BIFUNCTIONAL do2pa DERIVATIVES, CHELATES WITH METALLIC CATIONS AND USE THEREOF

Abstract

Disclosed are chelates resulting from the complexation of bifunctional do2pa derivatives ligands of formula (I), wherein the substituents R.sup.1, R.sup.1′, R.sup.2, R.sup.2′, R.sup.3, R.sup.3′, L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ are defined as in the claims, with metallic cations, especially Pb(II) and Bi(III). Also disclosed are bifunctional do2pa derivatives ligands of formula (I), as well as the use of chelates in nuclear medicine and the use of ligands in cations detection or epuration of effluents.

##STR00001##

Claims

1-14. (canceled)

15. A chelate resulting from the complexation of a ligand of formula (I) ##STR00043## wherein R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ each independently represents: a hydrogen atom; a coupling function, wherein the coupling function is selected from the group consisting of amine; isothiocyanate; isocyanate; activated ester; carboxylic acid; activated carboxylic acid; alcohol; alkyne; halide; azide; siloxy; phosphonic acid; thiol; tetrazine; norbornen; oxoamine; aminooxy; thioether; haloacetamide; glutamate; glutaric anhydride, succinic anhydride, maleic anhydride; aldehyde; ketone; hydrazide; chloroformate; and maleimide; or a bioactive group, wherein the bioactive group is selected from the group consisting of antibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine; solid support; fluorophore; chromophore; macrocyclic chelate; and combinations thereof; L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ each independently represents: a single bond; or a linker selected from the group consisting of alkyl, aryl, arylalkyl, alkylaryl, heteroaryl, heteroarylalkyl, alkylheteroaryl, alkenyl, and alkynyl; wherein alkyl moieties are optionally interrupted by one or more heteroatoms selected from the group consisting of O, N, and S; optionally additionally comprising a residue of a coupling function through which R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ are bounded to L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ respectively; R.sup.3 and R.sup.3′ each independently represents: a hydrogen atom; an activating function, wherein the activating function is selected from the group consisting of N-hydroxysuccinimide, N-hydroxyglutarimide, maleimide, halide, and —OCOR.sup.4, wherein R.sup.4 is selected from the group consisting of alkyl and aryl; or a bioactive group, wherein the bioactive group is selected from the group consisting of antibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine; solid support; fluorophore; chromophore; macrocyclic chelate; and combinations thereof; provided that at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ represents a linker with a coupling function or a linker with a bioactive group; with a metallic cation selected from the group consisting of bismuth (III), lead (II), copper (II), copper (I), gallium (III), zirconium (IV), technetium (III), indium (III), rhenium (VI), astatine (III), yttrium (III), samarium (III), actinium (III), lutetium (III), terbium (III), holmium (III), gadolinium (III), and europium(III).

16. The chelate according to claim 15, wherein, in the coupling function, the activated ester is selected from the group consisting of N-hydroxysuccinimide ester, N-hydroxyglutarimide ester, and maleimide ester; the activated carboxylic acid is selected from the group consisting of anhydride and acid halide; and the haloacetamide is selected from the group consisting of chloroacetamide, bromoacetamide, and iodoacetamide.

17. The chelate according to claim 15, wherein, in the bioactive group, the antibody is selected from the group consisting of polyclonal antibody, monoclonal antibody, hybrid antibody, chimeric antibody, single-domain antibody, dimeric antibody fragment construct, trimeric antibody fragment construct, and minibody; the chelate is spermine; and the solid support is selected from the group consisting of nanoparticle and polymeric microparticle.

18. The chelate according to claim 15, wherein the ligand is of formula (Ia) ##STR00044## wherein R.sup.1, R.sup.1′, L.sup.1 and L.sup.1′ are as defined in claim 15.

19. The chelate according to claim 15, wherein the metallic cation is a radioisotope.

20. The chelate according to claim 15, wherein the metallic cation is a radioisotope selected from the group consisting of .sup.212Bi (.sup.212Pb), .sup.213Bi (III), .sup.64Cu (II), .sup.67Cu (II), .sup.68Ga (III), .sup.89Zr (IV), .sup.99mTC (III), .sup.111In (III), .sup.186Re (VI), .sup.188Re (VI), .sup.211At (III), .sup.225Ac(III), .sup.90Y(III), .sup.177Lu(III), .sup.153Sm(III), .sup.149Tb(III), and .sup.166Ho(III).

21. The chelate according to claim 15, wherein the metallic cation is a radioisotope selected from the group consisting of .sup.212Bi(.sup.212Pb) and .sup.213Bi(III).

22. A pharmaceutical composition comprising the chelate according to claim 15, in association with at least one pharmaceutically acceptable excipient.

23. A ligand of formula (I) ##STR00045## wherein R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ each independently represents: a hydrogen atom; a coupling function, wherein the coupling function is selected from the group consisting of amine; isothiocyanate; isocyanate; activated ester; carboxylic acid; activated carboxylic acid; alcohol; alkyne; halide; azide; siloxy; phosphonic acid; thiol; tetrazine; norbornen; oxoamine; aminooxy; thioether; haloacetamide; glutamate; glutaric anhydride, succinic anhydride, maleic anhydride; aldehyde; ketone; hydrazide; chloroformate, and maleimide; or a bioactive group, wherein the bioactive group is selected from the group consisting of antibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine; solid support; fluorophore; chromophore; macrocyclic chelate; and combinations thereof; L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ each independently represents: a single bond; or a linker selected from the group consisting of alkyl, aryl, arylalkyl, alkylaryl, heteroaryl, heteroarylalkyl, alkylheteroaryl, alkenyl, and alkynyl; wherein alkyl moieties are optionally interrupted by one or more heteroatoms selected from the group consisting of O, N, and S; optionally additionally comprising a residue of a coupling function through which R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ are bounded to L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ respectively; R.sup.3 and R.sup.3′ each independently represents: a hydrogen atom; an activating function, wherein the activating function is selected from the group consisting of N-hydroxysuccinimide, N-hydroxyglutarimide, maleimide, halide, and —OCOR.sup.4, wherein R.sup.4 is selected from the group consisting of alkyl and aryl; or a bioactive group, wherein the bioactive group is selected from the group consisting of antibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine; solid support; fluorophore; chromophore; macrocyclic chelate; and combinations thereof; provided that at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ represents a linker with a coupling function or a linker with a bioactive group.

24. The ligand according to claim 23, wherein, in the coupling function, the activated ester is selected from the group consisting of N-hydroxysuccinimide ester, N-hydroxyglutarimide ester, and maleimide ester; the activated carboxylic acid is selected from the group consisting of anhydride and acid halide; and the haloacetamide is selected from the group consisting of chloroacetamide, bromoacetamide, and iodoacetamide.

25. The ligand according to claim 23, wherein, in the bioactive group, the antibody is selected from the group consisting of polyclonal antibody, monoclonal antibody, hybrid antibody, chimeric antibody, single-domain antibody, dimeric antibody fragment construct, trimeric antibody fragment construct, and minibody; the chelate is spermine; and the solid support is selected from the group consisting of nanoparticle and polymeric microparticle.

26. The ligand according to claim 23, wherein at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ is selected from formulae (a) and (b): ##STR00046## wherein n and m represent each independently an integer ranging from 1 to 10.

27. The ligand according to claim 23, wherein at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ is selected from formulae (a) and (b): ##STR00047## wherein n and m represent each independently 1, 2, 3, or 4.

28. The ligand according to claim 23, of formula (Ia) ##STR00048## wherein R.sup.1, R.sup.1′, L.sup.1 and L.sup.1′ are as defined in claim 23.

29. The ligand according to claim 23, selected from the group consisting of: 6,6′-((4-(4-isothiocyanatobenzyl)-10-methyl-1,4,7,10-tetraazacyclododecane-1,7-diyl)bis(methylene))dipicolinic acid; 6,6′-((4-(3-aminopropyl)-10-methyl-1,4,7,10-tetraazacyclododecane-1,7-diyl)bis(methylene))dipicolinic acid; and 6, 6′-((4-(4-(3-(4-(6-((aminooxy) carbonyl)-2-(16-((aminooxy)carbonyl)-1-(1H-imidazol-4-yl)-4,7,10,18-tetraoxo-3,8,11,17-tetraazanonadecan-19-yl)-21-(1H-imidazol-4-yl)-4,12,15,18-tetraoxo-2,5,11,14,19-pentaazahenicosyl)phenyl)thioureido)benzyl)-10-methyl-1,4,7,10-tetraazacyclododecane-1,7-diyl)bis(methylene))dipicolinate.

