AU647800B2 - Antiviral agents containing heteropolytungstate - Google Patents

Description

OPI DATE 23101/92 AOJP DATE 27/02/92 APPLN. D11 8006'4 91 PCT NUMBER PCT/AU91/00280 1 I NERATIJJ,,,L.~ ,a .t~njUt. it,. n .EATY (PCT) (51 Inerntioal atet Cassfictio International Publication Number: WVO 92/00078 A61K 31/66, 33/42, CGIG 41/02 Al (3 nentoa ulcto ae aur 92(90.2 COIG 49/04, 51/04, C07F 11/00 (3 nentoa ulcto ae aur 92(90.2 C07F 15/02, (21) International Application Number: PCT/AU91/00280 (74) Agents: CORBETT, Terence, Guy et al.; Davies Collison, I Little Collins Street, Melbourne, VIC 3000 (AU).

(22) International Filing Date: 28 June 1991 (28.06.91) (1 eintdSae:AA Erpa aetABB Priority data: (European patent), BF (QAPI patent), BG, BJ (GAP] PK 0917 29 June 1990 (29.06.90) AU patent), BR, CA, CF (OAPI paterit), CG (QAPI patent).

CH, CH (European patent), Cl (OAPI patent), CM (71) Applicants (for all designated States except US): COMMON- (QAPI patent), CS, DE, DE (European patent), DK, WEALTH SCIENTIFIC AND INDUSTRIAL RE- DK (European patent), ES, ES (European patent), Fl, SEARCH ORGANISATION [AU/AU]; Limestone FR (European patent), GA (QAPI patent), GB, G13 (Eu- Avenue, Campbell, ACT 2601 FAI!RIELD 11O ropean patent), ON (QAPI patent), GR (Europe:an pa- P4T-A-Be*fRl3-AU/AU];, -Be--R a-Fmiie~dt tent), HU, IT (European patent), JP, KP, KR, LK, LU, VI 04AJ- LU (European patent), MAC, MG, ML (OAP] patent), F.Q.-MEICALRESA-RGH-LI-MtED1ti7'tT; MR (QAPI patent), MW, NL, NL (European patent).

Yarra Bend Road. FfldAI47..(-4. NO, PL, RO, SD, SE, SE (European patent), SN (OAPI patent), SU, TD (OAPI patent), TG (OAPI patent), US.

(72) Inventors; and Inventors/Applicants (for US an lv) :WEIGOLD, Helmut Published [AU/AU]; 54 Leeds Road, Mt, Waverley, VIC 3149 1,1ith international search report.

NOLAN, George [AU/AU); 86 Were Street, rk\ 1h Brighton, VIC 3186 MARCUCCIO, Sebastian, 0Wb Mario [AU/AU]; 9 Stuart Avenue, Hamptoni Park, VIC 3976 BIRCH, Christopher, John [AU/AU]; I An- 4 derson Street, Surrey Hills, VIC 3127 GUST, Ian, David [AU/AU]; 8 The Boulevard, Ivanhoe, VIC 3079 SEC 113 (54) Title: ANTIVIRAL AGENTS CONTAINING H ETEROPOLYTUNGSTATE Am, ICPTPW 11

O

3 nH.,O [CpTiP 2

WI

7 061]. nH,0 A. [M 4

(H

2 0) 2

(PW

9 0)3) 2 nHO A. [M 4

(H

2 0) 2

(P

2 Wl 5 0 56 2 nHO A. [MAPW 27 0 11 9

H

1 nH 2

O

(57) Abstract Heteropolytungstate compounds of general formulae wherein M is Co, Fe, Zn, FeQA, FeO 1 Cp represents an optionally substituted cyclopentadienyl residue; A is a monovalent or divalent cation or mixture of such cations; m is the number of cations necessary for electrical neutrality of the molecule; or pharmaceutically acceptable derivatives thereof. Pharmaceutical compositions or methods for the treatment or prophylaxis of retrovirus-associated infection which invoplve the use of such compounds.

WO 92/00078 PCF/A U91 /00280 -1- ANTIVIRAL AGENTS CONTAINING HETEROPOLYTUNGSTATE The present invention relates to heteropolytungstates and pharmaceutically acceptable derivatives thereof, pharmaceutical compositions containing them, and to the use of these compounds in therapy, particularly for the treatment or prophylaxis of certain viral infections, for example retroviral infections such as Acquired Immune Deficiency Syndrome (AIDS).

Because of the difficulty of inhibiting viruses while leaving the non-infected cells unimpaired, few antiviral drugs are currently in widespread clinical use. This is especially true of viruses within the family Retroviridae. The causative agent for AIDS, the Human Immunodeficiency Virus, [also known as Human T-cell Lymphotropic Virus III (HLTV-III)] is a member of this family. This virus will be identified herein as HIV. Infection with HIV is associated with depletion of T4 lymphocytes, brain disease, and several types of cancer including Kaposis sarcoma.

Patients infected with the virus also have a high incidence of opportunistic infections and a significantly reduced life span. Another virus within the same family is the Human T-cell Lymphotropic Virus Type 1 (HLTV-1) the causative agent of Adult T-cell leukaemia, an infection with high mortality occurring in a distinctive geographical pattern.

All members of the family Retroviridae possess a unique enzyme, reverse transcriptase, which is necessary for their replication. Because this enzyme is not normally present in uninfected cells, it is considered a target for antiviral drugs.

Another virus utilising reverse transcriptase during replication is the Hepatitis B Virus (HBV). HBV causes widespread morbidity and mortality and is the main cause of primary hepatocellular carcinoma in individuals who are chronic carriers of the virus. Because of the common evolutionary origin of HBV and the retroviruses, treatment of HBV-infected individuals with the same regimen of WO 92/00078 PCT/AU91/00280 drugs proposed herein for the treatment of patients infected with HIV could be possible.

Background Heteropolytungstate compounds have been known for over 100 years. Most of their applications stem from their redox chemistry and also their high ionic weights and charges. Their redox chemistry has led to their most common use: catalysts for the oxidation of organic substrates such as, for example, propylene to acrylic acid, ethylene to acetaldehyde. In the biological field heteropolytungstates have found use as electron dense stains for electron microscopy, as analytical reagents for proteins and a few have also been shown to inhibit viral DNA and RNA polymerases Chermann et al., Biochem. Biophys. Res. Comm., 1975, 1229; M. Herve et al., ibid 1983, 116, 222). The antimony heteropolytungstate

(NH

4 ),g[NaSbgW, 1 (HPA-23), has also been shown to have marginal activity against HIV in permissive cell lines (MT-2, MT-4) however in these cases the cell toxicity is such that the therapeutic index' is low(about 10 or less) Inouye, et al., Chem. Pharm. Bull. 1990, 38, 285). Because few anti-HIV agents were available at the time HPA-23 was tested in clinical trials despite this low therapeutic index Rozenbaum et al, Lancet 1985, 11:450). HPA-23 has since been shown to be inactive in other in vitro screens and in subsequent clinical trials Balzarini et al., Int. J. Cancer, 1986, 37, 451; M. Burgard,et al., AIDS, 1989, 3, 665). The compounds of this invention are in general less toxic and more active than HPA-23 with antiviral indexes >20 and generally >50 and are active in all cell lines tested to date.

A recent report also cited that K,[PTi,W 1 oO40].6HO inhibits the replication of HIV in HTLV-1 carrying MT-4 cells Inouye, et al., Chem. Pharm. Bull. 1990, 38, 285).

Therapeutic index is defined as the ratio of the dose which is toxic to of the test cells to the dose required to reduce the virus count by WO 92/00078 PCT/AU91/00280 -3- The heteropolytungstates of this invention are based on Keggin-type and Dawson-type structures, carrying either one or three vacancies. These vacancies are created by extraction of WO 4 or W 3

O

6 +6 from the PWO2040 3 (Keggin) or

P

2

W

8 ,0 6 2 (Dawson) structure. Isomers of these unsaturated (lacunary) polyanions are possible, a consequence of the location of the vacancy. Massart et al., Inorg. Chem. 1977, 16, 2916; T.L. Jorris et al., J. Am. Chem. Soc. 1987, 109, 7402; T.J.R. Weakly, Polyhedron, 1987, 6, 931; R. Contant et al., J. Chem.

