- 學位論文
馬凡氏症候群流行病學、分子基礎醫學與血管張力素II受體阻斷劑療效之研究
Changing Epidemiology, Molecular Basis and Therapeutic Efficacy of Angiotension II Receptor Blockade in Marfan Syndrome
摘要
背景 馬凡氏症候群是一種全身性結締組織疾病，其中造成主動脈根部逐漸擴大，甚至發生主動脈剝離、破裂、或心包膜填塞等併發症，是導致馬凡氏症候群死亡的主要原因。致病原因主要是第15對染色體上的fibrillin-1 (FBN1)基因發生突變，導致體內轉型生長因子 (Transforming growth factor-β, TGF-β) 被大量活化，致使細胞外基質 (Extracellular matrix, ECM) 被分解，改變主動脈血管壁組織結構，最後形成主動脈瘤。 近年來，隨著科學與醫療技術的進步，馬凡氏症候群的預後已呈現穩定地改善；自1972年，患者的平均死亡與累積存活年齡分別為32及48歲，到1995年已延長至41及72歲。許多新的藥物治療及手術方式正陸續發展中，如何幫助這些病人，降低其死亡風險，延長生命週期，提升良好的生活品質，一直是醫療照護者的主要課題。 由於馬凡氏症候群的臨床表現多變，差異性大，增加了診斷的困難性，發生率或盛行率可能因此被錯誤的估算；在台灣，尚未有較完整的馬凡氏症候群疾病分析的相關資料；因此，本論文研究的第一部分，我們希望能建立台灣馬凡氏症候群的本土資料，利用健保資料庫，分析疾病的發生率、盛行率、心血管事件發生情形以及於各年齡層的分佈狀況，期望將來能透過預防醫學保健的觀點，幫助患者延長壽命，提高生活品質。Fibrillin-1基因相當巨大，包含65個外顯子 (exons)，目前約有超過1000種不同的fibrillin-1基因突變陸續被發現，然而，至今尚無明確的研究顯示突變位置與臨床表徵的相關性；因此，本論文研究的第二部分，我們希望收集足夠的基因突變資料，分析病患的基因型與表現型，試圖找出兩者間之關聯性，期待將來能作為疾病預後預測的參考。近年來，在馬凡氏症老鼠的動物模式成功被建立後，致病機轉逐漸清楚，藥物治療方面也有顯著的進步；動物實驗發現Losartan，一種血管張力素受體阻斷劑 (angiotensin receptor blocker)，有助於延緩主動脈持續擴大的情形，因此，本論文研究的第三部分，我們設計了一Losartan藥物臨床試驗，欲探討藥物之安全性與療效，希望能為病患證實與提供一個安全有效的治療方法。 研究方法與結果 第一部分：馬凡氏症候群在台灣的流行病學 自2000年1月1日至2012年12月31日健保資料庫中，總健保人數有22,765,535人，其中2329位病患 (58%為男性) 被診斷為馬凡氏症候群，其盛行率約為10.2 (95% CI, 9.8-10.7)/100,000，年齡層分佈則以15-19歲，10-14歲以及20-24歲為最多；我們再以現今20-29歲年齡層為對象，分析馬凡氏症候群的最低出生發生率約為23.3 (95% CI, 21.7-23.3)/100,000。追蹤期間共69人死亡，年平均死亡率約為0.23%，死亡原因主要與心臟有關，其中有40位病患是死於主動脈剝離或發生猝死；有226位病患 (9.7%, 61%為男性) 曾發生主動脈剝離，發生的平均年齡約為36.6 ± 10.7歲；發生主動脈剝離的機會在20、40、50歲的年齡層分別為0.6%、20.4%、34.5%。在69位死亡與226位主動脈剝離患者中，超過一半的病患其事件發生於40歲前；有360位病患曾接受心血管介入治療，包含心導管或手術治療，其中接受緊急手術治療者比接受非緊急手術治療者有較高的死亡率 (7.6%與0%)。 第二部分：馬凡氏症候群基因突變與臨床表現之關聯性 研究期間，我們納入了125位符合1996年Ghent準則且證實有fibrillin-1基因突變的馬凡氏症候群患者，共發現85種不同的基因突變，包括60種missense突變、9種nonsense突變、15種frame shift突變、以及一個全外顯子 (exon 36) 缺損突變。以突變位置而言，大部份的基因突變發生於外顯子11-24 (28.2%) 處，其次為外顯子25-40 (22.4%)；若以蛋白結構之特殊功能區域 (motif) 為單位，大部份(63.5%) 基因突變發生於鈣離子鍵結之類表皮生長因子區域 (calcium-binding epidermal growth factor (cbEGF)-like motif)，其次 (18.8%) 為類轉型生長因子區域(transforming growth factor (TGF)-β binding protein-like motif)。而年齡之分佈以及孩童患者 (小於10歲) 之分佈情形在fibrillin-1基因上呈現平均分佈狀態，並未發現好發於哪些特定區域。 在基因型與表現型之關係中，我們發現突變於外顯子1-24處與重大的眼睛問題相關，但骨骼系統影響較小；突變於外顯子41-57處與重大心血管及骨骼系統問題相關，但較不易影響眼睛；突變於外顯子58-65處，臨床症狀多較為輕微。若以蛋白結構之特殊功能區域 (motif) 為而言，突變於前端NH2處，其骨骼系統問題一般較為輕微；突變於混合區域 (hybrid motif)，多與重大心血管問題相關；突變於末端COOH區域，臨床症狀多較為輕微。若以突變方式而言；nonsense型突變多與重大心血管問題相關。 第三部分：馬凡氏症候群之藥物臨床試驗 自2007年5月至2010年12月我們共納入28位 (男/女：12/16，平均年齡：16.5 ± 6.6歲) 符合1996年Ghent準則合併主動脈擴大 (Z分數 > 2.0) 的馬凡氏症候群患者，進行隨機分配；一組繼續傳統乙型阻斷劑治療，另一組則加上試驗用藥Losartan，一種血管張力素受體阻斷劑 (angiotensin receptor blocker)。經過35個月藥物治療後，發現合併血管張力素受體阻斷劑與乙型阻斷劑治療比起單獨使用乙型阻斷劑，明顯地可以減緩主動脈的生長速度 (0.10公釐/年 vs 0.89公釐/年，p = 0.026)；此現象在aortic annulus及sinotubular junction處亦得到相似的結果 (p < 0.05)。然而，在主動脈彈性度方面，aortic distensibility以及cross-sectional compliance在兩組裡並無顯著的差異 (p = 0.462及0.535)。在試驗期間，無重大不良事件發生，不良反應於兩組間亦無顯著的差異。 結論 經研究我們發現，雖然近年來醫療技術的蓬勃發展，然而對於馬凡氏症候群患者仍有約十分之一的比例可能發生主動脈剝離，帶來較高的失能或死亡風險，因此早期診斷、早期接受治療越發顯得重要。透過基因檢測，除了有助於疾病之診斷外，亦能經由基因突變位置推測臨床表徵之嚴重度，可幫助病患決定長期追蹤的重點方向，建議其合適的運動方針，保持良好的生活體能；亦能應用於下一代，對胎兒或嬰幼兒進行基因檢測，有助於早期診斷與治療，也希望能運用於受孕前之基因篩檢，找出無基因突變之精卵進行人工受孕，以確保胎兒的健康。此外，我們也證實合併血管張力素受體阻斷劑 (Losartan) 與乙型阻斷劑治療，對於馬凡氏症候群患者提供了一種安全又有效的治療方式，能延緩主動脈持續擴大，降低主動脈剝離的風險。隨著馬凡氏症候群的致病機轉與病生理學越來越清楚，我們預期更多新的治療方法將陸續被研發，而病患的壽命與生活品質亦將獲得更好地改善。
並列摘要
