in the right precordial leads (V}

1 to V3)

Minor  Late potentials by SAECG in ≥1 of the following 3 parameters in the absence of a QRS duration of ≥110 _{ms on the standard ECG}

 Filtered QRS duration (fQRS) ≥114 ms

 Duration of terminal QRS <40 µV (low-amplitude signal duration) ≥38 ms  Root-mean-square voltage of terminal 40 ms ≤20 µV

 Terminal activation duration of QRS ≥55 ms measured from the nadir of the S wave to the end of the QRS, including R', in V1, V2, or V3, in the absence of complete right bundle-branch block

V. Arrhythmias

Major  Nonsustained or sustained ventricular tachycardia of left bundle-branch morphology with superior axis _{(negative or indeterminate QRS in leads II, III, and aVF and positive in lead aVL)}

Minor  Nonsustained or sustained ventricular tachycardia of RV outflow configuration, left bundle-branch block _{morphology with inferior axis (positive QRS in leads II, III, and aVF and negative in lead aVL) or of}

unknown axis

 >500 ventricular extrasystoles per 24 hours (Holter)

VI. Family history

Major  ARVC/D confirmed in a first-degree relative who meets current Task Force criteria _{ ARVC/D confirmed pathologically at autopsy or surgery in a first-degree relative}

 Identification of a pathogenic mutation¶_{categorized as associated or probably associated with ARVC/D}

in the patient under evaluation

Minor  History of ARVC/D in a first-degree relative in whom it is not possible or practical to determine whether _{the family member meets current Task Force criteria}

 Premature sudden death (<35 years of age) due to suspected ARVC/D in a first-degree relative  ARVC/D confirmed pathologically or by current Task Force Criteria in second-degree relative

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III. Repolarization abnormalities

Major  Inverted T waves in right precordial leads (V1, V2, and V3) or beyond in individuals >14 years of age (in

the absence of complete right bundle-branch block QRS ≥120 ms)

Minor  Inverted T waves in leads V_{bundle-branch block) or in V}1 and V2 in individuals >14 years of age (in the absence of complete right

4, V5, or V6

 Inverted T waves in leads V1, V2, V3, and V4 in individuals >14 years of age in the presence of complete

right bundle-branch block

IV. Depolarization/conduction abnormalities

Major  Epsilon wave (reproducible low-amplitude signals between end of QRS complex to onset of the T wave) _{in the right precordial leads (V}

1 to V3)

Minor  Late potentials by SAECG in ≥1 of the following 3 parameters in the absence of a QRS duration of ≥110 _{ms on the standard ECG}

 Filtered QRS duration (fQRS) ≥114 ms

 Duration of terminal QRS <40 µV (low-amplitude signal duration) ≥38 ms  Root-mean-square voltage of terminal 40 ms ≤20 µV

 Terminal activation duration of QRS ≥55 ms measured from the nadir of the S wave to the end of the QRS, including R', in V1, V2, or V3, in the absence of complete right bundle-branch block

V. Arrhythmias

Major  Nonsustained or sustained ventricular tachycardia of left bundle-branch morphology with superior axis _{(negative or indeterminate QRS in leads II, III, and aVF and positive in lead aVL)} Minor  Nonsustained or sustained ventricular tachycardia of RV outflow configuration, left bundle-branch block _{morphology with inferior axis (positive QRS in leads II, III, and aVF and negative in lead aVL) or of}

unknown axis

 >500 ventricular extrasystoles per 24 hours (Holter)

VI. Family history

Major  ARVC/D confirmed in a first-degree relative who meets current Task Force criteria _{ ARVC/D confirmed pathologically at autopsy or surgery in a first-degree relative}

 Identification of a pathogenic mutation¶_{categorized as associated or probably associated with ARVC/D}

in the patient under evaluation

Minor  History of ARVC/D in a first-degree relative in whom it is not possible or practical to determine whether _{the family member meets current Task Force criteria}

