Assessment of ATTR Cardiac Amyloidosis in Patients with Left Ventricular Hypertrophy Using Tc-99m Pyrophosphate Scintigraphy

Introduction

Amyloidosis occurs due to extracellular deposits of fibrillar proteins, which causes tissue damage and functional compromise. These abnormal fibrils are formed by the aggregation of misfolded proteins (which are soluble in their normal folded state). Cardiac amyloidosis (CA) arises when these fibrils lodge within the cardiac extracellular space, blood vessels, and the conduction system. If untreated, infiltrative cardiomyopathy, cardiac failure, and death result.[1] Consequently, early diagnosis, accurate classification, and specific management are key determinants in the prognosis of these patients.

CA is most often caused by acquired monoclonal immunoglobulin light-chain (AL) disorder or transthyretin (TTR) disorders. TTR amyloidosis, often referred to as ATTR amyloidosis, could be hereditary or nonhereditary. The nonhereditary form is due to age-related failure of homoeostatic mechanisms in wild-type ATTR amyloidosis (ATTRwt). The hereditary form occurs due to destabilizing mutations in hereditary TTR amyloidosis (ATTRv; v for variant). The rampant amyloid fibril deposition in the extracellular space compromises the structure of the affected tissue and impairs its function, which in turn makes the patient symptomatic. Cardiomyopathy is the most common manifestation of ATTRwt amyloidosis, while ATTRv amyloidosis presents with both polyneuropathy and cardiomyopathy.[2]

Because of the amyloid infiltrations, ventricular wall thickness and stiffness increase, and CA mimics left ventricular hypertrophy (LVH). As a result, CA is underdiagnosed in LVH patients.[3] Although CA has a worse prognosis than other LVH-related disorders, it is currently pharmacologically curable.[4] A definitive CA diagnosis needs evidence of amyloid depositions in cardiomyocytes via endomyocardial biopsy, which can be life-threatening.[5] Imaging using Tc-99 m pyrophosphate (Tc-99 m PYP), has made it possible to have a noninvasive approach to ATTR-CA diagnosis, which may now be performed in approximately 70% of instances without the need for histological proof of amyloid.[6]

One of the strong independent risk factors for the prediction of cardiovascular morbidity and mortality is the presence of LVH, which is a common echocardiographic finding in these patients. ATTR CA can be diagnosed noninvasively, even when the histological proof is not present in a patient with heart failure with an associated echocardiogram and a cardiac magnetic resonance imaging (MRI) that is consistent with amyloidosis, a Grade 2 or 3 tracer uptake in the heart on 99 m Tc-PYP nuclear scan, and absence of monoclonal gammopathy in the blood and urine.[7] If strongly positive, DNA sequencing can be performed to determine the TTR subtype.

In this study, the noninvasive assessment of ATTR-CA in patients with LVH was done by using a Tc-99 m PYP scan in our institution.

Materials and Methods

The study was conducted at the department of nuclear medicine in our hospital from January 2023 to June 2024. The study population consisted of patients referred with clinical suspicion of CA with left ventricular wall thickness >12 mm. This was a prospective observational study with a sample size of 44 patients, calculated based on the expected prevalence of 13% from the reference study by Watanabe et al. (External Validation of the Kumamoto Criteria in TTR Amyloid Cardiomyopathy Screening – A Retrospective Cohort Study).[7]

Eligible patients were selected according to the inclusion and exclusion criteria. Inclusion criteria included left ventricular wall thickness >12 mm and age >55 years. Patients were excluded if they had hypertensive LVH, aortic stenosis, or ischemic heart disease. The study maintained patient confidentiality, with personal details such as name, contact information, and address not being disclosed, and was approved by the institutional ethics committee.

Results

A total of 52 patients who underwent Tc-99 m PYP scintigraphy were included in this study.

The mean ± SD of age was 64.6 ± 7.87 years, and it ranges from 55 years to 91 years. Of these 52 patients, 36 (69.2%) and 16 (30.8%) were males and females, respectively [Table 2].

