Mélanie Bézard

L’amylose AL est une maladie systémique grave caractérisée par le dépôt d’agrégats de chaines légères (CL) d’immunoglobuline sous forme de fibrilles amyloïdes. Les organes les plus souvent atteints sont le rein et le cœur mais c’est l’atteinte cardiaque qui conditionne le pronostic. Le traitement consiste en une combinaison d’agents chimiothérapeutiques ayant pour cible le clone de plasmocytes producteur de la CL produite en excès. Une de ces molécules est la dexaméthasone ayant des propriétés immunosuppressives. Une étude préliminaire menée sur 9 patients a démontré que la dexaméthasone entraîne un doublement du NT-proBNP chez les patients atteints d’une amylose AL cardiaque sévère. Les objectifs de cette thèse sont 1) d’étudier l’impact de la dexaméthasone sur les décès précoces d’origine cardiovasculaire chez les patients atteints d’une amylose AL cardiaque sévère et 2) de mettre en place un modèle d’étude in vitro afin d’étudier l’impact et les mécanismes d’action, d’une part des chaines légères amyloïdes et d’autre part de la dexaméthasone.Le premier défi de cette thèse a été d’obtenir des chaines légères libres (CLL). Pour cela, nous avons mis au point un protocole de purification des CLL par chromatographie d’exclusion de taille, à partir d’urine de patients. Ceci nous a permis d’obtenir 4 CLL différentes (une lambda et 3 kappa) provenant de patients atteints d’une amylose AL cardiaque sévère. Pour étudier les effets et la toxicité des ces CLL, nous disposons d’un modèle cellulaire 2D et d’un modèle 3D de sphéroïdes cardiaques, tous deux constitués de cardiomyocytes dérivés d’hiPSC. Ces différents modèles nous ont permis de montrer que les CLL induisaient une désorganisation de la structure des cardiomyocytes, avec une diminution et ou une agrégation des protéines SERCA-2a et troponine T cardiaque, ainsi qu’une diminution de la connexine-43. De plus, nous avons observé une altération de la transitoire calcique dans les sphéroïdes cardiaques.Par ailleurs, nous avons eu accès à des biopsies cardiaques de patients atteints d’amylose AL cardiaque, ce qui nous a permis de montrer également une désorganisation de la structure des cardiomyocytes in vivo chez l’Homme en cas d’amylose AL. Nous avons également pu démontrer qu’une partie des CL était internalisée dans les cardiomyocytes.Nous avons voulu ensuite étudier la toxicité et le mécanisme d’action de certains traitements de l’amylose AL, en nous focalisant sur la dexaméthasone. Pour cela, nous avons d’abord étudié la cardiotoxicité de la dexaméthasone chez l’Homme. Nous avons donc effectué une analyse rétrospective de patients atteints d’une amylose AL cardiaque sévère. Ceci nous a permis de démontrer que la dexaméthasone est associée à la survenue de décès précoces d’origine cardiovasculaire.Par la suite, nous avons voulu étudier les mécanismes d’action de la dexaméthasone sur notre modèle d’étude 3D de sphéroïdes cardiaques. Les résultats très préliminaires n’ont pas permis de mettre en évidence un effet de la dexaméthasone sur la transitoire calcique. Cependant, la dexaméthasone rétablir la quantité de c-TnT dans les sphéroïdes préalablement traités avec une CLL, ainsi que la quantité de SERCA-2a dans les sphéroïdes préalablement traités avec les CLL.

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Transthyretin amyloid cardiomyopathy (ATTR-CM) is increasingly diagnosed at an early stage of the disease natural history, defined as National Amyloidosis Centre (NAC) ATTR Stage I. The natural history of early-stage ATTR-CM remains poorly characterized. A retrospective multi-centre observational study of 879 patients with ATTR-CM, either wild-type TTR genotype or carrying the p.V142I TTR variant, and NAC ATTR Stage I biomarkers at the time of diagnosis who did not receive disease-modifying therapy for amyloidosis. Disease characteristics at diagnosis that were independently associated with mortality by Cox regression analysis were N-terminal pro-B-type natriuretic peptide (NT-proBNP), TTR genotype, and troponin T. Patients were categorized into NAC ATTR Stage Ia, defined as a furosemide equivalent diuretic requirement of 100 months in the 12% with Stage Ia disease [hazard ratio for death 5.06 (95% confidence interval 1.23–20.87); P = 0.025] despite significant cardiovascular morbidity at the time of diagnosis which increased during follow-up, including among patients diagnosed in NAC ATTR Stage Ia. Estimated survival among UK NAC ATTR Stage Ia patients was comparable to UK general population controls (P = 0.297). Patients with NAC ATTR Stage I ATTR-CM can be further stratified according to NT-proBNP concentration and diuretic requirement at diagnosis. Patients with Stage Ia ATTR-CM have significant cardiovascular morbidity despite good short- and mid-term survival. Patients diagnosed with NAC ATTR Stage Ia amyloidosis have an estimated median survival in excess of 100 months in the absence of disease-modifying therapy despite significant cardiovascular morbidity which increases further during follow up.

