CA2065279A1 - Antiviral pyrimidine nucleosides - Google Patents

Abstract

Pyrimidine 4'-thionucleosides of formula (I), wherein Y is hydroxy or amino, and X is chloro, bromo, iodo, trifluoromethyl, C2-6 alkyl, C2-6 alkenyl, C2-6 haloalkenyl or C2-6 alkynyl and physiologically functional derivatives thereof. These compounds have utility as anti-viral agents.

Description

_ WO91/01326 PCT/GB90/01099

2~S~2~

The present invention relates to pyri~idine nucleo.~ides and their use in medical therapy particularly for the treatment ~r prophyl~xis of ~irus i~fections.
Of the DNA viruses, those of the herpes group are the ~ources of the ~ost co~mon vir~l illnesses ~n man. The group includes herpes ~i~plex vlru~ (BSV), vsricella zoster virus (~ZV), cytomegal~virus (CMV); Epst~in-Barr Yirus (EBV) and human herpes virus 6 (HHV6). HSY 1 ~nd HSV 2 are some of the most common infe~tious agents of ~a~. Most ~ these ~iruses are able to persist in the h~st's neur l cells: onc~
infected, individuals are at risk of recurrent clinical ma~ifestations of infection which can be both physically and psych~lcgieally distressing.
~ SV infection is often char~ctericed by extensive and 1~ debilit~ting lesions of the ~kin, ~outh ~nd~or genitals.
Pri~ary infections ~ay be subclinical ~lthough tend to be more ~evere th~n infeetions in individuals previously exposed to the virus. ocul r infecti~n ~y HsV can lead ~ ker2titis or catar~ct~ thereby ~ndangering the host'~ ~ght. Infection ~0 in the newborn, ~n i~mun~co~pro~ised patient~ or penetration of the infe~tion into the centr~l nervous ~ystem can prove fa~al.
~ rans~ on o~ the ~irus ~ by direct phy~ical W09l/01326 PCT/GB90/0l099 - 2 _ 2~a27~ :

contact between a host and a recipient: the spread of HSV
infection is therefore considered a very ciynificant ~ocial problem, particul~rly ~s no effecti~e vaccine i~ yet available.
Varicella zoster (~ZV) is ~ herpes virus which causes chickenpox and shingles. ChickenpDx is the pri~ary disease produced in a hDst without immunity and in young children is usually a mild illness ch~racteri~ed by a vesicular rash and fever. S~ingles or zoster is the recurr~nt f~rm of the disease which occurs in adults who were previously infected with varic~ z~ster virus. The clinic~l manifesticns of shingles are characterised by neuralgia and a vesicular skin rash that is unilateral and dermat~mal in distribution.
Spread of inflammation may lead to paralysis or convulsions. ;
Coma can occur if the meninges become affected. In immunodeficient patients YZV may disseminate causing serious or even fatal illness. V2Y is of serious concern in patients receiving immunosuppressive drugs for tr~nsplant purposes or for treatment ~f m~lign~nt ne~plasia and i~ a 6erious complic~tion of AIDS p~tients due to their imp~ired immune ~y~tem. -In common with other herpes viru~es, infection with CMV lea~s to a l~felong ~ssociation of ~irus and host and, : fDllowing ~ pri~ry lnfection, ~irU5 ~ay be hed for ~ number 25 ~f year~ Cong-nit~l infection fGllow.ing infection of the ~ -mother during pregnan~y ~ay give rise to cllnical effec~s ' ~

~ . . ~ . .
.~
Sp.s ;;1 ~

. wosl/0l326 PCT/GB90/01099 ~Q~52~9 such as death or gross disease (microcephaly, hepatosplenomegaly, ~aundice, mental retardation)~ retinitis leading to blindness or, in less ~evere f~rms, f~ilure to -' t~rive, and ~us~eptibility to ch~st and ear infections. CMVinfection in patients who ~re im~unoco~pri~ised for example DS a resul~ of ~ali~nancy, treatment with i~mun~suppressive drugs following transpla~t~tion or infection with ~uman Immunodeficiency virus ma~ give rise to retinitis, pneumoitis, gastrointestinal disorders ~nd neurological diseases. CMV infectîon in AIDS patients is a predominant cause of morbidity as, in 50-80~ of the ~dult population, it is present in a latent form and can be re-activated in immuno-compromised patients.
Epstein^Barr virus tEBv) causes infectious mononucleosi~, and is ~lso suggested as the causative agent of nasopharyngeal cancer, immunobl~stic lymphoma, Burkitt'~
lymphoma ~nd hairy leukoplaki~.
HBV is a vir~l pathogen of world-wide major importanoe. The virus is ~etiologically ~ss~ciated with pri~ary hepatocellul~r carcinoma ~nd i~ thought to cause 80~
of the world'c liver cancer. In the U~ited St~tes ~re than ~en thousand people ~re hospitali~e~ for H8V illness each ~^c year, and aYer~ge of 250 die wit~ ful~inAnt di~ea~e. The : .
United 8tates Gurrently contain~ an e5ti~ated po~l of ~00,000-1-million in~ect~ous c~ rriers. Chronic ~ctive hepatiti~ gener811y ~evelops in over 25% of c~rriers, ~nd 2U6527~P~r~GiB 9 G / 01 ~$1S
~ 1 October 1991 ~1 10 91 often progresses to cirrhosis. Clinical effects of infection with HBV ran~e from headache, fever, malaise, nausea, vomiting, anorexia and abdominal pains. Replication of the virus is usually controlled by the immune response, with a course of recovery lasting weeks or months i~ humans, but infection may be more severe leadinq to persistent chronic liver disease outlined above.
We have now found that certain pyrimidine 4'-thionucleosides have potent activity against herpes viruses.
Such compounds include pyrimidine 4'-thionucleosides of the following general formula ' .:
y : '.

N ~ X (I) o~l ~ . .
\ N

Ho___ ¦ .
:: i/S~

\ / ,''' ~ " ~.
OH

' wherein Y is hydroxy or~amino, and X is halo, trifluoromethyl, methyl, C2_6 alkyl, C2_6 alkenyl, C2_6 :

Uni'~ P ~.. ? ~t C~ ? ¦ ~ tr ~lT~l-rE

P~T!~ / ' ~ ~ 9 ~

~1 October 1~91 ~1 10 91 haloalkenyl,including 2-bromovinyl or C2_6 alkynyl, and physiologically functional derivatives thereof.
The invention thus provides pyrimidine 4'-thionucleosides of the formula (I) y N ~ X

HO ~ S ¦ :

. /\1 :'.' ~ ' ' OH
.

lS wherein Y is hydroxy or amino, and X is chloro, bromo, iodo tri~luoromethyl, C2_6 alkyl, C2_6 alkenyl, C2_6 haloalkenyl or C2~ alkynyl, and physiologically functional derivatives , thereof.

It will be appreciated that by virtue o~ the definition of the ~roup Y the compounds of formula (I) are derivatives either of uracil or of cytosine.
It will also be appreciated that the compounds of formula (I) may exist in various tautomeric forms.
The compounds of Formula I may exist as ~- or ~-, U ' ~C~ O~i D~ S ~ T ' F~ t~ . ~,;,, ,~ ~.
., ~ C ' , ;_ . -P~T/GB 90/~1 Q99 - 6 - ~31 October 1991 ~1 10 91 anomers; ~-anomers are preferred.
In the definition of formula (I), references to alkyl groups include groups which, when they contain at least three carbon atoms may be branched or cyclic ~ut which are preferably straight (particular alkyl groups include ethyl~;
references to alkenyl groups include groups which may be in the E- or Z- form or a mixture thereof and which, when they contain at least three carbon atoms, may be branched but are preferably straight; and references to alkynyl groups include groups which, whe~ they contain at least four carbon atoms may be branched but which are preferably straight: particular alkenyl groups include vinyl and E-~l-propen~l) and particular alkynyl groups include ethynyl and prop-l-ynyl.
References to halo-substituted groups include chloro, bromo, iodo and fluoro substituted groups and groups substituted with two or more halogens which may be the same or diffexent, for example perhalo substituted groups (particular haloalkenyl groups include E-(2-bromovinyl)).
Preferred compounds of the formula I include those in which the group X is C2_4 al~yl or haloalkenyl preferably C2_3 alkyl, C3_4 alkenyl or alkynyl, or hal~vinyl. Preferred ~aloalkenyl groups are straight chain haloalkenyl groups having a ~ingle halogen group on the terminal carbon. Also preferred are halo~lkenyl groups having ~ double bond in the - 25 l-position. iOf such compounds, those having a 2-halovinyl group which is in the E configuration are preferred.
Particular compounds of the invention are compounds of formula (I) and phy~iiologically acceptable derivatives ~ t~ om P.~o~t ~ ~ S~!~ST~

2 0 ~ a 2 7 ~&T~
~l October 199i ~1 10 91 :
thereof wherein the pyrimidine base is selected from:
1. 5-Iodouracil 2. 5-Iodocytosine

3. 5-Ethynyluracil 5 4. 5~Prop-l-ynyluracil 5. 5-Vinyluracil 6. E-5-(2-Bromovinyl)uracil 7. E-5-(1-Propenyl)uracil 8. 5-Ethyluracil 10 9. 5-Trifluoromethyluracil 10. E-5-(2-Brom~vinyl)cytosine 11. 5-Propyluracil and wherein the 4-thio sugar moiety is the 2-de~xy-4-thio-D-ribofuranose moiety.
Compounds of formula (I) havin~ the beta configuration which are of especial interest as ~ntiviral agents are: . .~
-- E-5-(2-bromovinyl)-21odeoxy-4 ~ -thiouridine " _t 2'-deoxy-5-iodo-4'-thiouridine _ 2'-deoxy-5-ethyl-4'-thiourid~ne ~i ..
5-bromo-2'-deoxy-4'-thiouridine ~ . -2'-deoxy-5-prQpynyl-4'-thiouridine ~ .
5-chloro-2'-deoxy-4'-thiouridine ~ :~
2'-deoxy-5-tri~luoromethyl-4'-thiouridine 2'-deoxy-5-ethy~yl-4l-thiouridine 2'-deoxy-5-~-(2 bromovinyl~-4'-thiocytidine ; 2'-deoxy-5-propyl-4'-thiouridine ~2'-deoxy-~-(propen-1-yl)o4'-thiouridine Preferred compounds of formula (I) are E-5-(?-- ` P~Tl~i~ a o~ GS~

31 10 91 ~ ;

bromovinyl)-2'-deoxy-4'-thio-~-uridine and 2'-d~oxy-5-ethyl-

4'-t~Aio-~-uridine. T~ese compounds are of particular use against HSV 1 and 2, and VZV infections.
Also preferred are 2'-deoxy-5-halo-4'-thiouridine ~-compounds, which are of particular use against CMV
infections.
The above-mentioned derivatives include the pharmaceutically acoeptable salts; esters and salts of esters, or any other compound which, upon administration to a human subject, is capable of providing (directly or indirectly) the antivirally active metabolite or residue thereof.
Preferred mono- and di-esters according to the ' in~ention include carboxylic acid esters in which the non-,, .
c~rbonyl moiety of the ester grouping is selected from . . I .
; straight or branched chain alkyl, (e.g. tertiarybutyl);
cyclic alkyl (e.g. cyclohexyl); alkoxyalkyl (e.g.
' methoxymethyl)~ carboxyalkyl (e.g. carboxyethyl), aralkyl , (e.g. benzyl), aryloxyalkyl (e.g. phenoxymethyl), aryl (e.g.
phenyl optionally substituted by halogen, Cl_4 alkyl or C
alkoxy); sulphonate esters such as alkyl- sr aralkyl- ' .
sulphonyl ~e.~. methanesulphonyl); mo~o-, di- or tri-phosphate esters which may or may not be blocked, amino acids '~ esters and nitrate esters. With regard to the above-- 25 described esters, unless otherwise specified, any alkyl moieties present in such esters advanta~eously contain 1 to 18 c~rbon atoms, particularly 1 to 4 carbon atoms, in the case of 6tra~ight chain alkyl groups, or 3 to 7 carbon atoms TC~ A~ T

PCT/~ 9~/~10~9 20~a~9 ~1 oclo~e~ 1~91 in the case of branched or cyclic alkyl groups. Any aryl moiety present in such esters advantageously comprises a phenyl group. Any reference to any of the above compounds also includes a reference to a physiologically acceptable ialt thereof.
Salts according to the invention which may he conveniently used in therapy include physiologically ~cceptable base salts, eg derived from an appropriate base, such as alkali metal (e.g. sodium), alkaline earth metal (e.g. magnesium) salts, ammonium and NR4 (wherein R is Cl_4 alkyl) salts. When Y represents an amino group, salts include physiologically acceptable acid addi~ion salts, including the hydrochloride and acetate salts.
Such nucleosides and their derivatives will be hereinafter referred to as the compounds according to the invention. The term "active ingredient" as used hereafter, unless the context requires otherwise, refers to a compound according to the invention.
The present invention further includes:
a~ compounds according to ~he invention ~or use in the treatment or prophylaxis ~f viral infections particularly herpes virus infections such as those mentioned above and moxe particularly HSV, VZV or CMV infections.
b) a method for the treatment or prophylaxis of a herpes virus infection such as those mentioned above in a mammal including man, particularly HSV, VZV or CMV infection which comprises Uri:e~ t~ m P~tent O~ice ~ InrT~ ' '~~ ' '~~~
~ pn~

PCT/EB ~0/01398 - 206~2~
13 1 October 1991 31 ~ 91 ~
treating a subject with an effective non-tsxic amount of a compound according to the invention.
c) use of a compound according to the invention in the manufacture of a medicament for use in the tre~tment cr prophylaxis of a herpes virus infection, such as those mentioned above, particularly HSV, væv or CMV infections.
Examples of the clinical conditions which may be treated in accordance with the invention include those infections caused HSV 1 ~ 2, VZV, CMV or HBV described above.
We have fou~d that ~ompounds according to the invention have high oral bioa~ailability and low toxicity.
This provides compounds with a favourable therapeutic index.
The compound of formula (I) wherein X is methyl and Y
is hydroxy has been found to have good activity against HSV1 and 2 but to b~ toxic.
The compounds according to the invention may be administered to mammals including humans by any route appropriate to the condition to be treated, suitable routes including oral, rectal, nasal, topical (including buccal and ~ublingual), vaginal and parenteral (includi~g subcutaneouci, intramuscul~r, intravenous, intradermal, intrathecal and epidural). It will be appreciated that the preferred route may vary with, for example, the condition of the recipient.
~ 25 For each of the above-indicated utilities and indications the amount required of the individual active ingredients will depend upon a number of factors including the severity of the condition to be treated and tbe identity J~ ^ ~ C~J~T~ Tr, ~
;pj:~l ic~tic".

2~S327T9~B 90/01 ~9~
~1 Oclobes 19~1 :
3 1 ~0 31 of the recipient and will ultimately be at the discretion of the attendant physician. In general, however, for each of these utilities and indications, a suitable, effective dose will be in the range 0.1 to 250 mg per kilogram body weight

5 of recipient per day, preferably in the range 1 to 10~ mg per kilogr~m body weight per day and most preferably in the range 5 to 30 mg per kilogram body weight per day; an optimum dose is about 15 mg per kilogram body weight per day (unless otherwice indicated all weights of active ingredient are 10 calculated as the parent compound, for salts and esters thereof the figures would be increased proportionately.) The desired dose may if desired be presented as two, three, four or more sub-doses administered at appropriate intervals throughout the day. These sub-doses may be administered in lS unit dosage ~orms, for example, containing 10 to 1000 mg, preferably 20 to 500 mg and most preferably 100 to 400 m~ of - ;
active ingredient per unit dosage form.
While it is possible for the compounds to be ~dministered alone it is preferable to present them as 20 pharmaceutical formulations. The formulations of the present ~ invention comprise at least one active ingredient, ~ ~bove ~ -? defined, together with one or more acoeptabl~ carriers ~ thereo~ and optionally other therapeutic ingredients. The .. .
carrier(s) must be "acceptable" in the senise of being compatible with the other ingredients of the formulation and not deleterious to the recipients thereof.
: ' .

Pl~ B ~ C /
2 0 ~ 5 2 ~ 9 ~ l Oc~obes The formulations include those suitable ~or oral, rectal, nasal, topical (including buccal and sublingual), ; vaginal or parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural) administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any of the 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 formulations are prepared by uniformly and intimately bringins into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product.
Formulations of the present invention suitable for ! oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient: as a powder or granules: as a solution or a suspension in an ~queous liquid or a 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 pas$e.
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 ` ' '; '~ ~ SU~S~ITUTE S' ~

' 2 0 6 ~ 2 ~ 9 P~ 0 / G 1 ~ 9 ~ ~:

- 13 - ~31 OctobeS,~991 sui*able machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder (e.g. povid3ne, gelatin, hydroxypropylmethyl cellulose), -lubricant, inert diluent, preservative, disintergrant (e.g.
s~dium starch glyoolate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose), sur~ace-active or dispersing agent. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound m~istened with an inert liquid diluent. The tablets may optionally be coated or scored an may be formulated so as to provide 810W
or controlled release of the active ingredient therein using for example, hydroxpropylmethylcellulose in varying ~
proporti~ns to provide desired release profile. ~:
For infections o~ the eye or other external tissues, e.g., mouth and skin, the formulations are pre~erably applied as a topical ointment or cream containing the active ingrediPnt in an amount Gf, for example, 0.075 to 20% w/w, preferably 0.2 to 15~ w/w and most preferably 0.5 to 10% w/w. .:
When formula~ed in an ointment, the active ingredients may be 20 employed with either a paraffinic or a water-miscible ~ :
.
ointment base. Alternatively, the active ingredients may be -~ormul~ted in a cream with an oil-in-water cream base.
If desired, the a~ueous phase cf the cream base may include, for example, at least 30% w/w o~ a polyhydric alcohol, i.e~ an alcohol having two or more hydroxyl groups such as propylene glycol, butane-1,3-diol, mannitol, ::

~ ~ C~ J ~
.

