Radiolabelling kit and method for radiolabelling with zirconium-89

Abstract

The present invention relates to methods and kits for radiolabelling an HBED or HBED-CC functionalized targeting agent with zirconium-89, using an easy to use cold-kit as well as the therapeutic and diagnostic use of such radiolabelled targeting agents.

Description

[0001] RADIOLABELLING KIT AND METHOD FOR RADIOLABELLING WITH ZIRCONIUM-89

[0002] TECHNICAL FIELD

[0003] The present invention is related to the field of nuclear medicine. More particular, the invention relates to radiolabelling of targeting agents, with radionuclides, more particular metal radio nuclides. The obtained radiolabelled targeting agents, may be used in therapeutic applications such as for treating tumours and cancer, such as prostate cancer or in imaging techniques, such as Positron Emission Tomography (PET) and Single-Photon Emission Computed Tomography (SPECT), for example in vivo imaging diagnosis, such as of tumours and cancer, such as prostate cancer.

[0004] BACKGROUND

[0005] As most radioisotopes used in nuclear medicine, and especially in the imaging applications, have a rather short half-life, the production of the radiolabel targeting agents is difficult to centralise, as the time to transport the radiolabelled targeting agents to the hospitals would take too long. Hence, most of the radiolabelled targeting agents are made on site, as also the radionuclides, especially metal radionuclides, can nowadays be generated on site by specifically therefore designed generators.

[0006] Generation of such radionuclides on site, prior to their use in the clinic, however requires adapted infrastructure, strong safety measures and expertise which is not available in every hospital setting around the world. It would hence be advantageous to be able to centrally produce prelabelled targeting agents, which can be shipped to, and directly used in the clinic. Most commonly used radionuclides for diagnosis and therapy however have a too short half-life, which makes it logistically non-practical or even impossible to pre-label targeting agents and ship them to the place where needed.

[0007] Enabling to centrally produce patient doses suitable for shipment to clinics in different countries, or to store the (batch) prepared radiolabelled targeting agents for a longer period of time before the radioactivity becomes too low for a patient dose would hence be beneficial in theragnostics.

[0008] One of the radionuclides that could be interesting in such a setting for both therapy and diagnosis, is zirconium-89, which has a half-life of approximately 78.4 hours (compared to the 68 minutes of e.g. gallium-68). Zr-89 is well-suited for integration into antibodies and other large molecules used in targeted therapy and diagnostic imaging. This compatibility is chiefly due to its suitable decay properties and its ability to provide clear and precise imaging results, especially in Positron Emission Tomography (PET) scans.

[0009] Zirconium-based radiopharmaceuticals can have dual utility in both diagnostic imaging and therapeutic applications. Due to the long half-life, they offer a non-invasive method to visualise and track biological processes at the molecular and cellular levels in real-time, which is increasingly important in personalised medicine. However, there are few chelators for zirconium known in the field that have been already clinically tested and used, DFO being one of them.

[0010] There is hence a need to develop new methods to radiolabel targeting agents with zirconium-89 and the present invention intends to solve at least some of the issues outlined above.

[0011] SUMMARY OF THE INVENTION

[0012] The present invention lies in adapting an existing radiolabelling method and kit for radiolabelling HBED or HBED-CC with gallium-68 to be suitable for labelling of said chelator with zirconium-89.

[0013] In order for this existing kit and method to work properly, a pre-treatment of the zirconium-89(from oxalate to chloride) and a post-treatment (purification step to capture free zirconium) of the radiolabelling mixture was developed.

[0014] The invention hence provides the following aspects:

[0015] Aspect 1 . A method for radiolabelling an HBED or HBED-CC functionalized targeting agent with a metal radionuclide being zirconium-89, comprising the steps of: a) providing an HBED or HBED-CC functionalized targeting agent; b) providing a buffering agent or buffer solution, allowing to maintain the pH in the range of 3.5 to 7, such as 3.5 to 6.5 or 4.0 to 5.5, more preferably 4.5 to 5.0; c) providing a suitable amount of zirconium-89-halide, preferably a suitable amount of zirconium-89-chloride; preferably obtained by converting a suitable amount of zirconium- 89-oxalate to zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4) by means of anion- exchange, or can be sourced from cyclotron production; d) eluting a suitable amount of zirconium-89-halide obtained in step c) into a mixture of a) and b) and allowing the radiolabelling reaction to take place, preferably at room temperature; e) optionally, purifying the radiolabelling mixture by eliminating unbound zirconium-89 from the radiolabelling mixture, thereby obtaining a zirconium-89 radiolabelled targeting agent.

[0016] Aspect 2. The method according to aspect 1 , wherein said buffering agent or buffer solution is a phosphate, nitrate, HEPES, acetate, formate, TRIS, ascorbate, and citrate or a mixture thereof, preferably an acetate or formate buffer, more preferably a sodium acetate or sodium formate buffer, most preferably a sodium acetate buffer.

[0017] Aspect 3. The method according to aspect 1 or 2, wherein the anion exchange step is performed on an anion exchange column, such as a HCO3‘ resin, column or cartridge, or a SepPak QMA anion exchange cartridge. In a preferred embodiment, said cartridge is first washed with de-ionized (Milli Q) water to remove the oxalate and contaminants and then eluted with HCI to elute the zirconium-chloride form, preferably about 1M HCI or more.

[0018] Aspect 4. The method according to any one of aspects 1 to 3, wherein the purification step e) is done by means of solid phase extraction or reverse phase extraction, preferably over a silica- based substrate with strong hydrophobicity such as C18-coated silica, C8-coated silica or C4- coated silica, depending on the targeting ligand to be labelled.

[0019] Typically, SPE substrates are made of high-purity silica, coated with C18, which is a hydrophobic and non-polar stationary phase, selectively retaining non-polar and moderately polar compounds such as lipids, steroids, and pesticides while allowing more polar compounds such as sugars and amino acids to pass through.

[0020] Aspect 5. The method according to any one of previous aspects, wherein additionally a metal inhibitor is added to the radiolabelling mixture, said metal inhibitor being a co-chelating agent, capable of inactivating metals other than radioactive metal without interfering with the chelation between the radioactive metal and the said chelate-functionalized targeting agent, under the conditions of the labelling reaction.

[0021] Aspect 6. The method according to any one of previous aspects, wherein said metal inhibitor is a sugar, preferably a short-chain sugar or oligosaccharide, such as comprising up to 7 monosaccharide units.

[0022] Aspect 7. The method according to any one of previous aspects, wherein said metal inhibitor is selected from the group comprising: monosaccharides and their derivatives, disaccharides and their derivatives, and cyclodextrins. Aspect 8. The method according to any one of previous aspects, wherein said metal inhibitor is selected from the group comprising: Glucose, D-Fructose, Beta-cyclodextrin, D-Mannose, and beta-cyclodextrin, more preferably D-mannose.

[0023] Aspect 9. The method according to any one of previous aspects, wherein said metal inhibitor and said functionalised agent are not chemically linked.

[0024] Aspect 10. The method according to any one of previous aspects, wherein said metal inhibitor and said functionalised agent are chemically linked, through a linker that is unstable in the radiolabelling conditions.

[0025] Aspect 11 . The method according to any one of previous aspects, wherein the chelate- functionalized targeting agent comprises a targeting moiety selected from the list comprising a peptide, a urea-based peptidomimetic, a polypeptide, a protein, a vitamin, a saccharide, an antibody, a nucleic acid, an aptamer, an antisense oligonucleotide, or an organic molecule.

[0026] Aspect 12. The method according to any one of previous aspects, wherein said chelate- functionalized targeting agent is Glu-urea-Lys-HBED-CC (gozetotide or PSMA-11).

[0027] Aspect 13. The method according to any one of previous aspects, wherein the radiolabelling reaction is carried out at ambient or room temperature (such as between 15 and 30°C).

[0028] Aspect 14. The method according to any one of the previous aspects, wherein the activity is up to 100 mCi, preferably from 3 mCi to 100 mCi.

[0029] Aspect 15. The method according to any one of aspects 1 to 14, wherein additionally a stabiliser is added to the radiolabelling mixture that prevents radiolysis (product degradation) of the chelate-functionalized targeting agent, wherein said stabiliser is selected from the group consisting of: ascorbic acid, dehydroascorbic acid, gentisic acid, cysteine, N-acetylcysteine, and methionine, or a salt thereof, preferably as a solution to the radiolabelling mixture prior to radiolabelling;

[0030] Aspect 16. The method according to aspect 15, producing a zirconium-89 radiolabelled targeting agent with an activity of at least 50.0 mCi, preferably at least 60.0 mCi, preferably at least 70.0 mCi, preferably at least 80.0 mCi, preferably at least 90.0 mCi, preferably at least 100.0 mCi, preferably at least 150.0 mCi, preferably at least 200.0 mCi, preferably at least 250.0 mCi, preferably at least 300.0 mCi, preferably at least 350.0 mCi, preferably at least 400.0 mCi, preferably at least 450.0 mCi, preferably at least 500.0 mCi, preferably at least 750.0 mCi, preferably at least 1000.0 mCi. Aspect 17. The method according to aspect 15 or 16, wherein the zirconium-89 is provided as a solution with an radioactive concentration of at least 5.0 mCi / ml, preferably at least 6.0 mCi / ml, preferably at least 7.0 mCi / ml, preferably at least 10.0 mCi / ml, preferably at least 15.0 mCi / ml, preferably at least 20.0 mCi / ml, preferably at least 25.0 mCi / ml, preferably at least 30.0 mCi / ml, preferably at least 35.0 mCi / ml, preferably at least 40.0 mCi / ml, preferably at least 45.0 mCi / ml, preferably at least 50.0 mCi / ml, preferably at least 75.0 mCi / ml, preferably at least 100.0 mCi / ml.

[0031] Aspect 18. The method according to any one of aspects 15 to 17, wherein the mixture radiolabelling mixture comprises at least 10.0 mg, preferably at least 15.0 mg, preferably at least 20.0 mg, preferably at least 25.0 mg, preferably at least 30.0 mg of stabiliser, more preferably of ascorbic acid.

