Combining Nuclear Medicine With Other Modalities: Future Prospect for Multimodality Imaging.

This meeting report summarizes a consultants meeting that was held at International Atomic Energy Agency Headquarters, Vienna, in July 2022 to provide an update on the development of multimodality imaging by combining nuclear medicine imaging agents with other nonradioactive molecular probes and/or biomedical imaging techniques.

Axial Phenoxylation of Aluminum Phthalocyanines for Improved Cannabinoid Sensitivity in OTFT Sensors.

Cannabis producers, consumers, and regulators need fast, accurate, point-of-use sensors to detect Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD) from both liquid and vapor source samples, and phthalocyanine-based organic thin-film transistors (OTFTs) provide a cost-effective solution. Chloro aluminum phthalocyanine (Cl-AlPc) has emerged as a promising material due to its unique coordinating interactions with cannabinoids, allowing for superior sensitivity. This work explores the molecular engineering of AlPc to tune and enhance these interactions, where a series of novel phenxoylated R-AlPcs are synthesized and integrated into OTFTs, which are then exposed to THC and CBD solution and vapor samples. While the R-AlPc substituted molecules have a comparable baseline device performance to Cl-AlPc, their new crystal structures and weakened intermolecular interactions increase sensitivity to THC. Grazing-incidence wide-angle X-ray scattering (GIWAXS) and atomic force microscopy (AFM) are used to investigate this film restructuring, where a significant shift in the crystal structure, grain size, and film roughness is detected for the R-AlPc molecules that do not occur with Cl-AlPc. This significant crystal reorganization and film restructuring are the driving force behind the improved sensitivity to cannabinoids relative to Cl-AlPc and demonstrate that analyte-semiconductor interactions can be enhanced through chemical modification to create more responsive OTFT sensors.

Tracking the fate of bacteria-derived site-specific immunomodulators by positron emission tomography.

INTRODUCTION: Site-specific immunomodulators (SSIs) are a novel class of therapeutics made from inactivated bacterial species designed to regulate the innate immune system in targeted organs. QBECO is a gut-targeted SSI that is being advanced clinically to treat and/or prevent inflammatory bowel disease, cancer, and serious infections of the gastrointestinal (GI) tract and proximal organs, and QBKPN is a lung-targeted SSI that is in clinical development for the treatment and/or prevention of chronic inflammatory lung disease, lung cancers and respiratory tract infections. While these SSIs have demonstrated both safety and proof-of-concept in preclinical and clinical studies, detailed understanding of their trafficking and biodistribution is yet to be fully characterized. METHODS: QBECO and QBKPN were radiolabeled with [89Zr] and injected subcutaneously into healthy mice. The mice underwent Positron Emission Tomography (PET) imaging every day for eight days to track biodistribution of the SSIs. Tissue from the site of injection was collected and immunohistologically probed for immune cell infiltration. RESULTS: Differential biodistribution of the two SSIs was seen, adhering to their site-specific targeting. QBKPN appeared to migrate from the site of injection (abdomen) to the cervical lymph nodes which are nearer to the respiratory tract and lungs. QBECO remained in the abdominal region, with lymphatic trafficking to the inguinal lymph nodes, which are nearer to GI-proximal tissues/organs. Immune infiltration at the site of injection comprised of neutrophils for both SSIs, and macrophages for only QBKPN. CONCLUSION: Radiolabeling of SSIs allows for longitudinal in vivo imaging of biodistribution and trafficking. PET imaging revealed differential biodistribution of the SSIs based on the organotropism of the bacteria from which the SSI is derived. Trafficking from the site of injection to the targeted site is in part mediated via the lymphatics and involves macrophages and neutrophils.

pUdOs: Concise Plasmids for Bacterial and Mammalian Cells.

