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Objective: We investigated whether the frequency, phenotype, and suppressive function of CD4+FOXP3+ regulatory T cells (Tregs) are altered in young TS patients with the 45,X karyotype compared to age-matched controls. Design and Methods: Peripheral blood mononuclear cells from young TS patients (n = 24, 17.4–35.9 years) and healthy controls (n = 16) were stained with various Treg markers to characterize their phenotypes. Based on the presence of thyroid autoimmunity, patients were categorized into TS (–) (n = 7) and TS (+) (n = 17). Tregs sorted for CD4+CD25bright were co-cultured with autologous CD4+CD25− target cells in the presence of anti-CD3 and -CD28 antibodies to assess their suppressive function. Results: Despite a lower frequency of CD4+ T cells in the TS (-) and TS (+) patients (mean 30.8% and 31.7%, vs. 41.2%; P = 0.003 and P < 0.001, respectively), both groups exhibited a higher frequency of FOXP3+ Tregs among CD4+ T cells compared with controls (means 1.99% and 2.05%, vs. 1.33%; P = 0.029 and P = 0.004, respectively). There were no differences in the expression of CTLA-4 and the frequency of Tregs expressing CXCR3+, and CCR4+CCR6+ among the three groups. However, the ability of Tregs to suppress the in vitro proliferation of autologous CD4+CD25− T cells was significantly impaired in the TS (–) and TS (+) patients compared to controls (P = 0.003 and P = 0.041). Meanwhile, both the TS (–) and TS (+) groups had lower frequencies of naïve cells (P = 0.001 for both) but higher frequencies of effector memory cells (P = 0.004 and P = 0.002) than did the healthy control group. Conclusions: The Tregs of the TS patients could not efficiently suppress the proliferation of autologous effector T cells, despite their increased frequency in peripheral CD4+ T cells. [ABSTRACT FROM AUTHOR]

Chimeric antigen receptor (CAR)‐T cell therapy has revolutionized cancer treatment. CARs are activated at the immunological synapse (IS) when their single‐chain variable fragment (scFv) domain engages with an antigen, allowing them to directly eliminate cancer cells. Here, an innovative IS biosensor based on fluorescence resonance energy transfer (FRET) for the real‐time assessment of CAR‐IS architecture and signaling competence is presented. Using this biosensor, scFv variants for mesothelin‐targeting CARs and identified as a novel scFv with enhanced CAR‐T cell functionality despite its lower affinity than the original screened. The original CAR promoted internalization and trogocytosis, disrupting stable IS formation and impairing functionality are further observed. These findings emphasize the importance of enhancing IS quality rather than maximizing scFv affinity for superior CAR‐T cell responses. Therefore, the FRET‐based IS biosensor is a powerful tool for predicting CAR‐T cell function, enabling the efficient engineering of next‐generation CARs with enhanced antitumor potency. [ABSTRACT FROM AUTHOR]

Background: The clinical outcomes and immunological features of coronavirus disease 2019 (COVID-19) patients receiving B-cell depletion therapy (BCDT), especially in Omicron variant era, have not been fully elucidated. We aimed to investigate the outcomes and immune responses of COVID-19 patients receiving BCDT during the Omicron period. Methods: We retrospectively compared clinical outcomes between COVID-19 patients treated with BCDT (the BCDT group) and those with the same underlying diseases not treated with BCDT (the non-BCDT group). For immunological analyses, we prospectively enrolled COVID-19 patients receiving BCDT and immunocompetent COVID-19 patients as controls. We measured humoral and cellular immune responses using the enzyme-linked immunosorbent assay and flow cytometry. Results: Severe to critical COVID-19 was more frequent in the BCDT group than in the non-BCDT group (41.9% vs. 28.3%, p =.030). BCDT was an independent risk factor for severe to critical COVID-19 (adjusted odds ratio [aOR] 2.21, 95% confidence interval [CI] 1.21–4.04, p =.010) as well as for COVID-19-related mortality (aOR 4.03, 95% CI 1.17–13.86, p =.027). Immunological analyses revealed that patients receiving BCDT had lower anti-S1 IgG titres and a tendency to higher proportions of activated CD4+ T-cells than the controls. Conclusions: BCDT was associated with worse COVID-19 outcomes in the Omicron period. Humoral immune response impairment and T-cell hyperactivation were the main immunological features of COVID-19 patients treated with BCDT, which may have contributed to the worse outcomes of COVID-19 in this population. [ABSTRACT FROM AUTHOR]

