Development of a generator for percussive ultrasonic drills used in asteroid exploration based on impedance characteristics analysis.

Percussive ultrasonic drills participate in asteroid exploration missions. Since space environments are complex and working loads vary dramatically, it is necessary to design a drive that better matches the percussive ultrasonic drill to make it achieve high-speed drilling with low power consumption. Impedance characteristics under different load conditions and the load varying in one drilling cycle are investigated based on analyzing the working principle of the percussive ultrasonic drill. An ultrasonic drill driver with automatic scanning resonant frequency and digital phase-locked loop frequency tracking control is designed according to the load characteristics. Series capacitance impedance matching ultrasonic drills is experimentally studied. Vibration amplitude and impedance characteristics of the ultrasonic drill driven by the designed driver are measured to evaluate the frequency tracking ability and the impedance matching. Finally, ultrasonic drilling experiments are conducted in room, low, and high temperature environments to investigate the driving performance. Under room temperature and 5 N drilling pressure, the speed of ultrasonic drilling into soft sandstone is 52 mm/min, and the power consumption is less than 70 W. The experimental results indicate that the designed driver can drive the percussive ultrasonic drill to achieve stable and high-speed drilling with low power consumption and low drilling pressure.

Magnetically actuated gearbox for the wireless control of millimeter-scale robots.

The limited force or torque outputs of miniature magnetic actuators constrain the locomotion performances and functionalities of magnetic millimeter-scale robots. Here, we present a magnetically actuated gearbox with a maximum size of 3 millimeters for driving wireless millirobots. The gearbox is assembled using microgears that have reference diameters down to 270 micrometers and are made of aluminum-filled epoxy resins through casting. With a magnetic disk attached to the input shaft, the gearbox can be driven by a rotating external magnetic field, which is not more than 6.8 millitesla, to produce torque of up to 0.182 millinewton meters at 40 hertz. The corresponding torque and power densities are 12.15 micronewton meters per cubic millimeter and 8.93 microwatt per cubic millimeter, respectively. The transmission efficiency of the gearbox in the air is between 25.1 and 29.2% at actuation frequencies ranging from 1 to 40 hertz, and it lowers when the gearbox is actuated in viscous liquids. This miniature gearbox can be accessed wirelessly and integrated with various functional modules to repeatedly generate large actuation forces, strains, and speeds; store energy in elastic components; and lock up mechanical linkages. These characteristics enable us to achieve a peristaltic robot that can crawl on a flat substrate or inside a tube, a jumping robot with a tunable jumping height, a clamping robot that can sample solid objects by grasping, a needle-puncture robot that can take samples from the inside of the target, and a syringe robot that can collect or release liquids.

Discovery of SARS-CoV-2 main protease covalent inhibitors from a DNA-encoded library selection.

Covalent inhibitors targeting the main protease (Mpro, or 3CLpro) of SARS-CoV-2 have shown promise in preclinical investigations. Herein, we report the discovery of two new series of molecules that irreversibly bind to SARS-CoV-2 Mpro. These acrylamide containing molecules were discovered using our covalent DNA-encoded library (DEL) screening platform. Following selection against SARS-CoV-2 Mpro, off-DNA compounds were synthesized and investigated to determine their inhibitory effects, the nature of their binding, and to generate preliminary structure-activity relationships. LC-MS analysis indicates a 1:1 (covalent) binding stoichiometry between our hit molecules and SARS-CoV-2 Mpro. Fluorescent staining assay for covalent binding in the presence of cell lysate suggests reasonable selectivity for SARS-CoV-2 Mpro. And lastly, inhibition of enzymatic activity was also observed against a panel of 3CLpro enzymes from different coronavirus strains, with IC50 values ranging from inactive to single digit micromolar. Our results indicate that DEL selection is a useful approach for identifying covalent inhibitors of cysteine proteases.

Dielectric Characterization and Multistage Separation of Various Cells via Dielectrophoresis in a Bipolar Electrode Arrayed Device.

