CA2086263A1 - Antiviral agents containing heteropolytungstate - Google Patents

Abstract

Heteropolytungstate compounds of general formulae (I), wherein M
is Co, Fe, Zn, FeOA, FeO1/2; 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 involve the use of such compounds.

Description

~ 2086263 ~ WO 92/00078 PCr/AU91/00280 ANTIYIRAL AGENTS CONTAINING ~ETEROPOLYTUN~STATE

The present invention relates to heteropolyturlgstates and pharrnaceutically acceptable derivatives thereo~, pharrnaceutical compositions containing them~ and to the use o~ these compounds in therapy, particularly for the treatment or prophvlaxis of certain viral infections, for example retroviral infections such as Acquired Lmmune Deficiency Syndrome (AIDS)~ .
Because of the difficulty of inhibiting viruses while lea- ing the non-infected cells unimpaired, few antiviral dru~s are currently in widespread clinical use. This is especially true of viruses within the family Re~roviridae. The causative agenl for AIDS, the Human Immunodeficiency Virus, [also known as Human T-cell 15 Lymphotropic Virus 111 (HLTV-III)] is a member of this family. This virus will be identified herein as HIV. Infection with HIV is associated with depletion of T~ :
lymphocytes, brain disease, and several types of cancer including ~aposis sarcoma~
Patients infected with the virus also have a high incidence of opportunistic .
infections and a significantlv reduced life span. Another virus within the same .
20 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 adistinctive geographical pattern.

All members of the family Retroviridae possess a unique enzyme, reverse ~ ~
''5 transcriptase, which is necessary for their replication. Because this enzyme is not - : .
normally present in uninfected ~ells, it is considered a target for antiviral drugs.
Another virus utilising reverse transcriptase during replica~ion is the Hepatitis B . -Virus (HBV). HB~ causes widespread morbidity and mortality and is the main cause of primary hepatocellular carcinoma in individuals who are chronic carriers 30 of the virus. Because of the common evolutionary origin of HBV and the retroviruses, treatment of HBV-infected individuals with the same regimen of 2~862~3 ,; ~
WO 92/00078 . . ; PCI/AU91/0028 drugs proposed herein for the treatment of patients infected with H~V could be possible.

Background Heteropolytungstate compounds have been known for over 100 ears. Most of their applications stem from their redox chernistry and also their high ionic weights and charges. Their redox chemistr,v has led to their most common use: catalYsts for the oxidation of organic substrates such as, for example, propylene to acrvlic 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 ha-e also been shown to inhibit viral DNA and Ri~'A
polymerases (J.C. Chermann e- al., Biochem. Biophys. Res. Comm., 1975, 65~
1''~9; M. Herve et al., ibid 1983,116, 222). The antimony heteropol~tungstate (NH~)18[NaSb9W.,086], (HPA-73), has also been shown to have marginal activity against HIV in perrnissive cell lines (MT-~. .UT-l) howe-er in these cases thc cell toxicity is such that the therapeutic inde~' is low(about 10 or less) (Y. Lnouye. et al., Chem. Pharm. Bull. 1990, 38, 285). Because few anti-HlV agents were available at the time HPA-23 was tested in clinical trials despite this low therapeutic index (W. Rozenbaum et al, Lancet 19B5, II:450). HPA-23 has since been shown to be inactive in other in vitro screens and in subsequent clinical trials (1. 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 K71PTi.W~oO4o].6H.O inhibits the replication of HIV
in HTLV-1 carrying MT-4 cells (Y. Inouye, et al., Chem. Pharm. Bull. 1990, 3~, ''85).

I Therapeutic index is defined as the ratio of the dose which is toxic to 50~7c of the test cells to the dose required to reduce the irus count b-y 50~c .

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~ wo 92/00078 2 0 8 6 2 6 3 PCT/A~;91/0028û

The heteropolytungstates of this in~ention are based on Keggin-t~pe and Dawson-type structures, carrying either one or three ~acancies. These ~,acanciesare created b~ extraction of WOJ~ or W,066+ from the PWl.O~ eg~in~ or P,Wl~O6,~ (Dawson) structure. Isomers of these unsaturated (lacunar~) pol~anionsS are possible, a consequence of the location of the ~,acanc~. (R. Massart et al..
Inorg. Chem. 1977, 16, ~916; T.I.. Jorris et al., J. Am. Chem. Soc. 1987. 109!
7401; T.J.R. Weakl~, Polyhedron, 1987, 6, g31; R. Contant e~ al., J. Chem.
Res.(S), 1977, ''2; R.G. Finke et al., Inorg. Chem. 1987, 26~ 3886; M.T. Pope, 'Heteropoly and Isopoly Oxometallates', Springer-Verlag, 1983). The posi~ion of 10 the vacanc~ in P.W,706,~ iS defined b~ the prefi.~ ~, for a belt acanc~ or c~. for a cap vacancy. The prefix a- or ~- refers to isomerisation associated with the relative rotation of the W~ triad cap. In the tri-acant pol~anions~ PW9049-, thevacancy 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~ oxide triadlS has been removed. The unsaturated heteropolytungstates can behave as pol~anion ligands and bond. at their vacant site, with metal ions. The more stable unsaturated polyanions appear to be the c~. (R. Contant et al. J. Chem. Res. (S)~ 1977. ''~:T.J.R. Weakley, Polyhedron, 1987, 6, 931) and B-tvpe (W.H. ~noth.
Organometallics 1985, 4, 6 ) isomers. Transition metal ion complexes of t~pe ~0 (M4tPW903.,)]'~ are known only with the B-~- isomer of the polyanion ligand. A
recent structure determination (T.J.R. Weakley et al., Inorg. Chem. 1990~ 29, 1'35) on Nal,Cu[Cu4(H,O),(P,Wl5056)t].53H.O has shown that the bonding of the P.W,5056'2- ligand to the set of four coplanar copper atoms is similar to that found in the corresponding PW90349- containing compounds. In particular, the 25 P.W,5O56'2~ polyanion is formed from a-P2W,8O6.~ b,v removal of one edge-sharing cap (containing three tungsten atoms).

It is lcnown that the A-a-PW903~9~ anion can form complexes with transition metal ions. With Co2+, the species formed is [Co~(H,0)3(PW90~,)2]l-~ which on 30 heating in aqueous solution is converted to the B-a- structure [Co4(H.O),(PW9034)]'~ (W.H. Knoth, et al., Organometallics 1985.4.6').

2086~3 ~ . ~ PCI/AU91/0028~' The preparation of the compounds A~O~M4(H,O),PW90~ .nH.O. A = ~ M =
Mn(II), Co(II), Ni(II), Cu(II), Zn(lI) was described in a note b~ T.J.R Weal;lev e~
al. J.C.S. Chem. Commun. 1973, 139. A full paper on the s~nthesis and structure of se-eral of the compounds by these workers appeared much more recently (H.T.
5 Evans et aL J.C.S. Dalton Trans. 1986, 2699). Alternate preparative procedures~
employing the preforrned PWgO?.,9- polyanion, were developed b~ R.G. Finke et al~ (J. Am. Chem. Soc. 1981. 103, 1~8;7; Inorg. Chem. 1987, 26, 3886). The structure of the compounds, as noted above, is based on the B-~- isomer of the PW903.,9- t~ivacant polyanion. The four transition metal atoms (.M.~) are 10 coordinated to, and are located be~een. the t~rvo B-a-PW90 ,9~ pol!anions.

