Medical radionuclides such as Ga-68, Cu-64 or Ac-225 are usually produced by irradiation of enriched target materials in cyclotrons or nuclear reactors. After irradiation, the radionuclides need to be separated from their target. While this is mostly done by ion-exchange chromatography, an emerging separation method includes the use of (microfluidic) solvent extraction. However, the extent to which the chelators and organic solvents used during solvent extraction contaminate the final radionuclide-containing solution, including their potential impact on subsequent radiolabeling applications, has not been studied in detail. In this study, the potential contaminants N-benzoyl-N-phenylhydroxilamine (BPHA), dithizone (DIZ) and di(2-ethylhexyl)phosphoric acid (D2EHPA) were investigated, and a microcolumn purification method is proposed. It was found that contaminations with two of these chelators, BPHA and DIZ, significantly interfered with DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) labeling. The applied microcolumn purification method eliminated the BPHA contamination from the Ga-68 solution completely, while simultaneously drastically reducing the total volume and acidity of the solution. It is therefore a promising purification method that can be included in an automated microfluidic solvent extraction procedure.

Background: The radionuclide Ga-68 is commonly used in nuclear medicine, specifically in positron emission tomography (PET). Recently, the interest in producing Ga-68 by cyclotron irradiation of [⁶⁸Zn]Zn nitrate liquid targets is increasing. However, current purification methods of Ga-68 from the target solution consist of multi-step procedures, thus, leading to a significant loss of activity through natural decay. Additionally, several processing steps are needed to recycle the costly, enriched target material. Results: To eventually allow switching from batch to continuous production, conventional batch extraction and membrane-based microfluidic extraction were compared. In both approaches, Ga-68 was extracted using N-benzoyl-N-phenylhydroxylamine in chloroform as the organic extracting phase. Extraction efficiencies of up to 99.5% ± 0.6% were achieved within 10 min, using the batch approach. Back-extraction of Ga-68 into 2 M HCl was accomplished within 1 min with efficiencies of up to 94.5% ± 0.6%. Membrane-based microfluidic extraction achieved 99.2% ± 0.3% extraction efficiency and 95.8% ± 0.8% back-extraction efficiency into 6 M HCl. When executed on a solution irradiated with a 13 MeV cyclotron at TRIUMF, Canada, comparable efficiencies of 97.0% ± 0.4% were achieved. Zn contamination in the back-extracted Ga-68 solution was found to be below 3 ppm. Conclusions: Microfluidic solvent extraction is a promising method in the production of Ga-68 achieving high efficiencies in a short amount of time, potentially allowing for direct target recycling. Graphical Abstract: [Figure not available: see fulltext.].

Scandium-47 (⁴⁷Sc) can be used in nuclear medicine as a therapeutic-diagnostic, or “theragnostic,” radioactive medical isotope for cancer detection and treatment. The ⁴⁷Sc isotope can be produced through the photonuclear reaction ⁴⁸Ti(γ,p)⁴⁷Sc by irradiating enriched ⁴⁸Ti target material. The enriched target material necessary for production is costly; ⁴⁸TiO₂ costs ~ $1550/g, and targets can be > 50 g ($77,500) to produce medically relevant amounts of ⁴⁷Sc. In order to keep costs low, a highly efficient separation of scandium from bulk titanium is desired, along with efficient methods for recycling the target material. This research is focused on evaluating efficient methods for the separation of scandium from bulk quantities of titanium using commercially available diglycolamide-based and hydroxamate-based extraction chromatography resins (DGA resin and ZR resin, respectively). The sorption of ⁴⁷Sc and Ti on these resins were investigated at varying concentrations of HNO₃, HCl, H₂SO₄, and HF to explore how they might be used in a large-scale production/processing setting.

Iron deficiency anemia can be treated with oral or intravenous Fe supplementation. Such supplementation has considerable effects on the human microbiome, and on opportunistic pathogenic micro-organisms. Molecular understanding of the control and regulation of Fe availability at the host-microbe interface is crucial to interpreting the side effects of Fe supplementation. Here, we provide a concise overview of the regulation of Fe by the opportunistic pathogen Staphylococcus aureus. Ferric uptake regulator (Fur) plays a central role in controlling Fe uptake, utilization and storage in order to maintain a required value. The micro-organism has a strong preference for heme iron as an Fe source, which is enabled by the Iron-regulated surface determinant (Isd) system. The strategies it employs to overcome Fe restriction imposed by the host include: hijacking host proteins, replacing metal cofactors, and replacing functions by non-metal dependent enzymes. We propose that integrated omics approaches, which include metalloproteomics, are necessary to provide a comprehensive understanding of the metal tug of war at the host-microbe interface down to the molecular level.

