Loss of Bcl-3 regulates macrophage polarization by promoting macrophage glycolysis.

M1/M2 macrophage polarization plays an important role in regulating the balance of the microenvironment within tissues. Moreover, macrophage polarization involves the reprogramming of metabolism, such as glucose and lipid metabolism. Transcriptional coactivator B-cell lymphoma-3 (Bcl-3) is an atypical member of the IκB family that controls inflammatory factor levels in macrophages by regulating nuclear factor kappa B pathway activation. However, the relationship between Bcl-3 and macrophage polarization and metabolism remains unclear. In this study, we show that the knockdown of Bcl-3 in macrophages can regulate glycolysis-related gene expression by promoting the activation of the nuclear factor kappa B pathway. Furthermore, the loss of Bcl-3 was able to promote the interferon gamma/lipopolysaccharide-induced M1 macrophage polarization by accelerating glycolysis. Taken together, these results suggest that Bcl-3 may be a candidate gene for regulating M1 polarization in macrophages.

The first-in-human preclinical evaluation of the new probe [123I]I-PSMA-7 for real-time intraoperative targeted biopsy and SPECT/CT imaging in prostate cancer.

PURPOSE: PSMA/PET has been increasingly used to detect PCa, and PSMA/PET-guided biopsy has shown promising results. However, it cannot be confirmed immediately whether the tissues are the targeted area. In this study, we aimed to develop a novel probe, [123I]I-PSMA-7. First, we hope that [123I]I-PSMA-7 can provide instant confirmation for prostate biopsy. Second, we hope it will help detect PCa. METHODS: We synthesized a high-affinity probe, [123I]I-PSMA-7, and evaluated its properties. We included ten patients with suspected PCa and divided them into two groups. The injection and biopsy were approximately 24 h apart. The activity in biopsy lesions was measured as the cpm by a γ-counter. Moreover, we enrolled 3 patients to evaluate the potential of [123I]I-PSMA-7 for detecting PCa. RESULTS: Animal experiments verified the safety, targeting and effectiveness of [123I]I-PSMA-7, and the tumor-to-muscle ratio was greatest at 24 h, which confirmed the results of this study in humans. After injection of 185MBq [123I]I-PSMA-7, 18/55 cores were positive, and the cpm was significantly greater (4345 ± 3547 vs. 714 ± 547, P < 0.001), with an AUC of 0.97 and a cutoff of 1312 (sens/spec of 94.40%/91.90%). At a lower dose, 10/55 biopsy cores were cancerous, and the cpm was 2446 ± 1622 vs. 153 ± 112 (P < 0.001). The AUC was 1, with a cutoff value of 490 (sens/spec of 100%). When the radiopharmaceuticals were added to 370 MBq, we achieved better SPECT/CT imaging. CONCLUSION: With the aid of [123I]I-PSMA-7 and via cpm-based biopsy, we can reduce the number of biopsies to a minimum operation. [123I]I-PSMA-7 PSMA SPECT/CT can also provide good imaging results. TRIAL REGISTRATION: Chinese Clinical trial registry ChiCTR2300069745, Registered 24 March 2023.

Synthesis and Bioevaluation of 2-Styrylquinoxaline Derivatives as Tau-PET Tracers.

This study focused on designing and evaluating Tau-PET tracers for noninvasive positron emission computed tomography (PET) imaging of neurofibrillary tangles (NFTs), a hallmark pathology of Alzheimer's disease (AD). The tracers were synthesized with a 2-styrylquinoxaline scaffold and varying lengths of FPEG chains. The compound [18F]15, which had two ethoxy units, showed high affinity for recombinant K18-Tau aggregates (Ki = 41.48 nM) and the highest selectivity versus Aß1-42 aggregates (8.83-fold). In vitro autoradiography and fluorescent staining profiles further validated the binding of [18F]15 or 15 toward NFTs in brain sections from AD patients and Tau-transgenic mice. In normal ICR mice, [18F]15 exhibited an ideal initial brain uptake (11.21% ID/g at 2 min) and moderate washout ratio (2.29), and micro-PET studies in rats confirmed its ability to penetrate the blood-brain barrier with the peak SUV value of 1.94 in the cortex. These results suggest that [18F]15 has the potential to be developed into a useful Tau-PET tracer for early AD diagnosis and evaluation of anti-Tau therapeutics.

