The primary molecular target of sitaxsentan is the endothelin A (ETA) receptor. Sitaxsentan acts as a competitive antagonist of endothelin-1 at the ETA receptor, with Ki values in the low nanomolar range (0.43 nM). The compound shows high selectivity for ETA over ETB receptors, with approximately 100- to 1000-fold selectivity depending on the assay system. By selectively blocking ETA receptors, sitaxsentan prevents the vasoconstrictive and proliferative effects mediated by endothelin-1, leading to vasodilation and reduced pulmonary vascular resistance. Sitaxsentan also inhibits ET-1-induced stimulation of phosphoinositide turnover with a Ki of 0.686 nM and pA2 of 8.0. The compound shows moderate inhibitory activity against PDE4A and demonstrates antiviral activity against SARS-CoV-2.

Sitaxentan sodium inhibits ET-1-induced stimulation of phosphoinositide turnover with a Ki of 0.69 nM and a pA2 of 8.0.
Kinase Assay: Binding studies are performed in a 30 mM HEPES buffer, pH 7.4, containing 150 mM NaCl, 5 mM MgCl2, and 0.05% bacitracin using 2 mg/tube (ETA) or 0.75 mg/tube (ETB) membrane. Sitaxentan sodium is dissolved in DMSO and diluted with the assay buffer to give a final concentration of 0.25% DMSO. Competitive inhibition experiments are performed in triplicate in a final volume of 200 μL containing 4 pM [125I]ET-1 (1.6 nCi). Nonspecific binding is determined in the presence of 100 nM ET-1. Samples are incubated for 16 hours−18 hours at 24 °C. One milliliter of PBS is then added and the assay centrifuged at 2000 g for 25 minutes at 4 °C. The supernatant is decanted and the membrane bound radioactivity counted on a Genesys gamma counter.
Cell Assay: TE 671 or transfected COS 7 cells are grown to confluence in six-well plates. Sixteen hours prior to use, the media in each well is replaced with 2 mL of inositol-free RPMI-164 (IF-RPMI) media containing 10% inositol-free FCS and 2 mCi [3H]myoinositol and incubated at 37 °C in the presence of 6% CO2. The media is aspirated, and the cells are washed twice with PBS. Cells are preincubated for 10 minutes in 1 mL of lithium buffer (15 μM HEPES, pH 7.4, 145 μM NaCl, 5.4 μM KCl, 1.8 μM CaCl2, 0.8 μM MgSO4, 1.0 μM NaH2PO4, 11.2 μM glucose, 20 μM LiCl) with or without Sitaxentan sodium prior to the addition of 100 μM of ET-1 at different concentrations. Cells are then incubated for an additional 45 minutes. The buffer is discarded, and the accumulated inositol phosphates are extracted with ice cold methanol. The total cell protein in each well is measured using the BCA assay after solubilizing the cells in 0.1 M NaOH.
In vitro studies demonstrate that sitaxsentan potently inhibits endothelin-1 binding to ETA receptors. The compound inhibits ET-1-induced stimulation of phosphoinositide turnover with a Ki of 0.686 nM and pA2 of 8.0, demonstrating potent functional antagonism. Sitaxsentan shows high selectivity for ETA over ETB receptors, ensuring that its vasodilatory effects are achieved without blocking ETB-mediated vasodilation and endothelin clearance. In cell-based assays, sitaxsentan inhibits endothelin-1-induced vasoconstriction and vascular smooth muscle cell proliferation. The compound's antiviral activity against SARS-CoV-2 has been demonstrated in vitro. Sitaxsentan shows moderate inhibitory activity against phosphodiesterase 4A (PDE4A), an enzyme involved in inflammatory signaling. These in vitro activities support its use as a research tool in cardiovascular and pulmonary research.

