Cationic liposome codelivering PI3K pathway regulator improves the response of BRCA1-deficient breast cancer cells to PARP1 inhibition

Although some progresses have been made in breast cancer therapy, effective treatment for BRCA1-deficient breast cancer remains to be a great challenge. It has been demonstrated that the PI3K pathway is inappropriately activated in BRCA1-deficient breast cancers which can be downregulated by microRNA 451 (miR-451). In addition, although PARP1 inhibitors showed relatively positive results in both preclinical and clinical studies, additional efforts to decrease drug resistance as well as reduce systematic toxicity need to be addressed. To this end, by encapsulating the miR-451 mimic and PARP1 inhibitor in the same cationic liposome, we examined the potential of enhancing the response of PARP1 inhibition on BRCA1-deficient breast cancer by regulating the PI3K pathway. Our results revealed that in BRCA1-deficient human breast cancer cell line, PARP1 inhibition resulted in DNA damage with viability decrease, G2/M arrest as well as apoptosis. In contrast, single PI3K inhibition induced G1 arrest along with retarded cell proliferation. However, it was noted that combination of PARP inhibitor and PI3K regulator could exert synergetic function to evidently decrease cell proliferation compared with PARP inhibition alone, which was also confirmed by in vivo antitumor assay using xenograft tumor models. Collectively, our results offer an alternative but superior strategy for the therapy of BRCA1-deficient human breast cancers which may benefit the clinical applications.     

Patterns of post-operative irradiation in breast cancer patients submitted to neoadjuvant chemotherapy

Background/AimPost-operative radiation therapy (PORT) is associated with improvement in loco-regional control and survival rates in early breast cancer. However, the evidence of benefit in patients after treatment with neoadjuvant chemotherapy (NAC) is poor. We aimed to assess the impact of the type of surgery in the PORT plan and the role of the PORT fields in clinical outcomes in breast cancer patients who had undergone NAC followed by surgery.Materials and methodsWe performed a retrospective analysis of all non-metastatic breast cancer patients treated between 2008 and 2014 at our institution who had received NAC and PORT.ResultsA total of 528 women were included of whom 396 were submitted to mastectomy or nipple-sparing/skin-sparing mastectomy. Most (92.8%) of the patients had locally advanced disease (clinical stage IIB to IIIC). All patients underwent irradiation for breast or chest wall. Most patients received PORT to the supraclavicular and axillary (levels II and III) nodes (87.1% and 86.4% for breast-conserving surgery and 95.1% and 93.8% for mastectomy and nipple-sparing/skin-sparing mastectomy, respectively). Irradiation of level I axillary and internal mammary nodes was uncommon. The disease-free survival and overall survival rates at 3 years were 72% and 85%, respectively. There were no statistically significant differences in clinical outcomes according to the use of nodal irradiation.ConclusionsAfter NAC, most patients received irradiation of the breast/chest wall and axillary and supraclavicular nodes. In this setting, PORT to breast/chest wall with or without regional nodal irradiation was safe and effective, with acceptable disease-free and overall survival rates reported in this high-risk population.     

巨噬细胞LSD1抑制H3K9Me2水平增强动脉粥样硬化中血清炎症因子的表达

目的:检测动脉粥样硬化患者巨噬细胞中组蛋白第9位赖氨酸的二甲基化(histone H3 lysine 9 dimethylation,H3K9Me2)的表达并探究其对炎症因子表达的影响及其机制。方法:选择厦门大学附属东南医院收治的动脉粥样硬化患者(n=20)与健康对照人群(n=22)作为研究对象,采用蛋白免疫印迹方法检测巨噬细胞中H3K9Me2的表达,并通过酶联免疫吸附的方法检测其血清炎症因子表达。随后,通过小干扰RNA(small interfering RNA,siRNA)敲减赖氨酸特异去甲基化酶1A(lysine-specific demethylase 1A,LSD1)后,在氧化低密度脂蛋白(oxidized low-density lipoproteins,ox-LDL)诱导的巨噬细胞中结合白免疫印迹和染色质免疫共沉淀的方法检测细胞整体H3K9Me2的表达及炎症因子启动子区H3K9Me2水平。同时,分析巨噬细胞中上清中炎症因子表达变化。结果:动脉粥样硬化病人巨噬细胞中H3K9Me2水平显著下调,并与血清中炎症因子表达呈负相关。在敲减LSD1后,ox-LDL诱导的巨噬细胞中H3K9Me2整体水平以及炎症因子启动子区H3K9Me2水平下调均被抑制。同时,核因子-κB(NF-κB)结合启动子区介导的炎症因子表达增强也被抑制。结论:巨噬细胞中H3K9Me2的表达与动脉粥样硬化病人血清中炎症因子的表达成负相关,而组蛋白去甲基化酶LSD1很可能在其中参与H3K9Me2水平调控,并通过影响NF-κB与启动子区结合调节炎症因子表达。     