30. A process for manufacturing a ligand according to claim 23, comprising starting from cyclen glyoxal: ##STR00049## and performing the following steps: (a1) reacting with compound M.sup.1-L.sup.1-X.sup.1 wherein L.sup.1 is as defined in formula (I); X.sup.1 represents a halogen atom; M.sup.1 represents R.sup.1, wherein R.sup.1 is as defined in formula (I); or M.sup.1 represents a precursor of a coupling function; (a1′) reacting with compound M.sup.1′-L.sup.1′-X.sup.1′ wherein L.sup.1′ is as defined in formula (I); X.sup.1′ represents a halogen atom; M.sup.1′ represents R.sup.1′, wherein R.sup.1′ is as defined in formula (I); or M.sup.1′ represents a precursor of a coupling function; (b1) reacting with compound of formula (i) ##STR00050## wherein L.sup.2 is as defined in formula (I); X represents a halogen atom; M.sup.2 represents R.sup.2, wherein R.sup.2 is as defined in formula (I); or M.sup.2 represents a precursor of a coupling function; M.sup.3 represents a protecting group selected from alkyl group; or R.sup.3, wherein R.sup.3 is as defined in formula (I), provided that it does not represents a hydrogen atom; (b1′) reacting with compound of formula (i′) ##STR00051## wherein L.sup.2′ is as defined in formula (I); X represents a halogen atom; M.sup.2′ represents R.sup.2′, wherein R.sup.2′ is as defined in formula (I); or M.sup.2′ represents a precursor of a coupling function; M.sup.3′ represents a protecting group selected from alkyl group; or R.sup.3′, wherein R.sup.3′ is as defined in formula (I), provided that it does not represents a hydrogen atom; (c) performing glyoxal bridge deprotection; said steps being performed in the following order: (a1), (a1′), (c), (b1) and (b1′); or alternatively (b1), (b1′), (c), (a1) and (a1′); to afford intermediate compound (ii) ##STR00052## wherein L.sup.1, L.sup.1′ L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.1, M.sup.1′, M.sup.2, M.sup.2′, M.sup.3 and M.sup.3′, are as defined above; and where needed, conducting on intermediate compound (ii) one or more subsequent step selected from: in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a precursor of a coupling function, converting the precursor to a coupling function to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a coupling function; in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a coupling function, introducing a bioactive group to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a bioactive group; in the case wherein M.sup.3 or M.sup.3′ represents a protecting group, deprotecting the acidic function, to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represent a hydrogen atom; and introducing an activating function or a bioactive group on the acidic function to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represents an activating function or a bioactive group; to afford compound of formula (I).

31. The process according to claim 30, wherein the alkyl group is selected from the group consisting of methyl, ethyl, and t-butyl.

32. The process according to claim 30, wherein X.sup.1 represents an halogen atom selected from the group consisting of Br and I; X.sup.1′ represents an halogen atom selected from the group consisting of Br and I; and X represents Cl.

33. A process for manufacturing a chelate according to claim 15 comprising reacting a ligand of formula (I) with a metallic cation selected from bismuth (III), lead (II), copper (II), copper (I), gallium (III), zirconium (IV), technetium (III), indium (III), rhenium (VI), astatine (III), yttrium (III), samarium (III), actinium (III), lutetium (III), terbium (III), holmium (III), gadolinium (III), and europium(III); wherein the ligand of formula (I) is defined as ##STR00053## wherein R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ each independently represents: a hydrogen atom; a coupling function, wherein the coupling function is selected from the group consisting of amine; isothiocyanate; isocyanate; activated ester; carboxylic acid; activated carboxylic acid; alcohol; alkyne; halide; azide; siloxy; phosphonic acid; thiol; tetrazine; norbornen; oxoamine; aminooxy; thioether; haloacetamide; glutamate; glutaric anhydride, succinic anhydride, maleic anhydride; aldehyde; ketone; hydrazide; chloroformate, and maleimide; or a bioactive group, wherein the bioactive group is selected from the group consisting of antibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine; solid support; fluorophore; chromophore; macrocyclic chelate; and combinations thereof; L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ each independently represents: a single bond; or a linker selected from the group consisting of alkyl, aryl, arylalkyl, alkylaryl, heteroaryl, heteroarylalkyl, alkylheteroaryl, alkenyl, and alkynyl; wherein alkyl moieties are optionally interrupted by one or more heteroatoms selected from the group consisting of O, N, and S; optionally additionally comprising a residue of a coupling function through which R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ are bounded to L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ respectively; R.sup.3 and R.sup.3′ each independently represents: a hydrogen atom; an activating function, wherein the activating function is selected from the group consisting of N-hydroxysuccinimide, N-hydroxyglutarimide, maleimide, halide, and —OCOR.sup.4, wherein R.sup.4 is selected from the group consisting of alkyl and aryl; or a bioactive group, wherein the bioactive group is selected from the group consisting of antibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine; solid support; fluorophore; chromophore; macrocyclic chelate; and combinations thereof; provided that at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ represents a linker with a coupling function or a linker with a bioactive group.

34. A pharmaceutical composition comprising the chelate according to claim 16, in association with at least one pharmaceutically acceptable excipient.

Description

DETAILED DESCRIPTION

[0072] Ligand

[0073] This invention relates to a bifunctional do2pa derivative ligand of formula (I):

##STR00004## [0074] wherein [0075] R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ each independently represents: [0076] a hydrogen atom; [0077] a coupling function, wherein the coupling function is selected from amine; isothiocyanate; isocyanate; activated ester such as for example N-hydroxysuccinimide ester, N-hydroxyglutarimide ester or maleimide ester; carboxylic acid; activated carboxylic acid such as for example acid anhydride or acid halide; alcohol; alkyne; halide; azide; siloxy; phosphonic acid; thiol; tetrazine; norbornen; oxoamine; aminooxy; thioether; haloacetamide such as for example chloroacetamide, bromoacetamide or iodoacetamide; glutamate; glutaric anhydride, succinic anhydride, maleic anhydride; aldehyde; ketone; hydrazide; chloroformate and maleimide; [0078] a bioactive group, wherein the bioactive group is selected from antibody, such as polyclonal or monoclonal antibody, hybrid or chimeric antibody, single-domain antibody, dimeric or trimeric antibody fragment construct or minibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine such as spermine; solid support such as nanoparticle or polymeric microparticle; fluorophore; chromophore; macrocyclic chelate; and combination thereof; [0079] L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ each independently represents: [0080] a single bond; [0081] a linker selected from alkyl, aryl, arylalkyl, alkylaryl, heteroaryl, heteroarylalkyl, alkylheteroaryl, alkenyl, alkynyl, wherein alkyl moieties are optionally interrupted by one or more heteroatoms selected from O, N and S; optionally additionally comprising a residue of a coupling function through which R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ are bounded to L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ respectively; [0082] R.sup.3 and R.sup.3′ each independently represents: [0083] a hydrogen atom; [0084] an activating function, wherein the activating function is selected from N-hydroxysuccinimide, N-hydroxyglutarimide, maleimide, halide and —OCOR.sup.4, wherein R.sup.4 is selected from alkyl, aryl; [0085] a bioactive group, wherein the bioactive group is selected from antibody, such as polyclonal or monoclonal antibody, hybrid or chimeric antibody, single-domain antibody, dimeric or trimeric antibody fragment construct or minibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine such as spermine; solid support such as nanoparticle or polymeric microparticle; fluorophore; chromophore; macrocyclic chelate; and combination thereof; [0086] provided that at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ represents a linker with a coupling function or a linker with a bioactive group.

[0087] In one embodiment, R.sup.3 and R.sup.3′ are both hydrogen atoms.

[0088] In one embodiment, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ represent both hydrogen atoms.

[0089] In one embodiment, in formula (I), at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ is selected from formulae (a) and (b):

##STR00005## [0090] wherein n and m represent each independently an integer ranging from 1 to 10, preferably 1, 2, 3 or 4.

[0091] According to an embodiment, preferred ligands of formula (I) are of formula (Ia):

##STR00006##

wherein R.sup.1, R.sup.1′, L.sup.1 and L.sup.1′ are as defined in formula (I).

[0092] According to one embodiment, in formula (Ia), L.sup.1′ is an alkyl linker and R.sup.1 is a hydrogen atom.

[0093] According to an embodiment, preferred ligands of formula (Ia) are of formula (Ib):

##STR00007##

wherein R.sup.1 and L.sup.1 are as defined in formula (I).

[0094] According to a specific embodiment, in formulae (Ia) and (Ib), -L.sup.1-R.sup.1 is selected from formulae (a) and (b):

##STR00008##

wherein n and m represent each independently an integer ranging from 1 to 10, preferably 1, 2, 3 or 4.

[0095] According to a specific embodiment, in formulae (Ia) and (Ib), -L.sup.1-R.sup.1 is selected from formulae (a1) and (b1):

##STR00009##

wherein n and m represent each independently an integer ranging from 1 to 10, preferably 1, 2, 3 or 4; and R.sup.1 is as defined in formula (I), preferably R.sup.1 is a bioactive group, more preferably R.sup.1 is a hapten group.

[0096] According to a specific embodiment, the ligand of formula (I) of the invention is grafted on nanoparticles.

