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.     

Role of tumor microenvironment in triple-negative breast cancer and its prognostic significance

Breast cancer has been shown to live in the tumor microenvironment,which consists of not only breast cancer cells themselves but also a significant amount of pathophysiologically altered surrounding stroma and cells.Diverse components of the breast cancer microenvironment,such as suppressive immune cells,re-programmed fibroblast cells,altered extracellular matrix (ECM) and certain soluble factors,synergistically impede an effective anti-tumor response and promote breast cancer progression and metastasis.Among these components,stromal cells in the breast cancer microenvironment are characterized by molecular alterations and aberrant signaling pathways,whereas the ECM features biochemical and biomechanical changes.However,triple-negative breast cancer (TNBC),the most aggressive subtype of this disease that lacks effective therapies available for other subtypes,is considered to feature a unique microenvironment distinct from that of other subtypes,especially compared to Luminal A subtype.Because these changes are now considered to significantly impact breast cancer development and progression,these unique alterations may serve as promising prognostic factors of clinical outcome or potential therapeutic targets for the treatment of TNBC.In this review,we focus on the composition of the TNBC microenvironment,concomitant distinct biological alteration,specific interplay between various cell types and TNBC cells,and the prognostic implications of these findings.     

POR overexpression induces tamoxifen-resistance in breast cancer through the STAT1/c-Myc pathway

Breast cancer is the most common cancer in women worldwide. Although tamoxifen (TAM), a selective estrogen receptor (ER) modulator, is widely used to treat ER-positive breast cancers, resistance to TAM remains a major clinical problem. NADPH-dependent cytochrome P450 reductase (POR) is known to participate in drug metabolism and steroid metabolism. Recent studies showed that high POR expression was correlated with poor outcomes in triple-negative breast cancer (TNBC), and POR might be a prognostic biomarker in TNBC. However, the role of POR in TAM resistance is still elusive. In this study, we found that high POR expression was associated with poor prognosis of ER-positive and TAM-treated breast cancer patients. In addition, COX analysis showed that POR expression was an independent prognostic biomarker for ER-positive as well as TAM-treated breast cancer patients. Furthermore, our results suggested that POR overexpression promoted TAM resistance by activating the STAT1/c-Myc pathway in ER-positive breast cancer cells. Immunohistochemical analysis showed that high POR/STAT1 expression was correlated with poor prognosis in TAM-treated breast cancer patients. Notably, combined treatment with TAM and a specific STAT1 inhibitor Fludarabine was more effective for inhibiting TAM-resistant breast cancer cells. Altogether, our findings suggested that POR overexpression induced TAM resistance through STAT1/c-Myc pathway and might serve as an independent prognostic biomarker in TAM-treated breast cancer patients. Combining TAM and STAT1 inhibitors might be an effective strategy for treating POR-induced TAM-resistant breast cancer.     

NuSAP1通过Wnt/β-catenin/EMT通路促进三阴性乳腺癌细胞增殖和侵袭及其对预后的预测价值

目的:分析核仁纺锤体相关蛋白(NuSAP1)的表达对乳腺癌（BC）患者预后的影响，探讨三阴性乳腺癌(TNBC)细胞株的致瘤机制。方法:我们从癌症基因组图谱(TCGA)下载了乳腺癌(BC)的RNA-seq数据，用R软件在此数据中筛选出了NuSAP1基因及BC患者的临床资料。生存曲线用Kaplan-Meier-plotter绘制。建立阴性对照组（NC）、敲减NuSAP1组（sh-NuSAP1）、过表达NuSAP1组(sh-NuSAP1+ov-NuSAP1),用细胞增殖毒性检测试剂盒(CCK8)和Transwell法检测各组细胞增殖和侵袭能力。实时定量聚合酶链反应（real-time quantitative polymerase chain reaction,qRT-PCR）和免疫印迹法（Western blotting）检测肿瘤细胞中NuSAP1、Wnt/β-catenin通路和上皮-间充质转化(epithelial-mesenchymal transition,EMT)相关蛋白的表达。结果:NuSAP1在BC中表达上调。NuSAP1表达差异与多种临床病理因素有关，且NuSAP1表达越高，生存期越短。在MDA-MB-231和BT-549细胞中，与NC组相比，sh-NuSAP1组细胞增殖和侵袭能力下降，CyclinD1、Vimentin、Slug、Twist、Wnt3a和p-β-catenin的表达减少，而E-cadherin的表达显著增加。与sh-NuSAP1组相比，sh-NuSAP1+ov-NuSAP1组结果与上述相反。结论:NuSAP1是一种致癌基因，预测乳腺癌的不良预后，并通过Wnt/β-catenin和上皮-间充质转化（EMT）途径促进肿瘤进展。     

