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AZD-3458 (PI3Kγ inhibitor 3) is a novel, potent, orally bioavailable and highly isoform selective PI3Kγ inhibitor with anticancer activity. It has pIC50s of 9.1, 5.1,<4.5, and 6.5 for PI3Kγ, PI3Kα, PI3Kβ, and PI3Kδ, respectively. It has ADME properties suitable for oral administration. Compound 15 was advanced into in vivo studies showing dose-dependent inhibition of LPS-induced airway neutrophilia in rats when administered orally.
Physicochemical Properties
| Molecular Formula | C20H23N3O4S2 |
| Molecular Weight | 433.544322252274 |
| Exact Mass | 433.112 |
| CAS # | 2132961-46-5 |
| PubChem CID | 134611894 |
| Appearance | Off-white to light yellow solid powder |
| LogP | 2 |
| Hydrogen Bond Donor Count | 1 |
| Hydrogen Bond Acceptor Count | 6 |
| Rotatable Bond Count | 5 |
| Heavy Atom Count | 29 |
| Complexity | 779 |
| Defined Atom Stereocenter Count | 1 |
| SMILES | S(C)(C1=CC(C2=C(C)N=C(NC(C)=O)S2)=CC2=C1C(N(C2)[C@@H](C)C1CC1)=O)(=O)=O |
| InChi Key | PAQUFWFUOVDUIO-NSHDSACASA-N |
| InChi Code | InChI=1S/C20H23N3O4S2/c1-10-18(28-20(21-10)22-12(3)24)14-7-15-9-23(11(2)13-5-6-13)19(25)17(15)16(8-14)29(4,26)27/h7-8,11,13H,5-6,9H2,1-4H3,(H,21,22,24)/t11-/m0/s1 |
| Chemical Name | N-[5-[2-[(1S)-1-cyclopropylethyl]-7-methylsulfonyl-1-oxo-3H-isoindol-5-yl]-4-methyl-1,3-thiazol-2-yl]acetamide |
| Synonyms | AZD-3458; PI3Kγ inhibitor 3; AZD3458 |
| 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 |
Phosphoinositide 3-kinase gamma (PI3Kγ) (pIC50 > 9.2 in enzyme assay). [1] Class II PI3K PI3KC2β (pIC50 = 7.5 in enzyme assay). [1] PI3Kδ (66% inhibition at 1 µM in a kinase panel, pIC50 = 6.5 in enzyme assay for close analog). [1] |
| ln Vitro |
Additionally, PI3KC2α, PI3KC2β, PI3KC2γ, and PI3KC3 are inhibited by AZD3458 (compound 15), with corresponding pIC50 values of <5, 7.5, 5.5, and 5.1 [1]. Compound 15 demonstrated potent inhibition of PI3Kγ enzyme activity with a pIC50 > 9.2. It showed excellent selectivity over other Class I PI3K isoforms (α, β, δ), with pIC50 values >100-fold lower or showing no significant inhibition in cell-based assays. [1] In a PI3Kγ target engagement cell assay measuring phosphorylation of AKT, compound 15 was highly active (pIC50 = 8.1). It showed >100-fold selectivity over PI3Kδ activity assessed in a human Jeko-1B cell line. The compound was inactive in cell assays assessing PI3Kα (MDA MB-468 cell line) and PI3Kβ (BT4 cell line) activity. [1] Kinase selectivity profiling against 395 kinases showed compound 15 inhibited only PI3Kγ (95%) and PI3Kδ (66%) at 1 µM among Class I/III kinases. Among Class II PI3Ks, it inhibited PI3KC2β (84%) and PI3KC2γ (71%) at 1 µM, with a determined pIC50 of 7.5 for PI3KC2β. No activity was detected against off-targets DRAK1 and CLK4, which were inhibited by an earlier analog. [1] A wide ligand profile screen (CEREP) against 89 targets showed compound 15 had a Ki below 1 µM only for the P2Y1 receptor. However, follow-up whole-cell P2Y1 agonist/antagonist assays showed no activity up to 100 µM. [1] |
| ln Vivo |
Oral administration of compound 15 significantly inhibited lipopolysaccharide (LPS)-induced neutrophil migration into the airways of rats in a dose-dependent manner. Doses of 0.1, 0.5, and 2.5 mg/kg were tested. [1] |
| Enzyme Assay |
A biochemical enzyme inhibition assay was used to measure inhibitory potency against PI3K isoforms (α, β, γ, δ). The assay format is not described in detail within the provided text. The reported pIC50 values are the mean of ≥3 replicates. [1] Selectivity profiling against Class II and Class III PI3Ks was also performed using comparable biochemical assays. The pIC50 values for these isoforms were determined, with the exact pIC50 for PI3Kγ reported as >9.2 due to assay format limitations. [1] |