[0097] Particularly preferred compounds of formula (I) of the invention are those listed in Table 1 hereafter:

TABLE-US-00002 TABLE 1 Cpd n° Structure Chemical name I-1 [00010] 6,6′-((4-(4-isothiocyanatobenzyl)- 10-methyl-1,4,7,10- tetraazacyclododecane-1,7- diyl)bis(methylene))dipicolinic acid I-2 [00011] 6,6′-((4-(3-aminopropyl)-10- methyl-1,4,7,10- tetraazacyclododecane-1,7- diyl)bis(methylene))dipicolinic acid I-3 [00012] 6,6′-((4-(4-(3-(4-(6- ((aminooxy)carbonyl)-2-(16- ((aminooxy)carbonyl)-1-(1H- imidazol-4-yl)-4,7,10,18-tetraoxo- 3,8,11,17-tetraazanonadecan-19- yl)-21-(1H-imidazol-4-yl)- 4,12,15,18-tetraoxo-2,5,11,14,19- pentaazahenicosyl)phenyl) thioureido)benzyl)-10-methyl- 1,4,7,10-tetraazacyclododecane- 1,7-diyl)bis(methylene))dipicolinate In Table 1, the term “Cpd” means compound.

[0098] The compounds of Table 1 were named using ChemBioDraw® Ultra version 12.0 (PerkinElmer).

[0099] Chelate

[0100] The present invention further relates to a chelate resulting from the complexation of a ligand of the invention of formula (I) as described above and a metallic cation, preferably lead (II) or bismuth (III). In one embodiment, the metallic cation is selected from the group consisting of Pb (II) and Bi(III).

[0101] Unless otherwise stated Pb (II) and Bi(III) encompass their radioactive isotopes, such as .sup.212Pb, .sup.212Bi and .sup.213Bi.

[0102] In an embodiment, the present invention relates to a chelate resulting from the complexation of a ligand of formula (I)

##STR00013## [0103] wherein [0104] R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ each independently represents: [0105] a hydrogen atom; [0106] a coupling function, wherein the coupling function is selected from amine; isothiocyanate; isocyanate; activated ester such as for example N-hydroxysuccinimide ester, N-hydroxyglutarimide ester or maleimide ester; carboxylic acid; activated carboxylic acid such as for example acid anhydride or acid halide; alcohol; alkyne; halide; azide; siloxy; phosphonic acid; thiol; tetrazine; norbornen; oxoamine; aminooxy; thioether; haloacetamide such as for example chloroacetamide, bromoacetamide or iodoacetamide; glutamate; glutaric anhydride, succinic anhydride, maleic anhydride; aldehyde; ketone; hydrazide; chloroformate and maleimide; [0107] a bioactive group, wherein the bioactive group is selected from antibody, such as polyclonal or monoclonal antibody, hybrid or chimeric antibody, single-domain antibody, dimeric or trimeric antibody fragment construct or minibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine such as spermine; solid support such as nanoparticle or polymeric microparticle; fluorophore; chromophore; macrocyclic chelate; and combination thereof; [0108] L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ each independently represents: [0109] a single bond; [0110] a linker selected from alkyl, aryl, arylalkyl, alkylaryl, heteroaryl, heteroarylalkyl, alkylheteroaryl, alkenyl, alkynyl, wherein alkyl moieties are optionally interrupted by one or more heteroatoms selected from O, N and S; optionally additionally comprising a residue of a coupling function through which R.sup.1, R.sup.1′, R.sup.2 and R.sup.2′ are bounded to L.sup.1, L.sup.1′, L.sup.2 and L.sup.2′ respectively; [0111] R.sup.3 and R.sup.3′ each independently represents: [0112] a hydrogen atom; [0113] an activating function, wherein the activating function is selected from N-hydroxysuccinimide, N-hydroxyglutarimide, maleimide, halide and —OCOR.sup.4, wherein R.sup.4 is selected from alkyl, aryl; [0114] a bioactive group, wherein the bioactive group is selected from antibody, such as polyclonal or monoclonal antibody, hybrid or chimeric antibody, single-domain antibody, dimeric or trimeric antibody fragment construct or minibody; hapten; peptide; protein; polysaccharide; fatty acid; liposome; lipid; polyamine such as spermine; solid support such as nanoparticle or polymeric microparticle; fluorophore; chromophore; macrocyclic chelate; and combination thereof; [0115] provided that at least one of -L.sup.1-R.sup.1, -L.sup.1′-R.sup.1′, -L.sup.2-R.sup.2 and -L.sup.2′-R.sup.2′ represents a linker with a coupling function or a linker with a bioactive group;
with a metallic cation selected from bismuth (III), lead (II), copper (II), copper (I), gallium (III), zirconium (IV), technetium (III), indium (III), rhenium (VI), astatine (III), yttrium (III), samarium (III), actinium (III), lutetium (III), terbium (III), holmium (III), gadolinium (III), europium (III); preferably selected from bismuth (III) and lead (II).

[0116] According to a preferred embodiment, the metallic cation is a radioisotope, preferably a radioisotope selected from .sup.212Bi (.sup.212Pb), .sup.213Bi(III), .sup.64Cu(II), .sup.67Cu(II), .sup.68Ga(III), .sup.89Zr(IV), .sup.99mTc(III), .sup.111In(III), .sup.186Re(VI), .sup.188Re(VI), .sup.211At(III), .sup.225Ac(III), .sup.90Y(III), .sup.177Lu(III), .sup.153Sm(III), .sup.149Tb(III) or .sup.166Ho(III), more preferably .sup.212Bi (.sup.212Pb), .sup.213Bi(III).

[0117] According to a preferred embodiment, the metallic cation is a radioisotope, preferably the metallic cation is the in situ .sup.212Pb/.sup.212Bi generator, more preferably the metallic cation is .sup.212Pb.

[0118] When the metallic cation is a radioisotope, the chelate of the invention is a radiopharmaceutical.

[0119] Preferred embodiments relative to the ligand of formula I described above apply to the chelate of the invention.

[0120] According to one embodiment, the ligand and/or the chelate of the invention may be grafted on solid support, such as for example nanoparticles, preferably gold, iron, or quantum dots According to one embodiment, the ligand and/or the chelate of the invention may be grafted on solid support, such as for example silicium, alumine or resin.

[0121] According to one embodiment, the ligand and/or the chelate of the invention may be linked to other ligands/chelates, such as for example porphyrines, cyclodextrines, calixarenes or azacycloalkanes. In this case, the resulting chelate may be used for bimodal imaging or for theragnostic.

[0122] Process of Manufacturing—Ligand and Chelate

[0123] The present invention further relates to a process for manufacturing the ligand of the invention. According to one embodiment, the ligand of formula (I) may be obtained starting from cyclen glyoxal:

##STR00014##

[0124] Starting from cyclen glyoxal, pendant arms -L.sup.1-R.sup.1 and -L.sup.1′-R.sup.1′ may first be introduced (steps (a1) and (a1′)), followed by picolinate arms (steps (b1) and (b1′)) or alternatively, picolinate arms may first be introduced (steps (b1) and (b1′)) before pendant arms -L.sup.1-R.sup.1 and -L.sup.1′-R.sup.1′ (steps (a1) and (a1′)). In both cases, a deprotection step of the glyoxal bridge (step (c)) should be performed after the functionalization of the 2 first amines and before the functionalization of the 2 remaining amines. These synthesis routes are summarized in the scheme below:

##STR00015## ##STR00016##

[0125] Therefore, according to one embodiment, the process for manufacturing the ligand of formula (I) of the invention comprises starting from cyclen glyoxal:

##STR00017##

and performing the following steps: [0126] (a1) reacting with compound M.sup.1-L.sup.1-X.sup.1 wherein [0127] L.sup.1 is as defined in formula (I); [0128] X.sup.1 represents a halogen atom, preferably Br or I; [0129] M.sup.1 represents R.sup.1, wherein R.sup.1 is as defined in formula (I); or M.sup.1 represents a precursor of a coupling function; [0130] (a1′) reacting with compound M.sup.1′-L.sup.1′-X.sup.1′ wherein [0131] L.sup.1′ is as defined in formula (I); [0132] X.sup.1′ represents a halogen atom, preferably Br or I; [0133] M.sup.1′ represents R.sup.1, wherein R.sup.1′ is as defined in formula (I); or M.sup.1′ represents a precursor of a coupling function; [0134] (b1) reacting with compound of formula (i)

##STR00018## [0135] wherein [0136] L.sup.2 is as defined in formula (I); [0137] X represents a halogen atom, preferably Cl; [0138] M.sup.2 represents R.sup.2, wherein R.sup.2 is as defined in formula (I); or M.sup.2 represents a precursor of a coupling function; [0139] M.sup.3 represents [0140] a protecting group selected from alkyl group, preferably methyl, ethyl or t-butyl, more preferably methyl; or [0141] R.sup.3, wherein R.sup.3 is as defined in formula (I), provided that it does not represents a hydrogen atom; [0142] (b1′) reacting with compound of formula (i′)

