FGFR3和PIK3CA基因在膀胱腫瘤中的作用和臨床生物學(xué)價值的研究
發(fā)布時間：2018-09-10 18:25
【摘要】：第一章膀胱癌中FGFR3和PIK3CA基因突變與預(yù)后關(guān)系的研究目的:檢測膀胱癌組織、配對癌旁組織及良性腫瘤組織中FGFR3基因和PIK3CA基因突變,同時分析FGFR3基因和PIK3CA基因突變與臨床病理參數(shù)之間的關(guān)系及相關(guān)性,為膀胱腫瘤的病因?qū)W研究提供一定的理論依據(jù),并探尋FGFR3基因和PIK3CA基因突變是否可作為膀胱腫瘤的有效治療靶點(diǎn)及預(yù)測腫瘤復(fù)發(fā)、腫瘤預(yù)后的標(biāo)志物。方法:分別從34例膀胱癌組織、7例配對癌旁組織、6例良性腫瘤組織中提取DNA,以FGFR3基因外顯子7、PIK3CA基因外顯子9、20設(shè)計引物,經(jīng)PCR擴(kuò)增,擴(kuò)增產(chǎn)物經(jīng)電泳確認(rèn)目的產(chǎn)物。應(yīng)用Sanger基因檢測方法檢測PCR擴(kuò)增產(chǎn)物中FGFR3和PIK3CA基因突變,探討膀胱癌組織中FGFR3和PIK3CA基因突變與臨床病理參數(shù)之間的關(guān)系,FGFR3和PIK3CA基因突變與膀胱癌的預(yù)后關(guān)系采用Kaplan-Meier生存曲線分析,單因素和多因素COX模型分析影響膀胱癌預(yù)后的各個因素。結(jié)果:FGFR3基因在膀胱癌中的突變是17.65%(6/34),癌旁組織是28.57%(2/7),良性腫瘤未見突變,三組FGFR3基因突變兩兩比較,差異無統(tǒng)計學(xué)意義(p0.05)。PIK3CA基因在膀胱癌中的突變是5.88%(2/34),癌旁組織中未見突變,良性腫瘤未見突變,膀胱癌組織分別與癌旁組織和良性腫瘤比較,差異無統(tǒng)計學(xué)意義(p0.05)。FGFR3基因突變在膀胱癌中,6例都是7號外顯子S249C位點(diǎn)突變,錯義突變;癌旁組織中,2例都是7號外顯子S249C位點(diǎn)突變,錯義突變,其中1例癌組織和配對癌旁組織都發(fā)生了7號外顯子S249C位點(diǎn)突變。PIK3CA基因突變在膀胱癌中,2例均為9號外顯子E545K位點(diǎn),錯義突變,未見外顯子20突變。FGFR3和PIK3CA基因突變與臨床病理分級分期都沒有相關(guān)性。34例膀胱癌中,術(shù)后復(fù)發(fā)5例,其中6例FGFR3基因突變的膀胱癌,術(shù)后復(fù)發(fā)3例,皮爾生相關(guān)系數(shù)(r)是0.46(p=0.01),FGFR3基因突變與腫瘤復(fù)發(fā)呈正相關(guān)。在FGFR3和PIK3CA基因突變對膀胱癌生存影響的研究中,通過Kaplan-Meier生存曲線進(jìn)行分析,FGFR3基因突變型和野生型生存期比較,差異無統(tǒng)計學(xué)意義(p0.05),而PIK3CA基因突變型和野生型生存期比較,差異有顯著的統(tǒng)計學(xué)意義(p0.05)。影響膀胱癌預(yù)后因素的單因素COX分析中,年齡是影響膀胱癌預(yù)后的重要因素(p0.05),而影響膀胱癌預(yù)后的因素進(jìn)行多因素COX分析,年齡也是影響膀胱癌預(yù)后的獨(dú)立因素(p0.05),與性別、腫瘤大小、數(shù)目、分期、分級以及FGFR3和PIK3CA蛋白表達(dá)都沒有相關(guān)性。結(jié)論:1.在膀胱腫瘤中PIK3CA/E545K是較常見基因突變類型,且提示是一個不利的預(yù)后因素。2.在膀胱腫瘤和癌旁組織中均存在FGFR3/S249C基因突變類型,故其突變不能作為診斷膀胱癌的指標(biāo),但可以預(yù)測腫瘤復(fù)發(fā)。第二章膀胱癌中FGFR3、PIK3CA蛋白表達(dá)的臨床生物學(xué)意義目的:探討膀胱癌、配對癌旁組織和良性腫瘤中FGFR3和PIK3CA蛋白表達(dá)情況,同時分析FGFR3和PIK3CA的表達(dá)與臨床病理參數(shù)之間的關(guān)系及其相互之間的關(guān)聯(lián)性,揭示FGFR3和PIK3CA蛋白在膀胱癌中表達(dá)的臨床意義,為進(jìn)一步探討膀胱腫瘤發(fā)生發(fā)展的分子機(jī)制奠定基礎(chǔ)。方法:應(yīng)用免疫組織化學(xué)的檢測42例膀胱癌組織、7例配對癌旁組織、5例良性膀胱腫瘤組織中FGFR3和PIK3CA蛋白表達(dá)情況,并計算蛋白表達(dá)強(qiáng)度。探討膀胱腫瘤組織中FGFR3和PIK3CA蛋白表達(dá)之間的關(guān)系及兩者與各臨床病理學(xué)參數(shù)之間的關(guān)系,FGFR3和PIK3CA蛋白表達(dá)與膀胱癌的預(yù)后關(guān)系采用Kaplan-Meier生存分析,單因素和多因素COX模型分析影響膀胱癌預(yù)后的因素。結(jié)果:FGFR3蛋白在膀胱癌中的表達(dá)是50.00%(21/42),癌旁組織是14.29%(1/7),良性腫瘤是20.00%(1/5),癌組織分別與癌旁組織和良性腫瘤組織比較FGFR3蛋白表達(dá),差異無統(tǒng)計學(xué)意義(p0.05)。PIK3CA蛋白在膀胱癌中的表達(dá)是64.28%(27/42),癌旁組織是14.29%(1/7),而良性腫瘤是20%(1/5),癌組織與癌旁組織比較差異有統(tǒng)計學(xué)意義(p0.05),而癌組織與良性腫瘤組織比較,差異無統(tǒng)計學(xué)意義(p0.05)。FGFR3蛋白表達(dá)在膀胱癌不同分期中,Ta-T1期是55.56%(10/18),T2-T4是45.83%(11/24),雖Ta-T1期蛋白表達(dá)要高于T2-T4期,但差異沒有統(tǒng)計學(xué)意義(P0.05)。FGFR3蛋白表達(dá)在膀胱癌不同病理分級中,低級別是53.85%(7/13),高級別是48.28%(14/29),低級別膀胱癌FGFR3蛋白表達(dá)高于高級別,但差異沒有統(tǒng)計學(xué)意義(P0.05)。PIK3CA蛋白表達(dá)在膀胱癌不同分期中,Ta-T1期是83.33%(15/18),T2-T4期是50.00%(12/24),Ta-T1期蛋白表達(dá)明顯高于T2-T4期,并且差異有顯著的統(tǒng)計學(xué)意義(p0.05)。PIK3CA蛋白表達(dá)在膀胱癌不同病理分級中,低級別是76.92%(10/13),高級別為58.62%(17/29),低級別膀胱癌PIK3CA蛋白表達(dá)高于高級別,差異沒有統(tǒng)計學(xué)意義(p0.05)。膀胱癌中FGFR3蛋白表達(dá)強(qiáng)度以2分為主,占57.14%(12/21);PIK3CA蛋白表達(dá)強(qiáng)度以3分為主,占50.00%(14/28)。