Diagnostic and Prognostic Value of Technetium-99m Pyrophosphate Uptake Quantitation for Transthyretin Cardiac Amyloidosis

Abstract

Background:

^99mTc-pyrophosphate imaging has emerged as an important non-invasive method to diagnose transthyretin cardiac amyloidosis (ATTR-CM). Quantitation of ^99mTc-pyrophosphate activity, on SPECT images, could be a marker of ATTR-CM disease burden. We assessed the diagnostic accuracy and clinical significance of ^99mTc-pyrophosphate quantitation.

Methods and Results

Patients who underwent ^99mTc-pyrophosphate imaging for suspected ATTR-CM were included. Using SPECT images, radiotracer activity in the myocardium was calculated using cardiac pyrophosphate activity (CPA) and volume of involvement (VOI), with thresholds for abnormal activity derived from LVBP activity. Diagnostic accuracy was assessed using area under the receiver operating characteristic curve (AUC).

In total, 124 patients were identified, mean age 73.9 ± 11.4, with ATTR-CM diagnosed in 43 (34.7%) patients. CPA had the highest diagnostic accuracy (AUC 0.996, 95% CI 0.987 – 1.00), and was significantly higher compared to the Perugini score (AUC 0.952, p=0.016). In patients with ATTR-CM, CPA was associated with reduced left ventricular ejection fraction (adjusted odds ratio 1.28, p=0.035) and heart failure hospitalizations (adjusted hazard ratio 1.29, p=0.006).

Conclusions:

Quantitative assessment of myocardial radiotracer activity with CPA or VOI have high diagnostic accuracy for ATTR-CM. Both measures are potential non-invasive markers to follow progression of disease or response to therapy.

BACKGROUND

Transthyretin amyloid cardiomyopathy (ATTR-CM) is an increasingly recognized cause of heart failure.(1) Diagnosis of ATTR-CM relies on a high clinical suspicion for disease in combination with cardiac imaging including echocardiography, cardiac magnetic resonance imaging, and nuclear cardiology imaging.(1) 99^m Technetium (Tc)-pyrophosphate imaging has emerged as a highly sensitive and specific tool for diagnosing ATTR-CM.(2) Initial studies described qualitative assessment of cardiac uptake relative to rib uptake.(2, 3) Subsequent studies have shown that heart to contralateral lung (H/CL) ratio on planar imaging may improve diagnostic accuracy compared to qualitative assessment and may have some prognostic utility.(4) However, the HCL ratio is influenced by regional rib radiotracer uptake, which may not be uniform, and radiotracer retention in the left ventricular blood pool (LVBP).(2) Examination of single-photon emission computed tomography (SPECT) imaging can improve diagnostic accuracy by differentiating myocardial from left ventricular blood pool (LVBP) activity (5).

In cardiac sarcoidosis, radiotracer activity in the LVBP is used as a reference to determine thresholds for abnormal myocardial activity.(6) This approach has theoretical benefits for quantification of ^99mTc-pyrophosphate uptake because it accounts for LVBP activity. Quantification from SPECT images with this method allows for exclusion of rib radiotracer activity, allowing direct myocardial quantification. Directly quantifying abnormal myocardial radiotracer activity could potentially provide closer associations with disease burden and clinical outcomes.

We performed a retrospective cohort study to determine the potential diagnostic and prognostic utility of direct quantitation of myocardial ^99mTc-pyrophosphate uptake, using thresholds for abnormal activity derived from the LVBP.

METHODS

Study Population

We identified patients who underwent ^99mTc-pyrophosphate imaging at the Foothills Medical Center between January 2016 and June 2019. For patients with more than one ^99mTc-pyrophosphate studies, only the first study was included for each patient.

