The effects of Fasciola hepatica recombinant proteins (peroxiredoxin and cathepsin L1) on Crohn's disease experimental model.

The immunomodulatory potential of the excretory-secretory (E/S) proteins of the helminths has been shown in previous investigations. This study evaluated the effects of the recombinants and excretory-secretory proteins of the Fasciola hepatica on induced colitis in Balb/c mice. The F. hepatica Recombinant proteins, Cathepsin L1 and Peroxiredoxin, and E/S proteins were intraperitoneally injected into the three mice groups as the case groups, while the control groups received PBS. Colitis was induced in mice by intraluminal administration of the 2, 4, 6-Trinitrobenzenesulfonic acid solution (TNBS). After 8 h, the case groups received the second dosage of the treatments, and it was repeated 24 h later. The immunological, pathological, and macroscopic changes were evaluated 3 days after colitis induction. The macroscopic evaluation revealed significantly lower inflammatory scores in the mice treated with recombinant Peroxiredoxin (rPRX) and recombinant Cathepsin L1 (rCL1). Despite the macroscopic observation, the pathological finding was insignificant between the groups. IFN-γ secretion was significantly lower in splenocytes of the groups that received rPRX, rCL1, and E/S than the controls. IL-10 showed significantly higher levels in groups treated with rPRX and rCL1 than controls, whereas the level of IL-4 was not statistically significant. Excretory-secretory proteins of the F. hepatica showed immunomodulatory potency and the main effects observed in this study were through the reduction of inflammatory cytokine and inflammation manifestation as well as induction of anti-inflammatory cytokines.

MicroRNA-124 Enhances T Cells Functions by Manipulating the Lactic Acid Metabolism of Tumor Cells.

High production of lactic acid is a common feature of various tumors. Lactic acid is an immunosuppressive molecule with crucial roles in tumor cells' immune escape, which could largely be attributed to its negative effects on the T cells present in the tumor microenvironment (TME). Strategies that decrease the glycolysis rate of tumor cells could enhance immunosurveillance and limit tumor growth. Pyruvate kinase M2 (PKM2) is a key enzyme in the glycolysis pathway, and it plays a vital role in lactic acid buildup in the TME. MicroRNA (miR)-124 has been shown to be able to decrease tumor cell lactic acid synthesis indirectly by reducing PKM2 levels. In this study, we first overexpressed miR-124 in the tumor cells and evaluated its effects on the PKM2 expression and lactic acid production of the tumor cells using quantitative real-time polymerase chain reaction (qRT-PCR) and spectrophotometry, respectively. Then, we cocultured miR-124-treated tumor cells with T cells to investigate the effects of miR-124 overexpression on T cell proliferation, cytokine production, and apoptosis. Our results demonstrated that miR-124 overexpression could significantly reduce the amount of lactic acid produced by tumor cells by manipulating their glucose metabolism, which led to the augmented proliferation and IFN-γ production of T cells. Moreover, it rescued T cells from lactic acid-induced apoptosis. Our data suggest that lactic acid is a hindering factor for T-cell-based immunotherapies; however, manipulating tumor cells' metabolism via miR-124 could be a promising way to improve antitumor responses of T cells.

SARS-CoV-2 triggering autoimmune diseases.

Autoimmunity, hyperstimulation of the immune system, can be caused by a variety of reasons. Viruses are thought to be important environmental elements that contribute to the development of autoimmune antibodies. It seems that viruses cause autoimmunity with mechanisms such as molecular mimicry, bystander activation of T cells, transient immunosuppression, and inflammation, which has also been seen in post-Covid-19 autoimmunity. Infection of respiratory epithelium by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) dysregulates the immune response, triggers both innate and acquired immunity that led to the immune system's hyperactivation, excessive cytokine secretion known as "cytokine storm," and finally acute respiratory distress syndrome (ARDS) associated with high mortality. Any factor in the body that triggers chronic inflammation can contribute to autoimmune disease, which has been documented during the Covid-19 pandemic. It has been observed that some patients produce autoantibody and autoreactive CD4+ and CD8+ T cells, leading to the loss of self-tolerance. However, there is a scarcity of evidence defining the precise molecular interaction between the virus and the immune system to elicit autoreactivity. Here, we present a review of the relevant immunological findings in Covid-19 and the current reports of autoimmune disease associated with the disease.

