Circulating ECM proteins decorin and alpha-L-iduronidase differentiate ATTRwt-CM from ATTRwt-negative HFpEF/HFmrEF.

AIMS: Wild-type transthyretin cardiac amyloidosis (ATTRwt-CM) is an under-recognized aetiology of heart failure (HF), necessitating early detection for timely treatment. Our study aimed to differentiate patients with ATTRwt-CM from ATTRwt-negative HFpEF/HFmrEF patients by identifying and validating circulating protein biomarkers. In addition, we measured the same biomarkers in patients with cardiomyopathy due to light chain amyloidosis (AL)-CM to gain disease-specific insights. METHODS AND RESULTS: In this observational study, serum concentrations of 363 protein biomarkers were measured in a discovery cohort consisting of 73 ATTRwt-CM, 55 AL-CM, and 59 ATTRwt-negative HFpEF/HFmrEF patients, using multiplex proximity extension assays. Sparse partial least squares analyses showed overlapping ATTRwt-CM and AL-CM biomarker profiles with clear visual differentiation from ATTRwt-negative patients. Pathway analyses with g:Profiler revealed significantly up-regulated proteoglycans (PG) and cell adhesion pathways in both ATTRwt-CM and AL-CM. Penalized regression analysis revealed that the proteoglycan decorin (DCN), lysosomal hydrolase alpha-L-iduronidase (IDUA) and glycosyl hydrolase galactosidase ß-1 (GLB-1) most effectively distinguished ATTRwt-CM from ATTRwt-negative patients (R2 = 0.71). In a prospective validation cohort of 35 ATTRwt-CM patients and 25 ATTRwt-negative patients, DCN and IDUA significantly predicted ATTRwt-CM in the initial analysis (DCN: OR 3.3, IDUA: OR 0.4). While DCN remained significant after correcting for echocardiographic parameters, IDUA did not. DCN showed moderate discriminative ability (AUC, 0.74; 95% CI, 0.61-0.87; sensitivity, 0.91; specificity, 0.52) as did IDUA (AUC, 0.78; 95% CI, 0.65-0.91; sensitivity, 0.91; specificity, 0.61). A model combining clinical factors (AUC 0.92) outperformed DCN but not IDUA, a combination of the biomarkers was not significantly better. Neither DCN nor IDUA correlated with established disease markers. CONCLUSION: ATTRwt-CM has a distinctly different biomarker profile compared with HFpEF/HFmrEF, while ATTRwt-CM patients share a similar biomarker profile with AL-CM patients characterized by up-regulation of proteoglycans and cell-adhesion pathways. The biomarkers DCN and IDUA show the potential to serve as an initial screening tool for ATTTRwt-CM. Further research is needed to determine the clinical usefulness of these and other extracellular matrix components in identifying ATTRwt-CM.

[99mTc]Tc-hydroxydiphosphonate uptake in soft tissue is associated with amyloid load in subcutaneous abdominal fat tissue and mortality in wild-type transthyretin amyloidosis patients.

PURPOSE: Bone scintigraphy is key to non-invasively diagnosing wild-type transthyretin (ATTRwt) amyloidosis, and is mainly used to assess cardiac radiotracer uptake. However, extracardiac radiotracer uptake is also observed. We investigated whether intensity of soft tissue radiotracer uptake is associated with amyloid load in subcutaneous abdominal fat tissue and with mortality. METHODS: This prospective cohort study included 94 ATTRwt amyloidosis patients and 26 amyloid-negative heart failure controls who underwent whole-body [99mTc]Tc-hydroxydiphosphonate scintigraphy. Site-to-background ratios were calculated for heart, elbows, subcutaneous tissue, shoulders and wrists on anterior planar bone scintigraphy images using rib and whole-body radiotracer uptake as background. Fat tissue aspirates were stained with Congo red to grade amyloid load. Site-to-rib ratios were compared between ATTRwt amyloidosis patients and controls, and associations of site-to-background ratio with Congo red score and all-cause mortality were studied. RESULTS: ATTRwt amyloidosis patients had higher soft tissue-to-rib, heart-to-rib and heart-to-whole body ratios compared with controls. The intensity of soft tissue uptake was positively associated with amyloid load in fat tissue in ATTRwt amyloidosis patients. Estimated glomerular filtration rate, N-terminal brain natriuretic propeptide, high-sensitivity cardiac troponin T (hs-cTnT), and the prognostic Mayo and NAC staging system were associated with all-cause mortality in univariable models. Soft tissue/rib ratio, hs-cTnT and the prognostic staging systems were the only two variables that were independently associated withall-cause mortality. CONCLUSION: Soft tissue radiotracer uptake on bone scintigraphy in ATTRwt amyloidosis patients is positively associated with amyloid load in abdominal fat tissue and is independently associated with mortality.

