AU2007202803B2 - Organo-phosphorous compounds for activating gamma/delta T cells - Google Patents

Description

P/00/011 Regulation 3.2 AUSTRALIA Patents Act 1990 COMPLETE SPECIFICATION STANDARD PATENT (ORIGINAL) Name of Applicant: Bioagcncy AG, of Schnackenburgallee 116a, Hamburg 22525, Germany Actual Inventors: ALTINCICEK, Boran EBERL, Mathias HINTZ, Martin JOMAA, Hassan KOLLAS, Ann-Kristin REICHENBERG, Armin WIESNER, Jochen WOLF, Oliver Address for Service: DAVIES COLLISON CAVE, Patent & Trademark Attorneys, of I Nicholson Street, Melbourne, 3000, Victoria, Australia Ph: 03 9254 2777 Fax: 03 9254 2770 Attorney Code: DM Invention Title: "Organo-phosphorous compounds for activating gamma/delta T cells" The following statement is a full description of this invention, including the best method of performing it known to us:- Organo-phosphorous compounds for activating gamma/delta T cells This is a divisional of Australian Patent Application No. 2002339423, the entire contents of which are incorporated herein by reference. Numerous diseases in humans and animals are caused by the abnormal functioning of the immune system. Consequently, there is a high demand for substances that are able to regulate the immune system. It is known how to employ the classical acetate/mevalonate pathway in the biosynthesis of isoprenoids (Beycia ED, Porter JW, Annu Rev Biochem. 10 1976;45:113-42), and an alternative method of biosynthesis is known that is independent of mevalonate, namely the 2-methyl-D-erythritol pathway (MEP, synonymous with DOXP) (Rohmer M. Nat Prod Rep. 1999 Oct;16(5):565-74). Both pathways lead to isopentenyl pyrophosphate (IPP), the common precursor of all higher isoprenoids. While the acetate/mevalonate pathway has been known for a long time and has been thoroughly explained, not all biosynthetic reaction steps 15 that occur along the MEP are as yet known. It is known that human gamnma/delta T-cells are activated by one or several intermediates along the MEP. This means that a selective proliferation and cytokine secretion of the gamma/delta T-cell population is brought about during the incubation of peripheral blood lymphocytes with extracts from organisms that 20 possess the MEP (Jomaa H, Feurle J, Luhs K, Kunzmann V, Tony HP, Herderich M, Wilhelm M, FEMS Imunol Med Microbiol, 1999 Sep;25(4):371-8). The exact chemical composition of this/these activating substance(s) is still unknown. The published data indicate that 3-formyl-l-butyl pyrophosphate is formed as a hypothetical intermediate of the MEP and that it plays a role in the activation of 25 gamma/delta T-cells (Belmant C, Espinosa E, Poupot R, Peyrat MA, Guiraud M, Poquet Y, Bonneville M, Fournie JJ, J Biol., Chem. 1999 Nov 5;274(45):32079 84). The present invention advantageously provides substances which are able to stimulate gamma/delta T-cells and thereby have a regulatory effect on the immune system. 30 2 Surprisingly, it has been found that compounds of formula (I) are eminently suitable for the activation of gamma/delta T-cells.

R

33 CH2 R1 2 X1-P -R3 I I R2 0 (I) 5 wherein R, is selected from the group comprising a methyl residue, a formyl residue, substituted and unsubstituted hydroxymethyl residues and CoH 2

R

3 1 , wherein R 31 is selected from the group comprising OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate and R 3 1 10 and R 2 cannot be present in the molecule at the same time,

R

33 is selected from the group comprising hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate,

R

3 is selected from the group comprising hydrogen, substituted and unsubstituted alkyl with 1 to 26 carbon atoms, substituted and unsubstituted hydroxyalkyl with I 15 to 26 carbon atoms, substituted and unsubstituted aryl, substituted and unsubstituted aralkyl, substituted and unsubstituted alkenyl with I to 26 carbon atoms, substituted and unsubstituted alkinyl with I to 26 carbon atoms, substituted and unsubstituted cycloalkyl, substituted and unsubstituted heterocyclic residues, substituted and unsubstituted phosphate, a silyl, a nucleoside, a nucleoside mono-, 20 di- or triphosphate, a deoxynucleoside, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids, and OR 34 , wherein R 34 is defined like R 3 ,

X

2 , inasmuch as a ring is formed between X 2 and C 1 , is defined like X 1 , and 25 otherwise X 2 is selected from the group comprising -OR 6

,

3 R7 0

-X

2 - P-O -Re, 0 wherein R7 and R 8 are defined like R 3 4 ,

-Z;-P--OR

4 0 wherein R 4 is defined like R 3 , and Z, is defined like X 1 , and X 3 , if it forms a ring 5 with C 1 , is defined like X, and, if does not form a ring with C 1 , corresponds to a group X4 1 4 0 wherein R 5 is defined like R 3 , and Z 2 and X 4 , which forms a ring with C 1 , are defined like X 1 , 10 R 2 is selected from the group comprising hydrogen, OH, alkoxy, phenoxy, benzyloxy, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate, X, can be oxygen or Ti

-C

1 Y2 15 wherein Yj and Y 2 can be the same or can be different and are selected from the group comprising H, OH, halogen, an amino residue, a Ci.

9 -alkoxy residue and a C I 9 -alkylthio residue, or together form an oxo group, and a double bond can be present between C 0 and C 1 , or between C 1 and C 2 , or between C 2 and C 3 . 20 4 Preference is given to compounds having the formula:

CH

3

RC

1 C

R

2 0 wherein a single or double bond is present between C 2 and C 3 , R, is selected from 5 the group comprising a methyl residue, a formyl residue and substituted and unsubstituted hydroxymethyl residues,

R

2 is selected from the group comprising hydrogen, hydroxyl, alkoxy, phenoxy and benzyloxy residues, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate, 10 X, is oxygen or corresponds to a group T,

Y

2 wherein Yj and Y 2 can be the same or can be different and are selected from the group comprising H, OH, halogen, amino and CI.

9 -alkoxy and C;.

9 -alkylthio residues, or together form an oxo group, 15 R 3 is selected from the group comprising hydrogen, substituted and unsubstituted alkyl with 1 to 26 carbon atoms, substituted and unsubstituted hydroxyalkyl with I to 26 carbon atoms, substituted and unsubstituted aryl, substituted and unsubstituted aralkyl, substituted and unsubstituted alkenyl with I to 26 carbon atoms, substituted and unsubstituted alkinyl with I to 26 carbon atoms, substituted 20 and unsubstituted cycloalkyl, substituted and unsubstituted heterocyclic residues, substituted and unsubstituted phosphate, a silyl, a nucleoside, a nucleoside mono-, di- or triphosphate, a deoxynucleoside, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds derived from 25 ethylenediamine or amino acids,

X

2 , inasmuch as a ring is formed between X 2 and C 1 , is defined like X 1 , and 5 otherwise X 2 corresponds to

--ZFP-OR

4 0 wherein R4 is defined like R 3 , and Z, is defined like XI, and X 3 , if it forms a ring with C 1 , is defined like X, and, if it does not form a ring with C 1 , corresponds to a 5 group x X4

-

2 -P-OR 0 wherein R 5 is defined like R 3 , and Z 2 and X 4 , which forms a ring with C 1 , are defined like X 1 . 10 Particular preference is given to compounds having formula (IIA) 0 Xg-P-OR 3 R 2 0R (IIA) wherein C 2 and C 3 are linked together by either a single or a double bond, R, is a methyl group or a substituted or unsubstituted hydroxymethyl group, R 2 is 15 hydrogen, OH, a substituted or unsubstituted phosphate or a substituted or unsubstituted pyrophosphate, X, and X 2 are selected from the group comprising 0, CHF, CHCI, CFCI, CH 2 , CF 2 or CCl 2 , and R 3 is selected from the group comprising hydrogen, substituted and unsubstituted phosphate, a nucleoside, a nucleoside mono-, di- or triphosphate, a deoxynucleoside, a cation of an organic or 20 inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids. Preference is also given to compounds having formula (IIB) 6 0?OR P-X2 0 X ~P\