A variety of polymer micelles are designed for the delivery of chemotherapeutic drugs to tumors. Although the promise of these nanocarriers is very high, in the clinic the effectivity is rather limited. Determining the in vivo fate of the micelles can greatly help to improve this treatment. Here, a simple and fast chelator-free method for radiolabeling of polymer micelles composed of different block copolymers is presented, which can allow evaluating the behavior of the nanocarriers in vivo using noninvasive nuclear imaging techniques (e.g., single photon computed tomography, SPECT). The radiolabeling method consists of adding the radioisotope ions, i.e., ¹¹¹In(III), resulting in a high radiolabeling efficiencies up to 90%. The results suggest that the radiolabeling efficiency depends on two important factors: the properties of the hydrophobic block in the block copolymer composing the micelle core, and the speciation of the radiometal salts. The formation of metal hydroxides and their precipitation in the core of the micelles appears to be a key factor for high stability. Moreover, the method can be applied to radiolabel the micelles in the presence of chemotherapeutic drugs. Finally, a SPECT study shows that the radiolabeled samples are stable in vivo without any evident loss of ¹¹¹In(III).

In certain tumor and diseased tissues, reactive oxygen species (ROS), such as H₂O₂, are produced in higher concentrations than in healthy cells. Drug delivery and release systems that respond selectively to the presence of ROS, while maintaining their stability in “healthy” biological conditions, have great potential as on-site therapeutics. This study presents polymer micelles with 4-(methylthio)phenyl ester functionalities as a ROS-responsive reactivity switch. Oxidation of the thioether moieties triggers ester hydrolysis, exposing a hydrophylic carboxylate and leading to micellar disassembly. At 37 °C, the micelles fall apart on a timescale of days in the presence of 2 mM H₂O₂ and within hours at higher concentrations of H₂O₂ (60-600 mM). In the same time frame, the nanocarriers show no hydrolysis in oxidant-free physiological or mildly acidic conditions. This logic gate cascade behavior represents a step forward to realize drug delivery materials capable of selective response to a biomarker input.

Polymersomes have the potential to be applied in targeted alpha radionuclide therapy, while in addition preventing release of recoiling daughter isotopes. In this study, we investigated the cellular uptake, post uptake processing and intracellular localization of polymersomes. Methods: High-content microscopy was used to validate polymersome uptake kinetics. Confocal (live cell) microscopy was used to elucidate the uptake mechanism and DNA damage induction. Intracellular distribution of polymersomes in 3-D was determined using super-resolution microscopy. Results: We found that altering polymersome size and concentration affects the initial uptake and overall uptake capacity; uptake efficiency and eventual plateau levels varied between cell lines; and mitotic cells show increased uptake. Intracellular polymersomes were transported along microtubules in a fast and dynamic manner. Endocytic uptake of polymersomes was evidenced through co-localization with endocytic pathway components. Finally, we show the intracellular distribution of polymersomes in 3-D and DNA damage inducing capabilities of²¹³Bi labeled polymersomes. Conclusion: Polymersome size and concentration affect the uptake efficiency, which also varies for different cell types. In addition, we present advanced assays to investigate uptake characteristics in detail, a necessity for optimization of nano-carriers. Moreover, by elucidating the uptake mechanism, as well as uptake extent and geometrical distribution of radiolabeled polymersomes we provide insight on how to improve polymersome design.

