Anxa2和P-gp蛋白相互作用对多药耐药乳腺癌细胞迁移与侵袭影响的研究

目的:探讨Anxa2和P-gp蛋白相互作用对多药耐药乳腺癌细胞迁移与侵袭的影响.方法:采用小干扰RNA技术下调人多药耐药乳腺癌细胞MCF-7/ADR中P-gp的表达,并筛选稳定的单克隆细胞株;利用噻唑蓝(MTT)比色法和Transwell实验研究P-gp的表达对MCF-7及其耐药细胞MCF-7/ADR的增殖、迁移和侵袭能力的影响;利用免疫沉淀和免疫荧光分析多药耐药细胞中Anxa2和P-gp的相互关系.结果:蛋白印迹法检测发现,Anxa2和P-gp在耐药乳腺癌细胞中均高表达;MCF-7/ADR与其亲本细胞MCF-7相比,迁移和侵袭能力明显增强;MCF-7/ADR细胞中P-gp的表达被抑制后,其迁移和侵袭能力显著下降,并且差异有统计学意义,P＜0.05.免疫沉淀证实,Anxa2和P-gp之间存在相互作用;免疫荧光发现,Anxa2和P-gp在细胞膜上存在很好的共定位.结论:降低P-gp的表达可以明显抑制耐药乳腺癌细胞MCF-7/ADR的迁移和侵袭能力,Anxa2和P-gp之间存在较好的共定位和相互作用.提示Anxa2和P-gp的相互作用有可能对增强多药耐药乳腺癌细胞的侵袭转移能力起到一个重要的作用.     

MDR1和Anxa2在乳腺癌组织中的表达与侵袭转移的关系研究

探讨乳腺癌组织中多药耐药基因（MDR1）和膜联蛋白（Anxa2）表达的相关性及其与乳腺癌转移的关系。方法：应用荧光定量PCR的方法检测20例配对的乳腺导管内癌、100例乳腺浸润性导管癌、70例癌旁正常组织中MDR1和Anxa2 mRNA的相对表达情况。结果：MDR1 mRNA在乳腺导管内癌中的表达水平明显高于癌旁正常组织（P=0.005）,乳腺浸润性导管癌组织中mRNA的表达明显高于癌旁正常组织（P=0.017）和导管内癌（P=0.019 6）。Anxa2 mRNA在乳腺导管内癌中表达与癌旁正常组织相比无显著性差异（P=0.188 9）,但是在乳腺浸润性导管癌组织中的表达明显高于导管内癌（P=0.000 8）和癌旁正常组织（P<0.000 1）;MDR1和Anxa2 mRNA的表达升高均与患者出现淋巴结转移有关（P<0.01）;2种基因在乳腺浸润性导管癌中的表达呈正相关（P<0.000 1）。结论：在肿瘤进展过程中,MDR1和Anxa2 mRNA表达上调与乳腺癌的淋巴结转移有关,二者之间表达具有正相关提示肿瘤细胞的多药耐药的获得和肿瘤侵袭转移之间有着密切联系。     

Augmentation by Bispecific F(ab')2 Reactive with P-Glycoprotein and CD3 of Cytotoxicity of Human Effector Cells on P-Glycoprotein Positive Human Renal Cancer Cells

A bispecific F(ba')₂ was constructed that was composed of two Fab fragments, one derived from anti-CD3 monoclonal antibody (mAb) (OKT3) and the other from anti P-glycoprotein mAb (MRK 16). This bispecific F(ab')₂ enhanced the binding and cytotoxicity of human peripheral blood mononuclear cells (PBMCs) on P-glycoprotein-positive human kidney cancer cells (ADMHK/E). It had no effect on the cytotoxicity of PBMCs on P-glycoprotein-negative HK/E cells [long-term cultured HK/E (LCHK/E)]. Control F(ab')₂ composed of OKT3 or MRK16 alone did not influence the cytotoxicity of PBMCs on ADMHK/E cells. These findings suggest that the MRK16-OKT3 bispecific F(ab')₂ may be therapeutically beneficial in treatment of human multidrug-resistant cancers.     