Isolation of microalgal cells is as an indispensable part of producing biofuels for energy security and detecting toxic contaminants for marine routine monitoring. Microalgae live together with various microalgae naturally, and abundant samples need to be tackled in practical applications. Therefore, effective separation technologies need to be developed urgently to achieve high-throughput separation of various microalgae. Herein, we develop a reliable device to characterize the dielectric response of microalgae and sequentially separate various microalgae utilizing dielectrophoretic force in a bipolar electrode (BPE) arrayed device. First, by investigating the array width extension (AWE) effect on the electric- and flow-field distributions, we explore consequences of incidental electrohydrodynamic mechanisms and axial flow rate on the separation. Second, based on device performance on sample characterizations, we demonstrate this technology by separating microparticles in three- and five-channel devices. Third, we discriminate dead and live cells to explore its capability using the cell viability test and illustrate the AWE influence on the separation. Fourth, we characterize dielectric responses of different microalgae and separate C. vulgaris and Oocystis sp. Finally, we extended BPEs in length and developed an arrayed device for sequential separation of various microalgae, and this platform is successfully engineered in high-throughput isolation of C. vulgaris from complex samples. This technology presents good potential in addressing depleting fossil fuel and burgeoning environmental concerns due to its performance in the separation of microalgal strains from complex samples.

68Ga-FAPI-04 vs. 18F-FDG in a longitudinal preclinical PET imaging of metastatic breast cancer.

PURPOSE: This longitudinal study aims to evaluate the performance of 68 Ga-FAPI-04 and 18F-FDG and to profile the dynamic process of tumor metastasis in a preclinical 4T1 breast cancer model. Although both of these two radioligands are wildly used in clinic, no study was reported on their performance in the longitudinal monitoring of tumor metastasis. Also, no correlation between the expression level of fibroblast activation protein (FAP) and the development of tumor metastasis has been elucidated previously. In this study, we evaluated the performance of 68 Ga-FAPI-04 and 18F-FDG PET during the entire process of tumor metastasis, and their potential for the early diagnosis of tumor metastasis. We also clarified the correlation of uptakes as well as the signal-to-background (S/B) ratios between these two probes at different stages of tumor metastasis. METHODS: Forty 4T1 metastatic breast cancer murine models were established using female BALB/c mice, followed by the longitudinal imaging with 68 Ga-FAPI-04 and 18F-FDG once a week for up to 6 weeks. In vitro hematoxylin and eosin (H&E) and immunochemistry (IHE) staining were performed to evaluate FAP expression on the metastatic lesions. Further statistical analysis was performed to evaluate the correlation of 68 Ga-FAPI-04 and 18F-FDG uptake (%ID/cc) at different stages of the metastasis. RESULTS: 68 Ga-FPAI-04 holds an advantage over 18F-FDG with higher sensitivity at the early stage of tumor metastasis. However, with the progress of tumor metastasis, uptake of 68 Ga-FAPI-04 decreases and becomes less sensitive than 18F-FDG. There is also no direct correlation between uptake or S/B ratios of 68 Ga-FAPI-04 and 18F-FDG during this dynamic process. CONCLUSION: 68 Ga-FAPI-04 is more sensitive than 18F-FDG in detecting the early stage of tumor metastasis, but becomes less sensitive than 18F-FDG at the late stage of tumor metastasis. We envision this result would be meaningful for the explanation of the 68 Ga-FAPI-04 and 18F-FDG imaging both in the future clinic and preclinic studies.

Dielectrophoresis Response of Water-in-Oil-in-Water Double Emulsion Droplets with Singular or Dual Cores.

Microfluidic technologies have enabled generation of exquisite multiple emulsion droplets, which have been used in many fields, including single-cell assays, micro-sized chemical reactions, and material syntheses. Electrical controlling is an important technique for droplet manipulation in microfluidic systems, but the dielectrophoretic behaviors of multiple emulsion droplets in electrical fields are rarely studied. Here, we report on the dielectrophoresis response of double emulsion droplets in AC electric fields in microfluidic channel. A core-shell model is utilized for analyzing the polarization of droplet interfaces and the overall dielectrophoresis (DEP) force. The water-in-oil-in-water droplets, generated by glass capillary devices, experience negative DEP at low field frequency. At high frequency, however, the polarity of DEP is tunable by adjusting droplet shell thickness or core conductivity. Then, the behavior of droplets with two inner cores is investigated, where the droplets undergo rotation before being repelled or attracted by the strong field area. This work should benefit a wide range of applications that require manipulation of double emulsion droplets by electric fields.

The optimization of a novel selective antagonist for human M2 muscarinic acetylcholine receptor.