Background and purpose: Glioblastoma multiforme (GBM) is the most aggressive subtype of malignant gliomas, with an average survival rate of 15 months after diagnosis. More than 90% of all GBMs have activating mutations in the MAPK/ERK pathway. Recently, we showed the allosteric MEK1/2 inhibitor binimetinib (MEK162) to inhibit cell proliferation and to enhance the effect of radiation in preclinical human GBM models. Because the free drug cannot pass the blood–brain barrier (BBB), we investigated the use of nanocarriers for transport of the drug through the BBB and its efficacy when combined with radiotherapy and temozolomide (TMZ) in glioma spheroids. Methods: In vitro studies were performed using multicellular U87 human GBM spheroids. Polymeric nanocarriers (polymersomes) were loaded with MEK162. The interaction between nanocarrier delivered MEK162, irradiation and TMZ was studied on the kinetics of spheroid growth and on protein expression in the MAPK/ERK pathway. BBB passaging was evaluated in a transwell system with human cerebral microvascular endothelial (hCMEC/D3) cells. Results: MEK162 loaded polymersomes inhibited spheroid growth. A synergistic effect was found in combination with fractionated irradiation and an additive effect with TMZ on spheroid volume reduction. Fluorescent labeled polymersomes were taken up by human cerebral microvascular endothelial cells and passed the BBB in vitro. Conclusion: MEK162 loaded polymersomes are taken up by multicellular spheroids. The nanocarrier delivered drug reduced spheroid growth and inhibited its molecular target. MEK162 delivered via polymersomes showed interaction with irradiation and TMZ. The polymersomes crossed the in vitro BBB model and therewith offer exciting challenges ahead for delivery of therapeutics agents to brain tumours.

The use of nanoparticles as tumor-targeting agents is steadily increasing, and the influence of nanoparticle characteristics such as size and stealthiness have been established for a large number of nanocarrier systems. However, not much is known about the impact of tumor presence on nanocarrier circulation times. This paper reports on the influence of tumor presence on the in vivo circulation time and biodistribution of polybutadiene-polyethylene oxide (PBd-PEO) polymersomes. For this purpose, polymersomes were loaded with the gamma-emitter¹¹¹In and administered intravenously, followed by timed ex vivo biodistribution. A large reduction in circulation time was observed for tumor-bearing mice, with a circulation half-life of merely 5 min (R² = 0.98) vs 117 min (R² = 0.95) in healthy mice. To determine whether the rapid polymersome clearance observed in tumor-bearing mice was mediated by macrophages, chlodronate liposomes were administered to both healthy and tumor-bearing mice prior to the intravenous injection of radiolabeled polymersomes to deplete their macrophages. Pretreatment with chlodronate liposomes depleted macrophages in the spleen and liver and restored the circulation time of the polymersomes with no significant difference in circulation time between healthy mice and tumor-bearing mice pretreated with clodronate liposomes (15.2 ± 1.2% ID/g and 13.6 ± 2.7% ID/g, respectively, at 4 h p.i. with p = 0.3). This indicates that activation of macrophages due to tumor presence indeed affected polymersome clearance rate. Thus, next to particle design, the presence of a tumor can also greatly impact circulation times and should be taken into account when designing studies to evaluate the distribution of polymersomes.

Increasing attention is given to personalized tumour therapy, where α-emitters can potentially play an important role. Alpha particles are ideal for localized cell killing because of their high linear energy transfer and short ranges. However, upon the emission of an α particle the daughter nuclide experiences a recoil energy large enough to ensure decoupling from any chemical bond. These ‘free’ daughter nuclides are no longer targeted to the tumour and can accumulate in normal tissue. In this paper, we used polymersomes as model carrier to evaluate the retention of recoiling daughters of ²²⁵Ac in vivo, and assessed their suitability as therapeutic agents. Vesicles containing ²²⁵Ac were injected intravenously in healthy mice, and intratumourally in tumour-bearing mice, and the relocation of free ²¹³Bi was assessed in different organs upon the injection [²²⁵Ac]Ac-polymersomes. The therapeutic effect of ²²⁵Ac-containing vesicles was studied upon intratumoural injection, where treatment groups experienced no tumour-related deaths over a 115 day period. While polymersomes containing ²²⁵Ac could be suitable agents for long-term irradiation of tumours without causing significant renal toxicity, there is still a significant re-distribution of daughter nuclides throughout the body, signifying the importance of careful evaluation of the effect of daughter nuclides in targeted alpha therapy.

