Genetic screening in heritable thoracic aortic disease—rationale, potentials and pitfalls

Abstract

Thoracic aortic aneurysms are silent yet deadly clinical entities which may elude detection until an acutely life-threatening aortic dissection or rupture occurs. Approximately 20% of patients with thoracic aortic aneurysms or dissection have a positive family history, indicating a strong genetic component to the aetiology. Genetic screening in such hereditary thoracic aortic disease (HTAD) may thus be beneficial in detecting causative genetic mutations in affected patients, identifying asymptomatic family members who may be at risk, and in guiding the optimal timing of preventative surgery in those with confirmed genetic aortopathy. Genetic screening can facilitate personalised aortic care tailored to an individual’s specific genetic abnormality, with the aim of mitigating the significant morbidity burden and premature mortality associated with HTAD. This review examines the rationale for genetic screening in HTAD, its potential applications, current limitations and potential future directions.

Thoracic aortic aneurysms: a silent killer

Thoracic aortopathy exists on a spectrum ranging from mild aortic dilatation and aneurysmal expansion to acutely life-threatening aortic syndromes comprising penetrating aortic ulcer, intra-mural haematoma and acute dissection. As dictated by their natural history, progressive dilatation of a thoracic aortic aneurysm (TAA) may eventually culminate in catastrophic acute dissection or rupture if left untreated, prior to which only 5% of affected individuals will have forewarning symptoms. Indeed, between 2.0 and 7.3% of sudden cardiac deaths in the general population have been attributed to aortic dissection and rupture [1–3], emphasising the “silent killer” nature of TAAs. Ninety-five percent of patients with non-syndromic TAAs are clinically silent entities, only manifesting after achieving very large dimensions, and in the vast majority, death is unfortunately the first symptom [4, 5]. Insights from family history studies conducted by multiple investigators over the last two decades have revealed a hereditary pattern of acquisition in some TAAs. Genetic aetiology is associated with significantly faster aortic dilatation, and aneurysms in patients with familial TAAs grow at 0.22 cm/year, whilst rapid expansion rates exceeding 1.0 cm/year are seen in the aggressive Loeys-Dietz syndrome [5, 6].

The silent, unpredictable and deadly nature of TAAs mandates their proactive identification as early as possible along the course of their natural history, in individuals who may be genetically susceptible to their development, along with screening of at-risk family members, to mitigate the significant related morbidity and mortality and thus afford them the best possible opportunity for treatment and survival.

The importance of pre-emptive diagnosis

Whilst TAAs are prevalent in approximately 1% of the general population [7] and considering their potentially life-threatening consequences, their pre-emptive detection represents a significant challenge despite our accruing knowledge of their natural history and underlying pathophysiology, alongside advancements in modern diagnostic imaging. Although timely recognition and treatment are of great importance, to date, no single, widely available screening test exists for early diagnosis, which is further hampered by their indolent growth preceding an aortic event. Thoracic aneurysms associated with genetic aortopathies are however associated with faster growth rates [5, 6].

TAAs are frequently diagnosed incidentally on radiological studies performed for an unrelated presentation. However, the presence of specific clinical signs and associated conditions can assist diagnosis in individuals harbouring an as yet asymptomatic aneurysm, or those susceptible to developing one in the future [8]. These include family history of aortic disease, bicuspid aortic valve, intra-cranial aneurysm, simple renal cysts, abdominal aortic aneurysm, a positive thumb-palm sign and temporal arteritis [8]. Approximately 20% of patients with TAAs have a positive family history, indicating a significant genetic component to aetiology [9]. Contemporary genetic testing can distinguish genetic mutations which may alter natural history processes, predict risk of earlier onset of aortic dissection at smaller diameters and thereby guide decision-making to distinguish patients at higher genetic risk who might benefit from earlier surgery to prevent an untimely death from lethal aortic events. Knowledge of an underlying genetic syndrome may also be of benefit in predicting the onset of related non-aortic conditions which may have not yet manifested clinically, and planning their management.

Genetic perspectives in TAAs

Whilst conventional cardiovascular risk factors such as smoking, hypertension, hypercholesterolaemia and atherosclerosis play a central role in the development of more distal aortic disease in older patients, aortopathy manifesting through aneurysm formation or dissection at a younger age, in conjunction with a positive family history of aortic events, should prompt clinicians to the strong possibility of an underlying genetic trigger.