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 in-ention provides a mcthod for the treatment or prophylaxis of a retrovirus-associated infection which in-olves the use of a heteropolytungstate compound of the general formulae I having a therapeutic index (as hereinbefore defined) of 50 or greater: .
~0 A" [CpTiPW~039]. nH.O I(a) Am [CpTiP2WI7 6J- nH~O I(b) Am [M4(H~O)~(PW903~).]. nH.O I(c) A", [M4(H20)~(P.Wl505~kl nH-O I(d) Am [Mgpsw27o~lgH~ nH.O 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;

. - . ............ .

-.
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~ WO 92/00~78 2 ~ 8 6 2 6 3 PCI/AU91/00280 m is ~he number of cations necessary for electrical neutrality of the molecule.

The compounds of the invention are polyanions with associated cations (A~ for 5 electrical neutrality. They crystallise with a tariable number of molecules. of water of crystallisation (n) dependent upon the conditions of product reco~,er~ and subsequent treatment; all such hydrate forms fall within the scope of this in-ention.

For mono~alent cations, the number of cations (m) for the compounds of Formula I10 is as follows:
Compo~nd m I(a) I(b) 7 I(c) 10 :, ' I(d) 16 I(e) 1 6 If diYalent cations, or a mi.~cture of monovalent and di~alent cations, are present then a lesser number will be sufficient for electrical neutrality and m will be ~0 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, ''S 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:
(1) they do not interfere with the ~-bonding of the cyclopentadiene ring to the titanium atom;
30 ('') they do not interfere with the formation of the compound of formula 1; and (3) they do not have a deleterious effect on the anti~iral properties of the compound.
~ 4., ,~. , WO 92/00078 . . ; ` ! . ..~ .,, PCr/AU91/00~8 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. (J. Amer. Chem. Soc. 198S, 107, 6714; ibid 1986~ 10~. 7951; ibid 1986~ lOS.
5 7957; J. Org. Chem. 1987, 52! ~571).

By "a pharmaceutically acceptable derivative" is meant any pha~naceutically acceptable salt, or any other compound which, upon administration to the recipient, is capable of providing (directly or indirectly) a heteropolytungstate of the general 10 formula I, or an antivirally active metabolite or residue thereof.

It will be appreciated that the compounds according to the invention ma- also beused in the manufacture of a medicament for the treatment or prophvlaxis of a retrovirus-associated infection.
1~ .
In another aspect, the prescnt in-ention provides a pharmaceutical composition for the treatment or prophylaxis of a retrovirus-associated infection, which comprises an effective amount of a compound of the ge~neral formula I, in association with a pharmaceutically acceptable carrier or diluent.
~0 The prese~nt invention also extends to a method for the treatment or prophylaxis of a retrovirus associated infection, which comprises administering to a patient inneed of such treatment or prophylaxis an effective amount of a compound of the general formula I.
~5 Examples of compounds of Formula 1 which fall within the ambit of the present invention include the following~:

- "18-C6K" means compounds containing Kt complexed by the cvclic polyether 18-Crown-6 , ` ' . :
- : . .. . : . . . . . .
- , . . : - - .
.. . .. . ~ :
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`~` 20862~3 :-i WO ~2/00078 PCr/AI~91/00280 1 K4 [CpTiPW,l039] nH.0 2 (N(n-C4Hg)l)4 ~CpTiPWI1039] nH.0 3 Na4 [CpTiPWIl039] nH.0 4 Li4 lCpTiPW,I039] nH.0 (NH4)4 [CpTiPWIl039] nH,0 6 K7 [CpTiP,WI706,] nH.0 7 Li7 [CpTiP,WI,06,~ nH.0 8 Na7 [CpTiP2WI706l] nH.0 9 (NHMe3)7 [CpTiP,WI706l] nH.0 (NH4)7 [CpTiP.WI706l] nH.0 11 K,olCo4(H.0). (PW90}4),] '~H.0 12 Lilo~Co~(H.O), (PW903,),] nH,0 13 Na10[Co4(H.0), (Pwso34)2] nH-0 1~ (NH1)lo[col(H~o)1 (PW9034)J nH,0 lS [NH(CH~)3]l0[Co4(H.0~, (PW90~4),] nH,0 16 [NH(C Hs)3]lo [Col(H.0). (PW9034),J nH,0 17 [NH~(C Hs)]lo [Co~(H.0). (PW9034)2] nH,0 18 Rb,o[Co4(H,0), (PW90?4),] nH.0 19 Cslo [Co,(H.0)2 (Pwso34)~] nH-0 ~0 '0 (18-C6K),o [Co4(H.0), (PW90l4),] nH,0 21 Cas [Co4(H20)2 (PWsO34)J nH-0 '' KIO[Zn4(H.0)~ (PW9034).] "H,0 ~3 LiloIZn4(H.0)2 (PwsO34)2] nH20 24 RblO[Zn4(H.Ok (PW9034)2] nH,0 CslO[Zn4(H.0)2 (PW9034)2] nH-0 26 Na10[Zn4(H,0), (Pwso34)~] nH~0 ~7 (NH4)l0[Zn4(H,0)~ (PW9034),] nH,0 '8 [NH(C~Hs)3]~0[7n4(H~0)2 (PW9034),] nH,0 29 [NH3(GHs)]~o [Z4(H,0)2 (PW9034)2] nH~0 30 (18-C6K)~o [Zn4(H,0)2 (PW9034)2] nH,0 31 Cas[Zn4(H~0)2 (PWsO34)~] nH 0 32 M&[Zn4(H,0), (PWsO34)~] nH-0 .

33. [NH(CH3)~],0[Zn4(H~O)~ (P~sO~)~] nH~0 34. K,O IFe4(H.O), (PW9Ol4),] .nH.0 35. Li,o[Fe4(H.O).(PW30~4)~].nH,0 36. Na,olFe4(H.o)~(pw9o34)~] nH~o 37. (NH4),0[Fe4(H.O), (PW90~4),].nH.0 37A. [NH(CH3)3],0[Fe,(H.O), (PWsO~ ]. nH-0 38. (18-C6K)~o [Fe,(H.O).(PWg0~4).].nH.O
39. K,6[Co4(H.O). (P.W~5O56).] .nH,0 40. (NH4)l6[co4(H~o)~(p~wl5o56)2].nH,O
41. Na,6[Co,(H.O), (P,W,sO56),] .nH.0 42. Li,6[Co.,(H.O),(P.W,5056),].nH.0 43. (NHMe3)l6[co~(H~o)~(p~wl5o56)~] nH~o 44. Mg8 [Co~(H~o)~(p~wl5o56)J .nH.O
45. K,6[Zn4(H.0). (P~W,5056)~] .nH.0 46. (NH4),6[Zn4(H.O)~(P7wl5o56)7] nH 0 17. ~la~6[Zn,(H.O).(P.w~50s6)~] nH-0 48- Li~6[Zn4(H.O). (P~WlsOs6)3l nH~0 ~9. (NHMe3),6[Zn4(H.O).(P.W,5056).].nH.0 Mgs[Zn4(H.O)~(P.W~sOs6)2] nH-0 68. Mg8 [Mn4(H.Ok (P~W~sOs6)2] nH-0 69. (NH4),6 [Cog P5W., 119 H~7].nH.0 70. K,6[Co9P5W, 0"9 H,7].nH.0 71. Li,6[Co9P5W, 0"9 H~7].nH.0 7~. Na,6 [Co9P5w, 119 Hl,].nH.0 73. Ca,3 [Co9P5W.7 119 Hl,].nH.O
121. Klo [(FeOK)4(PW9O34).].nH.O - ~ -127. Na~0 [(FeoNa)4(pwso34)~]~nH~o 1''3. Li,o[(FeoLi)4(pwso34)2]nH-o 124. ~NHMe3),0 [(FeO(NHMe34(PW9O34)2] nH~O
1~. Na~6[Fe4(H~o)~(p~w~5os6)2] nH7o 1''6. Li,6[Fe4(H~o)~(p~w,sos6)~]nH~o 1~7 (NH4)~o[(FeoNH~)4(pw9o34)~]nH~o .. - .