D-π-A-Based Trisubstituted Alkenes as Environmentally Sensitive Fluorescent Probes to Detect Lewy Pathologies.

Lewy pathologies, which mainly consist of insoluble α-synuclein (α-syn) aggregates, are the hallmarks of Parkinson's disease and many other neurodegenerative diseases termed "synucleinopathies". Detection of Lewy pathologies with optical methods is of interest for preclinical studies, while the α-syn fluorescent probe is still in great demand. By rational design, we obtained a series of D-π-A-based trisubstituted alkenes with acceptable optical properties and high binding affinities to α-syn fibrils. Among these probes, FPQXN and TQXN-2 exhibited high binding affinities (6 and 8 nM, respectively), significant fluorescence enhancements (17.2- and 26.6-fold, respectively), and satisfying quantum yields (36.5% and 10.4%, respectively), which met the need for the in vitro neuropathological staining of Lewy pathologies in the PD brain sections. In addition, TQXN-2 showed great potential in fluorescent discrimination of Lewy pathologies and Aß plaques. Our research provides flexible tools for in vitro detection of α-syn aggregates and offers new structural frameworks for the further development of α-syn fluorescent probes.

Synthesis and Evaluation of Fluorine-18 Labeled 2-Phenylquinoxaline Derivatives as Potential Tau Imaging Agents.

In this study, three pairs of optically pure 18F-labeled 2-phenylquinoxaline derivatives were evaluated as Tau imaging agents for the diagnosis of Alzheimer's disease (AD). The chiral 2-fluoromethyl-1,2-ethylenediol side chain was attached to the 2-phenylquinoxaline backbone to increase hydrophilicity, thereby improving the binding affinity of the probe to tangles and their selectivity toward Tau tangles over ß-amyloid plaques (Aß). These probes displayed excellent fluorescent properties and high selectivity for tangles on brain sections from transgenic mice (rTg4510) and AD patients. Quantitative binding assays with AD homogenates showed that the probes (R)-5 and (S)-16 have a high affinity (Ki = 4.1 and 10.3 nM, respectively) and high selectivity (30.5-fold and 34.6-fold, respectively) for tangles over Aß. The high affinity and selectivity of (R)-[18F]5 and (S)-[18F]16 for tangles were further confirmed with autoradiography on AD brain tissue in vitro. In addition, they displayed sufficient blood-brain barrier penetration (7.06% and 10.95% ID/g, respectively) and suitable brain kinetics (brain2 min/brain60 min = 10.1, 6.5 respectively) in normal mice. Ex vivo metabolism studies and micro-positron emission computed tomography (PET) revealed high brain biostability, good brain kinetic properties, and low nonspecific binding for (S)-[18F]16. Together, these results demonstrate that (R)-[18F]5 and (S)-[18F]16 are promising PET probes for Tau tangles imaging.

18F-labeled 2-phenylbenzoheterocycles with chiral dihydroxyl side chains as ß-amyloid imaging probes.

This study reported the design, synthesis and bio-evaluation of 2-phenylbenzoheterocycles with chiral dihydroxyl side chains as ß-amyloid (Aß) imaging probes. This strategy of introducing two hydroxyls offered a simplified method for effectively reducing the lipophilicity. The probes (R, S)/(S, R)-14-15 with benzothiazole scaffold displayed good binding affinities toward Aß1-42 aggregates with Ki values ranging from 47.63 to 56.28 nM. Further biological studies shown that (R, S)/(S, R)-[18F]14 have no obvious chirality-related discrepancy in binding ability and mice bio-distribution, while (S, R)-enantiomer exhibited slightly faster brain washout rate than (R, S)-enantiomer. Compared to the FDA approved [18F]Florbetapir and the fluoro-peglated 2-phenylbenzothiazole derivatives, (S, R)-[18F]14 displayed improved brain kinetics (6.40% ID/g at 2 min, brain2 min/brain60 min = 7.80) that is favorable for further application. In vitro autoradiography studies validated its high affinity and specificity to Aß plaques. Overall, (S, R)-[18F]14 deserved further detailed study as a potential PET imaging probe for AD early diagnosis.

