Characterization and natural history of different phenotypes in hereditary transthyretin amyloidosis: 40-year experience at a single Italian referral centre

Abstract

Aims

Hereditary transthyretin amyloidosis (ATTRv) is one of the leading aetiologies of systemic amyloidosis with more than 135 mutations described and a broad spectrum of clinical manifestations. We aimed to provide a systematic description of a population of individuals carrying pathogenic mutations of transthyretin (TTR) gene and to investigate the major clinical events during follow-up.

Methods and results

This was an observational, retrospective, cohort study including consecutive patients with mutations of TTR gene, admitted to a tertiary referral centre in Bologna, Italy, between 1984 and 2022. Three hundred twenty-five patients were included: 106 asymptomatic carriers, 49 cardiac phenotype, 49 neurological phenotype, and 121 mixed phenotype. Twenty-two different mutations were found, with Ile68Leu (41.8%), Val30Met (19%), and Glu89Gln (10%) being the most common. After a median follow-up of 51 months, 111 patients (38.3%) died and 9 (11.5%) of the 78 asymptomatic carriers developed ATTRv. Carriers had a prognosis comparable with healthy population, while no significant differences were seen among the three phenotypes adjusted by age. Age at diagnosis, New York Heart Association class III, left ventricular ejection fraction, modified polyneuropathy disability score IV, and disease-modifying therapy were independently associated with survival.

Conclusion

This study offers a wide and comprehensive overview of ATTRv from the point of view of a tertiary referral centre in Italy. Three main phenotypes can be identified (cardiac, neurological, and mixed) with specific clinical and instrumental features. Family screening programmes are essential to identify paucisymptomatic affected patients or unaffected carriers of the mutation, to be followed through the years. Lastly, disease-modifying therapy represents an evolving cornerstone of the management of ATTRv, with a great impact on mortality.

Graphical Abstract

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Introduction

Hereditary transthyretin amyloidosis (ATTRv) is a systemic disease characterized by autosomal dominant inheritance with incomplete penetrance and variable expressivity.^1,2 Mutations in the transthyretin (TTR) gene, a transport protein mainly synthesized by the liver, result in instability of the TTR tetramer, leading to the dissociation of the four monomers. These monomers undergo misfolding, generating mature insoluble amyloid fibres that deposit in various organs and tissues.³ To date, more than 135 mutations of the TTR gene have been described.⁴ Single-base substitutions resulting in missense mutations represent the majority of genetic alterations described. The most common mutations described worldwide are (in descending order) the Val30Met, Val122Ile, and Glu89Gln.⁵ In Italy, in addition to the Val30Met variant, endemic foci have been identified, particularly in the Tuscan–Emilian Apennines region and Sicily, for the mutations Ile68Leu, Glu89Gln, Phe64Leu, and Thr49Ala.^6,7 The clinical presentation can vary widely, ranging from phenotypes with exclusively neurological clinical presentation (such as Val30Met ‘early onset’) to predominantly cardiac phenotypes (mutations Thr60Ala, Leu111Met, Ile68Leu, and Val122Ile).⁸

We hereby present a monocentre retrospective study providing a systematic description of a population of individuals carrying pathogenic mutations for ATTRv, including both healthy carriers and those affected by neuropathy and/or cardiomyopathy. In particular, clinical and instrumental features of patients were analysed considering the phenotype at the time of presentation. Also, the onset of new clinical manifestations during follow-up and the progression of the disease in terms of survival and major clinical events were investigated in this study. Finally, the prognostic role of the analysed clinical variables and the impact of disease-modifying therapy (such as orthotopic heart or liver or combined transplantation and pharmacological strategies) on the prognosis of patients were examined.

