Long Non-Coding Ribonucleic Acid Forkhead Box P4-Antisense RNA 1 Targets microRNA-655-3p to Regulate the Proliferation, Transfer, and Invasion of Mammary Cancer Cells

LncRNA FOXP4-AS1 expresses at a higher level in gastric carcinoma cells and can promote proliferation and other biological behaviors. However, the effect of FOXP4-AS1 on mammary cancer cells has not yet been elucidated. Therefore, this article explores the influence of lncRNA FOXP4-AS1 on the proliferation and other biological processes of mammary cancer MDA-MB-231 cells via its regulation of miRNA-655-3p. Firstly, nanoPCR was used to quantify the expression of FOXP4-AS1 and miRNA-655-3p in mammary cancer tissues and adjacent tissues. Compared to the adjacent tissues, the expression level of FOXP4-AS1 in mammary cancer tissue was significantly increased, while that of miRNA-655-3p was substantially reduced. Then, si-FOXP4-AS1, miRNA-655-3p mimics, si-FOXP4-AS1 + anti-miRNA-655-3p were transfected into human mammary cancer MDA-MB-231 cells. The transfection of si-FOXP4-AS1 or miRNA-655-3p mimics considerably reduced cell viability and the protein levels of Ki-67 and MMPs. The transfection of si-FOXP4-AS1 or miRNA-655-3p mimics could reduce cell migration and invasion. The dual-luciferase reporter assays revealed that FOXP4-AS1 could target miRNA-655-3p. The co-transfection of si-FOXP4-AS1 and anti-miRNA-655-3p increased cell viability, migration, and invasion; the same co-transfection also elevated the protein levels of Ki-67 and MMPs. In conclusion, this study suggests that knocking down FOXP4-AS1’s expression can reduce mammary cancer cells’ ability to proliferate and execute other biological processes by targeting the expression of miRNA-655-3p.

Toxoplasma gondii destroys Her2/Neu-expressing mammary cancer cells in vitro using a continuous feed medium approach

Introduction: Toxoplasma gondii is an opportunistic protozoan and can be grown using several human cell lines. Breast cancer is the second leading cause of cancer death in women. Her2/Neu-expressing mammary cancer cell lines called TUBO can be grown in vitro. In recent years, protozoan parasites have become popular means of use in cancer therapy research. In this study, we analyzed whether T. gondii tachyzoites can destroy TUBO cells using a novel continuous feed medium approach. Methodology: Two sets of flasks (each containing four groups) containing TUBO cells were inoculated with T. gondii Ankara strain tachyzoites. First set containing 5×106 TUBO cells were inoculated with TUBO-tachyzoite ratios of 1:2, 1:1, 2:1, and 4:1 and second set containing 1×106 TUBO cells were inoculated with TUBO-tachyzoite ratios of 10:1, 100:1, 1000:1, and 2000:1. Thereafter, culture supernatants were harvested at various days until TUBO cells were destroyed and tachyzoites were counted. Results: In the first and second sets of flasks, TUBO cells were destroyed between days 8 to 12 and 12 to 25, respectively. In addition, the amount of tachyzoites increased 7- 43 and 595 to 112500 times in the first and second set of flasks, respectively. Conclusions: These results show that T. gondii tachyzoites successfully destroy Her2/Neu-expressing mammary cancer cells using a continuous feed medium approach. Although this idea may be too premature for the moment, the approach defined herein may support future researchers investigating the relationship between cancer and parasites which can make important progress toward saving cancer patient lives.

In Vitro Oncolytic activity of non-virulent Newcastle Disease Virus LaSota Strain against Mouse mammary adenocarcinoma

