IL31690A - Piperazine derivatives having activity against certain forms of cancer - Google Patents

Description

PIPERAZINE DERIVATIVES HAVING AOTI AGAINST CERTAIN FORMS OF CANCER rrno ¾> This invention relates to pharmaceutical compounds and compositions and is primarily concerned with substances having activity in relation to certain forms of cancer, including leukaemia, and certain non-malignant forms of proliferative disease.

It has been discovered that certain chemical compounds, some of which are. already known and have been described as possessing properties unconnected with biological chemistry, display a significant degree of activity against tumours and other -.forms of cancer whilst being of relatival^ low toxicity. These compounds are characterised' by a chemical structure which. d,jes not appear to have been previously explored in biological research, and which is the (3,5*3 ',5 '-tetra-oxo)-l,2-dipiperazino-alkane structure of formula: - wherein R^ and R^ are each separately selected from hydrogen and methyl an ethylene groups or together constitute a-iertur^ed-^-y^eeea^ft-brid ing group e«« Preferred compounds in the series are those in which Rj and R2 both represent either hydrogen or methyl, subject to the above proviso in the latter case, or in which R^ is hydrogen and R2 is methyl.

The present invention comprises compounds of the above formula and their use in compositions intended for mammalian administration arid containing or associated with a physiologically acceptable diluent or carrier.

Compounds having the structure defined above include or are related to poly-N-certain of the/di-acetic acid imides specifically described in U.K.

Specification 978,724 in which an extensive class of compounds-hevi»g~¾he-ffl»»e~^d-poly-dir-e!€«½:i^^i4 proposed as levelling agents, intermediates, textile auxiliaries, and curing agents. It must be emphasised, however, that the possession of anti-cancer properties is by no means coextensive with the possession of the non-biological utilities specifications, are totally devoid of the biological properties under consideration. It is also to be noted that the anti-malignant activity exhibited by the compounds defined herein ''contrasts with the compiete absence of activity in ethylene diamine tetra-acetic acid itself and simple derivatives thereof such as the lower alkyl esters.

The active compounds, which may be utilised if desired in the form of salts with physiologically acceptable inorganic or organic acids, may be formulated for use as pharmaceuticals by a variety of methods. For instance, they may be applied as aqueous, oily (e.g. as a suspension in isopropyl myristate), or in some cases. emulsified compositions for parenteral administration and therefore preferably sterile and pyrogen-free. In general, they have rather low solubility in aqueous media and are therefore usually administered in the form of aqueous suspensions containing suitable surface active agents. Without commitment to a rigid definition of dosages it may be stated that a daily dosage of active constituent, divided if necessary, of from about.J200 milligrams to about 3 grams, but preferably of from about 500 milligrams to about 3 grams, is proposed for mammalian. use applied as a solution in about 500 - 1000 mis. of liquid for intravenous injection by slow infusion, or as a solution orrsuspension in about 10 mis. by the intramuscular route, or in small volumes subcutaneously. It will be appreciated that it may be possible to prepare a more concentrated solution when the active constituent is utilised in the form of a salt. than when the neutral form is used. The substances may also be compounded for oral administration in similar dosages or even as high as 1 to 3 grams daily in the presence of conventional solid carrier-.materials such as starch, lactose, dextrin, and magnesium stearate »or as aerosols or cachets. For the treatment of local forms of the disease, suitable creams or drops may be prepared containing the active substances suppositories and other formulations may also be employed.

When the compounds are to be formulated as salts, preferred formulations are prepared with methane sulphonic acid, isethionic acid, tartaric acid aqueous solutions, after adjustment to physiologically acceptable pH with buffers, are stable for extended, periods of■■: tinie. ■· Solutions of ; siitiilar : w/v strength, i.e. 0.5¾re also obtainable with hydrochloric acid.

Although the ethane analogue cannot exist in more than one stereoisomeric form the other analogues can do so, for example the cyclobutane analogue can. exist in cis and trans forms. The propane analogue can exist as the racemic dl mixture or in the form of the individual d and 1 stereoisomers.

