Clinical Penetrance of the Transthyretin V122I Variant in Older Black Patients With Heart Failure: The SCAN-MP (Screening for Cardiac Amyloidosis With Nuclear Imaging in Minority Populations) Study.

Background Transthyretin amyloid cardiomyopathy (ATTR-CM) is an underdiagnosed cause of heart failure (HF) among patients ≥60 years of age. Although the V122I (valine to isoleucine substitution at position 122 of the transthyretin protein) variant associated with hereditary ATTR-CM is present in 3.4% of self-identified Black individuals in the United States (or 1.5 million people), the phenotypic penetrance is not known. Methods and Results The SCAN-MP (Screening for Cardiac Amyloidosis With Nuclear Imaging in Minority Populations) study is a currently accruing prospective multisite study designed to determine the prevalence of ATTR-CM using technetium-99m-pyrophosphate imaging in older (≥60 years of age) self-identified Black and Hispanic individuals with HF. Calculations of the penetrance and prevalence of the V122I allele, along with analyses of functional, biochemical, and echocardiographic parameters, were performed for the first 278 Black participants in SCAN-MP. The prevalence of ATTR-CM was 6.8% (95% CI, 4.2-10.5; n=19 cases), of whom 63% were ATTR wild-type. The prevalence of V122I was 6.5% (n=18 carriers), of whom 7 had ATTR-CM, yielding a phenotypic penetrance of 39% (95% CI, 17-64). V122I carriers with ATTR-CM evidenced more advanced HF than carriers without ATTR-CM. Prealbumin concentration was lowest among V122I carriers with ATTR-CM (12.9 mg/dL) versus carriers without ATTR-CM (21.0 mg/dL) and HF controls (25.0 mg/dL, P<0.0001). Conclusions Among older Black individuals with HF and increased left ventricular wall thickness, of those with ATTR-CM, 63% had wild-type, and of those with V122I, the phenotypic penetrance of ATTR-CM was 39% (95% CI, 17-64), suggesting that genotype alone is insufficient for diagnosis. Prealbumin concentration may be useful to identify V122I carriers with ATTR-CM. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT03812172.

Mapping cellular response to destabilized transthyretin reveals cell- and amyloidogenic protein-specific signatures.

BACKGROUND: In ATTR amyloidosis, transthyretin (TTR) protein is secreted from the liver and deposited as toxic aggregates at downstream target tissues. Despite recent advancements in treatments for ATTR amyloidosis, the mechanisms underlying misfolded TTR-mediated cellular damage remain elusive. METHODS: In an effort to define early events of TTR-associated stress, we exposed neuronal (SH-SY5Y) and cardiac (AC16) cells to wild-type and destabilized TTR variants (TTRV122I (p.V142I) and TTRL55P (p.L70P)) and performed transcriptional (RNAseq) and epigenetic (ATACseq) profiling. We subsequently compared TTR-responsive signatures to cells exposed to destabilized antibody light chain protein associated with AL amyloidosis as well as ER stressors (thapsigargin, heat shock). RESULTS: In doing so, we observed overlapping, yet distinct cell type- and amyloidogenic protein-specific signatures, suggesting unique responses to each amyloidogenic variant. Moreover, we identified chromatin level changes in AC16 cells exposed to mutant TTR that resolved upon pre-incubation with kinetic stabilizer tafamidis. CONCLUSIONS: Collectively, these data provide insight into the mechanisms underlying destabilized protein-mediated cellular damage and provide a robust resource representing cellular responses to aggregation-prone proteins and ER stress.

Role of complementarity-determining regions 1 and 3 in pathologic amyloid formation by human immunoglobulin κ1 light chains.

