賴美淑Lai, Mei-Shu臺灣大學:預防醫學研究所廖立人Liao, Li-JenLi-JenLiao2010-06-022018-06-292010-06-022018-06-292008U0001-2606200822223900http://ntur.lib.ntu.edu.tw//handle/246246/184793目的：研究p53 及 PTTG1 在口腔病變從正常口腔黏膜、癌前病變到口腔癌的表現及預後角色的相關性初步探討法：收集臺大醫院耳鼻喉科及口腔外科在1995年到 2001年的病人病理切片，共有141例，包括尚未接受治療之口腔癌前病變82例，及沒有遠隔轉移的口腔癌患者43例當做病患組，另選取2005年之正常的口腔黏膜病理切片16例，當做非病患組。採用石蠟包埋切片，做p53及PTTG1兩項生物標記的免疫組織染色，並定量此兩項生物標記在表皮細胞或腫瘤細胞染色表現的比例(Labeling indices, LI)。病人之資料收集採病歷回顧方式，記錄抽菸、喝酒、嚼檳榔等危險因子，及至2008年五月前所登錄，包括癌前病變病人轉變成口腔癌，或口腔癌病人術後的復發。死亡資料是以病人ID與國家死亡登錄資料庫碰檔取得。比較三組切片p53 與 PTTG1 LI之差異，並用線性回歸控制年齡、性別、生活型態等干擾因子。以原始之p53 與 PTTG1 LI值探討與癌前病變轉變為口腔癌，及口腔癌術後復發或存活之相關。在癌前病變轉變為口腔癌之p53 與 PTTG1之LI值，以ROC (Receiver operating characteristic) curve方法，選取最高的工具敏感度與特異性總和之最高值為切點，分成高LI與低LI組進行相關性分析。進一步以Cox 回歸模式及Kaplan-Meier curves計算兩組轉化為癌症之Hazard ratio，並做差異之檢定。本研究獲得台大醫院倫理委員會的同意。果：比較正常的口腔黏膜、口腔癌前病變及口腔癌之p53及PTTG1 LI值之差異; 發現兩種生物標記都會隨著口腔病變的嚴重程度而過度表現(over-expression)， p53 LI值之呈現在正常口腔黏膜為0，癌前病變為2.20±0.71，口腔癌的表現為23.40±5.43, 其差異有統計上之意義(p=0.0001); PTTG1 LI值之呈現在正常口腔黏膜、癌前病變及口腔癌分別是32.5±9.05, 68.20±3.20與85.46±2.68其差異同樣有統計上之意義(p=0.0001); 在控制可能的干擾因子後，仍有意義的過度表現之差異。82名癌前病變病人在3到71個月的追蹤期間，其中有20名有口腔癌轉變，Cox 回歸模式評估癌前病變病人的口腔癌轉變，發現PTTG1 LI的表現可能與口腔癌轉變有關，其Hazard ratio 為 1.03 (1.00~1.05); 以PTTG1 LI值80%，分成高值組與低值組兩組，評估癌前病變病人的口腔癌轉變，進行Cox 回歸模式多變項分析，高值組的Hazard Ratio為4.70(1.29-7.60), 有統計之差異( p=0.019)。口腔癌的復發及存活情形，與兩種生物標記之 LI值，並無明顯相關。論：p53 及PTTG1 LI值在癌化的過程會隨著口腔病變的嚴重而過度表現。 PTTG1是癌前病變病人口腔癌轉變的獨立預後因子，可能可以用來當做預測口腔癌前病變轉變為口腔癌的預後因子。Purpose:he aim of this study is to explore the expression of two molecular markers (p53 & PTTG1) in oral cancer tumorgenesis and to explore their association with the prognosis of patients with oral precancerous lesion and oral cancer.ethods:ne hundred and forty one cases of formalin-fixed, paraffin-embedded specimens were collected and classified as patients group, including primary oral precancerous lesions (n=82) and squamous cell carinomas without distant metastasis (n=43). These patients received operation at the Department of Otolaryngology or Oral and Maxillofacial Surgery in NTUH during 1995-2001. On the other hand, sixteen specimens of normal oral mucosa were obtained during extraction of impacted permanent mandibular tooth in 2005 and classified as non-patients group. The expression of two biomarkers, p53 and PTTG1 in epithelial cells and cancer cells was measured with immunohistochemical staining and scored as labeling indices (LI). Patients’ clinicopathological parameters were retrieved from hospital records, including habits of alcohol smoking, betel nut chewing and cigarette smoking and the record of malignant development in precancerous lesions, cancer recurrence till May 2008. Mortality data was obtained from national death registry database. The expression of p53 and PTTG1 LI in normal oral mucosa, precancerous lesions and cancer were compared and adjusted with age, gender and lifestyle in linear regression. We analyzed the relation of initial p53 and PTTG1 LI with cancer development in precancerous lesions and recurrence or survival in oral cancers. The significance of p53 and PTTG1 LI in precancerous lesion with subsequent cancer development was determined in receiver operating characteristic (ROC) curve method. The point at which (sensitivity + specificity ) is maximized on the curve is taken as the best cut-off point and classified into two groups. Cox regression and time to event analysis with Kaplan-Meier curves method were used to compare the hazard ratio and determine the significant difference of these two groups in cancer development in precancerous lesions. This study was approved by the IRB of the NTUH.esults:he difference of p53 and PTTG1 LI in normal oral mucosa, precancerous lesions and cancers were compared; the expression of these two biomarkers over-expressed with the progression of severity of oral lesions. The p53 LI was 0 in normal oral mucosa, 2.20±0.71 in precancerous lesions and 23.40±5.43 in oral cancer, the difference was significant(p=0.0001). The PTTG1 LI was also significantly different and was 32.50±9.05, 68.2±9.1 and 85.5±2.68 respectively