Automated Synthesis of (rac)‐, (R)‐, and (S)‐[18F]Epifluorohydrin and Their Application for Developing PET Radiotracers Containing a 3‐[18F]Fluoro‐2‐hydroxypropyl Moiety

To introduce the 3‐[¹⁸F]fluoro‐2‐hydroxypropyl moiety into positron emission tomography (PET) radiotracers, we performed automated synthesis of (rac)‐, (R)‐, and (S)‐[¹⁸F]epifluorohydrin ([¹⁸F] 1 ) by nucleophilic displacement of (rac)‐, (R)‐, or (S)‐glycidyl tosylate with ¹⁸F^? and purification by distillation. The ring‐opening reaction of (R)‐ or (S)‐[¹⁸F] 1 with phenol precursors gave enantioenriched [¹⁸F]fluoroalkylated products without racemisation. We then synthesised (rac)‐, (R)‐, and (S)‐ 2‐{5‐[4‐(3‐[¹⁸F]fluoro‐2‐hydroxypropoxy)phenyl]‐2‐oxobenzo[d]oxazol‐3(2H)‐yl}‐N‐methyl‐N‐phenylacetamide ([¹⁸F] 6 ) as novel radiotracers for the PET imaging of translocator protein (18 kDa) and showed that (R)‐ and (S)‐[¹⁸F] 6 had different radioactivity uptake in mouse bone and liver. Thus, (rac)‐, (R)‐, and (S)‐[¹⁸F] 1 are effective radiolabelling reagents and can be used to develop PET radiotracers by examining the effects of chirality on their in vitro binding affinities and in vivo behaviour.     

Comparison of in Silico,Electrochemical, in Vitro and in Vivo Metabolism of a Homologous Series of (Radio)fluorinated σ1 Receptor Ligands Designed for Positron Emission Tomography

The imaging of σ₁ receptors in the brain by fluorinated radiotracers will be used for the validation of σ₁ receptors as drug targets as well as for differential diagnosis of diseases in the central nervous system. The biotransformation of four homologous fluorinated PET tracers 1′‐benzyl‐3‐(ω‐fluoromethyl to ω‐fluorobutyl)‐3H‐spiro[2]benzofuran‐1,4′‐piperidine] ([¹⁸F] 1 – 4 ) was investigated. In silico studies using fast metabolizer (FAME) software, electrochemical oxidations, in vitro studies with rat liver microsomes, and in vivo metabolism studies after application of the PET tracers [¹⁸F] 1 – 4 to mice were performed. Combined liquid chromatography and mass spectrometry (HPLC–MS) analysis allowed structural identification of non‐radioactive metabolites. Radio‐HPLC and radio‐TLC provided information about the presence of unchanged parent radiotracers and their radiometabolites. Radiometabolites were not found in the brain after application of [¹⁸F] 2 – 4 , but liver, plasma, and urine samples contained several radiometabolites. Less than 2 % of the injected dose of [¹⁸F] 4 reached the brain, rendering [¹⁸F] 4 less appropriate as a PET tracer than [¹⁸F] 2 and [¹⁸F] 3 . Compounds [¹⁸F] 2 and [¹⁸F] 3 possess the most promising properties for imaging of σ₁ receptors in the brain. High σ₁ affinity (K_i=0.59 nm ), low lipophilicity (logD_7.4=2.57), high brain penetration (4.6 % of injected dose after 30 min), and the absence of radiometabolites in the brain favor the fluoroethyl derivative [¹⁸F] 2 slightly over the fluoropropyl derivative [¹⁸F] 3 for human use.     