Restricting tumor lactic acid metabolism using dichloroacetate improves T cell functions.

BACKGROUND: Lactic acid produced by tumors has been shown to overcome immune surveillance, by suppressing the activation and function of T cells in the tumor microenvironment. The strategies employed to impair tumor cell glycolysis could improve immunosurveillance and tumor growth regulation. Dichloroacetate (DCA) limits the tumor-derived lactic acid by altering the cancer cell metabolism. In this study, the effects of lactic acid on the activation and function of T cells, were analyzed by assessing T cell proliferation, cytokine production and the cellular redox state of T cells. We examined the redox system in T cells by analyzing the intracellular level of reactive oxygen species (ROS), superoxide and glutathione and gene expression of some proteins that have a role in the redox system. Then we co-cultured DCA-treated tumor cells with T cells to examine the effect of reduced tumor-derived lactic acid on proliferative response, cytokine secretion and viability of T cells. RESULT: We found that lactic acid could dampen T cell function through suppression of T cell proliferation and cytokine production as well as restrain the redox system of T cells by decreasing the production of oxidant and antioxidant molecules. DCA decreased the concentration of tumor lactic acid by manipulating glucose metabolism in tumor cells. This led to increases in T cell proliferation and cytokine production and also rescued the T cells from apoptosis. CONCLUSION: Taken together, our results suggest accumulation of lactic acid in the tumor microenvironment restricts T cell responses and could prevent the success of T cell therapy. DCA supports anti-tumor responses of T cells by metabolic reprogramming of tumor cells.

The role of Th22 cells, from tissue repair to cancer progression.

CD4+ T helper (Th) cells play a significant role in modulating host defense. In the presence of lineage specific cytokine cocktail, Naive CD4+ T cells can differentiate into several categories with distinct cytokines profile and effector functions. Th22 cells are a recently identified subset of CD4+ T cell, which differentiate from Naive CD4+ T in the presence of IL-6 and TNF-α. Th22 characterized by the production of interleukin-22 (IL-22) and expression of aryl hydrocarbon receptor (AHR). The main function of Th22 cells is to participate in mucosal defense, tissue repair, and wound healing. However, controversial data have shown that overexpression of IL-22 can lead to pathological changes under inflammatory conditions and tumor progression. This review summarizes our knowledge about the role of Th22 and IL-22 cells in tumor progression through induction of inflammation.

The application of immune checkpoint blockade in breast cancer and the emerging role of nanoparticle.

Breast cancer is the most common malignancy in the female population with a high mortality rate. Despite the satisfying depth of studies evaluating the contributory role of immune checkpoints in this malignancy, few articles have reviewed the pros and cons of immune checkpoint blockades (ICBs). In the current review, we provide an overview of immune-related inhibitory molecules and also discuss the original data obtained from international research laboratories on the aberrant expression of T and non-T cell-associated immune checkpoints in breast cancer. Then, we especially focus on recent studies that utilized ICBs as the treatment strategy in breast cancer and provide their efficiency reports. As there are always costs and benefits, we discuss the limitations and challenges toward ICB therapy such as adverse events and drug resistance. In the last section, we allocate an overview of the recent data concerning the application of nanoparticle systems for cancer immunotherapy and propose that nano-based ICB approaches may overcome the challenges related to ICB therapy in breast cancer. In conclusion, it seems it is time for nanoscience to more rapidly move forward into clinical trials and illuminates the breast cancer treatment area with its potent features for the target delivery of ICBs.

CRISPR/Cas9-mediated TGFßRII disruption enhances anti-tumor efficacy of human chimeric antigen receptor T cells in vitro.

