Target For Breast Cancer Therapy Research Articles

MiR-455 inhibits breast cancer cell proliferation through targeting CDK14

Breast cancer is the most frequently occurring cancer in women worldwide, microRNAs (miRNAs) play critical role in the initiation and progression of breast cancer. Here, we studied the effect of miR-455 on cell proliferation of breast cancer, and found that miR-455 was downregulated in breast cancer tissues and cells. Its overexpression inhibited cell proliferation, whereas its knockdown promoted cell proliferation of breast cancer. We found a Cdc2-related protein kinase CDK14 was the target of miR-455, when the 3’UTR of CDK14 was cloned into luciferase reporter vector and transfected into cells, miR-455 mimic could inhibit the luciferase activity in a dose-dependent manner, miR-455 inhibitor increased the luciferase activity, but the mutant miR-455 mimic couldn’t change the luciferase activity, suggesting miR-455 directly bound to the 3’UTR of CDK14. Meanwhile, we also found miR-455 inhibited Cyclin D1 expression and promoted p21 expression, confirming miR-455 inhibited cell proliferation. Double knockdown of miR-455 and CDK14 inhibited the proliferation of breast cancer cell, confirming miR-455 inhibiting cell proliferation by targeting CDK14. Moreover, miR-455 levels were negatively correlated with CDK14 levels in breast cancer tissues. Our finding revealed miR-455 inhibits breast cancer cell proliferation through targeting CDK14, it might be a target for breast cancer therapy.

Suppression of TAFI by siRNA inhibits invasion and migration of breast cancer cells.

Human thrombin activatable fibrinolysis inhibitor (TAFI), also known as carboxypeptidase B2 (CPB2), is a procarboxypeptidase enzyme. The purpose of the present study was to observe the expression of TAFI in breast cancer (BC) and breast cancer cell (BCC) lines and to investigate the effect of TAFI suppression by small interfering (si)RNA gene silencing on invasion and migration of BCC lines. A significant increase in TAFI level was identified by immunohistochemical analysis in BC tissues compared with normal breast tissues. TAFI suppression also inhibited cell viability, invasion and migration ability as demonstrated by MTT, Transwell chamber, and wound scratch assays, respectively (P<0.05). The data suggested that suppression of TAFI by siRNA inhibits invasion and migration of breast cancer cells and that TAFI may be a new target for breast cancer therapy.

Phosphatidylinositol 3-kinase regulatory subunit 1 and phosphatase and tensin homolog as therapeutic targets in breast cancer.

Breast cancer is the most commonly diagnosed cancer among women in Turkey and worldwide. It is considered a heterogeneous disease and has different subtypes. Moreover, breast cancer has different molecular characteristics, behaviors, and responses to treatment. Advances in the understanding of the molecular mechanisms implicated in breast cancer progression have led to the identification of many potential therapeutic gene targets, such as Breast Cancer 1/2, phosphatidylinositol 3-kinase catalytic subunit alpha, and tumor protein 53. The aim of this review is to summarize the roles of phosphatidylinositol 3-kinase regulatory subunit 1 (alpha) (alias p85α) and phosphatase and tensin homolog in breast cancer progression and the molecular mechanisms involved. Phosphatase and tensin homolog is a tumor suppressor gene and protein. Phosphatase and tensin homolog antagonizes the phosphatidylinositol 3-kinase/AKT signaling pathway that plays a key role in cell growth, differentiation, and survival. Loss of phosphatase and tensin homolog expression, detected in about 20%-30% of cases, is known to be one of the most common tumor changes leading to phosphatidylinositol 3-kinase pathway activation in breast cancer. Instead, the regulatory subunit p85α is a significant component of the phosphatidylinositol 3-kinase pathway, and it has been proposed that a reduction in p85α protein would lead to decreased negative regulation of phosphatidylinositol 3-kinase and hyperactivation of the phosphatidylinositol 3-kinase pathway. Phosphatidylinositol 3-kinase regulatory subunit 1 protein has also been reported to be a positive regulator of phosphatase and tensin homolog via the stabilization of this protein. A functional genetic alteration of phosphatidylinositol 3-kinase regulatory subunit 1 that results in reduced p85α protein expression and increased insulin receptor substrate 1 binding would lead to enhanced phosphatidylinositol 3-kinase signaling and hence cancer development. Phosphatidylinositol 3-kinase regulatory subunit 1 underexpression was observed in 61.8% of breast cancer samples. Therefore, expression/alternations of phosphatidylinositol 3-kinase regulatory subunit 1 and phosphatase and tensin homolog genes have crucial roles for breast cancer progression. This review will summarize the biological roles of phosphatidylinositol 3-kinase regulatory subunit 1 and phosphatase and tensin homolog in breast cancer, with an emphasis on recent findings and the potential of phosphatidylinositol 3-kinase regulatory subunit 1 and phosphatase and tensin homolog as a therapeutic target for breast cancer therapy.