##STR00019## [0143] wherein [0144] L.sup.2′ is as defined in formula (I); [0145] X represents a halogen atom, preferably Cl; [0146] M.sup.2′ represents R.sup.2′, wherein R.sup.2′ is as defined in formula (I); or M.sup.2′ represents a precursor of a coupling function; [0147] M.sup.3′ represents [0148] a protecting group selected from alkyl group, preferably methyl, ethyl or t-butyl, more preferably methyl; or [0149] R.sup.3′, wherein R.sup.3′ is as defined in formula (I), provided that it does not represents a hydrogen atom; [0150] (c) performing glyoxal bridge deprotection;
said steps being performed in the following order: (a1), (a1′), (c), (b1) and (b1′); or alternatively (b1), (b1′), (c), (a1) and (a1′);
to afford intermediate compound (ii)

##STR00020## [0151] wherein L.sup.1, L.sup.1′ L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.1, M.sup.1′, M.sup.2, M.sup.2′ M.sup.3 and M.sup.3′, are as defined above;
and where needed, conducting on intermediate compound (ii) one or more subsequent step selected from: [0152] in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a precursor of a coupling function, converting the precursor to a coupling function to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a coupling function; [0153] in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a coupling function, introducing a bioactive group to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a bioactive group; [0154] in the case wherein M.sup.3 or M.sup.3′ represents a protecting group, deprotecting the acidic function, to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represent a hydrogen atom; [0155] introducing an activating function or a bioactive group on the acidic function to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represents an activating function or a bioactive group;
to afford compound of formula (I).

[0156] In the present invention a “precursor of a coupling function” refers to a coupling function with a protective group or to a chemical moiety which may be interconverted to afford a coupling function. An example of a coupling function with a protective group is a protected amine group, wherein the amino-protecting group is of common knowledge for a person skilled in the art, such as for example a Boc or a Fmoc group. An example of chemical moiety which may be interconverted to afford a coupling function is a nitro group, which may be converted into amine or even into isothiocyanate coupling function.

[0157] According to one embodiment, step (c) of deprotection of the glyoxal bridge may be performed using hydrazine hydrate or ethylenediamine.

[0158] According to a preferred embodiment, step (c) of deprotection of the glyoxal bridge is performed using ethylenediamine. Preferably, deprotection using ethylenediamine is performed in mild conditions. Especially, deprotection may be performed in a solvent such as dichloromethane. Moreover, deprotection is preferably performed at room temperature.

[0159] When pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are different or when picolinate arms are different, the first substitution of cyclen glyoxal, through step (a1) or (b1), should be controlled in order to obtain the mono-substituted intermediate (iii) or (vi). In a preferred embodiment, reaction of cyclen glyoxal through step (a1) or (b1) is conducted in THF to favor mono-addition, leading to the precipitation of intermediate (iii) or (vi) respectively; and the second arm is introduced through step (a1′) or (b1′), using acetonitrile as solvent, leading to intermediate (iii′) or (vi′). In one embodiment, intermediate (iii) or (vi) is not further purified before undergoing step (a1′) or (b1′), respectively.

[0160] When pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are identical or when picolinate arms are identical, the substitution of cyclen glyoxal, through step (a1) or (b1) is performed in presence of an excess of reactant, to afford directly intermediate (iii′) or (vi′), respectively. Preferably, an excess of more than 2 molar equivalents is used, more preferably from 2 to 4 equivalents.

[0161] When picolinate arms are different or when pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are different, the first substitution of deprotected intermediate (iv) or (vii), through step (b1) or (a1), should be controlled in order to obtain the mono-substituted intermediate (v) or (viii). Mono-substitution is favored by using 1 equivalent of reactant. Preferably, the more bulky arm is first introduced, so that the steric hindrance favor the mono-addition. In one embodiment, intermediate (v) or (viii) is purified before undergoing step (b1′) or (a1′), respectively, preferably by column chromatography.

[0162] When picolinate arms are identical or when pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are identical, the substitution of deprotected intermediate (iv) or (vii), through step (b1) or (a1) is performed in presence of an excess of reactant, to afford directly intermediate (ii). Preferably, an excess of more than 2 molar equivalents is used, more preferably from 2 to 4 equivalents.

[0163] In the case wherein pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are different and picolinate arms are different, synthesis of intermediate (ii) may be performed either by steps (a1), (a1′), (c), (b1) and (b1′) or alternatively by steps (b1), (b1′), (c), (a1) and (a1′).

[0164] In the case wherein pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are different and picolinate arms are identical, synthesis of intermediate (ii) is preferably performed in the following order: (a1), (a1′), (c), (b1).

[0165] In the case wherein pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are identical and picolinate arms are different, synthesis of intermediate (ii) is preferably performed in the following order: (b1), (b1′), (c), (a1).

[0166] In the case wherein pendent arms -L.sup.1-M.sup.1 and L.sup.1′-M.sup.1′ are identical and picolinate arms are identical, synthesis of intermediate (ii) may be performed either by steps (a1), (c) and (b1) or alternatively by steps (b1), (c) and (a1).

[0167] According to a first embodiment, the process for manufacturing the ligand of formula (I) of the invention comprises reacting cyclen glyoxal:

##STR00021##

(a1) with compound M.sup.1-L.sup.1-X.sup.1 wherein [0168] L.sup.1 is as defined in formula (I); [0169] X.sup.1 represents a halogen atom, preferably Br or I; [0170] M.sup.1 represents R.sup.1, wherein R.sup.1 is as defined in formula (I); or M.sup.1 [0171] represents a precursor of a coupling function;
to afford compound (iii)

##STR00022## [0172] wherein L.sup.1 and M.sup.1 are as defined above;
(a1′) reacting (iii) with compound M.sup.1′-L.sup.1′-X.sup.1′ wherein [0173] L.sup.1′ is as defined in formula (I); [0174] X.sup.1′ represents a halogen atom, preferably Br or I; [0175] M.sup.1′ represents R.sup.1′, wherein R.sup.1′ is as defined in formula (I); or M.sup.1′ represents a precursor of a coupling function;
to afford compound (iii′)

##STR00023## [0176] wherein L.sup.1, L.sup.1′, M.sup.1 and M.sup.1′ are as defined above;
(c) deprotecting glyoxal bridge of compound (iii′), to afford compound (iv)

##STR00024## [0177] wherein L.sup.1, L.sup.1′, M.sup.1 and M.sup.1′ are as defined above;
(b1) reacting (iv) with compound of formula (i)

##STR00025## [0178] wherein [0179] L.sup.2 is as defined in formula (I); [0180] X represents a halogen atom, preferably Cl; [0181] M.sup.2 represents R.sup.2, wherein R.sup.2 is as defined in formula (I); or M.sup.2 represents a precursor of a coupling function; [0182] M.sup.3 represents [0183] a protecting group selected from alkyl group, preferably methyl, ethyl or t-butyl, more preferably methyl; or [0184] R.sup.3, wherein R.sup.3 is as defined in formula (I), provided that it does not represents a hydrogen atom;
to afford compound (v)

##STR00026## [0185] wherein L.sup.1, L.sup.1′ L.sup.2 are as defined in formula (I) and wherein M.sup.1, M.sup.1′, M.sup.2, M.sup.3 are as defined above;
(b1′) reacting (v) with compound of formula (i′)

##STR00027## [0186] wherein [0187] L.sup.2′ is as defined in formula (I); [0188] X represents a halogen atom, preferably Cl; [0189] M.sup.2′ represents R.sup.2′, wherein R.sup.2′ is as defined in formula (I); or M.sup.2′ represents a precursor of a coupling function; [0190] M.sup.3′ represents [0191] a protecting group selected from alkyl group, preferably methyl, ethyl or t-butyl, more preferably methyl; or [0192] R.sup.3′, wherein R.sup.3′ is as defined in formula (I), provided that it does not represents a hydrogen atom;
to afford intermediate compound (ii)

##STR00028## [0193] wherein L.sup.1, L.sup.1′ L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.1, M.sup.1′, M.sup.2, M.sup.2′, M.sup.3 and M.sup.3′, are as defined above;
and where needed, conducting on intermediate compound (ii) one or more subsequent step selected from: [0194] in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a precursor of a coupling function, converting the precursor to a coupling function to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a coupling function; [0195] in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a coupling function, introducing a bioactive group to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a bioactive group; [0196] in the case wherein M.sup.3 or M.sup.3′ represents a protecting group, deprotecting the acidic function, to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represent a hydrogen atom; [0197] introducing an activating function or a bioactive group on the acidic function to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represents an activating function or a bioactive group;
to afford compound of formula (I).

