【作者】 史恩溢；

【导师】 朱雄增； 侯英勇； 周晓燕；

【作者基本信息】 复旦大学 ， 肿瘤学， 2008， 博士

【摘要】 目的:探讨Fletcher和Miettinen两种分级标准对局限性胃肠道间质瘤(GIST)危险程度的评估效能;从组织病理形态、基因突变分型和免疫表型三个不同的层次研究与局限性GIST临床预后相关的参考指标,为补充和修订现行分级标准提供理论和实践依据。方法:(1)收集220例有完整临床病理及随访资料的局限性GIST,应用Fletcher和Miettinen两种分级标准对肿瘤的危险程度进行评估,生存分析比较两种标准在预测肿瘤恶性潜能中的意义;(2)在220例局限性GIST中观察与恶性生物学行为可能相关的组织病理形态学指标,生存分析这些指标在评估肿瘤危险程度中的意义;(3)应用PCR-直接测序的方法对168例局限性GIST进行c-kit和PDGFRA基因突变类型和方式的检测,生存分析突变的类型和方式在评估肿瘤危险程度中的意义;(4)收集160例局限性GIST肿瘤标本存档蜡块制作组织芯片,应用免疫组织化学方法检测Ki-67、p16、p53和CD44的表达状况,生存分析这些免疫标记物在评估肿瘤危险程度中的意义;(5)将组织病理形态、基因突变分型和免疫表型三个层次中经单因素生存分析显示与肿瘤临床预后相关的参考指标,纳入cox比例风险模型,对154例局限性GIST进行多因素生存分析,筛选影响肿瘤预后的独立因子;(6)将组织病理形态、基因突变分型和免疫表型三个层次中经单因素生存分析显示与肿瘤临床预后相关的参考指标,纳入cox比例风险模型,对具有c-kit基因第11号外显子突变表型的103例局限性GIST进行多因素生存分析,筛选影响该患者群预后的独立因子。结果:(1)220例局限性GIST按照Fletcher分级标准进行评估,高危组的总生存率和无病生存率显著低于极低、低和中危三组,但极低、低和中危三组之间总生存率和无病生存率差异无统计学意义。在高危组中,小肠和直肠GIST的总生存率和无病生存率显著低于胃GIST;而在中危组中,小肠GIST的无病生存率显著低于胃GIST。220例局限性GIST按照Miettinen分级标准进行评估,高危组的总生存率和无病生存率显著低于极低、低和中危三组,但极低、低和中危三组之间总生存率和无病生存率差异无统计学意义。在极低、低、中和高危各个危险组中,胃、小肠和直肠GIST之间总生存率及无病生存率差异均无统计学意义。(2)出现肿瘤细胞密集、肿瘤细胞明显异型、肿瘤性坏死、浸润性生长方式、古钱币样排列等组织病理形态的GIST组其总生存率和无病生存率显著低于相应的未出现以上组织病理形态的GIST组。(3)c-kit基因第9号外显子突变的GIST组其总生存率和无病生存率显著低于c-kit基因第11号外显子突变组和野生型组;c-kit基因第11号外显子发生缺失突变的GIST组其总生存率和无病生存率显著低于第11号外显子发生错义点突变和重复插入突变的GIST组;c-kit基因第11号外显子发生557-558位密码子缺失突变的GIST组其总生存率和无病生存率显著低于第11号外显子未发生557-558位密码子缺失突变的GIST组。(4)Ki-67阳性表达率＞5%的GIST组其总生存率和无病生存率显著低于Ki-67阳性表达率≤5%的GIST组;p16失表达的GIST组其总生存率和无病生存率显著低于p16表达的GIST组;p53低表达组和高表达组以及CD44表达评分≤3分组和＞3分组之间总生存率与无病生存率差异均无统计学意义。(5)多因素生存分析显示,Miettinen分级、肿瘤古钱币样排列和p16失表达是局限性GIST独立的预后影响因子;Miettinen分级、肿瘤古钱币样排列、p16失表达和c-kit基因第11号外显子557-558位密码子缺失是具有c-kit基因第11号外显子突变表型的局限性GIST独立的预后影响因子。结论:1.Fletcher分级标准简明易用,但Miettinen分级标准按不同发病部位评估危险程度能更准确地认识局限性GIST的生物学行为;2.肿瘤细胞密集度、肿瘤细胞异型性、肿瘤性坏死、浸润性生长方式、古钱币样排列等组织病理形态学指标与局限性GIST的临床预后相关;3.c-kit基因突变的类型及其方式与局限性GIST的临床预后相关;4.Ki-67标记指数和p16失表达与局限性GIST的临床预后相关;5.肿瘤古钱币样排列、p16免疫表型以及c-kit基因突变的类型和方式可作为补充和修订Miettinen分级标准的候选参考指标,对正确选择高危局限性GIST患者术后进行分子靶向辅助治疗具有重要意义。更多还原