Sitaxentan sodium has a serum half-life in the rat and the dog of 6 hours - 7 hours and 60−100% oral bioavailability. Orally administered Sitaxentan sodium is rapidly absorbed in both the rat and the dog with a t1/2(abs) of 0.7 hours and 0.3 hours, respectively. Peak plasma concentrations occurred between 2 hours and 3 hours postdosing in the rat and between 45 minutes and 90 minutes in the dog. The pulmonary vasoconstrictor response to acute hypoxia (10% O2 for 90 minutes) is prevented with Sitaxentan sodium (5 mg/kg infused i.v. 10 minutes prior to the onset of hypoxia). Sitaxentan sodium delivered i.v. 50 minutes after the onset of hypoxia reverses the established pulmonary vasoconstriction. Sitaxsentan blocks increased plasma endothelin levels. Sitaxsentan dose dependently (10 mg/kg and 50 mg/kg per day in the drinking water) attenuates right ventricular systolic pressure, right heart hypertrophy, and pulmonary vascular remodeling observed 3 weeks after a single subcutaneous injection of monocrotaline. Systemic administration of the ETA receptor antagonist Sitaxentan sodium significantly attenuates cerebral vasospasm after subarachnoid hemorrhage (SAH). Sitaxentan sodium reduces the development of hypoxic pulmonary vasoconstriction (HPV) in the pig. In addition, bolus injection of Sitaxentan sodium reverses already established HPV.

For ETA receptor binding assays, competition binding studies are performed using membrane preparations from cells expressing recombinant human ETA receptors. Membranes are incubated with radiolabeled endothelin-1 ([¹²⁵I]-ET-1, typically 20-50 pM) and varying concentrations of sitaxsentan (0.001 nM to 10 µM) in binding buffer (50 mM Tris-HCl, pH 7.4, containing 5 mM MgCl₂, 0.1% BSA, and protease inhibitors) at 25°C for 2-4 hours. Bound and free radioligand are separated by filtration through GF/B filters pre-soaked in 0.3% polyethylenimine, and radioactivity is counted by gamma counting. Specific binding is defined as total binding minus non-specific binding (determined in the presence of 100 nM unlabeled ET-1). Ki values are calculated from IC50 values using the Cheng-Prusoff equation. For functional assays, ET-1-induced stimulation of phosphoinositide turnover is measured in cells expressing ETA receptors. Cells are labeled with [³H]-myo-inositol, treated with sitaxsentan (0.001-10 µM) and ET-1 (1-100 nM), and [³H]-inositol phosphate accumulation is measured by ion-exchange chromatography.

For cell-based assays, vascular smooth muscle cells or endothelial cells are cultured in appropriate medium (e.g., DMEM or M199) with 10% FBS and antibiotics. Cells are seeded in 6-well or 96-well plates and grown to confluence. Sitaxsentan is dissolved in DMSO and diluted in culture medium to final concentrations (typically 0.001-10 µM). Cells are pre-treated with sitaxsentan for 30-60 minutes, then stimulated with endothelin-1 (10-100 nM) for 5-60 minutes. Intracellular calcium levels are measured using fluorescent calcium indicators (e.g., Fura-2 AM or Fluo-4). Phosphoinositide turnover is assessed by measuring [³H]-inositol phosphate accumulation as described above. Cell proliferation is assessed by [³H]-thymidine incorporation or BrdU incorporation assays. Apoptosis and survival signaling (e.g., Akt, ERK phosphorylation) are assessed by Western blot. For antiviral assays, SARS-CoV-2-infected Vero E6 cells are treated with sitaxsentan, and viral replication is measured by plaque assay or qRT-PCR.

5 mg/kg infused i.v.; 10 mg/kg and 50 mg/kg
Rats, dogs, pigs

Absorption, Distribution and Excretion
70-100% Kidneys (50% to 60%) Feces (40% to 50%) Metabolism/Metabolites Liver (CYP2C9 and CYP3A4 mediated) Biological Half-Life 10 hours
Pharmacokinetic data for sitaxsentan show a serum half-life ranging from 4.1 to 7.5 hours depending on the species. The compound has excellent oral bioavailability of 70-100%. Sitaxsentan is highly protein-bound (>99%). The drug undergoes hepatic metabolism primarily via CYP2C9 and CYP3A4. Sitaxsentan achieves once-daily dosing due to distinct metabolic pathways and extended duration of action, whereas bosentan requires twice-daily administration. The compound shows excellent pharmacokinetic profile and oral bioavailability. The drug's favorable pharmacokinetics supported its clinical development as a once-daily oral therapy for PAH. However, hepatotoxicity concerns led to its withdrawal from the market.

