SILICON-CONTAINING LIGAND COMPOUNDS

Abstract

Provided is a ligand compound, comprising (a) a targeting group, (b) one or more chelating groups, optionally containing a chelated radioactive or non-radioactive cation, and (c) a group carrying an Si—OH functional moiety. The ligand compound is suitable for use in therapeutic or diagnostic methods, especially in radiotherapy or radiodiagnosis of cancer, such as prostate cancer.

Claims

1. A ligand compound, comprising: (a) a targeting group, (b) one or more chelating groups, optionally containing a chelated radioactive or non-radioactive cation, and (c) a group carrying an Si—OH functional moiety.

2. The ligand compound in accordance with claim 1, wherein the group carrying an Si—OH functional moiety is a group of formula (S-1) ##STR00048## wherein R.sup.1S and R.sup.2S are independently a linear or branched C3 to C10 alkyl group; R.sup.3S is a C1 to C20 hydrocarbon group which comprises one or more aromatic and/or aliphatic units, and which optionally comprises up to 3 heteroatoms independently selected from O, N and S; and wherein the group carrying an Si—OH functional moiety of formula (S-1) is attached to the remainder of the compound via the bond marked by the dashed line.

3. The ligand compound in accordance with claim 1 or 2, wherein the targeting group is a PSMA binding group of formula (P-1) or a pharmaceutically acceptable salt thereof ##STR00049## wherein: R.sup.1P is CH.sub.2, NH or O; R.sup.3P is CH.sub.2, NH or O; R.sup.2P is C or P(OH); R.sup.4P is selected from a group —(CH.sub.2).sub.m—, wherein m is an integer of 2 to 6, and a group *—(CH.sub.2).sub.p—NH—C(O)—, wherein p is an integer of 1 to 5, and the bond marked with * faces upwards from R.sup.4P in formula (P-1); R.sup.5P is selected from a group —(CH.sub.2).sub.n—, wherein n is an integer of 1 to 6, and a group *—(CH.sub.2).sub.q—NH—C(O)—, wherein q is an integer of 1 to 5, and the bond marked with * faces upwards from R.sup.5P in formula (P-1); and wherein the PSMA binding group is attached to the remainder of the compound via the bond marked by the dashed line.

4. The ligand compound in accordance with any of claims 1 to 3, wherein the chelating group comprises at least one of (i) a macrocyclic ring structure with 8 to 20 ring atoms of which 2 or more, preferably 3 or more, are selected from oxygen atoms and nitrogen atoms; and (ii) an acyclic, open chain chelating structure with 8 to 20 main chain atoms of which 2 or more, preferably 3 or more are heteroatoms selected from oxygen atoms and nitrogen atoms.

5. The ligand compound in accordance with any of claims 1 to 4, wherein the targeting group is a PSMA binding group of formula (P-2) or a pharmaceutically acceptable salt thereof ##STR00050## wherein: m is an integer of 2 to 6, preferably 2 to 4, more preferably 2; n is an integer of 1 to 6, preferably 2 to 4, more preferably 2 or 4; R.sup.1P is CH.sub.2, NH or O, preferably NH; R.sup.3P is CH.sub.2, NH or O, preferably NH; R.sup.2P is C or P(OH), preferably C; wherein the PSMA binding group is attached to the remainder of the compound via the bond marked by the dashed line; wherein the chelating group is selected from a group of the formula (CH-1) or (CH-2), or a pharmaceutically acceptable salt thereof ##STR00051## which chelating group is attached by the bond marked by the dashed line to the remainder of the compound via an ester bond or an amide bond, preferably via an amide bond, and wherein the chelating group optionally contains a chelated radioactive or non-radioactive cation, and wherein the group carrying an Si—OH functional moiety is a group of formula (S-2) or (S-3) ##STR00052## wherein t-Bu indicates a tert-butyl group, and wherein the group carrying an Si—OH functional moiety of formula (S-2) and (S-3) is attached to the remainder of the compound via the bond marked by the dashed line.

6. The ligand compound in accordance with claim 1, which is a PSMA ligand compound of formula (I) or a pharmaceutically acceptable salt thereof ##STR00053## wherein, in formula (I), R.sup.1P is CH.sub.2, NH or O; R.sup.3P is CH.sub.2, NH or O; R.sup.2P is C or P(OH); R.sup.4P is selected from a group —(CH.sub.2).sub.m—, wherein m is an integer of 2 to 6, and a group *—(CH.sub.2).sub.p—NH—C(O)—, wherein p is an integer of 1 to 5, and the bond marked with * faces upwards from R.sup.4P in formula (I); R.sup.5P is selected from a group —(CH.sub.2).sub.n—, wherein n is an integer of 1 to 6, and a group *—(CH.sub.2).sub.q—NH—C(O)—, wherein q is an integer of 1 to 5, and the bond marked with * faces upwards from R.sup.5P in formula (I); X.sup.1A is selected from a covalent bond, an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, and an amine bond; L.sup.1 is a divalent linking group; X.sup.1B is selected from a covalent bond, an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, and an amine bond; R.sup.B is a trivalent linking group; X.sup.2A is selected from a covalent bond, an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, and an amine bond; L.sup.2 is a divalent linking group; X.sup.2B is selected from a covalent bond, an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, and an amine bond; or —X.sup.2B-L.sup.2 is absent, such that X.sup.2A is directly linked to R.sup.B R.sup.CH is a chelating group, optionally containing a chelated radioactive or non-radioactive cation; X.sup.3A is selected from a covalent bond, an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, an amine bond, and a group —NR.sub.2.sup.+—, wherein the groups R are each an alkyl group, preferably a methyl group; L.sup.3 is a divalent linking group; X.sup.3B is selected from a covalent bond, an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond and an amine bond; or —X.sup.3B-L.sup.3 is absent, such that X.sup.3A is directly linked to R.sup.B; R.sup.1S and R.sup.2S are independently a linear or branched C3 to C10 alkyl group; and R.sup.3S is a C1 to C20 hydrocarbon group which comprises one or more aromatic and/or aliphatic units, and which optionally comprises up to 3 heteroatoms selected from O, N and S.

7. The ligand compound in accordance with claim 6, wherein X.sup.2A is an ester bond, an amide bond, or a thiourea bond, wherein the chelating group R.sup.CH is a residue of a chelating agent selected from bis(carboxymethyl)-1,4,8,11-tetraazabicyclo[6.6.2]-hexadecane (CBTE2a), cyclohexyl-1,2-diaminetetraacetic acid (CDTA), 4-(1,4,8,11-tetraazacyclotetradec-1-yl)-methylbenzoic acid (CPTA), N′-[5-[acetyl(hydroxy)amino]-pentyl]-N-[5-[[4-[5-aminopentyl-(hydroxy)amino]-4-oxobutanoyl]amino]pentyl]-N-hydroxybutandiamide (DFO), 4,11-bis(carboxymethyl)-1,4,8,11-tetraazabicycle-[6.6.2]hexadecan (DO2A) 1,4,7,10-tetraazacyclododecan-N,N′,N″,N″′-tetraacetic acid (DOTA), 2-[1,4,7,10-tetraazacyclododecane-4,7,10-triacetic acid]-pentanedioic acid (DOTAGA), N,N′-dipyridoxylethylendiamine-N,N′-diacetate-5,5′-bis(phosphat) (DPDP), diethylenetriaminepentaacetic acid (DTPA), ethylenediamine-N,N′-tetraacetic acid (EDTA), ethyleneglykol-O,O-bis(2-aminoethyl)-N,N,N′,N′-tetraacetic acid (EGTA), N,N-bis(hydroxybenzyl)-ethylenediamine-N,N′-diacetic acid (HBED), hydroxyethyldiaminetriacetic acid (HEDTA), 1-(p-nitrobenzyl)-1,4,7,10-tetraazacyclo-decan-4,7,10-triacetate (HP-DOA3), 6-hydrazinyl-N-methylpyridine-3-carboxamide (HYNIC), 1,4,7-triazacyclononan-1-succinic acid-4,7-diacetic acid (NODASA), 1-(1-carboxy-3-carboxypropyl)-4,7-(carbooxy)-1,4,7-triazacyclononane (NODAGA), 1,4,7-triazacyclononanetriacetic acid (NOTA), 4,11-bis(carboxymethyl)-1,4,8,11-tetraaza-bicyclo[6.6.2]hexadecane (TE2A), 1,4,8,11-tetraazacyclododecane-1,4,8,11-tetra-acetic acid (TETA), terpyridine-bis(methyleneamine) tetraacetic acid (TMT), 1,4,7,10-tetraazacyclotridecan-N,N′,N″,N″′-tetraacetic acid (TRITA), and triethylenetetra-aminehexaacetic acid (TTHA), N,N′-bis[(6-carboxy-2-pyridil)methyl]-4,13-diaza-18-crown-6 (H.sub.2macropa), 4-amino-4-{2-[(3-hydroxy-1,6-dimethyl-4-oxo-1,4-dihydro-pyridin-2-ylmethyl)-carbamoyl]-ethyl} heptanedioic acid bis-[(3-hydroxy-1,6-dimethyl-4-oxo-1,4-dihydro-pyridin-2-ylmethyl)-amide] (THP), and 1,4,7-triazacyclononane-1,4,7-tris[methylene(2-carboxyethyl)phosphinic acid (TRAP), 2-(4,7,10-tris(2-amino-2-oxoethyl)-1,4,7,10-tetraazacyclododecan-1-yl)acetic acid (DO3AM), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis[methylene(2-carboxyethylphosphinic acid)](DOTPI), and S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane tetraacetic acid, and the bond X.sup.2A is formed using a functional group contained in the chelating agent, and wherein the chelating group R.sup.CH optionally contains a chelated radioactive or non-radioactive cation.