The plasmids from the Université d'Ottawa (pUdOs) are 28 small plasmids each comprising one of four origins of replication and one of seven selection markers, which together afford flexible use in Escherichia coli and several related gram-negative bacteria. The promoterless multicloning site is insulated from upstream spurious promoters by strong transcription terminators and contains type IIP or IIS restriction sites for conventional or Golden Gate cloning. pUdOs can be converted into efficient expression vectors through the insertion of a promoter at the user's discretion. For example, we demonstrate the utility of pUdOs as the backbone for an improved version of a Type III Secretion System reporter in Shigella. In addition, we derive a series of pUdO-based mammalian expression vectors, affording distinct levels of expression and transfection efficiency comparable to commonly used mammalian expression plasmids. Thus, pUdOs could advantageously replace traditional plasmids in a wide variety of cell types and applications.

Core-multi-shell design: unlocking multimodal capabilities in lanthanide-based nanoparticles as upconverting, T2-weighted MRI and CT probes.

Multimodal bioimaging probes merging optical imaging, magnetic resonance imaging (MRI), and X-ray computed tomography (CT) capabilities have attracted considerable attention due to their potential biomedical applications. Lanthanide-based nanoparticles are promising candidates for multimodal imaging because of their optical, magnetic and X-ray attenuation properties. We prepared a set of hexagonal-phase (ß)-NaGdF4:Yb,Er/NaGdF4/NaDyF4 core/shell/shell nanoparticles (Dy-CSS NPs) and demonstrated their optical/T2-weighted MRI/CT multimodal capabilities. A known drawback of multimodal probes that merge the upconverting Er3+/Yb3+ ion pair with magnetic Dy3+ ions for T2-weighted MRI is the loss of upconversion (UC) emission due to Dy3+ poisoning. Particular attention was paid to controlled nanoparticle architectures with tuned inner shell thicknesses separating Dy3+ and Er3+/Yb3+ to shed light on the distance-dependent loss of UC due to Yb3+ → Dy3+ energy transfer. Based on the Er3+ UC spectra and the excited state lifetime of Yb3+, a 4 nm thick NaGdF4 inner shell did not only restore but enhanced the UC emission. We further investigated the effect of the outer NaDyF4 shell thickness on the particles' magnetic and CT performance. MRI T2 relaxivity measurements in vitro at a magnetic field of 7 T performed on citrate-capped Dy-CSS NPs revealed that NPs with the thickest outer shell thickness (4 nm) exhibited the highest r2 value, with a superior T2 contrast effect compared to commercial iron oxide and other Dy-based T2 contrast agents. In addition, the citrate-capped Dy-CSS NPs were demonstrated suitable for CT in in vitro imaging phantoms at X-ray energies of 110 keV, rendering them interesting alternatives to clinically used iodine-based agents that operate at lower energies.

Cell-Impermeable Inhibitors Confirm That Intracellular Human Transglutaminase 2 Is Responsible for the Transglutaminase-Associated Cancer Phenotype.

Transglutaminase 2 (TG2) is a multifunctional enzyme primarily responsible for crosslinking proteins. Ubiquitously expressed in humans, TG2 can act either as a transamidase by crosslinking two substrates through formation of an Nε(ɣ-glutaminyl)lysine bond or as an intracellular G-protein. These discrete roles are tightly regulated by both allosteric and environmental stimuli and are associated with dramatic changes in the conformation of the enzyme. The pleiotropic nature of TG2 and multi-faceted activities have resulted in TG2 being implicated in numerous disease pathologies including celiac disease, fibrosis, and cancer. Targeted TG2 therapies have not been selective for subcellular localization, such that currently no tools exist to selectively target extracellular over intracellular TG2. Herein, we have designed novel TG2-selective inhibitors that are not only highly potent and irreversible, but also cell impermeable, targeting only extracellular TG2. We have also further derivatized the scaffold to develop probes that are intrinsically fluorescent or bear an alkyne handle, which target both intra- and extracellular TG2, in order to facilitate cellular labelling and pull-down assays. The fluorescent probes were internalized and imaged in cellulo, and provide the first implicit experimental evidence that by comparison with their cell-impermeable analogues, it is specifically intracellular TG2, and presumably its G-protein activity, that contributes to transglutaminase-associated cancer progression.

Direct mapping of kidney function by DCE-MRI urography using a tetrazinanone organic radical contrast agent.

