Cytotoxicity Of Tamoxifen Research Articles

Impact of PEGylated liposomes on cytotoxicity of tamoxifen and piperine on MCF-7 human breast carcinoma cells

Tamoxifen (TMF) is an anticancer agent used for managing estrogen receptor-positive breast cancer. It has limited therapeutic efficacy against breast cancer, which could be enhanced by the coadministration of herbal drugs like piperine (PIP). However, the hydrophobic nature of TMF and PIP restricts their therapeutic application. Therefore, the present study focuses on the impact of the anticancer activity of TMF in combination with PIP and after entrapping them into liposomes (TMF-PIP-LPs and TMF-PIP-PEG-LPs). The liposomes were prepared using the thin film hydration method. In addition, the morphology of the prepared liposomes was found spherical after SEM and TEM analyses. Further, the in vitro cytotoxicity (IC50) study of pure PIP and TMF was found to be 90.3 ± 10.2 μg/mL and 40.9 ± 5.9 μg/mL, respectively. Interestingly, an improved cytotoxicity (IC50) was observed when the TMF and PIP were loaded into liposomes (TMF-PIP-LPs: 21 ± 1.6 μg/mL and TMF-PIP-PEG-LPs: 10 ± 0.5 μg/mL). Also, the PEGylated liposomes showed improvement in cellular uptake as compared to liposomes without PEGylation in MCF-7 human breast carcinoma cells. Thus, the enhanced cellular uptake and improved cytotoxicity of PEGylated liposomes can be a suitable strategy for delivering TMF with PIP for breast cancer treatment.

2-methoxyestradiol sensitizes tamoxifen-resistant MCF-7 breast cancer cells via downregulating HIF-1α

The clinical studies for breast cancer (BC) are now assessing the efficacy of 2-Methoxyestradiol (2-ME), a naturally occurring derivative of estradiol. Our study aimed to explore the potential of combining the 2-ME and tamoxifen (TAM) on sensitization of TAM-resistant cells using LCC2 the TAM-resistant cells as a model and comparing the results to the sensitive cells MCF-7. Sulphorhodamine-B (SRB) assay is used to examine the 2-ME chemo-sensitizing impact on the cytotoxicity of TAM on LCC2 cells. Colorimetric assay kits were used to assess the level of the apoptosis-related markers caspases 3, Bcl2, and Bax in cell lysate. Hypoxia-inducible factor 1 alpha (HIF-1α) expression was measured using western blotting. Total cholesterol and triglyceride (TG) levels were examined colorimetrically, using the BIOLABO kit. The use of 2-ME enhanced the cytotoxic effects of TAM and effectively reversed TAM resistance. This was achieved by inhibiting the expression of HIF-1α, while concurrently increasing the levels of apoptotic marker caspase-3, as well as the pro-apoptotic protein Bax. Additionally, there was a reduction in the levels of Bcl2, an anti-apoptotic protein. Furthermore, a reduction in TG and cholesterol levels was noted. Our findings show that HIF-1α plays an important role in TAM resistance and that suppression of HIF-1α by 2-ME-mediated sensitization of BC-resistant cells to TAM. Therefore, the concurrent administration of TAM/2-ME might potentially serve as a viable therapeutic approach to address TAM resistance and enhance the overall therapy efficacy for patients with BC.

Hydroxycitric acid reverses tamoxifen resistance through inhibition of ATP citrate lyase