Monoclonal Anti-P-glycoprotein Antibody-dependent Killing of Multidrug-resistant Tumor Cells by Human Mononuclear Cells

Mouse monoclonal antibodies (MRK16 and MRK17) against human multidrug-resistant cancer cell lines were tested for antibody-dependent cytotoxicity mediated by human blood mononuclear cells, using a 4-h ⁵¹Cr release assay. MRK16 (IgG₂₈ isotype) was shown to be more effective than MRK17 (IgG₁ isotype). Moreover, when four pairs of drug-resistant and their parent sensitive human cancer cells were tested for antibody-dependent cell-mediated cytolysis (ADCC) using MRK16, only the drug-resistant cell lines were susceptible to ADCC reaction. When highly purified lymphocytes (>99%) and monocytes (>97%) were isolated from blood mononuclear cells by centrifugal elutriation and adherence, MRK16 promoted both lymphocyte- and monocyte-mediated tumor cell killing, whereas MRK17 induced only a lymphocyte-mediated ADCC reaction. These results suggest that MRK16 of IgG₂₈ subtype may be a useful therapeutic agent in eradication of drug-resistant cancer cells expressing P-glycoprotein through ADCC reaction.     

p-糖蛋白(pgp)在乳腺癌组织中的表达及与预后的关系

目的：研究P－糖蛋白（P－glycoprotein,Pgp)在乳腺癌组织中的表达,评估其在乳腺癌预后中的作用。方法：采用免疫组织化学方法（Immunohistochemistry,IHC)检测47例手术切除的乳腺癌组织中Pgp的表达,并分析其与临床、病理特征的关系及对预的的影响。结果：Pgp在乳腺癌组织中的总表达率为83．0％（39／47例）,高表达者占53．2％（25／47例）,其与月经状、肿瘤大小、淋巴结转移、组织分级和激素受体状况均无关（P＞0．05）;Kaplan-Meier生存分析结果表明Pgp表达与无病生存期和总生存期均显著相关（P＜0．01）,而多因素Cox分析却显著仅有Pgp和淋巴结转移是独立的预后因素,结论：Pgp在乳腺癌组织中具有较高的表达,有可能成为判断乳腺癌预后的有用指标。     

Potent Killing of Paclitaxel- and Doxorubicin-resistant Breast Cancer Cells by Calphostin C Accompanied by Cytoplasmic Vacuolization

Drug resistance is a major impediment to the successful treatment of breast cancer using chemotherapy. The photoactivatable drug calphostin C has shown promise in killing select drug-resistant tumor cells lines in vitro. To assess the effectiveness of this agent in killing doxorubicin- or paclitaxel-resistant breast tumor cells and to explore its mode of action, MCF-7 cells were exposed to increasing concentrations of either doxorubicin or paclitaxel until maximum resistance was obtained. This resulted in the creation of isogenic drug-resistant MCF-7TAX and MCF-7DOX cell lines, which were approximately 50- and 65-fold resistant to paclitaxel and doxorubicin, respectively. Interestingly, calphostin C was able to kill MCF-7TAX cells as efficiently as wildtype MCF-7 cells (IC50s were 9.2 and 13.2 nM, respectively), while MCF-7DOX cells required a 5-fold higher concentration of calphostin C to achieve the same killing (IC50 = 64.2 nM). Consistent with their known mechanisms of action, paclitaxel killed tumor cells by inducing mitotic arrest and cell multinucleation, while doxorubicin induced plasma membrane blebbing and decreased nuclear staining with propidium iodide. In contrast, cytoplasmic vacuolization accompanied cell killing by calphostin C in these cell lines, without the induction of caspase-8 or PARP cleavage or the release of cytochrome c from mitochondria. Calphostin C had little effect on the uptake of either paclitaxel or doxorubicin by the cells. Taken together, the above data suggests that calphostin C is able to potently kill drug-resistant breast tumor cells through a mechanism that may involve the induction of cytoplasmic vacuolization, without activation of typical apoptotic pathways. Consequently, calphostin C may prove useful clinically to combat tumor growth in breast cancer patients whose tumors have become unresponsive to anthracyclines or taxanes, particularly in association with photodynamic therapy.     

