Tolinapant
Based on 4 publication(s) in Google Scholar
Tolinapant (ASTX660) is an orally active cIAP1/2 and XIAP antagonist, with an IC50 of < 40 nM against XIAP and an IC50 of <12 nM against cIAP1. Tolinapant triggers TNFα-dependent Apoptosis in cancer cells. Tolinapant inhibits tumor growth in mouse xenograft models. Tolinapant can be used in research related to breast cancer, melanoma, lymphoma, multiple myeloma, acute lymphoblastic leukemia and advanced solid tumors.
For research use only. We do not sell to patients.
- Purity : 99.81%
- CAS No.: 1799328-86-1
- Formula: C30H42FN5O3
- Molecular Weight:539.68
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Storage:Powder -20°C, 3 years , 4°C, 2 years ; In solvent -80°C, 2 years , -20°C, 1 year
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
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Biological Activity
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Chemical Information
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Solvent & Solubility
- Protocols
- Purity & Documentation
- References
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Help & FAQs
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Publications Citing Use of MedChemExpress (MCE) Tolinapant
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Biological Activity
Description
IC50 & Target
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cIAP1 <12 nM (IC50) |
cIAP2 |
XIAP ＜40 nM (IC50) |
Cellular Effect
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Cell Line
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Type | Value | Description | References |
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| MDA-MB-231 | IC50 |
1.8 nM
Compound: 27; ASTX660
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Antiproliferative activity against human MDA-MB-231 cells after 72 hrs by Alamar blue assay
Antiproliferative activity against human MDA-MB-231 cells after 72 hrs by Alamar blue assay
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[PMID: 30091600] |
In Vitro
Tolinapant potently inhibits the binding of SMAC peptides to purified XIAP-BIR3 (IC50 <40 nM) and cIAP1-BIR3 (IC50 <12 nM) proteins by occupying the endogenous SMAC-binding pocket[1].
Tolinapant (0.001-10 μM; 5 min-16 h) dihydrochloride antagonizes endogenous XIAP in A375 melanoma cells, and displaces SMAC from XIAP at concentrations above 0.01 μM after 16 h, or within 5 min at a concentration of 1 μM[1].
Tolinapant (72 h) dihydrochloride inhibits the viability of most melanoma cell lines, with enhanced activity in the presence of TNFα, and exerts no significant effect on the viability of normal skin fibroblasts[1].
Tolinapant exhibits single-agent or TNFα-potentiated anti-survival activity in 43% of the tested triple-negative breast cancer (TNBC) cell lines, including MDA-MB-231, HCC38, HCC1806, Hs578T, BT549, HCC1395, DU4475, MDA-MB-453, and the mouse EMT6 cell line[3].
Tolinapant (1-3 μM; 24 h) dihydrochloride significantly enhances the lytic activity of NY-ESO-1 TCR-T cells against A375 melanoma cells, and this effect is dependent on TNF-α[4].
Tolinapant (0.01-1 μM; 48 h) dihydrochloride significantly enhances CD19 CAR-NK cell-mediated lysis of NALM6-Luc2 cells in a TNF-α-dependent manner[4].
Tolinapant (3 μM; for at least 33 days) dihydrochloride enhances the long-term proliferation and persistence of CD19 CAR-T cells, maintains their cytotoxic activity, and preserves the expression of the memory T cell marker CCR7 during repeated antigen stimulation[4].
Tolinapant inhibits the proliferation of various cancer cell lines in a manner dependent on the presence of inflammatory stimulation[5].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only. Further protocols information, click here.
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Cell Line:A375 human melanoma cells
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Concentration:0.001-10 μmol/L (16 h incubation); 1 μmol/L (short-term incubation)
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Incubation Time:16 h (0.001-10 μmol/L); 5-240 min (1 μmol/L)
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Result:Reduced SMAC levels immunoprecipitated with XIAP at concentrations above 0.01 μmol/L after 16 hours of treatment.
Antagonized the interaction of SMAC with XIAP within 5 minutes of exposure to 1 μmol/L.