Demographics, past medical history, and medical therapy at the time of imaging were determined through electronic medical records. Diagnosis of ATTR-CM was determined after reviewing all available clinical information. Previously described criteria were utilized to establish a diagnosis of ATTR-CM as one or more of the following: 1) endomyocardial biopsy positive for ATTR, 2) documented ATTR genetic mutation and evidence of cardiomyopathy (including increased LV mass, wall thickness, or clinical evidence of heart failure) without evidence of plasma cell dyscrasia, or 3) elevated H/CL ratio on ^99mTc-pyrophosphate imaging with evidence of cardiomyopathy and absence of plasma cell dyscrasia.(2, 7) Patients with evidence of plasma cell dyscrasia and evidence of cardiomyopathy (increased LV mass, wall thickness, or clinical evidence of heart failure) were considered to have AL-amyloidosis, unless there was an endomyocardial biopsy demonstrating ATTR-CM (n=1).(2, 7) Patients who did not meet any of the criteria above were classified as control patients.(2, 7) In total 23 patients had either endomyocardial or bone marrow biopsies including 14 with AL, 5 with ATTR-CM, and, and 4 patients classified as controls. Using Perugini grade 2 or 3 uptake in the absence of plasma cell dyscrasia as proposed by Gilmore et al.(8) would have reclassified one patient, who had positive SPECT visual interpretation and elevated CPA, from ATTR-CM to no ATTR-CM.

Image Acquisition

Imaging with ^99mTc-pyrophosphate was performed according to one of two protocols. All subjects underwent ^99mTc-PYP cardiac imaging on either a Ventri (GE, Boston, USA, n=16) or GE NM Discovery 670 (GE, Boston, USA, n=108) scanner. Patients received 740 or 770 Mbq (20 or 21 mCi) of ^99mTc-PYP intravenously and planar images were obtained at 1-hour and 3-hours post injection over 5–8 minutes duration, with a heart-centered field of view. SPECT images were acquired at 3-hours in all patients. Patients imaged with the Ventri camera system had tomogram images acquired over 60 steps, with a total acquisition time of 12 minutes using a low-energy high-resolution collimator, a 10% energy window and a 64 × 64 matrix. Patients imaged with the GE NM Discovery 670 had tomogram images acquired over 15 minutes using a low-energy high-resolution collimator, a 10% energy window and a 64 × 64 matrix. All images were reconstructed with filtered back projection.

Image Interpretation

Planar myocardial radiotracer retention was quantified using a H/CL ratio at the time of clinical interpretation. The H/CL ratio was calculated as the counts in a circular region of interest (ROI) drawn over the heart, divided by background counts in an identical size ROI over the contralateral chest from the 3-hour images.(5) The Perugini score, determined by comparing cardiac activity to rib activity (range 0—no uptake, to 3—uptake greater than bone), was also determined from 3-hour images at the time of clinical reporting.(5) To determine inter-rater variability for the Perugini score and H/CL ratio, all studies were over-read by a second physician blinded to the final clinical interpretation, H/CL ratio, clinical data and adjudication of TTR diagnosis. Lastly, the final clinical interpretation of the study (which includes consideration of the Perugini score, H/CL ratio, and SPECT imaging) was reported as positive, negative, or equivocal and included as “clinical interpretation”. This was included to compare to standard clinical practice. SPECT images at 3-hours were visually assessed to determine if radiotracer activity was present in the left ventricular blood pool (LVBP) or myocardium and recorded as “SPECT visual interpretation”.

SPECT images were quantified by two authors (RM and PP), blinded to results of the subjective visual assessment, H/CL ratio, clinical data and TTR adjudication with FusionQuant software (Cedars-Sinai Medical Center, Los Angeles, California). Examples of quantitation in a patient with ATTR-CM is shown in Figure 1 and in a patient without ATTR-CM in Figure 2. First, SPECT imaging was reviewed to identify myocardium and left ventricular blood pool (LVBP). Next, a spherical ROI with radius of 10-mm was placed in the center of the LVBP to determine background radiotracer activity. Maximal radiotracer counts of the LVBP was used as a marker of relative background activity.