Res.(S), 1977, 222; R.G. Finke et al., Inorg. Chem. 1987, 26, 3886; M.T. Pope, 'Heteropoly and Isopoly Oxometallates', Springer-Verlag, 1983). The position of the vacancy in P2W,70, 6 is defined by the prefix ac for a belt vacancy or ca, for a cap vacancy. The prefix a- or P- refers to isomerisation associated with the relative rotation of the W 3 triad cap. In the trivacant polyanions, PWO4'-, the vacancy is found in either of two locations. In the B-form, an edge-linked W, oxide triad has been lost, whereas in the A- form a corner linked W 3 oxide triad has been removed. The unsaturated heteropolytungstates can behave as polyanion ligands and bond, at their vacant site, with metal ions. The more stable unsaturated polyanions appear to be the c. Contant et al. J. Chem. Res. 1977, 222; T.J.R. Weakley, Polyhedron, 1987, 6, 931) and B-type Knoth, Organometallics 1985, 4, 62) isomers. Transition metal ion complexes of type

[M

4

(PW

9 O4)] 1 are known only with the B-a- isomer of the polyanion ligand. A recent structure determination Weakley et al., Inorg. Chem. 1990, 29, 1235) on Na 14 Cu[Cu 4

(H

2 0) 2 (P2W 1 5 0 6 2 ].53HO has shown that the bonding of the

PW

15 Os 6 12 ligand to the set of four coplanar copper atoms is similar to that found in the corresponding PW0349- containing compounds. In particular, the

PW

1 sO 56 1 2- polyanion is formed from a-PW 8 0 6 2 6 by removal of one edge-sharing cap (containing three tungsten atoms).

It is known that the A-a-PW0g3 9 anion can form complexes with transition metal ions. With Co 2 the species formed h, iCa 3 (H2O) 3

(PW

9 4) 2 1 2 which on heating in aqueous solution is converted to the B-a- structure [Co 4 (HaO),(PWgO)] 1 Knoth, et al., Organometallics 1985,4,62).

WO 92/00078 PCT/AU91/00280 -4- The preparation of the compounds Alo[M 4 (H20),PW034)2].nHO, A K, M Mn(II), Co(II), Ni(II), Cu(II), Zn(II) was described in a note by T.J.R Weaklev et al. J.C.S. Chem. Commun. 1973, 139. A full paper on the synthesis and structure of several of the compounds by these workers appeared much more recently (H.T.

Evans et al. J.C.S. Dalton Trans. 1986, 2699). Alternate preparative procedures, employing the preformed PW90349- polyanion, were developed by R.G. Finke et al. Am. Chem. Soc. 1981, 103, 1587; Inorg. Chem. 1987, 26, 3886). The structure of the compounds, as noted above, is based on the B-a- isomer of the

PW

9 0349- trivacant polyanion. The four transition metal atoms (M 4 are coordinated to and are located between, the two B-a-PW g

O,

9 polyanions.

It has now been discovered that phosphorous containing complex heteropolytungstates derived from Keggin and Dawson type structures carrying either one or three vacancies as described above are inhibitors of HIV infection.

Accordingly the present invention provides a method for the treatment or prophylaxis of a retrovirus-associated infection which involves the use of a heteropolytungstate compound of the general formulae 1 aving a therapeutic index (as hereinbefore defined) of 50 or greater: Am [CpTiPW 1

O,

39 nH20 I(a) Am [CpTiP 2

W

1 7 061]. nH2O I(b) Am [M 4

(H

2 0) 2 (PW0O34) 2 nH.,O I(c) A. [M 4

(H

2 0) 2

(P

2

WO

1 0,) 2 nHO I(d) Am [MgPsW270 1

,H

1 7 nH 2 0 I(e) Formula I; wherein: M is Co, Fe, Zn, FeOA, FeO,,; Cp represents an optionally substituted cyclopentadienyl residue; 'A is a monovalent or divalent cation or mixture of such cations; WO 92/00078 PCT/AU91/00280 m is the number of cations necessary for electrical neutrality of the molecule.

The compounds-of the invention are polyanions with associated cations for electrical neutrality. They crystallise with a variable number of molecules, of water of crystallisation dependent upon the conditions of product recovery and subsequent treatment; all such hydrate forms fall within the scope of this invention.

For monovalent cations, the number of cations for the compounds of Formula I is as follows: Compound m I(a) 4 I(b) 7 I(c) I(d) 16 I(e) 16 If divalent cations, or a mixture of monovalent and divalent cations, are present then a lesser number will be sufficient for electrical neutrality and m will be smaller. Preferred cations are sodium, potassium, lithium, ammonium, alkylammonium, cationic alkali metal crown ether complexes, magnesium, or calcium.

The term "optionally substituted", in relation to the cyclopentadienyl residue, means that the residue may contain one or more substituents. Suitable substituents will be chosen on the basis of the known chemistry of cyclopentadienyl-titanium complexes. In general, substituents will be selected on the basis that: they do not interfere with the n-bonding of the cyclopentadiene ring to the titanium atom; they do not interfere with the formation of the compound of formula I: and they do not have a deleterious effect on the antiviral properties of the compound.

WO 92/00078 PCT/AU91/00280 -6- A wide range of suitable substituents is known to those skilled in the art of cyclopentadienyl compounds. These are discussed, for example, in the series of Gmelin Handbooks on Titanium-Organic compounds, and in papers by Keana, et al. Amer. Chem. Soc. 1985, 107, 6714; ibid 1986, 108, 7951; ibid 1986, 108, 7957; J. Org. Chem. 1987, 52, 2571).

By "a pharmaceutically acceptable derivative" is meant any pharmaceutically acceptable salt, or any other compound which, upon administration to the recipient, is capable of providing (directly or indirectly) a heteropolytungstate of the general formula I, or an antivirally active metabolite or residue thereof.

It will be appreciated that the compounds according to the invention may also be used in the manufacture of a medicament for the treatment or prophylaxis of a retrovirus-associated infection.

In another aspect, the present invention provides a pharmaceutical composition for the treatment or prophylaxis of a retrovirus-associated infection, which comprises an effective amount of a compound of the general formula I, in association with a pharmaceutically acceptable carrier or diluent.

The present invention also extends to a method for the treatment or prophylaxis of a retrovirus associated infection, which comprises administering to a patient in need of such treatment or prophylaxis an effective amount of a compound of the general formula I.

Examples of compounds of Formula 1 which fall within the ambit of the present invention include the following 2 2 "18-C6K" means compounds containing K' complexed by the cyclic polyether 18-Crown-6 WO 92/00078 WO 9200078PCr/AU9I /00280 -7- 1. K 4

[CPTPW

11

O

39 .nH.,O 2. (N(n-C 4

H

9 4 4 [CpTiPW 1

O

39 .nH,O 3. Na 4

[CPTPW

11

O

39 .nH.,O 4. Li 4 [CpTiPW 1

O

39 nH 2

O

5. (NI- 4 4 [CpTiPW 1 0 39 1 .nHO 6. K7, [CPTP 2

W

17 0 6 11 nHO 7. Li 7 [CpTiPWI., 1 .nH,O 8. Na 7

[CPTP-'W

1 7

O

61 1 .nHO 9. (NHMe 3 [CPTiP'Wl 7

O

6 l] .nH,~O 10. (NH 4 7 [CPTiP2W 1 70 61 1 .nHO 11. Kj 0 [Co3 4 (HO0)2 (PW 9 O-1) 1 .22H.,O 12). Lil 0 [Co 4 (H2C)2 (PW 9

O

1 4) 2 .nHO 13. Na 10 [Co 4

(PW

9 O4) 2 .nHO 14. (NH 4 10 [Co 4 (HO)2 (PW 9 O34)21 .nH,O 15. [NH(CH 3 3 10 [Co 4 (HO)2 (PW 9 O4) 2 1 .nHO 16. 31 1 (Co 4

(H

2 10)2 (PW 9 O4) 2 J .nHO 17. [NH 3

(CHS)

10 (Co 4

(PW

9 O4) 2 .nHO 18. Rb 10 [C0 4 (H20)2 (PW 9 O4)2I .nHO 19. CS 10 [Co 4 (H,O0)2 (PW 9 O34) 2 .nH,O 20. (18-C6K~l 0 [Co 4

(H,O)

2

(PW

9 O4)2) .nH.,O 21. Ca 5 [Co 4 (H2O)2 (PW 9 O34)2 riH2O 2.K 10 [Zn 4

(HO)

2

(PW

9 O34) 2 .2211-LO 23. Li 10 [Zn 4

(H

2 0)2 (PW 9 O4) 2 .nHO 24. Rb 10 [Zn 4

(H

2 0)2 (PW 9 O4)2] .nH,O 25. Csl 0 [Zn 4 (H,O)2 (PW 9 0 34 2 1 .nHO 26. Na 10 [Zn 4

(H

1 0)2 (PW 9 O34) 1 .nHO 27. (NH 4 10 [Zn 4 (H,O)2 (PW 9 O34>] .nH,O 28. [NH(C-I 5 3 1 1 0 [Zn 4

(HO)

2

(PW

9

O

34 )2J .nH.,O 29. [NH 3 (CqH 5 10 [Zn 4

(PW

9 O4) 1 .nH.,O 30. (18-C6K) 0 [Zn 4

(HO)

