Current US Food and Drug Administration-approved chimeric antigen receptor (CAR) T cells harbor the T cell receptor (TCR)-derived ζ chain as an intracellular activation domain in addition to costimulatory domains. The functionality in a CAR format of the other chains of the TCR complex, namely CD3δ, CD3ε and CD3γ, instead of ζ, remains unknown. In the present study, we have systematically engineered new CD3 CARs, each containing only one of the CD3 intracellular domains. We found that CARs containing CD3δ, CD3ε or CD3γ cytoplasmic tails outperformed the conventional ζ CAR T cells in vivo. Transcriptomic and proteomic analysis revealed differences in activation potential, metabolism and stimulation-induced T cell dysfunctionality that mechanistically explain the enhanced anti-tumor performance. Furthermore, dimerization of the CARs improved their overall functionality. Using these CARs as minimalistic and synthetic surrogate TCRs, we have identified the phosphatase SHP-1 as a new interaction partner of CD3δ that binds the CD3δ-ITAM on phosphorylation of its C-terminal tyrosine. SHP-1 attenuates and restrains activation signals and might thus prevent exhaustion and dysfunction. These new insights into T cell activation could promote the rational redesign of synthetic antigen receptors to improve cancer immunotherapy.
© 2023. The Author(s).

COVID-19 can cause severe neurological symptoms, but the underlying pathophysiological mechanisms are unclear. Here, we interrogated the brain stems and olfactory bulbs in postmortem patients who had COVID-19 using imaging mass cytometry to understand the local immune response at a spatially resolved, high-dimensional, single-cell level and compared their immune map to non-COVID respiratory failure, multiple sclerosis, and control patients. We observed substantial immune activation in the central nervous system with pronounced neuropathology (astrocytosis, axonal damage, and blood-brain-barrier leakage) and detected viral antigen in ACE2-receptor-positive cells enriched in the vascular compartment. Microglial nodules and the perivascular compartment represented COVID-19-specific, microanatomic-immune niches with context-specific cellular interactions enriched for activated CD8+ T cells. Altered brain T-cell-microglial interactions were linked to clinical measures of systemic inflammation and disturbed hemostasis. This study identifies profound neuroinflammation with activation of innate and adaptive immune cells as correlates of COVID-19 neuropathology, with implications for potential therapeutic strategies.
Copyright © 2021 Elsevier Inc. All rights reserved.

The monoclonal antibody against CTLA-4, Ipilimumab, is a first-in-class immune-checkpoint inhibitor approved for treatment of advanced melanoma in adults but not extensively studied in children. In light of the fact that the immune response early in life differs from that of adults, we have applied a human in vitro model stimulating CD4+ T-cells from neonates, children (1-5 years), and adults antigen-specifically with Staphylococcus aureus (S. aureus) for assessment of CTLA-4 blockade early in life. We show that T-cell proliferation as well as frequencies of antigen-specific T-cells (CD40L+CD4+) were enhanced in neonatal T-cells upon CTLA-4 blockade showing a larger variance within the group (F-test p < .0001). Using machine learning algorithm Random Forest, adult and neonatal T-cell responses can be unambiguously categorized (F1 score-0.75) on the basis of their cytokine (co-)expression. Blockade of CTLA-4 enhanced frequencies of IL-8, IFNγ, and IL-10 producers among CD40L+ T-cells. Of note, antigen-specific T-cells from neonates displayed higher cytokine coproduction at baseline, while T-cells from children caught up to neonates, and adults to baseline of children upon CTLA-4 blockade. These findings reveal that in neonatal T-cells blockade of CTLA-4 mainly unleashes the antigen-specific capacity by increasing the numbers of responding T-cells, whereas in children and adults it promotes the coexpression of cytokines by individual T-cells. Thus, CTLA-4 blockade boosts antitumor immunity through different mechanisms depending on the patients' age. These data implicate a strong impact of the developmental stage of the T-cell compartment on the effects of immune-checkpoint therapy.
© 2021 The Author(s). Published with license by Taylor & Francis Group, LLC.

Control of visceral leishmaniasis (VL) caused by Leishmania donovani requires interferon-γ production by CD4+ T cells. In VL patients, antiparasitic CD4+ T-cell responses are ineffective for unknown reasons. In this study, we measured the expression of genes associated with various immune functions in these cells from VL patients and compared them to CD4+ T cells from the same patients after drug treatment and from endemic controls. We found reduced GATA3, RORC, and FOXP3 gene expression in CD4+ T cells of VL patients, associated with reduced Th2, Th17, and FOXP3+CD4+ T regulatory cell frequencies in VL patient blood. Interleukin 2 (IL-2) was an important upstream regulator of CD4+ T cells from VL patients, and functional studies demonstrated the therapeutic potential of IL-2 for improving antiparasitic immunity. Together, these results provide new insights into the characteristics of CD4+ T cells from VL patients that can be used to improve antiparasitic immune responses.
© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Exhausted CD8 T (Tex) cells are immunotherapy targets in chronic infection and cancer, but a comprehensive assessment of Tex cell diversity in human disease is lacking. Here, we developed a transcriptomic- and epigenetic-guided mass cytometry approach to define core exhaustion-specific genes and disease-induced changes in Tex cells in HIV and human cancer. Single-cell proteomic profiling identified 9 distinct Tex cell clusters using phenotypic, functional, transcription factor, and inhibitory receptor co-expression patterns. An exhaustion severity metric was developed and integrated with high-dimensional phenotypes to define Tex cell clusters that were present in healthy subjects, common across chronic infection and cancer or enriched in either disease, linked to disease severity, and changed with HIV therapy. Combinatorial patterns of immunotherapy targets on different Tex cell clusters were also defined. This approach and associated datasets present a resource for investigating human Tex cell biology, with implications for immune monitoring and immunomodulation in chronic infections, autoimmunity, and cancer.
Copyright © 2018 Elsevier Inc. All rights reserved.