Chronic kidney disease (CKD) and acute kidney injury (AKI) are ongoing global health burdens. Glomerular filtration rate (GFR) is the gold standard measure of kidney function, with clinical estimates providing a global assessment of kidney health without spatial information of kidney- or region-specific dysfunction. The addition of dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) to the anatomical imaging already performed would yield a 'one-stop-shop' for renal assessment in cases of suspected AKI and CKD. Towards urography by DCE-MRI, we evaluated a class of nitrogen-centered organic radicals known as verdazyls, which are extremely stable even in highly reducing environments. A glucose-modified verdazyl, glucoverdazyl, provided contrast limited to kidney and bladder, affording functional kidney evaluation in mouse models of unilateral ureteral obstruction (UUO) and folic acid-induced nephropathy (FAN). Imaging outcomes correlated with histology and hematology assessing kidney dysfunction, and glucoverdazyl clearance rates were found to be a reliable surrogate measure of GFR.

Silver nanoparticles with exceptional near-infrared absorbance for photoenhanced antimicrobial applications.

In this work, we outline a simple method for synthesizing decahedral and triangular silver nanoparticles using light to tune particle shape and spectral characteristics. Notably, we were able to generate triangular silver nanoparticles with exceptional absorbance in the near-infrared (NIR) region, with high spectral overlap with the biological window, making them particularly promising for biological applications. We further demonstrate that under complementary LED illumination, these excitable plasmonic particles display exceptional antibacterial properties, several orders of magnitude more potent than similar particles under dark conditions or under illumination that does not match particle absorbance. This work demonstrates the powerful effects that LED lights can have on the antibacterial activity of AgNPs, providing an inexpensive and easily implemented route to unlocking the full potential of AgNPs in photobiological applications.

16p11.2 haploinsufficiency reduces mitochondrial biogenesis in brain endothelial cells and alters brain metabolism in adult mice.

Neurovascular abnormalities in mouse models of 16p11.2 deletion autism syndrome are reminiscent of alterations reported in murine models of glucose transporter deficiency, including reduced brain angiogenesis and behavioral alterations. Yet, whether cerebrovascular alterations in 16p11.2df/+ mice affect brain metabolism is unknown. Here, we report that anesthetized 16p11.2df/+ mice display elevated brain glucose uptake, a phenomenon recapitulated in mice with endothelial-specific 16p11.2 haplodeficiency. Awake 16p11.2df/+ mice display attenuated relative fluctuations of extracellular brain glucose following systemic glucose administration. Targeted metabolomics on cerebral cortex extracts reveals enhanced metabolic responses to systemic glucose in 16p11.2df/+ mice that also display reduced mitochondria number in brain endothelial cells. This is not associated with changes in mitochondria fusion or fission proteins, but 16p11.2df/+ brain endothelial cells lack the splice variant NT-PGC-1α, suggesting defective mitochondrial biogenesis. We propose that altered brain metabolism in 16p11.2df/+ mice is compensatory to endothelial dysfunction, shedding light on previously unknown adaptative responses.

Strong Magnetic Field Annealing for Improved Phthalocyanine Organic Thin-Film Transistors.

Thin-film microstructure, morphology, and polymorphism can be controlled and optimized to improve the performance of carbon-based electronics. Thermal or solvent vapor annealing are common post-deposition processing techniques; however, it can be difficult to control or destructive to the active layer or substrates. Here, the use of a static, strong magnetic field (SMF) as a non-destructive process for the improvement of phthalocyanine (Pc) thin-film microstructure, increasing organic thin-film transistor (OTFTs) mobility by twofold, is demonstrated. Grazing incident wide-angle X-ray scattering (GIWAXS), X-ray diffraction (XRD), and atomic force microscopy (AFM) elucidate the effect of SMF on both para- and diamagnetic Pc thin-films when subjected to a magnetic field. A SMF is found to increase the concentration of oxygen-induced radical species within the Pc thin-film, lending a paramagnetic character to ordinarily diamagnetic metal-free Pc and resulting in magnetic field induced changes to its thin-film microstructures. In a nitrogen environment, without competing degradation effects of molecular oxygen, SMF processing is found to favorably improve charge transport characteristics and increase OTFT mobility. Thus, post-deposition thin-film annealing with a magnetic field is presented as an alternative and promising technique for future thin-film engineering applications.

