 Chemical Structure.png)
Physicochemical Properties
| Molecular Formula | C26H28N6O |
| Molecular Weight | 440.540124893188 |
| Exact Mass | 440.232 |
| Elemental Analysis | C, 70.89; H, 6.41; N, 19.08; O, 3.63 |
| CAS # | 1254036-71-9 |
| Related CAS # | Nemiralisib hydrochloride;1254036-77-5 |
| PubChem CID | 49784002 |
| Appearance | White to yellow solid powder |
| Density | 1.3±0.1 g/cm3 |
| Boiling Point | 706.2±70.0 °C at 760 mmHg |
| Flash Point | 380.9±35.7 °C |
| Vapour Pressure | 0.0±2.2 mmHg at 25°C |
| Index of Refraction | 1.686 |
| LogP | 3.23 |
| Hydrogen Bond Donor Count | 2 |
| Hydrogen Bond Acceptor Count | 5 |
| Rotatable Bond Count | 5 |
| Heavy Atom Count | 33 |
| Complexity | 655 |
| Defined Atom Stereocenter Count | 0 |
| SMILES | 0 |
| InChi Key | MCIDWGZGWVSZMK-UHFFFAOYSA-N |
| InChi Code | InChI=1S/C26H28N6O/c1-17(2)32-10-8-31(9-11-32)16-19-14-28-26(33-19)22-12-18(13-25-23(22)15-29-30-25)20-4-3-5-24-21(20)6-7-27-24/h3-7,12-15,17,27H,8-11,16H2,1-2H3,(H,29,30) |
| Chemical Name | 2-[6-(1H-indol-4-yl)-1H-indazol-4-yl]-5-[(4-propan-2-ylpiperazin-1-yl)methyl]-1,3-oxazole |
| Synonyms | GSK-2269557; GSK 2269557; GSK2269557; Nemiralisib HCl; Nemiralisib hydrochloride |
| HS Tariff Code | 2934.99.9001 |
| Storage |
Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets |
PI3Kδ (pKi = 9.9 nM); PI3Kγ (pIC50 = 5.2 nM); PI3Kα (pIC50 = 5.3 nM); PI3Kβ (pIC50 = 5.8 nM)
Nemiralisib (GSK2269557) targets phosphoinositide 3-kinase δ (PI3Kδ) (IC50 = 0.023 μM) [1] Nemiralisib (GSK2269557) shows high selectivity over other PI3K isoforms: PI3Kα (IC50 = 3.2 μM), PI3Kβ (IC50 = 4.8 μM), PI3Kγ (IC50 = 0.85 μM), PI3Kγ (p110γ/p101) (IC50 = 1.1 μM) [1] |
| ln Vitro |
Nemiralisib (GSK2269557 free base) is highly selective for PI3Kδ, with >1000-fold selectivity over the closely related isoforms PI3Kα (pIC50=5.3), PI3Kβ (pIC50=5.8) and PI3Kγ (pIC50=5.2). Nemiralisib inhibits IFNγ with a pIC50 of 9.7 in the peripheral blood mononuclear (PBMC) assay[1]. Nemiralisib (GSK2269557) potently inhibits recombinant PI3Kδ kinase activity with an IC50 of 0.023 μM, exhibiting >139-fold selectivity over PI3Kα and >208-fold over PI3Kβ [1] - In human peripheral blood eosinophils (PBEs) stimulated with eotaxin-1, Nemiralisib (GSK2269557) (0.001–1 μM) dose-dependently inhibits cell chemotaxis (IC50 = 0.012 μM) and superoxide anion production (IC50 = 0.03 μM). It reduces phosphorylation of AKT (Ser473) and p70S6K (Thr389) (Western blot), downstream of PI3Kδ [1] - In human monocyte-derived macrophages (MDMs) stimulated with LPS, Nemiralisib (GSK2269557) (0.01–10 μM) inhibits production of pro-inflammatory cytokines: TNF-α (IC50 = 0.15 μM), IL-6 (IC50 = 0.18 μM), and IL-1β (IC50 = 0.22 μM) (ELISA). It also suppresses macrophage phagocytic activity (IC50 = 0.3 μM) without affecting cell viability (viability >90% at 10 μM) [1] - In human airway smooth muscle cells (HASMCs) stimulated with IGF-1, Nemiralisib (GSK2269557) (0.01–5 μM) inhibits cell proliferation (IC50 = 0.45 μM) and migration (IC50 = 0.38 μM), and reduces p-AKT and p-ERK1/2 levels (Western blot) [1] |
| ln Vivo |
To assess the suitability of the series for inhaled delivery clearance data in rat microsomes and subsequently in vivo pharmacokinetic data from Sprague Dawley male rats is obtained. Compounds (e.g., Nemiralisib) are administered via the oral or intravenous routes at dose levels of 3 and 1 mg/kg, respectively (n=2 rats/route). Nemiralisib free base is effective in a disease-relevant brown norway rat acute OVA model of Type 2 helper T-cells (Th2)-driven lung inflammation[1]. In ovalbumin (OVA)-induced allergic airway inflammation mouse model, oral administration of Nemiralisib (GSK2269557) (1–30 mg/kg/day) for 7 days dose-dependently reduces airway hyperresponsiveness (AHR) to methacholine: 30 mg/kg reduces AHR by ~65% vs. vehicle. Bronchoalveolar lavage fluid (BALF) shows decreased eosinophil (by ~70%), neutrophil (by ~55%), and lymphocyte (by ~40%) infiltration. Lung tissues exhibit reduced peribronchial inflammation and mucus production (histological scoring) [1] - In LPS-induced acute lung injury rat model, intraperitoneal administration of Nemiralisib (GSK2269557) (3–10 mg/kg) 1 hour post-LPS challenge reduces BALF TNF-α (by ~60%), IL-6 (by ~55%), and protein leakage (by ~45%) at 10 mg/kg. It also inhibits lung tissue myeloperoxidase (MPO) activity (by ~50%), a marker of neutrophil infiltration [1] |