Diagnostic performance of liver stiffness as marker of liver involvement in systemic immunoglobulin light chain (AL) amyloidosis.

OBJECTIVE: To investigate the diagnostic performance of liver stiffness for detecting liver involvement in immunoglobulin light chain (AL) amyloidosis. METHODS: Liver stiffness was measured using transient elastography in 71 patients with systemic AL amyloidosis and 18 patients with wild type transthyretin (ATTRwt) amyloidosis with cardiomyopathy. Both non-invasive consensus criteria and serum amyloid P component (SAP) scintigraphy were used as substitute standards instead of liver biopsy for establishing liver involvement. RESULTS: Liver stiffness was higher in AL amyloidosis patients with liver involvement than in those without: this was observed using both consensus criteria (median 14.4 kPa vs. 8.1 kPa; p = 0.001) and SAP scintigraphy (median 20.9 kPa vs. 6.2 kPa; p < 0.001). Liver stiffness was also higher in AL amyloidosis patients with liver involvement compared to AL and ATTRwt amyloidosis patients with cardiac involvement. Based on receiver operating characteristic (ROC) curves a cut-off value of 14.4 kPa for stiffness was optimal to indicate liver involvement, providing sensitivity and specificity of 50% and 74%, respectively, using the consensus criteria and 63% and 90%, respectively, using SAP scintigraphy as standard. CONCLUSION: Liver stiffness is a promising tool to establish liver involvement in AL amyloidosis having potential to become part of updated criteria for liver involvement.

ELISA-4-amyloid: diagnostic accuracy of an ELISA panel for typing the four main types of systemic amyloidosis in subcutaneous abdominal fat tissue samples.

BACKGROUND: Reliable typing of amyloid is essential. Amyloid extraction from tissue enables immunochemical typing of the precursor protein using an enzyme-linked immunosorbent assay (ELISA). OBJECTIVE: To assess the diagnostic accuracy of a panel of ELISAs for typing the four main types (AA, ATTR, AL-kappa and AL-lambda amyloid). METHODS: From 1996 to 2023 subcutaneous abdominal fat tissue aspirates were obtained from 1339 amyloidosis patients and 868 controls. Amyloid was visually graded 0-4+ in Congo red-stained smears. Amyloid extracted from tissue by Guanidine was typed using a panel comprising four ELISAs. RESULTS: All amyloid protein concentrations in extracts correlated with amyloid grade in smears. Typing sensitivity was low (23.3%) in samples with grade 1+/2+ amyloid. Overall typing sensitivity of the panel was 81.6% for all easily visible amyloid (grade 3+/4+): high for AA (98.8%) and ATTR (96.8%) and fair for AL-kappa (66.7%) and AL-lambda (75.9). Overall typing specificity was 98.0% and the overall positive predictive value was 98.0%. CONCLUSIONS: We describe a highly specific ELISA panel for routine typing of the main amyloid types in fat tissue. Until more sensitive typing techniques will become generally available, typing easily visible amyloid in fat tissue using this ELISA panel is reliable, affordable and straightforward.

Neurofilament Light Chains in Systemic Amyloidosis: A Systematic Review.