Armeniaspirol analogues with more potent Gram-positive antibiotic activity show enhanced inhibition of the ATP-dependent proteases ClpXP and ClpYQ.

Antibiotics with fundamentally new mechanisms of action such as the armeniaspirols, which target the ATP-dependent proteases ClpXP and ClpYQ, must be developed to combat antimicrobial resistance. While the mechanism of action of armeniaspirol against Gram-positive bacteria is understood, little is known about the structure-activity relationship for its antibiotic activity. Based on the preliminary data showing that modifications of armeniaspirol's N-methyl group increased antibiotic potency, we probed the structure-activity relationship of N-alkyl armeniaspirol derivatives. A series of focused derivatives were synthesized and evaluated for antibiotic activity against clinically relevant pathogens including methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus. Replacement of the N-methyl with N-hexyl, various N-benzyl, and N-phenethyl substituents led to substantial increases in antibiotic activity and potency for inhibition of both ClpYQ and ClpXP. Docking studies identified binding models for ClpXP and ClpYQ that were consistent with the inhibition data. This work confirms the role of ClpXP and ClpYQ in the mechanism of action of armeniaspirol and provides important lead compounds for further antibiotic development.

A prodrug strategy for the in vivo imaging of aldehyde dehydrogenase activity.

Therapy resistance is one of the biggest challenges facing clinical oncology. Despite a revolution in new anti-cancer drugs targeting multiple components of the tumour microenvironment, acquired or innate resistance frequently blunts the efficacy of these treatments. Non-invasive identification of drug-resistant tumours will enable modification of the patient treatment pathway through the selection of appropriate second-line treatments. Here, we have designed a prodrug radiotracer for the non-invasive imaging of aldehyde dehydrogenase 1A1 (ALDH1A1) activity. Elevated ALDH1A1 activity is a marker of drug-resistant cancer cells, modelled here with matched cisplatin-sensitive and -resistant human SKOV3 ovarian cancer cells. The aromatic aldehyde of our prodrug radiotracer was intracellularly liberated by esterase cleavage of the geminal diacetate and specifically trapped by ALDH through its conversion to the charged carboxylic acid. Through this mechanism of action, ALDH-specific retention of our prodrug radiotracer in the drug-resistant tumour cells was twice as high as the drug-sensitive cells. Acylal masking of the aldehyde afforded a modest protection from oxidation in the blood, which was substantially improved in carrier-added experiments. In vivo positron emission tomography imaging of tumour-bearing mice produced high tumour-to-background images and radiotracer uptake in high ALDH-expressing organs but was unable to differentiate between drug-sensitive and drug-resistant tumours. Alternative strategies to protect the labile aldehyde are currently under investigation.

Phytoglycogen Encapsulation of Lanthanide-Based Nanoparticles as an Optical Imaging Platform with Therapeutic Potential.

Lanthanide-based upconverting nanoparticles (UCNPs) are largely sought-after for biomedical applications ranging from bioimaging to therapy. A straightforward strategy is proposed here using the naturally sourced polymer phytoglycogen to coencapsulate UCNPs with hydrophobic photosensitizers as an optical imaging platform and light-induced therapeutic agents. The resulting multifunctional sub-micrometer-sized luminescent beads are shown to be cytocompatible as carrier materials, which encourages the assessment of their potential in biomedical applications. The loading of UCNPs of various elemental compositions enables multicolor hyperspectral imaging of the UCNP-loaded beads, endowing these materials with the potential to serve as luminescent tags for multiplexed imaging or simultaneous detection of different moieties under near-infrared (NIR) excitation. Coencapsulation of UCNPs and Rose Bengal opens the door for potential application of these microcarriers for collagen crosslinking. Alternatively, coloading UCNPs with Chlorin e6 enables NIR-light triggered generation of reactive oxygen species. Overall, the developed encapsulation methodology offers a straightforward and noncytotoxic strategy yielding water-dispersible UCNPs while preserving their bright and color-tunable upconversion emission that would allow them to fulfill their potential as multifunctional platforms for biomedical applications.

