Quantitative detection of peripheral MDR genes in breast cancer before chemotherapy and their clinical significances

Objective： To explore expressions of multidrug resistance gene （MDR） product P170 and peripheral blood MDR1 mRNA in breast cancer tissue and their clinical significances. Methods： Fluorescent quantitative RT-PCR and immunohistochemistry were used to detect peripheral blood MDR1 mRNA before chemotherapy in breast cancer tissue samples from 32 cases with P170 positive （trial group） and 11 cases with P170 negative （control group）. Results： There were 14 cases （43.75%） peripheral blood MDR1 mRNA positive and 18 cases （56.25%） negative in 32 cases P170 positive breast cancer patients, while there were 6 cases （54.55%） peripheral blood MDR1 mRNA positive and 5 cases （45.45%） negative in 11 cases P170 negative breast cancer patients; there wasn＇t a significance difference with peripheral blood MDR1 mRNA express rate in P170 positive （43.75%） group and P170 negative （54.55%） group （x2 = 0.383, P 〉 0.05）. There were 12 cases （37.50%） Her-2 positive and 20 cases （62.50%） negative in P170 positive group; there were 2 cases （18.18%） Her-2 positive and 9 cases （81.82%） negative in P170 negative group; there wasn＇t a significance difference with Her-2 positive rate in P170 positive group （37.50%） and P170 negative （18.18%） group （χ^2= 1.391, P 〉 0.05）. There were 13 cases （40.63%） ER positive and 19 cases （59.37%） negative in P170 positive group; there were 7 cases （63.64%） ER positive and 4 cases （36.36%） negative in P170 negative group; there wasn＇t a significance difference with ER positive rate in P170 positive group （40.63%） and P170 negative （63.64） group （χ^2 = 1.742, P 〉 0.05）. There were 20 cases （62.50%） PR positive and 12 cases （37.50%） negative in P170 positive group; there were 4 cases （36.36%） PR positive and 7 cases （63.64%） negative in P170 negative group; there wasn＇t a significance difference with PR positive rate in P170 positive group （62.50%） and P170 negative （36.36%） group （χ^2 = 2.267, P 〉 0.05）. There were 9 cases （28.12%） with lymphaden metastasis and 23 cases （71.88%） without lymphaden metastasis in P170 positive group; there were 5 cases （45.45%） with lymphaden metastasis and 6 cases （54.55%） without lymphaden metastasis in P170 negative group; there wasn＇t a significance difference with lymphaden metastasis rate in P170 positive group （28.12%） and P170 negative （45.45%） group （χ^2 = 1.120, P 〉 0.05）. Conclusion： 1） There were possibly MDR expressing infrequently in carcinoma tissue and peripheral blood before chemotherapy. 2） FQ-RT-PCR for detecting of peripheral blood MDR1 mRNA is special, sensitive and reliable. It can be used as new molecular biology diagnostic maker dynamic detecting peripheral blood MDR1 mRNA for regulating chemotherapy proposal and elevating chemotherapy effect.

Congenital expression of mdr-1 gene in tissues of carcinoma and its relation with patho morphology and prognosis

AIM To detect the congenital expression patterns of mdr 1 gene in commonly encountered malignant tumors in clinic, and the relationship between the expression of mdr 1 gene and the prognostic morphology in esophageal carcinomas. METHODS A total of 151 resected samples of malignant tumors without preoperative treatment were taken from Anyang City Tumor Hospital. The congenital expression of their mdr 1 gene was detected with reverse transcription polymerase chain reaction (RT PCR) and was compared with each other. The positive incidence of mdr 1 gene in 46 samples of esophageal carcinoma was compared with their differentiated grades, TNM stages and macroscopic types, and the precautions and advantages of RT PCR were evaluated. RESULTS All the 151 samples were confirmed to be malignant histopathologically, including cancers of stomach and gastric cardia (n =51), esophagus ( n =46), colorectum ( n =16), breast ( n =15), thyroid ( n =10), lung ( n =9), uterine cervix ( n =24). The positive expression rate of their mdr 1 gene was 33 3%, 37%, 31 3%, 13 2%, 40%, 55%, and 0% respectively. All the 46 samples of esophageal carcinoma were pathologically confirmed to be squamous cell carcinoma. The total expression rate of their mdr 1 gene was 37% (17/46), 35% (6/17), 40% (8/20), and 33% (3/9) for differentiation grade Ⅰ, Ⅱ and Ⅲ respectively. The expression rate of TNM classification was 33% (6/18), 40% (5/12) and 37% (6/16) in stage Ⅱa, Ⅱb and Ⅲ. The expression rate was 33% (3/9) in ulcerous type, 37% (3/8) in constrictive one, 33% (5/15) in fungoid one, and 40% (6/14) in medullary one. No statistically significant difference was found. CONCLUSION Compared with other methods, RT PCR is more simple, reliable and accurate in detecting mdr 1 gene expression in tissues of tumor. The overexpression of mdr 1 gene in these neoplasms suggested that cases should be handled differently for chemotherapy with rational use of drugs. Excision is the chief treatment for carcinoma of esophagus. The expression of mdr 1 gene in tissues of esophageal cancer is correlated with the parameters of tumor molecular biology which are independent of histopathological morphology.