Over the last two decades, a great deal of effort has been directed to studying the genetic defects associated with heritable thoracic aortic disease and it is now well-recognised that several genetic syndromes exist which predispose to aortic aneurysm formation and can initiate aortic dissection in the absence of classical risk factors. To date, over 30 contributory genes have been identified, encoding for proteins involving the extra-cellular matrix (FBN1, FBN2, COL3A1, EFEMP2, ELN, EMILIN1, MFAP5, LOX, BGN), vascular smooth muscle cell contraction (MYH11, ACTA2, MYLK, PRKG1, FLNA, MAT2A, FOXE3) and transforming growth factor-ß (TGF-ß) signalling pathways (TGFRB1, TGFBR2, TGFB2, TGFB3, SMAD, SKI, SLC2A10) [10, 11].

Thoracic aortic aneurysm and dissection (TAAD) may be classified as syndromic or non-syndromic [12]. Syndromic aortopathy (Table 1) accounts for 5% of all TAADs and implies involvement of extra-aortic organ systems, whereas non-syndromic forms (Table 2) comprise 95% of all TAAD and are restricted to the aorta [10, 12, 13, 15]. Significant overlap can nevertheless exist between syndromic and non-syndromic TAAD variants. Non-syndromic TAAD may be further categorised as sporadic, which encompasses 80% of all thoracic aortic disease [6], or familial when more than one family member carries an aneurysm. Familial TAAs are often inherited in an autosomal dominant pattern, although with incomplete penetrance and variable expressivity, present at a younger age and grow faster than their non-familial counterparts [9, 16]. Owing to their lack of distinctive phenotypic characteristics, patients with non-syndromic thoracic aortopathy are at risk of delayed diagnosis and subsequent silent aneurysm formation prior to a dissection event. A high index of suspicion should be maintained and screening is thus of paramount importance in this vulnerable group.

Table 1.

Genes associated with syndromic thoracic aortic aneurysm and dissection [10, 13, 14]

Syndrome	Genes	Protein	Inheritance pattern
Marfan	FBN1	Fibrillin 1	Dominant
Loeys-Dietz	TGFBR2 TGFBR1	TGFß-R2 TGFß-R1	Dominant
Ehlers-Danlos	COL3A1	Type III collagen	Dominant
Arterial tortuosity syndrome	SLC2A10	GLUT10	Recessive
Aneurysm-osteoarthritis syndrome	Smad3	SMAD3	Dominant
Turner	45 XO or 45 XO mosaicism	–	Sex-linked

Table 2.

Genes associated with non-syndromic thoracic aortic aneurysm and dissection [10, 13, 14]

Genes	Protein	Inheritance
MYH11	Smooth muscle cell myosin heavy chain	Dominant
MYLK	Myosin light-chain kinase	Dominant
ACTA2	Smooth muscle α-actin	Dominant
MFAPS	Microfibril-associated glycoprotein 2	Recessive
MAT2A	Methionine adenosyl-transferase II alpha	Dominant
PRKG1	Type I cGMP-dependent protein kinase	Dominant
FOXE3	Forkhead box 3	Dominant
LOX	Lysyl oxidase	Dominant
NOTCH1	NOTCHI	Dominant

Syndromic heritable thoracic aortic disease

The most prevalent forms of syndromic TAAs are those related to Marfan syndrome (MFS), Loeys-Dietz syndrome (LDS), and vascular Ehlers-Danlos syndrome [17]. Prognosis in syndromic TAAs is generally worse than in non-syndromic cases, as reflected in guideline recommendations for prophylactic aortic surgery at more conservative diameters than the usual 5.0–5.5 cm cut-off [18, 19]. The diameter recommendations for ascending aortic intervention in syndromic aortopathy are shown in Fig. 1. Decision on a case-by-case basis is important, accounting for additional factors such as patient preference, age, body size, family history and aneurysm configuration.

Fig. 1.

Recommendations for ascending aortic intervention based on genetic diagnosis [20]. Abbreviations: ECM, extra-cellular matrix; SMC, smooth muscle cell; TAAD; thoracic aortic aneurysm and/or dissection; TGF, transforming growth factor

Other genetic syndromes affecting the aorta include Turner syndrome (partial or complete monosomy of the X chromosome), arterial tortuosity syndrome (SLC2A10 gene mutation) and aneurysm-osteoarthritis syndrome (SMAD3 gene mutation), amongst others [19].