'.',".''"'' .',."''".',.'," ~ .;` " :
,.. , .,, .... ", ! , .' ' . , . , . .. ' i, ; .. ' . ' , ~ , . ' ., . ~; ' . ' ' ' . ' . , . ' . ' , ' ' ' ' ' ' PCT/GB 90/0~9 2 0 6 5 2 ~ 9 ~1 oc~o~er 1991 - 14 ~ 0 31 sorbitol, glycerol and polyethylene glycol and mixtures , thereof. ~he topical formulations may desirably include a compound which enhances absorption ox penetration of the ; active ingredient throu~h the skin or other affected areas.
Examples of such dermal penetration enhancers include dimethylsulphoxide and related analogues.
The oily phase of the emulsions of this invention may be consti~uted from known ingredients in a known manner.
, While this phase may comprise merely an emulsifier (otherwise known as an emulgent), it desirably comprisPs a mixture of at .:
least one emulsifier with a fat or an oil or with both a fat and an oil. Preferably, a hydrophilic emulsifier is included together with a lipophilic emulsifier which acts as a sta~ilizer. It is also preferred to include both an oil and a fat. Together, the emulsifier(s) with or without stabilizer(s) make up the so-called emulsifying wax, and t~e wax toget~er with the oil and/or fat make up the so-called emulsifying ointment base which forms the oily dispersed phase of the cream formulations.
Emulgents and emul~ion stabilizers suitable for use in the formulation of the present invention include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl mono-stearate and sodium lauryl sulphate.
The choice of suitable oils or fats for the formulation is based on achieving the desired cosmetic -properties, since the solubility of the active compound in ~; rUn!l~d ~ d~;lt ~ S I ~-rT
f~T ~ n~.~.on~ ;3~i-at~ U~STITJTE 5....:.
7 ~ .~

P~ 1 099 20~ 3 2 7 gl O~o~e~ 19 31 lO 91 most oils likely to be used in pharmaceutical emulsion ~-formulations is very low. Thus the cream should preferably be a n~n-greasy, n~n-staining and washable product with ~uitable consistency to avoid leakage from tubes or other 5 containers. Straight or branched chain, mono- or dibasic ~lkyl esters such as di-isoadipate, i50cetyl stearate, propylene glycol diester ~f coconut fatty acids, isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitats ~r a blend ~ branched chain esters known as Crodamol CAP may be used, the last three being preferred esters. These may be used alone or in combination depending on the properties required. Alternatively, high melting point lipids such as white soft paraffin and/or liquid paraffin cr other mineral oil can be used.
Formulations suitable for topical administration to the eye also include eye drops wherein the active ingredient is dissolved or suspended in a suitable carrier, especially an aqueous sol~ent ~or the active ingredient. The active ingredient is preferably present in such formulations in a concentration of 0.5 to 20~, advantageously 0.5 to 10%
particularly 2bout 1.5% w/w.
Formulations suitable for topical administratlon in the mouth include lozenges comprising the active ingredient in a flavoured basis, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert basis such ~s ~elatin and glycerin, or ~ucrose and ~ 5 ~ r~ I s~'~r- 1 ~' 'T~ S' :-'T

," ' ' ' ' . . '- ,, "." ~. ' ' ' ' ', .'- ' , ' ' ' ' ~ " " ' '' ' . ,' : ' " ' ' '' . ' ' ' ',: , ' ''.' ' : ". . ' ' "

, '. ., , ;, . ' ' ' '` :.: ' . ' ' ' ' PGTI~ 90/~109~
~31 octo~eT 1991 - 16- 20~5279 .
~1 10 ~1 acacia; and mouth-washes comprising the active ingredient in a suitable liquid carrier.
:"
Formulations for rectal administration may be presented as a suppository with a suitable base comprising for example cocoa butter ~r a salicylate.
Formulations suitable for nasal administration wherein the carrier is a solid include a coarse powder having a particle size for example in the range 20 to 500 microns which is administered in the manner in which snuf is taken, i.e. by rapid inhalation through the nasal passage from a Gontainer of the powder held close up to the nose. Suitable formulations wherein the carrier is a liquid, for administration as for example a nasal spray or as nasal drops, include aqueous or oily s~lutions of the active ingredient.
Formulations suitable for vaginal administra~ion may be presented as pessaries, tampons, creams, gels, pastes, ~oams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.
~ ormulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats ~nd solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and no~-aqueous sterile ~uspensions which may include suspending agents and ~hickening agents, and liposomes or other microparticulate ~ SU~ST5TU~E S' ~

P&TI~ ~0/~1099 2 0 ~ 3 2 ~ October 1~91 ' systems which are designed to target the compound to blood Y! components or one ox more oryans. The formulations may be presented in unit-dose or multi-dose containers, for example -' sealed ampoules and vials, and may be stored in a ~reeze-dried (ly~philized) condition requiring only the addition of the sterile liquid carrier, f~r example water for injections, immediately prior to use. Injection solutions and suspensions may be prepared extemporaneously from sterile powders, granules and tablets of the kind previously described.
Preferred unit dosage formulations are those containing a daily dose or unit, daily sub-dose, as herein above recited, or an appropriate fraction thereof, of an active ingredient.
It should be understood that in addition to the ,' ' ingredients particularly mentioned a~ove the formulations of ~' this in~ention may include other agents convention~l in the ! art having regard to the type of formulation in question, for ~' example those suitable for oral administration may include flavGring agents.
The comp~unds of formula (I) may be produced by various methods known in the art of ~xganic chemistry in general and nu~leoside synthesis in particular. Starting ' materials are either known and readily,~vailable from ~ommercial sources ~r may themselves be produced by known and conventional techniques.
The pr~sent invention further provides a process for ~, m P~t~ ~ Sl~T, ~ ~ r ~. _T
~"l i...~.,,..~nal ~ c~

Pg~ 0 / ~ 9 .~ 1 october 1991 2Q~27~
producin~ a compound of formula (I) as hereinbefore defined which process c~mprises:
A)reacting a compound of formula (II) Y

N ~ - X
0~ ~

z50 1 !< y ~ .
~ '.
Oz3 wherein xl is a precursor for the group X as de~ined in relation to formula (I);
Y is as defined in relation to formula (I);
and Z3 and Z5 are the same or diff~rent and each is hydrogen or ~ hydroxyl-protecting group with a rea~ent or reagents serving to convert the group Xl to the desired group X:
B~ reacting a compound of formula (III) .

r~Jn..!c~ !'' . ~'om P~ t Off:~e Su S T! ~ 5! ~
L~!~ L; I ~ I J ~ . . .- .
.

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

2 ~ 5 ~ 2 7 9 - 1 9 - ~1 Oct~be~ 1~91 ~ 1 10 91 N~ '`';
I ~ X ~III) ~_\ ~ ..
N : :

wherein X and Y are as defined in relation to formula (I) or a protected form thereof with a 4-thio sugar compound serving to introduce the 4-thio sugar moiety, or a protected form ~-thereof, at the l-position of the compound of formula (III~;
5 a~d, where necessary or desired, thereafter effecting any one : .:
or more of the following further steps in any desired or necessary order:
a) removing each of the protecting groups, b) converting a compound of formula (I) or a protected form thereof into a further compound of formula ~I) or a protected form thereof, :
c) converting the compound of formula (I) or a protected ..
form thereof into a physiologically acceptable derivative of the compound of formula (I) or a protected form thereof, d) converting a physiologically acceptable derivative of the compound o~ formula (I) or a protected form ~ .:
thereof into the compound of formula (I) or a protected form, thereof, e) convertin~ a physiologically acceptable derivative of I Ijn~ )~i~ SU~ST`.T"TE S' '-,-T
rv~

P~TI~ 9D/~t ~59 ~ 1 Oct~ber 19~1 - 20 - 20~3279 ~1 10 91 ~ the compound of formula (I~ or a protected form ; thereo~ into another physiologically acceptable derivative of the compound of formula (I) or a protected form thereof, and f) where necessary, separating the ~ and ~ anomers of the compound of formula I or a protected derivative thereof or of a physioloqically acceptable derivative of a compound of formula (I) or a derivative thereof.
The term "4-thio sugar compound" is used herein to .
denote a compound containi~g the 2-deoxy-4-thio-D-ribofuranose ring wherein one or more of the hydroxyl groups thereof are optionally protected and wherein the l-position is optio~ally substituted by a leaving group.
Process B may be effected, for example, by ; a) reaction of the compound of formula (III), or a protected form thereof, with a 4-thio sugar compound of f ormula (IV).

Z /S\
: ~ ~ L (IV) \r-/ ~... .

~ wherein Z3 and Z5 are as defined abo~e and L is a leaving : ~roup, for example halogen, e.g. chlor~, ~cyloxy (e.g. Cl_6.~. . .
alkanoyloxy su~h as acetoxy), or S-benzyl. In formula (IV), the groups Z3 ~nd Z5 are preferably hydroxy protecting ~:: groups, particularly benzyl or tcluoyl groups. The reaction ,_ ____. , . I IJ,n~ r.1 P .o~t Qff.~,e r r. I~i~ - r~
PC~ ~U-,ST. ~ ~,T~

:

p~TI~ 2o/BlO~Y
2~6S27~ 91 - 21 - 3l October l9gl may be performed using standard methods including the use of a LRwis Acid ca~alyst such as mercuric chloride or bromide or stannic chloride or trimethylsilyltrifluoromethane-sulphonate in solvents such as acetonitrile, 1-2 dichloroethane, dichloromethane, chloroform or toluene at reduced, ambient or elevated temperature such as from -78 C
to reflux; or b) reaction of the compound Gf formula (III), or a protected form thereof, with a compound of formula (V) ';
ZS~ ~ (V~

z3 :

wherein Z3 and Z5 are as defined above and Py represents a pyrimidine base in the presence of a silylating agent such as N,O-~is-~trimethylsilyl)acetamide and in the presence of a ~:
Lewis Acid catalyst such as trimethylsilyltrifluoromethane :~
sulphatonate in a solvent such as acetonitrile. In the compound of formula (V), Py is preferably the uracil or thymine base.
T~e 4-thio-sugar compound may be produced ~y conven~tionsl methods prior to coupling with the ~ase or derived by modi~ication of another sugar moiety which is already part of a nu~leoside. Particular methods are as described in the Examples.

9 S.
L ~ ir~t!~n P~T/6~ 9 C/OI ~99 31 October 19il 20~32~9 3 1 10 91 Particular methods for producing the compounds of formula (I) in accordance with the above processes will be descri~ed below and these may be combined in order to produce further compounds within formula (I).
Reference may be made to the following texts:
Synthetic Procedures in Nucleic Acid Chemis~ry, Eds.
W.W. Zorbach R.S. Tipson, Vol. 1, Interscience, 1973;
Nucleic Acid Chemistry - Improved and New Synthetic Pr~cedures, Methods a~d Techniques, Eds. L.B. Townsend and ~.S.Tipson, Parts 1 and 2, Wiley-lnterscience, 1978 and Part 3, Wiley-Interscience, 1986;
Nucleoside Analogues-Ch~mistry, Biology and Medical Applicati~ns Eds R.T Walker, E. De Clercq ~ F. Eckstein, NAT0 AdYanced Study Institutes Series, Plenum Press, 1979;
Basic Principles in Nucleic Acid Chemistry, Eds. P.o.P
Ts'O, Academic Press, 1974.
With regard to the use of protecting groups as referred to above, it will be appreciated that the particular nature of such groups will b~ dependent on th~ identity and nature of the particular group(s) to be protected ~nd will ther-fore be selected in accordance with conventional tech~igues. Examples of pr~tecting groups that may be ~enerally used include acyl groups for example Cl_6 alkanoyl $(e.g. acetyl) or aroyl (e.g. benzoyl or toluoyl), ether groups such as tri-Cl_6alkylsilyl (e.g. trimethylsilyl) or tert-butyl diphenylsilyl; or arylmethyl groups such as henzyl ~ .i, .~`~ ~'J~ST~';J'T~
.. .. .. _ _ .... . _ , .. .. ~ :' ' ; ' ..... '.. ' ' . : . . .. ..

P~.T/GB Q O / ~ t ~9 2~5279 ~ t~r 1~9 ~ 1 10 91 . .' .
or triphenylmethyl groups.
The above groups may be removed in conventional manner, for example the acyl groups being removed advantageously under basic conditions (e.g. using sodium methoxide), the silyl ether groups being removed advantageously under aqueous or acidi~ conditions (e.g. using aqueous methanol to remove trimethylsilyl gr~ups) and the arylmethyl groups being removed advantageously under reducing conditions.
Protection of hydroxy groups with trialkylsilyl, eg. I
trimethylsilyl, grDups ~n the pyrimidine ring is conveniently achieved by reaction with (a) chlor~trimethylsilane together ~:
with triethylamine or with (b) hexamethyldisilazane, optionally together with chlorotrimethylsilane and/or ' 15 ammonium sulphate. . .
The following techniques are particularly convenient: .
X is.haloqen 5-Halopyrimidines are commercially available and may be coupled to the 4-thiosugar comp~und by conventional techniques, for instance by reacting a protected 5-halopyrimidine with a protected 4-thio sugar c~mpound having 2 leaving group in the l-p~siti~n. The leaving group on the 4-thio sugar compound may be a halogen, benzylthio or preferably acetate group.
Reac~ion of the protected 4-thio sugar.compound with.
; ~ the protected 5-halopyrimidine is c~nducted under conventional conditions using Lewis Acid catalysis such as by ~ ti~nal ~D~ r~ J~
.. .. . .. . ....... .

p~rl~B~Q/B~a 2~6~279 b 1 O~tober ~~t treatment with mercuric chloride or mercuric dibromide with cadmium carbonate or with stannic chloride, or preferably : ' ~
trimethylsilyltrifluoromethane sulphonate i~ toluene, acetonitrile, dichloromethane or l,i-dichloroethane, as ~olvent followed by treatment as neeessary with aqueous methanol (whi~h also serves to remove the protecting groups from any hydroxyls on the pyrimidine ring).
Protecting gr~ups may be removed by conventional techniques, for instance trimethylsilyl groups may be removed 10 from hydroxyl groups on the pyrimidine ring by treatment with :
aqueou~ methanol, benzyl gr~ups are removed from the hydroxyl groups on the 4-thio sugar compound by treat~ent with boron trichloride in dichloromethane at -78C, and p-toluyl groups are removed from the hydroxyl groups on the sugar by treatment with sodium methoxide in methanol at room :
temperature.
Alternatively the 5-halo substituent may be introduced into the pre-foxmed 5-unsubstituted 4'-thio-pyrimidine nucleosides having protected or unpr~tected hydroxyl group~.
When the hydroxyl groups on the 4-thio sugar compound : -are protected (f~r instance with ethers such as silyl ethers : or esters ~uch as acetate, benzoate or p-toluate ~sters), reaction with N-chlorosuccinimide in glacial acetic acid or ~E 25 with chlorine and iodobenzene and glacial acetic acid will .
introduce a 5-chloro substituent and reaction with iodine monochloride in dichlorometha~e will introduce a 5-iodo .
IJ~;it6~ , ;om r~ t~!~`t , ' " ' '; .' . ~ ; `

20 6 ~ 9 0 / 0 ~ 09~ :

- 25 - ~1 octobeI 1991 substituent, while reacting the unprotected 4~-thio sugar pyrimidine nucleoside with chlorine in carbon tetrachloride and acetic acid also introduces the 5-chloro substituent.
Reaction with iodine and nitric acid also introduces the 5-iodo ~ubstituent. Reaction with bromine and acetic acidintroduces a 5-~romo substituent to the unprotected nucleoside. Deprotection where nece~sary is by conventional techniques and is performed as the final step.
The 5-unsubstituted 4'-thionucleoside starting material of formula (II) may be produced as described ~bove by coupling a 5-unsubstituted pyrimidine to a 4-thio sugar compound. Protection of the hydroxy groups of the 4-thio sugar moiety may be effected at any convenient stage.
X is C2_6 alkynYl ~-Alkynyl compounds may be produced by reactinq a 5-iodo nucleoside of formula ~II) wherein the hydroxyl groups of the 4-thio sugar are optionally protected (for instance by reaction of the unprotected nucleoside with p-toluoylchloride~
in pyridine to introduce p-toluoyl ester groups on the hydroxyl groups of the 4-thio sugar) with an appropriate a}kynylating agent, for example trimethylsilyl acetylene or a terminal alkyne in the presence of a palladium catalyst such as ~is(triphenylphosphine~ palladium dichloride, and a ropper catalyst such as cuprous iodide and triethyl~mine and, where necessary, removal of the protectin~ ~roups using sodium methoxide in methanol [c.~. M.J. Robins et alr: Can. J. C~em., 60 554 (198~)].