Chitosan/pshRNA Plasmid Nanoparticles Targeting MDR1 Gene Reverse Paclitaxel Resistance in Ovarian Cancer Cells

In order to investigate the effect of chitosan/pshRNA plasmid nanoparticles targeting MDR1 genes on the resistance of A2780/TS cells to paclitaxel, chitosan/pshRNA plasmid nanoparti- cles were synthesized by means of a complex coacervation technique and transfected into A2780/TS cells. The cells transfected with MDRl-targeted chitosan/pshRNA plasmid nanoparticles were experimental cells and the cells transfected with chitosan/pGPU6/GFP/Neo no-load plasmid nanoparticles served as negative control cells. Morphological features of the nanoparticles were observed under transmission electron microscope （TEM）. MDR1 mRNA expression was assessed by RT-PCR. Half-inhibitory concentration （IC50） ofpaclitaxel for A2780/TS cells was determined by MTT method. TEM showed that the nanoparticles were round-shaped, smooth in surface and the diameters varied from 80 to 120 nm. The MDR1 mRNA in the transfected cells was significantly decreased by 17.6%, 27.8% and 52.6% on the post-transfection day 2, 4 and 7 when compared with that in A2780/TS cells control （P〈0.05）. MTT assay revealed that the relative reversal efficiency was increased over time and was 29.6%, 51.2% and 61.3% respectively in the transfected cells 2, 4, 7 days after transfection and IC_50 （0.197±0.003, 0.144±0.001, 0.120±0.004） were decreased with difference being significant when compared with that in A2780/TS （0.269±0.003） cells control （P〈0.05）. It was concluded that chitosan/pshRNA plasmid nanoparticles targeting MDR1 can effectively reverse the paclitaxel resistance in A2780/TS cells in a time-dependent manner.

PRELIMINARY STUDY OF RETROVIRAL MEDIATED TRANSFER OF THE HUMAN mdr-1 GENE INTO MURINE AND HUMAN HEMATOPOIETIC STEM/PROGENITOR CELLS

To investigate the characteristics of multidrugresistance and transplantation of modified stem/ progenitor cells by multidrugresistant gene (mdr1 gene), we established PA317/MDR1 cell line which producing retroviruses by transfecting the retroviral vector PHaMDR1/A into packging cell line PA317 by Lipofectin. The virus titer of the supernatants was 1.2×105 cfu/ml. We transfected the murine hematopietic cells collected from 5FU pretreated mice and they showed the ability to reconstitute the longterm hematopoiesis of preirradiated mice. After 4 months, both of bone marrow cells and peripheral blood cells of transplanted mice still contained mdr1 gene. We also transfered mdr1 gene into human bone marrow CD34+ cells selected by using magnetic cell sorting system. PCR analysis showed that transduced CD34+ cells maintained the mdr1 cDNA. A fraction of CFUGM originated from transfected CD34+ cells had the charactor of resistance to Taxol. It is indicated that mdr1 gene can be transduced into murine and human stem/proginitor cells through retroviral mediated gene transfer and it protects the transfected cells from cytotoxic drugs.

