IE912320A1 - Long-term prophylaxis against diseases caused by viruses or¹by unconventional viruses - Google Patents

Abstract

Sulphated polysaccharides and derivatives thereof are suitable for the long-term prophylaxis of diseases caused by viruses or by unconventional viruses.

Description

HOECHST AKTIENGESELLSCHAFT HOE 90/F 203 Dr. WN/fe Description Long-term prophylaxis against diseases caused by viruses or by unconventional viruses The present invention relates to long-term prophylaxis against diseases caused by viruses or by unconventional viruses. It has already been disclosed that sulfated polysaccharides and derivatives thereof have antiviral activity (DE 3,601,136, EP 0,270,317). It has also been disclosed that dextran sulfate and other sulfated polysaccharides have activity against scrapie (Ehlers, Diringer, J. Gen. Virol. 65/2 (423-428), 1984). There is a report in J. Gen. Virol. 65/8 (1325-1330), 1984 that there is an increase which, although extremely small, is still significant in the incubation time after scrapie infections when dextran sulfate has been administered 10 weeks before the infection.

It has now been found, surprisingly, that administration of sulfated polysaccharides or derivatives thereof prevents onset of a disease caused by viruses or unconventional viruses when the administration of the agent takes place a long interval of time before the infection. Administration of said agents protects the animals from onset of the infection for a long time, the result being that prophylactic administration of the agents resembles a vaccination (pseudovaccination). This is particularly surprising because no such observations have to date been made either with the chemotherapeutic treatment of viral diseases or with the treatment of unconventional viral diseases.

The method, according to the invention, for controlling viral diseases and unconventional viral diseases has the advantage that the compounds which can be used according to the invention are less toxic than other antiviral - 2 agents, and some of them are even virtually non-toxic. Furthermore, it is possible, for the long-term prophylaxis according to the invention, to divide the administrations of the polysaccharides into relatively small doses to be given more or less regularly, which allows unwanted side effects of the polysaccharides to be substantially or completely suppressed.

Accordingly, the present invention relates to the use of sulfated polysaccharides or derivatives thereof for the long-term prophylaxis of diseases caused by viruses or unconventional viruses.

Preferred for the use according to the invention are polysaccharides which have a polysaccharide framework which is composed of uniform or different monomer units selected from the following group of compounds, or comprises the following compounds: xylose, arabinose, rhamnose, fucose, glucose, mannose, galactose, fructose, glucosamine, galactosamine, mannosamine, glucuronic acid, galacturonic acid, mannuronic acid, carrageenans, fucoids, laminaran, alginate, lentinan, pluran, xylan, dextran, heparin, keratan sulfate, chondroitin 4- and 6sulfate, dermatan sulfate, heparin sulfate, hyaluronic acid, teichuronic acid and partial hydrolyzates of starch, cellulose, glycogen, chitin or pectin. Said compounds can be both of synthetic and of natural, i.e. in particular vegetable or microbial, origin.

Furthermore preferred for the use according to the invention are sulfated polysaccharides with lipophilic substituents. By sulfated polysaccharides with lipophilic substituents are meant in this connection particularly sulfated polysaccharides which can be linear or branched, are composed of uniform of different natural or synthetic monomers, which can also contain a replaced or unsubstituted amino group per monomer unit, and whose OH groups are on average between 5 and 80 % replaced by a group of the formula -X-B-Y where X is an oxy group or a group of the formula I (I) whose carbon atom is bonded to B, B is a non-aromatic hydrocarbon radical which has 2 to 30 carbon atoms and in whose alkyl chain up to 3 methylene units can be replaced by oxy groups, in which up to three C-C double bonds can be present and which can be substituted by up to three 0χ-04alkyl radicals and Y can - if X is an oxy group - be hydrogen, COOR or OSO3R1, in which R is a physiologically tolerated singly or doubly charged cation, or a hydrocarbon radical with up to 20 carbon atoms or a mono- or bisether residue with 3 to 10 carbon atoms, R1 is a physiologically tolerated cation, or Y is - if X is a radical of the formula I - hydrogen or COOR in which R has the abovementioned meanings, and where the OH groups of the abovementioned polysaccharides are between 10 and 95 % replaced by a group of the formula OSO3M where M is a physiologically tolerated cation and between 0 and 40 % of the OH groups are not replaced.