The Tc-99 m PYP scan results, based on visual interpretation grading and semi-quantitative grading, were divided into positive, negative, and equivocal scans [Figures 1-3]. SPECT imaging was done in all cases that showed myocardial uptake on planar scintigraphy. They were used to differentiate cardiac uptake from blood pool or overlying bone uptake.

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Figure 1 Case 1: A 76-year-old male presented with features of heart failure with preserved ejection fraction and severe restrictive cardiomyopathy. (a) Tc-99 m PYP whole body sweep and (b and c) anterior chest static images acquired at 1 h and 3 h show diffusely increased myocardial uptake more than rib (Grade 3)-black arrow. The H/CL ratio at 1 h is 1.7. Positive for transthyretin amyloidosis. (d) Single photon emission comuted tomography (SPECT) shows diffuse abnormal myocardial tracer activity in left ventricle (LV), left atrium, and involvement of right ventricle. (e) Two-dimensional transthoracic echocardiocardiograph parasternal long axis view shows left ventricular hypertrophy with speckled appearance (red star), right ventricular hypertrophy (red circle), left atrial dilatation (red arrow) and pericardial effusion (white arrow). (f) Strain rate imaging of LV showing abnormal global longitudinal strain with apical sparing–Cherry on top appearance. Cardiac magnetic resonance imaging shows (g) Sub-endocardial delayed enhancement not respecting coronary artery territory. LV: Left ventricle, LA: Left atrium, RA: Right atrium, RV: Right ventricle

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Figure 2 Case 2: A 91-year-old female presented with large pericardial effusion, left ventricular hypertrophy, and preserved ejection fraction. (a) Tc-99 m PYP whole body sweep and (b) anterior chest static images acquired at 1 h and 3 h do not show any significant increase in myocardial Tc-99 m PYP uptake. However, on (c) single-photon emission computed tomography (SPECT) images, we can see myocardial uptake equal to that of rib, and the H/CL ratio at 1 h is 1.2, equivocal for Transthyretin (TTR) amyloidosis. A diffuse area of photopenia is noted around the heart in the planar chest anterior static image, which is suggestive of pericardial effusion. Thus, SPECT is crucial in the diagnosis of ATTR cardiac amyloidosis

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Figure 3 Case 3: 46-year-old male with azotemia, proteinuria, and renal biopsy suggestive of amyloidosis. (a) Tc-99 m PYP whole body sweep and (b) anterior chest static images acquired at 1 h and 3 h show no increase in myocardial Tc-99 m PYP uptake. H/CL ratio at 1 h is 1.0-Negative for Transthyretin amyloidosis

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All the parameters of the data collected were grouped based on these findings and analyzed.

Patients who underwent Tc-99 m PYP scans were divided into two age groups 55–70 years old and >70 years old. Majority of the study population were seen between the ages of 55 and 70 years (81%) [Table 3]. A statistically significant difference in age group was found between the two groups (P = 0.010). It was found that out of the total 52 cases majority of our study population were males, 36 (69%). When they were grouped based on the scan findings and analyzed the data, the P = 0.092. We concluded that there was no significant association between gender in our study. It was found that dyslipidemia was the most common comorbidity, followed by coronary artery disease (CAD), diabetes, hypertension, and chronic kidney disease (CKD). On analysis, the P = 0.014 in CAD and 0.026 in dyslipidemia, which is statistically significant. Cardiac biomarkers N terminal pro-brain natriuretic peptide (NT pro-BNP) and Troponin I were not performed in all patients as it was a relatively expensive test. Of the total 29 who did NT pro-BNP, 9 (31%) had equivocal and positive scan findings. A total of 24 people underwent Troponin I, of which 5 (20.8%) were equivocal, and 7 (29.2%) had raised Troponin I levels [Table 3].

Echocardiography features in all patients with LVH of >12 mm. From Table 4, we could see that on grouping these parameters based on PYP scintigraphy findings, septal hypertrophy (P = 0.003), elevated LV filling pressure (P = 0.042), granular speckled appearance (P = 0.006), and pericardial effusion (P = 0.038) were all significant variables in a patient with ATTR CA [Table 4].