Different effects of genistein on molecular markers related to apoptosis in two phenotypically dissimilar breast cancer cell lines

The association between consumption of genistein-containing soybean products and lower risk of breast cancer suggests a cancer chemopreventive role for genistein. Consistent with this suggestion, exposing cultured human breast cancer cells to genistein inhibits cell proliferation, although this is not completely understood. To better understand how genistein works, the ability of genistein to induce apoptosis was compared in phenotypically dissimilar MCF-7 and MDA-MB-231 human breast cancer cells that express the wild-type and mutant p53 gene, respectively. After 6 days of incubation with 50 microM genistein, MCF-7 but not MDA-MB-231 cells, showed morphological signs of apoptosis. Marginal proteolytic cleavage of poly-(ADP-ribose)-polymerase and significant DNA fragmentation were also detected in MCF-7 cells. In elucidating these findings, it was determined that after 2 days of incubation with genistein, MCF-7 but not MDA-MB-231 cells, had significantly higher levels of p53. Accordingly, the expression of certain proteins modulated by p53 was studied next. Levels of p21 increased in both of the genistein-treated cell lines, suggesting that p21 gene expression was activated but in a p53-independent manner, whereas no significant changes in levels of the pro-apoptotic protein, Bax, were found. In MCF-7 cells, levels of the anti-apoptotic protein, Bcl-2, decreased slightly at 18-24 h but then increased considerably after 48 h. Hence, the Bax:Bcl-2 ratio initially increased but later decreased. These data suggest that at the genistein concentration tested, MCF-7 cells in contrast to MDA-MB-231 cells were sensitive to the induction of apoptosis by genistein, but Bax and Bcl-2 did not play clear roles.     

Chloroquine sensitizes breast cancer cells to chemotherapy independent of autophagy

Chloroquine (CQ) is a 4-aminoquinoline drug used for the treatment of diverse diseases. It inhibits lysosomal acidification and therefore prevents autophagy by blocking autophagosome fusion and degradation. In cancer treatment, CQ is often used in combination with chemotherapeutic drugs and radiation because it has been shown to enhance the efficacy of tumor cell killing. Since CQ and its derivatives are the only inhibitors of autophagy that are available for use in the clinic, multiple ongoing clinical trials are currently using CQ or hydroxychloroquine (HCQ) for this purpose, either alone, or in combination with other anticancer drugs. Here we show that in the mouse breast cancer cell lines, 67NR and 4T1, autophagy is induced by the DNA damaging agent cisplatin or by drugs that selectively target autophagy regulation, the PtdIns3K inhibitor LY294002, and the mTOR inhibitor rapamycin. In combination with these drugs, CQ sensitized to these treatments, though this effect was more evident with LY294002 and rapamycin treatment. Surprisingly, however, in these experiments CQ sensitization occurred independent of autophagy inhibition, since sensitization was not mimicked by Atg12, Beclin 1 knockdown or bafilomycin treatment, and occurred even in the absence of Atg12. We therefore propose that although CQ might be helpful in combination with cancer therapeutic drugs, its sensitizing effects can occur independently of autophagy inhibition. Consequently, this possibility should be considered in the ongoing clinical trials where CQ or HCQ are used in the treatment of cancer, and caution is warranted when CQ treatment is used in cytotoxic assays in autophagy research.     

乳腺癌细胞肿瘤标记物表达及其与临床病理特征相关性研究

目的 探讨肿瘤标记物14-3-3σ、Akt 和p27Kip1 蛋白在乳腺癌中的表达及其与临床病理特征及Her-2 的相关性.方法 选取43 份乳腺癌石蜡标本和10 份正常乳腺组织标本,采用免疫组织化学技术(SABC)检测组织中14-3-3σ、Akt 和p27Kip1 蛋白的表达,结合临床资料和随访资料,进行回顾性研究,...     

