IL40359A - 17-hydroxy-3-oxo-cyclopropa(6,7)17alpha-pregn-4-en-21-carboxylic acid and its salts - Google Patents

Description

n' 'o iKan 21 †K-4- JATS^ a 17 7,6/ 17-Hydroxy-3-oxo-cyclopropa 6,7 .l7Q:* pregn-4-en 21-carboxylic acid and its salts G.D. SEARLB & CO. 6 , 7-DIHYDRO-17-HYDROXY-3-OXO-3 ' H-CYCL0PR0PA[6 , 7 ]- 17ot-PREGN-lJ-ENE-21-CARBOXYLIC ACIDS AND SALTS This invention relates to 6 ,7-dihydro-17-hydroxy-3-0X0-3' H-cyclopropa[ 6,7 21-carboxylic acids and their salts, and .to preparations thereof. More particularly, this invention provides new, useful and unobvious chemical compounds of the formula CjH2CH2COOR wherein R represents hydrogen, an alkali or alkaline earth metal, or the ammonium radical, and the wavy lines indicate the presence of the a or 3 isomer.

The compounds of this invention are useful by reason of their valuable pharmacological properties. Thus, for example, they are unexpectedly potent and selective diuretics remarkably adapted to reverse the effect of aldosterone on the renal electrolyte ratio without producing the progestational side-effect which characterizes superficially-related prior art.

The potent diuretic utility of the instant compounds is evident from the results of an assay for their capacity to reduce the sodium-retaining effect of d-aldosterone , thereby reversing the renal electrolyte ratio, in dogs.. A group of 6 female mongrels weighing between 15 and 22 kg. is used in this assay, and compounds are tested at 2 dose1 levels in each animal, as also is a reference standard, spironolactone. Controls are provided by twice subjecting each animal to the test procedure without administering either compound or standard. Further controls are provided by likewise repeating the procedure without administering compound, standard, or aldosterone. The order in which the tests are carried out in the 6 test animals is randomized, and the tests are spaced at least 1 week apart to permit the animals to recover from the effect of the aldosterone previously administered. Spironolactone dosages are 2 and 8 mg./kg. (mpk.). Compound dosages are those which, on the basis of a preliminary assay in rats such as that described in U.S. 3 i ^ 2 , 096 , would be expected to produce responses roughly equivalent to those produced by the standard. In the event that any dose is insufficient to reduce the effect of the aldosterone, the assay is repeated at appropriately higher dose levels. The dogs are fed a regular diet at 4 : 00 p.m. on the day before each test and thereafter permitted tap water only (ad libitum) until the test is completed. Testing begins with the intramuscular Injection in a hind limb of 0.2 ml. of sesame oil in which is dissolved 10 yg. of d-aldosterone if such is to be administered. The bladder is catheterized 30 minutes later and then rinsed with 0 ml. of distilled water, whereupon compound or standard, if either, is orally administered in a gelatin capsule and 10 ml. /kg. of tap water is then immediately administered via stomach tube. Any urine excreted during the next 2 1/2 hr. is collected and combined with urine which, at the end of this time and following administration of 5 ml. /kg. of water as before, is removed from the bladder by catheterization.

Excreted urine, if any, is again collected for 2 1/2 hr. and combined with urine removed from the bladder by catheterization. The two urine specimens from each animal are analyzed for sodium and potassium; and the totals thereof are expressed as micro-equivalents per kg. Comparison of the mean log (Na x 10/ ) ratios ± 95? confidence limits determined from these totals serves as the index of antl-aldosterone potency in this assay.

When potassium 6a,7a-dihydro-17-hydroxy-3-oxo-3'H-cyclopropa[6 ,7]-17a-pregn-4-ene-21-carboxylate, the representative product of Example 1 hereinafter referred to as compound (A), was assayed by the foregoing procedure, the results were as shown in Table I.