Lipid metabolic reprogramming is involved in mediating tamoxifen (TAM) response in breast cancer cells. Published microarray data indicated that ATP citrate lyase (ACLY) is overexpressed in TAM-resistant BC cells. Hydroxycitric acid (HCA) is a powerful competitive inhibitor of the enzyme ACLY, which links carbohydrates and lipids metabolism. However, whether inhibition of ACLY could modulate TAM response in TAM-resistant BC cells remained unexplored. Thus the current study aimed to explore the effect of ACLY inhibition on TAM-resistant BC cells. The cytotoxicity of TAM and/or HCA on LCC2 and its TAM-sensitive counterpart MCF7 cells was evaluated. Also, the effect of TAM and/or HCA treatments on ACLY protein levels were investigated by western blotting. In addition, the effects of TAM and/or HCA on caspase-3, Bax, and Bcl2 levels were evaluated by ELISA.; besides, and flow cytometric analysis was performed for the detection of apoptosis. Moreover, cholesterol and triglyceride contents of LCC2 and MCF7 were quantified colorimetrically. Our results demonstrated that TAM/HCA co-treatment synergistically diminished LCC2 and MCF7 cell viability, with the effect being more significant on LCC2. Mechanistically, TAM/HCA co-treatment decreases the expression level of ACLY in LCC2 by 74%, while in MCF7 by only 59%. Moreover, apoptosis marker caspase-3 and Bax were increased, while the anti-apoptotic Bcl2 was decreased. Furthermore, the cholesterol and TG contents were increased in LCC2 than in MCF7. Our data revealed that ACLY plays a key role in TAM resistance and ACLY inhibition by HCA-mediated sensitization of BC-resistant cells to TAM.

Effect of nanovesicular surface-functionalization via chitosan and/or PEGylation on cytotoxicity of tamoxifen in induced-breast cancer model

AimsThe effect of surface-modification of Tamoxifen (Tam)-loaded-niosomes on drug cytotoxicity and bio-distribution, via functionalization with chitosan and/or PEGylation, was investigated. Materials and methodsTam-loaded hybrid-nanocarriers (Tam-loaded niosomes, chitosomes, PEGylated niosomes, and PEGylated chitosomes) were formulated and characterized. Key findingsChitosanization with/without PEGylation proved to selectively enhance Tam-release at the cancerous-acidic micromilieu. Cytotoxic activity study showed that Tam-loaded PEGylated niosomes had a lower IC50 value on MCF-7 cell line (0.39, 0.35, and 0.27 times) than Tam-loaded PEGylated chitosomes, Tam-loaded niosomes, and Tam-loaded chitosomes, respectively. Cell cycle analysis showed that PEGylation and/or Chitosanization significantly impact Tam efficiency in inducing apoptosis, with a preferential influence of PEGylation over chitosanization. The assay of Annexin-V/PI double staining revealed that chitosanized-nanocarriers had a significant role in increasing the incidence of apoptosis over necrosis. Besides, PEGylated-nanocarriers increased apoptosis, as well as total death and necrosis percentages more than what was shown from free Tam. Moreover, the average changes in both Bax/Bcl-2 ratio and Caspase 9 were best improved in cells treated by Tam-loaded PEGylated niosomes over all other formulations. The in-vivo study involving DMBA-induced-breast cancer rats revealed that PEGylation made the highest tumor-growth inhibition (84.9 %) and breast tumor selectivity, while chitosanization had a lower accumulation tendency in the blood (62.3 ng/ml) and liver tissues (103.67 ng/ml). The histopathological specimens from the group treated with Tam-loaded PEGylated niosomes showed the best improvement over other formulations. SignificanceAll these results concluded the crucial effect of both PEGylation and chitosan-functionalization of Tam-loaded niosomes in enhancing effectiveness, targetability, and safety.

Fractionated irradiation of MCF7 breast cancer cells rewires a gene regulatory circuit towards a treatment-resistant stemness phenotype.

Radiotherapy is the standard of care for breast cancer. However, surviving radioresistant cells can repopulate following treatment and provoke relapse. Better understanding of the molecular mechanisms of radiation resistance may help to improve treatment of radioresistant tumours. To emulate radiation therapy at the cellular level, we exposed MCF7 breast cancer cells to daily radiation doses of 2 Gy up to an accumulated dose of 20 Gy. Fractionally irradiated cells (FIR20) displayed increased clonogenic survival and population doubling time as compared with age‐matched sham‐irradiated cells and untreated parental MCF7 cells. RNA‐sequencing revealed a core signature of 229 mRNAs and 7 circular RNAs of which the expression was significantly altered in FIR20 cells. Dysregulation of several top genes was mirrored at the protein level. The FIR20 cell transcriptome overlapped significantly with canonical radiation response signatures and demonstrated a remarkable commonality with radiation and endocrine therapy resistance expression profiles, suggesting crosstalk between both acquired resistance pathways, as indicated by reduced sensitivity to tamoxifen cytotoxicity of FIR20 cells. Using predictive analyses and functional enrichment, we identified a gene‐regulatory network that promotes stemness and inflammatory signalling in FIR20 cells. We propose that these phenotypic traits render breast cancer cells more radioresistant but may at the same time serve as potential targets for combination therapies.

