沉默CXCR4的表达逆转肺癌细胞顺铂耐药

目的:探讨沉默CXCR4对肺癌细胞顺铂耐药的影响,并研究其分子作用机制。方法:利用RT-qPCR测定肺癌耐药（A549/DDP）及药物敏感细胞株（A549）中CXCR4 mRNA的表达水平;利用LipofectamineTM 2000将siRNA-CXCR4转染至肺癌耐药细胞株中,RT-qPCR及蛋白质印迹法（Western blot）验证siRNA对CXCR4基因的靶向沉默效率;利用CCK-8法检测沉默CXCR4后肺癌耐药细胞的增殖活性;利用流式细胞仪测定沉默CXCR4后肺癌耐药细胞的凋亡率;利用MTT法测定沉默CXCR4后肺癌耐药细胞对顺铂的敏感性;利用Western blot实验测定沉默CXCR4后CYP1B1蛋白的表达水平。结果:RT-qPCR实验结果显示,肺癌耐药细胞株A549/DDP中CXCR4 mRNA的表达量显著高于药物敏感细胞株（P＜0.01）;体外转染siRNA-CXCR4可明显下调A549/DDP细胞株中CXCR4的表达（P＜0.001）;CCK-8实验结果显示,沉默CXCR4的表达抑制了A549/DDP细胞的增殖活性（P＜0.01）;流式细胞仪实验结果显示,沉默CXCR4的表达促进了A549/DDP细胞凋亡（P＜0.01）;MTT实验结果显示,沉默CXCR4的表达增加了A549/DDP细胞对顺铂的敏感性（P＜0.01）;Western blot实验结果显示,沉默CXCR4后CYP1B1蛋白的表达水平显著降低（P＜0.05）。结论:沉默CXCR4的表达抑制肺癌耐药细胞增殖、促进凋亡,逆转肺癌细胞顺铂耐药,CXCR4正向调控CYP1B1的表达,可能是CXCR4逆转肺癌细胞顺铂耐药的作用机制。     

吉西他滨联合顺铂一线治疗晚期triple-negative乳腺癌41例临床研究

目的：观察吉西他滨联合顺铂（GP）一线治疗triple-negative（ER、PR、HER-2均阴性）晚期转移性乳腺癌的疾病进展时间、疗效和安全性。方法：2008年1月-2010年4月共41例经免疫组化检查证实为tri-ple-negative的晚期乳腺癌初治患者参与研究。患者接受吉西他滨联合顺铂方案治疗：吉西他滨1000mg/m2,静脉滴注30min,d1、d8;顺铂25mg/m2静脉滴注d1-3天,21天重复。结果：全组41例共完成160个周期的治疗,中位数4个周期,范围2-6个周期,均可评价疗效。完全缓解（CR）2例（4.88%）,部分缓解（PR）21例（51.22%）,病情稳定（SD）7例（17.07%）,病情进展（PD）11例（26.83%）。临床总缓解率（CR＋PR）56.10%;疾病控制率（CR＋PR＋SD）73.17%;中位疾病进展时间（mTTP）8.4个月,1年生存率65.85%。不良反应主要为Ⅰ-Ⅱ度骨髓抑制、末梢神经毒性、胃肠道反应、轻度肝功能损伤等。结论：吉西他滨联合顺铂一线治疗triple-negative晚期转移性乳腺癌患者,初步观察疗效较好,不良反应可耐受,值得进一步研究。     

吉西他滨联合顺铂一线治疗晚期triple-negative乳腺癌41例临床研究

目的:观察吉西他滨联合顺铂(GP)一线治疗triple-negative(ER、PR、HER-2均阴性)晚期转移性乳腺癌的疾病进展时间、疗效和安全性。方法:2008年1月-2010年4月共41例经免疫组化检查证实为tri-ple-negative的晚期乳腺癌初治患者参与研究。患者接受吉西他滨联合顺铂方案治疗:吉西他滨1000mg/m2,静脉滴注30min,d1、d8;顺铂25mg/m2静脉滴注d1-3天,21天重复。结果:全组41例共完成160个周期的治疗,中位数4个周期,范围2-6个周期,均可评价疗效。完全缓解(CR)2例(4.88%),部分缓解(PR)21例(51.22%),病情稳定(SD)7例(17.07%),病情进展(PD)11例(26.83%)。临床总缓解率(CR+PR)56.10%;疾病控制率(CR+PR+SD)73.17%;中位疾病进展时间(mTTP)8.4个月,1年生存率65.85%。不良反应主要为Ⅰ-Ⅱ度骨髓抑制、末梢神经毒性、胃肠道反应、轻度肝功能损伤等。结论:吉西他滨联合顺铂一线治疗triple-negative晚期转移性乳腺癌患者,初步观察疗效较好,不良反应可耐受,值得进一步研究。     

