Combined Effects of Annexin A5 Overexpression, 5-Fluorouracil Treatment, and Irradiation on Cell Viability of Caski Cervical Cancer Cell Line.

Cervical cancer is the fourth leading cause of cancer deaths in women globally. Combining gene therapy with chemo- and radiotherapy may improve cervical cancer treatment outcomes. This study evaluated the effects of Annexin A5(ANXA5) overexpression alongside 5-fluorouracil (5-FU) and irradiation on the viability of CaSki cervical squamous cell carcinoma (SCC) cells. pAdenoVator-CMV-ANXA5-IRES-GFP-plasmid and mock plasmid were transfected into CaSki cells using calcium-phosphate. Seventy-two hours post-transfection, GFP expression was quantified by fluorescence microscopy and flow cytometry to evaluate transfection efficiency. ANXA5 overexpression was confirmed via qPCR. Twenty-four hours post-transfection, cells received a single dose of 8 Gy and were treated with 1 and 2 µg/ml of 5-FU (IC50 = 2.783 µg/ml). Cell viability, apoptosis, cell cycle stage, and Bcl-2 and Bax gene expression were assessed via MTT, annexin V/7-AAD, PI staining, and qPCR assays, respectively. ANXA5 was overexpressed 31.5-fold compared to control (p < 0.0001). MTT assays showed ANXA5 overexpression dose-dependently reduced CaSki cell viability (p < 0.001). IC50 of 5-FU was reduced from 2.783 µg/mL to 1.794 µg/mL when combined with ANXA5 overexpression. Additive effects on cell death were observed for ANXA5 plus 5-FU or irradiation versus ANXA5 alone. Apoptosis assays indicated combinatorial treatment increased CaSki cell apoptosis over ANXA5 alone. Cell cycle analysis revealed ANXA5 arrested cell cycle at G1/S phases; the percentage of cells in the S phase further rose with combination treatment. Finally, combination therapy significantly decreased Bcl-2 expression and increased Bax versus control (p < 0.001). Altogether, ANXA5 overexpression alongside 5-FU and irradiation may improve cervical squamous cell carcinoma (SCC) treatment efficacy. Further, in vivo investigations are warranted to confirm these in vitro results.

Applications of single-cell omics for chimeric antigen receptor T cell therapy.

Chimeric antigen receptor (CAR) T cell therapy is a promising cancer treatment modality. The breakthroughs in CAR T cell therapy were, in part, possible with the help of cell analysis methods, such as single-cell analysis. Bulk analyses have provided invaluable information regarding the complex molecular dynamics of CAR T cells, but their results are an average of thousands of signals in CAR T or tumour cells. Since cancer is a heterogeneous disease where each minute detail of a subclone could change the outcome of the treatment, single-cell analysis could prove to be a powerful instrument in deciphering the secrets of tumour microenvironment for cancer immunotherapy. With the recent studies in all aspects of adoptive cell therapy making use of single-cell analysis, a comprehensive review of the recent preclinical and clinical findings in CAR T cell therapy was needed. Here, we categorized and summarized the key points of the studies in which single-cell analysis provided insights into the genomics, epigenomics, transcriptomics and proteomics as well as their respective multi-omics of CAR T cell therapy.

Outsmarting trogocytosis to boost CAR NK/T cell therapy.

Chimeric antigen receptor (CAR) NK and T cell therapy are promising immunotherapeutic approaches for the treatment of cancer. However, the efficacy of CAR NK/T cell therapy is often hindered by various factors, including the phenomenon of trogocytosis, which involves the bidirectional exchange of membrane fragments between cells. In this review, we explore the role of trogocytosis in CAR NK/T cell therapy and highlight potential strategies for its modulation to improve therapeutic efficacy. We provide an in-depth analysis of trogocytosis as it relates to the fate and function of NK and T cells, focusing on its effects on cell activation, cytotoxicity, and antigen presentation. We discuss how trogocytosis can mediate transient antigen loss on cancer cells, thereby negatively affecting the effector function of CAR NK/T cells. Additionally, we address the phenomenon of fratricide and trogocytosis-associated exhaustion, which can limit the persistence and effectiveness of CAR-expressing cells. Furthermore, we explore how trogocytosis can impact CAR NK/T cell functionality, including the acquisition of target molecules and the modulation of signaling pathways. To overcome the negative effects of trogocytosis on cellular immunotherapy, we propose innovative approaches to modulate trogocytosis and augment CAR NK/T cell therapy. These strategies encompass targeting trogocytosis-related molecules, engineering CAR NK/T cells to resist trogocytosis-induced exhaustion and leveraging trogocytosis to enhance the function of CAR-expressing cells. By overcoming the limitations imposed by trogocytosis, it may be possible to unleash the full potential of CAR NK/T therapy against cancer. The knowledge and strategies presented in this review will guide future research and development, leading to improved therapeutic outcomes in the field of immunotherapy.

The Role of the IL-33/ST2 Immune Pathway in Autoimmunity: New Insights and Perspectives.

Interleukin (IL)-33, a member of IL-1 cytokine family, is produced by various immune cells and acts as an alarm to alert the immune system after epithelial or endothelial cell damage during cell necrosis, infection, stress, and trauma. The biological functions of IL-33 largely depend on its ligation to the corresponding receptor, suppression of tumorigenicity 2 (ST2). The pathogenic roles of this cytokine have been implicated in several disorders, including allergic disease, cardiovascular disease, autoimmune disease, infectious disease, and cancers. However, alerted levels of IL-33 may result in either disease amelioration or progression. Genetic variations of IL33 gene may confer protective or susceptibility risk in the onset of autoimmune diseases. The purpose of this review is to discuss the involvement of IL-33 and ST2 in the pathogenesis of a variety of autoimmune disorders, such as autoimmune rheumatic, neurodegenerative, and endocrine diseases.