MFS is an autosomal dominant connective tissue disorder with high penetrance caused by pathogenic mutations in the FBN1 gene located on chromosome 15q15-31, leading to abnormalities of the fibrillin-1 protein within the extra-cellular matrix and resultant loss of elasticity and structural integrity of connective tissue. It accounts for 5% of all aortic dissections [21]. Besides aortic root aneurysm and aortic dissection, additional cardiovascular features of MFS include mitral valve prolapse, aortic insufficiency, pulmonary artery dilatation and ventricular dysfunction [10, 22, 23]. Other clinical features are ocular lens luxation, and skeletal manifestations such as arachnodactyly, pectus deformity, scoliosis, pes planus and increased arm span. Diagnosis is based on the revised Ghent criteria with emphasis on genetic testing of the FBN1 gene, family history and ocular, cardiovascular and skeletal features [24]. Replacement of the aneurysmal aorta is generally recommended in MFS at aortic diameters exceeding 5 cm [18, 19]. Even with optimal medical management, 90% of patients with MFS will require aortic surgery or develop aortic dissection in their lifetime [25].

LDS is an autosomal dominant connective tissue disorder sharing some features with Marfan syndrome, and characterised by arterial aneurysms and tortuosity, marfanoid habitus and distinctive cranio-facial anomalies such as hypertelorism, bifid uvula or cleft palate [26]. Although arteriopathy in LDS extends beyond the aorta, the rate of TAA growth and attendant complications is particularly worrying. TAA growth rates exceeding 1.0 cm/year have been observed in LDS, 10 times faster than those in MFS [5]. In one series, mean age of death amongst 90 patients with LDS was 26 years, with TAAD accounting for mortality in 67% [27]. The degree of cranio-facial dysmorphism in LDS can help to identify those at greater risk for early-onset arteriopathy, which is more aggressive than in MFS [27]. Five different subtypes exist, caused by mutations in TGFBR1 (type I) and TGBR2 (type II), SMAD3 (type III), TGFB2 (type IV) and TGFB3 (type V) [10, 23]. Current aortic guidelines recommend prophylactic aortic replacement at a diameter of 4.2 cm given the extremely high dissection risk associated with types I–III [18, 19].

Ehlers-Danlos syndrome (EDS) type IV (vascular type) is an autosomal dominant connective tissue disorder with high penetrance caused by mutations in the COL3A1 gene on chromosome 2q32.2 encoding type III procollagen [28, 29]. Delayed formation and destabilisation of the collagen triple helix and its impaired secretion results in significant vascular, cutaneous and hollow organ tissue fragility [29]. Affected individuals have thin, translucent skin, extensive bruising and characteristic facial appearances. Aortic dissection and rupture may occur without preceding aneurysm formation [28, 29]. The mean age for the first major arterial or gastrointestinal complication is 23 years, and median survival has been reported at 48 years, with 79% dying from arterial rupture, and the remainder from rupture of the uterus, left ventricle, liver, spleen or bowel [29]. Clinical diagnosis of vascular EDS is based on the revised Villefranche criteria [30] and non-invasive imaging, and confirmed by detection of COL3A1 mutation on genetic testing or protein-based testing via skin biopsy demonstrating abnormal type III procollagen [17, 28]. Due to its rarity and in the absence of family history, diagnosis is often made concurrent to a rupture event. Whilst surgery is extremely hazardous owing to vessel frailty, current aortic guidelines support ascending aortic replacement at a diameter of 5.0 cm [18, 19].

The significant phenotypic overlap between MFS, LDS and vascular EDS, along with their differing treatment options and prognosis, underpins the great importance of their accurate diagnosis.

Non-syndromic heritable thoracic aortic disease

The majority of patients with TAAD do not have a well-defined genetic syndrome and lack external pathognomonic features, but nevertheless may inherit a predisposition for TAAD. These non-syndromic TAAs may be classified as sporadic where only one person in a family harbours an aneurysm, representing 80% of all cases. Alternatively, they may occur as a heterogeneous group of aortopathies known as familial thoracic aortic aneurysms (FTAA), in which more than one family member is affected [11]. These are characterised by an autosomal dominant pattern of inheritance, variable clinical expressivity in terms of age of disease onset, aneurysm location and extent of aortic dilatation prior to dissection, and reduced penetrance [9, 31].

To date, mutations in over 9 contributory genes have been identified that predispose to FTAA. Mutations in the ACTA2 gene located on chromosome 10q23.31 encoding for smooth muscle–specific α-actin, account for 10–15% of all FTAA [32], and have been identified in patients with TAAD also presenting with coronary artery disease and stroke [33]. Early aortic replacement is advised for aneurysmal disease in patients with ACTA2 mutations, in whom the majority of aortic aneurysms were < 5.0 cm prior to dissection [17]. The MYH11 gene located on chromosome 16p13.11 encodes for smooth muscle–specific myosin heavy chain. Mutations in MYH11 account for 2% of non-syndromic TAAD [17], and are also associated with patent ductus arteriosus [34]. Acute aortic dissection has been known to occur with little or no pre-existing aortic enlargement in mutations of the MYLK gene on chromosome 3q21.1 which encodes myosin light-chain kinase, and accounts for 1% of FTAA. Surgical intervention is advised at aortic diameters of 4.5 cm for patients with MYLK mutations [20, 35]. PRKG1 is a gene found on chromosome 10 encoding a type 1 cyclic guanosine monophosphate–dependent protein kinase that regulates smooth muscle relaxation. Mutations in PRKG1 are associated with aortic dissection occurring at as young an age as 16 years, in addition to coronary artery aneurysms and dissection [36], although there is no consensus regarding the specific timing of prophylactic aortic repair.