Down-Regulation of CXCR4 Expression by siRNA Inhibits Invasive Ability of Breast Cancer Cells

OBJECTIVE To investigate the efficiency of gene silencing by CXCR4- siRNAs (small interfering RNA), and to examine the invasive ability and the expression of other metastatic-associated genes in siRNA-treated breast cancer cells. METHODS Three siRNAs were designed and cloned into the pSilenc ? 3.1-H1 neo vector. The reconstructed plasmids were purified and trans- fected into the T47D breast cancer cell line, which highly expressed CXCR4. The amount of CXCR4 expression in the transfected cells was measured by flow cytometry and Real-time PCR. Cell invasive ability was evaluated us- ing 24-well Matrigel invasion chambers. In addition, the expression of other metastatic-associated genes, such as E-cad, IGFBP-5, FN and MMP-2, was assessed by Real-time PCR. RESULTS The suppression rates of CXCR4 mRNA expression reached 95.7%, 85.9% and 98.3%compared with control-siRNA cells in the 3 CXCR4- siRNA T47D cells respectively. FCM assays for CXCR4 protein expression showed a similar inhibitory effect. The invasion indexes of these CXCR4- siRNA cells were 0.037, 0.290 and 0.188 respectively compared with control- siRNA cells. After treatment of the cells with CXCR4-siRNA, the expression of E-cad showed an upward tendency and that of IGFBP-5 had a downward trend, while alteration in expression of FN and MMP2 varied without a con- sistant effect. CONCLUSION CXCR4 plays an important role in modulating migra- tion of human breast cancer cells. Small interfering RNA can significantly silence the CXCR4 gene in the human T47D breast cancer cell line. The results of this study strengthen the need for further research on novel gene therapy against breast cancer metastasis.     

The Role of chemokine receptor CXCR4 in breast cancer metastasis

Breast cancer is one of the leading causes of cancer related deaths worldwide. Breast cancer-related mortality is associated with the development of metastatic potential of primary tumor lesions. The chemokine receptor CXCR4 has been found to be a prognostic marker in various types of cancer, including breast cancer. Recent advances in the field of cancer biology has pointed to the critical role that CXCR4 receptor and its ligand CXCL12 play in the metastasis of various types of cancer, including breast cancer. Breast tumors preferentially metastasize to the lung, bones and lymph nodes, all of which represent organs that secrete high levels of CXCL12. CXCL12 acts as a chemoattractant that drives CXCR4-positive primary tumor cells towards secondary metastatic sites leading to the onset of metastatic lesions. Since its discovery in 2001, the CXCR4 field has progressed at a very fast rate and further studies have pointed to the role of CXCR4 in dissemination of tumor cells from primary sites, transendothelial migration of tumor cells as well as the trafficking and homing of cancer stem cells. This review summarizes the information that has been obtained over the years regarding the role of CXCL12-CXCR4 signaling in breast cancer, discusses its potential application to the development of new therapeutic tools for breast cancer control, and elucidates the potential therapeutic challenges which lie ahead and the future directions that this field can take for the improvement of prognosis in breast cancer patients.     

CIP4 promotes metastasis in triple-negative breast cancer and is associated with poor patient prognosis

Signaling via epidermal growth factor receptor (EGFR) and Src kinase pathways promote triple-negative breast cancer (TNBC) cell invasion and tumor metastasis. Here, we address the role of Cdc42-interacting protein-4 (CIP4) in TNBC metastasis in vivo, and profile CIP4 expression in human breast cancer patients. In human TNBC cells, CIP4 knock-down (KD) led to less sustained activation of Erk kinase and impaired cell motility compared to control cells. This correlated with significant defects in 3D invasion of surrounding extracellular matrix by CIP4 KD TNBC cells when grown as spheroid colonies. In mammary orthotopic xenograft assays using both human TNBC cells (MDA-MB-231, HCC 1806) and rat MTLn3 cells, CIP4 silencing had no overt effect on tumor growth, but significantly reduced the incidence of lung metastases in each tumor model. In human invasive breast cancers, high CIP4 levels was significantly associated with high tumor stage, TNBC and HER2 subtypes, and risk of progression to metastatic disease. Together, these results implicate CIP4 in promoting metastasis in TNBCs.     