激素受体阳性晚期乳腺癌内分泌治疗选择

激素受体阳性乳腺癌占所有乳腺癌的70%。内分泌治疗是这个亚型乳腺癌的主要治疗手段,最常见药物有他莫昔芬和芳香酶抑制剂如阿拉曲唑、来曲唑和依西美坦。全文重点总结新型内分泌治疗药物,如雌激素受体降解剂(Fulvestrant),以及新的靶向药物如mTOR抑制剂(Everolimus)、CDK4/6抑制剂(Palbociclib、Ribociclib和Abemaciclib)和PI3K抑制剂(Alpelisib、Buparlisib和Pictilisib)等。新的靶向药物联合内分泌治疗已经改变了临床实践,延长激素受体阳性晚期乳腺癌患者的生存期。     

EGCG降低P糖蛋白表达逆转白血病多药耐药的研究

[目的]探讨表没食子儿茶素没食子酸酯[(-)epigallocatechin-3-gallate,EGCG]对人白血病细胞K562/A02多药耐药性的逆转作用及相关机制.[方法]采用MTT法确定EGCG的非细胞毒性剂量,以及对K562/A02细胞药物敏感性的影响.以流式细胞术测定EGCG作用前后细胞内阿霉素浓度的变化;同时采用半定量逆转录聚合酶链反应(RT-PCR)及Western blot方法分别检测经典多药耐药基因(MDR1)mRNA及其产物P糖蛋白的表达.[结果] EGCG在低于103.2μmol/L时对K562/A02细胞的生长抑制率小于10%.40μmol/L、60μmol/L及80μmol/LEGCG均可增加K562/A02细胞对阿霉素的敏感性,使阿霉素对K562/A02细胞的IC50由原来的113.34μg/ml降低至9.66μg/ml、7.67μg/ml和4.68μg/ml,其逆转倍数分别为11.73、14.77和24.23倍.流式细胞术结果表明EGCG可增加细胞内阿霉素浓度,并呈剂量依赖性.RT-PCR及Western blot结果显示不同浓度的EGCG均可抑制K562/A02细胞MDRlmRNA及P糖蛋白的表达.[结论]EGCG可部分逆转人白血病细胞K562/A02对阿霉素的耐药性,其逆转机制与增加细胞内化疗药物浓度、抑制经典多药耐药基因MDRlmRNA及其产物P糖蛋白的表达有关.     

化疗增敏剂影响P-糖蛋白依赖多药耐药性肿瘤细胞99Tcm-MIBI摄取的实验研究

目的观察化疗增敏剂对多药耐药性(Multi-drug resistance,MDR)肿瘤细胞P-糖蛋白(P-glycoprotein,P-gp)的影响及伴随99Tcm-MI BI摄取动力学的变化,以建立用99Tcm-MIBI来评价化疗增敏剂效果的方法。方法MDR人乳腺癌细胞MCF-7/Adr 37℃培养。(1)实验组和对照组细胞培养基中分别加入化疗增敏剂维拉帕米(10μmol/L) 和等体积培养液DMEM。99Tcm-MIBI与细胞共同孵育2 h后收集细胞,测定放射性活度和P-gp表达水平。(2)维拉帕米(10μmol/ L)加入细胞培养基中,与细胞孵育若干时间后99Tcm-MIBI加入细胞培养基中,与细胞一起培养2 h后收集细胞。测定放射性活度和P-gp表达水平。结果(1)维拉帕米作用2h后,细胞摄取99Tcm-MIBI较对照组显著增加(t=2.33,P<0.05),但P-gp表达水平差异无显著性(P> 0.05)。(2)肿瘤细胞摄取99Tcm-MIBI随维拉帕米作用时间的延长而增加,各时间点UR差异有显著性(F=58.2,P<0.05),并且, VRP作用时间在80 min内时,肿瘤细胞摄取99Tcm-MIBI与P-gp的表达水平无相关性(r=0.16,P>0.05),VRP作用时间大于8 h 时,肿瘤细胞摄取99Tcm-MIBI与P-gp的表达水平呈负相关(r=-0.73,P<0.01)。结论化疗增敏剂能够影响P-gp过度表达所致MDR肿瘤细胞对99Tcm-MIBI的摄取。     

泰索帝治疗晚期乳腺癌的临床观察

目的：观察泰索帝在治疗晚期乳腺癌中的临床疗效及毒性反应。方法：根据各个病例不同既往化疗史选择不同治疗方案。单用：泰索帝120mg，每1天静脉滴注1小时。顺铂40mg/d，第1天到3天静脉滴注。用泰索帝前24小时开始口服地塞米松7．5mg,2次／日，连服3天。化疗前半小时应用枢丹8mg静注或格拉司琼3mg静注。21天为1周期，治疗3周期评价疗效。结果：泰索帝治疗20例（共62疗程），19例可评价疗效。无完全缓解（CR），部分缓解（PR）4例，无变化（NR）13例和进展（PD）2例，总有效率（CR＋PR）为21％。骨髓抑制为主要毒性反应，20例中有18例至少发生一次Ⅲ度和Ⅳ度的白细胞减少并伴有发热，2例Ⅲ度血小板减少。未发现对红细胞的影响。非血液系统的毒性反应为19例Ⅲ度脱发。5例Ⅲ度／Ⅳ度口腔炎。2例Ⅱ度腹泻。2例双手指甲变色脱落。1例手足麻木。全部病例于应用泰索帝后有面部潮红，但无1例发生严重的过敏反应。大多数病例于应用泰索帝后有肌肉关节疼痛。未发现Ⅱ度以上的恶心呕吐。未发现与治疗有关的肝功能及心电图异常。结论 泰索帝在治疗晚期乳腺癌中具有较好的疗效。主要的毒性反应是白细胞减少伴发热。     