 Premature sudden death (<35 years of age) due to suspected ARVC/D in a first-degree relative  ARVC/D confirmed pathologically or by current Task Force Criteria in second-degree relative

Diagnostic terminology for revised criteria:

Defi nite diagnosis: 2 Major, OR 1 Major and 2 Minor criteria, OR 4 Minor from diff erent categories Borderline diagnosis: 1 Major and 1 Minor, OR 3 Minor criteria from diff erent categories Possible diagnosis: 1 Major, OR 2 Minor criteria from diff erent categories

PLAX indicates parasternal long-axis view; RVOT: RV outfl ow tract; BSA: body surface area; PSAX: parasternal short-axis view; aVF: augmented voltage unipolar left foot lead; aVL: augmented voltage unipolar left arm lead. * Hypokinesis is not included in this or subsequent defi nitions of RV regional wall motion abnormalities for the proposed modifi ed criteria.

¶ A pathogenic variant is a DNA alteration associated with ARVC/D that alters or is expected to alter the encoded protein, is unobserved or rare in a large non-ARVC/D control population, and either alters or is predicted to alter the structure or function of the protein or has demonstrated linkage to the disease phenotype in a conclusive pedigree. # Adapted from Marcus et al. (2010)10

Introduction and outline of the thesis

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1.3 Phospholamban

Phospholamban, encoded by the PLN gene (locus 6q22.31; OMIM #172405), is expressed in the sarcoplasmic reticulum (SR) membrane as a 52 aminoacids 30-kD homopentameric phosphoprotein. PLN is a reversible regulator of the sarcoplasmic reticulum Ca2+_{-ATPase (SERCA)}

pump.13,14_{It alters the Ca}2+_{affinity of SERCA in cardiac muscle (SERCA2a isoform) by a mechanism}

that depends on PLN’s phosphorylation. PLN inhibits the calcium uptake by SERCA2a in the non-phosphorylated state. When non-phosphorylated, by cAMP-dependent protein kinase (at Ser16; i.e. via the beta-adrenergic pathway, figure 1) or Ca2+_{/calmodulin-dependent protein kinase II (at Thr17;}

predominantly during pathophysiological conditions), PLN dissociates from SERCA2a resulting in a higher activation with increased SR Ca2+_{uptake, accelerated relaxation, enhanced SR Ca}2+

load and increased Ca2+_{release during systole. In this way, dynamic PLN/SERCA interaction}

plays an important role in regulating intracellular calcium homeostasis and subsequent cardiac contractility and relaxation.

Figure 1. Function of phospholamban (PLN). PLN is a reversibly phosphorylated transmembrane protein, which binds to and regulates the activity of SERCA2a, the sarcoplasmic reticulum Ca2+_{-ATPase pump. (From: MacLennan}

et al. Nat Rev Mol Cell Biol. 2003)13

Chapter 1

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1.4 Mutations in the PLN gene

As to be expected because of the key role of PLN in cardiac calcium cycling and pathophysiology, genetic variants in the PLN gene may cause human inherited cardiomyopathies. However, the exact underlying mechanism(s) have not been fully elucidated yet. So far, fi ve naturally occurring mutations have been identifi ed in humans in the coding region of PLN (table 2).

The fi rst mutation described in the PLN gene was c.146T>G (p.(Val49Gly)) which results in potent inhibition of the Ca2+_{affi nity of SERCA2a. Cardiac overexpression of the p.(Val49Gly) mutant}

PLN in mice led to super-inhibition of cardiac contractility and remodeling, with progression to DCM, heart failure and early death.15

The second PLN mutation, c.25C>T (p.(Arg9Cys)), was shown to have no eff ects on SERCA2a activity under basal conditions but appeared to prevent phosphorylation of endogenous PLN, resulting in chronic inhibition of SERCA2a activity by PLN. Such chronic inhibition, never being able to draw on the full cardiac reserve, resulted in DCM and heart failure.16

The third mutation, c.116T>G (p.(Leu39*)), results in a truncated form of PLN without inhibitory eff ect on the calcium affi nity of SERCA2a. Heterozygous carriers had hypertrophy but no signs of cardiac contractile dysfunction. However, in homozygous carriers onset of DCM and heart failure during the teenage years was observed.17

The fourth PLN mutation described, c.40_42delAGA, resulting in a deletion of Arg-14 (p.Arg14del). The current knowledge and new insights regarding this mutation will be discussed in this thesis (starting from chapter 1.5).