Background: We assesse the evolution and prognostic value of N-terminal pro-brain natriuretic peptide (NT-proBNP) and high-sensitivity troponin T (cTnT-HS) in transthyretin amyloid cardiomyopathy (ATTR-CA) before and after tafamidis treatment. Methods and Results: 454 ATTR-CA patients without tafamidis (Cohort A) and 248 ATTR-CA with tafamidis (Cohort B) were enrolled. Event-free survival (EFS) events were death, heart transplant, or acute heart failure. In Cohort A, 27% of patients maintained NT-proBNP < 3000 ng/L and 14% cTnT-HS < 50 ng/L at 12 months relative to baseline levels. In Cohort B, the proportions were 49% and 29%, respectively. In Cohort A, among the 333 patients without an increased NT-proBNP > 50% relative to baseline EFS was extended compared to the 121 patients with an increased NT-proBNP > 50% (HR: 0.75 [0.57; 0.98]; p = 0.032). In Cohort A, baseline NT-proBNP > 3000 ng/L and cTnT-HS > 50 ng/L and a relative increase of NT-proBNP > 50% during follow-up were independent prognostic factors of EFS. The slopes of logs NT-proBNP and cTnT-HS increased with time before and stabilized after tafamidis. Conclusion: ATTR-CA patients with increasing NT-proBNP had an increased risk of EFS. Tafamidis stabilize NT-proBNP and cTnT-HS increasing, even if initial NT-proBNP levels were >3000 ng/L. Thus suggesting that all patients, irrespective of baseline NT-proBNP levels, may benefit from tafamidis.

Background: Cardiac light chain amyloidosis (AL-CA) patients often die within three months of starting chemotherapy. Chemotherapy for non-immunoglobulin M gammopathy with AL-CA frequently includes bortezomib (Bor), cyclophosphamide (Cy), and dexamethasone (D). We previously reported that NT-ProBNP levels can double within 24h of dexamethasone administration, suggesting a deleterious impact on cardiac function. In this study, we evaluate the role of dexamethasone in early cardiovascular mortality during treatment. Methods and findings: We retrospectively assessed 100 de novo cardiac AL patients (62% male, mean age 68 years) treated at our institute between 2009 and 2018 following three chemotherapy regimens: CyBorDComb (all initiated on day 1; 34 patients), DCyBorSeq (D, day 1; Cy, day 8; Bor, day 15; 17 patients), and CyBorDSeq (Cy, day 1; Bor, day 8; D, day 15; 49 patients). The primary endpoint was cardiovascular mortality and cardiac transplantation at days 22 and 455. At day 22, mortality was 20.6% with CyBorDComb, 23.5% with DCyBorSeq, and 0% with CyBorDSeq (p = 0.003). At day 455, mortality was not significantly different between regimens (p = 0.195). Acute toxicity of dexamethasone was evaluated on myocardial function using a rat model of isolated perfused heart. Administration of dexamethasone induced a decrease in left ventricular myocardium contractility and relaxation (p<0.05), supporting a potential negative inotropic effect of dexamethasone in AL-CA patients with severe cardiac involvement. Conclusion: Delaying dexamethasone during the first chemotherapy cycle reduces the number of early deaths without extending survival. It is clear that dexamethasone is beneficial in the long-term treatment of patients with AL-CA. However, the initial introduction of dexamethasone during treatment is critical, but may be associated with early cardiac deaths in severe CA. Thus, it is important to consider the dosage and timing of dexamethasone introduction on a patient-severity basis. The impact of dexamethasone in the treatment of AL-CA needs further investigation.

Aims: Hereditary (ATTRv) and wild‐type (ATTRwt) transthyretin amyloidosis are severe and fatal systemic diseases, characterised by amyloid fibrillar accumulation principally in the heart or peripheral nerves (or both). Since 2012, tafamidis has been used in France to treat patients with ATTRv with neuropathy (alone or combined with cardiomyopathy). Recently, the Phase III ATTR‐ACT trial showed that tafamidis decreased the relative risk of mortality in ATTR amyloidosis with cardiomyopathy. The aims of this study were to assess the clinical characteristics of ATTR amyloidosis in a real‐life population in comparison to the population included in the ATTR‐ACT trial and to assess the impact of tafamidis treatment on major cardiovascular outcome (MCO)‐free survival time without cardiac decompensation, heart transplant, or death. Methods and results: From June 2008 to November 2018, 648 patients with ATTR amyloidosis (423 ATTRwt and 225 ATTRv) consecutively referred to the French Referral Center for cardiac amyloidosis were included. A total of 467 (72%) patients matched the inclusion criteria of the ATTR‐ACT trial. For the 631 patients with cardiomyopathy, tafamidis treatment was associated with a longer median MCO‐free survival time ( n = 98): 1565 (1010–2400) days vs. 771 (686–895) days without treatment (log‐rank P < 0.001). This association was confirmed after considering confounding factors (age at inclusion, N‐terminal pro‐B‐type natriuretic peptide and amyloidosis type) with a propensity score (hazard ratio 0.546; P = 0.0132). Conclusion: In a large cohort of ATTRwt and ATTRv patients, representative of the inclusion criteria of the ATTR‐ACT trial, the present results show an association between tafamidis treatment and a lower occurrence of cardiovascular outcomes in a real‐life population.