EXPRESSION OF mdr-1 GENE IN CANCER TISSUE AND ITS ASSOCIATION WITH MORPHOLOGICAL INDEXES OF ESOPHAGEAL CARCINOMA IN ANYANG

Objective: Overexpression of mdr-1 gene is associated with multidrug resistance (MDR) and aggressive characteristics of malignance. Our purposewas to detect the levels of P-gp expression in fresh untreated esophageal carcinomas, and to correlate these levels to current prognostic indicators of morphology.Methods: Reverse transcription polymerase chain reaction (RT-PCR) was used to investigate mdr-1 gene expression of 46 samples from untreated esophageal carcinoma, and compared the positive incidences among differentiated grades, TNM stages and macroscopic types.Results: All 46 samples were pathologically squamous cell carcinoma. The positive' incidences of mdr-1 gene expression were 37% (17/46) in whole group,35% (6/17), 40% (8/20), 33% (3/9), for Ⅰ,Ⅱ and Ⅲdifferentiated grades, respectively. The expression rates of 33% (6/18), 40% (5/12), and 37% (6/16), were found in Ⅱa, Ⅱ, and Ⅲ stage of TNM, respectively. In macroscopic type view, the positive incidence was 37%(3/8) in constrictive, 33% (5/15) in fungating, 40% (6/14)in marrowlike, 33% (319) in ulcerative type. There were no statistically significant differences among each category system of morphology.Conclusion: The result, high level expression of mdr-1 gene in untreated esophageal carcinoma, suggested the poor efficacy of chemotherapy for some esophageal carcinoma patients. And we should cautiously choose cases who will receive chemotherapy. Surgery is still the best treatment for carcinoma of esophagus. Besides, the data also revealed that the expression of mdr-1 gene in untreated esophageal cancer was independent of morphologic prognostic indexes, and that there were no correlation between mdr-1 gene expression and morphological indexes.

CONGENITAL EXPRESSION OF mdr-1 GENE IN FRESH CANCER TISSUES FROM SEVERAL HIGH-INCIDENCE NEOPLASMS WITHOUT PREOPERATIVE CHEMOTHERAPY

Objective: The purpose of the present study is to detect characteristics of primary expression of mdr 1 gene in several neoplasms which has high morbidity in clinic. Methods: 151 resected samples, which are pathologically malignant and clinically untreated before operation, were obtained from Anyang Cancer Hospital. All of them were investigated with RT PCR for the expression of mdr 1 gene and correlated each other. Besides, we evaluated the advantages of RT PCR in this study. Results: The mdr 1 gene expression rate of these 151 samples, including cancers of stomach and gastric cardia (n=51), esophagus (n=46), colorectum (n=16), breast (n=15), thyroid (n=10), lung (n=9), uterine cervix (n=4), was 33.3%, 37%, 31.3%, 13.2%, 40%, 55%, 0%, respectively. Conclusion: Compared with other methods, RT PCR for studying mdr 1 gene expression had certain advantages in simplicity, reliability, and accuracy. Overexpression of mdr 1 gene in these neoplasms suggested that cases should be distinguished before treatment according to MDR of tumor and to choose effective drugs for individual cancer patient.

EFFECTS OF IN VIVO GENE TRANSDUCTION OF ANTI-MDR1 RIBOZYME IN COMBINATION WITH CHEMOTHERAPY ON MULTIDRUG-RESISTANT HUMAN LYMPHOMA GROWTH IN MICE

Objective: To study the effect of adenovirus- mediated transfer of anti-MDR1 ribozyme on the reversal of multidrug resistant (MDR) phenotype of P-glycoprotein (P-gp)-positive Daudi human Burkitt lymphoma both in vitro and in vivo. Methods: A recombinant adenovirus expressing 196Rz (Adv-196Rz) was developed and functionally evaluated. SCID mice inoculated subcutaneously (s.c.) with 5?06 Daudi/MDR20 cells were locally treated with Adv-196Rz or mock virus (Adv-Mock) at the multiplicity of infection (MOI) of 400 PFU once a day for 3 consecutive days. Then the mice were intraperitoneally (i.p.) administrated with vincristine (VCR) 450ng/g for 5 consecutive days. Results: In vitro employment of Adv-196Rz was able to interrupt MDR1 transcription, to inhibit P-gp expression and to restore drug sensitivity to VCR of Daudi/MDR20 cells. In vivo, 87.5% (7/8) of Daudi/MDR20-inoculated mice treated with Adv-Mock+ VCR developed palpable tumor by the 6th week and died or were sacrificed (because of tumor weight > 10% of body weight) by the 11th week. In contrast, among 9 Daudi/MDR20-inoculated mice treated with Adv-196Rz + VCR, only 3 developed tumor by the 11th, 13th and 14th week, respectively. 66.7% of mice survived >120 days in tumor-free. The survival difference between the two groups was very significant (P<0.01). Conclusion: Adenovirus- mediated Transfer of 196Rz can revert drug resistance of MDR tumor cells both in vitro and in vivo. Adv-196Rz may prove useful as an adjuvant in the chemotherapy of P-gp mediated MDR human tumors.