8. The ligand compound in accordance with claim 6 or 7, wherein the chelating group contains a chelated cation selected from the cations of .sup.43Sc, .sup.44Sc, .sup.47Sc, .sup.51Cr, .sup.52mMn, .sup.58Co, .sup.52Fe, .sup.56Ni, .sup.57Ni, .sup.62Cu, .sup.64Cu, .sup.67Cu, .sup.66Ga, .sup.68Ga, .sup.67Ga, .sup.89Zr, .sup.90Y, .sup.86Y, .sup.94mTc, .sup.99mTc, .sup.97Ru, .sup.105Rh, .sup.109Pd, .sup.111Ag, .sup.110mIn, .sup.111In, .sup.113mIn, .sup.114mIn, .sup.117mSn, .sup.121Sn, .sup.127Te, .sup.142Pr, .sup.143Pr, .sup.147Nd, .sup.149Gd, .sup.149Pm, .sup.151Pm, .sup.149Tb, .sup.152Tb, .sup.155Tb, .sup.153Sm, .sup.156Eu, .sup.157Gd, .sup.161Tb, .sup.164Tb, .sup.161Ho, .sup.166Ho, .sup.157Dy, .sup.165Dy, .sup.166Dy, .sup.160Er, .sup.165Er, .sup.169Er, .sup.171Er, .sup.166Yb, .sup.169Yb, .sup.175Yb, .sup.167Tm, .sup.172Tm, .sup.natLu, .sup.177Lu, .sup.186Re, .sup.188Re, .sup.188W, .sup.191Pt, .sup.195mPt, .sup.194Ir, .sup.197Hg, .sup.198Au, .sup.199Au, .sup.212Pb, .sup.203Pb, .sup.211At, .sup.212Bi, .sup.213Bi, .sup.223Ra, .sup.224Ra, .sup.225Ac, and .sup.227Th, or a cationic molecule comprising .sup.18F, such as .sup.18F-[AIF].sup.2+.

9. The ligand compound in accordance with any of claims 6 to 8, wherein L.sup.1 comprises two or more subunits which form a chain of subunits between X.sup.1A and X.sup.1B, wherein the bond(s) between the subunits in the chain of subunits is (are) independently selected for each occurrence from an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond and an amine bond.

10. The ligand compound in accordance with any of claims 6 to 9, wherein —X.sup.2B-L.sup.2 is absent, or wherein the group L.sup.2 is an alkanediyl group, preferably a linear alkanediyl group, which may be substituted by one or more substituents independently selected from —OH, —OCH.sub.3, —COOH, —COOCH.sub.3, —NH.sub.2, —CONH.sub.2, —NHC(O)NH.sub.2, and —NHC(NH)NH.sub.2; and wherein one or more covalent bonds between carbon atoms in the alkanediyl group may be independently replaced by a bond selected from an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, and an amine bond.

11. The ligand compound in accordance with any of claims 6 to 10, wherein —X.sup.3B-L.sup.3 is absent, or wherein the group L.sup.3 is an alkanediyl group, preferably a linear alkanediyl group, which may be substituted by one or more substituents independently selected from —OH, —OCH.sub.3, —COOH, —COOCH.sub.3, —NH.sub.2, —CONH.sub.2, —NHC(O)NH.sub.2, and —NHC(NH)NH.sub.2; and wherein one or more covalent bonds between carbon atoms in the alkanediyl group may be independently replaced by a bond selected from an amide bond, an ether bond, a thioether bond, an ester bond, a thioester bond, a urea bond, a thiourea bond, and an amine bond.

12. The ligand compound in accordance with any of claims 6 to 11, wherein R.sup.B in formula (I) is a group of the formula (B-1): ##STR00054## wherein A is selected from N, CR.sup.B1 wherein R.sup.B1 is H or C.sub.1-C.sub.6 alkyl, and a 5 to 7 membered carbocyclic or heterocyclic group; the bond marked by the dashed line at (CH.sub.2).sub.a is formed with X.sup.1B, and a is an integer of 0 to 4; the bond marked by the dashed line at (CH.sub.2).sub.b is formed with X.sup.3B, if present, and otherwise with X.sup.3A, and b is an integer of 0 to 4; and the bond marked by the dashed line at (CH.sub.2).sub.c is formed with X.sup.2B, if present, and otherwise with X.sup.2A, and c is an integer of 0 to 4.

13. The ligand compound in accordance with any of claims 6 to 12, which is a compound of formula (II) or a pharmaceutically acceptable salt thereof ##STR00055## wherein, in formula (II), m, n, b, c, X.sup.1A, L.sup.1, X.sup.1B, X.sup.2B, L.sup.2, X.sup.2A, R.sup.CH, X.sup.3A, L.sup.3 and X.sup.3B are defined as in the preceding claims 6 to 12, and d is 0 or 1.

14. The ligand compound in accordance with any of claims 6 to 13, wherein —X.sup.2A—R.sup.CH is a group of the formula (XCH-1) or (XCH-2) ##STR00056## which is attached to the remainder of the compound via the bond marked by the dashed line, and wherein the chelating group R.sup.CH optionally contains a chelated radioactive or non-radioactive cation.

15. The ligand compound in accordance with any of claims 6 to 14, wherein the group —X.sup.1A-L.sup.1-X.sup.1B— in formula (I) is a group of any of the formulae (L-1) to (L-6):
*—NH—C(O)—R.sup.1L—C(O)—NH—R.sup.2L—NH—C(O)—  (L-1)
*—C(O)—NH—R.sup.3L—NH—C(O)—R.sup.4L—C(O)—NH—R.sup.5L—NH—C(O)—  (L-2)
*—C(O)—NH—R.sup.6L—NH—C(O)—R.sup.7L—NH—C(O)—R.sup.8L—NH—C(O)—R.sup.9L—NH—C(O)—  (L-3)
*—C(O)—NH—R.sup.10L—NH—C(O)—R.sup.11L—NH—C(O)—  (L-4)
*—C(O)—NH—R.sup.12L—NH—C(O)—R.sup.13L—C(O)—NH—R.sup.14L—NH—C(O)—R.sup.15L—NH—C(O)—  (L-5)
*—C(O)—NH—R.sup.16L—C(O)—NH—R.sup.17L—C(O)—NH—R.sup.18L—C(O)—NH—  (L-6) wherein each of R.sup.1L to R.sup.18L is independently an alkanediyl group containing 1 to 8 carbon atoms, preferably a linear alkanediyl group containing 1 to 8 carbon atoms, wherein each of R.sup.1L to R.sup.18L may be substituted by one or more substitutents independently selected from —OH, —OCH.sub.3, —COOH, —COOCH.sub.3, —NH.sub.2, —CONH.sub.2, —NHC(O)NH.sub.2, and —NHC(NH)NH.sub.2; and wherein * marks the bond corresponding to the X.sup.1A terminal bond of —X.sup.1A-L.sup.1-X.sup.1B—.

16. The ligand compound in accordance with any of claims 6 to 14, wherein the group —X.sup.1A-L.sup.1-X.sup.1B— in formula (I) is a group of the formula (L-7):
—NH—C(O)—R.sup.19L—NH—C(O)—R.sup.20L—NH—C(O)—R.sup.21L—NH—C(O)—  (L-7) wherein R.sup.19L is an alkanediyl group containing 1 to 8 carbon atoms, preferably a linear alkanediyl group containing 1 to 8 carbon atoms; R.sup.20L is an alkanediyl group containing 1 to 8 carbon atoms, a cycloalkanediyl group containing 3 to 6 carbon atoms or an alkanediyl-cycloalkanediyl group containing 5 to 8 carbon atoms, wherein each of R.sup.19L an R.sup.20L may be substituted by one or more substituents independently selected from —OH, —OCH.sub.3, —COOH, —COOCH.sub.3, —NH.sub.2, —CONH.sub.2, —NHC(O)NH.sub.2, —NHC(NH)NH.sub.2, aryl and aralkyl; R.sup.21L is an alkanediyl group containing 2 to 26 carbon atoms, preferably a linear alkanediyl group containing 2 to 26 carbon atoms, wherein one or more —CH.sub.2— groups may be replaced by —O—; and wherein * marks the bond corresponding to the X.sup.1A terminal bond of —X.sup.1A-L.sup.1-X.sup.1B—.

17. A ligand compound in accordance with any one of claims 1 to 16 for use in a therapeutic or diagnostic method.

18. A ligand compound in accordance with any one of claims 1 to 16 for use in a method of treating or diagnosing cancer.

19. A ligand compound for use in accordance with claim 18, wherein the cancer is prostate cancer.

Description

EXAMPLES

1. Materials and Methods

1.1 General Information

[0385] The protected amino acid analogs were purchased from Bachem (Bubendorf, Switzerland), Carbolution Chemicals (St. Ingbert, Germany) or Iris Biotech (Marktredwitz, Germany). The tritylchloride polystyrene (TCP) resin was obtained from Sigma-Aldrich (Steinheim, Germany). Chematech (Dijon, France) delivered the chelators DOTA, DOTA-GA and derivatives thereof. All necessary solvents and other organic reagents were purchased from either, Alfa Aesar (Karlsruhe, Germany), Sigma-Aldrich (Steinheim, Germany), Fluorochem (Hadfield, United Kingdom) or VWR (Darmstadt, Germany). Solid phase synthesis of the peptides was carried out by manual operation using an Intelli-Mixer syringe shaker (Neolab, Heidelberg, Germany). Analytical and preparative reversed-phase high-pressure chromatography (RP-HPLC) was performed using Shimadzu gradient systems (Shimadzu, Neufahrn, Germany), each equipped with a SPD-20A UV/Vis detector (220 nm, 254 nm). A Nucleosil 100 C18 (125×4.6 mm, 5 μm particle size) column (CS Chromatographie Service, Langerwehe, Germany) was used for analytical measurements at a flow rate of 1 mL/min. Both specific gradients and the corresponding retention times t.sub.R are cited in the text. Preparative RP-HPLC purification was done with a Multospher 100 RP 18 (250×10 mm, 5 μm particle size) column (CS Chromatographie Service, Langerwehe, Germany) at a constant flow rate of 5 mL/min. Analytical and preparative radio-RP-HPLC was performed using a Nucleosil 100 C18 (125×4.0 mm, 5 μm particle size) column (CS Chromatographie Service, Langerwehe, Germany). Eluents for all HPLC operations were water (solvent A) and acetonitrile (solvent B), both containing 0.1% trifluoroacetic acid. Electrospray ionization-mass spectra for characterization of the substances were acquired on an expression.sup.L CMS mass spectrometer (Advion, Harlow, United Kingdom). Radioactivity was detected through connection of the outlet of the UV-photometer to a HERM LB 500 NaI detector (Berthold Technologies, Bad Wildbad, Germany). NMR spectra were recorded on Bruker (Billerica, United States) AVHD-300 or AVHD-400 spectrometers at 300 K. pH values were measured with a SevenEasy pH-meter (Mettler Toledo, Gießen, Germany). Activity quantification was performed using a 2480 WIZARD.sup.2 automatic gamma counter (PerkinElmer, Waltham, United States). Radio-thin layer chromatography (TLC) was carried out with a Scan-RAM detector (LabLogic Systems, Sheffield, United Kingdom).