~ Uri~d ~ ,n ~ rte~t C!.i,ce ¦ ~;

P~B~O~Q~
- 26 - 2 0 6 ~ 2 7 9 ctober ,~9, Alternatively the ~-alkynyl qroup may be introduced by reacting a 5-iodo pyrimidlne with trimethylsilyl-acetylene or a ~erminal alkyne in the presence of bis(triphenylphosphine)palladium dichloride, cuprous iodide, triethylamine and dimethylformamide ~ollowed, where ;necessary, by removal of the protecting groups and rea~ting the ~-alkynyl pyrimidine of formula (III) in suitably protected form (for instance the trimethylsilyl-protected ~orm) with a protected 4-thio sugar comp~und as previ~usly described f~llowed by deprotection of the pyrimidine and sugar moieties as required.
X is C2_6 alkenyl 5-Alkenyl compounds may be produced by partial hydrogenation of the corresponding 5-alkynyl pyrimidine of 1~ formula (III) or of the nucleoside of formula (II) for instance using Lindlar catalyst poisoned with quinoline, and subsequently, in the case of the pyrimidine, coupling with a 4-thio sugar compound as described above.
AlternatiVely a ~-iodo nucleoside of formula ~II) may 20 be reacted with an appropriate alkenylating agent for example ~-a 2-alkenoic acid ester (for instance the methyl ester) in ;
the presence of palladium (III acetate and triphenylphosphine to ~orm the 5-(2-methoxycarbonyl alkenyl) derivative. The ester group is then removed by hydrolysis using sodium hydroxide forming the 2-carboxy alkenyl compound which itself is subjected to treatment with triethylamine in dimethylformamide at 100C to give the 5-vinyl analogue [c.f.

PCT/GB 9 O / ~ 1 0 9 2 U ~ ~ 2 7 ~9 ~3 l oc.ob~r l~i~
- 27 - 31 lO 91 S.G. Rahim et al., Nucleic Acids Research, 10(17):5285(1982)].
Yet another method for producing the 5-alkenyl compounds involves coupling the terminal alkene with a 5-iodo cr 5-chloromercuri nucleoside of formul~ (II) (formed by ~or example reaction of the 5-unsubstituted nucleoside with mercury (II) acetate and sodium chloride), in the presence of a palladium catalyst such as palladium (II) acetate and a copper salt such as copper (I) chloride, or pr,eferably a paladium catalyst such as dilithium palladium tetrachloride.
Reaction of a 5-iodo- or 5-chloromercuri-nucleoside of formula (II) with allyl halides such as chloride or bromide in the presence of dilithium pa-lladium tetrachloride leads to the formation of the corresponding 5-(alk-2-enyl) derivative which can be rearranged to form the 5-(alk-1-enyl) derivatives by treatmen~ with tris(triphenylphosphine)rhodium chloride. This process may also be applied to the free ~-pyrimidine base, whioh i~ subsequently condense~ with the 4-thio sugar compound.
:.
The ~bove processes are exemplified by J.L. Ruth & D.E.
~ Bergstrom, ~_OrG~. Chem, 43 (14!: 2870 (1978), J. Goodchild i~ et gL., J. Med. Chem, ~: (1983), D.E. Bergstrom & J.L. Ruth, .; ~ .
J. ~m.~Chem. S~g., 98: 1587 (1976) and D.E. Bergstrom & M.K.

Ogawa, J. Am. Chem. Soc., 100: 8106 (1978).

2S X is C2_6 haloalkenyl~

(~alDalkenyl) substituents may ~e introduced into a nucleoside o~ formula (II) by conventional methods. For .!s'~ s'-~1 p,~ t O,~.~e PCTt~ ~ O/ ~1 O~

20~279 3 l lO 91 - 28 - '31 Octobes 1991 example, in order to preare 5-(2-halovinyl) compounds the corresponding 5-(2-carboxyvinyl) nucleoside is treated with --an appropriate halogenating agent, for example N-halosuccinimide in aqueous potassium acetate, or with potassium carbonate in dimethylformamide when the halogen is ; bromo or iodo. A 5-(2-chlorovinyl) nucleoside may also be made from the corresponding 5-(2-carboxyvinyl) nucleoside using chlorine gas in, for example, dimethylformamide (DMF).
Alternatively the 5-haloalkenyl group may be introduced into the appropriate free pyrimidine base to form a compound of formula (III) which is subsequently coupled with a 4-thio compound as described above; this may be achieved for example by treating a 2,4-dimethoxy-protected 5-iodo- pyrimidine with an 2-alkenoic acid ester in the presence of palladium (II) acetate, triphenylphosphine and dioxane follow~d by removal of the methoxy protecting groups, ' ~ :
hydrolysis of the ester with sodium hydroxide and reaction of ,~
the resulting 5-(2-carboxyvinyl) derivative with N-halosuccinimide (where halo is ~romo or iodo) or chlorine gas ' :
- 20 (where halo is chloro) in the presence of a base such as ,.,-sodium hydrogen carbonate in dimethylformamide. The 5~
carboxyvinyl) compound may also be produced by treating an unprotected 5-(hydroxymethyl)pyrimidine o~ formula (III) with an oxidisins agent such as persulphate or m~nganese dioxide :, to form the corresponding aldehyde and followed ~y treatment of the aldehyde with malonic acid. The above processes are ~:' exempli~ed by A.S. Jones et ~1, Te~rah,e,dro.n_Letts, 4~; 441 ~ n~ r ... ,. . .... . ,. ._.. ... .......

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

-- PC~/G~ 90/01 G8S
206a279 - 29 - ~1 Octobes 1~91 (1979) and P.J. Barr et al, J. Chem. Soc. Perkin Trans 1, 1981, 1665.
The 5-(2-haloalkenyl) base may alternatively be made by a novel route starting with a 2,4-dimethoxy protected 5-bromopyrimidine. This may be converted to the corresponding5-lithium derivative by treatment with an organolithium reagent, preferably n-butyllithium at reduced temperature such as -70~C in an ethereal solvent such as diethylether.
Reaction of the llthio derivative i~ situ with an appropriate ester of formic acid, such as ethyl forma~e at reduced temperature such as -7~c gives rise to the corresponding 5-formyl compound. Treatment of the formyl compound with malonic acid as described ab~ve qive rise to the 5-(2-carboxyvinyl) derivative. Similar halogenation gives rise to the required 5-(2-haloalkenyl) compound which is in the 2,4-dimethoxy protected from~ Deprotection can then be carried out by conventional techniques.
5-Halovinyl compounds having more than one halogen I substituen~ may be produced from a 5-halo-substitued 2,4-dimethoxy protected pyrimidine of formula (III) by reaction with a strong base such as butyl lithium and the resulting lithio derivative treated with the appropriate haloalkene followed by removal of the protecting groups ~nd coupling to the 4-thio sugar compound as described above [c.f. P.L. Coe et al., J. Med. Chem. ~5:1329 (1982)].
Alternatively, the halogen atoms may be introduced se~uentially into a 5-substituent of the pyrimidine base.

~Uni;.~,d l' ^ :dc~m P~tent 0'.~"`9 ~

PCr/GB ~ t n g ~ .

2 0 ~ ~ 2 ~ October 1991 ~o 31 lD 91 Thus, ~or example treatmen~ of 5-acetyl uracil with a chlorinating agent such as phosphorus oxychloride provides the 5-(1-chlorovinyl) group wi~h simultaneous chlorination of the hydroxyl groups of the pyrimidine base. Treatment with potassium ethoxide then hydrogen chloride and finally bromine leads to bromination of the 5-unsaturated side chain of thé
pyrimidine base with simultaneous converr,ion of the 2,4-dichloro groups on the pyximidine ring to form the corresponding uracil derivative. The resulting pyrimidine base can then be coupled to the 4-thio sugar compound as described above [c.f. P.J. Barr et al. Nucleic Acids Res. 3:
2845 (1976) and P.J~ 8arr et al., J. Chem. Soc. Perkin Trans 1, 1981; 1665].
X is C2_6~ yl 5-C2_6 Alkyl eg. 5-ethyl substituted nucleosides may ~ ;
be produced by hydrogenation of the corr~sponding S-alkynyl or 5-alkenyl pyrimidine base followed by coupling to the 4-thio sugar compound. Conventional hydrogenation conditions, ~uch as hydrogen w er palladium/charco~l catalysts, may be 2 n adopted.
X is tri~luoro~ethyl -5-Trifluoromethyl uracil is commercially available and this may be condensed with a 4-thio ~ugar comp~und in accordance wi~h process B described above. The 5-trifluoromethyl cytosine analogue may be made from the uracilcompound/using a~ analogous procedure to th~t described by Sung as mentioned below.

S U
~ v~~ p,,"~ n I .

... . . . . . . ..

-- PCT/~ 90/01099 206~279 ~1 ~ctobe~ 199l The above reactions are all suitable for producing uracil nucleosides; most of such reactions may also be used to form cytosine nucleosides. When this is not convenient or possible, cytosine analogues can be prepared most conveniently from the uracil compounds using an analogous procedure to that described by W.L. ~ung, J. Chem. Soc. Chem.
Commum.. 1981, 1089]: for example the acetylated uracil nucleoside (produced for instance by rsactions as described above and acetylated using acetic anhydrid~ in pyridine) is ~;~
treated with p-chlorophenylphosphorodichloridate, 1,2,4-triazole and pyridine t~ produce the 4-iil,2,4-triazol-1-yl) derivative which is then treated with ammonia in dioxane (which also removes the 4-thio sugar protecting group(s)) to form the corresp~nding unprotected cytosine 4'-thionucleoside.
The derivatives of the compounds of formula (I) maybe prepared in conve~tional manner. For exampie, esters may be prepared by treating a compound of formula (I) with an appropria~e esterifying agent, for example, an acyl halide or anhydride. Salts may be prepared by treating a compound of formula ~I) with an appropriate bii se, for example an alkali metal, ~lkaline earth meta~ or ammonium hydroxide, or where necessary, ~n appropirate acid, such as hydr~cihloric acid or ~
an acetate, eg. sodium ac~tate.
The anomers of compounds of the formula I may be ~i : , -.
~ eparated by c~nventional means, for example by r"'~ 2~ ^r~ 2~q~ Off~e~ S~B~TI~

., ..; . i ~ tâ; ~ ~ n ~ . . . _ . ... _ .
- .

6 ~ 9 PCT/6B 9 O / ~ 1 09 - 32 - ~1 Octc~r 199l chromatography or fractional crystallisation.

The invention further relates to the intermediate .
' compounds of formula (IV') ` .

Z5 S -;

~ ~ L (I~) ~I .~
dZ3 ~::

.;
where Z3 and Z5 are benzyl and L is acetoxy, as well as compounds of formula (IVA) 1/ \ . :
S-CH2-Ar (IV) 3 ~ :
, .. :. ' , ` ' .
where Z3 and Z are hydroxy protecting groups and Ar is an optionally substituted aryl group. :
The compounds of formula (IV') are obtained by :
reaction of a oompound of formula (IVA) with an appropriate acylating agent such as acetic anhydride (optionally in the ,~ - . .
~ presence of acetic acid), in the presence of a mineral acid ~ .... .

~ . .
Lln,~ P~t O,~'ce ~ .. ..,;;i A,~;~.ica.ion -~ :
~3 ::

-~ 206S279 PCT/GB ~O/Ot O99 ~1 Oct9ber 1991 - 33 - 3 1 ~ 91 such as sulphuric acid.
The compounds of formula (IVA) may be produced by ring closure under basic conditions of ~e compound of formula (VI) Ar-CH2-S\ ~S-CH2-Ar ~ ~ ' C , f 2 ~c-oz3 (VI) A-O-CH
CH20Z5 ' :
, wherein Z3 and Z5 are hydroxyl protecting groups, Ar is an optionally substituted aryl group and A is an optionally :
: substituted alkyl- or aryl sulphonyl group. Preferably in the compound of formula (IVA), Z~ and Z5 are benzyl and Ar is phenyl.
.
: The compounds of formula (VI) may be produced by treating a compound of formula (VII) ~ :

Ar-CH2 -S \ f -CH2 -Ar ~ CH

-oz3 (VII) HO-CH
H20Zi5 wherein~Z3 and Z5 are hydroxy protecting groups and Ar is an 2 ~ t 20~;32~i~9 PC~/G~ ~Bl~t ~95 ~1 Ottobe~ 1991 _ 34 _ 3 1 1~ 91 optionally substituted aryl group with an optionally substituted alkyl- or aryl- sulphonyl halide; the compound of formula (VII) is optionally made by reaction of a compound of formula (VIII) Ar-CH2-S~_ S-CH2-Ar H ::

I (VIII) f 5 C}~20Z :
wherein Ar, Z3 and Z5 are as defined above, and M is a :
hydroxyl protecting group, under conditions which remove the group M and leave the groups -S-CH2-Ar, Z3 an~ Z5 in place;
the compound of formula (VIII) is optionally made by reaction .
of a compound of formula (IX) Ar-CH2- ~ S-CH2-Ar H
FH2 ' ' ~ ' ' '' HC-oz3 HC-OH (IX) CH20Z5 ' ~ ' .
wherein Ar, Z3 and Z5 are as defined above, by reaction with a derivative of the group M under neutral conditions in a ~; polar solvent; the compound of formula ~IX) is optionally made by reaction of a compound of formula (X) ' r~, ~ r-~ T~JTr~ T

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

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

~ 2~2~1Q~ ~ O / ~ t ~39~
~. .
l3 1 October 1091 Z50 _ S
.. ~ \~_,oR (~) oZ3 wherein 23 and Z5 are as defined above and ~ is a Cl_4 hydrocarbyl group, with a compound of formula Ar-CH2-SH where Ar is as defined above, under acidic conditions at elevated temperature; and the compound of formula ~X) is optionally made reaction of a compound for formula (XI) Ho -~ o ~ ~ R (XI) ''`1, ~

~1 H

wherein R is de~ined above, with reactive derivativ~is of the groups Z3 and Z5 in an organic solvent in the presence of a I suitable ~se, wh~rein the compound of ~ormula (XI) is made by reiction of 2-deoxy-D-ribose with an alcohol of f~rmula ~-~ 9H in the presence ~f an acid.

,1l ~, .
i, .

~ .._ um Pat~:lt C~f!ce el ~c~r n~l Appllc3tl0:~ ~ -J L~
''~ ~ : ' ~: . .

PGT/~B ~ 0/01 U9' 206a279 ~ Oclober 1991 In a further aspect of the invention, compounds of the formula (IV) may be made by ring closure of a compound of the formula (VI~

Ar-CH2- ~ ~ -CH2-Ar ~-'.

fH2 (VI) HC-oz3 A-O-CH

where Z3 and 25 are hydroxyl protecting groups such as benzyl optionally substituted on ~he phenyl ring by one or more halogen atoms, Cl_4 alkyl eg. methyl, cl_4 haloalkyl, Cl_4 alkoxy, nitro or amino groups. The group A is a leaving group, for example an organosulphonyl group such as an optionally substituted alkyl- or aryl- sulphonyl group, for ...
instance methanesulphonyl, a haloalkylsulphonyl group (eg. ::~
trifluoromethyl-sulphonyl) and optio~ally substituted phenylsulphonyl (eg. toluylsulphonyl or brom~benzenesulphonyl), and Ar is an optionally substituted aryl group, for example optionally substituted phenyl or toluyl. optional substituents o~ the aryl yroups include one ~r more halogen atoms, Cl_4 alkyl ~g. methyl, C1_4 haloalkyl, C1_4 alkoxy, nitro or amino groups. The ring closure may be E p2rformed under appropriate basic conditions. Suitable conditions include those described by J. Harness and N.A.
Hughes (Chem. Comm. 1971, 811), which includes the use o~ :
sodium iodide and_~arium_carho~ate.

... ~c~ ,. a~
~' . . ... . . . ... .. . .. . .