R

1

R

2 (IIB) wherein C 2 and C 3 are linked together by either a single bond or a double bond, R, 5 is a methyl group or a substituted or unsubstituted hydroxymethyl group, R 2 is H if R, is a substituted or unsubstituted hydroxymethyl and is OH, a substituted or unsubstituted phosphate or a substituted or unsubstituted pyrophosphate if R, is a methyl residue, X 1 , X 2 and X 3 are selected from the group comprising 0, CHF, CHCl, CFCl, CH 2 , CF 2 or CCl 2 , and R 3 and R 4 are selected from the group 10 comprising hydrogen, substituted and unsubstituted phosphate, a nucleoside, a nucleoside mono-, di- or triphosphate, a deoxynucleoside, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids. 15 Preference is likewise given to compounds having formula (IIC) 0

R

4 RO- -X 3 -psO R2 p_-OR 3 (IIC) 20 wherein a single or double bond can be present between C 2 and C 3 , R, is a methyl or a substituted or unsubstituted hydroxymethyl group, R 2 is H, OH, a substituted or unsubstituted phosphate or a substituted or unsubstituted pyrophosphate, X 1 , X 2 ,

X

3 and X 4 are selected from the group comprising 0, CHF, CHC, CFCI, CH 2 , CF 2 7 or CC 2 , and R 3 , R 4 and R 5 are selected from the group comprising hydrogen, substituted or unsubstituted phosphate, a nucleoside, a nucleoside mono-, di- or triphosphate, a deoxynucleoside, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, 5 ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids. Moreover, preferred compounds having formulae (II) and (IIA) to (IIC) are those wherein R, is a substituted or unsubstituted hydroxymethyl residue, particularly 10 hydroxymethyl itself or a hydroxymethyl residue substituted by phosphate, diphosphate or nucleoside diphosphate, for example a hydroxymethyl residue substituted by uridine diphosphate, and R 2 = H. Compounds which are likewise preferred are those having formulae (II) and (1IA) 15 to (IIC) in which R, is a methyl residue and R 2 is a hydroxyl residue, a substituted or unsubstituted phosphate residue or a substituted or unsubstituted diphosphate residue, particularly a nucleoside diphosphate residue, e.g. a uridine diphosphate residue. 20 The following compounds are particularly preferred: 8 OR OR OR ,P-O 0 P-O 0 P-C 0 0 P 0 P 0 P HO O OR 3 HOO OR O OR\ 1 2 HO 3 OR O O 0 -9-

R

4 0 OR O OR2HO HO R O OR RRO -s R O --OR5P -O O - ORO pP 3 00 7 8 wherein the residues R 3 , R 4 and Rs are selected from the group comprising hydrogen, ammonium, sodium or potassium. 5 Furthermore, preferred compounds also include those having the following formula; R O0 C( OOR 7R O wherin te rsidus R , R4and R5 ae seectd fr m te grup (mpII)n 9 wherein R 3 1 and R 2 , which cannot both be present in the molecule at the same time, are selected from the group comprising OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a double bond is formed between Ci and C 2 , and a double bond is 5 analogously formed between Co and C 1 if R 2 is present in the molecule; R 33 is selected from the group comprising hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; R 34 is selected from the group comprising hydrogen, substituted or unsubstituted alkyl with 1 to 26 carbon atoms, substituted or unsubstituted hydroxyalkyl with 1 to 26 carbon atoms, 10 substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with I to 26 carbon atoms, substituted or unsubstituted alkinyl with 1 to 26 carbon atoms, substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a deoxynucleoside, a nucleoside mono-, di- or 15 triphosphate, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium or ammonium compounds derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is defined analogously to R 34 , or may be R 7 02

-X

3 2

P-X

33 -Ra II 0 20 wherein R 7 and R 8 are defined like R 34 ; and X 1 , X 32 and X 33 can be the same or can be different and can be oxygen or a group

Y

1

Y

2 wherein Y, and Y 2 can be the same or can be different and are selected from the group comprising H, OH, halogen, an amino residue, a C 1

.

9 -alkoxy residue and a 25 CI.

9 -alkylthio residue, or together form an oxo group. Particularly preferred compounds are those having formula (IlIlA) 10

CH

3 R34 Ri I I C1 C3 0 Rar- CoH2 C2 X- P - XA2 -P-X 33 - Re H R 2 0 0 (1A) wherein R 31 and R 2 , which cannot be present in the molecule at the same time, are selected from the group comprising OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, 5 a double bond is formed between C 1 and C 3 , and a double bond is formed analogously between Co and CI if R 2 is present in the molecule; R 34 , R 7 and RS can be the same or can be different and are defined as above; and XI, X3 and X 3 3 can be the same or can be different and are defined as for compound (11). 10 Hydrogen substituents at C 1 , C 2 and C 3 are not explicitly indicated in formulae (1) to (IIIA) for reasons of clarity. However, carbon atoms are understood as being tetravalent. The missing substituents are therefore hydrogen radicals. Furthermore, preference is given to compounds having formula (lIHA) in which R 31 15 and R 2 are either OH or substituted or unsubstituted phosphate, R34, R 1 and Re are selected from the group comprising substituted and unsubstituted phosphate, a nucleoside, a deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and annonium 20 compounds derived from ethylenediamine or amino acids, and X 1 , X 32 and X 33 can be the same or can be different and are 0, CHF, CHCl, CFCl, CH 2 , CF 2 or CC1 2 . Other preferred compounds having formula (IHA) are those in which the phosphate groups are present as sodium, potassium or substituted or unsubstituted ammonium 25 salts. In another embodiment, the present invention provides a process P Ope\DAH~specMO22764A amended spec. Ispa doc.2V/7(/09 - 10A for the preparation of one compound of formula (III) or (IIIA)

R

33

CH

2

OR

34 C

C

3

H

2 1 I

R

3 1

-COH

2 C2 X -P-X2 H

R

2 O 111 wherein R 31 and R 2 , which cannot both be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate 5 and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a double bond is formed between Ci and C 2 , and a double bond is formed between Co and Ci if R 2 is present in the molecule; R 33 is selected from the group consisting of hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; R 34 is selected from the group consisting of hydrogen, 10 substituted or unsubstituted alkyl with I to 26 carbon atoms, substituted or unsubstituted hydroxyalkyl with 1 to 26 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with 1 to 26 carbon atoms, substituted or unsubstituted alkynyl with I to 26 carbon atoms, substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic 15 residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is 20 defined analogously to R 34 , or may be R7 0

-X

3 2

-P-X

3 3

-R

8

R

7 and R 8 being defined in the same way as R 34 , and X 1 , X 32 and X 33 , which can be P peADAMspme\3022764 amended spec Ispdoc.2310/2009 - 10B identical or different, can be oxygen or a group Y I

-C

I Y and Z, which can be identical or different, being selected from the group consisting of H, OH, halogen, amino, C 1

.

9 -alkoxy and C 1

.