Preclinical Evaluation of [18F]FACH in Healthy Mice and Piglets: An 18F-Labeled Ligand for Imaging of Monocarboxylate Transporters with PET

The expression of monocarboxylate transporters (MCTs) is linked to pathophysiological changes in diseases, including cancer, such that MCTs could potentially serve as diagnostic markers or therapeutic targets. We recently developed [¹⁸F]FACH as a radiotracer for non-invasive molecular imaging of MCTs by positron emission tomography (PET). The aim of this study was to evaluate further the specificity, metabolic stability, and pharmacokinetics of [¹⁸F]FACH in healthy mice and piglets. We measured the [¹⁸F]FACH plasma protein binding fractions in mice and piglets and the specific binding in cryosections of murine kidney and lung. The biodistribution of [¹⁸F]FACH was evaluated by tissue sampling ex vivo and by dynamic PET/MRI in vivo, with and without pre-treatment by the MCT inhibitor α-CCA-Na or the reference compound, FACH-Na. Additionally, we performed compartmental modelling of the PET signal in kidney cortex and liver. Saturation binding studies in kidney cortex cryosections indicated a K_D of 118 ± 12 nM and B_max of 6.0 pmol/mg wet weight. The specificity of [¹⁸F]FACH uptake in the kidney cortex was confirmed in vivo by reductions in AUC_0–60min after pre-treatment with α-CCA-Na in mice (−47%) and in piglets (−66%). [¹⁸F]FACH was metabolically stable in mouse, but polar radio-metabolites were present in plasma and tissues of piglets. The [¹⁸F]FACH binding potential (BP_ND) in the kidney cortex was approximately 1.3 in mice. The MCT1 specificity of [¹⁸F]FACH uptake was confirmed by displacement studies in 4T1 cells. [¹⁸F]FACH has suitable properties for the detection of the MCTs in kidney, and thus has potential as a molecular imaging tool for MCT-related pathologies, which should next be assessed in relevant disease models.     

Monitoring Therapeutic Responses to Silicified Cancer Cell Immunotherapy Using PET/MRI in a Mouse Model of Disseminated Ovarian Cancer

Current imaging approaches used to monitor tumor progression can lack the ability to distinguish true progression from pseudoprogression. Simultaneous metabolic 2-deoxy-2-[¹⁸F]fluoro-D-glucose ([¹⁸F]FDG) positron emission tomography (PET) and magnetic resonance imaging (MRI) offers new opportunities to overcome this challenge by refining tumor identification and monitoring therapeutic responses to cancer immunotherapy. In the current work, spatial and quantitative analysis of tumor burden were performed using simultaneous [¹⁸F]FDG-PET/MRI to monitor therapeutic responses to a novel silicified cancer cell immunotherapy in a mouse model of disseminated serous epithelial ovarian cancer. Tumor progression was validated by bioluminescence imaging of luciferase expressing tumor cells, flow cytometric analysis of immune cells in the tumor microenvironment, and histopathology. While PET demonstrated the presence of metabolically active cancer cells through [¹⁸F]FDG uptake, MRI confirmed cancer-related accumulation of ascites and tissue anatomy. This approach provides complementary information on disease status without a confounding signal from treatment-induced inflammation. This work provides a possible roadmap to facilitate accurate monitoring of therapeutic responses to cancer immunotherapies.     

MicroPET Imaging Assessment of Brain Tau and Amyloid Deposition in 6 × Tg Alzheimer’s Disease Model Mice

Alzheimer’s disease (AD) is characterized by the deposition of extracellular amyloid plaques and intracellular accumulation of neurofibrillary tangles (NFT). Amyloid beta (Aβ) and tau imaging are widely used for diagnosing and monitoring AD in clinical settings. We evaluated the pathology of a recently developed 6 × Tg − AD (6 × Tg) mouse model by crossbreeding 5 × FAD mice with mice expressing mutant (P301L) tau protein using micro-positron emission tomography (PET) image analysis. PET studies were performed in these 6 × Tg mice using [¹⁸F]Flutemetamol, which is an amyloid PET radiotracer; [¹⁸F]THK5351 and [¹⁸F]MK6240, which are tau PET radiotracers; moreover, [¹⁸F]DPA714, which is a translocator protein (TSPO) radiotracer, and comparisons were made with age-matched mice of their respective parental strains. We compared group differences in standardized uptake value ratio (SUVR), kinetic parameters, biodistribution, and histopathology. [¹⁸F]Flutemetamol images showed prominent cortical uptake and matched well with 6E10 staining images from 2-month-old 6 × Tg mice. [¹⁸F]Flutemetamol images showed a significant correlation with [¹⁸F]DPA714 in the cortex and hippocampus. [¹⁸F]THK5351 images revealed prominent hippocampal uptake and matched well with AT8 immunostaining images in 4-month-old 6 × Tg mice. Moreover, [¹⁸F]THK5351 images were confirmed using [¹⁸F]MK6240, which revealed significant correlations in the cortex and hippocampus. Uptake of [¹⁸F]THK5351 or [¹⁸F]MK6240 was highly correlated with [¹⁸F]Flutemetamol in 4-month-old 6 × Tg mice. In conclusion, PET imaging revealed significant age-related uptake of Aβ, tau, and TSPO in 6 × Tg mice, which was highly correlated with age-dependent pathology.     