Recommended aortic diameters for surgical intervention on the ascending aorta in non-syndromic aortopathy are shown in Fig. 1.

The rationale for genetic screening

Aortic dissection and rupture are devastating events which signify the endpoint of the uninterrupted and insidious natural history of TAAs. They impact a broad spectrum of patients with a great morbidity and mortality burden. Untreated, one-third of patients die within the first 24 h following an acute ascending aortic dissection and 50% die within 48 h [37]. Since the progressive and unchecked aneurysmal dilatation of the thoracic aorta can eventually culminate in these deadly complications, size-based criteria have been established in existing aortic guidelines which, for many years since their introduction, have served as a primary guide when selecting patients for prophylactic aortic repair. However, as is now a well-recognised phenomenon, aortic dissection or rupture may occur at significantly smaller diameters than those proposed for elective aneurysm surgery [38], especially in those with genetic predilection. Twenty percent of patients with non-syndromic TAAs have affected first-degree relatives, substantiating the notion that an underlying genetic mutation is implicated in many patients with TAA without a defined genetic syndrome [9]. Negative family history does not preclude genetic positivity. Whilst strict anti-hypertensive therapy is crucial in controlling this modifiable risk factor, aortic dissection may occur in younger populations without traditional cardiovascular risk factors, where an undisclosed genetic lesion should be highly suspected.

Thus, genetic screening could provide valuable information regarding the presence, or absence, of the aforementioned recognised mutations responsible for thoracic aortic disease (TAD), which in turn may influence treatment by stratifying risk, guiding the type of surgical repair to be performed, and aiding in prognostication. Once a genetic diagnosis is achieved in an individual, genetic testing can be extended to other family members, who may be asymptomatic, to assess their own risk from carrying the same genetic mutation. Genetic analysis could also enhance the discovery, and elucidate the contributory role, of novel mutations in the development of aortic disease. Ultimately, genetic screening can open avenues for personalised aortic care and specifically targeted interventions tailored according to the individual’s genetic abnormality. Genetic screening can help to identify high-risk patients allowing their close monitoring in dedicated surveillance programmes, and inform the appropriate timing of surgical intervention with the goal of decreasing morbidity and premature mortality from catastrophic aortic events.

Methods for genetic testing in HTAD

Gene panel testing is a useful and economical tool for identification of personal risk of hereditary thoracic aortic disease (HTAD) in probands, and should be performed under the supervision of a cardiologist or a clinical geneticist with experience in cardiovascular genetics and aortopathy. The correct interpretation of genetic tests in HTAD is important since diagnosis greatly influences the medical approaches for cardiovascular risk factor modification, and effectively dictates the timing of surgical intervention for aneurysm resection. Expert comprehension of the underlying genetics is crucial since the heterogeneity of pathologies that can be identified from genetic testing can hamper effective clinical management.

A positive genetic test indicates either a pathogenic or likely pathogenic (P/LP) variant, whilst a negative test signifies that no pathogenic genetic mutations are detected or only benign or likely benign (B/LB) variants are found. Detection of a P/LP variant may prompt additional analysis to test for segregation with TAAD. Individuals with a clearly syndromic form of HTAD who receive a negative genetic test should not be excluded from ongoing surveillance and management according to their presentation and family history. Occasionally, a variant of unknown significance (VUS) is identified, for which the paucity of available scientific data on its virulence means it cannot be assigned as either P/LP or B/LB. There is uncertainty amongst physicians as to whether they should disclose information about VUS to their patients, many of whom may not wish to be informed themselves [39].