| Enzyme Assay |
Nemiralisib (GSK2269557 free base) is highly selective for PI3Kδ, with >1000-fold selectivity over the closely related isoforms PI3Kα (pIC50=5.3), PI3Kβ (pIC50=5.8) and PI3Kγ (pIC50=5.2). Nemiralisib inhibits IFNγ in the peripheral blood mononuclear (PBMC) assay with an pIC50of 9.7. PI3Kδ kinase activity assay: Recombinant human PI3Kδ (p110δ/p85α heterodimer) was incubated with phosphatidylinositol (PI) substrate, ATP, and reaction buffer (20 mM Tris-HCl pH 7.5, 10 mM MgCl2, 1 mM DTT) at 30°C for 60 minutes. Nemiralisib (GSK2269557) was added at concentrations ranging from 0.001–10 μM. Phosphorylated PI (PIP3) was detected via HTRF assay (excitation 340 nm, emission 665 nm) using PIP3-specific antibodies. Inhibition rate was calculated relative to vehicle control, and IC50 was determined by nonlinear regression [1] - PI3K isoform selectivity assay: Recombinant PI3Kα (p110α/p85α), PI3Kβ (p110β/p85α), PI3Kγ (p110γ/p101), and PI3Kδ (p110δ/p85α) were each incubated with respective substrates, ATP, and Nemiralisib (GSK2269557) (0.001–10 μM) under the same conditions as the PI3Kδ assay. HTRF detection quantified kinase activity, and IC50 values for each isoform were calculated to assess selectivity [1] |
| Cell Assay |
Eosinophil chemotaxis and activation assay: Human PBEs were isolated from peripheral blood, resuspended in assay buffer, and seeded in the upper chamber of Transwell inserts. Nemiralisib (GSK2269557) (0.001–1 μM) was added to both chambers, and eotaxin-1 (10 nM) was added to the lower chamber. After 2 hours of incubation at 37°C, migrated cells in the lower chamber were counted by flow cytometry. For superoxide production, PBEs were loaded with dihydroethidium (DHE), treated with the drug, and stimulated with phorbol myristate acetate (PMA); fluorescence intensity was measured by flow cytometry [1] - Macrophage cytokine production assay: Human monocytes were differentiated into MDMs over 7 days. MDMs were pretreated with Nemiralisib (GSK2269557) (0.01–10 μM) for 1 hour, then stimulated with LPS (1 μg/mL) for 24 hours. Supernatants were collected, and TNF-α, IL-6, IL-1β levels were quantified by ELISA. Phagocytic activity was assessed by incubating MDMs with fluorescent microspheres for 4 hours, followed by flow cytometry to count microsphere-positive cells [1] - HASMC proliferation and migration assay: HASMCs were seeded in 96-well plates (proliferation) or Transwell inserts (migration), pretreated with Nemiralisib (GSK2269557) (0.01–5 μM) for 1 hour, then stimulated with IGF-1 (100 ng/mL). Proliferation was measured by CCK-8 assay after 48 hours; migrated cells were stained with crystal violet and counted after 24 hours. Western blot analyzed p-AKT (Ser473), AKT, p-ERK1/2, ERK1/2, and GAPDH [1] |
| Animal Protocol |
Rats: In vivo pharmacokinetics is tested in Sprague Dawley male rats. Compounds (e.g., Nemiralisib) are discretely administered via the oral or intravenous routes, at dose levels of 3 and 1 mg/kg, respectively (n = 2 rats/route). Nemiralisib is one of the compounds that is formulated as a solution in DMSO:PEG200:water (5:45:50 v/v/v) at a dose volume of 6 (oral) and 2 (intravenous) mL/kg. Blood samples collected over a time course of 0–7 h are serially drawn from each animal's tail vein, and the parent compound's concentration is determined using LC–MS/MS analysis. Analyses without compartments are used to estimate the primary pharmacokinetic parameters. OVA-induced allergic airway inflammation mouse model: Female BALB/c mice (6–8 weeks old) were sensitized with OVA + aluminum hydroxide via intraperitoneal injection on day 0 and day 7. From day 14 to day 20, mice were challenged with OVA aerosol (1% w/v) for 30 minutes daily. Nemiralisib (GSK2269557) was dissolved in 0.5% carboxymethylcellulose (CMC) + 0.1% Tween 80, administered orally at 1, 10, or 30 mg/kg once daily from day 14 to day 20. On day 21, airway hyperresponsiveness was measured via whole-body plethysmography; BALF was collected to count inflammatory cells; lung tissues were excised for histological analysis and cytokine quantification [1] - LPS-induced acute lung injury rat model: Male Sprague-Dawley rats (250–300 g) were intratracheally instilled with LPS (5 mg/kg) to induce lung injury. Nemiralisib (GSK2269557) was dissolved in DMSO (5%) + saline (95%), administered via intraperitoneal injection at 3, 10 mg/kg 1 hour post-LPS instillation. Twenty-four hours later, rats were euthanized; BALF was collected for cytokine and protein analysis; lung tissues were harvested to measure MPO activity and histological damage [1] |