PET/CT Imaging and Physiology of Mice on High Protein Diet

Background: High protein (HP) diets have been proposed to reduce body weight in humans. The diets are known to alter energy metabolism, which can affect the quality of [¹⁸F]FDG PET heart images. In this preclinical study, we therefore explore the impact of a prolonged HP diet on myocardial [¹⁸F]FDG uptake. Methods: C57BL/6J (Black six (Bl6)) and apolipoprotein E-deficient (apoE^−/−) mice were fed chow, a HP diet, or a low protein (LP) diet for 12 weeks. At baseline and after treatment, the animals were injected with 33.0 MBq of [¹⁸F]FDG and a 30 min PET/CT scan was made. Myocardial volume and [¹⁸F]FDG uptake were quantified using PET and the % of body fat was calculated from CT. Results: Myocardial [¹⁸F]FDG uptake was similar for all diets at the follow-up scan but an increase between baseline and follow-up scans was noticed in the LP groups. Myocardial volume was significantly smaller in the C57BL HP group compared to the other Bl6 groups. Body weight increased less in the two HP groups compared to the chow and LP groups. Body fat percentage was significantly higher in the LP groups. This effect was stronger in C57BL mice (28.7%) compared to apoE^−/− mice (15.1%). Conclusions: Myocardial uptake of [¹⁸F]FDG in mice is not affected by increased protein intake but [¹⁸F]FDG uptake increases when the amount of protein is lowered. A lower body weight and percentage of body fat were noticed when applying a HP diet.     

[18F]PRIMATX,a New Positron Emission Tomography Tracer for Imaging of Autotaxin in Lung Tissue and Tumor-Bearing Mice

Autotaxin (ATX) is a secreted enzyme with tissue levels associated with tissue injury, which increase during wound healing and chronic fibrotic diseases. We selected [¹⁸F](R,E)-3-(4-chloro-2-((5-methyl-2H-tetrazol-2-yl)methyl)phenyl)-1-(4-((5-(2-fluoroethoxy)pyridin-2-yl)methyl)-2-methylpiperazin-1-yl)prop-2-en-1-one ([¹⁸F]PRIMATX, [¹⁸F] 2 ), a tracer for positron emission tomography, to image ATX expression in vivo. It successfully differentiates expression levels in lung tissue samples from idiopathic pulmonary fibrosis patients, and allows the detection of ATX-expressing tumors in living mice, confirming its potential for development as a clinical imaging agent.     

18F Site-Specific Labelling of a Single-Chain Antibody against Activated Platelets for the Detection of Acute Thrombosis in Positron Emission Tomography

