Systemic lupus erythematosus therapeutic strategy: From immunotherapy to gut microbiota modulation.

Systemic lupus erythematosus (SLE) is characterized by a systemic dysfunction of the innate and adaptive immune systems, leading to an attack on healthy tissues of the body. During the development of SLE, pathogenic features, such as the formation of autoantibodies to self-nuclear antigens, caused tissue damage including necrosis and fibrosis, with an increased expression of type Ⅰ interferon (IFN) regulated genes. Treatment of lupus with immunosuppressants and glucocorticoids, which are used as the standard therapy, is not effective enough and causes side effects. As an alternative, more effective immunotherapies have been developed, including monoclonal and bispecific antibodies that target B cells, T cells, co-stimulatory molecules, cytokines or their receptors, and signaling molecules. Encouraging results have been observed in clinical trials with some of these therapies. Furthermore, a chimeric antigen receptor T cells (CAR-T) therapy has emerged as the most effective, safe, and promising treatment option for SLE, as demonstrated by successful pilot studies. Additionally, emerging evidence suggests that gut microbiota dysbiosis may play a significant role in the severity of SLE, and the use of methods to normalize the gut microbiota, particularly fecal microbiota transplantation (FMT), opens up new opportunities for effective treatment of SLE.

Artificial Extracellular Vesicles Generated from T Cells Using Different Induction Techniques.

Cell therapy is at the forefront of biomedicine in oncology and regenerative medicine. However, there are still significant challenges to their wider clinical application such as limited efficacy, side effects, and logistical difficulties. One of the potential approaches that could overcome these problems is based on extracellular vesicles (EVs) as a cell-free therapy modality. One of the major obstacles in the translation of EVs into practice is their low yield of production, which is insufficient to achieve therapeutic amounts. Here, we evaluated two primary approaches of artificial vesicle induction in primary T cells and the SupT1 cell line-cytochalasin B as a chemical inducer and ultrasonication as a physical inducer. We found that both methods are capable of producing artificial vesicles, but cytochalasin B induction leads to vesicle yield compared to natural secretion, while ultrasonication leads to a three-fold increase in particle yield. Cytochalasin B induces the formation of vesicles full of cytoplasmic compartments without nuclear fraction, while ultrasonication induces the formation of particles rich in membranes and membrane-related components such as CD3 or HLAII proteins. The most effective approach for T-cell induction in terms of the number of vesicles seems to be the combination of anti-CD3/CD28 antibody activation with ultrasonication, which leads to a seven-fold yield increase in particles with a high content of functionally important proteins (CD3, granzyme B, and HLA II).

Asymmetric T-cell division: insights from cutting-edge experimental techniques and implications for immunotherapy.

Asymmetric cell division is a fundamental process conserved throughout evolution, employed by both prokaryotic and eukaryotic organisms. Its significance lies in its ability to govern cell fate and facilitate the generation of diverse cell types. Therefore, attaining a detailed mechanistic understanding of asymmetric cell division becomes essential for unraveling the complexities of cell fate determination in both healthy and pathological conditions. However, the role of asymmetric division in T-cell biology has only recently been unveiled. Here, we provide an overview of the T-cell asymmetric division field with the particular emphasis on experimental methods and models with the aim to guide the researchers in the selection of appropriate in vitro/in vivo models to study asymmetric division in T cells. We present a comprehensive investigation into the mechanisms governing the asymmetric division in various T-cell subsets underscoring the importance of the asymmetry in fate-determining factor segregation and transcriptional and epigenetic regulation. Furthermore, the intricate interplay of T-cell receptor signaling and the asymmetric division geometry are explored, shedding light on the spatial organization and the impact on cellular fate.

Immunotherapy Strategy for Systemic Autoimmune Diseases: Betting on CAR-T Cells and Antibodies.