[0032] Aspect 19. The method according to any one of aspects 15 to 18, wherein the radiolabelling mixture comprises at most 100.0 mg, preferably at most 90.0 mg, preferably at most 80.0 mg, preferably at most 70.0 mg, preferably at most 60.0 mg, preferably at most 50.0 mg of stabiliser, more preferably of ascorbic acid.

[0033] Aspect 20. The method according to any one of aspects 15 to 19, wherein the radiolabelling mixture comprises at least 1 .0 mg, preferably at least 1 .2 mg, preferably at least 1 .4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of stabiliser per 50 mCi metal radionuclide in step f); more preferably wherein the radiolabelling mixture comprises of at least 1.0 mg, preferably at least 1.2 mg, preferably at least 1.4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of ascorbic acid per 50 mCi zirconium-89 in step f).

[0034] Aspect 21 . The method according to any one of aspects 15 to 20, wherein the radiolabelling mixture comprises at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of stabiliser per 50 mCi metal radionuclide in step f); more preferably wherein the radiolabelling mixture comprises of at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of ascorbic acid per 50 mCi zirconium-89 in step f). Aspect 22. The method according to any one of aspects 15 to 21 , wherein the amount of stabiliser in the radiolabelling mixture, is such that the weight ratio of the weight of stabiliser over the weight of the chelate-functionalized targeting agent in the radiolabelling mixture, is at least 40, preferably at least 80, preferably at least 150, preferably at least 300, preferably at least 600, preferably at least 1000, preferably at least 1500, preferably at least 2000.

[0035] Aspect 23. The method according to any one of aspects 15 to 22, wherein the solution of stabiliser has a concentration of at least 0.14 mg / ml, preferably at least 0.30 mg / ml, preferably at least 0.50 mg / ml, preferably at least 1 .00 mg / ml, preferably at least 2.00 mg / ml, preferably at least 4.00 mg / ml, preferably at least 5.00 mg / ml, preferably at least 7.00 mg / ml, preferably at least 9.00 mg / ml, preferably at least 10.00 mg / ml, preferably wherein the stabilizer is ascorbic acid.

[0036] Aspect 24. The method according to any one of aspects 15 to 23, wherein the stabiliser is ascorbic acid, dehydroascorbic acid or a salt thereof, and optionally wherein said stabiliser is provided together with a bisulfite and or a metabisulfite.

[0037] Aspect 25. The method according to any one of aspects 15 to 24, for providing at least 3.0, preferably at least 4.0, preferably at least 5.0, preferably at least 6.0, preferably at least 8.0, preferably at least 10.0, preferably at least 12.0, preferably at least 15.0, preferably at least 20.0 patient doses.

[0038] Aspect 26. A zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent obtained by the method according to anyone of aspects 1 to 25.

[0039] Aspect 27. The radiolabelled targeting agent according to aspect 26, which is zirconium-89 radiolabelled Glu-urea-Lys-HBED-CC (PSMA-11 or gozetotide).

[0040] Aspect 28. A radiolabelling kit, preferably suitable for producing a zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent with an activity of up to 50.0 mCi, comprising:

[0041] - an HBED or HBED-CC functionalized targeting agent, preferably in lyophilized form;

[0042] - a buffering agent or buffer solution, allowing to maintain the pH in the range of 3.5 to 7, such as 3.5 to 6.5 or 4.0 to 5,5, more preferably 4,5 to 5.0, optionally in lyophilized form;

[0043] - optionally, an anion exchange means suitable for converting zirconium-89-oxalate to zirconium-89-halide, preferably zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4);

[0044] - optionally, a purification means suitable for eliminating unbound zirconium-89 from the radiolabelling mixture after the radiolabelling reaction took place. Aspect 29. The kit according to aspect 28, additionally comprising a metal inhibitor, which is a co-chelating agent, capable of inactivating metals other than radioactive metal without interfering with the chelation between the radioactive metal and the said chelate-functionalized targeting agent, under the conditions of the labelling reaction.

[0045] Aspect 30. A high activity radiolabelling kit, preferably suitable for producing a zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent with an activity of at least 50.0 mCi, comprising:

[0046] - an HBED or HBED-CC functionalized targeting agent, preferably in lyophilized form;

[0047] - a buffering agent or buffer solution, allowing to maintain the pH in the range of 4.0 to 5.5, more preferably 4.5 to 5.0, optionally in lyophilized form;

[0048] - a stabiliser selected from the group consisting of: ascorbic acid, sodium ascorbate, dehydroascorbic acid, gentisic acid, cysteine and methionine, or a salt thereof, preferably as a solution;

[0049] - optionally, an anion exchange means suitable for converting zirconium-89-oxalate to zirconium-89-halide, preferably zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4);

[0050] - optionally, a purification means suitable for eliminating unbound zirconium-89 from the radiolabelling mixture after the radiolabelling reaction took place.

[0051] Aspect 31 . The kit according to any one of aspects 28 to 30, wherein said buffering agent or buffer solution is a phosphate, nitrate, HEPES, acetate, formate, TRIS, ascorbate, and citrate or a mixture thereof, preferably an acetate or formate buffer, more preferably a sodium acetate or sodium formate buffer, most preferably a sodium acetate buffer.

[0052] Aspect 32. The kit according to any one of aspects 28 to 31 , wherein said anion exchange means is a HCO3‘ resin, column or cartridge. In a preferred embodiment, said cartridge is first washed with de-ionized (Milli Q) water to remove the oxalate and contaminants and then eluted with 1 M HCI to elute the zirconium-chloride form.

[0053] Aspect 33. The kit according to any one of aspects 28 to 32, wherein the means is a solid phase extraction or reverse phase extraction means, preferably a silica-based substrate with strong hydrophobicity such as C18-coated silica. Different hydrophobic interaction resins with C4- , C6-, C8-, phenyl-, or benzyl- coated silica can be used as a substrate as well depending on the chemical character of the targeting agent. Typically, SPE substrates are made of high-purity silica, coated with C18, which is a hydrophobic and non-polar stationary phase, selectively retaining non-polar and moderately polar compounds such as lipids, steroids, and pesticides while allowing more polar compounds such as sugars and amino acids to pass through.

[0054] Aspect 34. The kit according to any one of aspects 28 to 33, wherein the HBED or HBED-CC- functionalized targeting agent and the buffer are provided in an lyophilized form, either in two separate or in one single vial. In one embodiment, also the metal inhibitor and / or stabilising agent are present in lyophilised form, either in a separate vial, or combined with either the buffer or targeting agent.

[0055] Aspect 35. The kit according to any one of aspects 30 to 34, suitable for radiolabelling a chelate-functionalized targeting agent with zirconium-89 thereby producing a zirconium-89 chelate-functionalized targeting agent with an activity of at least 50.0 mCi, preferably at least 100.0 mCi, preferably at least 150.0 mCi, preferably at least 200.0 mCi, preferably at least 250.0 mCi, preferably at least 300.0 mCi, preferably at least 350.0 mCi, preferably at least 400.0 mCi, preferably at least 450.0 mCi, preferably at least 500.0 mCi, preferably at least 750.0 mCi, preferably at least 1000.0 mCi.

[0056] Aspect 36. The kit according to any one of aspects 30 to 35, wherein the zirconium-89 is provided as a solution with an activity of at least 5.0 mCi / ml, preferably at least 7.0 mCi / ml, preferably at least 10.0 mCi / ml, preferably at least 15.0 mCi / ml, preferably at least 20.0 mCi / ml, preferably at least 25.0 mCi / ml, preferably at least 30.0 mCi / ml, preferably at least 35.0 mCi / ml, preferably at least 40.0 mCi / ml, preferably at least 45.0 mCi / ml, preferably at least 50.0 mCi / ml, preferably at least 75.0 mCi / ml, preferably at least 100.0 mCi / ml.

[0057] Aspect 37. The kit according to any one of aspects 30 to 36, comprising at least 10.0 mg, preferably at least 15.0 mg, preferably at least 20.0 mg, preferably at least 25.0 mg, preferably at least 30.0 mg of stabiliser, wherein said stabiliser preferably is ascorbic acid.

[0058] Aspect 38. The kit according to any one of aspects 30 to 37, wherein the kit comprises at most 100.0 mg, preferably at most 90.0 mg, preferably at most 80.0 mg, preferably at most 70.0 mg, preferably at most 60.0 mg, preferably at most 50.0 mg of stabiliser, more preferably of ascorbic acid.

[0059] Aspect 39. The kit according to any one of aspects 30 to 38, wherein the kit comprises at least 1 .0 mg, preferably at least 1 .2 mg, preferably at least 1 .4 mg, preferably at least 1 .6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of stabiliser per 50 mCi metal radionuclide; more preferably wherein the kit comprises of at least 1.0 mg, preferably at least 1.2 mg, preferably at least 1.4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of ascorbic acid per 50 mCi zirconium-89.

[0060] Aspect 40. The kit according to any one of aspects 30 to 39, wherein the kit comprises at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of stabiliser per 50 mCi metal radionuclide; more preferably wherein the kit comprises of at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of ascorbic acid per 50 mCi zirconium-89.

[0061] Aspect 41. The kit according to any one of aspects 30 to 40, comprising at least 10.0 pg, preferably at least 15.0 pg, preferably at least 20.0 pg, preferably at least 25.0 pg of said chelate- functionalized targeting agent.

[0062] Aspect 42. The kit according to any one of aspects 30 to 41 , comprising at least 2.0 pg, preferably at least 4.0 pg, preferably at least 6.0 pg, preferably at least 8.0 pg, preferably at least 10.0 pg of said metal inhibitor.

[0063] Aspect 43. The kit according to any one of aspects 30 to 42, comprising at least 50.0 mg, preferably at least 75.0 mg, preferably at least 100.0 mg, preferably at least 125.0 mg, preferably at least 150.0 mg of said buffering agent.

[0064] Aspect 44. The kit according to any one of aspects 30 to 43, wherein the amount of stabiliser in the kit, is such that the weight ratio of the weight of stabiliser over the weight of the chelate- functionalized targeting agent in the kit, is at least 40, preferably at least 80, preferably at least 150, preferably at least 300, preferably at least 600, preferably at least 1000, preferably at least 1500, preferably at least 2000. In a preferred embodiment, said stabiliser is ascorbic acid.