FGFR3蛋白表達(dá)與PIK3CA蛋白表達(dá)呈正的中等線性相關(guān)(r=0.845,p=0.001),隨著FGFR3蛋白表達(dá)和強(qiáng)度增加,PIK3CA表達(dá)和強(qiáng)度也隨之增加。FGFR3和PIK3CA蛋白表達(dá)與腫瘤復(fù)發(fā)無相關(guān)性。Kaplan-Meier生存曲線分析FGFR3和PIK3CA蛋白表達(dá)對生存期影響,FGFR3蛋白表達(dá)差異無統(tǒng)計學(xué)意義(p0.05),而PIK3CA蛋白表達(dá)差異有統(tǒng)計學(xué)意義(p0.05)。多因素COX分析FGFR3和PIK3CA基因突變和蛋白表達(dá)對膀胱癌預(yù)后影響,PIK3CA基因突變和蛋白表達(dá)對膀胱癌的預(yù)后影響,差異有統(tǒng)計學(xué)意義(p0.05);而FGFR3基因突變和表達(dá)對膀胱癌預(yù)后的影響,差異無統(tǒng)計學(xué)意義(p0.05)。影響膀胱癌預(yù)后因素的單因素COX分析中,年齡和PIK3CA蛋白表達(dá)強(qiáng)度是影響膀胱癌預(yù)后的因素(p0.05),PIK3CA蛋白表達(dá)可以作為影響膀胱癌預(yù)后的因素(p=0.05)。影響膀胱癌預(yù)后的多因素COX分析,PIK3CA蛋白表達(dá)是影響膀胱癌預(yù)后的因素(p0.05)。結(jié)論:1.FGFR3和PIK3CA蛋白表達(dá)不能作為膀胱癌診斷、治療、復(fù)發(fā)的指標(biāo)。2.FGFR3蛋白表達(dá)與PIK3CA蛋白表達(dá)呈線性相關(guān)。3.PIK3CA蛋白表達(dá)與腫瘤的分期相關(guān),PIK3CA蛋白表達(dá)是膀胱癌預(yù)后的一個不利因素。
[Abstract]:Chapter I Mutations of FGFR3 and PIK3CA genes in bladder cancer and their relationship with prognosis Objective: To detect mutations of FGFR3 and PIK3CA genes in bladder cancer tissues, matched adjacent tissues and benign tumor tissues, and to analyze the relationship between mutations of FGFR3 and PIK3CA genes and clinicopathological parameters. Methods: DNA was extracted from 34 cases of bladder cancer, 7 cases of matched adjacent tissues, 6 cases of benign tumors, and exon 7 of FGFR3 and PIK3CA genes were used as the effective therapeutic targets and prognostic markers. The primers of exon 9 and 20 were designed and amplified by PCR. The target products were identified by electrophoresis. The mutations of FGFR3 and PIK3CA genes in the PCR products were detected by Sanger gene detection method. The relationship between the mutations of FGFR3 and PIK3CA genes and clinicopathological parameters in bladder cancer tissues was discussed. The relationship between the mutations of FGFR3 and PIK3CA genes and the prognosis of bladder cancer was also discussed. Results: The mutations of FGFR3 gene in bladder cancer were 17.65% (6/34), 28.57% (2/7) in adjacent tissues and 28.57% (2/7) in benign tumors. There was no significant difference in the mutations of FGFR3 gene between the three groups (p0.05). There was no mutation in adjacent tissues and benign tumors. There was no significant difference between adjacent tissues and benign tumors (p0.05). The mutation of FGFR3 gene was found in 6 cases of bladder cancer. The mutation of S249C in exon 7 was missense in 2 cases of adjacent tissues. It is a mutation at exon 7, S249C, with missense mutation. In one case, the mutation at exon 7, S249C, occurred in both cancer tissues and matched adjacent tissues. In bladder cancer, the mutation of PIK3CA gene was found at exon 9, E545K in both cases. There was no mutation at exon 20. There were no mutations in FGFR3 and PIK3CA genes and no clinical pathological grading and staging. Correlation. Of 34 cases of bladder cancer, 5 had recurrence, 6 had recurrence, 3 had recurrence, the Pearson correlation coefficient (r) was 0.46 (p = 0.01), and the mutation of FGFR3 gene was positively correlated with recurrence. There