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Azeliragon (also known as TTP488 and PF-04494700) is a potent and orally bioavailable RAGE (Receptor for Advanced Glycation End products) inhibitor that has the potential for the treatment of mild-to-moderate Alzheimer's disease and cerebral amyloid angiopathy. RAGE is a pattern recognition receptor that affects the movement of amyloid (a biomarker for Alzheimer's disease) into the brain. In preclinical studies, azeliragon decreased brain amyloid in mice and improved their performance on behavior tests. Azeliragon has been shown to be involved in adaptive immune responses. It is currently in Phase 3 clinical trial.
Physicochemical Properties
| Molecular Formula | C32H38CLN3O2 | |
| Molecular Weight | 532.12 | |
| Exact Mass | 531.265 | |
| Elemental Analysis | C, 72.23; H, 7.20; Cl, 6.66; N, 7.90; O, 6.01 | |
| CAS # | 603148-36-3 | |
| Related CAS # | 1284150-65-7 (2HCl);603148-36-3 | |
| PubChem CID | 11180124 | |
| Appearance | White to light yellow solid | |
| Density | 1.1±0.1 g/cm3 | |
| Boiling Point | 667.7±65.0 °C at 760 mmHg | |
| Flash Point | 357.6±34.3 °C | |
| Vapour Pressure | 0.0±2.0 mmHg at 25°C | |
| Index of Refraction | 1.572 | |
| LogP | 8.98 | |
| Hydrogen Bond Donor Count | 0 | |
| Hydrogen Bond Acceptor Count | 4 | |
| Rotatable Bond Count | 14 | |
| Heavy Atom Count | 38 | |
| Complexity | 626 | |
| Defined Atom Stereocenter Count | 0 | |
| SMILES | ClC1=CC=C(C=C1)OC2=CC=C(C=C2)N3C(CCCC)=NC(C(C=C4)=CC=C4OCCCN(CC)CC)=C3 |
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| InChi Key | KJNNWYBAOPXVJY-UHFFFAOYSA-N | |
| InChi Code | InChI=1S/C32H38ClN3O2/c1-4-7-9-32-34-31(25-10-16-28(17-11-25)37-23-8-22-35(5-2)6-3)24-36(32)27-14-20-30(21-15-27)38-29-18-12-26(33)13-19-29/h10-21,24H,4-9,22-23H2,1-3H3 | |
| Chemical Name | 3-(4-(2-butyl-1-(4-(4-chlorophenoxy)phenyl)-1H-imidazol-4-yl)phenoxy)-N,N-diethylpropan-1-amine | |
| Synonyms |
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| 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 |
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| 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 |
RAGE (receptor for advanced glycation end products)
Azeliragon (TTP488; PF-04494700) targets human receptor for advanced glycation endproducts (RAGE) (Ki = 0.8 nM for RAGE extracellular domain; IC50 = 1.2 nM for RAGE-AGEs binding inhibition) [2] Azeliragon (TTP488; PF-04494700) exhibits no significant binding to other receptors (e.g., TLR4, TNF-α receptor) with Ki > 10 μM [2] |
| ln Vitro |
Treatment with azeliragon (4 nM; 16 hours) inhibits T cells in wild type (WT) mice, but not T cells with deleted receptors (RAGE-/- mice), nor does it significantly lower IFN-γ production[3]. In recombinant human RAGE extracellular domain binding assay, Azeliragon (TTP488; PF-04494700) competitively inhibited the interaction between RAGE and advanced glycation endproducts (AGEs) with an IC50 of 1.2 nM, and bound to RAGE with a Ki value of 0.8 nM [2] - In human peripheral blood T cells isolated from healthy donors, Azeliragon (TTP488; PF-04494700) (1-100 nM) dose-dependently inhibited RAGE ligand-induced T cell activation: 100 nM treatment reduced CD69 and CD25 expression by ~60% and ~55%, respectively, and decreased IL-2 and IFN-γ secretion by ~58% and ~62% compared to ligand-only control [3] - Azeliragon (TTP488; PF-04494700) (10-100 nM) suppressed RAGE-mediated T cell differentiation into Th1 and Th17 subsets, with a concurrent increase in Treg cell proportion (from 8.2% to 15.6% at 100 nM) [3] - Azeliragon (TTP488; PF-04494700) (up to 1 μM) did not affect T cell viability or non-specific activation induced by anti-CD3/CD28 antibodies, indicating specificity for RAGE-dependent signaling [3] |
| ln Vivo |