[0065] Aspect 45. The kit according to any one of aspects 30 to 44, wherein the solution of stabiliser, has a concentration of at least 0.14 mg / ml, preferably at least 0.30 mg / ml, preferably at least 0.50 mg / ml, preferably at least 1.00 mg / ml, preferably at least 2.00 mg / ml, preferably at least 4.00 mg / ml, preferably at least 5.00 mg / ml, preferably at least 7.00 mg / ml, preferably at least 9.00 mg / ml, preferably at least 10.00 mg / ml. In a preferred embodiment, said stabiliser is ascorbic acid.

[0066] Aspect 46. The kit according to any one of aspects 30 to 45, wherein the stabiliser is provided in the kit together with a bisulfite and or a metabisulfite.

[0067] Aspect 47. The kit according to any one of aspects 30 to 46, wherein the kit is suitable for providing at least 3.0, preferably at least 4.0, preferably at least 5.0, preferably at least 6.0, preferably at least 8.0, preferably at least 10.0, preferably at least 12.0, preferably at least 15.0, preferably at least 20.0 patient doses.

[0068] Aspect 48. The kit according to any one of aspects 28 to 47, wherein said metal inhibitor is a sugar, preferably a short-chain sugar or oligosaccharide, such as comprising up to 7 monosaccharide units.

[0069] Aspect 49. The kit according to anyone of aspects 28 to 48, wherein said metal inhibitor is selected from the group comprising: monosaccharides and their derivatives, disaccharides and their derivatives, and polysaccharides and cyclodextrins.

[0070] Aspect 50. The kit according to anyone of aspects 28 to 49, wherein said metal inhibitor is selected from the group comprising: Glucose, D-Fructose, Beta-cyclodextrin, and D-Mannose.

[0071] Aspect 51 . The kit according to any one of aspects 28 to 50, wherein said metal inhibitor and said functionalised agent are not chemically linked.

[0072] Aspect 52. The kit according to any one of aspects 28 to 51 , wherein said metal inhibitor and said functionalised agent are chemically linked, through a linker that is unstable in the radiolabelling conditions.

[0073] Aspect 53. The kit according to any one of aspects 28 to 52, wherein said targeting agent is Glu-urea-Lys-HBED-CC (gozetotide or PSMA-11).

[0074] Aspect 54. An imaging method, comprising the steps of:

[0075] 1) radiolabelling an HBED or HBED-CC functionalized targeting agent with zirconium-89 according to the method of any one of aspects 1 to 24, preferably by using a kit according to any one of claims 25 to 53,

[0076] 2) administering to a subject a diagnostic amount of said zirconium-89 radiolabelled targeting agent; and, 3) detecting said zirconium-89 radiolabelled targeting agent when bound to its target using PET or PET / CT imaging methods.

[0077] Aspect 55. A method of detecting a prostate tumour or cancer, comprising the steps of:

[0078] 1) radiolabelling Glu-urea-Lys-HBED-CC (PSMA-11 or gozetotide) with zirconium-89 according to the method of any one of aspects 1 to 24, preferably by using a kit according to any one of claims 25 to 53,

[0079] 2) administering to a subject a diagnostic amount of zirconium-89 radiolabelled PSMA-11 (gozetotide); and,

[0080] 3) detecting binding of said zirconium-89 radiolabelled PSMA-11 (gozetotide) using PET or PET / CT imaging methods.

[0081] Aspect 56. The method according to aspect 55, wherein said detection is used for:

[0082] (i) initial staging of prostate cancer into intermediate, unfavourable, high, or very high-risk prostate cancer,

[0083] (ii) detecting suspected recurrence of prostate cancer and / or detection of metastasis,

[0084] (iii) selection for radiotherapeutic treatment such as with Lutetium (177Lu) vipivotide tetraxetan (Pluvicto),

[0085] (iv) monitoring prostate cancer for progression into Non-Metastatic or Metastatic Castration-Resistant Prostate Cancer (nmCRPC or mCRPC), or

[0086] (v) determining response to (radio)therapy.

[0087] (vi) real-time imaging or diagnosis in a subject e.g. for use in PET scans for tracking slow biological processes or progression of those over a period of time. In some embodiments, this aspect can be carried out by a computer or processor and hence is directed by a system for in vivo diagnosis or for in vivo imaging in a subject, said system comprising a processing means (processor) configured to carry out the steps of receive real-time imaging data and presenting said imaging date in real-time to the user, thereby enabling in vivo imaging or in vivo diagnosis of said subject.

[0088] (vii) personalised medicine

[0089] Aspect 57. The method according to any one of aspects 54 to 56, wherein said detection method is used to replace the need for taking a prostate biopsy or is used in PET or PET / CT scan with MRI in clinically significant or intermediate favourable prostate cancers, or in MRI for active surveillance of prostate cancer.

[0090] Aspect 58. The zirconium-89 radiolabelled targeting molecule according to aspect 25 or 26, for use in radiotherapy.

[0091] Aspect 59. The zirconium-89 radiolabelled targeting molecule according to aspect 25 or 26, for use in radiodiagnosis, preferably for use in in vivo real time detection of a targeting agent.

[0092] Aspect 60. In any one of the above aspects relating to kits, said kit can comprise 2 vials, wherein: the first vial comprises 25.0 pg PSMA-11 , and 10.0 pg D-mannose as a lyophilised powder in a sterile 10 ml vial; the second vial comprises 150.0 mg anhydrous sodium acetate in 2.5 ml 0.292 M HCI solution in a sterile 10 ml vial;

[0093] Aspect 61 . A method for treatment of cancer, comprising the step of: administering an efficient amount of the zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent according to aspect 25 to a patient in need thereof. In some embodiments, said patient can have been diagnosed using the method of aspect 54 to 57.

[0094] Aspect 62. A method for treatment of cancer, comprising the step of: administering an efficient amount of the zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent according to aspect 26 to a patient in need thereof.

[0095] Aspect 63. In any one of the above aspects directed to methods, the amounts of reagents used can be as follows:

[0096] 25.0 pg PSMA-11 , and 10.0 pg D-mannose, both as a lyophilised powder;

[0097] 150.0 mg anhydrous sodium acetate in 2.5 ml 0.292 M HCI solution.

[0098] In any of the above aspects directed to “methods of treatment”, the corresponding “products for use in treating” or “use of product for the manufacture of a medicament for treating” formatting is also included. DETAILED DESCRIPTION

[0099] As used herein, the singular forms “a”, “an”, and “the” include both singular and plural referents unless the context clearly dictates otherwise.

[0100] The terms “comprising”, “comprises” and “comprised of’ as used herein are synonymous with “including”, “includes” or “containing”, “contains”, and are inclusive or open-ended and do not exclude additional, non-recited members, elements or method steps. The terms also encompass “consisting of’ and “consisting essentially of’.

[0101] The recitation of numerical ranges by endpoints includes all numbers and fractions subsumed within the respective ranges, as well as the recited endpoints.

[0102] The term “about” as used herein when referring to a measurable value such as a parameter, an amount, a temporal duration, and the like, is meant to encompass variations of and from the specified value, in particular variations of + / -10% or less, preferably + / -5% or less, more preferably + / -1 % or less, and still more preferably + / -0.1 % or less of and from the specified value, insofar such variations are appropriate to perform in the disclosed invention. It is to be understood that the value to which the modifier “about” refers is itself also specifically, and preferably, disclosed.

[0103] Whereas the term “one or more”, such as one or more members of a group of members, is clear per se, by means of further exemplification, the term encompasses inter alia a reference to any one of said members, or to any two or more of said members, such as, e.g., any >3, >4, >5, >6 or >7 etc. of said members, and up to all said members.

[0104] All documents cited in the present specification are hereby incorporated by reference in their entirety.

[0105] Unless otherwise specified, all terms used in disclosing the invention, including technical and scientific terms, have the meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. By means of further guidance, term definitions may be included to better appreciate the teaching of the present invention.

[0106] In the following passages, different aspects or embodiments of the invention are defined in more detail. Every aspect or embodiment so defined may be combined with each of the other aspects or embodiments unless stated otherwise. In particular, any feature indicated as being preferred or advantageous in one embodiment may be combined with any other embodiment or embodiments indicated as being preferred or advantageous. The present invention overcomes one or more of the problems identified and observed in the state of the art and allows stabilize radiolabelled chelate-functionalized targeting agents compositions with a high activity.

[0107] The present invention is related to a method for radiolabelling radiopharmaceuticals, preferably an HBED or HBED-CC functionalized targeting agent, with a metal radionuclide being zirconium- 89, comprising the steps of: a) providing an unlabelled radiopharmaceutical, preferably an HBED or HBED-CC functionalized targeting agent; b) providing a buffering agent or buffer solution, allowing to maintain the pH in the range of between 4.0 to 5.5, more preferably between 4.5 to 5.0; c) providing a suitable amount of zirconium-89-halide, preferably a suitable amount of zirconium-89-chloride; d) eluting a suitable amount of zirconium-89-halide, preferably zirconium-89-chloride, obtained in step c) into a mixture of a) and b) and allowing the radiolabelling reaction to take place, preferably at room temperature; e) optionally, purifying the radiolabelling mixture by eliminating unbound zirconium-89 from the radiolabelling mixture, thereby obtaining a zirconium-89 radiolabelled radiopharmaceutical, preferably a zirconium-89 radiolabelled targeting agent.

[0108] Preferably, said zirconium halide is obtained by converting a suitable amount of zirconium-89- oxalate to zirconium-89-halide, preferably converting a suitable amount of zirconium-89-oxalate to zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4); preferably by means of anion- exchange;

[0109] The present inventors have found an efficient way to use zirconium-89 as a radiolabel for radiopharmaceuticals, especially radiopharmaceuticals comprising a HBED, HBED-CC or a derivative thereof as chelator. More particular, [zirconium-89]-labelling of PSMA-11 (or Gozetotide) was demonstrated by the inventors to be efficient.

[0110] The longer half-life of zirconium-89 vs. e.g. gallium-68 also reduces the time pressure for in vivo diagnosis as compared to other radionuclides with a shorter half-life, thereby decreasing human error and increasing accuracy of diagnosis / therapy. The use of zirconium-89 may also allow for a centralized production of [zirconium-89]-labelled targeting agents such as PSMA11 with a sufficient high radioactivity, allowing transport to hospitals or other end users, without the need for expensive and highly regulated infrastructure and safety measures, expensive equipment and highly trained personnel.