Flexible multidentate benzyldiamine derivatives with high affinity for ß-amyloid in cerebral amyloid angiopathy.

Cerebral amyloid angiopathy (CAA) commonly found in the aged is pathologically characterized by ß-amyloid (Aß) deposition in the walls of arteries and capillaries of brain. In this study, four flexible multidentate benzyldiamine derivatives as potential probes for cerebrovascular Aß deposition were designed and synthesized. In in vitro inhibition assays, the ligands 18-21 displayed high affinities for Aß aggregates with Ki values of 1.45 ± 0.53 nM, 1.68 ± 0.35 nM, 1.16 ± 0.23 nM and 1.72 ± 0.19 nM, respectively. A significant improvement in the binding affinity over the monomer, compounds 9-12 or benzyldiamine derivatives, demonstrated the applicability of the multidentate approach. The underlying mechanism of these novel Aß agents was explored by molecular docking technique, which theoretically verified the high affinities of the multidentate benzyldiamine derivatives for Aß aggregates. Moreover, the molecular masses of the ligands 18-21 are more than 700 Dalton, which are believed to be hardly capable of penetrating blood brain barrier. In this regard, these ligands could be used to distinguish CAA from Alzheimer's disease which is another Aß-related disorder disease. To convert these ligands to positron emission tomography imaging agents, we attempted to radiosynthesize [18F]18. Though the radiolabeling was not very successful, the preliminary results suggested that these newly proposed multidentate benzyldiamine derivatives may be used as potential Aß imaging agents in cerebral amyloid angiopathy.

Discovery of Diphenoxy Derivatives with Flexible Linkers as Ligands for ß-Amyloid Plaques.

The highly rigid and planar scaffolds with π-conjugated systems have been widely considered to be indispensable for ß-amyloid (Aß) binding ligands. In this study, a library of diphenoxy compounds with different types of more flexible linkers as Aß ligands were synthesized and evaluated. Most of them displayed good affinity (Ki < 100 nM) for Aß1-42 aggregates, and some ligands even showed values of Ki less than 10 nM. Structure-activity relationship analysis revealed that modification on the linkers or substituents tolerated great flexibility, which challenged the long-held belief that rigid and planar structures are exclusively favored for Aß binding. Three ligands were labeled by iodine-125, and they exhibited good properties in vitro and in vivo, which further supported that this flexible scaffold was potential and promising for the development of Aß imaging agents.

Structure-Property Relationships of Polyethylene Glycol Modified Fluorophore as Near-Infrared Aß Imaging Probes.

To optimize the lipophilicity and improve in vivo pharmacokinetics of near-infrared probes targeted Aß plaques, we designed, synthesized, and evaluated a series of polyethylene glycol modified probes with hydroxyl and methoxyl terminals. The relationships between chemical structure and optical, biological properties were systemically elucidated. The results indicated that a desired Aß probe should keep a balance among molecular rigidity, size, and lipophilicity. Probe 12d displayed improved properties including intense and selective response to Aß1-42 aggregates ( Kd = 7.3 nM, 22-fold fluorescence enhancement and emission maxima at 715 nm upon interaction with Aß1-42 aggregates), sufficient blood-brain barrier penetration (3.04% ID/g), and fast wash out from the brain (brain2 min/brain60 min = 10.1). Clear fluorescence signals retention in transgenic mice than control mice in in vivo near-infrared imaging. Hence, polyethylene glycol modified probes retained favorable optical properties but displayed great improvement of biological properties for Aß detection.

Smart D-π-A Type Near-Infrared Aß Probes: Effects of a Marked π Bridge on Optical and Biological Properties.

To expand the scope of D-π-A based near-infrared (NIR) probes for detecting ß-amyloid (Aß) plaques and to systematically explore the relationship among their structural characteristics, optical properties, and biological properties, three series of smart NIR probes with different aromatic rings and up to seven trans double bonds were synthesized and evaluated. Marked correlations between the conjugated π system and properties of these probes, such as optical data, binding ability, and brain uptake, were observed. One probe, PHC-4, displayed improved properties as a NIR probe for the in vivo detection of Aß plaques.

