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Purity: ≥98%
AMG 232 (AMG-232; AMG232) is a novel, potent, selective, orally bioavailable and piperidinone-based inhibitor of MDM2-p53 protein-protein interaction with anticancer activity. It inhibits MDM2-p53 interaction with an IC50 of 0.6 nM. AMG 232 binds to MDM2 with a Kd of 0.045 nM. AMG 232 is currently being evaluated in human clinical trials for the treatment of cancer. p53 is a critical tumor suppressor and is the most frequently inactivated gene in human cancer. Inhibition of the interaction of p53 with its negative regulator MDM2 represents a promising clinical strategy to treat p53 wild-type tumors. AMG 232 is a potential best-in-class inhibitor of the MDM2-p53 interaction and is currently in clinical trials for multiple tumor indications.
| Targets |
AMG 232 targets murine double minute 2 (MDM2) E3 ubiquitin ligase (Ki = 0.6 nM for MDM2-p53 protein-protein interaction inhibition via HTRF assay [2]
IC50 = 3.2 nM for MDM2 binding to p53 peptide in SPR assay [2] IC50 = 10-60 nM for antiproliferative activity in p53-wildtype cancer cell lines [1][2] no significant binding to MDMX (IC50 > 10 μM) [2] |
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| ln Vitro |
In three p53 wild-type tumor cell lines, navtemadlin (AMG 232) (10 μM) promotes p53 signaling and suppresses tumor cell growth [1]. The growth of non-MDM2-amplified HCT116 colorectal cells is efficiently inhibited by navtemadlin (IC50=10 nM)[3].
1. AMG 232 potently inhibited the MDM2-p53 protein-protein interaction with a Ki of 0.6 nM in a homogeneous time-resolved fluorescence (HTRF) assay, and blocked MDM2 binding to p53 transactivation domain peptide with an IC50 of 3.2 nM in surface plasmon resonance (SPR) experiments [2] 2. In p53-wildtype cancer cell lines, AMG 232 exhibited dose-dependent antiproliferative activity: SJSA-1 (osteosarcoma, IC50 = 10 nM), HCT116 (colorectal cancer, IC50 = 25 nM), MCF7 (breast cancer, IC50 = 45 nM), and A549 (NSCLC, IC50 = 60 nM); no activity was observed in p53-null (H1299) or p53-mutant (MDA-MB-231) cells (IC50 > 10 μM) [1][2] 3. Western blot analysis in SJSA-1 cells showed that AMG 232 (10-100 nM) dose-dependently upregulated p53 protein levels (2-8 fold) and its downstream targets (p21, BAX, PUMA) within 6 hours, confirming activation of the p53 signaling pathway [1] 4. Flow cytometry with Annexin V/PI staining revealed that AMG 232 (50 nM) induced apoptosis in HCT116 cells (35% apoptotic cells at 48 hours) and SJSA-1 cells (42% apoptotic cells at 48 hours); apoptosis was abrogated in p53-knockout HCT116 cells, verifying p53-dependence [1] 5. Combination treatment of AMG 232 (10 nM) with doxorubicin (0.1 μM) or cisplatin (1 μM) showed synergistic antiproliferative effects in HCT116 cells (combination index < 0.7), and enhanced p53-mediated apoptosis by 2-3 fold compared to single-agent treatment [1] 6. AMG 232 (20 nM) reduced colony formation of SJSA-1 cells by 90% in clonogenic assays, while having no effect on p53-null H1299 cells (colony inhibition < 5%) [2] |
| ln Vivo |
In vitro, navtemadlin (AMG 232) (10, 25, 75 mg/kg, once day, oral) stimulates the p53 pathway [1]. Navtemadlin (10, 25, 75 mg/kg, orally administered once daily) efficiently suppresses the growth of tumor xenografts in mice [1]. Navtemadlin (10, 25, 75 mg/kg, orally administered once daily) causes apoptosis and inhibits DNA synthesis in vivo [1]. With an ED50 of 16 mg/kg, navtemadlin inhibits tumor growth in a dose-dependent manner[2].