Oestrogen Receptor Isoforms May Represent a Therapeutic Target in Oesophageal Adenocarcinoma.

Simple SummaryOesophageal adenocarcinoma is a lethal malignancy with limited treatment options. Recent studies have identified oestrogen receptors (ERs) in this cancer, which could represent a new target for therapy. In this study, we used laboratory models of oesophageal adenocarcinoma to look for the presence of variant forms of ERs. We also assessed the response to treatment with a drug that acts through these ERs. We found that variant forms of ERs do exist in this malignancy and that some of the variants appear to be important in order for the cells to respond to treatment. This could be due to interactions between different ERs, or between ERs and other molecules that are known to be important in cancer growth. Our findings are encouraging in that drugs that act through ERs might be useful for patients with oesophageal adenocarcinoma in the future.Oesophageal adenocarcinoma is a rapidly increasing problem in which treatment options are limited. Previous studies have shown that oesophageal adenocarcinoma cells and tissues express oestrogen receptors (ERs) and show growth suppression and apoptosis in response to ER modulator agents such as tamoxifen. ERs are known to be expressed in a number of isoforms that act together to regulate cell growth and cell death. In this study, we used western blotting to profile the expression of ERα and ERβ isoforms, and expression of the oncologically related molecules p53, HER2, and EGFR, in a panel of oesophageal adenocarcinoma cell lines. The cytotoxicity of tamoxifen in the cell lines was determined with Annexin V-FITC flow cytometry, and correlations between cytotoxicity and receptor expression were assessed using Spearman’s rank-order correlation. Oesophageal adenocarcinoma cell lines showed varying cytotoxicity in response to tamoxifen. The ER species ERα90, ERα50, and ERα46, as well as p53, were positively associated with a cytotoxic response. Conversely, ERα74, ERα70, and ERβ54 were associated with a lack of cytotoxic response. The ER species detected in oesophageal adenocarcinoma cells may work together to confer sensitivity to ER modulators in this disease, which could open up a new avenue for therapy in selected patients.

Overexpression of TMEM47 Induces Tamoxifen Resistance in Human Breast Cancer Cells.

Background:Tamoxifen (TAM) is the eminent first-line drug for endocrine therapy of hormone receptor positive premenopausal breast cancer and reduces the risk of recurrence by ∼50%. However, many patients developed TAM resistance and their diseases recurred. Our previous study on transcriptome profile of TAM resistant breast cancer cells revealed that the TMEM47 is one of the most significantly differentially expressed genes. The mechanism of how TMEM47 is involved in TAM resistance was not known.Methods:We constructed a mammal breast cancer cell line, in which TMEM47 was stably overexpressed (TMEM47-OE/MCF-7), to further verify the role of TMEM47 in TAM resistance. siRNA targeting TMEM47 was transfected into TAMR / MCF-7 cells by Liposome. TMEM47 expression was validated on mRNA and protein level by qRT-PCR and western blotting. We tested the cytotoxicity of TAM in the cells. Apoptosis was detected by flow cytometry.Results:Compared to the MCF7 cells, TMEM47 mRNA was significantly up regulated more than 6 folds in the TAMR/MCF7 cells and so its protein. TMEM47 expression level in TMEM47-OE/MCF-7 was similar as in the TAMR/MCF-7 cells. The 50% inhibitory concentration (IC50) value (mean ± SD) of TAM in MCF-7, TAMR/MCF-7 and TMEM47-OE/MCF-7 cells was 1.58 ± 0.19, 2.74 ± 0.24 and 3.12 ± 0.32 µγ/mL, respectively. The apoptosis rates of TAMR/MCF-7 and TMEM47-OE/MCF-7 cell lines were significantly lower than that of MCF-7 cells. After 24 and 48 hours TAM treatments, cell viability was significantly inhibitied in TMEM47 knockdown TAMR/MCF7 cells (P < 0.01). Consistant with the decreased cell viability, the apoptosis rate in TMEM47 knockdown TAMR/MCF-7 cells was significantly increased.Conclusions:Our results suggest that overexpression of TMEM47 in MCF-7 cells acquired TAM resistance to those cells, and knockdown of TMEM47 in TAMR/MCF-7 cells reversed their resistance to TAM. TMEM47 might confer TAM resistance on MCF-7 cells through the inhibition of apoptosis.