9 -alkylthio, or together forming an oxo 5 group, pharmaceutical salts, esters and spatial isomers thereof, provided that if R 2 is absent, R 33 is H and R 31 is OH, then the entity -XI-PO(OR 34

)-X

2 is not -O-PO(OH)-O-PO(OH) 2 or -O-PO(OH) 2 ; or R34 R7

CH

3 1 0 02 C C 3

H

2 I R3I--COH2 C2 XI--P-X32-P- X33-Rg H R 2 0 IIIA 10 wherein R 31 and R 2 , which cannot be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a double bond is formed between C 1 and C 2 , and a double bond is formed analogously between CO and C 1 if R 2 is present in the molecule; R 34 , R 7 and R8 can be the same or 15 can be different and are defined as in formula (III); and X 1 , X 32 and X 33 can be the same or can be different and are likewise defined as in formula (III), provided that if

R

2 is absent and R 31 is OH, then the entity -X 1

-PO(OR

34

)-X

32

-PO(OR

7

)-X

33

-R

8 is not

-O-PO(OH)-O-PO(OH)

2 ; wherein the lytB gene is deleted, inactivated or modified in selected cells or 20 organisms in order to increase the concentration of one of these compounds. In a further embodiment, the present invention provides a pharmaceutical composition comprising IL-2 and at least one compound of formula III or IIIA P \OperDAHspecUO22764 a e spcisp. doc-22lI/2UU9 -10C

R

33

CH

2

OR

3 4 C C 3 H2, I R3I- COH2 C 2 Xi-P- X 2 H R 2 III wherein R 31 and R 2 , which cannot both be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a 5 double bond is formed between C, and C 2 , and a double bond is formed between Co and C 1 if R 2 is present in the molecule; R 33 is selected from the group consisting of hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; R 34 is selected from the group consisting of hydrogen, substituted or unsubstituted alkyl with I to 26 carbon atoms, substituted or 10 unsubstituted hydroxyalkyl with 1 to 26 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with 1 to 26 carbon atoms, substituted or unsubstituted alkynyl with I to 26 carbon atoms, substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a 15 deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is defined analogously to R 34 , or may be R7 II 0 X32-P--X33-Rs 20 0

R

7 and R 8 being defined in the same way as R 34 , and X 1 , X 32 and X 33 , which can be identical or different, can be oxygen or a group POpeADAmspciG3O227641 named spec Isp doc-23110/2009 - 1OD Y If

-C

II Y and Z, which can be identical or different, being selected from the group consisting of H, OH, halogen, amino, C 1

.

9 -alkoxy and C 1

.

9 -alkylthio, or together forming an oxo group, 5 pharmaceutical salts, esters and spatial isomers thereof; or

R

34 R7

CH

3 01 02 ~C I C 3

-

2 III R3 I- COH2 C2 C 2X1-P- X32-- P--X33-Rs H R2 O 0 IIIA wherein R31 and R 2 , which cannot be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 3 1 is present in the molecule, a 10 double bond is formed between C, and C 2 , and a double bond is formed analogously between Co and C, if R2 is present in the molecule; R34, R 7 and R8 can be the same or can be different and are defined as in formula (III); and X 1 , X 32 and X 3 3 can be the same or can be different and are likewise defined as in formula (III). 15 In another embodiment, the present invention provides an isomerically pure compound of formula III or IIIA R33

CH

2 OR34 C I

C

3

H

2 I

R

31 - COH 2

C

2 Xi-P- X2 H R2 0 III wherein R31 and R2, which cannot both be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate P XOpcADAm4specOz2764$ smeded spec ispadoc-2zI/2cd9 - 10E and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a double bond is formed between Ci and C 2 , and a double bond is formed between Co and C, if R 2 is present in the molecule; R 33 is selected from the group consisting of hydrogen, OH, substituted and unsubstituted phosphate and substituted and 5 unsubstituted pyrophosphate; R 34 is selected from the group consisting of hydrogen, substituted or unsubstituted alkyl with 1 to 26 carbon atoms, substituted or unsubstituted hydroxyalkyl with I to 26 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with 1 to 26 carbon atoms, substituted or unsubstituted alkynyl with 1 to 26 carbon atoms, 10 substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, particularly a metal of the first, second or third main group of the Periodic Table, ammonium, substituted ammonium and ammonium compounds 15 derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is defined analogously to R 34 , or may be R7 O

X

32 - P- X 33

-R

8 0

R

7 and R 8 being defined in the same way as R 34 , and X 1 , X 32 and X 33 , which can be identical or different, can be oxygen or a group Y

-C

20 Z Y and Z, which can be identical or different, being selected from the group consisting of H, OH, halogen, amino, Ci.

9 -alkoxy and C 1

.

9 -alkylthio, or together forming an oxo group, pharmaceutical salts, esters and spatial isomers thereof, P OperDAH\specX30227648 amnded spe, Isp doc.27/10/2009 - IOF provided that if R 2 is absent, R 3 3 is H and R 3 1 is OH, then the entity -Xi-PO(OR 34

)-X

2 is not -O-PO(OH)-O-PO(OH) 2 or -O-PO(OH) 2 ; or R34 R7 CH3 I I 1 01 02 C I C 3

H

2 I

R

31 - COH 2

C

2 X -P-X 3 2 - P- X 3 3 -R H R 2 0 IIIA wherein R 31 and R 2 , which cannot be present in the molecule at the same time, are 5 selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 3 1 is present in the molecule, a double bond is formed between C 1 and C 2 , and a double bond is formed analogously between Co and C, if R 2 is present in the molecule; R 34 , R 7 and RS can be the same or can be different and are defined as in formula (III); and X 1 , X 32 and X 33 can be the 10 same or can be different and are likewise defined as in formula (III), provided that if

R

2 is absent and R3i is OH, then the entity -X 1

-PO(OR

34

)-X

32

-PO(OR

7

)-X

33 -Rs is not -0-PO(OH)-0-PO(OH) 2 . In a further embodiment, the present invention provides a kit for the activation of 7 6 15 T-cells comprising at least one compound of formula (III) or (IIIA), and IL-2. In another embodiment, the present invention provides a method of activating y8 T cells in-vitro, comprising contacting the cells with a compound of formula (III) or (IIIA) in the presence of human IL-2. 20 The following compounds are most suitable: 1l 0-01--P-ONa \ P-O-P-ONa HO I - NaO--O P- PONa ONa ONa NaO H ONa H 9 10 O 0 0 0 0 HO - -O-I-- -ONa ONa $Na ONa I I 11 12 --O--ONa ONa ONa Na QNa (Na OH N9O-P-0 13 14 aONa O 0 0 00 11 O -P -O 0 -- -O~ HO~ O~a ONa OH N-P 13 _ _ _ - V- C H - h - O N a HO O Na P-CHi-P-ONa HO OI 1O1 oa ~ _ ONa ONa OH 18 H 17 0 0 1 \ OH1--CH-P-ONa ONG HO ON ON. H0 H 19 OH 0 O -ONa -- -0Na ONa ONa ONa HO 21 22 Other embodiments of the invention are defined by the subordinate claims. 5 Peculiarities of the abovementioned definitions and suitable Examples of these will be given below: "Alkyl" is a straight-chain or branched-chain alkyl residue with up to 26 carbon atoms, unless otherwise stated, such as methyl, ethyl, propyl, isopropyl, butyl, 10 isobutyl, tert-butyl, pentyl, hexyl and the like.