for normal oral mucosa, precancerous lesions and cancer (p=0.0001). After adjusting possible confounding factors, the expression of p53 and PTTG1 still showed a significant increase with the progression of oral epithelial lesions. Twenty patients in 82 oral precancerous lesions had malignant development during follow-up period from 3 to 71 months. The PTTG1 LI was significantly associated with cancer development in precancerous lesions in Cox regression model (Hazard ratio=1.03, 1.00~1.05); PTTGI LI ≧80% had the maximum sensitivity and specificity in predicting malignant development in precancerous lesions. The precancerous lesions were classified as high PTTG1 LI (≧80%) and low PTTG1 LI. The high PTTG1 LI was significantly associated with malignant development in precancerous lesions in multivariate Cox regression analysis (Hazard ratio=4.70, 1.29~7.60, p=0.019). The cancer recurrence and mortality were not associated with these two biomarkers.onclusion:TTG1 and p53 LI increased from normal oral mucosa to oral precancerous lesions and further increased in oral cancers. PTTG1 LI was an independent prognostic factor in predicting cancer development when oral precancerous lesions occurred. PTTG1 may be a potential prognostic marker in oral precancerous lesions.中文摘要…………………………………………………………………………….i-iibstract………………………………………………………………………..........iii-vontents…………………………………………………………………………..vi-viiiigures...........................................................................................................................ixables.........................................................................................................................x-xi. Introduction: rationale of this study……………………………………………1. Literature Review: ………………………………………………………………2.1 Commonly used prognostic factors in oral precancerous lesion and oral cancer………………………………………………………………………2.2 Molecular alterations in oral cancer nature history………………………4.3 Expression and prognostic value of p53 and PTTG1 in various tumors……5.3.1 Expression of p53 in normal oral mucosa, oral precancerous lesions and oral squamous cell carcinoma…………………………………………………5.3.2 Prognostic value of p53 expression in oral precancerous lesions……………………9.3.3 Prognostic value of p53 expression in oral squamous cell carcinoma…12.3.4 Expression of PTTG1’s and prognostic values in various tumors……13. Aims of this study………………………………………………………………17. Materials and Methods:…………………………………………………………18.1 Specimen collection of oral lesions………… …………………………18.2 Tissue processing, measurement of p53 and PTTG1 expression…………19 .3 Obtain the clinical data and subsequent pathology report with rectrospective medical chart review………………………………………………………21.4 Cancer development and mortality data……………………………………22.5 Statistical analysis…………………………………………………………23. Results……………………………………………………………………………26.1 The expression of p53 and PTTG1 in normal oral mucosa, precancerous lesion and oral squamous cell carcinoma: descriptive analysis of cross-sectional data…… …………………………………………………………………………………26.2 Correlation between p53 and PTTG1 expression in the same specimen………28.3 Univariate analysis of risk factors and expression of p53 & PTTG1 in precancerous and cancer lesions………………………………………………28.4 Risk factors adjusted p53 and PTTG1 labeling indices analysis………………29.5 Follow-up study of patients with precancerous lesions…………………………30.5.1 Cox regression analysis of the risk factors in oral precancerous lesions with subsequent cancer development……………………………………………30.5.2 Receiver operating characteristic (ROC) curve method for best cut-off point selection………………………………………………………………………31.5.3 KM method in oral precancerous lesions with subsequently malignant development analysis……………………………………………………………32.5.4 Univariate analysis with COX regression of PTTG1 and p53 in oral precancerous lesion and subsequently malignant development ………………33.5.5 Multivariate COX regression in oral precancerous lesion cancer development………………………………………………………………33.6 Follow-up study of patients with cancerous lesions…………………………34.6.1 Cox