Evaluation of the 177mLu-concentration in in-house produced 177Lu-radiopharmaceuticals and commercially available Lutathera®

Background 177Lu-radiopharmaceuticals can contain the metastable impurity [177mLu]lutetium with a physical half-life of 160.4 days, in varying concentrations depending on the route of production of the radionuclidic precursor [177Lu]lutetium. Due to the long half-life of [177mLu]lutetium, difficulties with waste disposal or sterility testing could arise. Here, we analyzed several 177Lu-samples of different origins and suppliers regarding their 177mLu-concentration. Results All samples tested showed a 177mLu-concentration in the range that was stated on the certificate of analysis from the supplier which is in accordance with the European Pharmacopoeia. Conclusions Although all 177mLu-concentrations were in accordance with the European Pharmacopoeia, we need to take into account the respective national legislation regarding radioactivity release limits. With regard to the German legislation, several probes for sterility testing in external laboratories could not be released for transport due to the concentration of [177mLu]lutetium. Moreover, waste water tanks should specifically be monitored for 177mLu-concentration, when e.g. Lutathera® is administered in the clinic.

in Germany to distribute [ 177 Lu]lutetium, namely IDB Holland (Netherlands) and ITM Isotope Technologies (Germany).According to the European Pharmacopoeia, [ 177 Lu] lutetium for radiolabeling must have a radionuclidic purity of greater than 99%, whereas [ 176 Yb]ytterbium (< 0.1%), [ 177m Lu]lutetium (< 0.07%) and an overall radionuclidic impurity (without [ 176 Yb]ytterbium and [ 177m Lu]lutetium) of < 0.01% is allowed.In addition, radiopharmaceuticals for i.v.application must be sterile and therefore tested for sterility.In most radiopharmacies, producing in-house radiopharmaceuticals, the samples for sterility testing are sent to an external laboratory for analysis.The samples for sterility are therefore transported directly after synthesis, labeled as radioactive transport, to the external laboratory, or-as in most radiopharmacies-the samples are sent after the radioactivity reached the exemption limit.For [ 177m Lu]lutetium, the exemption limit is 1 × 10 6 Bq and 0.1 Bq/g for the unrestricted release of solid and liquid substances (Strahlenschutzverordnung Anlage 2018).
For the reactor production of The first approved drug that contains [ 177 Lu]lutetium is Lutathera ® .There are already several publications that describe the presence of 177m Lu-impurities in Lutathera ® (Brown 2020; Prevot et al. 2023).The problem that might arise in some nuclear medicine facilities is related to the exemption limit of 5 × 10 5 Bq/m 3 in the waste water (Strahlenschutzverordnung Anlage 2018).If this limit is reached it would result in waste Fig. 1 Production routes for lutetium-177 2 , created with biorender.commanagement problems like longer storage times.Worst case, if the contaminated water would enter unknowingly into waste water systems and could be harmful to the environment.Due to the recent approval of Pluvicto ® , an increased demand for [ 177 Lu]lutetium is assumed.Since nuclear medicine departments require an uninterrupted, stable, reliable and sufficient supply with [ 177 Lu]lutetium (Vogel et al. 2021), several additional suppliers of [ 177 Lu]lutetium might enter the market soon.
A recent study by Prevot et al. demonstrated that 45 vials of administered Lutathera ® contained 0.3% [ 177m Lu]lutetium (Prevot et al. 2023).The authors claim a waste disposal plan that anticipates a minimum of three years storage in terms of empty vials and biohazards (less than 1 MBq 177m Lutetium-content and less than 0.1 µSv/h at contact) and five years for partially filled vials.
In order to be able to estimate the possible effects of further approvals of 177 Lu-therapeutics on 177m Lu-concentrations in waste water, we evaluated 97 retention samples of 177 Lu-therapeutics that were produced in-house with approved EndolucinBeta.Furthermore, we evaluated the effect of [ 177m Lu]lutetium impurities on the realization of sterility controls as required by Ph.Eur. for sterile radiopharmaceuticals for i.v.application.In addition, we performed test synthesis with other [ 177 Lu]lutetium suppliers, i.e.IDB Holland, Eckert & Ziegler Radiopharma GmbH (EZAG), Novartis Pharma GmbH and Monrol Nuclear Products Co and analyzed the samples towards their 177m Lu-concentrations.Additionally, we analyzed 38 Lutathera ® probes.