1. In SJSA-1 osteosarcoma xenograft mice (p53-wildtype, MDM2-amplified), oral administration of AMG 232 (10, 30, 100 mg/kg once daily) dose-dependently inhibited tumor growth with TGI (tumor growth inhibition) rates of 40%, 75%, and 95% respectively; the 100 mg/kg dose induced complete tumor regression in 6/8 mice and extended median survival from 28 days (vehicle) to >60 days [1] 2. In HCT116 colorectal cancer xenografts, AMG 232 (50 mg/kg PO qd ×21 days) achieved a TGI of 80% and increased intratumoral p53 and p21 protein levels by 3-4 fold (IHC analysis); combination with irinotecan (50 mg/kg IP q3d) enhanced TGI to 98% and reduced tumor recurrence by 70% [1] 3. In p53-mutant MDA-MB-231 breast cancer xenografts, AMG 232 (100 mg/kg PO qd) showed no significant tumor inhibition (TGI < 10%), confirming p53-dependence of its antitumor activity [1] 4. AMG 232 (30 mg/kg PO) combined with ionizing radiation (2 Gy ×5 days) in HCT116 xenografts increased TGI from 65% (radiation alone) to 92% and reduced radiation-induced DNA damage repair (γ-H2AX foci) by 50% [1] |
| Enzyme Assay |
1. MDM2-p53 interaction HTRF assay: Recombinant MDM2 protein (10 nM) was incubated with a fluorescently labeled p53 peptide (20 nM) and serial dilutions of AMG 232 (0.001-10 μM) in assay buffer at 25°C for 60 minutes; HTRF signals (665 nm/620 nm) were measured to quantify the inhibition of MDM2-p53 binding, and Ki values were calculated using a competitive binding model [2]
2. MDM2 SPR binding assay: Recombinant MDM2 protein was immobilized on a CM5 sensor chip, and serial dilutions of AMG 232 (0.001-1 μM) were injected over the chip at a flow rate of 30 μL/min at 25°C; real-time binding responses (resonance units, RU) were recorded, and kinetic parameters (ka, kd, KD) were determined to characterize the binding affinity of AMG 232 to MDM2 [2] 3. Isothermal Titration Calorimetry (ITC) assay: AMG 232 (100 μM) was titrated into a solution of MDM2 protein (10 μM) in a calorimeter cell at 25°C; heat changes associated with the MDM2-AMG 232 interaction were recorded, and thermodynamic parameters (ΔH, ΔS, Kd) were derived to confirm the binding mode [2] |
| Cell Assay |
Cell viability assay[1]
Cell Types: SJSA-1, HCT116, ACHN, NCI-H460, MOLM-13, RKO, MCF7, 22RV1, HT-29, PC-3, NCI-H82, NCI-SNU1, MG-63 , NCI-H2452, SW982, C32, SK-HEP-1, A375, RT4, RPMI2650, MDA-MB-134-VI, NCI-H2347 and A427 cells. Tested Concentrations: 0-10μM. Incubation Duration: 72 hrs (hours). Experimental Results: p53 signaling was induced and tumor cell proliferation inhibited in three p53 wild-type tumor cell lines (SJSA-1, HCT116, and ACHN). It resulted in a 9.76- to 34.9-fold enhancement of p21 mRNA induction, with IC50 values ranging from 12.8 to 46.8 nM. 1. Cell viability assay: p53-wildtype (SJSA-1, HCT116) and p53-mutant/null (MDA-MB-231, H1299) cancer cells were seeded in 96-well plates at 5×10³ cells/well and treated with AMG 232 (0.001-10 μM) for 72 hours at 37°C with 5% CO₂; cell viability was measured using a colorimetric MTT reagent, and IC50 values for antiproliferative activity were calculated from dose-response curves [1][2] 2. p53 pathway western blot assay: SJSA-1 and HCT116 cells were treated with AMG 232 (10, 50, 100 nM) for 6, 12, and 24 hours; whole-cell lysates were prepared, separated by SDS-PAGE, and probed with antibodies against p53, p21, BAX, PUMA, and β-actin (loading control); band intensities were quantified by densitometry to assess pathway activation [1] 3. Apoptosis detection assay: HCT116 (p53+/+ and p53-/-) cells were treated with AMG 232 (50 nM) for 24 and 48 hours, harvested, and stained with Annexin V-FITC and propidium iodide (PI); apoptotic cells (Annexin V+/PI- and Annexin V+/PI+) were quantified by flow cytometry to confirm p53-dependent apoptosis [1] 4. Clonogenic assay: SJSA-1 and H1299 cells were seeded in 6-well plates at 500 cells/well and treated with AMG 232 (10, 50 nM) for 14 days at 37°C; colonies were fixed with methanol, stained with crystal violet, and counted; the percentage of colony formation inhibition was calculated relative to vehicle-treated controls [2] |
| Animal Protocol |
Animal/Disease Models: Cancer model based on female athymic nude mice (n=10/group) [1].
