Evolution of Microglia.

Microglial cells are unique tissue-resident macrophages located in the parenchyma of the central nervous system (CNS). A recent comparative transcriptional study on microglia across more than 20 species from leach across chicken and many more up to humans revealed multiple conserved features. The results indicate the imperative role of microglia over the last 500 million years (Geirsdottir et al. Cell 181:746, 2020). Improved understanding of microglial evolution provides essential insights into conserved and divergent microglial pathways and will have implications for future development of microglia-based therapies to treat CNS disorders. Not only therapeutic approaches may be rethought, but also the understanding of sex specificity of the immune system within the CNS needs to be renewed. Besides revealing the highly detailed characteristics of microglia, the former paradigm of microglia being the only CNS-resident immune cells was outdated by the identification of CNS-associated macrophages (CAMs) as CNS interface residents, who, most likely, accompanied microglia in evolution over the past million years.

Expansion of circulating stem-like CD8+ T cells by adding CD122-directed IL-2 complexes to radiation and anti-PD1 therapies in mice.

Combination of radiation therapy (RT) with immune checkpoint blockade can enhance systemic anti-tumor T cell responses. Here, using two mouse tumor models, we demonstrate that adding long-acting CD122-directed IL-2 complexes (IL-2c) to RT/anti-PD1 further increases tumor-specific CD8+ T cell numbers. The highest increase (>50-fold) is found in the blood circulation. Compartmental analysis of exhausted T cell subsets shows that primarily undifferentiated, stem-like, tumor-specific CD8+ T cells expand in the blood; these cells express the chemokine receptor CXCR3, which is required for migration into tumors. In tumor tissue, effector-like but not terminally differentiated exhausted CD8+ T cells increase. Consistent with the surge in tumor-specific CD8+ T cells in blood that are migration and proliferation competent, we observe a CD8-dependent and CXCR3-dependent enhancement of the abscopal effect against distant/non-irradiated tumors and find that CD8+ T cells isolated from blood after RT/anti-PD1/IL-2c triple treatment can be a rich source of tumor-specific T cells for adoptive transfers.

Adding Indoximod to Hypofractionated Radiotherapy with Anti-PD-1 Checkpoint Blockade Enhances Early NK and CD8+ T-Cell-Dependent Tumor Activity.

PURPOSE: There is growing interest in combinations of immunogenic radiotherapy (RT) and immune checkpoint blockade, but clinical responses are still limited. Therefore, we tested the triple therapy with an inhibitor of the indoleamine 2,3-dioxygenase pathway, which like immune checkpoints, downregulates the antitumor immune response. EXPERIMENTAL DESIGN: Triple treatment with hypofractionated RT (hRT) + anti-PD-1 antibody (αPD1) + indoximod was compared with the respective mono- and dual therapies in two syngeneic mouse models. RESULTS: The tumors did not regress following treatment with hRT + αPD1. The αPD1/indoximod combination was not effective at all. In contrast, triple treatment induced rapid, marked tumor regression, even in mice with a large tumor. The effects strongly depended on CD8+ T cells and partly on natural killer (NK) cells. Numbers and functionality of tumor-specific CD8+ T cells and NK cells were increased, particularly early during treatment. However, after 2.5-3 weeks, all large tumors relapsed, which was accompanied by increased apoptosis of tumor-infiltrating lymphocytes associated with a non-reprogrammable state of exhaustion, terminal differentiation, and increased activation-induced cell death, which could not be prevented by indoximod in these aggressive tumor models. Some mice with a smaller tumor were cured. Reirradiation during late regression (day 12), but not after relapse, cured almost all mice with a large B16-CD133 tumor, and strongly delayed relapse in the less immunogenic 4T1 model, depending on CD8+ T cells. CONCLUSIONS: Our findings may serve as a rationale for the clinical evaluation of this triple-combination therapy in patients with solitary or oligometastatic tumors in the neoadjuvant or the definitive setting.

Cisplatin Facilitates Radiation-Induced Abscopal Effects in Conjunction with PD-1 Checkpoint Blockade Through CXCR3/CXCL10-Mediated T-cell Recruitment.

