Characterization of breast precancerous lesions and myoepithelial hyperplasia in sclerosing adenosis with apocrine metaplasia

The identification as well as the molecular characterization of breast precancerous lesions in terms of increased risk of progression and/or recurrence is becoming a critical issue today as improved non‐surgical procedures are detecting cancer at an earlier stage. The strategy we have been pursuing to identify early apocrine breast lesions is based on the postulate that invasive apocrine carcinomas evolve from epithelial cells in terminal duct lobular units (TDLUs) in a stepwise manner that involves apocrine metaplasia of normal breast epithelia, hyperplasia, atypia, and apocrine carcinoma in situ. First, we identify specific protein biomarkers for benign apocrine metaplasia and thereafter we search for biomarkers that are highly overexpressed by pure invasive apocrine carcinomas. Here we present studies in which we have used antibodies against components of a benign apocrine signature that includes 15‐prostaglandin dehydrogenase (15‐PGDH), a protein that is expressed by all benign apocrine lesions, and markers that are highly overexpressed by pure invasive apocrine carcinomas such as MRP14 (S100A9), psoriasin (S100A7), and p53 to identify precancerous lesions in sclerosing adenosis (SA) with apocrine metaplasia. The latter is a benign proliferative lesion of the breast that exhibits an increase in the size of the TDLUs and characterized by retained two‐cell lining, and myoepithelial (ME) and stromal hyperplasia. SA with apocrine metaplasia, i.e. apocrine adenosis (AA), presents with a higher degree of atypical apocrine hyperplasia, and these lesions are believed to be precursors of apocrine carcinoma, in situ and invasive. Analysis of 24 selected SA samples with apocrine metaplasia revealed non‐obligate putative apocrine precancerous lesions that displayed some, or in same cases all the three markers associated with pure invasive apocrine carcinomas. These studies also revealed p53 positive, non‐apocrine putative precancerous lesions as well as novel phenotypes for ME and some luminal cells characterized by the expression of cytokeratin 15.     

Gene expression profiling of human lymph node metastases and matched primary breast carcinomas: Clinical implications

The genetic program that drives tumor metastasis and the mode and timing of its initiation are of great practical significance to clinical management. Modern technical advances open new opportunities for gaining useful relevant information. Gene expression profiles of histologically‐verified viable tissue from lymph node metastases were compared with those of matched primary breast cancers from 10 different patients, among samples from over 400 cases, using high‐throughput oligonucleotide arrays comprising probes for 22,000 genes. It was observed that metastases have very similar expression signatures to their parent tumors. However, detailed computational analysis revealed that a small number of genes were consistently differentially expressed between 100% of tumors and metastases, suggesting that these are mechanistically important. Lists of such candidate genes, of potential clinical interest, are provided. We interpret these results in the framework of a meta‐analysis of previous investigations by others and ourselves and of existing clinical knowledge on the behavior of human tumors. The collective data show that metastases resemble their primary tumors but the signatures obtained in different studies are not sufficiently reproducible or informative to be prognostically useful, although they do give valuable insights into the pathogenesis and biology of human tumor metastasis. The findings indicate that the genetic program encoding metastasis is implemented progressively over time although, occasionally, this evolution can occur rapidly, early in the life of the neoplasm. The important clinical significance of this deduction is that, in most patients, early detection provides time for appropriate therapeutic intervention to be effective in obstructing metastasis.     

Stromal mast cells in invasive breast cancer are a marker of favourable prognosis: a study of 4,444 cases