2

(PW

9 O3) 2 31. Ca 5 [Zn 4

(PW

9 04)2] .nH,O 32. Mg4[Zn 4 (H,0) 1

(PW

9 0 34 2 .nH,,O WO 92/00078 WO 9200078PCT/AU91/00280 -8 33. [NH(CH 3 3 1 10 [Zn 4 (H,0) 2

(PW

9 O34) 2 .nH 2

O

34. K 10 [Fe 4 2

(PW

9 O4) 1 .nH,O Lil 0 [Fe 4 (H,0)2 (PW 9 0) 2 .nH.0 36. Na 10 [Fe 4

(H

2 0) 2

(PW

9 O34),] .nHO 37. (NH 4 1 0 [Fe 4 (H,0)2 (PW 9 0-1) 2 .nH,0 37A. ENH(CHI)I] 10 [Fe 4

(H

1 0) 2

(PW

9 034)2,. nH,0 38. (18-C6K) 10 [Fe 4 (H,0) 2

(PW

9 04) 1 .nH.,0 39. K 1 6 [C0 4

(H

2 0) 2

(PW

15 0 5 6 1 iiH*)O

(NH

4 16 [Co 4 (H,0) 2 (P2, 1 0 56 .nH,O 41. Na 1 6 [Co 4

(H

2 0)2 (P 2

W

15 0 6 2 .nH,O 42. Li 16 [Co 4 (H,0)2 (PWI 5 0 6 2 .nH 1

O

43. (NHMe 3 16 [Co 4 (H,0)2 (P2W 15 0 56 .nH.,b 44. Mg 8 [C0 4 (H,0) 2

(P,W

1 5 0 56 2 .nH,)O

K

1 6 [Zn 4

(H

2 0) 2

(P

2

W

15 0 56 2 .nHO 46. (NH 4 16 [Zn 4 (H,0)2 (P 2

W

15 0 56 1 .nH-1 47. Na 16 [Zn 4 (H-10)2 (PW 15 0 56 .nH,O 48. LU 16 [Zn 4

(PW

15 0 56 )21 .nH 1

O

49. (NHMe 31 6 [Zn 4 (H.0) 2

(PWI

5 0 56 2 .nH,O Mg 8 [Zn 4

(H

2 0)2 (PI-W 15 0 56 2 .nHO 68. Mg 8 [Mn 4

(H

2 0) 2 (P2W 15 0 56 2 1 .nH,.O 69. (NH 4 16 [Co 9

P

5 W7 011 H 1 7 ].nH,,O

K

16 [Co 9

P

5

W-

27 0119 H 1 .nH,0 71. Li 1 6 [Co 9

P

5

W

27 0119 H 1 7 ].nHO 72. Na 16 [Co 9 P5W 27 0119 H 1 ].nH 2

O

73 Ca8 [Co 9

P

5

W

2 0119 H 17 1.nH 2 0 121. K 10 [(FeOK) 4

(PW

9 034) 1 .nH, 2

O

122. Na 10 [(FeONa) 4

(PW

9 034),].nH,0 123. Lil 0 [(Fe0Li) 4

(PW

9 03) 2 .nHO 12 4. (NHMe 3 10 [(Fe0(NHMe 3 4

(PW

9 O34) 2 ].nH,0 12-5. Na 16 [Fe 4 (H,0) 2

(PW

5

O

56 2 .nH 2

O

126. Li 16 [Fe 4 (H,0)(P 2

W

5

O

5 6 2 ].nH,0 127. (NH 4 10 [(Fe0NH 4 4

(PW

9 0.U4,].nH 2 0 WOQ 92/00078 PCT/AU9I/00280 -9- 128. Kl,[(FeO,,pD 4

((PW

9

O)

1 .nH,O Of the above listed compounds those numbered 1, 2, 6, 9, 11, 13, 22, 23, 26, 41, 47, 48 and 70 are known.

WO 92/00078 PCT/AU91/00280 10 The invention also provides novel compound according to formula, I and includes the specific compounds listed above, excluding those listed as being known.

The compounds according to the invention, also referred to herein as the "active ingredient(s)", may be administered for therapy by any suitable route, including oral, rectal, nasal, topical (including buccal and sublingual), vaginal and parenteral (including subcutaneous, intramuscular, intravenous and intradermal), Preferably, administration will be by the oral route, however it will be appreciated that the preferred route will vary with the condition and age of the recipient, and t'e nature of the chosen active ingredient.

In general a suitable dose will be in the range of 3.0 to 120 mg per kilogram body weight of the recipient per day, preferably in the range of 6 to 90 mg per kilogram body weight per day and most preferably in the range 15 to 60 mg per kilogram body weight per day. The desired dose is preferably presented as two, three, four, five, six or more sub-doses administered at appropriate intervals throughout the day. These sub-doses may be administered in unit dosage forms, for example, containing 10 to 1500 mg, preferably 20 to 1000 mg, and most preferably 50 to 700 mg of active ingredient per unit dosage form Ideally the active ingredient should be administered to achieve peak plasma concentrations of the active compound of from about 0.1 to about 75 mM, preferably about 2 to 50 mM. This may be achieved, for example, by the intravenous injection of a 0.1 to 5% solution of the active ingredient, optionally in saline, or orally administered as a bolus containing about 0.1 to about 100 mg/kg of the active ingredient. Desirable blood levels may be maintained by a continuous infusion to provide about 0.01 to about 5.0 mg/hour by intermittent infusions containing about 0.4 to about 15 mg/mg of the active ingredient.

While it is possible for the active ingredient to be administered alone, it is preferable to present it as a pharmaceutical composition. The compositions of the present invention comprise at least one compound of general formula together WO 92/00078 PCT/AU91/00280 11 with one or more pharmaceutically acceptably carriers thereof, and optionally other therapeutic agents. Each carrier must be pharmaceutically "acceptable" in the sense of being compatible with the other ingredients of the composition and not injurious to the patient. Compositions include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration. The compositions may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredient with the carrier which constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then if necessary shaping the product.

Compositions of the present invention suitable for oral administration may be presented as discrete units such as capsules, sachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient may also be presented as a bolus, electuary or paste.

A tablet may be made by compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder (e.g inert diluent, preservative disintegrant sodium starch glycollate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose) surface-active or dispersing agent. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for.example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally WO 92/00078 PCT/AU91/00280 12be provided with an enteric coating, to provide release in parts of the gut other than the stomach.

Compositions suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavoured basis, usually sucrose and acacia or tragacanth gum; pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia gum; and mouthwashes comprising the active ingredient in a suitable liquid carrier.

Compositions for rectal administration may be presented as a suppository with a suitable base comprising, for example, cocoa butter or a salicylate.

Compositions suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.

Compositions suitable for parenteral administration include aqueous and non-aqueous isotonic sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the composition isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The compositions may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

Preferred unit dosage compositions are those containing a daily dose or unit, daily sub-dose, as here ,i above recited, or an appropriate fraction thereof, of an active ingredient.

WO 92/00078 PCT/AU91/00280 13 The compounds according to the invention may also be presented for use in the form of veterinary compositions, which may be prepared, for example, by methods that are conventional in the art. Examples of such veterinary compositions include those adapted for: oral administration, external application, for example drenches aqueous or non-aqueous solutions or suspensions); tablets or boluses; powders, granules or pellets for admixture with feed stuffs; pastes for application to the tongue; parenteral administration for example by subcutaneous, intramuscular or intravenous injection, e.g. as a sterile solution or suspension; or (when appropriate) by intramammary injection where a suspension or solution is introduced into the udder via the teat; topical application, e.g. as a cream, ointment or spray applied to the skin; or intravaginally, e.g. as a pessary, cream or foam.

The administered ingredients may also be used in therapy in conjunction with other medicaments or in conjunction with other immune modulating therapy including bone marrow or lymphocyte transplants or medications such as levamisol or thymosin which would increase lymphocyte numbers and/or function as is appropriate.

It should be understood that in addition to the ingredients particularly mentioned above, the compositions of this invention may include other agents conventional in the art having regard to the type of composition in question, for example, those suitable for oral administration may include such further agents as sweeteiirs, thickeners and flavouring agents.

WO 92/00078 PCT/AU91/00280 14 GENERAL METHODS OF PREPARATION 1. Preparation of A, 1

[M

4 (H20)(PWgO 4 2 .nHO Two general methods of preparation of these compounds have been used; that due to H.T. Evans et al. (JCS Dalton Trans, 1986, 2699) or the method of R. G. Finke et al. (Inorg. Chem. 1987, 26, 3886) (All temperatures are in Celsius.) EXAMPLE OF A COMPOUND MADE BY THE EVANS ET AL PROCEDURE Preparation of Kjo[Co 4 (H2O) 2

(PW

9 Oz) 2 1.22H 2

O

6.11g (0.021 mol) Co(NO3) 2 .6H,O was dissolved in 110 mL H,O containing 9.8g (70% w/w) HNO 3 and brought to the boil. A solution of 29.7g (0.090 mol) Na 2

WO

4 .2H,O and 3.94g (0.011 mol) NaHPO 4 .12HO in 110 mL H,O was then slowly added. After about two thirds of this solution had been added to the refluxing cobalt nitrate solution, a precipitate formed which redissolved on continued addition of the solution. After 5h reflux the clear reddish solution was treated with 40g solid KCI. The precipitate formed was collected and washed well with H,O at room temperature. Yield of crude product 16.9g. [The washings gave 7.4g of crude K 16 [Co 9 Ps 5 W270 9

H

7 (see below).] The product was purified by twice dissolving in hot water and precipitating with excess KCI and then recrystallising from water at 850 and standing at room temperature. Yield of air dried product 14.6g blue-purple crystals.

