PUNAGLANDINS AND PHARMACEUTICAL USE THEREOF

Abstract: Novel punaglandins [5-(2,3-diacetoxy-6-methoxycarbonyl hexylidene)-2-chloro-4-)2,5-octadienyl)-4-hydroxy-2-cyclopentenone, or 5-(2,3-diacetoxy-6-methoxycarbonylhexy-lidene)-2-chloro-4-(2-octenyl)- 4-hydroxy-2-cyclopentenone] are isolated from a marine animal, Telesto riisei. These novel punaglandins have a pharmaceutical activity for treatment of tumors.

Description: PUNAGLANDINS AND PHARMACEUTICAL USE THEREOF * BACKGROUND OF THE INVENTION The present invention relates to punaglandins and the pharmaceutical use thereof, and more specifically relates to the novel compounds of punaglandins which are physiologically active substances derived from marine fauna and are useful as antitumor activity, and relates to the pharmaceutical use thereof. DESCRIPTION OF THE PRIOR ART Prostaglandins are natural substances which have a wide variety of pharmacological activities such as blood platelet aggregation inhibitory action, blood pressure lowering action, etc. and accordingly are valuable compounds to be used as an effective remedy for peripheral circulatory failure in recent medical treatment. Of the prostaglandins, prostaglandins A are known ' for having a double bond in the cyclόpentane ring in their molecules. For instance, prostaglan- din 2 is known as a compound having a function to lower the blood pressure (Amer. J. Med. Sci., 263, 335 (1972)). In recent years, prostaglandins A are expected to possibly have a function as antitumor drugs because of an admitted fact that prostaglandins A strongly suppress the formation of DNA (Biochem. Biophys. Res. Commun., vol. 87, p. 795 (1979)). Apart from the aforementioned prostaglandins, punaglandin 1 and punaglandin 2, which have a struc¬ ture similar to the above-mentioned prostaglandins A, are extracted and isolated from Telesto riisei, a marine animal which grows attached to a solid surface. They are expected to be physiologically active sub¬ stances of marine origin, and have their structure already established and identified (Gekkan Yakuji, or Monthly Drugs and Medical Instruments, vol. 24, No. 6, p. 37-43 (1982)). However, nothing has yet been disclosed concretely as to what pharmacological activ¬ ities these punaglandins have. SUMMARY OF THE INVENTION A primary object of the present invention is to provide novel punaglandins different, from puna¬ glandin 1 or 2. Another object of this invention is • to provide the pharmaceutical use of the punaglandins, especially their use as an antitumor drug. A further object of this invention is to provide novel punaglandins which are superior in antitumor activity to the hitherto known prostaglandins A. Other objects and advantages of this invention will become apparent from the following detailed description. These objects and advantages of this invention are achieved by the punaglandins which are expressed by the following formula (I): wherein R, represents a hydrogen atom, a C-,-C, 0 alkyl group, or one equivalent, of cation; R 2 , R 3 , and R 4 are identical or different and each represents a hydrogen atom or a C 2~ c χ o ac yl group; and the symbol - ^ π ^ r- indicates a single bond or a double bond. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS R, in the formula (I) represents a hydrogen atom, a C,-C 1Q alkyl group, or one equivalent, of cation. The C,-C, Q alkyl groups are linear or branched alkyl groups having 1 to 10 carbon atoms such as methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec- butyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, n-nonyl,and n-decyl. Examples of one equivalent of cations are ammon- 4+ lum cations such as NH , tetramethylammonium, mono- methylammonium, dimethylammonium, trimethyl-ammonium, benzylammonium, phenethylammonium, morpholium cation, monoethanolammonium, and piperidium cation, alkali metal cations such as Na and K ; and divalent or tπvalent metallic cations such as 1/2 Ca 2+, 1/2 Mg2.