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Tumour cells do not exist as isolated entities. Instead, they are surrounded by a variety of cells and extracellular matrix, which form the tumour microenvironment (TME). The interaction between cancer cells and their microenvironment is increasingly acknowledged as essential in dictating the outcome of the patients. The TME includes everything that surrounds tumour cells and is often highjacked by the latter to promote their growth, invasion, and immune escape. Immune cells and cancer-associated fibroblasts (CAFs) are essential components of the TME, and there is increasing evidence that their interaction constitutes a major player not only for tumour progression but also for therapy response.Recent work in the field of immuno-oncology resulted in the development of novel therapies that aim at activating immune cells against cancer cells to eliminate them. Despite their unprecedented success, the lack of response from a large portion of patients highlights the need for further progress and improvement. To achieve its ultimate goal, the interaction between cancer cells and the TME needs to be studied in-depth to allow the targeting of mechanisms that are involved in resistance or refractoriness to therapy. Moreover, predictive and prognostic biomarkers for patient stratification are still missing. In this review, we focus on and highlight the complexity of CAFs within the TME and how their interaction, particularly with immune cells, can contribute to treatment failure. We further discuss how this crosstalk can be further dissected and which strategies are currently used to target them.

Conformational change of the β2 integrin lymphocyte function-associated antigen 1 (LFA-1) is an early marker of T cell activation. A protocol using the mAb clone m24 recognizing the active, extended high-affinity conformation has been previously described for the assessment of functional CD4

Sequestration of Plasmodium falciparum(P. falciparum)-infected erythrocytes to host endothelium through the parasite-derived P. falciparum erythrocyte membrane protein 1 (PfEMP1) adhesion proteins is central to the development of malaria pathogenesis. PfEMP1 proteins have diversified and expanded to encompass many sequence variants, conferring each parasite a similar array of human endothelial receptor-binding phenotypes. Here, we analyzed RNA-seq profiles of parasites isolated from 32 P. falciparum-infected adult travellers returning to Germany. Patients were categorized into either malaria naive (n = 15) or pre-exposed (n = 17), and into severe (n = 8) or non-severe (n = 24) cases. For differential expression analysis, PfEMP1-encoding var gene transcripts were de novo assembled from RNA-seq data and, in parallel, var-expressed sequence tags were analyzed and used to predict the encoded domain composition of the transcripts. Both approaches showed in concordance that severe malaria was associated with PfEMP1 containing the endothelial protein C receptor (EPCR)-binding CIDRα1 domain, whereas CD36-binding PfEMP1 was linked to non-severe malaria outcomes. First-time infected adults were more likely to develop severe symptoms and tended to be infected for a longer period. Thus, parasites with more pathogenic PfEMP1 variants are more common in patients with a naive immune status, and/or adverse inflammatory host responses to first infections favor the growth of EPCR-binding parasites.

Simple Summary T cells bearing the co-receptor CD4 on their cell surface are a heterogeneous group of T lymphocytes that exert pro- or anti-inflammatory functions. Evidence from mouse models and cancer patients reveal that various CD4+ T cell subsets play an antagonistic role in the antitumor immune response. This review summarizes current knowledge on CD4+ T cell subsets, on how they impact tumor growth in patients, and which role these cells play in newest cancer immunotherapies. Abstract Cancer immunotherapy activates the immune system to specifically target malignant cells. Research has often focused on CD8+ cytotoxic T cells, as those have the capacity to eliminate tumor cells after specific recognition upon TCR-MHC class I interaction. However, CD4+ T cells have gained attention in the field, as they are not only essential to promote help to CD8+ T cells, but are also able to kill tumor cells directly (via MHC-class II dependent recognition) or indirectly (e.g., via the activation of other immune cells like macrophages). Therefore, immunotherapy approaches have shifted from only stimulating CD8+ T cells to targeting and assessing both, CD4+ and CD8+ T cell subsets. Here, we discuss the various subsets of CD4+ T cells, their plasticity and functionality, their relevance in the antitumor immune response in patients affected by cancer, and their ever-growing role in therapeutic approaches for human cancer.

