Formulation of patient dose of [177Lu]Lu-Trastuzumab using in-house developed freeze-dried kit: A path forward for clinical translation.

Trastuzumab is a US-FDA-approved humanized monoclonal antibody used for the treatment of human epidermal growth factor receptor 2 (HER2)-positive breast cancer. The aim of the present work is to optimize a freeze-dried formulation of DOTA-Trastuzumab conjugate for the preparation of patient doses of [177Lu]Lu-Trastuzumab for radioimmunotherapy of breast cancer. The formulation of [177Lu]Lu-Trastuzumab usually takes a long time, and thus, such a process is not suitable for the routine preparation of this agent in hospital radiopharmacies. To circumvent this, a pre-synthesized DOTA-Trastuzumab conjugate as a freeze-dried formulation is proposed. In the present work, DOTA-Trastuzumab conjugate was subjected to a freeze-drying process after the addition of optimized amounts of radioprotectant and cryoprotectant. [177Lu]Lu-DOTA-Trastuzumab was prepared by incubating the lyophilized powder of the kit vial with medium-specific activity 177LuCl3. The final radiochemical purity of [177Lu]Lu-DOTA-Trastuzumab, prepared using freeze-dried kit, was determined to be >95%. To ascertain the reproducibility of the procedure, six consecutive batches of the freeze-dried formulation were prepared, radiolabeled, and evaluated by carrying out both in vitro and ex vivo studies. The consistency of the results of all the six consecutive batches confirmed the robustness and utility of the in-house optimized freeze-dried formulation for the preparation of patient doses of [177Lu]Lu-Trastuzumab at hospital radiopharmacies.

A comparison of simplified protocols of personalized dosimetry in NEN patients treated by radioligand therapy (RLT) with [177Lu]Lu-DOTATATE to favor its use in clinical practice.

The role of internal dosimetry is usually proposed for investigational purposes in patients treated by RLT, even if its application is not yet the standard method in clinical practice. This limited use is partially justified by several concomitant factors that make calculations a complex process. Therefore, simplified dosimetry protocols are required. METHODS: In our study, dosimetric evaluations were performed in thirty patients with NENs who underwent RLT with [177Lu]Lu-DOTATATE. The reference method (M0) calculated the cumulative absorbed dose performing dosimetry after each of the four cycles. Obtained data were employed to assess the feasibility of simplified protocols: defining the dosimetry only after the first cycle (M1) and after the first and last one (M2). RESULTS: The mean differences of the cumulative absorbed doses between M1 and M0 were - 10% for kidney, - 5% for spleen, + 34% for liver, + 13% for red marrow, and + 37% for tumor lesions. Conversely, differences lower than ± 10% were measured between M2 and M0. CONCLUSION: Cumulative absorbed doses obtained with the M2 protocol resembled the doses calculated by M0, while the M1 protocol overestimated the absorbed doses in all organs at risk, except for the spleen.

Synthesis and evaluation of [177 Lu]Lu-labeled porphyrin loaded PAMAM dendrimer: Impact on tumor uptake and pharmacokinetics.

