结节性硬化症相关肾脏血管平滑肌脂肪瘤基因突变及mTOR通路研究

发布时间：2018-04-30 19:07

本文选题：结节性硬化症 + 血管平滑肌脂肪瘤　；参考：《北京协和医学院》2015年博士论文

【摘要】：第一部分 TSC-RAML患者TSC1/TSC2突变位点检测与分目的：明确TSC1/TSC2致病性突变在TSC-RAML患者中的分布,探索基因型与临床表型之间的关系。方法：本组研究对象共24例,其中,TSC确诊患者22例,TSC疑似患者2例；22例TSC确诊患者中TSC-RAML患者20例。经知情同意,取外周静脉血3m1,通过微阵列芯片捕获,在Illumina HiSeq2500平台上进行高通量测序,并将检测结果与LOVD数据库比对。结果：22例TSC确诊患者中,TSC1突变4例,TSC2突变15例,未检测到突变3例。20例TSC-RAML患者中,TSC突变率85%(17/20), TSC1:TSC2约为1：4.7(3vs 14),TSC1/TSC2突变位点的分布均较为弥散,突变类型以无义突变(41.2%)及框移突变(35.3%)为主。TSC1及TSC2突变患者中,最小年龄分别为14岁及8岁,RAML的最大直径分别为10.3cm及20cm, RAML最大直径≥10cm的例数分别为1例及8例。通过与LOVD数据库比对,检测出新发突变位点8例,TSCl突变1例,TSC2突变7例。3例家系中,先证者RAML最大直径分别为10、10.3及16cm,而家系成员RAML最大直径分别为0、0及1cm。结论：TSC-RAML患者基因突变以TSC2为主,突变位点弥散,以无义突变及框移突变为主；TSC2突变患者RAML发病年龄相对更早,病情相对更重；TSC-RAML临床表现个体化差异较大,但与突变位点及突变类型可能无关。第二部分 TSC-RAML组织中nTOR信号通路激活水平研究目的：研究mTOR信号通路在TSC-RAML组织中的激活水平。方法：研究组5例TSC-RAM IL标本,对照组10例S-RAML标本及10例非肾脏肿瘤患者的正常肾组织标本,以免疫组织化学技术分别检测p-mTOR、p-S6K1及p--4EBP1在上述3组标本中的表达。结果：p-mTOR在TSC-RAML中呈阳性或强阳性表达,在S-RAML中呈弱阳性表达；p-S6K1在TSC-RAM L中呈阳性表达,在S-RAML中呈弱阳性或阴性表达；p-4EBP1在TSC-RAML中呈阳性或弱阳性表达,在S-RAML中呈弱阳性表达；p-mTOR、p-S6K1及p-4EBP1在正常肾脏组织中均呈阴性表达。5例TSC-RAML标本中,p-mTOR及p-4EBP1在发生TSC2突变的3例患者标本中的表达水平相对高于2例发生TSC1突变患者的标本,而p-S6K1在TSC1突变及TSC2突变患者标本的表达无明显差别。结论：与S-RAML及正常肾脏组织相比,mTOR信号通路在TSC-RAML组织中的激活程度相对较高；与TSC1突变相比,mTOR信号通路在TSC2突变患者的RAML组织中激活程度相对较高。第三部分 mTOR抑制剂治疗TSC-RAML的疗效及安全性初步研究目的：评估mTOR抑制剂everolimus治疗TSC-RAML的疗效及安全性。方法：单中心、开放、非随机临床研究,研究对象为临床确诊的TSC-RAML患者,按照纳入标准及排除标准严格筛选后,符合标准的患者给予everolimus 10mg/d口服,治疗时间为一年,常规随访时间为开始服药后第3、6、12及24个月,治疗期间可根据患者耐受情况调整药物用量。详细评估和记录患者服药前基线数据及服药后病情变化,以及治疗相关不良事件。结果：截止目前,已有5例TSC-RAML患者入组并口服everolimus治疗3个月。与基线数据相比,每例患者的RAML均有明显减小,RAML最大直径平均缩小3.8(1.5-6)cm,最大直径平均减小百分比28.2%(15.4%-38%)。5例患者的面部血管纤维瘤也明显变淡变平。2例患者精神症状较前明显好转。口腔炎、咳嗽、上呼吸道感染、皮疹为最常见不良事件,其中仅1例3级不良事件,其余均为1～2级。结论：mTOR抑制剂everolimus在TSC-RAML治疗中疗效显著,TSC-RAML直径减小明显,不良事件以1-2级多见,安全性及患者耐受性良好。
[Abstract]:The first part of TSC-RAML patients TSC1/TSC2 mutation site detection and purpose: to clarify the distribution of TSC1/TSC2 pathogenic mutation in TSC-RAML patients and explore the relationship between genotype and clinical phenotype. Methods: this group of 24 cases, including 22 cases of TSC confirmed patients, 2 cases of suspected TSC patients, 22 cases of TSC confirmed patients 2 TSC-RAML patients 2 0 cases, after informed consent, take the peripheral venous blood 3M1, through microarray capture, high flux sequencing on the Illumina HiSeq2500 platform, and compare the results with the LOVD database. Results: in 22 cases of TSC confirmed patients, 4 cases of TSC1 mutation, 15 cases of TSC2 mutation, and 3 cases of.20 case TSC-RAML, TSC mutation rate 85% (17/20), T. T SC1:TSC2 is about 1:4.7 (3vs 14), and the distribution of TSC1/TSC2 mutation sites is more diffuse. The mutation type is mainly.TSC1 and TSC2 mutations in patients with.TSC1 and TSC2 mutations, with the minimum age of 14 and 8 years, respectively. The maximum diameter of RAML is 10.3cm and 20cm respectively, and the number of RAML maximum diameter > 10cm is 1 cases and 8 cases respectively. Compared with LOVD database, 8 new mutation sites, 1 cases of TSCl mutation, and 7 TSC2 mutations in.3 families were detected. The maximum RAML diameter of RAML was 10,10.3 and 16cm respectively, while the largest RAML diameter of family members was 0,0 and 1cm. conclusion: