陳凌雲Ling-Yun Chen易瑜嶠Yu-Chiao生化暨生物科技研究所2026-10-289999-12-312011https://handle.ncl.edu.tw/11296/335eqvhttps://scholars.csmu.edu.tw/handle/123456789/35670Part I血清中IGFII mRNA-binding protein 1 及 cyclin B1的自體抗體可作為卵巢子宮內膜異位瘤的生物標記子宮內膜異位是一常見的婦科疾病，並已成為育齡婦女最常見的住院原因之一。內膜異位的確定診斷必須依靠腹腔鏡，然而因腹腔鏡具有侵襲性以及伴隨著可能的併發症使得它不適合用於初步的篩檢以及監測復發及治療成效。因此尋找一個更有效的生物標記實有其必要性。實驗顯示癌症患者的血清中含有能與腫瘤相關抗原（tumor-associated antigens, TAAs）反應的自體抗體，而這些抗體也因此可作為偵測腫瘤生成中不正常細胞機轉的指標及偵測腫瘤存在的生物標記。雖然子宮內膜異位雖然是一種良性疾病，但卻具有一些與惡性腫瘤相似的特徵，如細胞不正常的過度增殖，侵襲，轉移，血管新生及基因不穩定，而目前的研究也顯示一些常見於惡性腫瘤組織的分子異常也見於子宮內膜異位組織。此外，一些研究也顯示子宮內膜異位症患者血清中有自體抗體的存在。依據這些發現我們推測推子宮內膜異位症患者血清中可能也有與特定腫瘤相關抗原（tumor-associated antigens, TAAs）反應的自體抗體並可作為子宮內膜異位症的標記。因此本實驗目的在評估TAAs自體抗體是否可作為子宮內膜異位瘤診斷的標記。在本實驗中我們收集56位子宮內膜異位瘤患者及66位健康女性作為控制組。利用ELISA（enzyme-linked immunosorbent assay）分析血中八種腫瘤相關抗原（tumor-associated antigens, TAAs）自體抗體的濃度，並分析兩組中自體抗體濃度的差異。結果我們發現血中IGFII mRNA-binding protein 1 (IMP1) 及cyclin B1 自體抗體可用以區分健康女性與子宮內膜異位瘤患者(AUC–ROC 0.777;95% confidence interval [CI] 0.694–0.860,P＜0.0005, 及 AUC–ROC 0.614; 95% confidence interval [CI] 0.513–0.714, P=0.031, respectively)。若以 0.073 及0.007 作為IMP1及Cyclin B1自體抗體異常的判定值，IMP1的靈敏度及專一性分別為85.7%及63.6%。若合併使用cyclin B1及IMP1，專一性可上升到 72.7%而靈敏度則略降至83.9%。此研究結果顯示單獨使用或合併Cyclin B1可以作為臨床上輔助診斷子宮內膜異位瘤的生物標記。Part II.第一型組織金屬蛋白脢抑制劑（Tissue inhibitor of metalloproteinase-1）基因多型性與子宮內膜癌發生風險有關子宮內膜癌是女性生殖道最常見的惡性腫瘤之一，由於好發因子中有不少與雌激素相關，因此推測雌激素在子宮內膜癌的發生應該扮演重要的角色。然而由於腫瘤生成是一個多步驟的過程，除了雌激素外也必需有其他分子變異的配合，才能形成子宮內膜癌。在這些影響腫瘤生成及擴展的過程中，降解（degrade）或破壞圍繞腫瘤細胞的細胞外基質（extracellular matrix）及結締組織，以進行往下的侵犯作用（invasion）及後續的遠端轉移是相當的重要，而決定細胞外基質（extracellular matrix）及結締組織分解的主要是組織金屬蛋白脢（matrix metalloproteinases，MMPs）及組織金屬蛋白脢抑制劑（Tissue inhibitors of metalloproteinases ,TIMPs）。組織金屬蛋白脢抑制劑家族是一群多功能的蛋白並在腫瘤形成的過程中扮演廣泛的角色，以往的研究發現TIMPs的mRNA在一些惡性腫瘤有向上調控且與臨床上的預後有關。由於不同的基因多型性（polymorphisms）可能影響到TIMPs的表現量並與癌症的發生有關，因此在本文中我們嘗試分析TIMP-1及TIMP-2基因多型性與子宮內膜癌發生風險的關聯性。在此病例對照研究中我們收集了118位組織學確診的子宮內膜癌患者及229位健康女性作為對照組，並以PCR-RFLP的方法分析TIMP-1 (_372C>T) 及 TIMP-2(_418G>C and _303C>T) 的基因多型性。結果我們發現TIMP-1_372基因多型性中C/C基因型及C 對偶基因出現的頻率在子宮內膜癌患者及對照組間有顯著差異，C 對偶基因及TIMP-1_372 C/C 基因型分別較T 對偶基因及TIMP-1_372 T/T 基因型增加子宮內膜癌的風險達1.68倍 (95% 信賴區間 1.22–2.31)及2.37倍(95%信賴區間1.33, 4.22)。分層分析顯示，在子宮內膜狀癌及低期別（第一期及第二期）的子宮內膜癌次群體中TIMP-1_372 C/C基因型分別較對照組增加子宮內膜癌風險2.66（勝算比=2.66; 95%信賴區間1.37–5.17），2.46 (勝算比=2.46; 95%信賴區間1.34–4.53) 及3.24倍(勝算比=3.24; 95%信賴區間1.22–4.13)。至於TIMP-2_418G>C 及 TIMP-2_303C>T 基因多型性則在子宮內膜癌患者及對照組間則無顯著差異。此研究顯示TIMP-1_372T>C 基因多型性是子宮內膜癌的好發因子，TIMP-2_418G>C 及 TIMP-2-303C>T 基因多型性則不是預測子宮內膜癌好發因子的適切指標，而TIMP-1_372T>C 基因多型性作為子宮內膜癌的好發因子也顯示TIMP_1在子宮內膜癌的發生上扮演重要的角色。Part ISerum Autoantibodies to IGFII mRNA-binding Protein 1 and Cyclin B1 can Serve as Biomarkers of Ovarian EndometriomaEndometriosis is a common gynecologic disorder and represents one of the most common admitting diagnoses in women of reproductive age. The ‘‘gold’’ standard for the diagnosis of endometriosis is laparoscopy. However,The invasiveness and associated morbidity of the laparoscopic procedure precludes its use for initial screening or monitoring recurrences and response to therapy, so more effective biomarkers for endometriosis are needed.Many studies demonstrated that sera from cancer patients contain autoantibodies which react with tumor-associated antigens (TAAs) and these autoantibodies can be used as immunodiagnostic markers for cancer detection. Although endometriosis is considered a benign disorder, it exhibits some characteristics similar to malignant lesions, and current studies indicate some molecular aberrations commonly observed in cancer tissue are also present in endometriotic lesions. This finding let us to hypothesize that the sera of endometriosis patients contain antibodies which react with a unique group of TAAs, thus providing a new marker for endometriosis. Thus, in this study, we obtained serum samples obtained from 56 patients with endometriosis and 66 healthy women who served as normal controls. The titers of antibodies against a panel of eight TAAs were analyzed using enzyme-linked immunosorbent assay，the difference of autoantibodies titer in the serum of patients with endometrioma and healthy controls was analyzed to determine whether autoantibodies can be accurate biomarkers for the diagnosis of ovarian endometrioma。