Figure 1:

Patient with ATTR-CM who was eventually diagnosed with ATTR-CM. The planar imaging was graded visually as grade 2 uptake, with a heart/contralateral lung (H/CL) ratio of 1.29. Quantitative analysis was positive, with confirmation of radiotracer in the LV myocardium more than blood pool. The patient had an endomyocardial biopsy confirming ATTR-CM after the ^99mTc-pyrophosphate study was reported as equivocal.

Figure 2:

Patient who was eventually diagnosed with AL amyloidosis. Planar imaging was interpreted as grade 2 uptake with a heart/contralateral lung (H/CL) ratio 1.34. Examination of SPECT imaging confirms the activity was in the left ventricular blood pool and the study was quantitatively negative.

We used scan-specific thresholds for abnormal myocardial activity, determined as 1.5 × LVBP maximal radiotracer counts. This threshold was chosen based on previous evidence for background activity in ¹⁸F-fluorodeoxyglucose activity in patients with cardiac sarcoidosis.(6, 9) Next, a polygonal ROI was placed to encompass the left and right ventricular myocardium, with exclusion of adjacent rib activity. Radiotracer activity in the myocardium was quantified as volume of abnormal activity (volume of involvement [VOI]). Cardiac pyrophosphate activity (CPA), which reflects the volume and intensity of abnormal activity, was calculated as:

$$CPA=\text{VOI}\times \frac{\text{mean radiotracer counts of regions with abnormal myocardial activity}}{\text{maximal LVBP radiotracer activity}}$$

CPA was normalized to LVBP radiotracer counts to account for differences in background activity between camera systems, study protocols and to account for soft-tissue attenuation.

One patient, ultimately diagnosed with ATTR-CM by endomyocardial biopsy, had two ^99mTc-pyrophosphate studies performed 18 months apart with both CPA and VOI increasing over time (Supplemental Figure 1). Only the first study was included in the analysis.

Analyses:

The primary analysis was diagnostic accuracy for ATTR-CM. The diagnostic criteria used for adjudication of ATTR-CM diagnosis are outlined above and consistent with previous studies.(2, 7) The secondary analysis was correlation with left ventricular ejection fraction (LVEF) and LV mass.(10) Increased wall thickness was defined using categories established for left ventricular hypertrophy.(10) LVEF and LV mass were assessed on the echocardiogram closest to the time of ^99mTc-pyrophosphate imaging (before or after). We also assessed interobserver variability for markers of ^99mTc-pyrophosphate activity.

Clinical Outcomes

The primary clinical outcome was association with a combined clinical outcome of cardiovascular death or heart failure hospitalization. This analysis was only performed in patients with a final adjudicated diagnosis of ATTR-CM, since^99mTc-pyrophosphate uptake would not be expected to be associated with clinical outcomes in patients without ATTR-CM. Admissions for heart failure were ascertained from electronic medical records based on a primary admitting diagnosis of heart failure using standardized ICD-10 codes (11). Cardiovascular mortality was ascertained from electronic medical records and included death related to heart failure, myocardial infarction, and sudden death.(12) Follow-up was censored at the last time that follow-up status could be verified. We also assessed associations with each component of the combined clinical outcome and pre-specified a sub-analysis in patients not receiving targeted ATTR-CM therapies during follow-up. Outcomes were adjudicated blinded to the ^99mTc-pyrophosphate results.

Statistical Analysis

Categorical variables were summarized as number (proportion) and compared with a Chi-square or Fisher exact test as appropriate. Continuous variables were summarized as mean (standard deviation) and compared with a student’s t-test if normally distributed and with a Wilcoxon rank sum test if not.

Interobserver variability was assessed with mean percent difference and limits of agreement and was visualized with Bland-Altman plots.(13) Receiver operating characteristic (ROC) curves were generated for each method of image interpretation (clinical interpretation, H/CL ratio, SPECT visual interpretation, CPA, and VOI) with respect to diagnosis of ATTR-CM. Area under the ROC curve (AUC) confidence intervals were established with bootstrapping and differences in AUC were compared using DeLong’s method.(14) Correlations with LVEF was assessed with Pearson’s correlation co-efficient. Associations with the presence of reduced LVEF, defined as LVEF <50%, and moderate or severe increased wall thickness were assessed with multivariable logistic regression adjusted for age, sex, and hypertension.