MDR基因及MDR基因编码产物在咽喉部恶性黑色素瘤中表达及其意义

目的 :研究多药耐药 (mnlti-drugresistance ,MDR)基因胎盘型谷胱甘肽 -S -转移酶 (GST -π)和DNA拓朴酶Ⅱ (TopoⅡ )以及MDR基因编码产物P -糖蛋白 (Pgp)、在咽喉部恶性黑色素瘤中的表达及其意义。方法 :应用链霉素亲生物素 -过氧化物酶标S -P法检测 2 8例咽喉部恶性黑色素瘤中Pgp、GST -π和TopoⅡ的表达 ,分析MDR基因及MDR基因编码产物阳性表达率与肿瘤主要临床病理特征的关系。结果 :2 8例标本中Pgp、GST -π和TopoⅡ的表达率分别为 35 .7%、5 7.1%和 4 6 .4 % ,相互间差异无显著性 (P >0 .0 5 )。Pgp、GST -π和TopoⅡ的表达与性别、年龄、肿瘤大小无明显相关 (P >0 .0 5 ) ,与AJC分级显著相关 (P <0 .0 5 )。结论 :Pgp、GST -π和TopoⅡ等多因素联合作用是咽喉部恶性黑色素瘤多药耐药的主要作用机理 ,其表达在化疗敏感性预测中具有必要性和可行性。

急性白血病患者bcl-2和bax基因表达的临床意义及其与mdr-1基因表达的关系

细胞凋亡受抑作为肿瘤细胞的耐药机制 ,是急性白血病 (AL)预后不良的原因之一。bcl 2家族是目前最受重视的调控细胞凋亡的基因家族 ,本研究为探讨bcl 2和bax基因在急性白血病表达的意义 ,应用逆转录 聚合酶链反应 (RT PCR)方法测定 70例初治AL患者及 2 0例正常人的bcl 2 ,bax ,mdr 1基因mRNA水平的表达 ,用流式细胞术检测其蛋白表达。结果表明 ,bcl 2和bax基因在AL患者广泛表达 ,bcl 2mRNA平均表达水平明显高于正常对照 (1.4 6vs 0 .71,P <0 .0 5 )。bcl 2和bax基因表达及bax bcl 2比值与AL患者的年龄、性别、血小板计数、血红蛋白水平、骨髓原始细胞百分率、FAB分型和S +G2 M %均未发现相关。Bcl 2蛋白表达 (34.6 %vs 6 9.2 %,P <0 .0 5 ) ,bax bcl 2mRNA比值 (37.1%vs 82 .9%,P <0 .0 1)决定AL对化疗的敏感性 ,与ALCR率密切相关。bax bcl 2mRNA比值还是影响总生存期的因素。bcl 2与bax基因表达和mdr 1表达两者无相关关系。

荧光定量RT-PCR检测mdr-1基因表达

目的 :建立荧光定量RT PCR检测肿瘤细胞mdr 1基因表达的方法 ,了解肺癌组织中mdr 1的表达水平。方法 :建立荧光定量RT PCR方法 ,在PE770 0型检测仪上定量检测K5 6 2 /ADM耐药株和K5 6 2不耐药株细胞mdr 1基因表达水平 ,同时检测 45例初治肺部肿瘤病人组织标本。结果 :荧光定量RT PCR检测K5 6 2 /ADM耐药株和K5 6 2不耐药株细胞mdr 1基因表达 ,重复 10次实验所得结果平均分别为 (6 86± 0 6 5 )× 10 7拷贝 /μgRNA和 (8 49± 0 6 7)× 10 5拷贝 /μgRNA ,两者相差 80 8倍 ,变异系数分别为 9 5 %和 7 9%。 45例肺部肿瘤中 ,有 12例检出有mdr 1基因不同程度的表达。结论 :荧光定量RT PCR检测mdr 1基因表达方法 ,检测结果用绝对拷贝数来表示 ,定量准确可靠 ,并有利于标准的统一。有 1/4未经化疗的肺癌病人有一定水平mdr

