Exploring the Role of the Voltage-Gated Potassium Channel Kv1.3 in Lymphocyte Function in vivo in Pathological Contexts

Regulatory T cells (Tregs) play a crucial role in the maintenance of self-tolerance and in the resolution of immune responses, thanks to their immunosuppressive activity. Their significance extends to various diseases, notably in cancer, where they exert a pivotal influence within the tumor microenvironment, impeding anti-tumor immunity and promoting tumor advancement. The voltage-gated potassium channel Kv1.3 exhibits heightened expression in lymphocytes, where it is localized both in the plasma membrane and inner mitochondrial membrane. In these cells, Kv1.3 regulates T cell proliferation and cytokines expression. However, the impact of Kv1.3 on Tregs in the context of tumor growth remains largely unexplored. Therefore, we developed a novel regulatory T cell-specific knockout mouse model to study the effects of Kv1.3 ablation in Tregs in the context of oncological diseases. Our findings reveal that genetic Kv1.3 ablation in Tregs impedes their suppressive activity and IL-10 secretion in vitro. Moreover, our results underscore the pivotal role of Kv1.3 in sustaining Treg suppressive function within the tumor microenvironment, as evidenced by reduced GITR expression in tumor-infiltrating knockout Tregs. Notably, tumors in knockout animals exhibit decreased infiltration of activated ICOS+ and GITR+ Tregs. Finally, we show that it is the plasma membrane, rather than the inner mitochondrial membrane Kv1.3 that regulates Tregs activity. These findings unveil the unexplored influence of Kv1.3 in modulating Treg function within tumors and highlight the potential of targeting Kv1.3 in Tregs as a promising strategy for reshaping the tumor microenvironment. The majority of the inflammatory infiltrate in the brain of individuals with multiple sclerosis (MS) is composed of activated effector memory T lymphocytes (TEM) that have undergone repeated antigen stimulation throughout the course of the disease. Therefore, there is a great need for therapies targeting chronically activated autoreactive T cells, the major mediators in MS pathogenesis, without causing general immunosuppression or toxicity. The plasma membrane-located Kv1.3 potassium channel is highly expressed by chronically activated TEMs and is crucial for their proliferation, while inhibition of the mitochondrial counterpart of Kv1.3 (mitoKv1.3) leads to apoptosis. Here, we explored a new strategy to selectively kill chronically activated TEMs, through pharmacological targeting of an intracellular ion channel, namely mitoKv1.3. The effect of PAPTP, a specific, mitoKv1.3 inhibitor was investigated on autoproliferative T cells from remitting-relapsing HLA-DRB1*15:01 positive MS patients treated with Natalizumab (NAT). A significant reduction of the percentage of autoproliferative TEM cells (on total CD4+ T cells) occurred following treatment with 1μM PAPTP. This decrease was mirrored by an increased percentage of apoptotic TEM cells upon treatment with PAPTP. The relevance of these findings was underlined showing that PAPTP was able to selectively kill activated TEMs and completely halted MS progression in the experimental autoimmune encephalitis (EAE) mouse MS model, strongly suggesting that elimination of autoreactive TEMs in the peripheral blood is sufficient to halt the disease. PAPTP did not cause toxicity and immune depletion. In addition, pre-treatment of whole T cell population with PAPTP in an adoptive transfer model, completely prevented EAE onset. Altogether, our data indicate that PAPTP is able to selectively eliminate autoproliferative effector memory T cells in the periphery and has a very strong beneficial effect on MS progression in the EAE mouse model as well as on blood cells of NAT-treated patients.