Tocilizumab potentiates cisplatin cytotoxicity and targets cancer stem cells in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is a very aggressive subtype with high recurrence rate and no molecular targets for therapies. This subtype is characterized by high expression/secretion of the proinvasive/metastatic interleukin-6 (IL-6) cytokine. In the present study, we have shown that tocilizumab inhibits the IL-6/STAT3 signaling and suppresses the cancer/inflammatory epigenetic IL-6/STAT3/NF-κB positive feedback loop. Furthermore, tocilizumab inhibited the proliferative and the migratory/invasiveness capacities as well as the epithelial-to-mesenchymal transition (EMT) process in TNBC cells. Importantly, tocilizumab suppressed the stemness-related characteristics of TNBC cells, through the inhibition of the Wnt/β-catenin breast cancer stem cell-related pathway. Additionally, we have shown that tocilizumab suppresses the paracrine activation of normal breast stromal fibroblasts to myofibroblats. Moreover, tocilizumab sensitized TNBC cells to the cytotoxic effect of cisplatin in vitro. Furthermore, pharmacological inhibition of IL-6 by tocilizumab had great inhibitory effect on tumor growth and the EMT process in humanized orthotopic breast tumors in mice. In addition, tocilizumab potentiated the proapoptotic effect of cisplatin in humanized breast tumors. Together, these findings indicate that tocilizumab can suppress the prometastatic capacity of TNBC cells and enhances the cytotoxic effect of cisplatin against these cells. Therefore, tocilizumab could be of great therapeutic value for these hard-to-treat TNBC patients.     

IAPs抑制剂APG1387联合EGFR抑制剂对三阴乳腺癌细胞的抑制作用及其机制

目的:探讨IAPs抑制剂APG1387联合EGFR抑制剂对三阴乳腺癌MDA-MB-231细胞的体外抑制作用及其可能的机制。方法:MTT法检测APG1387和EGFR抑制剂单药或联合用药对MDA-MB-231细胞的增殖抑制;用流式细胞技术检测细胞凋亡;Western blotting检测APG1387和EGFR抑制剂单药或联合用药对MDA-MB-231细胞凋亡通路蛋白的影响。结果:APG1387单药对MDA-MB-231细胞有一定的增殖抑制作用,EGFR抑制剂对MDA-MB-231细胞几乎没有影响,两药联用较单药组有明显的增殖抑制作用和促凋亡作用(P<0.01)。联合用药显著抑制了p-ERK1/2的水平,同时激活了Caspase-8及其下游的凋亡通路。结论:IAPs抑制剂APG1387和EGFR抑制剂联合应用在体外对乳腺癌MDA-MB-231细胞有明显的增殖抑制作用及促凋亡作用,其机制可能与联合用药降低p-ERK1/2水平以及激活凋亡通路有关。     

CXCL12，CXCR4及CXCR7表达检测对乳腺癌患者疾病预后意义探讨

目的：探讨乳腺癌患者肿瘤组织中CXCL12,CXCR4和CXCR7 mRNA表达情况在肿瘤转移和疾病预后中的价值。方法采用定量PCR方法检测115例乳腺癌,临近正常组织及乳腺癌肿瘤转移患者颈部淋巴结样本中CXCL12,CXCR4和CXCR7 mRNA表达情况。随访资料采用Kaplan-Meier生存分析,对影响生存质量的因素进行多重变量Cox回归分析。结果与正常组织相比,乳腺癌组织中CXCR4和CXCR7表达明显增加,差异均有统计学意义（P﹤0.001）,两种组织中CXCL12的表达差异无统计学意义（P﹥0.05）;CXCL12在肿瘤原发部位和淋巴结转移部位的表达差异有统计学意义（P﹤0.05）,转移瘤的CXCR4和CXCR7表达均增加（P﹤0.05）。Kaplan-Meier生存分析结果表明,与CXCR4和CXCR7低表达患者相比,高表达患者的总生存率较低（P﹤0.05）。Cox回归模型显示,CXCL12、CXCR4和CXCR7表达均为影响乳腺癌患者生存情况的独立因素。结论本研究结果表明CXCL12、CXCR4和CXCR7 mRNA表达在乳腺癌患者肿瘤发展和转移中发挥重要作用,可以作为乳腺癌患者疾病预后的生物标志物。     

三阴性乳腺癌患者的临床特征与预后分析

Objective  

We investigated the clinical characteristics, and the prognostic factors of triple-negative breast cancer.     

Significance of E-cadherin expression in triple-negative breast cancer

Purpose:

Triple-negative breast cancer (TNBC), a subtype of breast cancer that is oestrogen receptor (ER) negative, progesterone receptor (PR) negative, and human epidermal growth factor receptor 2 (HER2) negative, has a poor prognosis. Although a correlation between E-cadherin expression level and outcome has been demonstrated among all types of breast cancer, little is known about the significance of E-cadherin expression levels in TNBC.

