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Background Anti-TNF inhibitors successfully improve the quality of life of patients with inflammatory disease. Unfortunately, not all patients respond to anti-TNF therapy, and some patients show paradoxical immune side effects, which are poorly understood. Surprisingly, anti-TNF agents were shown to promote IL-17A production with as yet unknown clinical implications. Objective We sought to investigate the molecular mechanism underlying anti-TNF–driven IL-17A expression and the clinical implications of this phenomenon. Methods Fluorescence-activated cell sorting, RNA sequencing, quantitative real-time PCR, Western blotting, small interfering RNA interference, and kinase inhibitors were used to study the molecular mechanisms in isolated human CD4 + T cells from healthy donors. The clinical implication was studied in blood samples of patients with inflammatory bowel disease (IBD) receiving anti-TNF therapy. Results Here we show that anti-TNF treatment results in inhibition of the anti-inflammatory molecule TNF-α–induced protein 3 (TNFAIP3) /A20 in memory CD4 + T cells. We found an inverse relationship between TNFAIP3 /A20 expression levels and IL-17A production. Inhibition of TNFAIP3 /A20 promotes kinase activity of p38 mitogen-activated protein kinase and protein kinase C, which drives IL-17A expression. Regulation of TNFAIP3 /A20 expression and cognate IL-17A production in T cells are specifically mediated through TNF receptor 2 signaling. Ex vivo , in patients with IBD treated with anti-TNF, we found further evidence for an inverse relationship between TNFAIP3 /A20 expression levels and IL-17A–producing T cells. Conclusion Anti-TNF treatment interferes in the TNFAIP3 /A20-mediated anti-inflammatory feedback loop in CD4 + T cells and promotes kinase activity. This puts TNFAIP3 /A20, combined with IL-17A expression, on the map as a potential tool for predicting therapy responsiveness or side effects of anti-TNF therapy. Moreover, it provides novel targets related to TNFAIP3 /A20 activity for superior therapeutic regimens in patients with IBD. [ABSTRACT FROM AUTHOR]

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Background: Hematopoietic cell transplantation (HCT) is an effective treatment for pediatric patients with high-risk, refractory, or relapsed acute myeloid leukemia (AML). However, a large proportion of transplanted patients eventually die due to relapse. To improve overall survival, we propose a combined strategy based on cord blood (CB)-HCT with the application of AML-specific T cell receptor (TCR)-engineered T cell therapy derived from the same CB graft., Methods: We produced CB-CD8 + T cells expressing a recombinant TCR (rTCR) against Wilms tumor 1 (WT1) while lacking endogenous TCR (eTCR) expression to avoid mispairing and competition. CRISPR-Cas9 multiplexing was used to target the constant region of the endogenous TCRα ( TRAC ) and TCRβ ( TRBC ) chains. Next, an optimized method for lentiviral transduction was used to introduce recombinant WT1-TCR. The cytotoxic and migration capacity of the product was evaluated in coculture assays for both cell lines and primary pediatric AML blasts., Results: The gene editing and transduction procedures achieved high efficiency, with up to 95% of cells lacking eTCR and over 70% of T cells expressing rWT1-TCR. WT1-TCR-engineered T cells lacking the expression of their eTCR (eTCR -/- WT1-TCR) showed increased cell surface expression of the rTCR and production of cytotoxic cytokines, such as granzyme A and B, perforin, interferon-γ (IFNγ), and tumor necrosis factor-α (TNFα), on antigen recognition when compared with WT1-TCR-engineered T cells still expressing their eTCR (eTCR +/+ WT1-TCR). CRISPR-Cas9 editing did not affect immunophenotypic characteristics or T cell activation and did not induce increased expression of inhibitory molecules. eTCR -/- WT1-TCR CD8 + CB-T cells showed effective migratory and killing capacity in cocultures with neoplastic cell lines and primary AML blasts, but did not show toxicity toward healthy cells., Conclusions: In summary, we show the feasibility of developing a potent CB-derived CD8 + T cell product targeting WT1, providing an option for post-transplant allogeneic immune cell therapy or as an off-the-shelf product, to prevent relapse and improve the clinical outcome of children with AML., Competing Interests: Competing interests: JK reports grants from Gadeta, Novartis, and Miltenyi Biotec and is the inventor of patents dealing with γδT cell-related aspects, as well as the cofounder and shareholder of Gadeta. ZS is an inventor of patents dealing with γδT cell-related aspects., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Human leukocyte antigen (HLA) restriction of conventional T-cell targeting introduces complexity in generating T-cell therapy strategies for patients with cancer with diverse HLA-backgrounds. A subpopulation of atypical, major histocompatibility complex-I related protein 1 (MR1)-restricted T-cells, distinctive from mucosal-associated invariant T-cells (MAITs), was recently identified recognizing currently unidentified MR1-presented cancer-specific metabolites. It is hypothesized that the MC.7.G5 MR1T-clone has potential as a pan-cancer, pan-population T-cell immunotherapy approach. These cells are irresponsive to healthy tissue while conferring T-cell receptor(TCR) dependent, HLA-independent cytotoxicity to a wide range of adult cancers. Studies so far are limited to adult malignancies. Here, we investigated the potential of MR1-targeting cellular therapy strategies in pediatric cancer. Bulk RNA sequencing data of primary pediatric tumors were analyzed to assess MR1 expression. In vitro pediatric tumor models were subsequently screened to evaluate their susceptibility to engineered MC.7.G5 TCR-expressing T-cells. Targeting capacity was correlated with qPCR-based MR1 mRNA and protein overexpression. RNA expression of MR1 in primary pediatric tumors varied widely within and between tumor entities. Notably, embryonal tumors exhibited significantly lower MR1 expression than other pediatric tumors. In line with this, most screened embryonal tumors displayed resistance to MR1T-targeting in vitro MR1T susceptibility was observed particularly in pediatric leukemia and diffuse midline glioma models. This study demonstrates potential of MC.7.G5 MR1T-cell immunotherapy in pediatric leukemias and diffuse midline glioma, while activity against embryonal tumors was limited. The dismal prognosis associated with relapsed/refractory leukemias and high-grade brain tumors highlights the promise to improve survival rates of children with these cancers., Competing Interests: Competing interests: ZS and JK are inventors on different patents for γδ TCR sequences, recognition mechanisms and isolation strategies. JK is scientific cofounder and shareholder of Gadeta (www.gadeta.nl). The remaining authors declare no competing interests., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)