Methods

Setting and study design

We conducted an observational, retrospective, longitudinal study with the aim to analyse the clinical and instrumental data collected in a cohort of consecutive patients, admitted to the amyloidosis centre of the IRCCS Sant'Orsola Hospital of Bologna, Italy, between 1984 and 2022. Inclusion criteria were patients ≥18 years old with a pathogenic mutation of the TTR gene. Genomic DNA was isolated from whole peripheral blood by standard techniques. Exons 2, 3, and 4 of the TTR gene (accession number M11844) were amplified by polymerase chain reaction. Amplified DNA fragments were directly sequenced using ABI Prism 3130 automated sequence. Data collection was performed with the use of a centralized database, with anonymized data regarding clinical, electrocardiographic, echocardiographic, and nuclear medicine parameters at the time of the first evaluation and during the follow-up. All patients received optimal medical therapy according to scientific evidence and current practice. That may include medications for heart failure (HF), pacemaker implantation for bradyarrhythmia, anticoagulants for atrial fibrillation, and drugs for comorbidities, according to clinical judgement. Data regarding disease-modifying therapy such as orthotopic heart and/or liver transplantation and drug therapy were also collected. At presentation, all patients provided informed written consent for the anonymous collection of scientific data. This study conforms to the principles of the Helsinki declaration, and all authors guarantee the integrity of data from their respective institutions. The study was approved by the ethics committee of IRCCS Sant'Orsola Hospital—University of Bologna, study code: (478/2019/Oss/AOUBo).

Definitions and diagnostic criteria

The diagnosis of ATTRv was made by genetic analysis on a blood sample. The genetic study involved the amplification by polymerase chain reaction of exons 2, 3, and 4 of the TTR gene. Subsequently, amplified DNA fragments were sequenced using the automatic sequencer ABI Prism 3130. The analysis was performed both in symptomatic subjects, affected by polyneuropathy and/or amyloid cardiomyopathy, and in first-degree relatives for screening purposes.

According to the traditional definitions shared by expert consensus on the disease,⁹ the phenotypes at the time of diagnosis were defined according to the following criteria:

1. Cardiac phenotype: patients with maximum wall thickness of the left ventricle in end-diastole (ventricular septum and posterior wall) >12 mm in the absence of other plausible causes of left ventricular hypertrophy, together with the exclusion of neurological involvement after assessment performed by an experienced neurologist. In addition, other echocardiographic signs suggestive of cardiac amyloidosis (CA) were considered, such as the ‘granular sparkling’ appearance of the left ventricle, thickening of the atrioventricular valves and interatrial septum, and the presence of pericardial effusion. The diagnosis of CA was either confirmed by histological proof of amyloid deposits or following non-biopsy criteria (i.e. patients with echocardiographic features of CA, associated with a grade 2–3 bone scintigraphy scan, in the absence of plasma cell dyscrasia, excluded by normal values of serum free light chains, and negative serum and urine protein electrophoresis with immunofixation), according to current guidelines¹⁰

2. Neurological phenotype: patients with walking disabilities and symptoms of neurological relevance (early satiety, nausea, vomiting, involuntary weight loss, diarrhoea, or faecal incontinence), as evaluated by a trained neurologist. When indicated, neurophysiological studies such as nerve conduction study, electromyography, and cardiovascular reflex studies were performed. In these patients, diagnostic criteria for CA are absent. Severity of disease was defined using the modified polyneuropathy disability (mPND) score classification: 0, asymptomatic; I, sensory disturbance in lower limbs but preserved walking capacity; II, difficulties in walking but no need of a walking stick; IIIa, one stick or one crutch required for walking; IIIb, two sticks or two crutches required for walking; and IV, patient confined to a wheelchair or bed.¹¹

3. Mixed phenotype: patient presenting clinical and instrumental features of both phenotypes.

Twelve-lead electrocardiogram (ECG) analysis was based on standard definitions. Abnormal ECG was defined as the presence of one or more of the following features: rhythm abnormalities, conduction disturbances (atrioventricular block, right bundle branch block, left bundle branch block, left anterior or posterior haemiblock), low QRS voltages (QRS amplitude ≤ 0.5 mV in all limb leads or ≤1 mV in all precordial leads) discordant with left ventricular wall thickness in transthoracic echocardiography, ST- and T-wave abnormalities, and ‘pseudo-infarction’ pattern.