In the case of lipophilic polysaccharides whose polysaccharide framework already occurs in nature in sulfated form, it is possible for the sulfated polysaccharides to be used both with their natural sulfate content and after additional sulfation. The polysaccharide frameworks preferred for the lipophilic sulfated polysaccharides are the sulfated polysaccharides already mentioned above.

The OH groups of the lipophilic polysaccharides according to the invention are preferably 5 to 80 %, particularly preferably 5 to 50 % and especially 10 to 40 % replaced - 4 by a group of the formula -X-B-Y and, furthermore, preferably 10 to 95 %, particularly preferably 45 to 95 % and especially 50 to 90 % replaced by a group of the formula OSO3M, where preferably between 0 and 40 %, particularly preferably between 0 and 20 % and especially between 0 and 10 % of the OH groups are not replaced.

Preferred radicals B have 2 to 20 carbon atoms, have one or no double bond and are unsubstituted or monosubstituted by a Ci-C^-alkyl group.

Where X is an oxy group, Y is preferably H, COOR, OSO3R1, where R is an alkali metal cation, especially potassium or sodium or a branched or unbranched Ci-Ce-alkyl group, R1 is preferably an alkali metal cation, especially sodium.

If X is a radical of the formula I, Y is preferably H or a radical of the formula COOR, where R is an alkali metal cation, preferably sodium or potassium, a branched or unbranched Ci-Cg-alkyl group or a mono- or bisether residue with 6 to 9 carbon atoms.

Examples of radicals of the formula -X-B-Y are ethyloxy, propyloxy, isopropyloxy, butyloxy, hexyloxy, octyloxy, decyloxy, dodecyloxy, propionyloxy, butyryloxy, valeryloxy, hexanoyloxy, octanoyloxy, decanoyloxy, dodecanoyloxy, palmitoyloxy, stearoyloxy, 2-methylbutyryloxy, 9-octadecenoyloxy, linoleyloxy, linolenoyloxy, cholesteryloxy, hydroxyoxalyloxy, hydroxymalonyloxy, hydroxysuccinyloxy, hydroxyglutaryloxy, hydroxyadipoyloxy, crotonoyloxy, mesaconoyloxy or the esters of said dicarboxylic acid derivatives with unbranched Cx-C-alcohols or (1,3-diisopropoxy-2-propyloxycarbonyl)propanoyloxy.

The dodecanoyloxy and the stearoyloxy radicals have particular importance, especially on a xylan polysaccharide framework. - 5 The polysaccharides according to the invention in a form which is unsubstituted or has lipophilic substituents may have an average molecular weight which covers a wide range. The substituted polysaccharides preferably have an average molecular weight of up to 100 kD, particularly preferably of up to 40 kD, especially between 1 and 22 kD. Particularly important are substituted polysaccharides according to the invention whose polysaccharide framework is composed of xylan or dextran and which have an average molecular weight of 1 to 22 kD.

Pentosan polysulfate is very particularly preferred (polysulfated 0-(l-4)-D-xylopyranoside with an average molecular weight of 3,500-5,000 D) .

The preparation of the sulfated polysaccharides is state 15 of the art and is described, for example in EP 0,240,098, US 2,715,091, CH 293,566 or Carbohydr. Chem. 2, 298 (1963). The abovementioned sulfated polysaccharides with lipophilic substituents are prepared, for example, in the case of polysaccharides which are substituted by a group XBY where X is an oxy group, by partially etherifying the polysaccharides with an alkyl halide or an ω-halogeno carboxylic acid salt, with basic catalysis, followed, immediately or after alkoxylation, by total or partial sulfation.

If X is a group of the formula I, the polysaccharides are, for example, partially acylated with carboxylic acids or carboxylic acid derivatives, and the remaining OH groups of the polysaccharides are totally or partially sulfated.

