Home Apoptosis IAP antagonist - E3 Ligase inhibitor Xevinapant (AT406) For research use only.

Xevinapant (AT406)

Synonyms: ARRY-334543, Debio1143, SM-406

Xevinapant (AT406, ARRY-334543, Debio1143, SM-406) is a potent Smac mimetic and an antagonist of IAP (inhibitor of apoptosis protein via E3 ubiquitin ligase), binding to XIAP-BIR3, cIAP1-BIR3 and cIAP2-BIR3 with Ki of 66.4 nM, 1.9 nM, and 5.1 nM, 50- to 100-fold higher affinities than the Smac AVPI peptide. Phase 1.

Xevinapant (AT406) Chemical Structure

Xevinapant (AT406) Chemical Structure

CAS No. 1071992-99-8

Purity & Quality Control

Xevinapant (AT406) Related Products

Signaling Pathway

IAP Signaling Pathways

IAP Signaling Pathway

Cell Data

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
EVSA-T Cytotoxicity assay 72 hrs Cytotoxicity against human sensitive EVSA-T cells assessed as inhibition of cell growth after 72 hrs by Alamar Blue assay, EC50 = 0.0021 μM. 26218264
MDA-MB-231 Cytotoxicity assay 72 hrs Cytotoxicity against human sensitive MDA-MB-231 cells assessed as inhibition of cell growth after 72 hrs by Alamar Blue assay, EC50 = 0.019 μM. 26218264
HEK293 Function assay 2 hrs Antagonist activity at full-length FLAG-tagged XIAP (unknown origin) transfected in HEK293 cells assessed as inhibition of interaction with caspase 9 after 2 hrs by immunoprecipitation assay, EC50 = 0.034 μM. 26218264
SKOV3 Growth inhibition assay 4 days Growth inhibition of human SKOV3 cells after 4 days by WST8 assay, IC50 = 0.142 μM. 21443232
MDA-MB-231 Growth inhibition assay 4 days Growth inhibition of human MDA-MB-231 cells after 4 days by WST8 assay, IC50 = 0.144 μM. 21443232
MDA-MB-231 Function assay 10 nM Induction of degradation of cIAP1 in human MDA-MB-231 cells at 10 nM by Western blot analysis 21443232
MDA-MB-231 Function assay 100 nM 15 mins Induction of degradation of cIAP1 in human MDA-MB-231 cells at 100 nM after 15 mins by Western blot analysis 21443232
MDA-MB-231 Apoptosis assay 1.5 uM 12 hrs Induction of apoptosis in human MDA-MB-231 cells assessed as activation of caspase-3 activity at 1.5 uM after 12 hrs by Western blot analysis 21443232
MDA-MB-231 Apoptosis assay 1.5 uM 12 hrs Induction of apoptosis in human MDA-MB-231 cells assessed as activation of PARP cleavage at 1.5 uM after 12 hrs by Western blot analysis 21443232
MDA-MB-231 Antitumor assay 30 mg/kg 2 weeks Antitumor activity against human MDA-MB-231 cells xenografted in SCID mouse assessed inhibition of tumor growth at 30 mg/kg, po administered daily for 5 days/week for 2 weeks 21443232
MDA-MB-231 Antitumor assay 100 mg/kg 2 weeks Antitumor activity against human MDA-MB-231 cells xenografted in SCID mouse assessed inhibition of tumor growth at 100 mg/kg, po administered daily for 5 days/week for 2 weeks 21443232
MDA-MB-231 Antitumor assay 100 mg/kg every 3 days Antitumor activity against human MDA-MB-231 cells xenografted in Balb/c SCID mouse assessed as tumor growth inhibition at 100 mg/kg, po qd measured every 3 days 26218264
DAOY qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for DAOY cells 29435139
SJ-GBM2 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SJ-GBM2 cells 29435139
A673 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for A673 cells 29435139
SK-N-MC qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SK-N-MC cells 29435139
BT-37 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for BT-37 cells 29435139
NB-EBc1 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for NB-EBc1 cells 29435139
Saos-2 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Saos-2 cells 29435139
LAN-5 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for LAN-5 cells 29435139
BT-12 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for BT-12 cells 29435139
OHS-50 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for OHS-50 cells 29435139
SK-N-MC qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for SK-N-MC cells 29435139
TC32 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for TC32 cells 29435139
Click to View More Cell Line Experimental Data

