Dual silencing of tumor-intrinsic VISTA and CTLA-4 stimulates T-cell mediated immune responses and inhibits MCF7 breast cancer development.

BACKGROUND: As inhibitory immune checkpoint molecules, cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) and V-domain Ig suppressor of T-cell activation (VISTA) can be expressed in tumoral cells and facilitate immune evasion of tumoral cells. Herein, we studied the significance of tumor-intrinsic CTLA-4 and VISTA silencing in tumor development and inflammatory factors expression in a co-culture system with MCF7 and T-cells. METHODS: MCF7 cells were transfected with 60 pmol of CTLA-siRNA, VISTA-siRNA, and dual VISTA-/CTLA-4-siRNA. The MTT assay was performed to study the effect of CTLA-4 and VISTA knockdown on the viability of MCF7 cells. Colony formation and wound-healing assays were performed to investigate the effect of CTLA-4 and VISTA silencing on the clonogenicity and migration of MCF7 cells. Flow cytometry was used to study the significance of CTLA-4 and VISTA knockdown on the apoptosis and cell cycle of MCF7 cells. Also, a co-culture system with MCF7 and T-cells was developed to study the expression levels of IL-2, IFN-γ, TNF-α, TGF-ß, and IL-10 following CTLA-4 and VISTA knockdown. The expression levels of caspase3, Bax, Bcl2, and MMP-9 were also investigated using quantitative real-time PCR. Finally, the TCGA Breast Cancer and GSE45827 datasets were analyzed to study the potential prognostic values of VISTA and CTLA-4, their expression difference in luminal A breast cancer and non-tumoral tissues, and their correlation in luminal A breast cancer tissues. RESULTS: Combined knockdown of tumor-intrinsic VISTA and CTLA-4 is superior in upregulating IL-2, IFN-γ, and TNF-α, downregulating TGF-ß and IL-10 in T lymphocytes. Also, the combined silencing arrests the cell cycle at the sub-G1 phase, decreases migration, inhibits clonogenicity, and reduces cell viability of MCF7 cells. This combined treatment upregulates caspase 9 and BAX and downregulates MMP-9 in MCF7 cells. Our in-silico results have demonstrated a significant positive correlation between CTLA-4 and VISTA in luminal A breast cancer. CONCLUSION: The additive effect of the combined knockdown of tumor-intrinsic VISTA and CTLA-4 can substantially upregulate pro-inflammatory factors, downregulate anti-inflammatory factors, and inhibit tumor development in MCF7 cells. The significant positive correlation between VISTA and CTLA-4 in luminal A breast cancer might support the idea that a network of inhibitory immune checkpoint molecules regulates anti-tumoral immune responses; thus, combinational immune checkpoint molecules blockade can be suggested.

MicroRNAs and their vital role in apoptosis in hepatocellular carcinoma: miRNA-based diagnostic and treatment methods.

Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies with high invasive and metastatic capability. Although significant advances have been made in the treatment of HCC, the overall survival rate of patients is still low. It is essential to explore accurate biomarkers for early diagnosis and prognosis along with therapeutic procedures to increase the survival rate of these patients. Anticancer therapies can contribute to induce apoptosis for the elimination of cancerous cells. However, dysregulated apoptosis and proliferation signaling pathways lead to treatment resistance, a significant challenge in improving efficient therapies. MiRNAs, short non-coding RNAs, play crucial roles in the progression of HCC, which regulate gene expression through post-transcriptional inhibition and targeting mRNA degradation in cancers. Dysregulated expression of multiple miRNAs is associated with numerous biological processes, including cell proliferation, apoptosis, invasion and metastasis, epithelial-mesenchymal transition (EMT), angiogenesis, and drug resistance in HCC. This review summarizes the role and potential efficacy of miRNAs in promoting and inhibiting cell proliferation and apoptosis in HCC, as well as the role of miRNAs in therapy resistance in HCC.