The fi fth, most recently identifi ed, mutation c.73C>T p.Arg14del was found in a pedigree with DCM and malignant ventricular arrhythmia. Both SR Ca2+_{- uptake (super-inhibition of SERCA2a}

due to enhanced interaction between the p.(Arg25Cys) mutant PLN and SERCA2a) and SR Ca2+

-leak (increased due to increased Cam kinase II activity associated with hyper-phosphorylation of Serine 2814 in the ryanodine receptor) seem to be impacted in this mutation, leading to depressed myocyte contractile and Ca2+_{-kinetic parameters with increased arrhythmias.}18 Table 2. Overview of human PLN mutations and phenotypic characteristics

PLN, phospholamban; DCM, dilated cardiomyopathy; HF, heart failure; ACM, arrhythmogenic cardiomyopathy; VA, ventricular arrhythmia.

1

PLN mutation Phenotype Reference

c.146T>G, p.(Val49Gly) DCM, HF, premature death 15

c.25C>T, p.(Arg9Cys) DCM, HF, premature death 16

c.116T>G, p.(Leu39*) DCM, HF, hypertrophy 17

c.40_42delAGA,

p.(Arg14del) DCM, ACM, early myocardial fibrosis

19-25

c.73C>T, p.(Arg25Cys) DCM, VA 18

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1.5 The PLN p,Arg14del mutation

This mutation, a deletion of three nucleotides (c.40_42delAGA) in the coding region of the

PLN gene leading to deletion of amino acid arginine (p.Arg14del) in the PLN protein, was first

described in a large Greek family with hereditary DCM,19_{soon followed by others.}20,21_Heterozygous

carriers were shown to exhibit inherited DCM, both mild and severe forms were observed, showing episodic malignant ventricular arrhythmias and left-ventricular dilation with contractile dysfunction leading to overt heart failure in some cases.

Van der Zwaag et al. screened a large cohort of Dutch DCM and ARVC patients for PLN-mutations and found the pathogenic PLN p.Arg14del variant to be present in ≈15% of patients clinically diagnosed with idiopathic DCM and ≈12% of patients with ARVC.22_{Interestingly, a}

significant overlap between phenotypes was observed, in which left- or right-sided forms may predominate, compatible with the concept of ACM. Using haplotype analysis and postal code mapping, the pathogenic PLN p.Arg14del variant was characterized as a common Dutch founder mutation23_{(figure 2), but carriers have meanwhile also been identified in many other European}

countries (Germany, Belgium, Spain, the United Kingdom and Norway), Canada, and the USA. By now over 1000 carriers of the PLN p.Arg14del mutation have been identified (http://www. phorecast.nl), making this mutation the most prevalent single cardiomyopathy-related mutation identified in the Netherlands.

Figure 2. Postal code map illustrating the likely origin of the founder haplotype containing the PLN p.Arg14del mutation. The number of points based on the grandparents’ birthplaces is shown (in parenthesis: the number of postal code regions, 90 in total). On average, each region contains 180,000 inhabitants. The province of Friesland is enclosed by the bold border. (From: van der Zwaag et al. Neth Heart J. 2013; 21: 286-93.)