Therapeutic effect of β-blockers in triple-negative breast cancer postmenopausal women

Beta-blockers (BB) drugs have been used for decades worldwide, mainly to treat hypertension. However, in recent epidemiological studies, BBs were suggested to improve cancer prognosis. In the wake of this evidence, we evaluated the possible therapeutic effect of BBs in triple-negative breast cancer (TNBC) patients. We identified 800 postmenopausal women operated between 1997 and 2008 for early primary TNBC. The effect of BB intake on the risk of breast cancer (BC) recurrence and death was evaluated through competing risk and Cox regression survival models. At cancer diagnosis, 74 (9.3 %) women out of 800 were BBs users. Median age was 62 years in BB users and 59 years in non-users (P = 0.02). BB users and non-users were similarly distributed by all tumor characteristics. The 5-year cumulative incidence of BC-related events was 13.6 % in BB users and 27.9 % in non-users (P = 0.02). The beneficial impact of BBs remained statistically significant at multivariable analysis (HR, 0.52; 95 % CI 0.28–0.97), after the adjustment for age, tumor stage, and treatment, peritumoral vascular invasion and use of other antihypertensive drugs, antithrombotics, and statins. Adjusted HRs for metastases and for BC deaths were 0.32 (95 % CI 0.12–0.90) and 0.42 (95 % CI 0.18–0.97), respectively, in favor of BBs. Hypertension, other antihypertensive drugs, antithrombotics, and statins did not impact prognosis. In this series of postmenopausal TNBC patients, BB intake was associated with a significantly decreased risk of BC-related recurrence, metastasis, and BC death. Innovative therapeutic strategies including BBs should be urgently explored in cancer patients.     

环状RNA ciRS-7在三阴性乳腺癌中的表达及其对细胞侵袭和迁移的影响

 目的 探讨环状RNA ciRS-7在三阴性乳腺癌（TNBC）中的表达及其对细胞侵袭和迁移的影响。方法 通过qRT-PCR法检测乳腺癌组织和细胞中ciRS-7的表达水平,分析其与乳腺癌患者临床病理指标之间的关系。用划痕实验和Transwell实验检测沉默ciRS-7后MDA-MB-231和BT-549细胞的迁移和侵袭能力的变化。利用动物实验在体内进行验证。结果 TNBC组织中ciRS-7的相对表达量为（6.52±0.38）,显著高于Luminal型（1.56±0.17）（P<0.001）和HER2过表达型乳腺癌组织（2.27±0.66）（P<0.001）以及癌旁正常组织（0.83±0.09）（P<0.001）。ciRS-7表达与分子分型、肿瘤浸润和淋巴结转移明显相关（均P<0.05）;而与患者年龄、肿瘤直径和病理分级无关（均P>0.05）。沉默ciRS-7后,MDA-MB-231和BT-549细胞的迁移和侵袭明显减弱（均P<0.05）。ciRS-7敲低组肺转移瘤数明显减少（P<0.05）。结论 ciRS-7在TNBC中高表达,并可能增强TNBC细胞的侵袭和迁移。     

Breast cancer neoplastic seeding in the setting of image-guided needle biopsies of the breast

Purpose

Cathepsin S (CTSS) is expressed in a variety of cancers and stimulates tumor progression. However, the regulatory mechanism and role of CTSS in breast cancer progression are poorly understood. The aim of this study was to examine the relationships between CTSS expression and breast cancer grade and stage, and the signaling molecules involved in CTSS expression.