- - TABLE I Compound Dose Mean Aldosterone Compound Log (Na x 10/K) none (controls) 0 0 1.272 ± 0.105 none (controls) 10 yg. 0 0.528 ± 0.065 spironolactone ' 10 yg. 2 mpk. 0.900 ± Ο.ΙβΟ spironolactone 10 yg. 8 mpk. 1.127 ± 0.210 compound (A) 10 ug. 1 mpk. 1.012 ± 0.205 compound (A) 10 yg. 4 mpk. 1.248 + 0.170 It follows from the tabulated data that the anti-aldosterone response to the product of Example 1 is 304¾ of that to spironolactone in the described assay, fiducial limits being 233-434$. Reversal of aldosterone effect was statistically significant for the compound and standard (P < 0.05 by paired t-test, using each dog as its own control).

Tests for progestational activity can be carried out as described by Clauberg in C. Zentr . Gynakol . , 54 2757 (1930), modified as follows: To each of a group of 3-4 immature, female rabbits weighing about 1 kg. and primed with 173-estradiol by subcutaneously injecting yg. thereof per animal on each of 6 successive days, the test compound dissolved or suspended in corn oil is administered subcutaneously or bucally on each of 5 successive days beginning the next day after the last priming injection. Commonly, the initial daily dosage is 1 mg. of compound in 0.1 ml. of corn oil administered subcutaneously. Other animals likewise administered corn oil alone serve as controls. On the 6th day after the last priming injection, the animals are sacrificed; and a segment of each uterus is taken for histological examination whereby the degree of arborization of the endometrial glands is graded in accordance with a method similar to that described by McPhail in J . Physiol. , 83 » 1^5 (193^)· A +1 response represents the effect of estrogen priming alone and is indicated by the absence of glandular proliferation. A +2 response is induced in the estrogen-primed animals by approximately 0.05 mg. of progesterone administered subcutaneously , and is considered to represent minimal progestational activity. Responses in the range +3 to reflect potent progestational effects typical of those produced by larger doses (»0.1 mg. ) of subcutaneously administered progesterone. If the average rating for the test animals is less than +2 at the 1 mg. dose level, the compound is considered inactive. If the average rating at this dose is greater than +2, the test is repeated with compound administered in amounts decreased according to the progression 0.5, 0.2, 0.1, 0.05, 0.02, 0.01 . . . mg. until a dose insufficient to produce an average response of +2 can be determined. Potency of the compound in percent, relative to progesterone, is then calculated by dividing the next higher dose into 0.05 and multiplying by 100. Results of such tests on the product of Example 1, compound (A), are set forth in Table II. The compound was administered - - subcutaneously in corn oil.

TABLE II Compound Dose Animals Response (mg. ) (no.) (average) compound (A) 1 i» .1.5 This data shows that compounds of the instant invention, represented by the product of Example 1, do not produce the progestational side-effect in the described assay.

Those skilled in the art will recognize that observations of activity in standardized tests for particular biological effects are fundamental to the development of valuable new drug products, both veterinary and human.

The salts of this invention can be prepared by heating alcoholic solutions of 6 ,7-dihydro-17-hydroxy-3-oxo-3 ' H-cyclopropa [6,7 ]-17a-pregn-1*-ene-21-carboxylic acid γ-lactones with appropriate bases, for example, lithium hydroxide, sodium hydroxide, potassium hydroxide, barium hydroxide, strontium hydroxide, calcium hydroxide, or the like. The acids of the inven-tion can be prepared by precipitating aqueous solutions of the salts with excess hydrochloric acid. As an exception to the foregoing salt preparations, the corresponding ammonium salts are prepared by prolonged contact of the acids with saturated alcohol solutions of ammonia.

The following examples describe in detail compounds illustrative of the present invention and methods for their preparation. Throughout the examples temperatures are given in degrees Centigrade (C°.). and relative amounts of materials are indicated in parts by. weight, except as otherwise noted.

EXAMPLE 1 To a mixture of 100 parts of 6cx,7a-dihydro-17-hydroxy-3-oxo-3'H-cyclopropa[6 ^.l-^a-pregn-!l-ene-21-carboxylic acid γ-lactone and 6Ο parts of 2-propanol was added a solution of 15 parts of potassium hydroxide in 100 parts of water.. The resultant mixture was heated at the boiling point under reflux in a nitrogen atmosphere for 1 hour while solution occurred. Solvent was thereupon stripped by vacuum distillation, and the residue was cooled and slurried with ethyl acetate. The cyrstalline material isolated from the slurry by filtration was potassium 6a,7a-dihydro-17-hydroxy-3-oxo-3,H-cycl0propa[6,7]-17a-pregn-1i-ene-21-carb0xylate which, · dried in air, melts at 248-250-?-C. with decomposition.

