Interaction of bumetanide derivatives with hepatocellular bile acid uptake.

The loop diuretic bumetanide is an organic monocarboxylic organic anion assumed to be transported into hepatocytes by a transport system for bile acids. The structural requirements of 22 bumetanide analogues were analyzed for an interaction with bile acid uptake into isolated rat hepatocytes. Whereas bumetanide inhibited the hepatocellular uptake of [14C]cholate to the same degree as its own uptake, derivatization altered affinity and specificity and yielded compounds that selectively inhibited either cholate or taurocholate uptake or uptake of both. No correlation was found between the diuretic potency of bumetanide derivatives, reflecting the affinity to the Na(+)-K(+)-Cl- cotransporter, and their affinity to hepatic bile salt transport. Computer-aided model building combined with the calculation of potential energy maps showed a strictly amphipathic charge separation in bumetanide analogues as in bile acids. Ranking bumetanide compounds by their mean inhibitory concentration values, inhibition constants, and their type of competition, we conclude that at least three binding domains in the proteins are essential for recognition by bile acid transporters, namely two hydrophobic and an anionic side, and that for the anionic binding region a carbonyl atom in the ligands as an electron donor group is sufficient for ligand interaction.

Purification of proteins of the Na/Cl cotransporter from membranes of Ehrlich ascites cells using a bumetanide-sepharose affinity column.

Bumetanide-binding proteins were isolated from membranes of Ehrlich ascites tumor cells by affinity chromatography. An affinity column was constructed with the active moiety of bumetanide as a ligand using 4'-azidobumetanide, a photoactive analogue which inhibits Na/Cl cotransport in Ehrlich cells with high specificity. Covalent binding of the 4'-azidobumetanide with Sepharose was promoted by photolysis. Membranes isolated from Ehrlich cells were solubilized with n-octylglucoside. Solubilized proteins retarded by the affinity column were readily eluted by bumetanide. In reducing gels the major proteins eluted by bumetanide were approximately 76 kDa and 38-39 kDa. There were also two proteins of 32 to 35 kDa eluted in lesser amounts. No proteins retarded by the affinity column were eluted with extensive washing without bumetanide. Furthermore, bumetanide eluted no proteins from a "control" column lacking the specific ligand. Upon rechromatography with bumetanide in solution, bumetanide-eluted proteins were not retarded, but their purity was increased by the retardation of contaminating proteins. Bumetanide-binding protein purified in this manner were characterized further by electrophoresis in nonreducing, nondenaturing gels.

[Budd-Chiari syndrome caused by obstruction of the hepatic veins]. / Syndrome de Budd-Chiari par obstruction des veines sus-hépatiques.

A Budd-Chiari syndrome was observed in a 11 year-old girl who was admitted to hospital for abdominal pain and distension, soon followed by severe shock. X-ray investigations, surgery and autopsy showed partial stenosis of the common portion of the left hepatic veins associated with a complete obstruction of other hepatic veins but without alteration of the inferior vena cava.

Bumetanide--the way to its chemical structure.

The steps in the evolution of sulfamoyl diuretics in current clinical use are outlined. The development was initiated by the chance observation of a clinical side effect of sulfanilamide, which became the first sulfamoyl group (-SO2NH2)-bearing compound used for diuretic treatment of patients. Ensuing chemical synthesis over the past three decades led to the development of three types of sulfamoyl diuretics:carbonic anhydrase inhibitors, thiazides, and the loop or high-ceiling saluretic agents represented by furosemide and bumetanide. The structural relationship of furosemide to sulfanilamide and the thiazide-type diuretics and, for bumetanide, the more specific structure for loop diuretic activity are discussed.

cAMP-stimulated cation cotransport in avian erythrocytes: inhibition by "loop" diuretics.