Newcastle disease virus (NDV) is a wide-spectrum anti-tumor agent. The oncolytic selectivity of NDV, a family of Paramyxoviridae, depends on the differential type of inducing different death pathways. This work was conducted to further understand the oncolytic effect of LaSota strain. A mouse breast cancer model (Murine mammary adenocarcinoma cell line AMN3) was used in this study. Methyl Thiazolyl Tetrazolium (MTT) viability assay tested different NDV multiplicity of infection (MOI) values on mouse mammary adenocarcinoma cells incubated for 72 hours post-infection. The IC50 values and anti-tumor activity of LaSota strain against AMN3 cell line were determined. Following Hematoxylin and Eosin Stain, we examined the morphological modifications of IC50 along with 10 MOI values of NDV. The induction of NDV apoptosis in AMN3 cells was investigated using the technique of staining acridine orange and propidium iodide (AO / PI). Immunocytochemistry assay was performed using anti-NDV mAbs and caspases 8 and 9 to study NDV replication and apoptosis induction mechanisms. The lentogenic LaSota NDV strain, a live vaccine, demonstrated the oncolytic effect on mammary cancer cells of the AMN3 mouse and showed that LaSota strain triggered a dose-dependent increase in infected cells’ apoptosis relative to untreated mammary cancer cells. The immunocytochemistry study showed that NDV infected cells were positive for virus infection and that caspase9 in mouse mammary cancer cells after LaSota strain infection was significantly enhanced compared to caspase 8. In conclusions, NDV LaSota strain had oncolytic effects by destroying tumor cells and triggering the intrinsic apoptosis pathways in mouse mammary cancer cells. However, the mechanisms of the in vivo anti-tumor activity need to be better understood.

An epigenetic screening determines BET proteins as targets to suppress self-renewal and tumorigenicity in canine mammary cancer cells

Targeting self-renewal and tumorigenicity has been proposed as a potential strategy against cancer stem cells (CSCs). Epigenetic proteins are key modulators of gene expression and cancer development contributing to regulation and maintenance of self-renewal and tumorigenicity. Here, we have screened a small-molecule epigenetic inhibitor library using 3D in vitro models in order to determine potential epigenetic targets associated with self-renewal and tumorigenicity in Canine Mammary Cancer (CMC) cells. We identified inhibition of BET proteins as a promising strategy to inhibit CMC colonies and tumorspheres formation. Low doses of (+)-JQ1 were able to downregulate important genes associated to self-renewal pathways such as WNT, NOTCH, Hedgehog, PI3K/AKT/mTOR, EGF receptor and FGF receptor in CMC tumorspheres. In addition, we observed downregulation of ZEB2, a transcription factor important for the maintenance of self-renewal in canine mammary cancer cells. Furthermore, low doses of (+)-JQ1 were not cytotoxic in CMC cells cultured in 2D in vitro models but induced G2/M cell cycle arrest accompanied by upregulation of G2/M checkpoint-associated genes including BTG2 and CCNG2. Our work indicates the BET inhibition as a new strategy for canine mammary cancers by modulating the self-renewal phenotype in tumorigenic cells such as CSCs.

Melatonin decreases in vitro viability and migration of spheres derived from CF41.Mg canine mammary carcinoma cells

Background Mammary cancer is a common disease affecting female dogs, where approximately 50% of the cases are malignant. There is a subpopulation of cancer cells with stem cell-like features within the tumour microenvironment, which can form in vitro spheres, cell structures that grow in anchor-free conditions. This cell population shows resistance to conventional antitumor treatments explaining in part the recurrence of some type of cancers. It has been previously reported that spheres derived from CF41.Mg canine mammary carcinoma cells exhibit several stemness features. Melatonin has shown antitumor effects on cancer mammary cells; nevertheless, its effects have been poorly evaluated on canine mammary cancer stem-like cells. In this regard, it has described that melatonin decreases the expression of OCT-4 in CMT-U2229 mammary cancer cells, a transcription factor that participates in the modulation of self-renewal and drug resistance in cancer stem-like cells. The aim of this study was to compare the effects of melatonin on viability and migration of canine mammary carcinoma CF41.Mg-spheres, and CF41.Mg-parental cells. CF41.Mg cells were grown in DMEM high-glucose medium containing 10% bovine foetal serum. CF41.Mg-spheres were cultured in ultra-low attachment plates with serum-free DMEM/F12 containing several growth factors. Cell viability (MTS reduction) and migration (transwell) assays were conducted in presence of melatonin (0.01, 0.1 or 1 mM). Results Melatonin decreased cell viability at 1 mM (P < 0.05), with a significant reduction in spheres compared to parental cells at 24 and 48 h (P < 0.05). Cell migration was inhibited in response to non-cytotoxic concentration of melatonin (0.1 mM) (P < 0.05) in spheres and monolayer of cells, no significant differences were detected between both cell subtypes. Conclusions These results indicate that melatonin reduces viability and migration of CF41.Mg cells, where spheres exhibit greater sensitivity to the hormone. Thus, melatonin represents a valuable potential agent against mammary cancer cells, especially cancer stem-like cells.