Echocardiographic assessments and measurements were performed in keeping with the recommendations of the European Society of Cardiovascular Imaging.¹² Bone scintigraphy, performed with the aim of both confirming cardiac involvement and evaluating its appearance, consisted of the intravenous administration of 740 MBq of Tc-99m-3,3-diphosphono-1,2-propanodicarboxylicacid (DPD) with the subsequent late imaging evaluation (3 h after injection of the radiopharmaceutical) of the degree of myocardial uptake using the Perugini score.¹³

The presence of chronic kidney disease was defined as glomerular filtration rate (GFR) < 60 mL/min/1.73.

Follow-up

Time to follow-up started at the time of diagnostic genetic analysis for TTR mutation, and data recording was interrupted in August 2022. Ordinary timing of follow-up visit was every 18–24 months for unaffected carriers, every 9–12 months for affected patients with no or mild symptoms of the disease, and every 3–6 months for affected patients with moderate or severe symptoms or patients on disease-modifying therapy. Patients could also be visited at the clinic depending on individual clinical needs. The follow-up data derive from visits scheduled at the cardiology clinic of the IRCCS Sant'Orsola Hospital of Bologna, Italy, or through telephone contact with the patient himself and/or with the general practitioner in case of patients under treatment at other centres.

Carriers with a follow-up time of <12 months and patients with ATTRv with a follow-up of <6 months were excluded from the survival analysis. Major clinical events that occurred during the study period, such as death and worsening HF defined by at least one hospitalization for HF or urgent evaluation for HF requiring intravenous diuretic administration, were recorded.

Statistical analysis

Data suitable for our analysis were expressed as median and interquartile range (25th and 75th percentile) for the continuous quantitative variables. In case of qualitative and ordinal variables, they were expressed as absolute and relative frequencies.

In the contingency tables, the independence of categorical variables was tested using Fisher’s test or Pearson’s χ² test (according to Cochran’s rule). The independence of the continuous variables was analysed using the Mann–Whitney U/Kruskal–Wallis test. For multiple comparisons, we calculated P-values with the Bonferroni-adjusted test. Overall survival was analysed with Kaplan–Meier curves, whose statistical significance was evaluated using the log-rank test. Cox regression analysis was initially performed using clinical and instrumental variables in order to explore risk factors associated with overall mortality. Disease-modifying therapy was investigated as a time-varying parameter in a time-dependent Cox regression analysis. Then, a multivariate analysis was performed by entering the model the variables considered significant in the univariate analysis (P < 0.05). Statistical analyses were performed using Stata/IC 16.1 (StataCorp LP, College Station, TX, USA) and SPSS Statistics (version 27. IBM Corp., Chicago, IL). Data extraction was performed on 1 September 2022. For all tests, a two-sided P-value < 0.05 was required for statistical significance.

Results

Baseline characteristics

Three hundred twenty-five consecutive subjects carrying pathogenic mutations of the TTR gene were included in this study. Of these, 219 showed signs and symptoms of the disease and were divided into 3 phenotype categories based on the clinical–instrumental features recorded at first evaluation: 49 subjects presented a cardiac phenotype (15.1% of total population), 49 a neurological phenotype (15.1%), and 121 a mixed phenotype (37.2%). The enrolled population also included 106 subjects (32.6%) carrying a pathogenetic mutation of TTR gene in the absence of any clinical or instrumental manifestation of disease.