Combination of B7H6-siRNA and temozolomide synergistically reduces stemness and migration properties of glioblastoma cancer cells.

Glioblastoma multiforme (GBM) is among the malignant brain tumors of the central nervous system (CNS). The survival of this disease is about 14 months after diagnosis. To date, temozolomide is known as first-line therapy for glioma. Drug resistance and severe side effects against this drug are important obstacles to the effective treatment of this cancer. Small interfering RNA (siRNA) can adjust the expression of several genes and is used as a new method of gene therapy. Recent studies have shown that siRNAs can increase the sensitivity of cancer cells to chemotherapy drugs. This study aimed to understand the potential role and molecular mechanism of the combination therapy of B7H6-siRNA and temozolomide in glioblastoma cancer. U87 cells were treated with B7H6-siRNA and temozolomide, separately and in combination. Cell viability, stemness, cell migration, and apoptosis were measured. The results of this work presented the synergistic effect of B7H6-siRNA and temozolomide in inhibiting the cancerous features of the U87 cell line. Down-regulating B7H6-siRNA expression inhibited the cell viability of U87 glioblastoma cancer cells and increased their sensitivity to temozolomide. In addition, a noteworthy decrease in cell migration ability and stemness, an increase in apoptosis were observed in the combined groups compared to B7H6-siRNA and temozolomide individually. According to the results, a combination of B7H6-siRNA and temozolomide can be a promising strategy in glioblastoma cancer therapy.

Sodium metabisulfite as a cytotoxic food additive induces apoptosis in HFFF2 cells.

Sodium metabisulfite (SMB), an antioxidant agent, is extensively used as a preservative in food industry. The current study was aimed to clarify its potential toxic effects on human fetal foreskin fibroblasts (HFFF2) cells, in vitro. Subsequently, MTT results illustrated that exposure to SMB significantly (p < 0.0001) decreased HFFF2 cell viability in a dose-dependent manner, and the concentration of 25 µM reduced cell survival rates to 50% as the half-maximal inhibitory concentration of SMB. It was further shown that SMB exerted this cytotoxic effect on HFFF2 cells through apoptosis induction. qRT-PCR and western blotting results showed that treatment of HFFF2 cells with this food additive led to significant upregulation of Bax, caspase 8, and caspase 9 pro-apoptotic genes and downregulation of Bcl-2 expression as a pro-survival agent. Furthermore, SMB remarkably increased caspase 3 levels and promoted its activation through cleavage in treated cells. Besides, exposure to SMB increased ROS levels and activated autophagy in treated cells, which are considered as the other indicators for cell damage. Taken together, our findings suggested that SMB could exert remarkable toxic effects on human normal cells through multiple mechanisms, including apoptosis activation, and its widespread usage in food safety should be reconsidered.

Photodynamic therapy for cancer: Role of natural products.

Photodynamic therapy (PDT) is a promising modality for the treatment of cancer. PDT involves administering a photosensitizing dye, i.e. photosensitizer, that selectively accumulates in tumors, and shining a light source on the lesion with a wavelength matching the absorption spectrum of the photosensitizer, that exerts a cytotoxic effect after excitation. The reactive oxygen species produced during PDT are responsible for the oxidation of biomolecules, which in turn cause cell death and the necrosis of malignant tissue. PDT is a multi-factorial process that generally involves apoptotic death of the tumor cells, degeneration of the tumor vasculature, stimulation of anti-tumor immune response, and induction of inflammatory reactions in the illuminated lesion. Numerous compounds with photosensitizing activity have been introduced commercially. Although many papers have been published with regard to PDT in the last decade, there has been relatively little focus on natural medicinal plant extracts and compounds derived therefrom. Herbal plants and their extracts are natural substances, and in comparison with synthetic chemicals are considered "green". This review focuses on the different mechanisms of PDT and discusses the role of various plant extracts and natural compounds either alone or in combination for carrying out PDT on different types of cancers.