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| 5mg |
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| Targets |
SLC26A3 (downregulated in adenoma, DRA) anion exchanger - Inhibitor [3]
. - Estimated half-maximal inhibitory concentration (IC₅₀) values for Tenidap in various assays: - Inhibition of db-cAMP-stimulated [Cl⁻]ᵢ increase in noncapacitated mouse sperm: approximately 2.9 μM [3] . - Inhibition of capacitation-associated hyperpolarization in mouse sperm: approximately 2.6 μM [3] . - Inhibition of HCO₃⁻-induced hyperpolarization in mouse sperm: approximately 3.5 μM (estimated from text description) [3] . - Inhibition of capacitation-associated pHᵢ increase in mouse sperm: approximately 3.5 μM [3] . |
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| ln Vitro |
Effect on Intracellular Chloride Concentration ([Cl⁻]ᵢ): In noncapacitated mouse sperm loaded with the Cl⁻-sensitive fluorescent dye MQAE, the addition of db-cAMP (1 mM) and IBMX (100 μM) increased [Cl⁻]ᵢ by approximately 10 mM (from 25 to 35 mM). Preincubation with Tenidap inhibited this db-cAMP-stimulated [Cl⁻]ᵢ increase in a concentration-dependent manner, with an estimated IC₅₀ of 2.9 μM. At higher concentrations (e.g., 10 μM), Tenidap blocked approximately 50% of the response [3]
. - Effect on Membrane Potential (Em): Using the fluorescent dye DiSC₃(5) in mouse sperm populations, Tenidap was shown to inhibit the capacitation-associated hyperpolarization. Sperm incubated in capacitating medium for 60 min hyperpolarized from approximately -36 mV to -60 mV. Tenidap inhibited this hyperpolarization by about 30%, with an estimated IC₅₀ of 2.6 μM. Tenidap also inhibited the fast hyperpolarization induced by HCO₃⁻ addition to noncapacitated sperm, blocking approximately 50% of the response [3] . - Effect on Intracellular pH (pHᵢ): Using the pH-sensitive fluorescent dye BCECF, Tenidap was found to inhibit the capacitation-associated pHᵢ increase. Mouse sperm pHᵢ increased from approximately 6.4 (noncapacitated) to 6.8 after 60 min of capacitation. Tenidap inhibited this pHᵢ increase by approximately 34%, with an estimated IC₅₀ of 3.5 μM [3] . - Comparison with Other Inhibitors: The effects of Tenidap were compared with another SLC26A3 inhibitor (UK-5099), a CFTR inhibitor (inh-172), and SLC26A6 inhibitors (DOG and PMA). Tenidap and UK-5099 showed similar inhibitory profiles on [Cl⁻]ᵢ, Em, and pHᵢ, while DOG and PMA did not affect [Cl⁻]ᵢ or Em but did partially inhibit the pHᵢ increase [3] . |
| Cell Assay |
Sperm Preparation: Cauda epididymal mouse sperm were collected from CD1 retired male breeders and washed in Whitten Hepes-buffered medium. For noncapacitated conditions, sperm were used shortly after preparation. For capacitated conditions, sperm were incubated in complete Whitten Hepes-buffered medium supplemented with 5 mg/ml BSA and 24 mM NaHCO₃ for 60 min at 37°C [3]
. - Intracellular Chloride Measurement: Sperm were loaded with 10 mM MQAE (a Cl⁻-sensitive fluorescent dye) for 30 min at 37°C. Excess dye was removed by centrifugation. Fluorescence was recorded at 350/460 nm excitation/emission. Tenidap (at various concentrations) was added to the sperm suspension, followed by db-cAMP/IBMX to stimulate [Cl⁻]ᵢ increase. Fluorescence changes were converted to [Cl⁻]ᵢ using a calibration curve established with digitonin permeabilization and varying external Cl⁻ concentrations [3] . - Membrane Potential Measurement: Sperm were incubated in noncapacitating or capacitating conditions with or without Tenidap. Eight minutes before measurement, 1 μM DiSC₃(5) was added, followed by 1 μM CCCP (to collapse mitochondrial potential) 5 min later. Fluorescence was recorded at 640/670 nm excitation/emission. Calibration was performed by adding 1 μM valinomycin and sequential KCl additions to determine Em values [3] . - Intracellular pH Measurement: Sperm were loaded with 0.5 μM BCECF for 15 min at 37°C. Excess dye was removed by centrifugation. Cells were incubated under noncapacitating or capacitating conditions with or without Tenidap. Fluorescence was recorded at 510/450 nm excitation and 550 nm emission. Calibration was performed by adding 0.12% Triton X-100 and subsequent HCl additions, with pH measured by a conventional pH electrode at each step [3] . - Drug Preparation: Tenidap was prepared in dimethyl sulfoxide (DMSO) and stored at -20°C until use. Final DMSO concentrations were kept constant in all experiments with appropriate controls [3] . |
| References |
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| Additional Infomation |
Tenidap belongs to the indole, urea, thiophene, and organochlorine compounds class. It is a nonsteroidal anti-inflammatory drug and an EC 1.14.99.1 (prostaglandin intraperoxidase) inhibitor.