+, 1/2 Zn 2+ , and 1/3 A£ 3+ . R 2 , R 3 , and R 4 are identical or different and each represents a hydrogen atom or a c 2 " c ιo acyl group. Examples of the C 2 -C-, 0 acyl groups are acetyl, propionyl, n-butyryl, iso-butyryl, n-valeryl, iso- valeryl, caproyl, enanthoyl and benzoyl. R 2 and R 3 are preferably an acetyl group, and , is preferably a hydrogen atom. The symbol in the formula (I) represents a single or a double bond. In the aforementioned formula (I), if the symbol ........ is a double bond, the punaglandins of the present invention are punaglandins 3, and if the symbol is a single bond, the punaglandins of the present invention are punaglandins 4. Specific examples of the compounds of formula (I) are given below. (100) punaglandin 3 [5-(2,3-diacetoxy-6- methoxycarbonylhexylidene)-2-chloro-4-(2,5- octadienyl)-4-hydroxy-2-cyclopentenone] (102) punaglandin 4 [5-(2,3-diacetoxy-6- ethoxycarbonylhexylidene)-2-chloro-4- (2-octenyl)-4-hydroxy-2-cyclopentenone] (104) 5-(2,3-diacetoxy-6-carboxy- hexylidene ) -2-chloro-4- ( 2 , 5-octadienyl ) - 4-hydroxy-2 -cyclopentenone (106 ) 5- (2 , 3-diacetoxy-6-carboxy- hexylidene)-2-chloro-4-(2-octenyl)-4- hydroxy-2-cyclopentenone (108) 5-(2,3-diacetoxy-6-ethoxycarbonyl- hexylidene)-2-chloro-4-(2,5-octadienyl)- 4-hydroxy-2-cyclopentenone (110) 5-(2,3-diacetoxy-6-ethoxycarbonyl- hexylidene)-2-chloro-4-(2-octenyl)-4- hydroxy-2-cyclopentenone (112) 5-(2,3-diacetoxy-6-propoxycarbonyl- hexylidene)-2-chloro-4-(2,5-octadienyl)- 4-hydroxy-2-cyclopentene (114) 5-(2,3-diacetoxy-6-propoxycarbonyl- hexylidene)-2-chloro-4-(2-octenyl)-4- hydroxy-2-cyclopentenone (116) sodium salt of (104) (118) sodium salt of (106) (120) aluminum salt of (104) (122) aluminum salt of (106) The punaglandins of the present invention can be prepared according to the process mentioned below. The punaglandins expressed by the formula (I), wherein R-. is a methyl group, R 2 and R 3 are both acetyl groups, R 4 is a hydrogen atom, and the symbol represents a single or double bond (the compound is punaglandin 3 when the symbol is a double bond and the compound is punaglandin 4 when the symbol is a single bond), are extracted and isolated from a marine animal called Telesto riisei as follows.. The marine animal, Telesto riisei, is subjected to an extraction in a Soxhlet extractor first by use of hexane and then a mixture of methanol and water (methanol : water = 9 : 1) as the extracting reagents. In this extraction, about 5.7 g of an extract is obtained from about 360 g of freeze-dried Telesto riisei. About 2.4 g of aqueous methanolic residue is obtained from partition of this extract between the extraction conducted by use of hexane and the succeeding extraction conducted by use of a mixture of methanol and water (9 : 1). About 2.4 g of this extract is subjected . to column chromatography on silica gel using ethyl acetate - hexane (4 : 6) as an eluent to yield two fractions which exhibit the ultraviolet absorption. Of these two fractions, the fraction which is eluted earlier is further subjected to high pressure liquid chromatography by use of a mixture of water and methanol (3 : 7) as a developer to obtain firstly an eluate * of punaglandin 3 and then an eluate of puna¬ glandin 4. The respective eluates are concentrated to give desired punaglandin 3 and punaglandin 4. The punaglandins of the aforementioned