Muscarinic acetylcholine receptors (mAChRs) comprise five distinct subtypes denoted M1 to M5. The antagonism of M2 subtype could increase the release of acetylcholine from vesicles into the synaptic cleft and improve postsynaptic functions in the hippocampus via M1 receptor activation, displaying therapeutic potentials for Alzheimer's disease. However, drug development for M2 antagonists is still challenged among different receptor subtypes. In this study, by optimizing a scaffold from virtual screening, we synthesized two focused libraries and generated up to 50 derivatives. By measuring potency and binding selectivity, we discovered a novel M2 antagonist, ligand 47, featuring submicromolar IC50, high M2/M4 selectivity (~30-fold) and suitable lipophilicity (cLogP = 4.55). Further study with these compounds also illustrates the structure-activity relationship of this novel scaffold. Our study could not only provide novel lead structure, which was easy to synthesize, but also offer valuable information for further development of selective M2 ligands.

GluN2A-selective positive allosteric modulator-nalmefene-flumazenil reverses ketamine-fentanyl-dexmedetomidine-induced anesthesia and analgesia in rats.

Anesthetics are used to produce hypnosis and analgesic effects during surgery, but anesthesia for a long time after the operation is not conducive to the recovery of animals or patients. Therefore, finding appropriate treatments to counter the effects of anesthetics could enhance postoperative recovery. In the current study, we discovered the novel role of a GluN2A-selective positive allosteric modulator (PAM) in ketamine-induced anesthesia and investigated the effects of the PAM combined with nalmefene and flumazenil (PNF) in reversing the actions of an anesthetic combination (ketamine-fentanyl-dexmedetomidine, KFD). PAM treatment dose-dependently decreased the duration of the ketamine-induced loss of righting reflex (LORR). Compared with those in the KFD group, the duration of LORR and the analgesic effect of the KFD + PNF group were obviously decreased. Meanwhile, successive administration of PNF and KFD had no adverse effects on the cardiovascular and respiratory systems. Both the KFD group and the KFD + PNF group showed no changes in hepatic and renal function or cognitive function in rats. Moreover, the recovery of motor coordination of the KFD + PNF group was faster than that of the KFD group. In summary, our results suggest the potential application of the PNF combination as an antagonistic treatment strategy for anesthesia.

An Experimental Study of 3D Electrode-Facilitated Particle Traffic Flow-Focusing Driven by Induced-Charge Electroosmosis.

In this paper we present a novel microfluidic approach for continuous, rapid and switchable particle concentration, using induced-charge electroosmosis (ICEO) in 3D electrode layouts. Field-effect control on non-linear electroosmosis in the transverse direction greatly facilitates a selective concentration of biological yeast cells from a straight main microchannel into one of the three downstream branch channels in our microfluidic device. For the geometry configuration of 3D driving electrode plates on sidewalls and a 2D planar gate electrode strip on the channel bottom surface, we briefly describe the underlying physics of an ICEO-based particle flow-focusing method, and provide relevant simulation results to show how gate voltage amplitude can be used to guide the motion trajectory of the concentrated particle stream. With a relatively simple geometrical configuration, the proposed microfluidic device provides new possibilities to controllably concentrate micro/nanoparticles in continuous flow by using ICEO, and is suitable for a high-throughput front-end cell concentrator interfacing with various downstream biosensors.

Preclinical Evaluation of a Novel TSPO PET Ligand 2-(7-Butyl-2-(4-(2-[18F]Fluoroethoxy)phenyl)-5-Methylpyrazolo[1,5-a]Pyrimidin-3-yl)-N,N-Diethylacetamide (18F-VUIIS1018A) to Image Glioma.