The effect of a series of diuretically active substituted 3-aminobenzoic acid derivatives and related compounds was investigated on a cyclic AMP-activated Na+-K+ cotransport system in avian erythrocytes. A good correlation between the diuretic potency of this class of "loop" diuretics in the dog and their inhibition of cation cotransport in turkey erythrocytes was found. Selected thiazide-type diuretics were found to be ineffective. The most active compound tested (3-benzylamino-4-phenylthio-5-sulfamoylbenzoic acid) had an effective dose50 of 4.6 x 10(-8) M in the avian system, and was about 5 times more potent than bumetanide and 500 times more potent than furosemide in this regard. The diuretics appear to interact directly with the cation transport system itself, and not with some antecedent step in the sequence of events from intracellular cAMP accumulation to stimulation of transport. The compounds tested did not appear to compete at Na+- or K+-binding sites on the transport system. The similarity in the structure-activity relationship of these agents in the avian erythrocyte and the kidney suggests that the avian erythrocyte may be a useful model for analysis both of the diuretic-sensitive transport system of the mammalian kidney, and of the molecular mechanism of loop diuretic action.

Aminobenzoic acid diuretics. 9. 3,4-Disubstituted 5-acylaminobenzoic acids and related compounds.

A number of 3,4-disubstituted 5-acylamino-, 5-alkylamino-, and 5-ureidobenzoic acids corresponding to previously described 3,4-disubstituted 5-sulfamoylbenzoic acid diuretics were prepared and screened for their diuretic properties in dogs. The tabulated results reveal that several 3,4-disubstituted 5-formamido and 5-acetamidobenzoic acids possess considerable diuretic potency demonstrating that a 5-sulfamoyl or 5-methylsulfonyl substituent is not a necessity for potent diuretic activity of 3,4-disubstituted benzoic acids. 4-Benzoyl-3-benzyloxy-5-formamidobenzoic acid, one of the mo-t potent compounds of the present series, is approximately one-tenth as potent as bumetanide. The dose response and diuretic pattern indicate high-ceiling diuretic activity and suggest a mode of action similar to that of bumetanide.

Aminobenzioc acid diuretics. 8.(2) 3, 4-Disubstituted 5-methylsulfonylbenzioc acids and related compounds.

Various 2, 4- and 3, 4-disubstituted 5-methylsulfonylbenzoic acids were synthesized as methylsulfonyl analogues of previously described 5-sulfamoylbenzoic acid diuretics. The results of the diuretic screening in dogs reveal that substitution of the sulfamoyl group by the spatially and sterically similar methylsulfonyl group does not affect the diuretic pattern but leads generally to somewhat decreased potency. For the highly potent 3-benzylamino-4-phenoxy-5-methylsulfonylbenzoic acid the corresponding 5-methylthio and 5-methylsulfinyl analogs were prepared and found still to exhibit diuretic activity. Internal aldol condensation and subsequent dehydration of 3-benzylamino-and 3-n-butylamino-4-benzoyl-5-methylsulfonylbenzoic acid provided the corresponding inactive 4-alkylamino-6-carboxy-2, 3-dihydro-3-hydroxy-3-phenylbenzo[b]thiophene 1, 1-dioxides and 4-alkylamino-6-carboxy-3-phenyl-benzo[b]thiophene 1,1-dioxides.

Aminobenzoic acid diuretics. 7. 3-Substituted 4-phenyl-, 4-arylcarbonyl-, and 4-arylmethyl-5-sulfamoylbenzoic acids and related compounds.

Various 4-substituted 3-alkylamino-, 3-alkoxy-, 3-alkylthio-, and 3-alkyl-5-sulfamoylbenzoic acids related to known aminobenzoic acid diuretics were synthesized and screened for their diuretic properties in dogs. The tabulated results from a 3-hr test period revealed that generally the diuretic profile and potency could be retained when 3-alkoxy, 3-alkylthio, and 3-phenethyl were substituted for the 3-alkylamino moiety. The high potency of several 3-alkoxy-, 3-alkylthio-, and 3-phenethyl-4-benzoyl-5-sulfamoylbenzoic acids confirmed previous suggestions that the apparent diuretic effect of 4- and 5-alkylamino-6-carboxy-3-phenyl-1,2-benzisothiazole 1,1-dioxides originates from the corresponding 4-benzoyl-5-sulfamoylbenzoic acid derivatives due to an existing equilibrium in plasma. 4-Benzoyl-5-sulfamoyl-3-(3-thenyloxy) benzoic acid (118) is among the most potent benzoic acid diuretics hitherto synthesized and shows significant diuretic activity in dogs at 1 mug/kg. The results obtained with different 3-substituted 4-phenyl-5-sulfamoylbenzoic acids supported the earlier concept regarding the steric influence of the 4-substituent on the diuretic potency of sulfamoylbenzoic acid diuretics.