Overexpression of Bcl-xL in Human Breast Cancer Cells Enhances Organ-Selective Lymph Node Metastasis

Lymph node metastasis are the first prognostic factor in breast cancer diagnosis and an early event in metastatic spread. To assess the role of anti-apoptotic proteins in lymph node metastatic progression of human breast cancer cells we analyzed the metastatic activity of MDA-MB-435 cells transfected with the Bcl-xL gene, after orthotopic inoculation in Nude Balb/c and in SCID mice. The luciferase gene was introduced by permanent transfection in the 435/Bcl-xL and 435/Neo cells and used as a tumor marker to measure the number of tumor cells lodged in lymph nodes. We found that 435/Bcl-xL tumor cells had enhanced organ-specific metastatic activity, preferentially lodging in peripheral lymph nodes, where at 45 days post-implantation we found 7 x 10(6) +/- 6 x 10(6) 435/Bcl-xL.luc and 2 +/- 1.1 435/Neo.luc luciferase tagged tumor cell equivalents (TCEs). Metastases were abrogated in mice in which orthotopic tumors were induced with 435/Bcl-xL-antisense cells. Additionally, in vitro experiments show that in 435 cells Bcl-xL-antisense can override the emergence of resistance to apoptosis induced by TNF- alpha and TGF- beta in cells overexpressing Bcl-xL, increasing also adhesion to extracellular matrix proteins. These results point to the relevance of Bcl-xL overexpression inducing lymph node metastasis of breast cancer cells, and to the value of this gene as a target for therapy in order to prevent metastasis.     

乳腺癌脑转移68例临床特点及预后因素分析

[目的]分析乳腺癌脑转移患者的临床特点,探讨影响乳腺癌脑转移患者预后的因素。[方法]收集68例乳腺癌脑转移患者临床资料,采用单因素和多因素生存分析影响乳腺癌脑转移患者预后因素。[结果]乳腺癌脑转移患者多为未绝经、浸润性导管癌,首发症状多为颅内高压症状,多为多发颅内转移,多合并颅外转移。乳腺癌脑转移患者中位生存期8.82个月,1、2、3年生存率分别为39%、8%、8%。单因素分析显示KPS评分、合并肝转移、全身化疗、脑部放疗显著性影响乳腺癌脑转移患者的生存期;Cox多因素分析显示脑部放疗、KPS评分是影响乳腺癌脑转移生存的独立预后因素。[结论]脑部放疗、KPS评分是影响乳腺癌脑转移患者生存的独立预后因素。乳腺癌脑转移后的全身化疗价值需进一步探讨。     

微小RNA在乳腺癌转移及预后评估中的作用

微小RNA（miRNA）是近年来发现的一类内源性的、进化上高度保守的非编码单链小RNA,长度约为19~25个核苷酸,主要通过特异性识别并结合靶基因mRNA,抑制其翻译或者直接降解靶基因,在转录后水平调控基因表达。近来研究表明,miRNA在乳腺癌的侵袭和转移中发挥重要的作用,并有望成为乳腺癌预后评估的新型的潜在分子标志物。     

卡培他滨治疗蒽环类耐药的转移性乳腺癌

目的 研究卡倍他滨治疗既往蒽环类药物治疗失败的转移性乳腺癌患者的疗效和安全性。方法 17例具有可测量病灶的转移性乳腺癌患者接受卡培他滨（2510mg^2/天，口服14天，休息7天为一疗程）单药治疗2-6个疗程。结果 17例患者中CR1例，PR4例，SD4例和PD8例，有效率为29.3%。至疾病进展中位时间5个月（范围2-10^ 个月）。最常见的不良反应为手足综合征（64.7%)，皮肤色素沉着（58.8%)、白细胞减少（41.2%）和恶心呕吐消化道反应（17.6%），一般程度较轻，可耐受，Ⅲ度毒性反应为1例手足综合征，无IV度不良反应。应用Vitb6治疗能减轻手足综合征的症状。结论 卡培他滨治疗蒽环类药物治疗失败的转移性乳腺癌患者疗效好，不良反应少，且口服方便，对于晚期乳腺癌患者门诊治疗是一理想选择。     