Table 1 shows the distribution of TTR mutations in the study population according to the phenotype. Our cohort includes 22 different point mutations. The mutations Ile68Leu (136 subjects, 41.8%), Val30Met (62 subjects, 19%), and Glu89Gln (33 subjects, 10%) are the most represented. Of the 62 subjects with the Val30Met mutation, 47 were diagnosed with the ATTRv disease. Among the leading mutations, Ile68Leu was the one with the highest percentage of healthy carriers (46%), while Glu89Gln had the lowest (6%). Dominant mutations were found in patients with cardiac and neurological phenotype (Ile68Leu and Val30Met, respectively), while patients with mixed phenotype had marked genotypic variability.

Table 2 summarizes the main clinical and instrumental features found in the population study divided by phenotypes at the first evaluation. Patients with cardiac phenotype were mainly males (73%), just like the group with the mixed phenotype (69%); on the other hand, the groups with carrier subjects and neurological phenotypes are composed respectively of 54% and 51% of women.

Patients with exclusive cardiac involvement were diagnosed with ATTRv at a significantly older median age (73.2 years, IQR 66.5–78.1) than patients with mixed phenotype (58.4 years, IQR 47–66.3; P < 0.0001) and those with only neurological involvement (44 years, IQR 33.5–62.9; P < 0.001). The age difference was also significantly different between the group with mixed phenotype and with neurological phenotype (P < 0.0001).

Carpal tunnel syndrome appears to be significantly more represented in patients with CA (cardiological phenotype 57.1%, mixed 59.1%) than in subjects with neurological phenotype or carriers (22.4% and 20%, respectively).

All carrier subjects were identified following family screening, while patients affected by the disease were identified through various diagnostic pathways. Most patients with mixed phenotype received the diagnosis of the disease because of neurological symptoms (54.5%), while 37.2% of them were diagnosed after cardiological evaluation and, to a small extent, in the context of family screening (8.3%).

In most patients with predominantly cardiological or neurological phenotypes, the diagnostic pathway matched with the clinical symptoms of the phenotype (respectively, 82% and 69% of cases), although a more relevant group of subjects with a neurological phenotype received a diagnosis through familial screening (30.6%).

Notably, 6.1% of patients with cardiological phenotype were diagnosed after an incidental finding of cardiac uptake on bone scintigraphy performed for other reasons while one patient (0.02%) after haematological evaluation for suspected AL amyloidosis.

Furthermore, patients with CA show worse exercise tolerance at baseline, with a statistically significant higher proportion (83%) of patients in New York Heart Association (NYHA) functional class ≥ II, compared with patients with mixed phenotype (60%), neurological phenotype, or no signs of the disease. Similarly, patients with polyneuropathy associated or not with cardiomyopathy have greater difficulty in walking than patients with exclusively cardiologic involvement or carriers.

Regarding ECG, pseudo-infarction patterns, first-degree atrioventricular block, left anterior fascicular block, and atrial fibrillation are the most common findings in CA, obviously more frequent than in neurological phenotypes and healthy carriers. At last, cardiac and mixed phenotypes have comparable values of left ventricular ejection fraction (LVEF) (56% vs. 58%), pericardial effusion (44% vs. 54%), restrictive filling pattern (45% vs. 39%), and slightly higher values of mean left ventricular wall thickness (17 vs. 15.7 mm).

Disease-modifying therapy

During the follow-up, 40 patients received orthotopic liver transplantation: of these, 18 had a neurological phenotype at the baseline and 22 had a mixed phenotype. Furthermore, 13 patients received a combined orthotopic heart and liver transplant, including 2 patients with solely cardiac involvement and 11 with mixed phenotype.

Only one patient with a mixed phenotype received isolated heart transplantation, due to intraoperative complications that prevented combined liver transplantation.

Also, we analysed patients who received pharmacological treatment. In our population, 29 patients were treated with tafamidis 20 mg (7 with neurological involvement, 22 with mixed phenotype) with a median time of therapy of 36 months (IQR 14–55 months); 8 patients were treated with tafamidis 61 mg (3 with cardiac phenotype and 5 with mixed phenotype; the latter were switched form tafamidis 20 mg); and 10 patients (1 with neurological involvement and 9 with mixed phenotype) were treated with patisiran for a median of 8 months (IQR 6–19) and 2 patients (1 with neurological involvement and 1 with mixed phenotype) with Inotersen. Most of the patients received disease-modifying therapy during follow-up, and the median interval between date of diagnosis and beginning of therapy was 22 months.