With over 30 currently known genes predisposing to HTAD, the simultaneous panel testing of multiple genes by comprehensive whole exome sequencing (WES) is considered the gold standard technique [40]. It can diagnose both syndromic HTAD and approximately 30% of individuals with familial non-syndromic HTAD who will also test positive, suggesting that many more causative genes have yet to be discovered [20]. As opposed to performing multiple tests for individual genetic syndromes across various geographically distributed laboratories, often at a substantial cumulative cost, WES examines the entire exome for every patient for all mutations currently associated with HTAD and may represent a more cost-effective approach to genetic screening due to its high-throughput nature [40]. In a study by the Yale group, WES demonstrated a high “yield” for the identification of genetic mutations known to cause TAAD, with approximately 25% of patients undergoing WES noted to have deleterious genetic mutations [40]. Non-affected relatives with negative exome screening can be reassured in the knowledge that their risk of developing HTAD is very low or absent. WES also permits the genetic data to be retrieved for future examination as novel genes implicated in genetic aortopathy are discovered alongside advancements in the field [10], or retrospectively with greater scrutiny in those with aortic pathology but without genetic mutations detected on initial screening [40]. The Yale group are putting significant efforts into compiling a “dictionary” of genes linked to HTAD using WES in the clinical setting, which will serve as a highly valuable adjunct in genetic diagnosis [40, 41]. WES thus possesses enormous potential for the personalisation of aortic care, both currently and in the future, as our understanding of the genetics of aortic disease progresses further, and as exciting surgical innovations emerge.

Sanger single-site sequencing is a less complex and cheaper test that can be utilised for the more focused detection of harmful mutations in single genes for individuals with smaller TAAs who are first-degree relatives of probands carrying the same mutation [10]. It is often applied in other family members to sequence a culprit gene following its identification as an anomalous variant in a proband through WES. Multi-gene panel testing employs next-generation sequencing, whilst high-density targeted arrays are utilised when searching for specific deletions or duplications. Laboratory turnaround times are reported at 2–10 weeks, with an average of 4 weeks [42]. Patients should be informed as part of their pre-test counselling that mutation analysis is not 100% sensitive owing to the small possibility of technical and human error [42].

Genetic testing in HTAD is usually performed on deoxyribonucleic acid (DNA) extracted from the patient’s blood, although cell-based methods allow testing from a saliva sample or cheek swab.

When should genetic testing be offered?

Patients with (i) a medical history or physical examination compatible with syndromic HTAD; (ii) family history of TAD, sudden death or family members with syndromic features; and (iii) onset of TAD at age < 65 years especially in the absence of cardiovascular risk factors [42]. Combined involvement of the aortic root and ascending aorta, or arch and descending aorta, should also prompt genetic investigation. Patients with abnormal histopathological findings on the aorta, especially cystic medial degeneration, should be offered genetic testing as well.

Who should undergo genetic screening?

Factors associated with a genetic predilection for TAD include larger thoracic aortic aneurysms, an absence of traditional cardiovascular risk factors, positive family history and the presence of syndromic features. Non-genetic factors have been suggested as the major contributors to TAA with diameter < 4.5 cm, whereas a greater genetic role has been postulated in more severe presentations of aortic disease, such as aneurysms larger than 4.5 cm in diameter [16]. Besides genetic causes, TAAs may also be attributed to acquired factors, including increasing age, cardiovascular disease (systemic hypertension, dyslipidaemia, atherosclerosis), infectious disease (syphilis, tuberculosis) and autoimmune disease (giant cell arteritis, Takayasu arteritis, Behçet’s disease) and lifestyle (pregnancy, smoking, heavy weight-lifting, cocaine use). The likelihood of encountering a pathogenic mutation decreases with advancing age, and familial disease tends to present earlier [9, 43]. Thus, a genetic component should be suspected with TAD occurring in patients younger than 50 years of age irrespective of the presence of hypertension, and in hypertensive patients aged 50–60 years [16]. Positive family history describes when at least one first-degree relative has a thoracic aortic aneurysm or dissection, and greatly increases the likelihood of an underlying genetic aetiology. Similarly, the presence of characteristic syndromic features in TAD is strongly associated with a pathogenic gene mutation.

Genetic testing in HTAD may be offered to achieve a diagnosis in those who already have features of the disease or following an aortic event, or in a predictive capacity to determine which asymptomatic individuals might experience disease progression. Patients suspected to have HTAD should undergo a thorough clinical workup comprising (i) analysis of medical and family history; (ii) cardiological assessment with echocardiographic evaluation of the aortic annulus, aortic root and ascending aortic diameters, aortic valve morphology and function, mitral valve involvement and global cardiac size and function; (iii) cross-sectional imaging evaluation dependent on echocardiographic findings; and (iv) ophthalmology evaluation to confirm ocular manifestations of syndromic aortic disease [23]. Genetic testing is preferably performed after completion of the above clinical assessments.