Four electrode-based impedimetric biosensors for evaluating cytotoxicity of tamoxifen on cervical cancer cells.

In the current study, novel four electrode-based impedimetric biosensors have been fabricated using photolithography techniques and utilized to evaluate the cytotoxicity of tamoxifen on cervical cancer cell lines. The cell impedance was measured employing the electric cell-substrate impedance sensing (ECIS) method over the frequency range of 100 Hz to 1 MHz. The results obtained from impedimetric biosensors indicate that tamoxifen caused a significant reduction in the number of HeLa cells on the electrode surfaces in a dose-dependent manner. Next, the impedance values recorded by the fabricated biosensors have been compared with the results obtained from the different conventional techniques such as 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), live-dead cell assay, and flow cytometric analysis to estimate the cytotoxicity of tamoxifen. The impedimetric cytotoxicity of tamoxifen over the growth and proliferation of HeLa cells correlates well with the traditional methods. In addition, the IC50 values obtained from impedimetric data and MTT assay are comparable, signifying that the ECIS technique can be an alternative method to assess the cytotoxicity of different novel drugs. The working principle of the biosensor has been examined by scanning electron microscopy, indicating the detachment of cells from gold surfaces in a dose-dependent manner, signifying the decrease in impedance at higher drug doses.

Tamoxifen-loaded PLA/DPPE-PEG lipid-polymeric nanocapsules for inhibiting the growth of estrogen-positive human breast cancer cells through cell cycle arrest

The strategy of drug encapsulation with biocompatible polymeric nanocapsules could be a promising approach to improve the delivery of poorly water-soluble drugs for cancer therapy. The aim of this study was to investigate the drug delivery of tamoxifen from biocompatible nanocapsules made of poly(D,L-lactide) (PLA) core and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol)-2000] (DPPE-PEG) shell in the treatment of breast cancer. The analytical techniques such as FTIR, XRD, SEM, and TEM were used to investigate the structure of nanocapsules. Tamoxifen drug encapsulation efficiency and loading percentages as well as release kinetics at different pH values and temperatures were used to confirm the appropriate performance of these novel nanocapsules. The maximum tamoxifen release occurred at 40 °C, pH = 4. Based on the SEM and TEM images, the sizes of nanocapsules were between 57 and 159 nm. In order to investigate the anticancer effects and cytotoxicity of encapsulated tamoxifen in PLA/DPPE-PEG nanocapsules, the tetrazolium-based colorimetric (MTT) assay and cell cycle arrest analysis on MCF-7 breast cancer cell were performed. Also, cell cycle–related gene expressions of p53 and p21 in treated MCF-7 cells with tamoxifen-loaded PLA/DPPE-PEG nanocapsules were evaluated. The results of MTT cell proliferation experiments exhibited an increase in the cytotoxicity of tamoxifen nanocapsules on MCF-7 cell line compared with free tamoxifen. The 1.2-fold increase in the G1 stage compared with the control group in the cell cycle arrest protocol as well as the notable overexpression of p53 and p21 genes suggested that these nanocapsules arrested cell cycle in MCF-7 cells.