PURPOSE: There is an urgent need for the development of novel positron emission tomography (PET) tracers for glioma imaging. In this study, we developed a novel PET probe ([18F]VUIIS1018A) by targeting translocator protein (TSPO), an imaging biomarker for glioma. The purpose of this preclinical study was to evaluate this novel TSPO probe for glioma imaging. PROCEDURES: In this study, we synthesized [19F]VUIIS1018A and the precursor for radiosynthesis of [18F]VUIIS1018A. TSPO binding affinity was confirmed using a radioligand competitive binding assay in C6 glioma cell lysate. Further, dynamic imaging studies were performed in rats using a microPET system. These studies include displacement and blocking studies for ligand reversibility and specificity evaluation, and compartment modeling of PET data for pharmacokinetic parameter measurement using metabolite-corrected arterial input functions and PMOD. RESULTS: Compared to previously reported TSPO tracers including [18F]VUIIS1008 and [18F]DPA-714, the novel tracer [18F]VUIIS1018A demonstrated higher binding affinity and BPND. Pretreatment with the cold analog [19F]VUIIS1018A could partially block tumor accumulation of this novel tracer. Further, compartment modeling of this novel tracer also exhibited a greater tumor-to-background ratio, a higher tumor binding potential and a lower brain binding potential when compared with other TSPO probes, such as [18F]DPA-714 and [18F]VUIIS1008. CONCLUSIONS: These studies illustrate that [18F]VUIIS1018A can serve as a promising TSPO PET tracer for glioma imaging and potentially imaging of other solid tumors.

Evaluation of the novel TSPO radiotracer 2-(7-butyl-2-(4-(2-([18F]fluoroethoxy)phenyl)-5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-N,N-diethylacetamide in a preclinical model of neuroinflammation.

Translocator Protein (18 kDa, TSPO) is regarded as a useful biomarker for neuroinflammation imaging. TSPO PET imaging could be used to understand the role of neuroinflammation in brain diseases and as a tool for evaluating novel therapeutic effects. As a promising TSPO probe, [18F]DPA-714 is highly specific and offers reliable quantification of TSPO in vivo. In this study, we further radiosynthesized and evaluated another novel TSPO probe, 2-(7-butyl-2-(4-(2-[18F]fluoroethoxy)phenyl)-5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-N,N-diethylacetamide ([18F]VUIIS1018A), which features a 700-fold higher binding affinity for TSPO than that of [18F]DPA-714. We evaluated the performance of [18F]VUIIS1018A using dynamic in vivo PET imaging, radiometabolite analysis, in vitro autoradiography assays, biodistribution analysis, and blocking assays. In vivo study using this probe demonstrated high signal-to-noise ratio, binding potential (BPND), and binding specificity in preclinical neuroinflammation studies. Taken together, these findings indicate that [18F]VUIIS1018A may serve as a novel TSPO PET probe for neuroinflammation imaging.

Evaluation of TSPO PET Ligands [18F]VUIIS1009A and [18F]VUIIS1009B: Tracers for Cancer Imaging.

PURPOSE: Positron emission tomography (PET) ligands targeting translocator protein (TSPO) are potential imaging diagnostics of cancer. In this study, we report two novel, high-affinity TSPO PET ligands that are 5,7 regioisomers, [18F]VUIIS1009A ([18F]3A) and [18F]VUIIS1009B ([18F]3B), and their initial in vitro and in vivo evaluation in healthy mice and glioma-bearing rats. PROCEDURES: VUIIS1009A/B was synthesized and confirmed by X-ray crystallography. Interactions between TSPO binding pocket and novel ligands were evaluated and compared with contemporary TSPO ligands using 2D 1H-15N heteronuclear single quantum coherence (HSQC) spectroscopy. In vivo biodistribution of [18F]VUIIS1009A and [18F]VUIIS1009B was carried out in healthy mice with and without radioligand displacement. Dynamic PET imaging data were acquired simultaneously with [18F]VUIIS1009A/B injections in glioma-bearing rats, with binding reversibility and specificity evaluated by radioligand displacement. In vivo radiometabolite analysis was performed using radio-TLC, and quantitative analysis of PET data was performed using metabolite-corrected arterial input functions. Imaging was validated with histology and immunohistochemistry. RESULTS: Both VUIIS1009A (3A) and VUIIS1009B (3B) were found to exhibit exceptional binding affinity to TSPO, with observed IC50 values against PK11195 approximately 500-fold lower than DPA-714. However, HSQC NMR suggested that VUIIS1009A and VUIIS1009B share a common binding pocket within mammalian TSPO (mTSPO) as DPA-714 and to a lesser extent, PK11195. [18F]VUIIS1009A ([18F]3A) and [18F]VUIIS1009B ([18F]3B) exhibited similar biodistribution in healthy mice. In rats bearing C6 gliomas, both [18F]VUIIS1009A and [18F]VUIIS1009B exhibited greater binding potential (k 3/k 4)in tumor tissue compared to [18F]DPA-714. Interestingly, [18F]VUIIS1009B exhibited significantly greater tumor uptake (V T) than [18F]VUIIS1009A, which was attributed primarily to greater plasma-to-tumor extraction efficiency. CONCLUSIONS: The novel PET ligand [18F]VUIIS1009B exhibits promising characteristics for imaging glioma; its superiority over [18F]VUIIS1009A, a regioisomer, appears to be primarily due to improved plasma extraction efficiency. Continued evaluation of [18F]VUIIS1009B as a high-affinity TSPO PET ligand for precision medicine appears warranted.