Purpose We have previously demonstrated in a pilot study of 348 invasive breast cancers that mast cell (MC) infiltrates within primary breast cancers are associated with a good prognosis. Our aim was to verify this finding in a larger cohort of invasive breast cancer patients and examine the relationship between the presence of MCs and other clinical and pathological features. Experimental design Clinically annotated tissue microarrays (TMAs) containing 4,444 cases were constructed and stained with c-Kit (CD-117) using standard immunoperoxidase techniques to identify and quantify MCs. For statistical analysis, we applied a split-sample validation technique. Breast cancer specific survival was analyzed by Kaplan–Meier [KM] method and log rank test was used to compare survival curves. Results Survival analysis by KM method showed that the presence of stromal MCs was a favourable prognostic factor in the training set (P = 0.001), and the validation set group (P = 0.006). X-tile plot generated to define the optimal number of MCs showed that the presence of any number of stromal MCs predicted good prognosis. Multivariate analysis showed that the MC effect in the training set (Hazard ratio [HR] = 0.804, 95% Confidence interval [CI], 0.653–0.991, P = 0.041) and validation set analysis (HR = 0.846, 95% CI, 0.683–1.049, P = 0.128) was independent of age, tumor grade, tumor size, lymph node, ER and Her2 status. Conclusions This study concludes that stromal MC infiltration in invasive breast cancer is an independent good prognostic marker and reiterates the critical role of local inflammatory responses in breast cancer progression.     

乳腺癌组织中COX-2与MDR1/P-gp表达的相关性研究

目的:探讨环氧化酶-2(COX-2)蛋白与P糖蛋白(P-gp)在乳腺癌组织中表达的相关性。方法:应用免疫组织化学染色法,检测32例乳腺癌组织中COX-2蛋白与P-gp蛋白的表达情况。结果:32例乳腺癌组织中COX-2表达阳性率为62·5%(20/32),P-gp表达阳性率为46·9%(15/32),两者表达呈正相关性(r=0·598,P<0·05)。结论:乳腺癌组织中COX-2与P-gp表达呈正相关,COX-2可干预P-gp的表达并参与乳腺癌多药耐药(MDR)的调节。     

An association between invasive breast cancer and familial idiopathic hyperparathyroidism: a case series and review of the literature

The possibility of an association between primary hyperparathyroidism and breast cancer has been postulated. We report here a sibship with three premenopausal breast cancers and hyperparathyroidism, and no detectable BRCA or MEN1 gene mutations. We explore genetic and molecular rationales for an association between these metabolic and neoplastic processes.     

Relationship between mammographic density and the risk of breast cancer in japanese women: a case-control study

BACKGROUND: The relationship between mammographic density and the risk of breast cancer was examined in Japanese women. The study was a matched case-control study comparing the mammographic densities of both breast cancer cases and healthy controls. MATERIALS AND METHODS: We selected 237 women who were diagnosed with a histologically verified breast cancer, and who underwent surgery at Gihoku General Hospital in Gifu, from January, 1998 to December, 1999. During the time of this study, 3,650 people participated in breast cancer screening with mammography and ultrasound together. We selected 742 women as a control group from the screening participants and matched them by age and the number of deliveries with the cancer patients. The same mammography machine was used for both cases and controls. For evaluation, we used a visual method (Wolfe's classification) and a computer assisted method to classify the mammograms based on mammographic density. RESULTS: (1) According to Wolfe's classification, the DY group had a significantly increased breast cancer risk compared with the N1 group (Relative risk (RR)=2.20, 95% confidence interval (95%CI) (1.02-4.77). (2) The group showing a high mammographic density had a significantly increased risk of breast cancer compared with the group with low mammographic density (RR=2.83, 95%CI=1.33-5.98) as classified by the computer assisted method. CONCLUSION: It is suggested that women with high mammographic densities, classified visually or by computer, have an elevated risk of breast cancer compared with those with low mammographic densities.     