[0198] According to a second embodiment, the process for manufacturing the ligand of formula (I) of the invention comprises reacting cyclen glyoxal:

##STR00029##

(b1) with compound of formula (i)

##STR00030## [0199] wherein [0200] L.sup.2 is as defined in formula (I); [0201] X represents a halogen atom, preferably Cl; [0202] M.sup.2 represents R.sup.2, wherein R.sup.2 is as defined in formula (I); or M.sup.2 represents a precursor of a coupling function; [0203] M.sup.3 represents [0204] a protecting group selected from alkyl group, preferably methyl, ethyl or t-butyl, more preferably methyl; or [0205] R.sup.3, wherein R.sup.3 is as defined in formula (I), provided that it does not represents a hydrogen atom;
to afford compound (vi)

##STR00031## [0206] wherein L.sup.2 is as defined in formula (I) and wherein M.sup.2 and M.sup.3 are as defined above;
(b1′) reacting (vi) with compound of formula (i′)

##STR00032## [0207] wherein [0208] L.sup.2′ is as defined in formula (I); [0209] X represents a halogen atom, preferably Cl; [0210] M.sup.2′ represents R.sup.2′, wherein R.sup.2′ is as defined in formula (I); or M.sup.2′ represents a precursor of a coupling function; [0211] M.sup.3′ represents [0212] a protecting group selected from alkyl group, preferably methyl, ethyl or t-butyl, more preferably methyl; or [0213] R.sup.3′, wherein R.sup.3′ is as defined in formula (I), provided that it does not represents a hydrogen atom;
to afford intermediate compound (vi′)

##STR00033## [0214] wherein L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.2, M.sup.2′, M.sup.3 and M.sup.3′, are as defined above;
(c) deprotecting glyoxal bridge of compound (vi′), to afford compound (vii)

##STR00034## [0215] wherein L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.2, M.sup.2′, M.sup.3 and M.sup.3′, are as defined above;
(a1) reacting (vii) with compound M.sup.1-L.sup.1-X.sup.1 wherein [0216] L.sup.1 is as defined in formula (I); [0217] X.sup.1 represents a halogen atom, preferably Br or I; [0218] M.sup.1 represents R.sup.1, wherein R.sup.1 is as defined in formula (I); or M.sup.1 represents a precursor of a coupling function;
to afford compound (viii)

##STR00035## [0219] wherein L.sup.1, L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.1, M.sup.2, M.sup.2′, M.sup.3 and M.sup.3′, are as defined above;
(a1′) reacting (viii) with compound M.sup.1′-L.sup.1′-X.sup.1′ wherein [0220] L.sup.1′ is as defined in formula (I); [0221] X.sup.1′ represents a halogen atom, preferably Br or I; [0222] M.sup.1′ represents R.sup.1′, wherein R.sup.1′ is as defined in formula (I); or M.sup.1′ represents a precursor of a coupling function;
to afford compound (iii′)

##STR00036## [0223] wherein L.sup.1, L.sup.1′, M.sup.1 and M.sup.1′ are as defined above;
to afford intermediate compound (ii)

##STR00037## [0224] wherein L.sup.1, L.sup.1′ L.sup.2, L.sup.2′ are as defined in formula (I) and wherein M.sup.1, M.sup.1′, M.sup.2, M.sup.2′, M.sup.3 and M.sup.3′, are as defined above;
and where needed, conducting on intermediate compound (ii) one or more subsequent step selected from: [0225] in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a precursor of a coupling function, converting the precursor to a coupling function to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a coupling function; [0226] in the case wherein M.sup.1, M.sup.1′, M.sup.2 or M.sup.2′ represents a coupling function, introducing a bioactive group to afford compound of formula (I) wherein R.sup.1, R.sup.1′, R.sup.2 or R.sup.2′ respectively represents a bioactive group; [0227] in the case wherein M.sup.3 or M.sup.3′ represents a protecting group, deprotecting the acidic function, to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represent a hydrogen atom; [0228] introducing an activating function or a bioactive group on the acidic function to afford compound of formula (I) wherein R.sup.3 or R.sup.3′ represents an activating function or a bioactive group;
to afford compound of formula (I).

[0229] The present invention further relates to a process of manufacturing of the chelate of the invention.

[0230] According to one embodiment, the process for manufacturing a chelate according to the invention comprises reacting a ligand of formula (I) according to the invention with a metallic cation selected from bismuth (III), lead (II), copper (II), copper (I), gallium (III), zirconium (IV), technetium (III), indium (III), rhenium (VI), astatine (III), yttrium (III), samarium (III), actinium (III), lutetium (III), terbium (III), holmium (III), gadolinium (III), europium(III). In a specific embodiment, the process for manufacturing a chelate according to the invention comprises reacting a ligand of formula (I) according to the invention with a metallic cation selected from bismuth (III), lead (II), preferably lead (II).

[0231] In an embodiment, the process of manufacturing the chelate of the invention comprises reacting the ligand of formula (I) of the invention with a metallic cation in an aqueous medium, preferably by adjusting the pH around neutrality with KOH. The process of the invention is preferably conducted at a temperature ranging from room temperature to reflux, preferably at room temperature. Chelation process is generally performed for a period ranging from few minutes to 24 hours, preferably a few minutes.

[0232] In an embodiment, the metallic cation used in the process the invention is under the form of a salt, preferably perchlorate, chloride, bromide, nitrates, sulfates, acetate, triflate salts.

[0233] Use of the Chelate

[0234] The invention is further directed to the use of the chelates of the invention in nuclear medicine, preferably as imaging agents and/or medicaments, preferably as radiopharmaceuticals.

[0235] The chelates of the invention are useful as medicaments. In particular, chelates of radioisotopes, preferably chelates of .sup.212Bi (.sup.212Pb), .sup.213Bi, .sup.225Ac, .sup.67Cu, .sup.188Re, .sup.211At, .sup.90Y, .sup.153Sm, .sup.177Lu or .sup.149Tb may be used in RIT. Depending on the bioactive group present on the chelate, a broad variety of diseases may be targeted. For example, the following diseases may be targeted using specified bioactive groups:

TABLE-US-00003 Bioactive group Diseases name type lymphomes anti-CD20 antibody prostate cancer anti-CEA antibody bombesine peptide anti-PSMA antibody/ minibody Lys-urea-Asp sequence small molecule breast cancer anti-HER2 antibody colorectal cancer anti-EGFR antibody neuroendocrine tumors somatostatine analogues such peptide as octreotide, TATE, TOC tumoral RGD analogues (for integrin peptide neoangiogenesis targeting) bones cancer bisphosphonate derivatives small molecule medullar thyroid cancer minigastrin peptide melanoma α-MSH analogs peptide benzamide derivatives small molecule glioblastoma P substance peptide cancers (leukemia, chemokine antagonists peptides breast, prostate, pancreas, lung, ovarian, colorectal . . . )

[0236] The chelates of the invention are also useful as imaging agents. In particular, chelates of radioisotopes, preferably chelates of .sup.64Cu, .sup.68Ga, .sup.89Zr, .sup.99mTc, .sup.111In, .sup.186Re, or .sup.166Ho may be used in PET imaging and/or in SPECT imaging. Chelates of gadolinium (III) may be used in MRI imaging. Chelates of lanthanides, preferably chelates of Eu(III), Tb(III) or Yb(III), may be used for imaging by luminescence.

[0237] The invention thus provides methods of treatment and/or prevention of diseases, comprising the administration of a therapeutically effective amount of a chelate of the invention, preferably a chelate of a radioisotope, to a patient in need thereof.

[0238] The invention further provides the use of a chelate of the invention, preferably a chelate of a radioisotope, for the manufacture of a medicament, preferably a radiopharmaceutical.

[0239] According to one embodiment, the chelates of the invention may be administered as part of a combination therapy. Thus, are included within the scope of the present invention embodiments comprising co-administration of a compound of the present invention as active ingredient and additional therapeutic agents and/or active ingredients.

[0240] The present invention further relates to a pharmaceutical composition comprising the chelate of the invention in association with at least one pharmaceutically acceptable excipient.

[0241] The present invention further relates to a medicament comprising the chelate of the invention.

[0242] Generally, for pharmaceutical use, the chelates of the invention may be formulated as a pharmaceutical preparation comprising at least one chelate of the invention and at least one pharmaceutically acceptable carrier, diluent, excipient and/or adjuvant, and optionally one or more further pharmaceutically active compounds.