Multicomponent Aqueous Synthesis of Iodo-1,2,3-triazoles: Single-Step Models for Dual Modification of Free Peptide and Radioactive Iodo Labeling.

Iodo-1,2,3-triazoles are of considerable interest for chemical and biomedical applications. However, current synthetic methods for preparing iodo-1,2,3-triazoles cannot easily be applied to the direct modification of bioactive molecules in water. Through the combination of water-compatible oxidative iodination and the copper-catalyzed alkyne-azide cycloaddition reaction, a novel copper-catalyzed aqueous multicomponent synthetic method for the preparation of 5-iodo-1,2,3-triazoles has been developed. The method is highly effective and selective for substrates including biologically relevant compounds with nucleoside, sugar, and amino acid moieties. Based on this aqueous tandem reaction, a direct single-step multicomponent dual modification of peptide is developed from readily available starting materials. Furthermore, the method could also be applied to concise and fast multicomponent radioactive 125 I labeling from an aqueous solution of commercially available sodium 125 iodide as a starting material.

18F-Labeled indole-based analogs as highly selective radioligands for imaging sigma-2 receptors in the brain.

We have designed and synthesized a series of indole-based σ2 receptor ligands containing 5,6-dimethoxyisoindoline or 6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline as pharmacophore. In vitro competition binding assays showed that all ten ligands possessed low nanomolar affinity (Ki=1.79-5.23nM) for σ2 receptors and high subtype selectivity (Ki (σ2)/Ki (σ1)=56-708). Moreover, they showed high selectivity for σ2 receptor over the vesicular acetylcholine transporter (>1000-fold). The corresponding radiotracers [18F]16 and [18F]21 were prepared by an efficient one-pot, two-step reaction sequence with a home-made automated synthesis module, with 10-15% radiochemical yield and radiochemical purity of >99%. Both radiotracers showed high brain uptake and σ2 receptor binding specificity in mice.

Amyloid-ß Deposits Target Efficient Near-Infrared Fluorescent Probes: Synthesis, in Vitro Evaluation, and in Vivo Imaging.

The formation of extracellular amyloid-ß (Aß) plaques is a common molecular change that underlies several debilitating human conditions, including Alzheimer's disease (AD); however, the existing near-infrared (NIR) fluorescent probes for the in vivo detection of Aß plaques are limited by undesirable fluorescent properties and poor brain kinetics. In this work, we designed, synthesized, and evaluated a new family of efficient NIR probes that target Aß plaques by incorporating hydroxyethyl groups into the ligand structure. Among these probes, DANIR 8c showed excellent fluorescent properties with an emission maximum above 670 nm upon binding to Aß aggregates and also displayed a high sensitivity (a 629-fold increase in fluorescence intensity) and affinity (Kd = 14.5 nM). Because of the improved hydrophilicity that was induced by hydroxyls, 8c displayed increased initial brain uptake and a fast washout from the brain, as well as an acceptable biostability in the brain. In vivo NIR fluorescent imaging revealed that 8c could efficiently distinguish between AD transgenic model mice and normal controls. Overall, 8c is an efficient and veritable NIR fluorescent probe for the in vivo detection of Aß plaques in the brain.

Dual-Functional Nanoparticles for In Situ Sequential Detection and Imaging of ATP and H2 O2.

Within a complex biological sample, the in situ sequential detection of multiple molecules without any interference is greatly desirable. Dual-functional nanoparticles are constructed, with the enzyme-based core-shell structures, for the in situ sequential detection of ATP and H2 O2 within the same biological system.

99mTc-Labeled 2-Arylbenzothiazoles: Aß Imaging Probes with Favorable Brain Pharmacokinetics for Single-Photon Emission Computed Tomography.

