| Size | Price | Stock | Qty |
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| 1mg |
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| 5mg |
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| 10mg | |||
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| Targets |
EfpA (Mtb efflux pump). BRD-8000.3 specifically inhibits EfpA, an essential major facilitator superfamily (MFS) transporter in Mycobacterium tuberculosis that functions as a lipid flippase, playing a key role in cell envelope biogenesis and drug tolerance. The compound is an uncompetitive inhibitor, meaning it binds to the EfpA-substrate complex rather than the free enzyme. This inhibition disrupts lipid transport and efflux activity, leading to bacterial cell death.
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| ln Vitro |
BRD-8000.3 is a narrow-spectrum bactericidal antimycobacterial medication with good wild-type action that specifically inhibits EfpA [1].
In cell-free biochemical assays, BRD-8000.3 directly inhibits the EfpA efflux pump. Binding is studied using purified EfpA protein, typically reconstituted in a liposome system. Transport activity is measured by encapsulating a fluorogenic substrate like NBD-phosphatidylcholine or a pH-sensitive dye, and the compound's ability to block substrate translocation is monitored. Uncompetitive inhibition is confirmed by kinetic analysis showing that the inhibitor binds only to the enzyme-substrate complex, with a consequent decrease in both Km and Vmax. |
| ln Vivo |
BRD-8000.3 is bactericidal against replicating Mycobacterium tuberculosis with a minimum inhibitory concentration (MIC) of 2 ug/mL. It exhibits a narrow-spectrum activity, being inactive against a panel of Gram-positive and Gram-negative bacteria, consistent with the specificity of EfpA inhibition. The compound is effective against both drug-susceptible and drug-tolerant Mtb strains, making it a valuable tool for targeting persistent infections. It also shows activity in combination with first-line anti-TB drugs to enhance their efficacy.
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| Enzyme Assay |
To confirm binding to EfpA and assess uncompetitive inhibition, surface plasmon resonance (SPR) or isothermal titration calorimetry (ITC) can be used. Purified EfpA protein is immobilized on a chip or in a cell. Increasing concentrations of BRD-8000.3 (0.1-100 uM) are injected to measure binding affinity. For uncompetitive kinetics, EfpA activity is measured in the presence of a fixed concentration of inhibitor (e.g., 2 uM) and varying concentrations of a known substrate (e.g., a fluorescent lipid). The resulting Lineweaver-Burk plot will show parallel lines, characteristic of uncompetitive inhibition. The KI is calculated from the intercepts.
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| Cell Assay |
Determination of MIC values is performed using the microbroth dilution method in 96-well plates. Mycobacterium tuberculosis (e.g., H37Rv strain) is cultured in Middlebrook 7H9 broth supplemented with OADC enrichment and Tween 80. BRD-8000.3 is tested at concentrations ranging from 0.0625-32 ug/mL in 2-fold serial dilutions. The bacterial suspension is added to a final inoculum of ~5×10⁵ CFU/mL. Plates are incubated at 37degC for 7-14 days. The MIC is defined as the lowest concentration of compound that inhibits visible bacterial growth. For time-kill curves, bacteria are treated with 2-4× MIC, and aliquots are plated on agar at various time points to count CFU, confirming bactericidal activity.
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| Animal Protocol |
In vivo efficacy of BRD-8000.3 can be evaluated in a mouse model of chronic tuberculosis infection. Mice (e.g., BALB/c) are infected with M. tuberculosis via the aerosol route. After infection is established (e.g., 4 weeks), mice are treated daily with BRD-8000.3 by oral gavage or intraperitoneal injection at doses ranging from 10-100 mg/kg for 4-8 weeks. The primary endpoint is the bacterial load (CFU count) in the lungs and spleen, determined by homogenizing organs and plating serial dilutions on agar plates. Histopathological analysis of lung tissue is performed to assess inflammation and tissue damage.
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| ADME/Pharmacokinetics |
Formal pharmacokinetic studies for BRD-8000.3 have not been extensively published. The compound has a molecular weight of 401.30, a predicted LogP of ~4-5, and is soluble in DMSO at 50 mg/mL (124.6 mM). For in vivo use, it is typically formulated in a DMSO:PEG300:Tween-80:saline vehicle. Based on its chemical structure, it is expected to have moderate oral bioavailability and to distribute to tissues, including the lung, which is the primary site of Mtb infection. The compound has a sufficient PK profile to achieve therapeutic concentrations in mouse models.
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| Toxicity/Toxicokinetics |
Preclinical toxicology data for BRD-8000.3 are limited. In mouse efficacy studies at therapeutically relevant doses (e.g., 10-50 mg/kg), the compound is generally well-tolerated, with no significant body weight loss or signs of acute toxicity reported. Due to its narrow-spectrum activity against Mtb, off-target effects on the host microbiome are likely to be minimized. However, formal repeated-dose toxicity studies have not been published. As an experimental chemical, it should be handled with appropriate safety precautions, including the use of a fume hood and PPE, to avoid inhalation or skin contact.
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| References | |
| Additional Infomation |
BRD-8000.3 is also known by the chemical name (1S,3S)-N-[6-bromo-5-(pyrimidin-2-yl)pyridin-2-yl]-2,2-dimethyl-3-(2-methylprop-1-en-1-yl)cyclopropane-1-carboxamide. It is a specific inhibitor of the Mtb efflux pump EfpA and a narrow-spectrum bactericidal agent. The compound is used for research into tuberculosis, particularly in addressing drug tolerance. The EfpA protein structure at 2.9 Angstrom and 3.4 Angstrom resolutions has been determined in the presence of this inhibitor, revealing that EfpA forms an antiparallel dimer. BRD-8000.3 is not approved for human use and is intended for laboratory research only.
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| Molecular Formula |
C19H21BRN4O
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| Molecular Weight |
401.30
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| Exact Mass |
400.089
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| CAS # |
2365504-95-4
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| PubChem CID |
162641020
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| Appearance |
White to off-white solid powder
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| LogP |
4
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| Hydrogen Bond Donor Count |
1
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
4
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| Heavy Atom Count |
25
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| Complexity |
526
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| Defined Atom Stereocenter Count |
2
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| SMILES |
C/C(/C)=C/[C@@H]1C(C)(C)[C@H]1C(=O)NC1N=C(Br)C(C2=NC=CC=N2)=CC=1
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| InChi Key |
XIITZGMKUBIQRI-DZGCQCFKSA-N
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| InChi Code |
InChI=1S/C19H21BrN4O/c1-11(2)10-13-15(19(13,3)4)18(25)24-14-7-6-12(16(20)23-14)17-21-8-5-9-22-17/h5-10,13,15H,1-4H3,(H,23,24,25)/t13-,15+/m0/s1
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| Chemical Name |
(1S,3S)-N-(6-bromo-5-pyrimidin-2-ylpyridin-2-yl)-2,2-dimethyl-3-(2-methylprop-1-enyl)cyclopropane-1-carboxamide
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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 : ~50 mg/mL (~124.60 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (6.23 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. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.4919 mL | 12.4595 mL | 24.9190 mL | |
| 5 mM | 0.4984 mL | 2.4919 mL | 4.9838 mL | |
| 10 mM | 0.2492 mL | 1.2460 mL | 2.4919 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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