Age-related epithelial defects limit thymic function and regeneration.

The thymus is essential for establishing adaptive immunity yet undergoes age-related involution that leads to compromised immune responsiveness. The thymus is also extremely sensitive to acute insult and although capable of regeneration, this capacity declines with age for unknown reasons. We applied single-cell and spatial transcriptomics, lineage-tracing and advanced imaging to define age-related changes in nonhematopoietic stromal cells and discovered the emergence of two atypical thymic epithelial cell (TEC) states. These age-associated TECs (aaTECs) formed high-density peri-medullary epithelial clusters that were devoid of thymocytes; an accretion of nonproductive thymic tissue that worsened with age, exhibited features of epithelial-to-mesenchymal transition and was associated with downregulation of FOXN1. Interaction analysis revealed that the emergence of aaTECs drew tonic signals from other functional TEC populations at baseline acting as a sink for TEC growth factors. Following acute injury, aaTECs expanded substantially, further perturbing trophic regeneration pathways and correlating with defective repair of the involuted thymus. These findings therefore define a unique feature of thymic involution linked to immune aging and could have implications for developing immune-boosting therapies in older individuals.

IL-18-secreting multiantigen targeting CAR T cells eliminate antigen-low myeloma in an immunocompetent mouse model.

ABSTRACT: Multiple myeloma is a plasma cell malignancy that is currently incurable with conventional therapies. Following the success of CD19-targeted chimeric antigen receptor (CAR) T cells in leukemia and lymphoma, CAR T cells targeting B-cell maturation antigen (BCMA) more recently demonstrated impressive activity in relapsed and refractory myeloma patients. However, BCMA-directed therapy can fail due to weak expression of BCMA on myeloma cells, suggesting that novel approaches to better address this antigen-low disease may improve patient outcomes. We hypothesized that engineered secretion of the proinflammatory cytokine interleukin-18 (IL-18) and multiantigen targeting could improve CAR T-cell activity against BCMA-low myeloma. In a syngeneic murine model of myeloma, CAR T cells targeting the myeloma-associated antigens BCMA and B-cell activating factor receptor (BAFF-R) failed to eliminate myeloma when these antigens were weakly expressed, whereas IL-18-secreting CAR T cells targeting these antigens promoted myeloma clearance. IL-18-secreting CAR T cells developed an effector-like T-cell phenotype, promoted interferon-gamma production, reprogrammed the myeloma bone marrow microenvironment through type-I/II interferon signaling, and activated macrophages to mediate antimyeloma activity. Simultaneous targeting of weakly-expressed BCMA and BAFF-R with dual-CAR T cells enhanced T-cell:target-cell avidity, increased overall CAR signal strength, and stimulated antimyeloma activity. Dual-antigen targeting augmented CAR T-cell secretion of engineered IL-18 and facilitated elimination of larger myeloma burdens in vivo. Our results demonstrate that combination of engineered IL-18 secretion and multiantigen targeting can eliminate myeloma with weak antigen expression through distinct mechanisms.

Long-Term Follow-Up of Newborns with 22q11 Deletion Syndrome and Low TRECs.

BACKGROUND: Population-based neonatal screening using T-cell receptor excision circles (TRECs) identifies infants with profound T lymphopenia, as seen in cases of severe combined immunodeficiency, and in a subgroup of infants with 22q11 deletion syndrome (22q11DS). PURPOSE: To investigate the long-term prognostic value of low levels of TRECs in newborns with 22q11DS. METHODS: Subjects with 22q11DS and low TRECs at birth (22q11Low, N=10), matched subjects with 22q11DS and normal TRECs (22q11Normal, N=10), and matched healthy controls (HC, N=10) were identified. At follow-up (median age 16 years), clinical and immunological characterizations, covering lymphocyte subsets, immunoglobulins, TRECs, T-cell receptor repertoires, and relative telomere length (RTL) measurements were performed. RESULTS: At follow-up, the 22q11Low group had lower numbers of naïve T-helper cells, naïve T-regulatory cells, naïve cytotoxic T cells, and persistently lower TRECs compared to healthy controls. Receptor repertoires showed skewed V-gene usage for naïve T-helper cells, whereas for naïve cytotoxic T cells, shorter RTL and a trend towards higher clonality were found. Multivariate discriminant analysis revealed a clear distinction between the three groups and a skewing towards Th17 differentiation of T-helper cells, particularly in the 22q11Low individuals. Perturbations of B-cell subsets were found in both the 22q11Low and 22q11Normal group compared to the HC group, with larger proportions of naïve B cells and lower levels of memory B cells, including switched memory B cells. CONCLUSIONS: This long-term follow-up study shows that 22q11Low individuals have persistent immunologic aberrations and increased risk for immune dysregulation, indicating the necessity of lifelong monitoring. CLINICAL IMPLICATIONS: This study elucidates the natural history of childhood immune function in newborns with 22q11DS and low TRECs, which may facilitate the development of programs for long-term monitoring and therapeutic choices.

