The apoptotic effects of NK-92 cells stimulated with an anti-CD226 antibody on MDA-MB-231 triple-negative breast cancer cells.

Research on immunotherapy in breast cancer treatment has recently gained importance. In this context, natural killer (NK) cells have been shown to kill cancer cells without affecting normal cells. Our study used the NK-92 cells that were stimulated with anti-CD226 antibodies (sNK-92) to increase their activity to target MDA-MB-231 triple-negative breast cancer cells. MCF-12A normal breast cells were used as the control in all experiments. The cytotoxic effects of NK-92 and sNK-92 cells on MDA-MB-231 cells were investigated using lactate dehydrogenase tests. The sNK-92 cells were more cytotoxic than NK-92 cells on MDA-MB-231 cells. In contrast, a significant cytotoxic change was not observed in MCF-12A cells cocultured with NK-92 and sNK-92 cells. An increase in granzyme B levels after coculturing with sNK-92 cells was investigated using the granzyme B enzyme-linked immunosorbent assay. The sNK-92 cells secreted more granzyme B than NK-92 cells against MDA-MB-231 cells. This increase was not observed in MCF-12A, indicating that sNK-92 cells specifically target cancer cells. In addition, immunostaining was used to investigate the synthesis level of BAX, CASP3, and CASP9 proteins to determine whether the observed cytotoxic effect was due to apoptosis. These proteins were synthesized more in MDA-MB-231 cells cocultured with sNK-92 than with NK-92 cells. However, no increase in their synthesis was observed in normal breast cells cocultured with NK-92 and sNK-92 cells. In conclusion, NK-92 cells stimulated with anti-CD226 antibodies secrete more granzyme B, resulting in a greater cytotoxic effect by inducing programmed cell death (apoptosis). The fact that the observed effects on breast cancer cells were not observed in normal breast cells indicates that sNK-92 cells specifically target breast cancer cells. These results indicate the potential use of CD226-stimulated NK-92 cells in immunotherapy.

The effects of KIR2DL4 stimulated NK-92 cells on the apoptotic pathways of HER2 + /HER-breast cancer cells.

Natural killer (NK) cells are immune cells that have attracted significant attention due to their cytotoxic properties. They are believed to be highly effective in cancer therapy. In this study, anti-KIR2DL4 (Killer cell Immunoglobulin like Receptor, 2 Ig Domains and Long cytoplasmic tail 4) was used to stimulate the NK-92 activator receptor to increase their cytotoxicity on breast cancer cell lines. Unstimulated and stimulated NK-92 cells (sNK-92) were cocultured with breast cancer (MCF-7 and SK-BR-3) and normal breast (MCF-12A) cell lines at 1:1, 1:5, and 1:10 (Target:Effector) ratios. The most effective cell cytotoxicity ratio (1:10) was used in the immunostaining and western blot assays to evaluate apoptosis pathway proteins. The sNK-92 cells showed higher cytotoxic activity on breast cancer cells than NK-92 cells. sNK-92 cells had a selective significant cytotoxicity effect on MCF-7 and SK-BR-3 cells but not MCF-12A cells. While sNK-92 cells were effective at all cell concentrations, they were most effective at a 1:10 ratio. Immunostaining and western blots showed significantly higher BAX, caspase 3, and caspase 9 protein levels in all breast cancer cell groups cocultured with sNK-92 than with NK-92 cells. NK-92 cells stimulated with KIR2DL4 showed elevated cytotoxic activity. The cytotoxic activity of sNK-92 cells on breast cancer cells is via apoptosis pathways. However, their effect on normal breast cells is limited. While the obtained data contains only basic information, additional clinical studies are needed to provide a basis for a new treatment model.

Determination of Usnic Acid Responsive miRNAs in Breast Cancer Cell Lines.