Methods

Immunohistochemical staining was performed in tissue microarray sections of 1451 human invasive breast cancer samples to determine epithelial (E-CTSS) and stromal CTSS (S-CTSS) expression. Gene and protein expression levels in human breast cancer cell lines were measured by polymerase chain reaction and western blotting. Small interfering RNA transfection and a Matrigel transwell invasion assay were used to confirm the signaling pathways regulating CTSS expression.

Results

In patient tumor tissue blocks, high grade, late stage, and triple negativity were associated with elevated CTSS protein expression, and expression levels were related to the clinical outcomes of patients with invasive breast cancer. CTSS expression was also higher in triple-negative breast cancer (TNBC) cell lines than in hormone-responsive cells, and CTSS expression patterns matched those of tryptophan hydroxylase 1 (TPH1) and 5-hydroxytryptamine receptor 7 (5-HT₇). Treatment of TNBC cells (MDA-MB-231 and HCC-1395) with 5-HT significantly enhanced CTSS protein expression, whereas pharmacological inhibition or knockdown of 5-HT₇ significantly inhibited its expression. Correspondingly, cancer cell invasion was increased by 5-HT treatment and suppressed by 5-HT₇ knockdown. The expression of CTSS was regulated by PI3K/Akt and Ras/Raf/MAPK signaling pathways, and these signaling pathways were stabilized by HSP90 and enhanced by the 5-HT₇ receptor-dependent autocrine effect of 5-HT in TNBC cells.

Conclusion

Our findings suggest CTSS as a candidate target for development of a strategy to inhibit breast cancer invasion, and indicate that HSP90 and 5-HT₇ (regulators of CTSS) should be considered as alternative targets for the management of TNBC invasion and metastasis.

     

Role of epidermal growth factor receptor in breast cancer

Decades of research in molecular oncology have brought about promising new therapies which are designed to target specific molecules which promote tumor growth and survival. The epidermal growth factor receptor (EGFR) is one of the first identified important targets of these novel antitumor agents. Approximately half of cases of triple-negative breast cancer (TNBC) and inflammatory breast cancer (IBC) overexpress EGFR. Thus, EGFR inhibitors for treatment of breast cancer have been evaluated in several studies. However, results so far have been disappointing. One of the reasons for these unexpected results is the lack of biomarkers for predicting which patients are most likely to respond to EGFR inhibitors. Recent studies have shown that EGFR and its downstream pathway regulate epithelial-mesenchymal transition, migration, and tumor invasion and that high EGFR expression is an independent predictor of poor prognosis in IBC. Further, recent studies have shown that targeting EGFR enhances the chemosensitivity of TNBC cells by rewiring apoptotic signaling networks in TNBC. These studies indicate that EGFR-targeted therapy might have a promising role in TNBC and IBC. Further studies of the role of EGFR in TNBC and IBC are needed to better understand the best way to use EGFR-targeted therapy??e.g., as a chemosensitizer or to prevent metastases??to treat these aggressive diseases.     

Cell clusters overlying focally disrupted mammary myoepithelial cell layers and adjacent cells within the same duct display different immunohistochemical and genetic features: implications for tumor progression and invasion

Introduction

Our previous studies detected focal disruptions in myoepithelial cell layers of several ducts with carcinoma in situ. The cell cluster overlying each of the myoepithelial disruptions showed a marked reduction in or a total loss of immunoreactivity for the estrogen receptor (ER). This is in contrast to the adjacent cells within the same duct, which were strongly immunoreactive for the ER. The current study attempts to confirm and expand previous observations on a larger scale.

Methods

Paraffin sections from 220 patients with ER-positive intraductal breast tumors were double immunostained with the same protocol previously used. Cross-sections of ducts lined by ≥ 40 epithelial cells were examined for myoepithelial cell layer disruptions and for ER expression. In five selected cases, ER-negative cells overlying the disrupted myoepithelial cell layer and adjacent ER-positive cells within the same duct were separately microdissected and assessed for loss of heterozygosity and microsatellite instability.