EXAMPLES OF COMPOUNDS MADE BY THE FINKE ET AL PROCEDURE Preparation of Kio[Fe 4 (H20)(PW 9 O34) 2 ].nHO This method involves the reaction of preformed PWgO, 9 polyanion with a stoichiometric amount of metal salt. The PW0g3 9 anion was prepared by the method of R. Massart et al. (Inorg. Chem. 1977,16, 2916) as hydrated WO 92/00078 PC/AU91 /00280 15 A-Na 8 HPWgO which on heat treatment at 1400 is converted at least in part, to the B- form Knoth et al., Inorg. Chem. 1987, 26, 3886). This isomerization reaction is not completely understood. Following Finke et al., the polyanion used in this work was heated at 140-1450 for 1-2h, usually about 12/3 hours.

4.71g(12 mmol) (NH 4 )2Fe(SO 4 2 .6H 2 0 and 17.1g of heat treated Na 8 HPW,034.24H,O were placed in a 250 mL Schlenk flask. After degassing under vacuum and replacement of the air with argon, 120 mL of de-aerated water were added and the reaction mixture refluxed for 5h. Degassed KCI (ca. 30g) was then added and the mixture let stand at room temperature overnight. The brown solid was collected under argon to give a yield of 15.4g of crude product. The compound was redissolved in 100 mL of dc-aerated water at 80-850 under argon and treated with ca. 25g degassed KCI. The product was collected under argon from the cool reaction solution and then redissolved in 100 mL de-aerated water at 80-85° under argon and filtered hot through a G3 frit filter stick. On standing at room temperature, brown crystals formed which were collected under argon and dried under vacuum. Yield of brown crystalline product was 13.8g. It is air sensitive, oxidizing to a yellow material.

Preparation of Ko,[(FeOK) 4 (PWgO) 2 nH,O While the following preparation is typical and gives good yields, it is not mandatory to use preformed A-Na 8

HPW

9 04 19H 2 0 or an iron(III) carboxylate to obtain the product.

A mixture of 31 g (11.25 mmol) of A-NasHPW,034.19H 2 O, prepared following R.

G. Finke, M. Droege and P. J. Domaille, Inorg. Chem. 1987, 26, 3886-3896, and an iron(III) carboxylate (22.5 mmol based on iron) were mixed and 100 mL water added. The mixture was magnetically stired with gentle warming. The reaction mixture was refluxed for 8-12 hours, during which time the solution became somewhat lighter in colour. On addition of 35 g KC1, the solution solidified to a yellow mass which was collected yield 30.6 g) after the reaction mixture had WO 92/00078 PCT/AU91/00280 16cooled to room temp. The product was dissolved in ca. 275-300 mL boiling water and filtered hot through a Whatman No. 540 filter paper. 40g KCI was then added to the heated stirred solution. The compound (24.4 g) was collected at room temp.

It was dissolved again in ca. 250 mL water near boiling point, filtered through a Whatman No. 542 paper, and 5-8 g of KCI added. On cooling to room temp. the product separates and fills the volume of the solution. The compound is collected on a frit and washed with 3 x 25 mL water (at room temp.) and then with ethanol before air drying on the frit. Yield 22.5 g. Storing over silica gel under vacuum for several days resulted in a weight loss of about 0.3 g. The amount of water associated with the product is variable. Analyses by ICP indicated an atomic ratio of K 7 FePW 9 suggesting that the compound may have a formula of type

K,

4 [(FeO) 4 (PW90O4)2]. nH,O. Titration of the protonated form of the compound, prepared by elution through a column of "Amberlite" resin, type IR-120 in the H' form, with KOH, suggests that the compound is perhaps better formulated as Kio[(FeOK) 4

(PW

9 O34)2.nH20, since about 4 of the 14 protons are neutralized only when the pH of the solution exceeds The infra-red spectrum of the compound in KBr is characterized by a triplet between 1040-1080 cm 1 (phosphate stretches), and strong bands at 947, 873, 809 and 724 presumably arising from W-O stretches and also perhaps Fe-O stretching vibrations which would be expected to lie within this region (see D. M.

Kurtz, Jr. in Chem. Rev. 1990, 90, 585-606 The Na and NH 4 compounds can be obtained as long, thin, neadle-like crystals, greenish in color. Air drying, especially in the case of the Na compound, causes the crystals to collapse. The Na, Li and NH 4 salts are more readily soluble in water than either the corresponding K or NMe 3 H salt.

Compounds in which the cation is other than potassium can be obtained by ion exchange or addition of a large excess of the required salt (usually the chloride) instead of the KCI used in the examples above. Organic solvent (usually ethanol) was added to solutions of the very soluble compounds to effect their isolation.

Those compounds containing K* complexed by the cyclic polyether 18-Crown-6 WO 92/00078 PCT/AU91/00280 17 were made by addition of a slight excess of the ether to an aqueous slurry of the pure K 0 ioM 4

(H

2

O),(PW

9 04)2].nH 2 O compound. The isolated compound was then washed with aqueous ethanol.

EXAMPLE OF PREPARATION OF COMPOUND BY CATION EXCHANGE Preparation of (NH,) 16 [CoP s W,7O 11 9

H

1 7.nHO. The potassium salt of this compound is obtained as a side product in the synthesis of Klo[Co 4 (H0O) 2 (PW,034) 2 by the method of H.T. Evans et al Isolation of the crude K salt was as described in the preparation of Kio[Co 4 (H,O),(PW9034) 2 The product (7.4g) was dissolved in warm water (ca. 70mL) to give a red solution which was filtered and then treated with 16g

NH

4 CI. Pink product (5.3g) was obtained after standing 16h at 50. This step was repeated to give 5.1g of product which was dissolved in 20ml of water at 65-70°, filtered and kept at 50 for several days. The product was collected and air dried; yield 2.0g. More compound can be obtained by reducing the solvent or on addition of ethanol.

PREPARATION OF PWIsO 6 1 2

ANION

The compounds were prepared by the procedure of R.G. Finke et al.

a-KP 2

W

1 sO 6 2 was obtained by the slow addition of a near theoretical amount of a 1M solution of KHCO 3 to at,-KP 2

W,

8 062, reacidifying the reaction solution and precipitating with KC1. The potassium salt was converted to the sodium one using NaCIO 4 and the trivacant polyanion salt Na,, 2 PWs 5 0 56 .18HO was obtained on raising the reaction solution pH to 9 with NaCO 3 as described by R.G. Finke et al.

WO 92/00078 PCT/AU91/00280 18 EXAMPLE OF COMPLEXING OF POLYANION WITH A DIVALENT TRANSITION METAL Preparation of Na 16

[M

4

(HO)

2

(P

2

W

1 5 0 5 6 )2].nHO. The compound in which M Co was prepared by adding a-NaP2,W, 5

O

56 .xHO to a solution of CoCl,.6H,O (0.59g, 2.5mmol) and NaCI (3g) in distilled water 35-40°). The heteropolyanion dissolved over a few minutes; stirring was continued for 15 20 mins. before the solution was filtered and kept overnight at The grey coloured solid which formed in the reddish mother liquor was collected on a frit. Filtration was difficult and slow. After washing the product with ethanol and diethyl ether, 3.8g of air dried product was obtained. It was recrystallized from a little distilled water warmed to ca. 450.

The compounds in which M Zn, Cu, Ni or Mn, when prepared in a similar manner, yielded larger crystals which were much more readily collected by filtration than the corresponding cobalt compound. The 31 P nmr spectra of the compounds M Zn, Cu, Ni or Mn were indicative of just one isomer present in solution The 3 1 P nmr spectrum of the compound M Co had, however, up to 12 resonance peaks between -6.9 and 25.5ppm (85% H3PO 4 external reference).

On heating a solution of this compound for several hours at 80-900, the Pnmr spectrum simplified to one peak centred at 10.8ppm flanked by two very weak peaks (contained less than 2%of the total peak areas) at 9.5 and 11.9ppm. This species is referred to as the high temperature form of the cobalt compound.

Preparation of the high temperature form of Nal 6 [Co 4

(H.O)

2

(P

2 Wis0 56 2 ].nH,O.

The cobalt compound (ca. 12g), prepared as described above, was dissolved in distilled water (75mL) and heated in a water bath at 80-900 for 7h. Heating time was not optimized and a considerable shorter time may have been sufficient. After addition of NaCI the red solution was let stand at room temperature.