Positron emission tomography is the imaging modality of choice when it comes to the high sensitivity detection of key markers of thrombosis and inflammation, such as activated platelets. We, previously, generated a fluorine-18 labelled single-chain antibody (scFv) against ligand-induced binding sites (LIBS) on activated platelets, binding it to the highly abundant platelet glycoprotein integrin receptor IIb/IIIa. We used a non-site-specific bio conjugation approach with N-succinimidyl-4-[¹⁸F]fluorobenzoate (S[¹⁸F]FB), leading to a mixture of products with reduced antigen binding. In the present study, we have developed and characterised a novel fluorine-18 PET radiotracer, based on this antibody, using site-specific bio conjugation to engineer cysteine residues with N-[2-(4-[¹⁸F]fluorobenzamido)ethyl]maleimide ([¹⁸F]FBEM). ScFv_anti-LIBS and control antibody mut-scFv, with engineered C-terminal cysteine, were reduced, and then, they reacted with N-[2-(4-[¹⁸F]fluorobenzamido)ethyl]maleimide ([¹⁸F]FBEM). Radiolabelled scFv was injected into mice with FeCl₃-induced thrombus in the left carotid artery. Clots were imaged in a PET MR imaging system, and the amount of radioactivity in major organs was measured using an ionisation chamber and image analysis. Assessment of vessel injury, as well as the biodistribution of the radiolabelled scFv, was studied. In the in vivo experiments, we found uptake of the targeted tracer in the injured vessel, compared with the non-injured vessel, as well as a high uptake of both tracers in the kidney, lung, and muscle. As expected, both tracers cleared rapidly via the kidney. Surprisingly, a large quantity of both tracers was taken up by organs with a high glutathione content, such as the muscle and lung, due to the instability of the maleimide cysteine bond in vivo, which warrants further investigations. This limits the ability of the novel antibody radiotracer ¹⁸F-scFv_anti-LIBS to bind to the target in vivo and, therefore, as a useful agent for the sensitive detection of activated platelets. We describe the first fluorine-18 variant of the scFv_anti-LIBS against activated platelets using site-specific bio conjugation.     

Cannabidiol Treatment Improves Glucose Metabolism and Memory in Streptozotocin-Induced Alzheimer’s Disease Rat Model: A Proof-of-Concept Study

An early and persistent sign of Alzheimer’s disease (AD) is glucose hypometabolism, which can be evaluated by positron emission tomography (PET) with ¹⁸F-2-fluoro-2-deoxy-D-glucose ([¹⁸F]FDG). Cannabidiol has demonstrated neuroprotective and anti-inflammatory properties but has not been evaluated by PET imaging in an AD model. Intracerebroventricular (icv) injection of streptozotocin (STZ) is a validated model for hypometabolism observed in AD. This proof-of-concept study evaluated the effect of cannabidiol treatment in the brain glucose metabolism of an icv-STZ AD model by PET imaging. Wistar male rats received 3 mg/kg of STZ and [¹⁸F]FDG PET images were acquired before and 7 days after STZ injection. Animals were treated with intraperitoneal cannabidiol (20 mg/kg—STZ–cannabidiol) or saline (STZ–saline) for one week. Novel object recognition was performed to evaluate short-term and long-term memory. [¹⁸F]FDG uptake in the whole brain was significantly lower in the STZ–saline group. Voxel-based analysis revealed a hypometabolism cluster close to the lateral ventricle, which was smaller in STZ–cannabidiol animals. The brain regions with more evident hypometabolism were the striatum, motor cortex, hippocampus, and thalamus, which was not observed in STZ–cannabidiol animals. In addition, STZ–cannabidiol animals revealed no changes in memory index. Thus, this study suggests that cannabidiol could be an early treatment for the neurodegenerative process observed in AD.     

A Series of 18F‐Labelled Pyridinylphenyl Amides as Subtype‐Selective Radioligands for the Dopamine D3 Receptor

Synthesis, biological activity, and structure–selectivity relationship (SSR) studies of a novel series of potential dopamine D3 receptor radioligands as imaging agents for positron emission tomography (PET) are reported. Considering a structurally diverse library of D3 ligands, SSR studies were performed for a new series of fluorinated pyridinylphenyl amides using CoMFA and CoMSIA methods. The in vitro D3 affinities of the predicted series of biphenyl amide ligands 9 a – d revealed single‐digit to sub‐nanomolar potencies (K_i=0.52–1.6 nM ), displaying excellent D3 selectivity over the D2 subtype of 110‐ to 210‐fold for the test compounds 9 a – c . Radiofluorination by nucleophilic substitution of Br or NO₂ by ¹⁸F led to radiochemical yields of 66–92 % for [¹⁸F] 9 a – d . However, the specific activities of [¹⁸F] 9 b and [¹⁸F] 9 d were insufficient, rendering their use for in vivo studies impossible. Biodistribution studies of [¹⁸F] 9 a and [¹⁸F] 9 c using rat brain autoradiography revealed accumulation in the ventricles, thus indicating insufficient biokinetic properties of [¹⁸F] 9 a and [¹⁸F] 9 c for D3 receptor imaging in vivo.     