formula (I), where R, is a hydrogen atom, a C.,-C, 0 alkyl group other than methyl group, or one equivalent of cation; R 2 and R 3 are respectively a hydrogen atom or a c 2 ~ ιo ac Y 1 group other than acetyl groups; and R 4 is a C 2 ""C- IQ acyl group, can be obtained by subjecting the above-mentioned punaglandin compounds (punaglandin 3 and punaglandin 4) obtained by extrac¬ tion and isolation to the generally known hydrolysis reaction, esterification reaction, or salt-forming reaction, or a combination thereof. As the hydrolysis reaction, the hydrolysis reac¬ tion to be carried out in methanol or ethanol in the presence of sodium methoxide, potassium methoxide, sodium methoxide, or the like and the hydrolysis reaction to be conducted in an aqueous solution of sodium hydroxide, potassium hydroxide, or the like may be mentioned. Alternatively, enzymatic hydrolysis can also be applied to this case. As the esterification reaction generally adopted, there is a method in which punaglandins are allowed to react with such an alcohol compound as methanol, ethanol, propanol, butanol, etc., in the presence of such an acidic compound as sulfuric acid, hydrochloric acid, p-toluene sulfonic acid, pyridine-p-toluene sulfonic acid salt (PPTS), etc., or another method, in which punaglandins are made to react with such an acyl halide compound as acetyl chloride, propionyl chlor¬ ide, etc. As the salt-forming reaction, for instance, the neutralization reaction conducted by use of such a basic compound as sodium hydroxide, potassium hydrox¬ ide, ammonia, etc. may be mentioned. The punaglandins provided by the present inven¬ tion especially display a strong effect of suppressing the proliferation of cells even at low concentration when used against L1210 leukemia cells, thus amply proving that they are very useful as the antitumor drugs. The punaglandins of this invention can be admin¬ istered to a patient orally, or parenterally through percutaneous, subcutaneous, intramuscular, intra¬ venous, intrarectal, for example. Preparations for oral administration may be pre¬ pared, for instance, in the form of tablets, pills, granules, powders, solutions, suspensions, and capsules. In preparing the preparations in the form of tablets, tablets can be made according in the usual manner by use of such excipients as lactose, starch, calcium carbonate, crystalline cellulose, and silicic acid; such binders as carboxymethyl cellulose, methyl cellulose, potassium phosphate, and polyvinyl pyrro- lidone; such disintegrators as sodium alginate, sodium hydrogencarbonate, sodium lauryl sulfate, and ono- glyceride stearate; such moisturizers as glycerin, etc. ; such absorbents as kaolin, and colloidal silica; and such lubricants as refined talc and powdered boric acid. Preparations in the form of pills, powders and granules can also be prepared respectively according to the ordinary methods by use of the excipients, etc. mentioned above. Preparations in the form of solutions and suspen¬ sions can be prepared according to the ordinary method by use of, for instance, such glycerol esters as tricaprylin, triacetin, etc., purified water; and such alcohols as ethanol, etc. Preparations in the form of capsules can be prepared by filling gelatin capsules, etc. with granules, powders, or solutions prepared in the above. Preparations for percutaneous administration may be prepared, for instance, in the form of ointments and creams. Ointments can be prepared by use of such fatty oils as castor oil, olive oil, and vaseline, and creams by use of fatty oils and such emulsifying agents as diethylene glycol, sorbitan monofatty acid ester, etc. according to