Systemic autoimmune diseases (SAIDs), such as systemic lupus erythematosus (SLE), systemic sclerosis (SSc) and rheumatoid arthritis (RA), are fully related to the unregulated innate and adaptive immune systems involved in their pathogenesis. They have similar pathogenic characteristics, including the interferon signature, loss of tolerance to self-nuclear antigens, and enhanced tissue damage like necrosis and fibrosis. Glucocorticoids and immunosuppressants, which have limited specificity and are prone to tolerance, are used as the first-line therapy. A plethora of novel immunotherapies have been developed, including monoclonal and bispecific antibodies, and other biological agents to target cellular and soluble factors involved in disease pathogenesis, such as B cells, co-stimulatory molecules, cytokines or their receptors, and signaling molecules. Many of these have shown encouraging results in clinical trials. CAR-T cell therapy is considered the most promising technique for curing autoimmune diseases, with recent successes in the treatment of SLE and SSc. Here, we overview novel therapeutic approaches based on CAR-T cells and antibodies for targeting systemic autoimmune diseases.

Сytotoxic effect of CAR-T cells against modified MCF-7 breast cancer cell line.

The most often diagnosed and fatal malignancy in women is breast cancer. The International Agency for Research on Cancer (IARC) estimates that there are 2.26 million new cases of cancer in 2020. Adoptive cell therapy using T cells with chimeric antigen receptor shows potential for the treatment of solid tumors, such as breast cancer. In this work the effectiveness of CAR-T cells against monolayer and three-dimensional bioprinted tumor-like structures made of modified MCF-7 breast cancer cells was assessed. The cytokine profile of supernatants after co-cultivation of MCF-7 tumor cell models with CAR-T cells was also measured to reveal the inflammatory background associated with this interaction.

Evaluation of CAR-T Cells' Cytotoxicity against Modified Solid Tumor Cell Lines.

In recent years, adoptive cell therapy has gained a new perspective of application due to the development of technologies and the successful clinical use of CAR-T cells for the treatment of patients with malignant B-cell neoplasms. However, the efficacy of CAR-T therapy against solid tumor remains a major scientific and clinical challenge. In this work, we evaluated the cytotoxicity of 2nd generation CAR-T cells against modified solid tumors cell lines-lung adenocarcinoma cell line H522, prostate carcinoma PC-3M, breast carcinoma MDA-MB-231, and epidermoid carcinoma A431 cell lines transduced with lentiviruses encoding red fluorescent protein Katushka2S and the CD19 antigen. A correlation was demonstrated between an increase in the secretion of proinflammatory cytokines and a decrease in the confluence of tumor cells' monolayer. The proposed approach can potentially be applied to preliminarily assess CAR-T cell efficacy for the treatment of solid tumors and estimate the risks of developing cytokine release syndrome.

Recent Advances in the Development of Bioreactors for Manufacturing of Adoptive Cell Immunotherapies.

Harnessing the human immune system as a foundation for therapeutic technologies capable of recognizing and killing tumor cells has been the central objective of anti-cancer immunotherapy. In recent years, there has been an increasing interest in improving the effectiveness and accessibility of this technology to make it widely applicable for adoptive cell therapies (ACTs) such as chimeric antigen receptor T (CAR-T) cells, tumor infiltrating lymphocytes (TILs), dendritic cells (DCs), natural killer (NK) cells, and many other. Automated, scalable, cost-effective, and GMP-compliant bioreactors for production of ACTs are urgently needed. The primary efforts in the field of GMP bioreactors development are focused on closed and fully automated point-of-care (POC) systems. However, their clinical and industrial application has not yet reached full potential, as there are numerous obstacles associated with delicate balancing of the complex and often unpredictable cell biology with the need for precision and full process control. Here we provide a brief overview of the existing and most advanced systems for ACT manufacturing, including cell culture bags, G-Rex flasks, and bioreactors (rocking motion, stirred-flask, stirred-tank, hollow-fiber), as well as semi- and fully-automated closed bioreactor systems.

Knowns and Unknowns about CAR-T Cell Dysfunction.

