Mark Lazari

Titania-catalyzed radiofluorination of tosylated precursors in highly aqueous medium.

Nucleophilic radiofluorination is an efficient synthetic route to many positron-emission tomography (PET) probes, but removal of water to activate the cyclotron-produced [(18)F]fluoride has to be performed prior to reaction, which significantly increases overall radiolabeling time and causes radioactivity loss. In this report, we demonstrate the possibility of (18)F-radiofluorination in highly aqueous medium. The method utilizes titania nanoparticles, 1:1 (v/v) acetonitrile-thexyl alcohol solvent mixture, and tetra-n-butylammonium bicarbonate as a phase-transfer agent. Efficient radiolabeling is directly performed with aqueous [(18)F]fluoride without the need for a drying/azeotroping step to significantly reduce radiosynthesis time. High radiochemical purity of the target compound is also achieved. The substrate scope of the synthetic strategy is demonstrated with a range of aromatic, aliphatic, and cycloaliphatic tosylated precursors.

Fully automated production of diverse 18F-labeled PET tracers on the ELIXYS multireactor radiosynthesizer without hardware modification.

Fully automated radiosynthesizers are continuing to be developed to meet the growing need for the reliable production of PET tracers made under current good manufacturing practice guidelines. There is a current trend toward supporting kitlike disposable cassettes that come preconfigured for particular tracers, thus eliminating the need for cleaning protocols between syntheses and enabling quick transitions to synthesizing other tracers. Though ideal for production, these systems are often limited for the development of novel tracers because of pressure, temperature, and chemical compatibility considerations. This study demonstrated the versatile use of the ELIXYS fully automated radiosynthesizer to adapt and produce 8 different 18F-labeled PET tracers of varying complexity. Methods: Three-reactor syntheses of 2-deoxy-2-18F-fluoro-β-d-arabinofuranosylcytosine (d-18F-FAC), 2-deoxy-2-18F-fluoro-5-methyl-β-l-arabinofuranosyluracil (l-18F-FMAU), and 2-deoxy-2-18F-fluoro-5-ethyl-β-d-arabinofuranosyluracil (d-18F-FEAU) along with the 1-reactor syntheses of d-18F-FEAU, 18F-FDG, 3-deoxy-3-18F-fluoro-l-thymidine (18F-FLT), 18F-fallypride, 9-(4-18F-fluoro-3-hydroxymethylbutyl)-guanine (18F-FHBG), and N-succinimidyl-4-18F-fluorobenzoate (18F-SFB), were all produced using ELIXYS without the need for any hardware modifications or reconfiguration. Synthesis protocols were adapted and slightly modified from those in the literature but were not fully optimized. Furthermore, 18F-FLT, 18F-FDG, and 18F-fallypride were produced sequentially on the same day and used for preclinical imaging of A431 tumor–bearing severe combined immunodeficient mice and wild-type BALB/c mice. To assess future translation to the clinical setting, several batches of tracers were subjected to a full set of quality control tests. Results: All tracers were produced with radiochemical yields comparable to those in the literature. 18F-FLT, 18F-FDG, and 18F-fallypride were successfully used to image the mice, with results consistent with those reported in the literature. All tracers that were subjected to clinical quality control tests passed. Conclusion: The ELIXYS radiosynthesizer facilitates rapid tracer development and is capable of producing multiple 18F-labeled PET tracers suitable for clinical applications using the same hardware setup.

“Clicked” fluoropolymer elastomers as robust materials for potential microfluidic device applications

We report the design and synthesis of a new perfluoropolyether-based material, which has liquid-like viscosity and can be cured into a tough, highly durable elastomer when “clicked” with selected tri-pod organic small molecules. This highly fluorinated elastomer exhibits remarkable resistance to a variety of organic solvents, water, heat and even harsh acidic and basic conditions. Whereas PDMS-based microfluidic devices are commonly used for aqueous based applications, their limited chemical resistance and high swellability in many common organic solvents make it unfeasible for microfluidic applications involving organic solvents and/or harsh conditions. With excellent chemical resistance and low swellability, our newly synthesized fluoro-elastomers will hopefully provide an alternative material for organic based microfluidic devices. Furthermore, the alkyne–azide “click” chemistry employed in curing not only provides high efficiency of synthesis and ease of device fabrication, but, more importantly, produces 1,2,3-triazole linkages that are very stable against harsh acidic or basic conditions. This work has great potential to expand microfluidics to a series of novel applications especially in organic and medicinal chemistry.