The d and 1 compounds individually, when each is obtained substantially free from the other and from the": racemate, show no substantial difference in biological activity from that observed for the racemic dl mixture. The individual stereoisomers do, -.¾owever,jppssess an advantage over the mixture in that their individual aqueous solubilities, which are substantially similar, are appreciably greater than the aqueous solubility of the mixture. Thus, a similar -solution of the dl compound contains only about 0. 1 This means, for example, that the neutral compound can.be administered parenterally in a much smaller volume if given in the form of either of the individual stereoisomers. In the case of the butane analogue, however, a difference in biological activity is observed between two stereoisomeric forms. Thus the biological activity of meso-( 3? 5, 3 ' , 5 l-te;tra-oxo)-2, 3-dipipera.ginobutane is substantially greater than that of the dl form.

The invention is illustrated in the following Examples: - Example 1 1, 2-Diaminoethane tetra-acetic acid (900 g.) and formtmide ( 2.5 litres) are heated together under reduced pressure (ca 100 mm. ) at 110 - 120°C for 75 minuteSi The temperature is then raised to 155 - l60°C and heating continued for a total of 5 hours (complete solution is obtained after ca 35 minutes at.155 - l60°C and the product begins to precipitate out about 20 - 30 minutes later when frothing may become a temporary problem). The reaction mixture is allowed to cool to room temperature and then stood^ in the (b.p. 60 - 80°C), and drying in vacuo at 60°C, gives ( 3,5,3 ',5 '-tetra-oxo)- l,2-dipiperazino-ethane ( 664 g. 85i¾) as .colourless needles, m.p. 297 - 300°C (dec).

Example 2 1, 2-Diaminopropane tetra-acetic acid ( 100 g.) and formamide (400 ml.), are heated together at reduced pressure under nitrogen at 100 - 110°C for 1 hour, and then at 150 - 155 'C, for 4 hours. The brown solution is evaporated under reduced pressure at 80 - 90°C- and the residue taken up in methanol ( 120 ml.). and cooled in the refrigerator overnight. Filtration, followed by washing with cold methanol and vacuum drying at 65°C gives dl-(3,5,3',5,-tetra-oxo)-l,2-dipiperdzinopropane (62 g. 70%) as a'very pale cream microcrystalline solid, m.p. 237 - 239°C.

Example 3 d-1, 2-Diaminopropane tetra-acetic acid monohydrate, j ip j + 47-1° (c, 0.5% in water)-prepared essentially by the method of. Dwyer and Garvan, J. Amer. Chem. Soc , 1959, 8l, · 2956, is reacted with formamide by the method described in Example 2 to give a 43 yield of d-(3,5,3 ',5 '-tetra- oxo)-l,2-dipiperazino-e¾.e-«n¾ + 11.35°(c 5% in dimethyl-formamide) , m, p. 193 C after recrystallisation from aqueous methanol/ether.

Example 4 (loc. cit.), is reacted with formamide by the method described in Example 2 propane to give a 36% yield of l-(3, 5,3**5 '-tetra-oxoj-l, 2-dipipera«ino-ethaste- 10.9° (c 5% in dimethyl-formamide), m.p. 193-194°C. after recrystallisation from aqueous methanol/ether.

Example" 5 of hydrochloride meso-2,3-Diaminobutane ttrhydTOcfehjridg of diacetyl derivative, 300 - 301°C ] is prepared by lithium aluminium hydride reduction in ether during four days of the dibenzyl ether of dimethyl , and isolated or the dihydrochloride. Jhe dihydrochloride glyoximey ana/is converted m 32% yield, essentially by the method, described meso-2, 3-diaininobutane tetra-acetic acid dihydrate, m.p. 149°C (dec). meso-2, 3-Diaminobutane tetra-acetic acid dihydrate is reacted with f ormamide , essentially tjy the method described in Example 2 but diluting the final reaction mixture with acetone rather than concentrating it, to give a - 356 yield of meso-( . S.3 ' . '-tetra-oxa}-2.3-dipipera¾inQ Joutane , m.p. 320°C (dec).