~3 WO 92/00û78 2 0 8 6 2 6 3 PCr/AU91/00280 _ 9 _ 1~8. K,O[(FeO"2)4((PW90~.,).].nH.O

Of the abo~e listed compounds those numbered 1, ~, 6, g~ 13~ 3, ~6, 41, 47, 48 and 70 are known.

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" ,, ~ " ", ~ ,,~, ,,i, ' ~ i ' ;;. ~' ", ~ ""j,~ ' -2 0 8 6 2 6 3 ~ , PC~/AU91/0028~

-- 10 -- .
The in-ention also provides novel compound according to formula. I and includes the specific compounds listed above, e:Ycluding those listed as being known.

The compounds according to the im,ention, also refened to herein as the "active 5 ingredient(s)", may be administered for therapy by any suitable route~ including oral, rectal, nasal, topical (including buccal and sublingual)! ~aginal and parenteral (including subcutaneous, intramuscular, intravenous and intrade}mal~. PreferablS, administra~ion 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 the nature 10 of the chosen acti~e ingredien~.

In general a suitable dose will be in the range of 3.0 to 1~0 mg per }iilo_ram body weight of the recipient per da-, preferably in the range of 6 to 90 mg per ~iilogram body weight per day and most preferably in the range 15 to 60 mg per ~iilogram 15 body weight per day. The desired dose is preferably presented as two, three. foun fi-e, si~c or more sub-doses administered at appropriate intervals throuehout the day. These sub-doses may be administered in unit dosage forms, for e,~ample, containing 10 to lS00 mg, preferably ~0 to 1000 mg, and most preferably 50 to 700 mg of active ingredient per unit dosage form 'O
Ideally the active ingredient should be administered to achieve pea~i plasma concentrations of the acti-e compound of from about 0.1 to about 75 mM, preferably about ~ to 50 mM. This may be achieved, for e.~ample, by the intravenous injection of a 0.1 to 5% solution of the active ingredient, optionally in ''5 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 interrnittent infusions containing about 0.4 to about 15 mg/mg of the active ingredient.

30 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 (I). together ' ~ `

.~ "' .
. . ' ' ' ' ' . ' ' ' ' .

8~2~3 ;.-.~ WO 92/00078 PCI`/AU91/00280 with one or more pharmaceutically accep~ably carriers thereof, and optionally other therapeutic agents. Each carrier must be pharrnaceutically "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 intradennal~ administration~ The compositions maycon-eniently be presented in unit dosage fol~n and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the acti-e ingredient with the carrier which constitutes one 10 or more accessory in_redients. In general, the compositions are prepared by uniforrnly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both. and then if necessary shapin_ the product.

15 Compositions of the present in-ention suitable for oral administration may bepresented as discrete uni~s 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 ~0 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 ''S granules, optionally mixed with a binder (e.g inert diluent, preservative disintegrant (e.g. sodium starch glycollate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose) surface-active or dispersing agent. Moulded tablets maybe 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 30 scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethvl cellulose in varying proportions to provide the desired release profile. Tablets ma- optionally - - . . . . - ~ . ~ . .......... - . . . . ......... .

.. . ~ .. . . . , : . - - . ~ . . . , . . ;

2 08 62~ 3 . ; ;

be provided with an enteric coating, to pro-ide release in parts of the gut other than the stomach.

Compositions suitable for topical administration in the mouth include lozenges S comprising the acti~,e ingredient in a fla~oured basis~ usually sucrose and acacia or tragacanth gum; pastilles comprising the acti~e ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia gum; and mouthwashes comprising the actiYe ingredient in a suitable liquid carrier.

10 Compositions for rectal administration ma- be presented as a supposito~ with a suitable base comprising, for example. cocoa butter or a salicylate.

Compositions suitable for aginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition 15 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 ~0 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 (Iyophilized) condition requiring only the addition of the sterile liquid carrier, for exarnple water for '~S injections, immediately prior to use. Extemporaneous injection solutions andsuspensions 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 30 sub-dose, as herein abo~,e recited, or an appropriate fraction thereof, of an acti~,~e ingredient.

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~ ~VO 92/00~78 2 0 8 6 2 6 3 PCr/AU91/00280 The compounds according to the invention may also be presented for use in the form of veterinary compositions~ ~vhich may be prepared~ for e.~;ample~ bv methods that are conventional in the art. Examples of such etcrinary compositions include those adapted for:
S (a) oral administration! external application, for example drenches (e.g. aqueous or non-aqueous solutions or suspensions); tablets or boluses; powders!
granules or pellets for admixture with feed stuffs; pastes for application ~o the tongue;
(b) parenteral administration for example by subcutaneous, intramuscular or intravenous injection, e.g. as a sterile solution or suspension; or (when appropria~e) by intramammary injection where a suspension or solution is introduced into the udder via the teat: `~
(c) topical application, e.g. as a cream, ointment or sprav applied to the skin: or (d) intravaginally, e.g. as a pessary, cream or foam.
The administered ingredients mav also be used in therapv in conjunction with other medicaments or in conjunction with other immune modulating therapy includin&
bone marrow or Iymphocyte ~ransplants or medications such as le-amisol or thymosin which would increase Iymphocyte numbers and/or function as is ~0 appropriate~

It should be understood that in addition to the ingredients particularly mentioned above, the compositions of this invention may include other agents con-entional in the art having regard to the type of composition in question, for example, those~5 suitable for oral administration may include such further agents as sweeteners, thickeners and flavouring agents.

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wo 92/00078 ~ ; PCr/A~l91/00280 GFNER4L METHODS OF PR~:P.~R~ TION

1. Preparation of A~o [M4(H.O)(PW90-~).] .nH.O (1(c)) Two general methods of preparation of these compounds have been used: that due to H.T. Evans et aL (JCS Dalton Trans~ 1986. ~69g) or the method of R. G. Finke et aL (Inorg. Chem. 1987, 26, 3886) (All tempera~ures are in Celsius.) 10 EXAMPLE OF A COMPOUND ~DE BY THE EVANS ET~IL PROCEDURE

Preparation of K,o[Co4(H.O).(PWg03~)~].'~H.O

6.11g (0.0~1 mol) Co(NO3)..6H.O was dissol~ed in 110 mL H.O containing 9.8g 15 (70% w/w) HNOl and brought to the boil. A solution of ~9.7g (0.090 mol) Na.WO4~H.O and 3.9~g (0.011 mol) Na.HPO4.1~H.O in 110 mL H~O was then slowly added. After about two thirds of this solution had been added to the re~luxing cobalt nitrate solution, a precipitate formed which redissol~,ed on continued addition of the solution. After 5h reflux the clear reddish solution was ~0 treated with 40g solid ICCI. 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,6[Co9P5W~70"9H"] (see below).] The product was purified by twice dissolving in hot water and precipitating with excess KCI and then recrystallising from water at 85 and standing at room temperature. Yield of air''5 dried product = 14.6g blue-purple crystals.