Methods:

A total of 574 patients who had undergone a resection of a primary breast cancer except for invasive lobular carcinomas were enrolled in this study. Expressions of ER, PR, HER2, and E-cadherin were assessed by immunohistochemistry. We examined the association between TNBC and other clinicopathological variables and evaluated the significance of the E-cadherin expression.

Results:

Among the 574 breast cancer cases, 123 (21.4%) revealed a triple-negative phenotype. Patients with TNBC experienced more frequent lymph node metastasis (P=0.024) and a poorer prognosis (P<0.001) in comparison with non-TNBC patients. Triple-negative breast cancer was an independent prognostic factor. Reduced levels of E-cadherin were observed in 238 (41.5%) of the 574 breast cancer cases. E-cadherin reduction was significantly frequent in cases of TNBC (P<0.001) and lymph node metastasis (P=0.032). Furthermore, in the 123 TNBC cases, the prognosis of patients with an E-cadherin-negative expression was significantly worse than that of E-cadherin-positive patients (P=0.0265), especially for those in clinical stage II (P=0.002). A multivariate logistic regression analysis showed a reduction of the E-cadherin expression to be an independent prognostic factor (P=0.046).

Conclusion:

E-cadherin expression may be a useful prognostic marker for classifying subgroups of TNBC.     

晚期三阴乳腺癌挽救治疗进展

三阴乳腺癌约占全部乳腺癌病例的15％,具有恶性程度高,进展快,生存期短和预后较差的临床特点.因此近些年来,三阴乳腺癌受到越来越多的国际肿瘤学界的关注.化疗是晚期三阴乳腺癌治疗的核心,但目前没有统一标准的化疗方案.另外,靶向治疗是晚期三阴乳腺癌一个较有前途的治疗新策略.现对晚期三阴乳腺癌的挽救治疗做一综述.     

CXCL12- CXCR4在乳腺浸润性导管癌中的表达及临床意义

目的：研究乳腺浸润性导管癌中趋化因子 CXCL12及其对应的特异趋化因子受体 CXCR4的表达,分析其与浸润性导管癌临床病理学特征的关系。方法：用免疫组化法研究200例乳腺浸润性导管癌中 CX-CL12、CXCR4的表达情况,并探讨二者单一表达或共表达与临床病理因素的相关性。结果：CXCL12在40%（80/200）浸润性乳腺癌中阳性表达,其与 TNM 分期和肿瘤大小相关（P <0.05）;CXCR4在48%（96/200）浸润性乳腺癌中阳性表达,其表达与浸润性乳腺癌的 TNM 分期相关（P <0.05）。CXCL12与 CXCR4共表达于29%（58/200）的浸润性乳腺癌,二者的共表达与 TNM 分期和淋巴结转移情况相关（ P <0.01）。结论：CX-CL12与 CXCR4共表达很可能是协同乳腺浸润性导管癌进展及淋巴结转移的重要因素,检测 CXCL12与 CX-CR4共表达比检测 CXCL12或 CXCR4单一分子标记更有意义。     

三阴性乳腺癌精准治疗的机遇

三阴性乳腺癌（triple-negative breast cancer ,TNBC）是一类有着极大异质性的疾病,个体间生物学行为差异较大,根据基因表达谱分析,TNBC 细分为生物学行为各异的6 个类别,各类别对不同治疗表现出不同的敏感性。在医疗行业步入精准的时代,寻找TNBC 可治疗的各类靶点对不同类别TNBC 的精准靶向治疗可能具有指导作用。     

三阴乳腺癌的研究进展(英文)

Triple-negative breast cancer (TNBC) is a unique subgroup defined by a lack expression of ER (estrogen receptor), PR(progesterone receptor) and HER2 (human epidermal growth factor receptor 2), which has distinctly biological, clinical and pathological characteristics. This subgroup has close relationship with basal-like and BRCA1 (breast cancer susceptibility gene-1) breast cancers. Since endocrine and HER2-targered therapy can not be applied, chemotherapy is the major mean of therapy. Some studies show tha...     

Advantages of adjuvant chemotherapy for patients with triple-negative breast cancer at Stage II: usefulness of prognostic markers E-cadherin and Ki67

Introduction  

Triple-negative breast cancer (TNBC), which is characterized by negativity for estrogen receptor, progesterone receptor and human epidermal growth factor receptor 2 (HER2), is a high risk breast cancer that lacks specific targets for treatment selection. Chemotherapy is, therefore, the primary systemic modality used in the treatment of this disease, but reliable parameters to predict the chemosensitivity of TNBC have not been clinically available.     