Further details of the preparation of lipophilic sulfated polysaccharides are disclosed in German Patent Application P 3,921,761.2, to which express reference is made at this point. The compounds described as preferred in said patent application are also preferred for the present use according to the invention. - 6 Also suitable for the use according to the invention are polyelectrolyte complexes of a polyacid and a polybase. Preferred polyacids are the sulfated polysaccharides already described above, in substituted or unsubstituted form. Particularly preferred sulfated polysaccharides have a degree of sulfation of more than 10 %, particularly preferably 90 to 100 %.

The average molecular weight of the unsubstituted and substituted polysaccharides for the polyelectrolyte complexes should be between 500 and 80,000 D, preferably between 1,000 and 40,000 D, especially between 4,000 and 15,000 D.

The polyelectrolyte complexes according to the invention can be formed with the aid of different polybases. It is possible to employ, inter alia, proteins as polybases, such as, for example, hemoglobin. Further suitable examples are polylysine, lysine alkyl esters, chitosan, poly-(2-N,N-dimethylaminoethyl)-D,L-aspartamide, amino acids, poly(amino acids), aminated oligo- and polysac20 charides (for example aminated dextrans), poly-[a,0(spermidinyl)-D,L-aspartamide], gelatin and derivatives (for example Polygeline*, Behringwerke AG, Marburg), quaternary ammonium compounds (for example Luviquat*, BASF AG, Ludwigshafen or Micrapol A15®, Miranal Chem. Co., Inc. South Brunswick, New Jersey), C12-alkylsternamine, α,ω—bis—(N-propylamino)polyethylene glycol and aminated polyethylene glycol derivatives. It is also possible to use as polybases covalent compounds of polymers containing OH groups (for example dextran) and nitrogen crown ethers, and the metal complexes thereof.

It is additionally possible to optimize the absorbability of the polyelectrolyte complexes according to the invention by varying the substituent on the polyacids and polybases. Thus, for example, it is possible to provide the polyacids with different substituents as already explained above. The polybases can likewise be modified by substitution reactions. Particularly suitable substituents for the polybases are hydrophobizing in nature, such as, for example, alkyl chains, especially branched or unbranched alkyl chains with 10-20 carbon atoms, such as, for example, lysine octadecyl ester or cetyltrimethylammonium bromide. Other suitable substituents are, for example, steroid residues, especially cholesterol bonded via the alcohol group, or the cholesteryl ester of succinic acid or analogous steroids.

It is furthermore possible to employ tailored polyelectrolyte complexes prepared by reaction of a polyacid or a mixture of polyacids with a polybase or a mixture of polybases .

The polyelectrolyte complexes are prepared, for example, by adding either a) the polybase or b) the polyacid dropwise, preferably in aqueous solution at a suitable temperature and at a suitable pH to a) the polyacid or b) the polybase, preferably in aqueous solution.

Further details of the polyelectrolyte complexes are described in German Patent Application P 3,937,283.9, to which express reference is made at this point. The polyelectrolyte complexes which emerged as preferred in said patent application are also preferred for the present use according to the invention.

The compounds which can be used according to the invention are employed for the prophylaxis of diseases caused by viruses or unconventional viruses. They are particularly important for the long-term prophylaxis of diseases caused by herpesviruses or by retroviruses such as, for example, HTLV I and II, HIV, BLV, FLV, FeLV, FIV, VisnaMaedi viruses, goat arthritis viruses, human spumaviruses or by unconventional viruses.

They are very particularly important for the control of retroviral diseases and diseases connected with - 8 unconventional viruses, especially scrapie, bovine spongiform encephalopathy (BSE virus) and CreutzfeldtJakob syndrome.

Long-term prophylaxis means in the present connection 5 that the relevant compounds are given at least one, preferably three, especially five days before a potential infection.

