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The novel quinoline derivative SKA-346 as a KCa3.1 channel selective activator.
No full textThe calcium-activated KCa3.1 channel plays a crucial role in T-cell immune response. Genetic manipulation of T-cells to upregulate the expression of K+ channels has been shown to boost T-cell cytotoxicity in cancer. Here, we aimed to identify and characterize an activator that would augment KCa3.1 currents without affecting other channels. We synthesized five quinoline derivatives and used electrophysiology to screen them on KCa3.1 and a panel of 14 other ion channels. One quinoline derivative, SKA-346, activated KCa3.1 with an EC50 of 1.9 μM and showed selectivity against the other channels. In silico analysis using RosettaLigand and GLIDE demonstrated a well-converged pose of SKA-346 in a binding pocket at the interface between the calmodulin N-lobe and the S45A helix in the S4-S5 linker of the KCa3.1 channel. SKA-346 (30 mg kg-1), tolerated by mice after intra-peritoneal administration, exhibited a peak plasma concentration of 6.29 μg mL-1 (29.2 μM) at 15 min and a circulating half-life (t 1/2) of 2.8 h. SKA-346 could serve as a template for the development of more potent KCa3.1 activators to enhance T-cell cytotoxicity in cancer
Rearrangement of a unique Kv1.3 selectivity filter conformation upon binding of a drug
No full textWe report two structures of the human voltage-gated potassium channel (Kv) Kv1.3 in immune cells alone (apo-Kv1.3) and bound to an immunomodulatory drug called dalazatide (dalazatide–Kv1.3). Both the apo-Kv1.3 and dalazatide–Kv1.3 structures are in an activated state based on their depolarized voltage sensor and open inner gate. In apo-Kv1.3, the aromatic residue in the signature sequence (Y447) adopts a position that diverges 11 Å from other K(+) channels. The outer pore is significantly rearranged, causing widening of the selectivity filter and perturbation of ion binding within the filter. This conformation is stabilized by a network of intrasubunit hydrogen bonds. In dalazatide–Kv1.3, binding of dalazatide to the channel’s outer vestibule narrows the selectivity filter, Y447 occupies a position seen in other K(+) channels, and this conformation is stabilized by a network of intersubunit hydrogen bonds. These remarkable rearrangements in the selectivity filter underlie Kv1.3’s transition into the drug-blocked state
Imaging Kv1.3 Expressing Memory T Cells as a Marker of Immunotherapy Response
No full textImmune checkpoint inhibitors have shown great promise, emerging as a new pillar of treatment for cancer; however, only a relatively small proportion of recipients show a durable response to treatment. Strategies that reliably differentiate durably-responding tumours from non-responsive tumours are a critical unmet need. Persistent and durable immunological responses are associated with the generation of memory T cells. Effector memory T cells associated with tumour response to immune therapies are characterized by substantial upregulation of the potassium channel Kv1.3 after repeated antigen stimulation. We have developed a new Kv1.3 targeting radiopharmaceutical, [18F]AlF-NOTA-KCNA3P, and evaluated whether it can reliably differentiate tumours successfully responding to immune checkpoint inhibitor (ICI) therapy targeting PD-1 alone or combined with CLTA4. In a syngeneic colon cancer model, we compared tumour retention of [18F]AlF-NOTA-KCNA3P with changes in the tumour immune microenvironment determined by flow cytometry. Imaging with [18F]AlF-NOTA-KCNA3P reliably differentiated tumours responding to ICI therapy from non-responding tumours and was associated with substantial tumour infiltration of T cells, especially Kv1.3-expressing CD8+ effector memory T cells
Design and Structural Basis of Selective 1,4-dihydropyridine Inhibitors of the Calcium-activated Potassium Channel K\u3csub\u3eCa\u3c/sub\u3e3.1
No full textThe 1,4-dihydropyridines, drugs with well-established bioavailability and toxicity profiles, have proven efficacy in treating human hypertension, peripheral vascular disorders, and coronary artery disease. Every 1,4-dihydropyridine in clinical use blocks L-type voltage-gated calcium channels. We now report our development, using selective optimization of a side activity (SOSA), of a class of 1,4-dihydropyridines that selectively and potently inhibit the intermediate-conductance calcium-activated K+ channel KCa3.1, a validated therapeutic target for diseases affecting many organ systems. One of these 1,4-dihydropyridines, DHP-103, blocked KCa3.1 with an IC50 of 6 nM and exhibited exquisite selectivity over calcium channels and a panel of \u3e100 additional molecular targets. Using high-resolution structure determination by cryogenic electron microscopy together with mutagenesis and electrophysiology, we delineated the drug binding pocket for DHP-103 within the water-filled central cavity of the KCa3.1 channel pore, where bound drug directly impedes ion permeation. DHP-103 inhibited gain-of-function mutant KCa3.1 channels that cause hereditary xerocytosis, suggesting its potential use as a therapeutic for this hemolytic anemia. In a rat model of acute ischemic stroke, the second leading cause of death worldwide, DHP-103 administered 12 h postischemic insult in proof-of-concept studies reduced infarct volume, improved balance beam performance (measure of proprioception) and decreased numbers of activated microglia in infarcted areas. KCa3.1-selective 1,4-dihydropyridines hold promise for the many diseases for which KCa3.1 has been experimentally confirmed as a therapeutic target
