Boxer Cardiomyopathy (ARVC): Update on the Current Genetic Situation

Boxer cardiomyopathy (ARVC): update on the current genetic situation

Paul Wotton, Jo Dukes-McEwan, Hannah Copeland & Bruce Cattanach

Paul Wotton BVSc, PhD, DVC, MRCVS; University of Glasgow;

Acknowledgement: Dr. Kate Meurs kindly provided slides used in her presentation at the 2009 ACVIM Forum; a paper based on this work has been submitted for publication.

Cardiomyopathy in boxers

The first published description of ‘boxer cardiomyopathy’ (BCM) was in 1983 (Harpster1), and there have since been several clinical descriptions of this disease, mainly from the USA2-5. Three clinical categories of disease are classically described:

1)  ‘Concealed’ (occult or pre-clinical) disease: no signs, but arrhythmias present;

2)  ‘Overt’ disease: episodic syncope (usually with exertion or excitement) with arrhythmias. Various arrhythmias are seen, but VPCs with a left bundle-branch block (LBBB) pattern (implying right ventricular origin6) are common;

3)  ‘Myocardial dysfunction’ stage: congestive heart failure, esp. involving the left ventricle, resembling ‘classical’ dilated cardiomyopathy (DCM).

It is not known whether these 3 phases represent a continuum of the same disease, different variants of the same disease or different diseases altogether, but progression from 1) to 2) and sometimes 2) to 3) is seen.

Whereas in ‘classical’ DCM* histological changes in the myocardium are often mild or non-specific, out of proportion to the functional disturbances seen, the pathological and histological findings in BCM are striking, and involve7:

·  Right ventricular (RV) enlargement or aneurisms;

·  Early focal, later widespread, severe RV myocyte atrophy with replacement by fatty and fibro-fatty tissue, especially in the cranio-lateral and infundibular RV;

·  Focal fibro-fatty lesions are also present in both atria and the left ventricle;

·  Evidence of myocarditis may be seen especially in dogs that die suddenly, with patchy lysis, necrosis, haemorrhage and a mononuclear response.

(*DCM is probably not a single disease, but rather it probably represents a common phenotypic end-point for several pathological or genetic processes. Attenuated wavy fibres are seen histologically in a proportion of cases.)

Boxer cardiomyopathy is often referred to as arrhythmogenic right ventricular cardiomyopathy, ARVC, because of its resemblance to this condition in humans. This is also characterised by fibro-fatty replacement of the RV myocardium. It is familial, inherited as an autosomal dominant trait and a number of mutations have been identified, e.g. in the ryanodine receptor 2 and desmoplakin genes. Human ARVC results in ventricular tachyarrhythmias (with a LBBB morphology) and/or sudden unexpected death especially during exercise in young adults. Cardiomyopathies, including HCM & ARVC, are the most common causes of sudden death in young adults, especially athletes, when trauma and suicide are excluded. In some cases, however, CHF comes first8-9.

The search for a genetic marker in dogs

Much research effort, primarily by Meurs and associates, has been put into trying to identify a mutation or genetic abnormality linked to BCM, targeting especially the genes implicated in humans. Meurs & others10 looked at RV, IVS & LV myocardium from normal dogs and ARVC boxers and found that cardiac ryanodine receptor (RyR2) protein & RNA expression (implicated in ARVC2 in people) were lowest in the RV of normal dogs and were reduced in all chambers in ARVC dogs. This might explain the apparently increased susceptibility of the RV to an abnormality that affects the whole heart. However subsequent work failed to identify any genetic linkage with the RyR2 gene in cases of ARVC, and this may be a secondary rather than a primary abnormality.

Another research group11 has used immunofluorescence to look at the molecular composition of the intercalated disc structure in boxers with ARVC, as molecular remodeling of intercalated disc has been noted in human patients. Compared with control dogs, there was loss of staining for connexin43 at the intercalated discs. However gene sequencing failed to find mutations for desmoplakin, plakoglobin, connexin43, or plakophilin 2. Again, it is possible that these molecular changes might be secondary rather than primary. This group has also demonstrated ultrastructural changes in the desmosomes and gap junctions of affected dogs12.

Oyama and others13 have looked at the expression of the gene for calstabin2 (FKBP12.6), a regulatory molecule for RyR2 that stabilises its closed state in diastole, preventing calcium ‘leakage’. Diastolic leakage of calcium could trigger arrhythmias and reduces the efficiency of calcium cycling. They found that calstabin2 mRNA expression is significantly down-regulated in myocardium from ARVC boxers compared with healthy controls and dobermanns with DCM.