Unadjusted associations with the primary clinical outcome were assessed with Cox proportional hazards models using time to first event from the ^99mTc-pyrophosphate study date. Multivariable Cox proportional hazards models were adjusted for age, sex, and LVEF with each ^99mTc-pyrophosphate parameter tested separately. LVEF was included in the multivariable model as a marker of disease severity. The proportional hazards assumption was assessed with Schoenfeld residuals, with no significant violations identified.

All statistical tests were two-sided, with p<0.05 considered statistically significant. All analyses were performed with STATA version 13 (StataCorp, College Station, Texas). The study was approved by the institutional review board at the University of Calgary (REB19–1448) with waiver of the requirement for informed written consent.

RESULTS

Patient Population

In total, 124 patients were identified, with a mean age 73.9 ± 11.4 and 94 (73.4%) male. ATTR-CM was diagnosed in 43 (34.7%) patients. Baseline population characteristics stratified by ATTR-CM diagnosis are outlined in Table 1. Patients with ATTR-CM were older (mean age 79.3 vs 70.6, P<0.001)) and less likely to have diabetes (9.1% vs. 25.0%, p=0.035). Patients with ATTR-CM had a similar mean LVEF to patients without (49.0 vs 52.5, p=0.101), but higher prevalence of at least moderately increased wall thickness (63.6% vs 22.5%, p<0.001). Comparison of pyrophosphate imaging parameters are shown in Table 1. There were significant differences in all markers between patients with and without ATTR-CM. A summary of cases with discrepant clinical interpretation and final diagnosis are shown in Supplemental Table 1.

Table 1:

Baseline population characteristics

	No ATTR-CM (n=81)	ATTR-CM (n=43)	p-value
Clinical Characteristics
Age (years)	70.6 ± 11.6	79.3 ± 8.6	<0.001
Male	56(69.1)	36(83.7)	0.088
Hypertension	37(45.7)	26(60.4)	0.134
Diabetes	20(24.7)	4(9.3)	0.055
Dyslipidemia	12(14.8)	13(30.2)	0.059
LVEF	52.6 ± 11.8	48.8 ± 10.4	0.101
≥ Moderate increased wall thickness	18(22.2)	28(65.1)	<0.001
Pyrophosphate imaging
Clinical interpretation positive	2(2.5)	38(88.4)	<0.001
Perugini score 2/3	7(8.6)	40(93.0)	<0.001
H/CL ratio	1.18 ± 0.14	1.72 ± 0.25	<0.001
SPECT visual positive	1(1.2)	43(100.0)	<0.001
CPA	3 ± 24	409 ± 339	<0.001
VOI	2 ± 14	225 ± 140	<0.001

Baseline population characteristics in patients with and without a diagnosis of transthyretin amyloid cardiomyopathy (ATTR-CM). Continuous variables displayed as mean ± SD, categorical as number(proportion). For troponin and Nt-Pro BNP number in brackets is the number of patients with available information. CPA – cardiac pyrophosphate activity, H/CL – heart contralateral lung, LVEF – left ventricular ejection fraction, SD – standard deviation, VOI – volume of involvement.

Diagnostic Accuracy

ROC curves for diagnosis of ATTR-CM are shown in Figure 3. CPA had the highest diagnostic accuracy (AUC 0.996, 95% CI 0.987 – 1.00) and was significantly higher compared to overall clinical interpretation (AUC 0.925, 95% CI 0.874 – 0.977, p=0.006) or the Perugini score (AUC 0.952, 95% CI 0.917 – 0.988, p=0.016). H/CL ratio had similar diagnostic accuracy (AUC 0.964, 95% CI 0.930 – 0.999) to CPA (p=0.065). SPECT visual interpretation had similar diagnostic accuracy to CPA (AUC 0.994, 95% CI 0.982 – 1.00, p=0.379) and was also superior to clinical interpretation (p=0.020).