[0111] It may also allow to postpone imaging after the wash-out (clearance) from the patient of free zirconium-89]-labelled PSMA-11 (not bound to the target), which could increase the contrast and resolution.

[0112] In addition, the energy of the positrons emitted by zirconium-89 is slightly lower than e.g. gallium- 68, which allows for a slightly higher resolution. Hence, as an example the imaging of prostate cancer using [zirconium-89]-labelled PSMA-11 , may be more sensitive compared to previous methods known in the art.

[0113] Zirconium-89 is often commercially supplied in a complexed manner, meaning that Zirconium-89 is supplied in a complex with a polydentate ligand, such as oxalate or EDTA and the like. The inventors have found that Zirconium-89 in its complexed form (e.g. [89Zr]Zr-Oxalate) has a very low tendency to bind HBED, HBED-CC or a derivatives thereof. The inventors have found that converting the complexed zirconium-89 such as its chelate form, to a zirconium-89 halide (e.g. [89Zr]Zr-CI4) drastically increases the radiolabelling yields. Other forms of suitable halides would in principle be a fluoride, chloride, bromide, iodide or astatide, although these may be more difficult to use in the clinic.

[0114] The aspects and embodiment as described herein advantageously allow to obtain an appropriate chelation yield, particularly above 50%, more particular above 60%, yet more particular above 70%, still yet more particular above 75%, or even above 80%.

[0115] These chelation yields, optionally together with a preliminary or further final purification of the radiolabelled molecule, allow for a high radiochemical purity, e.g. of above 90%, above 95%, or even of and above 98% radiochemical purity.

[0116] Said high radiochemical purity can also be maintained over time when using high radioactivities of up to 500 mCi (from 10 mCi to 500 mCi).

[0117] As used herein, radioactivities are expressed in Curie [Ci] as unit. However, the conversion of Curie to Becquerel [Bq] is well known in the art, as 1 Ci = 3.7 ■ 1010Bq. Hence, 500 mCi is 1.85 ■ 1010Bq. In some embodiments, all kit components as described herein can be lyophilized altogether or frozen which ensures a longer shelf life.

[0118] Thus, the main advantages of the invention as disclosed herein that differentiate from the state of the art are:

[0119] - the radiolabelling method can be carried out centrally, for example at a specialised radiochemical laboratory or production site, or close to a [zirconium-89]-production site, such as a nuclear reactor or a cyclotron;

[0120] - the possibility of a radiolabelling without the need for heating; and / or,

[0121] - the production of highly active radiopharmaceuticals, allowing therapeutics and / or diagnostics, even with multiple acquisitions of images after one single administration.

[0122] - the possibility of radiolabelling without the need for an automated synthesizer, thereby opening up the possibility of using kits providing the necessary chemical, to be used with different sources of zirconium-89;

[0123] Furthermore, a metal inhibitor or scavenger can be used in the radiolabelling method for neutralizing, at least partially, interfering (radio)metal species and allowing the desired radioactive metal to react with the chelate-functionalized targeting agent. These metal inhibitors may temporarily or permanently remove metals that compete with radioactive metal for the reaction with the chelate-functionalized targeting agent. Said metal inhibitor is thus unable (or has a lower ability) to chelate the radioactive metal in the said conditions of the labelling reaction, but chelates other metals interfering with the chelation of radioactive metal by the chelate-functionalized targeting agent. The presence of metal inhibitors during the radiolabelling reaction provides an advantageous alternative to current approaches for managing the presence of metallic impurities such as increasing the amount of chelate-functionalized targeting agent or the pre-treatment of the eluate of the generator, these additional purification steps consume time (and radioactivity). Hence, in some embodiments, metal inhibitors are used in the present invention, which are selected for their ability to block the competing metals in the radiolabelling reaction without inhibiting the radioactive metal ions in their chelation reaction with the chelate-functionalized targeting agent. Indeed, these metal inhibitors should not interfere negatively on the main radiolabelling reaction or lead to the formation of secondary radiolabelled species. In other words, metal inhibitors should have a limited or no capacity to complex radioactive metal in the conditions used for the radiolabelling reaction. Limited means at least 100 times less than the chelating agent used for the radiolabelling of the chelate-functionalized targeting agent. It is interesting to note that the function of metal inhibitors in some embodiments of the present invention is the opposite of the function of the sequestering agents used in the prior art. Indeed, according to known methods, at the end of the labelling reaction, a sequestering agent having a particular affinity for e.g. the radioactive zirconium may be added to chelate the unreacted portion of the isotope, whereas, according to the present invention an agent capable of reducing the competition of metallic impurities other than the radioactive metal is added at the beginning of the reaction.

[0124] As used herein, an "inhibitor of metal" refers to any molecule capable of interacting with, or competing metals, or the chelating moiety of the chelate-functionalized targeting agent or with radioactive metal directly, to inhibit wholly or partially the chelation the chelate-functionalized targeting agent said competing metals and / or promote the chelating of radioactive metal by said targeting agent.

[0125] Metal inhibitors are preferably selected from the group of sugars. Sugars used as agents metal inhibitors in the kit of the invention are generally oligosaccharides (up to 6 or 7 monomeric sugar units or monosaccharides) and for example can be monosaccharides or disacharides or derivatives of monosaccharides such as tetracetose, pentacetose, hexacetose, tetrose, pentose, hexose, D-mannose, D-fructose, and derivatives; and / or disaccharides and their derivatives such as maltose and its derivatives; and I or, cyclodextrins and derivatives thereof.

[0126] Preferably, the metal inhibitor is present in the kit as described herein in micromolar amounts, preferably in nanomolar quantities, preferably in an amount of less than 500 nanomolar, still more preferably in an amount less than 100 nanomoles. In a preferred embodiment, said metal inhibitor is present in an amount of from 20 to 40 wt.% or from 25 to 35 wt.% based on the total weight of the chelate-functionalized targeting agent and metal inhibitor.

[0127] When used, the metal inhibitory agent is usually not bound to the chelate-functionalized targeting agent but may also be chemically bound to the chelate-functionalized targeting agent when the chemical bond is a labile (breakable) bond under the conditions of radiolabelling with the chelate- functionalized targeting agent being released in situ in the conditions of radiolabelling. In one embodiment, said metal inhibitory agent is not chemically bound to the chelate-functionalized targeting agent.

[0128] In case of upscaling the amount of activity to be used, adding a certain amount of stabiliser to the radiolabelling solution priorto the radiolabelling reaction enables to avoid radiolysis of the chelate- functionalized targeting agent making it possible to use high activity radiolabelling conditions in the clinic.

[0129] As used here, the term “stabilizer” refers to a compound with the ability to decrease or to prevent the radiolysis of the chelate-functionalized targeting agent and / or other compounds of the obtained radiolabelled chelate-functionalized targeting agent composition. Preferably, the stabilizer allows for a radiochemical purity of the radiolabelled chelate-functionalized targeting agent after 4 hours of at least 95%, and that preferably at radioactive concentrations higher than 7.0 mCi / ml. More preferably, the stabilizer allows for a radiochemical purity of the radiolabelled chelate-functionalized targeting agent after 6 hours of at least 95%, and that preferably at radioactive concentrations higher than 7.0 mCi / ml.

[0130] When used, the stabiliser is preferably selected from the group consisting of: ascorbic acid, dehydroascorbic acid, gentisic acid, cysteine and methionine, or a salt thereof. A typical salt of ascorbic acid that can be used as stabilizer is sodium ascorbate.

[0131] As used herein, a "chelate-functionalized targeting agent" refers to a targeting agent capable of being labelled with a radioisotope such as for example radioactive metal, by means of a chelation agent to which this targeting agent is bound.

[0132] Preferred chelation agents for functionalizing a targeting agent to be radiolabelled with radioactive metals are those which form stable complexes at least for a time sufficient for diagnostic investigations using radiolabelled targeting agents. Suitable chelating agents include aliphatic amines, linear or macrocyclic such as macrocyclic amines with tertiary amines. While these examples of suitable chelating agents are not limited, they preferably include N,N'-bis(2- hydroxybenzyl)ethylenediamine-N,N'-diacetic acid (HBED) or 3-[3-[4-[5-(2-carboxyethyl)-2- hydroxyphenyl]-1 ,4-bis(carboxymethylamino)butyl]-4-hydroxyphenyl]propanoic acid (HBED-CC), 1 ,4,7,10-tetraazacyclododecane-1 ,4,7,10-tetraacetic acid (DOTA), 1 ,4,7-triazacyclononane- 1 ,4,7-triacetic acid (NOTA), 1 ,4,7-triazacyclononane macrocycles substituted with phosphonic (NOTP) and phosphinic (TRAP) groups at the amine, tris(hydroxypyridinone) containing three 1 ,6- dimethyl-3-hydroxypyridin-4-one groups (THP) the hexadentate tris(hydroxamate) siderophore desferrioxamine-B (DFO), Ethylenediaminetetraacetic acid (EDTA), N,N’-bis(2,2-dimethyl-2- mercaptoethyl)ethylenediamine-N,N’-diacetic acid (6SS), 1-(4- carboxymethoxybenzyl)-N-Nz- bis[(2-mercapto-2,2-dimethyl)ethyl]-1 ,2- ethylenediamine-N,Nz-diacetic acid (B6SS), N,N'- dipyridoxylethylenediamine- N,N'-diacetic acid (PLED), 1 ,1 ,1-Tris-(aminomethyl)ethane (TAME), nitrilotrimethylphosphonic acid (NTP), 2-BAPEN, 2,2',2",2"'-(1 ,4,8, 11 - tetraazacyclotetradecane- 1 ,4,8, 11 -tetrayl)tetraacetic acid, tripodal tris(hydroxypyridinone) chelator, H3CP256 and its bifunctional maleimide derivative, H3YM103 (YM103), NTP(PRHP)3, H2dedpa, citrate and derivatives thereof. In a preferred embodiment, the chelator is HBED or a derivative thereof such as HBED-CC.