Biological Activity

Description Xevinapant (AT406, ARRY-334543, Debio1143, SM-406) is a potent Smac mimetic and an antagonist of IAP (inhibitor of apoptosis protein via E3 ubiquitin ligase), binding to XIAP-BIR3, cIAP1-BIR3 and cIAP2-BIR3 with Ki of 66.4 nM, 1.9 nM, and 5.1 nM, 50- to 100-fold higher affinities than the Smac AVPI peptide. Phase 1.
Targets
cIAP1-BIR3 [1]
(cell-free assay)		 cIAP2-BIR3 [1]
(cell-free assay)		 XIAP-BIR3 [1]
(cell-free assay)
1.9 nM(Ki) 5.1 nM(Ki) 66.4 nM(Ki)
In vitro
In vitro AT-406 is a Smac mimetic and appears to mimic closely the AVPI peptide in both hydrogen bonding and hydrophobic interactions with XIAP, with additional hydrophobic contacts with W323 of XIAP. AT-406 is more sensitive to these IAPs than Smac AVPI peptide with 50-100 fold binding affinities. AT-406 (at 1 μM) completely restores the activity of caspase-9, which is suppressed by 500 nM XIAP BIR3 in a cell-free system. In MDA-MB-231 cell, AT-406 induces rapid cellular cIAP1 degradation and also pulls down the cellular XIAP protein. AT-406 effectively inhibits lots of human cancer cell lines and shows IC50 of 144 and 142 nM in MDA-MB-231 cell and SK-OV-3 ovarian cell, with low toxicity against normal-like human breast epithelial MCF-12F cells and primary human normal prostate epithelial cells. AT-406 induces apoptosis in MDA-MB-231 cell by inducing activation of caspase-3 and cleavage of PARP. [1]
Kinase Assay Fluorescence Polarization Based Assays for XIAP, cIAP1, and cIAP2 BIR3 Proteins
FL-AT-406 (the fluorescently tagged AT-406) is employed to develop a set of new FP assays for determination of the binding affinities of Smac mimetics to XIAP, cIAP-1, and cIAP-2 BIR3 proteins. The Kd value of FL-AT-406 to each IAP protein is determined by titration experiments using a fixed concentration of FL-AT-406 and different concentrations of the protein up to full saturation. Fluorescence polarization values are measured using an Infinite M-1000 plate reader in Microfluor 2 96-well, black, round-bottom plates. To each well, FL-AT-406 (2, 1, and 1 nM for experiments with XIAP BIR3, cIAP-1 BIR3, and cIAP-2 BIR3, respectively) and different concentrations of the protein are added to a final volume of 125 μL in the assay buffer (100 mM potassium phosphate, pH 7.5, 100 μg/mL bovine γ-globulin, 0.02% sodium azide, with 4% DMSO). Plates are mixed and incubated at room temperature for 2-3 hours with gentle shaking. The polarization values in millipolarization units (mP) are measured at an excitation wavelength of 485 nm and an emission wavelength of 530 nm. Equilibrium dissociation constants (Kd) are then calculated by fitting the sigmoidal dose-dependent FP increases as a function of protein concentrations using Graphpad Prism 5.0 software. In competitive binding experiments for XIAP3 BIR3, AT-406 is incubated with 20 nM XIAP BIR3 protein and 2 nM FL-AT-406 in the assay buffer (100 mM potassium phosphate, pH 7.5; 100 μg/mL bovine γ-globulin; 0.02% sodium azide). In competitive binding experiments for cIAP1 BIR3 protein, 3 nM protein and 1 nM FL-AT-406 are used. In competitive binding experiments for cIAP2 BIR3, 5 nM protein and 1 nM FL-AT-406 are used. For each competitive binding experiment, polarization values are measured after 2-3 hours of incubation using an Infinite M-1000 plate reader. The IC50 value, the inhibitor concentration at which 50% of the bound tracer is displaced, is determined from the plot using nonlinear least-squares analysis. Curve fitting is performed using the PRISM software. A Ki value for AT-406 is calculated.
Cell Research Cell lines MDA-MB-231 breast cancer and SK-OV-3 ovarian cancer cell lines
Concentrations ~ 1 μM
Incubation Time 4 days
Method

Cells are seeded in 96-well flat bottom cell culture plates at a density of (3-4) × 103 cells/well with AT-406 and incubated for 4 days. The rate of cell growth inhibition after treatment with different concentrations of AT-406 is determined by assaying with (2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium monosodium salt (WST-8). WST-8 is added to each well to a final concentration of 10%, and then the plates are incubated at 37 °C for 2−3 hours. The absorbance of the samples is measured at 450 nm using a TECAN ULTRA reader. Concentration of AT-406 that inhibited cell growth by 50% (IC50) is calculated by comparing absorbance in the untreated cells and the cells treated with AT-406.
Experimental Result Images Methods Biomarkers Images PMID
Western blot cIAP-1 / XIAP / Mcl-1 / pS6K1 / Cleaved PARP 28036295
Growth inhibition assay Cell viability 28036295
In Vivo
In vivo AT-406 has good pharmacokinetic (PK) properties and oral bioavailability in mice, rats, non-human primates, and dogs. In the MDA-MB-231 xenograft, AT-406 effectively induces cIAP1 degradation and processing of procaspase-8, cleavage of PARP in tumor tissues at 100 mg/kg with well toleration even at 200 mg/kg. AT-406 induces significant tumor growth inhibition with p of 0.0012 at 100 mg/kg. [1]
Animal Research Animal Models MDA-MB-231 xenograft tumors in severe combined immune deficiency (SCID) mice
Dosages 10 mg/kg (i.v.), 10 mg/kg (p.o.), 30 mg/kg (p.o.) and 100 mg/kg (p.o.)
Administration Administered via intravenously (i.v.) or oral gavage (p.o.)
NCT Number Recruitment Conditions Sponsor/Collaborators Start Date Phases
NCT01265199 Terminated
Acute Myelogenous Leukemia (AML)
Ascenta Therapeutics|The Leukemia and Lymphoma Society
 February 2011 Phase 1
NCT01078649 Completed
Cancer|Solid Tumors|Lymphoma|Malignancy
Debiopharm International SA
 March 29 2010 Phase 1

Chemical Information & Solubility

Molecular Weight 561.71 Formula

C32H43N5O4
CAS No. 1071992-99-8 SDF Download Xevinapant (AT406) SDF
Smiles CC(C)CC(=O)N1CCC2CCC(N2C(=O)C(C1)NC(=O)C(C)NC)C(=O)NC(C3=CC=CC=C3)C4=CC=CC=C4
Storage (From the date of receipt)

In vitro
Batch:

DMSO : 112 mg/mL ( (199.39 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Ethanol : 112 mg/mL

Water : Insoluble


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