OBJECTIVE: Breast Cancer (BC) is the most common type of cancer diagnosed in women. A common treatment strategy for BC is still not available because of its molecular heterogeneity and resistance is developed in most of the patients through the course of treatment. Therefore, alternative medicine resources as being novel treatment options are needed to be used for the treatment of BC. Usnic Acid (UA) that is one of the secondary metabolites of lichens used for different purposes in the field of medicine and its anti-proliferative effect has been shown in certain cancer types, suggesting its potential use for the treatment. METHODS: Anti-proliferative effect of UA in BC cells (MDA-MB-231, MCF-7, BT-474) was identified through MTT analysis. Microarray analysis was performed in cells treated with the effective concentration of UA and UA-responsive miRNAs were detected. Their targets and the pathways that they involve were determined using a miRNA target prediction tool. RESULTS: Microarray experiments showed that 67 miRNAs were specifically responsive to UA in MDA-MB-231 cells while 15 and 8 were specific to BT-474 and MCF-7 cells, respectively. The miRNA targets were mostly found to play role in Hedgehog signaling pathway. TGF-Beta, MAPK and apoptosis pathways were also the prominent ones according to the miRNA enrichment analysis. CONCLUSION: The current study is important as being the first study in the literature which aimed to explore the UA related miRNAs, their targets and molecular pathways that may have roles in the BC. The results of pathway enrichment analysis and anti-proliferative effects of UA support the idea that UA might be used as a potential alternative therapeutic agent for BC treatment.

Determination of Vulpinic Acid Effect on Apoptosis and mRNA Expression Levels in Breast Cancer Cell Lines.

OBJECTIVE: Breast cancer is one of the most common diseases among women worldwide and it is characterized by a high ratio of malignancy and metastasis and low rate of survival of patients. Due to limited treatment options, the discovery of alternative therapeutic agents and clarifying the molecular mechanism of breast cancer development may offer new hope for its treatment. Lichen secondary metabolites may be one of these therapeutic agents. METHODS: In this study, the effects of Vulpinic Acid (VA) lichen secondary metabolite on the cell viability and apoptosis of breast cancer cells and non-cancerous cell line were investigated. Quantitative polymerase chain reaction was also performed to determine changes in the expression of apoptosis-related genes at a molecular level. RESULTS: The results demonstrated that VA significantly inhibited the cell viability and induced apoptosis of human breast cancer cells. The highest rates of decreased growth were determined using the IC50 value of VA for 48h on MCF-7 breast cancer cell. Interestingly, VA treatment significantly reduced cell viability in all examined breast cancer cell lines compared to their non-cancerous human breast epithelial cell line. This is the first study on the investigation of the effects of VA on the molecular mechanisms associated with the expression of apoptosis-related genes in breast cancer cell lines. Results demonstrated that the gene expression of P53 genes was altered up to fourteen-fold levels in SK-BR-3 cell lines whereas it reached 2.5-fold in the MCF-12A cell line after treatment with VA. These observations support that VA induces apoptosis on the breast cancer cells compared with the non-cancerous human breast epithelial cell line. CONCLUSION: It is implicated that VA may be a promising novel molecule for the induction of apoptosis on breast cancer cells.

Detection of hepatitis C virus RNA in paraffin-embedded tissues from patients with non-Hodgkin's lymphoma.

The aim of this study is to detect the possible role of hepatitis C Virus (HCV) in lymphomagenesis. HCV-RNA and anti-HCV antibodies were studied in tissue and serum samples taken from patients with non-Hodgkin's Lymphoma (NHL). The prevalence of HCV, the clinical presentation of these cases, and association with histologic subtypes were determined. RT-PCR was used to detect the HCV-RNA in serum and tissue samples. The anti-HCV antibodies were tested with microparticle enzyme immunoassay. Immunohistochemistry with the ABC method was used to detect the HCV core protein in HCV-RNA(+) cases. RNA could be detected in 30 of 35 cases, and other tests were performed in these 30 samples. HCV-RNA was detected in 11 tissue samples (11/30, 37%). HCV core protein was studied in 10 of 11 HCV-RNA(+) cases, and 1-3% nuclear staining was found in only 2 samples. Serologically, HCV-RNA was detected in 7 of 30 samples (23.3%) and anti-HCV antibody was detected in 3 of 30 samples (10%). Detection of HCV-RNA in 37% of the lymphoma tissue samples suggests that HCV may have a role or is a contributing factor in the pathogenesis of lymphoma. The very low HCV core protein in lymphoma tissues may be due to the low viral load in lymphoid tissues and/or higher sensitivity of the PCR method. Detection of anti-HCV antibody in only three cases may be associated with undetectable levels of antibodies due to the immune deficiency in cases with NHL.