Tamoxifen resistance is emerging as a big challenge in endocrine therapy of luminal A breast cancer patients. In this study, we aimed to determine the molecular changes of PI3K/AKT/PTEN signaling pathway during tamoxifen-resistance development using gradually increased doses of tamoxifen in one model, while fixing tamoxifen treatment dose at 35 μM for several times in the second model. An upregulation of AKT/PI3K genes was noticed at 30 μM tamoxifen concentration in cells treated with a gradual increase of tamoxifen doses. In the second model, significant upregulation of AKT1 was seen in cells treated with 35 μM tamoxifen for three times. All genes studied showed a significant increase in expression in resistant cells treated with 50 µM and 35 µM six times tamoxifen. These genes’ upregulation was accompanied by PTEN and GSK3 ß genes’ down-regulation, and it was in correlation to the changes in the metabolic rate of glucose in tamoxifen-resistant models. A significant increase in glucose consumption rate from culture media was observed in tamoxifen resistant cells with the highest consumption rate reported in the first day of culturing. Increased glucose consumption rates were also correlated with GLUL significant gene expression and non-significant change in c-MYC gene expression that may lead to increased endogenous glutamine synthesis. As a result, several molecular and metabolic changes precede acquired tamoxifen resistance could be used as resistance biomarkers or targets to reverse tamoxifen resistance.
BackgroundThe introduction of effective novel biomarkers of invasion and metastasis is integral for the advancement of breast cancer management. The present study focused on the identification and evaluation of calreticulin (CRT) as a potential biomarker for breast cancer invasion.MethodsTwo-dimensional gel protein electrophoresis and MALDI-TOF were utilized in the analysis of fresh-frozen invasive intra-ductal carcinoma specimens. Calreticulin-associated expression was analyzed using immunohistochemistry of FFPE non-malignant/malignant breast specimens. A CRT-knockdown model of MCF7 cell line was developed using siRNA and the CRT genotype/phenotype correlations based on migration and trans-well invasion assays were determined. Finally, microarray-based global gene expression profiling was conducted to elucidate the possible calreticulin pro-invasive regulatory pathways.ResultsTwo-dimensional gel protein electrophoresis and MALDI-TOF analysis showed upregulation of calreticulin expression in tumor tissues as compared to the normal adjacent tissues. Meta-analysis of the immunohistochemical results confirmed significantly higher expression of calreticulin (p < 0.05) in the stromal compartments of malignant tissues as compared to non-malignant tissues. Migration and transwell invasion assays showed significant loss in the migratory and invasive potential of CRT-knockdown cells (p < 0.05). Global gene expression profiling successfully identified various putative gene networks such as p53 and MAPK pathways that are involved in calreticulin breast cancer signaling.ConclusionBesides confirming calreticulin overexpression in invasive breast cancer tissues, this study reveals a calreticulin-dependent pro-invasive potential and suggests possible contributing pathways. Defining the mechanistic role of invasion and characterizing the possible calreticulin-dependent molecular targets will be the focus of future work.Electronic supplementary materialThe online version of this article (doi:10.1186/s12935-016-0329-y) contains supplementary material, which is available to authorized users.
Gold nanorods (GNRs) have gained pronounced recognition in the diagnosis and treatment of cancers driven by their distinctive properties. Herein, a gold-based nanosystem was prepared by utilizing a phospholipid moiety linked to thiolated polyethylene glycol, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-PEG-SH, as a surface decorating agent. The synthesized phospholipid-PEG-GNRs displayed good colloidal stability upon exposure to the tissue culture medium. Cytotoxicity of phospholipid-PEG-GNRs was investigated toward MCF-7 and T47D breast cancer cells using sulforhodamine B test. The results revealed that phospholipid-PEG-GNRs demonstrated high cytotoxicity to MCF-7 cells compared to T47D cells, and minimal cytotoxicity to human dermal fibroblasts. The cellular uptake studies performed by imaging and quantitative analysis demonstrated massive internalization of phospholipid-coated GNRs into MCF-7 cells in comparison to T47D cells. The cellular death modality of cancer cells after treatment with phospholipid-PEG-GNRs was evaluated using mitochondrial membrane potential assay (JC-1 dye), gene expression analysis, and flow cytometry study. The overall results suggest that phospholipid-modified GNRs enhanced mainly the cellular apoptotic events in MCF-7 cells in addition to necrosis, whereas cellular necrosis and suppression of cellular invasion contributed to the cellular death modality in the T47D cell line upon treatment with phospholipid-PEG-GNRs. The phospholipid-coated GNRs interact in a different manner with breast cancer cell lines and could be considered for breast cancer treatment.
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