[0133] The chelate-functionalized targeting agent can comprise as a targeting moiety a peptide, for example, a peptide comprising 2 to 20 amino acids, a urea-based peptidomimetic, a polypeptide, a protein, a vitamin, a saccharide, for example a monosaccharide or a polysaccharide, an antibody, nucleic acid, an aptamer, an antisense oligonucleotide, or an organic molecule. In a preferred embodiment, said targeting agent is urea-based peptidomimetic Glu-urea-Lys.

[0134] Chelate-functionalized targeting agent as described herein preferably have a capacity of biological targeting. Non-limiting examples of suitable targeting agents include molecules that target PSMA validated in prostate cancer, CAIX (carbonic anhydrase IX), a scientifically validated target in cell renal cell carcinoma (ccRCC), large amino acid transporter LAT 1 and LAT2 receptors validated targets that are highly expressed in several solid tumours, including malignancies of the central nervous system (CNS), cluster of differentiation 66 (CD66) for bone marrow conditioning, PDGFRa7 validated in soft tissue sarcoma (STS), VEGF receptors, analogues of bombesin or GRP receptor targeting molecules, molecules targeting somatostatin receptors, RGD peptides or molecules targeting av[33 and av[35, annexin V or molecules targeting the apoptotic process, molecules targeting oestrogen receptors, biomolecules targeting the plaque etc.. More generally, a list of targeting molecules, organic or not, functionalized by a chelating can be found in the journal of Velikyan et al., Theranostic 2014, Vol. 4, Issue 1 "Prospective of 68Ga- Radiopharmaceutical Development."

[0135] In a particularly preferred embodiment, said chelate-functionalized targeting agent can be an urea-based (di)peptide or peptidomimetic, in one example, said chelate-functionalized targeting agent is PSMA-11 (HBED-CC functionalised Glu-urea-Lys), e.g. Glu-urea-Lys-HBED-CC known as Gozetotide.

[0136] The term “radioactive metal” as used herein for radioactive labelling of the functionalised targeting agent(s) encompasses all radioactive metal ions suitable for use in medical imaging or radiotherapy and compatible with the HBED or HBED-CC and derivatives thereof. The radioactive metal typically is zirconium based radioisotope or radionuclide such as: zirconium-89. These radionuclides can be issued from nuclear reactor sub-products, cyclotron or from their specific radionuclide generator. They can be produced and supplied in a chelated or complexed form, such as in the form of an oxalate or complexed with EDTA. Typically prior to use in radiolabelling, the zirconium is converted into a halide form, more particularly a chloride form.

[0137] One of the primary methods for producing zirconium-89 is through cyclotron irradiation. The process involves bombarding a target, typically natural yttrium (Y-89), with protons in a cyclotron, which induces a nuclear reaction that transforms Y-89 into Zr-89. After irradiation, the Zr-89 is chemically separated from the yttrium target to produce high purity and specific activity suitable for medical use.

[0138] After addition of the radioactive metal solution to the mixture of chelate-functionalized targeting agent and buffer, optionally further comprising a stabilizer and / or a metal inhibitor, the solution obtained is left to the radiolabelling reaction for a short period of time, in particular between about 2 minutes and about 60 minutes, preferably from about 2 minutes to about 30 minutes, for example about 2 to 5, 2 to 10, or 2 to 15 minutes at room temperature.

[0139] The invention also discloses a radiolabelled targeting agent with radioactive metal, i.e. zirconium- 89, obtained by a method as described herein. Hence, the invention provides in [zirconium-89]- labelled PSMA-11 , obtained by a method as described herein.

[0140] The invention also provides for a radiolabelling kit, suitable for producing a zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent with an activity of up to 50.0 mCi, comprising:

[0141] - an HBED or HBED-CC functionalized targeting agent, preferably in lyophilized form;

[0142] - a buffering agent or buffer solution, allowing to maintain the pH in the range of 3.5 to 7, such as 3.5 to 6.5 or 4.0 to 5,5, more preferably 4,5 to 5.0, optionally in lyophilized form;

[0143] - optionally, an anion exchange means suitable for converting zirconium-89-oxalate to zirconium-89-halide, preferably zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4);

[0144] - optionally, a purification means suitable for eliminating unbound zirconium-89 from the radiolabelling mixture after the radiolabelling reaction took place.

[0145] The kit can additionally comprise a metal inhibitor as defined herein. Said kit can in some embodiments be a high activity radiolabelling kit, suitable for producing a zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent with an activity of at least 50.0 mCi, comprising:

[0146] - an HBED or HBED-CC functionalized targeting agent, preferably in lyophilized form;

[0147] - a buffering agent or buffer solution, allowing to maintain the pH in the range of 3.5 to 7, such as 3.5 to 6.5 or 4.0 to 5.5, more preferably 4.5 to 5.0, optionally in lyophilized form;

[0148] - a stabiliser selected from the group consisting of: ascorbic acid, sodium ascorbate, dehydroascorbic acid, gentisic acid, cysteine and methionine, or a salt thereof, preferably as a solution;

[0149] - optionally, an anion exchange means suitable for converting zirconium-89-oxalate to zirconium-89-halide, preferably zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4);

[0150] - optionally, a purification means suitable for eliminating unbound zirconium-89 from the radiolabelling mixture after the radiolabelling reaction took place.

[0151] The buffering agent or buffer solution in said kit can be a phosphate, nitrate, HEPES, acetate, formate, TRIS, ascorbate, and citrate or a mixture thereof, preferably an acetate or formate buffer, more preferably a sodium acetate or sodium formate buffer, most preferably a sodium acetate buffer.

[0152] The methods and kits described herein typically make use of ion exchange, more particular of anion exchange to convert the chelated or complexed zirconium-89 (e.g. in the oxalate-form) to its more reactive halide (such as chloride) form. Several anion exchange means and substrates may be used to this end. One particularly interesting substrate is a HCO3‘ resin, column or cartridge. In a preferred embodiment, said cartridge is first washed with de-ionized (Milli Q) water to remove the oxalate and contaminants and then eluted with higher or equal to 1 M HCI to elute the zirconium-chloride form. Typical, but non-limiting examples of such resins are: SepPak QMA anion exchange cartridge, AG 1-X8 Resin (250 mg, 200-400 dry mesh, 45-106 pm wet bead, BioRad, USA), Oasis® WAX (30mg, Waters, USA) and Chromafix® PS-HCO3 column (45 mg, Machery-Nagel, Duren, Germany). It will be clear for a person of skill in the art that equivalent resins can also be used.

[0153] In an alternative embodiment, the Zirconium-chloride [89Zr]ZrCI4is produced by a cyclotron.

[0154] In order to further increase the radiochemical purity of the end-product (the zirconium-89 radiolabelled targeting agent), the methods and kits described herein can further encompass or comprise a step or means for purifying the targeting agent. Exemplary means are solid phase extraction or reverse phase extraction means known in the art. Typically, SPE substrates are made of high-purity silica. For small peptides such as the PSMA-11 targeting agent, preferably a silica-based substrate with strong hydrophobicity such as C18-coated silica can be used, which is a hydrophobic and non-polar stationary phase, selectively retaining non-polar and moderately polar compounds such as lipids, steroids, and pesticides while allowing more polar compounds such as sugars and amino acids to pass through. For larger molecules such as polypeptides and antibodies, different hydrophobic interaction resins with C4-, C6-, C8-, phenyl-, or benzyl- coated silica can be used as a substrate depending on the chemical character of the targeting agent.

[0155] In the kit according to the invention, the HBED or HBED-CC-functionalized targeting agent and the buffer can be provided in an lyophilized form, either in two separate vials or in one single vial. In one embodiment, also the metal inhibitor and / or stabilising agent are present in lyophilised form, either in a separate vial, or combined with either the buffer or targeting agent. Said vials can be siliconized or can be acid-washed prior to use.

[0156] In a specific embodiment, the radiolabelling kit can comprise the following components:

[0157] Sterile Vial 1 comprising the chelate-functionalized targeting agent and the metal inhibitor, preferably in a sterile (e.g.10 mL) vial;

[0158] Sterile Vial 2 that comprises buffering agent, preferably in a sterile (e.g.10 mL) vial;

[0159] Optionally vial 3 comprising an aqueous solution of radiolysis stabiliser selected from ascorbic acid, dehydroascorbic acid, gentisic acid, cysteine and methionine, sodium ascorbate, or a salt thereof.

[0160] More preferably, said kit comprises:

[0161] Sterile Vial 1 , which comprises Gozetotide (PSMA-11) and D-mannose as a lyophilized powder in a sterile vial 10 mL vial, more preferably in an amount of 20 to 30 pg, most preferably of about 25 pg of Gozetotide and from 5 to 15 pg, more preferably about 10 pg of D-mannose; and / or

[0162] Sterile Vial 2, which comprises from 100 to 200 mg, preferably about 150 mg anhydrous sodium acetate in from 0,150 to 0,350 M HCI solution (2.5 mL volume to 6.4 mL volume); and / or Optionally sterile Vial 3, which comprises from 200 to 300 mg / mL, preferably about 250mg / mL of ascorbic acid solution, preferably in a glass vial in order to maintain shelflife of the stabiliser.

[0163] For use some cyclotrons (liquid or solid target) as source of gallium, or with IRE GalliEo type gallium generators (TiO2-based), Vial 2 typically requires a lower molarity of the HCL solution, such as from 0,150 to 0,200 M HCI, more preferably from 0,170 to 0,180 M HCI, most preferably of about 0,175 M HCI.

[0164] In some embodiments of the kits described herein, the amount of ascorbic acid, dehydroascorbic acid or a salt thereof in the kit, is so that the weight ratio of the weight of stabiliser over the weight of the buffering agent in the kit, is at least 0.006, preferably at least 0.010, preferably at least 0.030, preferably at least 0.050, preferably at least 0.100, preferably at least 0.150, preferably at least 0.200, preferably at least 0.250, preferably at least 0.300. In a preferred embodiment, said stabiliser is ascorbic acid.

[0165] In some embodiments of the kits described herein, the amount of stabiliser in the kit, is such that the weight ratio of the weight of stabiliser over the weight of the metal inhibitor in the kit, is weight of stabiliser over the weight of the metal inhibitor in the mixture obtained in step e), is at least 100, preferably at least 200, preferably at least 500, preferably at least 1000, preferably at least 2500, preferably at least 5000, preferably at least 10000, preferably at least 20000, preferably at least 50000. In a preferred embodiment, said stabiliser is ascorbic acid.