12 Unless otherwise stated, "alkenyl" includes straight-chain or branched-chain alkenyl groups with up to 26 carbon atoms, e.g. vinyl, propenyl (e.g. 1-propenyl, 2 propenyl), 1-methylpropenyl, 2-methylpropenyl, butenyl, 2-ethylpropenyl, pentenyl and hexenyl. 5 Unless otherwise stated, "alkinyl" includes straight-chain or branched-chain alkinyl groups with up to 26 carbon atoms. Cycloalkyl preferably refers to a C 3

-C

7 -cycloalkyl that may be substituted; alkyl, 10 alkoxy (e.g. methoxy, ethoxy, etc.), halogen (e.g. fluorine, chlorine, bromine, etc.), nitro and the like are suitable as possible substituents. Aryl is an aromatic hydrocarbon residue, such as phenyl, naphthyl, etc., which may have one or more suitable substituents such as alkoxy (e.g. methoxy, ethoxy, etc.), 1 5 halogen (e.g. fluorine, chlorine, bromine, etc.), nitro and the like. "Aralkyl" includes mono-, di- and triphenylalkyls such as benzyl, phenethyl, benzhydryl, trityl and the like, where the aromatic part may have one or more suitable substituents such as alkoxy (e.g. methoxy, ethoxy, etc.), halogen (e.g. 20 fluorine, chlorine, bromine, etc.), nitro and the like. "Alkoxy residue" relates to a straight-chain or branched-chain alkoxy residue with up to 26 carbon atoms, such as methoxy or ethoxy residues, etc., unless otherwise stated. It can be substituted for example by hydroxyl, amino, halogen and oxo 25 groups and alkoxy residues such as methoxy or ethoxy residues. Unless otherwise stated, "hydroxymethyl residue" relates to a residue that has a substituted or unsubstituted Ci-C-alkyl, aryl or aralkyl residue, e.g. methoxymethyl, ethoxymethyl, phenoxymethyl or benzoxymethyl, etc., attached to 30 the oxygen, or has a substituted or unsubstituted phosphate or pyrophosphate residue, such as adenosine diphosphate, uridine diphosphate, etc., attached to the oxygen. Unless otherwise stated, "alkylthio residue" relates to a straight-chain or branched- 13 chain alkylthio residue with up to 9 carbon atoms, such as thiomethyl or thioethyl residues, etc. It can be substituted e.g. by hydroxyl, amino, halogen and oxo groups and alkoxy residues such as methoxy or ethoxy residues. 5 "Silyl residues" can, for example, be substituted by the above-defined alkyl residues or cycloalkyl-(CO- 26 )-alkyl residues. "Silyl-(Co- 26 )-alkyl groups" are silyl residues that can also be bonded to the framework by means of an alkyl residue. The alkyl and silyl groups are defined as 10 above. The alkane and/or arene parts may, if desired, have at least one suitable substituent such as a halogen, alkoxy, hydroxyl, nitro or the like, in the case of the aforementioned esters. 15 Substituted and unsubstituted phosphate residues or substituted and unsubstituted pyrophosphate residues include salt compounds of the corresponding phosphoric acid derivatives with organic or inorganic bases (e.g. sodium salt, potassium salt, calcium salt, aluminium salt, ammonium salt, magnesium salt, triethylamine salt, 20 ethanolamine salt, dicyclohexylamine salt, ethylenediamine salt, N,N'-dibenzyl ethylenediamine salt, etc.), as well as amino acid salts (e.g. arginine salt, lysine salt, glycine salt, alanine salt, ornithine salt, etc.), and also residues in which the phosphate group forms esters with substituted or unsubstituted Ci-C 26 -alkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or 25 unsubstituted cycloalkyl or a substituted or unsubstituted heterocyclic residue, or with a nucleoside or a deoxynucleoside. "Nucleoside" is understood as meaning adenosine, guanosine, uridine, thymidine and cytidine, while "deoxynucleoside" is understood as meaning deoxyadenosine, 30 deoxyguanosine, deoxythymidine, deoxycytidine and deoxyuridine. The invention also relates to the pharmaceutical salts and esters of the salts. Moreover, it includes all spatial isomers of the compounds, both as pure substances and as mixtures thereof.

P pe\DAHspciUO227648 amended spec Ispdc-2210U/2fD9 - 13A In another embodiment, the present invention provides a use of a compound of formula o 0 -O-P--P-ONa, ONa ONa H for the preparation of a medicament for the prophylaxis and/or treatment of human or 5 animal diseases caused by viruses or bacteria. In a further embodiment, the present invention provides a use of a compound of formula o o HO -- O-P--P-ONa, ONa ONa H 10 for the preparation of a medicament for the prophylaxis and/or treatment of human or animal diseases for oral, inhalational, intravenous, parenteral, intravaginal, local or rectal administration. In another embodiment, the present invention provides a use of a compound of 15 formula o 0 y Il II -0- -0-P-ONa, ONa ONa H for the preparation of a medicament for the treatment or prophylaxis of immune diseases, autoimmune diseases, diseases of the respiratory tract or allergies of humans. 20 In a further embodiment, the present invention provides a use of a compound of formula P.OpWDAspcc\M022764R mended spei Ip.doc.-23/I2009 -13B o 0 -O-P-0-P-ONa, ONa ONa H for the preparation of a medicament for the treatment or prophylaxis of a disease selected from the group consisting of asthma, Crohn's disease, ulcerative colitis, multiple sclerosis, bone disease, especially osteoporosis, chronic bronchitis, 5 rheumatoid arthiritis, Hashimoto's thyroiditis, myasthenia gravis, lupus erythematosus, diabetes mellitus, primary biliary cirrhosis, active chronic hepatitis, adrenalitis/Addison's disease, polymyositis, dermatomyositis, autoimmune haemolytic anaemia, myocarditis, pericarditis, scleroderma, uveitis (phacouveitis, sympathetic ophthalmia), pemphigus vulgaris, pemphigoid, pernicious anaemia, 10 autoimmune atrophic gastritis, a benign tumour, a malignant tumour, hepatitis C virus, and helicobacter eradication therapy on ulcers of the gastrointestinal tract. In another embodiment, the present invention provides a use of a compound of formula o o O-P-0-P-ONa, ONa ONa 15 H for the preparation of a medicament for the prophylaxis and/or treatment of diseases of animals or humans wherein the therapeutic amount of active compound is comprised between 0.01 and 2000 pg. 20 In a further embodiment, the present invention provides a method of treatment and/or prophylaxis of immune diseases, autoimmune diseases, respiratory diseases or allergies comprising administering to a patient in need thereof a compound of formula O O I II -O--P--ONa, ONa ONa

H

P :cpeDAmspMcuO22764: amended speci ispdoc.231/flon 9 - 13C In another embodiment, the present invention provides a method of treatment or prophylaxis of a disease selected from the group consisting of asthma, Crohn's disease, ulcerative colitis, multiple sclerosis, bone disease, especially osteoporosis, 5 chronic bronchitis, rheumatoid arthiritis, Hashimoto's thyroiditis, myasthenia gravis, lupus erythematosus, diabetes mellitus, primary biliary cirrhosis, active chronic hepatitis, adrenalitis/Addison's disease, polymyositis, dermatomyositis, autoimmune haemolytic anaemia, myocarditis, pericarditis, scleroderma, uveitis (phacouveitis, sympathetic ophthalmia), pemphigus vulgaris, pemphigoid, pernicious anaemia, 10 autoimmune atrophic gastritis, a benign tumour, a malignant tumour, hepatitis C virus, and helicobacter eradication therapy on ulcers of the gastrointestinal tract, comprising administering to a human or animal in need thereof a compound of formula o 0 || II -O-P-0-P-ONa, Y- ONa ONa H 15 In a further embodiment, the present invention provides a method of treatment and/or prophylaxis of diseases of animals or humans comprising administering to a human or animal in need thereof a compound of formula o 0 IlII HO-P-0-P-ONa, ONa ONa H 20 wherein the therapeutic amount of active compound is comprised between 0.01 and 2000 pg. In another embodiment, the present invention provides a drug comprising a compound of formula P:OpcrDAmH3spe O227648 amended speci Ispa doc-22/U20o9 - 13D o 0 -O-P-0-P-ONa, ONa ONa H and a substance that can be recognized by the immune system as a foreign antigen or autoantigen.