Facile synthesis of SSR180575 and discovery of 7-chloro-N,N,5-trimethyl-4-oxo-3(6-[(18)F]fluoropyridin-2-yl)-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide, a potent pyridazinoindole ligand for PET imaging of TSPO in cancer.

A novel synthesis of the translocator protein (TSPO) ligand 7-chloro-N,N,5-trimethyl-4-oxo-3-phenyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide (SSR180575, 3) was achieved in four steps from commercially available starting materials. Focused structure-activity relationship development about the pyridazinoindole ring at the N3 position led to the discovery of 7-chloro-N,N,5-trimethyl-4-oxo-3(6-fluoropyridin-2-yl)-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide (14), a novel ligand of comparable affinity. Radiolabeling with fluorine-18 ((18)F) yielded 7-chloro-N,N,5-trimethyl-4-oxo-3(6-[(18)F]fluoropyridin-2-yl)-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide ([(18)F]-14) in high radiochemical yield and specific activity. In vivo studies of [(18)F]-14 revealed this agent as a promising probe for molecular imaging of glioma.

Preclinical imaging evaluation of novel TSPO-PET ligand 2-(5,7-Diethyl-2-(4-(2-[(18)F]fluoroethoxy)phenyl)pyrazolo[1,5-a]pyrimidin-3-yl)-N,N-diethylacetamide ([ (18)F]VUIIS1008) in glioma.

PURPOSE: Translocator protein (TSPO) concentrations are elevated in glioma, suggesting a role for TSPO positron emission tomography (PET) imaging in this setting. In preclinical PET studies, we evaluated a novel, high-affinity TSPO PET ligand, [(18)F]VUIIS1008, in healthy mice and glioma-bearing rats. PROCEDURES: Dynamic PET data were acquired simultaneously with [(18)F]VUIIS1008 injection, with binding reversibility and specificity evaluated in vivo by non-radioactive ligand displacement or blocking. Compartmental analysis of PET data was performed using metabolite-corrected arterial input functions. Imaging was validated with histology and immunohistochemistry. RESULTS: [(18)F]VUIIS1008 exhibited rapid uptake in TSPO-rich organs. PET ligand uptake was displaceable with non-radioactive VUIIS1008 or PBR06 in mice. Tumor accumulation of [(18)F]VUIIS1008 was blocked by pretreatment with VUIIS1008 in rats. [(18)F]VUIIS1008 exhibited improved tumor-to-background ratio and higher binding potential in tumors compared to a structurally similar pyrazolopyrimidine TSPO ligand, [(18)F]DPA-714. CONCLUSIONS: The PET ligand [(18)F]VUIIS1008 exhibits promising characteristics as a tracer for imaging glioma. Further translational studies appear warranted.

A peptide-based positron emission tomography probe for in vivo detection of caspase activity in apoptotic cells.

PURPOSE: Apoptosis, or programmed cell death, can be leveraged as a surrogate measure of response to therapeutic interventions in medicine. Cysteine aspartic acid-specific proteases, or caspases, are essential determinants of apoptosis signaling cascades and represent promising targets for molecular imaging. Here, we report development and in vivo validation of [(18)F]4-fluorobenzylcarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone ([(18)F]FB-VAD-FMK), a novel peptide-based molecular probe suitable for quantification of caspase activity in vivo using positron emission tomography (PET). EXPERIMENTAL DESIGN: Supported by molecular modeling studies and subsequent in vitro assays suggesting probe feasibility, the labeled pan-caspase inhibitory peptide, [(18)F]FB-VAD-FMK, was produced in high radiochemical yield and purity using a simple two-step, radiofluorination. The biodistribution of [(18)F]FB-VAD-FMK in normal tissue and its efficacy to predict response to molecularly targeted therapy in tumors was evaluated using microPET imaging of mouse models of human colorectal cancer. RESULTS: Accumulation of [(18)F]FB-VAD-FMK was found to agree with elevated caspase-3 activity in response to Aurora B kinase inhibition as well as a multidrug regimen that combined an inhibitor of mutant BRAF and a dual PI3K/mTOR inhibitor in (V600E)BRAF colon cancer. In the latter setting, [(18)F]FB-VAD-FMK PET was also elevated in the tumors of cohorts that exhibited reduction in size. CONCLUSIONS: These studies illuminate [(18)F]FB-VAD-FMK as a promising PET imaging probe to detect apoptosis in tumors and as a novel, potentially translatable biomarker for predicting response to personalized medicine.