We have previously shown that conformational change in the β2-integrin is a very early activation marker that can be detected with fluorescent multimers of its ligand intercellular adhesion molecule (ICAM)-1 for rapid assessment of antigen-specific CD8+ T cells. In this study, we describe a modified protocol of this assay for sensitive detection of functional antigen-specific CD4+ T cells using a monoclonal antibody (clone m24 Ab) specific for the open, high-affinity conformation of the β2-integrin. The kinetics of β2-integrin activation was different on CD4+ and CD8+ T cells (several hours vs. few minutes, respectively); however, m24 Ab readily stained both cell types 4–6 h after antigen stimulation. With this protocol, we were able to monitor ex vivo effector and memory CD4+ and CD8+ T cells specific for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), cytomegalovirus (CMV), Epstein–Barr virus (EBV), and hepatitis B virus (HBV) in whole blood or cryopreserved peripheral blood mononuclear cells (PBMCs) of infected or vaccinated individuals. By costaining β2-integrin with m24 and CD154 Abs, we assessed extremely low frequencies of polyfunctional CD4+ T cell responses. The novel assay used in this study allows very sensitive and simultaneous screening of both CD4+ and CD8+ T cell reactivities, with versatile applicability in clinical and vaccination studies.

We have previously shown that beta2-integrin conformational change is a very early activation marker that can be detected with fluorescent multimers of its ligand ICAM-1 for a rapid assessment of antigen-specific CD8+ T cells. Here, we describe a modified protocol of this assay for sensitive detection of functional antigen-specific CD4+ T cells using a monoclonal antibody (clone m24) specific for the open, high affinity conformation of the beta2-integrin. Kinetics of beta2-integrin activation were different on CD4+ and CD8+ T cells (several hours vs. few minutes, respectively), however, m24 antibody readily stained both cell types 4-6 hours after antigen stimulation. With this protocol, we were able to monitor CD4+ and CD8+ virus-specific T cells specific for CMV, EBV, HBV, and SARS-CoV-2 in whole blood or cryopreserved PBMCs of infected or vaccinated individuals. By costaining with m24 and CD154 antibodies, we assessed extremely low frequencies of polyfunctional CD4+ T cell responses to SARS-CoV-2 derived peptides. Our novel assay thus allows very sensitive and simultaneous screening of both CD4+ and CD8+ T cell reactivities with versatile applicability in clinical and vaccination studies, and for epitope discovery.

Antimetabolites are small molecules that inhibit enzymes by mimicking physiological substrates. We report the discovery and structural elucidation of the antimetabolite 7-deoxy-sedoheptulose (7dSh). This unusual sugar inhibits the growth of various prototrophic organisms, including species of cyanobacteria, Saccharomyces, and Arabidopsis. We isolate bioactive 7dSh from culture supernatants of the cyanobacterium Synechococcus elongatus. A chemoenzymatic synthesis of 7dSh using S. elongatus transketolase as catalyst and 5-deoxy-d-ribose as substrate allows antimicrobial and herbicidal bioprofiling. Organisms treated with 7dSh accumulate 3-deoxy-d-arabino-heptulosonate 7-phosphate, which indicates that the molecular target is 3-dehydroquinate synthase, a key enzyme of the shikimate pathway, which is absent in humans and animals. The herbicidal activity of 7dSh is in the low micromolar range. No cytotoxic effects on mammalian cells have been observed. We propose that the in vivo inhibition of the shikimate pathway makes 7dSh a natural antimicrobial and herbicidal agent., Mother Nature is a valuable resource for the discovery of drug and agricultural chemicals. Here, the authors show that 7-deoxy-sedoheptulose produced by a cyanobacterium is an antimicrobial and herbicidal compound that acts through inhibition of 3-dehydroquniate synthase in the shikimate pathway.