UBE2C Overexpression Aggravates Patient Outcome by Promoting Estrogen-Dependent/Independent Cell Proliferation in Early Hormone Receptor-Positive and HER2-Negative Breast Cancer

We previously showed that UBE2C mRNA expression is significantly associated with poor prognosis only in patients with hormone receptor (HR)+/human epidermal growth factor receptor 2 (HER2)– breast cancer. In this study, we further reanalyzed the correlation between UBE2C mRNA expression and clinical outcomes in patients with HR+/HER2– breast cancer, and we investigated the molecular mechanism underlying the role of UBE2C modulation in disease progression in this subgroup of patients. Univariate and multivariate analyses showed that high UBE2C expression was associated with significantly shorter survival of breast cancer patients with pN0 and pN1 tumors but not pN2/N3 tumors (P < 0.05). In vitro functional experiments in HR+/HER2– breast cancer cells showed that UBE2C expression is a tumorigenic factor, and that estrogen upregulated UBE2C mRNA and protein by directly binding to the UBE2C promoter region. UBE2C knockdown inhibited cell proliferation by affecting cell cycle progression, and UBE2C overexpression was associated with estrogen-independent growth. UBE2C depletion markedly increased the cytotoxicity of tamoxifen by inducing apoptosis. The present findings suggest that UBE2C overexpression is correlated with relapse and promotes estrogen-dependent/independent proliferation in early HR+/HER2– breast cancer.

17β-estradiol at low concentrations attenuates the efficacy of tamoxifen in breast cancer therapy

Tamoxifen has been applied widely in the treatment of estrogen receptor (ER)-positive breast cancer. The impact of low concentrations of 17β-estradiol (E2) (a pervasive environmental pollutant) on its effectiveness was studied in vitro using an MCF-7 cell line. Cell proliferation, migration, invasion, and apoptosis were studied along with cell cycle progression, reactive oxygen species generation and mitochondrial membrane potentials repression. The signaling pathways involved were identified. Typical concentrations of E2 in the environment (10−10 to 10−8 M) were observed to promote cell growth and protect MCF-7 cells from tamoxifen's cytotoxicity. Cell migration, invasion, cell cycle progression and apoptosis all involved in reducing tamoxifen's cytotoxicity. E2 at environmental concentrations induced PI3K/Akt and MAPK/ERK signal transduction through the estrogen receptor pathways to affect cell proliferation. Taken together, the results explain how E2 in the environment may attenuate the efficacy of tamoxifen in ER-positive breast cancer therapy. They provide considerable support for E2's adverse effects on human health and cancer management.

Development of a dual fluorescence system for simultaneous detection of two cell populations in a 3D coculture

• A dual fluorescent system (green + red) was developed for cell detection in coculture. • A polymer-based 3D platform was used for high throughput drug screening. • Cytotoxicity of tamoxifen on breast cancer MCF-7 cells was evaluated. • Down-regulation effects of doxorubicin on survivin expression were assessed. A dual fluorescence system consisting of enhanced green fluorescent protein (EGFP) and red fluorescent protein (Ds-red) signals was developed for monitoring cellular activities. For system validation, MCF-7 breast cancer cells were engineered to express EGFP and Ds-red, respectively. Cells were cultured in three-dimensional (3D) polymer-based scaffolds on a 40-microbioreactor platform (40-MBR) with real-time monitoring of fluorescence signals without significant background fluorescence noises from culture medium and 3D scaffold. The fluorescence kinetics study showed that both EGFP and Ds-red signals in the 3D coculture could be simultaneously detected without interference with each other. The cytotoxicity study of tamoxifen and the survivin promoter test of doxorubicin demonstrated that both EGFP and Ds-red signals provided high sensitivity to evaluate drug effectiveness. These studies suggested that the novel dual fluorescence system could provide a simple, real-time, and rapid cell detection in 3D coculture assay, which could be used for in vitro high throughput drug screening.