[0243] By means of non-limiting examples, such a formulation may be in a form suitable for oral administration, for parenteral administration (such as by intravenous, intramuscular, intradermic or subcutaneous injection or intravenous infusion), for intralesional administration, for submucosal administration, for intra-articular administration, for intra-cavitary administration, for topical administration (including ocular), for artery embolization, for administration by inhalation, by a skin patch, by an implant, by a suppository, etc. Such suitable administration forms—which may be solid, semi-solid or liquid, depending on the manner of administration—as well as methods and carriers, diluents and excipients for use in the preparation thereof, will be clear to the skilled person; reference is made to the latest edition of Remington's Pharmaceutical Sciences. Some preferred, but non-limiting examples of such preparations include tablets, pills, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols, ointments, creams, lotions, soft and hard gelatin capsules, suppositories, drops, sterile injectable solutions and sterile packaged powders (which are usually reconstituted prior to use) for administration as a bolus and/or for continuous administration, which may be formulated with carriers, excipients, and diluents that are suitable for such formulations, such as salts (especially NaCl), glucose, lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, polyethylene glycol, cellulose, (sterile) water, methylcellulose, methyl- and propylhydroxybenzoates, talc, magnesium stearate, edible oils, vegetable oils and mineral oils or suitable mixtures thereof. The formulations can optionally contain other substances that are commonly used in pharmaceutical formulations, such as buffers, antioxidants, lubricating agents, wetting agents, emulsifying and suspending agents, dispersing agents, desintegrants, bulking agents, fillers, preserving agents, sweetening agents, flavoring agents, flow regulators, release agents, etc. The compositions may also be formulated so as to provide rapid, sustained or delayed release of the active compound(s) contained therein.

[0244] The pharmaceutical preparations of the invention are preferably in a unit dosage form, and may be suitably packaged, for example in a box, blister, vial, bottle, sachet, ampoule or in any other suitable single-dose or multi-dose holder or container (which may be properly labeled); optionally with one or more leaflets containing product information and/or instructions for use.

[0245] Use of the Ligand

[0246] According to an embodiment, the ligand of the invention is used for the synthesis of a chelate according to the present invention.

[0247] According to an embodiment, the ligand of the invention may be used as chelating agent to form chelates which may be used as imaging agents or medicaments in nuclear medicine.

[0248] According to an embodiment, the ligand of the invention may be used as scavenging agent.

[0249] According to an embodiment, the ligand of the invention is used for depollution of liquid medium by trapping of metallic cations.

[0250] According to a specific embodiment, the ligand of the invention may be used in epuration of effluents contaminated with metals. Especially, the ligand of the invention may be used to trap lead or radioactive elements. In a preferred embodiment, the ligand of the invention is used for ultrapurification of liquids. In the present invention, “ultrapurification” refers to the purification of a contaminated solution to a level of contaminant which is much less than 1 ppm (part per million), and generally in the range of ppb (part per billion), ppt (part per trillion), or lower i.e. an ultrapure solution.

[0251] According to another embodiment, the ligand of the invention may be used in cation detection, preferably in detection of traces of metallic cations.

Examples

[0252] The present invention will be better understood with reference to the following examples. These examples are intended to representative of specific embodiments of the invention, and are not intended as limiting the scope of the invention.

[0253] I. Materials and Methods

[0254] All commercial reagents were used as received from the suppliers unless otherwise indicated. The solvents were freshly distilled prior to use and according to literature procedures. Cyclen glyoxal (perhydro-2a,4a,6a,8a-tetraazacyclopenta[f,g] acenaphthylene) was synthesized according to Le Baccon et al., New. J. Chem., 2001, 25, 1168-1174. 6-(Chloromethyl)pyridine-2-carboxylic acid methyl ester was synthesized according to Mato-Iglesias et al., Inorg. Chem. 2008, 47, 7840-7851. Spectral data were in accordance with the literature.

[0255] Analytical .sup.1H and .sup.13C NMR spectra were recorded on a Bruker AMX3-300 MHz spectrometer operating at 300.17 and 75.48 MHz, for .sup.1H and .sup.13C, respectively. All experiments were performed at 25° C. The signals are indicated as follows: chemical shift (ppm), multiplicity (s, singlet; br. s, broad singlet; d, doublet; t, triplet; m, multiplet; quint, quintuplet), coupling constants J in hertz (Hz). The abbreviations Am, Ar, Cq, Pic and Ph stand for aminal, aromatic, quaternary carbon, picolinate and phenyl, respectively.

[0256] Chromatographic methods: All reactions were monitored by thin-layer chromatography, which was performed on aluminium sheets covered with silica gel 60 F254. Visualization was realized by UV-light irradiation (254 and 365 nm) and/or developed with Dragendorf stain.

[0257] High resolution mass spectra (HRMS-ESI) were performed in positive Electrospray Ionisation (ESI+) mode by the Mass Spectrometry service of the ICOA (Institut de Chimie Organique et Analytique), Orleans, France.

[0258] II. Synthesis of the Ligands

[0259] II.1. Synthesis of Ligand I-1

##STR00038## ##STR00039##

Intermediate 2

[0260] A solution of glyoxal cyclen 1 (5.14 g, 26.46 mmol) and 4-nitrobenzyl bromide (5.71 g, 26.46 mmol) in 30 mL of dry THF was stirred for 2 days at room temperature. The resulting white precipitate was collected by filtration and washed with dry THF to give a white solid (10.38 g, 96% yield).

[0261] .sup.1H NMR (D.sub.2O, 300 MHz): 8.34 (d, 2H, .sup.3J=8.7), 7.85 (d, 2H, .sup.3J=8.1), 5.06 (d, 1H, CH.sub.2Ph, .sup.2J=13.2), 4.85 (d, 1H, CH.sub.2Ph, .sup.2J=13.3), 4.26 (t, 1H, .sup.3J=9.2), 4.12 (s, 1H, H.sub.am), 3.82 (s, 1H, H.sub.am), 3.72-3.52 (m, 4H), 3.33-3.17 (m, 5H), 2.97-2.77 (m, 4H), 2.57 (br. s, 2H). .sup.13C NMR (D.sub.2O, 300 MHz): 148.8 (CqNO.sub.2), 133.6 (CqCH.sub.2), 133.5 (2×CH.sub.ar), 124.2 (2×CH.sub.ar), 83.0 (CH.sub.am), 71.4 (CH.sub.am), 61.4 (CH.sub.2Ph), 59.9, 57.0, 51.1, 48.0, 47.9, 47.3, 43.5. HRMS (ESI): calculated for C.sub.17H.sub.25N.sub.5O.sub.2.sup.+ [M]: 330.192451; found: 330.192631.

Intermediate 3

[0262] Methyl iodide (1.05 mL, 16.93 mmol) was added to a stirred suspension of 2 (6.04 g, 14.72 mmol) in 25 mL of dry acetonitrile at room temperature. After stirring for 12 hours, the suspension was filtered and the filter cake washed with dry THF and then dried to obtain the desired compound pure (yellow solid, 7.73 g, 95%).

[0263] .sup.1H NMR (D.sub.2O, 300 MHz): 8.35 (d, 2H, 2×H.sub.ar, .sup.3J=7.5), 7.93 (d, 2H, 2×H.sub.ar, .sup.3J=7.7), 5.20 (d, 1H, CH.sub.2Ph, .sup.3J=13.2), 4.99 ((d, 1H, CH.sub.2Ph, .sup.3J=13.4), 4.86 (d, 2H, 2×H.sub.am), 4.40 (t, 1H, .sup.3J=8.8), 4.14-3.92 (m, 4H), 3.76-3.46 (m, 8H), 3.53 (s, 3H, CH.sub.3) 3.33-3.24 (m, 3H). .sup.13C NMR (D.sub.2O, 300 MHz): 151.9 (CqNO.sub.2), 136.7 (2×CH.sub.ar), 136.0 (CqCH.sub.2), 127.5 (2×CH.sub.ar), 80.9 (CH.sub.am), 80.8 (CH.sub.am), 67.7 (CH.sub.2Ph), 64.1, 62.4, 61.9, 57.8, 49.5, 49.2, 49.0, 45.8, 45.4. HRMS (ESI): calculated for C.sub.18H.sub.29N.sub.5O.sub.2 [M+2H].sup.2+: 172.607689; found: 172.608108.

Intermediate 4

[0264] To a suspension of 3 (7.73 g, 14.02 mmol) in 40 mL of EtOH, was added 1,2-ethylenediamine (2.81 mL, 42.07 mmol). The reaction mixture was stirred for 6 hours at room temperature. The solvent was then removed under reduced pressure and the resulting residue was partitioned between CH.sub.2Cl.sub.2 and water. The aqueous layer was further extracted twice with CH.sub.2Cl.sub.2, and the organic layers were combined and dried over anhydrous MgSO.sub.4. The solvent was removed under vacuum to yield intermediate 4 as a pale yellow solid (5.07 g, 100%). No purification step was required.

[0265] .sup.1H NMR (CDCl.sub.3, 300 MHz): 8.07 (d, 2H, 2×H.sub.ar, .sup.3J=8.7), 7.46 (d, 2H, 2×H.sub.ar, .sup.3J=8.7), 5.93 (s, 2H, 2NH), 3.77 (s, 2H, CH.sub.2Ph), 2.73-2.64 (m, 12H), 2.57-2.53 (m, 4H), 2.41 (s, 3H, CH.sub.3). .sup.13C NMR (CDCl.sub.3, 300 MHz): 147.1 (Cq), 146.9 (Cq), 129.4 (2×CH.sub.ar), 123.4 (2×CH.sub.ar), 60.2 (CH.sub.2Ph), 53.5, 52.2, 46.2, 44.2 (CH.sub.3). HRMS (ESI): calculated for C.sub.16H.sub.28N.sub.5O.sub.2 [M+H].sup.+: 322.223752; found: 322.224088.