was no significant difference in survival time between R3 mutant and wild type (p0.05), but there was significant difference in survival time between PIK3CA mutant and wild type (p0.05). Age was an important prognostic factor in univariate COX analysis of prognostic factors of bladder cancer (p0.05). Multivariate COX analysis showed that age was also an independent prognostic factor for bladder cancer (p0.05). There was no correlation between age, sex, tumor size, number, stage, grade, and the expression of FGFR3 and PIK3CA protein. The mutation of FGFR3/S249C gene can not be used as a marker for the diagnosis of bladder cancer, but can predict the recurrence of bladder cancer. Chapter 2 The clinical biological significance of the expression of FGFR3 and PIK3CA proteins in bladder cancer. The expression of fibroblast growth factor receptor 3 and PIK3CA in bladder cancer was analyzed. The correlation between the expression of fibroblast growth factor receptor 3 and PIK3CA and clinicopathological parameters was analyzed. The clinical significance of the expression of fibroblast growth factor receptor 3 and PIK3CA protein in bladder cancer was revealed. The expression of FGFR3 and PIK3CA protein in tissues, 7 matched paracancerous tissues and 5 benign bladder tumors was measured. The relationship between the expression of FGFR3 and PIK3CA protein in bladder tumors and the clinicopathological parameters was investigated. The relationship between the expression of FGFR3 and PIK3CA protein and the prognosis of bladder cancer was studied. Results: The expression of FGFR3 protein in bladder cancer was 50.00% (21/42), 14.29% (1/7) in adjacent tissues, and 20.00% (1/5) in benign tumors. There was no statistical difference in the expression of FGFR3 protein between adjacent tissues and benign tumors. The expression of PIK3CA protein in bladder cancer was 64.28% (27/42), paracancerous tissue was 14.29% (1/7), and benign tumor was 20% (1/5). There was significant difference between cancer tissue and paracancerous tissue (p0.05). There was no significant difference between cancer tissue and benign tumor tissue (p0.05). The expression of FGFR3 protein in different stages of bladder cancer. The expression of Ta-T1 was 55.56% (10/18) and T2-T4 was 45.83% (11/24). Although the expression of Ta-T1 was higher than that of T2-T4, the difference was not statistically significant (P 0.05). The expression of FGFR3 protein was 53.85% (7/13) in low grade and 48.28% (14/29) in high grade of bladder cancer. The expression of FGFR3 protein in low grade bladder cancer was higher than that in high grade. The expression of PIK3CA protein in different stages of bladder cancer, Ta-T1 stage was 83.33% (15/18), T2-T4 stage was 50.00% (12/24), the expression of Ta-T1 stage was significantly higher than that in T2-T4 stage, and the difference was statistically significant (p0.05). The expression of PIK3CA protein in different pathological grades of bladder cancer was 76.92% (10/13) in low grade and 