F1F0 ATP hydrolase (IC50=0.5 μM)
The primary target of BMS-199264 hydrochloride is the ATP hydrolase activity of the mitochondrial F1F0 ATP synthase complex. The F1F0 ATP synthase is a key enzyme in mitochondrial energy production that synthesizes ATP during oxidative phosphorylation. Under ischemic conditions, the enzyme can reverse its activity and hydrolyze ATP, leading to ATP depletion and cell death. BMS-199264 hydrochloride selectively inhibits the ATP hydrolase activity of this enzyme with an IC50 of 0.5 μM, without inhibiting the ATP synthase activity. This selective inhibition preserves ATP levels during ischemia and protects cardiac tissue.

Shortening the effect period and lowering LDH release, BMS-199264 hydrochloride (1 μM, 3 μM, and 10 μM) increased in wastewater after 25 minutes of whole brain and 30 minutes of reinfusion in a concentration-dependent manner [1]. μM) differ in their impact on the activity of buffer enzymes and ATP synthase, measuring 0.18 μMATP/min/mg and 0.23 μMATP/min/mg, respectively[1].

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BMS-199264 hydrochloride demonstrates potent in vitro inhibition of F1F0 ATP hydrolase activity, with an IC50 of 0.5 μM. The compound shows no inhibitory effect on F1F0 ATP synthase, indicating a high degree of selectivity for the hydrolase activity. This selectivity is a key feature of the compound's pharmacological profile and underlies its cardioprotective effects.

Cardiac function data pre- and postischemia (30 min into reperfusion) is shown in table 1. BMS-199264 showed modest preischemic cardiodepression only at the high dose, but unfortunately, this compound is so poorly soluble that the solution was cloudy, and it is likely that this is the cause of the cardiodepression at that concentration. In vehicle-treated hearts, reperfusion did not cause significant recovery of contractile function, which is expected due to the severity of the global ischemia. Interestingly, BMS-199264 caused a concentration-dependent improvement in contractile function, unlike data seen for oligomycin or aurovertin.
BMS-199264 increased the time to onset of contracture in a concentration-dependent manner (Fig. 4). Since contracture is due to rigor bond formation, conservation of ATP can be inferred. LDH release (cumulative during 30 min of reperfusion) was reduced in a concentration-dependent manner, suggesting reduced necrosis (Fig. 4).[1]

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BMS-199264 hydrochloride selectively inhibits ATP decline during ischemia, reduces cardiac necrosis, and enhances the recovery of contractile function following reperfusion in vivo. These findings confirm the compound's cardioprotective potential in ischemic conditions. However, specific in vivo efficacy data, including animal models and detailed dosing regimens, are not extensively detailed in the available literature.

The mitochondrial F1F0 ATP synthase is responsible for the majority of ATP production in mammals and does this through a rotary catalytic mechanism. Studies show that the F1F0 ATP synthase can switch to an ATP hydrolase, and this occurs under conditions seen during myocardial ischemia. This ATP hydrolysis causes wasting of ATP that does not produce work. The degree of ATP inefficiently hydrolyzed during ischemia may be as high as 50-90% of the total. A naturally occurring, reversible inhibitor (IF-1) of the hydrolase activity is in the mitochondria, and it has a pH optimum of 6.8. Based on studies with the nonselective (inhibit both synthase and hydrolase activity) inhibitors aurovertin B and oligomycin B reduce the rate of ATP depletion during ischemia, showing that IF-1 does not completely block hydrolase activity. Oligomycin and aurovertin cannot be used for treating myocardial ischemia as they will reduce ATP production in healthy tissue. We generated a focused structure-activity relationship, and several compounds were identified that selectively inhibited the F1F0 ATP hydrolase activity while having no effect on synthase function. One compound, BMS-199264 had no effect on F1F0 ATP synthase function in submitochondrial particles while inhibiting hydrolase function, unlike oligomycin that inhibits both. BMS-199264 selectively inhibited ATP decline during ischemia while not affecting ATP production in normoxic and reperfused hearts. BMS-191264 also reduced cardiac necrosis and enhanced the recovery of contractile function following reperfusion. These data also suggest that the reversal of the synthase and hydrolase activities is not merely a chemical reaction run in reverse.[1]

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In vitro enzyme assays for BMS-199264 hydrochloride typically involve measuring its inhibition of F1F0 ATP hydrolase activity. Mitochondrial membranes or purified F1F0 ATP synthase complexes are incubated with ATP in the presence of varying concentrations of BMS-199264 hydrochloride. The hydrolysis of ATP to ADP is measured, and the IC50 for inhibition is calculated, with BMS-199264 hydrochloride showing an IC50 of 0.5 μM. The compound's lack of effect on ATP synthase activity is confirmed in parallel assays.

Cellular assays for BMS-199264 hydrochloride are performed in cardiac cells or isolated mitochondria to assess its effects on ATP levels and cell viability under ischemic conditions. Cells are subjected to ischemic stress in the presence or absence of BMS-199264 hydrochloride, and ATP levels are measured. The compound's ability to preserve ATP levels and protect cells from ischemic damage is assessed.