EXAMPLES OF COMPOUNDS MADE BY THE FINKE ET.'.~. PROCEDURE
Preparation of K!0[Fe4(H.O)(PW9O34).~.nH.O

30 This method in-olves the reaction of preformed PW90349- polyanion with a stoichiometric amount of metal salt. The PW90},9- anion was prepared bv the method of R. Massart e~ aL (Inorg. Chem. 1977,16, ~916) as hydrated - . :. ., - ......... :
, . . ~ - :. . - --. :- .
. , . . ~ -. ' .
. . . . .

- . -, - ~. . .,: . . ; . ~ . . . ..

2~86263 A-Na8HPW90~ which on heat treatment at 1~0 is converted at least in part~ to the B- forrn (W.H. Knoth et al., Inorg. Chem. 1987, 26, 3886). This isomerization reaction is not completely understood. Following Finke et al., the pol~ranion used in this work was heated at 1~0-14~ for l-'h, usually about 1-l~ hours.

~.71g(1~ mmol) (NH4).FetSO~k.6H,O and 17.1g of heat treated Na8HPWg03~.~4H.O were placed in a ~50 mL Schlen~; flasl;. After degassing under u,acuum and replacement of the air with argon, 1'0 mL of de-aerated water were added and the reaction mixture refluxed for 5h. Degassed KCI (ca. 30g) was 10 then added and the mixture let stand at room temperature o~erni~ht. The brownsolid was collected under argon to gi~e a ~ield of 15.~g of crude product. The compound was redissol~ed in 100 mL of de-aerated water at 80-85 under argon and treated with ca. ~Sg degassed KCI. The product was collected under argon from the cool reaction solution and then redissolved in 100 mL de-aerated water at 15 80-85 under argon and filtered hot through a G3 frit filter stic};. On standing at room temperature, brown crystals tormed which were collected under ~rgon and dried under vacuum. Yield of brown crystalline product was 13.8g. It is air sensiti~e, oxidizing to a yellow material.

''0 Preparation of Klc[(FeOK).,(PW90~,).]. nH,O

While the following preparation is typical and gives good yields, it is not mandatory to use preformed ~-Na8HPW9034.19H~O or an iron(III) carboxylate to ~.
obtain the product. ~.-''5 A mixture of 31 g (11.~5 mmol) of ~-Na8HPW90}~.19H.O, prepared following R.
G. Finke, M. Droege and P. J. Domaille, Inorg. Chem. 1987, 26, 3886-3896, and an iron(IlI) carboxylate (''''.5 mmol based on iron) were mixed and 100 mL wateradded. The mixture was magnetically stired with gentle warming. The reaction .
30 mixture was refluxed for B-1~ hours, during which time the solution became somewhat lighter in colour. On addition of 35 g KCI, the solution solidified to a yellow mass which was collected ( yield 30.6 g) after the reaction mixture had .

. .
... , .. , . , , , . ~ .

. ~. .

- : : . . .
. ., . ~ ., ~ . ;
.. . .. . .. ~ ... . - . , . ~

20862~ 3 - `^
~VO 92/00078 PCT/AU91 /0028~i cooled to room temp. The product was dissolved in ca~ '75-300 mL boiling water and filtered hot through a Whatman No. 540 filter paper. ~0g 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. ~50 mL water near boiling point, filtered through a 5 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 ~5 mL water (at room temp.) and then ~vith ethanolbefore air drying on the frit. Yield ~''.5 g. Storing over silica gel under v acuum for several days resulted in a weight loss of about 0.3 g. The amount of water 10 associated with the product is ariable. Analyses bv ICP indicated an atomic ratio of K7Fe.PW9, suggesting that the compound may ha-e a formula of type K~.,[(FeO)4tPWg03~),]. nH,O. Titration of the protonated fonn of the compound~
prepared by elution through a column of "Amberlite" resin~ type IR-1'0 in the H~form, with KOH, suggests that the compound is perhaps better formulated as 15 K,0[(FeOK)4(PWgO~)~].nH.O, since about ~ of the 14 protons are neutralized only when the pH of the solution e:sceeds 3.5.

The infra-red spectrum of the compound in KBr is characterized by a triplet between 1040-1080 cm-' (phosphatc stretches), and strong bands at 947~ 873, 809 ''O and 7''4 cm~~, 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, 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 S Na, Li and NH4 salts are more readily soluble in water than either the corresponding K or NMe}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) 30 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 :, .:

2~8626~

were made by addition of a slight e~ccess of the ether to an aqueous slurry of the .~ ~v pure K,O[M4(H.O),(PW90~ ].nH.O compound. The isolated compound was then washed with aqueous ethanol. .

S EXAMPLE OF PREPAR~TION OF CO;~/SPOUND BY CATION EXCH~NGE

Preparation of (NH4),6 ~Co9P5W~70~9H~7~.nH.O. (1(e)) The potassium salt of this compound is obtained as a side product in the s~nthesis 10 of E~lo[C~(H-)-(P Ws3~)~] H-O by the method of H T E~ans et al . Isolalion of the crude ~ salt was as described in the preparation of K,O[Co4(H.O),(PWg03.,).].nH.O. The product (7.4g) was dissolved in warm water (ca. 70mL) to give a red solu~ion which was filtered and then treated with 16g NH4CI. Pink product (5.3g) was obtained after standing 16h ar 5. This step was 15 repeated to give 5.1g of product which was dissolved in Oml of water at 65-70, filtered and kept at 5 for se-eral da~s. The produc~ was collected and air dried:
yield ~.0g. More compound can be obtained by reducing the solvent o~ on addition of ethanol.

The compounds were prepared by the procedure of R.G. Finke et al.
a-K6P.W,8O62 was obtained by the slow addition of a near theoretical amount of alM solution of KHCO3to a"B-K6P2W,8O62, reacidifying the reaction solution and ~S precipitating with KCI The potassium salt was converted to the sodium one using NaClO4 and the trivacant polyanion salt Na,.P.W,5O56.18H~O was obtained on raising the reaction solution pH to 9 with Na.CO~ as described by R.G. Finke et al.

.
,, . . , .-. . ~ . .. .
: : . . , , ~, - - - -: , :,. . --. -. : . - . . . .

~u~o~ ~
WO 92/00078 PCl /AU91/0028 EXAMPLE OF COMPLEXING OF POLYANION WITH A DIVALE~'T
TRANSITION ~ETAL

Preparation of Na,6~M4(H.O)~(P.w,5O56)~] nH~O (l(d)~

The compound in which M = Co was prepared by adding c~-Na,.P.W,5O5O.:;H.O
(5g) to a solution of CoCl..6H.O (0.59g, '.~rnmol) and NaCl (3g) in distilled water (50mL, 35-~0). The heteropolvanion dissolved over a few minutes: stirring was continued for 15 - ~0 mins. before the solution was filtered and kept o-ernight at 10 5. 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 recrvstallized from a little distilled water warmed to ca. ~.