Since the majority of HTAD exhibits an autosomal dominant inheritance pattern conferring a 50% chance of acquiring the genetic lesion, the European Society of Cardiology (ESC) guidelines recommend that first-degree relatives (parents and siblings) of individuals affected by HTAD are investigated to exclude a familial form, and offered genetic counselling and molecular testing if a familial variant is highly suspected (class 1, level of evidence C) [19]. The ESC guidelines also recommend screening of healthy, at-risk relatives every 5 years until a clinical or molecular diagnosis is either confirmed or excluded (class 1, level of evidence C) [19].

The American College of Cardiology (ACC)/American Heart Association (AHA) recommend genetic counselling and testing for first-degree relatives of patients with FBN1, TGFBR1, TGFBR2, COL3A1, ACTA2, and MYH11 associated with aortic aneurysm or dissection (class 1, level of evidence C) [18]. They also propose that testing for ACTA2 mutation is reasonable in patients with family history of TAAD to determine whether this is responsible for the aortopathy (class IIa, level of evidence B) [18]. Sequencing of other genes (TGFBR1, TGFBR2, MYH11) may be considered in patients with a family history of TAAD and clinical features associated with these mutations, and referral to a geneticist may be considered if ≥ 1 first-degree relative of the affected patient with TAAD is found to have thoracic aortic dilatation, aneurysm or dissection (class IIb, level of evidence C) [18].

The Canadian Cardiovascular Society (CCS) recommends genetic screening for Marfan syndrome when suspected to confirm the diagnosis, and for genes associated with TAD in non-bicuspid aortic valve (BAV) aortopathy cases [44]. They also recommend genetic counselling and testing in first-degree relatives of patients with an identified causal mutation of a TAD-associated gene [44].

The Japanese Circulation Society have published guidelines for the diagnosis and treatment of aortic aneurysm and aortic dissection but no specific details are presented regarding which patients require genetic testing in either a diagnostic or a predictive capacity [45].

An expert European committee has more recently proposed their own consensus recommendations for the cardiogenetic care of patients with TAD and their first-degree relatives [16]. They advise that clinicians should be alerted to a possible genetic aetiology in individuals affected by aortopathy at younger than 50–60 years of age and/or a positive family history with aortic aneurysm or dissection in a first- or second-degree relative or in the presence of syndromic features [16]. Importantly, patients may present with cerebral, coronary or other non-aortic aneurysms in the presence of a syndromic aortopathy with reduced expression of aortic disease. In such circumstances, referral for genetic assessment should be considered, even in the apparent absence of an aortic aneurysm [23]. They further advise that targeted analysis of the FBN1 gene should be performed where a strong suspicion of Marfan syndrome exists to achieve molecular diagnosis [16]. Otherwise, genetic testing may proceed with next-generation sequencing utilising a targeted or exome-based gene panel. These recommendations are summarised in Fig. 2.

Fig. 2.

Algorithm for genetic testing in hereditary thoracic aortic disease [16]. Abbreviations: CT, computed tomography; FBN1, fibrillin 1 gene; ID, intellectual disability; MCA, multiple congenital abnormalities; MRI, magnetic resonance imaging; TAA, thoracic aortic aneurysm; TTE, trans-thoracic echocardiography

A contemporary systematic review evaluating existing screening regimens in non-syndromic TAD concluded that the risk of an acute aortic event, as determined by the underlying genetic mutation, extended not only to first-degree relatives, but also concerns second- and third-degree relatives [46]. The findings suggest that first-degree, second-degree and possible third-degree relatives are candidates for genetic screening. Since patients with non-syndromic HTAD receive a genetic diagnosis on average 10 years later than those with syndromic aortopathies [47], the authors advise that screening should commence 10–15 years earlier than the first occurrence of an aneurysm, dissection or sudden death within the family [46]. Figure 3 depicts a proposed screening pathway for relatives of patients with non-syndromic TAD [46].

Fig. 3.

Proposed flow chart for a dedicated screening program for relatives affected by non-syndromic diseases of the thoracic aorta [46]

Pitfalls and controversies specific to genetic screening for HTAD

For an effective screening programme, the related disease should be relatively common with significant associated morbidity and mortality such that it constitutes a serious burden to public health, and be treatable with a potential for cure if detected early enough [48]. There are additional criteria describing the test for the specific condition, which must be widely available, safe, cost-effective and capable of detecting pre-clinical disease in a large proportion of patients, and lead to enhanced health outcomes [48].