Here we report on how metastable supramolecular gels can be formed through seeded self-assembly of multicomponent gelators. Hydrazone-based gelators decorated with non-ionic and anionic groups are formed in situ from hydrazide and aldehyde building blocks, and lead through multiple self-sorting processes to the formation of heterogeneous gels approaching thermodynamic equilibrium. Interestingly, the addition of seeds composing of oligomers of gelators bypasses the self-sorting processes and accelerates the self-assembly along a kinetically favored pathway, resulting in homogeneous gels of which the network morphologies and gel stiffness are markedly different from the thermodynamically more stable gel products. Importantly, over time, these metastable homogeneous gel networks are capable of converting into the thermodynamically more stable state. This seeding-driven formation of out-of-equilibrium supramolecular structures is expected to serve as a simple approach towards functional materials with pathway-dependent properties.

A recent addition to the treatment options in external beam therapy, so-called FLASH radiotherapy, shows remarkable healthy-tissue-sparing properties in a number of pre-clinical studies without impacting the overall treatment efficacy. Its potential in clinical applications is attracting a great deal of interest in the medical community. The use of ultra-high dose rates at extremely short irradiation times has been shown to significantly enhance the differential effects between normal and tumor tissue. This makes it possible to increase treatment doses without further harming the surrounding healthy tissue. While most studies to date have focused on the use of electron beams, X-ray and proton FLASH radiotherapy have also shown beneficial effects, although for these latter two the results still need to be independently confirmed. Furthermore, the mechanisms underlying the biological effects remain to be elucidated. Very recently, the FLASH effect has been demonstrated in the first human patient, with promising results, supporting further clinical studies. This review will present an overview of the investigations into FLASH radiotherapy to date.

Alpha emitters have great potential in targeted tumour therapy, especially in destroying micrometastases, due to their high linear energy transfer (LET). To prevent toxicity caused by recoiled daughter atoms in healthy tissue, alpha emitters like ²²⁵Ac can be encapsulated in polymeric nanocarriers (polymersomes), which are capable of retaining the daughter atoms to a large degree. In the translation to a (pre-)clinical setting, it is essential to evaluate their therapeutic potential. As multicellular tumour spheroids mimic a tumour microenvironment more closely than a two-dimensional cellular monolayer, this study has focussed on the interaction of the polymersomes with U87 human glioma spheroids. We have found that polymersomes distribute themselves throughout the spheroid after 4 days which, considering the long half-life of ²²⁵Ac (9.9 d) (Vaidyanathan and Zalutsky, 1996), allows for irradiation of the entire spheroid. A decrease in spheroidal growth has been observed upon the addition of only 0.1 kBq ²²⁵Ac, an effect which was more pronounced for the ²²⁵Ac in polymersomes than when only coupled to DTPA. At higher activities (5 kBq), the spheroids have been found to be destroyed completely after two days. We have thus demonstrated that ²²⁵Ac containing polymersomes effectively inhibit tumour spheroid growth, making them very promising candidates for future in vivo testing.

Alpha-emitting radionuclides like actinium-225 (²²⁵Ac) are ideal candidates for the treatment of small metastasised tumours, where the long half-life of ²²⁵Ac enables it to also reach less accessible tumours. The main challenge lies in retaining the recoiled alpha-emitting daughter nuclides, which are decoupled from targeting agents upon emission of an alpha particle and can subsequently cause unwanted toxicity to healthy tissue. Polymersomes, vesicles composed of amphiphilic block copolymers, are capable of transporting (radio)pharmaceuticals to tumours, and are ideal candidates for the retention of these daughter nuclides. In this study, the Geant4 Monte Carlo simulation package was used to simulate ideal vesicle designs. Vesicles containing an InPO₄ nanoparticle in the core were found to have the highest recoil retention, and were subsequently synthesized in the lab. The recoil retention of two of the daughter nuclides, namely francium-221 (²²¹Fr) and bismuth-213 (²¹³Bi) was determined at different vesicle sizes. Recoil retention was found to have improved significantly, from 37 ± 4% and 22 ± 1% to 57 ± 5% and 40 ± 2% for ²²¹Fr and ²¹³Bi respectively for 100 nm polymersomes, as compared to earlier published results by Wang et al. where ²²⁵Ac was encapsulated using a hydrophilic chelate (Wang et al. 2014). To better understand the different parameters influencing daughter retention, simulation data was expanded to include vesicle polydispersity and nanoparticle position within the polymersome. The high retention of the recoiling daughters and the ²²⁵Ac itself makes this vesicle design very suitable for future in vivo verification.