Electrocarcdiograph (ECG) features in the study population were analyzed according to scintigraphy. LVH with strain pattern, followed by low voltage QRS and atrial fibrillation were the most common ECG findings.

A serum-free AL assay revealed elevated kappa and lambda ALs in 63.6% (n = 21 out of 32 available data). Serum immunofixation (SIFE) was positive in 34.5% (n = 10 out of 29 available data), demonstrating immunoglobulin G (IgG) lambda monoclonal gammopathy in most cases. Urine immunofixation was positive in 35% (n = 7 out of 20 available data).

Discussion

CA causes an increase in ventricular wall thickness and stiffness, mimicking LVH. Therefore, CA can be underdiagnosed in patients with LVH and requires appropriate screening to determine those who require further workup.[8] A landmark study by Gillmore et al. showed that ATTR-CA could be diagnosed noninvasively using a 99 m Tc-PYP scan in the absence of an endomyocardial biopsy.[6]

We performed our study by selecting patients with LV wall thickness >12 mm on echocardiography who underwent Tc99 m PYP scan based on clinical features, ECG and Echo. The male-to-female ratio of 9:4 and most of them were in the age group 55–70 years old. A total of 52 patients were divided into three groups based on their visual grading and ratio as negative n = 28, equivocal n = 15, and positive = 9. Cardiac biomarkers, ECG, and echo features were grouped and analyzed according to the Tc99 m PYP scan findings.

The assessment of demographic data showed that the majority of our study population lies between the age group of 55 and 70 years old (n = 42). Out of the remaining 10 patients in the age group of > 70 years, 50% had positive and 20% had equivocal PYP scan findings, respectively. Furthermore, the P < 0.005 (0.010), which is significant.

Bokhari et al.[9] showed that out of the 45 patients with CA, were predominately male (84%) older adults with a mean of 70 ± 2 years old. The age group is comparable to our study; however, females had a higher scan positivity rate than males in our study (25% against 13.9% males in positive PYP findings and 43.8% against 22.2% males in equivocal PYP scan findings). This female predominance is similar to some of the previous studies on the Indian population by Jain et al.[10] and an Italian and Spanish study by González-López et al.[11] In contrast, male predominance is reported by other Indian studies[1213] and Western studies.[9] These differences may be due to bias from referral centers and the use of Tc99 m PYP scan in other studies, and our study has increased the rate of diagnosis of female ATTR patients.

Among the associated comorbidities, CAD is seen in 38.4% of the study population (P = 0.014), which is significant. The majority of these patients presented with anginal pain and was labeled as CAD on CT angiography. Five out of 20 patients had equivocal Tc-99 m PYP scan findings, who had only mild or insignificant CAD and were on medical management. Al Suwaidi et al.[14] showed that amyloidosis can result in CAD by deposition of amyloid fibrils leading to atherosclerosis. Dorbala et al.[15] postulated that there is an accumulation of amyloid within the walls of small coronary arteries and none in the epicardial arteries; obstruction of these small coronaries may lead to ischemia. Further, amyloid deposits in the interstitial and perivascular regions of the heart increase coronary microvascular resistance and LV filling pressures, leading to coronary microvascular dysfunction and a higher risk of ischemia. Other comorbidities seen are diabetes mellitus (46.1%), dyslipidemia (61.6%), chronic kidney disease (32.6%), hypothyroidism (25%), and sarcoidosis (17%).