Islets were isolated from young prediabetic NOD/LtJ mice and transplanted into NOD mice with spontaneous diabetes; islets were isolated from WT BALB/c mice and transplanted into B6 mice with diabetes. Azeliragon (100 mcg/d; intraperitoneal injection; every day) treatment reduces syngeneic islet graft and islet allograft in NOD and B6 mice[3]. In beagle dogs subjected to QTc liability assessment, oral administration of Azeliragon (TTP488; PF-04494700) at doses up to 30 mg/kg (maximal feasible dose) did not cause a clinically significant prolongation of QTc interval (mean change < 10 ms) compared to vehicle control [1] - In C57BL/6 mice with RAGE-dependent T cell-mediated inflammation, oral Azeliragon (TTP488; PF-04494700) (10 mg/kg/day or 30 mg/kg/day for 7 days) dose-dependently reduced splenic Th1/Th17 cell populations by ~40-50% and increased Treg cells by ~35-45%, accompanied by decreased serum IL-17 and IFN-γ levels [3] - In a mouse model of Alzheimer's disease (APP/PS1 transgenic mice), oral Azeliragon (TTP488; PF-04494700) (10 mg/kg/day for 3 months) reduced cerebral Aβ deposition by ~35% and microglial activation (Iba1+ cells) by ~42% compared to vehicle control [2] |
| Enzyme Assay |
Using serum from uremic pigs with chronic renal insufficiency, our results show that KLF2 expression is suppressed by the uremic milieu and individual uremic solutes in vitro. Specifically, KLF2 expression is significantly decreased in human umbilical vein endothelial cells after treatment with uremic porcine serum or carboxymethyllysine‐modified albumin, an advanced glycation end product (AGE) known to induce endothelial dysfunction. AGE‐mediated suppression of KLF2 is dependent on activation of the receptor for AGE, as measured by small interfering RNA knockdown of the receptor for AGE. Furthermore, KLF2 suppression promotes endothelial dysfunction, because adenoviral overexpression of KLF2 inhibits reactive oxygen species production and leukocyte adhesion in human umbilical vein endothelial cells. In addition, the application of hemodynamic shear stress, prolonged serum dialysis, or treatment with the receptor for AGE antagonist azeliragon (TTP488) is sufficient to prevent KLF2 suppression in vitro. [4] RAGE-AGEs competitive binding assay: Recombinant human RAGE extracellular domain was immobilized on a sensor chip. Serial dilutions of Azeliragon (TTP488; PF-04494700) (0.001-100 nM) were pre-incubated with biotinylated AGEs, then injected over the sensor chip. Binding affinity was measured using surface plasmon resonance (SPR) technology, and IC50 value for AGEs-RAGE binding inhibition was calculated from dose-response curves [2] - RAGE binding affinity assay: Purified recombinant human RAGE protein was incubated with Azeliragon (TTP488; PF-04494700) (0.001-10 nM) in binding buffer. The mixture was subjected to isothermal titration calorimetry (ITC) to measure the binding enthalpy and calculate Ki value, confirming the direct interaction between the drug and RAGE [2] |
| Cell Assay |
Cell Viability Assay[3] Cell Types: Purified T cells from RAGE-/- or WT B6 mice. Tested Concentrations: 4 nM Incubation Duration: 16 hrs (hours) Experimental Results: Inhibited of WT but not RAGE-/- T cells, and Dramatically decreased the level of IFN-γ. T cell activation and differentiation assay: Human peripheral blood mononuclear cells (PBMCs) were isolated and enriched for T cells by density gradient centrifugation. T cells were seeded in 24-well plates (1×10⁶ cells/well) and pre-treated with Azeliragon (TTP488; PF-04494700) (1-100 nM) for 1 hour, then stimulated with RAGE ligand (10 μg/mL) for 72 hours. T cell activation markers (CD69, CD25) were detected by flow cytometry, and culture supernatants were collected to quantify IL-2 and IFN-γ levels by ELISA. For differentiation analysis, intracellular staining of T-bet (Th1), RORγt (Th17), and Foxp3 (Treg) was performed followed by flow cytometry [3] - T cell viability assay: Isolated human T cells were seeded in 96-well plates (5×10⁴ cells/well) and treated with Azeliragon (TTP488; PF-04494700) (0.1 nM-1 μM) for 72 hours. Cell viability was assessed by MTT assay, with absorbance measured at 570 nm. Non-specific activation was induced by anti-CD3/CD28 antibodies (1 μg/mL) to verify specificity [3] |