Figure 3:

Diagnostic accuracy for diagnosis of transthyretin amyloid cardiomyopathy. Differences as denoted in the figure. CI – confidence interval, CPA – cardiac pyrophosphate activity, H/CL – heart contralateral lung.

Correlation with Left Ventricular Morphology and Function

In patients with ATTR-CM, there was no correlation between H/CL ratio (r=0.129, p=0.405) or Perugini score (r=0.263, p=0.085) and LVEF. There was moderate correlation between CPA and LVEF (r=−0.420, p=0.005). Association between LVEF and CPA is shown in Supplemental Figure 2. There was also moderate correlation between LVEF and VOI (r=−0.422, p=0.004). In patients with ATTR-CM, CPA (r=0.309, p=0.080), H/CL raito (r=0.156, p=0.139) and Perugini score (r=0.225, p=0.141) were not significantly correlated with LV mass.

Unadjusted and adjusted associations with reduced LVEF and moderate or greater increased wall thickness are shown in Table 2. Higher CPA was associated with moderate or greater increased wall thickness after adjustment for age, sex, and history of hypertension (adjusted odds ratio [OR] 1.64 per 100 units*cm³, 95% CI 1.16 – 2.31, p=0.005). Similar results were seen for VOI. Neither Perugini score (adjusted OR 3.06, p=0.056), nor H/CL ratio (adjusted OR 11.9, p=0.112), were associated with the presence of at least moderately increased wall thickness. Higher CPA was also associated with an increased likelihood of reduced LVEF (defined as LVEF <50%, adjusted odds ratio [OR] 1.28 per 100 units*cm³, 95% CI 1.02 – 1.62, p=0.035). Similar results were seen with VOI. Neither Perugini score (adjusted OR 3.12, p=0.076), nor H/CL ratio (adjusted OR 1.96, p=0.604), were associated with the presence of reduced LVEF.

Table 2:

Associations with ventricular morphology

	Unadjusted OR (95% CI)	p-value	Adjusted OR (95% CI)	p-value
Reduced LVEF (<50%)
Clinical interpretation positive	1.58(0.24 – 10.5)	0.637	1.17(0.15 – 9.03)	0.879
Perugini score	3.59(1.07 – 12.0)	0.039	3.12(0.89 – 11.0)	0.076
H/CL ratio	2.23(0.20 – 24.5)	0.510	1.96(0.15 – 25.0)	0.604
CPA (per 100 units*cm3)	1.09(1.02 – 1.18)	0.017	1.28(1.02 – 1.61)	0.035
VOI (per 100 cm3)	1.59(1.20 – 2.11)	0.001	1.59(1.17 – 2.16)	0.003
≥ Moderate Increased Wall Thickness
Clinical interpretation positive	1.19(0.18 – 8.0)	0.858	2.02(0.24 – 16.9)	0.516
Perugini score	2.73(0.95 – 7.86)	0.063	3.06(0.97 – 9.61)	0.056
H/CL ratio	9.56(0.57 – 160)	0.116	11.9(0.56 – 255)	0.112
CPA (per 100 units*cm3)	1.15(1.04 – 1.26)	0.005	1.64(1.16 – 2.31)	0.005
VOI (per 100 cm3)	2.25(1.58 – 3.19)	<0.001	2.57(1.72 – 3.85)	<0.001

Associations with reduced left ventricular ejection fraction (LVEF) and moderate or greater increased wall thickness in patients with ATTR-CM. Model adjusted for age, sex, and history of hypertension with each ^99mTc-pyrophosphate parameter assessed separately. SPECT visual interpretation excluded because all patients with ATTR-CM were positive. Reduced LVEF defined as values <50%. CI – confidence interval, CPA – cardiac pyrophosphate activity, H/CL – heart/contralateral lung, OR – odds ratio.