Impacts of in vivo and in vitro exposures to tamoxifen: Comparative effects on human cells and marine organisms

Tamoxifen (TAM) is a first generation-SERM administered for hormone receptor-positive (HER+) breast cancer in both pre- and post-menopausal patients and may undergo metabolic activation in organisms that share similar receptors and thus face comparable mechanisms of response. The present study aimed to assess whether environmental trace concentrations of TAM are bioavailable to the filter feeder M. galloprovincialis (100 ng L−1) and to the deposit feeder N. diversicolor (0.5, 10, 25 and 100 ng L−1) after 14 days of exposure. Behavioural impairment (burrowing kinetic), neurotoxicity (AChE activity), endocrine disruption by alkali-labile phosphate (ALP) content, oxidative stress (SOD, CAT, GPXs activities), biotransformation (GST activity), oxidative damage (LPO) and genotoxicity (DNA damage) were assessed. Moreover, this study also pertained to compare TAM cytotoxicity effects to mussels and targeted human (i.e. immortalized retinal pigment epithelium – RPE; and human transformed endothelial cells – HeLa) cell lines, in a range of concentrations from 0.5 ng L−1 to 50 μg L−1. In polychaetes N. diversicolor, TAM exerted remarkable oxidative stress and damage at the lowest concentration (0.5 ng L−1), whereas significant genotoxicity was reported at the highest exposure level (100 ng L−1). In mussels M. galloprovincialis, 100 ng L−1 TAM caused endocrine disruption in males, neurotoxicity, and an induction in GST activity and LPO byproducts in gills, corroborating in genotoxicity over the exposure days. Although cytotoxicity assays conducted with mussel haemocytes following in vivo exposure was not effective, in vitro exposure showed to be a feasible alternative, with comparable sensitivity to human cell line (HeLa).

Overexpression of the erythropoietin receptor in RAMA 37 breast cancer cells alters cell growth and sensitivity to tamoxifen.

Erythropoietin (EPO) is the main regulator of erythropoiesis, and its receptor (EPOR) is expressed in various tissues, including tumors. Expression of EPOR in breast cancer tissue has been shown to correlate with expression of the estrogen receptor (ER). However, EPOR promotes proliferation in an EPO-independent manner. In patients with breast cancer, EPOR is associated with impaired tamoxifen response in ER-positive tumors, but not in ER-negative tumors. Furthermore, a positive correlation between EPOR/ER status and increased local cancer recurrence has been demonstrated, and EPOR expression is associated with G-protein coupled ER (GPER). Herein, we assessed the effects of EPOR on cell physiology and tamoxifen response in the absence of EPO stimulation using two cell lines that differ only in their EPOR expression status: RAMA37 cells (low EPOR expression) and RAMA37-28 cells (high EPOR expression). Alterations in cell growth, morphology, response to tamoxifen cytotoxicity, and EPOR-activated signal transduction were observed. RAMA37 cells showed higher proliferation capacity without tamoxifen treatment, while RAMA37-28 cells were more resistant to tamoxifen and proliferated more rapidly in the presence of tamoxifen. EPOR overexpression induced cell-morphology changes upon tamoxifen treatment, which resulted in the production of cell protrusions and subsequent cell death. Short-term treatment with tamoxifen (6h) prompted RAMA37 cells to acquired longer protrusions than RAMA37-28 cells, which indicated a pre-apoptotic stage. Furthermore, prolonged treatment with tamoxifen (72h) caused a greater reduction in RAMA37 cell numbers, which indicated a higher rate of cell death. RAMA37-28 cells showed prolonged activation of AKT signaling. We propose sustained AKT phosphorylation in EPOR-overexpressing cells as a mechanism that can lead to EPOR-induced tamoxifen resistance.

Chitosan-palmitic acid based polymeric micelles as promising carrier for circumventing pharmacokinetic and drug delivery concerns of tamoxifen

Being a BCS class II drug and a good substrate for microsomal enzymes, tamoxifen (TAM) offers a scope for research owing to poor aqueous solubility and compromised bioavailability. The present study designs a novel copolymer derived from palmitic acid and chitosan, and evaluate the derived TAM-loaded micelles for various delivery attributes. The nanometric micellar carriers not only substantially loaded the drug, but also controlled the rate of release of TAM. The designed nanocarrier significantly enhanced the cytotoxicity of TAM on MCF-7 cancer cells. The developed system was designed for intravenous route and was observed to be substantially haemo-compatible with an enhancement of approx. 5 times in AUC vis-a-vis plain drug. The findings employing new polymer-based carrier are promising in nature for the better delivery of similar drugs.