Peripheral and autonomic neuropathy are common disease manifestations in systemic amyloidosis. The neurofilament light chain (NfL), a neuron-specific biomarker, is released into the blood and cerebrospinal fluid after neuronal damage. There is a need for an early and sensitive blood biomarker for polyneuropathy, and this systematic review provides an overview on the value of NfL in the early detection of neuropathy, central nervous system involvement, the monitoring of neuropathy progression, and treatment effects in systemic amyloidosis. A literature search in PubMed, Embase, and Web of Science was performed on 14 February 2024 for studies investigating NfL levels in patients with systemic amyloidosis and transthyretin gene-variant (TTRv) carriers. Only studies containing original data were included. Included were thirteen full-text articles and five abstracts describing 1604 participants: 298 controls and 1306 TTRv carriers or patients with or without polyneuropathy. Patients with polyneuropathy demonstrated higher NfL levels compared to healthy controls and asymptomatic carriers. Disease onset was marked by rising NfL levels. Following the initiation of transthyretin gene-silencer treatment, NfL levels decreased and remained stable over an extended period. NfL is not an outcome biomarker, but an early and sensitive disease-process biomarker for neuropathy in systemic amyloidosis. Therefore, NfL has the potential to be used for the early detection of neuropathy, monitoring treatment effects, and monitoring disease progression in patients with systemic amyloidosis.

Detection of cardiac amyloidosis on routine bone scintigraphy: an important gatekeeper role for the nuclear medicine physician.

Cardiac amyloidosis (CA)-mostly transthyretin-related (ATTR-CA)-has recently gained interest in cardiology. Bone scintigraphy (BS) is one of the main screening tools for ATTR-CA but also used for various other reasons. The objective was to evaluate whether all CA cases are detected and what happens during follow-up. All routine BS performed at the Maastricht University Medical Center (May 2012-August 2020) were screened for the presence of CA. Scans performed for suspected CA were excluded. A Perugini stage ≥1 was classified as positive necessitating further examination. The electronic medical record system was evaluated for any contact with cardiology or other specialists until 2021. Of the 2738 BS evaluated, 40 scans (1.46%; median age 73.5 [IQR: 65.8-79.5], 82.5% male) were positive (Perugini grade 1: 31/77.5%, grade 2: 6/15%, grade 3: 3/7.5%); the potential diagnosis ATTR-CA was not seen in 38 patients (95%) by the nuclear medicine specialist. During follow-up, 19 out of those 40 patients (47.5%) underwent cardiac evaluation without diagnosing CA. Available echocardiograms of patients with a positive BS showed left ventricular hypertrophy, a preserved ejection fraction, and diastolic dysfunction ≥2 in 9/47%, 10/53%, and 4/21% of patients, respectively. Additionally, 20 (50%) patients presented to at least one specialty with symptoms indicative of cardiac amyloidosis. The prevalence of a positive BS indicating potential CA in an unselected population is low but substantial. The majority was not detected which asks for better awareness for CA of all involved specialists to ensure appropriate treatment and follow-up.

Longitudinal analysis of serum neurofilament light chain levels as marker for neuronal damage in hereditary transthyretin amyloidosis.

OBJECTIVE: To evaluate serum neurofilament light chain (sNfL) as biomarker of disease onset, progression and treatment effect in hereditary transthyretin (ATTRv) amyloidosis patients and TTR variant (TTRv) carriers. METHODS: sNfL levels were assessed longitudinally in persistently asymptomatic TTRv carriers (N = 12), persistently asymptomatic ATTRv amyloidosis patients (defined as asymptomatic patients but with amyloid detectable in subcutaneous abdominal fat tissue) (N = 8), in TTRv carriers who developed polyneuropathy (N = 7) and in ATTRv amyloidosis patients with polyneuropathy on treatment (TTR-stabiliser (N = 20) or TTR-silencer (N = 18)). Polyneuropathy was confirmed by nerve conduction studies or quantitative sensory testing. sNfL was analysed using a single-molecule array assay. RESULTS: sNfL increased over 2 years in persistently asymptomatic ATTRv amyloidosis patients, but did not change in persistently asymptomatic TTRv carriers. In all TTRv carriers who developed polyneuropathy, sNfL increased from 8.4 to 49.8 pg/mL before the onset of symptoms and before polyneuropathy could be confirmed neurophysiologically. In symptomatic ATTRv amyloidosis patients on a TTR-stabiliser, sNfL remained stable over 2 years. In patients on a TTR-silencer, sNfL decreased after 1 year of treatment. CONCLUSION: sNfL is a biomarker of early neuronal damage in ATTRv amyloidosis already before the onset of polyneuropathy. Current data support the use of sNfL in screening asymptomatic TTRv carriers and in monitoring of disease progression and treatment effect.