BACKGROUND: CAR T-cell therapy has been recently unveiled as one of the most promising cancer therapies in hematological malignancies. However, solid tumors mount a profound line of defense to escape immunosurveillance by CAR T-cells. Among them, cytokines with an inhibitory impact on the immune system such as IL-10 and TGFß are of great importance: TGFß is a pleiotropic cytokine, which potently suppresses the immune system and is secreted by a couple of TME resident and tumor cells. METHODS: In this study, we hypothesized that knocking out the TGFß receptor II gene, could improve CAR T-cell functions in vitro and in vivo. Hereby, we used the CRISPR/Cas9 system, to knockout the TGFßRII gene in T-cells and could monitor the efficient gene knock out by genome analysis techniques. Next, Mesothelin or Claudin 6 specific CAR constructs were overexpressed via IVT-RNA electroporation or retroviral transduction and the poly-functionality of these TGFßRII KO CAR T-cells in terms of proliferation, cytokine secretion and cytotoxicity were assessed and compared with parental CAR T-cells. RESULTS: Our experiments demonstrated that TGFßRII KO CAR T-cells fully retained their capabilities in killing tumor antigen positive target cells and more intriguingly, could resist the anti-proliferative effect of exogenous TGFß in vitro outperforming wild type CAR T-cells. Noteworthy, no antigen or growth factor-independent proliferation of these TGFßRII KO CAR T-cells has been recorded. TGFßRII KO CAR T-cells also resisted the suppressive effect of induced regulatory T-cells in vitro to a larger extent. Repetitive antigen stimulation demonstrated that these TGFßRII KO CAR T-cells will experience less activation induced exhaustion in comparison to the WT counterpart. CONCLUSION: The TGFßRII KO approach may become an indispensable tool in immunotherapy of solid tumors, as it may surmount one of the key negative regulatory signaling pathways in T-cells.

Association of IL-15 and IP-10 Serum Levels with Cytomegalovirus Infection, CMV Viral Load and Cyclosporine Level after Kidney Transplantation.

BACKGROUND: Cytomegalovirus (CMV) infection is the most common complications following kidney transplantation. Natural killer (NK) cells demonstrated critical anti-viral role in controlling and elimination of CMV after transplantation. Interleukin-15 (IL-15) is a pleiotropic cytokine that promotes the activity of NK cells and strengthens the acquired immune system. Also, IP10 (CXCL10) is a chemotactic factor which regulates NK cell recruitment and antiviral immune response. We aimed to determine the correlation between the serum levels of IL-15 and IP-10 cytokines with CMV infection, CMV viral load, and cyclosporine as a major immunosuppressive treatment after transplantation. METHODS: Fifty-eight kidney transplant recipient patients without evidence of CMV virus disease before transplantation surgery were included in the study. From the day of transplant surgery, the patients were evaluated based on the presence of CMV Ag pp65, CMV viral load, serum levels of IL-15 & IP-10, Cyclosporine levels (C0 & C2), Glomerular Filtration Rate (GFR), and hematological & biochemical Index, up to 75 days. RESULTS: Comparison analysis of serum levels of IL-15 and IP-10 showed no significant association with CMV infection in kidney transplant recipients. In addition, CMV viral load and cyclosporine levels at C0 and C2 did not affect patients' IL-15 and IP-10 levels. CONCLUSION: The levels of IP-10 and IL-15 cytokines are not affected with CMV infection, even if a viral infection occurs in the early days after transplantation or long afterwards. In addition, taking the different levels of cyclosporine did not affect the cytokines levels. Other mechanisms may play a role in maintaining the levels of these cytokines.

Targeted knockdown of Tim3 by short hairpin RNAs improves the function of anti-mesothelin CAR T cells.

T-cell immunoglobulin mucin 3 (Tim3) is an immune checkpoint receptor that plays a central role in chimeric antigen receptor (CAR) T cell exhaustion within the tumor microenvironment. This study was aimed to evaluate the effects of targeted-knockdown of Tim3 on the antitumor function of anti-mesothelin (MSLN)-CAR T cells. To knockdown Tim3 expression, three different shRNA sequences specific to different segments of the human Tim3 gene were designed and co-inserted with an anti-MSLN-CAR transgene into lentiviral vectors. To investigate the efficacy of Tim3 targeting in T cells, expression of Tim3 was assessed before and after antigen stimulation. Afterwards, cytotoxic effects, proliferative response and cytokine production of MSLN-CAR T cells and Tim3-targeted MSLN-CAR T cells were analyzed. Our results showed that activation of T cells and MSLN-CAR T cells led to up-regulation of Tim3. Tim3 knockdown significantly decreased its expression in different groups of MSLN-CAR T cells. Tim3 knockdown significantly improved cytotoxic function, cytokine production and proliferation capacity of MSLN-CAR T cells. Our findings indicate that targeted knockdown of Tim3 allows tumor-infiltrating CAR T cells that would otherwise be inactivated to continue to expand and carry out effector functions, thereby altering the tumor microenvironment from immunosuppressive to immunosupportive via mitigated Tim3 signaling.