Immunotherapy using chimeric antigen receptor (CAR) T cells is a promising option for cancer treatment. However, T cells and CAR-T cells frequently become dysfunctional in cancer, where numerous evasion mechanisms impair antitumor immunity. Cancer frequently exploits intrinsic T cell dysfunction mechanisms that evolved for the purpose of defending against autoimmunity. T cell exhaustion is the most studied type of T cell dysfunction. It is characterized by impaired proliferation and cytokine secretion and is often misdefined solely by the expression of the inhibitory receptors. Another type of dysfunction is T cell senescence, which occurs when T cells permanently arrest their cell cycle and proliferation while retaining cytotoxic capability. The first section of this review provides a broad overview of T cell dysfunctional states, including exhaustion and senescence; the second section is focused on the impact of T cell dysfunction on the CAR-T therapeutic potential. Finally, we discuss the recent efforts to mitigate CAR-T cell exhaustion, with an emphasis on epigenetic and transcriptional modulation.

Promising New Tools for Targeting p53 Mutant Cancers: Humoral and Cell-Based Immunotherapies.

Transcription factor and oncosuppressor protein p53 is considered as one of the most promising molecular targets that remains a high-hanging fruit in cancer therapy. TP53 gene encoding the p53 protein is known to be the most frequently mutated gene in human cancers. The loss of transcriptional functions caused by mutations in p53 protein leads to deactivation of intrinsic tumor suppressive responses associated with wild-type (WT) p53 and acquisition of new pro-oncogenic properties such as enhanced cell proliferation, metastasis and chemoresistance. Hotspot mutations of p53 are often immunogenic and elicit intratumoral T cell responses to mutant p53 neoantigens, thus suggesting this protein as an attractive candidate for targeted anti-cancer immunotherapies. In this review we discuss the possible use of p53 antigens as molecular targets in immunotherapy, including the application of T cell receptor mimic (TCRm) monoclonal antibodies (mAbs) as a novel powerful approach.

Adoptive Immunotherapy beyond CAR T-Cells.

Adoptive cell immunotherapy (ACT) is a vibrant field of cancer treatment that began progressive development in the 1980s. One of the most prominent and promising examples is chimeric antigen receptor (CAR) T-cell immunotherapy for the treatment of B-cell hematologic malignancies. Despite success in the treatment of B-cell lymphomas and leukemia, CAR T-cell therapy remains mostly ineffective for solid tumors. This is due to several reasons, such as the heterogeneity of the cellular composition in solid tumors, the need for directed migration and penetration of CAR T-cells against the pressure gradient in the tumor stroma, and the immunosuppressive microenvironment. To substantially improve the clinical efficacy of ACT against solid tumors, researchers might need to look closer into recent developments in the other branches of adoptive immunotherapy, both traditional and innovative. In this review, we describe the variety of adoptive cell therapies beyond CAR T-cell technology, i.e., exploitation of alternative cell sources with a high therapeutic potential against solid tumors (e.g., CAR M-cells) or aiming to be universal allogeneic (e.g., CAR NK-cells, γδ T-cells), tumor-infiltrating lymphocytes (TILs), and transgenic T-cell receptor (TCR) T-cell immunotherapies. In addition, we discuss the strategies for selection and validation of neoantigens to achieve efficiency and safety. We provide an overview of non-conventional TCRs and CARs, and address the problem of mispairing between the cognate and transgenic TCRs. Finally, we summarize existing and emerging approaches for manufacturing of the therapeutic cell products in traditional, semi-automated and fully automated Point-of-Care (PoC) systems.

Application of CAR-T Cell Therapy beyond Oncology: Autoimmune Diseases and Viral Infections.

Adoptive cell transfer (ACT) has long been at the forefront of the battle with cancer that began last century with the therapeutic application of tumor-infiltrating lymphocytes (TILs) against melanoma. The development of novel ACT approaches led researchers and clinicians to highly efficient technologies based on genetically engineered T lymphocytes, with chimeric antigen receptor (CAR)-T cells as the most prominent example. CARs consist of an extracellular domain that represents the single-chain variable fragment (scFv) of a monoclonal antibody (mAb) responsible for target recognition and the intracellular domain, which was built from up to several signaling motifs that mediated T cell activation. The number of potential targets amenable for CAR-T cell therapy is expanding rapidly, which means that the tremendous success of this approach in oncology could be further translated to treating other diseases. In this review, we outlined modern trends and recent developments in CAR-T cell therapy from an unusual point of view by focusing on diseases beyond cancer, such as autoimmune disorders and viral infections, including SARS-CoV-2.