Finally, we analysed the prognostic impact of disease-modifying therapies in a time-dependent analysis (from date of diagnosis to initiation of therapy) among patients of our cohort, regardless of the disease phenotype. Figure 1 shows that patients treated with surgical and/or pharmacological strategies have a higher total survival (P < 0.00001) than patients treated exclusively with supportive therapy. No significant difference was found in single subgroups of patients treated with tafamidis 20 mg (P = 0.079), tafamidis 61 mg (P = 0.586), and patisiran (P = 0.098).

Follow-up analysis based on phenotypes

The study population of 325 subjects was observed for a median follow-up of 51 months (IQR 24–90 months). Of these, 21 healthy carrier subjects with a follow-up time of <12 months and 3 patients with cardiac and 1 mixed phenotype with follow-up <6 months were excluded from the survival analysis. Overall, 10 patients were lost to follow-up (6 healthy carriers, 2 patients with cardiac phenotype, and 2 with mixed phenotype).

During the observation period, 9 (11.5%) of the 78 healthy mutation carriers included in the analysis were diagnosed with ATTRv. Of these, three developed a cardiological phenotype (Thr59Lys, Arg34Thr, and Ile68Leu mutations), three developed a neurological phenotype (two with Ile68Leu mutation and one patient with Gly47Ala), and three people with Glu89Gln, Ala36pro, and Gly47Ala mutations developed a mixed phenotype. The median age of patients at diagnosis of ATTRv was 50 years (IQR 44–60) after a median follow-up of 65 months (IQR 43–109 months) from the first clinical evaluation.

Furthermore, we analysed the onset of new signs and symptoms of the disease: four patients with the Ile68Leu mutation, initially evaluated as suffering from isolated cardiac amyloidosis and then developed signs of neurological involvement after a median time of 31 months (IQR 17–44 months) and two patients with exclusively neurological involvement with Val30Met and Ala36Pro mutations developed amyloid cardiomyopathy (Figure 2).

At the end of the follow-up time, 290 patients were included in the final analysis (70 healthy carriers, 43 cardiac phenotype, 50 neurological phenotype, and 127 mixed phenotype). Of these, 111 (38.3%) died: 2 (2.8%) healthy carriers, 21 (48.8%) with cardiac phenotype, 19 (38%) with neurological, and 69 (54.3%) with mixed phenotype. At the time of death, the patients with the mixed phenotype had a median age of 61.4 years (IQR 53.5–69.6 years), those with the neurological phenotype were 60 years old (IQR 50.3–68.1), while the patients with the cardiac phenotype were significantly older with a median age of 77.7 years (IQR 71.3–81.3). No significant differences were found between the three phenotypes in the interval between symptom onset and death. Twelve patients (4.1%) presented worsening HF (four patients with cardiac phenotype and eight with mixed phenotypes).

Figure 3 shows the overall survival of the study population divided by phenotypes. Survival curves are adjusted by age, and no significant differences are seen among the three phenotypes of the disease. Subjects without signs and symptoms of disease have a total and event-free survival comparable with the healthy population.

The main clinical and instrumental parameters (sex, GFR, NYHA functional class at presentation, mPND score, age at diagnosis, disease-modifying therapy, LVEF, and phenotype and main TTR gene mutations) were initially investigated with a univariate analysis Cox regression aimed at identifying the predictors of death from any cause. A multivariate analysis model was subsequently performed with the parameters resulting statistically significant in the univariate analysis.

