Development and Validation of an Analytical HPLC Method to Assess Chemical and Radiochemical Purity of [68Ga]Ga-NODAGA-Exendin-4 Produced by a Fully Automated Method

Migliari, Silvia; Sammartano, Antonino; Boss, Marti; Gotthardt, Martin; Scarlattei, Maura; Baldari, Giorgio; Bonadonna, Riccardo C.; Ruffini, Livia

doi:10.3390/molecules27020543

Cited by 9 publications

(3 citation statements)

References 35 publications

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“…Molar activity (GBq/µmol) is an important parameter in PET imaging. When the biological target concentration is minimal, image quality and quantification can be improved by a high MA, as has been shown for GLP-R (Velikyan 2015 ; Velikyan et al 2017 , 2008 ; Migliari et al 2022 ; Eriksson et al 2014 ) for insulinoma imaging using ligands such as exendin-4. In particular, the presence of unlabelled peptide can reduce the concentration of radioactivity in the target tissue due to competition with the labeled peptide for the same receptor.…”

Section: Discussionmentioning

confidence: 99%

Fully automated radiolabeling of [68Ga]Ga-EMP100 targeting c-MET for PET-CT clinical imaging

Rusu,

Delion,

Pirot

et al. 2023

EJNMMI radiopharm. chem.

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Background c-MET is a transmembrane receptor involved in many biological processes and contributes to cell proliferation and migration during cancer invasion process. Its expression is measured by immunehistochemistry on tissue biopsy in clinic, although this technique has its limitations. PET-CT could allow in vivo mapping of lesions expressing c-MET, providing whole-body detection. A number of radiopharmaceuticals are under development for this purpose but are not yet in routine clinical use. EMP100 is a cyclic oligopeptide bound to a DOTA chelator, with nanomolar affinity for c-MET. The aim of this project was to develop an automated method for radiolabelling the radiopharmaceutical [68Ga]Ga-EMP100. Results The main results showed an optimal pH range between 3.25 and 3.75 for the complexation reaction and a stabilisation of the temperature at 90 °C, resulting in an almost complete incorporation of gallium-68 after 10 min of heating. In these experiments, 90 µg of EMP-100 peptide were initially used and then lower amounts (30, 50, 75 µg) were explored to determine the minimum required for sufficient synthesis yield. Radiolysis impurities were identified by radio-HPLC and ascorbic acid and ethanol were used to improve the purity of the compound. Three batches of [68Ga]Ga-EMP100 were then prepared according to the optimised parameters and all met the established specifications. Finally, the stability of [68Ga]Ga-EMP100 was assessed at room temperature over 3 h with satisfactory results in terms of appearance, pH, radiochemical purity and sterility. Conclusions For the automated synthesis of [68Ga]Ga-EMP100, the parameters of pH, temperature, precursor peptide content and the use of adjuvants for impurity management were efficiently optimised, resulting in the production of three compliant and stable batches according to the principles of good manufacturing practice. [68Ga]Ga-EMP100 was successfully synthesised and is now available for clinical development in PET-CT imaging.

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Section: Discussionmentioning

confidence: 99%

Fully automated radiolabeling of [68Ga]Ga-EMP100 targeting c-MET for PET-CT clinical imaging

Rusu,

Delion,

Pirot

et al. 2023

EJNMMI radiopharm. chem.

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“…For the development of a feasible UV-HPLC method to detect HEPES, its chemical and physical properties need to be considered, such as a UV/vis absorption wavelength that is below 200 nm, or 220 nm for [ 68 Ga]Ga-radiopharmaceuticals [ 21 , 22 , 23 , 24 , 25 , 26 ]. We started from a ‘‘mother’’ solution with the highest concentration (250 μg/mL) of HEPES, obtained by dissolving the analyte in phosphate buffer solution (PBS) with 10% ethanol and serial dilutions (100, 80, 60, 40, 20, 10, 5, 3 μg/mL).…”

Section: Methodsmentioning

confidence: 99%

“…We have synthesized PET radiopharmaceuticals such as [ 68 Ga]Ga-DOTATOC, [ 68 Ga]Ga-HBEDD-CC-PSMA [ 21 ]; [ 68 Ga]Ga-DOTATATE [ 22 ]; [ 68 Ga]Ga-DOTA-Pentixafor [ 23 , 24 ]; and [ 68 Ga]Ga-NODAGA-Exendin-4 [ 25 , 26 ] using the automated synthesis module Scintomics GRP TM (Scintomics GmbH, Fürstenfeldbruck, Germany) and tested them to assess all release criteria with a validated QCs system.…”