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Cell Line:human CD19 CAR-T cells, NALM6-Luc2 (B-ALL)
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Concentration:3 μM
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Incubation Time:at least 33 days
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Result:Increased CD19 CAR-T cell growth rate relative to DMSO control, with CAR-T cells persisting for at least 33 days (compared to decline by Day 18 in controls).
Maintained specific lysis of NALM6-Luc2 cells at consistently high 80-90% throughout the study period.
Preserved expression of the memory T cell marker CCR7.
In Vivo
Tolinapant (10-20 mg/kg; p.o.; daily; 14 days) dihydrochloride significantly inhibits A375 melanoma xenograft growth in nude mice[1].
Tolinapant (30-100 mg/kg; p.o.; daily; 7-14 days) dihydrochloride achieves complete on-target engagement via full cIAP1 protein degradation in cynomolgus NHP PBMCs[2].
Tolinapant (16 mg/kg; p.o.; once daily; 21 days) dihydrochloride enhances the in vivo anti-tumor efficacy of low-dose MMG49 CAR-T cells in a disseminated multiple myeloma xenograft model by accelerating CAR-T cell proliferation and persistence, enabling tumor control equivalent to a 10-fold higher dose of CAR-T cells alone[4].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.
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Animal Model:BALB/cJHanHsd-Prkdcscid (male)[1]
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Dosage:5 mg/kg; 10 mg/kg; 20 mg/kg; 20 mg/kg (intermittent schedule)
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Administration:p.o.; daily; 25 days; p.o.; 7 days on/7 days off; 2 cycles
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Result:Significantly inhibited tumor growth (P < 0.05 from days 15, 18, and 11 for 5, 10, and 20 mg/kg daily dosing, respectively).
Achieved significant tumor growth inhibition (P < 0.05 vs. vehicle from day 8) with intermittent 20 mg/kg schedule, equivalent to continuous 20 mg/kg daily schedule.
Distributed to tumors with detectable compound levels up to 168 hours post-dose (AUC_last = 129 μmol/L/h/mL).
Induced rapid cIAP1 degradation, reduced XIAP:SMAC association (persisting up to 3 days), and increased cleaved PARP and cleaved caspase-3 within 1 hour post-dose.
Showed no excessive body weight loss or adverse effects.
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Animal Model:BALB/cOlaHsd-Foxn1nu (male)[1]
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Dosage:10 mg/kg; 20 mg/kg
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Administration:p.o.; daily; 14 days
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Result:Caused significant tumor growth inhibition (P < 0.05 from day 8).
Showed no excessive body weight loss or adverse effects.
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Animal Model:NOD/SCID/IL2Rγcnull (NOG) (6- to 8-week-old female; disseminated multiple myeloma xenograft model)[4]
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Dosage:16 mg/kg
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Administration:p.o.; once daily; 21 days
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Result:Attenuated tumor expansion by day 11 and reduced tumor load thereafter when combined with 1×105 MMG49 CAR-T cells, resulting in lower tumor load than treatment with 1×106 MMG49 CAR-T cells alone by day 21.
Accelerated in vivo MMG49 CAR-T cell expansion, with significantly higher CAR-T cell numbers in the combination group than in the CAR-T monotherapy group between days 10 and 17.
Showed no anti-tumor effect when used alone.
Clinical Trial
| NCT Number | Sponsor | Condition | Start Date |
Phase
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|---|---|---|---|---|
| NCT01329991 | Plexxikon| | 2011-05 | PHASE1 |
Chemical Information
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CAS No. 1799328-86-1
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Appearance Solid
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Molecular Weight 539.68
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Formula C30H42FN5O3
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Color White to off-white
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SMILES
C[C@H]1N(C[C@@H]2N(CC(N3CC(C)(C)C4=NC(CO)=C(CC5=CC=C(F)C=C5)C=C43)=O)C[C@@H](C)NC2)CCOC1
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Synonyms
ASTX660
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Shipping
Room temperature in continental US; may vary elsewhere.