11C‐ and 18F‐Labeled Radioligands for P‐Glycoprotein Imaging by Positron Emission Tomography

P‐Glycoprotein (P‐gp) is an efflux transporter widely expressed at the human blood–brain barrier. It is involved in xenobiotics efflux and in onset and progression of neurodegenerative disorders. For these reasons, there is great interest in the assessment of P‐gp expression and function by noninvasive techniques such as positron emission tomography (PET). Three radiolabeled aryloxazole derivatives: 2‐[2‐(2‐methyl‐(¹¹C)‐5‐methoxyphenyl)oxazol‐4‐ylmethyl]‐6,7‐dimethoxy‐1,2,3,4‐tetrahydroisoquinoline ([¹¹C]‐ 5 ); 2‐[2‐(2‐fluoromethyl‐(¹⁸F)‐5‐methoxyphenyl)oxazol‐4‐ylmethyl]‐6,7‐dimethoxy‐1,2,3,4‐tetra‐hydroisoquinoline ([¹⁸F]‐ 6 ); and 2‐[2‐(2‐fluoroethyl‐(¹⁸F)‐5‐methoxyphenyl)oxazol‐4‐ylmethyl]‐6,7‐dimethoxy‐1,2,3,4‐tetrahydroisoquinoline ([¹⁸F]‐ 7 ), were tested in several in vitro biological assays to assess the effect of the aryl substituent in terms of potency and mechanism of action toward P‐gp. Methyl derivative [¹¹C]‐ 5 is a potent P‐gp substrate, whereas the corresponding fluoroethyl derivative [¹⁸F]‐ 7 is a P‐gp inhibitor. Fluoromethyl compound [¹⁸F]‐ 6 is classified as a non‐transported P‐gp substrate, because its efflux increases after cyclosporine A modulation. These studies revealed a promising substrate and inhibitor, [¹¹C]‐ 5 and [¹⁸F]‐ 7 , respectively, for in vivo imaging of P‐gp by using PET.     

Synthesis and Pharmacological Evaluation of Fluorinated Quinoxaline-Based κ-Opioid Receptor (KOR) Agonists Designed for PET Studies

κ-Opioid receptors (KORs) play a predominant role in pain alleviation, itching skin diseases, depression and neurodegenerative disorders such as multiple sclerosis. Therefore, imaging of KOR by a fluorinated PET tracer was envisaged. Two strategies were followed to introduce a F atom into the very potent class of cis,trans-configured perhydroquinoxalines. Whereas the synthesis of fluoroethyltriazole 2 has already been reported, fluoropyrrolidines 14 (1-[2-(3,4-dichlorophenyl)acetyl]-8-[(R)-3-fluoropyrrolidin-1-yl]-perhydroquinoxalines) were prepared by S_N2 substitution of a cyclic sulfuric acid derivative with hydroxypyrrolidine and subsequent transformation of the OH moiety into a F substituent. Fluoropyrrolidines 14 showed similar low-nanomolar KOR affinity and selectivity to the corresponding pyrrolidines, but the corresponding alcohols were slightly less active. In the cAMP and β-arrestin assay, 14b (proton at the 4-position) exhibited similar KOR agonistic activity as U-50,488. The fluoro derivatives 14b and 14c (CO₂CH₃ at the 4-position) revealed KOR-mediated anti-inflammatory activity as CD11c and the IFN-γ production were reduced significantly in mouse and human dendritic cells. Compounds 14b and 14-c also displayed anti-inflammatory and immunomodulatory activity in mouse and human T cells. The PET tracer [¹⁸F]- 2 was prepared by 1,3-dipolar cycloaddition. In vivo, [¹⁸F]- 2 did not label KOR due to very fast elimination kinetics. Nucleophilic substitution of a mesylate precursor provided [¹⁸F]- 14c . Unfortunately, defluorination of [¹⁸F]- 14c occurred in vivo, which was analyzed in detail by in vitro studies.     

