Mtb F1FO-ATP synthase
TDR M. tuberculosis strains are inhibited in growth by bedaquiline, with MIC values ranging from 0.125 to 0.5 mg/L[1]. With MIC50 and MIC90 values of 0.03 and 16 mg/L, respectively, bedaquiline has the strongest activity against Mycobacterium avium among slowly growing mycobacteria (SGM). Among the mycobacteria that grow quickly (RGM), Mycobacterium abscessus subsp.With MIC50 and MIC90 values of 0.13 and >16 mg/L, respectively, for both species, abscessus (M. abscessus) and Mycobacterium abscessus subsp. massiliense (M. massiliense) appear to be more susceptible to bedaquiline than Mycobacterium fortuitum. Moderate in vitro activity of bedaquiline against NTM species is also demonstrated[2]. In vitro activity of bedaquiline against Mycobacterium tuberculosis, including multidrug-resistant M tuberculosis, is very good[3].

TDR M. tuberculosis strains are inhibited in growth by bedaquiline, with MIC values ranging from 0.125 to 0.5 mg/L[1]. With MIC50 and MIC90 values of 0.03 and 16 mg/L, respectively, bedaquiline has the strongest activity against Mycobacterium avium among slowly growing mycobacteria (SGM). Among the mycobacteria that grow quickly (RGM), Mycobacterium abscessus subsp.With MIC50 and MIC90 values of 0.13 and >16 mg/L, respectively, for both species, abscessus (M. abscessus) and Mycobacterium abscessus subsp. massiliense (M. massiliense) appear to be more susceptible to bedaquiline than Mycobacterium fortuitum. Moderate in vitro activity of bedaquiline against NTM species is also demonstrated[2]. In vitro activity of bedaquiline against Mycobacterium tuberculosis, including multidrug-resistant M tuberculosis, is very good[3].

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Bedaquiline exhibited bactericidal activity against Mycobacterium tuberculosis H37Rv in time-kill assays. Treatment over 21 days with concentrations equivalent to 3-, 30-, or 300-fold the MIC90 resulted in a significant reduction in viable bacterial counts (CFU/ml).[2]
The MIC90 of Bedaquiline against Mycobacterium smegmatis was reported as 100 nM, and against Mycobacterium bovis BCG as 70 nM.[2]
Bedaquiline displayed protonophore (uncoupler) activity. In experiments with inverted membrane vesicles from M. smegmatis, 15 µM Bedaquiline completely eliminated the succinate-generated transmembrane pH gradient, as measured by the quenching reversal of the pH-sensitive fluorophore ACMA. This activity was dose-dependent.[2]
In whole-cell Mycobacterium bovis BCG, treatment with 1 µM Bedaquiline caused a rapid intracellular pH increase of approximately 0.35 units within 20 minutes, indicating its ability to cross membranes and act as a weak base.[2]

BDQ was highly efficacious in a zebrafish model of M. abscessus infection. Remarkably, a very short period of treatment was sufficient to protect the infected larvae from M. abscessus-induced killing. This was corroborated with reduced numbers of abscesses and cords, considered to be major pathophysiological signs in infected zebrafish. [7]

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The study primarily reports in vitro data. It cites a separate preliminary report (Philley et al.) that demonstrated potential clinical and microbiological activity of bedaquiline in patients with advanced M. avium complex or M. abscessus disease. Conversely, it also references a nude mouse model experiment which indicated that bedaquiline alone did not prevent death, potentially due to high minimal bactericidal concentrations. The current study did not conduct its own in vivo experiments.[4]

Intracellular ATP quantification. Intracellular ATP levels were determined using a 96-well flat-bottom plate, as described previously for M. tuberculosis. M. abscessus was exposed to BDQ or amikacin (negative control) and incubated for 180 min at 32°C. Twenty-five microliters of M. abscessus culture was mixed with an equal volume of the BacTiter-Glo reagent in 96-well flat-bottom white plates and incubated for 5 min in the darkness. Luminescence was detected using a BioTek Cytation 3 multimode reader, and the values obtained were plotted using GraphPad Prism 6 software.[7]

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Protonophore Activity Assay using Inverted Membrane Vesicles: To assess the uncoupler activity of Bedaquiline, inverted vesicles were prepared from the plasma membrane of Mycobacterium smegmatis. The pH-sensitive fluorescent dye ACMA was used to monitor changes in the transmembrane pH gradient. Vesicles were energized by adding an electron donor, either 0.5 mM succinate or 2 mM NADH, leading to proton pumping and establishment of a pH gradient, visualized as quenching of ACMA fluorescence. Upon addition of Bedaquiline at various concentrations, its protonophore activity was measured by the resulting dequenching (increase) of fluorescence, indicating dissipation of the pH gradient. The known protonophore SF6847 was used as a positive control to fully collapse the gradient at the end of each run.[2]