初发及复发转移大肠癌患者外周血MDR-1基因表达的研究

目的 :检测初发及复发转移大肠癌患者外周血中多药耐药基因 (MDR 1)的表达 ,探讨其与病理类型及化疗疗效的关系。方法 :应用逆转录多聚酶链反应 (RT -PCR)法对 5 6例初发大肠癌及 4 0例经术后辅助化疗后复发、转移大肠癌患者的外周血进行检测 ,并与其病理类型及化疗疗效作对比研究。结果 :5 6例初发大肠癌患者外周血中MDR 1阳性表达率为2 7 2 % ,与病理类型无显著相关 (P >0 0 5 ) ;但与肠系膜淋巴结转移密切相关 ,有淋巴结转移者阳性表达率显著高于无淋巴结转移者 (P <0 0 5 ) ;4 0例化疗后复发、转移患者外周血MDR 1阳性表达率为 72 5 % ,与初发大肠癌患者的MDR 1阳性表达率相比差异有显著性 (P <0 0 5 ) ,且MDR 1的表达与化疗疗效呈负相关 ,MDR 1阳性表达者化疗疗效明显低于阴性表达者(P <0 0 5 )。结论 :大肠癌存在先天性和获得性多药耐药性 ;检测外周血MDR 1表达情况可以帮助预测化疗疗效 ,对制定临床化疗方案有指导意义。

MDR1基因的表达与肝癌生物学行为指标的关系

目的:通过对肝癌(hepatocellular carcinoma,HCC)中多药耐药基因(multid-rug resistance gene 1,MDR1)表达和生物学行为指标HBV、AFP、AST的检测,探讨肝癌中MDR1的表达特点及其与肝癌生物学行为的相互关系。方法:运用荧光定量PCR(flurescence quantitative-PCR,FQ-PCR)技术检测MDR1基因在51例肝癌组织和10例正常肝组织中的表达,分析MDR1基因的表达与肝癌生物学特性之间的关系。结果:MDR1在肝癌中的表达为0.55±0.27,正常肝组织中的表达为0.23±0.10,(P<0.05)。表达与肿瘤Ed-mondson分级呈正相关(P<0.05)。HBV感染、AFP阳性、AST升高与MDR1基因表达明显呈正相关(r=0.463、0.473、0.299)。结论:肝癌中存在原发性耐药的现象,MDR1基因高表达在肝癌原发性耐药中可能起重要作用;HBV的检测可以作为肝癌化疗耐药的参考指标之一,血浆中AFP水平、AST变化可以作为动态监测MDR1表达的参考指标。

急性白血病患者bcrp、mdr-1、mrp-1和lrp耐药基因表达的临床意义

目的:定量检测急性白血病(AL)患者乳腺癌耐药相关蛋白基因(bcrp)、多药耐药基因(mdr-1)、多药耐药相关蛋白基因(mrp-1)及肺耐药相关蛋白基因(lrp)表达及其与临床的关系。方法:应用实时荧光定量逆转录-聚合酶链反应RT-PCR(real time fluorescent quantitative RT-PCR,FQ RT-PCR)检测105例AL患者bcrp、mdr-1、mrp-1、lrp等基因的拷贝数,分析其与临床预后的关系。结果:AL组bcrp基因拷贝数(2.8×103)±(8.4×103)明显高于正常对照组(7.6×102)±(2.3×103),P<0.05;mdr-1基因拷贝数(1.3×105)±(2.2×105)及lrp基因拷贝数(8.3×105)±(1.0×106)显著高于正常对照组(P<0.01)。复发组AL患者的mdr-1拷贝数(3.7×104)±(1.1×105)明显高于初治组患者(1.1×104)±(3.6×104),P<0.05。在AL耐药组mdr-1拷贝数(2.1×104)±(9.1×104)明显高于敏感组(1.0×104)±(3.8×104),P<0.05)。经直线相关分析显示mdr-1和mrp-1,mdr-1和lrp,mrp-1和lrp之间呈显著正相关,经等级相关分析显示mdr-1和lrp的表达与耐药性密切相关。结论:急性白血病多药耐药的出现与bcrp、mdr-1、mrp-1、lrp中一项或几项过度表达有关,mdr-1较其它基因更具有预测临床预后的意义。