Brown-red, shiny, platelet-like crystals collected on the bottom of the flask and WO 92/00078 PCT/AU91/00280 19 were readily isolated by filtration to give 9.8g of air dried product. This was dissolved in 7mL distilled water, filtered and set aside to cool. The crystals were collected and air dried, yield 8.5g, n is ca. 52.

EXAMPLE OF PREPARATION OF COMPOUNDS CONTAINING A CYCLOPENTADIENYL GROUP Preparation of ac-A 4 [rl-C H 5 )TiPW 11 039] .nHO. The [a-(Tis-CsHs)TiPWi 1

O

39 4 polyanion was made first by R.K.C. Ho and W.

Klemperer Am. Chem. Soc. 1978, 100, 6772) by reaction of a-[(n-C 4

H

9 4

N]

4

H

3

PWO

1 1 0 3 with (rs-CH s )TiCI 3 under anhydrous conditions.

[Full description of preparation in R.K.C. Ho Ph.D Thesis, Columbi, University, 1979, pp 82 83 W. Knoth Am. Chem. Soc. 1979, 101, 759) independently prepared the polyanion under aqueous conditions from (tlS-C 5 H)TiCl 3 and

PW

1 204 3 (at pH obtaining two isomers. More recently J.F.W. Keana et al. (J.

Amer. Chem. Soc. 1985, 107, 6714; ibid 1986, 108, 7951; ibid 1986, 108, 7957; J.

Org. Chem. 1987, 52, 2571) have made this polyanion and a number of substituted cyclopentadienyl analogues for use as electron microscopy labels for biological molecules.

The general synthetic method of Ho and Klemperer was used to make this compound. 30g NaWO 4 .2HO and 2.97g Na 2

HPO

4 .12H,O were dissolved in H2O in a beaker. To the stirred solution was added, dropwise, 7.2ml 10N HC1. A white precipitate formed. Stirring was continued and after lh sufficient 10N HCI was added to solubilize the compound (ca. 6.5mL). The pH of this solution was between 5 and 6. On addition of 12g solid (n-C 4

H

9 4 NBr a white precipitate formed which was treated slowly, especially after the reaction solution pH reached 4, and with vigorous stirring, with 3N HCI until the pH reached 0.7 to 0.8.

Approximately 15ml 3N HCI were required. The product was collected, washed with H,O and ether and dried. Yield WO 92/00078 PC/AU91/00280 20 10.94g (0.0030mol) a-[(n-C 4

H)

4

N]

4

H

3 PW,1039 and 0.70g (0.0032mmol) (rs-CHs)TiCl 3 were placed under N, into a 150mL Schlenk flask and 120mL 1,2-CH 4 C1 2 (distilled under argon from P,0 5 was added. The solution was stirred and warmed to 600 under an argon flow for 2-3h. The reaction solution colour changed from orange to yellowish to a greenish colour during the course of the reaction. On cooling to room temperature the solution was filtered and then treated with 200mL dry ether (about half the 1,2-CH 4 C1 2 was expelled with the HCI during the course of the reaction by the argon stream). The product was collected, washed well with water and ether and air dried. Yield 8.5g. This was dissolved in ca. 55mL acetone, filtered, and the yellow solution was treated with toluene until cloudy. 7g of yellow crystals of a-[(n-C4H,) 4

N]

4 [(rlS-C 5

H

5 )TiPW ,0391] were obtained.

6.4g of this tetra-n-butylammonium salt was dissolved in 100mL 1,2-CH,C1l and a saturated aqueous solution of KI (10g) was added. CHC3 was then added to the stirred mixture until the upper layer was nearly colourless. The organic layer was removed and the aqueous solution (ca. 6mL) together with some organics was centrifuged. Some solids had separated and therefore the aqueous solution was diluted to 15mL, filtered and placed in a vacuum dessicator over PO, After the solution had concentrated to ca. 10mL, orange crystals formed which were collected when the solution volume was ca. 5mL. Yield 2.9g orange a-K 4 [('q-CsHs)TiPW 1 ,03 9 ].nHO.

Preparation of cL-AT[(ls-C 5 sH)TiPW 7

O

6 1 ].nHO Compounds containing the heteropolyanion [(t 5

-C

5 Hs)TiP,W 7 0 6 1 7 were first pared by J.F.W. Keana et al. Am. Chem. Soc. 1985, 107, 6714; ibid. 1986, 7951; ibid. 1986, 108, 7957; J. Org. Chem. 1987, 52, 2571). In many of the pounds substituted cyclopentadienyl groups were incorporated and these compounds were designed for use as labels for biological compounds in transmission electron microscopy. The unsubstituted compound was prepared in high yield from 5

-C

5 H,)TiCl 3 by the method of W. Knoth Am. Chem. Soc.

WO 92/00078 PCT/AU91/00280 21 1979, 101, 759) but using o.,-KoP,2W 1 0 6 1 instead of the Keggin-type mono-vacant polyanion. The substituted cyclopentadienyl compounds were obtained by Keana et al. from (TlS-C 5

H

4 R)Ti(NMe)3 and o-KioPW 7 0 6 in DMF/benzene and adding a little aqueous acid during the reaction.

We have prepared these type of compounds from the readily available (l -CsH), TiCI 2 and ((rqs-CH 4

CH

3 ),TiCI 2 by reaction with a warm aqueous solution of ct-KoP 2

WT

1 7 0 61 Two of the compounds, ct,-(NMe 3 H)7(^ 5

-C

5

H

5 )TiPWO7 61 and the corresponding potassium salt, have been reported by J. F. W. Keana et al. (J.

Am. Chem. Soc., 1985, 107, 6714; ibid. 1986, 108, 7951).

Preparation of ca,-[NMe 3

H]

7 [(rS-CsH)TiPW, 7 6 1 ,].nH,O a-K 1 oP 2

W

1 7 0 61 .22HO (24.7g, Smmol) is dissolved in 100-150mL water. To this stirred solution, held at a temperature of 70-80 0 C but may be lower, was added a solution of bis(Tq'-cyclopentadienyl)titanium dichloride (1.3-1.4g; approx.

in an organic solvent usually tetrahydrofuran, but others of lower boiling point such as CH2C1, can also be used. The organometallic is added to the reaction solution at a rate commensurate with the expulsion of organic solvent from the reaction solution to minimize precipitation of heteropolytungstate. A flow of gas through the solution can be used to accellerate the rate of organic solvent evaporation. At the completion of the reaction an orange coloured solution is obtained which may contain a little precipitate.

Product purification is effected by either decanting or filtering the reaction solution at room temperature onto an alumina column (For example, Merck, Type 1077, 50-75g, washed with 1M sodium acetate solution acidified with acetic acid to around pH 5.6, and then water) and eluted with water. On treatment of the orange solution with trimethylammonium chloride (25g) a solid product is obtained, which is collected, washed with cold water and recrystallized from water (ca. 750, Ig product in 7-8ml water. The orange crystals are collected and air dried. Yield WO 92/00078 PCT/AU91/00280 (not optimized) 7-8g. The product is readily soluble in DMSO and hot water, sparingly in cold water.

'H nmr (CH 6 -DMSO, TMS 60.000); 62.91(s,63H, 66.38(s,5H,C 5 Hs), 68.9(b,NH).

'H nmr (D 2 0, 2

H

4 -TMSP(Na salt) d0.00); 62.99 (s,63H,CH3), 66.78(s,5H,CH,).

Alkali metal salts of the heteropolyanion are obtained by eluting the reaction product from the alumina column with the required alkali metal acetate solution (ca. pH 5.6) and precipitating the product with ethanol followed by recrystallization.

The'following examples are intended for illustration only and are not intended to limit the scope of the invention in any way. The term "active ingredient" as used in the Examples means a compound of formula or a pharmaceutically acceptable derivative thereof.

