Radiolabeling and quality control of therapeutic radiopharmaceuticals: optimization, clinical implementation and comparison of radio-TLC/HPLC analysis, demonstrated by [177Lu]Lu-PSMA

Abstract

Background Radiopharmaceuticals are considered as regular medicinal products and therefore the same regulations as for non-radioactive medicinal products apply. However, specific aspects should be considered due to the radiochemical properties. Radiopharmaceutical dedicated monographs are developed in the European Pharmacopoeia to address this. Currently, different quality control methods for non-registered radiopharmaceuticals are utilized, often focusing on radio-TLC only, which has its limitations. When the radiochemical yield (RCY) is measured by radio-TLC analysis, degradation products caused by radiolysis are frequently not detected. In contrast, HPLC analysis defines the radiochemical purity (RCP), allowing for detection of peak formation related to radiolysis. During the introduction and optimization phase of therapeutic radiopharmaceuticals, significant percentages of impurities, like radiolysed construct formation, may have consequential impact on patient treatment. Since more hospitals and institutes are offering radiopharmaceutical therapies, such as [177Lu]Lu-PSMA with an in-house production, the demand for adequate quality control is increasing. Here we show the optimization and implementation of a therapeutic radiopharmaceutical, including the comparison of ITLC and HPLC quality control. Results Downscaled conditions (74 MBq/μg) were in concordance to clinical conditions (18 GBq/250 µg, 5 mL syringe/100 mL flacon); all results were consistent with an > 98% RCY (radio-TLC) and stability of > 95% RCP (HPLC). Radio-TLC did not identify radiolysis peaks, while clear identification was performed by HPLC analysis. Decreasing the RCP with 50%, reduced the cell-binding capacity with 27%. Conclusion This research underlines the importance of the radiolabeling and optimization including clinical implementation and clarifies the need for cross-validation of the RCY and RCP for quality control measurements. Only HPLC analysis is suitable for identification of radiolysis. Here we have proven that radiolysed [177Lu]Lu-PSMA has less binding affinity and thus likely will influence treatment efficacy. HPLC analysis is therefore essential to include in at least the validation phase of radiopharmaceutical implementation to ensure clinical treatment quality.

Improved Quality Control of [177Lu]Lu-PSMA I&T

“…In the case of PSMA I&T it was striking that radiolysis was more pronounced as compared to PSMA-617 radiolabelled under the same conditions. These results are in accordance with ndings of Hooijman (14), who observed as well an increasing side peak over time and eluting before the main peak. Whereas they report a difference in retention time of less than 1 min without baseline separation, in our case this was more than 4 min, supporting the improved high resolution and speci city of our method.…”

“…Quality of such a radiopharmaceutical, in particular radiochemical purity, is of utmost importance, especially considering the therapeutic application. Several groups have published methods to determine the quality of this preparation (11)(12)(13)(14)(15)(16), with a focus on RCP testing. However, the quality is not only related to the RCP alone, HPLC analysis allows obtaining conformation of identity of the radiopharmaceutical and, typically in the form of a limit test, also to determine the amount of precursor and related substances, which should be limited in relation to toxicity concerns.…”

“…However, so far, no formal quality standards are available. A variety of methods have been reported to determine the radiochemical purity of [ 177 Lu]Lu-PSMA I&T. These methods are based on reversed phase high performance liquid chromatography (RP-HPLC) employing standard conditions with C-18 columns and acetonitrile (ACN) /water mixtures containing 0.1% TFA as counterion (11)(12)(13)(14)(15). Recently, Aalbersberg et al (16) reported unspeci c retention of [ 177 Lu]Lu-PSMA I&T on RP-C-18 HPLC columns using TFA containing solvents, that could be overcome by spiking quality control samples with PSMA I&T precursor.…”

Improved quality control of [177Lu]Lu-PSMA I&T

“…Quality of such a radiopharmaceutical, in particular radiochemical purity, is of utmost importance, especially considering the therapeutic application. Several groups have published methods to determine the quality of this preparation (Orhon et al 2022 ; Weineisen et al 2014 ; Iorio et al 2022 ; Hooijman et al 2022 ; Vyas et al 2022 ; Aalbersberg et al 2022 ), with a focus on RCP testing. However, the quality is not only related to the RCP alone, HPLC analysis allows obtaining confirmation of identity of the radiopharmaceutical and, typically in the form of a limit test, also to determine the amount of precursor and related substances, which should be limited in relation to toxicity concerns.…”

“…However, so far, no formal quality standards are available. A variety of methods have been reported to determine the radiochemical purity of [ 177 Lu]Lu-PSMA I&T. These methods are based on reversed-phase high-performance liquid chromatography (RP-HPLC) employing standard conditions with C-18 columns and acetonitrile (ACN) /water mixtures containing 0.1% Trifluoroacetic acid (TFA) as counterion (Orhon et al 2022 ; Weineisen et al 2014 ; Iorio et al 2022 ; Hooijman et al 2022 ; Vyas et al 2022 ). Recently, Aalbersberg et al ( 2022 ) reported unspecific retention of [ 177 Lu]Lu-PSMA I&T on RP-C-18 HPLC columns using TFA containing solvents, that could be overcome by spiking quality control samples with PSMA I&T precursor.…”