Background/Purpose: Phosphatidylinositol (3,5)-bisphosphate (PI(3,5)P₂) is the most recently-identified phospholipid component of cellular membrane. Phosphatidylinositol 3-phosphate 5-kinase (PIKfyve) is a critical enzyme for the synthesis of PI(3,5)P₂ from phosphatidylinositol 3-monophosphate (PI3P), which has been implicated in intracellular trafficking events, but little is known about its biological function. We discovered a lead-compound APY0201 which is a potent, highly selective PIKfyve kinase inhibitor. In this study, we characterized anti-inflammatory properties of APY0201 in vitro and in vivo.

Methods: In vitro study: Mouse macrophages (thioglycolate-induced peritoneal exudate cells) and human peripheral blood mononuclear cells (PBMCs) stimulated by IFN-γ and heat-killed whole bacteria (Staphylococcus aureusCowan strain I (SAC)) were used in this assay.

In vivo study: The imiquimod (IMQ)-induced psoriasis model was used. Female Balb/c mice received daily topical application of approx. 16 mg 0.5% IMQ cream & Vaseline mix (1:1, equivalent to 0.4 mg IMQ) on the right ear for 4 days. APY0201 or vehicle was orally or topically administered once daily. The severity of inflammation was assessed by daily ear thickness and ear weight after 4 days of IMQ-treatment.

Results: APY0201 inhibited the conversion of PI3P to PI(3,5)P₂ in the presence of recombinant human PIKfyve with an IC₅₀ value of 8.9 nM. It is highly selective for PIKfyve compared to various receptors, channels and enzymes including 25 lipid kinases and 83 protein kinases. Interestingly, APY0201 demonstrated potent in vitro inhibitory activity against interleukin(IL)-12p70 production from mouse macrophages and human PBMCs with an IC₅₀ value of 8.4 nM and 9.9 nM, respectively. Furthermore, IL-12/23 production but not other inflammatory mediators, such as TNF-α and MCP-1 from these cells were inhibited by APY0201. Downregulation of PIKfyve using siRNA of PIKfyve inhibited expression of IL-12 subunit mRNA in IFN-γ/SAC-activated mouse macrophages. These results indicate APY0201 exerts its inhibitory activity of IL-12/23 production via PIKfyve inhibition. In ex vivostudy, IL-12p70 production was significantly inhibited in IFN-γ/SAC-activated whole blood from mice orally administered with APY0201. Finally, daily oral or topical administration of APY0201 significantly ameliorated skin inflammation in mouse psoriasis-like model.

Conclusion: The therapeutic potential of IL-12/23 inhibition in Th1/Th17-mediated immunological disorders, such as psoriasis has been clinically validated on the efficacy of ustekinumab, anti-IL-12/23 antibody. Our results clearly indicate that PIKfyve inhibition contribute to the suppression of inflammation in both in vitro and in vivo. Consequently APY0201, highly selective PIKfyve kinase inhibitor might be a promising medication for the treatment of patients with psoriasis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/a-novel-orally-active-phosphatidylinositol-3-phosphate-5-kinase-pikfyve-inhibitor-ameliorates-mouse-psoriasis-like-model-by-inhibition-of-interleukin-12-and-interleukin-23-production-from-macrophage/