15 The compounds in which M = Zn. Cu, Ni or Mn. when prepared in a similar marmer, yielded lar~er crystals which were much mor~ readil- collected bv filtration than the corresponding cobalt compound. The 31p nmr spectra of the compounds M = Zn. Cu, Ni or Mn were indicati-e of just one isomer present in solution (D.O). The ~P rLmr spectrum of the compound M = Co had~ however. up ~0 to 1~ resonance peaks between -6.9 and 75.5ppm (85% H~PO~ e.~ternal reference).
On heating a solution of this compound for several hours at 80-90, the 3'Pnmr spectrum simplified to one peak centred at 10.8ppm flanked by two ver~ 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 temperatureform of the cobalt compound.
''5 Preparation of the high temperature form of Na,6[Co4(H.O).(P.W,5O56),].nH.O.

The cobalt compound (ca. 12g), prepared as described above, was dissolved in distilled water (75mL) and heated in a water bath at 80-90 for 7h. Heating time30 was not optimized and a considerable shorter time may ha-e been sufficient. After addition of NaCl (6g). 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 PCr/."U91/00280 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 crvstals were collected and air dried, ~ield 8.5g, n is ca. 5.'.

S EXAMPLE OF PREPARATIOl~ OF COMPOUNDS CONTA~NING
CYCLOPENTADIENYL GROUP
Preparation of a-A4[r,5-C5H5)TiPWI,O39] .nH.O. (1(a)) The [c~ 5-C5H5)TiPW110;9] polvanion was made first by R.K.C. Ho and W.
10 Klemperer (J. Arn. Chem. Soc. 1978~ 100, 677') by reaction of a-[(n-C,H9)4N],H.PW,10 9 with (~5-C5H5)TiCl. under anhydrous condi~ions.
[Full description of preparation in R.K.C. Ho Ph.D Thesis, Columbia Ijniversi~y 1979, pp8~.83.] W. Knoth (J. Am. Chem. Soc. 1979,101, 759) independently prepared the polyanion under aqueous conditions from (~5-C5H5)TiCI. and 15 PW,,O 03- (at pH 7), obtaining two isomers. More recently J.F.W. Keana et al. (J.
Amer. Chem. Soc. 1985, 107. 6714; i'oid 1986~ 108, 79~1: ibid 1986~ lOS. 79~7: JOrg. Chem. 1987, ~2, ~571) ha-e made this polyanion and a number of substituted cyclopentadienvl analoeues for use as electron microscopy labels for biological molecules.
The general synthetic method of Ho and Klemperer was used to make this compound. 30g Na WO4.~H.O and ~.97g Na HPO4.12H,O were dissolved in 60mL
H.O in a beaker. To the stirred solution was added, dropwise, 7.'ml 10N HCI. A
white precipitate formed. Stirring was continued and after lh sufficient 10N HCI~5 was added to solubilize the compound (ca. 6.5mL). The pH of this solution wasbetween 5 and 6. On addition of 1~g solid (n-C4Hg)4NBr a white precipitate formed which was treated slowly, especially after the reaction solution pH reached 4, and with ~igorous stirring, with 3N HCI until the pH reached 0.7 to 0.8.
Approximately 15ml 3N HCI were required. The product was collected, washed 30 with H.O and ether and dried. Yield 30g.

.. ..
. - . - : . .

208~2S 3 - -WO 92/000?8 PCT/AU91/0028~
-- O -- . `
10.94g (0.0030mol) cL-[(n-C4H9)~N~,H,PW"039 and 0.70g (0.003 mmol) (~5-C5H5)TiCl3 were placed under ~. into a 150mL Schlen~i flas~; and 1'0mL
l~-C,H4Cl. (distilled under argon from P.O5) was added. The solution was stirredand warmed to 60 under an argon flow for ~-3h. The reaclion solution colour S changed from orange to yellowish to a greenish colour during the course of thereaction. On cooling to room temperature the solution was filtered and then treated with 'OOmL dry ether (about half the 1.'-C~H4Cl. was expelled with the HCI
during the course of the reaction bv the argon stream). The product was collected.
washed well with water and ether and air dried. Yield 8.5g. This was dissolved in 10 ca. 55mL acetone, filtered! and the yello~ solution was treated with toluene until cloudy. 7g of yellow crystals of ~-[(n-C,Hg)~N].,[(-l5-C~H~)TiPWllO g] were obtained.

6.4g of this tetra-n-butylammonium salt was dissol~,ed in 100mL 1~-CH,Cl, and 15 a saturated aqueous solution of KI (lOg) was added. CHCl3 was then added to thc stirred mixture until the upper la~er was nearly colourless. The organic la~er 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 l~mL, ~iltered and placed in a vacuum dessicator ov,er P.O5. After the ''O solution had concentrated to ca. 10mL, orange crvstals formed which were collected when the solution ~rolume was ca. SmL. Yield ''.9g orange a-K4[t~5-C5H5)TiPWIl039].nH.O.

Preparation of ch-A7[(~5-C5H5)TiP.W,706,].nH,O (1(b)) ~5 Compounds containing the heteropolyanion [(~j-C5H5)TiP,W,70~jl]7- were first prepared by J.F.W. Keana et aL (J. Am. Chem. Soc. 1985, 107, 671~; ibid. 1986, 108, 7951; i~id 1986, 108, 79S7; I. Org. Chem. 1987, 52, ~571). In many of the compounds substituted cyclopentadienyl groups were incorporated and these 30 compounds were designed for use as labels for biolo~ical compounds in transmission electron microscopy. The unsubstituted compound was prepared in high yield from (~5-C5H5)TiCl3 by the method of W. Knoth (J. Am. Chem. Soc.

r~- 2 0 8 6 2 6 ~ ; ~

1979, 101, 759) but using ch-K,oP.W,7061 instead of the ~eggin-t,vpe mono-~acant polyanion. The substituted cyclopentadienyl compounds were obtained by Keana et al. from (~5-C5HIR)Ti(l`~Me~)~ and a.-K,OP~W,.O~ in DMF/benzene and adding a little aqueous acid during the reaction.
We hav,e prepared these type of compounds from the readily a-ail.,ble (~'-C5H5).TiCI. and t(~5-C5H4CH3).TiCI. by reaction with a warm aqueous solution of ch-K,oP,W,7O~,,. Two of the compounds! c~-(NMe3H)7(~s-C5H5)TiP.W,7O6,] and :
the corresponding potassium salt. have been reported by J. F. W. E~eana er al. (J.
10 Am. Chem. Soc., 1985,107, 671~: ibid 1986.108. 7951).