Development of erythropoietin receptor-targeted drug delivery system against breast cancer using tamoxifen-loaded nanostructured lipid carriers.

Tamoxifen (TAM) has been used in the treatment of breast cancers and is supplemented with erythropoietin (EPO) to alleviate the cancer-related anemia. The purported deleterious effects caused by the use of EPO with chemotherapeutic agents in the treatment of cancer-related anemia vary across studies and remain controversial. The use of nanoparticles as a drug delivery system has the potential to improve the specificity of anticancer drugs. In this study, we simultaneously incorporated two pharmacological active ingredients in one nanocarrier to develop EPO-conjugated TAM-loaded lipid nanoparticles (EPO-TAMNLC), a targeted delivery system, to enhance the cytotoxic activity while reducing the side effects of the ingredients. The effect of temperature in modulating the thermodynamic parameters associated with the binding of EPO and TAMNLC was assessed using isothermal titration calorimetry, while the unfolding of EPO structure was determined using fluorescence-quenching approach. The association efficiency of EPO and TAMNLC was 55.43%. Unlike binding of albumin to TAMNLC, the binding of EPO to TAMNLC occurred through endothermic and entropy-driven reaction. The EPO-TAMNLC formulation was stable because of the hydrophobic interaction and the high free energy, suggesting the spontaneity of the interactions between EPO and TAMNLC. The EPO-TAMNLC enhanced the in vitro cytotoxicity of TAM to MCF-7 cells. The EPO surface-functionalized TAMNLC could sequentially deliver EPO and TAM as well as improving site-specific delivery of these therapeutic compounds.

Salinomycin co-treatment enhances tamoxifen cytotoxicity in luminal A breast tumor cells by facilitating lysosomal degradation of receptor tyrosine kinases.

Luminal A breast cancer is the most common breast cancer subtype which is usually treated with selective estrogen receptor modulators (SERMS) like tamoxifen. Nevertheless, one third of estrogen receptor positive breast cancer patients initially do not respond to endocrine therapy and about 40% of luminal A breast tumors recur in five years. In this study, we investigated an alternative treatment approach by combining tamoxifen and salinomycin in luminal A breast cancer cell lines. We have found that salinomycin induces an additional cytotoxic effect by inhibiting the ligand independent activation of ERα. Thereby salinomycin increases the intracellular calcium level. This leads to a premature fusion of endosomes with lysosomes and thus to the degradation of Egfr family members. Since this process is essential for luminal A breast cancer cells to circumvent tamoxifen treatment, the combination of both drugs induces cytotoxicity in tamoxifen sensitive as well as resistant luminal A breast cancer cell lines.

ALOE EMODIN ENHANCES TAMOXIFEN CYTOTOXICITY EFFECT ON ERa-POSITIVE BREAST CANCER CELLS, MCF-7, THROUGH DOWNREGULATION OF MEK1 AND MEK2