14 The substances according to the invention can be obtained from bacteria, algae, plants and protozoa, including those in which the lytB gene has been deleted, and purified (Example 1). Purification can be effected by means of HPLC or by other methods known per se, such as electrophoresis, precipitation (e.g. as a barium salt) 5 or other chromatographic techniques. Various applications of the compounds are possible. Accordingly, it has been shown, for example, that the substances can be employed in the activity testing of the enzymes GcpE and LytB, as well as in test systems for the measurement of 10 gamma/delta T-cell activation (see Examples 2, 5). The substances according to the invention can either be chemically synthesized (Example 3) or be obtained from bacteria, algae, plants and protozoa and purified (Example 4). Purification can be effected by means of HPLC or by other methods 15 known per se, such as electrophoresis, precipitation (e.g. as a barium salt) or other chromatographic techniques. Furthermore, the substances according to the invention can be used in a screening procedure for the identification of GcpE and LytB enzyme inhibitors, as they are 20 intermediates of the MEP. This method of determining the activity of the enzymes is based on the measurement of differences in the concentration of the enzyme substrates and products under suitable reaction conditions. By bringing suitable test substances into contact with the enzymes during activity determination, inhibitors can be identified by the reduction in the observed enzyme activity. The inhibitors 25 are suitable as herbicides and as active ingredients with antibacterial, antiparasitic and antiviral activity in humans and animals. The compounds according to the invention can also be used in the production of medicines. The efficacy of the compounds is based on the activation of 30 gamma/delta T-cells. Depending on the field of application, the immunological defence mechanism can thereby be strengthened or an immunological tolerance can be induced towards autoantigens and allergens. The fields of application are the treatment of immune and autoimmune diseases 15 and allergies in humans and animals. Examples of these are: allergies, multiple sclerosis, rheumatoid arthritis, Hashimoto's thyroiditis, myasthenia gravis, lupus erythematosus, diabetes mellitus, primary biliary cirrhosis, active chronic hepatitis, adrenalitis/Addison's disease, polymyositis, dermatomyositis, autoimmune 5 haemolytic anaemia, myocardial inflammation and inflammation of the heart membrane, scleroderma, uveitis (phacouveitis, sympathetic ophthalmia), pemphigus vulgaris, pemphigoid, pernicious anaemia, autoimmune atrophic gastritis, inflammatory diseases of the intestine, such as Crohn's disease and ulcerative colitis, and inflammatory diseases of the lung, such as asthmatic and 10 bronchitic ailments. The preferred fields of application are Crohn's disease, ulcerative colitis, multiple sclerosis, asthma, chronic bronchitis and allergies. 15 Furthermore, it has been shown that the substances according to the invention can be successfully employed in the treatment of diseases which are caused by viruses, bacteria and parasites. In particular, the substances according to the invention may be suitable for the 20 prevention and treatment of tumours that are caused by microorganisms. Bacteria, such as Helicobacter pylori (e.g. tumours of the gastrointestinal tract), and papilloma viruses (e.g. tumours of the female genitalia), belong to this group of microorganisms. 25 The compounds defined in the claims are particularly suitable for the prophylaxis and treatment of one of the aforesaid diseases as well as hepatitis C virus infections and benign and malignant tumours, particularly those caused by papilloma viruses, and for helicobacter eradication therapy in cases of ulceration of the gastrointestinal tract. 30 For medicinal purposes, pharmaceutical preparations can be used on their own or in combination with other medicines and can contain either the isolated substances according to the invention or living or dead organisms containing the substances. They are preferably used in combination with substances that are recognized by the 16 immune system as being foreign antigens or autoantigens. Examples of these are myelin basic protein (MBP) and other extracts of the tissue of the nervous system, type I, II or III collagen, thyroglobulin, acetylcholine 5 receptor protein, DNA, islet cell extracts, human insulin, liver extracts, hepatocellular extracts, adrenocortical extracts, skin extracts, heart extracts, muscle extracts, skin cell extracts, haemopoietic line cell extracts, eye lens proteins, S antigens, S-antigen mixtures, stomach cell extracts, parietal cell extracts, intrinsic factor and intestinal extracts. 10 Preferred forms of administration are oral, inhalational, intravenous, parenteral, intracisternal, intravaginal, intraperitoneal, local (powder, ointment, drops) and rectal administration, as well as application to the skin or mucous membranes. The invention includes the administration of an inhalant containing at least one of the 15 substances according to the invention for the treatment of human diseases, particularly allergies and diseases of the respiratory tract such as asthma and chronic bronchitis. Suitable pharmaceutical compositions are moreover: tablets, retard tablets, dragees, 20 capsules, premixes, pills, pellets, boli, aerosols, granules, suppositories, solutions, concentrates, suspensions and emulsions, pastes, ointments, gels, creams, lotions, powders, infusions and sprays. The pharmaceutical formulations may correspond to a fraction or a multiple of a single dose. Dosage units can be 1, 2, 3 or 4 times a single dose, for example, or may contain 1/2, 1/3 or 1/4 of a single dose. A single 25 dose preferably contains the quantity of active ingredient which is used for one administration and which usually corresponds to a whole, a half, a third or a quarter of the daily dosage. Tablets, dragees, capsules, pills and granules may contain the active ingredients in 30 addition to the usual excipients such as (a) fillers and diluents, e.g. starches, lactose, cane sugar, glucose, mannitol and silicic acid, (b) binders, e.g. carboxymethyl cellulose, alginates, gelatine and polyvinylpyrrolidone, (c) humectants, e.g. glycerine, (d) disintegrating agents, e.g. agar-agar, calcium carbonate and sodium carbonate, (e) solution retarders, e.g. paraffin, and (f) 17 absorption accelerators, e.g. quaternary ammonium compounds, (g) wetting agents, e.g. cetyl alcohol and glycerol monostearate, (h) adsorbents, e.g. kaolin and bentonite, and (i) lubricants, e.g. talcum, calcium and magnesium stearate and solid polyethylene glycols, or mixtures of the substances listed under (a) to (i). 5 Moreover, the compounds according to the invention can also be incorporated into other carrier materials such as plastics (plastic chains for local therapy), collagen or bone cement. The tablets, dragees, capsules, pills and granules may be provided with the usual 10 coatings and envelopes optionally containing opaquing agents, and can be prepared in such a way that the active ingredients are released, optionally with a delay, only in the intestinal tract or, preferably, in a particular part of the intestinal tract, it being possible to use e.g. polymer substances and waxes as embedding compounds. 15 The active ingredients can also be in microencapsulated form, optionally with one or more of the aforesaid excipients. In addition to the active ingredients, suppositories may contain the usual water soluble or water-insoluble excipients, e.g. polyethylene glycols, fats, e.g. cacao fat, 20 and higher esters (e.g. a C 4 alcohol with a C 16 fatty acid), or mixtures of these substances. In addition to the active ingredient(s), ointments, pastes, creams and gels can contain the usual excipients, e.g. animal and vegetable fats, waxes, paraffins, 25 starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acids, talcum and zinc oxide, or mixtures of these substances. In addition to the active ingredient(s), powders and sprays may contain the usual excipients, e.g. lactose, talcum, silicic acid, aluminium hydroxide, calcium silicate 30 and polyamide powder, or mixtures of these substances. Additionally, sprays may also contain the usual propellant, e.g. chlorofluorocarbons. In addition to the active ingredients, solutions and emulsions may contain the usual excipients such as solvents, solubilizers and emulsifying agents, e.g. water, ethyl 18 alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils, particularly cottonseed oil, peanut oil, maize oil, olive oil, castor oil and sesame oil, glycerine, glycerol formal, tetrahydrofurfuryl alcohol, polyethylene glycols and 5 fatty acid esters of sorbitol, or mixtures of these substances. There are very marked differences in the amounts of the individual derivatives that are necessary in order to achieve the desired effect. Generally speaking, both in human as well as in veterinary medicine, it has proved to be advantageous if the 10 active ingredient(s) of formula (I) are administered in total amounts of approximately 0.01 to about 2000 pg every 24 hours, if necessary in the form of several single doses, in order to achieve the desired results. A single dose preferably contains the active ingredient(s) in amounts of approximately 0.01 to about 2000 pg. However, it may be necessary to deviate from the abovementioned 15 dosages, depending on the type and body weight of the person to be treated, the nature and severity of the disease, the type of preparation and the administration of the medicine, as well as the period or interval over which the preparation is administered. 20 Consequently, it may be sufficient in a number of cases to manage with less than the abovementioned quantity of active ingredient, while in other cases the afore mentioned quantity of active ingredient will have to be exceeded. The optimum dosage required and the type of administration of the active ingredients can be determined by those skilled in the art on the basis of their specialist knowledge. 25 In the treatment of animals, the compounds to be used according to this invention can be given in the usual concentrations and preparations together with the food or food preparations or with the drinking water. 30 Example I Purification of gamma/delta T-cell activating compounds Various gamma/delta T-cell activating compounds were isolated from Coryne bacterium ammoniagenes. 28 kg of the cell mass were digested with a Dynax mill 19 in 50 mM ammonium formate buffer (pH 8.0). After preabsorption on a hydrophobic polystyrene matrix, the digested material was loaded onto an anion exchanger and eluted with a stepped gradient (100, 300, 500 mM ammonium formate, pH 8.0). The 300 mM eluate was passed through a C- 18 matrix and then 5 through a 3 kDa hollow fibre filter for ultrafiltration. The filtrate was diluted with water to 30 mM ammonium formate and loaded once more onto an anion exchanger. Elution then took place with a linear gradient of 30 to 500 mM ammonium formate. Individual fractions were tested for their ability to activate gamma/delta T-cells. Then some of the active compounds were precipitated as 10 barium salts by the admixture of 100 mM BaCl 2 and 80% EtOH. The precipitates were dissolved in 20 mM ammonium formate buffer (pH 8.0) and rechromatographed on an anion exchanger. In this way it is possible to isolate compounds 1 to 6. 15 Example 2 Activation of gamma/delta T-cells by enriched MEP intermediates Lymphocytes were obtained from the peripheral blood of healthy donors by Ficoll density gradient centrifugation. For each test, 2 x 105 of the cells so obtained were 20 seeded in a volume of 0.2 ml of RPMI-1640 medium (Life Technologies) that was enriched with 25 mM HEPES, 2 mM L-glutamine, 0.025 mg/ml of gentamicin, 100 U/ml of human interleukin-2 (IL-2) (all from Life Technologies) and 10% human AB serum (Bavarian Red Cross). The test fractions were added in various dilutions, and isopentenyl diphosphate (IPP) from Sigma was used in a final 25 concentration of 10 pM as a positive control. Incubation was effected in the incubator with 5% CO 2 at 37*C. After 72 hours the cells were harvested and analysed in a flow cytometer. In so doing, the expression of the CD25 activation marker on the surface of V gamma 9* T-cells was measured with the aid of the monoclonal antibodies CD25-PE (Bl.49.9), V gamma 9-FITC (Immu360) and 30 CD3-PC5 (UCHT1) supplied by Beckman-Coulter. The results showed that compound 1 was approximately 750 times more active than IPP, while compound 2 was about 400 times and compounds 3, 5 and 6 were about 100 times more active than IPP.

