Topiroxostat (FYX-051, compound 39) has strong and longer-lasting effects that have been verified by XOR-Topiroxostat complex crystallographic investigation. The binding activity between Topiroxostat and XOR has been observed to be significantly influenced by the cyano group of Topiroxostat. Asn 768 of XOR and the cyano group of Topiroxostat have formed a hydrogen bond, which is responsible for this[1].

In a rat model of potassium oxonate-induced hyperuricemia, topiroxostat (FYX-051; 0.03-10 mg/kg; oral administration; for 1 hour; male Wistar/ST strain rats) treatment demonstrates a strong and long-lasting hypouricemic effect[2]. Topiroxostat (FYX-051, compound 39) has a Cmax of 4.62 μg/mL (3 mg/kg) and a bioavailability of 69.6%, respectively. Additionally, Topiroxostat's t1/2 value is 19.7 hours[1].

Animal/Disease Models: Male Wistar/ST strain rats (7 weeks old) injected with potassium oxonate[2]
Doses: 0.03 mg/kg, 0.1 mg/kg, 0.3 mg/kg, 1 mg/kg, 3 mg/kg, 10 mg/kg
Route of Administration: Oral administration; for 1 hour
Experimental Results: Caused a dose-dependent decrease in serum urate levels with an extremely low ED50 of 0.15 mg/kg, evaluated at 1 h after oral administration.

Absorption, Distribution and Excretion
The time to reach peak plasma concentration of 229.9 ng/mL was 0.67 hour following a single oral dose of 20mg topiroxostat. The oral bioavailability in male rats was 69.6% after oral administration of a single dose of 1mg/kg.
Urinary excretion and fecal excretion of radiolabeled topiroxostat are 30.4% and 40.9% of total dose of 1mg/kg administered to rats, respectively. Within 24 h after a single oral administration of 120mg of topiroxostat, the main metabolites of topiroxostat, N-oxide, N1-gluculonide, and N2-gluculonide, are excreted into urine about 4.8, 43.3, and 16.1 % of the dose, respectively. Unchanged topiroxostat and the hydroxide metabolite was 0.1% or less.
The distribution of 14C-topiroxostat (20, 200, and 2000 ng/mL) in human blood cells was 6.7% to 12.8%.
The apparent total body clearance rate is 89.5 L/h and the renal clearance rate is 17.4 mL/h following a single oral dose of 20mg topiroxostat.

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Metabolism / Metabolites
Topiroxostat is mainly inactivated by hepatic metabolism. 2-hydroxy topiroxostat is formed from primary hydroxylation of the drug by xanthine oxidase and still retains an inhibitory activity on the enzyme. Topiroxostat N-oxide is another major metabolite that can be detected in plasma and urine. It is determined that the N-oxide and hydroxide metabolites are pyridine N-oxide and pyridine 2 (or 6)-hydroxide, respectively. Topiroxostat is mainly inactivated by hepatic metabolism where it undergoes glucuronidation. The metabolism of topiroxostat to N1-and N2-glucuronide conjugates is mainly mediated by UGT1A1, 1A7, and 1A9, with UGT1A9 being the most predominant.
FYX-051 has known human metabolites that include 4-[2-[(3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxyoxan-2-yl]-5-pyridin-4-yl-1,2,4-triazol-3-yl]pyridine-2-carbonitrilium and (2S,3S,4S,5R)-6-[3-(2-cyanopyridin-4-yl)-5-pyridin-4-yl-1,2,4-triazol-1-yl]-3,4,5-trihydroxyoxane-2-carboxylic acid.

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Biological Half-Life
The mean half life of topiroxostat after a single oral dose of 20mg topiroxostat is 5 hours under fasting condition. The complex of molybdenum (IV)- topiroxostat has an approximate half life of 20.4 hours.

Protein Binding
The mean protein binding of radiolabeled (14C)-topiroxostat in human plasma is >97.5% at 20ng/mL, 98.8% at 200ng/mL, and 98.4% at 2000ng/mL. Binding to serum albumin is most predominant with 92.3-93.2%, and mean protein binding to α1-acid protein and γ-globulin is 12.3% to 16.8% and 34.7% to 40.4%, respectively.