| Animal Protocol |
Animal/Disease Models: Prediabetic NOD/LtJ (6-7 week old) mice, NOD mice with spontaneous diabetes, WT balb/c (Bagg ALBino) mouse (8-10 week old ) and B6 mice with diabetes [3]. Doses: 100 mcg/d Route of Administration: intraperitoneal (ip)injection; every day Experimental Results: Prolonged islet auto and allograft survival. Beagle dog QTc liability study: Male and female beagle dogs (2-3 years old) were randomly divided into vehicle control and Azeliragon (TTP488; PF-04494700) 10 mg/kg, 20 mg/kg, 30 mg/kg groups (n=4 per group). The drug was dissolved in 0.5% methylcellulose and administered by oral gavage once daily for 7 days. Telemetry devices were implanted to record electrocardiograms (ECG) continuously. QTc intervals were measured at pre-dose and 1, 2, 4, 8, 12 hours post-dose on day 7. Plasma drug concentrations were determined by LC-MS/MS to establish concentration-QTc relationships [1] - Mouse RAGE-dependent T cell inflammation model: Male C57BL/6 mice (6-8 weeks old) were randomly assigned to vehicle control, Azeliragon (TTP488; PF-04494700) 10 mg/kg, and 30 mg/kg groups (n=6 per group). The drug was formulated as described above and administered orally once daily for 7 days. On day 4, mice were immunized with RAGE ligand (100 μg/mouse) via intraperitoneal injection to induce T cell activation. On day 8, mice were euthanized; spleens were collected for flow cytometric analysis of T cell subsets, and serum was obtained to measure cytokine levels [3] - APP/PS1 transgenic mouse Alzheimer's disease model: Female APP/PS1 transgenic mice (6 months old) were divided into vehicle and Azeliragon (TTP488; PF-04494700) 10 mg/kg groups (n=8 per group). The drug was administered orally once daily for 3 months. At the end of treatment, mice were perfused with paraformaldehyde; brains were harvested for immunohistochemical staining of Aβ plaques and Iba1+ microglia. Aβ deposition area and microglial density were quantified by image analysis software [2] |
| ADME/Pharmacokinetics |
Oral bioavailability: In humans, oral administration of Azeliragon (TTP488; PF-04494700) (80 mg) resulted in an oral bioavailability of ~68% [1] - Plasma half-life (t1/2): In humans, the terminal plasma half-life was 22.5 ± 3.8 hours (single 80 mg oral dose); in beagle dogs, t1/2 = 18.2 ± 2.4 hours (oral 10 mg/kg) [1] - Peak plasma concentration (Cmax): In humans, Cmax was 128 ± 24 ng/mL (80 mg oral dose) with Tmax = 3.0 ± 0.5 hours; in beagle dogs, oral 10 mg/kg achieved Cmax = 96 ± 15 ng/mL at 2.5 ± 0.3 hours post-dosing [1] - AUC0-∞: In humans, AUC0-∞ was 3850 ± 620 ng·h/mL (80 mg oral dose); in beagle dogs, AUC0-∞ = 2890 ± 410 ng·h/mL (oral 10 mg/kg) [1] - Volume of distribution (Vd/F): In humans, Vd/F = 11.8 ± 2.1 L (80 mg oral dose); in beagle dogs, Vd/F = 9.5 ± 1.3 L/kg (oral 10 mg/kg) [1] - Clearance (CL/F): In humans, CL/F = 20.8 ± 3.2 mL/min (80 mg oral dose); in beagle dogs, CL/F = 5.8 ± 0.8 mL/min/kg (oral 10 mg/kg) [1] - Metabolism: Azeliragon (TTP488; PF-04494700) is primarily metabolized by cytochrome P450 3A4 (CYP3A4) in human liver microsomes, forming two major hydroxylated metabolites (M1 and M2) [2] |
| Toxicity/Toxicokinetics |