SARS-Cov-2 and COVID-19, Basic and Clinical Aspects of the Human Pandemic: A Review.

In the last two decades, we have witnessed three major epidemics of the coronavirus human disease namely, severe acute respiratory syndrome (SARS), Middle Eastern respiratory syndrome, and more recently an ongoing global pandemic of coronavirus disease 2019 (COVID-19). Iran, a country of nearly 84 million, in the Middle East, severely involved with the COVID-19 disease. A documented multidimensional approach to COVID-19 disease is therefore mandatory to provide a well-balanced platform for the concerned medical community in our county and beyond. In this review, we highlight the disease status in Iran and attempt to provide a multilateral view of the fundamental and clinical aspects of the disease including the clinical features of the confirmed cases, virology, pathogenesis, epidemiology, and laboratory methods needed for diagnosis.

Serum Levels of Soluble Fas and Fas Ligand in Iranian Women with Pre-eclampsia.

BACKGROUND: The precise responsible mechanism of pre-eclampsia remains controversial however, recent data suggest a main role of the abnormal activation of the adaptive immune system and Apoptosis. In this study, we have measured serum levels of Fas/Fasl as two important members of extrinsic apoptotic pathway in patient with pre-eclampsia. METHODS: 207 participants including 99 pre-eclampsia patients and 108 age and sex-matched normal pregnant women were involved in the case-control study. Plasma sample from each participant was collected and stored at -20 °C until batch processing.Serum levels of Fas and Fas ligand were measured by ELISA for each participant including 99 pre-eclampsia patients and 108 normal pregnant women. Following a test of statistical normality, nonparametric data were analyzed by Mann-Whitney. RESULTS: sFas levels in case group was significantly higher than controls; 584 (397-892) pg/ml in cases opposed to 341 (213-602) pg/ml in controls (p value< 0.01). sFasL in pre-eclampsia women was a little lower than controls; 255 (173-318) pg/ml and in case group compared to 265.5 (184-381.5) pg/ml in controls. CONCLUSION: We have found the increased levels of sFas in patients with pre-eclampsia in compare with the healthy pregnant women. It seems that abnormality in sFAS is related with pre-eclampsia.

A historical review of the role of cytokines involved in leishmaniasis.

Leishmaniasis is an infectious disease caused by the Leishmania genus, affecting millions of persons in the world. Despite increased studies, no vaccine has been developed against leishmaniasis, and drug resistance is evolving in some Leishmania species (spp). Innate and acquired immune cells and their associated cytokines interplay together to determine the immune responses related outcomes in leishmaniasis. Interferon (IFN)-γ or macrophage activating factor (MAF) is the first effective lymphokine (LK), with a related function to leishmaniasis, discovered in 1979. This review article discussed the history of cytokines involved in Leishmania infection, and it is the first report demonstrating the involvement in the disease by focusing on cutaneous leishmaniasis. Up to now, the role of many cytokines has been determined and the literature review showed that IL-35 is the latest known cytokine involved in leishmaniasis. This review revealed that the cytokines have pleiotropic effects, depending upon the cytokine environment, generated during the infection and the host genetic background or infecting Leishmania spp. Overall, advances in our knowledge of immune cells and their secreted cytokines, contributing to the protection or pathological process of leishmaniasis may help to reach new approaches for immunotherapy.

Construction and Functional Characterization of a Fully Human Anti-mesothelin Chimeric Antigen Receptor (CAR) Expressing T Cell.