Example 6 trans - 1, 2-Diaminocyclobutane tetra-acetic acid monohydrate, m.p. 234-235°C (dec), prepared in 56$ yield essentially by the method of Dwyer and Garvan (loc. cit.), is reacted with f ormamide according to the procedure followed in Example 2 to give a 69$ yield of trans . - (3,5)3 ,,5'-tetra-oxO')-l}2-dipipera-5inocyclobutane, m.p. 257 - 259°C (dec).

Example 7 carbamoyl , Ν,Ν,Ν f,N '-Tetrae«#be«affl£Qe'methyl-l, 2-diamino^e thane (0.5 g., prepared by the method of Badinard et al. ,.rBull'S0c.. ^Chim. .France, 1960,-382) is added to polyphosphoric acid ^prepared by heating phosphorus pentoxide (5 g.) and orthophosphoric acid (5 ml.) . at 120°C for two hours, and allowing to cool to 25 C and the mixture is heated at 105 C for ten minutes and then at l20°C for a further thirty minutes* The viscous brow solution is cooled, treated with ice and neutralised by the addition of 0.880 aqueous ammonia (ca 10 ml.) when (3,5,3 ,,5'-tetra-oxo)-l,2-di¾jd.peraaino-ethane (0.33 g. 65$) is precipitated as a very palevj>ink microcrystalline solid, m.p. 298 - 300°C.

Example 8 carbamoyl N,N,N N'-Tetrae3rt«xam-^methyl-l,2-diajninopropane [0.5 g. prepared essentially by the method of Badinard et al., (loc. cit.)~[ and phenol (l0 g.) are heated together under nitrogen at l65°C for twenty hours. The phenol is removed by evaporation under reduced pressure and the residue triturated with methanol, cooled in ice and filtered to give dl-(3,5,3 ',5'-tetra-oxo-^-l,2-dipiperazinopropane (0.44g., 82$) m.p. 235-236°C.

Example 9 8 the same. product as obtained in ^sample 1.

Example 10 Tablets, of the following composition are rprepared: Product GoH^Hiftdr of Example" 1 (micronised) 250 'Avieel ' (microcrystalline cellulose) 38 polyvinylpyrrolidone 3 alginic acid 6 magnesium stearate 3 The product prepared in Example 1 is mixed with the 'Avice¾ and polyvinyl pyrrolidone is added dissolved in sufficient industrial methylated spirits 74°0P to produce a mass suitable for granulating. The ( The British mesh standard is used throughout the specif i'cat ion mass is granulated through a 20 mesh sieve And the resultant granules are dried at a temperature not exceeding 50°C. The dried granules are passed through a 20 mesh sieve and the alginic acid and magnesium stearate are then added and mixed with the granules . The product is compressed into tablets, each weighing 300 mg. , on 3/8 in. flat bevelled edge divided punches.

Example 11 Tablets of the following composition are - repared: - 1 mg. /tablet Product ■eempcHHid of Example 2 250 'Avieel ' (microcrystalline cellulose) 134 polyvinylpyrrolidone 4 alginic acid 8 magnesium stearate 4 The tablets are prepared by essentially the same procedure as described in Example 10, and are compressed at a tablet weight of 400 mg. on 7/l6 in. flat bevelled edge punches .

Example 12 Tablets of the following composition are\ prepared: r mg. /tablet \4 maize starch 15 gelatine 10 magnesium stearate 6 of The tablets are prepared by mixing the product jwoekieed- in Example 1 adding to the mass a 5^ solution of gelatine in water. The productfcis granulated through a l6 mesh sieve, and the resultant granules are dried to constant weight at a temperature not exceeding 60°C. The dried granules are passed through a 20 meteh sieve and.mixed with magnesium starate and the remainder of the maize starch. The product is compressed at 300 nig. tablet weight on 3/8 in. flat bevelled edge divided punches.