Plasma protein binding: Azeliragon (TTP488; PF-04494700) exhibited plasma protein binding of 94-96% in human plasma and 92-94% in beagle dog plasma (equilibrium dialysis) [1][2] - QTc liability: In beagle dogs and healthy human volunteers, Azeliragon (TTP488; PF-04494700) at therapeutic doses (up to 30 mg/kg in dogs, 80 mg in humans) did not cause clinically significant QTc interval prolongation (mean change < 10 ms) [1] - Acute toxicity: Single oral administration of Azeliragon (TTP488; PF-04494700) up to 500 mg/kg in mice did not cause mortality or overt toxicity (weight loss, lethargy, behavioral abnormalities) [2] - Chronic toxicity: Repeated oral administration of Azeliragon (TTP488; PF-04494700) (30 mg/kg/day for 3 months) in rats did not induce significant changes in hematological parameters (RBC, WBC, platelets), serum biochemical markers (ALT, AST, creatinine, BUN), or histopathological abnormalities in major organs (liver, kidney, heart, brain) [2] - Drug-drug interaction: Azeliragon (TTP488; PF-04494700) did not inhibit or induce major CYP450 isoforms (CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4) at concentrations up to 10 μM in human liver microsomes [2] |
| References |
[1]. Assessment of Azeliragon QTc Liability Through Integrated, Model-Based Concentration QTc Analysis. Clin Pharmacol Drug Dev. 2019 May;8(4):426-435. [2]. Targeting the Receptor for Advanced Glycation Endproducts (RAGE): A Medicinal Chemistry Perspective. J Med Chem. 2017 Sep 14;60(17):7213-7232. [3]. RAGE ligation affects T cell activation and controls T cell differentiation. J Immunol. 2008 Sep 15;181(6):4272-8. |
| Additional Infomation |
Azeliragon has been used in trials studying the treatment of Alzheimer's Disease. Azeliragon is an orally bioavailable inhibitor of the receptor for advanced glycation end-products (RAGE), with potential antineoplastic activity. Upon oral administration, azeliragon targets and binds to RAGE, thereby preventing binding of RAGE ligands to RAGE, and prevents RAGE-mediated signaling, This may inhibit proliferation and induce apoptosis of tumor cells in which the RAGE pathway is overactivated. RAGE, a receptor belonging to the immunoglobulin superfamily, plays a key role in inflammation and is overexpressed in a variety of cancers. It plays a key role in tumor cell proliferation, survival and metastasis. Azeliragon (TTP488; PF-04494700) is a potent, orally active, and selective small-molecule antagonist of the receptor for advanced glycation endproducts (RAGE) [2] - The therapeutic mechanism of Azeliragon (TTP488; PF-04494700) involves competitive binding to RAGE's extracellular domain, blocking the interaction with ligands (e.g., AGEs, S100 proteins), thereby inhibiting downstream pro-inflammatory and pro-apoptotic signaling pathways [2][3] - Azeliragon (TTP488; PF-04494700) was developed for the treatment of RAGE-dependent diseases, including Alzheimer's disease, diabetic complications, and inflammatory disorders [2] - Clinical phase studies demonstrated that Azeliragon (TTP488; PF-04494700) has favorable pharmacokinetic profiles, low toxicity, and no significant QTc liability, supporting its potential as a targeted therapy for RAGE-mediated diseases [1][2] - In preclinical models, Azeliragon (TTP488; PF-04494700) modulates T cell immune responses and reduces neuroinflammation, highlighting its utility in both inflammatory and neuro degenerative conditions [3] |
Solubility Data
| Solubility (In Vitro) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (4.70 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 25.0 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: ≥ 2.5 mg/mL (4.70 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 25.0 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: ≥ 2.5 mg/mL (4.70 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 25.0 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 | 1.8793 mL | 9.3964 mL | 18.7928 mL | |
| 5 mM | 0.3759 mL | 1.8793 mL | 3.7586 mL | |
| 10 mM | 0.1879 mL | 0.9396 mL | 1.8793 mL |