20652~9PG~/GB ~ Q / o ~ 09 ~
~1 October Iq91 - 37 - ~1 10 91 The compound of the formula (VI) may be made from a compound of formula (VII) -Ar-CH2- ~ S-CH2-Ar H

Hof_oZ3 (VII) H-O-CH

where Ar, Z3 and Z5 are as defined above. Conversion of a compound of formula (~II) to a compound of formula (VI) is carried out according to standard procedures such as treatment with an appropriate optionally substituted alkyl-or aryl- sulphonyl halide, eg. methanesulphonylchloride in a basic solvent such as pyridine.
The compound of formula (VII) may be made from a compound of formula (VIII):

Ar-CH2-S\ S-CH2-Ar C}~
CH~

Hf-OZ3 (~III) M-O-CH

where Ar, Z3 and Z5 are as defined a~ove and M is a hydroxyl protecting group which may be removed under conditions which leave the -S-CH2-Ar groups and the groups Z3 and Z5 in place.
Preferably, the group M is a group of the formula Arl-CO- where Ar1 is a phenyl group which may be cptionally substituted ~y any of the ~ubstituents described above for p~ o~ o~
~1 10 91 6 ~ 2 7 9 131 October 1991 the group Ar. Removal of the group M may be performed under standard conditions, for example with a base such as an alkali metal alkoxide, for instance sodium methoxide in methanol.
The compounds of formula ~VIII) may be ~btained by the concomitant inversion and derivatization of the 4-hydroxy group of a compound of formula (IX) Ar-C~2-S \ f -CH2-Ar ¦
fH2 HC-oZ3 (IX) HC-OH

wherein Ar, Z3 and Z5 are as defined abo~e. The inversion and derivatiza,tion may be effected by reacting the compound of foxmula ~IX) with a derivative of the group M, such as an acid of the formula Arl-COOH, for example benzoic acid (or a reactive derivative thereof) where Arl is as defined abo~e. ~-The reaction is performed typically at ro~m temperature and under neutral conditions in a suitable polar solvent, ~or instance tetrahydrofuran. Preferably the Mitsunobu reaction i8 used for the inversion and derivatization, diethyl azodicarboxylate (DEAD) and triphenylphosphine are used as coreactants together with the acid ArlCOOH.
The compound of formula (IX) may be màde from a glycoside compou~d of formula (X) , - -' ~ 0 ~ ~ 2 7 9P~T/~ ~ O l l~
39 ~1 OCtober 1991 Z5O ~ O
~R (X) dZ3 wh~re Z3 and Z5 are as defined above And R is a Cl_4 hydrocarbyl group, eg. a Cl_4 alkyl group, preferably methyl.
The compound of the formula IX is reacted under acid conditions at an elevated temperature with a compound of ~ormula Ar-CH2-SH, where Ar is as defined above. Suitably, hydrochloric acid is used as the acid which may be in aqueous or anhydrous form. Prefera~ly the elevated temperature is from 30C to 60~C, f~r example 40OC. When Ar i5 a phenyl group, the c~mpound Ar-CH2-SH will be benzyl thiol.
Comp~unds of the formula (X) may be made from a compound of ~ormula (XI) HO ~ / o\
R tXI) ' bH
where R is a defined above. The hydroxyl groups of the compound of formula tXI) are protected under conventional conditions with the reactive derivative of the groups Z3 and Z5. Suitably, the bromo derivative may be used. Thus when Z3 and P5 are benzyl groups, benzyl ~romide may be used. The reaction may be per~ormed in an organic solvent ~uch as tetr~hydrofuran in the presence of a suitable base such as sodium hydride and a phase transfer catalyst such as tetrabutylam~onium iodide.

n~=.................................. .. .
~ ~CT ;;..-..;i.~,~2!,~

P~T/f'f~B ~ 0 ~ 1~ 7 ~
- 20~a279 31 10 91 ~f I Ocl'ober 1 ~f9 Ccfmpounds of the formula (XI) may be made by standard techniques fxom 2-Deoxy-D-ribose, which is commercially available. 2-Deoxy-D-ribose may be reacted with an alcohol ;
of ~ormula R-OH (where R is as defined above) in the presence of an ~cid. ~ydrochlorif- acid is suitable. When 2 is a methyl group, the alcohol R-O~ will be methanol.
The conversion of 2-deoxy-D-ribose to a compound c~f formula (XI) will also produce a small prcfportion of the corresponding pyranoside compound, substituted at the 1-position by the group -OR. This may remain in the reactiofn mixture during the converions of (XI) to ~X), (X) to (IX) and the subsequent reactions described above and it will undergo analogous r actions. These by-products may be separated at any convenient step by conventional means, eg.
chromatography.
The compound of the formula VII may also be made directly from the compound of formula IX using a Mitsunobu reaction under conditions analogous to those described by D.R. Williams et al, JACS 51990) 112, 4552~

' .

~, .

.

: 20~a2~T/~B 90/~1G99 ~1 O~ober 1~91 ~ he invention is illustrated by the following non-limiting Examples Example A
Preparation o~ Methyl 3.5-di-0-benzyl-2-deoxv-D-erythE~o-Dentoside To a solution of 2-deoxy-D-ribose (SOg, 373 mmol) in dry methanol (9OOml) was added a 1~ solution of dry hydrogen chloride in methanol (lOOml). The mixture was kept in a ~- stoppered flask for 30 minutes after which the reaction was stopped by adding, with vigorous stirring, silver carb~na~e (lOg). The mixture was filtered by gravity and the colourless filtrate evaporated to a syrup using a dry rotary evaporator. ~esidual methanol was then removed by repeated ~: evaporation with dry THF. The syrup was then dissolved in dry THF (47~ml). Under an atmosphere of dry nitrogen, at O D C, with stirring sodium hydride in a 50% oil-dispersion ; (39.4g, 821 mmol) was slowly added to the THF mixture. Next, ~ -dry tetrabutylammonium iodide (30 3g, 82.1 mmol) was added followed by b~nzyl bromide (140g, 821 mmol~, which was added over 1 hour. After stirring for 60 hours at room temperature, with exclusion of moisture, TLC (hexane-ethyl acetate [4:1]) showed almost complete conversion to two ~aster moving components (Rf 0.47 and 0.36). The THF was . ( .
removed i~ vacuo, the residue dissolved in dichloromethane and then poured into ice/water. The dichloromethane solution was extracted from this mixture and then dried over magnesium sulphate. The dichloromethane was evaporated under reduced PCT/GB 90J~1 ~99:
29~5279 bl o~obe~ 199 - 42 - ~1 10 91 pressure and the resulting residue applied tu a silica gel column eluted with hexane-ethyl acetate ~4:1). Combination of the appropriate fractions gave the ~ (RfO.36) and ~
(R~0.47) isomers of the title product as a clear, colourless syrup ~MR SPECTRA
- isomer ~1H) ~ (d6DMSO):7.56 - 7.17 (lOH,d,aromatic), 5.12- ~ `
5.00 ~lH,q;H-l), 4.60-4.45 (4H,m, PhCH20), 4.40-3.86 (2H,m,X'-3, H-4), 3.58-3.42 (2H,d,H-5), 3.40 (3H,s,CH3),2.40-1.80 (2H,m,H-2).
(13C) ~ (CDC13): 128.3-127.6 (aromatic), 105.2 (C-1), 82.1 (C-3 or C-4), 78.6 (C-3 or C-4), ~3;4 (PhCH20), 71.5 ,:
(PhCH20), 70.2 (C-5~, 55.1 (OMe), 38.9 (C-2).
~ - isome~
(lH) ~ (d6DMSO):7.50 - 7.20 (lOH,d,aromatic), 5.18-5.02 (lH,q,H-1), 4.65-4.43, (4H,d, PhCH20), 4.43-4.00 ~2H,m,H-3,H-4), 3.60-3.42 (2H,m,H-5)l 3.30 (3H,s,CH3), 2.45-2.05 (2H,m,H-2).
: 20 (13C) ~ (CDCl3):128.3-12?.6 (aromatic), 105.4 tC-l), 82.8 (C-3 or C-4), 80.0 (C-3 or C-4), 73.3 (Ph~20), 72.0 :~
(PhC~20), 70.2 (C-5), 54.9 (OMe), 39.3 (C-2).
: : Preparation of 3.5-di-0-ben~yl -2-deoxy-D-erythro-pen~ose ~- 25 dibenzyl dithioacetal Concentrated hydrochloric acid (lSOml) was added dropwise to a stirred mixture of methyl 3,5-di-Q-~enzyl-2-deoxy D-erythro-pentoside (77-5S~, 236 mmol) and benzyl thiol DC~ .r ~ .r A~

P~T/GB 90/01099 206~279 ~1 October 1991 - 43 - 31 1~ 91 (147g, l.l9 mol) at room temperature. The temperature was then raised to 40 C and the mixture ~tirred for 18 hours. At the end of this time TLC (hexane-ethyl acetate [4:l]) showed two faster moving minor component~ (~f 0.58 and 0.53), a major component (~fo.2s) and a slower moving minor component (R~0.22). The mixture was dissolved in chloroform, poured into ice/water, neutralised with sodium hydrogen carbonate and extracted with chloroform. The chloroform extracts were dried over magnesium sulphate and the chloroform was evaporated under reduced pressure. The residue was applied to a silica gel column which was eluted with hexane-ethyl acetate (4:l). The first component to be eluted from the column, as a clear colourless syrup was the ~, B anomers of ; benzvl 3.5-di-0-benzvl-2-deoxy-l-thio-D-erythro-lS pentofuranoside.
NMR SPECTRUM
(lH) ~ (d6DMSO):7.43-7~15 (15H,m,aromatic), 5.12-4.94 (lH,m,H l), 4~.59-4.36 (4H,m,PhCH20), 4.12-3.30 (6H,m,H-; 3,H-4,H-5,PhÇ~2S), 2.35-1.35 (2H,m,H-2).
~LEMENTAL ANALy~IS
~ounB: C,74.4; H,6.5. C26H2803S requires C,74.3;
H,6.7%
MASS SP~CTRUM
m/z 420 M+, 297 [~-S8n]+, 3-~oba matrix.
The title product was the second component to be eluted from the column as a clear syrup (lOgg,~%).
~MR SPECTRUM

C~ r - -I P~T l..~ IaL-o~ r~,~lication ~1 ' ~';~''`` ~ ~ -.- ' ;.. ' '. .. ' ',''. ' ,` '".,`'.. ' PCT/~ 9 G f 2 0 6 a 2 7 9~ 1 ~ctobe~ 199 ~ t lû
(lH~ ~ (d6DMSO)o7.35~7.05 (20~,m,aromatic), 4.97-4.g5(lH,d,OH-4), 4.47-3.95 (4H,m,PhCH20),3.81-3.66 :1 (7H,m,H-l,H-3,~-4, PhCH2S), 3.44-3.32 (2H,d,~-5), 2.10-1.83 (2H,m,H-2).

Found: C,73.0; H,6.5. C33H3603S2 requires C,72.8: ~ -H,6.7%.
MASS SPECTRUM
m/z 298 [M-2.SBn]~, 3-noba matrix.
SP~CIFIC ROTATION
[~)25 = -101.~ (c1.2 in EtOH) :

preparation of 4-0-benzoyl 3.5-di-0-benz~1-2-deoxy-L-thre~
pentose dibenzyl dithioacetal To a solution of 3,5-di-Q-benzyl-2-deoxy-~-eryt~kro-pentose dibenzyl dithioacetal (54.1g,99.3 mmol),triphenyl-phosphine (39.1g, 149 mmol) and benzoic acid (18.2g, 149 mmol) in dry THF (800ml) was added a solution of DEAD (26.0g, 149 mmol) in dry THP (200ml) dropwise, with stirriny, a~ room ~emperature.
After stirring at room temperatur~ for ~8 hours, TLC
(hexane-et~yl acetate [4:1~) revealed a faster moving component (Rf 0.56) and slower moving starting material (Rf 0.36~. The THF was removed in ~3ç~_ and the residue applied to a silica gel column eluted w-ith hexane-ethyl acetate (85:15). Co~bination of the appropriate fractions gave the title product as a white solid. Starting material could also ' ~.r ~ ..,c~l A p'l~,atton I

PCr/~ 9~/~1o99 206a27~ 3~ 10 ~.1 _ 45 _ ~1 October 1991 be recovered.
NMR SPEC~UM
(lH) ~ (d6DMS0):7.99-6.~8 (25H,m,aromatic), 5.39-5.22 (lH,m,H-4), 4.54-4.04 (4H,m,PhCH20), 4.01-3.62 (8H,m,H-l,H-3,H-5,PhCH25), 2.17-1.84 (2H,m,H-2).
ELEMENTAL A~LYSIS
FQund: C,74.3; H,6.4. C40H4004S2 requires C,74.0;
H,6.2% MASS SPECTRUM
m/z 525 [M-SBn]+, 435 [M-2.SBn]+, glycerol matrix.
SPECIFIC R~TATION

[~]2~ = 51.6~ (cO.7 in CH2C12).
D
; Preparation of 3.5-di-~-benzyl-2-deoxv - L - threo-~entose dibenzvl dithioacetal To a solution of 4-0-benzoyl-3,5-di-0-benzyl-2-deoxy-L-threo-pentose dibenzyl dithioacetal (8~.8g, 137 mmol) in dichloromethane (500ml) was added a solution of sodium methoxide (ll.lg, 20Ç mmol) in methanol (205ml) dr~pwi~e, with stirring, at OsC. The reaction mixture was then allowed t~ warm to ro~m temperature ~ver ~ period of 3 hours. At the end of this time TLC ~hexane-ethyl acetate t4-1]) revealed complete convexsion t~ a slower m~ving component ~Rf Q.31). The mixture was then poured int~ a 5~
solution o~ NaH2P04 and extracted with dichloromethane. The dichloromethane extract ~ ere then washed with a 5% solution of sodium hydrogen carbonate and water, dried ~magnesium sulphate) and evaporated. The crude title pr~duct was ~ p.~ ,~ n I ` ~ ' ` `

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

PCT/~B ~ 0/~1 0 - 46 - ~1 Octobes 1991 applied to a silica gel column eluted with hexane-ethyl acetate (4:1~. Combination of the appropriate fractions gave the title product as a clear colourless syrup.

(1H) ~ (d6DMSO):7.34-7.06 (20H,m,aromatic), 4.88-4.86 (lH,d,OH-4), 4.55-4.00 (4H,m,PhCH20~, 4.83-3.32 (9H,m, H-l,H-3,H-4,H-5, PhCH25), 2~08-1.84 (2H,m,H-2).
ELEMENTAL ANA~YSIS

Found: C,72.6: H,6.9. C33H3603S2 re~uires C,72.8;

! 10 H,6.7~
MASS SPECTRUM
m/z 297 [M-2.SBn ~ H)~, glycerol matrix.
SP~CIFIC ROTATION ~-[~]25 = -75.6~ (c1.9 in EtOH).
Preparation of 3 5-di-0-benzyl-2-deoxy-4-0-methanç-sul~honvl-L-threo-pentose dib~nzyl dithioace~al To a solution of 3,5-di-Q-benzyl-2-deoxy - L - threo-pentose dibenzyl dithi~acetal (61.4g, 113 mmol) in dry pyridine (700ml) was added methanesulphonyl chloride (19.4g, 169 mm~l) in dry pyridine (200ml~ dropwise, with stirring, at 0'C. The temperature ~f the mixture was raised to room temperature and stirring continued for 18 hour~. The pyridine was then removed in Yacuo and the residue dissolved in dichlorometha~e. The dichloromethane extracts were then successively washed with 2M hydrochlcric acid, 1~ scdium - carbonate and water, dried ~magnesium sulphate) and ~' ~ S~ U ,. _ .~T
L~ qtional Applicatlon ' , ' . ' ~ ' . ' , ' ' . : ' ., . ! ', 20$5279 PC~/~B a ~
31 lo 91 _ 47 _ ~31 O~tober 1991 evaporated t~ give the title product as a thick viscous syrup. ~ sample of this was crystallised from hexane~ethyl acetate to ~ive the title product as white crystals, m.p 82-83-C.
N~R SPECTRUM
(lH) ~ (d6DMSO): 7.64-6.89 (20H,m,aromatic), 4.88-4.64 .(lH,m,H-4),4.60-4.09 t4H,m,PhCH20), 4-05-3-43 (8H,m,H~ -3,H-5,PhCH2S), 3.11 (3H,s,CH3), 2.12-1.80 (2H,m,H-2) 1 0 " , EL~MENTAL ANALYSIS
Found: C,65.5; H,6.1 C34H3805S3 requires C,65.6;
H,6.2~
.:
~ ~ASS SPEC~RUM
:! 15 m/z 499 [M-SBn]+, 393 [M-SBn-OBn ~ H]+, glycerol matrix.
SPECI~IC ~OTA~ON
[~25 = -58.4' (c2.4 in CH2C12).

PreparatiDn of benzyl 3.5-di-0-benzyl-2-deoxy 1.4-dit~io-p-ervthro~pentofuranoside A ~uspension of 3,~-di-0-benzyl-2-deoxy-4-o-methanesulphonyl L-threo-pentose dibenzyl dithioacietal (29.4g, ~ .4 mmol), sodium iodide (74.0g, 494 mmol), barium car~o~ate (148g, 750 mmol) and dry acetone (lL) was boiled under re~lux for 42 hours. At the end cf this time the ~:

= ~`''''~

,. . .. ..... ..... . . . .... ... . . . .. . . ..... ..
. ! , , , . . ' ' . . ~ . . ` . , ., ', ' . ~ , . . ' , .