58.92% (10/13) in high grade. The expression of PIK3CA protein in low grade bladder cancer was higher than that in high grade bladder cancer (p 0.05). The expression of FGFR3 protein in bladder cancer was mainly 2 points (57.14% (12/21); the expression of PIK3CA protein was mainly 3 points (50.00% (14/28). The expression of FGFR3 protein was positively correlated with the expression of PIK3CA protein (r = 0.845, p = 0.0). There was no correlation between the expression of FGFR3 and PIK3CA protein and tumor recurrence. Kaplan-Meier survival curve showed that the expression of FGFR3 and PIK3CA protein had no significant effect on survival, but the expression of PIK3CA protein had no statistical difference (p0.05). Multivariate COX analysis showed that mutation and protein expression of FGFR3 and PIK3CA gene had significant effect on the prognosis of bladder cancer, and mutation and protein expression of PIK3CA gene had significant effect on the prognosis of bladder cancer (p0.05). However, mutation and expression of FGFR3 gene had no significant effect on the prognosis of bladder cancer (p0.05). Age and PIK3CA protein expression were the prognostic factors of bladder cancer (p0.05). PIK3CA protein expression could be a prognostic factor of bladder cancer (p = 0.05). Multivariate COX analysis showed that PIK3CA protein expression was a prognostic factor of bladder cancer (p0.05). Conclusion: 1. The expression of 3CA protein can not be used as an indicator of diagnosis, treatment and recurrence of bladder cancer. 2. The expression of FGFR3 protein is linearly correlated with the expression of PIK3CA protein. 3. The expression of PIK3CA protein is correlated with the stage of bladder cancer. The expression of PIK3CA protein is a disadvantageous factor for the prognosis of bladder cancer.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R737.14
[Abstract]:Chapter I Mutations of FGFR3 and PIK3CA genes in bladder cancer and their relationship with prognosis Objective: To detect mutations of FGFR3 and PIK3CA genes in bladder cancer tissues, matched adjacent tissues and benign tumor tissues, and to analyze the relationship between mutations of FGFR3 and PIK3CA genes and clinicopathological parameters. Methods: DNA was extracted from 34 cases of bladder cancer, 7 cases of matched adjacent tissues, 6 cases of benign tumors, and exon 7 of FGFR3 and PIK3CA genes were used as the effective therapeutic targets and prognostic markers. The primers of exon 9 and 20 were designed and amplified by PCR. The target products were identified by electrophoresis. The mutations of FGFR3 and PIK3CA genes in the PCR products were detected by Sanger gene detection method. The relationship between the mutations of FGFR3 and PIK3CA genes and clinicopathological parameters in bladder cancer tissues was discussed. The relationship between the mutations of FGFR3 and PIK3CA genes and the prognosis of bladder cancer was also discussed. Results: The mutations of FGFR3 gene in bladder cancer were 17.65% (6/34), 