Background Marfan syndrome is a multisystem connective tissue disorder, which may lead to progressive aortic root dilation or even aortic dissection, rupture, or cardiac tamponade, the most life-threatening complications. It is caused by mutations in fibrillin-1 (FBN1) gene, which is located in chromosome 15q21.1, and leads to excessive transforming growth factor-β (TGF-β) signaling and results in aberrant thickening of the aortic media with fragmentation and disarray of elastic fiber. Such changes would contribute to the formation of aortic aneurysms. With the advance in medical and surgical management, the life expectancy and outcomes of Marfan syndrome increased significantly over the last few decades. The mean age of mortality and cumulative survival age are 32 and 48 years in 1972 and 41 and 72 years in 1995, respectively. Cardiovascular manifestations of Marfan syndrome remain among the central issues in diagnosis and management, how to reduce the risks of morbidity and mortality, prolong the lifespan and provide a good life quality are incumbents and most important issues on healthcare professionals. Marfan syndrome has wide disease spectrum from mild to severe, which increase the difficulty in correct diagnosis. The incidence or prevalence rate may be misestimated. Such data concerning Marfan syndrome in Taiwan are still scanty. Therefore, in the first part of this thesis, we wish to explore the current epidemiological profile, including incidence, prevalence, cardiovascular event, age distribution, and long-term outcomes of Marfan syndrome in a general population by using the database from Taiwan National Health Insurance. Fibrillin-1 gene is a huge gene and consists of 65 exons. Up to date, more than 1000 fibrillin-1 gene mutations have been described which leads to variable clinical manifestations. However, there are no established correlations between the genotype and the phenotype. Therefore, in the second part of this thesis, we collected the clinical and genetic information of all patients with FBN1 gene mutation and fulfilling the 1996 Ghent criteria in our hospital, try to find the correlations between FBN1 genotype and phenotype in an Asia cohort. Recently, a mouse model of Marfan syndrome have been created successfully, which promises to provide insights into the pathogenesis of Marfan syndrome. Losartan, an angiotensin II receptor blockade (ARB), had been shown to prevent aortic root dilation in the mouse model of Marfan syndrome. However, the effects in human have never been assessed. In the third part of this thesis, we conducted a clinical trial of Losartan to investigate the safety and efficacy of ARB added-on beta-blockade on attenuation of aortic root dilation in patients with Marfan syndrome. Methods and Results Part I: The epidemiology profile of Marfan syndrome in Taiwan To explore the current epidemiological profile of Marfan syndrome in Taiwan, we used Taiwan National Health Insurance database from January 1, 2000 through December 31, 2012 (population 22,765,535) for analysis. We identified 2329 patients (58% male) with Marfan syndrome. The overall prevalence rate was 10.2 (95% CI, 9.8-10.7)/100,000 with the peaks at 15-19, 10-14 and 20-24 years. Minimal birth incidence of 23.3 (95% CI, 21.7-23.3)/100,000 was estimated from those aged 20-29 years. The average annual mortality was 0.23% (69 deaths), mostly due to cardiac causes (including dissection and sudden death in 40 patients). Aortic dissection occurred in 226 (9.7%) patients (61% male) at a mean age of 36.6 ± 10.7 years. The freedom from dissection was 99.4%, 79.6%, and 65.5% at the age 20, 40 and 50 years, respectively. Among the 69 deaths and 226 dissections during the follow up period, over half of cases occurred before the age of 40 years. Cardiovascular intervention was performed in 360 patients, with higher early mortality (7.6%) in the emergent surgery group as compared to that in elective group (0%). Part II: The correlations between fibrillin-1 genotype and phenotype in Marfan syndrome A total of 85 mutations were identified