Since diagnosis of ATTR-CM requires specific laboratory and imaging diagnostic tools beyond basic routine tests,2 a high level of clinical suspicion is essential.2 International guidelines recommend a system of red flags to raise the index of clinical suspicion for this condition, although there is no consensus, possibly due to different country contexts.27 The 2021 position statement of the European Society of Cardiology (ESC) Working Group on Myocardial and Pericardial Diseases is the most widely used guide among Portuguese clinicians for the diagnosis and treatment of cardiac amyloidosis.28 However, it remains unclear how many red flags should trigger a diagnostic work-up, whether some present higher predictive value than others, and how to combine them, suggesting an unmet need for a suspicion algorithm tailored to Portugal's national context.

The following red flags were deemed by the authors to be the most important, considering the situation in Portugal and their experience/expertise: unexplained LVH ≥12 mm, reduced global left ventricular (LV) longitudinal strain with apical sparing pattern, discordance between QRS voltage and LV wall thickness, late gadolinium enhancement with diffuse transmural or subendocardial pattern, increased signal on T1 mapping and/or extracellular volume (ECV), signs and symptoms of HF, newly developed intolerance to previously tolerated antihypertensive drugs, carpal tunnel syndrome (CTS), distal biceps tendon rupture, polyneuropathy, and orthostatic hypotension (Table 1 and online Supplementary Materials). These red flags have also been previously highlighted by other authors.2,3,5,27,28 Some were considered to have a greater predictive impact, prompting suspicion even when presenting alone, as is the case of unexplained LVH ≥12 mm and other suggestive findings from electrocardiography, echocardiography or cardiac magnetic resonance (CMR) such as global LV longitudinal strain with apical sparing pattern, discordance between QRS voltage and LV wall thickness, late gadolinium enhancement with diffuse transmural or subendocardial pattern, increased signal on T1 mapping and/or ECV (see online survey results, Supplementary Materials, and Table 2). Additionally, given the increasing known prevalence of ATTRwt-CM,12 physicians are encouraged to maintain a low threshold for suspicion, meaning that they should suspect ATTR-CM even in patients with one or few red flags. This is in line with a recent prospective observational study, which showed that most ATTR-CM patients with LVH ≥12 mm referred to a center of excellence presented three or fewer red flags.29

Data from the online survey also indicated that Portuguese patients suspected of having ATTR-CM are most frequently referred by other medical specialties within the same institution, a scenario frequently seen in public hospitals and other public healthcare facilities. However, referrals also come from primary healthcare providers and private hospital settings (Supplementary Materials).

The suspicion algorithm proposed in Figure 1 reflects the national panorama and outlines four distinct scenarios in which ATTR-CM may be suspected, leading to a recommendation for the patient to be referred to a specialized CM clinic.

Figure 1.

Algorithm for referring patients with suspicion of transthyretin amyloid cardiomyopathy to a specialized cardiomyopathy clinic. 99mTc-DPD/HMDP: 99mTc-3,3-diphosphono-1,2-propanodicarboxylic acid or 99mTc-hydroxymethylene diphosphonate; ATTR-CM: transthyretin amyloid cardiomyopathy; ATTRv: variant ATTR amyloidosis; BNPs: brain natriuretic peptides; CM: cardiomyopathy; CMR: cardiac magnetic resonance; CV: cardiovascular; ECG: electrocardiogram; HF: heart failure; LVH: left ventricular hypertrophy; NT-proBNP: N-terminal pro brain-type natriuretic peptide; TTE: transthoracic echocardiogram; TTR: transthyretin. aE.g., hypertension, diabetes, obesity47,48; bSee Table 2 for details; cLVH not explained by hypertension or valvular disease.

positive: Perugini grade 2–3 scintigraphy.31

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The two most common scenarios encompass distinct clinical settings: centers with a specialized CM clinic and those without. These scenarios differ in their typical patient profiles, leading to differences in the most commonly observed red flags, as shown in Figure 1.

