Identification of WP1066, an inhibitor of JAK2 and STAT3, as a Kv1.3 potassium channel blocker.

BACKGROUND AND PURPOSE Kv1.3 potassium channels play a predominant role in regulating calcium signaling that is essential for the activation and proliferation of effector memory T (TEM ) cells. The channel has been recognized as a promising therapeutic target against various autoimmune diseases. EXPERIMENTAL APPROACH In a high-throughput screening campaign, WP1066 was identified as a Kv1.3 channel inhibitor. Using molecular biology and electrophysiology methods, the mechanism of WP1066 blocking Kv1.3 channels was investigated. Using TEM cell proliferation assay and mouse delayed-type hypersensitivity (DTH) model, the effects of WP1066 were examined. KEY RESULTS WP1066 blocked Kv1.3 channels in a dose-dependent manner with an IC50 of 3.2 ± 0.04 μM and induced a hyperpolarizing shift of the steady-state inactivation curve. In addition, WP1066 blocked Kv1.3 channels use-dependently by preferentially interacting with channels in their open-state instead of closed-state and inactivated-state. When the residues located in the S6 domain scaffolding the inner vestibule were sequentially mutated, the potencies of WP1066 were significantly impaired, especially by mutations A413C and I420C, indicating a higher affinity of interacting sites for WP1066. Moreover, WP1066 effectively suppressed mouse TEM cell proliferation in vitro and mouse DTH reaction in vivo. CONCLUSIONS AND IMPLICATIONS In conclusion, the present study identified WP1066 as a Kv1.3 potassium channel blocker with an open-state dependent property, which provides fundamental evidences for the application of WP1066 in further immunomodulatory studies targeting Kv1.3 potassium channels.