Age at diagnosis, exertional dyspnoea (NYHA class III), severe gait disturbances (mPND score IV), LVEF, and the use of disease-modifying therapy were found to be independently associated with prognosis (Table 3), while the presence of a specific genotype (Ile68Leu, Val30Met, other mutations) was not.

Discussion

Our study includes a large series of patients affected by ATTRv or healthy carriers of a mutation of the TTR gene and provides an accurate description of the clinical and instrumental features of the three main disease phenotypes. Also, we aimed to describe the phenotype–genotype correlation and to compare outcomes according to clinical manifestations and disease-modifying therapy.

It is common knowledge that ATTRv is a multi-organ disease characterized by an extreme clinical heterogeneity.³ In our population, three main disease phenotypes could be identified at baseline evaluation, with the mixed phenotype resulting the most prevalent (55.2% of enrolled patients). This result appears in keeping with previous literature, showing a high frequency of a multisystemic involvement in ATTRv cohorts. The presence in the enrolled population of 106 (32.6%) subjects carrying the mutation in the absence of disease was also remarkable. All carriers were identified thanks to the family screening programme, which underlines a greater sensitivity and awareness of the disease.

Among patients with amyloid cardiomyopathy, there is a clear predominance of the male sex, both in patients with isolated cardiac phenotype and in those with simultaneous neurological involvement; on the contrary, no gender difference emerged among patients with neurological phenotype and in carrier subjects. This data supports the hypothesis of a greater susceptibility of the male sex regarding the cardiac involvement of the disease.¹⁴

Comparing the three phenotype groups, it emerges an important difference in the age of onset of symptoms. In fact, subjects with the cardiac phenotype develop the first symptoms of the disease 15 years later [73.2 (66.5–78.1) years old] than patients with the mixed phenotype [58.4 (47–66.3) years old] and 29 years later than patients with the neurological phenotype [44 (33.5–62.9) years old]. If the difference between the cardiac and neurological phenotypes can be partly explained by the age-related penetrance of the different mutations (i.e. the high proportion of subjects with the Val30Met mutation with early onset of neurological symptoms), the reason for this time age gap appears less clear in patients with mixed phenotype. In fact, this group develops the first symptoms of the disease and are diagnosed with ATTRv at a younger age than patients with the cardiac phenotype, as the first symptom is neurological in 54.5% of cases and cardiological in only 37.2%. This differences in age observed in our cohort are in keeping with the published literature. Rapezzi et al.⁷ hypothesized that one of the reasons for this time gap is that myocardial infiltration in ATTR requires longer than the time required to develop axonal neuropathy. Another possible explanation could be that the absence of neurological symptoms causes patients to seek medical care later, but this hypothesis appears less likely as in our cohort there is no significant difference between the three groups in the time interval between symptom onset and diagnosis of the disease. This finding is in keeping with the higher prevalence of history of HF hospitalizations in subjects with isolated cardiac amyloidosis compared with the other phenotypes (P < 0.0001).

Regarding instrumental data, CA is confirmed to be characterized by a symmetrical increase in the wall thicknesses of the left ventricle with an ejection fraction that is only slightly impaired and by the coexistence of diastolic dysfunction and pericardial effusion. There are no substantial differences in the instrumental features of cardiac amyloidosis associated with neurological involvement or not.

Among the mixed phenotype population, we observed a lower posterior wall thickness and lower mean thickness, together with a smaller atrial diameter: this could be related to an earlier stage of the disease or to a lower presence of comorbidities such as hypertension, diabetes mellitus, and renal insufficiency. In favour of this hypothesis, there is a higher frequency of advanced NYHA class, history of hospitalization for HF, and atrial fibrillation in patients with the cardiac phenotype compared with those with the mixed phenotype (56% vs. 23%, P < 0.0001).

