IPI-3063 is a potent and selective p110δ inhibitor with biochemical IC50 of 2.5?±?1.2 nM and IC50 values for the other class I PI3K isoforms (p110α, p110β, p110γ) are at least 400-fold higher.
IPI-3063 is a potent and selective p110δ inhibitor with biochemical IC50 of 2.5?±?1.2 nM and IC50 values for the other class I PI3K isoforms (p110α, p110β, p110γ) are at least 400-fold higher.
IPI-3063 is a p110δ selective compound with an IC50?=?0.1?nM in p110δ-specific cell-based assays and cellular IC50 values for the other class I PI3K isoforms are at least 1,000-fold higher. IPI-3063 potently reduces mouse B cell proliferation, survival, and plasmablast differentiation[1].
IPI-3063 has good pharmacokinetics in mice[1].
[1] Honyin Chiu, et al. Front Immunol. 2017, 8: 747. [2] Winkler DG, et al. Chem Biol. 2013, 20(11):1364-74.
Kinase experiment: | Human recombinant PI3K-α, PI3K-β, PI3K-δ, and PI3K-γ are used. Phosphatidylinositol 4,5 bis phosphate (diC8-PtdIns(4,5)P2) is used. PI3K-α, β, and δ are heterodimers consisting of full length p110α, p110β, or p110δ catalytic subunit and the p85α regulatory subunit. PI3K-γ is a monomer of the p110γ catalytic subunit. Samples of kinase (10?nM-α, β, and δ; 20?nM-γ) are incubated with IPI-3063 for 30?min at room temperature in reaction buffer (15?mM HEPES pH 7.4, 20?mM NaCl, 1?mM EGTA, 0.02% Tween 20, 10?mM MgCl2, 0.2?mg/mL bovine-γ-globulins) followed by addition of ATP/diC8-PtdIns(4,5)P2 mixture to give final concentrations of 3?mM ATP and 500?μM diC8-PtdIns(4,5)P2. Reactions are incubated at room temperature for 2?h, with PI3K activity is assessed. Plates are read on plate reader in luminescence mode[1]. |
Cell experiment: | Peripheral blood mononuclear cells (PBMCs) are first purified from blood by density gradient centrifugation. Human B cells are then purified from PBMCs by negative selection. B-cell purity is increased from 4% to >70% as measured by FACS analysis using anti-CD19 PE conjugated antibody. Purified B cells are seeded at a final concentration of 0.1×106?cells/mL and cultured with 2?μg/mL human CD40L+5?μg/mL anti-human IgM/IgG+100?μg/mL hIL-2+100?μg/mL hIL-21. All B cells are cultured in RPMI 1640 supplemented with 10% (vol/vol) heat-inactivated FCS, 5?mM Hepes, 2?mM L-glutamine, 100?U/mL Penicillin, 100?μg/mL Streptomycin, 50?μM 2-mercaptoethanol. Purified human B cells are pretreated with IPI-3063 (0.1, 1, 10, and 100?nM) for 30?min, then stimulated with human CD40L+anti-human IgM/IgG+human IL-2+human IL-21 for 120?h[1]. |
参考文献: [1]. Chiu H, et al. The Selective Phosphoinoside-3-Kinase p110δ Inhibitor IPI-3063 Potently Suppresses B Cell Survival, Proliferation, and Differentiation. Front Immunol. 2017 Jun 30;8:747. | |
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