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17 Analogous to other inherited cardiomyopathies (see 1.1), the natural course of the disease is age- related (“age-related penetrance”); after a presymptomatic phase of variable length many carriers progress to overt disease, and are diagnosed with ACM. Symptomatic carriers were found to often exhibit a malignant arrhythmogenic phenotype, refl ected by high rates of VT/VF as a presenting symptom, appropriate ICD interventions, and a positive family history for premature sudden cardiac death (SCD).22_{Besides the high risk for malignant ventricular arrhythmias, a high}

risk of end-stage heart failure with subsequent high mortality was observed with a poor prognosis from late adolescence.24_{Left ventricular ejection fraction of <45% and sustained or nonsustained}

ventricular tachycardia were identifi ed as independent risk factors for malignant ventricular arrhythmias. Electrocardiographically, the p.Arg14del pathogenic variant is characterized by low amplitudes of the QRS complexes on the surface ECG and repolarization abnormalities (present in 41-46 % of carriers).21, 23, 24_{These ECG changes, which often occur early, are likely a refl ection of}

myocardial fi brosis.

Haghighi et al. extensively studied the eff ects of the PLN p.Arg14del pathogenic variant in an overexpression mouse model.19_{Transgenic mice overexpressing this variant exhibited}

depressed cardiac function, histopathological abnormalities (i.e. extensive myocardial fi brosis), and premature death, recapitulating the phenotype of the human mutation carriers. It remains, however, unclear how exactly the PLN p.Arg14del mutation leads to such severe cardiomyopathy and arrhythmia.

Experimental evidence from the mouse model has suggested a link between the PLN p.Arg14del mutation and impairment of cardiac Ca2+_{cycling: SERCA2a super inhibition due to a}

disturbance in the structure of PLN (partial destabilization of PLN’s pentameric structure, leading to the production of highly inhibitory monomers and a persistent PLN-SERCA association).19, 25_{As a}

result, phosphorylation is still possible but the inhibitory eff ect is no longer relieved. Coexpression of wild-type PLN and mutant PLN-R14Del in human embryonic kidney (HEK) cells, confi rmed the super inhibition of the SERCA channel. The dominant inhibitory eff ect of this mutant was not alleviated by phosphorylation, leading to myocellular calcium dysregulation, calcium overload, cardiomyocyte damage, and eventually to myocardial fi brosis.

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Aims and outline of this thesis

This thesis presents multiple clinical and morphological studies in PLN p.Arg14del mutation carriers. Considering the ever-growing number of mutation carriers identified and the importance of more detailed phenotyping PLN p.Arg14del cardiomyopathy to improve early diagnostic-, risk stratification- and treatment options, the aims of this thesis were:

To study the morphological characteristics and molecular signature of PLN p.Arg14del cardiomyopathy; and

To further elucidate the clinical phenotype using different imaging modalities; and

To evaluate the clinical implications of these findings on prognostication, i.e. follow-up study, and treatment, i.e. intervention trial.

Part I provides a general introduction. Chapter 2 and 3 describe the genetics, pathology, pathogenesis, translational aspects and the clinical utility of genetic testing in ACM.

Part II of this thesis focuses on the morphological features of phospholamban p.Arg14del cardiomyopathy. In Chapter 4 and 5 the characteristics and localization of PLN protein aggregates in complete heart, LV myocardial apex samples and right ventricular endomyocardial biopsy samples are shown, studied using both light- and electron microscopy. In Chapter 6 and 7 distinct pathological characteristics of PLN p.Arg14del cardiomyopathy are presented. In complete heart specimens we studied differential protein distribution patterns, using multiple immunohistochemical markers, and the fibrosis pattern in PLN p.Arg14del cardiomyopathy. In a selected group the presence of fibrosis was quantified using a high resolution systematic digital quantification technique.