1.2 Solid Phase Peptide Synthesis

1.2.1 TCP-Resin Loading (General Procedure 1 (GP1))

[0386] Loading of the tritylchloride polystyrene (TCP) resin with a Fmoc-protected amino acid (AA) was carried out by stirring a solution of the TCP-resin (1.60 mmol/g) and Fmoc-AA-OH (1.5 eq.) in anhydrous DCM with DIPEA (3.8 eq.) at room temperature (rt) for 2 h. Remaining tritylchloride was capped by the addition of methanol (2 mL/g resin) for 15 min. Subsequently the resin was filtered and washed with DCM (2×5 mL/g resin), DMF (2×5 mL/g resin), methanol (5 mL/g resin) and dried in vacuo. Final loading/of Fmoc-AA-OH was determined by the following equation:

[00001]$\begin{array}{cc}& m_2=\mathrm{mass}\mathrm{of}\mathrm{loaded}\mathrm{resin}\left[g\right]\\ l\left[\frac{m\mathrm{mol}}{g}\right]=\frac{\left(m_2-m_1\right)\times 1000}{\left(M_W-M_{\mathrm{HCl}}\right)m_2}& m_1=\mathrm{mass}\mathrm{of}\mathrm{unloaded}\mathrm{resin}\left[g\right]\\ & M_W=\mathrm{molecular}\mathrm{weight}\mathrm{of}\mathrm{AA}\left[g/\mathrm{mol}\right]\\ & M_{\mathrm{HCl}}=\mathrm{molecular}\mathrm{weight}\mathrm{of}\mathrm{HCl}\left[g/\mathrm{mol}\right]\end{array}$

1.2.2 On-Resin Amide Bond Formation (GP2)

[0387] For conjugation of a building block to the resin-bound peptide, a mixture of TBTU with HOBt or HOAt is used for pre-activation of the carboxylic with DIPEA or 2,4,6-trimethylpyridine as a base in DMF (10 mL/g resin). After 5 min at rt, the solution was added to the swollen resin. The exact stoichiometry and reaction time for each conjugation step is given in the respective synthesis protocols. After reaction, the resin was washed with DMF (6×5 mL/g resin).

1.2.3 On-Resin Fmoc-Deprotection (GP3)

[0388] The resin-bound Fmoc-peptide was treated with 20% piperidine in DMF (v/v, 8 mL/g resin) for 5 min and subsequently for 15 min. Afterwards, the resin was washed thoroughly with DMF (8×5 mL/g resin).

1.2.4 On-Resin Dde-Deprotection (GP4)

[0389] The Dde-protected peptide was dissolved in a solution of 2% hydrazine monohydrate in DMF (v/v, 5 mL/g resin) and shaken for 20 min (GP4a). In the case of present Fmoc-groups, Dde-deprotection was performed by adding a solution of imidazole (0.92 g/g resin), hydroxylamine hydrochloride (1.26 g/g resin) in NMP (5.0 mL/g resin) and DMF (1.0 mL/g resin) for 3 h at room temperature (GP4b). After deprotection the resin was washed with DMF (8×5 mL/g resin).

1.2.5 Peptide Cleavage from the Resin (GP5)

[0390] The fully protected resin-bound peptide was dissolved in a mixture of TFA/TIPS/water (v/v/v; 95/2.5/2.5) and shaken for 30 min. The solution was filtered off and the resin was treated in the same way for another 30 min. Both filtrates were combined, stirred for additional 1-24 h at rt. Product formation was monitored by HPLC. After removing TFA under a stream of nitrogen, the residue was dissolved in a mixture of tert-butanol and water and freeze-dried.

1.3 Synthesis of Functional Building Blocks

[0391] 1.3.1 (S)-5-(tert-Butoxy)-4-(3-((S)-1,5-di-tert-butoxy-1,5-dioxopentan-2-yl)ureido)-5-oxopentanoic acid ((tBuO)EuE(OtBu).sub.2) (prepared according to WO 2019/020831):

##STR00031##

1.3.2 4-(Di-tert-butylfluorosilyl)benzoic acid (SiFA-BA) (prepared according to WO 2019/020831):

##STR00032##

1.3.3 4-(Di-tert-butyl(hydroxy)silyl)benzoic acid (SiOH)

##STR00033##

[0392] Synthesis of 4-(Di-tert-butylfluorosilyl)benzoic acid was performed according to WO 2019/020831. For hydrolysis, 4-(Di-tert-butylfluorosilyl)benzoic acid (50 mg, 0.177 mmol, 1.0 eq.) was dissolved in a 4:1 mixture (v/v) of DMF and water, whereupon KOH (100 mg, 1.79 mmol, 10 eq.) was added. After stirring the reaction mixture for 4 h at 40° C., the reaction mixture was neutralized by the addition of 1 M aq. HCl. and extracted with Et.sub.2O (5×50 mL). The combined organic fractions were dried, filtered and concentrated in vacuo to give the crude product (95%), which was used without further purification. HPLC (50 to 100% B in 15 min): t.sub.R=6.0 min. Calculated monoisotopic mass (C.sub.15H.sub.24O.sub.3Si): 280.2 found: m/z=281.6 [M+H].sup.+.

1.3.4 (4-(Bromomethyl)phenyl)di-tert-butylsilanol

##STR00034##

[0393] Synthesis of (4-(bromomethyl)phenyl)di-tert-butylsilanol was performed in analogy to a previously published procedure (Kostikov A P, Iovkova L, Chin J, et al., J Fluor Chem. 2011; 132:27-34). HPLC (50 to 100% B in 15 min): t.sub.R=11.2 min. Calculated monoisotopic mass (C.sub.15H.sub.24BrFSi): 328.09 found: m/z=not detectable.

1.4 Reference Ligand Compounds

[0394] 1.4.1 rhPSMA-7.3 (Prepared According to WO 2019/020831):

##STR00035##

1.4.2 rhPSMA-10 (Prepared According to WO 2019/020831):

##STR00036##

1.4.3 PSMA I&T (Weineisen et al.; Journal of Nuclear Medicine 55, 1083-1083 (2014):

[0395] ##STR00037##

1.4.4 PSMA-617 (Benešová et al., Journal of Nuclear Medicine 56, 914-920 (2015)):

[0396] ##STR00038##

1.5 Synthesis of PSMA-SiOH Ligand Compounds

1.5.1 PSMA-7.3-SiOH

[0397] ##STR00039##

[0398] The compound was synthesized, applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-D-Orn(Dde)-OH. After cleavage of the Fmoc group with piperidine in DMF (GP3), (tBuO)EuE(OtBu).sub.2 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 4.5 h (GP2). After cleavage of the Dde-group with a mixture hydrazine in DMF (GP4a), a solution of succinic anhydride (7.0 eq.) and DIPEA (7.0 eq.) in DMF was added and left to react for 2.5 h (GP2). Subsequently, the conjugated succinic acid was pre-activated, by adding a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF. After 20 min, Fmoc-D-Lys(OtBu)-HCl (2.0 eq.) dissolved in DMF was added and left to react for 2.5 h (GP2). Subsequent cleavage of the Fmoc-group was performed, by adding a mixture piperidine in DMF (GP3). Fmoc-D-Dap(Dde)-OH (2.0 eq.) was pre-activated in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF and added to the resin-bound peptide for 2.5 h (GP2). Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). SiOH (1.5 eq.) was reacted with the free amine of the side chain with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.), as activation reagents in DMF for 2 h (GP2). After Fmoc-deprotection with piperidine (GP3), (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). Cleavage from the resin with simultaneous deprotection of acid labile protecting groups was performed in TFA for 6 h (GP5). After HPLC-based purification PSMA-7.3-SiOH was obtained as a colorless solid (35%). HPLC (10 to 70% B in 15 min): t.sub.R=8.6 min. Calculated monoisotopic mass (C.sub.63H.sub.100N.sub.12O.sub.26Si): 1468.7; found: m/z=1470.0 [M+H].sup.+, 735.4 [M+2H].sup.2+.

1.5.2 PSMA-10-SiOH

[0399] ##STR00040##

[0400] PSMA-10-SiOH was synthesized in analogy to PSMA-7.3-SiOH, by using DOTA instead of DOTA-GA. The tert-butyl protected chelator, DOTA(tBu).sub.3 was conjugated to the free N-terminus with a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) for 2 h in DMF (GP2). Cleavage from the resin and deprotection of acid labile protecting groups was performed in TFA for 6 h (GP5). After HPLC-based purification, PSMA-10-SiOH (34%) was obtained as a colorless solid. HPLC (10 to 70% B in 15 min): t.sub.R=8.1 min. Calculated monoisotopic mass (C.sub.60H.sub.96FN.sub.12O.sub.24Si): 1396.4; found: m/z=1398.0 [M+H].sup.+, 699.5 [M+2H].sup.2+.