In the most promising development to date, Meurs and others14 have now identified (the following is quoted directly from the research abstract) ‘a 7 base pair deletion within a non-coding conserved element in a regulatory region of a calcium modulating gene (striatin) [was identified] and observed to be highly associated with the disease status (p<0.0001). The deletion was not observed in unaffected Boxer dogs or in any of 31 dogs from other breeds tested. Affected dogs were either homozygous or heterozygous for the deletion. Homozygous dogs (N=15) had a statistically greater number of VPC/24 hours than heterozygous (mean of 7860 and 2845, respectively) (p<0.05). We conclude that this 7 base pair deletion is associated with the development of Boxer ARVC and that dogs that are homozygous for the mutation appear to have a more severe degree of arrhythmias.’

The identification of this striatin gene mutation has resulted in widespread screening for this mutation in affected and apparently normal boxers. Unfortunately striatin genotype does not appear to correlate with phenotype in UK dogs, and similar findings have been made in other countries, including the USA. Thus it appears that the striatin gene mutation may merely enhance of modify expression of an ARVC gene, and further searches for a primary causative mutation are under discussion.

Diagnosis of BCM

As discussed by Meurs4, the diagnosis of BCM may be challenging, and may often be a diagnosis of exclusion. Changes in the criteria for diagnosis of ARVC in humans have recently been made due to a number of factors, including increasing recognition of the familial nature of the disease and early left ventricular involvement on some cases15. Holter ECG recordings are often regarded as a gold standard for clinical diagnosis, but considerable variability in arrhythmia frequencies has been found16. Serum levels of cardiac troponin I (cTnI) appeared to be a better marker for BCM than plasma BNP concentrations in work performed by Meurs’ group17, 18. The potential value of cTnI as a screening test for boxer cardiomyopathy in the UK, especially in trying to pick out the best dogs to target for closer examination (e.g. a Holter ECG), is currently being assessed, but has been frustrated by lack of follow-up/definitive diagnoses. Magnetic resonance imaging has been found to be useful in humans with ARVC19, especially for early detection of RV fatty infiltrates, but this has not so far been found to be useful in boxer dogs20.

References

1)  Harpster, N. (1983). Boxer cardiomyopathy. In: Kirk R, editor. Current Veterinary Therapy VIII. Philadelphia, WB Saunders. 329-337

2)  Wotton, P.R. (1999). Dilated cardiomyopathy (DCM) in closely related boxer dogs and its possibly resemblance to arrhythmogenic right ventricular cardiomyopathy (ARVC) in humans. Proceedings of the 17th Annual Veterinary Medical Forum, ACVIM, Chicago. p88-89

3)  Meurs, K.M. et al. (1999). Familial ventricular arrhythmias in boxers. J Vet Intern Med 14, 437-439

4)  Meurs, K.M. (2004). Boxer dog cardiomyopathy: an update. Vet Clin N Am Small Anim Pract 34, 1235-1244

5)  Baumwart, R.D. et al. (2005). Clinical, echocardiographic and electrocardiographic abnormalities in boxers with cardiomyopathy and left ventricular systolic dysfunction: 48 cases (1985-2003). J Am Vet Med Assoc 226, 538-541

6)  Kraus, M.S. et al. (2002). Morphology of ventricular arrhythmias in the Boxer as measured by 12-lead electrocardiography with pace-mapping comparison. J Vet Intern Med 16, 153-8

7)  Basso, C. et al. (2004). Arrhythmogenic right ventricular cardiomyopathy causing sudden cardiac death in boxer dogs. A new animal model of human disease. Circulation 109, 1180-1185

8)  Calabrese, F. et al. (2006). Arrhythmogenic right ventricular cardiomyopathy/dysplasia: is there a role for viruses? Cardiovascular Pathology 15, 11-17

9)  Corrado, D. et al. (2009). Arrhythmogenic right ventricular cardiomyopathy: an update. Heart 95, 766-773

10) Meurs, K.M. et al. (2006). Differential expression of the cardiac ryanodine receptor in normal and arrhythmogenic right ventricular cardiomyopathy canine hearts. Human Genetics 120, 111-118

11) Oxford, E.M. et al. (2007). Molecular composition of the intercalated disc in a spontaneous canine animal model of arrhythmogenic right ventricular dysplasia/cardiomyopathy. Heart Rhythm 4, 1196-1205

12) Oxford, E.M. et al. (2009). Phenotypic differences in the ultrastructure of cardiomyocytes from boxer dogs afflicted with arrhythmogenic right ventricular cardiomyopathy (ARVC). Proceedings ACVIM Forum 2009, p685

13) Oyama, M.A. et al. (2008). Arrhythmogenic right ventricular cardiomyopathy in Boxer dogs is associated with calstabin2 deficiency. J Vet Cardiol 10, 1-10

14) Meurs, K.M. et al. (2009). Genome-wide association identifies a mutation for arrhythmogenic right ventricular cardiomyopathy in the Boxer. Proceedings ACVIM Forum 2009, p684-685