BACKGROUND: Immunoglobulin light chain (LC) amyloidosis is a life-threatening disease complicated by vast numbers of patient-specific mutations. We explored 14 patient-derived and engineered proteins related to κ1-family germline genes IGKVLD-33*01 and IGKVLD-39*01. METHODS: Hydrogen-deuterium exchange mass spectrometry analysis of conformational dynamics in recombinant LCs and their fragments was integrated with studies of thermal stability, proteolytic susceptibility, amyloid formation and amyloidogenic sequence propensity. The results were mapped on the structures of native and fibrillary proteins. RESULTS: Proteins from two κ1 subfamilies showed unexpected differences. Compared to their germline counterparts, amyloid LC related to IGKVLD-33*01 was less stable and formed amyloid faster, whereas amyloid LC related to IGKVLD-39*01 had similar stability and formed amyloid slower, suggesting different major factors influencing amyloidogenesis. In 33*01-related amyloid LC, these factors involved destabilization of the native structure and probable stabilization of amyloid. The atypical behavior of 39*01-related amyloid LC stemmed from increased dynamics/exposure of amyloidogenic segments in ßC'V and ßEV that could initiate aggregation and decreased dynamics/exposure near the Cys23-Cys88 disulfide. CONCLUSIONS: The results suggest distinct amyloidogenic pathways for closely related LCs and point to the complementarity-defining regions CDR1 and CDR3, linked via the conserved internal disulfide, as key factors in amyloid formation.

Design and Rationale the SCAN-MP (Screening for Cardiac Amyloidosis With Nuclear Imaging in Minority Populations) Study.

Background Transthyretin amyloid cardiomyopathy (ATTR-CM) is an important cause of heart failure in older individuals. Misfolding and deposition of transthyretin or prealbumin protein causes ATTR-CM in the context of a normal (wild-type) or variant TTR sequence. Variant ATTR-CM is most commonly caused by the substitution of valine for isoleucine at position 122 in transthyretin (Val122Ile or pV142I, almost exclusively observed in individuals of West African ancestry), demonstrated in 3.4% of self-identified Black individuals in the United States with an estimated 1.5 million carriers. Despite the large number of known pV142I carriers, the proportion of older Black patients with heart failure attributable to ATTR-CM remains unknown. Methods To address this knowledge gap, the SCAN-MP (Screening for Cardiac Amyloidosis with Nuclear Imaging in Minority Populations) study was funded by the National Institutes of Health/National Heart, Lung, and Blood Institute (R01HL139671) to enroll a targeted population of self-identified, community-dwelling Black or Caribbean Hispanic patients (many of whom are of West African ancestry) >60 years of age with heart failure and identify ATTR-CM by noninvasive nuclear imaging. The principal objective of SCAN-MP is to determine the prevalence of ATTR-CM in this population. Secondary objectives will explore TTR genotype, demographics, progression of variant versus wild-type ATTR-CM, and biochemical mechanisms of transthyretin amyloid fibril formation. Conclusions The SCAN-MP study is the largest, prospective study of cardiac amyloidosis in Black and Hispanic individuals. Both wild-type and variant ATTR-CM are now treatable with the US Food and Drug-approved drug tafamidis. The insights gained from SCAN-MP are likely to improve those at risk for or afflicted with ATTR-CM. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT03812172.

Role of Complementarity-Determining Regions 1 and 3 in Pathologic Amyloid Formation by Human Immunoglobulin κ1 Light Chains.

Immunoglobulin light chain (LC) amyloidosis is a life-threatening disease whose understanding and treatment is complicated by vast numbers of patient-specific mutations. To address molecular origins of the disease, we explored 14 patient-derived and engineered proteins related to κ1-family germline genes IGKVLD-33*01 and IGKVLD-39*01. Hydrogen-deuterium exchange mass spectrometry analysis of local conformational dynamics in full-length recombinant LCs and their fragments was integrated with studies of thermal stability, proteolytic susceptibility, amyloid formation, and amyloidogenic sequence propensities using spectroscopic, electron microscopic and bioinformatics tools. The results were mapped on the atomic structures of native and fibrillary proteins. Proteins from two κ1 subfamilies showed unexpected differences. Compared to their germline counterparts, amyloid LC related to IGKVLD-33*01 was less stable and formed amyloid faster, whereas amyloid LC related to IGKVLD-39*01 had similar stability and formed amyloid slower. These and other differences suggest different major factors influencing amyloid formation. In 33*01-related amyloid LC, these factors involved mutation-induced destabilization of the native structure and probable stabilization of amyloid. The atypical behaviour of 39*01-related amyloid LC tracked back to increased dynamics/exposure of amyloidogenic segments in ßC' V and ßE V that could initiate aggregation, combined with decreased dynamics/exposure near the Cys23-Cys88 disulfide whose rearrangement is rate-limiting to amyloidogenesis. The results suggest distinct amyloidogenic pathways for closely related LCs and point to the antigen-binding, complementarity-determining regions CDR1 and CDR3, which are linked via the conserved internal disulfide, as key factors in amyloid formation by various LCs.