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Storage
Powder -20°C 3 years 4°C 2 years In solvent -80°C 2 years -20°C 1 year
Publications (4)
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Journal Impact Factor
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Most Recent
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Cell Rep Med
Pharmacological Activation of Non-canonical NF-κB Signaling Activates Latent HIV-1 Reservoirs In Vivo. [Abstract]2020 Jun 23;1(3):100037. PMID: 33205060 -
Cancer Gene Ther
b-AP15 enhances TRAIL-induced cell death in HNSCC via the induction of ROS/JNK/DR5 signalling. [Abstract]2026 May 2. PMID: 42069980 -
Blood Res
Translational investigation of tolinapant (ASTX660) in acute myeloid leukemia by integrating clinical, bioinformatic and pharmacological approaches. [Abstract]2025 Dec 30;60(1):67. PMID: 41469824 -
bioRxiv
2024 Nov 28:2024.11.25.625306. PMID: 39651304
Solvent & Solubility
In Vitro:
DMSO : 100 mg/mL (185.29 mM; Need ultrasonic; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.
Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
In Vivo:
Select the appropriate dissolution method based on your experimental animal and administration route.
- For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
- To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for In Vivo experiments, it is recommended to prepare freshly and use it on the same day.
- The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.
Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline
Solubility: ≥ 2.5 mg/mL (4.63 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.
Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
Add each solvent one by one: 10% DMSO 90% (20% SBE-β-CD in Saline)
Solubility: ≥ 2.5 mg/mL (4.63 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C, storage for one week): 2 g SBE-β-CD powder is dissolved in 10 mL Saline, completely dissolve until clear.
In Vivo Dissolution Calculator
Please enter the basic information of animal experiments:
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Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.
Please enter your animal formula composition:
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%DMSO +
Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
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%+
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+%Tween-80 + +
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%Saline +
The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).
Working solution concentration: 0.22 mg/mL
Method for preparing stock solution: mg drug dissolved in μL DMSO. Stock solution concentration: mg/mL.
1. Take μL DMSO stock solution;
2. Add μL .
μL , mix evenly;
3. Then add μL Tween 80, mix evenly;
4. Then add μL
Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.
Protocols
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Apoptosis
Apoptosis, also called programmed cell death, is generally characterized by distinct morphological characteristics.
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TUNEL staining for apoptotic DNA fragmentation
TUNEL staining detects DNA strand breaks by using terminal deoxynucleotidyl transferase to add labeled nucleotides to exposed 3′-OH DNA termini, generating either microscopic staining in fixed cells or tissue sections, or fluorescence/cytometric signal in cell suspensions. TUNEL positivity reflects DNA fragmentation but should not be interpreted alone as definitive apoptosis, because TUNEL can also label necrotic, autolytic, mechanically damaged, or DNA-repair-associated DNA breaks.
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Cell Cytotoxicity Assay
Cytotoxicity assays are usually based on the assessment of cell membrane damage, which can also be indirectly detected by measuring cell viability. Detection methods include MTT assay, CKK-8 assay, LDH assay and ATP assay, etc.
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Annexin V plus membrane-impermeant dye apoptosis staining
Annexin V-based apoptosis assays rely on the detection of phosphatidylserine (PS) externalization from the inner leaflet of the plasma membrane to the outer leaflet, an early biochemical hallmark of apoptosis. Fluorescently labeled Annexin V binds PS in a calcium-dependent manner, enabling identification of early apoptotic cells by flow cytometry or fluorescence microscopy. When combined with a membrane-impermeant DNA-binding dye (e. g. , propidium iodide), this approach allows discrimination between viable (Annexin V−/dye−), early apoptotic (Annexin V+/dye−), and late apoptotic or necrotic (Annexin V+/dye+) cell populations by assessing membrane integrity and PS exposure.
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Patient-Derived Xenograft (PDX)
Patient-derived xenograft (PDX) models are generated by engrafting primary human tumor tissue directly into immunodeficient mice, allowing in vivo propagation of patient tumor biology without initial in vitro adaptation. These models are used to preserve key histopathological and molecular characteristics of the original tumor and enable assessment of tumor growth dynamics and therapeutic response in a living organism. The biological readout is tumor engraftment and subsequent growth in the murine host, which reflects the ability of human tumor cells to survive, vascularize, and expand in an immunocompromised microenvironment.