Detection of pathological response of axillary lymph node metastasis after neoadjuvant chemotherapy in breast cancer using multiphoton microscopy

Neoadjuvant treatment is often considered in breast cancer patients with axillary lymph node involvement, but most of patients do not have a pathologic complete response to therapy. The detection of residual nodal disease has a significant impact on adjuvant therapy recommendations which may improve survival. Here, we investigate whether multiphoton microscopy (MPM) could identify the pathological changes of axillary lymphatic metastasis after neoadjuvant chemotherapy in breast cancer. And furthermore, we find that there are obvious differences in seven collagen morphological features between normal node and residual axillary disease by combining with a semi-automatic image processing method, and also find that there are significant differences in four collagen features between the effective and no-response treatment groups. These research results indicate that MPM may help estimate axillary treatment response in the neoadjuvant setting and thereby tailor more appropriate and personalized adjuvant treatments for breast cancer patients.     

Correlation between baseline 18F-FDG PET/CT features and pathological complete response after neoadjuvant chemotherapy in early triple negative breast cancer

Aim of the studyTo evaluate correlation between metabolic and textural parameters on baseline ¹⁸F-FDG PET/CT and pathological response after neoadjuvant chemotherapy in non-metastatic triple negative breast cancer (TNBC).MethodsAll consecutive non-metastatic TNBC women treated by neoadjuvant chemotherapy followed by breast surgery who underwent ¹⁸F-FDG PET/CT examination at diagnosis between 2012 and 2018 were retrospectively included. Metabolic parameters (SUVmax, SUVmean, SUVpeak, MTV, TLG) of the primary tumour and lymph nodes, and textural features (entropy, homogeneity, SRE, LRE, LGZE, HGZE) of the primary tumour were collected. Pathological response was defined according to Sataloff classification.ResultsSeventy-four patients were enrolled. In univariate analysis, metabolic and textural features of the primary breast lesion or metabolic parameters of regional lymph nodes were not predictive of pathological complete response after neoadjuvant chemotherapy.ConclusionMetabolic and textural features of baseline PET/CT do not seem to predict pathological response after neoadjuvant chemotherapy in non-metastatic triple negative breast cancer.     

Expression and clinicopathological significance of lipin-1 in human breast cancer and its association with p53 tumor suppressor gene

Breast cancer (BC) is an important cause of female cancer-related death. It has recently been demonstrated that metabolic disorders including lipid metabolism are a hallmark of cancer cells. Lipin-1 is an enzyme that displays phosphatidate phosphatase activity and regulates the rate-limiting step in the pathway of triglycerides and phospholipids synthesis. The objective of this study was to evaluate lipin-1 expression, its prognostic significance, and its correlation with p53 tumor suppressor in patients with BC. In this study, 55 pairs of fresh samples of BC and adjacent noncancerous tissue were used to analyze lipin-1, using quantitative real-time polymerase chain reaction and immunohistochemistry (IHC) staining. The expression of other clinicopathological variables and p53 was also examined using IHC technique. The cell migration was studied in MCF-7 and MDA-MB231 cells following the inhibition of lipin-1 by propranolol. Our results show that the relative expression of lipin-1 messenger RNA was significantly higher in BC tissues compared with the adjacent normal tissue and its inhibition reduced cell migration in cancer cells. This upregulation was negatively correlated with histological grade of tumor and p53 status (p = .001 and p = .034) respectively and positively correlated with the tumor size (p = .006). Our results also seem to indicate that the high lipin-1 expression is related to a good prognosis in patients with BC. The expression of lipin-1 may be considered as a novel independent prognostic factor. The inhibition of lipin-1 may also have therapeutic significance for patients with BC. The correlation between lipin-1 and p53 confirms the role of p53 in the regulation of lipid metabolism in cancer cells.     

Long non‐coding RNA SNHG14 induces trastuzumab resistance of breast cancer via regulating PABPC1 expression through H3K27 acetylation

Currently, resistance to trastuzumab, a human epidermal growth factor receptor 2 (HER2) inhibitor, has become one major obstacle for improving the clinical outcome of patients with advanced HER2⁺ breast cancer. While cell behaviour can be modulated by long non‐coding RNAs (lncRNAs), the contributions of lncRNAs in progression and trastuzumab resistance of breast cancer are largely unknown. To this end, the involvement and regulatory functions of lncRNA SNHG14 in human breast cancer were investigated. RT‐qPCR assay showed that SNHG14 was up‐regulated in breast cancer tissues and associated with trastuzumab response. Gain‐ and loss‐of‐function experiments revealed that overexpression of SNHG14 promotes cell proliferation, invasion and trastuzumab resistance, whereas knockdown of SNHG14 showed an opposite effect. PABPC1 gene was identified as a downstream target of SNHG14, and PABPC1 mediates the SNHG14‐induced oncogenic effects. More importantly, ChIP assays demonstrated that lncRNA SNHG14 may induce PABPC1 expression through modulating H3K27 acetylation in the promoter of PABPC1 gene, thus resulting in the activation of Nrf2 signalling pathway. These data suggest that lncRNA SNHG14 promotes breast cancer tumorigenesis and trastuzumab resistance through regulating PABPC1 expression through H3K27 acetylation. Therefore, SNHG14 may serve as a novel diagnostic and therapeutic target for breast cancer patients.     