1.5.3 C007-SiOH

[0401] ##STR00041##

[0402] C007-SiOH was synthesized applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-D-Orn(Dde)-OH. After cleavage of the Fmoc group with piperidine in DMF (GP3), (tBuO)EuE(OtBu).sub.2 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 4.5 h (GP2) and the Dde-group was subsequently cleaved with a mixture of hydrazine in DMF (GP4a). Fmoc-β-Ala-OH (2.0 eq.), Fmoc-β-Ala-OH (2.0 eq.) and Fmoc-D-Ser(tBu)-OH (2.0 eq.) were then conjugated to the resin-bound peptide. Each coupling was performed for 2.0 h (GP2) after pre-activating the respective amino acid with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF, each coupling was followed by Fmoc-removal with piperidine (GP3). Fmoc-D-Dap(Dde)-OH (2.0 eq.) was pre-activated in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF and added to the resin-bound peptide for 2.5 h (GP2). Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). SiOH (1.5 eq.) was reacted with the free amine of the side chain with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.), as activation reagents in DMF for 2 h (GP2). After Fmoc-deprotection with piperidine (GP3), (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). Cleavage from the resin with simultaneous deprotection of acid labile protecting groups was performed in TFA for 23 h (GP5). After HPLC-based purification C007-SiOH was obtained as a colorless solid (7%). HPLC (10 to 70% B in 15 min): t.sub.R=8.6 min. Calculated monoisotopic mass (C.sub.62H.sub.99N.sub.13O.sub.26Si): 1469.7; found: m/z=1470.4 [M+H].sup.+, 736.0 [M+2H].sup.2+.

1.5.4 P105-SiOH

[0403] ##STR00042##

[0404] P105-SiOH was synthesized, applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-D-Orn(Dde)-OH. After cleavage of the Fmoc group with piperidine in DMF (GP3), (tBuO)EuE(OtBu).sub.2 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 4.5 h (GP2). After cleavage of the Dde-group with a mixture of hydrazine in DMF (GP4a), a solution of Fmoc-6-Ahx-OH (2.0 eq.) in DMF pre-activated for 5 min in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) was added to the resin-bound peptide for 2.5 h. After Fmoc-deprotection with piperidine (GP3), Dde-D-Dap(Fmoc)-OH (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). After subsequent Fmoc-deprotection with piperidine (GP3), Fmoc-D-Glu-OtBu (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) for 2.5 h (GP2). Final Fmoc-deprotection was again done with piperidine (GP3) and SiOH (1.5 eq.) was conjugated with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.) for 2.5 h (GP2). P105-SiOH was cleaved from the resin within 18 h with simultaneous deprotection of acid labile protecting groups using TFA (GP5). After HPLC-based purification P105-SiOH was obtained as a colorless solid (16%). HPLC (10 to 70% B in min): t.sub.R=9.2 min. Calculated monoisotopic mass (C.sub.64H.sub.102N.sub.12O.sub.26Si): 1482.7; found: m/z=1483.4 [M+H].sup.+, 742.3 [M+2H].sup.2+.

1.5.5 P110-SiOH

[0405] ##STR00043##

[0406] P110-SiOH was synthesized, applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-D-Orn(Dde)-OH. After cleavage of the Fmoc group with piperidine in DMF (GP3), (tBuO)EuE(OtBu).sub.2 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 4.5 h (GP2). After cleavage of the Dde-group with a mixture of hydrazine in DMF (GP4a), a solution of Fmoc-6-Ahx-OH (2.0 eq.) in DMF pre-activated for 5 min in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) was added to the resin-bound peptide for 2.5 h. After Fmoc-deprotection with piperidine (GP3), Fmoc-D-Orn(Dde)-OH (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 2.0 h (GP2). Following orthogonal Fmoc-deprotection was done using piperidine (GP3) and (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). After subsequent Dde-deprotection using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b), N,N-Dimethylglycine (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) for 2.5 h (GP2). After final coupling of (4-(bromomethyl)phenyl)di-tert-butylsilanol (3.0 eq.) with 2,4,6-trimethylpyridine (6.0 eq.) in DCM for 18.5 h, P110-SiOH was cleaved from the resin within 21.5 h with simultaneous deprotection of acid labile protecting groups using TFA (GP5). After HPLC-based purification P110-SiOH was obtained as a colorless solid (32%). HPLC (10 to 70% B in 15 min): t.sub.R=8.9 min. Calculated monoisotopic mass (C.sub.65H.sub.109N.sub.12O.sub.23Si.sup.+): 1453.8; found: m/z=1455.3 [M+H].sup.+, 728.0 [M+2H].sup.2+.

1.5.6 E102-SiOH

[0407] ##STR00044##

[0408] The compound was synthesized, applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-D-Orn(Dde)-OH. After cleavage of the Fmoc-group with piperidine in DMF (GP3), (tBuO)EuE(OtBu).sub.2 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 4.5 h (GP2). After cleavage of the Dde-group with a mixture hydrazine in DMF (GP4a), a solution of succinic anhydride (7.0 eq.) and DIPEA (7.0 eq.) in DMF was added and left to react for 2.5 h (GP2). Subsequently, the conjugated succinic acid was pre-activated, by adding a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF. After 20 min, Fmoc-L-Lys(OtBu)-HCl (2.0 eq.) dissolved in DMF was added and left to react for 2.5 h (GP2). Subsequent cleavage of the Fmoc-group was performed, by adding a mixture of piperidine in DMF (GP3). Fmoc-Gly-OH (2.0 eq.) was pre-activated in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF and added to the resin-bound peptide for 2 h (GP2). The Fmoc-group was cleaved by adding a mixture of piperidine in DMF (GP3). Fmoc-D-Dap(Dde)-OH (2.0 eq.) was pre-activated in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF and added to the resin-bound peptide for 2.5 h (GP2). Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). SiOH (1.5 eq.) was reacted with the free amine of the side chain with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.), as activation reagents in DMF for 2 h (GP2). After Fmoc-deprotection with piperidine (GP3), (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). Cleavage from the resin with simultaneous deprotection of acid labile protecting groups was performed in TFA for 6 h (GP5). After HPLC-based purification the fluoride-containing ligand E102-SiOH was obtained as a colorless solid (19%). HPLC (10 to 90% B in 15 min): t.sub.R=7.4 min. Calculated monoisotopic mass (C.sub.65H.sub.103N.sub.13O.sub.27Si): 1525.7; found: m/z=1526.9 [M+H].sup.+, 763.9 [M+2H].sup.2+.

1.5.7 E104-SiOH

[0409] ##STR00045##

[0410] The compound was synthesized, applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-L-Lys(Dde)-OH. The Dde-group was cleaved by adding imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h in DMF (GP4b). (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). After removal of the Fmoc-group (GP3), Fmoc-Gly-OH (2.0 eq.) was coupled by pre-activation in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF and addition to the resin-bound peptide for 2 h (GP2). The Fmoc-group was cleaved by adding a mixture of piperidine in DMF (GP3). Fmoc-D-Dap(Dde)-OH (2.0 eq.) was pre-activated in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF and added to the resin-bound peptide for 2.5 h (GP2). Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). SiOH (1.5 eq.) was reacted with the free amine of the side chain with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.), as activation reagents in DMF for 2 h (GP2). After Fmoc-deprotection with piperidine (GP3), Fmoc-Ahx-OH (1.5 eq.) was activated with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.) and added to the resin-bound peptide. After Fmoc-removal (GP3), Fmoc-L-Glu-OtBu (1.5 eq.) was pre-activated with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.) and coupled to the resin-bound peptide. The Fmoc-group was removed with piperidine in DMF (GP3) and (tBuO)EuE(OtBu).sub.2 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 4.5 h (GP2). Cleavage from the resin with simultaneous deprotection of acid labile protecting groups was performed in TFA for 6 h (GP5). After HPLC-based purification the fluoride-containing ligand E104-SiOH was obtained as a colorless solid (16%). HPLC (10 to 90% B in 15 min): t.sub.R=7.6 min. Calculated monoisotopic mass (C.sub.71H.sub.114N.sub.14O.sub.28Si): 1638.8; found: m/z=1639.6 [M+H].sup.+, 820.8 [M+2H].sup.2+.

1.5.8 A204-SiOH

[0411] ##STR00046##

[0412] The compound was synthesized, applying the standard Fmoc-SPPS protocol described above, starting from resin bound Fmoc-L-Lys(Dde)-OH. The Fmoc-group was cleaved by adding piperidine in DMF (GP3). Di-tert-butyl (1H-imidazole-1-carbonyl)-L-glutamate (2.0 eq.) was coupled to the resin-bound amino acid, similar to the synthesis of (tBuO)EuE(OtBu).sub.2, in DCE with TEA (3.0 eq.) at 40° C. for 16 h. Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). Fmoc-L-2-NaI—OH (2.0 eq.) was coupled after pre-activation with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF for 2 h. After the removal of the Fmoc-group (GP3), Fmoc-Txa-OH (2.0 eq.) was pre-activated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF and coupled to the resin-bound peptide for 2 h. The Fmoc-group was removed according to GP3, Fmoc-NH-PEG.sub.8-COOH (2.0 eq.) was pre-activated with HOAt (2.0 eq.), TBTU (2.0 eq.) and DIPEA (6.0 eq.) in DMF and coupled for 3 h. Fmoc-D-Dap(Dde)-OH (2.0 eq.) was pre-activated in a mixture of HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF and added to the resin-bound peptide for 2.5 h (GP2). Following orthogonal Dde-deprotection was done using imidazole and hydroxylamine hydrochloride dissolved in a mixture of NMP and DMF for 3 h (GP4b). SiOH (1.5 eq.) was reacted with the free amine of the side chain with HOAt (1.5 eq.), TBTU (1.5 eq.) and DIPEA (4.5 eq.), as activation reagents in DMF for 2 h (GP2). After Fmoc-deprotection with piperidine (GP3), (S)-DOTA-GA(tBu).sub.4 (2.0 eq.) was conjugated with HOAt (2.0 eq.), TBTU (2.0 eq.) and 2,4,6-trimethylpyridine (6.7 eq.) in DMF for 2.5 h (GP2). Cleavage from the resin with simultaneous deprotection of acid labile protecting groups was performed in TFA for 6 h (GP5). After HPLC-based purification the fluoride-containing ligand A204-SiOH was obtained as a colorless solid (12%). HPLC (10 to 90% B in 15 min): t.sub.R=9.0 min. Calculated monoisotopic mass (C.sub.89H.sub.140N.sub.12O.sub.30Si): 1885.0; found: m/z=1886.6 [M+H].sup.+, 944.5 [M+2H].sup.2+.