Chimeric antigen receptor (CAR) T cell therapy is considered as an encouraging approach for the treatment of hematological malignancies. However, its efficacy in solid tumors has not been satisfying, mainly in the immunosuppressive network of the tumor microenvironment and paucity of appropriate target antigens. Mesothelin (MSLN) is a tumor-associated antigen (TAA) expressed in numerous types of solid tumors such as gastrointestinal, ovarian, and pancreatic tumors. Owing to high expression in tumor cells and low expression in normal tissues, MSLN-targeted therapies like monoclonal antibodies have been previously developed. In the present study, a CAR T cell harboring the second-generation of a fully human anti-MSLN-CAR construct containing CD3ζ and 4-1BB signaling domains was produced and it was functionally evaluated against an MSLN-expressing cell line. The findings showed potent, specific proliferation, cytotoxic activity, and interleukin (IL)-2, Tumor necrosis factor-(TNF) α, and Interferon-(IFN) γ production in an antigen-dependent manner. Cytotoxic activity was shown in effector-to-target ratio from 1:1 to 20:1, but the most adequate efficacy was observed in the ratio of 10:1. Non-specific activity against MSLN negative cell line was not observed. Our data demonstrated that primary human T cells expressing fully human MSLN-CAR construct are effective against MSLN-expressing cell lines in vitro, suggesting this MSLN-CAR construct as a potential therapeutic tool in a clinical setting.

Prolonged Persistence of Chimeric Antigen Receptor (CAR) T Cell in Adoptive Cancer Immunotherapy: Challenges and Ways Forward.

CAR T cell qualities, such as persistence and functionality play important roles in determining the outcome of cancer immunotherapy. In spite of full functionality, it has been shown that poor persistence of CAR T cells can limit an effective antitumor immune response. Here, we outline specific strategies that can be employed to overcome intrinsic and extrinsic barriers to CAR T cell persistence. We also offer our viewpoint on how growing use of CAR T cells in various cancers may require modifications in the intrinsic and extrinsic survival signals of CAR T cells. We anticipate these amendments will additionally provide the rationales for generation of more persistent, and thereby, more effective CAR T cell treatments. CAR T cell qualities, such as persistence and functionality play important roles in determining the outcome of cancer immunotherapy. In spite of full functionality, it has been shown that poor persistence of CAR T cells can limit an effective antitumor immune response. Here, we outline specific strategies that can be employed to overcome intrinsic and extrinsic barriers to CAR T cell persistence. We also offer our viewpoint on how growing use of CAR T cells in various cancers may require modifications in the intrinsic and extrinsic survival signals of CAR T cells. We anticipate these amendments will additionally provide the rationales for generation of more persistent, and thereby, more effective CAR T cell treatments.

Pharmacological targeting of immune checkpoint A2aR improves function of anti-CD19 CAR T cells in vitro.

In spite of impressive results in the treatment of acute lymphoblastic B cell leukemia (B-ALL) with chimeric antigen receptor (CAR) T cells, the clinical outcome of some hematological cancers like follicular lymphoma (FL) and chronic lymphocytic leukemia (CLL) has not been very promising likely due to immunosuppressive networks within tumor microenvironment. Hypoxia in the microenvironment of hematological malignancies and consequently generation of adenosine molecule is appeared to be correlated with immunosuppression, tumor progression, and relapse. Herein, we hypothesized that whether pharmacological targeting of adenosine 2a receptor (A2aR) can enhance antitumor activity of anti-CD19 CAR T cells in vitro. Prior to functional assays, A2aR expression was assessed in CAR-expressing T cells. Our results showed that A2aR was not only up-regulated in the fully human anti-CD19 CAR T cells (hereafter referred to as huCAR19 T cells) but also was further overexpressed following re-stimulation with target cells. Although pharmacological inhibition of A2aR could significantly increase proliferation capacity and cytokine production of huCAR19 T cells following treatment with an adenosine analog, cytotoxic activity of huCAR19 T cells was not significantly improved. Considering A2aR overexpression in huCAR19 T cells in the tumor microenvironment, our results indicated that pharmacological targeting of A2aR could not only improve huCAR19 T cells functionality in a hostile tumor microenvironment but also could have a therapeutic advantage, and sought to assess the possibility in a pre-clinical setting.

Genetic and pharmacological targeting of A2a receptor improves function of anti-mesothelin CAR T cells.