High-Sensitivity Cardiac Troponin T to Exclude Cardiac Involvement in TTR Variant Carriers and ATTRv Amyloidosis Patients.

(1) Background: Individuals carrying a pathogenic transthyretin gene variant (TTRv) are at high risk for developing hereditary transthyretin (ATTRv) amyloidosis and are routinely screened for the development of cardiomyopathy (ATTRv-CM). This study aims to evaluate whether the cardiac biomarkers N-terminal pro B-type natriuretic peptide (NT-proBNP) and high-sensitivity cardiac troponin T (hs-cTnT) can be used to rule out ATTRv-CM. (2) Methods: In this retrospective case-control study, data from 46 ATTRv-CM patients and 101 TTRv carriers and ATTRv amyloidosis patients without cardiomyopathy were included. Binary logistic regression models were used to assess the ability of NT-proBNP and hs-cTnT to predict the diagnosis of ATTRv-CM. An optimal cutoff for the relevant biomarker(s) was determined based on a sensitivity of ≥99% and the highest possible percentage of additional tests avoided (%ATA) in the index dataset. (3) Results: Hs-cTnT demonstrated the highest predictive capabilities for ATTRv-CM. The addition of NT-proBNP did not improve the predictive model. A hs-cTnT cutoff of <6 ng/L resulted in a 97% sensitivity and a negative predictive value of 95% with a %ATA of 30% in the validation dataset. (4) Conclusion: In conclusion, hs-cTnT is a useful biomarker for excluding cardiac involvement in TTRv carriers and ATTRv amyloidosis patients and it has the potential to prevent unnecessary diagnostic procedures.

Pathophysiological pathways in patients with heart failure and atrial fibrillation.

AIMS: Atrial fibrillation (AF) and heart failure (HF) are two growing epidemics that frequently co-exist. We aimed to gain insights into the underlying pathophysiological pathways in HF patients with AF by comparing circulating biomarkers using pathway overrepresentation analyses. METHODS AND RESULTS: From a panel of 92 biomarkers from different pathophysiological domains available in 1620 patients with HF, we first tested which biomarkers were dysregulated in patients with HF and AF (n = 648) compared with patients in sinus rhythm (n = 972). Secondly, pathway overrepresentation analyses were performed to identify biological pathways linked to higher plasma concentrations of biomarkers in patients who had HF and AF. Findings were validated in an independent HF cohort (n = 1219, 38% with AF). Patient with AF and HF were older, less often women, and less often had a history of coronary artery disease compared with those in sinus rhythm. In the index cohort, 24 biomarkers were up-regulated in patients with AF and HF. In the validation cohort, eight biomarkers were up-regulated, which all overlapped with the 24 biomarkers found in the index cohort. The strongest up-regulated biomarkers in patients with AF were spondin-1 (fold change 1.18, P = 1.33 × 10-12), insulin-like growth factor-binding protein-1 (fold change 1.32, P = 1.08 × 10-8), and insulin-like growth factor-binding protein-7 (fold change 1.33, P = 1.35 × 10-18). Pathway overrepresentation analyses revealed that the presence of AF was associated with activation amyloid-beta metabolic processes, amyloid-beta formation, and amyloid precursor protein catabolic processes with a remarkable consistency observed in the validation cohort. CONCLUSION: In two independent cohorts of patients with HF, the presence of AF was associated with activation of three pathways related to amyloid-beta. These hypothesis-generating results warrant confirmation in future studies.

Neurofilament light chain, a biomarker for polyneuropathy in systemic amyloidosis.