An additive destabilising effect of compound T60I and V122I substitutions in ATTRv amyloidosis.

BACKGROUND: The amyloidogenic transthyretin (TTR) variant, V122I, occurs in 4% of the African American population and frequently presents as a restricted cardiomyopathy. While heterozygosity for TTR V122I predominates, several compound heterozygous cases have been previously described. Herein, we detail features of ATTRv amyloidosis associated with novel compound heterozygous TTR mutation, T60I/V122I and provide evidence supporting the amyloidogenecity of T60I. METHODS: A 63-year-old African American female presented with atrial fibrillation, congestive heart failure, autonomic and peripheral neuropathy. In vitro studies of TTR T60I and V122I were undertaken to compare the biophysical properties of the proteins. RESULTS: Congophilic deposits in a rectal biopsy were immunohistochemically positive for TTR. Serum screening by isoelectric focussing revealed two TTR variants in the absence of wild-type protein. DNA sequencing identified compound heterozygous TTR gene mutations, c.239C > T and c.424G > A. Adipose amyloid deposits were composed of both T60I and V122I. While kinetic stabilities of T60I and V122I variants were similar, distinct thermodynamic stabilities and amyloid growth kinetics were observed. CONCLUSIONS: This report provides clinical and experimental results supporting the amyloidogenic nature of a novel TTR T60I variant. In vitro data indicate that the destabilising effect of individual T60I and V122I variants appears to be additive rather than synergistic.

A novel substitution of proline (P32L) destabilises ß2-microglobulin inducing hereditary systemic amyloidosis.

BACKGROUND: ß2-microglobulin amyloidosis was first described in the 1980s as a protein deposition disease associated with long-term haemodialysis. More recently, two inherited forms resulting from separate point mutations in the ß2-microglobulin gene have been identified. In this report, we detail a novel ß2M variant, P32L, caused by a unique dinucleotide mutation that is linked to systemic hereditary ß2-microglobulin amyloidosis. METHODS: Three family members from a Portuguese kinship featured cardiomyopathy, requiring organ transplantation in one case, along with soft tissue involvement; other involvements included gastrointestinal, neuropathic and sicca syndrome. In vitro studies with recombinant P32L, P32G, D76N and wild-type ß2-microglobulin were undertaken to compare the biophysical properties of the proteins. RESULTS: The P32L variant was caused by the unique heterozygous dinucleotide mutation c.154_155delinsTT. Amyloid disease featured lowered serum ß2-microglobulin levels with near equal amounts of circulating P32L and wild-type proteins; amyloid deposits were composed exclusively of P32L variant protein. In vitro studies of P32L demonstrated thermodynamic and chemical instability and enhanced susceptibility to proteolysis with rapid formation of pre-fibrillar oligomeric structures by N- and C-terminally truncated species under physiological conditions. CONCLUSIONS: This work provides both clinical and experimental evidence supporting the critical role of P32 residue replacement in ß2M amyloid fibrillogenesis.

Neurological manifestations of hereditary transthyretin amyloidosis: a focus on diagnostic delays.