选择性COX-2抑制剂塞来昔布抑制B细胞淋巴瘤细胞株MDR-1及Bcl-2的mRNA表达并增强表柔比星的抗肿瘤作用

背景与目的:部分非霍奇金淋巴瘤(non-Hodgkin’s lymphoma,NHL)具有高表达环氧合酶-2(cyclooxygenase-2,COX-2)的特征,而后者与P-糖蛋白及Bcl-2表达相关,可能导致NHL对化疗耐药。本研究旨在探讨B细胞淋巴瘤细胞株中COX-2的表达以及选择性COX-2抑制剂塞来昔布增强淋巴瘤细胞对表柔比星抗肿瘤效应的敏感性及其可能机制。方法:用荧光定量PCR(q RT-PCR)及蛋白[质]印迹法(Western blot)分别检测Raji、Jeko-1和Namalwa等淋巴瘤细胞株以及正常人外周血B细胞的COX-2表达;以梯度浓度的塞来昔布作用于淋巴瘤细胞株,CCK-8方法检测细胞增殖的抑制程度,q RT-PCR检测各细胞株MDR-1 mRNA及Bcl-2 mRNA表达的变化;表柔比星单独或联合不同浓度的塞来昔布处理Raji细胞株72 h后,CCK-8方法分析塞来昔布对表柔比星的增敏作用。结果:各淋巴瘤细胞株及正常对照外周血B细胞均不表达COX-2。塞来昔布单药即可对各淋巴瘤细胞株产生程度不同的抗增殖效应;随着塞来昔布作用浓度的增加,除Jeko-1细胞不表达MDR-1外,其余细胞株MDR-1 mRNA及Bcl-2 mRNA表达水平逐渐下降;塞来昔布明显增强表柔比星对Raji细胞的抗肿瘤活性,两者之间具有协同作用。结论:选择性COX-2抑制剂塞来昔布下调B细胞淋巴瘤细胞株的MDR-1 mRNA及Bcl-2 mRNA水平,并且增强表柔比星对淋巴瘤细胞的抗肿瘤效应。

MDR相关新基因CA916798真核表达载体的构建及其对H446细胞增殖的影响

目的构建新基因CA916798真核表达载体,并观察其对H446增殖的影响。方法以人A549细胞总RNA为模板,用RT-PCR技术扩增获得CA916798开放阅读框序列,连接入pBluescriptⅡSK克隆载体上,将重组质粒转化入大肠杆菌DH5α内并提取质粒,酶切与DNA测序鉴定准确,再用双酶切方法将CA916798开放阅读框序列定向连接到真核表达载体pQE-Trisystem中,构成pQE-Tri-CA916798,将pQE-Tri-CA916798转化入大肠杆菌DH5α内并提取质粒,双酶切鉴定、DNA测序鉴定准确,脂质体转染人小细胞肺癌细胞系H446细胞,选取稳定转染不同质粒的H446细胞,采用MTT绘制细胞增殖曲线,流式细胞仪检测细胞周期和细胞凋亡率。结果扩增出CA916798开放阅读框序列,大小约为350bp,重组质粒的酶切鉴定结果与预期一致,测序结果完全正确。转染了pQE-Tri-CA916798的H446细胞顺铂作用后增殖速度加快,凋亡率下降。结论成功构建了pQE-Tri-CA916798真核表达质粒,并初步证实了其与顺铂诱导的肺癌多药耐药相关。