[0166] In some embodiments of the methods described herein, the amount of stabiliser in the mixture obtained in step e), is so that the weight ratio of the weight of stabiliser over the weight of the buffering agent in the mixture obtained in step e), is at least 0.006, preferably at least 0.010, preferably at least 0.030, preferably at least 0.050, preferably at least 0.100, preferably at least 0.150, preferably at least 0.200, preferably at least 0.250, preferably at least 0.300.

[0167] In some embodiments of the methods described herein, the amount of stabiliser in the mixture obtained in step e), is so that the weight ratio of the weight of stabiliser over the weight of the metal inhibitor in the mixture obtained in step e), is at least 100, preferably at least 200, preferably at least 500, preferably at least 1000, preferably at least 2500, preferably at least 5000, preferably at least 10000, preferably at least 20000, preferably at least 50000.

[0168] In some embodiments of the use described herein, the amount of stabiliser in the radiolabelled chelate-functionalized targeting agent composition, is so that the weight ratio of the weight of stabiliser over the weight of the buffering agent in the radiolabelled chelate-functionalized targeting agent composition, is at least 0.006, preferably at least 0.010, preferably at least 0.030, preferably at least 0.050, preferably at least 0.100, preferably at least 0.150, preferably at least 0.200, preferably at least 0.250, preferably at least 0.300. In a preferred embodiment, said stabiliser is ascorbic acid.

[0169] In some embodiments of the use described herein, the amount of stabiliser in the radiolabelled chelate-functionalized targeting agent composition, is so that the weight ratio of the equivalent weight of stabiliser over the weight of the metal inhibitor in the radiolabelled chelate-functionalized targeting agent composition, is at least 100, preferably at least 200, preferably at least 500, preferably at least 1000, preferably at least 2500, preferably at least 5000, preferably at least 10000, preferably at least 20000, preferably at least 50000. In a preferred embodiment, said stabiliser is ascorbic acid.

[0170] Zirconium-89 is interesting because of its longer half-life, suitability for labelling larger molecules, and high-quality imaging capabilities. Due to its slightly lower positron energy, it can also result in higher resolution imaging. It can be used in imaging for tracking slow biological processes, particularly in oncology, in real-time.

[0171] Due to the longer half-life, imaging can be postponed for a couple of hours to allow washing out (clearance) of non-target bound radiochemicals, thereby increasing the resolution and making it possible to detect smaller target sites even more precisely and clear.

[0172] Zirconium-based radiotracers can for example be used in oncology, cardiology, and neurology.

[0173] In oncology for example, targeted imaging of Zr-89 labelled radiotracers can lead to targeting specific cancer cells, enabling the visualisation of tumours and metastases with high specificity. In addition, Zr-89 labelled radiotracers can be used for monitoring treatment response and efficacy of therapies, e.g. by tracking the biodistribution of targeting agents in cancer treatment. Due to the longer half-life, Zr-89 labelled radiotracers can be used in longer observation windows, allowing for prolonged imaging periods, suitable for slow biological processes like tumour accumulation and ligand-targeting. One example is its use in breast cancer patients, where Zr-89 labelled trastuzumab was used to detect HER2-positive metastases that were not easily detectable using other radioimaging methods. Also, Zr-89 labelled rituximab was used to evaluate its distribution in patients, providing information on the biodistribution and tumour uptake of rituximab, aiding in personalised treatment. In cardiology for example, Zr-89 labelled radiotracers can be used to detect areas of inflammation in cardiovascular diseases, in imaging of atherosclerotic plaques, potentially helping in assessing the risk of cardiac events.

[0174] In neurology, Zr-89 labelled radiotracers can be used for brain tumour imaging, especially in assessing the efficacy of novel therapies. Additionally, Zr-89 labelled radiotracers can be used to study neuroinflammatory processes in conditions like Alzheimer’s disease.

[0175] In addition to imaging, zirconium-89-labelled compounds can be used in targeted radiotherapy, by delivering a therapeutic dose of radiation directly to cancer cells while minimising damage to healthy tissues. This is currently being tested in breast cancer treatment, such as HER2 positive cancers. By attaching Zr-89 to antibodies targeting these markers, a targeted radiation dose can be delivered to the cancer cells. Also in prostate cancer, studies are trying to target prostatespecific membrane antigen (PSMA), to deliver targeted radiotherapy to prostate cancer cells.

[0176] Zr-89 labelled radiotracers are being developed for combined diagnostics and therapeutics (Theranostics). Due to its dual functionality, i.e. being able to be used for PET imaging and as a potential targeted therapeutic make Zr-89 labelled radiotracers ideal candidates for developing theranostic agents. Zr-89 labelled theranostics can be used to diagnose and assess the extent of the disease and deliver targeted therapy to the same sites, allowing for more personalised and effective treatment plans.

[0177] EXAMPLES

[0178] The following I lluccix® PSMA-11 radiolabelling cold kits (Configuration A or B) from Telix Pharmaceuticals were used in the examples below: llluccix® PSMA-11 radiolabelling kit Configuration A:

[0179] The kit comprises 2 vials, wherein: the first vial comprises 25.0 pg PSMA-11 , and 10.0 pg D-mannose as a lyophilised powder in a sterile 10 ml vial; the second vial comprises 150.0 mg anhydrous sodium acetate in 2.5 ml 0.292 M HCI solution in a sterile 10 ml vial;

[0180] Optionally, there may be a the third vial, a sterile vacuumed vial, to collect and store the [89Zr]ZrCI4solution, to be used in the labelling method. llluccix® PSMA-11 radiolabelling kit Configuration B:

[0181] The kit comprises 2 vials, wherein: the first vial comprises 25.0 pg PSMA-11 , and 10.0 pg D-mannose as a lyophilised powder in a sterile 10 ml vial; the second vial comprises 150.0 mg anhydrous sodium acetate in 6.4 ml 0.175 M HCI solution in a sterile 10 ml vial;

[0182] Optionally, there may be a the third vial, a sterile vacuumed vial, to collect and store the [89Zr]ZrCI4solution, to be used in the labelling method.

[0183] Preparation of the [89Zr]ZrCI4solution (conversion)

[0184] 213 MBq [89Zr]Zr-oxalate was obtained from Cyclotron B.V. (Amsterdam, The Netherlands), as a 0.186 ml aqueous solution. The [89Zr]Zr-oxalate solution was directly eluted onto a Chromafix 30PS-HCO3‘ cartridge (45 mg, Synthra, Hamburg, Germany) and the cartridge was rinsed with 10 mL Milli Q water to wash away the oxalate and any contaminations. The [89Zr]ZrCI4was then eluted from the cartridge with 0.5 mL 1 M HCI, which was collected in a vial in a lead-shielded pot, resulting in 201 .4 MBq [89Zr]ZrCI4in 5 ml. Said solution was further diluted with Milli Q water to 0.1 M HCI (equals to a total volume of 20 mL) before the labelling experiments (Examples 1 to 6) were performed. Hence, the [89Zr]ZrCI4solution used in Examples 1-6 has an radioactive concentration of 10.07 MBq / mL.

[0185] Examples 1-7

[0186] For each Example, the llluccix® PSMA-11 radiolabelling kit was provided in one of the configurations A or B. For each Example, the content of the second vial was added to the content of the first vial, yielding a PSMA-11 solution with a pH between 4.5 and 5.0. The vial was placed in a lead casing. If needed, the pH can be adjusted by adding extra buffer or by other pH adjusting means in case after elution the pH would be clearly outside said range.

[0187] Various amounts of the above prepared [89Zr]ZrCI4solution were added to the vial, as indicated in Table 1 , below. The vial was closed with a screw cap, and the content was briefly shaken. The vial was placed back in the lead casing at room temperature, or in a heating block at 90°C (cf. Table 1) and the radiolabelling reaction allowed to proceed for 30 minutes (for Examples 1-6) or 60 min (for Example 7).

[0188] Then, samples of 50 pL were taken for HPLC analysis. Every sample was pretreated by the addition of 2 pL of a 4 mM solution of diethylenetriaminepentaacetic acid (DTPA). HPLC measurements were performed by injection of 100-300 kBq per injection. The HPLC method was as follows:

[0189] ACE Equivalence 3 C18 150 x 3.0 mm column (Avantor) and a gradient profile of 0-8 min from 0% towards 75% B, and 8-12 min at 75% solvent B, at a flow rate of 0.8 mL / min. Solvent A was water + 0.1 % trifluoracetic acid (TFA) (v / v), and solvent B was acetonitrile (MeCN). Detection was done either by UV absorption at 254 nm, or radioactivity measured by a POMO radio detector (Elisia Raytest). Free zirconium captured with DTPA elutes at the injection peak.

[0190] The final reaction mixture was subsequently eluted over a Sep-Pak C18 cartridge (Sep-Pak C18 Plus Short Cartridge, 360 mg Sorbent per Cartridge, 55 - 105 pm (Waters)). Which was then eluted with 1 ml methanol.

[0191] Finally, the total eluted activity was determined by measuring the activity in the vial by a dose calibrator to determine the radiolabelling yield compared to the initial amount of activity added to the vial. From the above experiments it follows that the llluccix ® kit of configuration A, used at room temperature performs best in these reaction conditions, resulting in a radiolabelling yield of 81 %. This opens the possibility of using the existing llluccix ® kit, designed for gallium-68 radiolabelling of HBED-linked targeting agents exemplified here with PSMA-11.

[0192] Example 8: Stability test

[0193] The 1 ml [89Zr]-labelled PSMA-11 solution in methanol, obtained in Example 7, is further diluted with water to a total volume of 8 ml. this is split in two solutions of 4 ml each. To the first solution, 1 ml MilliQ water is added. To the second solution, 1 ml MilliQ water is added, which comprises 100 mg / mL of ascorbic acid. Both solutions are stored for 3 days. At several timepoints, the radioactive purity is analysed by the HPLC method described above.