Dower et al. showed that NT pro-BNP, pro-BNP, and Troponin I were statistically significantly higher in patients with positive PYP scans.[16] In our study, we find similar raised cardiac biomarkers, which are shown in Table 3. Of the total 29 who did NT pro-BNP, 31% had equivocal and positive scan findings. A total of 24 people underwent Troponin I, of which 20.8% equivocal and 26% positive scan findings were having raised Troponin I levels. Persistent elevated cardiac biomarkers that are disproportionate to the clinical severity must raise the suspicion of CA. Ogasawara et al. reported that NT pro-BNP and highly sensitive troponin T were disproportionately elevated to the clinical severity, and these were the best predictors of mortality in ATTR-CA patients with advanced heart failure due to cardiomyopathy.[17]

LVH with strain pattern was the most common electrocardiographic (ECG) finding in 55.7% (n = 29) of our study, as shown in Table 5. Other common ECG features seen were low voltage QRS (QRS amplitude <5 mm in limb leads or <10 mm in precordial leads), pseudo-infarct pattern, and QTc prolongation (QTc interval >440 ms in men and > 460 ms in women). Bundle branch block AV conduction defects were seen in 17.3% (n = 9) and 23% (n = 12), respectively. Atrial fibrillation with a fast ventricular rate requiring electrical cardioversion was seen in nearly 32.1% of the cases.

All of the above ECG changes are well documented in the literature, which reflects the infiltrative nature of CA, as shown by Garcia-Pavia et al.[18] One of the important features of CA is disproportionately low QRS voltage and LVH which can be attributed to the loss of viable myocardium due to diffuse amyloid deposit in the heart. Pseudo-infarct pattern on ECG leads to an initial misdiagnosis of CAD and the presence of AV block in older patients with LVH should always raise the suspicion of CA.[3] While ECG is a helpful initial screening test to suspect CA, it can be nonspecific and difficult to interpret in isolation and cannot identify CA.[19]

Detailed Echocardiography with 3D Echo, tissue Doppler, and speckled tracking was done and reviewed for LVH, ejection fraction, valvular thickening, diastolic dysfunction, myocardial speckled appearance, and global longitudinal strain (GLS). Table 4 demonstrates the various echocardiographic features in our study.

Septal hypertrophy (interventricular septal thickness >11 mm) was seen in 71.1% of the study population, of which 16.2% had equivocal and 24.3 had positive scintigraphy findings, respectively. A statistically significant difference in interventricular septal thickness (P = 0.003) was noted between the different groups. This is similar to a study conducted by Saleem et al.[20] did a retrospective study by dividing the patients into three groups based on the scintigraphy findings and showed a statistically significant difference in septal thickness and posterior wall thickness, and a final diagnosis was found between the groups. A recent study by Grigoratos et al. showed that on consolidating different Perugini scores to the evolution of the disease, amyloid deposition in the myocardium progresses from the septum to the inferior wall, followed by the lateral wall and apex of the heart. This segmental analysis can be particularly helpful in patients with Grade 1 Perugini uptake. Apical sparing pattern in CA may be explained by the late involvement of the apex.[21]

The other statistically significant difference in echocardiographic parameters found between the groups in our study is a GLS with a relative apical sparing pattern in 67%, P = 0.05, granular speckling of the LV myocardium in 53.1%, P = 0.006, elevated filling pressures in 40.3%, P = 0.04 and pericardial effusion in 26.9%, P = 0.038. Biventricular hypertrophy is seen in 23%, RV dysfunction in 15%, and LV dysfunction in 46.1%. Atrial involvement is seen in 55.7% of which 30.7% of patients have biatrial involvement. Although these findings are characteristic of CA and have been useful in selecting patients for further confirmatory tests, they cannot accurately distinguish between AL-CA and ATTR-CA.

Cardiac MRI (cMRI) was done in 10 patients, revealing concentric LV hypertrophy, late gadolinium enhancement in sub-endocardium of ventricles, and atrial and ventricular dilatation in 40% of our study population (n = 4 out of 10) [Figure 1]. Nearly 75% of the patients with positive and equivocal PYP scans showed characteristic features of CA. Even though CMR is an excellent modality for the diagnosis of CA, it cannot be used to differentiate between ATTR-CA and AL-CA. Contraindications such as pacemakers, claustrophobia, and severe kidney dysfunction (preventing the use of contrast, which is crucial for diagnosis) limit its usage. Image quality in these patients can be significantly affected due to limited breath-holding ability or cardiac arrhythmias. Furthermore, obtaining delayed enhancement imaging, which is unique to CMR, can be challenging depending on the experience of operators.[22] Furthermore, it is expensive and requires expert opinion for diagnosis; we need another reliable imaging modality that is noninvasive, easy to interpret, and inexpensive. Thus, a 99 m-Tc PYP scan can be used for an earlier and more accurate diagnosis of ATTR CA.