In this cohort, the presence of mono- or bilateral carpal tunnel syndrome is confirmed to be a possible ‘red flag’ of cardiac involvement,¹⁵ as it is significantly more present in patients with cardiomyopathy than in the other groups (57.1% in the group with cardiac phenotype, 59.1% in the group with the mixed phenotype, and 20% and 22.4%, respectively, in the healthy carriers and in the group with the neurological phenotype). Notably, in our population, the association between amyloid cardiomyopathy and low voltages on 12-lead ECG does not appear a useful marker to identify subjects at greater risk of cardiac involvement.

Our study cohort includes a total of 22 different mutations of the TTR gene. The most represented mutations are Ile68Leu (136 subjects, 42%) followed by Val30Met (62 subjects, 19%) and Glu89Gln (33 subjects, 10%). Although the patients enrolled in this study were referred to our centre from several Italian regions, the frequency analysis of the different mutations is clearly influenced by the geographical location of our clinic near the Tuscan–Emilian Apennines, which is known to be endemic for Ile68Leu mutation.¹⁶ This mutation appears to be one of the major causes of CA and accounts for 82% of cardiac phenotypes in this cohort. However, despite being known as a ‘cardiogenic mutation’, it has been previously demonstrated that in these patients, neurological involvement is not uncommon,¹⁷ especially autonomic dysfunction.¹⁸ As a matter of fact, 24% of the mixed phenotypes and 8% of the neurological phenotypes in our cohort are related to Ile68Leu mutations.

Val30Met mutation appears to be as one of the most frequent mutations, and it is strongly associated with the neurological manifestations of the disease, as it is found in 53% of neurological phenotypes and 17% of mixed phenotypes, while Glu89Gln mutation shows a mixed expression in 79% of cases. Regarding the remaining mutations, a certain clinical variability can be observed and so it is not possible to detect a close genotype–phenotype correlation.

The high clinical heterogeneity and the frequent multisystem involvement found in all forms of ATTRv confirm the need for a multidisciplinary, cardiological, and neurological evaluation, in all subjects, regardless of the mutation found.

Healthy carriers represent a relevant part of the study population, as a result of our extensive screening programme for relatives of affected patients. Ile68Leu is the mutation with the highest prevalence of asymptomatic carriers (46%), probably given to the elder age of disease onset in this specific subset, contrariwise to some other mutations such as Glu89Gln (6% of asymptomatic carries), more aggressive since young adult age, as already acknowledged.¹⁹

At the end of the follow-up of 51 months (IQR 24–90 months), 9 of the 78 healthy carrier subjects (11.5%) included in the analysis developed signs and symptoms of the disease, after a median time of 65 months (Figure 2). We did not observe neither a prevalent clinical presentation nor a prominent mutation, as three patients presented the onset of disease with cardiac amyloidosis, three with sensorimotor polyneuropathy, and three with both. This finding emphasizes the importance of following carriers through the years, as previously stated by experts’ consensus.^10,20 Hereditary transthyretin amyloidosis is also confirmed to be an adult disease, as the median age of the patients at the time of disease onset was 50 years (IQR 44–60). In only two cases, the disease began before the age of 40: at 28 years with a mixed phenotype in a patient carrying the Ala36Pro mutation and at 38 years with a neurological phenotype in a woman with the Gly47Ala mutation.

From the survival analysis, it emerged that 111 (38.3%) of the 290 patients included died, while 123 patients (42.4%) experienced a major clinical event between death and hospitalization for HF. As expected, carriers who have not developed ATTRv at the end of follow-up have a survival comparable with that of the general healthy population.

