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Arginase activity and arginine metabolism in immune cells have important consequences for health and disease. Their dysregulation is commonly observed in cancer, autoimmune disorders and infectious diseases. Following the initial description of a role for arginase in the dysfunction of T cells mounting an antitumour response, numerous studies have broadened our understanding of the regulation and expression of arginases and their integration with other metabolic pathways. Here, we highlight the differences in arginase compartmentalization and storage between humans and rodents that should be taken into consideration when assessing the effects of arginase activity. We detail the roles of arginases, arginine and its metabolites in immune cells and their effects in the context of cancer, autoimmunity and infectious disease. Finally, we explore potential therapeutic strategies targeting arginases and arginine.

Here we report the identification of human CD66b−CD64dimCD115−neutrophil-committed progenitor cells (NCPs) within the SSCloCD45dimCD34+and CD34dim/−subsets in the bone marrow. NCPs were either CD45RA+or CD45RA−, and in vitro experiments showed that CD45RA acquisition was not mandatory for their maturation process. NCPs exclusively generated human CD66b+neutrophils in both in vitro differentiation and in vivo adoptive transfer experiments. Single-cell RNA-sequencing analysis indicated NCPs fell into four clusters, characterized by different maturation stages and distributed along two differentiation routes. One of the clusters was characterized by an interferon-stimulated gene signature, consistent with the reported expansion of peripheral mature neutrophil subsets that express interferon-stimulated genes in diseased individuals. Finally, comparison of transcriptomic and phenotypic profiles indicated NCPs represented earlier neutrophil precursors than the previously described early neutrophil progenitors (eNePs), proNeus and COVID-19 proNeus. Altogether, our data shed light on the very early phases of neutrophil ontogeny.

Inflammatory responses rapidly detect pathogen invasion and mount a regulated reaction. However, dysregulated anti-pathogen immune responses can provoke life-threatening inflammatory pathologies collectively known as cytokine release syndrome (CRS), exemplified by key clinical phenotypes unearthed during the SARS-CoV-2 pandemic. The underlying pathophysiology of CRS remains elusive. We found that FLIP, a protein that controls caspase-8 death pathways, was highly expressed in myeloid cells of COVID-19 lungs. FLIP controlled CRS by fueling a STAT3-dependent inflammatory program. Indeed, constitutive expression of a viral FLIP homolog in myeloid cells triggered a STAT3-linked, progressive, and fatal inflammatory syndrome in mice, characterized by elevated cytokine output, lymphopenia, lung injury, and multiple organ dysfunctions that mimicked human CRS. As STAT3-targeting approaches relieved inflammation, immune disorders, and organ failures in these mice, targeted intervention towards this pathway could suppress the lethal CRS inflammatory state.

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ABSTRACTCancer cells favor the generation of myeloid cells with immunosuppressive and inflammatory features, including myeloid-derived suppressor cells (MDSCs), which support tumor progression. The anti-apoptotic molecule, cellular FLICE (FADD-like interleukin-1β-converting enzyme)-inhibitory protein (c-FLIP), which acts as an important modulator of caspase-8, is required for the development and function of monocytic (M)-MDSCs. Here, we assessed the effect of immune checkpoint inhibitor (ICI) therapy on systemic immunological landscape, including FLIP-expressing MDSCs, in non-small cell lung cancer (NSCLC) patients. Longitudinal changes in peripheral immunological parameters were correlated with patients’ outcome. In detail, 34 NSCLC patients were enrolled and classified as progressors(P) or non-progressors(NP), according to the RECIST evaluation. We demonstrated a reduction in pro-inflammatory cytokines such as IL-8, IL-6, and IL-1β in only NP patients after ICI treatment. Moreover, using t-distributed stochastic neighbor embedding (t-SNE) and cluster analysis, we characterized in NP patients a significant increase in the amount of lymphocytes and a slight contraction of myeloid cells such as neutrophils and monocytes. Despite this moderate ICI-associated alteration in myeloid cells, we identified a distinctive reduction of c-FLIP expression in M-MDSCs from NP patients concurrently with the first clinical evaluation (T1), even though NP and P patients showed the same level of expression at baseline (T0). In agreement with the c-FLIP expression, monocytes isolated from both P and NP patients displayed similar immunosuppressive functions at T0; however, this pro-tumor activity was negatively influenced at T1 in the NP patient cohort exclusively. Hence, ICI therapy can mitigate systemic inflammation and impair MDSC-dependent immunosuppression.

