Molecule

Description

AS-604850 is a selective, ATP-competitive PI3Kγ inhibitor with IC50 of 250 nM.

Targets

IC₅₀

In Vitro

AS-604850 is ATP-competitive PI3Kγ inhibitor with K_i values of 0.18 μM. AS-604850 is isoform selective inhibitor of PI3Kγ with over 30-fold selectivity for PI3Kγ and β, and 18-fold selectivity over PI3Kα. (PI3Kα: IC50 = 4.5 μM, PI3Kγ and β: IC50 > 20μM) AS-604850 is capable of inhibiting C5a-mediated PKB phosphorylation in RAW264 mouse macrophages with an IC50 with 10 μM. AS-604850 blocks MCP-1–mediated chemotaxis in Pik3cg ^+/+ monocytes in a concentration- dependent manner, with an IC50 of 21 mM, but dosn’t affect chemotaxis in Pik3cg^-/- cells, indicating that AS-604850 acts through PI3Kγ. ^[1] AS-604850 diminishes glycochenodeoxycholate (GCDC) induced Akt-phosphorylation and apoptosis in rat hepatocytes. AS-604850 diminishes bile salt-induced apoptosis in HepG2 Ntcp and Huh7-Ntcp cells. ^[2] AS604850 causes a concentration-dependent suppression of chemotactic responses of EoL-1 cells and blood eosinophils to platelet-activating factor (PAF). ^[3]

In Vivo

AS-604850 reduces RANTES-induced peritoneal neutrophil recruitment with a ED50 of 42.4 mg/kg. In the thioglycollate-induced peritonitis model, oral administration of 10 mg/kg AS-604850 results in a 31% reduction of neutrophil recruitment. ^[1]

Clinical Trials

Features

In vitro PI3Kγ Kinase Assay

Human PI3Kγ (100 ng) is incubated at RT with kinase buffer (10 mM MgCl₂, 1 mM β-glycerophosphate, 1 mM DTT, 0.1 mM Na₃VO₄, 0.1% Na Cholate and 15 M ATP/100 nCi γ[³³]ATP, final concentrations) and lipid vesicles containing 18 M PtdIns and 250 M of PtdSer (final concentrations), in the presence of AS-252424 or DMSO. Kinase reaction is stopped by adding 250 g of Neomycin-coated Scintillation Proximity Assay (SPA) beads and preceded.

Cell Lines

HepG2 Ntcp and Huh7-Ntcp cells

Concentrations

2.5 μM

Incubation Time

2 - 4 hours

Methods

Hepatocyte cultures are treated with diluent (DMSO), 25 μM TLC, 250 μM TCDC, 50 μM GCDC, or 50 ng/ml Fas for 2–4 hours HepG2-Ntcp and Huh7-Ntcp cells are treated with DMSO, 20 μM TLC, 75 μM TCDC or GCDC, 200 μM etoposide or 200 ng/ml TNFa plus 28 ng/ml actinomycin D for 2–4 hours. The number of apoptotic cells is determined morphologically using fluorescent staining and expressed as % of cells. Apoptosis is confirmed in human cell lines by determination of caspase-3/-7 activity and in rat hepatocytes by detection of the 17 kDa proteolytic cleavage fragment of caspase-3 by immunoblotting; equal protein loading is monitored by immunoblotting for actin.

Animal Models

RANTES (0.5 mg/kg in 200 ml saline) or thioglycollate (40 ml/kg) are intraperitoneally injected to C3H mice to induce peritonitis mouse models

Formulation

AS-604850 is dissolved in vehicle (0.5% carboxymethylcellulose/0.25% Tween-20) and adjusted the solution to 10 mL/kg of body weight.

Doses

0, 1, 3, 10 or 30 mg/kg for RANTES, 10 mg/kg for thioglycollate

Administration

Oral administration 30 or 15 minutes before injection of RANTES or thioglycollate