Emerging Radionuclides in a Regulatory Framework for Medicinal Products – How Do They Fit?

Recent years have seen the establishment of several radionuclides as medicinal products in particular in the setting of theranostics and PET. [177Lu]Lutetium Chloride or [64Cu]Copper Chloride have received marketing authorization as radionuclide precursor, [68Ga]Gallium Chloride has received regulatory approval in the form of different 68Ge/68Ga generators. This is a formal requirement by the EU directive 2001/83, even though for some of these radionuclide precursors no licensed kit is available that can be combined to obtain a final radiopharmaceuticals, as it is the case for Technetium-99m. In view of several highly promising, especially metallic radionuclides for theranostic applications in a wider sense, the strict regulatory environment poses the risk of slowing down development, in particular for radionuclide producers that want to provide innovative radionuclides for clinical research purposes, which is the basis for their further establishment. In this paper we address the regulatory framework for novel radionuclides within the EU, the current challenges in particular related to clinical translation and potential options to support translational development within Europe and worldwide

Radiopharmaceutical Precursors for Theranostics

Due to the complex nomenclature used in various regulations and guidance documents, the understanding of radiopharmaceutical precursor’s definition might be challenging. Depending on the context it could be interpreted as the substance which becomes a radiopharmaceutical after radiolabeling with a radionuclide of choice or a radionuclide which is used for radiolabeling of that substance. In this Chapter we present and discuss the requirements for precursors which are used in the preparation of theranostic radiopharmaceuticals, in particular for preparation of new radiopharmaceuticals for clinical trials within the EU. In discussion on the available methods for assessing the quality of radiopharmaceutical precursors and on the specified limits the reference to Ph. Eur. is made. Since the EANM guidelines for in-house preparation of radiopharmaceuticals also specify the need for testing the quality of radiopharmaceutical precursors, information provided herein might help the radiopharmacist working on the development of new theranostic agents to adequately define identity, strength, quality, purity and stability of the final radiopharmaceutical preparation

Influence of a novel, versatile bifunctional chelator on theranostic properties of a minigastrin analogue.

BACKGROUND: 6-[Bis(carboxymethyl)amino]-1,4-bis(carboxymethyl)-6-methyl-1,4-diazepane (AAZTA ) is a promising chelator with potential advantages over 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) for radiopharmaceutical applications. Its mesocyclic structure enables fast radiolabelling under mild conditions with trivalent metals including not only (68)Ga for positron emission tomography (PET) but also (177)Lu and (111)In for single-photon emission computed tomography (SPECT) and radionuclide therapy. Here, we describe the evaluation of a bifunctional AAZTA derivative conjugated to a model minigastrin derivative as a potential theranostic agent. METHODS: An AAZTA derivative with an aliphatic C9 chain as linker was coupled to a minigastrin, namely [AAZTA(0), D-Glu(1), desGlu(2-6)]-minigastrin (AAZTA-MG), and labelled with (68)Ga, (177)Lu and (111)In. The characterisation in vitro included stability studies in different media and determination of logD (octanol/PBS). Affinity determination (IC50) and cell uptake studies were performed in A431-CCK2R cells expressing the human CCK2 receptor. μPET/CT and ex vivo biodistribution studies were performed in CCK2 tumour xenograft-bearing nude mice and normal mice. RESULTS: AAZTA-MG showed high radiochemical yields for (68)Ga (>95 %), (177)Lu (>98 %) and (111)In (>98 %). The logD value of -3.7 for both [(68)Ga]- and [(177)Lu]-AAZTA-MG indicates a highly hydrophilic character. Stability tests showed overall high stability in solution with some degradation in human plasma for [(68)Ga]- and transchelation towards DTPA for and [(177)Lu]-AAZTA-MG. An IC50 value of 10.0 nM was determined, which indicates a high affinity for the CCK2 receptor. Specific cell uptake after 60 min was >7.5 % for [(68)Ga]-AAZTA-MG and >9.5 % for [(177)Lu]-AAZTA-MG, comparable to other DOTA-MG-analogues. μPET/CT studies in CCK2 receptor tumour xenografted mice not only revealed high selective accumulation in A431-CCK2R positive tumours of (68)Ga-labelled AAZTA-MG (1.5 % ID/g in 1 h post injection) but also higher blood levels as corresponding DOTA-analogues. The (111)In-labelled peptide had a tumour uptake of 1.7 % ID/g. Biodistribution in normal mice with the [(177)Lu]-AAZTA-MG showed a considerable uptake in intestine (7.3 % ID/g) and liver (1.5 % ID/g). CONCLUSION: Overall, AAZTA showed interesting properties as bifunctional chelator for peptides providing mild radiolabelling conditions for both (68)Ga and trivalent metals having advantages over the currently used chelator DOTA. Studies are ongoing to further investigate in vivo targeting properties and stability issues and the influence of spacer length on biodistribution of AAZTA