Myeloid cells can restrain antitumor immunity by metabolic pathways, such as the degradation of l-arginine, whose concentrations are regulated by the arginase 1 (ARG1) enzyme. Results from preclinical studies indicate the important role of arginine metabolism in pancreatic ductal adenocarcinoma (PDAC) progression, suggesting a potential for clinical application; however, divergent evolution in ARG1 expression and function in rodents and humans has restricted clinical translation. To overcome this dichotomy, here, we show that neutrophil extracellular traps (NETs), released by spontaneously activated neutrophils isolated from patients with PDAC, create a microdomain where cathepsin S (CTSS) cleaves human (h)ARG1 into different molecular forms endowed with enhanced enzymatic activity at physiological pH. NET-associated hARG1 suppresses T lymphocytes whose proliferation is restored by either adding a hARG1-specific monoclonal antibody (mAb) or preventing CTSS-mediated cleavage, whereas small-molecule inhibitors are not effective. We show that ARG1 blockade, combined with immune checkpoint inhibitors, can restore CD8+T cell function in ex vivo PDAC tumors. Furthermore, anti-hARG1 mAbs increase the frequency of adoptively transferred tumor-specific CD8+T cells in tumor and enhance the effectiveness of immune checkpoint therapy in humanized mice. Thus, this study shows that extracellular ARG1, released by activated myeloid cells, localizes in NETs, where it interacts with CTSS that in turn cleaves ARG1, producing major molecular forms endowed with different enzymatic activity at physiological pH. Once exocytosed, ARG1 activity can be targeted by mAbs, which bear potential for clinical application for the treatment of PDAC and require further exploration.

In the colonic environment, sustained Wnt–β-catenin activation in regulatory T cells promotes epigenetic rewiring toward proinflammatory RORγt+Tregcells, whose expansion parallels the disease progression from inflammatory bowel disease (IBD) to manifest colorectal cancer (CRC).

Breast tumor infiltrating lymphocytes (TIL) are enriched in tumor-specific cytotoxic T lymphocytes (CTL), and may represent a superior source of CTL compare to peripheral blood lymphocytes (PBL), for adoptive T cell immunotherapy of breast cancer. However, the immunocompetence of TIL and the possibility to consistently restore their tumor-specific lytic activity in vitroremains an open issue. In this study we evaluated the potential of tumor antigen-pulsed fully mature dendritic cell (DC) stimulation in restoring tumor-specific cytotoxicity in anergic TIL populations from advanced breast cancer patients. In addition we have compared tumor-specific T cell responses induced by tumor antigen-loaded DC stimulation of TIL to responses induced from PBL. Although TIL were consistently non-cytotoxic after isolation or culture in the presence of interleukin-2 (IL-2), in matched experiments from three consecutive patients, tumor-lysate-pulsed DC-stimulated CD8+ T cell derived from TIL were found to be significantly more cytotoxic than PBL (p< 0.05). In addition, cytotoxicity against autologous tumor cells was more significantly inhibited by an anti-HLA class I (W6/32) MAb in TIL compared to PBL (p< 0.05). CTL populations derived from TIL and PBL did not lyse autologous EBV-transformed lymphoblastoid cell lines, and showed negligible cytotoxicity against the NK-sensitive cell line K562. Furthermore, in both CD8+ T cell populations the majority of the tumor-specific CTL exhibited a Th1 cytokine bias (IFN-γhigh/IL-4low). Taken together, these data show that tumor lysate-pulsed mature DC can consistently restore tumor-specific lytic activity in non-cytotoxic breast cancer TIL. These results may have important implications for the treatment of chemotherapy resistant breast cancer with active or adoptive immunotherapy.

ABSTRACTMeningiomas WHO grade I and II are common intracranial tumors in adults that normally display a benign outcome, but are characterized by a great clinical heterogeneity and frequent recurrence of the disease. Although the presence of an immune cell infiltrate has been documented in these tumors, a clear phenotypical and functional characterization of the immune web is missing. Here, we performed an extensive immunophenotyping of peripheral blood and fresh tumor tissue at surgery by multiparametric flow cytometry in 34 meningioma patients, along with immunosuppressive activity of sorted cells of myeloid origin. Four subsets of myeloid cells, phenotypically corresponding to myeloid-derived suppressor cells (MDSCs) are detectable in the blood and in the tumor tissue of patients and three of them are significantly expanded in the blood of patients, but show no evidence of suppressive activity. At the tumor site, a large leukocyte infiltrate is present, predominantly constituted by CD33+myeloid cells, largely composed of macrophages endowed with suppressive activity and significantly expanded in grade II meningioma patients as compared to grade I.