The objective of the present work is to evaluate the ability of the radiolabeled PAMAM dendrimers (polyamidoamine) towards facilitating the delivery of an in-house synthesized porphyrin derivative in the tumorous lesions to evaluate their candidature for possible application in endo-radionuclide therapy. For this, PAMAM particles were conjugated with a porphyrin derivative namely, 5,10,15,20-tetrakis-(4-carboxymethyleneoxyphenyl)porphyrin (STAP), synthesized in-house following a two-step reaction. The average number of porphyrin molecules loaded per PAMAM particle was evaluated using ultraviolet-visible spectrophotometry and was found to be approximately 2. STAP conjugated PAMAM particles were further conjugated with p-NCS-benzyl-DOTA (subsequently referred as DOTA) to facilitate radiolabeling with 177 Lu. On an average, two p-NCS-benzyl-DOTA molecules were observed to be attached per PAMAM-STAP particle. DOTA-PAMAM-STAP conjugate was radiolabeled with 177 Lu with a final radiochemical purity of >95%, which was determined by paper chromatography using two different mobile phases viz. 0.1 M sodium citrate buffer (pH 5.0) and 10 mM DTPA. Biological behavior of [177 Lu]Lu-DOTA-PAMAM-STAP conjugate was investigated in fibrosarcoma bearing Swiss mice model wherein accumulation of radiolabeled particles was observed in liver, GIT, spleen, and kidneys at 3 h post-administration. However, accumulated activity exhibited rapid clearance from majority of the organs at 24 h post-administration. [177 Lu]Lu-DOTA-PAMAM-STAP conjugate exhibited an appreciable uptake in tumor mass [6.09 ± 1.22 percentage injected activity/organ (% IA/organ)] at 3 h post-administration (p.i.) which was found to reduce to 1.05 ± 0.13 % IA/organ at 24 h post-administration. The results obtained in biodistribution studies were further corroborated through scintigraphic imaging performed in the same animal model. Despite of an appreciable accumulation in tumor mass, the lower retention of the [177 Lu]Lu-DOTA-PAMAM-STAP conjugate therein, at longer time point (24 h p.i.) may limit its possible potential as a radio-therapeutic agent and indicates towards need for further structural manoeuvring to attain favorable in vivo performance.

Technical note: Quantifying radionuclide residues in hospital wastewater: A case study on [131I]I and [177mLu]Lu/[177Lu]Lu.

BACKGROUND: Historically, [131I]I has been a common isotope in radionuclide therapy, with [177Lu]Lu-labelled radiopharmaceuticals now seeing a surge in use. These can include no-carrier-added or carrier-added [177Lu]Lu with slight impurities of [177mLu]Lu with a significantly longer half-life than [131I]I. Wastewater from therapy wards can contain a mixture of these radioisotopes. In some countries, national regulations require wastewater to be stored in dedicated systems before it is discharged into the public sewage system. To fulfill legal requirements, the nuclide specific activity concentration must be verified. PURPOSE: We evaluate a method for determining the activity concentration of [177mLu]Lu /[177Lu]Lu at equilibrium and [131I]I in pure and mixed samples in order to prove that the determined values are reliably below the limits for release. METHODS: We analysed the emitted energy spectrum of 1 L samples with a wastewater counter using an energy window-based approach by evaluating measurements from two different time points. Based on the law of decay and the time and energy-dependent measured values, equation systems were set up to calculate the count rates for [131I]I and [177mLu]Lu, which were converted into activity concentration using calibration factors. RESULTS: There is strong linear correlation between the nominal and determined activity concentrations (correlation coefficients R = 0.99; coefficient of determinations R2 = 0.99). We underestimate the actual activity concentration by a median of -1.4% for [177mLu]Lu and overestimate the activity concentration for [131I]I by a median of 7.1%. CONCLUSION: We show that an undercut of the clearance levels for material release is measurable. We analyse and determine activity concentrations of mixed samples consisting of [131I]I and [177mLu]Lu/[177Lu]Lu in equilibrium. The method is simple to implement using a conventional wastewater counter, however with a slightly increased effort, as two samples and measurements are required. The methodology can be adapted for the analysis of other nuclide mixtures.

Technical note: Assessment of radiation measurement devices for the detection of [177 Lu]Lu-labeled radiopharmaceuticals containing [177m Lu]Lu-impurities.