15) Marcus, F.I. et al. (2010). Diagnosis of arrhythmogenic right ventricular cardiomyopathy/ dysplasia. Proposed modification of the task force criteria. Circulation 121 (ePub. ahead of publication)

16) Spier, A.W. & Meurs, K.M. (2004). Evaluation of spontaneous variability in the frequency of ventricular arrhythmias in Boxers with arrhythmogenic right ventricular cardiomyopathy. J Am Vet Med Assoc 224, 538-541

17) Baumwart, R.D. & Meurs, K.M. (2005). Assessment of plasma brain natriuretic peptide concentration in Boxers with arrhythmogenic right ventricular cardiomyopathy. Am J Vet Res. 66, 2086-2089

18) Baumwart, R.D., Orvalho, J. & Meurs, K.M. (2007). Evaluation of serum cardiac troponin I concentration in Boxers with arrhythmogenic right ventricular cardiomyopathy. Am J Vet Res. 68, 524-528

19) Conen, D. et al. (2006). Value of repeated cardiac magnetic resonance imaging in patients with suspected arrhythmogenic right ventricular cardiomyopathy. Journal of Cardiovascular Magnetic Resonance 8, 361-366

20) Baumwart, R.D. et al. (2009). Magnetic resonance imaging of right ventricular morphology and function in boxer dogs with arrhythmogenic right ventricular cardiomyopathy. J Vet Intern Med. 23, 271-4

APPENDIX: BCM DIAGNOSIS, SAMPLE SUBMISSION & CONSENT FORMS

To submit blood samples to LUPA the UK DNA archive form is available at the following weblink
http://www.liv.ac.uk/dna_archive_for_companion_animals/PDFDocs/More%20canine/New_Boxer_cardiomyopathy_form__doc.pdf

SAMPLES WITH THIS FORM can be sent directly to:

VETERINARY DNA ARCHIVE
Centre for Integrated Genomic Medical Research
University of Manchester
Stopford Building
Oxford Road
Manchester,
M13 9PT

EDTA blood samples and pedigrees may also be submitted with the form shown below to Dr. Bruce Cattanach (address on the form), or to Paul Wotton:

Dr. Paul Wotton BVSc, PhD, DVC, MRCVS

Small Animal Hospital

Faculty of Veterinary Medicine

University of Glasgow

Bearsden Road

Glasgow, G61 1QH

Confirmation of diagnosis form: ‘Boxer cardiomyopathy’

(upper section to be completed by owner)

Registered name …………………………………………………… Pet name…………………………………………

Date of birth…………………………… Sex (D/B)…………… Colour……………………………………......

Sire…………………………………………………………………………………………………………………………….

Dam……………………………………………………………………………………………………………………………

Copy of full pedigree provided – yes/no; Date (mth/yr) of final diagnosis………………………………..

Attending veterinary surgeon…………………………………………………………………………………......

Consulting cardiologist…………………………………………………………………………………………………….

Owner consent

I agree that stored blood samples may be used for genetic (DNA) research

I agree that the above information, along with relevant clinical details, may be shared with other researchers/geneticists for the study of cardiomyopathy in Boxers. I am aware that this information may be published, but without names of individual dogs or owners. The identities of individual dogs will be known to geneticists, breeders and cardiologist researchers to facilitate genetic counseling and to limit the impact of this disease on the breed in the future.

Owner name/address (please print) …………………………………………………………………………......

…………………………………………………………………………………………………………………………………

Telephone/email ……………………………………………………………………………………………………………

Signature……………………………………………………………………...... Date………………………

Confirmation of diagnosis (to be completed by cardiologist)

I confirm that I have examined the above-named Boxer and/or examined its records and that the clinical signs and results of diagnostic tests are consistent with a diagnosis of cardiomyopathy.

Cardiologist name/address (please print)………………………………………… ……………………………………

…………………………………………………………………………………………………………………………......

Telephone/email ……………………………………………………………………………………………………......

Major sign(s) …………………………………………………………………………………………………………….....

Criteria used for diagnosis (please circle); please add these details if possible/available - further details may be added overleaf if required:

Echo changes: abnormal RV ……………………….; abnormal LV ………………..……..; FS/EF low …….…%

Arrhythmia: ECG findings ………………………; Holter…………………………………; Post mortem? ………

Troponin: Result:………..ng/ml Laboratory used:………………………… Reference range:……………………

Signature……………………………………………………………. Date …………………………………………

CertSAC/CertVC/DVC/DipECVIM/other ………………… (please circle)

Please return the completed form to:
Dr BM Cattanach, Downs Edge, Reading Road, Harwell, Oxfordshire OX11 0JJ

Tel/Fax: 01235 835410; email