Claims

CLAIMS1. A method for radiolabelling an HBED or HBED-CC functionalized targeting agent with a metal radionuclide being zirconium-89, comprising the steps of: a) providing an HBED or HBED-CC functionalized targeting agent; b) providing a buffering agent or buffer solution, allowing to maintain the pH in the range of 3.5 to 7, such as 3.5 to 6.5 or 4.0 to 5.5, more preferably 4.5 to 5.0; c) providing a suitable amount of zirconium-89-halide, preferably a suitable amount of zirconium-89-chloride; preferably obtained by converting a suitable amount of zirconium- 89-oxalate to zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4) by means of anion- exchange, or sourced from cyclotron production; d) eluting a suitable amount of zirconium-89-halide obtained in step c) into a mixture of a) and b) and allowing the radiolabelling reaction to take place, preferably at room temperature; e) optionally, purifying the radiolabelling mixture by eliminating unbound zirconium-89 from the radiolabelling mixture, thereby obtaining a zirconium-89 radiolabelled targeting agent.

2. The method according to claim 1 , wherein said buffering agent or buffer solution is a phosphate, nitrate, HEPES, acetate, formate, TRIS, ascorbate, and citrate or a mixture thereof, preferably an acetate or formate buffer, more preferably a sodium acetate or sodium formate buffer, most preferably a sodium acetate buffer.

3. The method according to claim 1 or 2, wherein the anion exchange step is performed on an anion exchange column, such as a HCO3‘ resin, column or cartridge, or a SepPak QMA anion exchange cartridge. In a preferred embodiment, said cartridge is first washed with de-ionized (Milli Q) water to remove the oxalate and contaminants and then eluted with a HCI to elute the zirconium-chloride form, preferably about 1 M HCI or more.

4. The method according to any one of claims 1 to 3, wherein the purification step e) is done by means of solid phase extraction or reverse phase extraction, preferably over a silica-based substrate with strong hydrophobicity such as C18-coated silica, C8-coated silica, C6-coated silica or C4-coated silica, depending on the targeting ligand to be labelled.

5. The method according to any one of previous claims, wherein additionally a metal inhibitor is added to the radiolabelling mixture, said metal inhibitor being a co-chelating agent, capable ofinactivating metals other than radioactive metal without interfering with the chelation between the radioactive metal and the said chelate-functionalized targeting agent, under the conditions of the labelling reaction.

6. The method according to any one of previous claims, wherein said metal inhibitor is a sugar, preferably a short-chain sugar or oligosaccharide, such as comprising up to 7 monosaccharide units.

7. The method according to any one of previous claims, wherein said metal inhibitor is selected from the group comprising: monosaccharides and their derivatives, disaccharides and their derivatives, and cyclodextrins.

8. The method according to any one of previous claims, wherein said metal inhibitor is selected from the group comprising: Glucose, D-Fructose, Beta-cyclodextrin, D-Mannose, and beta-cyclodextrin, more preferably D-mannose.

9. The method according to any one of previous claims, wherein said metal inhibitor and said functionalised agent are not chemically linked.

10. The method according to any one of previous claims, wherein said metal inhibitor and said functionalised agent are chemically linked, through a linker that is unstable in the radiolabelling conditions.11 . The method according to any one of previous claims, wherein the chelate-functionalized targeting agent comprises a targeting moiety selected from the list comprising a peptide, a urea- based peptidomimetic, a polypeptide, a protein, a vitamin, a saccharide, an antibody, a nucleic acid, an aptamer, an antisense oligonucleotide, or an organic molecule.

12. The method according to any one of previous claims, wherein said chelate-functionalized targeting agent is Glu-urea-Lys-HBED-CC (gozetotide or PSMA-11).

13. The method according to any one of previous claims, wherein the radiolabelling reaction is carried out at ambient or room temperature (such as between 15 and 30°C).

14. The method according to any one of the previous claims, wherein the activity is up to 100 mCi, preferably from 3 mCi to 100 mCi.

15. The method according to any one of claims 1 to 14, wherein additionally a stabiliser is added to the radiolabelling mixture that prevents radiolysis (product degradation) of the chelate- functionalized targeting agent, wherein said stabiliser is selected from the group consisting of:ascorbic acid, dehydroascorbic acid, gentisic acid, cysteine, N-acetylcysteine, and methionine, or a salt thereof, preferably as a solution to the radiolabelling mixture prior to radiolabelling;16. The method according to claim 15, producing a zirconium-89 radiolabelled targeting agent with an activity of at least 50.0 mCi, preferably at least 60.0 mCi, preferably at least 70.0 mCi, preferably at least 80.0 mCi, preferably at least 90.0 mCi, preferably at least 100.0 mCi, preferably at least 150.0 mCi, preferably at least 200.0 mCi, preferably at least 250.0 mCi, preferably at least 300.0 mCi, preferably at least 350.0 mCi, preferably at least 400.0 mCi, preferably at least 450.0 mCi, preferably at least 500.0 mCi, preferably at least 750.0 mCi, preferably at least 1000.0 mCi.

17. The method according to claim 15 or 16, wherein the zirconium-89 is provided as a solution with an radioactive concentration of at least 5.0 mCi / ml, preferably at least 6.0 mCi / ml, preferably at least 7.0 mCi / ml, preferably at least 10.0 mCi / ml, preferably at least 15.0 mCi / ml, preferably at least 20.0 mCi / ml, preferably at least 25.0 mCi / ml, preferably at least 30.0 mCi / ml, preferably at least 35.0 mCi / ml, preferably at least 40.0 mCi / ml, preferably at least 45.0 mCi / ml, preferably at least 50.0 mCi / ml, preferably at least 75.0 mCi / ml, preferably at least 100.0 mCi / ml.

18. The method according to any one of claims 15 to 17, wherein the mixture radiolabelling mixture comprises at least 10.0 mg, preferably at least 15.0 mg, preferably at least 20.0 mg, preferably at least 25.0 mg, preferably at least 30.0 mg of stabiliser, more preferably of ascorbic acid.

19. The method according to any one of claims 15 to 18, wherein the radiolabelling mixture comprises at most 100.0 mg, preferably at most 90.0 mg, preferably at most 80.0 mg, preferably at most 70.0 mg, preferably at most 60.0 mg, preferably at most 50.0 mg of stabiliser, more preferably of ascorbic acid.

20. The method according to any one of claims 15 to 19, wherein the radiolabelling mixture comprises at least 1.0 mg, preferably at least 1.2 mg, preferably at least 1.4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of stabiliser per 50 mCi metal radionuclide in step f); more preferably wherein the radiolabelling mixture comprises of at least 1.0 mg, preferably at least 1.2 mg, preferably at least 1.4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of ascorbic acid per 50 mCi zirconium-89 in step f).

21. The method according to any one of claims 15 to 20, wherein the radiolabelling mixture comprises at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of stabiliser per 50 mCi metal radionuclide in step f); more preferably wherein the radiolabelling mixture comprises of at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of ascorbic acid per 50 mCi zirconium-89 in step f).

22. The method according to any one of claims 15 to 21 , wherein the amount of stabiliser in the radiolabelling mixture, is such that the weight ratio of the weight of stabiliser over the weight of the chelate-functionalized targeting agent in the radiolabelling mixture, is at least 40, preferably at least 80, preferably at least 150, preferably at least 300, preferably at least 600, preferably at least 1000, preferably at least 1500, preferably at least 2000.

23. The method according to any one of claims 15 to 22, wherein the solution of stabiliser has a concentration of at least 0.14 mg / ml, preferably at least 0.30 mg / ml, preferably at least 0.50 mg / ml, preferably at least 1.00 mg / ml, preferably at least 2.00 mg / ml, preferably at least 4.00 mg / ml, preferably at least 5.00 mg / ml, preferably at least 7.00 mg / ml, preferably at least 9.00 mg / ml, preferably at least 10.00 mg / ml, preferably wherein the stabilizer is ascorbic acid.

24. The method according to any one of claims 15 to 23, wherein the stabiliser is ascorbic acid, dehydroascorbic acid or a salt thereof, and optionally wherein said stabiliser is provided together with a bisulfite and or a metabisulfite.

25. The method according to any one of claims 15 to 24, for providing at least 3.0, preferably at least 4.0, preferably at least 5.0, preferably at least 6.0, preferably at least 8.0, preferably at least 10.0, preferably at least 12.0, preferably at least 15.0, preferably at least 20.0 patient doses.

26. A zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent obtained by the method according to anyone of claims 1 to 25.

27. The radiolabelled targeting agent according to claim 26, which is zirconium-89 radiolabelled Glu-urea-Lys-HBED-CC (PSMA-11 or gozetotide).

28. A radiolabelling kit, preferably suitable for producing a zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent with an activity of up to 50.0 mCi, comprising:- an HBED or HBED-CC functionalized targeting agent, preferably in lyophilized form;- a buffering agent or buffer solution, allowing to maintain the pH in the range of 3.5 to 7, such as 3.5 to 6.5 or 4.0 to 5,5, more preferably 4,5 to 5.0, optionally in lyophilized form;- optionally, an anion exchange means suitable for converting zirconium-89-oxalate to zirconium-89-halide, preferably zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4);- optionally, a purification means suitable for eliminating unbound zirconium-89 from the radiolabelling mixture after the radiolabelling reaction took place.

29. The kit according to claim 28, additionally comprising a metal inhibitor, which is a cochelating agent, capable of inactivating metals other than radioactive metal without interfering with the chelation between the radioactive metal and the said chelate-functionalized targeting agent, under the conditions of the labelling reaction.

30. A high activity radiolabelling kit, preferably suitable for producing a zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent with an activity of at least 50.0 mCi, comprising:- an HBED or HBED-CC functionalized targeting agent, preferably in lyophilized form;- a buffering agent or buffer solution, allowing to maintain the pH in the range of 4.0 to 5.5, more preferably 4.5 to 5.0, optionally in lyophilized form;- a stabiliser selected from the group consisting of: ascorbic acid, sodium ascorbate, dehydroascorbic acid, gentisic acid, cysteine and methionine, or a salt thereof, preferably as a solution;- optionally, an anion exchange means suitable for converting zirconium-89-oxalate to zirconium-89-halide, preferably zirconium-89-chloride ([89Zr]Zr-Oxalate to [89Zr]ZrCI4);- optionally, a purification means suitable for eliminating unbound zirconium-89 from the radiolabelling mixture after the radiolabelling reaction took place.