Protein Binding
99%+
Sitaxsentan demonstrated hepatotoxicity in clinical use, leading to its voluntary withdrawal from the market in 2010. The drug was associated with elevated liver enzymes (ALT and AST) in a significant proportion of patients, and cases of liver failure were reported. This hepatotoxicity is likely related to the sulfonamide moiety of the drug and its metabolism via CYP2C9, which may generate reactive intermediates. Sitaxsentan shows moderate liver toxicity with elevated ALT and AST levels. The compound is contraindicated in patients with pre-existing liver disease. Other potential adverse effects include headache, peripheral edema, and hypotension. The safety concerns ultimately outweighed the therapeutic benefits, leading to the drug's withdrawal. Despite its withdrawal, sitaxsentan remains a valuable research tool for studying endothelin receptor pharmacology and serves as a reference compound for developing safer ETA receptor antagonists.

J Med Chem.1997;40(11):1690-7;Pulm Pharmacol Ther.2000;13(2):87-97;Neurosurgery.1998;43(6):1409-17.

Sitaxentan belongs to the benzodioxane class of drugs. It was previously marketed under the brand name Thelin by Encysive Pharmaceuticals for the treatment of pulmonary arterial hypertension (PAH) until Pfizer acquired Encysive in February 2008. In 2010, Pfizer withdrew Sitaxentan from the market due to concerns about its hepatotoxicity. Drug Indications It has been studied for the treatment of pulmonary arterial hypertension, connective tissue diseases, hypertension, and congestive heart failure. Mechanism of Action Sitaxentan is a competitive antagonist of endothelin-1 at both endothelin A (ET-A) and endothelin B (ET-B) receptors. Normally, the binding of endothelin-1 to either ET-A or ET-B receptors leads to pulmonary vasoconstriction. Sitaxentan reduces pulmonary vascular resistance by blocking this interaction. Sitaxentan has a higher affinity for endothelin A (ET-A) than for endothelin B (ET-B).

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Pharmacodynamics
Sitasentan belongs to the class of endothelin receptor antagonists (ERAs). Patients with pulmonary arterial hypertension (PAH) have elevated levels of endothelin (a potent vasoconstrictor) in their plasma and lung tissue. Sitasentan blocks the binding of endothelin to its receptors, thereby eliminating the harmful effects of endothelin.

Sitaxsentan (IPI-1040, TBC-11251) is a selective endothelin A (ETA) receptor antagonist that was developed for the treatment of pulmonary arterial hypertension. It was marketed as Thelin by Encysive Pharmaceuticals until Pfizer acquired the company in 2008. The drug achieved once-daily dosing due to its favorable pharmacokinetic profile, including high oral bioavailability (70-100%) and extended duration of action. However, sitaxsentan was voluntarily withdrawn from the market in 2010 due to hepatotoxicity, including elevated liver enzymes and liver failure. Despite this setback, the compound remains an important research tool for studying endothelin receptor pharmacology and serves as a reference compound for developing safer ETA receptor antagonists. Sitaxsentan also shows moderate inhibitory activity against PDE4A and antiviral activity against SARS-CoV-2. It is now used exclusively for research purposes.

C₁₈H₁₅CLN₂O₆S₂

454.90

454.006

184036-34-8

Sitaxsentan sodium;210421-74-2

216235

Typically exists as solid at room temperature

1.6g/cm3

600.4ºC at 760 mmHg

316.9ºC

2.25E-14mmHg at 25°C

1.687

5.115

1

9

6

29

720

0

O=S(C1=C(C(CC2=CC3=C(C=C2C)OCO3)=O)SC=C1)(NC4=C(C(C)=NO4)Cl)=O

PHWXUGHIIBDVKD-UHFFFAOYSA-N

InChI=1S/C18H15ClN2O6S2/c1-9-5-13-14(26-8-25-13)7-11(9)6-12(22)17-15(3-4-28-17)29(23,24)21-18-16(19)10(2)20-27-18/h3-5,7,21H,6,8H2,1-2H3

N-(4-chloro-3-methyl-1,2-oxazol-5-yl)-2-[2-(6-methyl-1,3-benzodioxol-5-yl)acetyl]thiophene-3-sulfonamide

IPI-1040Thelin Sitaxentan sodiumIPI-1040 TBC11251IPI1040TBC-11251 IPI 1040TBC 11251 TBC-11251 sodium salt, Sitaxsentan

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

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.)