Microwave-assisted, one-pot reaction of 7-azaindoles and aldehydes: a facile route to novel di-7-azaindolylmethanes.

A novel and highly efficient synthetic method leveraging microwave-assisted organic synthesis (MAOS) to yield di-7-azaindolylmethanes (DAIMs) is reported. Under MAOS conditions, reaction of 7-azaindole with aldehydes resulted predominantly in DAIMs, as opposed to the expected 7-azaindole addition products that form at ambient temperature. Based upon studies of different indoles and azaindoles with various aromatic and aliphatic aldehydes, we herein propose a mechanism where rapid and efficient microwave heating promotes nucleophilicity of 7-azaindoles towards the corresponding alkylidene-azaindolene intermediate to form the DAIM. This sequence provides a versatile approach to efficiently synthesize novel DAIMs that may be useful pharmaceuticals.

Synthesis and structure-activity relationships of 5,6,7-substituted pyrazolopyrimidines: discovery of a novel TSPO PET ligand for cancer imaging.

Focused library synthesis and structure-activity relationship development of 5,6,7-substituted pyrazolopyrimidines led to the discovery of 2-(5,7-diethyl-2-(4-(2-fluoroethoxy)phenyl)pyrazolo[1,5-a]pyrimidin-3-yl)-N,N-diethylacetamide (6b), a novel translocator protein (TSPO) ligand exhibiting a 36-fold enhancement in affinity compared to another pyrazolopyrimidine-based TSPO ligand, 6a (DPA-714). Radiolabeling with fluorine-18 ((18)F) facilitated production of 2-(5,7-diethyl-2-(4-(2-[(18)F]fluoroethoxy)phenyl)pyrazolo[1,5-a]pyrimidin-3-yl)-N,N-diethylacetamide ((18)F-6b) in high radiochemical yield and specific activity. In vivo studies of (18)F-6b were performed which illuminated this agent as an improved probe for molecular imaging of TSPO-expressing cancers.

Quantitative preclinical imaging of TSPO expression in glioma using N,N-diethyl-2-(2-(4-(2-18F-fluoroethoxy)phenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-3-yl)acetamide.

UNLABELLED: There is a critical need to develop and rigorously validate molecular imaging biomarkers to aid diagnosis and characterization of primary brain tumors. Elevated expression of translocator protein (TSPO) has been shown to predict disease progression and aggressive, invasive behavior in a variety of solid tumors. Thus, noninvasive molecular imaging of TSPO expression could form the basis of a novel, predictive cancer imaging biomarker. In quantitative preclinical PET studies, we evaluated a high-affinity pyrazolopyrimidinyl-based TSPO imaging ligand, N,N-diethyl-2-(2-(4-(2-(18)F-fluoroethoxy)phenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-3-yl)acetamide ((18)F-DPA-714), as a translational probe for quantification of TSPO levels in glioma. METHODS: Glioma-bearing rats were imaged with (18)F-DPA-714 in a small-animal PET system. Dynamic images were acquired simultaneously on injection of (18)F-DPA-714 (130-200 MBq/0.2 mL). Blood was collected to derive the arterial input function (AIF), with high-performance liquid chromatography radiometabolite analysis performed on selected samples for AIF correction. Compartmental modeling was performed using the corrected AIF. Specific tumor cell binding of DPA-714 was evaluated by radioligand displacement of (3)H-PK 11195 with DPA-714 in vitro and displacement of (18)F-DPA-714 with an excess of DPA-714 in vivo. Immediately after imaging, tumor and healthy brain tissues were harvested for validation by Western blotting and immunohistochemistry. RESULTS: (18)F-DPA-714 was found to preferentially accumulate in tumors, with modest uptake in the contralateral brain. Infusion with DPA-714 (10 mg/kg) displaced (18)F-DPA-714 binding by greater than 60% on average. Tumor uptake of (18)F-DPA-714 was similar to another high-affinity TSPO imaging ligand, (18)F-N-fluoroacetyl-N-(2,5-dimethoxybenzyl)-2-phenoxyaniline, and agreed with ex vivo assay of TSPO levels in tumor and healthy brain. CONCLUSION: These studies illustrate the feasibility of using (18)F-DPA-714 for visualization of TSPO-expressing brain tumors. Importantly, (18)F-DPA-714 appears suitable for quantitative assay of tumor TSPO levels in vivo. Given the relationship between elevated TSPO levels and poor outcome in oncology, these studies suggest the potential of (18)F-DPA-714 PET to serve as a novel predictive cancer imaging modality.