BACKGROUND: The recent availability of disease-modifying therapies for hereditary transthyretin amyloid (ATTRv) amyloidosis warrants urgency for earlier diagnosis and timely identification of active disease state among genetic carriers. METHODS: We reviewed clinical neurological data of all patients with ATTRv amyloidosis with initial visits at our amyloidosis centre between January 2016 and December 2018. We abstracted the signs and symptoms of neurological manifestations, as well as rates and patterns of diagnostic testing. RESULTS: Of 92 patients with 19 different transthyretin (TTR) mutations, 66 and 36% had symptoms attributed to large-fibre and small-fibre neuropathy, respectively, compared to 75 and 66% with corresponding examination findings. Thirty-six patients with V122I ATTR mutation had asymptomatic polyneuropathy identified on neurological examination, eight without concurrent cardiac disease. Seventy-three percent of patients had symptoms of carpal tunnel syndrome (CTS), while 26% had dysautonomia. The average delays between the onset of symptoms of large fibre neuropathy (LFN) or CTS to ATTRv amyloidosis diagnosis were 2.9 and 6.7 years, respectively. DISCUSSION: Our study found higher rates of polyneuropathy by examination than patient-reported symptoms, especially among those with V122I TTR amyloidosis, signalling asymptomatic polyneuropathy. Our findings suggest the need for routine neurological examinations and other testing for genetic carriers to achieve earlier identification of active disease state.

Correlation Between 24-Hour Urine Protein and Random Urine Protein-Creatinine Ratio in Amyloid Light-Chain Amyloidosis.

Rationale & Objective: Test the feasibility of replacing 24-hour urine collection with a single voided urinary protein-creatinine ratio (UPCR) in patients with amyloid light-chain (AL) amyloidosis. Study Design: Retrospective study examining the correlation between a 24-hour urine measurement and UPCR at various proteinuria levels using a linear regression analysis with Pearson's correlation coefficient (r). We assessed how using these 2 different measurements would alter the diagnosis, staging, and kidney response assessment in patients with AL amyloidosis. Setting & Participants: We included 265 patients with systemic AL amyloidosis who visited the Amyloidosis Center at Boston University between July 2018-January 2020 and had proteinuria measurement by both methods on the same day. Tests Compared: 24-hour urine collection for protein versus UPCR. Results: The correlation between 24-hour urine and UPCR was moderate in patients with proteinuria levels of 500-3,000 mg/day and >3,000 mg/day, with r values of 0.57 and 0.62, respectively. Replacing the 24-hour urine collection with UPCR changed kidney staging in 10% of the patients: 77% were reclassified to a worse kidney stage and 23% to a more favorable stage. The majority of changes (85%) in kidney staging occurred in the >3,000 mg/day cohort. There were 35 patients whose kidney response was assessed by concomitant 24-hour urine collection and UPCR with visits at least 6 months apart. Of these patients, 20% had discordance between the 24-hour urine collection and UPCR that changed their definition of organ response. Limitations: Given the rarity of AL amyloidosis, our sample size is small and from a single referral center. Conclusions: Although the 24-hour urine collection is cumbersome, we continue to recommend it in patients with AL amyloidosis because replacing the 24-hour urine collection with UPCR would change kidney staging and organ response in 10%-20% of patients. In addition, the correlation between the 2 modalities was moderate at best in patients with nephrotic-range proteinuria.

Diflunisal treatment is associated with improved survival for patients with early stage wild-type transthyretin (ATTR) amyloid cardiomyopathy: the Boston University Amyloidosis Center experience.

BACKGROUND: Diflunisal is a non-steroidal anti-inflammatory drug that stabilises transthyretin (TTR) and reduces neurologic deterioration in patients with polyneuropathy caused by hereditary transthyretin amyloidosis (ATTRv). METHODS: We conducted a retrospective cohort study of patients with wild-type transthyretin cardiac amyloidosis (ATTRwt-CM) treated with diflunisal for at least one year between 2009 and 2016 at the Boston University Amyloidosis Centre. Baseline and one year follow up characteristics were measured, including plasma chemistries and echocardiography. Cox proportional hazards analysis assessed the primary outcome of all-cause mortality. RESULTS: A total of 104 ATTRwt-CM patients were evaluated with 35 patients receiving diflunisal. Patients in the diflunisal group were younger (73.8 vs 76.8 years, p = 0.034), with lower B-type natriuretic peptide (BNP, 335 +/- 67 vs. 520 +/- 296 pg/mL, p = 0.006), similar troponin I (0.1 +/- 0.1 vs 0.2 +/- 0.3 ng/mL, p = 0.09), and better renal function (eGFR 67 +/- 17 vs 53 +/- 18 mL/min/1.73m2, p = 0.0002) at baseline. Over a median follow-up of 3.2 years, 52 deaths occurred. Diflunisal administration was associated with improved survival in unadjusted analysis (HR 0.13, 95% CI 0.05 - 0.36, p < 0.001) that persisted after adjustment for age, baseline BNP, eGFR, troponin I, interventricular septal thickness, and left ventricular ejection fraction (HR 0.18, 95% CI 0.06 - 0.51, p = 0.0006). Over the observation period, no significant changes in BNP, troponin I, interventricular septal thickness or left ventricular ejection fraction were observed with diflunisal treatment. A total of 14 patients (40%) discontinued diflunisal in this study, but only 3 within the first year. Mean eGFR in treated patients was 59 ml/min/1.73m2 at 1 year (change from baseline p = 0.03). CONCLUSION: Diflunisal administration in ATTRwt-CM was associated with improved survival and overall stability in clinical and echocardiographic markers of disease with decrement renal function.

