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| 5mg |
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| 25mg |
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| 50mg |
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| 100mg |
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Purity: ≥98%
| Targets |
The primary target of 2-hydroxyflutamide is the androgen receptor (AR). It acts as a competitive antagonist, binding to the AR and preventing the binding of endogenous androgens such as testosterone and 5α-dihydrotestosterone (DHT). This inhibits AR-mediated gene transcription and downstream signaling pathways, thereby suppressing the growth of androgen-dependent cells, such as those found in prostate cancer. 2-Hydroxyflutamide exhibits significantly higher binding affinity for the AR compared to its parent compound flutamide.
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| ln Vitro |
2-Hydroxyflutamide is a potent androgen receptor antagonist. Multiple studies have reported an IC₅₀ value of approximately 700 nM for the androgen receptor, which is approximately 4-fold less potent than bicalutamide. However, an IC₅₀ of 15 nM has also been reported in certain cellular systems. In human AR binding assays, a Ki value of approximately 430 nM was determined via displacement of [³H]testosterone. In androgen-sensitive prostate cancer cell lines such as LNCaP, 2-hydroxyflutamide downregulates the expression of AR target genes and prevents androgen-dependent stabilization of the AR protein. It also modulates the phosphorylation of cadherin/catenin complexes and the AR via PI3K/Akt and MAPK/ERK1/2 signaling pathways.
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| ln Vivo |
2-Hydroxyflutamide is a more potent antiandrogen in vivo than flutamide, exhibiting higher binding affinity for the androgen receptor. In preclinical efficacy studies, 2-hydroxyflutamide effectively suppresses tumor growth in prostate cancer xenograft models by inhibiting AR signaling. In clinical studies, intraprostatic injection of 2-hydroxyflutamide formulated as Liproca Depot (NanoZolid sustained-release delivery system) demonstrated promising antitumor effects in patients with localized prostate cancer. At 5 months post-injection, serum PSA decreased by an average of 14%, 95% of patients experienced PSA reduction, 78% showed a decrease in prostate volume from baseline, and MRI and biopsy confirmed stabilization or reduction of lesion size. The treatment was safe and well tolerated, with no hormonal side effects observed.
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| Enzyme Assay |
A typical cell-free AR binding assay protocol is as follows: Human androgen receptor protein (recombinant or tissue-derived) is incubated with a radiolabeled ligand such as [³H]-testosterone or [³H]-DHT. Various concentrations of 2-hydroxyflutamide (typically 1 nM–100 μM, serial dilutions) are added, followed by incubation at 4°C or room temperature for approximately 2–4 hours. After incubation, bound and free radioligands are separated by rapid filtration (e.g., using glass fiber filters), followed by washing to remove unbound radioactivity. After drying, the bound radioactivity on the filters is quantified using a scintillation counter. The percentage of inhibition at each concentration is calculated, a competition binding curve is generated, and IC₅₀ and Ki values are determined by nonlinear regression analysis. This protocol can be referenced from published methods (e.g., Bioorg Med Chem 2008, 16: 6799–6812).
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| Cell Assay |
Using LNCaP cells (androgen-sensitive prostate cancer cells) as an example, the protocol is as follows: Cells are cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum at 37°C in 5% CO₂. Prior to the assay, cells are transferred to charcoal-stripped serum-containing medium (to remove endogenous androgens) for 24–48 hours. Cells are seeded in 96-well plates (approximately 1×10⁴ cells/well) and allowed to attach overnight. Various concentrations of 2-hydroxyflutamide (0.1–100 μM) and control compounds (e.g., DHT as an agonist) are added, and incubation continues for 24–72 hours. Cell viability is assessed using CCK-8, MTT, or CellTiter-Glo assays. AR transcriptional activity can be measured using a luciferase reporter gene driven by an androgen response element (ARE). Western blot can be used to detect changes in AR target protein expression (e.g., PSA), and flow cytometry can be employed to analyze cell cycle and apoptosis.
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| Animal Protocol |
The prostate cancer xenograft model is commonly used to evaluate the in vivo antitumor activity of 2-hydroxyflutamide. Male immunodeficient mice (e.g., BALB/c nude mice or SCID mice) are subcutaneously inoculated with LNCaP or other AR-positive prostate cancer cells (approximately 5×10⁶ cells/mouse). When tumors reach approximately 100–200 mm³, animals are randomized into treatment and control groups. The treatment group receives 2-hydroxyflutamide by oral gavage, intraperitoneal injection, or subcutaneous injection (typically 10–100 mg/kg/day in an appropriate vehicle such as DMSO/PEG or corn oil), while the control group receives vehicle alone. Treatment lasts 2–6 weeks, with tumor volume (length × width²/2) and body weight measured 2–3 times weekly. At the end of the study, blood samples are collected for PSA measurement and pharmacokinetic analysis, and tumor tissues are harvested for histopathological analysis, immunohistochemistry for AR downstream markers (e.g., Ki-67, PSA), and TUNEL apoptosis assays.