20 Example 3 The synthesis was effected in the manner described in diagram 1:

R

1 O R 2

-*

1 0 X1-- P-Xar- OR4

OR

2

OR

3 5 Ia: R, = benzyl; R 2 = OH Ila: Ri = benzyl; X, = X32= 0; R 2

-R

4 = H Ib: R, = benzyl; R 2 Br lIb: R, = benzyl, Xi = 0; X 32 = CH 2 or

CF

2 or CHF; R 2

-R

4 = H Ic: Ri = acetyl; R 2 = Br Ilc: R, = acetyl; X,= X 32 = CH 2 ; R 2

-R

4 = 10 ethyl 0 R10 X1-P-X3r- -OH OH OH 15 Ila: X, = X32 = 0 II1b: X, = 0; X 32 = CH 2 or CF 2 or CHF IIIc: Xi = X32 = CH 2 Diagram 1: Synthesis plan 20 1. Preparation of compounds la to Ic Compounds la and lb were prepared in an analogous manner to that described in K. Sato, S. Inoue, Y. Takagi, S. Morii, Bull. Chem. Soc. Jpn., 1976, 49(11), 3351 3351. 25 The preparation of compound Ic is analogous to that described in H. Kunio, H. Kazushige, Chem. Pharm. Bull., 1994, 42, 4, 786-791. 2. Syntheses of compounds Ila to HIe 21 Compound Ila was prepared according to current methods which are known to those skilled in the art, such as have been described in e.g. B. Woodside, Z. Huang, C. Poulter, Org. Synth. 1988, 66, 211-219, starting from compound lb. 5 Compound Ilb was prepared starting from compound Ia. Ia was first converted to the corresponding tosylate and then reacted e.g. with tns(tetra-n-butylammonium) hydrogenomethylenediphosphate. The synthesis was carried out in a manner analogous to that described in WOOO/59916 and the publications cited therein. 10 Compound Ile in turn was prepared from compound Ic. The syntheses were carried out in the manner described in R.C. McClard and T.S. Fujita, J. Am. Chem. Soc., 1987, 109, 5544-5545. Compound Ilc could be obtained in a low yield and was immediately hydrolysed in 15 order to obtain compound Ic. 3. Syntheses of compounds 1IIa to IIIc In order to prepare compounds Illa to Ic, 500 mg of the corresponding precursors Ila and Ilb were each dissolved in 5 ml of methanol and treated with 10 mol% of 20 hydrogenation catalyst. Then hydrogen was introduced at room temperature and the uptake of hydrogen was measured. After the appropriate amount of hydrogen had been taken up, the mixture was filtered and the solvent was stripped off. The required products Illa and IIlb were obtained with a good degree of purity. Further purification can be achieved by chromatographic methods. Compound IIc was 25 obtained from compound Ile. 200 mg of compound Ile were dissolved in absolute methylene chloride (3 ml) in a heated, argon-flushed flask and 10 eq. of trimethylbromosilane were added at 0 0 C. After stirring for one hour at 0*C, stirring was continued for a further 12 h at room temperature. Finally, an aqueous work-up yielded the required product IIc, which was purified by ion exchange 30 chromatography. In order to test the activation of gamma/delta T-cells, either the isomerically pure compounds or E/Z mixtures of the compounds were used.