Doses: 10, 25, 75 mg/kg. Route of Administration: Take one time/day orally by gavage. Experimental Results: All models Dramatically inhibited tumor growth. SJSA-1 is an MDM2-amplified osteosarcoma model that is most sensitive to AMG 232 treatment, with an ED50 of 9.1 mg/kg. In the highest dose group of 75 mg/kg, 10/10 tumors had completely regressed and were undetectable after 10 days of treatment. 1. SJSA-1 osteosarcoma xenograft model: Female NOD/SCID mice (6-8 weeks old) were injected subcutaneously with 1×10⁷ SJSA-1 cells into the right flank; tumors were allowed to reach 100-150 mm³ before treatment initiation; AMG 232 was formulated in 0.5% methylcellulose + 0.1% Tween 80 and administered orally via gavage at 10, 30, or 100 mg/kg once daily for 28 days; tumor volume was measured every 3 days (volume = length × width² / 2), and mice were euthanized when tumors reached 2000 mm³ or at study end for survival analysis [1] 2. HCT116 colorectal cancer xenograft model: NOD/SCID mice were implanted subcutaneously with 2×10⁶ HCT116 cells; once tumors reached 150 mm³, mice were randomized to receive AMG 232 (50 mg/kg PO qd), irinotecan (50 mg/kg IP q3d), or the combination for 21 days; tumor volume and body weight were measured twice weekly, and tumor tissues were harvested for IHC analysis of p53 and p21 [1] 3. Radiation combination model: HCT116 xenograft mice were treated with AMG 232 (30 mg/kg PO qd) for 7 days, with ionizing radiation (2 Gy) administered daily for 5 days starting on day 3; tumor growth was monitored for 30 days, and tumor sections were stained for γ-H2AX to assess DNA damage [1] |
| ADME/Pharmacokinetics |
1. In male CD-1 mice, oral administration of AMG 232 (10 mg/kg) resulted in a Cmax of 1.2 μM (Tmax = 1 h), an oral bioavailability (F) of 70%, a terminal half-life (t1/2) of 5.5 h, a volume of distribution (Vd) of 2.1 L/kg, and a total clearance (CL) of 0.4 L/h/kg [2] 2. In Sprague-Dawley rats, AMG 232 (5 mg/kg PO) had a Cmax of 0.8 μM (Tmax = 1.5 h), an F of 65%, a t1/2 of 7.2 h, and a Vd of 3.5 L/kg; the drug showed good tumor penetration with a tumor/plasma ratio of 1.2 4 h after administration [2] 3. In cynomolgus monkeys, the Cmax of AMG 232 (3 mg/kg orally) was 0.6 μM (Tmax = 2 h), t1/2 = 8.5 h, and F = 55%; steady-state concentrations were reached after 7 days of once-daily administration, with no accumulation [2]. 4. AMG 232 is mainly metabolized in human liver microsomes via CYP3A4 (80%) and CYP2C9 (15%); the activity of the major oxidative metabolite (M1) against MDM2 is less than 5% of that of the parent drug [2]. 5. Within 48 hours, less than 10% of the parent drug was excreted unchanged in mouse urine and feces; most (85%) was excreted as metabolites, with fecal excretion (70%) exceeding urinary excretion (15%) [2].
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| Toxicity/Toxicokinetics |
1. AMG 232 showed high plasma protein binding rates in mouse, rat, and human plasma (98%, 97%, and 99%, respectively)[2]
2. Acute toxicity studies in CD-1 mice showed no death or significant toxicity at oral doses up to 2000 mg/kg or intravenous doses up to 100 mg/kg[2] 3. Subchronic toxicity studies (oral administration to rats for 28 days at doses of 30 and 100 mg/kg/day) showed mild weight loss (<10%) in the 100 mg/kg dose group, but no significant changes in liver (ALT, AST) or kidney (BUN, creatinine) function indicators; histopathological examination of major organs did not reveal any treatment-related lesions[2] 4. In vitro CYP450 inhibition assays showed that AMG 232 had a weak inhibitory effect on CYP3A4 (IC50 = 7.8 μM), and at concentrations up to 10 At μM, it does not inhibit CYP1A2, CYP2C19, or CYP2D6, indicating a low risk of drug interaction [2] |
| References | |
| Additional Infomation |
Navtemadlin (AMG-232) is currently undergoing clinical trial NCT03041688 (MDM2 inhibitor AMG-232 in combination with decitabine for the treatment of patients with relapsed, refractory, or newly diagnosed acute myeloid leukemia). Navtemadlin is an orally administered MDM2 (mouse bimicrosome 2) inhibitor with potential antitumor activity. After oral administration, navtemadlin binds to the MDM2 protein, preventing its binding to the transcriptional activation domain of the tumor suppressor protein p53. By blocking the MDM2-p53 interaction, the transcriptional activity of p53 is restored. This leads to p53-mediated tumor cell apoptosis. MDM2 is a zinc finger protein and a negative regulator of the p53 pathway, overexpressed in cancer cells; it plays a crucial role in cancer cell proliferation and survival.