Quantitative, preclinical PET of translocator protein expression in glioma using 18F-N-fluoroacetyl-N-(2,5-dimethoxybenzyl)-2-phenoxyaniline.

UNLABELLED: Translocator protein (TSPO), also referred to as peripheral benzodiazepine receptor (PBR), is a crucial 18-kDa outer mitochondrial membrane protein involved in numerous cellular functions, including the regulation of cholesterol metabolism, steroidogenesis, and apoptosis. Elevated expression of TSPO in oncology correlates with disease progression and poor survival, suggesting that molecular probes capable of assaying TSPO levels may have potential as cancer imaging biomarkers. In preclinical PET studies, we characterized a high-affinity aryloxyanilide-based TSPO imaging ligand, 18F-N-fluoroacetyl-N-(2,5-dimethoxybenzyl)-2-phenoxyaniline (18F-PBR06), as a candidate probe for the quantitative assessment of TSPO expression in glioma. METHODS: Glioma-bearing rats were imaged with 18F-PBR06 in a small-animal PET system. Dynamic images were acquired simultaneously on injection of 18F-PBR06 (70-100 MBq/0.2 mL). Over the course of scanning, arterial blood was collected to derive the input function, with high-performance liquid chromatography radiometabolite analysis performed on selected samples for arterial input function correction. Compartmental modeling of the PET data was performed using the corrected arterial input function. Specific tumor cell binding of PBR06 was evaluated by radioligand displacement of 3H-PK 11195 with PBR06 in vitro and by displacement of 18F-PBR06 with excess PBR06 in vivo. Immediately after imaging, tumor tissue and adjacent healthy brain were harvested for assay of TSPO protein levels by Western blotting and immunohistochemistry. RESULTS: 18F-PBR06 was found to preferentially accumulate in tumors, with modest uptake in the contralateral brain, facilitating excellent contrast between tumor and adjacent tissue. Infusion with PBR06 (10 mg/kg) displaced 18F-PBR06 binding by approximately 75%. The accumulation of 18F-PBR06 in tumor tissues and adjacent brain agreed with the ex vivo assay of TSPO protein levels by Western blotting and quantitative immunohistochemistry. CONCLUSION: These preclinical studies illustrate that 18F-PBR06 is a promising tracer for visualization of TSPO-expressing tumors. Importantly, the close correlation between 18F-PBR06 uptake and TSPO expression in tumors and normal tissues, coupled with the high degree of displaceable binding from both tumors and the normal brain, represents a significant improvement over other TSPO imaging ligands previously evaluated in glioma. These data suggest the potential of 18F-PBR06 to elucidate the role of TSPO in oncology, as well as its potential development as a cancer imaging biomarker.

Microwave-assisted Organic Synthesis of a High-affinity Pyrazolo-pyrimidinyl TSPO Ligand.

We herein report a dramatically improved total synthesis of the high-affinity translocator protein (TSPO) ligand DPA-714, featuring microwave-assisted organic synthesis (MAOS). Compared with previously described approaches, our novel MAOS method dramatically reduces overall reaction time without adversely effecting reaction yields. We envision that the described MAOS protocol may be suitably applied to high-throughput, diversity-oriented synthesis of novel compounds based on the pyrazolo-pyrimidinyl scaffold. Such an approach could accelerate the development of focused libraries of novel TSPO ligands with potential for future development as molecular imaging and therapeutic agents.