22 Example 4 Purification of gamma/delta T-cell activating compounds Various gamma/delta T-cell activating compounds were isolated from Coryne bacterium ammoniagenes. 28 kg of the cell mass were digested with a Dynax mill 5 in 50 mM ammonium formate buffer (pH 8.0). After preabsorption on a hydrophobic polystyrene matrix, the digested material was loaded onto an anion exchanger and eluted with a stepped gradient (100, 300, 500 mM ammonium formate, pH 8.0). The 300 mM cluate was passed through a C-18 matrix and then through a 3 kDa hollow fibre filter for ultrafiltration. The filtrate was diluted with 10 water to 30 mM ammonium formate and loaded once more onto an anion exchanger. Elution then took place with a linear gradient of 30 to 500 mM ammonium formate. Individual fractions were tested for their ability to activate gamma/delta T-cells. Then some of the active compounds were precipitated as barium salts by the admixture of 100 mM BaCl 2 and 80% EtOH. The precipitates 15 were dissolved in 20 mM ammonium formate buffer (pH 8.0) and rechromatographed on an anion exchanger. In this way it was possible to isolate compounds I to 6, 13 and 14. Example 5 20 Activation of gamma/delta T-cells by enriched MEP intermediates Lymphocytes were obtained from the peripheral blood of healthy donors by Ficoll density gradient centrifugation. For each test, 2 x 10s of the cells so obtained were seeded in a volume of 0.2 ml of RPMI-1640 medium (Life Technologies) that was enriched with 25 mM HEPES, 2 mM L-glutamine, 0.025 mg/ml of gentamicin, 100 25 U/ml of human interleukin-2 (IL-2) (all from Life Technologies) and 10% human AB serum (Bavarian Red Cross). The test fractions were added in various dilutions, and isopentenyl diphosphate (IPP) from Sigma was used in a final concentration of 10 pM as a positive control. Incubation was effected in the incubator with 5% CO 2 at 37'C. After 72 hours the cells were harvested and 30 analysed in a flow cytometer. In so doing, the expression of the CD25 activation marker on the surface of V gamma 9' T-cells was measured with the aid of the monoclonal antibodies CD25-PE (Bl.49.9), V gamma 9-FITC (Immu360) and CD3-PC5 (UCHTl) supplied by Beckman-Coulter.

23 The results showed that compound 9 was approximately 10,000 times more active than IPP, compounds 15, 17 and 19 were about 500 times more active, compound 10 was around 1000 times more active and compound 12 was about 50 times more active than IPP. 5 Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps. The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is not, and should not be taken as an acknowledgment or admission or any form of suggestion that that prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates.

Claims (28)

1. Use of a compound of formula o 0 O-P-0-P-ONa, ONa ONa H 5 for the preparation of a medicament for the prophylaxis and/or treatment of human or animal diseases caused by viruses or bacteria.

2. Use of a compound of formula o 0 -O-P-0-P-ONa, Hl ONa ONa H 10 for the preparation of a medicament for the prophylaxis and/or treatment of human or animal diseases for oral, inhalational, intravenous, parenteral, intravaginal, local or rectal administration.

3. The use according to claim 2 for the preparation of a medicament for the treatment 15 of diseases caused by viruses, bacteria or parasites for oral, inhalational, intravenous, parenteral, intravaginal, local or rectal administration.

4. The use according to claim 2 or 3, for the preparation of a medicament for oral administration. 20

5. The use according to claim 2 or 3, for the preparation of a medicament in the form of tablets, retard tablets, dragees, capsules, premixes, pills, pellets, boli aerosols, granules, suppositories, solutions, concentrates, suspensions and emulsions, pastes, ointments, gels, creams, lotions, powders, infusions or sprays. 25

6. The use according to claim 5, for the preparation of a medicament in the form of a tablet. p %OpeAHsp=.s"0227648 .- MW~k Spec, Isp doc.2711M02019 -25

7. The use according to claim 5, for the preparation of a medicament in the form of a capsule. 5

8. Use of a compound of formula o 0 IlII O-P-O-P-ONa, ONa ONa H for the preparation of a medicament for the treatment or prophylaxis of immune diseases, autoimmune diseases, diseases of the respiratory tract or allergies of humans. 10

9. Use of a compound of formula o 0 Il II -0- P -0--P-ONa, ONa ONa H for the preparation of a medicament for the treatment or prophylaxis of a disease selected from the group consisting of asthma, Crohn's disease, ulcerative colitis, 15 multiple sclerosis, bone disease, chronic bronchitis, rheumatoid arthiritis, Hashimoto's thyroiditis, myasthenia gravis, lupus erythematosus, diabetes mellitus, primary biliary cirrhosis, active chronic hepatitis, adrenalitis/Addison's disease, polymyositis, dermatomyositis, autoimmune haemolytic anaemia, myocarditis, pericarditis, scleroderma, uveitis (phacouveitis, sympathetic ophthalmia), pemphigus 20 vulgaris, pemphigoid, pernicious anaemia, autoimmune atrophic gastritis, a benign tumour, a malignant tumour, hepatitis C virus, and helicobacter eradication therapy on ulcers of the gastrointestinal tract.

10. The use according to claim 9, wherein said disease is a benign tumour, a 25 malignant tumour or hepatitis C virus.

11. The use according to claim 9, wherein said tumour is caused by papilloma P peDAflAspcuO22764 smendkd specl spadoc.22t1z/2 O9 - 26 viruses.

12. The use according to claim 9, wherein said treatment is helicobacter eradication therapy on ulcers of the gastrointestinal tract. 5

13. The use according to claim 2, for the preparation of a medicament for the inhalational treatment of allergies and diseases of the respiratory tract.

14. The use according to claim 13, wherein the allergies and diseases of the 10 respiratory tract are asthma or chronic bronchitis.

15. Drug comprising a compound of formula o 0 -0-P--P-ONa, ONa ONa H and a substance that can be recognized by the immune system as a foreign antigen or autoantigen. 15

16. The drug according to claim 15, wherein said substance recognized by the immune system as a foreign antigen is selected from the group consisting of Myelin Basic Protein (MBP), extracts of the nervous system, type I, I or III collagen, thyroglobulin, acetylcholine receptor protein, DNA, islet cell extracts, haemetopoietic 20 line cell extracts, eye lens proteins, S-antigens, S-antigen mixtures, stomach cell extracts, parietal cell extracts, intrinsic factor and intestinal extracts.

17. The drug according to claim 15, wherein said substance recognized by the immune system as a foreign antigen is Myelic basis Protein (MBP). 25

18. Use of a compound of formula 0 0 IlI| O-P-0-P-ONa, ONa ONa H P:pe,'\DAH\spccM022764S amended spei apadoc-27/10/2009 - 27 for the preparation of a medicament for the prophylaxis and/or treatment of diseases of animals or humans wherein the therapeutic amount of active compound is comprised between 0.01 and 2000 pg. 5

19. The use according to claim 18, wherein the therapeutic amount of the compound can be administered one, two, three or four times a day.

20. A method of treatment and/or prophylaxis of immune diseases, autoimmune diseases, respiratory diseases or allergies comprising administering to a patient in 10 need thereof a compound of formula o O O--P-O--ONa, ONa ONa H

21. A method of treatment or prophylaxis of a disease selected from the group consisting of asthma, Crohn's disease, ulcerative colitis, multiple sclerosis, bone disease, chronic bronchitis, rheumatoid arthiritis, Hashimoto's thyroiditis, myasthenia 15 gravis, lupus erythematosus, diabetes mellitus, primary biliary cirrhosis, active chronic hepatitis, adrenalitis/Addison's disease, polymyositis, dermatomyositis, autoimmune haemolytic anaemia, myocarditis, pericarditis, scleroderma, uveitis (phacouveitis, sympathetic ophthalmia), pemphigus vulgaris, pemphigoid, pernicious anaemia, autoimmune atrophic gastritis, a benign tumour, a malignant tumour, 20 hepatitis C virus, and helicobacter eradication therapy on ulcers of the gastrointestinal tract, comprising administering to a human or animal in need thereof a compound of formula o o -O-P-0-P-ONa, ONa ONa H 25

22. A method of treatment and/or prophylaxis of diseases of animals or humans comprising administering to a human or animal in need thereof a compound of formula P OpeeDAspeciu1022764BamendeWdspec Ilpac- 2 7 fO/ 2 00 9 -28 o 0 -H-P-0-P-ONa, H ONa ONa H wherein the therapeutic amount of active compound is comprised between 0.01 and 2000 pg. 5