Preparation of c~.-[NMe3H]7 [(~5-C5H5)TiP.W~7O61].nH~O

15 a-K,OP,W,706,.~'H.O ('1.7g, 5mmol) is dissolved in 100-150mL water. To this - stirred solution. held at a temperature of 70-80C but may be lower~ was added a solution of bis(~5-cyclopentadienyl)titanium dichloride (1.3-1.~g; approx.
S.~mmol) in an organic solvent usuall~ tetrahydrofuran, but others of lower boiling point such as CH.CI. can also be used. The organometallic is added to the reaction 'O solution at a rate commensurate with the e.~pulsion of organic sol~ent from the reaction solution to minimize precipitation of heteropolvtungstate. A flow of gas through the solution can be used to accellerate the rate of organic solvent e~,aporation. At the completion of the reaction an orange coloured solution is obtained which may contain a little precipitate.
~5 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 lM sodium acetate solution acidified with acetic acid to around pH 5.6, and then water) and eluted with water. On treatment of the orange30 solution with trimethylammonium chloride ('5g) a solid product is obtained, which is collected, washed with cold water and recrystallized from water (ca. 75", lg product in 7-8ml water. The orange crystals are collected and air dried. Yield - . - . - ~

(not optimized) 7-8g. The product is readily soluble in DMSO and hot water~
sparingly in cold water.
IH nmr (~H6-DMSO, TMS ~ ?: o~.91(s~63~I~ CH3)~ o6.38(s~5H,C5H~) o8~9(b,NH).
5 ~H nmr (D~O, ~H~-TMSP(I~a salt) dO.OO); o~.99 (s~63H~CH~)~ o6.78(s~5H~C5H5~.

Alkali metal salts of the heteropolvanion 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 b~
10 recrystallization.

The following examples are intended for illustration onl~ and are not intended to limit the scope of the im,~ention in an~ wa~. The term "ac~i~e ingredient" as used in the Examples means a compound of formula (I) or a pharmaceuticall~
15 acceptable deri~ati-~e thereof.

-- '3 --EX, lMPL ~ 1 Using the appropriate general methods described abo~e ~he follo~in~ compounds were synthesised Compound l~o. Formula K4 [CpTiPWI,O;9] nH.0 (N(n-C4H9)4)l ICpTiPWI1039] nH.0 6 K7 [CpTiP,W~706~] nH.0 7 Li7 ICpTiP.W1706~] nH.0 8 Nla7 [CpTiP,W17061] nH.0 9 (NHMe )7 [CpTiP.WI.O~] nH.0 (NH4)7 [CpTiP.W,706,] nH,0 11 Klo [Co4(H.O). (PwsO34)2] ''H-0 1~ Lilo [Co,(H.0). (PWsO34h] nH-0 13 ~a~O[Co4(H-0)~ (PWsO34h] nH-0 14 (~H4),0[Co (H~0). (PW~0~,).] nH.0 16 [~H(C Hs)~]~o[col(H.oh (PW9034).~ nH.0 17 [NH3tC H5)],0 lCo,(H.0), (PW90~4),] nH.0 18 Rb,o[Co,(H.0). (PW903,),] nH.0 ~0 19 Cs~o[Co4(H.0). (Pwso34h] nH~0 (18-c6K)~olco4(H~o)~ (PW9034)2] nH.0 ~1 Ca5 [Co4(H~0)- (PW9034).] nH-0 ''' Klo[Zn4(H.Oh (PW9034).] ~H.0 ~3 Li~O[Zn4(H,Ok (PWsO34)2] nH-0 ~5 ~4 Rb~O[Zn4(H.Oh (PW903~).] nH.0 ~5 CslO [Zn4(H.O)~ (PW9034).] nH,0 ~6 Na~0[Zn4(H.0). (PW9034).] nH.0 ~7 (NH4)~0 [Zn~(H.Oh (PW9034),] nH,0 ~9 [NH3(C Hs)]lo[zn4(H~oh (PW90~4),] nH,0 (18-C6K)~O[Zn4(H.0), (PW903,),] nH.0 31 Cas[zn4(H~o)~ (PWsO34)~] nH-0 .. .. . . . . . .

. . , . . .... ~ . :, . . : ~

208626 3 - ` :
W O 92/00078 PC~r/AU91/0028 - ~4 -33 [NH(CH3)3]l0 [ZnJ(H,O), (PW9034),~ .nH,O
34. Klo [Fe4(H,O), (PW903~),] .nH,O
35. Li,o EFe4(H,O), (PWgO.~,),] .nH,O
36. NalO[Fe4(H20), (PW903,),] nH~O
37. (NH4),0 [Fe4(H,0)2 (PW90~4),~ .nH.O
40. (NH4),6 lCo4(H,O), (P2W,5056)2] .nH,O
~1. Na,6 [Co4(H,0)7 (P,W,50s6),] ~nH,O
4'~. Li,6[Co~(H.O). (P~wlsos6)~] nH-O
69. (NH4),6 [Co9 P5W~, 0,19 H,7].nH.O
70. ~16 [Cog P5W77 01l9 Hl7].nH.O
121. K~O[(FeOE~) (PWg03,).].nH,O
1''''. NalO [(FeONa),(PWg0~4).].nH.O
123. Lilo[(FeoLi)4(p W9~4)2]-nH~
12~. (N H Me3)lo[(Feo(N H Me3))4(p W934)2]nH~
125. Nal6 [Fe.,(H.O).(P.WI5056)2].nH.O
126. Li,6 ~Fe,(H,O).(P.W,5056).].nH.O
127. (NH4),O[(FeoNHJ)4(pw9o34)~ nH~o 1~8. K,o[(Feo~r~,((pwso3~)~] nH~o 2 0 8 ~ 2 ~ 3 ; ! ~
WO 92/00078 PCl`/A~'9I/00280 ~ ~ _ EX,IMPLE 2: Anti-H~YActivitv The anti-HlV activity of compounds 1 to ~1 of E~ample 1 has been assessed by the following test: `

Approximately one million human continuous Iymphoc-tes (Mr) were e~posed to each test compound at concentrations pre-iollsl- demonstrated to be non-toxic toMT_ cells (that is no drug related effect on cell numbers or cell morpholog-) for hours at 37~C. They were then infected with Human Irnmunodeficiency Virus (HIV strain 237288, ~000-10,000 TCID50 per million MT~ cells) and cultured at 10 37C in the presence of the test compound. Fresh amounts of test compound were added at the appropriate dilution 3 da-s later. Two types of controls were run with each batch of tests:
A. H~V infected non-drug treated, B. Uninfected non-drug treated.
15 ta) All cultures wete monitored for the presence or absence of virus-specjfjccvtopathic effects daily from day 3. The e.Ytent of ~,irus-specific cytopathic effects (cpe) associated with each concentration of test compound was rated according to the following scale:
4+ 75-1009'o of Mr cells showing cpe 3+ 50-75% Of Mr cells showing cpe 2+ ~5-50% of Mr cells showing cpe + 5-~5% of MT~ cells showing cpe .
+/- less than 5% of Mr cells showing cpe -ve no observable cpe in Mr cells ~5 (b) The activity of the test compounds was also assessed by degree of the inhibition of reverse transcriptase activity at each concentration. After ~-7 days incubation as described above, supernatant fluids from each flask were removed and tested for ~irion associated reverse transcriptase (RT~ activity using a standard 30 method.
The cpe effects and RT activity for each compound at various dose rates are gi-en in Tables 1, lA and lB below. Greater than 90% inhibition of re-erse . -: ~ . .- .................... . ~, :- -- - : . - . . : -, .