28.57% (2/7) in adjacent tissues and 28.57% (2/7) in benign tumors. There was no significant difference in the mutations of FGFR3 gene between the three groups (p0.05). There was no mutation in adjacent tissues and benign tumors. There was no significant difference between adjacent tissues and benign tumors (p0.05). The mutation of FGFR3 gene was found in 6 cases of bladder cancer. The mutation of S249C in exon 7 was missense in 2 cases of adjacent tissues. It is a mutation at exon 7, S249C, with missense mutation. In one case, the mutation at exon 7, S249C, occurred in both cancer tissues and matched adjacent tissues. In bladder cancer, the mutation of PIK3CA gene was found at exon 9, E545K in both cases. There was no mutation at exon 20. There were no mutations in FGFR3 and PIK3CA genes and no clinical pathological grading and staging. Correlation. Of 34 cases of bladder cancer, 5 had recurrence, 6 had recurrence, 3 had recurrence, the Pearson correlation coefficient (r) was 0.46 (p = 0.01), and the mutation of FGFR3 gene was positively correlated with recurrence. There was no significant difference in survival time between R3 mutant and wild type (p0.05), but there was significant difference in survival time between PIK3CA mutant and wild type (p0.05). Age was an important prognostic factor in univariate COX analysis of prognostic factors of bladder cancer (p0.05). Multivariate COX analysis showed that age was also an independent prognostic factor for bladder cancer (p0.05). There was no correlation between age, sex, tumor size, number, stage, grade, and the expression of FGFR3 and PIK3CA protein. The mutation of FGFR3/S249C gene can not be used as a marker for the diagnosis of bladder cancer, but can predict the recurrence of bladder cancer. Chapter 2 The clinical biological significance of the expression of FGFR3 and PIK3CA proteins in bladder cancer. The expression of fibroblast growth factor receptor 3 and PIK3CA in bladder cancer was analyzed. The correlation between the expression of fibroblast growth factor receptor 3 and PIK3CA and clinicopathological parameters was analyzed. The clinical significance of the expression of fibroblast growth factor receptor 3 and PIK3CA protein in bladder cancer was revealed. The expression of FGFR3 and PIK3CA protein in tissues, 7 matched paracancerous tissues and 5 benign bladder tumors was measured. The relationship between the expression of FGFR3 and PIK3CA protein in bladder tumors and the clinicopathological parameters was investigated. The relationship between the expression of FGFR3 and PIK3CA protein and the prognosis of bladder cancer was studied. Results: The expression of FGFR3 protein in bladder cancer was 50.00% (21/42), 14.29% (1/7) in adjacent tissues, and 20.00% (1/5) in benign tumors. There was no statistical difference in the expression of FGFR3 protein between adjacent tissues and benign tumors. The expression of PIK3CA protein in bladder cancer was 64.28% (27/42), paracancerous tissue was 