BACKGROUND: Nuclear medicine therapies using [177 Lu]Lu-labeled radiopharmaceuticals have increased significantly in recent years. Some of these radiopharmaceuticals contain long-lived impurities of [177m Lu]Lu. Identification of this specific contamination and its quantification is important in different scenarios. PURPOSE: In this study, standard measurement hardware used for handling radioisotopes was evaluated for the measurement of [177m Lu]Lu and [177 Lu]Lu at equilibrium. Device-specific detection limits (LODs) for [177m Lu]Lu were determined according to international standards and then validated. METHODS: The LODs were determined according to the international standard ISO 11929-1 for [177m Lu]Lu for five identical portable contamination monitors (PCMs), a wastewater counter (WWC), and a release counter system (RCS) at different measuring times. Subsequent activity measurements of the defined samples were used to validate the linearity of the measurement instruments down to the LOD for each system. RESULTS: The average LOD across all PCMs was 0.249 ± 0.009 and 0.129 ± 0.005 Bq/cm2 for 10 and 30 s measurements, respectively. The LODs of WWC varied between 3.3 and 4.7 Bq/L for measurement times of 300 s and 0.8-1.3 Bq/L for 3600 s depending on the energy window studied. The LOD of the RCS depended on the container volume and was 0.08 Bq/g for the 50 L container at 60 s measurement. The measurements for all examined devices were linear down to the LOD (correlation coefficient R ≥ 0.96; coefficient of determination R2  ≥ 0.92). CONCLUSIONS: All investigated measuring instruments (PCM, WWC, RCS) were suitable for the determination of [177m Lu]Lu at equilibrium, and their specific LODs were determined. Based on the measurements performed, activity is overestimated; however, this is tolerable because assumptions and measurements in the context of radiation protection should be conservative.

Bioinformatic pipelines combining denoising and clustering tools allow for more comprehensive prokaryotic and eukaryotic metabarcoding.

Environmental DNA metabarcoding is a powerful tool for studying biodiversity. However, bioinformatic approaches need to adjust to the diversity of taxonomic compartments targeted as well as to each barcode gene specificities. We built and tested a pipeline based on read correction with DADA2 allowing analysing metabarcoding data from prokaryotic (16S) and eukaryotic (18S, COI) life compartments. We implemented the option to cluster amplicon sequence variants (ASVs) into operational taxonomic units (OTUs) with swarm, a network-based clustering algorithm, and the option to curate ASVs/OTUs using LULU. Finally, taxonomic assignment was implemented via the Ribosomal Database Project Bayesian classifier (RDP) and BLAST. We validated this pipeline with ribosomal and mitochondrial markers using metazoan mock communities and 42 deep-sea sediment samples. The results show that ASVs and OTUs describe different levels of biotic diversity, the choice of which depends on the research questions. They underline the advantages and complementarity of clustering and LULU-curation for producing metazoan biodiversity inventories at a level approaching the one obtained using morphological criteria. While clustering removes intraspecific variation, LULU effectively removes spurious clusters, originating from errors or intragenomic variability. Swarm clustering affected alpha and beta diversity differently depending on genetic marker. Specifically, d-values > 1 appeared to be less appropriate with 18S for metazoans. Similarly, increasing LULU's minimum ratio level proved essential to avoid losing species in sample-poor data sets. Comparing BLAST and RDP underlined that accurate assignments of deep-sea species can be obtained with RDP, but highlighted the need for a concerted effort to build comprehensive, ecosystem-specific databases.

Expand, relocate, or underground? Social acceptance of upgrading wastewater treatment plants.

Securing a moderate level of social acceptance for obnoxious facilities, public facilities that have negative effects, such as odors, noise, or other disruptions, is critical to infrastructure plans. For wastewater treatment plant (WWTP), also obnoxious facilities, upgrading and expanding the capacity of existing WWTP, are more important than the construction of new plants, in some regions. This study analyzes and compares the social acceptance of different types of WWTP upgrades and capacity expansion projects. Contingent valuation method is used to elicit South Korean households' willingness to pay (WTP) for preventing the expansion of a WWTP. The aggregated WTP is interpreted from the perspective of social conflict costs. The results show that the annual mean WTP of South Korean households to prevent WWTP expansion ranges from KRW 32,058 (US $27.61) to KRW 45,793 (US $39.44) depending on spatial location, which implies that the social conflict costs for the WWTP expansion in South Korea are considerable. It is also found that an underground WWTP at current site is a best alternative to lower the social conflict costs; it is even better than relocation an existing WWTP to another area. Several related policy implications are provided based on the analysis results.