Centers without a specialized CM clinic have fewer resources and diagnostic tools, such as advanced imaging techniques, and their health professionals are less specialized in CM, often including those in primary healthcare settings. Although the complementary diagnostic tools to confirm a diagnosis are often unavailable, raising awareness and promoting early suspicion among GPs is essential, since they monitor many of the patients at risk for ATTR-CM. These centers will mainly refer patients with echocardiographic findings suggestive of ATTR-CM, such as unexplained LVH, to CM clinics.

Centers with a specialized CM clinic, which have access to advanced resources such as 99mTc-3,3-diphosphono-1,2-propanodicarboxylic acid or 99mTc-hydroxymethylene diphosphonate (99mTc-DPD/HMDP) scintigraphy, CMR and endomyocardial biopsies, will mainly refer to the CM clinic those patients with suspicion of ATTR-CM based on basic or advanced cardiac imaging, as well as patients with an established diagnosis of ATTR-CM.

Two less frequent but important scenarios were also considered sufficient to raise suspicion of ATTR-CM (Figure 1). The first encompasses patients who present a diagnosis of ATTR variant (ATTRv) amyloidosis due to non-p.V50M TTR variants, carriers of non-p.V50M TTR variants or individuals with a family history of ATTRv amyloidosis due to non-p.V50M TTR variants. The second consists of an incidental finding of grade 2 or 3 myocardial uptake of the radiotracer (99mTc-DPD/HMDP) on bone scintigraphy with planar and/or single-photon emission computed tomography (SPECT), performed for non-cardiac reasons.3,28

Importantly, unexplained LVH ≥12 mm was considered a red flag of high predictive value, which, alone or in conjunction with other red flags, should always be investigated and is therefore sufficient to trigger suspicion of ATTR-CM and referral of the patient to a specialized CM clinic (Figure 1). An LV thickness of ≥12 mm is the consensus threshold value in several guidelines,2,27,28 and is a cutoff above the gender-specific upper limit of normal. Women have lower LV wall thickness values than men, which may lead to underdiagnosis of ATTR-CM in women.30 LVH has the advantage of being detected by TTE, which is widely available.

After suspicion of ATTR-CM has been raised (Figure 1 and Table 2), the non-invasive diagnostic algorithm requires grade 2–3 99mTc-DPD/HMDP myocardial uptake in bone scintigraphy with SPECT, echocardiographic or CMR evidence of cardiac involvement by ATTR amyloidosis and exclusion of immunoglobulin light chain (AL) amyloidosis, in order to establish a diagnosis of ATTR-CM.

Exclusion of AL amyloidosis is mandatory, as these conditions have overlapping clinical, electrocardiographic and cardiac imaging features.2 The differential diagnosis may be performed by invasive and/or non-invasive diagnostic criteria.28

AL amyloidosis is excluded if all the following hematologic tests yield negative results, i.e., no monoclonal protein was detected: serum free light chain (FLC) assay; serum immunofixation electrophoresis (IFE); and urine IFE.3,28,31 The serum FLC assay measures the relative proportion of kappa and lambda light chains (κ/λ ratio), with monoclonality assumed by an abnormal ratio.27,28 Reference values for the κ/λ ratio depend on the estimated glomerular filtration rate (eGFR),32 as the κ/λ ratio increases with advancing chronic kidney disease due to the higher production of kappa light chains by the bone marrow and lower renal clearance of light chains in the presence of kidney disease. Reference values for the κ/λ ratio are depicted in Figure 2. Serum and urine IFE both measure the presence or absence of a monoclonal protein.2,3 Immunofixation must always be performed since electrophoresis alone lacks sensitivity.33 Hematology test results should be quickly performed and reviewed. This review should not be deferred due to the unavailability of scintigraphy results, as an abnormal light chain ratio or monoclonal protein detection should prompt a hematology consultation and potentially an endomyocardial biopsy to exclude AL cardiac amyloidosis (AL-CA).1–3

Figure 2.