the ordinary methods, respectively. As preparations for subcutaneous, intramascular, or intravenous administration, there are injections formulated into solutions and suspensions. In the preparation of solutions and suspensions, propylene glycol, polyethylene glycol, olive oil, and ethyl oleate, for instance, are usually used and some anti¬ septics, stabilizers, etc. are added thereto, if necessary. Injections can be sterilized by filtration through the bacterial filter and by addition of a • bactericide thereto. For rectal administration, ordinary suppositories formulated into soft gelatin capsules is used. The punaglandins which are the active ingredients of the present invention can also be contained in the preparations as their inclusion compounds formed with any of α-, β-, γ-cyclic dextrins or their methylated cyclic dextrins. The effective dose of the punaglandins provided with this invention varies with the age, sex, and conditions of a patient; however, it is ordinarily, in the range of 10 2-2x105 μg/kg/day, preferably in the range of 5x10 2-104 μg/kg/day. As described in detail in the above, the present invention provides the novel compounds of punaglandins which are useful as antitumor drugs by themselves, and antitumor preparations which contain punaglandins as an active ingredient. The following examples are " given to illustrate the present invention in more detail. Example 1 360 g of a freeze-dried marine animal Telesto riisei was subjected to an extraction in a Sexhlet extractor first by use of hexane and then a mixture of methanol and water (methanol : water = 9 : 1) as the extracting reagents and 5.7 g of an extract was obtained. In the course of this extraction, 2.4 g of aqueous methanolic residue was obtained from parti¬ tion of this extract between hexane and a mixture of methanol and water. This extract thus obtained was chromatographed on a column of silica gel using a mixture of ethyl acetate and hexane (4 : 6) as an eluent. Of the eluates eluted from the column, two fractions which exhibited the ultraviolet absorp¬ tion were collected. Of these two fractions, the fraction which was eluted earlier was further subjected to C-18 reversed- phase high-pressure liquid chromatography by use of a mixture of water and methanol (3 : 7) to obtain punaglandin 3 from the first eluate and punaglandin 4 from the succeeding eluate. Physical properties of punaglandin 3: MW: 496 C 25 H 33 C 0 8 ( α D =+66 - 8 °' CO.54, MeOH) UV: λ ™ __ v = 238 nro (ε=8600, MeOH) IR(CHC£ 3 , cm "1 ): 3100-3600(broad), 2960, 2940, 1725, 1680(shoulder), 1378, 1235(broad), 990-1110(broad) 'HNMR(360MH 2 , in CDCJ£ 3 , δppm): 7.27(1H, H.^, s), 6.35(1H, H ? , d, J=9.1, 4.3), 5.53(1H, H 15 ), 5.41(1H, H 18 ), 5.35-5.20 (3H, H 14 , .H 1? , H 5 ), 3.65(3H, OMe), 3.01(1H, H 13a , dd, J=8.'l, 14.3), 2.74(2H, H 16 , dd, J=7.1, 7.1), 2.68(1H, H 13b , dd, J=7.0, 14.3), 2.31(2H, H 2 , n), 2.09(3H, OAc , s ) , 2.05(5H, OAc, H 19 ), 1.6-1.75(4H, H 3 , H 4 ), 0.94(3H, H 2Q , t, J=7.5) Physical properties of punaglandin 4: MW: 498 c 25 H 35 -θ 8 ad), 2930, 2860(shoulder), 1725, 1675(shoulder), 1375, 1150-1300(broad), 1000-1100(broad) • HNMR(300MH 2 , in CDCJ- 3 , αpp ) : 7.25(1H, H 11# s), 6.33(1H, H ? , d, J=9.1), 6.01(1H, H & , dd, J=9.1, 4.3), 5.52(1H, H 15 , dt, J=10.9, 6.7), 5.20-5.30 (2H, H 14 , H 5 ), 3.63(3H, OMe, s), 2.95(1H, ≡ 13a , dd, J-9.5, 13.5), 2.62(1H, H 13b , dd, J=7.2, 13.5),2.30(2H, H 2 , m), 2.10(3H, OAc, s), 1.95- 2.05(5H,OAc, H 16 ), 1.60-1.70(4H, H 3 , H 4 ), 1.20- 1.40(6H,H 17 -H 19 ), 0.86(3H, H 2Q , t, J=7.1) Mass(70eV, m/2): 420(M + -Ac0H-H 0), 387(M + -C Q H C ), + + a • -> 378(M -2AC0H), 360(M -2AcOH-H 