Transthyretin Amyloidosis Cardiomyopathy in Greece: Clinical Insights from the National Referral Center.

BACKGROUND: Clinical characteristics and outcomes of patients with transthyretin amyloidosis cardiomyopathy (ATTR-CM) vary by region, necessitating the acquisition of country-specific evidence for proper management. METHODS: This is an observational study including sequential patients presenting in the Amyloidosis Reference Center of Greece, from 01/2014 to 12/2022. ATTR-CM was diagnosed by positive scintigraphy and exclusion of light-chain amyloidosis or positive biopsy typing. Genetic testing was performed in all cases. RESULTS: One-hundred and nine ATTR-CM patients were included (median age, 81 years) of which 15 carried TTR mutations (27% Val30Met). Most patients (82%) presented with heart failure and 59% with atrial fibrillation, while 10% had aortic stenosis. Importantly, 78 (71.6%) had clinically significant extracardiac manifestations (45% musculoskeletal disorder, 40% peripheral neuropathy and 33% gastrointestinal symptoms). Sixty-five (60%) received disease-specific treatment with tafamidis. Estimated median survival was 48 months; advanced NYHA class, National Amyloidosis Center stage, eGFR<45 ml/kg/1.73m2, NT-pro-BNP>5000 pg/mL were associated with worse survival, while tafamidis treatment was associated with improved survival in patients with IVS≥ 12 mm. DISCUSSION: These are the first data describing the characteristics, management, and outcomes of patients with ATTR-CM in Greece, which could influence local guidelines. SHORT TITLE: Transthyretin cardiomyopathy in Greece.

Variant transthyretin amyloidosis (ATTRv) polyneuropathy in Greece: a broad overview with a focus on non-endemic unexplored regions of the country.

Comprehensive data on variant transthyretin amyloidosis polyneuropathy (ATTRv-PN) in Greece are lacking. We presently provide an overview of ATTRv-PN in Greece, focusing on unexplored non-endemic regions of the country. In total, we identified 57 cases of ATTRv-PN diagnosed over the past 25 years, including 30 from the island of Crete, an apparent endemic region. Patients carried 10 different TTR mutations (C10R; P24S; V30M; R34G; R34T; I68L; A81T; E89Q; E89K and V94A). Carriers of the common V30M mutation constituted 54.3 % of the cohort. A known founder effect for the V30M mutation was present on the island of Crete. Non-endemic cases identified outside the island of Crete are presently reported in more detail. The age of onset ranged from 25 to 77 years, with a mean of 51.1 years. A mean diagnostic delay of 3.2 years was observed. V30M patients had earlier onset and less cardiac involvement than patients carrying other mutations. Genotype-phenotype correlations were largely consistent with published data. We conclude that, with the exception of the Cretan cluster, ATTRv-PN is not endemic in the Greek population. This makes timely diagnosis more challenging, yet absolutely essential given the availability of therapies that can alter the long-term course of the disease.

Next generation flow cytometry for MRD detection in patients with AL amyloidosis.

The treatment of AL amyloidosis aims to eradicate the plasma cell clone and eliminate toxic free light chain production. Only in a minority of patients the plasma cell clone is completely eradicated; residual light chain production may still exist while clonal relapse may occur. We used sensitive next-generation flow cytometry (NGF) to detect minimal residual disease (MRD) in AL amyloidosis patients at complete haematologic response. MRD evaluation was feasible in 51 of 52 (98%) tested patients and at a median sensitivity of 2.3 × 10-6 MRD was undetectable in 23 (45%). An organ response occurred in 86% of MRDneg vs 77% in MRDpos; renal response in 15/17(88%) of MRDneg vs in 14/16(87.5%) of MRDpos and cardiac response in 10/10(100%) of MRDneg vs 11/15(73%) of MRDpos patients. After a median follow-up of 24 months post MRD testing, no MRDneg patient had a haematologic relapse vs 6/28(21%) MRDpos (p = .029). Pooling haematologic and organ progressions, 9 (32%) MRDpos patients had disease progression vs only 1 (4%) MRDneg patient (p = .026). In conclusion, MRD detection using NGF has profound clinical implications, so that AL patients with undetectable MRD have a very high probability of organ response and a very low probability of haematologic relapse.

Timing and impact of a deep response in the outcome of patients with systemic light chain (AL) amyloidosis.

