CS276046B6 - Polyanhydroglucuronic acid derivatives and process for their preparation - Google Patents

Abstract

Derivatives of polyanhydroglucuronic acid, which contain diammine or amino complexes of platinum or palladium chemically bound to reactive carboxyl or hydroxyl groups of a hydrolyzed glucuronic polymer, the Pc of which is 10 to 200. The reactive groups of this polymer may be partially saturated with a suitable inorganic or organic cation to increase the solubility of the drug. The polymer drug is prepared by controlled hydrolysis of a glucuronic polymer obtained by oxidation of alpha-cellulose with nitrogen oxides, and after optional fractionation of the hydrolysis product, by its reaction with a solution of the corresponding dinitratolatto or dinitratolatto amino complex. The drug is easily resorbable, while the cytostatic activity of the diammine or amino complex of platinum or palladium is not reduced by its binding to the glucuronic polymer.

Description

The invention relates to polyanhydroglucuronic acid derivatives containing diamine or amino complexes of platinum or palladium in the macromolecule and to a process for their preparation.

In the last twenty years, one of the greatest successes of cytostatic research has been the discovery of significant antitumor activity of platinum compounds. However, the cis-diamine-dichloroplatinum complex (cis-DDP, cis-platinum) has been shown to have undesirable side effects, especially GI toxicity and nephrotoxicity, which cannot be fully managed in clinical practice. Efforts to increase the value of the therapeutic index of this type of cytostatics, expressing the ratio of therapeutic effect to toxicity, led to the preparation of other platinum coordination compounds referred to as Pt - so-called II cytostatics. generation, where NH ₃ groups of cis-DDP were replaced by various types of amino ligands, especially cycloaliphatic ones. This group may include, for example, Pt complexes with amino ligands based on cyclohexane, cycloheptane or cyclopentane. Acetates, haloacetates, lactates, glycolates, oxalates, malonates, substituted malonates and the like have been used as leaving anionic ligands (USP 4,322,362, 4,359,425).

Significant disadvantages of some complexes include the relatively low solubility in water, so that it is sometimes necessary to carry out the application in a less suitable suspension form, while reducing their effect. Although an increase in solubility can be achieved for complexes containing a polyfunctional acid as an acid ligand, such as trimellitic acid, using sodium bicarbonate solution, this complex is not very stable and the acid is liberated from the complex due to ionization of all carboxyls.

A common disadvantage of the above-mentioned cytostatics is, in addition to hematoxicity, also the overall strong load on the organism during their application. Due to the relatively small molecules of these drugs, their relatively free random passage through cell membranes through uncontrolled diffusion, rapid glomerular filtration and a rapid decrease in plasma cytostatic concentrations occur.

Efforts to further reduce toxicity, increase the therapeutic index, prevent rapid penetration of biological barriers, and thereby achieve uniform and controlled release of the active drug form have led to the binding of the low molecular weight cytostatite to various types of polymers. It is known that in some cases these efforts are unsuccessful, because the binding of a Pt cytostatic to, for example, some natural or synthetic polymers, proteins, DNA or some polyamino acids reduces toxicity but also reduces the disappearance of cytostatic activity.

These disadvantages of the hitherto known cytostatics containing diamine or amino complexes of platinum or palladium are largely eliminated by the polyanhydroglucuronic acid derivatives of the general formula I,

where

X represents, independently of one another, hydrogen, a diamine or amino complex of platinum or palladium, the amino ligand of which consists of - two molecules of ammonia or an aliphatic amine with a branched or linear solution of 1 to 6 carbon atoms, or

a cycloaliphatic 1,2-diamine containing 4 to 7 carbon atoms in the ring, or

a branched or linear aliphatic diamine of 1 to 6 carbon atoms, or

- a diamine of formula II,

wherein and R ₂ are linear or branched alkyls or cycloalkyls of 3 to 7 carbon atoms, a sodium, calcium, potassium, lithium or magnesium cation, or an organic, for example alkylammonium, alkanolammonium, pyridinium or hexamethylenetetraammonium cation, n is a small integer from the range of 10 to 200, containing 1 to 33.50% by weight of platinum or 1 to 21.65% by weight of palladium.