Part III presents phenotypical insights into phospholamban p.Arg14del cardiomyopathy using different imaging modalities. In Chapter 8 the results of a large multicenter cardiac magnetic resonance (CMR) study, consisting of mainly presymptomatic mutation carriers, are described. Our main focus for this study was the presence of myocardial fibrosis in this group, and the association between myocardial fibrosis, electrocardiographical findings and the occurrence of ventricular arrhythmia. Chapter 9 presents a echocardiographic study in presymptomatic PLN p.Arg14del mutation carriers where we investigated whether subtle abnormalities in cardiac structure and function can already be observed in this group. In Chapter 10, the follow-up results of the CMR cohort (chapter 8) are shown. Our aim was to investigate whether the presence of late gadolinium enhancement on CMR is of Incremental prognostic value in early-stage phospholamban p.Arg14del cardiomyopathy.

Part IV, Chapter 11, part IV, describes the design and rationale of iPHORECAST (intervention in PHOspholamban RElated CArdiomyopathy Study). Cardiac fibrosis appears to be an early feature of PLN p.Arg14del cardiomyopathy, occurring in many presymptomatic mutation carriers before onset of overt disease. No proven treatment is available for this group. We designed and initiated iPHORECAST to demonstrate that pre-emptive treatment of presymptomatic PLN p.Arg14del mutation-carriers with eplerenone reduces disease progression and postpones onset of overt disease. The study has a multicenter, prospective, randomized, open-label, blinded endpoint (PROBE) design with a follow-up time of three years.

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References

McKenna WJ, Maron BJ, Thiene G. Classifi cation, Epidemiology, and Global Burden of Cardiomyopathies. Circ Res. 2017 Sep 15;121(7):722-30.

Global, regional, and national incidence, prevalence, and years lived with disability for 301 acute and chronic diseases and injuries in 188 countries, 1990-2013: a systematic analysis for the Global Burden of Disease Study 2013.”. Lancet 2013; 386 (9995): 743–800. doi:10.1016/s0140- 6736(15)60692-4 Brochure Hartfalen & Cardiomyopathie. Versie 4.7. Den Haag: Nederlandse Hartstichting; februari 2018. Watkins H, Ashrafi an H, Redwood C. Inherited cardiomyopathies. N Engl J Med. 2011 Apr 28;364(17):1643-56. Geisterfer-Lowrance AA, Kass S, Tanigawa G, Vosberg HP, McKenna W, Seidman CE, et al. A molecular basis for familial hypertrophic cardiomyopathy: a beta cardiac myosin heavy chain gene missense mutation. Cell. 1990 Sep 7;62(5):999-1006.

Jacoby D, McKenna WJ. Genetics of inherited cardiomyopathy. Eur Heart J. 2012 Feb;33(3):296- 304. Ackerman MJ, Priori SG, Willems S, Berul C, Brugada R, Calkins H, et al. HRS/EHRA expert consensus statement on the state of genetic testing for the channelopathies and cardiomyopathies: this document was developed as a partnership between the Heart Rhythm Society (HRS) and the European Heart Rhythm Association (EHRA). Europace. 2011 Aug;13(8):1077-109.

Corrado D, Link MS, Calkins H. Arrhythmogenic Right Ventricular Cardiomyopathy. N Engl J Med. 2017 Jan 5;376(1):61-72.

Dalal D, Nasir K, Bomma C, Prakasa K, Tandri H, Piccini J, et al. Arrhythmogenic right ventricular dysplasia: a United States experience. Circulation. 2005 Dec 20;112(25):3823-32.

Marcus FI, McKenna WJ, Sherrill D, Basso C, Bauce B, Bluemke DA, et al. Diagnosis of arrhythmogenic right ventricular cardiomyopathy/dysplasia: proposed modifi cation of the Task Force Criteria. Eur Heart J. 2010 Apr;31(7):806-14.

Groeneweg JA, van der Heijden JF, Dooijes D, van Veen TA, van Tintelen JP, Hauer RN. Arrhythmogenic cardiomyopathy: diagnosis, genetic background, and risk management. Neth Heart J. 2014 Aug;22(7-8):316-25.