Algorithm for the diagnosis of transthyretin amyloid cardiomyopathy. AL: light chain amyloidosis; ATTR-CM: transthyretin amyloid cardiomyopathy; ATTRv: variant ATTR amyloidosis; CM: cardiomyopathy; CMR: cardiac magnetic resonance; eGFR: estimated glomerular filtration rate; FLC: free light chain; IHC: immunohistochemistry; LVH: left ventricular hypertrophy; MGUS: monoclonal gammopathy of undetermined significance; MS: mass spectrometry; TTE: transthoracic echocardiogram; TTR: transthyretin.

positive;

negative;

99mTc-DPD/HMDP(99mTc-3,3-diphosphono-1,2-propanodicarboxylic acid or 99mTc-hydroxymethylene diphosphonate) scintigraphy with single-photon emission computed tomography (SPECT): Perugini grade 2–331;

biopsy: Congo red staining of amyloid deposits showing green birefringence when viewed under cross-polarized light28,39,40; aSee details in Table 2; $In the presence of the p.V50M variant, a biopsy is often required since 99mTc-DPD/HMDP scintigraphy can yield false negatives for this variant34–36; $Endomyocardial biopsy is the gold standard. An extracardiac biopsy may be considered and is helpful if positive, but the presence of a negative result should prompt further assessment through endomyocardial biopsy2,41; *Refined κ/λ FLC limits for normal ratio32; # Endomyocardial biopsy is the gold standard. An extracardiac biopsy may be considered and is helpful if positive, but the presence of a negative result should prompt further assessment through endomyocardial biopsy.2,41

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Grade 2–3 99mTc-DPD/HMDP myocardial uptake in bone scintigraphy with SPECT and negative results on all three hematologic tests have demonstrated specificity and positive predictive values of 100% (confidence interval 98.0–100.0) for ATTR-CM in a large cohort.31 However, there are situations that can lead to false positives on 99mTc-DPD/HMDP bone scintigraphy31 (Table 3).

The proposed diagnostic algorithm is based on the situation in Portugal, the authors’ expertise and experience, and current international guidelines (Figure 2). In Portugal, this algorithm is primarily implemented in national reference centers for familial amyloid polyneuropathy due to the p.V50M variant or in centers with specialized CM clinics.

Two possible initial scenarios were considered: (1) patients with a diagnosis of ATTRv amyloidosis due to the p.V50M variant, carriers of the p.V50M variant, or individuals with a family history of ATTRv amyloidosis due to the p.V50M variant and (2) patients with suspicion of ATTRv-CM due to other TTR variants or ATTRwt-CM.

In the first scenario, patients are referred to Portuguese reference centers for familial amyloid polyneuropathy due to the p.V50M variant. Cardiac imaging features suggestive of ATTR-CM would raise suspicion of the condition and a diagnosis of ATTR-CM could be confirmed through the non-invasive diagnostic algorithm or, if necessary, extracardiac or endomyocardial biopsy (Figure 2). In the case of the TTR variant being p.V50M, a biopsy is often required to confirm a diagnosis of ATTR-CM, since 99mTc-DPD/HMDP bone scintigraphy has low sensitivity and can yield false negatives for this variant, particularly in patients with early-onset disease.34–36

In the second scenario, patients are referred to specialized CM clinics, where the non-invasive diagnostic algorithm is applied, including the hematologic tests mentioned above along with 99mTc-DPD/HMDP scintigraphy with SPECT. Based on the results of these tests, there are four potential patient pathways that could be followed (Figure 2). Despite the differences in access to diagnostic tools, physicians working in centers without a specialized CM clinic (including those in primary healthcare settings) are encouraged to order all possible exams simultaneously with the patient's referral to a specialized CM clinic, if this helps to speed up diagnosis.