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Apoptosis Solutions
Apoptosis is a regulated, generally non-lytic cell-death pathway that removes unwanted, damaged, infected, or abnormal cells through coordinated morphological changes, caspase activation, DNA fragmentation, and membrane remodeling. The intrinsic apoptosis pathway is controlled mainly by mitochondrial outer membrane permeabilization, BCL-2 family proteins, cytochrome c release, apoptosome formation, caspase-9 activation, and downstream executioner caspase-3/7 activation. The extrinsic apoptosis pathway is initiated by death receptors such as Fas, TNFR, and TRAIL receptors, which recruit adaptor proteins and activate caspase-8 before engaging executioner caspases or mitochondrial amplification through BID cleavage. Apoptosis is linked to many phenotypes, including cancer cell killing, tissue homeostasis, immune regulation, neurodegeneration, infection response, and treatment-induced cytotoxicity; unresolved questions include how apoptosis interacts with necroptosis, pyroptosis, ferroptos
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Patient-Derived Orthotopic Xenograft (PDOX)
Patient-derived orthotopic xenograft (PDOX) modeling implants fresh patient tumor tissue or patient-derived tumor cells into the anatomically corresponding organ or tissue site of immunodeficient mice, usually by surgical orthotopic implantation, to preserve patient tumor histology, local microenvironmental context, invasion, metastatic behavior, and treatment-response features better than subcutaneous implantation. PDOX readouts include tumor engraftment, orthotopic tumor growth, local invasion, metastasis, recurrence after resection, histologic similarity to the donor tumor, biomarker retention, molecular concordance, survival, and response or resistance to therapy. PDOX models are used for preclinical drug testing and individualized therapy evaluation, but engraftment success varies by tumor type and specimen quality.
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Splenic/Portal-Vein Liver Metastasis Xenograft
Splenic and portal-vein liver metastasis xenograft models deliver tumor cells into the portal circulation so that cells reach the liver first and form hepatic metastatic lesions; splenic injection uses the spleen as an access route to the portal system, while direct portal-vein injection introduces cells into the portal vein without requiring splenectomy. The assay detects liver colonization, intrahepatic tumor growth, tumor distribution, treatment response, survival, and liver-metastasis microenvironment changes; readouts include bioluminescence or fluorescence imaging, gross liver nodule counts, liver weight or tumor burden, histology, and survival.
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Subcutaneous Cell-Line-Derived Xenograft
Subcutaneous cell-line-derived xenograft (CDX) models are established by implanting cultured human cancer cell lines into immunodeficient mice, where the injected cells form localized tumors that can be monitored in vivo as a measure of tumorigenic potential, growth kinetics, and treatment response. These models are widely used in oncology research because they allow reproducible tumor formation and enable comparative assessment of tumor growth between different cell lines or genetic manipulations in a controlled in vivo microenvironment. Subcutaneous implantation of cancer cells in immunodeficient mice is a standard approach for evaluating tumor growth behavior and therapeutic response across multiple cancer types, including prostate, esophageal, pancreatic, and colon cancer models.
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Breast Cancer Modeling
Breast cancer is a heterogeneous cancer, and it has been distinguished into four subtypes: luminal A, luminal B, HER2-positive and basal-like. Molecular mutations, epigenetic alterations, hormone exposure and immune microenvironment are related to the progression of breast cancer.