The Need to Pair Molecular Monitoring Devices with Molecular Imaging to Personalize Health.

By enabling the non-invasive monitoring and quantification of biomolecular processes, molecular imaging has dramatically improved our understanding of disease. In recent years, non-invasive access to the molecular drivers of health versus disease has emboldened the goal of precision health, which draws on concepts borrowed from process monitoring in engineering, wherein hundreds of sensors can be employed to develop a model which can be used to preventatively detect and diagnose problems. In translating this monitoring regime from inanimate machines to human beings, precision health posits that continual and on-the-spot monitoring are the next frontiers in molecular medicine. Early biomarker detection and clinical intervention improves individual outcomes and reduces the societal cost of treating chronic and late-stage diseases. However, in current clinical settings, methods of disease diagnoses and monitoring are typically intermittent, based on imprecise risk factors, or self-administered, making optimization of individual patient outcomes an ongoing challenge. Low-cost molecular monitoring devices capable of on-the-spot biomarker analysis at high frequencies, and even continuously, could alter this paradigm of therapy and disease prevention. When these devices are coupled with molecular imaging, they could work together to enable a complete picture of pathogenesis. To meet this need, an active area of research is the development of sensors capable of point-of-care diagnostic monitoring with an emphasis on clinical utility. However, a myriad of challenges must be met, foremost, an integration of the highly specialized molecular tools developed to understand and monitor the molecular causes of disease with clinically accessible techniques. Functioning on the principle of probe-analyte interactions yielding a transducible signal, probes enabling sensing and imaging significantly overlap in design considerations and targeting moieties, however differing in signal interpretation and readout. Integrating molecular sensors with molecular imaging can provide improved data on the personal biomarkers governing disease progression, furthering our understanding of pathogenesis, and providing a positive feedback loop toward identifying additional biomarkers and therapeutics. Coupling molecular imaging with molecular monitoring devices into the clinical paradigm is a key step toward achieving precision health.

Surface engineering of zinc phthalocyanine organic thin-film transistors results in part-per-billion sensitivity towards cannabinoid vapor.

Phthalocyanine-based organic thin-film transistors (OTFTs) have been demonstrated as sensors for a range of analytes, including cannabinoids, in both liquid and gas phases. Detection of the primary cannabinoids, Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD), is necessary for quality control and regulation, however, current techniques are often not readily available for consumers, industry, and law-enforcement. The OTFT characteristics, X-ray diffraction (XRD) spectra, and grazing incident wide angle x-ray scattering (GIWAXS) spectra of two copper and three zinc phthalocyanines, with varying degrees of peripheral fluorination, were screened to determine sensitivity to THC vapor. Unsubstituted ZnPc was found to be the most sensitive material and, by tuning thin-film morphology, crystal polymorphs, and thickness through altered physical vapor deposition conditions, we increased the sensitivity to THC by 100x. Here we demonstrate that deposition conditions, and the resulting physical film characteristics, play a significant role in device sensitization.

Mapping vitamin B6 metabolism by hydrazoCEST magnetic resonance imaging.

A new CEST-MRI contrast agent, 2-HYNIC, capable of sensing aromatic aldehydes is reported. Pyridoxal 5'-phosphate, a key Vitamin B6 metabolite necessary for >140 biotransformations was mapped by CEST-MRI in vitro and in vivo in lung cancer. 2-HYNIC provided access to this key biomarker associated with a variety of human diseases.

Synthesis of natural/13C-enriched d-tagatose from natural/13C-enriched d-fructose.

A concise, easily scalable synthesis of a rare ketohexose, d-tagatose, was developed, that is compatible with the preparation of d-[UL-13C6]tagatose. Epimerization of the widely available and inexpensive ketohexose d-fructose at the C-4 position via an oxidation/reduction (Dess-Martin periodinane/NaBH4) was a key step in the synthesis. Overall, fully protected natural d-tagatose (3.21 g) was prepared from d-fructose (9 g) on a 50 mmol scale in 23% overall yield, after five steps and two chromatographic purifications. d-[UL-13C6]Tagatose (92 mg) was prepared from d-[UL-13C6]fructose (465 mg, 2.5 mmol) in 16% overall yield after six steps and four chromatographic purifications.