Clinical Outcomes:

During a median follow-up time of 1.2 years, 10 (18.2%) patients with ATTR-CM experienced cardiovascular mortality and 14 (31.8%) were admitted for decompensated heart failure. Unadjusted and adjusted assocations with the composite clinical outcome of heart failure hospitalization or all-cause mortality are shown in Table 3. CPA was not associated with an increased risk of the combined outcome of cardiovascular death or admission for heart failure (adjusted HR per 100 unit increase 1.17, 95% CI 0.98 – 1.38, p=0.076) after adjusting for age, sex, and LVEF. However, CPA (adjusted HR per 100 unit increase 1.29, p=0.001) and VOI (adjusted HR per 100 cm³ 1.81, p<0.001) were associated with HF hospitalization. Clinical interpretation, Perugini score, and H/CL ratio were not associated with the composite clinical outcome or its components. In patients not receiving targeted ATTR-CM therapies, CPA was significantly associated with the composite outcome (adjusted HR per 100 unit increase 1.28, 95% CI 1.05 – 1.56, p=0.014).

Table 3:

Unadjusted and adjusted associations

	Unadjusted HR (95% CI)	p-value	Adjusted HR (95% CI)	p-value
Cardiovascular Death or HF hospitalization
Clinical interpretation positive	0.60(0.17–2.16)	0.439	0.67(0.18–2.49)	0.548
Perugini score	1.10(0.50–2.40)	0.816	1.02(0.45–2.32)	0.957
H/CL Ratio	1.57(0.23–10.9)	0.649	2.45(0.31–19.3)	0.396
CPA (per 100 units*cm3)	1.13(0.96–1.34)	0.137	1.17(0.98–1.38)	0.076
VOI (per 100 cm3)	1.22(0.80–1.85)	0.350	1.31(0.85–2.00)	0.219
Cardiovascular Death
Clinical interpretation positive	0.33(0.06–1.80)	0.199	0.14(0.02–1.15)	0.068
Perugini score	0.45(0.18–1.13)	0.088	0.31(0.09–1.01)	0.061
H/CL Ratio	0.25(0.01–6.96)	0.417	0.25(0.01–8.82)	0.350
CPA (per 100 units*cm3)	0.93(0.71–1.21)	0.592	0.99(0.79–1.24)	0.921
VOI (per 100 cm3)	0.93(0.67–1.28)	0.648	0.98(0.66–1.44)	0.900
HF Hospitalization
Clinical interpretation positive	1.19(0.18–8.0)	0.858	1.86(0.22–15.6)	0.568
Perugini score	2.73(0.95–7.86)	0.063	3.30(1.05–10.3)	0.041
H/CL Ratio	9.56(0.57–160)	0.116	14.8(0.67–330)	0.088
CPA (per 100 units*cm3)	1.26(1.06–1.51)	0.010	1.29(1.08–1.55)	0.006
VOI (per 100 cm3)	1.58(1.00–2.50)	0.050	1.81(1.09–3.00)	0.022

Associations with clinical outcomes in patients with an adjudicated diagnosis of ATTR-CM. Model adjusted for age, sex, and history of hypertension with each ^99mTc-pyrophosphate parameter assessed separately. SPECT visual interpretation excluded because all patients with ATTR-CM were positive. CI – confidence interval, CPA – cardiac pyrophosphate activity, HCL – heart/contralateral lung, HR – hazard ratio.

Interobserver Variability

Interobserver variability for H/CL ratio at 3-hours, CPA, and VOI are shown in Supplemental Figures 3 to 5. Interobserver variability for CPA (mean difference 1.0%, limits of agreement −19.4% to 21.4%) and VOI (mean difference 0.6%, limits of agreement −17.2% to 18.4%) were low, and comparable to H/CL ratio (mean difference 2.2%, limits of agreement −20.5% to 24.9%). Agreement on absence of activity by CPA and VOI were excellent (kappa =1.00).