Distinguishing between different types of ATTR-CM by TTR genetic testing is important regardless of age, as about 5% of ATTR-CM patients aged ≥70 years (10% among females) have ATTRv.37 Genetic testing is also recommended for relatives of patients with ATTRv-CM, along with genetic counseling for patients and their families.27,28

Although they are unable to identify the type of amyloid, imaging techniques like CMR can provide helpful additional tissue characterization and high-resolution morphologic and functional assessment,38 and are thus recommended here as a complementary diagnostic method. Multimodality imaging techniques such as echocardiography, CMR, and radionuclide imaging are becoming more widely available. These non-invasive methods offer comprehensive cardiac imaging and could potentially be useful in monitoring amyloid burden and assessing its impact on cardiac function, for instance, in response to treatment.39

A biopsy positive for amyloid deposits has the pathognomonic histologic property of green birefringence when viewed under cross-polarized light after staining with Congo red.28,39,40 Although endomyocardial biopsy is highly specific, it is not widely available, requires technical expertise, and is an invasive procedure with potential risk for complications. Hence, cardiologists are often hesitant to proceed to biopsy in older adult patients or in those with early symptoms.39 Extracardiac biopsies, such as those taken from the abdominal fat pad, salivary glands, skin, peripheral nerve, kidneys, or bone marrow, are easier to perform.27,39,41 However, they have low sensitivity, and their yield in diagnosing ATTR amyloidosis is limited, particularly in the wild-type form.42 Extracardiac biopsies may be used as an alternative to endomyocardial biopsy, but given the high number of false negatives, further assessment is warranted in the presence of a negative result.2,41

Amyloid typing should be performed by immunohistochemistry (IHC) or mass spectrometry (MS) on a tissue biopsy by a trained clinical pathologist.28,39–41 IHC is a widely available technique, but it may lack specificity and sensitivity.40 MS is a more labor-intensive technique but has shown around 100% specificity and sensitivity in identifying the amyloid type, and should be used in cases of inconclusive IHC results.40,43 Reports have emerged of multiple myeloma patients with features of AL-CA (including abnormal serum FLC results), but ATTR-CM was in fact the confirmed diagnosis after biopsy and MS.44,45 Moreover, monoclonal gammopathy of undetermined significance (MGUS) has been estimated to affect 6.6% of people over the age of 80 years,46 which can lead to an increased risk of misdiagnosis in this population. Data from our online survey indicate that patients with abnormal findings suggestive of both ATTR-CM and AL-CA are the cases that give rise to the most uncertainty among experts (Supplementary Materials). Whenever this is the case, MS should be used for amyloid typing.

This algorithm shares several features with those from international societies and national consensus documents, namely: (1) the importance of differentiating between ATTR-CM and AL cardiac amyloidosis and the cornerstone techniques for doing so through a non-invasive diagnosis; (2) the possible diagnoses arising from different combinations of results (ATTR-CM, AL-CA, MGUS); and (3) the need for TTR genetic testing in patients with a definite diagnosis of ATTR-CM.2,3,5,27,28 Differences occur, for example, in the timings of the diagnostic techniques. The diagnostic algorithms of the World Heart Federation consensus, the ACC expert consensus, and the Amyloidosis Research Consortium expert consensus all suggest performing the hematologic tests first and, depending on their results, proceeding with 99mTc-DPD/HMDP bone scintigraphy or a biopsy,2,3,41 whereas in the present algorithm, hematologic tests and scintigraphy are recommended simultaneously. This diagnostic algorithm more closely resembles that of the position statement of the ESC Working Group on Myocardial and Pericardial Diseases.28 However, an important difference is that those guidelines rely more heavily on CMR results for diagnosis.28 Moreover, guidelines differ on when to opt for an extracardiac or an endomyocardial biopsy.2,3,5,27,28 Particularly in patients with a positive TTR genetic test and with no other cause for the cardiac phenotype, an extracardiac biopsy of an affected organ may be considered and is helpful if positive, but a negative result should prompt further assessment through endomyocardial biopsy.