[1]. Sato T, et al. Discovery of 3-(2-cyano-4-pyridyl)-5-(4-pyridyl)-1,2,4-triazole, FYX-051 - a xanthine oxidoreductase inhibitor for the treatment of hyperuricemia [corrected]. Bioorg Med Chem Lett. 2009 Nov 1;19(21):6225-9.
[2]. Matsumoto K, et al. FYX-051: a novel and potent hybrid-type inhibitor of xanthine oxidoreductase. J Pharmacol Exp Ther. 2011 Jan;336(1):95-103.

Topiroxostat is a selective xanthine oxidase inhibitor developed for treatment and management of hyperuricemia and gout. Xanthine oxidase, or xanthine oxidoreductase (XOR), regulates purine metabolism, and inhibition of the enzyme results in efficacious reduction of serum urate levels. Xanthine oxidase inhibitors are classified into two groups; purine analogs such as [DB00437] and [DB05262], and non-purine agents which includes topiroxostat. While [DB00437] is considered a first-line therapy in treating hyperuricemic conditions, it is often associated with side effects and ineffective in reducing uric acid levels under recommended dosing regimens. Renal complications are major comorbidities that limit the [DB00437] therapy as dose reductions are recommended. Topiroxostat and its metabolites are shown to be unaffected by renal complications, thus may be effective in patients with chronic kidney diseases. Approved for therapeutic use in Japan since 2013, topiroxostat is marketed under the name Topiloric and Uriadec and is orally administered twice daily.

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Drug Indication
Indicated for the treatment of gout and hyperurcemia in Japan.

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Mechanism of Action
Uric acid synthesis depends on the action of xanthine oxidase activity in the conversion of hypoxanthine to xanthine, followed by the conversion of xanthine to uric acid. Xanthine oxidase consists of a molybdenum ion as cofactor in the active center that has different redox states upon substrate binding. When a substrate such as hypoxanthine or xanthine binds, xanthine oxidase hydroxylates it and molybdenum ion is reduced from hexavalent, Mo(VI), to tetravalent form, Mo(IV). Molybdenum ion is reoxidized into hexavalent state once the hydroxylated substrate, xanthine or uric acid, dissociates from the active site. Topiroxostat is shown to interact with multiple amino acid residues of the solvent channel and additionally forms a reaction intermediate by covalent binding with molybdenum (IV) ion via an oxygen atom. It also forms hydrogen bonds with molybdenum (VI) ion, suggesting that it has multiple inhibition modes to xanthine oxidase. Enhanced binding interactions to xanthine oxidase achieves delayed dissociation of topiroxostat from the enzyme. 2-hydroxy-topiroxostat, the metabolite formed by primary hydroxylation of topiroxostat by xanthine oxidase, also causes time and concentration-dependent inhibition of the enzyme. Topiroxostat is shown to inhibit ATP-binding cassette transporter G2 (ABCG2) in vitro, which is a membrane protein responsible for recovering uric acid in the kidneys and secreting uric acid from the intestines.

C13H8N6

248.24

248.081

577778-58-6

Topiroxostat-d4;2732868-49-2

5288320

Typically exists as solid at room temperature

1.5±0.1 g/cm3

594.7±60.0 °C at 760 mmHg

175.3±18.1 °C

0.0±1.7 mmHg at 25°C

1.697

1.35

1

5

2

19

344

0

N1([H])C(C2C([H])=C([H])N=C([H])C=2[H])=NC(C2C([H])=C([H])N=C(C#N)C=2[H])=N1

UBVZQGOVTLIHLH-UHFFFAOYSA-N

InChI=1S/C13H8N6/c14-8-11-7-10(3-6-16-11)13-17-12(18-19-13)9-1-4-15-5-2-9/h1-7H,(H,17,18,19)

4-(5-pyridin-4-yl-1H-1,2,4-triazol-3-yl)pyridine-2-carbonitrile

FYX051; FYX-051; FYX 051; Trade names: Topiloric; Uriadec; Topiroxostat.

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)