However, it is preferable to administer the relevant agents long before a potential infection after admini10 stration of one or more initial doses at regular intervals in a relatively low dose which has few side effects to the organisms at risk of infection. In this way the relevant organisms are optimally protected against the onset of an infection without unwanted side effects occurring. The compounds can be administered preferably at an interval of 90 days, particularly preferably of 80 days, especially of 60 days. The dosage of the individual administrations is, in terms of amount, below the dose which would be required to control an infection after onset. The preferred initial dose is 0.1-10, particularly preferably 3-9, especially about 4 mg/kg body weight of a mammalian organism. The preferred maintenance dose, which should be administered after a time interval as explained above, is 0.1-5, particularly preferably 0.5-2, especially about 1.5 mg/kg body weight.

The described dosages apply to administration by injection; it may be necessary to increase the doses, for example by 5-10 times, for a different mode of administration.

The compounds which can be used according to the invention can be administered, for example, orally, intravenously, intramuscularly, intraperitoneally, subcutaneously or rectally.

The present invention also relates to pharmaceuticals for the long-term prophylaxis of diseases caused by viruses - 9 or unconventional viruses, which contain compounds which can be used according to the invention. The pharmaceuticals according to the invention are prepared by converting at least one substituted polysaccharide or one of its derivatives, where appropriate with other auxiliaries and/or excipients, into a suitable dosage form. The auxiliaries and excipients are derived from the group comprising vehicles, preservatives and other customary auxiliaries.

For example, it is possible to use for oral dosage forms auxiliaries such as starches, for example potato, corn or wheat starch, cellulose or derivatives thereof, especially microcrystalline cellulose, silica, various sugars such as lactose, magnesium carbonate and/or calcium phosphates. It is furthermore advantageous to add to the oral dosage forms auxiliaries which improve the tolerability of the medicaments, such as, for example, swelling agents and resins. It is also possible for the purpose of improving tolerability to administer the medicaments in the form of enteric capsules. It may furthermore be advantageous to add a retarding agent to the dosage form, where appropriate in the form of permeable membranes, for example those based on cellulose or polystyrene resin, or ion exchangers. It may furthermore be worthwhile to administer the compounds which can be used according to the invention in the form of an implant.

The intention of the exemplary embodiments which follow is to explain the present invention in more detail.

The present invention is explained by the following exemplary embodiments and by the patent claims.

Example 1: Effect of prophylactic administration of pentosan polysulfate (MW about 5,000 dalton) on splenomegaly after - 10 infection with Friend leukemia virus (FLV). groups each of 10 female NMRI mice, body weight 18-20 g, were treated in each case 50, 40 and 30 days before infection by intraperitoneal injection of pentosan polysulfate in physiological saline as follows: Group 1: Group 2: Group 3: Group 4: mg/kg 50 mg/kg 250 mg/kg 500 mg/kg (control) The infection was brought about by intravenous injection of 0.2 ml of a 1:10,000 dilution of serum from FLVinfected NMRI mice. 14 Days after infection, the spleens of the experimental animals were weighed. The spleen weight served as a measurement unit for the replication of FLV in the experimental animals (Chirgos M.A., Luber, E., March, R., Pettigrew, H. (1965) Antiviral chemotherapeutic assay with Friend leukemia virus in mice. Cancer Chemother. Rep. 45, 29-33). The result is summarized in the following table (1): 0 Group Dosage mg/kg Spleen weight ± S.D.

Survivors 1 0 2.54 + 0.37 10 2 50 1.77 ± 0.61 8 3 250 1.67 ± 0.89 4 25 4 500 1.56 ± 0.53 5 Whereas the spleen weight of uninfected mice is about 0.1 g, FLV-infected mice develop splenomegaly with spleen weights above 2 g (Group 1) within 2 weeks.

Pretreatment with pentosan polysulfate resulted in a 30 dose-dependent and statistically significant (p < 0.05) reduction in the spleen weight compared with the untreated control (Group 1). It can be concluded from this that pretreatment of the mice with pentosan polysulfate has influenced subsequent infection with FLV. The deaths - 11 observed among the experimental animals during the experiment occurred as a consequence of the anticoagulant effect of pentosan poly sulfate at high dosage even before the infection.