Although TAAs are a prevalent disease, those with a hereditary component that could be detected pre-emptively via genetic screening are much rarer. Indeed, the estimated incidences of the most frequently encountered genetic disorders are 1:5000 for MFS and 1:90,000 for vascular EDS, whilst LDS is even rarer with prevalence estimated at < 1:100,000. Certainly, these are highly aggressive genetic diseases as emphasised by the devastating aorto-vascular complications affected individuals suffer at a young age with a detrimental impact on life expectancy. However, the rarity of these HTADs implies that their genetic testing does not represent a common clinical requirement within the general population [49]. Whilst great strides have been made in elucidating the non-syndromic aortopathies associated with TGFBR1, TGFBR2, MYH11 and ACTA2 mutations, our accrued knowledge of these rare conditions is compiled from a very limited case population and requires further validation.

Furthermore, genetic sequencing is not essential for diagnosing MFS, for which the revised Ghent nosology additionally incorporates clinical features, anthropometric variables and family history elements [24]. This is important since some patients with a recognised FBN1 mutation do not express a Marfan phenotype [50], and similarly, the presence of clinical features of Marfan syndrome does not always assume an inherent mutation on genetic testing [51, 52].

Genetic testing in HTAD is a complex process, and many hundreds of mutations may exist for a given gene, all of which require careful examination as part of screening. The tests themselves are not widely available and patients and their relatives may need to travel significant distances to a tertiary or quaternary aortic centre or genetic clinic for counselling, testing and ongoing surveillance. Patients may also have to bear the expense of multiple gene panel tests themselves, since insurance companies may not provide coverage if genetic screening tests are not reasonably priced. The advent of next-general sequencing has, however, substantially reduced the costs associated with WES in recent years. In any case, the costs implicated in genetic screening for HTAD need to be justified by their accurate detection of a high proportion of responsible mutations in individuals who have not yet manifest an aneurysm or suffered an aorta-related adverse event. Yet despite continued progress in our understanding of the genetic pathogenesis of HTAD, currently identified mutations account for only 20% of familial TAAD. Thus, genetic screening may not achieve a high “yield” due to our presently incomplete profile of causative mutations.

The establishment of a genetic mutation by molecular testing only has limited bearing on determining phenotype or prognosis. Significant genetic heterogeneity exists in HTAD with large numbers of recognised mutations occurring for a particular gene. Furthermore, there is considerable intra- and inter-familial variability regarding disease expressivity in HTADs, with individuals sharing the same genetic mutation developing phenotypic manifestations at different ages, and on a spectrum of severity [53]. Risk analysis is more difficult in cases of sexual dimorphism, a phenomenon whereby women have a later onset of aortopathy than men, together with a lower overall lifetime risk [54]. Underlying genetic mutations, detected through screening processes, may induce only subtle variations in gene expression and protein function [55]. These observations underline the importance of the degree of clinical expression with HTAD genes in defining prognosis, which does not depend exclusively on the presence of the mutation in isolation [56]. Such clinical and genetic heterogeneity complicates the translation of novel findings on genetic screening to pertinent decisions affecting management and prognosis.

Genetic testing does not provide meaningful patient-specific information on the predicted rate of aortic growth or likelihood of progression to acute aortic events, which are intrinsically linked to aortic prognosis. This is instead largely determined practically by aortic size in the context of patient symptoms, physical evaluation and imaging findings. Again, the threat posed by a hereditary aneurysm is dependent more on the clinical expression of the disease, rather than the precise mutation [55]. Supporting these notions, international guidelines for the management of TAD originally devised one decade ago continue to rely heavily on diameter as a surrogate marker of aortic dimensions when selecting patients with inherited aneurysms for prophylactic surgery [18, 19]. The results of genetic testing, therefore, do not significantly influence surgical decision-making. In extension, identification of a deleterious genetic mutation in the relatives of an affected individual does not preclude them from serial cross-sectional imaging studies as part of a surveillance programme, and the associated risks of cumulative exposure to ionising radiation.

The interpretation of predictive genetic testing is not always straightforward. For example, the ambiguity of a VUS genetic screening result is a problematic issue for both clinicians and their patients, as encountered in 18% of syndromic aortopathies [40]. Patients with VUS may be cross-referenced with affected family members to check for presence of the same variant, in which case the VUS may be reclassified as pathogenic, and vice versa [57]. At the same time, identification of a VUS cannot be extrapolated to denote predisposition to aortopathy. The question is then raised as to whether the same VUS discovered in an affected patient should be tested for in asymptomatic relatives, since its implications for treatment remain unclear. The uncertain clinical connotations could have an adverse impact on a patient’s insurability. Controversy regarding the significance of a VUS could lead to incorrect assignment to a pathogenic category, and the attendant risks of unnecessary investigation, or even surgical intervention. In contrast, inadvertent underestimation of a dangerous VUS could lead to a dangerous situation whereby the patient is submitted to interval surveillance instead of undergoing earlier surgery.