1.6 Synthesis of .sup.natLu-PSMA Ligand Compounds

[0413] The corresponding .sup.natLu-complexes were prepared from a 2 mM aqueous solution of the indicated PSMA ligand compound (1.0 eq.) with a 20 mM solution of LuCl.sub.3 (2.5 eq.), heated to 95° C. for 30 min. After cooling, the .sup.natLu-chelate formation was confirmed using HPLC and MS. If required, the complexed compound was purified by RP-HPLC.

[0414] .sup.natLu-PSMA-7.3-SiOH: HPLC (10 to 70% B in 15 min): t.sub.R=8.3 min. Calculated monoisotopic mass (C.sub.63H.sub.97LuN.sub.12O.sub.26Si): 1640.6; found: m/z=821.8 [M+2H].sup.2+.

[0415] .sup.natLu-PSMA-10-SiOH: HPLC (10 to 70% B in 15 min): t.sub.R=8.6 min. Calculated monoisotopic mass (C.sub.60H.sub.93LuN.sub.12O.sub.24Si): 1568.6; found: m/z=786.2 [M+2H].sup.2+.

[0416] .sup.natLu-C007-SiOH: HPLC (10 to 70% B in 15 min): t.sub.R=8.3 min. Calculated monoisotopic mass (C.sub.62H.sub.96LuN.sub.13O.sub.26Si): 1641.6; found: m/z=821.8 [M+2H].sup.2+.

[0417] .sup.natLu-P105-SiOH: HPLC (10 to 70% B in 15 min): t.sub.R=8.9 min. Calculated monoisotopic mass (C.sub.64H.sub.99LuN.sub.12O.sub.26Si): 1654.6; found: m/z=1655.5 [M+H].sup.+, 828.2 [M+2H].sup.2+.

[0418] .sup.natLu-P110-SiOH: HPLC (10 to 70% B in 15 min): t.sub.R=9.1 min. Calculated monoisotopic mass (C.sub.65H.sub.106LuN.sub.12O.sub.23Si.sup.+): 1625.7; found: m/z=1626.2 [M]F, 813.2 [M+H].sup.2+.

[0419] .sup.natLu-E102-SiOH: HPLC (10 to 90% B in 15 min): t.sub.R=7.2 min. Calculated monoisotopic mass (C.sub.65H.sub.100LuN.sub.13O.sub.27Si): 1697.6; found: m/z=850.4 [M+2H].sup.2+.

[0420] .sup.natLu-E104-SiOH: HPLC (10 to 90% B in 15 min): t.sub.R=t.b.d. min. Calculated monoisotopic mass (C.sub.71H.sub.111LuN.sub.14O.sub.28Si): 1810.7; found: m/z=t.b.d. [M+2H].sup.2+.

[0421] .sup.natLu-A204-SiOH: HPLC (10 to 90% B in 15 min): t.sub.R=9.0 min. Calculated monoisotopic mass (C.sub.89H.sub.137LuN.sub.12O.sub.30Si): 2056.9; found: m/z=1030.3 [M+2H].sup.2+.

1.7 .SUP.177.Lu-Labelling of PSMA-SiOH Ligand Compounds

[0422] For .sup.177Lu-labelling a previously published procedure was applied with minor modifications (Sosabowski, J. K.; Mather, S. J., Conjugation of DOTA-like chelating agents to peptides and radiolabeling with trivalent metallic isotopes. Nat Protoc. 2006, 1: 972-976). The labelling precursor (1.0 nmol, 10 μL, 1 mM in DMSO) was added to 10 μL of 1.0 M aq. NH.sub.4OAc buffer (pH 5.9). Subsequently, 20 to 110 MBq .sup.177LuCl.sub.3 (Specific Activity (S.sub.A)>3000 GBq/mg, 740 MBq/mL, 0.04 M HCl, ITG, Garching, Germany) were added and the mixture was filled up to 100 μL with water (Tracepur®, Merck, Darmstadt, Germany). The reaction mixture was heated for 30 min at 95° C. and the radiochemical purity was determined using radio-HPLC and radio-TLC (Silica gel 60 RP-18 F.sub.254s, 3:2 mixture (v/v) of MeCN in H.sub.2O, supplemented with 10% of 2 M NaOAc solution and 1% of TFA).

1.8 .SUP.125.I-Labelling

[0423] ##STR00047##

[0424] [.sup.125I]NaI as a basic solution (74 TBq/mmol, 3.1 GBq/mL, 40 mM NaOH) was purchased from Hartmann Analytic (Braunschweig, Germany). The reference ligand for in vitro studies ([.sup.125I]I-BA)KuE was prepared according to a previously published procedure (Weineisen, M.; Simecek, J.; Schottelius, M.; Schwaiger, M.; Wester, H. J., Synthesis and preclinical evaluation of DOTAGA-conjugated PSMA ligands for functional imaging and endoradiotherapy of prostate cancer. EJNMMI Res 2014, 4; 63; Vaidyanathan, G.; Zalutsky, M. R., Preparation of N-succinimidyl 3-[*I]iodobenzoate: an agent for the indirect radioiodination of proteins. Nat Protoc. 2006, 1, 707-713). Briefly, approximately 0.1 mg of the stannylated precursor SnBu.sub.3-BA-(OtBu)KuE(OtBu).sub.2 (Vaidyanathan, G. et al, loc. cit.) was dissolved in a mixture of 20 μL peracetic acid, 5.0 μL [.sup.125I]NaI (40 mM in NaOH, 15±5 MBq, 74 TBq/mmol), 20 μL MeCN and 10 μL acetic acid. The reaction solution was incubated for 10 min at rt, and loaded on a Sep-Pak C18 Plus Short cartridge (360 mg, 55-105 μm, Waters), which was initially preconditioned with 10 mL MeOH followed by 10 mL water). After purging with 10 mL of water, the cartridge was eluted with 2.0 mL of a 1:1 mixture (v/v) of EtOH and MeCN. The eluate was evaporated to dryness under a gentle nitrogen stream at 70° C. and treated with 500 μL TFA for 30 min. After removing TFA in a stream of nitrogen, the crude product was purified by RP-HPLC, yielding ([.sup.125I]I-BA)KuE (5±2 MBq). HPLC (20% to 40% B in 20 min): t.sub.R=13.0 min.

1.9 In Vitro-Experiments

1.9.1 Cell Culture

[0425] PSMA-positive LNCAP cells (300265; Cell Lines Service, Eppelheim, Germany) were cultivated in Dulbecco modified Eagle medium/Nutrition Mixture F-12 with Glutamax (1:1) (DMEM-F12, Biochrom, Berlin, Germany) supplemented with fetal bovine serum (10%, FBS Zellkultur, Berlin, Germany) and kept at 37° C. in a humidified CO.sub.2 atmosphere (5%). A mixture of trypsin and EDTA (0.05%, 0.02%) in PBS (Biochrom) was used in order to harvest cells. Cells were counted with a Neubauer hemocytometer (Paul Marienfeld, Lauda-Königshofen, Germany).

1.9.2 Affinity Determinations (IC.SUB.50.)

[0426] For PSMA affinity (IC.sub.50) determinations, the respective ligand was diluted (serial dilution 10.sup.−4 to 10.sup.−10) in Hank's balanced salt solution (HBSS, Biochrom). In the case of metal-complexed ligands, the crude reaction mixture was diluted analogously, without further purification. Cells were harvested 24±2 hours prior to the experiment and seeded in 24-well plates (1.5×10.sup.5 cells in 1 mL/well). After removal of the culture medium, the cells were carefully washed with 500 μL of HBSS, supplemented with 1% bovine serum albumin (BSA, Biowest, Nuaillé, France) and left 15 min on ice for equilibration in 200 μL HBSS (1% BSA). Next, 25 μL per well of solutions, containing either HBSS (1% BSA, control) or the respective ligand in increasing concentration (10.sup.−1-10.sup.−4 M in HBSS) were added with subsequent addition of 25 μL of [.sup.125I]-BA-KuE (2.0 nM) in HBSS (1% BSA). After incubation on ice for 60 min, the experiment was terminated by removal of the medium and consecutive rinsing with 200 μL of HBSS (1% BSA). The media of both steps were combined in one fraction and represent the amount of free radioligand. Afterwards, the cells were lysed with 250 μL of 1 M aqueous NaOH for at least 10 min. After a washing step (250 μL of 1 M NaOH), both fractions, representing the amount of bound ligand, were united. Quantification of all collected fractions was accomplished in a γ-counter. PSMA-affinity determinations were carried out at least three times per ligand.

1.9.3 Internalization Studies

[0427] For internalization studies, LNCaP cells were harvested 24±2 hours before the experiment and seeded in poly-L-lysine coated 24-well plates (1.25×105 cells in 1 mL/well, Greiner Bio-One, Kremsmünster, Austria). After removal of the culture medium, the cells were washed once with 500 μL DMEM-F12 (5% BSA) and left to equilibrate for at least 15 min at 37° C. in 200 μL DMEM-F12 (5% BSA). Each well was treated with either 25 μL of either DMEM-F12 (5% BSA, control) or 25 μL of a 100 μM PMPA (2-(Phosphonomethyl)-pentandioic acid, Tocris Bioscience, Bristol, UK) solution in PBS, for blockade. Next, 25 μL of the radioactive-labelled PSMA inhibitor (10.0 nM in PBS) was added and the cells were incubated at 37° C. for 60 min. The experiment was terminated by placing the 24-well plate on ice for 3 min and consecutive removal of the medium. Each well was carefully washed with 250 μL of ice-cold HBSS. Both fractions from the first steps, representing the amount of free radioligand, were combined. Removal of surface bound activity was accomplished by incubation of the cells with 250 μL of ice-cold PMPA (10 μM in PBS) solution for 5 min and rinsed again with another 250 μL of ice-cold PBS. The internalized activity was determined by incubation of the cells in 250 μL 1 M aqueous NaOH for at least 10 min. The obtained fractions were combined with those of the subsequent wash step with 250 μL 1 M aqueous NaOH. Each experiment (control and blockade) was performed in triplicate. Free, surface bound and internalized activity was quantified in a γ-counter. All internalization studies were accompanied by external reference studies, using ([.sup.125I]l-BA)KuE (c=0.2 nM), which were performed analogously. Data were corrected for non-specific binding and normalized to the specific-internalization observed for the radioiodinated reference compound.