Inflammatory peroxidases promote breast cancer progression in mice via regulation of the tumour microenvironment.

Myeloperoxidase (MPO) and eosinophil peroxidase (EPO) are heme-containing enzymes, well known for their antimicrobial activity, are released in high quantities by infiltrating immune cells in breast cancer. However, the functional importance of their presence within the tumour microenvironment is unclear. We have recently described a new role for peroxidases as key regulators of fibroblast and endothelial cell functionality. In the present study, we investigate for the first time, the ability of peroxidases to promote breast cancer development and progression. Using the 4T1 syngeneic murine orthotopic breast cancer model, we examined whether increased levels of peroxidases in developing mammary tumours influences primary tumour growth and metastasis. We showed that MPO and EPO stimulation increased mammary tumour growth and enhanced lung metastases, effects that were associated with reduced tumour necrosis, increased collagen deposition and neo-vascularisation within the primary tumour. Invitro, peroxidase treatment, robustly stimulated human mammary fibroblast migration and collagen typeI and typeVI secretion. Mechanistically, peroxidases induced the transcription of pro-tumorigenic and metastatic MMP1, MMP3 and COX-2 genes. Taken together, these findings identify peroxidases as key contributors to cancer progression by augmenting pro-tumorigenic collagen production and angiogenesis. Importantly, this identifies inflammatory peroxidases as therapeutic targets in breast cancer therapy.

Vitamin D receptor as a target for breast cancer therapy.

Considerable epidemiological evidence suggests that high levels of circulating vitamin D (VD) are associated with a decreased incidence and increased survival from cancer, i.e., VD may possess anti-cancer properties. The aim of this investigation was therefore to investigate the anti-cancer potential of a low calcaemic vitamin D analogue, i.e., inecalcitol and compare it with the active form of vitamin D, i.e., calcitriol, in a panel of breast cancer cell lines (n = 15). Using the MTT assay, IC50 concentrations for response to calcitriol varied from 0.12 µM to >20 µM, whereas those for inecalcitol were significantly lower, ranging from 2.5 nM to 63 nM (P = 0.001). Sensitivity to calcitriol and inecalcitol was higher in VD receptor (VDR)-positive compared to VDR-negative cell lines (P = 0.0007 and 0.0080, respectively) and in ER-positive compared to ER-negative cell lines (P = 0.043 and 0.005, respectively). Using RNA-seq analysis, substantial but not complete overlap was found between genes differentially regulated by calcitriol and inecalcitol. In particular, significantly enriched gene ontology terms such as cell surface signalling and cell communication were found after treatment with inecalcitol but not with calcitriol. In contrast, ossification and bone morphogenesis were found significantly enriched after treatment with calcitriol but not with inecalcitol. Our preclinical results suggest that calcitriol and inecalcitol can inhibit breast cancer cell line growth, especially in cells expressing ER and VDR. As inecalcitol is significantly more potent than calcitriol and has low calcaemic potential, it should be further investigated for the treatment of breast cancer.

Abstract P6-08-06: A positive feedback loop couples CD44s and HAS2 for sustained Akt activation and tumor cell survival