Expression of the stromelysin-3 gene in fibroblastic cells of invasive carcinomas of the breast and other human tissues: a review

Summary Stromelysin-3 (ST3) is a putative new matrix metalloproteinase (MMP) which may play a role in the progression of human carcinomas, and exhibits unique structural and functional characteristics among the MMP family. The ST3 gene, which is generally not expressed at significant levels in benign breast tumors, has been found to be expressed in all invasive breast carcinomas tested so far. The gene is also expressed in somein situ breast carcinomas, which have a higher probability to become invasive. ST3 RNA and protein are specifically found in fibroblastic cells immediately surrounding the neoplastic cells, both in invasive andin situ breast carcinomas. The same expression pattern is observed in other types of human carcinomas, and the highest ST3 RNA levels are observed in tumors that exhibit high local invasiveness. The ST3 gene is also expressed in fibroblastic cells during the inflammatory phase of wound healing, which suggests that ST3 gene expression in stromal fibroblasts may be under the control of factors produced by inflammatory cells during wound healing, and by cancer cells during carcinoma progression. ST3 may thus represent a stroma-derived factor necessary for the progression of epithelial malignancies, and its manipulation may possibly be used to develop new anti-cancer agents.     

Retracted: Glabridin attenuates the migratory and invasive capacity of breast cancer cells by activating microRNA‐200c

Current treatments for breast cancer, a common malignancy in human females, are less than satisfactory because of high rates of metastasis. Glabridin (GLA), which acts through the FAK/ROS signaling pathway, has been used as an antioxidant and anti‐metastatic agent. However, little is known regarding the effect of microRNA (miRNA) on GLA's anti‐metastatic activity. The miRNA‐200 family, which is frequently expressed at low levels in triple negative breast cancers, inhibits metastasis by blocking the epithelial–mesenchymal transition. Here, we found that GLA attenuated the migratory and invasive capacity of breast cancer cells by activating miR‐200c. GLA induced the mesenchymal–epithelial transition in vitro and in vivo, as determined by increased expression of the epithelial marker, E‐cadherin, and decreased expression of the mesenchymal marker, vimentin. Overexpression of miR‐200c enhanced the expression of E‐cadherin and decreased the expression of vimentin. Furthermore, in MDA‐MB‐231 and BT‐549 breast cancer cells exposed to GLA, knockdown of miR‐200c blocked the GLA‐induced mesenchymal–epithelial transition and alleviated the GLA‐induced inhibition of migration and invasion. Thus, elevation of miR‐200c by GLA has considerable therapeutic potential for anti‐metastatic therapy for breast cancer patients.     

内分泌及化疗对ER+乳腺癌CD44+CD24-/low细胞亚群的实验与临床研究

目的:探讨内分泌治疗及化疗对雌激素受体(estrogen receptor,ER)阳性的乳腺癌CD44+CD24-/low细胞亚群的影响.方法:用3个浓度的雌二醇(estradiol,E2)和E2+他莫西芬(tamoxifen,TAM)处理乳腺癌MCF-7细胞,用工作浓度的化疗药物(多西他赛、紫杉醇、表柔比星和5-氟尿嘧啶)处理MCF-7细胞,FCM法检测各组细胞中CD44+CD24-/low的细胞比例,并检测ER+的转移性乳腺癌(metastatic breast cancer,MBC)患者化疗前、后外周静脉血中CD45-CD44+CD24-/low的细胞比例.结果:不同浓度E2培养MCF-7细胞10和20 d后,CD44+ CD24-/low细胞比例均高于对照组,E2+TAM组低于相应浓度E2组.各化疗药物作用MCF-7细胞24 h后,CD44+CD24-/low细胞比例均上升;而继续培养至20 d后,CD44+ CD24-/low细胞比例降低.部分缓解和疾病稳定的MBC患者,化疗后外周血CD45-CD44+CD24-/low细胞比例降低;而疾病进展者化疗后外周血CD45-CD44+CD24-/low细胞比例上升.结论:E2促进MCF-7细胞中CD44+CD24-/low细胞亚群的生长,TAM可抑制MCF-7细胞中CD44+CD24-/low细胞亚群的生长;MCF-7细胞中CD44+CD24-/low细胞亚群对多西他赛、紫杉醇、表柔比星和5-氟尿嘧啶均有抵抗能力;化疗有效患者外周血中CD45-CD44+CD24-/low细胞比例降低.     