1.9.4 Octanol-Water Partition Coefficient

[0428] Approximately 1 MBq of the labelled tracer was dissolved in 1 mL of a 1:1 mixture (v/v) of PBS (pH 7.4) and n-octanol in a reaction vial (1.5 mL). After vigorous mixing of the suspension for 3 minutes at room temperature, the vial was centrifuged at 15000 g for 3 minutes (Biofuge 15, Heraeus Sepatech, Osterode, Germany) and 100 μL aliquots of both layers were measured in a gamma counter. The experiment was repeated at least six times.

1.9.5 Determination of Human Serum Albumin (HSA) Binding by High Performance Affinity Chromatography (HiPAC)

[0429] HSA binding of the PSMA-addressing ligands was determined according to a previously published procedure via HPLC (Valko, K.; Nunhuck, S.; Bevan, C.; Abraham, M. H.; Reynolds, D. P., Fast gradient HPLC method to determine compounds binding to human serum albumin. Relationships with octanol/water and immobilized artificial membrane lipophilicity. J Pharm Sci. 2003, 92, 2236-2248). A Chiralpak HSA column (50×3 mm, 5 μm, H13H-2433, Daicel, Tokyo, Japan) was used at a constant flow rate of 0.5 mL/min at rt. Mobile phase A was a freshly prepared 50 mM aqueous solution of NH.sub.4OAc (pH 6.9) and mobile phase B was isopropanol (HPLC grade, VWR). The applied gradient for all experiments was 100% A (0 to 3 min), followed by 80% A (3 to 40 min). Prior to the experiment, the column was calibrated using nine reference substances with a HSA binding, known from literature, in the range of 13 to 99% (Valko, K.; Nunhuck, S.; Bevan, C.; Abraham, M. H.; Reynolds, D. P., J Pharm Sci. 2003, 92, 2236-2248; Yamazaki, K.; Kanaoka, M., Computational prediction of the plasma protein-binding percent of diverse pharmaceutical compounds. J Pharm Sci. 2004, 93, 1480-1494). All substances, including the examined PSMA ligands, were dissolved in a 1:1 mixture (v/v) of isopropanol and a 50 mM aqueous solution of NH.sub.4OAc (pH 6.9) with a final concentration of 0.5 mg/mL. Non-linear regression was established with the OriginPro 2016G software (Northampton, United States).

[0430] FIG. 1 is an exemplary sigmoidal plot, showing the correlation between human serum albumin (HSA) binding of selected reference substances and retention time (t.sub.R). The underlying values of HSA binding also shown in the corresponding table and were obtained from literature (Valko, K.; Nunhuck, S.; Bevan, C.; Abraham, M. H.; Reynolds, D. P., J Pharm Sci. 2003, 92, 2236-2248; Yamazaki, K.; Kanaoka, M., J Pharm Sci. 2004, 93, 1480-1494). Log t.sub.R: logarithmic value of experimentally determined retention time. Log K HSA: logarithmic value of HSA binding values.

1.9.6 Determination of Human Serum Albumin (HSA) Binding by Radio Inversed Affinity Chromatography (RIAC)

[0431] A gel filtration column Superdex 75 Increase 10/300 GL (GE Healthcare, Uppsala, Sweden) was beforehand calibrated following the producer's recommendations with a commercially available gel filtration calibration kit (GE Healthcare, Buckinghamshire, UK) comprising conalbumin (MW: 75 kDa), ovalbumin (44 kDa), carbonic anhydrase (29 kDa), ribonuclease A (13.7 kDa) and aprotinin (6.5 kDa) as reference proteins of known molecular weight. RIAC experiments were conducted using a constant flow rate of 0.8 mL/min at rt. A solution of HSA in PBS at physiological concentration (700 μM) was used as the mobile phase. PSMA ligands were labelled as described with molar activities of 10-20 GBq/μmol. Probes of 1.0 MBq of the radioligand were injected directly from the labelling solution. HSA binding was expressed as an apparent molecular weight MW calculated from the retention time of the radioligand using the determined calibration curve.

[0432] FIG. 2 shows a calibration plot of Superdex 75 Increase gel filtration column using a low molecular weight gel filtration calibration kit. The following table summarizes the underlying data. MW: molecular weight. t.sub.R: experimentally determined retention time. V: elution volume. K.sub.av: partition coefficient.

TABLE-US-00001 reference MW t.sub.R V K.sub.av blue dextran 10.034 8.027 2000 conalbumin 75000 11.859 9.487 0.0914 ovalbumin 44000 13.034 10.427 0.1503 carbonic 29000 14.912 11.930 0.2443 anhydrase ribonuklease A 13700 17.398 13.918 0.3688 aprotinin 6500 20.531 16.425 0.5257

[0433] The RIAC method is based on the Hummel-Dreyer-method, which displays ligand-protein-binding in a gel filtration chromatographic experiment using protein containing samples and a ligand containing mobile phase (Soltes L. The Hummel-Dreyer method: impact in pharmacology. Biomed Chromatogr. 2004; 18:259-271). RIAC, in an inversed way, comprises ligand samples (radiolabeled PSMA ligands) and a protein (HSA) containing mobile phase. Ligand-specific retention is caused by interaction of the ligand probe and HSA in the mobile phase. Unbound species, due to a molecular weight beneath the molecular cut off of the gel filtration column, are maximally retained while bound species are eluted in accordance to the higher molecular weight of the ligand-HSA-complex (MW.sub.HSA: 66.47 kDa). Experiments show a single radiopeak indicating a dynamic process of formation and dissociation of the ligand-HSA-complex throughout the passage through the column bed. Accordingly, the absolute retention time is the result of the ligand's mean time of abidance at HSA which is directly defined by the ligand's binding kinetics and affinity toward HSA. For evaluation, experimentally determined retention times t.sub.R are first converted into elution volumes V.sub.e by multiplying with the flow rate and thereafter converted into partition coefficients K.sub.av following the equation

[00002]$K_{\mathrm{av}}=\frac{V_e-V_0}{V_c-V_0}$

where V.sub.0 is the column void volume (8.027 mL) and V.sub.c is the geometric column volume (24 mL). Using the equation given by the trend line plot of the column calibration

K.sub.av=−0.18 ln(MW)+2.0967

the apparent molecular weight MW is calculated as

[00003]$\mathrm{MW}=e^{\frac{2.0967-K_{\mathrm{av}}}{0.18}}$

[0434] The retention time of HSA determined via UV-detection with PBS as mobile phase and a HSA-containing protein sample was found to be t.sub.R(HSA)=11.792 min, the retention time of [.sup.18F]fluoride under standard experimental conditions serving as a non-binding sample was found to be t.sub.R([.sup.18F]fluoride)=24.351 min opening a detection window of 70.2 kDa (apparent MW of HSA) to 2.3 kDa (apparent MW of [.sup.18F]fluoride; beneath molecular cut-off of the gel filtration column) defining maximal and minimal HSA-binding of evaluated PSMA-ligands.

1.10 In Vivo Experiments

1.10.1 General Information

[0435] All animal experiments were conducted in accordance with general animal welfare regulations in Germany (German animal protection act, as amended on 18 May 2018, Art. 141 G v. 29.3.2017|626, approval no. 55.2-1-54-2532-71-13) and the institutional guidelines for the care and use of animals. To establish tumor xenografts, LNCaP cells (approx. 10.sup.7 cells) were suspended in 200 μL of a 1:1 mixture (v/v) of DMEM F-12 and Matrigel (BD Biosciences, Germany), and inoculated subcutaneously onto the right shoulder of 6-8 weeks old CB17-SCID mice (Charles River, Sulzfeld, Germany). Mice were used for experiments when tumors had grown to a diameter of 5-10 mm (3-6 weeks after inoculation).

1.10.1 Biodistribution Studies

[0436] Approximately 5-10 MBq (0.1-0.2 nmol) of the radioactive-labelled PSMA inhibitors were injected into the tail vein of LNCaP tumor-bearing male CB-17 SCID mice and sacrificed after 24 h post injection. Selected organs were removed, weighted and measured in a γ-counter.

2. Results

2.1 Affinity

2.1.1 Half Maximal Inhibitory Concentration (IC.SUB.50.)

[0437] FIG. 3 and the following table shows the binding affinities (IC.sub.50 in nM) of novel ligand compounds and references to PSMA. Affinities were determined using LNCaP cells (150000 cells/well) and ([.sup.125I]I-BA)KuE (c=0.2 nM) as the radioligand (1 h, 4° C., HBSS+1% BSA). Data are expressed as mean±SD (n=3).

TABLE-US-00002 compound IC.sub.50 [.sup.natLu] - x PSMA-SiOH 7.3-SiOH 5.9 ± 1.4 10-SiOH 11.1 ± 0.5  C007-SiOH 7.1 ± 0.9 P105-SiOH 9.8 ± 0.4 P110-SiOH 42.2 ± 6.2  E102-SiOH 8.4 ± 2.4 E104-SiOH 9.0 ± 2.2 A204-SiOH 13.5 ± 3.2  PSMA-617 3.8 ± 1.7 PSMA I&T 7.9 ± 2.4 rhPSMA-7.3 8.0 ± 1.6 rhPSMA-10 2.8 ± 0.7

2.1.2 Discussion of Results

[0438] In the present investigation, the substitution of a fluoride by a hydroxide at the SiFA-benzoyl unit had little impact on the affinity comparing 7.3-SiOH and 10-SiOH with their radiohybride (rh) counterparts, respectively. The determined IC50 values of the majority of the SiOH-ligands were in a one-digit region or only slightly worse. Putting these findings into a context of ongoing clinical trials with the PSMA ligand DCFPyL (18F-DCFPyL Positron Emission Tomography (PET) in Intermediate or High Risk Prostate Cancer, https://clinicaltrials.gov/ct2/show/NCT04727736), exhibiting an IC50 value of 12.3±1.2 nM in the same assay (Robu et al., EJNMMI (2018) 8:30), PSMA affinity in this scope proofs to be sufficient.