Influence of a novel, versatile bifunctional chelator on theranostic properties of a minigastrin analogue

Background: 6-[Bis(carboxymethyl)amino]-1,4-bis(carboxymethyl)-6-methyl-1,4-diazepane (AAZTA ) is a promising chelator with potential advantages over 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) for radiopharmaceutical applications. Its mesocyclic structure enables fast radiolabelling under mild conditions with trivalent metals including not only 68Ga for positron emission tomography (PET) but also 177Lu and 111In for singlephoton emission computed tomography (SPECT) and radionuclide therapy. Here, we describe the evaluation of a bifunctional AAZTA derivative conjugated to a model minigastrin derivative as a potential theranostic agent. Methods: An AAZTA derivative with an aliphatic C9 chain as linker was coupled to a minigastrin, namely [AAZTA0, D-Glu1, desGlu2\u20136]-minigastrin (AAZTA-MG), and labelled with 68Ga, 177Lu and 111In. The characterisation in vitro included stability studies in different media and determination of logD (octanol/PBS). Affinity determination (IC50) and cell uptake studies were performed in A431-CCK2R cells expressing the human CCK2 receptor. \u3bcPET/CT and ex vivo biodistribution studies were performed in CCK2 tumour xenograft-bearing nude mice and normal mice. Results: AAZTA-MG showed high radiochemical yields for 68Ga (>95 %), 177Lu (>98 %) and 111In (>98 %). The logD value of 123.7 for both [68Ga]- and [177Lu]-AAZTA-MG indicates a highly hydrophilic character. Stability tests showed overall high stability in solution with some degradation in human plasma for [68Ga]- and transchelation towards DTPA for and [177Lu]-AAZTA-MG. An IC50 value of 10.0 nM was determined, which indicates a high affinity for the CCK2 receptor. Specific cell uptake after 60 min was >7.5 % for [68Ga]-AAZTA-MG and >9.5 % for [177Lu]-AAZTA-MG, comparable to other DOTA-MG-analogues. \u3bcPET/CT studies in CCK2 receptor tumour xenografted mice not only revealed high selective accumulation in A431-CCK2R positive tumours of 68Ga-labelled AAZTA-MG (1.5 % ID/g in 1 h post injection) but also higher blood levels as corresponding DOTA-analogues. The 111In-labelled peptide had a tumour uptake of 1.7 % ID/g. Biodistribution in normal mice with the [177Lu]-AAZTA-MG showed a considerable uptake in intestine (7.3 % ID/g) and liver (1.5 % ID/g). Conclusion: Overall, AAZTA showed interesting properties as bifunctional chelator for peptides providing mild radiolabelling conditions for both 68Ga and trivalent metals having advantages over the currently used chelator DOTA. Studies are ongoing to further investigate in vivo targeting properties and stability issues and the influence of spacer length on biodistribution of AAZTA

Guidance on current good radiopharmacy practice (cGRPP) for the small-scale preparation of radiopharmaceuticals

This guidance is meant as a guidance to Part B of the EANM “Guidelines on Good Radiopharmacy Practice (GRPP)” issued by the Radiopharmacy Committee of the EANM (see www.eanm.org), covering the small-scale “in house” preparation of radiopharmaceuticals which are not kit procedures. The aim is to provide more detailed and practice-oriented guidance to those who are involved in the small-scale preparation of, for example, PET, therapeutic or other radiopharmaceuticals which are not intended for commercial purposes or distribution

EANM guideline on the validation of analytical methods for radiopharmaceuticals

Contains fulltext : 219679.pdf (publisher's version ) (Open Access)BACKGROUND: To fulfil good manufacturing requirements, analytical methods for the analysis of pharmaceuticals for human and vetinary use must be validated. The International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) has published guidance documents on the requirements for such validation activities and these have been adopted by the European Medicines Agency, The U.S. Food and Drug Administration (FDA) and other regulatory bodies. These guidance documents do not, however, fully address all the specific tests required for the analysis of radiopharmaceuticals. This guideline attempts to rectify this shortcoming, by recommending approaches to validate such methods. RESULTS: Recommedations for the validation of analytical methods which are specific for radiopharmaceutials are presented in this guideline, along with two practical examples. CONCLUSIONS: In order to comply with good manufacturing practice, analytical methods for radiopharmaceuticals for human use should be validated