Isolation and transplantation of highly purified autologous peripheral CD34<Superscript>+</Superscript> progenitor cells: purging efficacy,hematopoietic reconstitution following high dose chemotherapy in patients with breast cancer: results of a feasibility study in Japan

BACKGROUND: High-dose chemotherapy with autologous stem cell support may have some therapeutic impact on certain groups of the patients with advanced breast cancer(BRCA). Since stem cell contamination by tumor cells might contribute to relapse, development of a tumor cell purging technique would improve the clinical outcome. The present study was undertaken to evaluate the purging efficacy of autologous mobilized CD34+peripheral stem cells in patients with breast cancer (BRCA) in an advanced stage or relapse. METHODS: CD34+cells were selected from autologous peripheral blood stem cells (PBSC) using a clinical scale of magnetic-activated cell sorting system (CliniMACS), followed by high-dose chemotherapy with transplantation of CD34+ selected cells. Amplification of cytokeratin 19 (CK19) and 20 (CK20) gene in leukapheresis products were measured to evaluate the performance of tumor cell elimination. RESULTS: Seven patients were entered into this study. After leukopheresis, 1 patient was dropped form this study due to poor mobilization. Among 6 patient, a total of 8 CD34+ selection was performed. The median purity and recovery rate of the CD34+ cells post selection was 85.1% (range 62.5-98.1%) and 74.2% (range 50.2-90.2%), respectively. After isolation of CD34+cells, the elimination rate in the logarithmic transformation of CK19 was 2.77 log, and that of CK20 were 2.43 log and 2.53 log. In 4 patients, high-dose chemotherapy was performed, followed by the transplantation of the isolated CD34+cells. Rapid neutrophil recovery, as well as platelet recovery was seen with a median time to reach 0.5 x 109/l neutrophils of 9 days(range 8-9), and 20 x 109/l platelets of 12 days (range 10-13). There was no treatment related death and no serious adverse events directly associated with the selection procedure or infusion of selected cells. CONCLUSIONS: The present study demonstrated that the CliniMACS system is a highly effective positive selection method and that a high purging efficacy could be obtained without compromising the hematopoietic reconstitution capacity of the graft in BRCA patients undergoing high-dose chemotherapy.     

hsa-miRNA27a和hsa-miRNA451通过调控MDRl/P-gp的表达和功能参与卵巢癌和乳腺癌细胞耐药

背景与目的：肿瘤细胞耐药是临床化疗失败的主要原因,微小RNA(microRNA,miRNA)在肿瘤细胞中的异常表达与耐药关系密切。本研究旨在探讨卵巢癌及乳腺癌细胞中hsa-miRNA27a和hsa-miRNA451的表达差异及其与耐药的关系。方法：用浓度递增法建立卵巢癌耐紫杉醇细胞系A2780/Taxol;颈环状引物实时定量聚合酶链反应(stem-loop quantitative real-time PCR,stem-loop RT-PCR)检测卵巢癌耐紫杉醇细胞A2780/Taxol和亲本细胞A2780以及乳腺癌耐阿霉素细胞MCF-7/ADM和亲本细胞MCF-7中hsa-miRNA27a和hsa-miRNA451的表达;利用Lipofectamine^TM 2000分别将成熟miRNA27a的模拟物、阻遏物及阴性对照(negative control,NC)RNA转染A2780和A2780/Taxol细胞,将成熟miRNA451的模拟物及NC转染MCF-7/ADM细胞;RT-PCR技术检测细胞MDR1 mRNA表达;蛋白[质]印迹法(Western blot)检测细胞中P-糖蛋白(P-glycoprotein,P-gp)的表达;采用四甲基偶氮唑蓝(MTT)法检测细胞增殖情况。结果：miRNA27a在A2780/Taxol细胞中高表达,与A2780细胞相比,表达增高2.2±0.30倍,差异有统计学意义(P<0.05);miRNA451在MCF-7/ADM细胞中低表达,与MCF-7细胞相比,表达降低84%,差异有统计学意义(P<0.05)。A2780/Taxol细胞转染miRNA27a阻遏物后,MDR1 mRNA表达明显下降,与转染NC组相比,表达下降(39±0.14)%,差异有统计学意义(P<0.05)。P-gp相对表达量［(26±5.3)%)］与转染NC组的P-gp相对表达量［(43±6.7)%］比较,下降39%,差异有统计学意义(P<0.05)。对紫杉醇的敏感性增加,半数抑制浓度(IC₅₀)为0.53 μmol/L,与转染NC组IC50(6.8 μmol/L)相比,差异有统计学意义(P<0.05)。A2780细胞转染miRNA27a模拟物后,细胞的MDR1 mRNA表达升高,与转染NC组相比,升高(121±0.11)%,差异有统计学意义(P<0.05);细胞对紫杉醇的敏感性下降,IC₅₀为0.2 μmol/L,与转染NC组IC₅₀(0.06 μmol/L)相比,差异有统计学意义(P<0.05)。MCF-7/ADM细胞转染miRNA451模拟物后,MDR1 mRNA表达明显下降,与转染NC组细胞相比,表达下降(65±12)%,差异有统计学意义(P<0.05);P-gp相对表达量［(31±19)%)］与转染NC组细胞P-gp相对表达量［(83±12)%］相比,下降62%,差异有统计学意义(P<0.05);对阿霉素的敏感性增加,IC₅₀为4.61 μmol/L,与转染NC组细胞IC₅₀(26 μmol/L)相比,差异有统计学意义(P<0.05)。结论：在卵巢癌耐紫杉醇细胞A2780/Taxol和乳腺癌耐阿霉素细胞MCF-7/ADM中,miRNA27a和miRNA451分别异常表达,它们可能分别通过间接或直接作用于MDR1/P-gp,参与肿瘤细胞耐药的发生、发展。     