PC~/g~ 9 ~/o1 o~

206~279 bl3bcto~r 19911 suspension was filtered and the s~lids were washed with chloroform. The filtrate was sequentially washed with water, ~odium thiosulphate solution (5%) and water, dried (magnesium sulphate) and evaporated. The resultant residue was applied to a silica gel c~lumn, eluted with hexane-ethyl acetate (9:1j. Combination of the appropriate fracti~ns gave the title product as a clear, sliqhtly yellow, syrup, and recovered starting material. ~ -~MR SPECTRA
(lH) ~ (d6DMS0):7.50-7.12 (lSH,m,ar~matic), 4.66-4.13 (6H,m,H-l,H-4,PhCH~0), 4.09-3.35 (SH,m,H-3,H-~,PhCH2S), 2.44-1.94 (2H,m,H-2).
~aior anomer (13C) ~ (CDC13):129.0-127.1 (aromatic), 83.04 (C-l), 73.1 (PhCH20), 73.1 (PhCH20), 71.0 (C-3), 53.2 (PhCH2S), 49.9 (C-4~, 41.3 (C-5), 37.0 (C-2).
Minor a~omer (13C) ~ (CDC13):129.0-127.1 (aromatic), 82.7 (C-l), 72.9 (PhCH20), 72.9 (PhCH20), 71.6 (C-3), 53.0 (PhS~2S), 49.0 (C-4), 41.0 (C-5), 37.0 (C-2).
ELEMENTAL ANALYSIS
Pound C~7l~8;H~6-7;sll4-4- C26H282S2 requires C,71.5;H,6.5;5/14.7%
. ~ ~ASS SP CTRy~
~ m/z 437 [M+H3+, 345 [M-Bn]+, 329 [M-9Bn]+, 313 [M-S3n~, 223 ~M-SBn-Bn+H]+, glycerol matrix.

I PCT Inir..:~t~ ~

- PCTIGB ~ O I ~ ~ O 9 9 ~ 7 ~ ~ l Oc~o~er 1~91 _ 49 _ 3~ 1~ 91 `

~reparation of 3' 5'-di-0-benzyl~4'-thio-thvmidine and its ~-anomer A suspension of benzyl 3,5-di-Q-benzyl-2-deoxy-1,4-dithio-D-ervthro-p~ntofuranoside (22.~g, ~1.6 m~ol), bis TMS-S thymine (46g, 17~ mmol), mercuric bromide (20.5g, 56.7 mmol),cadmium carbonate (29.3g, 170 mmol) and dry toluene (lL) was boiled under reflux, with stiring, for 24 hours. The hot mixture was then filtered and the solids were washed with ;
toluene. The filtrate was successively washed with potassium iodide solution (30%) and wat~r and then evaporated. The residue was taken up in 4:1 methanol-water, stirred for 30 minutes, the suspension filtered and the filtrate e~aporated.
The residue was applied to a silica gel column (hexane-ethyl acetate (1:1)) and combination of the appropriate ~ractions gave the title product as a cl~ar colourless syrup. lX NMR
indicated the ratio of ~- to B- anomers to be 2.8:1. Purther column chromatography gave more of the ~eparated anomers.
The first compound to be eluted from the olumn was 3'.5'-~i-0-benzyl-4'-thio-thym~dine as a c~lourless syrup. This could be crystallised from methanol to give colourless crystals m.p. 1~0-142~C.
NMR SPEC~
(lH) ~ (d6DMS0):11.36 (lH,s,N~ .69 (lH,s,H-6),7.48-7.22 (lOH,mtaromatic), 6.33-6.~7 (lH,t,H-l') 4.61-4.51 (4~,m,Ph~20), 4 30 (lH,s,~-3'), 3.76-3.66 (3H,m,H-4',H-5')2.42-2.32 (2H,m,~-2),1.66 [3H,s,CH3).

¦ U~ited Kin~dom P;t;3~t Ofr:c ~ C~ r-r-F"

.

PCT/~B 9 Gl~l ~9 2~527g ~1 octOh~ 1991 31 10 ~
UV SPECTRUM
max 269.1nm (~,14,300).
` ELEMENTAL ANALYSIS
Found: C,66.0;H,6.0;N,6-3- C24H26N204S requires S C,65.7;H,6.0;N,6.4%
MASS SPECTRUM
m/z 439 [M+H]+, 347 [M-Bn]~, 331 [M-OBn]~, 3-noba matrix.
The next component to be eluted from the column was the ~-anomer, as a cslourless syrup.
NMR SPECTR~
(lH) ~ (d6DMSO): 11.28 (lH,s,NH), 7.95 (lH,s,H-6),

7.43-7.20 (lOH,m, aromatic), 6.25-6~21 (lH,d,H-l'), 4.66-4.47 (7H,m, Ph C~20), 4.25 (lH,s,H-3'), 4.10-4.06 (lH,m,H-4~), 3.57-3.42 (2H,~,H-5'), 2.68-2.26 (2H,m,H-2'), 1.55 (3H,s, CH3)-UV SPECTRUM
Max 268.1 nm (~, 10,300).
ELEMENTAL ANA~ S
Founa: C,65.4; H,6.1;N,6.7:S,7.4. C24H26N2~4S
re~uires C,65.7; H, 6.0; N 6.4; S,7.3~.
MASS SPEIB2~
m/z 439 [M+H]+, 461 [M+Na]+, 3-noba matrix.
Pr~paration of ~ -4'-thio-thymidin~
, ~ ~5 To a 2M boron trichloride solution in dry dichloromethane (55ml) cooled to -78-C, was added a solution of ~-3',5'- di-P-benzyl-4'-thio-thymidine (1.6g, 3.7mmol) in ,~
.. I llrite;' ISin~om F~n. Off~^ ~
~: L~

PCT/GB 9 ~ 9 2 0 6 a 2 7 ~ ~ 1 OCtober 1991 dry dichloromethane (30ml). Stirring was continued for 5 hours at -78-C. This was then followed by the dropwise addition of a 1:1 methanol-dichloromethane solution ~200ml) over 40 minutes. The reaction mixture was allowed to warm to room temperature over 1 hour and the solvent removed in Y~YQ
and coevaporated with dry methanol (3 x 30 ml). The residue was applied to a silica gel column eluted with chloroform-methanol (85:15) to give the title title product. This could be crystallised from methanol to give oolourless crystals m.p. 208-209C.
NMR SPECTRUM
(lH) ~ (d6DMSO) 11.34 (lH,s,NH), 7.8i (lH,s,H-6), 6.32-6.26 (lH,t,H-l'), 5.26-5.25 (lH,d,OH-3'~, 5.20-5.16 (lH,t, OH~S'), 4.40-4.35 (lH,m,H-3') 3.18-3.16 (3~,m,~-4', H-15 5 2.25-2.13 (2H,m,H-2'), 1.80 (3H,s,CH3).
UV SPECTRUM
max 270.5nm (~,10,300).
ELEMENTAL ANALYSIS
~ .
Fou~d C,46.2: H,5.3;N,10-6 ClOH14N204S requ~res 20 C,46.5: H,5.5; N,10.9~.
MASS 5PECTR~
m/z 259 ~M+H]~, 3-noba matrix.
Prepa~ation of benzyl 2-deoxy-1.4-dithio-3~-di-O~p-toluoyl-D-erythro-pentofuFanosi-de To a 2~ boron trichloride solution in dry dichloromethane ~lSOml) cooled to -78-C, was added a solution of benzyl 3,5-di-Q- ~:
~ s~ ................................ s.: ~S , "
~ Câti~n . .

P~/GB 90/11~ &~9 2 0 6 ~ 2 7 9 ~ 1 Octobe~ 1991 benzyl-2-deoxy-1,4-dithio-D-erythro -pentofuran~side 14.2g, lOmmol~ in dry dichlormethane (lOOml), dropwise, over 30 mi~utes. Stirring was continued for 5 ~ours at -78-C. This wa then followed by the dropwise addition of a 1:1 methanol-dichloromethane solution ~200ml) over 40 minutes. Thereaction mixture waC allowed to warm to r~om temperature over 1 hour and the solvent removed n y~Q and coevaporated with dry methanol (3x30ml). The crude residue was diss~lved in dry pyridine (25ml), cooled tG O'C, and a solution of p-toluoyl chloride (4.6g, 30 mmol) in dry pyridine (25ml)added, dropwisej with stirring. The pyridine was removed 1~
vacuo, the residue extracted with chlorof~rm, and the extract successively washed with 2M hydrochloric acid, lM sodium carbonate and waterj dried (magnesium sulphate) and evaporated. The residue was applied to a ~ilica gel column elu~ed with hexane-ethyl acetate (9:1) to give the title product as a clear, slightly yell~w, syrup (2.5g, 53%).

~E~ , . . .
(lH) ~ (d6DNSO): 7.94-7.25 (13H,m,aromatic), 5-68-5.62 (lH,m,H-l') 4.74-4.66 (lH,m,H-3'), 4.39-3.83 (6Htm,H-3',H-4',H-51, PhCH2S), 2~51-2.25~2H,m,H-2'), 2.39 (6H,s,CH3).
EL~ENTAL ANALYSIS
Found: C,67.2;H,5.7. C2~2804S2.1/2H20 r~quires - C,67.0:H,5.8%.
.,';' ~
; 25 ~ASS SPECTRUM

m/z 515 [M+Na]+,401 ~M-Bn]+,369~M-SBn]+,357 r:it~ m ~ ,t Oi$,ce e~ CT~ T

:` ' ' ' ' i, . . ' . , , , ~ . . '' . . , ' ' ~ . .' ` ' ' .

2 0~ 5 2 79 ~l October 1 3 1 10 9~ ~
[M-OpTol]~, 3-noba matrix.
Pre~aration of E-5(2-bromovinvl-2'-deoxy-4l-thio-3'.5'd~
D-toluovl-uridine and its ~ -anomer To a solution of benzyl 2-deoxy-1,4-dithio-3,5-di-0-~-toluoyl-D-erythr~o-pentofuranoside (1.4g,2.8 mmol) in carbon tetrachloride (15ml) was added a ~olution of bromine (0.49g, 3.1mmol) in carbon tetrachloride (l~ml) with stirring at room temperature. After 5 minutes the mixture was concentrated under diminished pressure and then carbon tetrachlorid2 (5ml) was added and the mixture was evaporated to remove the excess bromine. The eYaporation procedure was repeated four times.
The resulting syrupy bromide was unstable and was used , directly in the next step.
! To a solution of the bromide in carbon tetrachloride (lOml) was added the bis TMS-derivative of E-5-(2-bromovinyl)ur~cil (1.7g, 4.7mmol) in carbon tetrachloride ; (lOml). The mixture was stirred until hcmogenous, evap~rated and the residue heated for 1 hour at 90-lOODC.. The cooled, dark residue was dissolved in 4:1 methanol-water (30ml), the 20 s~lution boiled for 15 minutes under reflux and then evaporated. The residue was triturated with chloroform (~Oml) and the solid 5-(2-bromovinyl) uracil that separated filtered off. The filtrate was successively wa~hed with aqueous sodium hydrogen carbonate and water, dried (sodium sulphate) and evaporated. The residue was applied to a silica gel column eluted with hexane-ethyl acetate (3:2).
Combination of the apporpirate fractions gave the title ~ ,; ? ~ SU~C~T~

PC~B ~ & ~ 9 - 206~279 ~~ C ', product as a white solid. lH NMR indicated the ratio of ~ -to ~ -anomers to be 1.8:1. Further column chromatography (chloroform-propan-2-ol (98:1)~ gave more separated anomers.
The first compound to be eluted from the column was E-5~L~
bromovinyl)-2'-deoxy-4'-thio-3l5ldi-o-p-toluoyl-uEidine which could be crystallis~d from methanol to give colourless crystals m.p.
182-184-C.
NMR SPECTRUM
(1H)~(d6DMSO): 11.73 ~lH,s,NH),8.10 tlH,s,H-6), 7-94~
7.86 (4H,m,aromatic), 7.39-7.19(SH,m, aromatic and vinylic . . .
N), 6.89 (lH,d,vinylic H,J=5Hz), 6.45-6.40 (lH,t,H-l'), 5.85-5.80 (lH,m,H-3'), 4.71-4.53 (2H,m, H-5'), 4.00-3.92 (lH,m, H-4'), 2.83-2.50 (2H,m, H-2'), 2.39 (6H,s, CH3).
UV SPECTRUM
max 241.6nm (~,34,960), 296.9nm (E,10,100 min 271.4nm (~,7,~00).
MASS SPECTRUM
; m~z~586 lM~Hj~, thioglycerol matrix.
;20~ The~-next~component~to be eluted from the column was the ~ -anom-t~w~ich could~be crystalli~sed from methanol to give `colo`urless crystals~m.p.l76-172 C.
NMR~SPECTR~M
d6DMSO)~11.64~1H,s,NH),~8.36 (lH,s,H-6), 7.91-25~ 7.77~ 4H,m,aromatic), 7.36-7.22 (SH,m,aromatic, vinylic 6.~80 ~1H~,~d,vinylic~H~ JcSHz)~, 6.29-6.27 (lH,d,Hl') 5.74-5.62 ;(lH,m,~H-3')4~8-4~.39~ 3H,m,~H-4',~H-5'), 2.94-2.~5 (2H,m,H-2'), -~
n'^~c~ lom P~tent~Off~ce ~ ~ .;r I~ a~ ~pp!iQ tion ~

PC~ D/~l~9' 20~2 ~9 _ 55 _ ~1 Oc~ober 1991 ~1 10 91 2.37 (6H,s,CH3).
UV SPECTRUM `
max 241.6nm (~,47,000) 296.1nm (~,12,500).
min 273.1nm (~,lO,OOo) ;~-ELEMENTAL ANALYSIS
~ound C,5404; H,4.4;N,4.5. C27H2~BrN2o6sl/2~2o requires C,54.6;H,4.2;N,4.7%.
MASS SPECTRUM
m/z 586 [M~H]+, thioglycerol matrix.

Pre~aration of E-5-(2-bromovinyl-?'-_eoxy-4'~thi~urid~e_and its ~ -anomer E-5-(2-bromovinyl)-2'-deoxy-4'-thio-3,'5'-di-0-~-toluoyl-uridine (200mg, 0.34mmol) was dissolved in a solution of sodium methoxide in methanol (7.Sml,O.lm) and the mixture allowed to stand at 22-C for 24 hours. ~he solution was n~utralised by careful addition of Dowex 50 ion exchange resin (~form) to p~6. The resin was filtered off and washed with methanol and the filtrate and washings evaporated to a white solid. This was ~pplied to a silica gel column eluted with chloroform-methanol f9:1). Combination of the appropriate fractions gave E-S- f 2-bromovinyl ) -2 ' -deoxy-4'thio-uridine (9Omg, 75%) which was crystallised from methanol-water to give colourless crystals, m.p. 190-191-C.
r~j SU~ S:; ~r-'r ~n :- P~ B 9~/~
2 0 S ~ 2 7 9 ~ I Oc~obe~ 1991 ~MR SPECTRUM
(lH) ~ (d6DMSO). 11.63 (lH,s,NH), 8.20 (lH,s,H-6), 7.30 (lH,d,vinylic H,J=5Hz), 6.97 (~H,d, vinylic H,J=5Hz), 6.27-6.22 (lH,t,H-l'), 5.29-5.28 (lH,d,OH-3'),5.24-5.20 (lH,t,OH-S') 4.40-4.32 (lH,m,H-3~),3.69-3.16 (3H,m H-4',H-5'), 2.30-2.15(2H,m,H-2') yV SPEI~UM
max 249.7 nm (~,16,000), 297.3 (~,14,300) min 271.2 nm (~,7,700).
EL~MENTAL ANALYSIS
F~und c,37.42: H, 3.72; N,7-76%- CllH13~rN2O45 requires C,37.82: H, 3.72: N,~.02~.
MASS $PEC~RVM
m/z 350 [M~H~, glycerol matrix The ~-anomer could be deblocked in an analogus manner ; ~ to ~ive E-5-!2-~r~mDvi~Yl)-l-r2-deoxY-4'-thio-~-D-erythro-pentofuranosyl)uracil. This was crystallised from methanol:
m.p. 186-187-.
~ :
~20 NMR SP~c~R~M

H)6(d6DM50): 11.56 (lH,s,NH), 8.44 (lH,s,H-6), 7.24 (lH,d,vinylic H, J-5Hz), 6.86 (lH,d,vinylic H,J-5Hz), 6.13-~;~ 6.09 ~lH,q,H-l'), 5.46-5.45 (lH,d,OH-3'), 5.06-5.02 ~lH,t,OH-; `5i~, 4.3-4.24 (lH,m, H 3'); 3.63-3.16 (3H,m, H-4'H-5'), 2.17-2.09 (2H,m,H-2'). ~
.
United K~ m P,~.te~t o~cel SU~3S~lTUTE S~ .-2 0 ~ ~ 2 7 9 ~ 9 ~ ~ ~ 09 ~
~ 57 ~ 3~ 10 91 UV SPECTRUM
max 250.9nm ~,16,500), 2s6.2nm (~,14,300) min 271.7nm (~,8,600).
~ASS SPECTRUM
m/z 350 [M+H]~, glycerol matrix.