Abstract Breast cancer is the second leading cause of cancer death in women worldwide. Despite recent advances in breast cancer treatment, tumor metastasis and recurrence remain the most significant clinical obstacles, largely due to the resistance of residual breast tumor cells to existing therapies. RNA alternative splicing is a fundamental mechanism for gene regulation and protein diversity. Despite the fact that nearly all human genes are regulated by alternative splicing, very limited studies have addressed the role of alternative splicing in tumorigenesis. Our lab uses the cell adhesion molecule CD44 as a model to investigate the role of alternative splicing in breast cancer metastasis. CD44 undergoes extensive alternative splicing, giving rise to the standard (CD44s) and variant (CD44v) isoforms. Previous work from our lab demonstrated that the CD44s isoform is required for Epithelial to Mesenchymal transition (EMT) and breast tumor metastasis. Mechanistically, we have shown that CD44s, but not CD44v, potentiates Akt signaling, promoting cell survival and chemoresistance in breast cancer cells. To better understand CD44s-mediated downstream pathways critical for breast tumor metastasis, we analyzed CD44s-dependent gene signature and identified Hyaluronan Synthase 2 (HAS2), an enzyme that produces hyaluronan (HA), as one of the most significantly upregulated genes in response to CD44s expression. Interestingly, HA is the major ligand for CD44, and HA/CD44 interaction promotes Akt activation. These results led us to test a hypothesis that a positive feedback loop, involving CD44s and HAS2, sustains Akt signaling and promotes tumor cell survival. Indeed, our results show that CD44s-mediated Akt activation stimulates HAS2 transcription. This increase in HAS2 transcription is mediated by Akt phosphorylation of FOXO1, a transcription repressor inactivated by phosphorylation. We further show that HAS2 and HA activate Akt signaling in a CD44s-dependent manner. Importantly, disruption of this feed-forward loop inhibits EMT and triggers cell death. PI3K/Akt signaling is hyperactive in more than 70% of breast tumors. Hence, the proteins involved in this positive feedback loop represent very attractive and promising drug targets for breast cancer therapy. Our work has demonstrated how aberrant expression of alternative spliced isoform in breast cancer, through a positive feedback loop, leads to sustained Akt signaling, enhanced cell survival, and a metastatic phenotype. Disrupting this positive feedback loop may provide a foundation for an innovative approach to treat metastatic breast cancer patients. Citation Format: Liu S, Cheng C. A positive feedback loop couples CD44s and HAS2 for sustained Akt activation and tumor cell survival [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-08-06.

Abstract P3-05-08: Steroid receptors and steroidogenic enzymes in human breast cancer: Associations with breast cancer subtypes and clinical outcome

Abstract Introduction: The majority of breast cancer (BC) cases are believed to be highly steroid dependent. Therefore exploitation of currently untapped steroid pathways could potentially contribute to our search for novel targeted breast cancer therapies. Given the growing understanding of the importance of both intratumoral steroid metabolism and impact of alternate steroid receptors in BC biology, we evaluated selected steroidogenic pathways in a cohort of 140 BC cases from Akershus University Hospital, Norway. In this study we immunolocalized both the androgen receptor (AR) and glucocorticoid receptor (GR) in addition to steroid sulfatase (STS) and 17β hydroxysteroid dehydrogenase 2 (17βHSD2). We also evaluated immunoreactivity of Ki-67 and CYP19/aromatase. We correlated these factors with the occurrence of distant metastasis or local relapse, and survival adjusted by age at surgery. We also examined whether any of these pathways were enriched in the classical breast cancer subtypes (Luminal A, Luminal B, HER-2 positive, triple-negative BC). Logistic and Cox regression analysis was employed to examine interactions between the above factors. Results: In an analysis looking at possible interactions between recurrence and the factors above, the status of intratumoral STS was significantly inversely associated with distant metastasis (OR=0.17, p&amp;lt;0.001) and local relapses (OR=0.17, p&amp;lt;0.001), while that of 17βHSD2 trended towards correlation with the presence of metastasis (OR=3.47, p=0.055) and relapse (OR=3.14, p=0.07). Interestingly, ER and PR were not correlated with metastasis or relapse, but HER2 status positively correlated with distant metastasis during follow-up (OR=2.15, p=0.02) and trended to be associated with local relapses as well (OR=1.99, p=0.054). When evaluated according to the established breast cancer subtypes the only significant changes detected between subgroups were for AR and STS. AR was positive in a lesser proportion of the cases in the TNBC subgrouping (p=0.01), while STS was depleted in the luminal B and enriched in the HER2 subtypes (p=0.027). In an analysis of survival, STS was the only factor examined that was significantly associated with survival outcomes of the patients. Positive intratumoral STS status was significantly associated with improved survival (HR=0.27, p&amp;lt;0.001). The inverse of this was also noticed for 17βHSD2 but the correlation did not reach statistical significance (HR=2.1, p=0.16). Discussion and Conclusions: The results of our present study suggest that the intratumoral metabolism of estrogens through STS is associated with significantly lower incidence of relapse and/or distant metastasis and correspondingly a better prognosis. The inverse is also true when evaluating the trends noted with the expression of 17βHSD2 adding consistency to our finding above. The enrichment of STS in the HER2 overexpressing subtype of breast cancer is intriguing, especially given the possible role of HER-2 positivity in endocrine resistance. All in all, our results suggest that extended endocrine information about the intratumoral steroid metabolism of a given tumour may have so far unused prognostic potential and impact on clinical decision making. Citation Format: McNamara KM, Guestini F, Sauer T, Lindstrøm JC, Sasano H, Geisler J. Steroid receptors and steroidogenic enzymes in human breast cancer: Associations with breast cancer subtypes and clinical outcome [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P3-05-08.