Prediction of chemotherapy response in breast cancer patients at pre-treatment using second derivative texture of CT images and machine learning

Although neoadjuvant chemotherapy (NAC) is a crucial component of treatment for locally advanced breast cancer (LABC), only about 70% of patients respond to it. Effective adjustment of NAC for individual patients can significantly improve survival rates of those resistant to standard regimens. Thus, the early prediction of NAC outcome is of great importance in facilitating a personalized paradigm for breast cancer therapeutics. In this study, quantitative computed tomography (qCT) parametric imaging in conjunction with machine learning techniques were investigated to predict LABC tumor response to NAC. Textural and second derivative textural (SDT) features of CT images of 72 patients diagnosed with LABC were analysed before the initiation of NAC to quantify intra-tumor heterogeneity. These quantitative features were processed through a correlation-based feature reduction followed by a sequential feature selection with a bootstrap 0.632+ area under the receiver operating characteristic (ROC) curve (AUC0.632+) criterion. The best feature subset consisted of a combination of one textural and three SDT features. Using these features, an AdaBoost decision tree could predict the patient response with a cross-validated AUC0.632+ accuracy, sensitivity and specificity of 0.88, 85%, 88% and 75%, respectively. This study demonstrates, for the first time, that a combination of textural and SDT features of CT images can be used to predict breast cancer response NAC prior to the start of treatment which can potentially facilitate early therapy adjustments.     

8-氯腺苷诱导的组蛋白H3K79双甲基化促进NBS1和p21的基因转录激活

组蛋白H3第79位赖氨酸甲基化（H3K79me）修饰有单甲基、双甲基及三甲基3种形式,是常染色质的标志.然而,对于组蛋白H3K79三种甲基化各自在基因转录、DNA损伤修复中所起的作用尚不十分清楚.本研究以8-氯腺苷（8-Cl-Ado）为DNA双链断裂（DNA double-stranded breaks,DSB）诱导剂,采用Western 印迹,在人肺癌细胞H1299检测出了DNA修复分子NBS1、细胞周期检验点相关分子p21,并发现H3K79me1、H3K79me2和H3K79me3三种甲基化修饰的组蛋白明显增加;染色质免疫共沉淀结合实时定量PCR实验显示,只H3K79me2与DNA损伤检验点分子p21、DNA修复分子NBS1的启动子区域相结合,说明H3K79双甲基化修饰与这些基因的转录激活有关.结果提示,在8-氯腺苷引起 DSB时,是H3K79me2、而不是H3K79me1和H3K79me3参与NBS1和p21基因转录激活时的染色质重塑.8-氯腺苷诱导H3K79双甲基化增强、促进H3K79me2所在染色质区域的NBS1和p21基因转录激活可能是8-Cl-Ado抑制肿瘤细胞生长作用机制之一.     

Proteomic approach reveals FKBP4 and S100A9 as potential prediction markers of therapeutic response to neoadjuvant chemotherapy in patients with breast cancer

Although doxorubicin (Doxo) and docetaxel (Docet) in combination are widely used in treatment regimens for a broad spectrum of breast cancer patients, a major obstacle has emerged in that some patients are intrinsically resistant to these chemotherapeutics. Our study aimed to discover potential prediction markers of drug resistance in needle-biopsied tissues of breast cancer patients prior to neoadjuvant chemotherapy. Tissues collected before chemotherapy were analyzed by mass spectrometry. A total of 2,331 proteins were identified and comparatively quantified between drug sensitive (DS) and drug resistant (DR) patient groups by spectral count. Of them, 298 proteins were differentially expressed by more than 1.5-fold. Some of the differentially expressed proteins (DEPs) were further confirmed by Western blotting. Bioinformatic analysis revealed that the DEPs were largely associated with drug metabolism, acute phase response signaling, and fatty acid elongation in mitochondria. Clinical validation of two selected proteins by immunohistochemistry found that FKBP4 and S100A9 might be putative prediction markers in discriminating the DR group from the DS group of breast cancer patients. The results demonstrate that a quantitative proteomics/bioinformatics approach is useful for discovering prediction markers of drug resistance, and possibly for the development of a new therapeutic strategy.