Drug susceptibility testing. [7] The CLSI guidelines were followed to determine the MICs based on the broth microdilution method in CaMHB using an inoculum containing 5 × 106 CFU/ml in the exponential-growth phase. Bacteria (100 μl) were seeded in 96-well plates, and 2 μl of drug at its highest concentration was added to the first wells containing double the volume of bacterial suspension (200 μl). Twofold serial dilutions were then carried out, and incubation with drugs was performed at 30°C for 3 to 5 days. MICs were recorded by visual inspection and by absorbance at 560 nm to confirm visual recording. Experiments were done in triplicate on three independent occasions. Time-kill assay.[7] Microtiter plates were set up as for MIC determination. Serial dilutions of the bacterial suspension were plated after 0, 24, 48, 72, and 96 h of exposure to different drug concentrations. CFU were enumerated after 4 days of incubation at 30°C.

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Intracellular pH Measurement in Whole Bacterial Cells: The effect of Bedaquiline on the intracellular pH of Mycobacterium bovis BCG was measured using the pH-sensitive fluorophore CMFDA. Bacterial cultures were loaded with the dye. In the absence of an external pH gradient, the cells were treated with different concentrations of Bedaquiline. Changes in the fluorescence intensity of CMFDA, which correlates with intracellular pH, were monitored over time (up to 20 minutes). The protonophore CCCP was used as a positive control for intracellular acidification. This assay demonstrated Bedaquiline's ability to rapidly alkalinize the bacterial cytoplasm due to its membrane-permeant weak base properties.[2]
Time-Kill Assay against M. tuberculosis: The bactericidal activity of Bedaquiline was evaluated against Mycobacterium tuberculosis H37Rv. Bacterial cultures were treated with Bedaquiline at concentrations equivalent to 3, 30, and 300 times its MIC90. Cultures were incubated over a 21-day period. At specified time points, samples were taken, serially diluted, and plated on solid medium to enumerate colony-forming units (CFU). The reduction in CFU/ml over time was plotted to generate kill curves, which were compared to those of the analog TBAJ-876 and a drug-free control.[2]

Assessment of BDQ efficacy in infected zebrafish. [7] Rough M. abscessus CIP104536T (ATCC 19977T) carrying pTEC27 (plasmid 30182; Addgene) and expressing the red fluorescent protein tdTomato was prepared and microinjected in zebrafish embryos, according to procedures described earlier. Briefly, mid-log-phase cultures of M. abscessus expressing tdTomato were centrifuged, washed, and resuspended in phosphate-buffered saline (PBS) supplemented with 0.05% Tween 80 (PBS-T). Bacterial suspensions were then homogenized through a 26-gauge needle and sonicated, and the remaining clumps were allowed to settle down for 5 to 10 min. Bacteria were concentrated to an optical density at 600 nm (OD600) of 1 in PBS-T and injected intravenously (≈2 to 5 nl containing 50 to 300 CFU) into the caudal vein in 30-h-postfertilization (hpf) embryos previously dechorionated and anesthetized. To follow infection kinetics and embryo survival, infected larvae were transferred into 24-well plates (2 embryos/well) and incubated at 28.5°C. The CFU numbers in the inoculum were determined by injection of 2 nl of the bacterial suspension in sterile PBS-T and plating on 7H10 with 500 μg/ml hygromycin.

Effects During Pregnancy and Lactation
◉ Overview of Drug Use During Lactation
Data from two women taking bedaquiline and their breastfed infants showed that the infants were exposed to high drug concentrations through breast milk, with one infant reaching therapeutic serum drug concentrations. The clinical consequences of this exposure are unclear. This drug may protect infants from multidrug-resistant tuberculosis infection, but it may also cause adverse reactions. Breastfeeding should not be discontinued if the mother is taking bedaquiline. Monitor the breastfed infant for adverse reactions such as insufficient weight gain, hepatotoxicity, nausea, arthralgia, headache, hemoptysis, and chest pain.
◉ Effects on Breastfed Infants
A woman co-infected with HIV and rifampicin-resistant tuberculosis was taking bedaquiline (dosage not specified) as part of her anti-tuberculosis treatment regimen, which also included pyrazinamide and other unnamed drugs. At one-month follow-up, the infant was underweight and had slow growth, but the mother experienced nausea due to drug treatment and also experienced weight loss. Six months later, after the mother completed her treatment, the baby's weight began to increase, following a normal growth curve trajectory and reaching developmental milestones.
◉ Impact on breastfeeding and breast milk
As of the revision date, no relevant published information was found.

[1]. Bedaquiline susceptibility test for totally drug-resistant tuberculosis Mycobacterium tuberculosis. J Microbiol. 2017 Apr 20.

[2]. TBAJ-876 displays Bedaquiline-like mycobactericidal potency without retaining the parental drug's uncoupler activity. Antimicrob Agents Chemother. 2019 Nov 11.