WO 92/00078 WO 9200078PCT'/ALJ91/OO2AFO 23 EXAMPLE 1 Using the appropriate general methods described above the following compounds were synthesised: Compound No. Formula 1. K 4 tCpTiPW 1

O

39 .nHO 2. (N(n-C 4

H

9 4 4 [CpTiPW 1

O

39 .nH,0 6. K 7 [CpTiPW 7

O

61 .nHO 7. Li 7 [CpTipl-W 17

O

61 1 .nHO O8. Na 7 [CpTiP-lW 17 0 6 1 .nHO 9. (NHMe 3 7 [CpTiPW 17

O

61 .nHO

(NH

4 7 [CpTiPW 17

O

61 .nH.0 11. K 10 [Co 4

(H

2 0) 2

(PW

9 O.1) 2 .22-H,O 12. Li 10 [Co 4 (H,0) 2

(PW

9 O4) 2 .nH 1

O

13. Na 10 [Co 4 (H,0)2 (PW 9 .nH,O 14. (NH 4 1 0 [Co 4 (H,0)2 (PW 9 .nH 1

O

16. INH(C2H 5 3 1 0 [Co 4

(HO)

2

(PW

9 O4)2J .nHO 17. [NH 3 (C2H 5 1 0 [Co 4 (H,0) 2

(PW

9 O4) 2 *nHO 18. Rb 10 [Co 4

(H

2 0) 2

(PW

9 0 3 4) 2 nHO .0 19. C$-1 [Co 4

(H

2 0) 2

(PW

9 O34) 2 J nHL{.O (18-C6K)I 0 [Co 4

(H

2 0) 1

(PW

9 O4) 2 .nH,O 21. Ga 5 (Co 4 (HO0) 2

(PW

9 O4) 2 J .nH,O 22. K 10 [Zn 4 (H,0)2 (PW 9 O34),] .22'-H,O 23. Li 10 [Zn 4 2

(PW

9 0 4 2 .nH,O 24. Rb 10 [Zn 4 (H,0)2 (PW 9 O4)2] .nHO 2 5. Cs 10 [Zn 4 (H20)2 (PW 9 O4) 2 .nHO 2 6. Na 10 [Zn 4 (HO0) 2

(PW

9 .nH.,O 27. (NH 410 [Zn 4

(H,O)

1

(PW

9 O34) 2 1 nHO 29. [NH 3

(C

1

HS)J

10 tZn 4

(PW

9 O4)2] .nH,O 30. (18-C6K) 10 [Zn 4 (H,O)2 (PW 9 O4)2] .nH,O 31. Ca5[Zn 4 (H,0)2 (PW90O42 .nHO WO 92/00078 WO 9200078PCTF/AU91/00280 24 33. [NH(CH 3 3 1 0 [Zn 4

(H,O)

2

(PW

9 04) 1 .nH,O 34. K 10 [Fe 4 (H,0) 2

(PW

9 O4) 1 .nHO Lil 0 [Fe 4

(H

2 -0) 2

(PW

9 0 34 2 .nH 1

O

36. Na 10 [Fe 4

(H

2

(PW

9 O,.4) 2 1 .nH 1

O

37. (NH 4 1 0 [Fe 4 2

(PW

9 O4) 1 .nHO

(NH

4 1 6 [Co 4 (lj 1

(P

1

.W

15 0 56 2 J .nH,0 41. Na 16 [Co 4 (HO2)2, 15 0 56 )21 .nH,)O 42. Li 16 [Co 4 (H,0) 2

(PIWI

5 0 56 2 .nH 1

O

69. (NH 4 2 6 [Co 9

P

5 W2 7 01 g H 1 ].nH 2

O

K

16 [Co 9

P

5

W

2 7 0119 H 1 .nH 1

O

121. Kj 0 [(FeOK) 4

(PW

9 0 34 )jI.nHO 122. Na 10 [(Fe0Na) 4

(PW

9 03)2].nH 2

O

123. Lil 0 [(FeOLi) 4

(PW

9 034) 2 ].nH,O 124. (NHMe 3 1 0 [(Fe0(NHMe 3 4

(PW

9 O34) 2 j.nH,0 125. Na 16 [Fe 4 (H,0) 2

(PW

15 0 56 1 .nH 2

O

126. Li 1 6 [Fe 4

(H

1 0)2(P 2

-W

1 5 0 56 )2].nHO 127. (NH 4 1 [(Fe0NH 4 4

(PW

9 04).nH,O 128. K 10 [(FeOIP) 4

((PW

9 O4) 1 .nH 2

O

WO 92/00078 PC/AU91/00280 25 EXAMPLE 2: Anti-HIV Activity The anti-HIV activity of compounds 1 to 21 of Example 1 has been assessed by the following test: Approximately one million human continuous lymphocytes (MT2) were exposed to each test compound at concentrations previously demonstrated to be non-toxic to MT2 cells (that is no drug related effect on cell numbers or cell morphology) for 2 hours at 37 0 C. They were then infected with Human Immunodeficiency Virus (HIV strain 237288, 2000-10,000 TCIDso per million MT2 cells) and cultured at 37 0 C in the presence of the test compound. Fresh amounts of test compound were added at the appropriate dilution 3 days later. Two types of controls were run with each batch of tests: A. HIV infected non-drug treated, B. Uninfected non-drug treated.

All cultures were monitored for the presence or absence of virus-specific cytopathic effects daily from day 3. The extent of virus-specific cytopathic effects (cpe) associated with each concentration of test compound was rated according to the following scale: 4+ 75-100% of MT2 cells showing cpe 3+ 50-75% of MT2 cells showing cpe 2+ 25-50% of MT2 cells showing cpe 5-25% of MT2 cells showing cpe less than 5% of MT2 cells showing cpe -ve no observable cpe in MT2 cells The activity of the test compounds was also assessed by degree of the inhibition of reverse transcriptase activity at each concentration. After 5-7 days incubation as described above, supernatant fluids from each flask were removed and tested for virion associated reverse transcriptase (RT) activity using a standard method.

The cpe effects and RT activity for each compound at various dose rates are given in Tables 1, 1A and 1B below. Greater than 90% inhibition of reverse WO 92/00078 PCT/AU91/00280 26 transcriptase activity represents effective control of virus replication and hence HIV infection.

WO 92/00078 WO 9200078PCI'/A U91 /00280 27 TAIBLE 1 Concentration ([tg/mL) 1 0.1 Therapeutic Index 0.01 HPA 23 Prior art compound for comparison Inhibition of RT activity of control) 62 0 11 cpe 3+ 4+ 4+ .0 Co2mpoun1 Inhibition of RT activity of control) 91.6 88.8 82.3 16.6 >1000 cpc y 3+ 6 RT activity of control) 99.7 97.4 98.8 82.5 >100 cpe -ye -ye 1+ 1, Cmpound9 Inhibition of RT activity of control 97.6 98.9 87.4 0 >200 cpe -ye -ye 4+ Cmpoun 1 RT activity of control) 97.1 88.7 58.1 58.4 >1000 cpe -ye 2+ 3+ WO 92/00078 W092/0078PCT/AU91 /00280 28 TABLE 1 (continued) Concentration tg/mL) 1 0.1 Therapeutic Inde~x Compound I' Inhibition of RT activity of control) cpe 99.3 -ye 99.4 -ye >100 Comp-id23 RT activity of conitrol) 98.8 85.8 12 cpe 2- 3+ £QD.Pund 27 Inhibition of RT activity of control 99.7 99.2 89.3 60.1 1500 cpe -ye -ye 1 4+ CompouInd34 Inhibition of RT activity of control) 9W6 81A4 26.2 cpe -ye 1+ Compound Inhibition of RT activity of control) 99.3 10.3 cpe -ye 4+ £CQmpu d 69 Inhibition of RT activity of control) 93.2 98.5 93.7 cpe -ye -ye WO 92/00078 PCT/AU9 1/00280 29 TABLE 1 (continued) Concentration .tg/ml,) i0 1 0.1 Therapeutic Index £ompnd Inhibition of RT activity %oi ntrol cpe 2+ 2+ Compound 122 Inhibition of RT activity of control) 98.6 84 >500 cpe -ye 1+ WO 92/00078 WO 9200078PCr/AU9I /00280 30 TABLE 1A Concentration ([tg/mL) s0 5 0.5 Therapeutic Tndex 0.05 Compound I Inhibition of RT activity of control) 96.1 cpe -ye 75.7 0 1+ 3+ Compoundl14 Inhibition of RT activity of control) 99.7 99.3 93.6 31.7 >200 cpe -ye -ye 1+ 4+ Compound 16 Inhibition of RT activity of control) cpe 2+ 4+ 4+ Compund 18 Inhibition of RT activity of control) cpe 2+ 4+ Compound 19 RT activity of control) 99.6 91.4 12 cpe -ye -ye 3+ Comnpound 21 RT activity of control) 19.4 7.7 cpe 3+ 4+ Crnp-Qflfl31 Inhibition of RT activity of control) 97 79 8 cpe -ye 4+ WO 92/00078 WO 9200078PCr/AU91 /00280 31 TABlLE 1A (=Q Concentration ([tg/mL) 1 n5 n5 Therapeutic Tnripy Compound 22 Inhibition of RT activity of control) 94.6 40 0 cpe -ye 2+ 4+ Comipound .0 inhibition of RT activity of control) 33.6 0 0 cpe 2+ 4+ 4+ Compound Inhibition of RT activity of control) 95A 60.3 15.4 >100 cpe 2+ 4+ Compund 33 Inhibition of RT activity of control) cpe 3+ 4+ Comp~und Inhibition of RT activity of control) 97 85 0 >100 cpe -ye 1+ 3+ Compolund 127 W0 Inhibition of RT activity of control) 97.5 97.3 76.2 >100 cpe -ye -ye WO 92/00078 WO 9200078PCTF/AU91/00280 32 TABLE 1A (continued) Concentration tg/mL) 0.5 0.05 0.005 Therapeutic Index Compou~nd 7 Inhibition of RT activity of control) cpe 91.3 18.4 3+ 22. 4+ 110 Comnpound 8 Inihibition of RT activity of control) 98.0 93.3 0 20 >100 cpe -ye 1- 3+ 4+ Compound Inhibition of RT activity of control) 97.1 93.5 23 7.8 >100 cpe -ye 3+ 4+ Compound 121 Inhibition of RT activity of control) 92.9 63.8 0 cpe 1+ 2+ Compound 123 Inhibition of RT activity of control) 74 15.1 23.4 cpe 1+ 3+ 4+ Compound 124 Inhibition of RT activity of control) cpe 2+ 4+ Compound 128 Inhibition of RT activity of control) 84 77 30 >100 cpe -ve 2)+ WO 92/00078 PCT/AU9I /00280 33 TABLE 18 (continued) Concentration ([tg/mL) 0.05 0.005 Therapeutic Index Compund 2 Inhibition of RT activity of control) cpe 2+ 4+ 4+ 4+ Com~pouLnd 37 .0 Inhibition of RT activity of control) cpe 3+ 4+ 4+ WO 92/00078 WO 9200078PCT/AU9I /00280 34 TABLE 1 Concentration (tg/mL) 175 n nl, Therapeutic Tnrli-v Compund13 Inhibition of RT activity of control) cpe 2+ 3+ Compound 17 .0 Inhibition of RT activity of control) cpe 4+ Cmpond 24 Inhibition of RT activity of control) 83 0 0 cpe 3+ 4+ Compound26 Inhibition of RT activity of control) cpe 2+ Compound 36 2 5 Inhibition of RT activity of control) 99.8 Coe -ye 70.4 27 3+ WO 92/00078 PCT/AU91/00280 35 EXAMPLE 3 The following formulation A may be prepared by wet granulation of the ingredients with a solution of povidone, followed by addition of magnesium stearate and compression.