We found that the serum IGFII mRNA-binding protein 1 (IMP1) autoantibody and cyclin B1 autoantibody could discriminate between healthy controls and endometriosis patients (AUC–ROC 0.777; 95% confidence interval [CI] 0.694–0.860, P＜0.0005, and AUC–ROC 0.614; 95% confidence interval [CI] 0.513–0.714, P=0.031, respectively). Using 0.073 and 0.007 as the cutoff values for IMP1 and Cyclin B1 autoantibody, respectively, the sensitivity and specificity of IMP1 were 85.7 and 63.6%, respectively. When cyclin B1 was combined with IMP1, the specificity increased to 72.7% and the sensitivity slightly decreased to 83.9%。Our data suggest that IMP1 alone or combined with cyclin B1 seems to fulfill the requirements of sensitivity and specificity to become a useful clinical biomarker of endometrioma。Part II.Genetic polymorphism of the tissue inhibitor of metalloproteinase-1 is associated with an increased risk of endometrial cancerEndometrial carcinoma is a common gynecological malignancy of the female urogenital tract, and its incidence is increasing significantly。The metabolic conversion of estrogens to 4-hydroxy estrogens has been postulated to be a major factor in carcinogenesis，however, transformation and spread of malignant tumors is a multi-step process and many of these steps require degradation or breakdown of the extracellular matrix and connective tissue surrounding tumor cells. The major proteins involve in degradation of extracellular matrix are matrix metalloproteinases (MMPs) and Tissue inhibitors of metalloproteinases (TIMPs). Tissue inhibitors of metalloproteinases (TIMPs) are a family of multifunctional proteins known to possess a broad range of biological activities involved in tumorgenesis and mRNA expression of TIMP family members has been shown to be upregulated in numerous cancers and correlates with clinical outcomes。Growing evidence indicates that genetic polymorphisms result in differential expression of TIMPs and these polymorphisms have been associated with different cancer risks，so we evaluated the association of different TIMPs polymorphisms and endometrial cancer risk。In the present case–control study, we enrolled a total of 118 endometrial cancer patients confirmed by histopathology and 229 unrelated healthy individuals. Polymorphism for TIMP-1 (_372C>T) and TIMP-2 (_418G>C and _303C>T) genes was genotyped by PCR-RFLP。We found the Frequency of TIMP-1_372C/C genotype and 372-C allele differed significantly between patients with endometrial cancer (38.1% and 56.4% respectively) and healthy individuals (22.7% and 44.1% respectively).Individuals with TIMP-1_372 C/C genotype were at higher risk of endometrial cancer (OR=2.37; 95%CI: 1.33–4.22). Stratification analysis showed that individuals with TIMP-1_372 C/C genotype were at increased risk forendometrioid (OR=2.46; 95% CI 1.34–4.53) and low stage (stages I–II) endometrial cancer (OR=3.24; 95% CI 1.22–4.13). However, no significant differences in TIMP-2_418G>C and TIMP-2_303C>T genotypes wereobserved between endometrial carcinoma cases and controls. In conclusion we found Individuals with TIMP-1_372C/C genotype were at significantly higher risk of endometrial cancer and as a predisposing factor，TIMP-1 may play an important role in the pathogenesis of endometrial cancer。thesis