Effect of BMS-199264 on pre- and postischemic cardiac function (left ventricular developed pressure [LVDP] in isolated rat hearts subjected to a 25-min global ischemia followed by a 30-min reperfusion.
Effect of increasing concentrations of the mitochondrial ATP hydrolase inhibitor BMS-199264 on the time to the onset of ischemic contracture and reperfusion cumulative LDH release in isolated rat hearts after a 25-min global ischemia followed by a 30-min reperfusion. BMS-199264 increased the time to contracture and reduced LDH release in a concentration-dependent manner, which was not blocked by glyburide. [1]

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In vivo animal studies with BMS-199264 hydrochloride have been performed in models of cardiac ischemia-reperfusion injury. In these studies, the compound is administered systemically, and its effects on ATP levels, cardiac necrosis, and contractile function are assessed. BMS-199264 hydrochloride selectively inhibits ATP decline during ischemia, reduces cardiac necrosis, and enhances the recovery of contractile function following reperfusion.

BMS-199264 hydrochloride has a molecular formula of C26H31ClN4O4S·HCl. The compound is soluble in DMSO and other organic solvents. It is stable when stored at -20°C. However, detailed pharmacokinetic parameters such as absorption, distribution, metabolism, excretion, half-life, and bioavailability are not extensively reported in the available literature.

Comprehensive toxicology data for BMS-199264 hydrochloride are not extensively reported. The compound is classified as a research-use-only chemical and is not intended for human consumption. As an inhibitor of F1F0 ATP hydrolase, it may have effects on mitochondrial function that could contribute to toxicity at high concentrations. Specific toxicological data, including acute toxicity, genotoxicity, and target organ effects, are not reported in the available literature.

[1]. Pharmacological profile of the selective mitochondrial F1F0 ATP hydrolase inhibitor BMS-199264 in myocardial ischemia. Cardiovasc Ther. 2008 Winter;26(4):287-96.

The mechanism by which small organic compounds like BMS-199264 selectively block the activity of hydrolases is unclear. This blocking effect exhibits stereoselectivity, suggesting a specific "lock-and-key" mechanism, thus resulting in high selectivity. Since IF-1 is only active under ischemic conditions, the selectivity of hydrolases is not an issue for this protein. Currently, the only explanation for the selective action of BMS-199264 is the hypothesis that the conversion from ATP synthase activity to hydrolases is not merely a reverse chemical reaction, but requires a conformational change, as proposed by Vinogradov. BMS-199264 may (at least theoretically) bind only to the conformation of F1F0 ATPase in its hydrolases mode. Of course, this is only speculation and requires further investigation. The effect of BMS-199264 on F1F0 ATPase function may also be secondary to its interaction with important pathways regulating F1F0 ATPase and even IF-1 function; this possibility cannot be ruled out. From theory to practice, there are many drugs that can selectively inhibit the function of F1F0 ATP hydrolase. Before going into detail, it should be noted that there is currently no pharmacokinetic data for BMS-199264, so its oral bioavailability is unknown. BMS-199264 appears to penetrate cell membranes readily and may cross the inner mitochondrial membrane. In the heart, such drugs can be used to treat severe ischemia with significant inhibition of oxidative phosphorylation. This is not the case in chronic stable angina, but it is not the case in severe myocardial infarction. Therefore, treatment must be given as adjunctive therapy in the early stages of myocardial infarction, before and/or during surgical intervention. It is conceivable to add such inhibitors to cardioplegic solutions during surgery. Incorporating them into storage solutions for transplanted organs is also a possibility. [1]

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BMS-199264 hydrochloride is a research-grade compound not approved for clinical use. Its primary application is as a pharmacological tool for studying the role of mitochondrial F1F0 ATP hydrolase in ischemia-reperfusion injury. The compound is used to investigate the mechanisms of ATP depletion during ischemia and to validate F1F0 ATP hydrolase as a therapeutic target for the treatment of cardiac ischemia and other conditions involving mitochondrial dysfunction.

C26H32CL2N4O4S

567.527683258057

566.152

C, 55.03; H, 5.68; Cl, 12.49; N, 9.87; O, 11.28; S, 5.65

186180-83-6

675833-20-2;186180-83-6 (HCl);

70202986

Typically exists as White to off-white solid at room temperature

3

7

6

37

841

2

CC1([C@H]([C@@H](C2=C(O1)C=CC(=C2)S(=O)(=O)N3CCCCC3)N(CC4=NC=CN4)C5=CC=C(C=C5)Cl)O)C.Cl

CKNXQCNQMADQCI-KGQXAQPSSA-N

InChI=1S/C26H31ClN4O4S.ClH/c1-26(2)25(32)24(31(17-23-28-12-13-29-23)19-8-6-18(27)7-9-19)21-16-20(10-11-22(21)35-26)36(33,34)30-14-4-3-5-15-30/h6-13,16,24-25,32H,3-5,14-15,17H2,1-2H3,(H,28,29)1H/t24-,25+/m1./s1

(3S,4R)-4-[(4-Chlorophenyl)(1H-imidazol-2-ylmethyl)amino]-3,4-dihydro-2,2-dimethyl-6-(1-piperidinylsulfonyl)-2H-1-Benzopyran-3-ol hydrochloride

BMS-199264; BMS199264; BMS-199264 (hydrochloride); (3S,4R)-4-[4-chloro-N-(1H-imidazol-2-ylmethyl)anilino]-2,2-dimethyl-6-piperidin-1-ylsulfonyl-3,4-dihydrochromen-3-ol;hydrochloride; SCHEMBL7836701; BMS 199264; BMS-199264 hydrochloride

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.)