Results

Of the 220 cases with 5698 duct cross-sections examined, 94 showed disrupted myoepithelial cell layers with 405 focal disruptions. Of the 94 cases, 79 (84%) contained only ER-negative cell clusters, nine (9.6%) contained both ER-negative and ER-positive cell clusters, and six (6.4%) contained only ER-positive cell clusters overlying disrupted myoepithelial cell layers. Of the 405 disruptions, 350 (86.4%) were overlain by ER-negative cell clusters and 55 (13.6%) were overlain by ER-positive cell clusters (P < 0.01). Microdissected ER-negative and ER-positive cells within the same duct from all five selected cases displayed a different frequency or pattern of loss of heterozygosity and/or microsatellite instability at 10 of the 15 DNA markers.

Conclusions

Cells overlying focally disrupted myoepithelial layers and their adjacent counterparts within the same duct displayed different immunohistochemical and molecular features. These features potentially represent an early sign of the formation of a biologically more aggressive cell clone and the myoepithelial cell layer breakdown possibly associated with tumor progression or invasion.     

The clinical relevance of unfolded protein response signaling in breast cancer

Protein homeostasis regulated by the Endoplasmic Reticulum (ER) is a recognized process involved in cancer progression. ER stress activates the Unfolded Protein Response (UPR) and has been implicated in a variety of cancers. Given the role of the UPR activation in carcinogenesis, we hypothesized that UPR activation could be associated with pathological progression, higher clinical stage, and worse survival in breast cancer. A total of 4,416 breast cancer patients from multiple independent cohorts were analyzed. We defined the UPR pathway score by the degree of enrichment by Gene Set Variant Analysis and median was used to divide high vs. low score groups in each cohort. High UPR breast cancer significantly enriched not only cell proliferation-related but also other pro-cancerous gene sets consistently in both METABIC and {"type":"entrez-geo","attrs":{"text":"GSE96058","term_id":"96058"}}GSE96058 cohort. Majority of UPR pathway score high cells in the bulk tumor were tumor cells compared to other cells, including stromal, T-, B-, and myeloid-cells (P<0.001). UPR score was significantly associated with advanced stage, high grade, and triple negative breast cancer (TNBC) (all P<0.001). High UPR breast cancer was associated with worse patient survival in both cohorts (all P<0.001). Among breast cancer subtype, ER-positive/HER2-negative breast cancer with high UPR was significantly associated with worse survival, but neither HER-positive nor TNBC. High UPR ER-positive/HER2-negative breast cancer was infiltrated with high level of Th1 and Th2 cells, M1 macrophage, and plasma cells. On the other hand, they were significantly infiltrated with high level of several types of stromal cells in tumor microenvironment (all P<0.001). Finally, high UPR metastatic breast cancer was also associated with worse patient survival (P=0.041). UPR signaling is associated with cancer aggressiveness, and worse survival, especially ER-positive/HER2-negative breast cancer subtype.     

The Effects of Low HER2/neu Expression on the Clinicopathological Characteristics of Triple-Negative Breast Cancer Patients

Background: Triple-negative breast cancer (TNBC) is an aggressive type of breast cancer (BC), and its diagnosis is associated with negative expression of hormone receptors and HER2/neu. It consists of 10-20% of all BCs diagnosed. Methods and materials: This study focuses on three groups with different pathology: group one showed complete triple-negative HER2 expression with IHC of BC; groups two and three included patients with ER-, PR-, and HER21+, and ER-, PR-, and HER22+ with a negative FISH test. These three groups were compared from the point of prognosis, which consisted of tumor size, patients’ age, lymphatic, vascular and perineural invasion, organ metastasis, number of lymph nodes involvement, and the survival rate. Results: A total of 459 TNBC patients were enrolled, of which 268 were placed in the HER20 group, 146 in the HER21+ group, and 45 in the HER22+ group. Distant metastasis and recurrence rate were more common in HER20 patients, but bone metastasis was more common in patients with low HER2 expression. All patients with HER20 had a smaller tumor size at the time of BC diagnosis in comparison to patients in the low HER2 expression group. Patients with HER22+ had less lymphatic and vascular invasion as well as axillary lymph nodes involvement, but larger tumor size at presentation, resulting in a lower rate of recurrence and higher overall survival. Conclusion: The findings revealed that patients with HER22+ had better outcome in comparison to the patients with HER20 and HER21+. Furthermore, the results showed that many patients with HER22+ expression were not basal-like and had good prognosis amongst TNBC patients.     

