Development of a sea anemone toxin as an immunomodulator for therapy of autoimmune diseases
Highlights
► ShK-186 is a 37-residue synthetic derivative of ShK, a peptide toxin from the sea anemone Stichodactyla helianthus. ► ShK-186 blocks Kv1.3 potassium channels with picomolar affinity and with a high degree of specificity. ► Kv1.3 potassium channels play a critical role in regulating the function of effector-memory T cells and class-switched memory B cells that are implicated in autoimmune diseases. ► ShK-186 has a good safety profile in rats and it ameliorates disease in rat models of multiple sclerosis and rheumatoid arthritis. ► ShK-186 is being developed as a therapeutic for autoimmune diseases and phase 1 human trials are planned for the near future.
Section snippets
The clinical problem – autoimmune diseases
Nearly 80 different autoimmune disorders are known, affecting more than 125 million people worldwide. Autoimmune diseases involve virtually every organ system in the body including joints (e.g. rheumatoid arthritis [RA], ankylosing spondylitis), the central nervous system (multiple sclerosis [MS]), endocrine organs (type-1 diabetes mellitus [T1DM], Hashimoto's thyroiditis) (Leyendeckers et al., 2002) and skin (psoriasis). Tissue destruction is mediated by autoreactive (self-reactive) immune
K+ channels promote calcium influx in lymphocytes
Calcium signaling is essential for lymphocytes to activate, synthesize and secrete cytokines (or antibodies), migrate in vivo, and proliferate. In the case of T cells, antigen binding to the T-cell receptor causes the generation of IP3, which releases Ca2+ from the ER Ca2+ store (Fig. 1). Depletion of the ER store triggers an ER protein called STIM1 to cluster under the plasma membrane and activate Ca2+ influx through CRAC channels formed from Orai1 subunits (Lioudyno et al., 2008, Luik et al.,
The discovery of ShK and development of ShK-186
Stichodactyla helianthus is a common species of sea anemone in the Caribbean Sea around Cuba. In 1995, Olga Castaneda, Evert Karlsson, Alan Harvey, Reto Stöcklin and their colleagues found that S. helianthus extracts administered to mice by intraperitoneal injection, induced hypersensitivity to touch and sound, excessive salivation, lacrimation, sweating, motor incoordination and paralysis, reminiscent of poisoning by cholinesterase inhibitors (Castaneda and Harvey, 2009, Castaneda et al., 1995
In vitro studies
ShK-186 inhibits Ca2+ signaling in human TEM clones in a dose-dependent fashion with an IC50 ≈ 200 pM (Beeton et al., 2006). ShK-186 is significantly more effective in suppressing IL-2 and IFN-γ production by human CCR7− TEM cells than CCR7+ naïve/TCM cells, but it is less effective in suppressing the production of TNF-α and IL-4 by either subset (Fig. 5) (Beeton et al., 2006). ShK-186 is ≈10-fold more effective in suppressing proliferation ([3H]thymidine incorporation) by human CCR7− TEM cells
Manufacture and formulation of ShK-186
ShK-186 has been manufactured under full GMP regulations at up to the 30 g scale using standard Fmoc-tBu solid-phase synthesis methods. Synthesis at this scale demonstrates the robustness of the manufacturing process, and is adequate to support clinical development through the expected range of human doses (approximately 1–15 μg/kg/week). Disulfide bond formation is achieved by air oxidation or using a glutathione exchange reaction. The manufacturing process provides a consistently pure and
Overview and future directions
Three ion channels—Kv1.3, KCa3.1, Stim/Orai—regulate calcium signaling during the activation process in lymphocytes. All three of these channels cluster at the immunological synapse, the narrow cleft formed between the antigen-presenting cell (e.g. dendritic cell or macrophage or class-switched memory B cell) and the T lymphocyte. Kv1.3 and KCa3.1 regulate the membrane potential of lymphocytes and allow efflux of positively-charged ions (K+) to counterbalance the influx of Ca2+ through CRAC
Conflict of interest statement
The authors (KGC, RSN, MWP, HW, CB) are co-founders of Airmid Inc., which has licensed a patent on PAP-1 from the University of California. Airmid has sub-licensed the patent to Circassia Inc., who is developing the drug as a therapeutic for autoimmune disease (B S-F).
The authors (SI, ET, LL, B S-F) are co-founders of Kineta Inc., which has licensed a patent on ShK-186 from the University of California. Kineta is developing the drug as a therapeutic for autoimmune disease.
The authors (KGC, MWP,
Acknowledgments
The work was supported by grant numbers: NIH RO1 NS48252 (KGC), NIH 1R43AI085691 (SI, KGC), NIH RO1 GM076063 (HW), UC Discovery 445160 (KGC, SI), Iacocca Foundation 485160 (SI, JTL, KGC), and fellowship support from the National Health and Medical Research Council of Australia (RSN).
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2022, Journal of Biological ChemistryCitation Excerpt :The activation of MAPK pathway is necessary during DTH (36, 37), which directly or indirectly mediates the production of inflammatory cytokines such as TNF-α in vivo (38–40). The Kv1.3 blocking peptides such as MgTx, KTX, ShK-186, and BmKTX-D33H ameliorate autoimmune DTH reaction characterized by tissue swelling and infiltration in the subcutaneous layer and have no obvious side effects in vivo (9, 13, 22, 41–43). Consistent with these findings and its effects in Jurkat T cells, like MgTx, FS48 markedly suppresses the ear swelling (Figs. 6, A and B and S1), the production of TNF-α and IL-2 and inflammatory infiltrate in an OVA-induced DTH response (Fig. 6, C–E).