Example 2 Prophylactic effect of pentosan polysulfate (MW: 5,000 dalton) on scrapie infections in mice.

In 2 independent experiments, groups each of 6-8 mice were pretreated by intraperitoneal administration of, in each case, 10 mg/mouse of pentosan polysulfate on the following days before the infection with the scrapie agent: Experiment 1 - 84 Experiment 2 - 70 15 - 77 - 60 - 71 - 50 Corresponding control groups were treated at the stated times with phosphate-buffered saline.

The infection was brought about by intraperitoneal injection of 0.1 ml of homogenate from scrapie-infected mice in a dilution of Is 100 to 1:1 million. The observation period lasted 384 (Experiment 1) or 307 days (Experiment 2), during which the experimental animals were examined twice a week, each day on onset of the symptoms, for their state of health and possible signs of scrapie infection. The survival time and the number of surviving animals served as a measure of the effect of the prophylaxis with pentosan polysulfate.

The result of the experiment is summarized in the follow30 ing Table (2): - 12 Incubation time (days) and survival rate Infective Experiment 1 Experiment 2 dose Control Treated Control Treated 10-2 15511 23919 16712 22016 0/7 0/6 0/6 1/7 10-3 160+2 248+9 17413 268115* 0/7 1/6 0/7 5/8 10-4 184+5 - 18713 267 + 0/7 6/6 1/7 7/8 10-5 18714 - 18612 - 2/6 6/6 4/7 Ill 10-6 - - 7/7 7/7 *1 -1 n = 3, one animal died prematurely with other symptoms It was possible to show that in both experiments the pretreatment with pentosan polysulfate extended the incubation time compared with the relevant control or, with a lower infective dose, prevented the onset of scrapie.

Further analyses of the animals from Experiment 2 showed that the animals which did not become diseased during the experimental period had no amyloid protein whereas amyloid was detectable in all the controls and the treated diseased mice. Accumulation of amyloid protein in the brain is regarded as an important criterion for the presence of scrapie.

Claims (10)

1. The use of sulfated polysaccharides or derivatives thereof for the long-term prophylaxis of diseases caused by viruses or unconventional viruses. 5

2. The use as claimed in claim 1, wherein the sulfated polysaccharide is composed of or comprises uniform or different monomer units selected from the following group of compounds: xylose, arabinose, rhamnose, fucose, glucose, mannose, galactose, fructose, glucosamine, 10 galactosamine, mannosamine, glucoronic acid, galacturonic acid, mannuronic acid, carrageenans, fucoids, laminaran, alginate, lentinan, pluran, xylan, dextran, heparin, keratan sulfate, chondroitin 4- and 6-sulfate, dermatan sulfate, heparin sulfate, hyaluronic acid, teichuronic 15 acid and partial hydrolyzates of starch, cellulose, glycogen, chitin or pectin.

3. The use as claimed in claim 1, wherein the derivative of the sulfated polysaccharide is a lipophilic sulfated polysaccharide. 20

4. The use as claimed in claim 1, wherein the derivative of the sulfated polysaccharide is a polyelectrolyte complex.

5. The use as claimed in one or more of claims 1-4, wherein the virus is a retrovirus, especially HIV. 25

6. The use as claimed in one or more of claims 1-5, wherein the unconventional virus is the causative agent of scrapie, of Creutzfeldt-Jakob syndrome or of spongiform encephalopathy.

7. The use of sulfated polysaccharides or derivatives 30 thereof for preparing a long-term prophylactic against diseases caused by viruses or by unconventional viruses. - 14

8. A pharmaceutical containing a sulfated polysaccharide or one of its derivatives for the long-term prophylaxis of diseases caused by viruses or by unconventional viruses. 5

9. A method for the long-term prophylaxis of diseases caused by viruses or unconventional viruses, which comprises administering a sulfated polysaccharide or one of its derivatives. - 15

10. A pharmaceutical as claimed in claim 8, hereinbefore described. substantially as