EXAMPLE 2 "' Substitution of 100 parts of 68 ,7B-dihydro-17-hydroxy-3-oxo-31 H-cyclopropa[ 6 , 7 ]-17a-pregn-4-ene-21-carboxylic acid γ-lactone for the 6a,7a-epimer called for in Example 1 afforded, by the procedure there detailed, potassium 68,78-dihydro-17-hydroxy-3-oxo-3'H-cyclo-propa[6,†]-17ci-pregn-4-ene-21-carboxylate/.m.p. 177-179 °C. with decompositi EXAMPLE 3 To a mixture of 100 parts of 6a,7a-dihydro-17-hydroxy-3-oxo-3'H-cyclopropa[6,7]-17a-pregn-iJ-ene- 21-carboxylic acid γ-lactone and 96Ο parts of 2-pro-panol was added a solution of 11 parts of sodium hydroxide in 100 parts of water. The resultant mixture was heated at the boiling point under reflux in a nitrogen atmos-phere for 1 hour, whereupon solvent was stripped by vacuum distillation. The residue was cooled and slurried with ethyl acetate. The insoluble crystalline product isolated from the slurry by filtration was sodium 6a , 7a-dihydro-17-hydroxy-3-oxo-3 ' H-cyclopropaC 6 , 7 ]-17a-pregn-^-ene-21-carbox late.

EXAMPLE l\ A mixture of 10 parts of 6a,7a-dihydro-17-hydroxy-3-oxo-3 ' H-cyclopropa[6 , 7 ]-17a-pregn-4-ene— 21-carboxylic acid γ-lactone, 2 parts of calcium hydroxide, and JJOO parts of 2-propanol was stirred and heated at the boiling point under reflux in a nitrogen atmosphere for 4 hours. The resultant mixture was stripped -of solvent by vacuum distillation, and the residue was slurried with ethyl acetate. The slurry was filtered.

The crystalline product thus isolated was calcium 6a, 7a-dihydro-17-hydroxy-3-oxo-3 ' H-cyclopropa[6 , 7 ]-17a-pregn-Jj-ene-21-carboxylate .

EXAMPLE 5 To a solution of 10 parts of potassium 6a, 7a-dihydro-17-hydroxy-3-oxo-3'H-cyclopropa[6,73-17a-pregn-4-ene-21-carboxylate in 1000 parts of water was added 18 parts of 5 % hydrochloric acid. The colorless preci- pitate thrown down was filtered off, washed with water, and dried in air. The product thus isolated was 6a, 7a-dihydro-17-hydroxy-3-oxo-3' H-cyclopropa[6 ,7]-17cx-pregn-4-ene-21-carboxylic acid.

EXAMPLE 6 To 20 parts of 2-propanol saturated with ammonia was added 1 part of 6a,7a-dihydro-17-hydroxy-3-OXO-3 ' H-cyclopropa[6,7]-17a-pregn- -ene-21-carboxylic acid. The resultant mixture was allowed to stand at room temperature for 24 hours at which point solvent was removed by vacuum distillation. The residue was washed with ethyl acetate and dried in air. The product thus isolated was ammonium 6 ,7a-dihydro-17-hydroxy-3-oxo-31 H-cyclopropa[6 ,7]-17a-pregn-4-ene-21-carboxylate ·

Claims (4)

WHAT WE CLAIM IS:

1. Compounds of the general formula wherein R is a hydrogen atom, or an alkali or alkaline earth metal, or an ammonium radical; and the wavy lines indicate the presence of the a or $ isomer.

2. Compounds of the general formula OR wherein R is an alkali metal. 40359/2

3. Potassium 6a,7a-dihydro-17-hydroxy-3-oxo- 3·' H-cyclopropa[6 ,7 ]-17x-pregn-ii-ene-21-carboxylate .

4. Potassium 6β,. 7g-dihydro-17-hydroxy-3-oxo- 3 Ή-cyclopropa [6 , 7] -17a-pregn-4-ene-21-carboxylate . HE :mr