DISCUSSION

We performed a retrospective cohort study to determine the potential diagnostic and prognostic utility of quantitation of myocardial ^99mTc-pyrophosphate uptake compared to clinical interpretation, Perugini score and H/CL ratio. Quantitative assessment of myocardial radiotracer activity with CPA or VOI had high diagnostic accuracy for ATTR-CM. The inter-observer variability in quantifying CPA and VOI were lower compared to H/CL ratio, and there was excellent agreement regarding the presence of absence of activity. Most importantly, in this preliminary study we demonstrated significant correlations between direct quantitation of myocardial radiotracer uptake and ventricular morphology as well as associations with the incidence of heart failure hospitalization. Quantitative assessment of ^99mTc-pyrophosphate uptake could potentially have a clinical role as a non-invasive marker in patients with ATTR-CM to follow progression of disease or response to medical therapy.

We investigated a novel method for assessing ^99mTc-pyrophosphate images with direct myocardial radiotracer quantitation. However, previous studies have assessed other methods to directly measure myocardial ^99mTc-pyrophosphate activity. Scully et al. recently demonstrated high diagnostic accuracy for myocardial SUV_MAX (15)_. Direct myocardial quantification has also been reported by two studies which used a ratio of myocardial to LVBP activity. Sperry et al. reported myocardial uptake for each segment in a 17-segment model using the ratio of mean myocardial counts divided by mean blood pool counts in 54 patients with ATTR-CM(16). They found no association between all-cause mortality and total myocardial to blood pool ratio(16). However, they did find that an apical-sparing ratio >2.75 was associated with all-cause mortality(16). Glaudemans et al. showed that a similar myocardial to LVBP ratio correlated with left ventricular mass index (17). In comparison to methods which derive ratios for activity levels, we determined thresholds for abnormal activity relative to the LVBP and quantified total abnormal activity (CPA) and volume of abnormal activity (VOI). We demonstrated that CPA and VOI were correlated with LVEF and associated with an increased incidence of heart failure hospitalizations, and therefore may be useful markers of disease severity.

Direct myocardial quantification has theoretical benefits compared to existing methods. Qualitative scoring systems and indirect quantitation of myocardial activity are influenced by regional difference in rib uptake and LVBP activity. Direct myocardial radiotracer quantitation, with CPA or VOI, accounts for these sources of error by excluding rib activity from regions of interest and using LVBP activity to determine thresholds for abnormal activity. Visual differentiation of LVBP from myocardial radiotracer activity on SPECT images also accounts for these sources of error (5), and had similarly high diagnostic accuracy in our study. In fact, given the high diagnostic accuracy of visual SPECT interpretation it is unlikely that quantifying ^99mTc-pyrophosphate uptake could improve diagnostic accuracy. However, it may have a role as a marker of disease burden. Direct myocardial quantitation was associated with lower interobserver variability compared to H/CL ratio. Additionally, the relative limits of agreement for H/CL ratio in our study (−20.5% to 24.9%) were lower compared to absolute limits of agreement in a recent study by Masri et al. (−0.4 to 0.4) (18). Importantly, there was uniform agreement on the presence of absence of abnormal activity using direct myocardial quantification. Reducing interpretation variability is potentially more important for centers with less expertise in evaluating ^99mTc-pyrophosphate studies. Regardless, recent myocardial infarction will impact all methods for interpretation and evaluation of uptake pattern is important to ensure high diagnostic accuracy.

There has been mixed evidence for the prognostic utility of ^99mTc-pyrophosphate imaging. Single center studies did not identify an association between Perugini score or H/CL ratio and cardiovascular events in patients with ATTR-CM (19). However, in a multicenter study with 171 patients H/CL ratio >1.6 was associated with worse survival in several multivariable models.⁴ Similar to Sperry et al. (16), we did not identify significant associations with cardiovascular mortality. However, we identified a significant association between CPA and VOI with heart failure hospitalizations after adjusting for age, sex, and LVEF. Additionally, in patients not receiving targeted ATTR-CM therapies, CPA was associated with the composite outcome of heart failure hospitalization or cardiovascular mortality. However, our study was under-powered and larger prospective studies would be required to validate the prognostic utility of CPA and VOI. However, if prospective studies can demonstrate these findings then CPA and VOI may have significant clinical value as markers of disease burden.