A Conservative Point Mutation in a Dynamic Antigen-binding Loop of Human Immunoglobulin λ6 Light Chain Promotes Pathologic Amyloid Formation.

Immunoglobulin light chain (LC) amyloidosis (AL) is a life-threatening human disease wherein free mono-clonal LCs deposit in vital organs. To determine what makes some LCs amyloidogenic, we explored patient-based amyloidogenic and non-amyloidogenic recombinant LCs from the λ6 subtype prevalent in AL. Hydrogen-deuterium exchange mass spectrometry, structural stability, proteolysis, and amyloid growth studies revealed that the antigen-binding CDR1 loop is the least protected part in the variable domain of λ6 LC, particularly in the AL variant. N32T substitution in CRD1 is identified as a driver of amyloid formation. Substitution N32T increased the amyloidogenic propensity of CDR1 loop, decreased its protection in the native structure, and accelerated amyloid growth in the context of other AL substitutions. The destabilizing effects of N32T propagated across the molecule increasing its dynamics in regions ∼30 Å away from the substitution site. Such striking long-range effects of a conservative point substitution in a dynamic surface loop may be relevant to Ig function. Comparison of patient-derived and engineered proteins showed that N32T interactions with other substitution sites must contribute to amyloidosis. The results suggest that CDR1 is critical in amyloid formation by other λ6 LCs.

Marked progress in AL amyloidosis survival: a 40-year longitudinal natural history study.

The recent decades have ushered in considerable advancements in the diagnosis and treatment of systemic light chain (AL) amyloidosis. As disease outcomes improve, AL amyloidosis-unrelated factors may impact mortality. In this study, we evaluated survival trends and primary causes of death among 2337 individuals with AL amyloidosis referred to the Boston University Amyloidosis Center. Outcomes were analyzed according to date of diagnosis: 1980-1989 (era 1), 1990-1999 (era 2), 2000-2009 (era 3), and 2010-2019 (era 4). Overall survival increased steadily with median values of 1.4, 2.6, 3.3, and 4.6 years for eras 1-4, respectively (P < 0.001). Six-month mortality decreased over time from 23% to 13%. Wide gaps in survival persisted amid patient subgroups; those with age at diagnosis ≥70 years had marginal improvements over time. Most deaths were attributable to disease-related factors, with cardiac failure (32%) and sudden unexpected death (23%) being the leading causes. AL amyloidosis-unrelated mortality increased across eras (from 3% to 16% of deaths) and with longer-term survival (29% of deaths occurring >10 years after diagnosis). Under changing standards of care, survival improved and early mortality declined over the last 40 years. These findings support a more optimistic outlook for patients with AL amyloidosis.

Race/ethnicity in systemic AL amyloidosis: perspectives on disease and outcome disparities.