Said platinum and palladium contents in the polymer correspond to the degree of substitution of the reactive groups of the hydrolyzed glucuronic polymer containing the full theoretical amount of carboxyl groups, in the case of platinum 0.03 to 1 and in the case of palladium 0.046 to 1.

The invention also relates to a process for the preparation of the compounds according to item 1 by oxidation of alpha-cellulose with a mean degree of polymerization Pc in the range of 200 to 2,400 with nitrogen oxides and subsequent controlled hydrolysis, wherein the glucuronide polymer formed by oxidation of cellulose has a mean degree of uron group content 3 to 25.5% by weight, and degree of order up to 90%. The essence of this process according to the invention is that the prepared crude glucuronic polymer is, after optional fractionation, contacted with a solution of dinitrate-platinum or dinitrate-palladium amino complex and precipitating the product in solution with a water-miscible solvent such as acetone, ethanol or isopropanol. of reactive groups of the polyanhydroglucuronic polymer by an inorganic or organic cation is carried out during the hydrolysis. The corresponding water-soluble salts or hydroxides, such as sodium hydroxide, calcium acetate, potassium bicarbonate, lithium carbonate, ammonium hydroxide or magnesium nitrate, or organic bases, for example hexamethylenetetramine or triethylamine, are used for saturation.

The preparation of a suitable glucuronic polysaccharide as a carrier of cytostatically acting diamine and amino metal complexes can be carried out, for example, according to MS. No. 242 920. This process, using hydrolysis in solutions of salts or bases both organic and inorganic, and fractional precipitation of the produced crude uron polysaccharide guarantees the preparation of a polymer with a very narrow distribution curve of polymerization degree values, defined degree of order and uniform distribution of carboxyl groups. in the molecule. In particular, the amorphous form of the product guarantees uniform degradation in the organism.

A significant advantage of the polymeric drug according to the invention is that the basic hydrolyzed uronic polymer is completely compatible with the organism, even when administered parenterally or orally, which has been confirmed by extensive analyzes and physiological tests.

For compatibility tests, resorption capacity and histological examinations, 8-day-old HAN-Wistar rats and 1 to 2-year-old beagle dogs were used in the application of the basic uronic hydrolyzed polysaccharide. Based on the results, it was found that the hydrolyzed uron polysaccharide does not adversely affect the metabolic functions of the organism. No changes in the red blood count, increased leaching of immature myeloid elements into the peripheral blood or changes in the morphology of circulating leukocytes were observed during hematological examination. During pathological-anatomical and histological examination, the leukocyte response to the presence of hydrolyzed uronic polysaccharide was shown to be small or non-existent. Ultrastructural examination of the liver and kidneys of the examined animals showed that the applied uronic polymer showed neither hepatotoxic nor nephrotoxic effect.

Significant advantages of the polymeric drug of the invention also include the fact that the uronic polymer is in some cases capable of forming a thin continuous film on biological surfaces, such as the endothelium of a tumor vessel, thereby allowing the active cytostatic to be evenly distributed in the tumor.

It has also been shown that the activity of the cytostatic diamine or amino metal complex of metals has not disappeared by binding to the uron polymer, but this polymeric drug has sufficient activity, which is another advantage of the invention.

The advantages of the polymeric cytostatics according to the invention further include that the base polymer is resorbable and that the polymeric drug can be prepared according to the intended application either in water-soluble form or in microdisperse form, for example with a particle size of 0.1 to 10 .mu.m. The size of the average degree of polymerization, the content of carboxyl groups, the content and type of cytostatically active diamine or amino metal complex as well as the degree of uronic polymer can be controlled by solubility requirements of the polymer drug according to the invention, its rate of degradation the drug itself as well as the process for its preparation according to the invention.

Although a polymer with a degree of polymerization in the range of 10 to 200 can be prepared, due to the possible immune response of the organism, which is determined not only by the chemical structure of the macromolecule but also by its size and arrangement, uron polymers according to the invention are preferred. A polymer with a higher degree of polymerization can cause problems by uncontrolled blockage of capillaries or can be metabolized in an uncontrolled manner in lung tissue, as is known from the practice of using other types of polymers, for example fatty acid esters and glycerol.