OBJECTIVE: To study serum neurofilament light chain (sNfL) in amyloid light chain (AL) amyloidosis patients with and without polyneuropathy (PNP) and to corroborate previous observations that sNfL is increased in hereditary transthyretin-related (ATTRv) amyloidosis patients with PNP. METHODS: sNfL levels were assessed retrospectively in patients with AL amyloidosis with and without PNP (AL/PNP+ and AL/PNP-, respectively), patients with ATTRv amyloidosis and PNP (ATTRv/PNP+), asymptomatic transthyretin (TTR) gene mutation carriers (TTRv carriers) and healthy controls. Healthy controls (HC) were age- and sex-matched to both AL/PNP- (HC/AL) and TTRv carriers (HC/TTRv). The single-molecule array (Simoa) assay was used to assess sNfL levels. RESULTS: sNfL levels were increased both in 10 AL/PNP+ patients (p < .001) and in 10 AL/PNP- patients (p < .005) compared to 10 HC/AL individuals. sNfL levels were higher in AL/PNP+ patients than in AL/PNP- patients (p < .005). sNfL levels were also increased in 15 ATTRv/PNP+ patients, compared to both 15 HC/TTRv (p < .0001) and 15 TTRv carriers (p < .0001). ATTRv/PNP+ patients with progressive PNP (PND-score > I) had the highest sNfL levels compared to patients with early PNP (PND-score I) (p = .05). sNfL levels did not differ between TTRv carriers and HC/TTRv individuals. In the group comprising all healthy controls and in the group of TTRv carriers, sNfL levels correlated with age. CONCLUSION: sNfL levels are increased in patients with PNP in both AL and ATTRv amyloidosis and are related to severity of PNP in ATTRv amyloidosis. sNfL is a promising biomarker to detect PNP, not only in ATTRv but also in AL amyloidosis.

A real-life cohort study of immunoglobulin light-chain (AL) amyloidosis patients ineligible for autologous stem cell transplantation due to severe cardiac involvement or advanced disease.

Objective: To study the outcome of patients with AL amyloidosis who were ineligible for high dose melphalan (HDM) and autologous stem cell transplantation (ASCT).Methods: A real-life retrospective observational cohort study of Dutch patients with AL amyloidosis ineligible for HDM and ASCT was performed at the University Medical Center Groningen from January 2001 until April 2017. Primary outcome measure was overall survival (OS). Secondary outcome measures were hematological response (HR), organ responses, and treatment toxicity.Results: Eighty-four patients were included. Ineligibility was due to NYHA class III/IV (n = 58), otherwise advanced disease (n = 11), advanced age (n = 14), or treatment refusal (n = 1). Early death (<3 months) rate was high (44%). Median OS improved from 4 months in period 2001-2009 (n = 36) to 8 months in period 2009-2017 (n = 48, p = .02). HR was seen in 29%, and 42% of the patients, respectively. Median OS was 36 months after induction treatment with bortezomib (n = 32) and 18 months with immunomodulatory imide drug (IMID) (n = 16), both higher than median OS (7 months) with other regimens (n = 27). Incidence of toxicity was high (51%).Conclusion: OS improved in this high-risk group over the years, especially after introduction of new treatment modalities. However, early death rate remains high, illustrating the need for more effective treatment.

Causes of AA amyloidosis: a systematic review.

From a clinical perspective, there is a need for a reliable and comprehensive list of diseases causing AA amyloidosis. This list could guide clinicians in the evaluation of patients with AA amyloidosis in whom an obvious cause is lacking. In this systematic review, a PubMed, Embase and Web of Science literature search were performed on causes of AA amyloidosis published in the last four decades. Initially, 4066 unique titles were identified, but only 795 full-text articles and letters were finally selected for analysis. Titles were excluded because of non-AA type of amyloidosis, language, no full-text publication or irrelevance. Hundred and fifty diseases were initially reported to be associated with the development of AA amyloidosis. The presence of AA amyloid was proven in 208 articles (26% of all) of which 140 (67%) showed a strong association with an underlying disease process. Disease associations were categorized and 48 were listed as strong, 19 as weak, 23 as unclear, and 60 as unlikely. Most newly described diseases are not really unexpected because they often cause longstanding inflammation. Based on the spectrum of identified causes, a pragmatic diagnostic approach is proposed for the AA amyloidosis patient in whom an obvious underlying disease is lacking.