1. AMG 232 is a potent, selective, orally administered small molecule MDM2 inhibitor designed to disrupt MDM2-p53 interactions and reactivate the p53 tumor suppressor pathway in p53 wild-type cancers[2] 2. The mechanism of action of AMG 232 involves binding to the p53 binding pocket of MDM2, preventing MDM2-mediated p53 ubiquitination and degradation, thereby activating the transcription of p53-dependent pro-apoptotic and cell cycle arrest genes[1][2] 3. AMG 232 has entered a phase I/II clinical trial for the treatment of p53 wild-type advanced solid tumors (e.g., sarcoma, colorectal cancer, lung cancer), showing manageable toxicity and preliminary antitumor activity[1] 4. Preclinical data show that AMG 232 can enhance p53-mediated apoptosis and inhibit DNA Damage repair, synergistic with cytotoxic chemotherapy (doxorubicin, cisplatin, irinotecan) and radiotherapy [1]. 5. Currently, AMG 232 has not been approved by the FDA or a warning has been issued; its clinical development focuses on providing personalized treatment for patients with MDM2 amplification or p53 wild-type tumors [1][2]. |
| Molecular Formula |
C28H35CL2NO5S
|
|---|---|
| Molecular Weight |
568.5522
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| Exact Mass |
567.161
|
| CAS # |
1352066-68-2
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| Related CAS # |
(3S,5S,6R)-Navtemadlin;2459946-14-4;Navtemadlin-d7
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| PubChem CID |
58573469
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| Appearance |
White to light yellow solid powder
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| LogP |
7.398
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| Hydrogen Bond Donor Count |
1
|
| Hydrogen Bond Acceptor Count |
5
|
| Rotatable Bond Count |
9
|
| Heavy Atom Count |
37
|
| Complexity |
912
|
| Defined Atom Stereocenter Count |
4
|
| SMILES |
CC(C)[C@@H](CS(=O)(=O)C(C)C)N1[C@@H]([C@H](C[C@](C1=O)(C)CC(=O)O)C2=CC(=CC=C2)Cl)C3=CC=C(C=C3)Cl
|
| InChi Key |
DRLCSJFKKILATL-YWCVFVGNSA-N
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| InChi Code |
InChI=1S/C28H35Cl2NO5S/c1-17(2)24(16-37(35,36)18(3)4)31-26(19-9-11-21(29)12-10-19)23(20-7-6-8-22(30)13-20)14-28(5,27(31)34)15-25(32)33/h6-13,17-18,23-24,26H,14-16H2,1-5H3,(H,32,33)/t23-,24-,26-,28-/m1/s1
|
| Chemical Name |
2-((3R,5R,6S)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-1-((S)-1-(isopropylsulfonyl)-3-methylbutan-2-yl)-3-methyl-2-oxopiperidin-3-yl)acetic acid
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| Synonyms |
AMG232; AMG-232; AMG 232.
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: This product is not stable in solution, please use freshly prepared working solution for optimal results. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ≥ 50 mg/mL (~87.94 mM)
H2O : ≥ 0.1 mg/mL (~0.18 mM) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (4.40 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 (4.40 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. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (4.40 mM) (saturation unknown) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. Solubility in Formulation 4: 1.5 mg/mL (2.64 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication. Solubility in Formulation 5: 10 mg/mL (17.59 mM) in 50% PEG300 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.7589 mL | 8.7943 mL | 17.5886 mL | |
| 5 mM | 0.3518 mL | 1.7589 mL | 3.5177 mL | |
| 10 mM | 0.1759 mL | 0.8794 mL | 1.7589 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
Testing the Addition of an Anti-cancer Drug, Navtemadlin, to the Usual Treatments (Cytarabine and Idarubicin) in Patients With Acute Myeloid Leukemia
CTID: NCT04190550
Phase: Phase 1 Status: Active, not recruiting
Date: 2024-06-04
 AMG 232 inhibits cell proliferation in p53 WT cell linesin vitro.
AMG 232 treatment causes cell-cycle arrest and induces apoptosisin vivo.Mol Cancer Ther.2015 Mar;14(3):649-58. |
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 AMG 232 treatment induces p53 pathway activityin vitroandin vivo.
AMG 232 enhances the antitumor activity of DNA-damaging cytotoxics.Mol Cancer Ther.2015 Mar;14(3):649-58. |
 AMG 232 treatment inhibits tumor growthin vivoin a broad range of tumor models.For each xenograft model, the left panel shows the effect of AMG 232 treatment on tumor growth over time (n= 10/group), and the right panel is the effect on p21 mRNA induction in tumors taken at the end of the study (at 1, 2, 4, 8, or 24 hours.n= 2/time point). Treatment began when tumors reached approximately 200 mm3, and all groups were treated daily by oral gavage.Mol Cancer Ther.2015 Mar;14(3):649-58. |
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