in 125 proved MFS patients. The mutation comprised 60 missense, 9 nonsense, 15 frame shift, and 1 whole exon 36 deletion. Most mutations located in exon 11-24 (28.2%), then exon 25-40 (22.4%), and in cbEGF (calcium-binding epidermal growth factor) like motif (63.5%), then TGFBP (transforming growth factor-β binding protein) like motif (18.8%). The distributions of age and young patients number were even in the whole gene without statistical significance. The correlation of genotype and phenotype based on the exonic location showed that mutations in exon 1-24 were associated with major ocular symptoms and less skeletal involvement. Mutations in exon 41-57 were associated with major cardiovascular and skeletal symptoms but less ocular involvement. Mutations in exon 58-65 were associated with minor symptoms in three systems. Based on the classification of domain, mutations in NH2 region were associated with minor skeletal symptoms. Mutations in hybrid motif were associated with major cardiovascular symptoms. Mutations in COOH region were associated with minor symptoms in three systems. Based on mutation type, nonsense mutations were associated with major cardiovascular symptoms. Part III: Clinical trial in patients with Marfan syndrome From May 2007 to Dec 2010, twenty-eight patients (M/F: 12/16, 16.5 ± 6.6 years) diagnosed as Marfan syndrome with recognized aortic root dilation (Z score >2.0) and under beta-blockade were enrolled and randomized into two groups. One group continued previous beta-blocker therapy. The other group received a combination therapy with beta-blocker and Losartan (an angiotensin II receptor blockade). After 35 months of treatment, the combination therapy with an angiotensin II receptor blockade and beta-blockade effectively slowed down the growth rate of aortic root as compared to beta-blockade alone (0.10 mm/yr versus 0.89 mm/yr, p = 0.026). The mean aortic diameter at sinus of Valsalva, annulus, and sinotubular junction also showed a reduced rate of change in diameter under combination therapy, but still keep growing under beta-blockade treatment (p < 0.05). However, the aortic distensibility and cross-sectional compliance changed insignificantly between these two groups (p = 0.462 and 0.535, respectively). The adverse effects from the combination therapy were not significantly different between the two groups. Conclusions Despite medical advances, aortic dissection still occurs in about one-tenth of the Marfan syndrome patients and carries a high mortality risk. Early diagnosis and timely medical interventions are warranted. We demonstrated the preliminary correlations between fibrillin-1 genotype and phenotype. Genetic testing for fibrillin-1 mutation will provide helpful information beyond disease diagnosis. It can help predict the severity of clinical manifestations as a reference for medical follow-up and exercise capacity to improve patients’ physical fitness. Besides, genetic testing can be applied in newborn or prenatal screening to confirm or rule out the diagnosis of Marfan syndrome if parents with history of Marfan syndrome. Even it may be applied in preimplantation genetic diagnosis (PGD). We also proved that Losartan add-on beta-blockade therapy is safe and provides more effective protection to slow the progression of aortic root dilatation than sole beta-blockade treatment in Marfan syndrome patients. With the advance in the pathogenesis of Marfan syndrome, many clinical trials are under way for promising new treatment. We believed that prophylactic monitoring and therapies will result in a nearly normal life expectancy and bring favorable functional outcomes.