A rapid and deep haematologic response is fundamental in order to improve outcomes of patients with AL amyloidosis. We evaluated the impact of timing and depth of haematologic response at early time points (at 1 and 3 months from the start of therapy) in 227 consecutive previously untreated AL patients, who received bortezomib-based primary therapy. After 1 month of therapy, 30.5% had ≥VGPR, 28% PR and 36% no response (NR), with 11% having iFLC <20 mg/L and 15% dFLC <10 mg/L. Deep haematologic response at 1 month (either ≥VGPR or iFLC <20 mg/L or dFLC <10 mg/L), was associated with a high organ response rate. The survival of patients with ≥VGPR was significantly better than those with PR and NR at 1-month landmark (p < .001) but this benefit was mainly driven by those with iFLC <20 mg/L. The depth of haematologic response at 1 month was significant across all Mayo stages. At 3 months, 46% of the patients had not significantly improved the depth of their response but even patients that improved their response from an iFLC ≥20 mg/L at 1 month to iFLC <20 mg/L at 3 months still had inferior outcome to those with an early deep response. Thus, in patients with AL amyloidosis, a very rapid and deep response is crucial, especially for those at high risk, targeting very low FLC levels within the first month of therapy.

Familial MTC with RET exon 8 Gly533Cys mutation: origin and prevalence of second malignancy.

INTRODUCTION: High prevalence of RET p.Gly533Cys (c.1597G > T) has been found in familial MTC in Greece (exon 8 fMTC). We studied their origin and compared clinical characteristics with non-exon 8 fMTC. METHODS: 102 fMTC (FMTC and MEN2A) patients (31.4% males) were followed for 2.9-37 years (median 6 years). Fifty-one carried the RET exon 8 mutation; the remaining were non-exon 8 fMTC (exons 10, 11, 13, 14). Pre-, post-operative calcitonin, disease extent at diagnosis and follow-up and families' place of origin were recorded. RESULTS: Exon 8 fMTC were older (42.3 ± 13.3 vs 30.8 ± 17.8 years, P < 0.001), including index cases (P = 0.016). In index cases, the stage at diagnosis was more favorable in exon 8 fMTC compared to non-exon 8 fMTC (stage I and II: 65% vs 23.8%, stage III: 25% vs 57.1%, stage IV: 10% vs 19%, P = 0.025). More favorable outcome was noted in exon 8 fMTCs (remission: 72.5% vs 45.8%, stable disease: 27.5% vs 41.7%, progression: 0.0% vs 12.5%, P = 0.001). Exon 8 fMTC patients carried more frequently a second malignancy (25.5% vs 6.3%, P = 0.009); 69% of these were PTCs. Exon 8 fMTC patients were significantly older at diagnosis compared to non-exon 8 moderate-risk RET carriers and presented more favorable clinical outcome (remission: 72.5% vs 50%, stable disease: 27.5% vs 41.7%, progression: 0.0% vs 8.3%, P = 0.021). This difference remained when only index cases were analyzed. 'Hot spots' in the origin of exon 8 fMTCs families were recognized. No phenotype or outcome differences were found between the exon 8 families from the various regions. CONCLUSIONS: In exon 8 fMTCs' older age, favorable disease stage at diagnosis and favorable outcome suggest slow disease progression compared to non-exon 8 fMTC. Compared with moderate-risk RET mutation carriers, exon 8 fMTC patients have a more favorable clinical outcome. The higher prevalence of second malignancies, especially PTC, not previously reported, merits further investigation. Increased awareness for inherited disease is required for patients with apparently sporadic MTC originating from recognized 'hot spots', as the age at presentation is usually delayed.

Evidence for the founder effect of RET533 as the common Greek and Brazilian ancestor spreading multiple endocrine neoplasia 2A.

OBJECTIVES: About one-quarter of patients with medullary thyroid cancer (MTC) have inherited disease due to mutations in the RET gene. A rare mutation in exon 8 (G533C) of RET, previously described in a large Brazilian family with MEN2A, also appeared to be clustering in Greece, whereas it was rarely reported in other ethnic groups. The aim of this study was to identify a possible common ancestry between these carriers. PATIENTS AND METHODS: Twelve RET G533C mutation carriers, four randomly selected from the Brazilian cohort and eight from apparently unrelated Greek families, were studied for a possible common ancestral origin. RET flanking microsatellite markers at chromosome 10q (D10S197, D10S196, D10S1652 and D10S537) were used. RESULTS: Genomic DNA analysis using these markers showed that many of these apparently unrelated individuals shared a common haplotype indicating a common ancestral origin. CONCLUSION: Our data suggest that Brazilian and Greek patients with MTC carrying the G533C mutation in exon 8 of RET gene originate from a common ancestor. Due to historical reasons, we speculate that the more plausible explanation for the origin of this mutation is in Greece.

Medullary thyroid carcinoma: the influence of policy changing in clinical characteristics and disease progression.