31. The kit according to any one of claims 28 to 30, wherein said buffering agent or buffer solution is a phosphate, nitrate, HEPES, acetate, formate, TRIS, ascorbate, and citrate or a mixture thereof, preferably an acetate or formate buffer, more preferably a sodium acetate or sodium formate buffer, most preferably a sodium acetate buffer.

32. The kit according to any one of claims 28 to 31 , wherein said anion exchange means is a HCO3‘ resin, column or cartridge. In a preferred embodiment, said cartridge is first washed withde-ionized (Milli Q) water to remove the oxalate and contaminants and then eluted with 1 M HCI to elute the zirconium-chloride form.

33. The kit according to any one of claims 28 to 32, wherein the means is a solid phase extraction or reverse phase extraction means, preferably a silica-based substrate with strong hydrophobicity such as C18-coated silica, C8-coated silica, C6-coated silica or C4-coated silica, depending on the targeting ligand to be labelled.

34. The kit according to any one of claims 28 to 33, wherein the HBED or HBED-CC- functionalized targeting agent and the buffer are provided in an lyophilized form, either in two separate or in one single vial. In one embodiment, also the metal inhibitor and / or stabilising agent are present in lyophilised form, either in a separate vial, or combined with either the buffer or targeting agent.

35. The kit according to any one of claims 30 to 34, suitable for radiolabelling a chelate functionalized targeting agent with zirconium-89 thereby producing a zirconium-89 chelate functionalized targeting agent with an activity of at least 50.0 mCi, preferably at least 100.0 mCi preferably at least 150.0 mCi, preferably at least 200.0 mCi, preferably at least 250.0 mCi, preferably at least 300.0 mCi, preferably at least 350.0 mCi, preferably at least 400.0 mCi, preferably at least 450.0 mCi, preferably at least 500.0 mCi, preferably at least 750.0 mCi, preferably at least 1000.0 mCi.

36. The kit according to any one of claims 30 to 35, wherein the zirconium-89 is provided as a solution with an activity of at least 5.0 mCi / ml, preferably at least 7.0 mCi / ml, preferably at least10.0 mCi / ml, preferably at least 15.0 mCi / ml, preferably at least 20.0 mCi / ml, preferably at least25.0 mCi / ml, preferably at least 30.0 mCi / ml, preferably at least 35.0 mCi / ml, preferably at least40.0 mCi / ml, preferably at least 45.0 mCi / ml, preferably at least 50.0 mCi / ml, preferably at least75.0 mCi / ml, preferably at least 100.0 mCi / ml.

37. The kit according to any one of claims 30 to 36, comprising at least 10.0 mg, preferably at least 15.0 mg, preferably at least 20.0 mg, preferably at least 25.0 mg, preferably at least 30.0 mg of stabiliser, wherein said stabiliser preferably is ascorbic acid.

38. The kit according to any one of claims 30 to 37, wherein the kit comprises at most 100.0 mg, preferably at most 90.0 mg, preferably at most 80.0 mg, preferably at most 70.0 mg, preferably at most 60.0 mg, preferably at most 50.0 mg of stabiliser, more preferably of ascorbic acid.

39. The kit according to any one of claims 30 to 38, wherein the kit comprises at least 1 .0 mg, preferably at least 1.2 mg, preferably at least 1.4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of stabiliser per 50 mCi metal radionuclide; more preferably wherein the kit comprises of at least 1.0 mg, preferably at least 1.2 mg, preferably at least 1.4 mg, preferably at least 1.6 mg, preferably at least 1.8 mg, preferably at least 2.0 mg, preferably at least 2.2 mg, preferably at least 2.4 mg, preferably at least 2.5 mg of ascorbic acid per 50 mCi zirconium-89.

40. The kit according to any one of claims 30 to 39, wherein the kit comprises at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of stabiliser per 50 mCi metal radionuclide; more preferably wherein the kit comprises of at most 100.0 mg, preferably at most 80.0 mg, preferably at most 60.0 mg, preferably at most 40.0 mg, preferably at most 20.0 mg, preferably at most 15.0 mg, preferably at most 10.0 mg, preferably at least 7.5 mg, preferably at most 5.0 mg of ascorbic acid per 50 mCi zirconium-89.41 . The kit according to any one of claims 30 to 40, comprising at least 10.0 pg, preferably at least 15.0 pg, preferably at least 20.0 pg, preferably at least 25.0 pg of said chelate-functionalized targeting agent.

42. The kit according to any one of claims 30 to 41 , comprising at least 2.0 pg, preferably at least 4.0 pg, preferably at least 6.0 pg, preferably at least 8.0 pg, preferably at least 10.0 pg of said metal inhibitor.

43. The kit according to any one of claims 30 to 42, comprising at least 50.0 mg, preferably at least 75.0 mg, preferably at least 100.0 mg, preferably at least 125.0 mg, preferably at least 150.0 mg of said buffering agent.

44. The kit according to any one of claims 30 to 43, wherein the amount of stabiliser in the kit, is such that the weight ratio of the weight of stabiliser over the weight of the chelate-functionalized targeting agent in the kit, is at least 40, preferably at least 80, preferably at least 150, preferably at least 300, preferably at least 600, preferably at least 1000, preferably at least 1500, preferably at least 2000. In a preferred embodiment, said stabiliser is ascorbic acid.

45. The kit according to any one of claims 30 to 44, wherein the solution of stabiliser, has a concentration of at least 0.14 mg / ml, preferably at least 0.30 mg / ml, preferably at least 0.50 mg / ml, preferably at least 1.00 mg / ml, preferably at least 2.00 mg / ml, preferably at least 4.00 mg / ml, preferably at least 5.00 mg / ml, preferably at least 7.00 mg / ml, preferably at least 9.00 mg / ml, preferably at least 10.00 mg / ml. In a preferred embodiment, said stabiliser is ascorbic acid.

46. The kit according to any one of claims 30 to 45, wherein the stabiliser is provided in the kit together with a bisulfite and or a metabisulfite.

47. The kit according to any one of claims 30 to 46, wherein the kit is suitable for providing at least 3.0, preferably at least 4.0, preferably at least 5.0, preferably at least 6.0, preferably at least 8.0, preferably at least 10.0, preferably at least 12.0, preferably at least 15.0, preferably at least 20.0 patient doses.

48. The kit according to any one of claims 28 to 47, wherein said metal inhibitor is a sugar, preferably a short-chain sugar or oligosaccharide, such as comprising up to 7 monosaccharide units.

49. The kit according to anyone of claims 28 to 48, wherein said metal inhibitor is selected from the group comprising: monosaccharides and their derivatives, disaccharides and their derivatives, and polysaccharides and cyclodextrins.

50. The kit according to anyone of claims 28 to 49, wherein said metal inhibitor is selected from the group comprising: Glucose, D-Fructose, Beta-cyclodextrin, and D-Mannose.

51. The kit according to any one of claims 28 to 50, wherein said metal inhibitor and said functionalised agent are not chemically linked.

52. The kit according to any one of claims 28 to 51 , wherein said metal inhibitor and said functionalised agent are chemically linked, through a linker that is unstable in the radiolabelling conditions.

53. The kit according to any one of claims 28 to 52, wherein said targeting agent is Glu-urea- Lys-HBED-CC (gozetotide or PSMA-11).

54. An imaging method, comprising the steps of:1) radiolabelling an HBED or HBED-CC functionalized targeting agent with zirconium-89 according to the method of any one of claims 1 to 24, preferably by using a kit according to any one of claims 25 to 53,2) administering to a subject a diagnostic amount of said zirconium-89 radiolabelled targeting agent; and,3) detecting said zirconium-89 radiolabelled targeting agent when bound to its target using PET or PET / CT imaging methods.

55. A method of detecting a prostate tumour or cancer, comprising the steps of:1) radiolabelling Glu-urea-Lys-HBED-CC (PSMA-11 or gozetotide) with zirconium-89 according to the method of any one of claims 1 to 24, preferably by using a kit according to any one of claims 25 to 53,2) administering to a subject a diagnostic amount of zirconium-89 radiolabelled PSMA-11 (gozetotide); and,3) detecting binding of said zirconium-89 radiolabelled PSMA-11 (gozetotide) using PET or PET / CT imaging methods.

56. The method according to claim 55, wherein said detection is used for:(i) initial staging of prostate cancer into intermediate, unfavourable, high, or very high-risk prostate cancer,(ii) detecting suspected recurrence of prostate cancer and / or detection of metastasis,(iii) selection for radiotherapeutic treatment such as with Lutetium (177Lu) vipivotide tetraxetan (Pluvicto),(iv) monitoring prostate cancer for progression into Non-Metastatic or Metastatic Castration-Resistant Prostate Cancer (nmCRPC or mCRPC), or(v) determining response to (radio)therapy,(vi) real-time imaging or diagnosis in a subject e.g. for use in PET scans for tracking slow biological processes or progression of those over a period of time, or(vii) personalised medicine.

57. The method according to any one of claims 54 to 56, wherein said detection method is used to replace the need for taking a prostate biopsy or is used in PET or PET / CT scan with MRI in clinically significant or intermediate favourable prostate cancers, or in MRI for active surveillance of prostate cancer.

58. The zirconium-89 radiolabelled targeting molecule according to claim 25 or 26, for use in radiotherapy.

59. The zirconium-89 radiolabelled targeting molecule according to claim 25 or 26, for use in radiodiagnosis, preferably for use in in vivo real time detection of a targeting agent.

60. A system comprising a computer or processor configured to carry out the steps of the method of in vivo imaging or diagnosis according to any one of claims 54 to 57, preferably being able to receive real-time imaging data and presenting said imaging date in real-time to the user, thereby enabling in vivo imaging in or in vivo diagnosis of said subject.61 . A method for treatment of cancer, comprising the step of: - administering an efficient amount of the zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent according to claim 25 to a patient in need thereof. In some embodiments, said patient can have been diagnosed using the method of claim 54 to 57.

62. A method for treatment of cancer, comprising the step of: administering an efficient amount of the zirconium-89 radiolabelled HBED or HBED-CC functionalized targeting agent according to claim 26 to a patient in need thereof.