Human Double-Negative Regulatory T-Cells Induce a Metabolic and Functional Switch in Effector T-Cells by Suppressing mTOR Activity

The recently discovered population of TCRαβ+ CD4–/CD8– (double-negative, DN) T-cells are highly potent suppressor cells in mice and humans. In preclinical transplantation models, adoptive transfer of DN T-cells specifically inhibits alloreactive T-cells and prevents transplant rejection or graft-vs.-host disease (GvHD). Interestingly, clinical studies in patients who underwent allogeneic stem cell transplantation reveal an inverse correlation between the frequency of circulating DN T-cells and the severity of GvHD, suggesting a therapeutic potential of human DN T-cells. However, their exact mode of action has not been elucidated yet. Investigating the impact of DN T-cells on conventional T-cells, we found that human DN T-cells selectively inhibit mTOR signaling in CD4 T-cells. Given that mTOR is a critical regulator of cellular metabolism, we further determined the impact of DN T-cells on the metabolic framework of T-cells. Intriguingly, DN T-cells diminished expression of glucose transporters and glucose uptake, whereas fatty acid uptake was not modified, indicating that DN T-cells prevent metabolic adaptation of CD4 T-cells upon activation (i.e., glycolytic switch) thereby contributing to their suppression. Further analyses demonstrated that CD4 T-cells also do not upregulate homing receptors associated with inflammatory processes. In contrast, expression of central memory-cell associated cell surface markers and transcription factors were increased by DN T-cells. Moreover, CD4 T-cells failed to produce inflammatory cytokines after co-culture with DN T-cells, whereas IL-2 secretion was enhanced. Taken together DN T-cells impair metabolic reprogramming of conventional CD4 T-cells by abrogating mTOR signaling, thereby modulating CD4 T-cell functionality. These results uncover a new mechanism of DN T-cell-mediated suppression, pointing out that DN T-cells could serve as cell-based therapy to limit alloreactive immune response

Selective PRMT5 Inhibitors Suppress Human CD8+ T Cells by Upregulation of p53 and Impairment of the AKT Pathway Similar to the Tumor Metabolite MTA

Genetic alterations in tumor cells provide promising targets for antitumor therapy. Recently, loss of methylthioadenosine phosphorylase (MTAP), a deletion frequently occurring in cancer, has been shown to create vulnerability to the inhibition of the protein arginine methyltransferase 5 (PRMT5). MTAP deficiency leads to accumulation of methylthioadenosine (MTA), which reduces PRMT5 activity, and thus, sensitizes the tumor cells to selective PRMT5 inhibitors (PRMT5i). PRMT5i are investigated as a new strategy to selectively kill MTAP-deficient tumor cells by blocking residual PRMT5 activity, but also to treat PRMT5-overexpressing tumors. Although many studies investigated the role of PRMT5 in cancer, only little data exist about the effect of PRMT5 inhibition on immune cells. As we could show that the tumor metabolite MTA suppresses T cells, we asked whether selective PRMT5 inhibition is detrimental for T-cell immune responses. Therefore, we examined the effect of the synthetic PRMT5 inhibitor EPZ015666 on human CD8(+) T cells in direct comparison with the naturally occurring PRMT5-inhibiting molecule MTA. Both compounds reduced T-cell proliferation, viability, and functionality. In addition, T-cell metabolism was impaired upon PRMT5 inhibition. These effects coincided with the induction of p53 expression and reduced AKT/mTOR signaling. Our data clearly demonstrate that PRMT5 activity is involved in various cellular processes of human CD8(+) T cells associated with essential T-cell functions. Therefore, not only tumor cells, but also antitumor immune responses, are compromised by PRMT5 inhibitors. This emphasizes the importance of considering side effects on the immune system when developing new strategies to specifically target not only MTAP-deficient tumors

Structural insights into TRPV2 activation by small molecules

Transient receptor potential vanilloid 2 (TRPV2) is involved in many critical physiological and pathophysiological processes, making it a promising drug target. Here we present cryo-electron microscopy (cryo-EM) structures of rat TRPV2 in lipid nanodiscs activated by 2-aminoethoxydiphenyl borate (2-APB) and propose a TRPV2-specific 2-ABP binding site at the interface of S5 of one monomer and the S4-S5 linker of the adjacent monomer. In silico docking and electrophysiological studies confirm the key role of His521 and Arg539 in 2-APB activation of TRPV2. Additionally, electrophysiological experiments show that the combination of 2-APB and cannabidiol has a synergetic effect on TRPV2 activation, and cryo-EM structures demonstrate that both drugs were able to bind simultaneously. Together, our cryo-EM structures represent multiple functional states of the channel, providing a native picture of TRPV2 activation by small molecules and a structural framework for the development of TRPV2-specific activators

Structural insights into TRPV2 activation by small molecules.

Transient receptor potential vanilloid 2 (TRPV2) is involved in many critical physiological and pathophysiological processes, making it a promising drug target. Here we present cryo-electron microscopy (cryo-EM) structures of rat TRPV2 in lipid nanodiscs activated by 2-aminoethoxydiphenyl borate (2-APB) and propose a TRPV2-specific 2-ABP binding site at the interface of S5 of one monomer and the S4-S5 linker of the adjacent monomer. In silico docking and electrophysiological studies confirm the key role of His521 and Arg539 in 2-APB activation of TRPV2. Additionally, electrophysiological experiments show that the combination of 2-APB and cannabidiol has a synergetic effect on TRPV2 activation, and cryo-EM structures demonstrate that both drugs were able to bind simultaneously. Together, our cryo-EM structures represent multiple functional states of the channel, providing a native picture of TRPV2 activation by small molecules and a structural framework for the development of TRPV2-specific activators