Abstract OT3-01-05: Functional MRI signatures of immune response to targeted breast cancer therapy

Abstract Background: Emerging data has shown that the immune system participates in both tumor development and tumor elimination and control. In breast cancer, activation of the immune system may mediate the effects of several anticancer drugs. In HER2+ and triple negative breast cancers, even 10% increases in tumor infiltrating lymphocytes (TILs) predict improved prognosis to adjuvant chemotherapy. Furthermore, the monoclonal antibody trastuzumab is known to increase peripheral Th1 immunity in women with HER2+ breast cancer, which may be reflected by increased TILs. An imaging approach to measure and monitor immune response would provide an essential non-invasive tool to select appropriate therapies based on an individual's response. Dynamic contrast-enhanced (DCE) MRI, reflecting blood flow and capillary permeability, and diffusion weighted (DW) MRI, reflecting cellularity, hold unique potential to provide quantitative imaging markers of therapy-induced immune response in breast tumors. Trial Design: We propose to use trastuzumab to invoke immune response in patients with HER2+ tumors. Each subject will undergo a multiparametric MRI exam (with DCE and DW-MRI) and tissue sampling before and after a single dose of targeted anti-HER2 therapy with trastuzumab, prior to surgery. Pretreatment tissue will be obtained form the diagnostic core biopsy, and post-trastuzumab tissue will be collected from a research biopsy or the surgical excision, requiring at most a single additional biopsy. A variety of quantitative parameters will be extracted from the MRI scans. Whole tumor characterization will be performed, to calculate histogram-based metrics and second order textural features. Functional MRI features will be compared with histologic assessment of TILs, VEGF expression, and other immune response markers in tumors treated with HER2-targeted therapy. After the study window, patients will undergo standard of care treatment at the discretion of their physician. Specific Aims: The primary objective will be to identify quantitative MRI markers of immune response to HER2-targeted treatment. We will implement a high-resolution multiparametric MRI approach and determine whether early changes on MRI after a preoperative run-in dose of trastuzumab correlate with tumor immune response markers as measured by histologic assessment. Statistical Methods: The magnitude of response will be determined for individual MRI and histologic markers following one cycle of trastuzumab, and Pearson and Spearman correlation coefficients will be used to assess associations between changes in MRI and histologic markers. With 50 patients, we will have 80% power to detect a significant correlation between MRI and histologic markers as small as r = 0.39 (R2 = 15%), based on a 2-sided test of the null hypothesis r = 0 at the 0.05 significance level. Accrual: Three patients have been accrued to date, with a target of 50 HER2+ patients. Patients may consent to either 1) preoperative run-in of a single cycle of anti-HER2 treatment or 2) neoadjuvant targeted anti-HER2 only treatment regimen (e.g., co-enrollment in TBCRC 026 or comparable trial), either of which allows for monitoring at pre and post 1 cycle of anti-HER2 treatment. Contact information: VK Gadi (vkgadi@uw.edu) or Savannah Partridge (scp3@uw.edu). Citation Format: Gadi VK, Stanton S, Dintzis SM, Calhoun KE, Hippe DS, Partridge SC. Functional MRI signatures of immune response to targeted breast cancer therapy [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr OT3-01-05.

The innate and adaptive infiltrating immune systems as targets for breast cancer immunotherapy.

A cancer cell-centric view has long dominated the field of cancer biology. Research efforts have focussed on aberrant cancer cell signalling pathways and on changes to cancer cell DNA. Mounting evidence demonstrates that many cancer-associated cell types within the tumour stroma co-evolve and support tumour growth and development, greatly modifying cancer cell behaviour, facilitating invasion and metastasis and controlling dormancy and sensitivity to drug therapy. Thus, these stromal cells represent potential targets for cancer therapy. Among these cell types, immune cells have emerged as a promising target for therapy. The adaptive and the innate immune system play an important role in normal mammary development and breast cancer. The number of infiltrating adaptive immune system cells with tumour-rejecting capacity, primarily, T lymphocytes, is lower in breast cancer compared with other cancer types, but infiltration occurs in a large proportion of cases. There is strong evidence demonstrating the importance of the immunosuppressive role of the innate immune system during breast cancer progression. A consideration of components of both the innate and the adaptive immune system is essential for the design and development of immunotherapies in breast cancer. In this review, we focus on the importance of immunosuppressive myeloid-derived suppressor cells (MDSCs) as potential targets for breast cancer therapy.