.
EXAM~L~B -~
3',5'-Di-O-benzvl-2'-deoxv-5-iodo-B-4'-thiouridine Mercuric bromide (370 mg; 1.03 mmol) and cadmium carbonate (4&0 mg: 2.8 mmol) were added to a s irred solution, protected from moisture, of benzyl 3,5-di-0-benzyl-2-deoxy-1,4-dithio-D-erythro-pentofur~n~side (436 mg;
1.0 mmol) in dry MeCN (3 ml). A soluti~n of 5-iodo-bis-O-trimethylsilyluracil (3 mmol) in MeCN (12 ml) was added via syringe. The progress of the reaction was monitored by analytical HPLC while the mixture was heated under reflux for 1 h. When c~oled to ambient temperature, water (200 ~1) was add~d and after stirring for 30 min. the suspe~sion was filtered. The filtrate was evaporated and redissolved in dry MeCN, the precipitated 5-iodouracil was removed by f'iltration. The filtrate was purified by preparative HPLC on a 2.5 cm (1 in.) Zorbax C8 reverse phase column eluted at 20 ml min 1 with a gradient ~0 - 95% MeCN-water containing a constant 0.2% trifluoroacetic acid] over 20 min.; half-minute , ~;~ .
fractions were collected. Fractions containi~g pure produ~
were pooled and evaporated tQ yield 330 mg ~f product as a gum. The anomer ratio was 2.~ as determined by 200 .~,., . : .
llnl~Ed i ~cdo:n PrteDt O:f!c- Irr~ ., , r~
PC,T i~ ....;cton:l Ar~?!icatioll S~J ...- ; ~

, ", ., ,,;~ ~, 1 ;"; .. ;,,, ., , ., ", ~ " ,,,, .. ", ,, ~ , " " ., , " . .~ /,, .,, .. " . " , PE~ 01 agg 2 0 6 5 2 7 9 ~ l October ~9;~1 - 58 - 3 1 lO 9~

MHz lH-NMR. [CDC13 ~ :8.72 (s, 0.26H, ~-6-H); 8.55 (s, 0.74H, ~-6-H); 6.35 (t, 0.26H, ~ H); 6.24 (dd , 0.74H, ~ H)]

2'-Deoxy-5-iodo-4'-thiouridîne The above product (240 mg; 0.44 mmol) dissolved in dry CH2C12 (10 ml + 2 ml rinse) was added over 30min to a stirred lM
solution of BC13 in CH2C12 (lR ml, 18 mmol) at -78~C under N2. The reaction was followed by analytical HPLC. After 6h at -78C, MeO~-CH2C12 (1:1, 18 ml) was added slowly and the mixture allowed to warm to ambien~ temperature then evaporated. The residue was re-evaporated ~rDm MeOH
(3x) whereupon partial crystallisation occurred. The residue was taken up in MeO-CHC13 (1:1, 15ml) and the s~lid collected by filtration, yield 7.7 mg of the desired ~-anomer of the product. Mass spectrum m/z 370 (10%) for CgHllIN204S:200 MHz H-NMR ~ : 11.2 (br 5, lH, NH); 8.48 (s, lH, 6-H); 6.2 (t, lH, l'-H);5.23 (m,2H, 2xOH);4.35 (m, lH 3'-H); 3.60 (t, 2H, 5'-H2): 3.2 (4'-H, partly obscured by DOH): 2.20 (m, 2H, 2'-~2) The filtrate yielded a further 40 mg of mixed ~
anomers (ca. 1:1) from which pure ~-anomer could be obtained by preparati~e HPLC.

~' EXAMPhF 3a --'-Deoxy-5-ethyl-4'th~uridine ~
Benzyl 3,~-di-0-benzyl-2-deoxy-1,4-dithio-D-erythro-pentofuranoside (5.6 mmol) was dissolved in CC14 (30 ml) and ¦ PCl Int~.r~ n~' Applicatlon ~
.

PCT/~ 9 0/
205~279 ~1 OetobeS 1~91 _ 59 _ ~ 1 10 91 bromine (6.2 mmol) in CC14 (30 ml) was added. Af~er stirring for 5 min. at ambient temperature the solvent was evaporated ! and the residue re-evaporated from CC14 (10 ml) to remoYe ex~ess bromine. To a solution o~ this crude l-bromo-t~iosugar in CC14 (15 ml) was added bi~-0-trimethylsilyl-5-ethyl ura~il (16.6 mmol~ ~prepared by refluxing ~-ethyluracil (16.6 mmol) in a mixture o~ hexamethyldisilazane (50 ml) and chlorotrimet~ylsilane (5 ml) for 2h., and evaporation of the solvents], HgBr2 (1.99 g; 5.5 mmol) and CdC03 (2.36 g; 16.6 mmol). The solvent was evaporated and the residue heated at lOO-C for lh. The residue was worked up as for the thymidine analogue and the product purified by column chromatography.
The benzyl ether protecting groups were removed by treatment with BC13 as described for the 5-iodo analcgue.
.

~XAMPLE__3b Separation of ~nomers of 2'-deoxv-5-ethvl-4'thiQuridine The sample of the mixed ~,~-anomers of the 5-ethyl compound were separated by prepar3tive reverse phase HPLC on a 2.5 cm (lin.) Zorbax C8 ~olumn eluted with MeCN-~20 (1:9, v~v);
h21f-minute fractions were collected. The separate anomer poQls were freeze dried.

-Anomer: Yield 23mg; retention time: 6.2min.; mass spe~trum m/z 272; calc. for CllH16N204SØ3H~o:C, 47.47; H,6.03; N;

10.06; found, C, 47.48; H, 5.71: N,9.96%: 200 MHz 1H~NMR, DMS0-d6, ~ : 11.28 (br s, lH, NH); 7.8 (s, lH, 6-H) 6.3 (t, ~'r~;

~" , ", ,, ,"~ ,;, ,.,,, ", ", ~ " "" ~"~ ",; ,"~

PCT/GB ~ al~ 1 ~8t 2~5279 3 1 10 ~1 - 60 - ~l octobes 1 lH, 1'-H); 5.24 (d, lH, 3'-OH); 5.16 (t, lH, 5'0H): 4.37 (nl, -~
lH, 3'-H); 3.62 (m, 2H, 5'-H2); 3.3 (4'-H, partly obscured by DOH): 2.25-2.4 (q+m, 4H, CH2CH3 + 2'-H2); 1.05 (t, 3H, CH2CH3 ) -~-Anomer: Yield 15.8mg; retention time: 7.5min.; mass spectrum m/z 272; calc- for C11H16N2O4S-H20 C~ 45-51 H~
6.25; N. 9.65; found, C, 45.54: H, 5.7~; N, 9.33%. 200 MHz H-NMR, DMSO-d6, ~ : 11.1 tbr s, lH, N~); 8.12 (s, lH,6-H);
6.2(q, lH, l'-H); 5.50 (d, lH, 3'-OH); 5.00 (t, lH, 5'-OH):
4.34 (m, lH, 3'-H); 3.45-3.70 (m, 3H 4'-H + 5'-H2); 2.55 (m~
lH, 2l-H, partly obscured by solvent); 2.25 (q 2H, CH2CH3)2.05 (dt, lH, 2'-H) 1.05 (t, 3H,CH2CH3).

EXAM~LE C
3'5'-Di-O-benzvl-5-bromo-2'deoxv'B-4'~hiourid~ne.
This compound was p~epared by a method similar t~ the iodo compound above with the following modifications:
1. The total solvent (MeCN) volume for the reaction was 3 ml.
2. The CdCO3 was omitted.
20 3. The excess of 5-bromo-bis-0-trimethylsilyuracil was ~.
reduced to 1.5 mole equi~alents.

. . .
~- ~ . . ' ~ dol.l P~t-~t ~f~l Sl.~.~B~'T 'T_ 5 -<

-' 2 0 ~ 5 2 7 P~ ~

~1 OcSob~ 1991 The yield of HPLC-purified compound was 193 mg; mass spectrum gave m/z 502 expected for C23H23BrN2O4S-~XAMPLE 4 5-Bromo-2'-deoxy-4'-thiouridine :-.
, 5 The BC13 deprotection was conducted as for the iodo-compound.
After HPLC purification a sample of the bromo-derivative (3.3 mg) of anomer ratio 3.~ was obtained. Mass spectrum showed the expected molecular ions at 322(1%) and 324(0.8%);
10 the 200 MHz lH-NMR spectrum was consistent with the structure ~.
(DMS0-d6, :8.72(0.22H, s, ~-H6);8.48(0.78H,s, ~-~6) EXAMPLE D
l-Acetoxv-3.5-di-p-tolu~yl-2-deoxy-4-thio-D-erythro-~entofuranoside A solution of benzyl 3,5-di-0-benzyl 2-deoxy-1-4-dithio-D-erythro-pentofuranoside (3.68 g; 8.76 mmol) in dry CH2CH2 .:
(20ml) was added dropwise to a stirred lM solution of BC13 in :
CH2C12 ~125 ml; 0.125 ml; 0.125 mol) at 78-C u~der N2. The .

mixture was stirred at -78 C for 4.5h, then a mixture of :
Me~H-CH2Cl2(1:1,v/v) was added slowly. After warming to room temperature the solvents were evaporated to give the crude O-debenzylated ~hiosugar. The gum was dissolved in dry pyridine at O'C under N2 3nd a soluti~n of p-toluoyl chloride (3.47 ml; 26.3 mmol) was addPd slowly. The mixture was stirred at O~C for 3 hours the~ the solvents were evaporated~

L P.,T ~ at.G~ A~ c~t~r~. l .
P~/G~ 9 206Y~279 3 1 10 9 ~1 October 1199 The residue was dissolved in CH2C12, washed with 2M HCl, lM
Na2CO3 and water, dried over MgSO4 and evaporated. The residue was purified by flash chromatography on sio2 eluted with EtOAc-hexane (1:9, v/v) to give the bis-toluoylthiosugar derivative (2.18 y;ca.50%): mass spectrum m/z 492. This product was dissolved in acetic anhydride (16 ml~ and stirred at O-C. Conc. H2SO4 (8 ~1) was added ~ollowed after 10 min.
by a second aliquot (8 ~1); the reaction was monitored by TLC. After a further 55 min. stirring NaHCO3 (100 mg.) was added and after 20 min. the mixture was cautiously poured into ice-water containing NaHCO3. The product was extracted into CH2C12, dried and evaporated. The residue was purified by flash chromatography on SiO2 eluted with 20-25% EtOAc-hexane. Yi~ld 0.97g:- 200 MHz lH-NMR DMSO-d6, ~:~.7-

8.1(m,4H,ArH); 7.1-7.4(m, 4H + s41vent, ArH); 6.35 (dd,0.55H, ... .
~i 1-H);6.27(q,0.45H, l-H);5.7-5.9 (m,lH, 3-H); 3.7-4.7(m,3H,5-H2+4-H);2.2-2.7(m+2.2-2.7(m+2xs, 8H, 2xArCH3 ~ 2-H~); 2.0-2.1 (2xs, 3H, CH3CO~ ). The anomer ratio was approximately 1.1~1: the material was sufficiently pure for use.
, ' . .
EXAM~L~ 5 2'-Deoxy-~-pr~pYnvl-4'-thiouridine 5-Propynyluracil(0.112 g; 0.~5 mmol) was hieated in , hexamethyldisilazane (3 ml) containing trimethylsilyl ,.
chloride (1 ml)~until the solid dissolved (4h). The solvents were evaporated and the residue dissolved in dry NeC~ (6 ml).

The solution was added, under N2, to a stirred solution of . ~
IJni~ed K.n~ ~ P.^t~nt Office Cl Inc~ r Fv1 In. ..~.ional Application ,~UJ
`

P~r/~ 9 ~
~0~5279 ~ 1 tO 9 - 63 - ~ 1 Oclober 1991 ', the above thiosugar ester (0.2 g; O.5 mmol) in MeCN (10 ml) at O C. Trim~thylsilyl triflate (0.096 ml; 0.5 mmol) was added and the mixture stirred for 15 min. The mixture was diluted with CH2C12 (20 ml), poured into saturated aqueous 5 NaHC03 and the organic layer separated. The aqueous layer was further extracted with CH2C12 and the combined organic~
dried and evaporated. Flash chromatography on SiO2 eluted with EtOAc-hexane (3.2, v/v) gave the protected thionucleoside as a mixture of anomers (i.47/1 ~
contaminated with a little propynyluracil. Yield 0.21 g.
This material (0.206 g; 0.397 mmol) was dissolved in MeOH (15 ml) containing NaOMe (0.021 g; 0.397 mmol) and the mixture kept at ambient temperature overnight. The solution was neutralised with Dowex 50(Hf)ion-exchange resin, filtered and the filtrate evaporated to dryness. The solid was washed with ether (3 x 4 ml) and digested with hot acetone to give the required product as a white solid. Yield 100 mg.
Methanol was added to the mixture and the solid was filtered off to give the pure ~-anomer, 30 mg. The filtrate was proce sed by HPLC as dei~cribed above to give a further 6 mg.
of the ~-an~mer and a quantity of the ~-anomer.
~-Anomer: mass spectrum m/z 282;200 MHz lH-NMR,DMSO-d6, ~:
11.55 ~br s, lH, NH); 8.7 (s, lH, 6-H); 6.25 (t, lH, 1'-H~ ;5.2~m, 2H, 2 x OH); 4.3 (m, lH, 3'-H): 3.6(m, 2H, 5'-H);3.3 (4 ~ , partially obscured by DOH); 2.15, (m, 2~, 2'-H);2.0(s, 3H, C~CCH3)-5-Propynyluracil may be obtained from 5-iodouracil using the ~=1 ` ~i ~ ' . . . s. ._ET
.

P~ n l ~ 9 20~52~il9 - 64 - ~1 October 1991

9~
methodology analogous to that described by M.J. Robins et al (ibid).

2~-Deoxv-5-chloro-4'-thiouridine:
Starting with ~-chlorouracil, this compound was prepared in a similar manner to that described in Example S. 5-chlorouracil is commercially available.
The compound was purified by HPLC as described above and obtained as mixture ~f anomers ~/a ca. 3:1 1H-200 M~z NMR DMSO-d6, C:11.8 (br s, lH, NH); 8.65 (s, 0.25H, ~-6-H); 8.4 (s, 0.75H, ~-6-H); 6.1-6.3 (t+m, lH, l'-H);5.55(d,0.25H, ~-3'-OH);5.2-5.3(m, 1.5H,~-3'-OH + ~-5'-OH); 5.05 (t,0.25H, ~-5'-OH); 4.3-4.45 (m, lH, 3'-H); 3.6-3.7 (m,2H, 5' H2);2.1-2.4(m, 2H, 2'-H2); 4'-H obscured by : 15 solvent.

Mass spectrum: ~bserved m/z 278 and 280 for CgHllC1~2O4S.
,.

EXAMP~E 7 2 '-Deoxy-5-trifluoromethvl-4 '-thiouridine:
~ .
Starting with 5-trifluoromethyluracil, this compound was prepared in a similar manner to that described in Example 5.
5-trifluoromethyluracil is commercially available.
A sample of this compound of anomer ratio ~/~ ca. 8:1 was obtained by trituration of the crude deprotected nucleoside mixture with acetone, filtration and evaporation.
1H-200 MHz N~R DMSO-d6, ~:11.8 (br s, lH, NH); 8.83 (s, lH, I uni~e~l ''."`'.

- 20~27~ P~T/~d go/~ ~
- 65 - ~1 Octob~r It ~-6-H); 6.2 (t, lH, ~-l'-H); 5.2-5.4 (m, 2H, ~-3'+ 5~-OH);

4.25-4.4 (m, lH, ~-3~-H); 3.5-3.8 (m, 2H, ~5'H); 3.0-3.5 (m, , 4'H obscured by solvent): 2.2-2.4 (m, 2H, ~-5'-H2); small signals indicative of the ~-anomer content were also observed.
Mass spectrum: observed m/~ 312 ~or CloHl1F3N204S.

2'-Deoxy-5-ethy~vl-4'-thiourldine:
Starting with 5-ethynyluracil, this compound was prepared in

10 a similar manner to that described in Example 5. 5- -ethynyluracil may be prepared from 5-ioduracil usin~ the methodology analogous to that described by M.J. Robins et al ' (ibid).

A sample of the pure ~-anomer of this compound was obtained ~ :
by boiling the crude anomer mixture with MeOH and filtering off the product.
H-200 MHz NNR DMSO-d6 ~: 11.6 (br s, lH, NH); 8.42 (s, lH, ~-6-H~; 6.23(t, lH, ~ H); 5.1-5.35 (m, 2H, ~-3'~5'-OH):
4.25-4.45 (m, lH, ~-3'-H); 4.15 (s, lH ~CH); 3.55-3.75 (m,2H, ,; 20 ~-5'-H); 3.1-3.5 (~-4'H, obscured by DOH~: 2.1-2.4 (m,2H, ,j~ 5 ~2~
,~ , Mass spectrum: observed m/z 26~ for CllHllN~04S.

I--~d ~ dom P.te~,t Of~ic~ v~i'i-~T_ 5. .-. . - ,, .