23. A process for the preparation of one compound of formula (III) or (IIIA) R33 CH 2 I OR 34 C I C 3 H 2 1 R 31 - COH 2 C 2 X,-P-X2 H R 2 O III wherein R 3 1 and R 2 , which cannot both be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 3 1 is present in the molecule, a 10 double bond is formed between C 1 and C 2 , and a double bond is formed between Co and Ci if R 2 is present in the molecule; R 33 is selected from the group consisting of hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; R 34 is selected from the group consisting of hydrogen, substituted or unsubstituted alkyl with I to 26 carbon atoms, substituted or 15 unsubstituted hydroxyalkyl with 1 to 26 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with I to 26 carbon atoms, substituted or unsubstituted alkynyl with I to 26 carbon atoms, substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a 20 deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is defined analogously to R 34 , or may be P:OpeDAHspccMO227648amecnddspco lspa.doc-23/10/2009 - 29 R 7 0 -- X32-P-X33-R8 R 7 and R 8 being defined in the same way as R 34 , and Xi, X 32 and X 33 , which can be identical or different, can be oxygen or a group Y -C 5 Y and Z, which can be identical or different, being selected from the group consisting of H, OH, halogen, amino, CI. 9 -alkoxy and CI. 9 -alkylthio, or together forming an oxo group, pharmaceutical salts, esters and spatial isomers thereof, provided that if R 2 is absent, R 33 is H and R 31 is OH, then the entity -XI-PO(OR 34 )-X 2 10 is not -O-PO(OH)-O-PO(OH) 2 or -O-PO(OH) 2 ; or R 34 R7 CH 3 1 01 02 ~C 1 C 3 H 2 II R 3 1 - CO 2 C 2 X 1 -P- X 32 -P- X 3 3-R H R2 O IIA wherein R31 and R2, which cannot be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R31 is present in the molecule, a 15 double bond is formed between C, and C 2 , and a double bond is formed analogously between CO and C, if R2 is present in the molecule; R 34 , R 7 and R8 can be the same or can be different and are defined as in formula (III); and X 1 , X 32 and X 33 can be the same or can be different and are likewise defined as in formula (111), provided that if R 2 is absent and R31 is OH, then the entity -X 1 -PO(OR 34 )-X 32 -PO(OR 7 )-X 33 -R 8 is not 20 -O-PO(OH)-O-PO(OH) 2 ; P:OpeiDAHspci30227648 amended speci Ispa.doc-27/10/2009 - 30 wherein the lytB gene is deleted, inactivated or modified in selected cells or organisms in order to increase the concentration of one of these compounds.

24. A pharmaceutical composition comprising IL-2 and at least one compound of 5 formula III or IIIA R33 CH 2 OR 34 C C 3 H 2 I R 3 1 - COH 2 C 2 X1-P-X2 H R 2 O 111 wherein R 3 1 and R 2 , which cannot both be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 3 1 is present in the molecule, a 10 double bond is formed between C 1 and C 2 , and a double bond is formed between CO and C, if R 2 is present in the molecule; R 33 is selected from the group consisting of hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; R 34 is selected from the group consisting of hydrogen, substituted or unsubstituted alkyl with 1 to 26 carbon atoms, substituted or 15 unsubstituted hydroxyalkyl with 1 to 26 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with 1 to 26 carbon atoms, substituted or unsubstituted alkynyl with I to 26 carbon atoms, substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a 20 deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is defined analogously to R 34 , or may be F .Opc DAA ,cc3Q22764S mmndcd spwi Ispdoc-291GnW209 - 31 R7 II 0 -X32-P-X33-Rs R 7 and R 8 being defined in the same way as R 34 , and X 1 , X 32 and X 33 , which can be identical or different, can be oxygen or a group Y -C 5 Y and Z, which can be identical or different, being selected from the group consisting of H, OH, halogen, amino, C. 9 -alkoxy and C 1 . 9 -alkylthio, or together forming an oxo group, pharmaceutical salts, esters and spatial isomers thereof; or R34 R7 CH 3 1 01 02 1C I C 3 H 2 I R31- COH2 C2 X 1 -P- X 32 -P- X 33 -R H R 2 0 O IIIA 10 wherein R 31 and R 2 , which cannot be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a double bond is formed between C 1 and C 2 , and a double bond is formed analogously between CO and C 1 if R 2 is present in the molecule; R 34 , R 7 and R 8 can be the same or 15 can be different and are defined as in formula (III); and X 1 , X 32 and X 33 can be the same or can be different and are likewise defined as in formula (III).

25. A kit for the activation of y8 T-cells comprising at least one compound of formula (Ill) or (IIIA), as defined in claim 23, and IL-2. 20 P :prDAHVpcci\227648 amended sP~ i Ispe doc.27/110/2009 -32

26. A method of activating y T-cells in-vitro, comprising contacting the cells with a compound of formula (III) or (IIIA), as defined in claim 24, in the presence of human IL-2. 5

27. An isomerically pure compound of formula III or IIIA R33 CH 2 I OR 3 4 C I C 3 H 2 I R31-COH2 C2 X1-P-X2 H R 2 II wherein R 31 and R 2 , which cannot both be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a 10 double bond is formed between C 1 and C 2 , and a double bond is formed between Co and C, if R 2 is present in the molecule; R 33 is selected from the group consisting of hydrogen, OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; R 34 is selected from the group consisting of hydrogen, substituted or unsubstituted alkyl with 1 to 26 carbon atoms, substituted or 15 unsubstituted hydroxyalkyl with I to 26 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkenyl with I to 26 carbon atoms, substituted or unsubstituted alkynyl with I to 26 carbon atoms, substituted or unsubstituted cycloalkyl, a substituted or unsubstituted heterocyclic residue, substituted or unsubstituted phosphate, a silyl, a nucleoside, a 20 deoxynucleoside, a nucleoside mono-, di- or triphosphate, a cation of an organic or inorganic base, ammonium, substituted ammonium and ammonium compounds derived from ethylenediamine or amino acids; X 2 is either -OR 6 , wherein R 6 is defined analogously to R 34 , or may be - 33 R7 -- X 32 -P- X 33 -R R 7 and R 8 being defined in the same way as R 3 4 , and X 1 , X 32 and X 33 , which can be identical or different, can be oxygen or a group Y -C I 5 Y and Z, which can be identical or different, being selected from the group consisting of H, OH, halogen, amino, CI. 9 -alkoxy and Ci. 9 -alkylthio, or together forming an oxo group, pharmaceutical salts, esters and spatial isomers thereof, provided that if R 2 is absent, R 33 is H and R 31 is OH, then the entity -XI-PO(OR 34 )-X 2 10 is not -O-PO(OH)-O-PO(OH) 2 or -O-PO(OH) 2 ; or R 34 R7 CH 3 I 01 02 C C 3 H 2 I R31- COH2 C 2 Xi-P- X 32 - P- X 33 -R H R 2 O IIIA wherein R 3 1 and R 2 , which cannot be present in the molecule at the same time, are selected from the group consisting of OH, substituted and unsubstituted phosphate and substituted and unsubstituted pyrophosphate; if R 31 is present in the molecule, a 15 double bond is formed between Ci and C 2 , and a double bond is formed analogously between Co and C, if R 2 is present in the molecule; R 34 , R 7 and R 8 can be the same or can be different and are defined as in formula (III); and XI, X 32 and X 33 can be the same or can be different and are likewise defined as in formula (III), provided that if R 2 is absent and R 31 is OH, then the entity -XI-PO(OR 34 )-X 32 -PO(OR 7 )-X 33 -R 8 is not 20 -O-PO(OH)-O-PO(OH) 2 . P:\OperDAH\spcsakO22764K m ded spcci Ispa doc-22/10/2009 - 34

28. The use according to any one of claims 1, 2, 8, 9 and 18, the drug according to claim 15, the method according to any one of claims 20 to 22 or the process according to claim 23, substantially as hereinbefore described and/or exemplified.