Fluorine‐18 Radiolabeling and Radiopharmacological Characterization of a Benzodioxolylpyrimidine‐based Radiotracer Targeting the Receptor Tyrosine Kinase EphB4

Members of the Eph receptor tyrosine kinase family play essential roles in the pathogenesis of cancer and are therefore promising candidates for molecular imaging by positron emission tomography (PET), for example. In this regard, radiochemical access to novel PET radiotracers derived from potent inhibitors that target the EphB4 kinase domain and which bear a benzodioxolylpyrimidine structural motif was developed. A synthetic route was established for a new fluorine‐18‐containing radiotracer and for the desired precursor based on a high‐affinity benzodioxolylpyrimidine receptor tyrosine kinase inhibitor lead structure. The radiotracer [¹⁸F] 15 was obtained in 16 % radiochemical yield with a specific activity of ～7 GBq μmol^?1 and >95 % radiochemical purity. Due to the implication of EphB4, particularly in the progression, angiogenesis, and metastasis of melanoma, EphB4‐overexpressing human melanoma cells were generated and used as a novel in vitro model for radiopharmacological evaluation of the radiotracer. We demonstrate that the corresponding non‐radioactive reference compound regained its functionality as an inhibitor for both EphB4 receptor tyrosine kinase and Src kinase. EphB4 was significantly inhibited at compound concentrations >1 μM . Cellular uptake studies with [¹⁸F] 15 revealed substantial uptake in both EphB4‐overexpressing and control cells. Moreover, NMRI nu/nu mice bearing both EphB4‐overexpressing tumors and control tumors were used for radiopharmacological characterization by biodistribution studies ex vivo and by dynamic small‐animal PET experiments in vivo. Despite the high metabolic stability of the novel radiotracer observed in vivo, no substantial binding or accumulation in EphB4‐overexpressing and control tumors was observed. Nevertheless, we point out that the approach presented herein gives convenient access to novel ¹⁸F‐labeled benzodioxolylpyrimidines and is a promising strategy for the further development of novel radiotracers for imaging Eph receptor tyrosine kinases in cancer.     

Radiofluorinated Pyrimidine‐2,4,6‐triones as Molecular Probes for Noninvasive MMP‐Targeted Imaging

Matrix metalloproteinases (MMPs) are zinc‐ and calcium‐dependent endopeptidases. Representing a subfamily of the metzincin superfamily, MMPs are involved in the proteolytic degradation of components of the extracellular matrix. Unregulated MMP expression, MMP dysregulation and locally increased MMP activity are common features of various diseases, such as cancer, atherosclerosis, stroke, arthritis, and others. Therefore, activated MMPs are suitable biological targets for the specific visualization of such pathologies, in particular by using radiolabeled MMP inhibitors (MMPIs). The aim of this work was to develop a radiofluorinated molecular probe for noninvasive in vivo imaging for the detection of up‐regulated levels of activated MMPs in the living organism. Fluorinated MMPIs ( 26 , 31 and 38 ) based on the pyrimidine‐2,4,6‐trione lead structure RO 28‐2653 ( 1 ) were synthesized, and their MMP inhibition potency was evaluated in vitro. The radiosynthesis and the in vivo biodistribution of the first ¹⁸F‐labeled prototype, MMP‐targeted tracer [¹⁸F] 26 , suitable for molecular imaging by means of positron emission tomography (PET) were realized.     