[3]. Bedaquiline: a novel diarylquinoline for multidrug-resistant tuberculosis. Ann Pharmacother. 2014 Jan;48(1):107-15.

[4]. In Vitro Activity of Bedaquiline against Nontuberculous Mycobacteria in China. Antimicrob Agents Chemother. 2017 Apr 24;61(5).

[5]. Ann Pharmacother.2014 Jan;48(1):107-15.

[6]. Antimicrob Agents Chemother.2017 Apr 24;61(5). pii: e02627-16.
[7]. Antimicrob Agents Chemother. 2017 Nov; 61(11): e01225-17.

Bedaquiline fumarate is a fumarate salt prepared from equimolar amounts of bedaquiline and fumaric acid. It treats multidrug-resistant tuberculosis (MDR-TB) by inhibiting ATP synthase (an enzyme crucial for mycobacterial replication) and is used in combination with other drugs. It is both an anti-tuberculosis drug and an ATP synthase inhibitor. It contains bedaquiline (2+) ions. Bedaquiline fumarate is the fumarate form of bedaquiline, a highly bioavailable, orally bioavailable diarylquinoline anti-mycobacterial drug used to treat multidrug-resistant tuberculosis (MDR-TB). After oral administration, bedaquiline specifically binds to the C subunit of ATP synthase in Mycobacterium tuberculosis, thereby inhibiting ATP synthase activity. This product inhibits ATP synthesis in Mycobacterium tuberculosis, thereby blocking its energy metabolism and killing the bacteria. See also: Bedaquiline (with active ingredient).

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Drug Indications
Sirturo is indicated for the combined treatment of multidrug-resistant tuberculosis (MDR-TB) in adult and adolescent patients (aged 12 to 18 years and weighing at least 30 kg) for the treatment of cases where an effective treatment regimen cannot be established due to drug resistance or tolerance. Official guidelines for the rational use of antimicrobial drugs should be considered.
Treatment of Multidrug-Resistant Tuberculosis

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Bedaquiline (Sirturo) is a diarylquinoline drug used to treat multidrug-resistant tuberculosis. [2]
This study revealed that bedaquiline has a dual mechanism of action: 1) inhibition of F1F0-ATP synthase; 2) proton carrier activity that can uncouple oxidative phosphorylation. [2]
This uncoupling activity is thought to contribute to the bactericidal effect of the drug against Mycobacterium tuberculosis. [2] This study compared bedaquiline with its analogue TBAJ-876, noting that TBAJ-876 had significantly reduced proton carrier activity due to its lower lipophilicity (clogP 5.15). While its activity was 7.25 times that of bedaquiline, its bactericidal efficacy was similar, suggesting that uncoupling activity may not be a necessary condition for diarylquinoline drugs to exert their antimycobacterial effects. [2]

C36H35BRN2O6

671.59

670.167

C, 64.38; H, 5.25; Br, 11.90; N, 4.17; O, 14.29

845533-86-0

Bedaquiline;843663-66-1;(Rac)-Bedaquiline;654655-80-8

24812732

White to off-white solid powder

6.842

3

8

10

45

834

2

OC(/C=C/C(O)=O)=O.BrC1=CC=C(N=C(OC)C([C@H]([C@@](C2=CC=CC3=C2C=CC=C3)(O)CCN(C)C)C4=CC=CC=C4)=C5)C5=C1

ZLVSPMRFRHMMOY-WWCCMVHESA-N

InChI=1S/C32H31BrN2O2.C4H4O4/c1-35(2)19-18-32(36,28-15-9-13-22-10-7-8-14-26(22)28)30(23-11-5-4-6-12-23)27-21-24-20-25(33)16-17-29(24)34-31(27)37-3;5-3(6)1-2-4(7)8/h4-17,20-21,30,36H,18-19H2,1-3H3;1-2H,(H,5,6)(H,7,8)/b;2-1+/t30-,32-;/m1./s1

(1R,2S)-1-(6-Bromo-2-methoxy-3-quinolyl)-4-dimethylamino-2-(1-naphthyl)-1-phenyl-butan-2-ol fumarate

R207910 fumarate; TMC-207 fumarate; R-207910; TMC 207; R 207910; TMC207 fumarate; Bedaquiline fumarate; trade name: Sirturo

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

DMSO : ~100 mg/mL ( ~148.9 mM )
Ethanol : ~4 mg/mL

Solubility in Formulation 1: ≥ 2.75 mg/mL (4.09 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 27.5 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.

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Solubility in Formulation 2: 2.75 mg/mL (4.09 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), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 27.5 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.

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Solubility in Formulation 3: ≥ 2.75 mg/mL (4.09 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 27.5 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: ≥ 2.75 mg/mL (4.09 mM) (saturation unknown) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 5: ≥ 2.75 mg/mL (4.09 mM) (saturation unknown) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
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.

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