| Cell Assay |
A PI3Kγ target engagement cellular assay was performed by measuring the phosphorylation level of AKT (a downstream signaling protein) in a relevant cell line. The detailed protocol is not provided in the main text but is referenced to be in the Supporting Information. [1] Isoform selectivity was further confirmed in mechanism-based cellular assays. PI3Kδ activity was assessed using a human Jeko-1B cell line. PI3Kα and PI3Kβ activities were assessed using human MDA MB-468 and BT4 cell lines, respectively. The details of these cell-based readouts are not specified in the main text. [1] |
| Animal Protocol |
An LPS-induced acute inflammation model in rats was used to assess in vivo efficacy. Female Wistar rats were challenged with LPS. Compound 15 was administered orally (po) 2 hours prior to the LPS challenge. The vehicle group and treated groups (n=8-10) were sacrificed 4 hours after the challenge. Bronchoalveolar lavage fluid was collected, and differential cell counts (specifically neutrophils) were performed using an automated hematology analyzer. [1] |
| ADME/Pharmacokinetics |
Compound 15 showed favorable ADME properties suitable for oral administration. Its kinetic solubility at pH 7.4 was 303 µM. [1] The logD at pH 7.4 was 2.4, indicating reduced lipophilicity compared to earlier analogs. [1] Intrinsic clearance in rat hepatocytes was low (1.7 (µL/min)/10^6 cells). [1] Plasma protein binding (PPB) was low across species: human (12% free), rat (12% free), dog (22% free). [1] In vivo pharmacokinetics in rats showed a clearance (CL) of 6.3 (mL/min)/kg, a half-life (t1/2) of 2.3 hours, a volume of distribution (Vd) of 1.2 L/kg, and an oral bioavailability (F) of 51%. [1] In dogs, in vivo PK parameters were: CL = 3.3 (mL/min)/kg, t1/2 = 3.9 hours, Vd = 1.1 L/kg, and F = 82%. [1] |
| Toxicity/Toxicokinetics |
Compound 15 showed a clear margin against hERG channel inhibition with a pIC50 of 4.5. [1] Cytotoxicity assessed in THP1 cells gave a pIC50 of 4.1. [1] No mitochondrial toxicity was observed. [1] Plasma protein binding data is listed under ADME/Pharmacokinetics. [1] |
| References |
[1]. Discovery of Highly Isoform Selective Orally Bioavailable Phosphoinositide 3-Kinase (PI3K)-γ Inhibitors. J Med Chem. 2018 Jun 28;61(12):5435-5441. |
| Additional Infomation |
The series was discovered from a high-throughput screening (HTS) hit featuring an isoindolinone core. The remarkable PI3Kγ selectivity originates from an N-alkyl tail (methylcyclopropyl) that extends deep into the ATP-binding pocket, interacting uniquely with the kat3-helix region, a feature not seen in other reported PI3Kγ inhibitors. [1] The initial HTS hit was optimized using structure-based design. Introduction of a methylsulfonyl group at the 7-position of the core improved physicochemical properties (solubility, lipophilicity) while maintaining potency and selectivity. [1] Compound 15 was identified as the lead candidate due to its combination of high potency, excellent isoform selectivity, favorable pharmacokinetics, and clean in vitro safety profile. [1] |
Solubility Data
| Solubility (In Vitro) | DMSO : ~250 mg/mL (~576.65 mM) |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (4.80 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 20.8 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: ≥ 2.08 mg/mL (4.80 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 20.8 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. Solubility in Formulation 3: ≥ 2.08 mg/mL (4.80 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.3066 mL | 11.5330 mL | 23.0659 mL | |
| 5 mM | 0.4613 mL | 2.3066 mL | 4.6132 mL | |
| 10 mM | 0.2307 mL | 1.1533 mL | 2.3066 mL |