Targeted breast cancer therapy by harnessing the inherent blood group antigen immune system

Cancer gene therapy has attracted increasing attention for its advantages over conventional therapy in specific killing of tumor cells. Here, we attempt to prove a novel therapeutic approach that targets tumors by harnessing the blood antigen immune response system, which is inherently present in patients with breast cancers. Breast cancer MDA-MB-231 cells expressed blood group H antigen precursor. After ectopic expression of blood group A glycosyltransferase, we found that the H precursor was converted into the group A antigen, appearing on the surface of tumor cells. Incubation with group B plasma from breast cancer patients activated the antigen-antibody-complement cascade and triggered tumor cell killing. Interestingly, expression of blood A antigen also reduced tumorigenesis in breast cancer cells by inhibiting cell proliferation, migration, and tumor sphere formation. Cell cycle analysis revealed that cancer cells were paused at S phase due to the activation of cell cycle regulatory genes. Furthermore, pro-apoptotic genes were unregulated by the A antigen, including BAX, P21, and P53, while the anti-apoptotic BCL2 was down regulated. Importantly, we showed that extracellular HMGB1 and ATP, two critical components of the immunogenic cell death pathway, were significantly increased in the blood A antigen-expressing tumor cells. Collectively, these data suggest that blood antigen therapy induces specific cancer cell killing by activating the apoptosis and immunogenic cell death pathways. Further translational studies are thereby warranted to apply this approach in cancer immuno-gene therapy.

Glutamic Pyruvate Transaminase GPT2 Promotes Tumorigenesis of Breast Cancer Cells by Activating Sonic Hedgehog Signaling.

Increased glutamine metabolism is a hallmark of cancer. Mitochondrial glutamic pyruvate transaminase (GPT2) catalyzes the reversible transamination between alanine and α-ketoglutarate (α-KG), also known as 2-oxoglutarate, to generate pyruvate and glutamate during cellular glutamine catabolism. However, the precise role of GPT2 in tumorigenesis remains elusive. Here, we report that in breast cancer tissue samples and breast cancer cell lines, GPT2 expression level was markedly elevated and correlated with the pathological grades of breast cancers. GPT2 overexpression increased the subpopulation of breast cancer stem cells in vitro and promoted tumorigenesis in mice. GPT2 reduced α-KG level in cells leading to the inhibition of proline hydroxylase 2 (PHD2) activity involved in the regulation of HIF1α stability. Accumulation of HIF1α, resulting from GPT2-α-KG-PHD2 axial, constitutively activates sonic hedgehog (Shh) signaling pathway. Overall, GPT2 promotes tumorigenesis and stemness of breast cancer cells by activating the Shh signaling, suggesting that GTP2 is a potential target for breast cancer therapy.

G Protein-Coupled Estrogen Receptor Is a Critical Regulator in Metastasis of Breast Cancer Cells

Estrogen signaling via GPER in breast cancers has been intensively discussed over years, but the underlying molecular mechanism remains to be fully elucidated, especially for the transcriptional profiles of the GPER under estrogen stimulation. In this study, we evaluated the potential role GPER in regulating invasion of metastatic breast cancer cell line MDA-MB-231 via transcriptional way. First, with primary breast cancer tissue samples, we found that the expression of GPER significantly coordinated with another membrane receptor-CXCR1. Besides, the expression level of these two proteins was associated with the development of the primary breast cancers. Second, to dissect the cross talk between GPER and CXCR1, we further our studies to detect the activation of ERK, Akt and transcriptional factor NF-kB. We found that upon estrogen stimulation, the phosphorylation level of ERK and Akt were rapidly increased and then resulted in the activation and nucleus translocation of NF-kB. When we considered the sub-sequence of the NF-kB activation, we found the autocrine of IL-8 was boosted by stimulation of the estrogen and followed by the promoted invasion of the MDA-MB-231 cells. In conclusion, our data demonstrated the estrogen-mediated GPER stimulation would regulate the invasive activities of metastatic breast cancer cell line MDA-MB-231 coupled with another membrane receptor CXCR1 via transcriptional pathway, the cross talk between GPER and CXCR1 may be another target for breast cancer therapies.