FOXM1 Inhibitors as Potential Diagnostic Agents: First Generation of a PET Probe Targeting FOXM1 To Detect Triple-Negative Breast Cancer in vitro and in vivo

The FOXM1 protein controls the expression of essential genes related to cancer cell cycle progression, metastasis, and chemoresistance. We hypothesize that FOXM1 inhibitors could represent a novel approach to develop ¹⁸F-based radiotracers for Positron Emission Tomography (PET). Therefore, in this report we describe the first attempt to use ¹⁸F-labeled FOXM1 inhibitors to detect triple-negative breast cancer (TNBC). Briefly, we replaced the original amide group in the parent drug FDI-6 for a ketone group in the novel AF-FDI molecule, to carry out an aromatic nucleophilic (¹⁸F)-fluorination. AF-FDI dissociated the FOXM1-DNA complex, decreased FOXM1 levels, and inhibited cell proliferation in a TNBC cell line (MDA-MB-231). [¹⁸F]AF-FDI was internalized in MDA-MB-231 cells. Cell uptake inhibition experiments showed that AF-FDI and FDI-6 significantly decreased the maximum uptake of [¹⁸F]AF-FDI, suggesting specificity towards FOXM1. [¹⁸F]AF-FDI reached a tumor uptake of SUV=0.31 in MDA-MB-231 tumor-bearing mice and was metabolically stable 60 min post-injection. These results provide preliminary evidence supporting the potential role of FOXM1 to develop PET radiotracers.     

Harnessing the Maltodextrin Transport Mechanism for Targeted Bacterial Imaging: Structural Requirements for Improved in vivo Stability in Tracer Design

Diagnosis and localization of bacterial infections remains a significant clinical challenge. Harnessing bacteria‐specific metabolic pathways, such as the maltodextrin transport mechanism, may allow specific localization and imaging of small or hidden colonies. This requires that the intrabacterial tracer accumulation provided by the transporter is matched by high serum stability of the tracer molecule. Herein, radiolabeled maltodextrins of varying chain lengths and with free nonreducing/reducing ends are reported and their behavior against starch‐degrading enzymes in the blood, which compromise their serum stability, is evaluated. Successful single‐photon emission computed tomography (SPECT) imaging is shown in a footpad infection model in vivo by using the newly developed model tracer, [^99mTc] MB1143 , and the signal is compared with that of ¹⁸F‐fluorodeoxyglucose positron emission tomography ([¹⁸F]FDG‐PET) as a nonbacterial specific marker for inflammation. Although the [^99mTc] MB1143 imaging signal is highly specific, it is low, most probably due to insufficient serum stability of the tracer. A series of stability tests with different ¹⁸F‐labeled maltodextrins finally yielded clear structural guidelines regarding substitution patterns and chain lengths of maltodextrin‐based tracers for nuclear imaging of bacterial infections.     

An In Vivo Study of a Rat Fluid-Percussion-Induced Traumatic Brain Injury Model with [11C]PBR28 and [18F]flumazenil PET Imaging

Traumatic brain injury (TBI) modelled by lateral fluid percussion-induction (LFPI) in rats is a widely used experimental rodent model to explore and understand the underlying cellular and molecular alterations in the brain caused by TBI in humans. Current improvements in imaging with positron emission tomography (PET) have made it possible to map certain features of TBI-induced cellular and molecular changes equally in humans and animals. The PET imaging technique is an apt supplement to nanotheranostic-based treatment alternatives that are emerging to tackle TBI. The present study aims to investigate whether the two radioligands, [¹¹C]PBR28 and [¹⁸F]flumazenil, are able to accurately quantify in vivo molecular-cellular changes in a rodent TBI-model for two different biochemical targets of the processes. In addition, it serves to observe any palpable variations associated with primary and secondary injury sites, and in the affected versus the contralateral hemispheres. As [¹¹C]PBR28 is a radioligand of the 18 kD translocator protein, the up-regulation of which is coupled to the level of neuroinflammation in the brain, and [¹⁸F]flumazenil is a radioligand for GABA_A-benzodiazepine receptors, whose level mirrors interneuronal activity and eventually cell death, the use of the two radioligands may reveal two critical features of TBI. An up-regulation in the [¹¹C]PBR28 uptake triggered by the LFP in the injured (right) hemisphere was noted on day 14, while the uptake of [¹⁸F]flumazenil was down-regulated on day 14. When comparing the left (contralateral) and right (LFPI) hemispheres, the differences between the two in neuroinflammation were obvious. Our results demonstrate a potential way to measure the molecular alterations in a rodent-based TBI model using PET imaging with [¹¹C]PBR28 and [¹⁸F]flumazenil. These radioligands are promising options that can be eventually used in exploring the complex in vivo pharmacokinetics and delivery mechanisms of nanoparticles in TBI treatment.     