Lazzarini E, Jongbloed JD, Pilichou K, Thiene G, Basso C, Bikker H, et al. The ARVD/C genetic variants database: 2014 update. Hum Mutat. 2015 Apr;36(4):403-10.

MacLennan DH, Kranias EG. Phospholamban: a crucial regulator of cardiac contractility. Nat Rev Mol Cell Biol. 2003 Jul;4(7):566-77.

Young HS, Ceholski DK, Trieber CA. Deception in simplicity: hereditary phospholamban mutations in dilated cardiomyopathy. Biochem Cell Biol. 2015 Feb;93(1):1-7.

Haghighi K, Schmidt AG, Hoit BD, Brittsan AG, Yatani A, Lester JW, et al. Superinhibition of sarcoplasmic reticulum function by phospholamban induces cardiac contractile failure. J Biol Chem. 2001 Jun 29;276(26):24145-52.

Schmitt JP, Kamisago M, Asahi M, Li GH, Ahmad F, Mende U, et al. Dilated cardiomyopathy and heart failure caused by a mutation in phospholamban. Science. 2003 Feb 28;299(5611):1410-3.

Haghighi K, Kolokathis F, Pater L, Lynch RA, Asahi M, Gramolini AO, et al. Human phospholamban null results in lethal dilated cardiomyopathy revealing a critical diff erence between mouse and human. J Clin Invest. 2003 Mar;111(6):869-76.

Liu GS, Morales A, Vafi adaki E, Lam CK, Cai WF, Haghighi K, et al. A novel human R25C- phospholamban mutation is associated with super-inhibition of calcium cycling and ventricular arrhythmia. Cardiovasc Res. 2015 Jul 1;107(1):164-74.

Haghighi K, Kolokathis F, Gramolini AO, Waggoner JR, Pater L, Lynch RA, et al. A mutation in the human phospholamban gene, deleting arginine 14, results in lethal, hereditary cardiomyopathy. Proc Natl Acad Sci U S A. 2006 Jan 31;103(5):1388-93.

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DeWitt MM, MacLeod HM, Soliven B, McNally EM. Phospholamban R14 deletion results in late- onset, mild, hereditary dilated cardiomyopathy. J Am Coll Cardiol. 2006 Oct 3;48(7):1396-8.

Posch MG, Perrot A, Geier C, Boldt LH, Schmidt G, Lehmkuhl HB, et al. Genetic deletion of arginine 14 in phospholamban causes dilated cardiomyopathy with attenuated electrocardiographic R amplitudes. Heart Rhythm. 2009 Apr;6(4):480-6.

van der Zwaag PA, van Rijsingen IA, Asimaki A, Jongbloed JD, van Veldhuisen DJ, Wiesfeld AC, et al. Phospholamban R14del mutation in patients diagnosed with dilated cardiomyopathy or arrhythmogenic right ventricular cardiomyopathy: evidence supporting the concept of arrhythmogenic cardiomyopathy. Eur J Heart Fail. 2012 Nov;14(11):1199-207.

van der Zwaag PA, van Rijsingen IA, de Ruiter R, Nannenberg EA, Groeneweg JA, Post JG, et al. Recurrent and founder mutations in the Netherlands-Phospholamban p.Arg14del mutation causes arrhythmogenic cardiomyopathy. Neth Heart J. 2013 Jun;21(6):286-93.

van Rijsingen IA, van der Zwaag PA, Groeneweg JA, Nannenberg EA, Jongbloed JD, Zwinderman AH, et al. Outcome in Phospholamban R14del Carriers: Results of a Large Multicentre Cohort Study. Circ Cardiovasc Genet. 2014 7(4):455-65.

Haghighi K, Pritchard T, Bossuyt J, Waggoner JR, Yuan Q, Fan GC, et al. The human phospholamban Arg14-deletion mutant localizes to plasma membrane and interacts with the Na/K- ATPase. J Mol Cell Cardiol. 2012 Mar;52(3):773-82. 20 21 22 23 24 25 Chapter 1

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