EANM guideline on the validation of analytical methods for radiopharmaceuticals

BACKGROUND: To fulfil good manufacturing requirements, analytical methods for the analysis of pharmaceuticals for human and vetinary use must be validated. The International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) has published guidance documents on the requirements for such validation activities and these have been adopted by the European Medicines Agency, The U.S. Food and Drug Administration (FDA) and other regulatory bodies. These guidance documents do not, however, fully address all the specific tests required for the analysis of radiopharmaceuticals. This guideline attempts to rectify this shortcoming, by recommending approaches to validate such methods.RESULTS: Recommedations for the validation of analytical methods which are specific for radiopharmaceutials are presented in this guideline, along with two practical examples.CONCLUSIONS: In order to comply with good manufacturing practice, analytical methods for radiopharmaceuticals for human use should be validated.</p

Selection of the first 99m^{99m} Tc-Labelled somatostatin receptor subtype 2 antagonist for clinical translation : preclinical assessment of two optimized candidates

Recently, radiolabelled antagonists targeting somatostatin receptors subtype 2 (SST2) in neuroendocrine neoplasms demonstrated certain superior properties over agonists. Within the ERA-PerMED project &ldquo;TECANT&rdquo; two 99mTc-Tetramine (N4)-derivatized SST2 antagonists (TECANT-1 and TECANT-2) were studied for the selection of the best candidate for clinical translation. Receptor-affinity, internalization and dissociation studies were performed in human embryonic kidney-293 (HEK293) cells transfected with the human SST2 (HEK-SST2). Log D, protein binding and stability in human serum were assessed. Biodistribution and SPECT/CT studies were carried out in nude mice bearing HEK-SST2 xenografts, together with dosimetric estimations from mouse-to-man. [99mTc]Tc-TECANT-1 showed higher hydrophilicity and lower protein binding than [99mTc]-TECANT-2, while stability was comparable. Both radiotracers revealed similar binding affinity, while [99mTc]Tc-TECANT-1 had higher cellular uptake (&gt;50%, at 2 h/37 &deg;C) and lower dissociation rate (&lt;30%, at 2 h/37 &deg;C). In vivo, [99mTc]Tc-TECANT-1 showed lower blood values, kidney and muscles uptake, whereas tumour uptake was comparable to [99mTc]Tc-TECANT-2. SPECT/CT imaging confirmed the biodistribution results, providing the best tumour-to-background image contrast for [99mTc]Tc-TECANT-1 at 4 h post-injection (p.i.). The estimated radiation dose amounted to approximately 6 &micro;Sv/MBq for both radiotracers. This preclinical study provided the basis of selection of [99mTc]Tc-TECANT-1 for clinical translation of the first 99mTc-based SST2 antagonist

From preclinical development to clinical application : kit formulation for radiolabelling the minigastrin analogue CP04 with In-111 for a first-in-human clinical trial

Introduction A variety of radiolabelled minigastrin analogues targeting the cholecystokinin 2 (CCK2) receptor were developed and compared in a concerted preclinical testing to select the most promising radiotracer for diagnosis and treatment of medullary thyroid carcinoma (MTC). DOTA-DGlu-DGlu-DGlu-DGlu-DGlu-DGlu-Ala-Tyr-Gly-Trp-Met-Asp-Phe-NH2 (CP04) after labelling with 111In displayed excellent characteristics, such as high stability, receptor affinity, specific and persistent tumour uptake and low kidney retention in animal models. Therefore, it was selected for further clinical evaluation within the ERA-NET project GRAN-T-MTC. Here we report on the development of a pharmaceutical freeze-dried formulation of the precursor CP04 for a first multi-centre clinical trial with 111In-CP04 in MTC patients. Materials and methods The kit formulation was optimised by adjustment of buffer, additives and radiolabelling conditions. Three clinical grade batches of a final kit formulation with two different amounts of peptide (10 or 50 μg) were prepared and radiolabelled with 111In. Quality control and stability assays of both the kits and the resulting radiolabelled compound were performed by HPLC analysis. Results Use of ascorbic acid buffer (pH 4.5) allowed freeze-drying of the kit formulation with satisfactory pellet-formation. Addition of methionine and gentisic acid as well as careful selection of radiolabelling temperature was required to avoid extensive oxidation of the Met11-residue. Trace metal contamination, in particular Zn, was found to be a major challenge during the pharmaceutical filling process in particular for the 10 μg formulation. The final formulations contained 10 or 50 μg CP04, 25 mg ascorbic acid, 0.5 mg gentisic acid and 5 mg l-methionine. The radiolabelling performed by incubation of 200-250 MBq 111InCl3 at 90°C for 15 min resulted in reproducible radiochemical purity (RCP) > 94%. Kit-stability was proven for > 6 months at + 5°C and at + 25°C. The radiolabelled product was stable for > 4 h at + 25°C. Conclusion A kit formulation to prepare 111In-CP04 for clinical application was developed, showing high stability of the kit as well as high RCP of the final product