mg/tablet Formulation A Active ingredient Lactose B.P.

Povidone B.P.

Sodium starch glycollate Magnesium stearate 50 250 10 26 L5 9 20 12 5 3 500 300 The following formulation B, may be prepared by admixed ingredients.

Formulation B Active ingredient Pregelatinised starch NF15 direct compression of the mg/capsule 250 150 400 WO 92/00078 PCT/AU91/00280 36 Formulation C (Controlled release formulation) This formulation may be prepared by wet granulation of the ingredients (below) with a solution of povidone followed by the addition of magnesium stearate and compression.

mg/tablet Active ingredient 500 Hydroxypropylmethylcellulose 112 (methocel K4M Premium) Lactose B.P. 53 Povidone B.P.C. 28 Magnesium stearate 7 700 EXAMPLE 4: Capsule Formulations Formulation A A capsule formulation may be prepared by admixing the ingredients of Formulation B in Example 3 above and filling into a two-part hard gelatin capsule.

Formulation B (infra) may be prepared in a similar manner.

Formulation B mg/capsule Active ingredient 250 Lactose B.P. 143 Sodium starch glycollate Magnesium stearate 2 420 931129,q:\oper\tgc,80064-91.L,37 -37- Formulation C (Controlled release capsule) The following controlled release capsule formulation may be prepared by extruding ingredients and using an extruder, followed by spheronisation of the extrudate and drying. The dried pellets may then be coated with release-controlling membrane and filled into a two-piece, hard gelatin capsule.

(a) (b) (c) (d) Active ingredient Microcrystalline cellulose Lactose B.P.

Ethyl cellulose rng/capsule 250 125 125 13 C C 513 EXAMPLE 5: Injectable Formulation Formulation: Active ingredient 0.200 g Hydrochloric acid solution, 0.1M qs to pH 5.0-7.0 Sodium hydroxide solution, 0.1M qs to pH 5.0-7.0 Sterile water qs to 10 ml The active ingredient may be dissolved in most of the water (35 "-40 and the pH adjusted to between 5.0 and 7.0 with the hydrochloric acid or the sodium hydroxide as appropriate. The batch may then be made up to volume with the water and filtered through a sterile micropore filter into a sterile ml amber glass vial (type 1) and sealed with sterile closures and overseals.

Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.

Claims (21)

1. A method for the treatment or prophylaxis of a retrovirus-associated infection characterised in that it involves the use of a heteropolytungstate compound of the general formulae I: Am [CpTiPWn 1 O 39 nH,O I(a) Am [CpTiP 2 W 1 7 O61]. nH z O I(b) Am [M 4 (H 2 0) 2 (PW 9 nHO I(c) A. [M 4 (H 2 0) 2 (P 2 W 1 5 0 6 nH,O I(d) Am [MgP s W-,7On 1 H 17 nH 2 O I(e) Formula I; wherein: M is Co, Fe, Zn, FeOA, FeOl Cp represents an optionally substituted cyclopentadienyl residue; A is a monovalent or divalent cation or mixture of such cations; m is the number of cations necessary for electrical neutrality of the molecule; or a pharmaceutically acceptable derivative thereof.

2. A method as claimed in Claim 1, characterised in that the cation A is sodium, potassium, lithium, ammonium, alkylammonium, a cationic alkali metal crown ether complex, magnesium, or calcium. NVO 92/00078 WO 9200078PCT/AU91/00280 39

3. A method as claimed in Claim 1, c-haracterised in that the compound of formula I is 1. 3.

4.

6.

7.

8.

9.

11.

13.

14.

16. 1,.

18.

19.

21.

22.

23.

24.

26.

27.