regression analysis of the correlation between local recurrence and risk factors, molecular markers…………………………………………………34.6.2 Time to event analysis with K-M method and Log-Rank test in oral cancer local recurrence analysis………………………………………………………35.6.3 Cox regression analysis of the correlation between overall survival and risk factors, molecular markers …………………………………………………35.6.4 Time to event analysis with K-M method and Log-Rank test in overall survival analysis………………………………………………………………36. Discussion…………………………………………………………………………37. Conclusion………………………………………………………………………40. References…………………………………………………………………………41iguresigure 2.1 Molecular progression model in oral cancer carcinogenesis......................46igure 4.1 Example of measurement of p53 expression in different severity of oral lesions………………………………………………………………………………47igure 4.2 Example of measurement of PTTG1 expression in different severity of oral lesions…………………………………………………………………………………48igure 5.1 Scatter plot showing the expression of p53 (A) and PTTG1 (B) from normal oral mucosa, different degree of precancerous lesions and oral cancer………49igure 5.2 The expression of p53 (A) and PTTG1 (B) in different degrees of precancerous lesions with epithelial dysplasia………………………………………49igure 5.3 Correlation between p53 and PTTG1 labeling indices in 141 specimens..50igure 5.4 ROC for PTTG1 & p53 LI in predicting oral precancerous lesion with subsequent cancer development………………………………………………………50igure 5.5 Kaplan-Meier curve for cancer free proportions…………………………52igure 5.6 Kaplan-Meier curve for local regional recurrence………………………54igure 5.7 Kaplan-Meier curve for overall survival in cancer patients………………56ablesable 2.1 Pooled prevalence estimate for p53 overexpression among normal oral mucosa (NOM), oral precancerous lesion (OPL) and oral squamous cell carcinoma (OSCC)………………………………………………………………………………57able 2.2 Meta-analysis for the expression of p53 in nature history of oral cancer....57able 4.1 Demography and life style data of study cases……………………………58able 4.2 Clinicopathologic features of oral precancerous lesion……………………58able 4.3 Characteristics of cancer stage and pathologic features……………………59able 5.1 Mean p53 and PTTG1 expression labeling indices (LI) in normal, precancerous and malignant oral mucosa……………………………………………60able 5.2 Relationship of p53, PTTG1 expression and different severity in oral cancer patients………………………………………………………………………………60ables 5.3 Correlation between p53 and PTTG1 expression in the same specimens……………………………………………………………………………61able 5.4 Univariate analysis of risk factors and p53 & PTTG1 expression in oral precancerous lesion……………………………………………………………………61able 5.5 Univariate analysis of risk factors and p53 & PTTG1 expression in cancer patients……………………………………………………………………………62able 5.6 Estimated parameters in linear regression model for p53 and PTTG1 labeling indices………………………………………………………………………63able 5.7 Correlation of p53 and PTTG1 initial labeling indices and subsequent cancer development in patients with precancerous lesion……………………………64able 5.8 Cox regression analysis in comparison of risk factors and cancer development…………………………………………………………………………64able 5.9 Comparison of different cut-off point of PTTG1 LI in predicting cancer development………………………………………………………………………65able 5.10 Cox univariate analysis in oral precancerous lesion with subsequent malignant development, with category data…………………………………………65able 5.11 Age adjusted and multivariate-adjusted Cox regression in oral precancerous lesion cancer development……………………………………………65able 5.12 Factors related to local regional recurrence—analysis with univariable Cox regression with continuous variable (p53 LI and PTTG1 LI)…………………………66able 5.13 Factors related to overall survival in oral cancer patients—analysis with univariable Cox regression……………………………………………………………67application/pdf1224860 bytesapplication/pdfen-US口腔癌前病變口腔癌PTTG1p53預後因子oral precancerous lesionoral squamous cell carcinomaprognostic factor[SDGs]SDG3thesishttp://ntur.lib.ntu.edu.tw/bitstream/246246/184793/1/ntu-97-R95846013-1.pdf