2.2 Internalization

[0439] FIG. 4 and the following table shows a summary of the internalized activity (c=1.0 nM) at 1 hour as % of the reference ligand ([.sup.125I]I-BA)KuE (c=0.2 nM), determined on LNCaP cells (37° C., DMEM F12+5% BSA, 125000 cells/well). Data is corrected for non-specific binding (10 μmol PMPA) and expressed as mean±SD (n=3-6):

TABLE-US-00003 compound Internalization (% IBA) [.sup.177Lu] - x PSMA-SiOH 7.3-SiOH 225 ± 23 10-SiOH 203 ± 14 C007-SiOH 185 ± 15 P105-SiOH 185 ± 4  P110-SiOH 46 ± 2 E102-SiOH 203 ± 15 E104-SiOH 103 ± 2  A204-SiOH 85 ± 5 PSMA-617 160 ± 2  PSMA I&T 76 ± 2 rhPSMA-7.3 162 ± 14 rhPSMA-10 231 ± 57

[0440] FIG. 5 and the following table summarize log P.sub.O/W values of the synthesized radiolabeled .sup.177Lu-PSMA ligand compounds and .sup.177Lu-references (n=6).

TABLE-US-00004 compound Log P.sub.O/W [.sup.177Lu] - x PSMA-SiOH 7.3-SiOH −3.9 ± 0.1 10-SiOH −3.8 ± 0.1 C007-SiOH −4.28 ± 0.09 P105-SiOH −3.94 ± 0.07 P110-SiOH −4.11 ± 0.07 E102-SiOH −4.14 ± 0.08 E104-SiOH −3.77 ± 0.16 A204-SiOH −2.36 ± 0.17 PSMA-617 −4.1 ± 0.1 PSMA I&T −4.1 ± 0.1 rhPSMA-7.3 −3.8 ± 0.1 rhPSMA-10 −3.6 ± 0.1

2.4 Binding to Human Serum Albumin (HSA)

2.4.1 HSA Binding by High-Performance Affinity Chromatography (HiPAC)

[0441] FIG. 6 and the table in 2.4.2 below shows the HSA-binding of lutetium-complexed PSMA-ligand compounds and reference compounds determined by HiPAC on a Chiralpak HSA column (50×3 mm, 5 μm, H13H-2433).

2.4.2 HSA Binding by Radio Inversed Affinity Chromatography (RIAC)

[0442] FIG. 7 and the table below show the results HSA-binding of lutetium-complexed PSMA-ligands and reference compounds determined by RIAC on a Superdex 75 Increase 10/300 GL column 700 μM HSA in PBS as solvent using a constant flow rate of 0.8 mL/min (plotted data, extracted from FIG. 8).

[0443] FIG. 8 illustrates the determination of HSA binding via radio inverse affinity chromatography (RIAC). Correlation of apparent molecular weight (MW) in Dalton (Da) and retention time of .sup.177Lu-labeled samples, determined on a Superdex 75 Increase 10/300 GL with 700 μM HSA in PBS as solvent using a constant flow rate of 0.8 mL/min.

TABLE-US-00005 HSA-binding compound HSA-binding RIAC [kDa] [Lu]PSMA HiPAC [%] (see FIG. 7) 7.3-SiOH 96.4 10.1 10-SiOH 86.0 7.3 P105-SiOH 97.8 10.4 P110-SiOH 31.3 5.2 C007-SiOH 86.3 8.0 E102-SiOH 95.1 9.3 E104-SiOH 85.8 7.8 A204-SiOH 87.3 19.2 PSMA I&T 78.6 5.3 PSMA-617 74.7 13.7 Alb-02 (10) 97.7 31.6 rhPSMA-7.3 98.5 30.2 rhPSMA-10 94.0 25.4

2.4.3 Discussion of Results

[0444] The analyzed SiOH-PSMA ligands mostly show high HSA-binding of above 85% when determined by High Performance HSA Affinity Chromatography (HiPAC), indicating stronger interaction with HSA than for PSMA-617 and PSMA-I&T (74.7% and 78.6%, respectively). Ligand P110-SiOH constitutes the only exception (31.3%), which can be attributed to a positive charge within the SiFA/in-group probably hampering the ligands interaction with the basic drug binding sites I and II of HSA (Ghuman, 2005, doi: 10.1016/j.jmb.2005.07.075). Comparison of compounds PSMA-7.3-SiOH and PSMA-10-SiOH with their respective radiohybrid counterparts reveals nearly identical (96.4% vs 98.5%) and slightly decreased (86.0% vs 94.0%) HSA-binding for PSMA-7.3-SiOH and PSMA-10-SiOH, respectively. At first glance, these results seem to reflect the high similarity in structure and lipophilicity of radiohybrid compounds and their respective SiOH-analogs. However, in contrast to these findings, surprising differences in HSA-binding of rhPSMA and SiOH-PSMA ligands were found when measured with Radio Inverse Affinity Chromatography (also named AMSEC for Albumin Mediated Size Exclusion Chromatography). While high apparent Molecular Weights of 30.2 and 25.4 kDa were determined for rhPSMA-7.3 and rhPSMA-10, respectively, MW.sub.app obtained for the analog SiOH-PSMA ligands was only 10.1 and 7.3 kDa, respectively. Thus, instead of showing similarity to their radiohybrid analogs, these apparent Molecular Weights are significantly lower. This trend was further corroborated by MW.sub.app of approximately 5-10 kDa found for the remaining SiOH-PSMA compounds discussed herein. It becomes apparent, that the HSA-binding capacity of rh/SiOH-PSMA ligands in terms of apparent Molecular Weight is predominantly governed by the differing HSA-binding properties of the SiFA-group and the SiOH-group.

[0445] While both methods, HiPAC and RIAC, suggest different strength of HSA-binding for SiOH-PSMA ligands compared to rhPSMA and reference compounds, several inherent features of RIAC might qualify this methodology to predict in vivo albumin-binding more precisely. Firstly, protein-ligand-interactions in RIAC take place in solution and at physiologic pH (pH(PBS)=7.4) and HSA-concentration (700 μM), while HiPAC is based on interaction of ligands with immobilized HSA in an isopropanol containing eluent. Additionally, the output dimension of Molecular Weight [kDa] in RIAC might allow for estimation of implications on plasma half-life and excretion in vivo. In a similar fashion, apparent molecular size of PASylated proteins with expanded hydrodynamic volume was analyzed by Schlapschy et al. (Schlapschy, 2013, doi: 10.1093/protein/gzt023) Apparent molecular sizes determined in SEC experiments exceeded actual molecular sizes of the PASylated proteins and were correlated to significantly increased plasma half-life in vivo (compared to un-PASylated proteins). We therefore concluded that the surprising effect of the SiOH-group on the in vitro HSA-binding of SiOH-PSMA ligands can be exploited to create SiOH-PSMA ligands with more rapid and complete excretion kinetics (decreased residual binding to circulating HSA) compared to analog rhPSMA compounds.

[0446] FIG. 9 summarizes the Apparent Molecular Weight [kDa] as measured by Radio Inverse Affinity Chromaotography (RIAC) versus lipophilicity (log P.sub.OW=octanol/water partition coefficient) of the Lu-177 labeled SiOH-ligands and the Lu-177 labeled rhPSMA ligands [Lu-177]rhPSMA7-3 and [Lu-177]rhPSMA-10

[0447] Comparative biodistribution studies of [.sup.177Lu]-rhPSMA-7.3 vs [.sup.177Lu]-PSMA-7.3-SiOH and [.sup.177Lu]-rhPSMA-10 vs [.sup.177Lu]-PSMA-10-SiOH 24h p.i. confirm the hypothesis of more rapid and complete excretion kinetics for SiOH-PSMA ligands compared to their rhPSMA counterparts

2.5 Biodistribution in LNCaP-Tumor-Bearing Mice (24 h Post Injection)

[0448] 2.5.1 Biodistribution: rhPSMA-7.3 Vs. PSMA-7.3-SiOH

[0449] FIG. 10 and the following table show the biodistribution data 24 h p.i. of .sup.177Lu-labeled compounds in LNCaP tumor-bearing CB17-SCID mice. Data are expressed as mean±SD (n=4-5).

TABLE-US-00006 [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] organs PSMA I&T PSMA-617 rh7.3 7.3-SiOH blood 0.012 ± 0.01  0.006 ± 0.01  0.004 ± 0.002 0.002 ± 0.001 heart 0.05 ± 0.03 0.01 ± 0.01 0.03 ± 0.01 0.01 ± 0.01 lung 0.16 ± 0.03 0.04 ± 0.01 0.06 ± 0.02 0.03 ± 0.01 liver 0.05 ± 0.01 0.12 ± 0.06 0.22 ± 0.03 0.08 ± 0.03 spleen 1.94 ± 1.01 0.08 ± 0.01 0.51 ± 0.22 0.21 ± 0.11 pancreas 0.05 ± 0.02 0.01 ± 0.01 0.02 ± 0.01 0.01 ± 0.01 stomach 0.05 ± 0.02 0.02 ± 0.01 0.05 ± 0.01 0.24 ± 0.36 intestine 0.12 ± 0.06 0.12 ± 0.08 0.12 ± 0.03 0.45 ± 0.39 kidneys 34.7 ± 17.2 1.4 ± 0.4 13.2 ± 6.7  3.7 ± 1.9 adrenals 1.06 ± 0.24 0.13 ± 0.12 — — muscle 0.01 ± 0.01 0.01 ± 0.01 0.012 ± 0.004 0.006 ± 0.005 bone 0.01 ± 0.01 0.03 ± 0.01 0.02 ± 0.01 0.01 ± 0.01 tumor 4.1 ± 1.1 7.5 ± 0.9 7.6 ± 0.5 7.3 ± 0.8 parotis 0.22 ± 0.16 submandibularis 0.05 ± 0.02
2.5.2 Biodistribution: rhPSMA-10 vs. PSMA-10-SiOH

[0450] FIG. 11 and the following table show the biodistribution data 24 h p.i. of .sup.177Lu-labeled compounds in LNCaP tumor-bearing CB17-SCID mice. Data are expressed as mean±SD (n=4-5).