In marked contrast to multiple myeloma, racial/ethnic minorities are underrepresented in publications of systemic light-chain (AL) amyloidosis. The impact of race/ethnicity is therefore lacking in the narrative of this disease. To address this gap, we compared disease characteristics, treatments, and outcomes across racial/ethnic groups in a referred cohort of patients with AL amyloidosis from 1990 to 2020. Among 2416 patients, 14% were minorities. Non-Hispanic Blacks (NHBs) comprised 8% and had higher-risk sociodemographic factors. Hispanics comprised 4% and presented with disproportionately more BU stage IIIb cardiac involvement (27% vs. 4-17%). At onset, minority groups were younger in age by 4-6 years. There was indication of more aggressive disease phenotype among NHBs with higher prevalence of difference between involved and uninvolved free light chains >180 mg/L (39% vs. 22-33%, P = 0.044). Receipt of stem cell transplantation was 30% lower in Hispanics compared to non-Hispanic White (NHWs) on account of sociodemographic and physiologic factors. Although the age/sex-adjusted hazard for death among NHBs was 24% higher relative to NHWs (P = 0.020), race/ethnicity itself did not impact survival after controlling for disease severity and treatment variables. These findings highlight the complexities of racial/ethnic disparities in AL amyloidosis. Directed efforts by providers and advocacy groups are needed to expand access to testing and effective treatments within underprivileged communities.

Glycosylation of Serum Clusterin in Wild-Type Transthyretin-Associated (ATTRwt) Amyloidosis: A Study of Disease-Associated Compositional Features Using Mass Spectrometry Analyses.

Wild-type transthyretin-associated (ATTRwt) amyloidosis is an age-related disease that causes heart failure in older adults. This disease frequently features cardiac amyloid fibril deposits that originate from dissociation of the tetrameric protein, transthyretin (TTR). Unlike hereditary TTR (ATTRm) amyloidosis, where amino acid replacements destabilize the native protein, in ATTRwt amyloidosis, amyloid-forming TTR lacks protein sequence alterations. The initiating cause of fibril formation in ATTRwt amyloidosis is unclear, and thus, it seems plausible that other factors are involved in TTR misfolding and unregulated accumulation of wild-type TTR fibrils. We believe that clusterin (CLU, UniProtKB P10909), a plasma circulating glycoprotein, plays a role in the pathobiology of ATTRwt amyloidosis. Previously, we have suggested a role for CLU in ATTRwt amyloidosis based on our studies showing that (1) CLU codeposits with non-native TTR in amyloid fibrils from ATTRwt cardiac tissue, (2) CLU interacts only with non-native (monomeric and aggregated) forms of TTR, and (3) CLU serum levels in patients with ATTRwt are significantly lower compared to healthy controls. In the present study, we provide comprehensive detail of compositional findings from mass spectrometry analyses of amino acid and glycan content of CLU purified from ATTRwt and control sera. The characterization of oligosaccharide content in serum CLU derived from patients with ATTRwt amyloidosis is novel data. Moreover, results comparing CLU oligosaccharide variations between patient and healthy controls are original and provide further evidence for the role of CLU in ATTRwt pathobiology, possibly linked to disease-specific structural features that limit the chaperoning capacity of CLU.

Expression of Amyloidogenic Transthyretin Drives Hepatic Proteostasis Remodeling in an Induced Pluripotent Stem Cell Model of Systemic Amyloid Disease.

The systemic amyloidoses are diverse disorders in which misfolded proteins are secreted by effector organs and deposited as proteotoxic aggregates at downstream tissues. Although well described clinically, the contribution of synthesizing organs to amyloid disease pathogenesis is unknown. Here, we utilize hereditary transthyretin amyloidosis (ATTR amyloidosis) induced pluripotent stem cells (iPSCs) to define the contribution of hepatocyte-like cells (HLCs) to the proteotoxicity of secreted transthyretin (TTR). To this end, we generated isogenic, patient-specific iPSCs expressing either amyloidogenic or wild-type TTR. We combined this tool with single-cell RNA sequencing to identify hepatic proteostasis factors correlating with destabilized TTR production in iPSC-derived HLCs. By generating an ATF6 inducible patient-specific iPSC line, we demonstrated that enhancing hepatic ER proteostasis preferentially reduces the secretion of amyloidogenic TTR. These data highlight the liver's capacity to chaperone misfolded TTR prior to deposition, and moreover suggest the potential for unfolded protein response modulating therapeutics in the treatment of diverse systemic amyloidoses.

Patient outcomes in light chain (AL) amyloidosis: The clock is ticking from symptoms to diagnosis.