Results
The experiments were performed following the schematic protocol described in Fig. 2 and Fig. 3.In total, 138 samples from year 2021 were analyzed by gamma spectroscopy and analyzed with the program Apex-Gamma (Version 1.4.1) for their 177m Lu-content.The results obtained were normalized by recalculation to the activity and volume on the calibration date.For the retention samples, the exact amount of [ 177m Lu]lutetium in the volume in the retention sample was calculated.

EndolucinBeta ® (probes 1-97)
Figure 4 shows the 177m Lu-activity that was found in the EndolucinBeta ® (ITM) samples in the respective months of the year 2021 (January until December).In January, we measured an 177m Lu-activity of 1075 Bq/g.From February until May, we measured an 177m Lu-activity of approximately 482 Bq/g and from June until December, the 177m Lu-activity was around 175 Bq/g.The mean 177m Lu-activity in the samples of EndolucinBeta ® was 381 ± 511 Bq/g, which represents a radionuclidic impurity of < < 0.01% (approx.0.0000037 ± 0.0000053%) which is very far below the limit of 0.01% stated on the certificate of analysis (see Table 1).
Regarding waste management, we evaluated the volume in the retention samples of the EndolucinBeta ® probes for their 177m Lu-content.Here, we observed a 177m Lu-content of 61 ± 101 Bq/g.

Non-approved [ 177 Lu]lutetium (probes 98 and 99)
For the test syntheses with non-approved [ 177 Lu]lutetium probes at the date of testing (EZAG and Monrol), we found 9521 Bq/g (± 371 Bq/g with an measurement error of 3.9%) which results in a radionuclidic impurity of < < 0.01% (approx.0.0000453%) of [ 177m Lu]lutetium for EZAG and 39 Bq/g (± 1 Bq/g with an measurement error of 3.7%) and < < 0.01% (approx.0.0000002%) of [ 177m Lu]lutetium for Monrol (see Table 1).Both values are very far below the stated limits on the certificate of analysis.For the test synthesis with approved LuMark ® (IDB Holland), we found 706,594 Bq/g (± 21,198 Bq/g with an measurement error of 3.0%) resulting in radionuclidic impurity of < 0.01% (approx.0.008248%) of [ 177m Lu]lutetium which is in the range of the stated 177m Lu-content on the certificate of analysis, when the measurement error of 3.0% is taken into account.

Lutathera ® (probes 101-138)
For Lutathera ® (Novartis), we analyzed a total of 38 samples-29 empty Lutathera ® vials and 9 samples with residual activity that was filled into extra vials before the patient received a reduced Lutathera ® activity.We found 29,512 ± 6491 Bq/g [ 177m Lu] lutetium in Lutathera ® , corrected to the time of the first measurement before application, resulting in an radionuclidic impurity of < 0.01% (approx.0.0096%) of [ 177m Lu] lutetium.Novartis Pharma GmbH replied to our inquiry that the ratio of [ 177m Lu] lutetium in Lutathera ® is below 0.015% and the absolute [ 177m Lu]lutetium-activity is about 1 MBq.

Discussion
We analyzed 97 EndolucinBeta ® probes regarding their 177m Lu-content via gamma spectroscopy.First, we evaluated the results with respect to the stated 177m Luimpurity on the certificate of analysis.The specifications and measured activities of [ 177m Lu]lutetium were within the criteria limits and the stated values were successfully reproduced.Secondly, we evaluated the 177m Lu-content and consequences for sterility testing or waste disposal with regard to the retention samples.The retention samples of EndolucinBeta ® (n = 97), showed an average 177m Lu-content of 61 Bq/g ± 101 Bq/g in a volume ranging from 0.4 mL to 11.1 mL.With a physical half-life of 160.4 days, it will take approximately 5 years until the 177m Lu-content undercuts the exemption limit of 0.1 Bq/g.Consequently, the retention samples for sterility testing would need to be transported with a pricy radioacitivity transport to an external laboratory for analysis as well as the laboratory must have a permit for handling.To prevent this, an in-house sterility testing might need to be established.In 13 out of 97 probes, no 177m Lu-content was detected.The storage period for retention samples of radiopharmaceuticals is 6 months according to GMP guidelines (Commission 2008).Thus, an extra 4.5 years is necessary until the retention samples can be safely disposed.
The other three 177 Lu-samples from different suppliers (EZAG, Monrol, IDB Holland) were analyzed regarding their 177m Lu-content without patient application.To note, for each supplier, we only had one probe (n = 1) in contrast to EndolucinBeta ® (n = 97).As seen in Table 1, Monrol had the lowest 177m Lu-content of 39 Bq/g and 0.00002 ppm of the total 177 Lu-activity of 21.21 GBq.On the other hand, the c.a. production from IDB Holland clearly showed an elevated concentration of [ 177m Lu]lutetium.Here, we found 0.8 ppm of [ 177m Lu]lutetium.
For Lutathera ® , we found higher 177m Lu-concentrations compared to n.c.a.produced [ 177 Lu]lutetium.In 38 analyzed samples, we stayed within the Ph.Eur.required 177m Luactivity level of 0.07%.