三阴性乳腺癌的临床病理特点及预后分析研究

目的:探讨三阴性乳腺癌患者临床病理特点、生存率以及影响预后的因素。方法:回顾性分析30例三阴性乳腺癌患者的资料,分析其临床病理特点、免疫组化指标和各种治疗方式对生存率的影响。结果:全组30例患者均为女性,占同期收治乳腺癌患者的5.7%,中位年龄50岁(39-76岁)。其中,浸润性导管癌21例(70.0%),单纯癌4例(13.3%),髓样癌2例(6.7%),管状腺癌2例(6.7%),导管内癌1例(3.3%)。Ⅰ期患者5例(16.7%),Ⅱ期15例(50.0%),Ⅲ期8例(26.7%),Ⅳ期2例(6.7%)。核分级1级4例(13.3%),2级11例(36.7%),3级15例(50.0%)。伴有脉管瘤栓10例(33.3%),伴有神经浸润5例(16.7%)。中位生存时间3.8年(1.6-7.5年)。单因素分析结果显示,影响总生存的因素为肿瘤大小、淋巴结状态、临床分期、脉管瘤栓和神经浸润。多因素分析结果显示,肿瘤大小和淋巴结状态是影响预后的独立因素。3年总生存率65.7%。结论:三阴性乳腺癌发病率较低,组织学分级较高,多为浸润性导管癌,易较早出现复发和转移。影响生存率的因素为肿瘤大小、淋巴结状态、临床分期、脉管瘤栓和神经浸润,其中影响预后的独立因素是肿瘤大小和淋巴结状态。     

Unique sphingolipid signature identifies luminal and triple-negative breast cancer subtypes

Breast cancer (luminal and triple-negative breast cancer [TNBC]) is the most common cancer among women in India and worldwide. Altered sphingolipid levels have emerged as a common phenomenon during cancer progression. However, these alterations are yet to be translated into robust diagnostic and prognostic markers for cancer. Here, we present the quantified sphingolipids of tumor and adjacent-normal tissues from patients of luminal (n = 70) and TNBC (n = 42) subtype from an Indian cohort using targeted liquid chromatography mass spectrometry. We recorded unique sphingolipid profiles that distinguished luminal and TNBC tumors in comparison to adjacent normal tissue by six-sphingolipid signatures. Moreover, systematic comparison of the profiles of luminal and TNBC tumors provided a unique five-sphingolipid signature distinguishing the two subtypes. We further identified key sphingolipids that can stratify grade II and grade III tumors of luminal and TNBC subtype as well as their lymphovascular invasion status. Therefore, we provide the right evidence to develop these candidate sphingolipids as widely acceptable marker/s capable of diagnosing luminal vs TNBC subtype of breast cancer, and predicting the disease severity by identifying the tumor grade.     