Clinical efficacy of local targeted chemotherapy for triple-negative breast cancer

Background

The aim of the study was to evaluate the clinical efficacy of superselective intra-arterial targeted neo-adjuvant chemotherapy in the treatment of estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and human epidermal growth factor receptor 2 (HER2)-negative (triple-negative) breast cancer.

Patients and methods.

A total of 47 triple-negative breast cancer patients (29 at stage II, 13 at stage III and 5 at stage IV) were randomly assigned to two groups: targeted chemotherapy group (n=24) and control group (n=23). Patients in the targeted chemotherapy group received preoperative superselective intra-arterial chemotherapy with CEF regimen (C: cyclophosphamide [600 mg/m²]; E: epirubicin [90 mg/m²]; F: 5-fluorouracil [600 mg/m²]), and those in the control group received routine neoadjuvant chemotherapy with CEF. The duration of the treatment, changes in lesions and the prognosis were determined.

Results

The average course of the treatment was 15 days in the targeted chemotherapy group which was significantly shorter than that in the control group (31 days) (P<0.01). The remission rate of lesions was 91.6% in the targeted chemotherapy group and 60.9% in the control group, respectively. Among these patients, 9 died within two years, including 2 (both at IV stage) in the targeted chemotherapy group and 7 (2 at stage II, 4 at stage III and 1 at stage IV) in the control group.

Conclusions

As an neoadjuvant therapy, the superselective intra-arterial chemotherapy is effective for triple-negative breast cancer, with advantages of the short treatment course and favourable remission rates as well as prognoses.     

Inhibition of Notch1 reverses EMT and chemoresistance to cisplatin via direct downregulation of MCAM in triple-negative breast cancer cells

Resistance to chemotherapy continues to be a critical issue in the clinical therapy of triple-negative breast cancer (TNBC). Epithelial–mesenchymal transition (EMT) is thought to contribute to chemoresistance in several cancer types, including breast cancer. Identification of the key signaling pathway that regulates the EMT program and contributes to chemoresistance in TNBC will provide a novel strategy to overcome chemoresistance in this subtype of cancer. Herein, we demonstrate that Notch1 positively associates with melanoma cell adhesion molecule (MCAM), a unique EMT activator, in TNBC tissue samples both at mRNA and protein levels. High expression of Notch1 and MCAM both predicts a poor survival in basal-like/TNBC patients, particularly in those treated with chemotherapy. The expression of Notch1 and MCAM in MDA-MB-231 cells gradually increases in a time-dependent manner when exposing to low dose cisplatin. Moreover, the expressions of Notch1 and MCAM in cisplatin-resistant MDA-MB-231 cells are significantly higher than wild-type counterparts. Notch1 promotes EMT and chemoresistance, as well as invasion and proliferation of TNBC cells via direct activating MCAM promoter. Inhibition of Notch1 significantly downregulates MCAM expression, resulting in the reversion of EMT and chemoresistance to cisplatin in TNBC cells. Our study reveals the regulatory mechanism of the Notch1 pathway and MCAM in TNBC and suggesting that targeting the Notch1/MCAM axis, in conjunction with conventional chemotherapies, might be a potential avenue to enhance the therapeutic efficacy for patients with TNBC.     

CXCR4和Survivin启动子的克隆及其在乳腺癌细胞系的转录特异性分析

目的:评估CXCR4和Survivin启动子在乳腺癌细胞系中的特异转录活性.方法:采用PCR方法扩增CXCR4和Survivin启动子序列,将其分别克隆到舍有报告基因增强绿色荧光蛋白(EGFP)的质粒载体pEGFP-1和含有荧光素酶(LUC)的质粒载体pGL3-Basic,经脂质体分别转染乳腺癌细胞系(MCF-7MDA-MB-231和T-47D)和乳腺上皮细胞(HBL-100).荧光显微镜下观察EGFP的表达情况,并测定2个启动子在乳腺癌细胞中的荧光素酶活性.结果:CXCR4和Sur-vivin启动子在3种乳腺癌细胞中均表现特异的转录活性.CXCR4启动子在MDA-MB-231中的活性高于Survivin启动子(P=0.004);而在MCF-7细胞中活性低于Survivin启动子(P=0.006),两者在T-47D细胞中的启动子活性相当,P=0.615.结论:CXCR4和Survivin启动子在乳腺癌细胞中具有强的特异转录活性,且在不同乳腺癌细胞类型表现的转录特异性有一定差异.