Silencing HS6ST3 inhibits growth and progression of breast cancer cells through suppressing IGF1R and inducing XAF1

Heparan sulfate 6-O-sulfation is biologically edited by 6-O-sulfotransferases (HS6STs) within heparan sulfate chains. Three isoforms of HS6ST have been identified. These enzymes are found to be differentially expressed in a variety of tissues. Recently, several studies have shown that dysregulation of 6-O-sulfotransferases could be involved in tumorigenesis of several cancers. This study aimed to analyze the expression and function of HS6ST3 in breast cancer. HS6ST3 was found up-regulated in T47D, MCF7 and MDA-MB231 breast cancer cell lines. HS6ST3 was then silenced in T47D and MCF7 using siRNA. Silencing HS6ST3 diminished tumor cell growth, migration and invasion, but enhanced cell adhesion and apoptosis in breast cancer. Gene microarray analysis revealed that silencing HS6ST3 significantly changed the expression of IGF1R and XAF1 in breast cancer cells. Further functional studies showed that the cellular processes were mediated by IGF1R and XAF1 after silencing HS6ST3 in breast cancer cells. Together these results indicate that HS6ST3 might be involved in the tumorigenesis of breast cancer and it could be a promising target in breast cancer therapy.

The effect of transmembrane 7 superfamily member 4 gene on behavior of breast cancer cell

Objective To investigate the effect of transmembrane 7 superfamily member 4 (TM7SF4) gene on behavior of breast cancer cell. Methods The TM7SF4 lentivirusis constructed and used to transfect MCF-7 human breast cancer cells, and then down-regulate TM7SF4 gene. Investigate the effect of TM7SF4 gene on the proliferation, apoptosis, cell cycle and cell proliferation of MCF-7 cells by Cellomics, flow cytometry, propidium iodide-fluorescence activated cell sorter (PI-FACS) and clone formation. Results (1) Effect on proliferation of MCF-7 cells: the number of MCF-7 cellsof experimental group in 5 days cultured were: (239.1±37.7), (508.4±52.8), (282.2±84.0), (308.0±167.1), (131.6±82.6) cell/well; the number of MCF-7 cells of no-treatment control group in 5 days cultured were: (382.8±38.0), (1 115.0±155.9), (1 426.2±192.4), (2 468.4±232.6), (2 903.0±165.7) cell/well; the number of MCF-7 cells of plasmid-treatment group in 5 days cultured were: (380.8±39.0), (1 090.0±145.9), (1 426.0±180.3), (2 379.0±228.6), (2 855.0±156.6) cell/well. results showed that the number of cells and rate of cells growth of experimental group were significantly inhibited (P<0.05); (2) Effect on cell cycle of MCF-7 cells: the number of MCF-7 cells of experimental group at G0/G1 stage, S stage, G2/M stage were (61.87±0.27), (20.31±0.11), (17.83±0.35) cell/well, respectively; and that of no-treatment control group were (57.46±1.40), (30.42±0.36), (12.83±0.55) cell/well, respectively, and that of plasmid-treatment group were (57.78±1.23), (30.11±0.47), (13.30±0.79) cell/well, respectively. Statistical differences were found between experimental group and no-treatment control group, between experimental group and plasmid-treatment group (P<0.05); (3) Effect on apoptosis of MCF-7 cells: MCF-7 cell apoptosis rate of experimental group, no-treatment control group and plasmid-treatment group was: (9.94±0.57)%, (5.88±0.23)%, (5.80±0.24)% in scatter diagram and (27.37±1.11)%, (23.34±0.50)%, (23.37±0.46)% in peak valuesdiagram, respectively; There were statistical difference between experimental and no-treatment control group, between experimental group and plasmid-treatment group both in scatter diagram (P<0.05) and peak values diagram (P<0.05), respectively; (4) Effect on clone formation of MCF-7 cells: number of the colony MCF-7 cells in the experimental group, no-treatment control group and plasmid-treatment control was 11±3, 34±2, 29±1, respectively. There were statistical difference between experimental group and no-treatment control group (P<0.05), between experimental group and plasmid-treatment control group (P<0.05), respectively. Conclusion It was confirmed that the TM7SF4 gene had the ability to promote the proliferation and clone formation of MCF-7 breast cancer cells, and to inhibit the apoptosis and cell cycle progression. TM7SF4 gene played a role in promoting the development of breast cancer cells. A new theoretical basis and experimental data were presented for early diagnosis and targeted therapy of breast cancer. Key words: Breast cancer; Transmembrane 7 superfamily member 4; Apoptosis; Proliferation

ERBB2 mutation is associated with a worse prognosis in patients with CDH1 altered invasive lobular cancer of the breast.