20862~3 ~

-- '6 --transcriptase activity represenls effective control of v itus replication and hence HrV
infection~

~ 2086263 ~
wo 92/00078 pcr/Al~i9l /oo28o -- _7 --Concentra~ion (ug/mL) Therapeutic 10 1 0.1 0.01 Inde~
S HPA '3 - Prior art compound for comparison Inhibition of RT aeti~,ity (~o of eontrol) 6' 0 11 <10 epe 3+ ~+ 1+

10 Compound 1 Inhibition of RT activitv (% of control~ 91.6 88.8 8'.3 16.6 >1000 epe -ve +!- '+ 3+
:
Compound 6 RT aetivity (% of eontrol) 99.7 97.~ 98.8 8~.5 >100 epe -ve -~e 1+

~0 Compound 9 Inhibition of RT aetivit~ .
% of eontrol 97.6 98.9 87.~ 0 >200 epe -ve -ve + ~+

. 5 Compound 11 ., RT aetivity (% of eontrol) 97.1 88.7 S8.1 58.1 >1000 epe -ve +/- 2+ 3+

. . . . . . . . - ~ .. - .
- : : . . . .

- - . . . . , .. , ; . . ,, .. ~
- - ~

208~2~.3 ~

TABLF 1(continued!
Concentration (~lg/mL) Therapeutic lOO l O l Q. l Inde~
CompQun~ 1~
5 Inhibition of RT activity (% of control) 99.3 99.~ 0 >100 cpe -~ e -~ e 4+
Compound ~3 RT activity (% of control) 98.8 85.8 1. >50 cpe +~- ~+ 3+

Compound''7 Inhibition of RT activitv % of control 99.7 99.~ 89.3 60.1 >1500 cpe -- e -ve 1+ 4+
. ~ .
Compound 34 ~0 Inhibition of RT activit~
(~o of control) 98.6 81.4 ~6.~ >75 cpe -- c +/- l+
.
Compound 40 ''5 ~nhibition of RT acti~,ity : :.
(% of control) 99.3 10.3 >50 cpe -ve 4+ -Inhibition of RT activity (% of control) 93.~ 98.5 93.7 >50 cpe -ve -ve +/-.

.. . . , . . . , ., . ,.. . .. , . . - . , -- . .. j .. .

i~ U ~ b ~ b ~
~ W O 92/00078 P ~ /AU91/00280 _ ~9 _ TAP~T.F. 1(continued?
Concentration (~lg/mL) Therapeutic 100 10 1 0.1 Inde:Y
Compound 70 5 Inhibition of RT acti~ity qo of control cpe +l- ~+ ~+ >~0 Compound 1~
10 Inhibition of RT activity (% of control) 98.6 8~ >~00 cpe -~ e l+

.. ... . ..... ... .... . .
- - ~., :

... - ........... .. ~: . . .
- .;,; . . -.

2~8626 3 ` `
W O 92/00078 P ~ /AU91/00280 TA~I.E lA
Concentration (ju~/mL) Therapeutic ().~ 0.05 Inde:~
Compound ~
Inhibition of RT activity (% of control) 96.1 75.7 0 >40 cpe -~, e 1+ 3+
Compound 1~
Inhibition of RT activity ~ -(% of control! 99.7 99.3 93.6 31.7 >'00 cpe -ve --e 1+ 4+
Compound 1 6 Inhibition of RT activity (% of control) cpe ~+ 4+ 4+ ~-Compound 18 Inhibition of RT activity (% of control) cpe ~+ ~+ >~o ~L2 ,, ''~ RT activity (% of control) 99.6 91.4 12 >20 cpe -ve -ve 3+
Compound . 1 RT activity :
(% of control) 19.4 7.7 >20 cpe 3+ 4+
Compound 31 Inhibition of RT activity (% of control) 97 79 8 >20 cpe -ve +/- 4+

, . ~ v .

:.

2~2~3 ,........................................... . . . ..

T~BT .E 1 A (cont ) Concen~ration (~lg/mL) Therapeutic 0 5 0.05 Index Compound ~
Inhibition of RT activity (~o of control) 94.6 40 0 >~0 cpe -ve ~+ ~ +
Compound . .5 Lnhibition of RT activit-(% of control) 33.6 0 0 >~0 epe ~+ ~+ ~+ : -Compound 30 Inhibition of RT aetivity (% of eontrol) 95.160.3 15. I >100 cpe +/- + 4+
.
Compound 33 ~0 Inhibition of RT activity . .
(% of control) cpe 3+ 4+ >5 .

~5 Lnhibition of RT aetivity (% of eontrol) 97 85 0 >100 epe -ve 1+ 3+
~ -Compound 1~7 30 Lnhibition of RT aetivity (% of eontrol) 97.597.3 76. >100 epe -ve -ve +/-.
. ~ -... . - . ~ .

WO 92/00078 : PCI/AU91/00280/~
-- 3_ --TAl"T.F. 1 A (contirlued) Concentration (~lg/mL) Therapeu~ic ~ 0.0~0.00~ Index CompounçL 7 S Inhibition of RT activity (% of control) 91.3 18.~_.5 ~o cpe +/_ 3+ ~+

Compound 8 ' 10 Inhibition of RT activit-(% of control) 98.0 93.3 0 ~0 >100 cpe -ve +~- 3+ 4+

Compound 10 Lnhibition of RT activity (5~o of control) 97.1 93.5 '3 7.8 >100 cpe -ve +/- 3+ 4+
. _ .
CompQund 1''1 Inhibition of RT activity (% of control) 92.9 63.8 0 ~0 cpe +I- 1 + ~+
Compound 1~3 ''S Inhibition of RT acti~ity (% of control) 74 15.1 '3.4 >50 cpe 1+ 3+ 4+
Compound 124 Inhibition of RT activity (% of control) cpe +/- 2+ 4+ >~o Compound 128 Inhibition of RT activity (% of control) 84 77 30 >100 cpe -ve +/- ~+
' , ., .~ 20862~3 -~' W O 92/00078 PCT/A U91/00280 TABT F 1 A (continued~
Concentration (!lg/mL) Therapeutic 0.5 0.0~ 0.005 Inde~
CompQUn~
o Inhibition of RT acti-i (% of control) cpe '+ l+ 4+ 4+ ~o Compound 37 Inhibition of RT acti~ity (% of control~
cpe +/- 3+ 4+ 4+ ,~o .

- 20862~3 `:
WO 92/00078 PCr~AU91/0028(:~i --3~ -TAP~I F 1 B
Concentration (llg/mL) Therapeutic -5 '' 5 0.75 _ 0.0''5 Inde~; .
Cornpou~ld 13 5 Inhibition of RT acti~,ity (% of control) cpe +!- ~+ 3+ >O
. :.
Compound 17 10 Inhibition of RT actin,ity t% of control) ~'0 :
cpe +I- 3+ 4+
Cornpound '4 Inhibition of RT activity (% of control) 83 0 0 ~'0 cpe +/- 3+ 4+

Compound ''6 ''O Inhibition of RT acti~itS
(% of con~rol) ~-cpe +/~ +

Comp.o~ 30 ~5 Inhibition of RT acti~ity (% of control) 99.8 70.1 7 0 cpe -n, e +/- 3+ 4+