A series of 2-arylbenzothiazole derivatives conjugated with bis(aminoethanethiol) (BAT) chelating groups were designed and synthesized. A competitive binding assay-based screening was used to select seven rhenium complexes with potent binding affinity toward Aß1-42 aggregates (Ki < 50 nM) for 99mTc labeling and further evaluation. The 99mTc-labeled probes showed good affinity and specificity to Aß plaques in Tg mouse brain tissue in in vitro autoradiography studies. Moreover, [99mTc]14b exhibited favorable brain pharmacokinetics in normal mice (2.11% ID/g at 2 min and 0.62% ID/g at 60 min). Ex vivo autoradiography revealed extensive labeling of Aß plaques by [99mTc]14b in the brain of Tg mice. Furthermore, we performed the first single-photon emission computed tomography (SPECT) imaging study in nonhuman primates with 99mTc-labeled Aß probes. The semiquantitative data showed that [99mTc]14b penetrated the brains of rhesus monkeys. These results indicate that [99mTc]14b could be utilized as a SPECT imaging probe for Aß plaques.

Synthesis and Monkey-PET Study of (R)- and (S)-18F-Labeled 2-Arylbenzoheterocyclic Derivatives as Amyloid Probes with Distinctive in Vivo Kinetics.

This study describes an effective strategy to improve pharmacokinetics of Aß imaging agents, offering a novel class of (R)- and (S)-18F-labeled 2-arylbenzoheterocyclic derivatives which bear an additional chiral hydroxyl group on the side chain. These ligands displayed binding abilities toward Aß aggregates with Ki values ranging from 3.2 to 195.6 nM. Chirality-related discrepancy was observed in biodistribution, and (S)-2-phenylbenzoxazole enantiomers exhibited vastly improved brain clearance with washout ratios higher than 20. Notably, (S)-[18F]28 possessed high binding potency (Ki = 7.6 nM) and exceptional brain kinetics (9.46% ID/g at 2 min, brain2min/brain60min = 27.8) that is superior to well-established [18F]AV45. The excellent pharmacokinetics and low nonspecific binding of (S)-[18F]28 were testified by dynamic PET/CT scans in monkey brains. In addition, (S)-[18F]28 clearly labeled Aß plaques both in vitro and ex vivo. These results might qualify (S)-[18F]28 to detect Aß plaques with high signal-to-noise ratio.

99mTc(CO)3-Labeled Benzothiazole Derivatives Preferentially Bind Cerebrovascular Amyloid: Potential Use as Imaging Agents for Cerebral Amyloid Angiopathy.

Cerebral amyloid angiopathy (CAA) is a disorder affecting the elderly that is characterized by amyloid-ß (Aß) deposition in blood vessel walls of the brain. A series of 99mTc(CO)3-labeled benzothiazole derivatives as potential SPECT imaging probes for cerebrovascular Aß deposition is reported. Rhenium surrogate displayed high affinities to Aß aggregates with Ki values ranging from 106 to 42 nM, and they strongly stained Aß deposits in transgenic mice (Tg) and Alzheimer's disease (AD) patients. In vitro autoradiography on brain sections of Tg and AD patients confirmed that [99mTc]24 possessed sufficient affinity for Aß plaques, and [99mTc]24 could only label Aß deposition in blood vessels but not Aß plaques in the parenchyma of the brain of AD patients. Moreover, [99mTc]24 possessed favorable initial uptake (1.21% ID/g) and fast blood washout (blood2 min/blood60 min=23) in normal mice. These preliminary results suggest that [99mTc]24 may be used as an Aß imaging probe for the detection of CAA.

Smart near-infrared fluorescence probes with donor-acceptor structure for in vivo detection of ß-amyloid deposits.