E-cadherin (CDH1) is a glycoprotein that mediates adhesion between epithelial cells and also suppresses cancer invasion. Mutation or deletion of the CDH1 gene has been reported in 30–60% cases of invasive lobular carcinoma (ILC). However, little is known about genomic differences between ILC with and without a CDH1 alteration. Therefore, we analyzed whole genome sequencing data of 169 ILC cases from The Cancer Genome Atlas (TCGA) to address this deficiency. Our study shows that CDH1 gene was altered in 59.2% (100/169) of ILC. No significant difference was identified between CDH1-altered and -unaltered ILC cases for any of the examined demographic, clinical or pathologic characteristics, including histologic grade, tumor stage, lymph node metastases, or ER/PR/HER2 states. Seven recurrent mutations (PTEN, MUC16, ERBB2, FAT4, PCDHGA2, HERC1 and FLNC) and four chromosomal changes with recurrent copy number variation (CNV) (11q13, 17q12-21, 8p11 and 8q11) were found in ILC, which correlated with a positive or negative CDH1 alteration status, respectively. The prevalence of the most common breast cancer driver abnormalities including TP53 and PIK3CA mutations and MYC and ERBB2 amplifications showed no difference between the two groups. However, CDH1-altered ILC with an ERBB2 mutation shows a significantly worse prognosis compared to its counterparts without such a mutation. Our study suggests that CDH1-altered ILC patients with ERBB2 mutations may represent an actionable group of patients who could benefit from targeted breast cancer therapy.

Multiple molecular targets in breast cancer therapy by betulinic acid.

Breast cancer is the second most common type of cancer in the world, and is by far the most prevalent form of cancer in women. However, the efficacy of current treatments for breast cancer is limited. In addition to the high risk of recurrence, some of these have side effects that significantly reduce the quality of life. Therefore, new avenues of treatment for breast cancer are needed. Betulinic acid (BA), a pipeline anticancer drug, exerts anti-proliferative effects on breast cancer cells is mainly through inhibition of cyclin and topoisomerase expression, leading to cell cycle arrest. It induces apoptosis through mitochondrial pathway and anti-angiogenesis effect by inhibiting the expression of transcription factor nuclear factor kappa B (NF-κB), specificity protein (Sp) transcription factors, and vascular endothelial growth factor (VEGF) signaling. In addition, it exerts anti-metastatic effect by inhibiting the expression of matrix metalloproteases. The specific targets of BA in breast cancer are reported to be the estrogen receptor and various multidrug resistance proteins. Synergistically interactions of BA with other chemotherapeutics are also described in the literature. In this review, we describe the detailed published mechanisms of action of BA, a pentacyclic triterpene with a lupine skeleton, on multiple molecular targets to treat breast cancer. We hope that this review will provide basic information in support of future studies of effects of BA on breast cancer cells.

Albumin hybrid nanoparticles loaded with tyrosine kinase A inhibitor GNF-5837 for targeted inhibition of breast cancer cell growth and invasion

Novel albumin hybrid nanoparticles (Alb-HNPs) loaded with tyrosine kinase A (TrkA) inhibitor GNF-5837 were prepared and evaluated for antineoplastic efficacy in a panel of breast cancer cell lines. The nanomedicines (GNF-Alb-HNPs, hydrodynamic diameter ∼150nm) were formed through a unique polyelectrolyte complexation process where albumin and GNF-5837 were encapsulated by a stabilizing layer of oppositely charged chitosan and dextran sulfate polysaccharides. GNF-Alb-HNPs showed an excellent colloidal stability and a sustained drug release over more than 24h. We found that these nanomedicines inhibited TrkA phosphorylation and downstream mitogen-activated protein kinase (MAPK) signaling in breast cancer cells specifically, resulting in anti-proliferative and pro-apoptotic effects. Moreover, the migration and invasion activities of cancer cells were dramatically suppressed and the inhibitory effects were much more prominent with GNF-Alb-HNPS than the drug alone. These results show that the GNF-Alb-HNPs may represent a novel approach for targeted breast cancer therapy.

Leptin promotes epithelial-mesenchymal transition of breast cancer via the upregulation of pyruvate kinase M2.