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| ADME/Pharmacokinetics |
Metabolism / Metabolites
2-Hydroxyflutamide is a known metabolite of flutamide in the human body. Following oral administration, flutamide is rapidly absorbed from the gastrointestinal tract with a tmax of approximately 2 hours. Flutamide undergoes extensive first-pass metabolism, and its primary active metabolite is 2-hydroxyflutamide. After oral administration of 250 mg flutamide, the mean steady-state plasma concentration of 2-hydroxyflutamide is approximately 0.94 μg/mL (steady state achieved within 2–4 days). 2-Hydroxyflutamide is eliminated monoexponentially, with elimination half-lives ranging from 4.3 to 21.9 hours (substantial interindividual variability). In humans, the elimination half-life of 2-hydroxyflutamide is approximately 8 hours. In rats, the AUC of 2-hydroxyflutamide is 219±22 mg·L⁻¹·h, Cmax is 8.6±0.6 mg·L⁻¹, and the elimination rate constant K(m) is 0.07±0.01 h⁻¹; elimination is significantly inhibited in hepatic injury rats. |
| Toxicity/Toxicokinetics |
2-Hydroxyflutamide exhibits significant hepatocytotoxicity and mitochondrial toxicity. Studies have shown that 2-hydroxyflutamide significantly reduces both basal and maximal respiration in HepG2 cells. It inhibits respiratory complex I (similar to the parent drug flutamide) and additionally inhibits complexes II and V—mitochondrial liabilities not demonstrated by the parent compound. These additional mitochondrial liabilities may fundamentally contribute to the idiosyncratic drug-induced liver injury (DILI) observed clinically with flutamide. In primary rat hepatocytes, 50 mg/L of 2-hydroxyflutamide for 8 hours increases ALT and AST activities, decreases GSH content, and induces CYP1A2 mRNA expression (approximately 3.5-fold), though its cytotoxic effects are weaker than those of flutamide. According to safety data sheets, 2-hydroxyflutamide causes skin and eye irritation, is harmful if swallowed, may cause damage to fertility or the unborn child, and is toxic to aquatic life with long-lasting effects.
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| References |
Ball AL, Kamalian L, Alfirevic A, Lyon JJ, Chadwick AE. Identification of the Additional Mitochondrial Liabilities of 2-Hydroxyflutamide When Compared With its Parent Compound, Flutamide in HepG2 Cells. Toxicol Sci. 2016 Oct;153(2):341-51. doi: 10.1093/toxsci/kfw126.
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| Additional Infomation |
2-Hydroxyflutamide is a metabolite of flutamide. Flutamide is an oral nonsteroidal anti-androgen drug primarily used to treat prostate cancer. It competes with testosterone and its potent metabolite dihydrotestosterone (DHT) for binding to androgen receptors in the prostate. In this way, it can prevent androgen receptors from stimulating the growth of prostate cancer cells. Due to fewer side effects, flutamide has been superseded by bicalutamide, a newer member of this class of drugs. (Wikipedia)
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| Molecular Formula |
C11H11F3N2O4
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|---|---|
| Molecular Weight |
292.21
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| Exact Mass |
292.067
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| CAS # |
52806-53-8
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| Related CAS # |
52806-53-8
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| PubChem CID |
91649
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| Appearance |
Light yellow to yellow solid powder
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| Density |
1.5±0.1 g/cm3
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| Boiling Point |
443.8±45.0 °C at 760 mmHg
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| Melting Point |
125-130℃
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| Flash Point |
222.2±28.7 °C
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| Vapour Pressure |
0.0±1.1 mmHg at 25°C
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| Index of Refraction |
1.546
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| LogP |
3.16
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
7
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| Rotatable Bond Count |
2
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| Heavy Atom Count |
20
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| Complexity |
392
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
YPQLFJODEKMJEF-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C11H11F3N2O4/c1-10(2,18)9(17)15-6-3-4-8(16(19)20)7(5-6)11(12,13)14/h3-5,18H,1-2H3,(H,15,17)
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| Chemical Name |
2-hydroxy-2-methyl-N-(4-nitro-3-(trifluoromethyl)phenyl)propanamide
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| Synonyms |
2-HOF 2HOF 2 HOF
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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 |
| 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 : ~100 mg/mL (~342.22 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (8.56 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 (8.56 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. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (8.56 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.4222 mL | 17.1110 mL | 34.2220 mL | |
| 5 mM | 0.6844 mL | 3.4222 mL | 6.8444 mL | |
| 10 mM | 0.3422 mL | 1.7111 mL | 3.4222 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.
| NCT Number | Recruitment | interventions | Conditions | Sponsor/Collaborators | Start Date | Phases |
| NCT00913263 | COMPLETEDWITH RESULTS | Drug: 2-hydroxyflutamide (2-HOF) [Liproca Depot] Drug: 2-Hydroxyflutamide |
Prostate Cancer | Lidds AB | 2009-06 | Phase 1 Phase 2 |
| NCT02341404 | COMPLETED | Drug: 2-hydroxyflutamide (2-HOF) | Prostate Cancer | Lidds AB | 2012-05 | Phase 2 |
| NCT03348527 | COMPLETED | Drug: 2-Hydroxyflutamide Depot | Prostate Cancer | Lidds AB | 2017-05-12 | Phase 2 |
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