INTRODUCTION: Symptomology of AL amyloidosis can be vague, with a broad range of manifestations and potential etiologies. We sought to determine whether time from initial patient-reported symptom onset to diagnosis was associated with survival. METHODS: The Boston University Amyloidosis Patient Database was queried for patients with AL amyloidosis who presented to the Center for initial evaluation from 2010 to 2015. RESULTS: A total of 324 patients with AL amyloidosis were evaluated for initial evaluation. The median time to diagnosis from initial symptom onset was 7.1 months (range, 0-61). At data cutoff, 60.2% (n = 195) of patients were alive; of those, the majority were diagnosed <6 months from initial symptoms (52.3%, n = 102). In contrast, time to diagnosis from symptom onset was >6 months in 63.6% (n = 82) of patients who did not survive at the time of data cutoff (P = .0005). Survival analysis of time from diagnosis to death or data cutoff stratified by time from patient-reported symptom onset to diagnosis (<6, 6-12, and >12 months) showed significant differences among groups (P = .001). Additionally, multivariable regression demonstrated that an increase in time from self-reported symptom onset to diagnosis was significantly associated with an increased risk of death (HR = 1.02, 95% CI = 1.01-1.04, P = .002). CONCLUSION: These results support the importance of early diagnosis for patients with AL amyloidosis.

Structural Characterization of Cardiac Ex Vivo Transthyretin Amyloid: Insight into the Transthyretin Misfolding Pathway In Vivo.

Structural characterization of misfolded protein aggregates is essential to understanding the molecular mechanism of protein aggregation associated with various protein misfolding disorders. Here, we report structural analyses of ex vivo transthyretin aggregates extracted from human cardiac tissue. Comparative structural analyses of in vitro and ex vivo transthyretin aggregates using various biophysical techniques revealed that cardiac transthyretin amyloid has structural features similar to those of in vitro transthyretin amyloid. Our solid-state nuclear magnetic resonance studies showed that in vitro amyloid contains extensive nativelike ß-sheet structures, while other loop regions including helical structures are disrupted in the amyloid state. These results suggest that transthyretin undergoes a common misfolding and aggregation transition to nativelike aggregation-prone monomers that self-assemble into amyloid precipitates in vitro and in vivo.

Stabilization of Cardiac Function With Diflunisal in Transthyretin (ATTR) Cardiac Amyloidosis.

BACKGROUND: Transthyretin amyloidosis cardiomyopathy (ATTR-CM) is an underappreciated cause of heart failure that results from misfolded TTR (prealbumin) protein. Diflunisal is an approved non-steroidal anti-inflammatory drug that stabilizes TTR, with limited data available regarding effects on cardiac structure and function. METHODS AND RESULTS: ATTR-CM patients (n=81, 41% treated with 250 mg twice-daily diflunisal by clinical practice) were retrospectively identified with baseline and follow-up (median interval 1 year) serum biomarker and echocardiographic data compared, including global longitudinal strain (GLS). Chi-squared and Wilcoxon tests assessed differences between subjects, divided by treatment group, and univariable and multivariable linear regression was performed. At baseline, patients treated with diflunisal were younger (68 vs 77 years, P = .0001), with lower B-type natriuretic peptide (BNP; 249 vs 545 pg/mL, P = .009) and serum creatinine (1.1 vs 1.2 mg/dL, P = .04), but similar TTR concentration (P = .31), cardiac troponin I (P = .06), and GLS (P = .67). At follow-up, diflunisal untreated versus treated patients showed differences in TTR concentration (19 vs 33 mg/dL, P = .01) and favorable differences in left atrial volume index (+4.6 vs -1.4 mL/m2, P = .002) and cardiac troponin I (+0.03 vs -0.01 ng/mL, P = .01) for the entire cohort. Among the subset with wild-type ATTR (n=53), diflunisal treatment was associated with differences in GLS (+1.2% untreated vs +0.1% treated, P = .03). Changes in wall thickness (P = .2), left ventricular ejection fraction (P = .71), and BNP (P = .42) were similar between groups. CONCLUSIONS: In ATTR-CM, diflunisal treatment resulted in measurable differences in some parameters of cardiac structure and function after only 1 year of administration. Further longer-term analysis is warranted.