BackgroundAccumulating researches have shown that epithelial-mesenchymal transition (EMT) contributes to tumor metastasis. Leptin, a key adipokine secreted from adipocytes, shapes the tumor microenvironment, potentiates the migration of breast cancer cells and angiogenesis, and is also involved in EMT. However, the potential mechanism remains unknown. This study aims to explore the effect of leptin on EMT in breast cancer cells and the underlying mechanism.MethodsWith the assessment of EMT-associated marker expression in MCF-7, SK-BR-3, and MDA-MB-468 cells, the effect of leptin on breast cancer cells was analyzed. Besides, an array of pathway inhibitors as well as RNA interference targeting pyruvate kinase M2 (PKM2) were used to clarify the underlying mechanism of leptin-mediated EMT in vitro and in vivo.ResultsThe results demonstrated that leptin promoted breast cancer cells EMT, visibly activated the PI3K/AKT signaling pathway, and upregulated PKM2 expression. An antibody against the leptin receptor (anti-ObR) and the PI3K/AKT signaling pathway inhibitor LY294002 significantly abolished leptin-induced PKM2 expression and EMT-associated marker expression. SiRNA targeting PKM2 partially abolished leptin-induced migration, invasion, and EMT-associated marker expression. In vivo xenograft experiments indicated that RNA interference against PKM2 suppressed breast cancer growth and metastasis.ConclusionsOur data suggest that leptin promotes EMT in breast cancer cells via the upregulation of PKM2 expression as well as activation of PI3K/AKT signaling pathway, and PKM2 might be one of the key points and potential targets for breast cancer therapy.

TIPE2 suppresses the tumorigenesis, growth and metastasis of breast cancer via inhibition of the AKT and p38 signaling pathways.

Breast cancer is the second leading cause of cancer‑related deaths in female patients, and the main reasons are late diagnosis, limited therapeutic options and metastasis. Therefore, development of molecular therapeutic targets for breast cancer to suppress tumorigenesis, growth and metastasis may improve the therapeutic options and be of great benefit to patients. Tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) is a novel molecule for maintaining immune homeostasis and is involved in cancer development. Previous studies have shown that TIPE2 plays a suppressive role in the development of several types of human cancers. However, its role in breast cancer is still not clear. In the present study, we detected TIPE2 expression in human breast cancer and adjacent normal tissues. The expression of TIPE2 was reduced in breast cancer tissues compared to the level in adjacent normal tissues. We then established a breast cancer cell line stably expressing TIPE2 to investigate the role of TIPE2 in breast cancer carcinogenesis. Our results showed that overexpression of TIPE2 significantly inhibited the proliferation of MDA-MB‑231 cells as detected by Cell Counting Kit-8 assay and suppressed the migration and invasion of breast cancer cells as detected by Transwell migration and invasion assays invitro. TIPE2 also promoted cell apoptosis as detected by flow cytometry analysis. Moreover, TIPE2 inhibited the tumorigenesis of breast cancer invivo. Mechanistically, TIPE2 inhibited the phosphorylation of AKT and p38 as detected by western blot analysis. Taken together, TIPE2 suppressed breast cancer tumorigenesis, growth and metastasis possibly via regulation of the AKT and p38 signaling pathways. The results indicate that TIPE2 may be a potential therapeutic target for breast cancer therapy.

MiR-9 and miR-200 Regulate PDGFRβ-Mediated Endothelial Differentiation of Tumor Cells in Triple-Negative Breast Cancer.

Organization of cancer cells into endothelial-like cell-lined structures to support neovascularization and to fuel solid tumors is a hallmark of progression and poor outcome. In triple-negative breast cancer (TNBC), PDGFRβ has been identified as a key player of this process and is considered a promising target for breast cancer therapy. Thus, we aimed at investigating the role of miRNAs as a therapeutic approach to inhibit PDGFRβ-mediated vasculogenic properties of TNBC, focusing on miR-9 and miR-200. In MDA-MB-231 and MDA-MB-157 TNBC cell lines, miR-9 and miR-200 promoted and inhibited, respectively, the formation of vascular-like structures in vitro Induction of endogenous miR-9 expression, upon ligand-dependent stimulation of PDGFRβ signaling, promoted significant vascular sprouting of TNBC cells, in part, by direct repression of STARD13. Conversely, ectopic expression of miR-200 inhibited this sprouting by indirectly reducing the protein levels of PDGFRβ through the direct suppression of ZEB1. Notably, in vivo miR-9 inhibition or miR-200c restoration, through either the generation of MDA-MB-231-stable clones or peritumoral delivery in MDA-MB-231 xenografted mice, strongly decreased the number of vascular lacunae. Finally, IHC and immunofluorescence analyses in TNBC specimens indicated that PDGFRβ expression marked tumor cells engaged in vascular lacunae. In conclusion, our results demonstrate that miR-9 and miR-200 play opposite roles in the regulation of the vasculogenic ability of TNBC, acting as facilitator and suppressor of PDGFRβ, respectively. Moreover, our data support the possibility to therapeutically exploit miR-9 and miR-200 to inhibit the process of vascular lacunae formation in TNBC. Cancer Res; 76(18); 5562-72. ©2016 AACR.