The coordination of the metal amino complex to the uron polymer causes crosslinking of the uron polymer in some cases. Due to steric hindrance according to the type of amino ligands, not all carboxyl groups need to be blocked by this amino complex. The occupancy of uronic carboxyl groups can be controlled by molar ratios, type of amino ligands, pH of the reaction medium and the like. The free carboxyl groups can then be advantageously used to increase their solubility by occupying them, for example, with Na, NH 4, K cations, but also with some organic cations. As noted above, these cations can be retained in the polymer after hydrolysis, prior to treating the uronic polymer with a suitable type of diamine or amino metal complex. When the above-mentioned cations in the form of salts act on the uronic polymer or its metal coordination complex, it is possible to use salts of acids whose pK value is equal to or greater than the pK value of polyuronic acid or salts of volatile acids, e.g. acetic acid, carbonic acid. sulfurous and the like.

Recently, a methodology for the treatment of tumors called the microembolization technique, which reduces the secondary effects of cytostatics on healthy tissue, has come to the fore. In particular, the polymeric drug according to the invention can be applied to this method in particular in microdisperse form alone or in combination with other microembolizing agents and cytostatics.

The advantage of the polymeric drug according to the invention is that all preparations, even those which are not readily soluble in water, can be converted into a stable solution by dissolving them in isotonic sodium bicarbonate solution.

Other advantages include that an amino complex with already known cytostatic activity and toxicity can be bound to the uronic polysaccharide, and that the complex formed by the uronic polysaccharide and the aminoligand-bearing metal is stable with a virtually unmeasurable hydrolysis half-life, as confirmed by extensive conductivity measurements of aqueous solutions and suspensions of polymeric drugs according to the invention.

Other advantages of the polymeric drug according to the invention include that the glucuronic polysaccharide polymer itself already contains reactive groups capable of binding a cytostatically active metal complex, so that these groups do not have to be laboriously introduced into the polymer, and that free hydroxyl groups and optionally free uronic carboxyl groups Other suitable groups may be attached to the polymer, preferably affecting the properties of the drug, for example its solubility, or the uronic polysaccharide polymer itself may suitably affect the therapeutic effect.

A not insignificant advantage of the medicament according to the invention is the fact that the hydrolytic degradation of the polymer in the organism produces glucose-based substances which are well tolerated by the organism.

Exemplary embodiments

Example 1

Alpha-cellulose No. 1,150 with a proportion of individual fractions in the range of No. 220 to 2,000 is subjected to oxidation in 78% by weight of nitric acid, containing 3.0% by weight, of nitrogen oxides. A crude uron polymer with a carboxyl group content of C8 4.88 meq is obtained. g ¹ and 0.8% by weight of nitrogen, which in the next step is subjected to a controlled hydrolysis in a sodium hydroxide solution according to MS. Author's Certificate No. 242 920 at 2 'c. After hydrolysis, the solid is filtered off and the filtrate is precipitated by adding ethanol after reaching an ethanol concentration in the hydrolyzate of 23% by weight.

The precipitated polymer is filtered off. The filtrate is reprecipitated with ethanol until complete precipitation of the polymer. The polymer obtained by filtration is dissolved in water and dialyzed against redistilled water for 6 days. Then alcohol precipitation, filtration and decantation with isopropanol are performed. The polymer obtained is dried under vacuum.

Analysis: Fc = 16% COOH = 5.22 meq. .g ¹ % N ₂ = 0.015 radiograph - completely amorphous.

Preparation of cis-diamine-dinitrate-platinum complex:

2.84 g of cis-platinum (0.00946 mol) are suspended together with 3.2 g (0.0188 mol) of silver nitrate in 40 ml of water and, after mixing, left overnight until exposed to light. The suspension is then filtered to remove silver chloride and unreacted cis-platinum residues. The volume of the filtrate is adjusted with redistilled water to a total volume of 60 ml.