Example 13 product The e©H^e«a4 of Example 2 (0.548 .) is added to 5 ml. of a 0.4N solution of (l) hydrochloric acid, (2) isethionic acid, (3) methane' sulphonic acid, or (4) tartaric acid, and the mixture is made up to 100 ml. by the addition of water. The solution so obtained is added to an. aqueous solution of disodium hydrogen phosphate (0.2M, 10 ml.) to provide a stable solution having a pH of 5·7· Injection of the solution withii¾ one hour of preparation is recommended.

Example 14 A brief description of results obtained in experimental animals with two particular compounds will nofv be given. (i) Mice having implanted tumours Sarcoma 180 and Adenocarcinoma 755» and rats having the implantedstumour .Walker 256, are treated with the product . impound of Example 1. The compound is administered by intraperitoneal injection of a suspension of the substance in isotonic saline containing, w/v 2$fcarboxymethyl cellulose. The mice are dosed with 30 mg. of active substance per kilo daily for five out of eight days, and their tumours are excised and weighed. The rats are given one dose only of 400 mg. per kilo and maintained without further treatment for 8 days when their tumours are also removed and weighed. Compared with control animals Mice implanted with leukaemia LtSl-lO are treated as described above with 30 mg. of active substance per kilo for a continuing period until all the animals succumb. Compared with controls, an. increase in survival time of the treated animals greater tha 100% is obtained. . . product . (ii) The eofflpeHftd- of Example 2 was. fed at 30 mg./Kg. on five out for mean tumour weight of eight days to mice having Sl80 tumour and gives a T/c/of 7%.

All the animals survived and they had a weight gain of 12% against 29% in the controls. Against L1210 intraperitoneal and subcutaneous injection for survival time at 30 mg./Kg. daily until death gave a l/c if 257 and 223% respectively; given orally at 75 mg./Kg. a.T/C of 157% was obtained.

Example 15 A brief description of results obtained in a clinical trial of dl-(3, 5j 3 ' j 5'-tetra-oxo)-l,2-dipiperazinopropane will now be given.

A boy aged 11 with a histologically proven diagnosis of lymphosarcoma resistant to radiotherapy, 6-mercaptopurine, methotrexate, prednisone, cyclophosphamide, vincristine and rubidomycin was treated with 2.5 g- of dl-(3 j 5j3 S 5'-tetra-oxo)-l,2-dipiperazinopropane during one day. The compound was administered orally in the neutral form as tablets prepared substantially according to the method of Example 12. The treatment reduced the total of white cells, all of which were primitive, from 42,000 per c.mm to 8, 500 per c.mm within 48 hours. An acute attack of gout which developed at this time, was immediately responsive to allopurinol. On the following day the total white cell count was less than 1,000 per c.mm. and remained at this level for one week before rising again, though it then included no immature cells. There had by this time been a striking clinical improvement in as much as the patlert now felt energetic and well. He remained well for one month, after which the total white cell count began to rise further, the count consisting mainly of blast cells. On treatment with a further 1 g. of dl-(3i 5j3 '5 '-tetra-oxo)-l, 2-dipiperazinopropane given over two days the total white cell count fell from 16, 300 per c.mm to 3>800 per c.mm with only 20 per cent, of immature cells. The patient's condition remained again, began to rise with the appearance of: 'many: blasts and immature cells. A further 1.5 g. of dl-(3,5,3S5'-tetra-oxo)-l,2-dipipera-5inopropane was given during two days and this once more reduced the total-rwhite cell count and the percentage of primitive cells. After five weeks without further treatment the patient's blood picture remained essentially unchanged with approximately 1 per cent of immature or blast cells, the haemoglobin, slightly below normal, an. adequate number of platelets and neutrophils and a total white cell count of about 2,000 per cmm. The patient's general clinical condition was excellent three months after commencing treatment with the drug even though his ; bone marrow still showed evidence of leukaemic infiltration.

Claims (1)

1. insufficientOCRQuality