Intermediate 5

[0266] To a mixture of 4 (0.50 g, 1.55 mmol) and methyl 6-(chloromethyl)-2-pyridinecarboxylate (0.58 g, 3.11 mmol) in dry acetonitrile (10 mL) was added K.sub.2CO.sub.3 (0.90 g, 6.53 mmol). The solution was stirred under nitrogen at room temperature for 7 days. The undissolved materials were removed by filtration. Upon the evaporation of the solvent in vacuo, the crude product was dissolved in CH.sub.2Cl.sub.2 and the white precipitate was filtered off. The solvent was removed by evaporation under vacuum to afford the crude product (yellow oil, 0.74 g, 88%), which was taken onto the next step without further purification.

[0267] .sup.13C NMR (CDCl.sub.3, 300 MHz): 165.1 (2×CO), 159.0 (Cq.sub.ar), 147.1 (Cq.sub.ar), 146.3 (Cq.sub.ar), 140.6 (CH.sub.ar), 138.5 (CH.sub.ar), 131.6 (CH.sub.ar), 127.5 (CH.sub.ar), 123.6 (CH.sub.ar), 122.9 (CH.sub.ar), 59.3 (2×CH.sub.2Pic), 55.6 (CH.sub.2PhNO.sub.2), 53.4, 52.8 (2×CO.sub.2CH.sub.3), 50.5 (br), 44.0 (CH.sub.3). HRMS (ESI): calculated for C.sub.32H.sub.42N.sub.7O.sub.6 [M+H].sup.+: 620.319109; found: 620.318734.

Intermediate 6

[0268] Tin(II) chloride (3.22 g, 14.28 mmol) was added to a stirred solution of the crude intermediate 5 (1.77 g, 2.86 mmol) in concentrated hydrochloric acid (40 mL) and MeOH (4 mL). The resulting solution was stirred at room temperature for 2 hours and the completion of reaction was monitored by mass spectrometry. The reaction mixture was neutralized at 0° C. by cautiously adding saturated aqueous Na.sub.2CO.sub.3. The product was then extracted with CH.sub.2Cl.sub.2 (3×50 mL). The organic layer was dried over anhydrous MgSO.sub.4, filtered and evaporated under reduced pressure. The crude was purified by flash chromatography on alumina (CH.sub.2Cl.sub.2.fwdarw.CH.sub.2Cl.sub.2:MeOH 98:2) to give 6 as yellow oil (1.15 g, 68% yield).

[0269] .sup.1H NMR (CDCl.sub.3, 300 MHz): 7.88-7.79 (m, 4H, 4×H.sub.ar), 7.55 (d, 2H, 2×H.sub.ar, .sup.3J=7.5), 6.52 (d, 2H, 2×H.sub.ar, .sup.3J=8.4), 6.40 (d, 2H, 2×H.sub.ar, .sup.3J=8.1), 3.84 (s, 2H), 3.73 (s, 2H), 3.35 (s, 6H, 2×CO.sub.2CH.sub.3), 3.15 (s, 2H, CH.sub.2PhNH.sub.2), 3.01-2.09 (m, 16H), 1.73 (s, 3H, CH.sub.3). .sup.13C NMR (CDCl.sub.3, 300 MHz): 164.6 (CO), 158.5 (Cq), 145.9 (Cq), 145.6 (Cq), 138.1 (CH.sub.ar), 130.9 (CH.sub.ar), 127.0 (CH.sub.ar), 123.1 (CH.sub.ar), 121.5 (Cq), 113.9 (CH.sub.ar), 58.7, 55.4, 52.1 (2×CO.sub.2CH.sub.3), 43.6 (CH.sub.3). HRMS (ESI): calculated for C.sub.32H.sub.44N.sub.7O.sub.4 [M+H].sup.+: 590.344929; found: 590.344450.

[0270] Ligand I-1

[0271] Intermediate 6 (1.30 g, 2.20 mmol) was dissolved in 5 mL of hydrochloric acid (6 M), and refluxed 48 h to hydrolyse the methyl esters. The reaction mixture was allowed to cool down to ambient temperature, and after being diluted by water (5 mL) and CH.sub.2Cl.sub.2 (5 mL), thiophosgene (2.53 mL, 33.06 mmol) was added carefully. The mixture was kept under vigorous stirring for 12 hours, followed by decanting of the organic phase with a pipet to remove excess thiophosgene. The aqueous layer was washed with CH.sub.2Cl.sub.2 (3×5 mL) by vigorous biphasic stirring. The aqueous layer was loaded directly onto a RP-HPLC. The solvent system used had the following time profile with a run time of 25 min: 0-3 min: 90% solvent A (0.1% HCOOH in water) and 10% of solvent B (0.1% HCOOH in MeCN); 3-25 min: 90% of solvent A declining to 45% with the solvent B rising to 55% over the same time interval. Flow rate: 5 mL/min. Detector: 254 nm. Retention time: 15 min. Removal of solvents by lyophilization gave the desired ligand as pale yellow powder.

[0272] .sup.13C NMR (D.sub.2O, 300 MHz): 168.3 (CO), 158.3 (Cq.sub.ar—CH.sub.2), 148.0 (Cq.sub.ar—CO.sub.2H), 144.2 (CH.sub.ar), 139.3 (CN), 135.8, 135.1, 131.9, 131.2, 129.7, 128.6, 128.3, 128.2, 126.6, 65.1 (2×CH.sub.2Pic), 64.7 (CH.sub.2PhNCS), 58.9, 57.8, 55.7, 52.7, 50.4, 46.3 (CH.sub.3). HRMS (ESI): calculated for C.sub.31H.sub.38N.sub.7O.sub.4S [M+H].sup.+: 604.270050; found: 604.269622.

[0273] II.2. Synthesis of Ligand I-2

##STR00040## ##STR00041##

Intermediate 7

[0274] A solution of glyoxal cyclen 1 (1.00 g, 5.15 mmol) and tert-butyl n-(3-iodopropyl)carbamate (1.64 g, 5.66 mmol) in 10 mL of dry THF was stirred for 4 days at room temperature. The resulting precipitate was recovered by filtration and washed with small portions of dry THF, to give a white solid (2.25 g, 91% yield).

[0275] .sup.1H NMR (D.sub.2O, 300 MHz): 3.95-3.41 (m, 10H), 3.27-3.17 (m, 6H), 2.95-2.76 (m, 4H), 2.58-2.47 (m, 2H), 2.14-2.00 (m, 2H, CH.sub.2CHNHBoc), 1.44 (s, 9H, C(CH.sub.3).sub.3). .sup.13C NMR (D.sub.2O, 300 MHz): 161.3 (CO), 87.1 (CH.sub.am), 84.6 (C(CH.sub.3).sub.3), 75.0 (CH.sub.am), 65.3, 60.3, 9.3 (CH.sub.2NHCO), 54.5, 51.7, 51.4, 51.0, 50.9, 46.9, 40.3 (CH.sub.2NHCO), 31.1, 26.7 (CH.sub.2CH.sub.2NHCO). HRMS (ESI): calculated for C.sub.18H.sub.34N.sub.5O.sub.2 [M+H].sup.+: 352.270702; found: 352.270888.

Intermediate 8

[0276] A solution of 7 (0.62 g, 1.29 mmol) and iodomethane (0.085 mL, 1.36 mmol) in 10 mL of dry acetonitrile was allowed to stir overnight at room temperature. The resulting precipitate was recovered by filtration and washed with small portions of dry acetonitrile, to yield a white powder (0.74 g, 92%).

[0277] .sup.1H NMR (D.sub.2O, 300 MHz): 4.57 (s, 2H, 2×H.sub.am), 4.10-3.39 (overlapping m, 14H), 3.43 (s, 3H, CH.sub.3), 3.27-3.10 (m, 6H), 2.22-1.99 (m, 2H, CH.sub.2CHNHBoc), 1.46 (s, 9H, C(CH.sub.3).sub.3). .sup.13C NMR (D.sub.2O, 300 MHz): 161.0 (CO), 84.2 (C(CH.sub.3).sub.3), 81.4 (CH.sub.am), 80.9 (CH.sub.am), 67.8, 64.5, 62.0, 58.6, 57.9, 49.6 (CH.sub.3), 49.2, 45.8, 45.4, 39.8 (CH.sub.2NH), 30.7 (C(CH.sub.3).sub.3), 26.2 (CH.sub.2CH.sub.2NHCO). HRMS (ESI): calculated for C.sub.19H.sub.37N.sub.5O.sub.2.sup.2+ [M+2H].sup.2+: 183.646814; found: 183.647112.