~ 2~86263 `
:, ? WO 92/0007B PCr/AU91/00280 E~XAMPLE 3 The following fonnulation A may be prepared by wet granulation of the ingredients with a solution of povidone, followed by addition of magnesium stearate and compression.
mgltablet Formulation A
(a3 Active ingredient 50 ~50 (b) Lactose B.P. 210 '6 (c) Povidone B.P. 15 9 (d) Sodium starch glycollate ~0 1' (e) Magnesium stearate 5 15 The following formulation B, may be prepared by direct compression of the admixed ingredients.
Formulation B mglcapsule Active ingredient ~sO
Pregelatinised starch NF15 150 " ' . ' " ', ' ` ' i ' , ~ ~ , :
'`' , ' 20~6263 . ~`.` "
WO 92/00078 PCr/AU91/0028 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 steara~e and compression.
mgilablet (a) Active ingredicnt 500 (b) Hydroxypropylmethylcellulose 11' (methocel K4M Premium) (c) Lactose B.P. 53 (d) Povidone B.P.C. '8 (e) Magnesium stearate 7 15 ~XIMPLE 4: Capsule Formulations Formulation,1 A capsule formulation may be prepared b~ admixing the ingredients of FormulationB in Exarnple 3 above and filling into a two-part hard gelatin capsule.
Formulation B (in~ra) may be prepared in a similar marmer.
Formulation B ~ ~ .
mg/capsule (a) Active ingredient 250 (b~ Lactose B.P. 143 (c) Sodium starch glycollate '5 (d) Magnesium stearate ~''0 -~ WO 92/00078 2 0 8 6 2 6 3 pcr/Au9l/oo28n Formulation C (Controlled release capsule) The following controlled release capsule formulalion may be prepared by extruding ingredients (a), (b) and (c) using an e:Ytruder~ followed by spheronisation of the extrudate and drying. The dried pellets ma- then be coated with S release-controlling membrane (d) and filled into a two-piece! hard gela~in capsule.

mg/capsll le (a) Acti-e ingredient ~50 (b) Microcr-stalline cellulose 125 (c) Lactose B.P. 1'5 (d) Ethyl cellulose 13 ~13 EXAMPLE 5: Injectable Formula~ion Formulation:
Active ingredient 0 ~00 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 (35-40C) 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 volurne with the ''5 water and filtered through a sterile micropore filter into a sterile 10 ml arnber glass vial (type 1~ and sealed with sterile closures and overseals.

.,.. ,.. ..... ... ,... , ,. . ~ .

Claims (8)

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 [CpTiPW11,O39]. nH2O I(a) Am [CpTiP2W17O61]. nH2O I(b) Am [M4(H2O)2(PW9O34)2]. nH2O I(c) Am [M4(H2O)2(P2W15O56)2]. nH2O I(d) Am [M9P5W27O119H17]. nH2O I(e) Formula I;

wherein:
M is Co, Fe, Zn, FeOA, FeO1/2;
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.

3. A method as claimed in Claim 1, characterised in that the compound of formula I is one of the compounds listed below:

1. K4[CpTiPW11O39].nH2O

2. (N(n-C4H9)4)4[CpTiPW11O39].nH2O

3. Na4[CpTiPW11O39].nH2O
4. Li4[CpTiPW11O39].nH2O
5. (NH4)4[CpTiPW11O39].nH2O
6. K7[CpTiP2W17O61].nH2O
7. Li7[CpTiP2W17O61].nH2O
8. Na7[CpTiP2W17O61].nH2O
9. (NHMe3)7[CpTiP2W17O61].nH2O
10. (NH4)7[CpTiP2W17O61].nH2O
11. K10[Co4(H2O)2(PW9O34)2].22nH2O
12. Li10[Co4(H2O)2(PW9O34)2].nH2O
13. Na10[Co4(H2O)2(PW9O34)2].nH2O
14 (NH4)10[Co4(H2O)2(PW9O34)2].nH2O
[NH(CH3)3]10[Co4(H2O)2(PW9O34)2].nH2O
16. [NH(C2H5)3]10[Co4(H2O)2(PW9O34)2].nH2O
17. [NH3(C2H5)]10[Co4(H2O)2(PW9O34)2].nH2O
18. Rb10[Co4(H2O)2(PW9O34)2].nH2O
19. Cs10[CO4(H2O)2(PW9O34)2].nH2O
20. (18-C6K)10[Co4(H2O)2(PW9O34)2].nH2O
21. Ca5[Co4(H2O)2(PW9O34)2].nH2O
22. K10[Zn4(H2O)2(PW9O34)2].22H2O
23. Li10[Zn4(H2O)2(PW9O34)2].nH2O
24. Rb10[Zn4(H2O)2(PW9O34)2].nH2O
25. Cs10[Zn4(H2O)2(PW9O34)].nH2O
26. Na10[Zn4(H2O)2(PW9O34)2].nH2O
27 (NH4)10[Zn4(H2O)2(PW9O34)2].nH2O
28 [NH(C2H5)3]10[Zn4(H2O)2(PW9O34)2].nH2O
29. [NH3(C2H5)]10[Zn4(H2O)2(PW9O34)2].nH2O

30. (18-C6K)10[Zn4(H2O)2 PW9O34)2].nH2O
31. Ca5[Zn4(H2O)2(PW9O34)2].nH2O
32. Mg5[Zn4(H2O)2(PW9O34)2].nH2O
33. [NH(CH3)3]10[Zn4(H2O)2(PW9O34)2].nH2O
34. K10[Fe4(H2O)2(PW9O34)2].nH2O
35. Li10[Fe4(H2O)2(PW9O34)2].nH2O
36. Na10[Fe4(H2O)2(PW9O34)2].nH2O
37. (NH4)10 [Fe4(H2O)2(PW9O34)2].nH2O
37A. [NH(CH3)3]10[Fe4(H2O)21(PW9O34)2].nH2O
38. (18-C6K)10[Fe4(H2O)2(PW9O34)2].nH2O
39. K16[CO4(H2O)2(P2W15O2)2].nH2O
40. (NH4)16[Co4(H2O)2(P2W15O26)2].nH2O
41. Na16 Co4(H2O)2(P2W15O56)2].nH2O
42. Li16[Co4(H2O)2(P2W15O26)2].nH2O
43. (NHMe3)16[Co4(H2O)2(P2W15O56)2].nH2O
44. Mg8[Co4(H2O)2(P2W15O56)2].nH2O
K16 [Zn4(H2O)2(P2W15O56)2].nH2O
46. (NH4)16[Zn4(H2O)2(P2W15O56)2].nH2O
47. Na16[Zn4(H2O)2(P2W15O56)2].nH2O
48. Li16 [Zn4(H2O)2(P2W15O56)2].nH2O
49. (NHMe3)16[Zn4(H2O)2(P2W15O26)2].nH2O
50. Mg8[Zn4(H2O)2(P2W15O56)2].nH2O
68. Mg8[Mn4(H2O)2(P2W15O56)2].nH2O
69. (NH4)16[CO9P5W27O119H17].nH2O
70. K16[Co9P5W27O119H17].nH2O
71. Li16[Co9P5W27O119H17].nH2O
72. Na16[Co9P5W27O119H17].nH20 73. Ca8[Co9P5W27O119H17].nH20 121. K10[(FeOK)4(PW9O34)2].nH2O
122. Na10[(FeONa)4(PW9O34)2].nH2O
123. Li10[(FeOLi)4(PW9O34)2].nH2O

124. (NHMe3)10[(FeO(NHMe3))4(PW9O34)2].nH2O
125. Na16[Fe4(H2O)2(P2W15O56)2].nH2O
126. Li16[Fe4(H2O)2(P2W15O56)2].nH2O
127. (NH4)10[(FeONH4)4(PW9O34)2].nH2O
128. K10[(FeO1/2)4((PW9O34)2].nH2O
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 orgreater.

5. 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 retrovirusassociated 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, 11, 13, 22, 23, 26, 41, 47, 48 and 70.