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Orthotopic Cell-Line Xenograft
Orthotopic cell-line xenograft models involve implantation of human cancer cell lines into the anatomically corresponding organ of immunodeficient mice to reproduce tumor growth within a native microenvironment, enabling more clinically relevant tumor behavior compared with subcutaneous models. These models are widely used because orthotopic placement better recapitulates tumor progression, including invasion and metastatic spread, which are often underrepresented in heterotopic implantation systems. Compared with conventional xenografts, orthotopic implantation is described as more technically complex but provides improved simulation of tumor-microenvironment interactions and metastatic behavior, making it particularly valuable for translational oncology research. Surgical orthotopic implantation approaches have been emphasized as enabling faithful reproduction of clinical cancer features, including metastasis and disease progression patterns that align with the tumor’s organ of origi
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Intraperitoneal/Peritoneal Dissemination Xenograft
Intraperitoneal (IP) or peritoneal dissemination xenograft models are based on the introduction of human cancer cells into the peritoneal cavity of immunodeficient mice, where they attach to peritoneal surfaces, form multicellular aggregates or spheroids, and progressively generate disseminated tumor nodules that mimic advanced peritoneal metastatic disease. These models are widely used to study ovarian cancer progression, tumor-microenvironment interactions, and intraperitoneal therapeutic responses, often incorporating bioluminescence or fluorescence imaging to longitudinally monitor tumor burden in vivo. The biological principle relies on the capacity of tumor cells such as SKOV3 or related ovarian carcinoma lines to survive in suspension, aggregate within ascites-like fluid, adhere to mesothelial surfaces, and invade peritoneal organs, thereby recapitulating human peritoneal carcinomatosis patterns observed in advanced disease.
Purity & Documentation
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Data Sheet (301 KB)
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SDS (396 KB)
- English - EN (396 KB)
- Français - FR (396 KB)
- Deutsch - DE (396 KB)
- Norwegian - NO (396 KB)
- Español - ES (396 KB)
- Swedish - SV (396 KB)
- Italian - IT (396 KB)
- Korean - KR (396 KB)
- Portuguese - PT (396 KB)
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Handling Instructions (2659 KB)
References
[1]. Ward GA, et al. ASTX660, a Novel Non-peptidomimetic Antagonist of cIAP1/2 and XIAP, Potently Induces TNFα-Dependent Apoptosis in Cancer Cell Lines and Inhibits Tumor Growth. Molecular cancer therapeutics. 2018 Jul;17(7):1381-1391. [Content Brief]
[2]. Martins V, et al. The preclinical pharmacokinetics of Tolinapant-A dual cIAP1/XIAP antagonist with in vivo efficacy. Pharmacology research & perspectives. 2024 Dec;12(6):e70030. [Content Brief]
[4]. Tazuru K, et al. The IAP antagonist tolinapant enhances the anti-tumor activity of cell therapies. European journal of pharmacology. 2025 May 15;995:177400. [Content Brief]
[5]. Mita MM, et al. A Phase I Study of ASTX660, an Antagonist of Inhibitors of Apoptosis Proteins, in Adults with Advanced Cancers or Lymphoma. Clinical cancer research : an official journal of the American Association for Cancer Research. 2020 Jun 15;26(12):2819-2826. [Content Brief]
Complete Stock Solution Preparation Table
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.
| Optional Solvent | Concentration Solvent Mass | 1 mg | 5 mg | 10 mg | 25 mg |
|---|---|---|---|---|---|
| DMSO | 1 mM | 1.8529 mL | 9.2647 mL | 18.5295 mL | 46.3237 mL |
| 5 mM | 0.3706 mL | 1.8529 mL | 3.7059 mL | 9.2647 mL | |
| 10 mM | 0.1853 mL | 0.9265 mL | 1.8529 mL | 4.6324 mL | |
| 15 mM | 0.1235 mL | 0.6176 mL | 1.2353 mL | 3.0882 mL | |
| 20 mM | 0.0926 mL | 0.4632 mL | 0.9265 mL | 2.3162 mL | |
| 25 mM | 0.0741 mL | 0.3706 mL | 0.7412 mL | 1.8529 mL | |
| 30 mM | 0.0618 mL | 0.3088 mL | 0.6176 mL | 1.5441 mL | |
| 40 mM | 0.0463 mL | 0.2316 mL | 0.4632 mL | 1.1581 mL | |
| 50 mM | 0.0371 mL | 0.1853 mL | 0.3706 mL | 0.9265 mL | |
| 60 mM | 0.0309 mL | 0.1544 mL | 0.3088 mL | 0.7721 mL | |
| 80 mM | 0.0232 mL | 0.1158 mL | 0.2316 mL | 0.5790 mL | |
| 100 mM | 0.0185 mL | 0.0926 mL | 0.1853 mL | 0.4632 mL |
Tolinapant Related Classifications
HY-109565 Related Classifications
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