2 0), 347(M -2ACOH- OMe), 327(M + -C 8 H 15 -AcOH), 285, 267(M + -CgH 15 - 2AcOH),253, 235. Example 2 (i) Determination of antitumor activity in vitro L1210 carcinoma cells were grown in the RPM11640 medium containing 10% fetal calf serum, and the concentration of the cells was adjusted to 1x10 cells/m . Punaglandin 3 and punaglandin 4 were respectively dissolved in 99.5% ethanol. Prior to use, the final concentration of the ethanol solutions was adjusted to less than 0.1%, and it was then added to the respective culture medium. The culture medium was maintained at 37°C in a stationary condition for 4 days. As a control, 0.1% ethanol was used. After the cultivation, the number of surviving cells were measured after dyeing with Trypan Blue. The concentration (IC 5Q ) of punaglandin 3 to inhibit the proliferation of L1210 carcinoma cells was 0.10 μg/mJ-., and IC 5Q of punaglandin 4 was 0.030 μg/m . (ii) Determination of antitumor activity in vivo 5 1x10 Ehrlich ascites carcinoma cells were intraperitoneally administered to ICR mice, and 24 hours thereafter, 5 mg/kg/day of punaglandin 3 and punaglandin 4 were respectively intraperitoneally administered to the mice for 9 days. The period of survival of these animals was observed. The average survival period of the mice treated with punaglandin 3 was 28.6±4.5 days and the increased life span (ILS%) was higher than that of the control by 50.5% and the number of the mice living more than 45 days was 1/6 the treated group. The average survival period of punaglandin 4 was 34.0±4.5 days, the ILS% was higher by 78.9%, and the number of the mice living more than 45 days was 1/6 the treated group. Example 3 Punaglandin 3 was dissolved in fractionated coco¬ nut oil in concentration of 100 mg/m£. Soft capsules were prepared in the usual manner with a soft capsule- making machine in accordance with the below-mentioned recipe, each capsule being made to contain 10 mg of punaglandin 3 as active ingredient. Gelatin 10 parts by weight Glycerin 5 parts by weight Sorbic acid 0.08 parts by weight Purified water 14 parts by weight

Claims: CLAIMS 1. Punaglandins expressed by the following formula ( I ) : wherein R, represents a hydrogen atom, a C,-C-, Q alkyl group, or one equivalent of cation; R_, R 3 , and R 4 are identical or different and each represents a hydrogen atom or a C 2 ~ C 10 ac yl group; and the symbol indicates a single bond or double bond. 2. The punaglandin according to ■ claim 1, wherein in the above-mentioned formula (I) R 2 and R 3 are respec¬ tively an acetyl group and R 4 is a hydrogen atom. 3. A pharmaceutical composition for treatment of malignant tumors comprising punaglandins expressed by the following formula (I) and a pharmaceutically ac¬ ceptable carrier wherein R, represents a hydrogen atom, a alkyl group, or one equivalent of cation; R~, R~, and R 4 are identical or different and each represents a hydrogen atom or a C~-C-. n acyl group; and the symbol indicates a single bond or a double bond. 4. A medicament in unit dosage form comprising the pharmaceutical composition of claim 3. 5. A method for treatment of malignant tumors com¬ prising the administration of punaglandins in pharmaceutically effective amount. 6. The method of claim 5 wherein said pharmaceuti- cally effective amount is m the range of 10 2 to 2xl0 5 μg/kg/day. 3 '.Jι it-9 - Λ j -_ __« H£ET

Inventors: FUKUSHIMA MASANORI, KUROZUMI SEIZI, SCHEUER PAUL J, YU PATRICK T K, FUKUSHIMA, MASANORI, KUROZUMI, SEIZI, SCHEUER, PAUL, J., YU, PATRICK, T., K.
Applicants: HAWAII UNIVERSITY
Application year: 1985
Main classification: C07C177
IPC codes: A61K31, C07C67, C07C401, C07C405, C12P31