The positive response to tamoxifen in ERa-positive breast cancer patients is usually of a short duration as manyof the patients eventually develop resistance. Our preliminary results show that aloe emodin extracted fromthe leaves of the Aloe barbadensis Miller demonstrated a cytotoxicity that is selective to ERa-positive breastcancer cells (MCF-7), but not to ERa-negative breast cancer cells (MDA-MB-231) and to the control cells (MCF-10A). The objective of this study was to test the hypothesis that aloe emodin may enhance the response ofMCF-7 cells to treatment with tamoxifen. MCF-7 cells were treated with aloe emodin alone, tamoxifen aloneor a combination of emodin and tamoxifen, at their respective IC50 concentrations and at different time pointsof 24 hours, 48 hours and 72 hours. The respective IC50s were the concentrations of aloe emodin and tamoxifenrequired to achieve 50% inhibition of the cells in the study. Cell viability and apoptosis were determined usingtrypan blue exclusion and DNA fragmentation assays, respectively. The involvement of RAS/MEKs/ERKs genesof MAPK signalling pathways with aloe emodin was determined using QuantiGene 2.0 Plex assay. Data wasevaluated using the one-way ANOVA test. Our findings showed that aloe emodin enhanced the cytotoxicity oftamoxifen on MCF-7 cells through apoptosis by downregulation of MEK1/2 genes. Our research may provide arational basis for further in vivo studies to verify the efficacy of a combination of aloe emodin and tamoxifenon the viability of ERa-positive-breast cancer cells.

Prognostic value of protein inhibitor of activated STAT3 in breast cancer patients receiving hormone therapy.

BackgroundDeregulated signal transducer and activator of transcription 3 (STAT3) signaling has been well documented in certain cancers. Alterations in specific negative regulators, such as protein inhibitor of activated STAT3 (PIAS3), may contribute to cancer development.MethodsThe expression of total PIAS3 was determined in 100 paired cancerous and non-cancerous breast tissues by immunoblotting and was statistically analyzed along with the clinicopathological characteristics and overall survival of the patients. XTT, immunoblotting, and chromatin immunoprecipitation (Chip) were used to examine the biological effect of PIAS3 in breast cancer cells.ResultsHormone therapy failed to improve the overall survival in patients presenting with increased PIAS3 expression. Ectopic PIAS3 overexpression increased the proliferation and expression of cyclin D1 in estrogen receptor (ER)-positive MCF-7 and T47D cells, but decreased those in ER-negative MDA-MB-231 and SKBR3 cells. Furthermore, PIAS3 overexpression attenuated cytotoxicity of tamoxifen and increased proliferation and cyclin D1 expression in MCF-7 cells. PIAS3 also decreased the binding of itself on the cyclin D1 promoter and this decreased binding was not affected by tamoxifen.ConclusionPIAS3 may serve as a biomarker for predicting hormone therapy stratification, although it is limited to those breast cancer patients receiving hormone therapyElectronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2063-1) contains supplementary material, which is available to authorized users.

Deciphering the Nongenomic, Mitochondrial Toxicity of Tamoxifens As Determined by Cell Metabolism and Redox Activity.

Tamoxifen is not only considered a very potent chemotherapeutic adjuvant for estrogen receptor positive breast cancers but also a very good chemo-preventive drug. Recently, there has been a rising amount of evidence for a nongenomic cytotoxicity of tamoxifen, even in estrogen receptor negative cells, which has greatly confounded researchers. Clinically, the side effects of tamoxifen can be very serious, ranging from liver steatosis to cirrhosis, tumorigenesis, or onset of porphyrias. Herein, we deciphered the nongenomic, mitochondrial cytotoxicity of tamoxifen in estrogen receptor positive MCF7 versus triple-negative MDA-MB-231 cells, employing the mitochondrial complex III quinoloxidizing-center inhibitor myxothiazol. We showed a role for hydroxyl-radical-mediated lipid peroxidation, catalyzed by iron, stemming from the redox interactions of tamoxifen quinoid metabolites with complex III, resulting in Fenton-capable reduced quinones. The role of tamoxifen semiquinone species in mitochondrial toxicity was also shown together with evidence of mitochondrial DNA damage. Tamoxifen caused an overall metabolic (respiratory and glycolytic) rate decrease in the Pasteur type MCF cells, while in the Warburg type MDA-MB-231 cells the respiratory rate was not significantly affected and the glycolytiv rate was significantly boosted. The nongenomic cytotoxicity of tamoxifens was hence associated with the metabolic phenotype and redox activity of the cells, as in the present paradigm of Pasteur MCF7s versus Warburg MDA-MB-231 cells. Our present findings call for caution in the use of the drugs, especially as a chemopreventive and/or in cases of iron overload diseases.