MAPK信号激活抑制三阴性乳腺癌细胞的迁移和侵袭

背景与目的：三阴性乳腺癌具有高复发和转移风险,除了化疗,临床上无特定的靶向治疗。因此,研究三阴性乳腺癌复发转移机制对提高患者生存率具有重要意义,本研究旨在探讨丝裂原激活的蛋白激酶(MAPK)信号激活对三阴性乳腺癌细胞迁移和侵袭的影响。方法：首先用细胞划痕和细胞小室(Transwell)试验分析和比较肺高转移三阴性乳腺癌细胞系231-HM及其父代肺低转移细胞系231-p的体外迁移和侵袭性差异;然后用蛋白质印迹法(Western blot)检测转移相关蛋白和MAPK分子激活状态;最后用MAPK抑制剂处理231-p细胞,测定MAPK抑制情况下的细胞迁移、侵袭和相关蛋白变化。结果：与231-p细胞相比,231-HM细胞迁移和侵袭性明显增强;Western blot检测发现,231-HM细胞中促细胞迁移和侵袭蛋白Caveolin-1和β-catenin升高,MAPK通路相关蛋白P38、Erk1/2和MEK的磷酸化水平明显降低;用P38/MAPK磷酸化抑制剂(SB202190)处理231-p细胞后发现,其细胞迁移和侵袭性明显增强,Caveolin-1和β-catenin表达水平上升。结论：MAPK信号激活抑制三阴性乳腺癌的迁移和侵袭。     

年轻三阴性乳腺癌患者的临床病理特征及预后分析

目的 分析年轻三阴性乳腺癌(TNBC)患者的临床病理特征和预后因素.方法 1999年1月至2007年12月间,中国医学科学院肿瘤医院收治的年龄≤35 岁的年轻TNBC患者94例,所有患者肿瘤组织中雌激素受体(ER)、孕激素受体(PR)和人表皮生长因子受体2(Her-2)均为阴性,收集患者的临床病理资料和生存情况,分析其临床病理特点和预后因素.结果 全组94例年轻TNBC患者,占同期收治年轻乳腺癌患者的12.0%.主要病理类型为浸润性导管癌,共81例,占86.2%.分期为Ⅰ、Ⅱ、Ⅲ和Ⅳ期的患者分别有17例(18.1%)、48例(51.1%)、28例(29.8%)和1例(1.1%).14例(14.9%)患者存在脉管瘤栓.全组94例患者的1、3、5和7年无病生存率(DFS)分别为88.3%、66.9%、59.7%和59.7%,1、3、5和7年总生存率(OS)分别为98.9%、85.6%、72.9%和69.6%.单因素预后分析结果显示,肿瘤大小、腋淋巴结状态、临床分期以及有无脉管瘤栓对年轻TNBC患者的5年OS具有显著影响(均P<0.05).多因素预后分析结果显示,有无脉管瘤栓是影响年轻TNBC患者预后的独立因素(P<0.05).在随访期出现复发和(或)转移的33 例患者中,29例发生于术后3年内,其余4例发生于术后3～4年间.结论 年轻TNBC 患者具有独特的临床、病理和预后特点,需要探索更为合理的个体化治疗方案.     

三阴性乳腺癌的临床病理特点及预后分析研究

目的：探讨三阴性乳腺癌患者临床病理特点、生存率以及影响预后的因素。方法：回顾性分析30例三阴性乳腺癌患者的资料,分析其临床病理特点、免疫组化指标和各种治疗方式对生存率的影响。结果：全组30例患者均为女性,占同期收治乳腺癌患者的5.7%,中位年龄50岁（39-76岁）。其中,浸润性导管癌21例（70.0%）,单纯癌4例（13.3%）,髓样癌2例（6.7%）,管状腺癌2例（6.7%）,导管内癌1例（3.3%）。Ⅰ期患者5例（16.7%）,Ⅱ期15例（50.0%）,Ⅲ期8例（26.7%）,Ⅳ期2例（6.7%）。核分级1级4例（13.3%）,2级11例（36.7%）,3级15例（50.0%）。伴有脉管瘤栓10例（33.3%）,伴有神经浸润5例（16.7%）。中位生存时间3.8年（1.6-7.5年）。单因素分析结果显示,影响总生存的因素为肿瘤大小、淋巴结状态、临床分期、脉管瘤栓和神经浸润。多因素分析结果显示,肿瘤大小和淋巴结状态是影响预后的独立因素。3年总生存率65.7%。结论：三阴性乳腺癌发病率较低,组织学分级较高,多为浸润性导管癌,易较早出现复发和转移。影响生存率的因素为肿瘤大小、淋巴结状态、临床分期、脉管瘤栓和神经浸润,其中影响预后的独立因素是肿瘤大小和淋巴结状态。