Section: Introductionmentioning

confidence: 99%

A Specific HPLC Method to Determine Residual HEPES in [68Ga]Ga-Radiopharmaceuticals: Development and Validation

et al. 2022

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Background: Nowadays, in Nuclear Medicine, clinically applied radiopharmaceuticals must meet quality release criteria such as high radiochemical purity and radiochemical yield. Many radiopharmaceuticals do not have marketing authorization and have no dedicated monograph within European Pharmacopeia (Ph. Eur.); therefore, general monographs on quality controls (QCs) have to be applied for clinical application. These criteria require standardization and validation in labeling and preparation, including quality controls measurements, according to well defined standard operation procedures. However, QC measurements are often based on detection techniques that are specific to a certain chromatographic system. Several radiosyntheses of [68Ga]Ga-radiopharmaceuticals are more efficient and robust when they are performed with 2-[4-(2-hydroxyethyl)piperazin-1-yl] ethanesulfonic acid (HEPES) buffer, which is considered as an impurity to be assessed in the QC procedure, prior to clinical use. Thus, Ph. Eur. has introduced a thin-layer chromatography (TLC) method to quantify the HEPES amount that is present in [68Ga]Ga-radiopharmaceuticals. However, this is only qualitative and has proven to be unreliable. Here we develop and validate a new high-performance liquid chromatography (UV-Radio-HPLC) method to quantify the residual amount of HEPES in 68Ga-based radiopharmaceuticals. Method: To validate the proposed UV-Radio-HPLC method, a stepwise approach was used, as defined in the guidance document that was adopted by the European Medicines Agency (CMP/ICH/381/95 2014). The assessed parameters are specificity, linearity, precision (repeatability), accuracy, and limit of quantification. A range of concentrations of HEPES (100, 80, 60, 40, 20, 10, 5, 3 μg/mL) were analyzed. Moreover, to test the validity and pertinence of our new HPLC method, we analyzed samples of [68Ga]Ga-DOTATOC; [68Ga]Ga-PSMA; [68Ga]Ga-DOTATATE; [68Ga]Ga-Pentixafor; and [68Ga]Ga-NODAGA-Exendin-4 from different batches that were prepared for clinical use. Results: In the assessed samples, HEPES could not be detected by the TLC method that was described in Ph. Eur. within 4 min incubation in an iodine-saturated chamber. Our developed HPLC method showed excellent linearity between 3 and 100 μg/mL for HEPES, with a correlation coefficient (R2) for calibration curves that was equal to 0.999, coefficients of variation (CV%) < 2%, and percent deviation value of bias from 100% to 5%, in accordance with acceptance criteria. The intra-day and inter-day precision of our method was statistically confirmed and the limit-of-quantification (LOQ) was 3 μg/mL, confirming the high sensitivity of the method. The amount of HEPES that was detected with our developed HPLC method in the tested [68Ga]Ga-radiopharmaceuticals resulted well below the Ph. Eur. limit, especially for [68Ga]Ga-NODAGA-Exendin-4. Conclusions: The TLC method that is described in Ph. Eur. to assess residual HEPES in [68Ga]-based radiopharmaceuticals may not be sufficiently sensitive and thus unsuitable for QC release. Our new HPLC method was sensitive, quantitative, reproducible, and rapid for QCs, allowing us to exactly determine the residual HEPES amount in [68Ga]Ga-radiopharmaceuticals for safe patient administration.

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Automated radiosynthesis of pharmaceutical grade [68Ga]Ga-NODAGA-Lys40-Exendin-4 and demonstration of its efficacy for use in patients

Menon,

Mitra,

Chakraborty

et al. 2024

J Radioanal Nucl Chem

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The radiosynthesis of pharmaceutical grade [68Ga]Ga-NODAGA-Lys40-Exendin-4 was accomplished in a fixed tubing-based module, EZ Modular-Lab Standard. Purification of the product via hydrophilic-lipophilic balance cartridge, yielded satisfactory results with radiochemical purity > 98%. The decay-corrected radiochemical yield (RCY) with 50 µg of Exendin-4[Lys40(NODAGA)] was (78.7 ± 0.8)%, which, by far, to the best of our knowledge, is the highest RCY reported till date, using an automated synthesizer. The quality control parameters were in accordance with that of gallium (68Ga) Edotreotide® Injections, featuring in the European Pharmacopoeia. Pre-clinical dosimetry studies in animal models, with co-administration of kidney protectant, demonstrates promising potential of the product towards clinical translation to insulinoma patients. Graphical abstract

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Development and Validation of an Analytical HPLC Method to Assess Chemical and Radiochemical Purity of [68Ga]Ga-NODAGA-Exendin-4 Produced by a Fully Automated Method

Cited by 9 publications

References 35 publications

Fully automated radiolabeling of [68Ga]Ga-EMP100 targeting c-MET for PET-CT clinical imaging

Fully automated radiolabeling of [68Ga]Ga-EMP100 targeting c-MET for PET-CT clinical imaging

A Specific HPLC Method to Determine Residual HEPES in [68Ga]Ga-Radiopharmaceuticals: Development and Validation

Automated radiosynthesis of pharmaceutical grade [68Ga]Ga-NODAGA-Lys40-Exendin-4 and demonstration of its efficacy for use in patients

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