218. 29. one of the compounds listed below: K. 4 [CpTiPW 1 1 03] .nHO (N(n-C 4 H 9 4 4 [CpTiPW 1 O 39 .nH,,O Na 4 [CpTiPW 1 O 39 nI- 2 0 Li 4 [CpTPW 11 O 39 nH 2 O (NH 4 4 [CpTL WV 1 0 39 .nHO K7, [CpTiP 2 W 17 O 6 .nHO Li.7 [CpTiPW 7 O 1 .nHO Na., [CpTiPW 7 O 61 .nHO (NHMe,) 7 [CpTiPW 1 O 6 ,1 .nIIO (NH 4 [CpTiPW 7 O 61 .nHO K 1 0 [Co0 4 (H 2 0) 2 (PW 9 O34),] .22H 2 Li 10 [Co 4 (H 2 0) 2 (PW 9 O34) 2 .nH 2 O Na, 10 [Co 4 (H2)2) (PW 9 O4) 2 .nH.,O (NH 4 10 [Co 4 (PW 9 O4)2] .nH 2 O [NH(CH 3 )3j 10 [Co 4 (H 2 0) 2 (PW 9 O4) 2 .nHO [NH(C 2 H5) 3 [Co 4 (H 2 0) 2 (PWON)2] .nlf,O Rbl 0 (Co 4 (PW 9 O34) 2 .nH,O CSI [Co 4 (H 2 0) 2 (PW 9 034] .nH 2 O (18-C6K) 1 0 [CO4(H 2 (PW 9 Q34) 2 .nH 2 0 Ca 5 [Co 4 (H 2 0) 2 (PW 9 O4)2] K 10 [Zn 4 2 (PW 9 O4) 2 .22H 2 0 Li 10 [Zn 4 (H20) 2 (PW 9 O34) 2 .nHO Rb 10 [Zn 4 (H,O) 2 (PW 9 O34), .nHO CSI, [Zn 4 (H,O)2 (PW 9 O4) 2 J .nHO Na 10 [Zn 4 2 (PW 9 O34) 2 .nH 2 O (NH 4 10 [Zn 4 (H,O)2 (PW.VJX),]2 .nHO [NH(C.,H 5 )3jJO [Zn 4 fPW 9 O4) 2 .nHO [NH 3 (C2H5)] 1 0 tZn 4 (H.,Oy2 (PW 9 .nH.,O WO 92/00078 WO 9200078PCTF/AU9I /00280 40 (18-C6K)I 0 [Zn 4 (H 2 0) 2 (PW 9 O4) 2 .nH.,O 31. Ca,[Zn 4 (H,O) 2 (PW 9 O4) 2 .nH.,O 32. Mg4[Zn 4 (H 2 0) 2 (PW 9 O34) 2 1 *nH 2 O 33. [NH(CH 3 3 10 [Zn 4 (H,O) 2 (PW 9 O4) 2 .nH.,O 34. K 10 [Fe 4 (H,O)2 (PW 9 O4) 2 .nH2,O Lil 0 [Fe 4 (H 2 0) 2 (PW 9 O4)2] .nH 2 O 36. Nal 0 [Fe 4 (H,0) 2 (PW 9 O4) 2 .nH.,O 37. (NH 410 [Fe 4 (H 2 ,0)2 (PW 9 O4) 2 .nHO 37A. [NH(CH 3 3 10 [Fe 4 (H 2 0) 2 (PW 9 O.1) 2 nHO 38. (18-C6K) 10 [Fe 4 (PW 9 O4) 2 .nHO 39, K 16 [Co 4 (HO) 2 (P 2 Wl 5 0 56 .nHO (NH 4 1 6 [Co 4 (H 2 0) 2 (P- 2 W 15 0 56 2 .nH 2 O 41. Na 16 [Co 4 (H20) 2 (P 2 Wl 5 0 56 2 .nHO 42. Li 16 [Co 4 (H 2 0) 2 (P 2 W 15 0 56 2 .nH 2 O 43. (NHMe 3 16 [Co 4 (H 2 O)2 (P.-W 1 5 0 56 2 1 .nHO 44. Mg 8 [Co 4 (H,0) 2 (P 2 W 15 0 56 2 .nHO K 1 6 [Zn 4 (H 2 0) 2 (P'.Wl 5 0 56 2 .nHO 46. (NH 41 6 [Zn 4 (H 2 0) 2 (P 2 WI 5 0 56 2 .nHZO 47. Na 16 [Zn 4 (H 2 0) 2 (P 2 WI 5 0 56 2 48. Li 1 6 [Zn 4 (H 2 0) 2 (P 2 -WI 5 0 56 2 .nH 1 O 49. (NHMe 3 16 [Zn 4 (H 2 0) 2 (P2W 15 0 56 2 1 .nH 2 0 Mg 8 [Zn 4 (H 2 0) 2 (P 2 W 15 0 56 2 .nHO 68. Mg 8 [Mn 4 (H 2 0) 2 (P 2 W 15 0 56 2 .nHO 69. (NH 4 16 [Co 9 P 5 W27 01,9 H 1 .nH 2 O 70. K 16 [Co 9 P5W 27 0119 H 17 ,nI{,O 71. Li 1 6 [Co 9 P 5 W27 0119 Hf 1 7 .nH 2 0 72. Na 1 6 [Co 9 P 5 W 27 0119 H 17 ].-nH 2 0 73. Ca 8 [Cog P 5 W27 0119 H 17 ].nH 2 0 121. K 10 [(Fe 0K) 4 (P W 9 034),].nH )O 122. 10l [(Fe0Na) 4 (PW 9 031) 2 ].nH 2 0 123. Li 10 [(Fe0Li) 4 (PW 9 034)2].nHO WO 92/00078 PCT/AU91/00280 41 124. (NHMe 3 )o [(FeO(NHMe 3 4 (PW 9 ,O4),].nH,O 125. Na 16 [Fe 4 (HO) 2 (P2W,50 56 2 ].nHO 126. Li 1 6 [Fe 4 (HO0) 2 (P 2 W 1 5 0 56 2 ].nHO 127. (NH 4 1 o[(FeONH 4 4 (PWO934),].nHO 128. Ko[(FeOi 4 ((PW 9 gO4) 2 ].nHO or a pharmaceutically acceptable derivative thereof. 4. A method as claimed in any one of Claims 1 to 3, characterised in that the compound of formula I has a therapeutic index (as hereinbefore defined) of 50 or greater. The use in the manufacture of a medicament for the treatment or prophylaxis of a retrovirus-associated infection of a compound of formula I, as defined in any one of Claims 1 to 4. 6. A method for the treatment or prophylaxis of a retrovirus associated infection, characterised in that it comprises administering to a patient in need of such treatment or prophylaxis an effective amount of a compound of the general formula I, as defined in any one of Claims 1 to 4. 7. A pharmaceutical composition for the treatment or prophylaxis of a retrovirus associated infection, characterised in that it comprises a compound of the general formula I, as defined in any one of Claims 1 to 4, in association with a pharmaceutically acceptable carrier or diluent. 8. A compound of formula I as defined in any one of Claims 1 to 4, excluding those numbered 1, 2, 6, 9, 1 1 41, 47, 48 and those numbered 1, 2, 6, 9, lljl3 3 41, 47, 48 and INTERNATIONAL SEARCH REPORT International Application No. PCT'/A 91/00280 I. CLASSIFICATION OF SUBJECT MATER (if several classification symbols apply, indicate all) 6 According to International Patent Classification (IPC) or to both National Classification and IPC Int. Cl. A61K 31/66, 33/42; C01G 41/02, 49/04, 51/04; C07F 11/00, 15/02, 15/06 II. FIELDS SEARED Minimum Documentation Searched 7 SClassification System j Classification Symbols IPC I A61K 31/66, 33/42; C01G 41/00, 41/02; CO7F 11/00, 15/02, 15/06 Documentation Searched other than Minimum Documentation to the Extent that such Documents are Included in the Fields Searched 8 AU: IPC as above III. DOCUMENTS CONSIDERED TO BE RELEVANT 9 Category* I Citation of Document, with indication, where appropriate, I Relevant to of the relevant passages 12 Claim No 13 E,X AU,A, 70944/91 (JOHNSON MATITEY PUBLIC LIMITED COMPANY) 22 August 1991 (22.08.91) 1-8 P,A AU,A, 51410/90 (JOHNSON MATTHEY PUBLIC LIMITED COMPANY) 20 September 1990 (20.09.90) 1-8 A AU,A, 43168/89 (R F SCHINAZI C L HILL) 5 April 1990 (05.04.90) 8 P,A Chemical Abstracts, Volume 113, No. 22, issued 1990, 26 November (Columbus, Ohio, USA), M A Fedotov, B 2 Pertsikov D K Danovich, 'Oxygen-17, Phosphorus-31 and tungsten 183 NMR spectra of paramagnetic complexes with heteropolytungstate anion', see page 824, column 2, abstract no. 203399m, Polyhedron 1990, 9(10), 1249-56 (Eng) 8 A Chemical Abstracts, Volume 98, no. 14, issued 1983, 4 April (Columbus, Ohio, USA), R G Finke MW Droege, 'Trivant Sheteropolytungstate derivatives', see page 715, column 1, abstract no. 118514x, Inorg Chem 1983, 22(6), 1006-8 (BEg) 8 X J Chem Soc Dalton Trans, No 12, issued 1986 (Royal Society of SChemistry, London), H T Evans, CM Tourne, G F Tourne Special categories of cited documents: 10 later document published after the international filing date or priority date S"A" document defining the general state of the and not in conflict with the application but art which is not considered to be of cited to understand the principle or theory particular relevance underlying the invention earlier document but published on or document of particular relevance; the after the international filing date claimed invention cannot be considered novel document which may throw doubts on priority or cannot be considered to involve an claim(s) or which is cited to establish the inventive step publication date of another citation or document of particular relevance; the other special reason (as specified) claimed invention cannot be considered to document referring to an oral disclosure, involve an inventive step when the document use, exhibition or other means is combined with one or more other such document published prior to the documents, such combination being obvious to international filing date but Later than a person skilled in the art. the priority date claimed document member of the same patent family IV. CERTIFICATION SDate of the Actual Completion of the Date of Mailing of this International International Search Search Report September 1991 (05.09.91)I q 5 g 4 ,j 9, International Searching Authority I ignature of Auth'-iid Officer Australian Patent Office J BODEGRAVEN Form PCT/ISA/210 (second sheet) (January 1985) International Apr :ation No. PCT/AU 91/00280 III. DOCUMENTS CONSIDERED TO BE RELEVANT (CONTINUED FROM THE SECOND SHEET) Category* I Citation of Document, with indication, where appropriate, Relevant to of the relevant passages I Claim NO ST J R Weakley, 'X-Ray crystallographic and tungsten-183 NMR Structural studies of heteropolytungstate anions', see pages 2699 to 2705 1 7 X J Am Chem Soc, Volume 103, issued 1981 (American Chemical Society, Columbus, Ohio), R G Finke, M Droege, JR Hutchinson 0 Gansow, 'Trivacant Heteropolytungstate Derivatives', see pages 1587 to 1589 7 A J Am Chem Soc, Volume 108, issued 1986 (American Chemical Society, S Columbus, Ohio), R G Finke, B Rapko, R J Saxton P J Droaille, 'Trisubstituted heteropolytungstates as soluble metal oxide S analogues', see pages 2947 to 2960 8 Form PCTIIS; /210 (extra sheetn (January 19851 ANNEX TO THE INTERNATIONAL SEARCH REPORT ON ITERNATIONAL APPLICATION NO. PCT/AU 91/00280 This Annex lists the known publication level patent family members relating to the patent documents cited in the above-mentioned international search report. The Australian Patent Office is in no way liable for these particulars which are merely given for the purpose of information. Patent Document Cited in Search Patent Family Members Report AU 51410/0 Q CA 2012153 EP 390365 HU 53655 JP 3047130 NO 901232 NZ 232929 PT 93478 ZA 9001993 AU 43168/g' EP 360619 NZ 230745 WO 9003176 ZA 8907260 END OF ANNEX