PURPOSE: Localized radiotherapy can cause T-cell-mediated abscopal effects on nonirradiated metastases, particularly in combination with immune checkpoint blockade (ICB). However, results of prospective clinical trials have not met the expectations. We therefore investigated whether additional chemotherapy can enhance radiotherapy-induced abscopal effects in conjunction with ICB. EXPERIMENTAL DESIGN: In three different two-tumor mouse models, triple therapy with radiotherapy, anti-PD-1, and cisplatin (one of the most widely used antineoplastic agents) was compared with double or single therapies. RESULTS: In these mouse models, the response of the nonirradiated tumor and the survival of the mice were much better upon triple therapy than upon radiotherapy + anti-PD-1 or cisplatin + anti-PD-1 or the monotherapies; complete regression of the nonirradiated tumor was usually only observed in triple-treated mice. Mechanistically, the enhanced abscopal effect required CD8+T cells and relied on the CXCR3/CXCL10 axis. Moreover, CXCL10 was found to be directly induced by cisplatin in the tumor cells. Furthermore, cisplatin-induced CD8+T cells and direct cytoreductive effects of cisplatin also seem to contribute to the enhanced systemic effect. Finally, the results show that the abscopal effect is not precluded by the observed transient radiotherapy-induced lymphopenia. CONCLUSIONS: This is the first report showing that chemotherapy can enhance radiotherapy-induced abscopal effects in conjunction with ICB. This even applies to cisplatin, which is not classically immunogenic. Whereas previous studies have focused on how to effectively induce tumor-specific T cells, this study highlights that successful attraction of the induced T cells to nonirradiated tumors is also crucial for potent abscopal effects.

Structure of the malaria vaccine candidate antigen CyRPA and its complex with a parasite invasion inhibitory antibody.

Invasion of erythrocytes by Plasmodial merozoites is a composite process involving the interplay of several proteins. Among them, the Plasmodium falciparum Cysteine-Rich Protective Antigen (PfCyRPA) is a crucial component of a ternary complex, including Reticulocyte binding-like Homologous protein 5 (PfRH5) and the RH5-interacting protein (PfRipr), essential for erythrocyte invasion. Here, we present the crystal structures of PfCyRPA and its complex with the antigen-binding fragment of a parasite growth inhibitory antibody. PfCyRPA adopts a 6-bladed ß-propeller structure with similarity to the classic sialidase fold, but it has no sialidase activity and fulfills a purely non-enzymatic function. Characterization of the epitope recognized by protective antibodies may facilitate design of peptidomimetics to focus vaccine responses on protective epitopes. Both in vitro and in vivo anti-PfCyRPA and anti-PfRH5 antibodies showed more potent parasite growth inhibitory activity in combination than on their own, supporting a combined delivery of PfCyRPA and PfRH5 in vaccines.

Establishing High Dimensional Immune Signatures from Peripheral Blood via Mass Cytometry in a Discovery Cohort of Stage IV Melanoma Patients.

The identification of blood-borne biomarkers correlating with melanoma patient survival remains elusive. Novel techniques such as mass cytometry could help to identify melanoma biomarkers, allowing simultaneous detection of up to 100 parameters. However, the evaluation of multiparametric data generated via time-of-flight mass cytometry requires novel analytical techniques because the application of conventional gating strategies currently used in polychromatic flow cytometry is not feasible. In this study, we have employed 38-channel time-of-flight mass cytometry analysis to generate comprehensive immune cell signatures using matrix boolean analysis in a cohort of 28 stage IV melanoma patients and 17 controls. Clusters of parameters were constructed from the abundance of cellular phenotypes significantly different between patients and controls. This approach identified patient-specific combinatorial immune signatures consisting of high-resolution subsets of the T cell, NK cell, B cell, and myeloid compartments. An association with superior survival was characterized by a balanced distribution of myeloid-derived suppressor cell-like and APC-like myeloid phenotypes and differentiated NK cells. The results of this study in a discovery cohort of melanoma patients suggest that multifactorial immune signatures have the potential to allow more accurate prediction of individual patient outcome. Further investigation of the identified immune signatures in a validation cohort is now warranted.