乳腺癌原发灶和淋巴结转移灶MDR1 mRNA表达及与临床指标的相关性分析

目的:通过对乳腺癌原发灶和淋巴结转移灶的多药耐药基因1(multi-drug resistance gene 1,MDR1)mRNA表达差异及与临床指标相关性进行分析,探讨肿瘤多药耐药和侵袭转移之间可能存在的相关性。方法:应用实时荧光定量PCR(real-time fluorescence quantitative PCR,RFQ-PCR)方法检测21例浸润性乳腺癌患者的原发灶和淋巴结转移灶MDR1 mRNA的表达。结果:MDR1 mRNA在淋巴结转移灶中的表达水平明显高于原发灶(P<0.05),并且在淋巴结转移灶和原发灶中的表达有相关性(r=0.795,P<0.05)。MDR1 mRNA表达与雌激素受体(estrogen receptor,ER)、孕激素受体(progesterone receptor,PR)、CerbB2、肿瘤大小和淋巴结转移个数等临床指标无相关性(P>0.05)。结论:MDR1 mRNA表达可能与肿瘤的侵袭转移有关。

多药抗药基因Mdrl探针的克隆及初步应用

多药抗药基因Ｍｄｒｌ的表达水平与细胞的耐药性直接相关，检测Ｍｄｒｌ的表达水平可预测化疗的效果以及预后，用分子原位杂交的方法可检测单个细胞中Ｍｄｒｌ的表达水平．用ＰＣＲ扩增方法获得了一段特异的ＤＮＡ片段，并将其克隆到ｐＵＣ１８载体中，经ＤＮＡ序列分析证明与文献报道一致，此探针可用于临床标本的分子杂交检测．

RNA干扰沉默mdrl基因联合阿霉素对卵巢癌耐药细胞株SKOV_3/ADM增殖的抑制作用

目的:观察RNA干扰沉默mdrl基因后化疗药物对卵巢癌耐药细胞株SKOV3 /ADM增殖的影响。方法:RT -PCR检测靶向于mdrl基因的小发夹状RNA(pshRNA -mdrl)对SKOV3 /ADM细胞内mdr1mRNA表达的影响;采用MTT法及软琼脂克隆形成实验测定pshRNA -mdrl联合化疗药物阿霉素对SKOV3 /ADM的增殖抑制作用。结果:pshRNA -mdrl能抑制SKOV3 /ADM细胞mdr1mRNA的表达,pshRNA -mdr1联合阿霉素明显抑制SKOV3 /ADM细胞的增殖。结论:靶向于mdr1的pshRNA联合阿霉素能显著抑制SKOV3 /ADM的增殖。

初发及复发转移大肠癌组织与外周血MDR-1基因表达相关性研究

目的 探讨初发及复发转移大肠癌组织与外周血MDR 1基因表达的相关性。方法 采用RT PCR方法分别检测了 5 6例初发大肠癌组织及外周血和 4 0例术后化疗的复发转移大肠癌病灶组织及外周血的MDR 1表达情况。结果 无论是组织中 ,还是外周血中经过化疗的大肠癌患者MDR 1的阳性表达率均明显高于初发大肠癌患者 ,统计学上有显著性差异 (P <0 .0 5 )。结论 可以用外周血代替癌组织来检测MDR

RNAi抑制肾癌细胞MDR1基因表达及对顺铂化疗耐药的逆转

目的:研究RNA干扰(RNA interference,RNAi)对肾癌细胞多药耐药基因1(multidrug resistance,MDR1)表达的抑制作用,并分析干扰前后肾癌细胞对顺铂(cisplatin)敏感度的变化。方法:根据MDR1基因序列设计3条小干扰RNA(small in-terfering RNA,siRNA),转染肾癌A498细胞;RT-PCR法检测转染前后MDR1 mRNA表达水平的变化,筛选出干扰效率最高的siRNA序列;进而利用慢病毒包装siRNA重组质粒,感染A498细胞,RT-PCR法筛选沉默效果最好的细胞株进行克隆;Western印迹法检测MDR1蛋白表达水平;MTT法检测干扰前后顺铂对半数细胞抑制浓度(half inhibitory concentration,IC50)的变化。结果:3条siRNA序列均能不同程度地抑制细胞MDR1基因的表达,其中siRNA-1序列能更有效地封闭MDR1基因,使MDR1 mRNA表达水平下降67%;筛选得到稳定转染的细胞株与未干扰细胞株相比,MDR1蛋白表达量明显下降(P<0.01),并使顺铂对细胞的IC50值降低约83.37%(P<0.01)。结论:RNAi能有效抑制肾癌A498细胞MDR1基因的表达,并可显著增加其对顺铂的敏感度,从而使肾癌细胞化疗耐药逆转成为可能。