The importance of accurate diagnosis of ATTR-CM is driven by the evolving therapeutic options for patients with the disease, with increasing use of ^99mTc-pyrophosphate imaging (20). While the classical therapy for this condition was liver transplantation,¹⁹ recent studies have demonstrated the utility of medical therapies aimed at stabilizing misfolded transthyretin precursors.²⁰ In the ATTR-ACT trial, treatment with Tafamadis significantly reduced the incidence of all-cause mortality and cardiovascular hospitalization compared to placebo in patients with ATTR-CM.²⁰ Patisiran improved echocardiographic parameters and decreased a combined end-point of all-cause mortality or cardiovascular hospitalization compared to placebo.(21) While these drugs have clearly demonstrated benefit, given the associated costs it is important to accurately identify which patients should be considered for therapy.(22) Methods to quantify disease burden such as quantification with CPA or VOI, could potentially help select patients who benefit most from targeted therapies or be used to monitor the efficacy of therapy.

Our study has a few important limitations. We combined patients imaged with two separate camera systems and acquisition protocols. While this may have increased the variability of CPA calculation, it does increase the generalizability of the results. Additionally, although the ROI locations were not standardized the interobserver variability for low and comparable to that for the H/CL ratio. Studies investigating potential differences in quantitative parameters between camera systems and acquisition protocols could be informative. We quantified all images at 3-hours using ^99mTc-pyrophosphate. It is unknown whether the results can be extrapolated to centers only performing imaging at 1 hour (18), or using 3,3-diphosphono-1,2-propanodicarboxylic acid radiotracers (23)³. Additionally, it is likely that myocardial quantitation may not be as accurate in patients with genetic mutations associated with type B fibrils (24). Echocardiograms were acquired during routine clinical care and strain measurements were not uniformly available to assess correlations with CPA and VOI. We relied on a combination of criteria to adjudicate a diagnosis of ATTR-CM similar to previous studies. This was based on the initial clinical interpretation of studies, which would tend to underestimate the diagnostic accuracy of the novel quantitative measures. While a pathologic diagnosis would provide greater certainty regarding adjudication of ATTR-CM, this is not reflective of clinical practice which has shifted to increasingly non-invasive diagnosis (18) Studies are needed to demonstrate the test retest reliability and repeatability between camera systems. Lastly, larger prospective studies would be required to assess the independent associations between CPA and VOI with clinical outcomes after adjustment for all important confounding factors.

CONCLUSIONS

Quantitative assessment of myocardial radiotracer activity with CPA or VOI have high diagnostic accuracy for ATTR-CM. The inter-observer variability in quantifying CPA and VOI were lower compared to H/CL ratio, and there was excellent agreement regarding the presence or absence of activity. Most importantly, CPA and VOI are associated with reduced left ventricular ejection fraction and incidence of heart failure hospitalization and are therefore potential non-invasive markers in ATTR-CM to follow progression of disease or response to medical therapy.

NEW KNOWLEDGE GAINED

Cardiac pyrophosphate activity and volume of involvement have low interobserver variability and high diagnostic accuracy for ATTR-CM. Both measures are correlated with left ventricular ejection fraction and associated with heart failure hospitalizations and therefore are potentially useful non-invasive markers of disease burden in ATTR-CM.

ABBREVIATIONS

ATTR-CM

transthyretin cardiac amyloidosis

AUC

area under the receiver operating characteristic curve

CI

confidence interval

CPA

cardiac pyrophosphate activity

H/CL

heart contralateral lung

LVBP

left ventricular blood pool

SPECT

single photon emission computed tomography

VOI

volume of involvement