OBJECTIVE: Medullary thyroid carcinoma (MTC) has varying clinical course. We assessed trends in MTC presentation during the last 34 years. DESIGN: Retrospective study. METHODS: One hundred and fifty one patients (44.4% males) were followed for 0.934 years. Patients were classified according to year of diagnosis: group 1, 1977-2000 (n=53) and group 2, 2001-2011 (n=98). Extent of disease at diagnosis, during follow-up, number of surgeries, and pre- and postoperative calcitonin levels were recorded. RESULTS: In total, 48.34% reported family history of MTC. Group 1 had larger tumors (median 1.70 (intraquartile range (IQR) 1.7) vs 1.1 (1.2) cm, P=0.045, Mann-Whitney), they presented less frequently micro-MTCs (27.8 vs 46.1%, P=0.045), and underwent more multiple surgeries (63.3 vs 20.0%, P<0.001). Group 1 had more frequently progressive disease (35.8 vs 12.2%, P=0.003) and distant metastasis at follow-up (39.7 vs 17.4%, P=0.017). Chronological group (HR 0.15, 95% CI 0.03-0.68, P=0.015) and distant metastases at follow-up (HR 0.07, 95% CI 0.015-0.30, P=0.001) were independently associated with 10-year disease progression (P<0.001). In sporadic cases, cervical lymph node invasion and distant metastases at diagnosis were more frequent in group 1 (72.7 vs 45.5%, P=0.032 and 27.3 vs 5%, P=0.019 respectively); disease stage at diagnosis was more advanced (P=0.004). They underwent more multiple surgeries (P<0.001), presented more frequently distant metastasis at follow-up (67.7 vs 20.0%, P=0.002), had less frequently remission, and more frequently progressive disease (21.4 vs 58.0% and 64.3 vs 14.0% respectively, P<0.001). Postoperative calcitonin levels were higher (P=0.024). CONCLUSIONS: Recently, an increase in micro-MTCs is observed, while indices of invasiveness and persistence of disease are better. Increased awareness in familial cases, routine calcitonin measurements, and improved surgical procedures could be responsible.

Severity of coronary artery disease in postmenopausal women: association with the androgen receptor gene (CAG)n repeat polymorphism.

OBJECTIVE: Androgen may adversely affect vascular health in women. We investigated the associations between the androgen receptor gene (CAG)n repeat polymorphism, which affects androgen receptor transcriptional activity, and the severity of coronary artery disease (CAD) in women undergoing coronary angiography. METHODS: We examined 131 postmenopausal women (46-82 y). CAD severity was assessed by the number of vessels with greater than 50% stenosis. The history of angina, myocardial infarctions, and biochemical parameters were recorded. CAG repeats ranged between 13 and 24 in the shorter allele and 16 and 28 in the longer allele. The mean lowest quartile corresponded to 19, and the highest, to 22 repeats. RESULTS: Carriers of 19 repeats or less in their shorter allele had severe disease (≥2 vessels affected) and a history of angina more frequently than those carrying 22 or more (39.2% vs 9.5%, P = 0.009 and 80.8% vs 55%, P = 0.037, respectively, using the Fisher exact test). A higher percentage of women carrying 19 repeats or less had one and two myocardial infarctions (28.6% and 10.7%, respectively) compared with women with more than 19 repeats (18.2% and 1.45%, respectively, P = 0.019). Women homozygous for two longer alleles (≥22 repeats) had less severe CAD, significantly higher sex hormone-binding globulin levels, and less frequent antilipid drug therapy compared with those homozygous for shorter alleles (P < 0.05). Total cholesterol and low-density lipoprotein cholesterol levels were negatively correlated with the number of repeats in the shorter allele (P < 0.04). CONCLUSIONS: Shorter polyglutamine stretch in the androgen receptor correlates with more severe CAD and worse predisposing factors in postmenopausal women undergoing coronary angiography. This association may support the adverse cardiovascular effect of lifelong androgenic exposure in this selected group of women.

Alterations in signal transduction pathways implicated in tumour progression during multistage mouse skin carcinogenesis.

The multistage mouse skin carcinogenesis model, although an artificial one, is an ideal system to study the timing of qualitative and quantitative alterations which take place during the different stages of chemical carcinogenesis, allowing analysis of the events that lead to the transition from the stage of initiation to the stage of promotion and finally to the progression of carcinogenesis. In this review we focus on the role of the H-ras gene and its target molecules during mouse skin carcinogenesis. Besides H-ras, which is a critical target of chemical carcinogens, we report alterations in oncosuppressor genes. Finally, we examine the potential suppression of metastatic dynamics of spindle cells after biological or chemical inhibition of the signalling cascades involved in mouse skin carcinogenesis.