TABLE-US-00007 [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] organs PSMA I&T PSMA-617 rh10 10-SiOH blood 0.012 ± 0.01  0.006 ± 0.01  0.004 ± 0.001 0.002 ± 0.002 heart 0.05 ± 0.03 0.01 ± 0.01 0.03 ± 0.01 0.01 ± 0.01 lung 0.16 ± 0.03 0.04 ± 0.01 0.05 ± 0.01 0.02 ± 0.01 liver 0.05 ± 0.01 0.12 ± 0.06 0.28 ± 0.13 0.05 ± 0.01 spleen 1.94 ± 1.01 0.08 ± 0.01 0.22 ± 0.11 0.07 ± 0.04 pancreas 0.05 ± 0.02 0.01 ± 0.01 0.02 ± 0.01 0.01 ± 0.01 stomach 0.05 ± 0.02 0.02 ± 0.01 0.04 ± 0.01 0.21 ± 0.25 intestine 0.12 ± 0.06 0.12 ± 0.08 0.11 ± 0.06 0.66 ± 0.48 kidneys 34.7 ± 17.2 1.4 ± 0.4 4.1 ± 2.3 0.69 ± 0.44 adrenals 1.06 ± 0.24 0.13 ± 0.12 0.09 ± 0.06 0.01 ± 0.01 muscle 0.01 ± 0.01 0.01 ± 0.01 0.010 ± 0.003 0.005 ± 0.002 bone 0.01 ± 0.01 0.03 ± 0.01 0.02 ± 0.01 0.02 ± 0.01 tumor 4.1 ± 1.1 7.5 ± 0.9 10.2 ± 2.8  5.9 ± 1.0

2.5.3 Biodistribution: C007-SiOH

[0451] FIG. 12 and the following table show the biodistribution data 24 h p.i. of .sup.177Lu-labeled compounds in LNCaP tumor-bearing CB17-SCID mice. Data are expressed as mean±SD (n=4-5).

TABLE-US-00008 [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] organs PSMA I&T PSMA-617 C007-SiOH blood 0.012 ± 0.01  0.006 ± 0.01  0.003 ± 0.001 heart 0.05 ± 0.03 0.01 ± 0.01 0.02 ± 0.01 lung 0.16 ± 0.03 0.04 ± 0.01 0.04 ± 0.01 liver 0.05 ± 0.01 0.12 ± 0.06 0.40 ± 0.13 spleen 1.94 ± 1.01 0.08 ± 0.01 0.40 ± 0.10 pancreas 0.05 ± 0.02 0.01 ± 0.01 0.01 ± 0.01 stomach 0.05 ± 0.02 0.02 ± 0.01 0.04 ± 0.01 intestine 0.12 ± 0.06 0.12 ± 0.08 0.13 ± 0.08 kidneys 34.7 ± 17.2 1.4 ± 0.4 3.5 ± 1.4 adrenals 1.06 ± 0.24 0.13 ± 0.12 0.20 ± 0.01 muscle 0.01 ± 0.01 0.01 ± 0.01 0.004 ± 0.001 bone 0.01 ± 0.01 0.03 ± 0.01 0.02 ± 0.01 tumor 4.1 ± 1.1 7.5 ± 0.9 5.4 ± 0.8 parotis 0.06 ± 0.03 submandibularis 0.03 ± 0.01

2.5.4 Biodistribution P105-SiOH

[0452] FIG. 13 and the following table show the biodistribution data 24 h p.i. of .sup.177Lu-labeled compounds in LNCaP tumor-bearing CB17-SCID mice. Data are expressed as mean±SD (n=4-5).

TABLE-US-00009 [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] organs PSMA I&T PSMA-617 P105-SiOH blood 0.012 ± 0.01  0.006 ± 0.01  0.004 ± 0.003 heart 0.05 ± 0.03 0.01 ± 0.01 0.01 ± 0.01 lung 0.16 ± 0.03 0.04 ± 0.01 0.04 ± 0.01 liver 0.05 ± 0.01 0.12 ± 0.06 0.37 ± 0.07 spleen 1.94 ± 1.01 0.08 ± 0.01 0.31 ± 0.10 pancreas 0.05 ± 0.02 0.01 ± 0.01 0.01 ± 0.01 stomach 0.05 ± 0.02 0.02 ± 0.01 0.02 ± 0.01 intestine 0.12 ± 0.06 0.12 ± 0.08 0.08 ± 0.05 kidneys 34.7 ± 17.2 1.4 ± 0.4 4.2 ± 2.5 adrenals 1.06 ± 0.24 0.13 ± 0.12 0.12 ± 0.01 muscle 0.01 ± 0.01 0.01 ± 0.01 0.004 ± 0.001 bone 0.01 ± 0.01 0.03 ± 0.01 0.02 ± 0.01 tumor 4.1 ± 1.1 7.5 ± 0.9 6.3 ± 2.1 parotis 0.07 ± 0.03 submandibularis 0.03 ± 0.01

2.5.5 Biodistribution: E102-SiOH

[0453] FIG. 14 and the following table show the biodistribution data 24 h p.i. of .sup.177Lu-labeled compounds in LNCaP tumor-bearing CB17-SCID mice. Data are expressed as mean±SD (n=4-5).

TABLE-US-00010 [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] organs PSMA I&T PSMA-617 E102-SiOH blood 0.012 ± 0.01  0.006 ± 0.01  0.002 ± 0.001 heart 0.05 ± 0.03 0.01 ± 0.01 0.02 ± 0.02 lung 0.16 ± 0.03 0.04 ± 0.01 0.04 ± 0.02 liver 0.05 ± 0.01 0.12 ± 0.06 0.05 ± 0.02 spleen 1.94 ± 1.01 0.08 ± 0.01 0.13 ± 0.10 pancreas 0.05 ± 0.02 0.01 ± 0.01 0.01 ± 0.01 stomach 0.05 ± 0.02 0.02 ± 0.01 0.08 ± 0.07 intestine 0.12 ± 0.06 0.12 ± 0.08 0.43 ± 0.31 kidneys 34.7 ± 17.2 1.4 ± 0.4 3.3 ± 2.9 adrenals 1.06 ± 0.24 0.13 ± 0.12 0.05 ± 0.03 muscle 0.01 ± 0.01 0.01 ± 0.01 0.001 ± 0.001 bone 0.01 ± 0.01 0.03 ± 0.01 0.03 ± 0.01 tumor 4.1 ± 1.1 7.5 ± 0.9 4.4 ± 0.6 parotis — — 0.02 ± 0.01 submandibularis — — 0.01 ± 0.01

2.5.6 Biodistribution A204-SiOH

[0454] FIG. 15 and the following table show the biodistribution data 24 h p.i. of .sup.177Lu-labeled compounds in LNCaP tumor-bearing CB17-SCID mice. Data are expressed as mean±SD (n=4-5).

TABLE-US-00011 [.sup.177Lu] [.sup.177Lu] [.sup.177Lu] organs PSMA I&T PSMA-617 A204-SiOH blood 0.012 ± 0.01  0.006 ± 0.01  0.061 ± 0.027 heart 0.05 ± 0.03 0.01 ± 0.01 0.04 ± 0.01 lung 0.16 ± 0.03 0.04 ± 0.01 0.15 ± 0.01 liver 0.05 ± 0.01 0.12 ± 0.06 0.34 ± 0.03 spleen 1.94 ± 1.01 0.08 ± 0.01 0.15 ± 0.04 pancreas 0.05 ± 0.02 0.01 ± 0.01 0.01 ± 0.01 stomach 0.05 ± 0.02 0.02 ± 0.01 0.06 ± 0.02 intestine 0.12 ± 0.06 0.12 ± 0.08 0.22 ± 0.15 kidneys 34.7 ± 17.2 1.4 ± 0.4 0.92 ± 0.45 adrenals 1.06 ± 0.24 0.13 ± 0.12 0.0 ± 0.0 muscle 0.01 ± 0.01 0.01 ± 0.01 0.008 ± 0.003 bone 0.01 ± 0.01 0.03 ± 0.01 0.02 ± 0.01 tumor 4.1 ± 1.1 7.5 ± 0.9 3.5 ± 0.3 parotis 0.06 ± 0.01 submandibularis 0.04 ± 0.01

2.5.7 Discussion of Results

[0455] The examined .sup.177Lu-labelled inhibitor compounds showed the typical uptake pattern of PSMA-addressing ligands in mice 24 h p.i. with high uptake in PSMA-expressing tissues like kidneys and tumor but also in spleen and adrenal glands. The .sup.177Lu—SiOH-based ligand compounds showed lower accumulation in most of the analysed tissues and blood pool compared to the .sup.177Lu-rhPSMA compounds These results indicate a substantially decreased HSA binding and confirm the results of the RIAC-based determinations in which SiOH-PSMAs demonstrated significantly lower HSA binding compared to rhPSMAs. The tumor uptakes of the novel SiOH-based ligand compounds were lower compared to .sup.177Lu-rhPSMA-7.3 and -10 l, which can most probably be attributed to the decreased plasma protein binding and the resulting faster excretion observed for SiOH-PSMAs. Nevertheless, tumor uptakes were still in the same range or higher as determined for the state-of-the-art reference compound .sup.177Lu-PSMA I&T. Unexpectedly, all SiOH-PSMA ligands showed very low accumulation in kidneys 24 h post injection (<5% ID/g), confirming fast excretion kinetics.