Role in This Study: Tenidap was used as a pharmacological tool to investigate the functional role of the SLC26A3 anion exchanger in mouse sperm capacitation. Along with another inhibitor (UK-5099), it helped establish that SLC26A3, together with CFTR, participates in the regulation of [Cl⁻]ᵢ, membrane potential, and pHᵢ during capacitation [3] . - SLC26A3 Inhibitor Profile: Tenidap is described as an antagonist of SLC26A3. Its effects were compared with those of UK-5099 (another SLC26A3 inhibitor), inh-172 (a CFTR inhibitor), and DOG/PMA (compounds that indirectly inhibit SLC26A6 via PKC activation). The similar inhibitory profiles of Tenidap and UK-5099, distinct from those of DOG and PMA, supported the specific involvement of SLC26A3 in the observed capacitation-associated changes [3] . - Source: Tenidap used in this study was kindly provided by Pfizer Inc [3] . - Significance: By demonstrating that Tenidap partially inhibits key capacitation-associated events ([Cl⁻]ᵢ increase, hyperpolarization, pHᵢ increase), the study provided evidence that SLC26A3-mediated Cl⁻/HCO₃⁻ exchange is an important component of the ion transport mechanisms underlying sperm capacitation in mice [3] . |
| Molecular Formula |
C14H9CLN2O3S
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|---|---|
| Molecular Weight |
320.747
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| Exact Mass |
320.002
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| CAS # |
120210-48-2
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| Related CAS # |
Tenidap-d3;142741-60-4
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| PubChem CID |
60712
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| Appearance |
White to yellow solid powder
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| Density |
1.58g/cm3
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| Boiling Point |
523.9ºC at 760mmHg
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| Melting Point |
230° (dec)
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| Flash Point |
270.7ºC
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| Vapour Pressure |
7.42E-13mmHg at 25°C
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| Index of Refraction |
1.756
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| LogP |
4.018
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
2
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| Heavy Atom Count |
21
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| Complexity |
450
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
IZSFDUMVCVVWKW-UHFFFAOYSA-N
|
| InChi Code |
InChI=1S/C14H9ClN2O3S/c15-7-3-4-9-8(6-7)11(13(19)17(9)14(16)20)12(18)10-2-1-5-21-10/h1-6,19H,(H2,16,20)
|
| Chemical Name |
5-chloro-2-hydroxy-3-(thiophene-2-carbonyl)indole-1-carboxamide
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| Synonyms |
Tenidap CP66,248 CP-66248CP-66,248 CP 66,248 CP 66248 CP66248
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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 : ~25 mg/mL (~77.94 mM)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: 2.5 mg/mL (7.79 mM) in 10% DMSO + 40% PEG300 +5% Tween-80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
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. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.1177 mL | 15.5885 mL | 31.1769 mL | |
| 5 mM | 0.6235 mL | 3.1177 mL | 6.2354 mL | |
| 10 mM | 0.3118 mL | 1.5588 mL | 3.1177 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.
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