Tumor acidosis, a common phenomenon in solid cancers such as breast cancer, is caused by the abnormal metabolism of cancer cells. The low pH affects cells surrounding the cancer, and tumor acidosis has been shown to inhibit the activity of immune cells. Despite many previous studies, the immune surveillance mechanisms are not fully understood. We found that the expression of PD-L1 was significantly increased under conditions of extracellular acidosis in MDA-MB-231 cells. We also confirmed that the increased expression of PD-L1 mediated by extracellular acidosis was decreased when the pH was raised to the normal range. Gene set enrichment analysis (GSEA) of public breast cancer patient databases showed that PD-L1 expression was also highly correlated with IL-6/JAK/STAT3 signaling. Surprisingly, the expression of both phospho-tyrosine STAT3 and PD-L1 was significantly increased under conditions of extracellular acidosis, and inhibition of STAT3 did not increase the expression of PD-L1 even under acidic conditions in MDA-MB-231 cells. Based on these results, we suggest that the expression of PD-L1 is increased by tumor acidosis via activation of STAT3 in MDA-MB-231 cells. [ABSTRACT FROM AUTHOR]

Background: Practical guidance is needed regarding the vaccination of coronavirus disease 2019 (COVID-19) convalescent individuals in resource-limited countries. It includes the number of vaccine doses that should be given to unvaccinated patients who experienced COVID-19 early in the pandemic. Methods: We recruited COVID-19 convalescent individuals who received one or two doses of an mRNA vaccine within 6 or around 18 months after a diagnosis of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection. Their samples were assessed for IgG-binding or neutralizing activity and cell-mediated immune responses against SARS-CoV-2 wild-type and variants of concern. Results: A total of 43 COVID-19 convalescent individuals were analyzed in the present study. The results showed that humoral and cellular immune responses against SARS-CoV-2 wild-type and variants of concern, including the Omicron variant, were comparable among patients vaccinated within 6 versus around 18 months. A second dose of vaccine did not significantly increase immune responses. Conclusion: One dose of mRNA vaccine should be considered sufficient to elicit a broad immune response even around 18 months after a COVID-19 diagnosis. [ABSTRACT FROM AUTHOR]

• COVID-19 can deteriorate at the 2nd week of illness despite decreasing viral loads. • We analyzed PBMCs of severe or mild COVID-19 cases at the 1st and 3rd week of illness. • Cellular responses were not different between two groups at the 1st week of illness. • Higher T-cell proliferation, activation, and cytotoxicity was noted in severe cases at the 3rd week. • Aberrant hyperactivation of cytotoxic T-cell may contribute to severe COVID-19 pneumonia. We hypothesized that immune response may contribute to progression of coronavirus disease-19 (COVID-19) at the second week of illness. Therefore, we compared cell-mediated immune (CMI) responses between severe and mild COVID-19 cases. We examined peripheral blood mononuclear cells of laboratory-confirmed COVID-19 patients from their first and third weeks of illness. Severe pneumonia was defined as an oxygen saturation ≤93% at room air. Expressions of molecules related to T-cell activation and functions were analyzed by flow cytometry. The population dynamics of T cells at the first week were not different between the two groups. However, total numbers of CD4+ and CD8+ T cells tended to be lower in the severe group at the third week of illness. Expressions of Ki-67, PD-1, perforin, and granzyme B in CD4+ or CD8+ T cells were significantly higher in the severe group than in the mild group at the third week. In contrast to the mild group, the levels of their expression did not decrease in the severe group. Severe COVID-19 had a higher degree of proliferation, activation, and cytotoxicity of T-cells at the late phase of illness without cytotoxic T-cell contraction, which might contribute to the development of severe COVID-19. [ABSTRACT FROM AUTHOR]