14.29% (1/7), and benign tumor was 20% (1/5). There was significant difference between cancer tissue and paracancerous tissue (p0.05). There was no significant difference between cancer tissue and benign tumor tissue (p0.05). The expression of FGFR3 protein in different stages of bladder cancer. The expression of Ta-T1 was 55.56% (10/18) and T2-T4 was 45.83% (11/24). Although the expression of Ta-T1 was higher than that of T2-T4, the difference was not statistically significant (P 0.05). The expression of FGFR3 protein was 53.85% (7/13) in low grade and 48.28% (14/29) in high grade of bladder cancer. The expression of FGFR3 protein in low grade bladder cancer was higher than that in high grade. The expression of PIK3CA protein in different stages of bladder cancer, Ta-T1 stage was 83.33% (15/18), T2-T4 stage was 50.00% (12/24), the expression of Ta-T1 stage was significantly higher than that in T2-T4 stage, and the difference was statistically significant (p0.05). The expression of PIK3CA protein in different pathological grades of bladder cancer was 76.92% (10/13) in low grade and 58.92% (10/13) in high grade. The expression of PIK3CA protein in low grade bladder cancer was higher than that in high grade bladder cancer (p 0.05). The expression of FGFR3 protein in bladder cancer was mainly 2 points (57.14% (12/21); the expression of PIK3CA protein was mainly 3 points (50.00% (14/28). The expression of FGFR3 protein was positively correlated with the expression of PIK3CA protein (r = 0.845, p = 0.0). There was no correlation between the expression of FGFR3 and PIK3CA protein and tumor recurrence. Kaplan-Meier survival curve showed that the expression of FGFR3 and PIK3CA protein had no significant effect on survival, but the expression of PIK3CA protein had no statistical difference (p0.05). Multivariate COX analysis showed that mutation and protein expression of FGFR3 and PIK3CA gene had significant effect on the prognosis of bladder cancer, and mutation and protein expression of PIK3CA gene had significant effect on the prognosis of bladder cancer (p0.05). However, mutation and expression of FGFR3 gene had no significant effect on the prognosis of bladder cancer (p0.05). Age and PIK3CA protein expression were the prognostic factors of bladder cancer (p0.05). PIK3CA protein expression could be a prognostic factor of bladder cancer (p = 0.05). Multivariate COX analysis showed that PIK3CA protein expression was a prognostic factor of bladder cancer (p0.05). Conclusion: 1. The expression of 3CA protein can not be used as an indicator of diagnosis, treatment and recurrence of bladder cancer. 2. The expression of FGFR3 protein is linearly correlated with the expression of PIK3CA protein. 3. The expression of PIK3CA protein is correlated with the stage of bladder cancer. The expression of PIK3CA protein is a disadvantageous factor for the prognosis of bladder cancer.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R737.14
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