The deposition of ß-amyloid (Aß) plaques in the parenchymal and cortical brain is accepted as the main pathological hallmark of Alzheimer's disease (AD); however, early detection of AD still presents a challenge. With the assistance of molecular imaging techniques, imaging agents specifically targeting Aß plaques in the brain may lead to the early diagnosis of AD. Herein, we report the design, synthesis, and evaluation of a series of smart near-infrared fluorescence (NIRF) imaging probes with donor-acceptor architecture bridged by a conjugated π-electron chain for Aß plaques. The chemical structure of these NIRF probes is completely different from Congo Red and Thioflavin-T. Probes with a longer conjugated π system (carbon-carbon double bond) displayed maximum emission in PBS (>650 nm), which falls in the best range for NIRF probes. These probes were proved to have affinity to Aß plaques in fluorescent staining of brain sections from an AD patient and double transgenic mice, as well as in an in vitro binding assay using Aß(1-42) aggregates. One probe with high affinity (K(i) = 37 nM, K(d) = 27 nM) was selected for in vivo imaging. It can penetrate the blood-brain barrier of nude mice efficiently and is quickly washed out of the normal brain. Moreover, after intravenous injection of this probe, 22-month-old APPswe/PSEN1 mice exhibited a higher relative signal than control mice over the same period of time, and ex vivo fluorescent observations confirmed the existence of Aß plaques. In summary, this probe meets most of the requirements for a NIRF contrast agent for the detection of Aß plaques both in vitro and in vivo.

Synthesis and evaluation of a 18F-labeled spirocyclic piperidine derivative as promising σ1 receptor imaging agent.

Several spirocyclic piperidine derivatives were designed and synthesized as σ1 receptor ligands. In vitro competition binding assays showed that the fluoroalkoxy analogues with small substituents possessed high affinity towards σ1 receptors and subtype selectivity. Particularly for ligand 1'-((6-(2-fluoroethoxy)pyridin-3-yl)methyl)-3H-spiro[2-benzofuran-1,4'-piperidine] (2), high σ1 receptor affinity (Ki=2.30 nM) and high σ1/σ2 subtype selectivity (142-fold) as well as high σ1/VAChT selectivity (234-fold) were observed. [18F]2 was synthesized using an efficient one-pot, two-step reaction method in a home-made automated synthesis module, with an overall isolated radiochemical yield of 8-10%, a radiochemical purity of higher than 99%, and specific activity of 56-78GBq/µmol. Biodistribution studies of [18F]2 in ICR mice indicated high initial brain uptake and a relatively fast washout. Administration of haloperidol, compound 1 and different concentrations of SA4503 (3, 5, or 10 µmol/kg) 5 min prior to injection of [18F]2 significantly decreased the accumulation of radiotracer in organs known to contain σ1 receptors. Ex vivo autoradiography in Sprague-Dawley rats demonstrated high accumulation of radiotracer in brain areas with high expression of σ1 receptors. These encouraging results prove that [18F]2 is a suitable candidate for σ1 receptor imaging with PET in humans.

Screening of [18F]Florbetazine for Aß Plaques and a Head-to-Head Comparison Study with [11C]Pittsburgh Compound-B ([11C]PiB) in Human Subjects.

Positron emission tomography (PET) imaging of amyloid-ß (Aß) has emerged as a crucial strategy for early diagnosis and monitoring of therapeutic advancements targeting Aß. In our previous first-in-human study, we identified that [18F]Florbetazine ([18F]92), featuring a diaryl-azine scaffold, exhibits higher cortical uptake in Alzheimer's disease (AD) patients compared to healthy controls (HC). Building upon these promising findings, this study aimed to characterize the diagnostic potential of [18F]92 and its dimethylamino-modified tracer [18F]91 and further compare them with the benchmark [11C]PiB in the same cohort of AD patients and age-matched HC subjects. The cortical accumulation of these tracers was evident, with no significant radioactivity retention observed in the cortex of HC subjects, consistent with [11C]PiB images (correlation coefficient of 0.9125 and 0.7883 between [18F]Florbetazine/[18F]91 and [11C]PiB, respectively). Additionally, quantified data revealed higher standardized uptake value ratios (SUVR) (with the cerebellum as the reference region) of [18F]Florbetazine/[18F]91 in AD patients compared to the HC group ([18F]Florbetazine: 1.49 vs 1.16; [18F]91: 1.33 vs 1.20). Notably, [18F]Florbetazine exhibited less nonspecific bindings in myelin-rich regions, compared to the dimethylamino-substituted [18F]91, akin to [11C]PiB. Overall, this study suggests that [18F]Florbetazine displays superior characteristics to [18F]91 in identifying Aß pathology in AD. Furthermore, the close agreement between the uptakes in nontarget regions for [18F]Florbetazine and [11C]PiB in this head-to-head comparison study underscores its suitability for both clinical and research applications.