Conclusions
We can conclude, that the measured 177m Lu-impurity concentrations of all suppliers were within the limits stated on the respective certificate of analysis.We also detected [ 177m Lu]lutetium in n.c.a. 177Lu-samples.This can be explained by the target material, which is usually > 99%, but not 100% pure.Regarding the elevated 177m Lu-content in Lutathera ® and LuMark ® , each nuclear medicine facility and radiopharmacy needs to pay very close attention to a safe waste disposal and shipping for e.g.sterility testing with respect to their local and national radiation safety regulations and radioactivity release limits.We did not include Pluvicto ® in our studies, as there was no sample available in our institution.But we would like to highlight, that in the smPC of Pluvicto ® it is stated, that the radioactive precursor might either produced by the direct or the indirect route (smPC. of Pluvicto 2022) and that special attention should be drawn to the respective batch release certificate.No such comment was found in the smPC of Lutathera ® (smPC. of Lutathera 2022).

Probes
The probes originate from in-house-or validation productions of 177 Lu-radiopharmaceuticals.The [ 177 Lu]LuCl 3 used as starting material, was purchased either from ITM Garching (EndolucinBeta All probes were decaying for at least 85 days so that the 177 Lu content is low enough to measure metastable contamination with gamma spectroscopy.

Gamma spectroscopy
Gamma radiation was measured for 60 min with deadtime correction using a standard coaxial Germanium-detector [HPGe detector, GC2020, Mirion Technologies (Canberra) GmbH].The output is connected to a LYNX multichannel analyzer.The measured spectra were analyzed with the program Apex-Gamma (Version 1.4.1) for their 177m Lu-content.The analysis program calculates the activity from the measured energy peaks, the probability of emission and the energy-dependent efficiency.In the report it lists the mean activity of all analyzed energy peaks, which are given in the nuclide library.The report also shows the measurement error.It is multifactorial and includes the uncertainty of the instrument, the uncertainty of the Poisson statistics and the differences between the activities calculated from the individual peaks.The measurement is limited by the influence of residual 177 Lu, which affects the activity calculation of 177m Lu.However, since the energy spectrum of the two nuclides differs by the energy 228 keV, which is only emitted by 177m Lu, this error is also represented by the listed measurement errors (Table 1), since they include the scatter of the calculated activities of the different energy peaks.The detection limit is depending on the background and the nuclide, for the measured Lutathera Vials the mean detection limit for 177m Lu is 29.8 ± 22.2 Bq (values reach from 12.8 Bq to 94.1 Bq).To ensure the reproducibility of the gamma spectroscopy measurements monthly quality assurances are performed.Data was recorded and stored on a APEX PC, Windows 10 Pro (2019) and processed and analyzed with Apex Gamma software.
In our experiments, we used correction factors to increase the accuracy of our measurements.Only the 100 ml Kautex vials are calibrated on our system.Consequently, we had to calculate a correction factor for each different vial type.First, the probes were analyzed as described below.Secondly, the probe (e.g.27 mL vial) was rinsed with approximately 50 mL of demineralized water and refilled up to 100 mL into a 100 mL plastic Kautex vial and the activity was measured by gamma spectroscopy.For the 27 mL Lutathera ® vial (n = 29), we calculated a correction factor of 0.66 (the vials with the reduced dose had a correction factor of 0.55).For EndolucinBeta (n = 97), IDB Holland (n = 1) and EZAG (n = 1), the correction factor was 0.48.
From 34 received batches Lutathera ® (in 52 vials), we used the released documents to calculate the amount of [ 177m Lu]lutetium according to the manufacturer's measurements to 0.0087% ± 0.0019% of the total activity or 28,063 ± 5884 Bq/g at the planned time of injection.

Fig. 2
Fig. 2 Schematic overview of experimental setup, created with biorender.com

Fig. 3 Fig. 4 .
Fig. 3 Gamma spectrometer a full view of automatic sample changer (left) sample arm (middle) and lead shielded detector (right) and b sample tray