Quercetin shortened survival of radio-resistant B-1 cells in vitro and in vivo by restoring miR15a/16 expression

Chronic lymphocytic leukemia (CLL) is a malignancy disease characterized by the expansion of CD5⁺ B-1 cells. The NZB mouse model of CLL shows similarities to human CLL, has age-associated increase in malignant B-1 clones and decreased expression of miR-15a/16. It was demonstrated that systemic lentiviral delivery of miR-15a/16 ameliorates disease manifestations in this mouse model. Nowadays, new therapeutic approaches have been focus on miRNA in cancer cells. Natural compounds like quercetin can modulate these miRNAs, consequently, suppress oncogenes or stimulate tumor suppressor genes by altering miRNA expressions. Here we investigate the effects of quercetin on miRNA15a/16 expression by radio-resistant B-1 cells. It has been described that a small percentage of B-1 cell survives to irradiation in vitro, and these cells show similarities to B-CLL cells. In these cells, the level of miR15a/16 is diminished and Bcl-2 is overexpressed. Quercetin treatment restore both, miR15a/16 and Bcl-2, to normal levels. Furthermore, transference of radioresistant B-1 cells to NOD/SCID mice causes an expansion of this population and also a migration to the liver. However, after quercetin treatment, even radioresistant B-1 cells are not able to expand or disseminate in vivo, and the levels of miR15a/16 and Bcl-2 are also normalized. Our data support the hypothesis that quercetin is an important adjuvant molecule that acts on miRNA15a/16 level and leads cells more permissive to apoptosis. This work could help to design new approaches to therapy in CLL patients.     

Relationship between staining by digital subtraction angiography and vascularity in breast cancer: Analysis of the time-density curve and the results of staining for factor VIII-related antigen/von Willebrand factor

Summary We performed intravenous digital subtraction angiography (IV-DSA) in 31 patients with primary invasive breast cancer, and calculated the maximum density of tumor staining from the time-density curve obtained. Specimens resected from the same patients were stained for factor VIII-related antigen/von Willebrand factor (vWF), and the amount of microvessels was calculated by image processing. A significant correlation (correlation coefficient: 0.85) was found between the maximum density of tumor staining and the vascularity determined by staining for vWF, indicating that the maximum density indirectly reflects the vascularity in the tumor.