At the same time, 4.06 g of the prepared sodium salt of the uron polymer are dissolved in 40 ml of redistilled water and the solution is left until the next day. To this polymer solution, 60 ml of the prepared solution of cis-diamine dinitrate-platinum complex are gradually metered in at 20 DEG C. with stirring. The immediately formed precipitate is left in the reaction medium in the absence of light for 12 hours at 5 ‘c. It is then filtered, washed until the reaction with NO 2 is removed with 20% ethanol and decanted with isopropanol. After vacuum drying, 5.55 g of uronic polymer containing diamoplatinum

of that complex. Analysis:% C = 32.21 IR spectrum: 3 mg / 1 g KBr % H = 4.47 3,400 (OH), 3,320 (NH), 1,615 (C = O); % N = 3.45 1,404, 1,056, 790 cm ^-1 % Pt = 25.48

Example 2

The same alpha-cellulose as in Example 1 was oxidized in 59.5% nitric acid containing 3.95% nitrogen oxides. A crude uron polymer with a carboxyl group content of 2.98 meq is obtained. g ¹ , which is subjected to hydrolysis in solution, filtration and fractional precipitation according to the already cited MS. copyright certificate No. 242 920.

Analysis: Pc = 40% COOH = 2.88 meq. g ^-1 % n ₂ = 0.010% aldehyde, groups = O X-ray diffraction pattern CrKd = 2.28530 K: peakD.

11,105,93

12,425,31

15,334,32

17,153,87 • 26,152,59

1.42 g (0.00473 mol) of cis-platinum are converted into cis-diamine dinitrate-platinum complex with 1.6 g (0.0094 mol) of silver nitrate in 20 ml of water with stirring overnight. After separation of unreacted cis-platinum and silver chloride, 30 ml of complex solution are obtained.

0.82 g of the prepared uron polymer is dissolved in 40 ml of water and left to stand overnight. After filtration, 30 ml of a solution of cis-diamine dinitrate-platinum complex are added to the polymer solution.

The resulting dispersion was stirred without access for light for 48 hours and then dialyzed against water for 5 days. The dispersion is filtered off with a frit S3, washed with 2 x 30 ml of ethanol and 2 x 30 ml of acetone. A preparation with a particle size of 2 to 5 .mu.m is obtained.

Analysis: % C = 26.00 IR spectrum: 3 mg / 1 g KBr % H = 3.49 3,420 (OH), 3,310 (NH), 1,628 (C = O) % N = 3.29 1615 (COO ^-); % Pt = 15.6 1,400, 1,059, 780 cm- ¹

Example 3

Alpha-cellulose is oxidized in 65% nitric acid containing nitrogen oxides. A uron polymer with a carboxyl group content of 3.68 meq is obtained. g ^-1 , which is subjected to subsequent hydrolysis in hexamethylenetetramine solution. After separation of the unreacted fraction by filtration and subsequent fractionation with isopropanol, a polymer fraction with the following parameters is obtained:

Analysis:

Pc = 51% COOH = 3.70 meq. g ^-1 % N ₂ = 8.7% aldehyde, groups = 0 X-ray diffraction pattern CrKd = 2.28503 K: j * - peakD.

11,05,98

12,255,38

15,354,32

16,953.92 ml of a solution of the cis diamine dinitrate-platinum complex, prepared as described in Example 2, are added to 20 ml of the polymer solution prepared above with stirring. The resulting reaction mixture was allowed to stand for 2 hours. The dispersion is then filtered through a frit S3. The dispersion is repeated three more times. The isolated polymer is washed with 2 x 30 ml of ethanol and filtered off

CS 276046 B6 6 x 30 ml of anhydrous acetone.

After vacuum drying, an ivory powder of the following composition is obtained:

% C = 23.69 IR spectrum: 3 mg / 1 g KBr% H = 3.75 3,410 (OH), 3,310 (NH), 1,621 (C = O)% N = 6.26 1,405.1 055, 790 cm ^-1 % Pt = 20.15

Example 4

3.46 g of the hydrolysed uronic polymer prepared according to Example 2, but in the amorphous state, are dissolved in 80 ml of water and left to stand for 24 hours. Simultaneously, 1.9 g of cis-1,2-diaminocyclohexanedichloroplatinum complex are weighed into a second flask, which is suspended together with 1.65 g of silver nitrate in 25 ml of water. The mixture was stirred in the absence of light for 24 hours. The unreacted residues of the original complex and the silver chloride formed are then separated by filtration. The solution of the resulting diamino-cyclohexanedinitrate-platinum complex is added with stirring to the previously prepared polymer solution in the absence of light. After stirring for 30 minutes, a white bulky precipitate formed. The mixture is left in the dark for 12 hours. After filtration, the precipitate is dispersed in 100 ml of 25% ethanol and filtered after standing for 3 hours. The dispersion in 25% ethanol is repeated twice more. The separated polymer was washed with 2 x 50 ml of ethanol and decanted with 2 x 50 ml of anhydrous acetone. It is then vacuum dried. 4.15 g of product are obtained. The polymeric drug containing the cis-1,2-diamino-cyclohexaneplatinum complex in the form of a white microdisperse powder with a particle size of 1 to 5 .mu.m contains:

% C = 32.5 IR spectrum: 3 mg / 1 g KBr% H = 4.45 3,420 (OH), 2,980 (CH), 1,725 (C = O);

% N = 2.21. 1615 (COO ^-);

% Pt = 15.6 1 405, 1 050, 780 cm ^-1

1.5 g of the prepared product are dissolved in 100 ml of 1% sodium bicarbonate solution. The solution is precipitated with a 1: 1 by volume mixture of isopropanol and ethanol. The precipitate is filtered off, washed several times with 50% ethanol solution, decanted with isopropanol and dried in vacuo. An aqueous solution with a concentration of 1.5 mg of product was prepared from the product. ml ¹ . The stability of the dissolved complex was monitored by measuring conductivity at 20 'ca 37 ° C. During the 20 days was not practically observed change in the originally measured conductivity of 8.1 mS cm x "} · / 20 DEG C. and 11.5 mS cm x ^1/37 C .

Example 5

Alpha-cellulose is oxidized in 70% by weight of nitric acid containing nitrogen oxide. A crude uron polymer containing 3.98 meq is obtained. g ” ¹ · carboxyl groups. This polymer is subjected to hydrolysis in a solution of triethylamine followed by filtration of the unreacted portion with precipitation of the dissolved polymer with isopropanol.

This is followed by vacuum distillation. After distilling off two-thirds of the original volume of the mixture, the uron polymer is separated by filtration, decanted with anhydrous acetone and dried in vacuo.

Analysis: Fc = 25% COOH = 3.90 meq. g ^-1 % N ₂ = 0.2% aldehyde, groups = 0

2.5 g of the polymer obtained are dispersed in 100 ml of a solution of the cis-diamine dinitrate-palladium complex (1.32 g) in water in the absence of light. The reaction mixture was stirred at 25 ° C for 24 hours. The polymer is then filtered off, dispersed in 100 ml of 25% ethanol, the mixture is stirred for 2 hours and filtered. On the filter, the polymer is washed with 2 x 50 ml of ethanol, decanted with 2 x 30 ml of isopropanol and dried in vacuo.

CS 276046 B6 *

A preparation is obtained with the composition: - % C = 32.7 % N = 4.45 IR spectrum: 3 mg / 1 g KBr 3,410 (OH), 3,310 (NH), 1,638 (C = O) ' % H = 4.35 % Pd = 17.0 1615 (COO ^-); 1,405, 1,057, 780 cm- ¹

Example 6

Polyanhydroglucuronic acid (4.0 g) obtained by oxidation of cellulose and further treatments according to Example 1 was reacted with cis-di (methylamine) dinitrate-platinum complex (0.2 g) according to the procedure of Example 1. After isolation, a preparation of the determined composition was obtained. :

% C = 33.80 % H = 4.80 % N = 0.22 · % Pt = 2.2 IR spectrum: 1 mg / 1 g KBr 3,415 (OH), 2,907 (CH), 1,725 (C = O), 1,612 (COO ^- ); 1,383, 1,058, 849, 781 cm- ¹

Example 7

Polyanhydrogluorocaric acid (5.0 g) was reacted with di (hexylamine) dinitrate-platinum complex (0.7 g) according to the procedure of Example 1. After isolation, a preparation of the following composition was obtained:

% C = 35.20 % H = 4.30 % N = 0.18 % Pt = 2.90 IR spectrum: 3 mg / 1 g KBr 3,340 (OH), 2,910 (CH), 1,727 (C = O), 1,609 (COO-); 1,458, 1,058, 825, 790 cm- ¹

Example 8

Polyanhydroglucuronic acid (5.0 g) was reacted with ethylenediamine dinitrate-platinum complex (2.6 g) according to the procedure of Example 1. After isolation, a preparation of the following composition was obtained:

% C = 23.70 % H = 4.10 % N = 5.80 % Pt = 23.70 IR spectrum: 3 mg / 1 g KBr 3,390 (OH), 3,316 (NH), 2,905 (CH); 2,829, 2,723, 2,625, 2,377, 1,727, 1,609, 1,458, 1,400, 1,157, 1,058, 976, 835, 792 cm ¹

Example 9

Polyanhydroglucuronic acid (3.0 g) was reacted with N, N'-diethylenediamine dinitrate-platinum complex (1.3 g) according to the procedure of Example 1. After isolation, a product of the composition:

% C = 28.50 % H = 4.15 % N = 4.75 % Pt = 20.60 IR spectrum: 3 mg / 1 g KBr 3,415 (OH), 3,310 (NH), 2,907 (CH), 1,612 (COO); . 1,383, 1,058, 849, 781 cm- ¹

Example 10

Polyanhydroglucuronic acid (3.5 g) was reacted with 2,2-dimethyl-1,3-diaminopropanedinitrate-platinum complex (4.2 g) according to the procedure of Example 1. After isolation, a preparation having the composition was obtained:

% C = 31.40 % H = 4.20 % N = 4.50 % Pt = 29.80 IR spectrum: 3 mg / 1 g KBr 3,431 (OH), 3,305 (NH), 2,894 (CH), 1,618 (COO); 1,404, 1,056, 835, 790 cm- ¹

CS 276046 B6 8

Example 11

Uron polymer containing 15.6 wt. Pt in the form of the cis-1,2-diaminocyclohexane platinum complex prepared according to Example 4 was tested in vitro and in vivo on L 1 210 leukemia cells in comparison with cis-platinum. In vitro comparisons were made in suspension culture and in softagar. % For value reached

The ID 2 of the uron polymer cis-platinum, according to Example 4 of suspension 1, μmol. 1 Pt 0.4 .urnol. 1 Pt '-1' -1 softagar 0,84 ^ urnol. 1 Pt 0.08 μmol. 1 Pt

For in vivo testing, a ^{200 g DBA-2 J} mouse was used. A T / C value of 150% was achieved with uron polymer at a dose of 15 mg of the product of Example 4 per kg of body weight.

Claims (3)

PATENT CLAIMS 1. Polyanhydroglucuronic acid derivatives of general formula I, where X represents, independently of one another, hydrogen, a diamine or amino complex of platinum or palladium, the amino ligand of which consists of - two molecules of ammonia or an aliphatic amine with a branched or linear chain of 1 to 6 carbon atoms, or a cycloaliphatic 1,2-diamine containing 4 to 7 carbon atoms in the ring, or - a branched or linear aliphatic diamine of .1 to 6 carbon atoms, or> - a diamine of formula II, (II) wherein R 1 and R ₂ are linear or branched alkyls or cycloalkyls of 3 to 7 carbon atoms, a sodium, calcium, potassium, lithium or magnesium cation, or an organic, for example alkylammonium, alkanolammonium, pyridinium or hexamethylenetetrammonium cation, n is a small integer in the range of 10 to 200, containing 1 to 33.50% by weight, platinum, or 1 to 21.65% by weight, palladium. 2. A process for the preparation of the compounds according to item 1 by oxidation of alpha-cellulose with a mean polymerization degree Pc in the range of 200 to 2,400 by the action of nitrogen oxides and subsequent controlled hydrolysis, wherein the glucuronic polymer formed by oxidation of alpha-cellulose has a mean polymerization degree uronic groups of 3 to 25.5% by weight, and a degree of order of up to 90%, characterized in that the prepared crude polymer is contacted with a solution of dinitrate-platinum or dinitrate-palladium amino complex after optional fractionation and precipitates the product in solution with a water-miscible solution. a solvent, for example acetone, ethanol, or isopropanol, wherein the saturation of the reactive groups of the polyanhydroglucuron polymer with the inorganic or organic cation is carried out during the hydrolysis. 3. The process of claim 2, wherein the saturation of the reactive groups of the polyanhydroglucuron polymer with an inorganic or organic cation is accomplished by treatment with water-soluble salts or hydroxides such as sodium hydroxide, calcium acetate, potassium bicarbonate, lithium carbonate, ammonium hydroxide or magnesium nitrate, or by the action of organic bases, for example hexamethyltetramine or triethylamine.