Further, AL CA was ruled out using a monoclonal protein assay with serum and urine-free ALs and serum and urine immunofixation studies. Serum-free AL assay was done in 33 of 52 cases. Normal k/l (kappa/lambda) ratios were noted in 88.8% (8/9) of positive PYP cases and 92.3% (12/13) of equivocal PYP cases. SIFE studies showed absent monoclonal gammopathy in all positive PYP cases and monoclonal protein in the gamma region present in only one equivocal PYP case. This patient was subjected to a gastric biopsy and was diagnosed with both ATTR and AL-positive CA. Similar dual-positive CA was also reported by previous studies.[16232425] This is similar to a study by Quarta et al.[25] showed that in a cohort of AL-CA patients, approximately 40% of cases showed cardiac uptake on PYP scan. Those with both AL and ATTR-CA were associated with poorer cardiac function and outcome.

A 40% (4/10) of patients in the negative PYP scan group showed elevated kappa and lambda AL and positive SIFE, demonstrating IgG lambda monoclonal gammopathy in most cases. Two of these patients were later proven to be AL-Amyloidosis on biopsy, thus excluding ATTR-CA in these patients. A combination of serum-free AL proteins and urine and SIFE studies are needed to rule out plasma cell dyscrasias/AL-CA and confirm the diagnosis of ATTR CA. In cases of normal or equivocal results, a biopsy is necessary to arrive at the diagnosis. Hence, we can conclude that a 99 m-Tc PYP scan is essential in a nonbiopsy diagnosis of ATTR CA.

Extra-cardiac tissue biopsy was done to confirm amyloid, which includes renal (n = 3), gastric (n = 2) and abdominal fat pad biopsy (n = 1). Endomyocardial biopsy was suggested in one patient, but the patient refused as it is invasive and has significant morbidity. Bone marrow aspiration and bone marrow biopsy were performed in 11 out of 52 patients, demonstrating plasmacytosis and amyloid deposits in 50% (6 out of 11 patients). Apple green birefringence of amyloid deposits on polarized microscopy on Congo red stain is seen in these biopsy findings. Serum protein electrophoresis and FISH for multiple myeloma panel confirmed the diagnosis of multiple myeloma in two patients.

We can see in our study that there is a statistical difference between the Tc 99 m PYP scan group and the age of the population, highlighting the importance of a high index of suspicion in the older age group and a female predominance. Further, ECG and echocardiogram serve as necessary initial tools in helping us identify the important parameters such as Left ventricular wall thickness, interventricular septal thickness, and GLS with relative apical sparing, which prompts us to do a Tc99 m PYP scan and diagnose ATTR-CA earlier and noninvasively. Absence of monoclonal gammopathy further concludes the diagnosis, and this helps us to start appropriate therapy. Those with raised ALs were started on cyclophosphamide, bortezomib, and dexamethasone regimens. One patient with ATTR CA received targeted Tafamidis therapy.

The major limitations are that our study is a single-center study with a small sample size; a large-scale multicenter study is required to assess the prevalence of the disease. Further, due to financial constraints, the data pertaining to cardiac biomarkers, AL assay, cMRI, and genetic testing were unavailable to all patients.

Conclusion

Tc-99 m PYP imaging, along with a negative free AL assay, is highly useful in the early detection of ATTR CA and prompt initiation of specific targeted therapy. Our study demonstrates that unexplained LVH in the elderly is one of the most important parameters to assess for ATTR CA. The presence of heart failure with preserved ejection fraction, pseudo-infarct pattern, atrial arrhythmias, low ECG voltages despite LVH, restrictive features on echocardiography, and awareness of the red flag signs helped us keep a high index of suspicion for CA.