Oxidation of p-[125I]Iodobenzoic Acid and p-[211At]Astatobenzoic Acid Derivatives and Evaluation In Vivo

The alpha particle-emitting radionuclide astatine-211 (²¹¹At) is of interest for targeted radiotherapy; however, low in vivo stability of many ²¹¹At-labeled cancer-targeting molecules has limited its potential. As an alternative labeling method, we evaluated whether a specific type of astatinated aryl compound that has the At atom in a higher oxidation state might be stable to in vivo deastatination. In the research effort, para-iodobenzoic acid methyl ester and dPEG₄-amino acid methyl ester derivatives were prepared as HPLC standards. The corresponding para-stannylbenzoic acid derivatives were also prepared and labeled with ¹²⁵I and ²¹¹At. Oxidization of the [¹²⁵I]iodo- and [²¹¹At]astato-benzamidyl-dPEG₄-acid methyl ester derivatives provided materials for in vivo evaluation. A biodistribution was conducted in mice with coinjected oxidized ¹²⁵I- and ²¹¹At-labeled compounds. The oxidized radioiodinated derivative was stable to in vivo deiodination, but unfortunately the oxidized [²¹¹At]astatinated benzamide derivative was found to be unstable under the conditions of isolation by radio-HPLC (post animal injection). Another biodistribution study in mice evaluated the tissue concentrations of coinjected [²¹¹At]NaAtO₃ and [¹²⁵I]NaIO₃. Comparison of the tissue concentrations of the isolated material from the oxidized [²¹¹At]benzamide derivative with those of [²¹¹At]astatate indicated the species obtained after isolation was likely [²¹¹At]astatate.     

Theranostic Small-Molecule Prodrug Conjugates for Targeted Delivery and Controlled Release of Toll-like Receptor 7 Agonists

We previously reported the design and synthesis of a small-molecule drug conjugate (SMDC) platform that demonstrated several advantages over antibody–drug conjugates (ADCs) in terms of in vivo pharmacokinetics, solid tumor penetration, definitive chemical structure, and adaptability for modular synthesis. Constructed on a tri-modal SMDC platform derived from 1,3,5-triazine (TZ) that consists of a targeting moiety (Lys-Urea-Glu) for prostate-specific membrane antigen (PSMA), here we report a novel class of chemically identical theranostic small-molecule prodrug conjugates (T-SMPDCs), [^18/19F]F-TZ(PSMA)-LEGU-TLR7, for PSMA-targeted delivery and controlled release of toll-like receptor 7 (TLR7) agonists to elicit de novo immune response for cancer immunotherapy. In vitro competitive binding assay of [¹⁹F]F-TZ(PSMA)-LEGU-TLR7 showed that the chemical modification of Lys-Urea-Glu did not compromise its binding affinity to PSMA. Receptor-mediated cell internalization upon the PSMA binding of [¹⁸F]F-TZ(PSMA)-LEGU-TLR7 showed a time-dependent increase, indicative of targeted intracellular delivery of the theranostic prodrug conjugate. The designed controlled release of gardiquimod, a TLR7 agonist, was realized by a legumain cleavable linker. We further performed an in vivo PET/CT imaging study that showed significantly higher uptake of [¹⁸F]F-TZ(PSMA)-LEGU-TLR7 in PSMA⁺ PC3-PIP tumors (1.9 ± 0.4% ID/g) than in PSMA⁻ PC3-Flu tumors (0.8 ± 0.3% ID/g) at 1 h post-injection. In addition, the conjugate showed a one-compartment kinetic profile and in vivo stability. Taken together, our proof-of-concept biological evaluation demonstrated the potential of our T-SMPDCs for cancer immunomodulatory therapies.