Bcl-2,Bax在乳腺癌中表达的临床意义

目的　探讨Bcl 2 ,Bax在乳腺癌中表达的临床意义。方法　免疫组化链霉菌素 生物素 (S P)法对术前未使用过化疗、放疗及免疫治疗的乳腺癌 82例 (其中有腋窝淋巴结转移的 61例 ) ,15例乳腺良性肿瘤和 15例正常乳腺组织作对照 ,检测Bcl 2 ,Bax的表达情况。结果　Bcl 2在癌组织中的表达 ( 5 6 10 % )与正常乳腺组织 ( 3 3 3 3 % )和良性肿瘤 ( 4 0 0 0 % )中的表达无统计学差异 ,在不同种类、不同分级的浸润性导管癌中无明显差异 (P >0 0 5 ) ,在有转移 ( 4 7 5 4% )和无转移组 ( 80 95 % )中有明显差异 (P <0 0 1)。Bax在癌组织中的 ( 60 98% )表达与正常乳腺组织 ( 73 3 3 % )和良性肿瘤中 ( 60 0 0 % )的表达无统计学差异 ,在不同种类癌中无明显差异 (P>0 0 5 ) ,在不同分级的浸润性导管癌中有显著差异 (P <0 0 5 ) ,在有转移 ( 67 2 1% )和无转移组 ( 4 2 86% )中有显著差异 (P <0 0 5 )。两者在乳腺癌中的表达有相关 (P <0 0 5 )。结论　检测乳腺癌Bcl 2 ,Bax基因的表达情况 ,有助于预测乳腺癌患者的淋巴结转移情况及预后     

Role of HER2 in Brain Metastasis of Breast Cancer: a Systematic Review and Meta-Analysis

Background: Breast cancer is one of the most common cancers among women worldwide and the HER2receptor plays an important role in its development and progression. This systematic review aimed to summarizethe role of HER2 in brain metastasis in patients with breast cancer. Materials and Methods: We conducted aliterature search by advanced search in title field using the Scopus, Pubmed, and Google scholar databasesuntil the end of June 2014. With metastasis, metastatic, HER2, brain, and breast cancer, as terms of search weselected 31 articles, which were reviewed by two independent and blinded expert reviewers. The studies werefirst selected according to their titles and abstracts. Quality of the studies were then assessed using the STROBE(Strengthening the Reporting of Observational Studies in Epidemiology) protocol for observational studiesand CONSORT(Consolidation of Standards for Reporting Trials) protocol for clinical trials. For statisticalanalyses, we used STATA, version 11.0 software. Forest and funnel diagrams were drawn and for heterogeneity,index was also considered. Also we used meta regression analysis. Results: Finally, we reviewed 10 studies. Theprevalence of brain metastasis in HER2- positive breast cancer patients was 24.9%. There was publication biasin the reviewed studies. Meta regression analysis showed that follow up time had no significant effect (p=0.396)on the prevalence of brain metastasis. Conclusions: The results showed a high prevalence of brain metastasis inHER2 positive breast cancer patients.     

乳腺癌细胞对八种抗癌药物的化疗敏感性

背景与目的:本文研究了不同个体乳腺癌细胞对八种抗癌药物的化疗敏感性。材料与方法:根据临床血浆高峰浓度(1PPC),41例乳腺癌细胞受8种抗癌药物分别作用48 h,采用MTT法的肿瘤细胞抑制率分析了化疗敏感性。结果:在临床血浆高峰浓度(1 PPC)和细胞数为1×105/孔的条件下,乳腺癌细胞的平均抑制率(％)为表柔芘星(EADM)44.30±13.40和米托蒽醌(MTZ)44.14±13.46,其化疗敏感性比其它药物较高,但无统计学意义。8种抗癌药物在肿瘤临床分期和病理分型、淋巴结转移时的化疗敏感性差异有显著性。结论:41例乳腺癌细胞对8种抗癌药物有不同的化疗敏感性。体外MTT法对不同个体乳腺癌进行化疗敏感性预测是必要的。