P&T/6~ ~O/~&9 - 66 - ~1 ~tObleO 9 EXAMPLE g 2'-~eoxy-5-E-(2-bromovinyl~-4'-thiocytidine.
To a solution of benzyl 3,5-di-O-benzyl-2-deoxy-1,4-dithio-P-erythro-pentofuranose ~4 g: 9.5 mmol) in acetic ~cid (50 ml) and acetic anhydride (50 ml) was added conc. sulphuric acid (50 ~1) and the mixture stirred at ambient temperature for 30 min. when TLC (EtOAc-hexane, 1:4, v/v) showed complete c~nversion to a m~re polar sugar. The mixture was poured into excess sodium bicarbonate Na2HCO3, extracted with CHC13, 10 the extracts dried over MgSO4 and evaporated. The residue -was purified by column chromatography on SiO2 in the TLC
solvent to give the l'-acetoxy-di-O-benzylthiosugar derivative (1.84 g; 54~) which was used directly below.
To the above derivative (0.33 g, 0.89 mmol) in dry CH2C12 (3 ml) at OC was added S~C14 (0.33 g; 0.89 mmol) in dry CH2C12 ~3 ml) and E-5-(2-bromovinyl)-2,4-dimethoxypyrimidine (0.218 g; 0.89 mmol) in dry CH2C12 (3 ml). The stirred mixture was allowed to warm t~ ambient temperature and stirred for a further 4 h. The mixture was poured onto water, washed with ~ -~0 saturated NaHCO3 and dried o~er MgSO~. After evaporation,the residue was chromatographed on SiO2 in toluene-acetone (9:1, v/v) to give the pure ~-anomer of the protected t~ionucleoside, which was crystallised from MeOH (ca 40 mg).
NNR, DMSO-d6, ~:8.31 (s, lH, H-6); 7.39-7.28 (s, lOH, 2Ph);
7.0 (d, lH, vinyl H, Jz 14 Hz); 6.85 (d, lH, ~inyl H,J= 14 ~: Hz): 6.27 (t, lH, l'-H); 4.56 (s, 4H, 2PhCH2); 4.33 (m, lH, 3'-H~; 3.90 (5, 3H, OC~3); 3.77 (m, 2H, 5'-H2); 3.63 (m, lH, r; - d ~ p~nt ff~e1 ~U esl.,V .~ ~

- . .

P~T/~ ~aJ~I ~9;
206a279 3 1 10 91 -:
- 67 - ~1 October 1991 4'-H), 2.50 (m, 2H, 2'-H2) The above methoxy derivative of the protected thionucleoside was converted to t~e cytidine anal~gue by dissolution in NH3/MeOH at ambient temperature for 2d. The product was --isolated by column chromatography ~n SiO2 eluted with CHC13-MeOH (9:1, v/v), then depr~tected directly with BC13 as described above. The product was essentially pure and consisted of the HCl salt ~f the mixture of anomers in the approximate ratio 95:5, ~:~ by HPLC on Zorbax C8 eluted with a 0-95% gradient of MeCN in water over 15 min. followed by 95% MeCN: retention times ~ 16.3 min.; ~ 18.11 min. 200 MHz 1 H-NMR, DMSO-d6, ~: 8.55 (s, lH, 6-H); 7.25 (d, lH, Jtrans 13.8 Hz, vinyl H): 6.95 (d, lH, Jtrans 13.8 Hz, vinyl H);
6.18 (t, lH, l'-H~). Mass spectrum (EI): no molecular ion was observed but characteristic ions for the base ~215,217 for C6H6BrN3O; 136 for C6H6N3O] and the thiosugar [85,C5H3S~
were seen.
A pure sample ~f the free base of the ~-anomer was obtained by~pr-parative HPLC as described above.
200 NHz 1 H-NNR, DMSO-d6, ~: 8.18 (s, lH, 6-H); 7.1-7.4 (br s, 2H, NH2); 7.05 (d, lH, Jtrans 14.5 Hz, vi~yl H); 6.85 (d, lH, Jtrans 14.5 Hz, vinyl H); 6.30 (t, lH, l'-H); 5.1-5.25 tm, 2H, 2 x OH); 4.3-4.45 (m, lH, 3'-H); 3.55-3.7 (m,2H, 5'-, ` H~); 3.0-3~4 (4'-H obs~ured by DOH): 2.1-2.35 (m,2H, 2'-H2).

; ~-~~ ~~p~ent Oi ~ S~n~ J~E 5~
~ d D~

2 0 6 a 2 7 9 ~ 1 qO ~1g `
- 68 ~ c~obe~ ~992 ~'-peoxy~5-propyl-4'~h~Quridine:
2'-Deoxy-5-propynyl-4'thiouridine, B-anomer, (26 mg) and 5~
Pd/C ~40 mg) in MeOH (80 ml) was stirred in an atmosphere of hydrog n for 45 min. ~PLC analysis showed complete conversion to a more lipophilic compound. The mixture was filtered and ~vaporated to give a gum, yield 25 mg.
Trituration with ether-hexane gave the product as a white solid. 200 MHz lH-NMR, DMSO-d6, ~ 5 (br s, lH, NH);
7.80 (s, lH, ~-6-H): 6.27 (t, lH, ~-l'H); 5.22 (d, lH, ~-3'-OH); 5.14 (m, lH, ~-5'-OH); 4.3-4.45 (m, lH, ~-3'-H); 3.55-3.75 (m, 2H, ~-5'-H); 3.2-3.4 (~-4'~, obscured by DO~); 2.1-2.35 (m, 4H, ~-5'-H2 ~ CH2CH2Me); 1.45 ~m, 2H, CH2CH2Me); : .
0.88 (t, 3~, CH3).
Mass spectrum: m/z 286 (M~) for C12H18N24S
' . ' ~:
~XAMPLE ~l E-2'-~eox~-5-(pro~en-1-yl)-4'-thiouridine ta~ 5-Allyluracil.
. Uracil (1 g; ~ mmol) was dissolved in water (200 ml) at 70-C
and Hg(OAc)2 t2.9 g; 9.1 mmol) was added. The mixture was :, stirred at 70-C for 1 week. After cooling to ambient ~, temperature NaCl (1.5 g) was added and the mixture ~tirred . . for 4h. The resulting thick suspension of 5-chlor~mercury-uracil was filtered, the solid washed with 0.1 M NaCl . 25 solution and dried i~ vacuo at 85C for 2 days (2.27 q)~ To the crude solid (1 g; 2.9 mmol) in MeCN (25 ml) was added ~:
?~ .t O~ri~,e I sug .

2~63279~/GB 9 n / ~ t Q~ ~
31 10 91:
- 69 - ~1 Oclober 199~ :

Li2PdC14 (0.76 g) and allyl chloride (2.9 ml) and the mixture stirred at ambient temperature for 1 week. The suspension was ~iltered and the filtrate evaporated to dryness. The residue wa~ dissolved in MeO~ (75 ml) and treated with H2S
gas; the black precipitate of HgS was removed by filtration and the filtrate was evaporated t~ ~eave a white solid. The .
desired product was isolated by flash chromatsgraphy on sio2 ~ :
eluted with 8% MeOH-CH2C12 (v/~). Yield (85 mg, 20%): Mass spectrum gave m/z 152 for C7H8N202 (M~); 200 MHz lH-NMR
DMSO-d6, ~: 10.9 (br s, lH, N~); 7.16 (s, lH, 6-H~; 5.7-6.0 (m, lH, -CH=); 4.95-~.15 (m, 2H =CH2); 4.33 (br s, lH, NH);
2.92 (d, 2H, CH2). :
(b) 5-rE-propen-~-yl) uracil To a solution of 5-allyluracil (80 mg; 0.5 mmol) in 95% aq.
EtOH (50 ml) was added (Ph3P)3RhCl (90 mg; 0.1 mmol) and the mixture heated under reflux for 3 days. The solvent was evaporated and the product isolated by flash chromatography on Sio2 eIuted with 5% MeOH-CH2C12. Yield 56 mg 70%; 200 MHz : lH-NMR DMSO-d6, ~: 11.0 (br s, lH, NH): 7.42 (s, lH, 6-H)7 6.35-6.55 (qq, lH ~CH-Me); 5.95-6.1 (dd, lH, -C~s): 1.74 ~dd, 3H, C~3)-(c~ E~ DeoxY-5-(pro~en-1-Yl)-4'-thiouridine ; 5-(E-propen-l-yl)uracil ~110 mg; 0.78 mmol) was converted to the bis-TM~-ether, coupled with the protected thiosugar and deprotected with methoxide as described for the ~-propynyl analogue. The ¢rude produot was purified by chromatography on SiO2 eluted with 5% MeOH-CH2C12. Yield 11.8 mg of mixed ~G~ Sl)BSTI~
.

~.. ,.. ,",. .,. , . . ,, . .. , . ... , .,. .. ... i . .. . . .... . . .. . . . .. .. . ... .... .
. ~ . .. - . .. , . ~ ,.. . .. . , , (.. . . .. . . .... ... . . .
- - ,. ., . ~... . ,i. . .. , . , . . - ,. .. .
,. . ~ . . i ... . i . .. .... ... . . . . .. .. .. .. . .. . .

~CT/GB ~ t~
~1 10 ~1 2 ~ ~ ~ 2 7 9 I :
anomers in the ratio 1.2~ . 200 MHz lH-NMR DMSO-d6, ~:

11.35 (~r s, lH, NH); 8.35 (s, 0.55H, ~-6-H); 8.05 (s, 0.45H, ~-6-H); 6.0-6.6 (m, 3H, l'-H + -CH=CH-); 5.5 (d, 0.55H, ~-3'-OH); 5.1-5.3 (d+t, 0.9H, ~-5~-OH ~ ~-3'-OH); 5.0 (t, 0.55H, ~-5'-OH); 4.38 (m, lH, 3~-H); 3.1-3.7 (m, 5'-H2 + 4'-H, partially obscured by DO~; 2.0-2.6 (m, 2'-~2, partially obscured by solvent); 1.75 (d, 3H, CH3).
Nass spectrum: m/z 284 (M+) for C12H16N2O4S

EXAMPLE E
Preparati~n of E-5-(2-bromovinvl~-Uracil-5-Bromo-2~4-dimethoxypyrimidine A solution of 5-br~mo-2,4-dichloropyrimidine (16 g: 70.2 mmol) [D.M. Mulvey et al. J. Het. Chem., 1973,p79] in dry MeOH (55 ml) was added slowly to a stirred soluticn ~f sodium (3.23 g: 140.4 mmol) in MeOH (55 ml) at O C ~ver 30 min. The ice-bath was rem~ved and the reaction mixture stirred at ambient temperature for 18 h. The precipitated salt was removed by filtration and the filtrate evaporated to give an oil. To this was added an aqueous solution ~f NaOH (30 ml:
30~ w/v); the product separated as an upper layer and was extracted into Et2O. The organic extracts were dried over MgSO4 and evaporated. The ~esidue was crystallised from thanol to give the product as c~lourless plates, yield 14.3 g, 93%, mp 62-63-C. Mass spectrum, elm/z 219 (M~, 11%).
Analysis, found: C,33.20, H,3.26, Br 36.90, N, 12.7%; ~
C6~7BrN2O2 requires: C,32.90, H,~.33, Br 36.50, N, 12.80%. ~ -r~-~ S ~ ,E 5~ ~rT

--- PG~/~B ~Q/ a 2065279 ;~ i lD 91 - 71 - 31 October 1~91 5-Formyl-2.4-dimethoxvpryrimidine ; A solution of 1.6 M n-Buli in hexane (48 ml, 73.6 mm~l) was added over 5 min. to a stirred suspension of 5-bromo-2,4-dimethoxypyrimidine (16 g; 72.9 mmol) in dry Et2O (240 ml) at -70-C under an atmosphere of dry N2. Dry ethyl formate (28 g: 377 mmol) was added and the orange solution ~tirred at -70~C for 1 h then allowed to warm slowly to ambient temperature. water (400 ml) was added and the aqueous layer separated and extracted with ~t2O (3 x 200 ml). The ether layer was combined with the extracts and dried over MgS04, filtered and evaporated. The residue was purified by column chromatography by preloading i~ SiO2 and eluting with EtOAc-hexane (3:7l v/v). Product fractions were com~ined and evaporated to give fine white needles, yield 6.89 g, (56%).
lS Mass spectrum m/z 169 (M~H)+ Analysis, f~und: C, 50:1:H,4.5;N,16.9%;C7H8N203 requires C,50.00; H,4.79; N, lS.66~

E-5-~2-çarboxvviny1)-?.4-dimethoxypvrimidi~e Malonio acid (13.03 g; 126.2 mmol) ~nd redistilled piperidine -( 2ml) were added to a s~lution cf ~-formyl-2,4,dimethoxypyrimidine (10.52 g: 6.2.6 mmol) in dry pyridine (60 ml). The mixture was heated on a steam bat~ for 10 h -the~ the solvent was removed by distillati~n under reduced pressure. The residual oil was re-evap~rated from water (3 x 25 ml) and the solid thus obtained recrystalli~ed firstly from water and then from dry methanol t~ give the product as d~ Pa~er~i~j Sl~ TE S~''ET
PCT l"a. ;~tio.;~l Aprli~ation ~, , , ~
,~ .

--- P~ B ~ t Q
~065279 3 1 1~ 9~
- 72 - ;~1 OctobeI 1991 white needles, yield 6.45 g; a second crop was obtained from the filtrate (1.08 g). ~otal yield 7.53g (57~). Mass spectrum: (El) m/z 210 (M+). Analysis, found:
C,52.1;~,4.8;N, 13.1%: CgHl~N204 requires: C, 52~43: H, 4.79;
N, 13.33%.

E-5-(2-Bromovinvl)-2.4-dimethoxypyrimidin~e To a solution of E-5-(2-carboxyvinyl)-2,4-dimethoxypyrimidine (0.300 g: 1.43 mmol) in dry DMF 15 ml) was added K2CO3 (0.45 g: 5.25 mmol). After stirring at ambient temperature for 15 min. a solution of N.-bromosuccinimide (0.258 g; 1.45 mmol) in dry DMF ~4 ml) was added dropwise over 10 min. The suspension was immediately filtered, the solid washed with DMF and the filtrate evaporated in high vacuum. The solid residue was purified by column chromatography by preloading on SiO2 and eluting with EtOAc-hexane (7:3, v/v). Product fractions were pooled and evaporated to give ~ine white crystals, yield 0.561 g (45%).
FAB mass spectrum: m/z245 and 247 (M+H)+. Analysis, fou~d:
C~39~9r H, 3.6: N, 11.5% CgHgBrN2o2 requires C, 40.20: H, 20 3 ~ 70 N. 11. 43%

E-5-(2-BromovinYl)Uraci ., :
To a solution of E-5-(2-bromovinyl)-2,4-dimethoxypyrimidi (2.45 g; 10 mmol) in AcOH (10 ml) was added NaI (3.3 g; 2.2 e~.; 22 mmol) and the solution heated under reflux for 3h.

L'-~,r t.... ;~
~ :.

2 0 6 5 ~y~ 9 ~ 9 9 :

- 73 ~ C~9~ 9~1 The hot mixture was filtered and diluted with water (15 ml).
After cooling, the precipitated product was filtered off, washed with acetone (50 ml) and ether (20 ml) and dried to give a pale yellow powder (1.40 g, 65%). Mp >320-C: 60 MHz 1H-NMR, DMS0-6d, ~: 7.60 (s, lH, H-6); 7.30 (d, lH, J=13 Hz, vinyl H); 6.80 (d, lH, J-13 Hz, vinyl H).
BIOLO~ICAL DATA
a) Anti-~SV Activitv Herpes Simplex Virus types 1 (HSV 1) and 2 ~HSV2 were assayed in monolayers of Vero cells in multiwell trays.
The virus strains used were SC16 and 186 for HSV-1 and HSV-2 respectively. Activity of compounds was determined in the plaque reduction assay, in which a cell monolayer was infected with a suspensi~n of the appropirate HSV, and then overlaid with nutrient agarose in the form of a gel to ensure that there was no spread of virus throughout the culture. A
range of concentrations of compound of known molarity was incorporated in the nutxient agarose overlay. Plaque numbers -~t each concentration were expressed as percentages of the c~ntrol and a dose-response curve was drawn. From this curve the 50% inhibitory concentration (IC50) was estimated to be 0.66 ~m for the csmpound of formula (I) in which X represents a 2-bromovinyl group.
b) Anti-CMY activity Human cytomogalovirus (HCMV) was assayed in monolayer~ of either MRC5 cells (human embrycnic lung) in multiwell tray~. ~he standard CMV strain AD 15~ was used.

~ A ~

.. . . . ~ .. .

P~T/~ 9 632~9 ~ 1 10 91 ~1 Oclober 1991 Activity of compounds is determined in the plaque reduction assay, in which a cell monolayer is infected with a suspension of HCMV, and then overlaid with nutrient agarose in the form of a gel to ensure tha~ there is no ~pr~ad of virus throughout the culture. A range of concentrations of compound of known molarity was incorporated in the nutrient agarose overlay. Plaque numbers at each concentration of drug are expressed as percentage of the control and a dose-response curve is drawn.
c~Anti-VZV Activitv Clinical isolates of varicella zoster virus ~VZV) were assayed in monolayers of MRC-5 cells. ~RC-5 cells are derived from human embyonic lung tissue. A plaque reduction assay was used in which a suspension of the virus stock was lS used to infect monolayers of the cells in multiwell trays. a range of concentrations of the compound under test of known molarity was added to the wells. Plaque numbers at each concnetration were expressed as percentages of the control and a dose response curve was constructed. From th~se curves ~ -tht~ 50% inhibitory concentration of each drug was determined.