Thin-Film Engineering of Solution-Processable n-Type Silicon Phthalocyanines for Organic Thin-Film Transistors.

Metal and metalloid phthalocyanines are an abundant and established class of materials widely used in the dye and pigment industry as well as in commercial photoreceptors. Silicon phthalocyanines (SiPcs) are among the highest-performing n-type semiconductor materials in this family when used in organic thin-film transistors (OTFTs) as their performance and solid-state arrangement are often increased through axial substitution. Herein, we study eight axially substituted SiPcs and their integration into solution-processed n-type OTFTs. Electrical characterization of the OTFTs, combined with atomic force microscopy (AFM), determined that the length of the alkyl chain affects device performance and thin-film morphology. The effects of high-temperature annealing and spin coating time on film formation, two key processing steps for fabrication of OTFTs, were investigated by grazing-incidence wide-angle X-ray scattering (GIWAXS) and X-ray diffraction (XRD) to elucidate the relationship between thin-film microstructure and device performance. Thermal annealing was shown to change both film crystallinity and SiPc molecular orientation relative to the substrate surface. Spin time affected film crystallinity, morphology, and interplanar d-spacing, thus ultimately modifying device performance. Of the eight materials studied, bis(tri-n-butylsilyl oxide) SiPc exhibited the greatest electron field-effect mobility (0.028 cm2 V-1 s-1, a threshold voltage of 17.6 V) of all reported solution-processed SiPc derivatives.

Engineering Cannabinoid Sensors through Solution-Based Screening of Phthalocyanines.

Organic thin-film transistors (OTFTs) have shown promise for a range of sensing applications, with phthalocyanine-based OTFTs demonstrated as sensors for atmospheric parameters, volatile gases, and small organic molecules including cannabinoids. However, the process of fabricating, testing, and optimizing OTFTs in a laboratory setting requires highly specialized equipment, materials, and expertise. To determine if sensor development can be expedited and thus reduce manufacturing burden, spectroelectrochemistry is applied to rapidly screen for molecular interactions between metal-free phthalocyanines and a variety of metal phthalocyanines (MPcs) and the cannabinoids Δ9-tetrahydrocannabinol (THC) or cannabidiol (CBD), with and without a cannabinoid-sensitive chromophore (Fast Blue BB). Spectral analyses are corroborated by 2D-NMR and related to measured OTFT performance. Spectroelectrochemical changes to the Q band region of the phthalocyanine spectra in the presence of analytes can be used to predict the response of OTFTs. Thus, with spectroelectrochemistry, a range of potential materials for OTFT small organic molecule-sensing applications can be quickly analyzed, and phthalocyanines with a preferred response can be selected.

On-the-Spot Detection and Speciation of Cannabinoids Using Organic Thin-Film Transistors.

Quality control is imperative for Cannabis since the primary cannabinoids, Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD), elicit very different pharmacological effects. THC/CBD ratios are currently determined by techniques not readily accessible by consumers or dispensaries and which are impractical for use in the field by law-enforcement agencies. CuPc- and F16-CuPc-based organic thin-film transistors have been combined with a cannabinoid-sensitive chromophore for the detection and differentiation of THC and CBD. The combined use of these well-characterized and inexpensive p- and n-type materials afforded the determination of the CBD/THC ratio from rapid plant extracts, with results indistinguishable from high-pressure liquid chromatography. Analysis of the prepyrolyzed sample accurately predicted postpyrolysis THC/CBD, which ultimately influences the psychotropic and medicinal effects of the specific plant. The devices were also capable of vapor-phase sensing, producing a unique electrical output for THC and CBD relative to other potentially interfering vaporized organic products. The analysis of complex medicinal plant extracts and vapors, normally reserved for advanced analytical infrastructure, can be achieved with ease, at low cost, and on the spot, using organic thin-film transistors.