TSC-RAML patient gene mutation was mainly TSC2, the mutation site dispersed, with nonsense mutation and frame shift mutation. The age of RAML in TSC2 mutation is relatively early and the condition is relatively heavier; the individual difference of TSC-RAML clinical manifestation is larger, but it may not be related to the mutation site and mutation type. The purpose of the study on the activation level of nTOR signaling pathway in the second part of TSC-RAML tissue is to study the activation level of mTOR signaling pathway in the TSC-RAML tissue. Method: 5 specimens of TSC-RAM IL in the study group, 10 cases of S-RAML specimens in the control group and 10 normal renal tissue specimens of non renal tumor patients, the expression of p-mTOR, p-S6K1 and p--4EBP1 in the above 3 specimens were detected by immunohistochemistry. Results: p-mTOR was positive or strongly positive in TSC-RAML, and was weakly positive in S-RAML. P-S6K1 was positive in TSC-RAM L and expressed weakly positive or negative in S-RAML; p-4EBP1 was positive or weakly positive in TSC-RAML, and expressed weakly positive in S-RAML; p-mTOR, p-S6K1 and p-4EBP1 were negative expression in.5 TSC-RAML specimens in normal kidney tissues, and 3 cases suffered from mutation The expression level in the specimens was higher than that of the 2 cases with TSC1 mutation, while there was no significant difference in the expression of p-S6K1 in TSC1 and TSC2 mutations. Conclusion: compared with S-RAML and normal renal tissue, the activation degree of mTOR signaling pathway in TSC-RAML tissues is relatively higher; mTOR signaling pathway is in T compared with TSC1 mutation. The degree of activation in RAML tissues of patients with SC2 mutation is relatively high. Third the efficacy and safety of mTOR inhibitors for the treatment of TSC-RAML. Objective: To evaluate the efficacy and safety of mTOR inhibitor everolimus in the treatment of TSC-RAML. Methods: single center, open, non randomized clinical study, and clinical diagnosis of TSC-RAML patients. After the inclusion criteria and exclusion criteria were strictly screened, the patients in accordance with the standard were given everolimus 10mg/d oral administration for one year. The routine follow-up time was 3,6,12 and 24 months after the beginning of the medicine. The dosage of the drug could be adjusted according to the patient's tolerance. Results: up to now, 5 patients with TSC-RAML were enrolled and treated with everolimus for 3 months. Compared with the baseline data, the RAML of each case decreased significantly, the maximum diameter of RAML decreased by 3.8 (1.5-6) cm, the maximum diameter decreased by 28.2% (15.4%-38%).5 patient's facial blood. The mental symptoms of.2 patients were obviously better than before. Stomatitis, cough, upper respiratory tract infection, and rash were the most common adverse events, of which only 1 cases were grade 3 and the rest were 1~2. Conclusion: mTOR inhibitor everolimus has a significant effect in the treatment of TSC-RAML, the diameter of TSC-RAML decreases obviously, and the adverse event is obvious. It is more common in class 1-2, safety and patient tolerance.

【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R737.11

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