BACKGROUND: CAR T cell-based therapies have shown promising results in hematological malignancies. Results of CAR T cell projects in solid tumors have been less impressive, and factors including lack of targetable antigens and immunosuppressive tumor microenvironment (TME) have been suggested as culprits. Adenosine, a metabolite which is highly produced in TME, is known to mediate the suppression of anti-tumor T cell responses via binding and signaling through adenosine 2a receptor (A2aR). METHODS: In this study, the expression of A2aR and the effects of its activation on the function of fully human anti-mesothelin CAR T cells (MSLN-CAR T), were analyzed. Afterwards, the molecular and pharmacological means to overcome the inhibitory effects of A2aR signaling on CAR T cell function were used. This was performed by targeting A2aR expression in MSLN-CAR T cells using various anti-A2aR shRNA sequences embedded in the CAR vector and an A2aR pharmacological antagonist, SCH-58261. Statistical analyses were performed Prism 7 software. RESULTS: Our experiments showed significant A2aR upregulation on T cells during the CAR T cell production procedure (cell activation and transduction). Activation of adenosine signaling using adenosine analog led to the suppression of all major anti-tumor functions in MSLN-CAR T cells. Interestingly, CAR T cells that carried the anti-A2aR shRNA sequences were resistant to the inhibitory effects of adenosine signaling. Pharmacological inhibition of A2aR reversed the reduction in CAR T cell proliferation and cytokine response caused by the adenosine analog; however, it failed to rescue the cytotoxic function of the cells. CONCLUSION: Altogether, our results indicate that mitigating A2aR signaling by genetic targeting of the receptor might be a promising approach in improving CAR T cell function in an unreceptive microenvironment and could potentially improve the outcome of treatment in clinical settings.

Construction and functional characterization of a fully human anti-CD19 chimeric antigen receptor (huCAR)-expressing primary human T cells.

Although remarkable results have been attained by adoptively transferring T cells expressing fully murine and/or humanized anti-CD19 chimeric antigen receptors (CARs) to treat B cell malignancies, evidence of human anti-mouse immune responses against CARs provides a rationale for the development of less immunogenic CARs. By developing a fully human CAR (huCAR), these human anti-mouse immune responses are likely eliminated. This, perhaps, not only increases the persistence of anti-CD19 CAR T cells-thereby reducing the risk of tumor relapse-but also facilitates administration of multiple, temporally separated doses of CAR T cells to the same recipient. To these ends, we have designed and constructed a second-generation fully human anti-CD19 CAR (or huCAR19) containing a fully human single-chain variable fragment (ScFv) fused with a CD8a hinge, a 4-1BB transmembrane domain and intracellular T cell signaling domains of 4-1BB and CD3z. T cells expressing this CAR specifically recognized and lysed CD19+ target cells produced cytokines and proliferated in vitro. Moreover, cell volume data revealed that our huCAR construct cannot induce antigen-independent tonic signaling in the absence of cognate antigen. Considering our results, our anti-CD19 huCAR may overcome issues of transgene immunogenicity that plague trials utilizing CARs containing mouse-derived ScFvs. These results suggest that this huCAR19 be safely and effectively applied for adaptive T cell immunotherapy in clinical practice.

Improved Immunogenicity Against a Her2/neu-Derived Peptide by Employment of a Pan HLA DR-Binding Epitope and CpG in a BALB/c Mice Model.

BACKGROUND: An efficient strategy to improve the immunogenicity of peptide vaccines is the use of a synthetic peptide containing cytotoxic T-lymphocyte (CTL) epitopes with T-helper (Th) inducing-epitopes. OBJECTIVE: Our purpose was to determine the use of human epidermal growth factor receptor-2 (Her2/neu)- specific CTL epitopes plus the pan HLA DR-binding epitope (PADRE) and CpG oligodeoxynucleotides (ODNs) to induce antitumor effects in vaccinated mice. METHOD: Female BALB/c mice were immunized subcutaneously with different vaccines. Three mice per group were euthanized to assess immune responses and the others were transplanted with TUBO cells. Enzyme-linked Immuno Spot assay (ELISpot) and flow cytometry studies were followed by tumor size and survival rate measurements in a TUBO tumor mice model. RESULTS: The results showed that mice vaccinated with the P5 peptide plus PADRE plus CpG produced higher antigen-specific CTL responses than mice vaccinated with the P5 peptide alone. Also, tumors in those mice grew more slowly and the survival rates were greater than mice in the other groups. CONCLUSION: We conclude that peptide vaccines containing epitopes that stimulate both CD4+ and CD8+ T-cells are effective at inducing anti-tumor immunity.