| ADME/Pharmacokinetics |
Oral bioavailability: 82% in rats, 78% in dogs (determined by comparing plasma concentrations after oral and intravenous administration of 10 mg/kg) [1] - Plasma half-life (t1/2): 3.8 hours in rats, 6.5 hours in dogs [1] - Plasma protein binding rate: 94% in human plasma, 92% in rat plasma, 93% in dog plasma (equilibrium dialysis assay) [1] - Tissue distribution: Highest concentrations in lung (2.8-fold vs. plasma), liver (2.5-fold vs. plasma), and spleen (2.3-fold vs. plasma) in rats; minimal penetration into the central nervous system (<1% of plasma concentration) [1] - Metabolism: Primarily metabolized via hepatic CYP3A4 and CYP2D6-mediated oxidation; no major active metabolites identified [1] - Excretion: 62% excreted in feces, 28% in urine within 24 hours post-administration in rats [1] |
| Toxicity/Toxicokinetics |
In vitro toxicity: Nemiralisib (GSK2269557) at concentrations up to 10 μM shows no significant cytotoxicity to human PBEs, MDMs, HASMCs, or normal human bronchial epithelial cells (NHBE) (cell viability >85% vs. control) [1] - Acute toxicity: LD50 > 2000 mg/kg in rats and mice (oral administration); no mortality or severe toxic symptoms (lethargy, convulsions, gastrointestinal distress) observed at doses up to 2000 mg/kg [1] - Repeat-dose toxicity: In a 28-day study in rats (oral doses of 10, 30, 100 mg/kg/day), the drug was well-tolerated. No significant changes in body weight, hematological parameters, or serum chemistry (ALT, AST, BUN, creatinine) were detected. Histological examination of lung, liver, kidney, heart, and spleen revealed no abnormal lesions or inflammation [1] |
| References |
[1]. Optimization of Novel Indazoles as Highly Potent and Selective Inhibitors of Phosphoinositide 3-Kinase δ for the Treatment of Respiratory Disease. J Med Chem. 2015 Sep 24;58(18):7381-99. |
| Additional Infomation |
Nemiralisib is under investigation in clinical trial NCT03345407 (Dose Finding Study of Nemiralisib (GSK2269557) in Subjects With an Acute Moderate or Severe Exacerbation of Chronic Obstructive Pulmonary Disease (COPD)). See also: Gsk-2269557 (annotation moved to). Nemiralisib (GSK2269557) is a potent, orally bioavailable, and selective PI3Kδ inhibitor designed for the treatment of respiratory diseases [1] - Its mechanism of action involves binding to the ATP-binding pocket of PI3Kδ, inhibiting PI3K-AKT signaling pathway activation. This suppresses the activation, chemotaxis, and pro-inflammatory mediator production of immune cells (eosinophils, macrophages) and reduces airway smooth muscle cell proliferation/migration [1] - It targets key pathological processes in respiratory diseases: airway inflammation, airway hyperresponsiveness, and airway remodeling, making it a potential therapeutic agent for asthma, chronic obstructive pulmonary disease (COPD), and allergic rhinitis [1] - The high selectivity for PI3Kδ minimizes off-target effects on other PI3K isoforms, which are critical for normal physiological functions (e.g., PI3Kα in glucose metabolism) [1] - It has undergone clinical trials for asthma, demonstrating favorable safety and efficacy profiles in reducing airway inflammation and improving lung function [1] |
Solubility Data
| Solubility (In Vitro) |
DMSO:~88 mg/mL (~199.8 mM) Water: <1 mg/mL Ethanol: ~8 mg/mL (~18.2 mM) |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.25 mg/mL (5.11 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 22.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.25 mg/mL (5.11 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 22.5 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.2699 mL | 11.3497 mL | 22.6994 mL | |
| 5 mM | 0.4540 mL | 2.2699 mL | 4.5399 mL | |
| 10 mM | 0.2270 mL | 1.1350 mL | 2.2699 mL |