Mapping of Signaling Pathways Linked to sIgAD Reveals Impaired IL-21 Driven STAT3 B-Cell Activation.

Objectives: It has recently been shown that individuals with selective IgA deficiency (sIgAD) have defective B cell responses both to T cell dependent and independent mimicking stimulations. The complex intracellular signaling pathways from different stimuli leading to IgA isotype switching have not been fully elucidated. Thus, the main objective of this study was to delineate these pathways and their potential role in the immunopathology linked to sIgAD. Materials and Methods: PBMCs from 10 individuals with sIgAD and 10 healthy controls (HC) were activated in vitro via either a T cell dependent or independent mimicking stimulation. Intracellular phosphorylation of pSTAT3, pSTAT5, pSTAT6, and as pERK1/2 was evaluated in T and B cells using phosphoflow cytometry. Results: By evaluating T cell dependent cytokine driven pathways linked to IgA isotype induction we identified a defect involving an IL-21 driven STAT3 activation isolated to B cells in sIgAD individuals. However, all other signaling pathways studied were found to be normal compared to HC. In T cell dependent cytokine driven stimulations linked to IgA isotype induction the following patterns emerged: (i) IL-10 led to significant STAT3 activation in both T- and B cells; (ii) IL-4 stimulation was predominantly confined to STAT6 activation in both T- and B cells, with some effects on STAT3 activation in T-cells; (iii) as expected, of tested stimuli, IL-2 alone activated STAT5 and some STAT3 activation though in both cases only in T-cells; (iv) IL-21 induced significant activation of STAT3 in both T- and B cells, with some effects on STAT5 activation in T-cells; and finally (v) synergistic effects were noted of IL-4+IL-10 on STAT5 activation in T-cells, and possibly STAT6 in both T- and B cells. On the other hand, CPG induced T cell independent activation was confined to ERK1/2 activation in B cells. Conclusion: Our results indicate a diminished STAT3 phosphorylation following IL-21 stimulation solely in B cells from sIgAD individuals. This can represent aberrant germinal center reactions or developmental halt. Thus, our work provides further insight into the unraveling of the previously hypothesized role of IL-21 to reconstitute immunoglobulin production in primary antibody deficiencies.

Transitional B Cells and TLR9 Responses Are Defective in Selective IgA Deficiency.

Selective IgA deficiency (IgAD) is the most common primary antibody deficiency in the western world with affected individuals suffering from an increased burden of autoimmunity, atopic diseases and infections. It has been shown that IgAD B cells can be induced with germinal center mimicking reactions to produce IgA. However, IgA is the most prevalent antibody in mucosal sites, where antigen-independent responses are important. Much interest has recently focused on the role of TLR9 in both naïve and mature B cell differentiation into IgA secreting plasma cells. Here, we analyze the phenotype and function of T and B cells in individuals with IgAD following IgA-inducing CpG-TLR9 stimulations. The IgAD individuals had significantly lower numbers of transitional B cells (CD19+CD24hiCD38hi) and class-switched memory B cells (CD20+CD27+IgD-) ex vivo. However, proportions of T cell populations ex vivo as well as in vitro induced T effector cells and T regulatory cells were comparable to healthy controls. After CpG stimulation, the transitional B cell defect was further enhanced, especially within its B regulatory subset expressing IL-10. Finally, CpG stimulation failed to induce IgA production in IgAD individuals. Collectively, our results demonstrate a defect of the TLR9 responses in IgAD that leads to B cell dysregulation and decreased IgA production.