When comparing the Kaplan–Meier survival curves adjusted by age (Figure 3), no significant differences were seen among all phenotypes. Surprisingly, cardiac involvement was not significantly associated with mortality in our cohort. To the best of our knowledge, the main study focused on the prognostic impact of cardiac phenotype in ATTRv was conducted by Damy et al.²¹ within the THAOS registry. In that analysis, cardiac phenotype was associated with a worst outcome, but notably, ATTRwt patients were also included. The lack of significance in our analysis is influenced by several factors and limitations, including the nature of our study (retrospective and single centre cohort) and by the fact that many of our patients with exclusive neurological phenotype belong to decades ago (mainly 1980s and 1990s); thus, their prognosis could have been affected by the poor knowledge of the disease but still represents a real-life experience of a long-term follow-up in a tertiary referral centre. In our multivariate analysis, age at diagnosis along with three easy-to-obtain parameters reflecting disease severity emerged as independent variables of death: NYHA functional class and LVEF as measures of cardiomyopathy and HF severity and mPND classification as a parameter of motor disability. These findings corroborate the idea that severity of organ involvement impacts more on prognosis than the bare presence of cardiac or neurological disease. Notably, the presence of a specific genotype (Ile68Leu, Val30Met, other mutations) was not associated with any of the pre-specified endpoint. In another study about ATTRv published by researchers from the National Amyloidosis Centre in UK, patients with V122I-related ATTR-CA were more impaired functionally (P < 0.001) and had worse measures of cardiac disease (P < 0.001) at the time of diagnosis, a greater decline in quality of life, and poorer survival (P < 0.001) in comparison with the other subgroups.²² Those differences underline the heterogeneity of the impact of various TTR mutations, which may carry a worse prognostic role even within the same phenotype category (e.g. Val122Ile vs. Ile68Leu). Lastly, in the multivariate Cox regression model, starting a disease-modifying therapy (pharmacological or not) was strongly and independently associated with a better prognosis, as graphically confirmed by Kaplan–Meier survival curves in Figure 1. This observation is limited by some bias mainly related to the retrospective nature of the study, the initiation of therapy during follow-up, and the variability of the applied treatment (surgical or pharmacological). Although, this evidence of efficacy on survival still represents an important remark, given the lack of data essentially based on single studies about liver transplantation, especially in Val30Met,²³ and tafamidis on ATTR-CA.²⁴

Conclusion

This retrospective study offers a wide and comprehensive overview of ATTRv from the point of view of a tertiary referral centre in Italy. We performed an analysis on a large cohort of patients affected by ATTRv or healthy carriers of mutation of the TTR gene, which has allowed us to confirm the clinical heterogeneity of the disease, splitable into three main phenotypes (cardiac, neurological, and mixed) with specific clinical and instrumental features. Family screening programmes are essential to identify paucisymptomatic affected patients or unaffected carriers of the mutation, to be followed through the years. Lastly, disease-modifying therapy represents an evolving cornerstone of the management of ATTRv, with a great impact on mortality.

Study limitations

This study presents a large series of patients with ATTRv, but it represents the experience of a single centre, located in a geographic area which is endemic for a specific mutation and with a relatively small absolute number of patients for each phenotype. Furthermore, the prolonged interval of enrollment made the diagnostic techniques and therapeutic strategies applied in the study not homogeneous. For the same reason, we had to exclude from the analysis some important echocardiographic parameters such as a comprehensive evaluation of diastolic function, mitral annular systolic velocities, atrial volumes, and global longitudinal strain.

Finally, the evaluation of the independent prognostic factors is limited by the absence in our series of complete data for some variables with known prognostic impact [N-terminal prohormone of brain natriuretic peptide (NTproBNP), troponin] which, therefore, were not included in the multivariate analysis model.

Acknowledgements

We would like to acknowledge all participants from the present study.

Author contribution

A.G.C. and M.S. were responsible for conceiving the study, interpreting the results, and drafting the manuscript. A.A., Giul.S., A.P., A.G., P.M., I.R., Gius.S., A.Z., V.P., R.B., P.G., C.G., E.B., P.C., N.G., and S.L. were responsible for data collection, data interpretation, and editing of the manuscript.

Funding

The work reported in this publication was funded by the Ministero della Salute, RC-2022-2773270 project.

Data availability

The data underlying this article will be shared on reasonable request to the corresponding author.

Previous presentation

Part of this work has been presented to the ESC Congress 2023 Amsterdam.

References

Author notes