Genetic testing in children importantly requires consideration of autonomy. It may be appropriate to defer screening until adulthood when the patient can participate more actively in decisions about their suspected aortopathy, unless aortic surgery is already indicated in childhood. A positive genetic test in teenagers and younger adults could affect their opportunities to participate in high-impact sports and affect future career choices. Consensus is lacking on the optimal age at which asymptomatic children should undergo molecular testing, and when to refer children carrying a pathogenic mutation for preventative surgery. Children and teenagers with aortic dilatation may “grow into” their aorta and acquire normalised aortic diameters when indexed to their height and/or body surface area [57], suggesting that genetic screening around the time of growth spurts may be unreliable.

The realisation stemming from predictive genetic testing that an asymptomatic individual or their close relatives are likely to acquire a potentially lethal aneurysm at an early age carries an immense psychological burden which cannot be underestimated. These anxieties may provoke demand from some patients to undergo prophylactic surgery at perhaps inappropriately early intervals. The psychological effects of genetic screening on patients and its impact on quality of life is poorly studied [46].

Finally, in the currently adopted guidelines on the management of thoracic aortic disease [18, 19], the proposed genetic screening recommendations are based on expert opinion and small group studies only, rather than high-quality data [46]. Despite their widespread implementation, the need for extended screening to accommodate second- and third-degree relatives, sequencing of rarer genetic mutations and ideal interval for the genetic screening of healthy at-risk relatives remain unspecified [46].

Future directions in genetic screening of HTADs

As our comprehension of the intricate pathophysiological mechanisms underlying HTAD advances further concurrent with the enhanced capabilities of genetic sequencing, so the emergent era of precision medicine holds much promise for the increasingly personalised management of this complex genetic disease targeted to the responsible genetic mutation.

The more widespread adoption of WES as the preliminary genetic testing strategy, and data sharing collaboratives by international working groups, will permit the faster identification of multiple novel pathogenic variants contributing to hereditary aortopathy, although clarification of their specific roles in phenotype manifestation represents another vital yet laborious task. This necessitates the standardisation of reporting methods for gene-disease relationships so that mutations can be linked to well-defined phenotypes to overcome the problem of syndromic overlap in genetic testing [58].

Further refinements in the sensitivity and specificity of genetic testing can be anticipated in the near future. This would potentially spare healthy relatives of an affected patient from the stresses of regular genetic screening as well as radiological investigations with the hazards of repeated exposure to ionising radiation from a relatively young age. Research efforts could be directed at addressing the functional relevance of VUS in HTAD through the development and validation of high-throughput assays. Exploring the mechanisms supporting epigenetic change, such as histone modification, DNA methylation and non-coding ribonucleic acid (RNA), could improve our understanding of incomplete penetrance and variable expressivity, and the resulting implications for the natural history of specific HTAD syndromes.

The necessity for the pre-emptive detection of TAAs has stimulated great interest in reliable, blood-based biomarkers which can facilitate the identification of at-risk individuals by means of a point-of-care test. In early trials, RNA signature sequences consisting of a 41-single nucleotide polymorphism panel have demonstrated 80% accuracy in distinguishing patients with TAAs [59]. Work is currently underway to formulate a straightforward blood test permitting the detection of hereditary aneurysms, building on from these very promising initial results.

Conclusions

TAAs represent a substantial burden to human health worldwide. The vast majority are clinically silent and grow insidiously until the point of a potentially fatal dissection or rupture, for which emergent surgery carries increased substantial peri-operative risks. Yet a significant 20% of cases are associated with an underlying genetic abnormality, providing a unique and crucial opportunity through screening for the early detection of asymptomatic individuals at risk of developing dangerous aneurysms, their surveillance and timely selection for safer prophylactic surgery. Over 30 genetic mutations have been identified to date to account for syndromic and non-syndromic HTADs. Detailed analyses have provided valuable insights into the complex molecular mechanisms initiating aneurysm formation in these genetic aortopathies, their characteristic phenotypic manifestations and altered natural histories. Whilst a great deal evidently remains to be elucidated, genetic screening is slowly but surely fulfilling its promise of affording personalised aortic management, customised to the individual’s genetic findings, in the special population with HTAD. The routine and widespread adoption of predictive genetic screening will have the enormous potential to save countless lives through the pre-emptive recognition of genetically susceptible patients before they suffer a catastrophic aortic complication arising within an undiagnosed aneurysm.

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The authors declare that they have no conflict of interest.

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