【Abstract】 Objective: To evaluate the implication of Fletcher and Miettinen biological potential grading criteria in native localized GISTs. Meanwhile, from histologic features, mutational types and immunophenotypes, to study the risk assessment parameters and provide new clue and theory for complement of the biological potential grading criteria.Methods: (1) Two hundred and twenty localized GISTs with complete clinicopathologic and follow-up date were evaluated their biologic potential by Fletcher and Miettinen grading criteria. The implications of the two grading criteria were compared by survival analysis. (2) The correlation between histologic features and clinical prognosis are studied by survival analysis in two hundred and twenty localized GISTs. (3) One hundred and sixty-eight localized GISTs were evaluated for the presence of c-kit and PDGFRA mutations by PCR amplication and direct sequencing. The correlation between mutational types and clinical prognosis are studied by survival analysis. (4) One hundred and sixty formalin-fixed paraffin-embedded localized GISTs were selected and Tissue microarry (TMA) block and section slides were prepared. Sections were immunohistochemically stained with antibodies against Ki-67, p16, p53 and CD44. The correlation between the protein expression and clinical prognosis are studied by survival analysis. (5) Cox’s multivariate regression analysis were performed to evaluate the effect of all significant markers about histology, mutational types and immunophenotypes on prognosis in one hundred and fifty-four localized GISTs. (6) Cox’s multivariate regression analysis were performed to evaluate the effect of all significant markers about histology, mutational types and immunophenotypes on prognosis in one hundred and three localized GISTs with c-kit exon 11 mutations.Results: (1) Evaluated by Fletcher grading criteria, the overall and disease-free survival rate of high risk GISTs were lower than that of very-low, low and intermediate GIST; while the overall and disease-free survival rate of very-low, low and intermediate risk GISTs had no statistical difference. In the high risk GISTs, the overall and disease-free survival rate of small intestinal and rectal GISTs was lower than that of gastric GISTs; while in the intermediate risk GISTs, the disease-free survival rate of small intestinal GISTs was lower than that of gastric GISTs. Evaluated by Miettinen grading criteria, the overall and disease-free survival rate of high risk GISTs were lower than that of very-low, low and intermediate GIST; while the overall and disease-free survival rate of very-low, low and intermediate risk GISTs had no statistical difference. In the risk subgroup of GISTs, the overall and disease-free survival rate of gastric, small intestinal and rectal GISTs had no statistical difference. (2) Univariate analysis demonstrated the following as poor prognostic histologic factors for overall and disease-free survival: high cellularity, nuclear pleomorphism, tumor necrosis, tumor invasion and perivascular collar-like pattern. (3) the overall and disease-free survival rate of the GISTs with c-kit exon 9 mutations was lower than that the GISTs with c-kit exon 11 mutations and wide-type GISTs. the overall and disease-free survival rate of the GISTs with deletions in c-kit exon 11 was lower than that the GISTs with duplications or missence in c-kit exon 11 mutations, the overall and disease-free survival rate of the GISTs with deletions affecting codons 557 to 558 in c-kit exon 11 was lower than that the GISTs without deletions affecting codons 557 to 558 in c-kit exon 11. (4) Univariate analysis demonstrated the following as poor prognostic immunophenotypes for overall and disease-free survival in the protein expression of Ki-67, p16, p53 and CD44: Ki-67 labeling index>5% and pl6 loss. (5) Multivariate analysis indicated that Miettinen high risk index, perivascular collar-like histologic pattern and p16 loss were the independent poor prognostic factor of localized GISTs. Multivariate analysis indicated that Miettinen high risk index, perivascular collar-like histologic pattern, pl6 loss and deletions affecting codons 557 to 558 in c-kit exon 11 were the independent poor prognostic factor of localized GISTs with c-kit exon 11 mutations.Conclusions: (1) Fletcher grading criteria is simple and easy to use, while Miettinen grading criteria for evaluating biological potential by anatomic site is more critical. (2) the histologic features of high cellularity, nuclear pleomorphism, tumor necrosis, tumor invasion and perivascular collar-like pattern are poor prognostic factors of localized GISTs. (3) The mutational types of c-kit gene in localized GISTs have effect on patient’s prognosis. (4) Ki-67 labeling index>5% and pl6 loss are poor prognostic factors of localized GISTs. (5) perivascular collar-like histologic pattern, pl6 loss and mutational types of c-kit gene may be the risk parameters for the completement of Miettinen grading criteria and have important reference implication for the selection of high risk patients for targeted adjuvant treatment.更多还原