Intermediate 9

[0278] To a suspension of 8 (0.58 g, 0.93 mmol) in 15 ml of CH.sub.2Cl.sub.2, was slowly added 1,2-ethanediamine (0.25 mL, 3.73 mmol) at room temperature. The reaction mixture was stirred overnight. 1,4,5,8-Tetrazaperhydronaphthalene (tetrazadecaline) appeared as a white precipitate which was removed by filtration. The filtrate was concentrated in vacuo to provide the pure compound without recourse to chromatography (white solid, 0.32 g, quantitative yield).

[0279] .sup.1H NMR (CDCl.sub.3, 300 MHz): 3.12 (m, 4H, CH.sub.2NH & 2H), 2.65-2.43 (m, 18H, 9CH.sub.2), 2.28 (s, 3H, CH.sub.3), 1.64 (quint, 2H, CH.sub.2CH.sub.2NHBoc), 1.40 (s, 9H, C(CH.sub.3).sub.3). .sup.13C NMR (CDCl.sub.3, 300 MHz): 156.3 (CO), 78.7 (Cq), 54.0, 53.3 (CH.sub.2(CH.sub.2).sub.2NHBoc), 51.4, 46.3, 46.1, 44.0, 44.0 (CH.sub.3), 37.5 (CH.sub.2NHBoc), 28.2 (C(CH.sub.3).sub.3), 27.8 (CH.sub.2CH.sub.2NHBoc). HRMS (ESI): calculated for C.sub.17H.sub.38N.sub.5O.sub.2 [M+H].sup.+: 344.302002; found: 344.302210.

Intermediate 10

[0280] To a solution of 9 (0.45 g, 1.31 mmol) and methyl 6-(chloromethyl)-2-pyridinecarboxylate (0.51 g, 2.76 mmol) in dry acetonitrile was added K.sub.2CO.sub.3 (0.91 g, 6.58 mmol). The solution was stirred at room temperature for 5 days. The undissolved materials were filtered off and the solvent was removed under reduced pressure. The residue was taken up with CH.sub.2Cl.sub.2. After removal of the white precipitate by filtration, the product was simply obtained by evaporating the solvent (yellow oil, 0.74 g, 88%). The crude product was used for next step without further purification.

[0281] .sup.1H NMR (CDCl.sub.3, 300 MHz): 8.04-7.91 (m, 4H, 4H.sub.ar), 7.58 (d, 1H, H.sub.ar, .sup.3J=7.5), 3.87 (s, 6H, 2×CO.sub.2CH.sub.3), 3.13-2.16 (m, 20H), 1.98 (s, 3H, CH.sub.3), 1.55-1.47 (m, 2H, CH.sub.2CH.sub.2NHBoc), 1.34 (s, 9H, C(CH.sub.3).sub.3). .sup.13C NMR (CDCl.sub.3, 300 MHz): 164.9 (CO), 158.9, 146.6, 138.2, 127.4, 123.5, 78.2 (C(CH.sub.3).sub.3), 60.6 (2×CH.sub.2Pic), 52.4 (2CO.sub.2CH.sub.3), 50.6, 50.4, 42.8 (NCH.sub.3), 38.3 (CH.sub.2NH.sub.2), 27.2 (C(CH.sub.3).sub.3), 23.5 (CH.sub.2CH.sub.2NHBoc). HRMS (ESI): calculated for C.sub.33H.sub.52N.sub.7O.sub.6 [M+H].sup.+: 642.397359; found: 642.396805.

Intermediate 11

[0282] To a solution of 10 (0.23 g, 0.36 mmol) in CH.sub.2Cl.sub.2 (5 mL) at room temperature was added dropwise aqueous phosphoric acid (85 wt %) (0.15 mL, 2.15 mmol). The mixture was vigorously stirred for 3 h, and TLC showed reaction completion. The supernatant was removed and the solid gum was washed with CH.sub.2Cl.sub.2 (2×5 mL). 5 mL of MeOH was then added to dissolve the solid residue and an aqueous 6N NaOH solution was added slowly to adjust the pH to 7. The precipitated salt was removed and the mixture was concentrated in vacuo. The resulting residue was taken up with CH.sub.2Cl.sub.2 and the white precipitate removed by filtration. The filtrate was dried under reduced pressure to give the desired product as a light yellow foam (0.64 g, 91%). No purification was deemed necessary.

[0283] .sup.1H NMR (CDCl.sub.3, 300 MHz): 8.05 (d, 2H, 2×H.sub.ar, .sup.3J=6.0 Hz), 7.94 (t, 2H, 2×H.sub.ar, .sup.3J=6.0), 7.61 (d, 2H, 2×H.sub.ar, .sup.3J=5.7), 4.13 (m, 2H, CH.sub.2Pic), 3.98 (s, 6H, 2×CO.sub.2CH.sub.3), 3.66 (m, 2H, CH.sub.2Pic), 2.91-2.40 (overlapping m, 20H), 2.02-1.88 (m, 2H, CH.sub.2CH.sub.2NH.sub.2), 1.67 (s, 3H, CH.sub.3). .sup.13C NMR (CDCl.sub.3, 300 MHz): 164.7 (CO), 158.0 (C.sub.ar), 146.7, 138.0 (C.sub.ar), 127.7 (C.sub.ar), 123.6 (C.sub.ar), 61.7 (2×CH.sub.2Pic), 53.5, 52.4 (2×CO.sub.2CH.sub.3), 51.7, 51.5, 50.5, 50.3, 42.1 (CH.sub.3), 39.4 (CH.sub.2NH.sub.2), 23.6 (CH.sub.2CH.sub.2NH.sub.2). HRMS (ESI): calculated for C.sub.28H.sub.44N.sub.7O.sub.4 [M+H].sup.+: 542.344929; found: 542.344902.

[0284] Ligand I-2.

[0285] Intermediate 11 was dissolved in hydrochloric acid (6 M), and refluxed 48 h to hydrolyze the methyl esters. The reaction mixture was allowed to cool down to ambient temperature. Removal of solvents under reduce pressure gave the desired ligand under its chlorhydrate form.

[0286] II.3. Conjugation of I-1 to Di-HSGL Affording 1-3

[0287] Ligand I-1 (Me-do2pa-PhNCS) was coupled to di-HSGL hapten, a synthetic peptide of low molecular weight, leading to ligand I-3, as presented in the scheme below:

##STR00042##

3.5 equivalents of (Me-do2pa-PhNCS).sup.2− and 1 equivalent of di-HSGL were dissolved in a 0.2 M MES buffer solution (2-morpholino ethanesulfonic acid monohydrate) at pH 6.1. The solution was incubated at room temperature and stirred for 16 h.

[0288] The coupling was analyzed by HPLC, with a C.sub.18 symmetry shield column (isocratic mode 15% ACN/85% TFA (0.5% in water), UV detection λ=254 nm, 1 mL/min). A main peak at rt=14.5 min that corresponds to the coupling product between (Me-do2pa-PhNCS).sup.2− and di-HSFL was detected.

[0289] The coupling product between (Me-do2pa-PhNCS).sup.2− and di-HSGL (i.e. ligand I-3) was purified by HPLC, in the same conditions as described above.

[0290] III. Synthesis of the Chelates

[0291] Formation of the Lead Chelates.

[0292] The ligand is dissolved in H.sub.2O and PbCl.sub.2 (1 eq) is added to the solution followed if necessary by a few drops of concentrated HCl until complete dissolution of PbCl.sub.2. The solution is stirred at room temperature for 30 min. The pH is raised to pH=7 by addition of KOH. The resulting mixture is stirred for a few hours under reflux. The reaction mixture is cooled down to room temperature and MeOH is added. The precipitate is filtered off. The filtrate is evaporated and washed several times with MeOH to yield to lead chelate.

[0293] Formation of the Bismuth Chelates.

[0294] The ligand is dissolved in 3 mL of H.sub.2O and concentrated HCl in order to have a starting pH around 1. Then, 1 eq. of Bi(NO.sub.3).sub.3.5H.sub.2O is added. The mixture is stirred at room temperature for 30 min. The pH is raised to pH=4 by addition of KOH. A white precipitate appears. The resulting mixture is stirred for a few hours under reflux. The reaction mixture is cooled down to room temperature and MeOH is added. The precipitate is filtered off. The filtrate is evaporated and washed several times with MeOH to give the bismuth chelate.

[0295] Radiolabeling of di-HSGL ligand I-3 with 213Bi.

[0296] Bismuth-213 was eluted in 600 μL of water. A solution comprising 5 μL of ligand I-3 (0.195 nmol), 60 μL tris pH7 (2M), 20 μL NaOH (2.5 M) and 500 μL of elute (440 μCi) was prepared and incubated 15 min at 92° C. Radiolabeling was analyzed by ITLC-SG, with a 0.1M citrate buffer pH5. The radiolabeling yield was of 50% leading to a specific activity of 42 MBq/nmol.

[0297] The resulting .sup.213Bi radiolabeled chelate was purified on Sep-Pak Oasis (Waters).