' . -i :

~;~;;~ Sl.l~STITI~E S~

.

-` P~ B

31 Oct~ber 1991 Ta~le l shows the activity of compounds of the invention.
~A~LE l Activity of ~omp~unds of the inventio~n against HSvl, ~sv2 and VSV. Data relates to the ~-anomers.

. _ COMPOUND HSVl¦ HSV2 VZV CCID50 . _. _. . L .. _ _ : X Y IC50 (~M) ~M
_ .__ CH=CHBr OHO. 6 >10, <100 O. i8 >500 ._ . ._ ., CH2CH3 OH0.17, 5, 2.3 0.79,0.99 ~l00, 0 25, 253 .
CH=CHBr NH? -2 . 3 >10 ~, , .
, . ' ..1 EXAMPLES
The fi3110wing examples illustrate pharmaceutical formulations according to the invention in which the active ingredient is a compound of formula (I). :
~: Formulation Ex~mple A Tablet ~ ~ .

.. ' A~ti~e ingredient lO0 mg Lactose 200 mg '1 . .
Starch 50 mg Polyvinylpyrrolidone 5 mg~$~.
Magnesium stearate : 359 mg ~ r~r.ii~ } ~ . OfT~,c~ Sl n~ r ~ 1 P~iT ~ ;ai ~pyil'C~io!l ~;.. ~. ~.i ;.. l ~ S. . ~ -~Tt~ ~ 01 Qt 0 2~6a27~ 31 10 ~1 - 76 - 31 Oc~ober 1991 Tablets are prepared from the foregoing ingredients by wet granulation followed by compression.

Formulation ExamPle B O~halmic Solution Active ingredient 0.5 g Sodium chloride, analytical grade o.g g Thiomersal 0.001 g Purified water to 100 ml pH adjusted to 7.5 Formulation Exam~le C: Tablet For~ulat ons The following formulations a and b are prepared ~y wet ~ranulation of the ingredients with a solution of povidone, followed by addition of magnesium stearate and compression.

~.-~ .

' -., :..

'PCT/~ 9 ` 2 ~ ~ ~ 2 7 9 ~ I October l99 Tablet Formulation a ~g~tablet~/tablet (a) Active ingredient 250 250 (b) Lactose 8.P. 210 26 (c) Povidone B.P. 15 9 (d) Sodium Starch Glyc~late 20 12 (e) Magnesium Stearate 5 3 Tablet Formulation b ma/tablet ~/tablet ta) Active ingredie~t 250 250 (b) Lactose 150 (c) Avicel PH 101 60 26 (d) Povidone B.P. 15 9 (e) Sodium Starch Glycollate 20 12 (f) Magne~ium Stearate 5 3 Tablet Formulation c ~g/~ablet Active ingredient 100 L2ctose 200 Starch 50 Povidone Magnesium stearate 4 ! ` 359 ' ~ ¦ Uni~ c:~ P.~ t C.~iC ~ ~ r ~ C~ l hj/?l.~ S~l ~TlTI i ~- _ -- !

~G~/~B ~ C ~ ~, i G

- 78 _ 2~6~279 31 31 ~ctober 1991 The following formulations, D and E, are prepared by direct~:
compression of the admixed ingredients. The lactose used in formulation F iS of the direct compression type.

Ta~let ~ormulation d m~capsule :~
Act1ve Ingredient 250 Pregelatinised Starch NF15 150 400 .
, ', ' , .
ablet Formulation e m~/capsule Active Ingredient 250 Lactose 150 Avicel lO0 500 ~: .

~ablet Formulation f ~Controlled Release Formulation2 The formulation is prepared ~y wet gr~nulation of the ingredients (below~ with a solution o~ povidone followed by: - .
the addition of magnesium stearate and compression.
~t~blet (a) Active Ingredient 500 I ~ ~ (b) Hydroxpropylmethylcellulose 112 . (Methocel K4M Premiu ~ ~

- :~ (c) Lactose B.P. 53 ~ (d) Povidone B.P.C. 28 , ,. .. _ ~ . .
Un~ m P-to!~t ~,..&f~ S~ ~-`tC'TIT' ' ~
~ ' f.~s~ ~ " ~

- - P~T~ 9 ~
_ 79 20~a279 31 10 91 (e) Nagnesium Stearate 7 ~1 Ctober ~991 Drug release takes place over a period of about 6-8 hours and was c~mplete after 12 hours~

Formulation ~xam~le D: Ca~sule Formulati~s Capsule Formulation a A capsule formulati~n is prepared by admixing the ingredients ~ F~rmulati~n D in Example C above and filling into a two-part hard gelatin capsule. Formulation B (infra) is prepared in ~ similar manner.
Capsule Formulation b mQ/capsule (a) Active ingredient 250 (b) Lactose B.P. 143 (c) Sodium Starch Glyc~llate 25 (d) Magnesium Stearate _ 2 - :
Capsule ~ormulat~n c mq/caDsule (a~ Acti~e ingredient 250 (b) Nacrog~l 4000 BP 350 ~ .
.

Capsules are prepared by melting the Macrogol 4000 ~P, disper~ing the active ingredient in the melt and filli~g the [~¦ SU~S~IIUTE S; --~' P~T/~ ~ 91 û 1 -- ~1 10 9 ~ - 80 20 6 a 2 7 9 ~ 1 Octo~er 199 melt into a two-part hard gelatin capsule.

,, ,CaPsule Fo~mulation d m~/~a~sule ' Active ingredient 250 Lecithin ~oo Arachis Oil 10Q

.
Capsules are prepared by dispersing the active ingredient in ~',.
the lecithin and arachis oil and filling the dispersion into :-~oft, elastic gelatin capsules. -,Capsule For~ulation e fControlled Release CaApsule~
The following controlled release capsule formulation is prepared by extruding ingredients a, b, and c using an extruder, followed by spheronisation of the extrudate and ' drying. The drled pellets are then coated with release~
controlling membrane (d) and filled into a two-piece, hard gelatin capsule.

., i .
: :

e d ~ S U ~ S 5 ~

2 0 6 ~ ~ 7 9 ~ / ~
ctober 1991 mg/capsule (a) Active Ingredient 250 (b~ Microcrystalline Cellulose 125 (c) Lactose BP 125 (d) Ethyl Cellulose ~

513 ~ ~:
-:
Fo,rmulation Exam~le E: In~ectable ~o_mu_ation ~ctive inaredient 0.200 g Sterile, pyrogen free phosphate buffer (pH 7.0) to lO ml The aetive ingredient is dissolved in most of the phGsphate buffer (3~-40~C), then made up to volume and filtered through a sterile micropore filter into a sterile lOml amber glass vial (type l) and sealed with sterile closures and overseals.

Formulation Example F: ,Intramuscular in~ec~ion Active Ingredient 0.20 g Benzyl Alcohol O.lO ~
Glucofurol 75 l.45 g Water for In~ection q.s. ~o 3.00 ml The active ingredient is dissolved in the glyco~urol. The benzyl alcohol is then added and dissolved, and water added to 3 ml. The mixture is then filtered through a sterile micropQre filter and sealed in sterile 3 ml glass vials (type 1) .

[= ¦ S~ E ~ E~

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

~ 19 91 PG~!G~ 9 ~
.
- ~2~ 5 a 2 7 9 ~ I Octobes 1~91 Formulation Example G: Syrup Suspension Active ingredient 0.2500 g S~rbitol S~lution l.5000 g Glycerol 2.0000 g Dispersible Cellulose 0.0750 g S~dium Benzoate 0.0050 g Flavour, Peach 17.42.3l6g 0.0125 ml Purified Water q.s. to 5.0000 ml The sodium benzoate is dis~olved in a portion of the purified .
water and the sorbitol solution added. The active ingredient -is added and di~persed. In the glycerol is dispersed the thickener (dispersibl~ cellulose). The two ~ispersions are mixed and made up to the required volume with the purified .. -water. Further thickening is achieved as required by extra shearing of the suspension.
_ormulation Example H: SUPD~SitOrY
mc~/supDository Active Ingredient (63~!m)* ~50 Hard Fat, BP (Witepsol Hl5 -Dynamit NoBel) 1?70 2020 ~ :

' , . ~ . : . ....
Ur~ d !'.~ n p s~ af~ic~. SUr~z' p~ ~ ficn ,5.3 '.

20~52'~9 - 8 3 - 31 October l~l The active ingredient is used as a powder wherein ~ le1Qt 9 s0~ Df the particles are of 63~m diame~er or less.
One-fifth of the Witepsol H15 is melted in a steam-jacketed pan at 45C maximum. The active ingredient is sifted through a 200~m sieve and added to the molten base with mixing, using a silverson fitted with a cutting head, un il a sm~oth dispersi~n is achieved. Maintaining the mixture at 45~C, the remaining Witepsol H15 is added to the suspension and stirred to ensure a homogenous mix. The entire suspension is passed through a 250~m stainless steel screen and, with c~ntinuous stirring, is allowed to cool to 40C. At a temperature ~f 38~C to 40-C 2.02g of the mixture is filled into suita~le plastic moulds. The suppositories are allowed to cool to room temperature.

F~rmulation Example I: Pessaries m~Pessary Acti~e ingredient 63~m 250 Anydrate Dextr~se 380 Potato Starch 363 Magnesium Stearate 7 1000 .,:
~ ~.
~he above ingredients are mixed directly and pessaries ~-prepared by direct compression of the resulting mixture. ~ .

... . . .. . .. ~. . . : . . .

Claims (23)

1. A pyrimidine 4'-thionucleoside of the formula I
(I) wherein Y is hydroxy or amino, and X is chloro, bromo, iodo, trifluoromethyl, C2-6 alkyl, C2-6 alkenyl, C2-6 haloalkenyl or C2-6 alkynyl or a physiologically functional derivative thereof.

2. A compound according to claim 1 wherein X is C2-3 alkyl, C3-4 alkenyl, halovinyl or C3-4 alkynyl.

3. A compound according to claim 1 or 2 wherein the pyrimidine 4'-thionucleoside is E-5-(2-bromovinyl)-2'-deoxy-4'-thiouridine 2'-deoxy-5-iodo-4'-thiouridine 2'-deoxy-5-ethyl-4'-thiouridine SET A

5-bromo-2'-deoxy-4'-thiouridine 2'-deoxy-5-propynyl-4'-thiouridine 5-chloro-2'-deoxy-4'-thiouridine 2'-deoxy-5-trifluoromethyl-4'-thiouridine 2'-deoxy-5-ethynyl-4'-thiouridine E-5-(2-bromovinyl)-2'-deoxy-4'-thiocytidine 2'-deoxy-5-propyl-4'-thiouridine or E-2'-deoxy-5-(propen-1-yl)-4'-thiouridine.

4. A compound according to any one of claims 1 to 3 wherein the pyrimidine 4'-thionucleoside is the .beta.-anomer.

5. E-5-(2-bromovinyl)-2'-deoxy-4'-thio-.beta.-uridine.

6. 2'-Deoxy-5-ethyl-4'-thio-.beta.-uridine.

7. A physiologically functional derivative of a pyrimidine nucleoside of Formula (I) according to any one of claims 1 to 6.

8. A derivative according to claim 7 which is an alkali metal, alkali earth metal, ammonium, tetra (C1-4 alkyl) ammonium, hydrochloride or acetate salt, or a mono- or di-carboxylic acid ester or an alkali metal, alkali earth metal, ammonium or tetra (C1-4) alkyl ammonium salt.

9. A composition comprising a compound according to any one of claims 1 to 8 in association with a pharmaceutically acceptable carrier or diluent.

10. A compound according to any one of claims 1 to 8 or a composition according to claim 9 for use in a method of treatment SET A

or prophylaxis of virus infections.

11. Use of a compound according to any one of claims 1 to 8 or a composition according to claim 9 in the manufacture of a medicament for use in the treatment or prophylaxis of virus infections.

12. A process for the production of a pyrimidine 4'-thionucleoside of the formula (I) (I) wherein Y is hydroxy or amino, and X is chloro, bromo, iodo, trifluoromethyl, C2-6 alkyl, C2-6 alkenyl, C2-6 haloalkenyl or C2-6 alkynyl or a physiologically functional derivative thereof, which process comprises:
A) reacting a compound of formula (II), SET A

(II) wherein X1 is a precursor for the group X as defined in relation to formula (I); Y is as defined in relation to formula (I); and Z3 and Z5 are the same or different and each is hydrogen or a hydroxyl-protecting group: with a reagent or reagents serving to convert the group X1 to the desired group X; or B) reacting a compound of formula (III) (III) wherein X and Y are as defined in relation to formula (I) or a protected form thereof with a 4-thio sugar derivative serving to SET A

introduce the 4-thio sugar moiety, or a protected form thereof, of formula (I) at the 1-position of the compound of formula (III);
and, where necessary or desired, thereafter optionally effecting any one or more of the following further steps in any desired or necessary order:
a) removing one or more protecting groups, b) converting a compound of formula (I) or a protected form thereof into a further compound of formula (I) or a protected form thereof, c) converting the compound of formula (I) or a protected form thereof into a physiologically functional derivative of the compound of formula (I) or a protected form thereof, d) converting a physiologically functional derivative of the compound of formula (I) or a protected form thereof into the compound of formula (I) or a protected form thereof, e) converting a physiologically functional derivative of the compound of formula (I) or a protected form thereof into another physiologically acceptable derivative of the compound of formula (I) or a protected form thereof, f) separating the .alpha. and .beta. anomers of the compound of formula I or a protected derivative thereof or of a physiologically functional derivative of a compound of formula (I) or a derivative thereof.

13. A process according to claim 12 for the production of a compound according to any one of claims 2 to 8.

14. A process according to claim 12 or claim 13 wherein SET A

PCT/GB90/0109?

the 4-thio sugar derivative is a compound of formula (IV) (IV) where Z3 and Z5 are hydroxy protecting groups and L is a leaving group.

15. A process according to any one of claims 12 to 14 wherein the 4-thio sugar derivative is a 1-acetoxy-4-thio sugar derivative.

16. A compound of formula (IV') ( IV') where Z3 and Z5 are hydroxy protecting groups and L is acetoxy.

17. A compound according to claim 16 wherein Z3 and Z5 are benzyl.

18. A process for the production of a compound of formula (IV') as defined in claim 16 comprising conversion of a compound of formula (IV A) SET A

(IV A) where Z3 and Z5 are hydroxy protecting groups and Ar is an optionally substituted aryl group, by reaction with an appropriate acylating agent (optionally in the presence of acetic acid), in the presence of a mineral acid such as sulphuric acid.

19. A compound of the formula (IV A) (IV A) wherein Z3 and Z5 are hydroxy protecting groups and Ar is an optionally substituted aryl group.

20. A compound according to claim 19 wherein Z3 and Z5 are benzyl and Ar is optionally substituted phenyl.

21. A process for the production of a compound of formula (IV A) ( IV A) where Z3 and Z5 are hydroxy protecting groups and Ar is an SET A

optionally substituted aryl group, which comprises ring closure under basic conditions of a compound of formula (VI) (VI) wherein Z3 and Z5 are hydroxyl protecting groups, Ar is an optionally substituted aryl group and A is an optionally substituted alkyl- or aryl-sulphonyl group.

22. A process for the production of a compound of formula (VI) (VI) wherein Z3 and Z5 are hydroxyl protecting groups, Ar is an optionally substituted aryl group and A is an optionally substituted alkyl- or aryl-sulphonyl group which comprises treatment of a compound of formula (VII) SET A

(VII) wherein Z3 and Z5 are as hydroxy protecting groups and Ar is an optionally substituted aryl group with an optionally substituted alkyl- or aryl- sulphonyl halide:
wherein the compound of formula (VII) is optionally made by reaction of a compound of formula (VIII) (VIII) where Ar, Z3 and Z5 are as defined above, and M is a hydroxyl protecting group, under conditions which remove the group M and leave the groups -S-CH2-Ar, Z3 and Z5 in place;
wherein the compound of formula (VIII) is optionally made by reaction of a compound of formula (IX) SET A

PCT/GB90/0109?

(IX) where Ar, Z3 and Z5 are as defined above, by reaction with a derivative of the group M under neutral conditions in a polar solvent;
wherein the compound of the formula (IX) is optionally made by reaction of a compound of formula (X) (X) where Z3 and Z5 are as defined above, and R is a C1-4 hydrocarbyl group, with a compound of formula Ar-CH2-SH where Ax is as defined above, under acidic conditions at elevated temperature;
wherein the compound of the formula (X) is optionally made by reaction of a compound of formula (XI) (XI) where R is as defined above, with reactive derivatives of the SET A

_ 94 -groups Z3 and Z5 in an organic solvent in the presence of a suitable base, wherein the compound of the formula (XI) is made by reaction of 2-deoxy-D-ribose with an alcohol of formula R-OH in the presence of an acid.

23. A process according to claim 21 wherein the compound of formula (VI) is produced by a process according to claim 22.

SET A