ELIXYS - a fully automated, three-reactor high-pressure radiosynthesizer for development and routine production of diverse PET tracers

BackgroundAutomated radiosynthesizers are vital for routine production of positron-emission tomography tracers to minimize radiation exposure to operators and to ensure reproducible synthesis yields. The recent trend in the synthesizer industry towards the use of disposable kits aims to simplify setup and operation for the user, but often introduces several limitations related to temperature and chemical compatibility, thus requiring reoptimization of protocols developed on non-cassette-based systems. Radiochemists would benefit from a single hybrid system that provides tremendous flexibility for development and optimization of reaction conditions while also providing a pathway to simple, cassette-based production of diverse tracers.MethodsWe have designed, built, and tested an automated three-reactor radiosynthesizer (ELIXYS) to provide a flexible radiosynthesis platform suitable for both tracer development and routine production. The synthesizer is capable of performing high-pressure and high-temperature reactions by eliminating permanent tubing and valve connections to the reaction vessel. Each of the three movable reactors can seal against different locations on disposable cassettes to carry out different functions such as sealed reactions, evaporations, and reagent addition. A reagent and gas handling robot moves sealed reagent vials from storage locations in the cassette to addition positions and also dynamically provides vacuum and inert gas to ports on the cassette. The software integrates these automated features into chemistry unit operations (e.g., React, Evaporate, Add) to intuitively create synthesis protocols. 2-Deoxy-2-[18F]fluoro-5-methyl-β-l-arabinofuranosyluracil (l-[18F]FMAU) and 2-deoxy-2-[18F]fluoro-β-d-arabinofuranosylcytosine (d-[18F]FAC) were synthesized to validate the system.Resultsl-[18F]FMAU and d-[18F]FAC were successfully synthesized in 165 and 170 min, respectively, with decay-corrected radiochemical yields of 46% ± 1% (n = 6) and 31% ± 5% (n = 6), respectively. The yield, repeatability, and synthesis time are comparable to, or better than, other reports. d-[18F]FAC produced by ELIXYS and another manually operated apparatus exhibited similar biodistribution in wild-type mice.ConclusionThe ELIXYS automated radiosynthesizer is capable of performing radiosyntheses requiring demanding conditions: up to three reaction vessels, high temperatures, high pressures, and sensitive reagents. Such flexibility facilitates tracer development and the ability to synthesize multiple tracers on the same system without customization or replumbing. The disposable cassette approach simplifies the transition from development to production.

The separation and detection of PET tracers via capillary electrophoresis for chemical identity and purity analysis.

CE coupled with UV detection was assessed as a possible platform for the chemical identity and purity analysis of positron emission tomography (PET) tracers using [(18)F]FAC and [(18)F]FLT as examples. Representative samples containing mixtures of the tracers plus well-known impurities, as well as real radioactive samples (formulated for injection), were analyzed. Using MEKC with SDS in a neutral phosphate buffer, the separation of all compounds in the samples was achieved with baseline resolutions in less than 4.5min and 3min for FLT and FAC samples, respectively. In comparison to the gold-standard for chemical analysis (i.e. HPLC/UV), we have demonstrated improvements in analysis times, and comparable LOD. Although the reproducibility in migration time is slightly lower than that of the HPLC, identification of the compounds was still possible due to good peak separation. In addition, we show that CE can be used to identify and quantify Krytofix2.2.2 (a toxic and commonly used phase transfer catalyst) in less than 2min and with a LOD of 45μg/mL (non-optimized). These results demonstrate adequate performance for chemical identity and purity analysis. Combined with the potential for miniaturization into a microchip format, these results suggest the potential of CE as an integral part of a miniaturized quality control system for PET tracers.

Simplified programming and control of automated radiosynthesizers through unit operations.

BackgroundMany automated radiosynthesizers for producing positron emission tomography (PET) probes provide a means for the operator to create custom synthesis programs. The programming interfaces are typically designed with the engineer rather than the radiochemist in mind, requiring lengthy programs to be created from sequences of low-level, non-intuitive hardware operations. In some cases, the user is even responsible for adding steps to update the graphical representation of the system. In light of these unnecessarily complex approaches, we have created software to perform radiochemistry on the ELIXYS radiosynthesizer with the goal of being intuitive and easy to use.MethodsRadiochemists were consulted, and a wide range of radiosyntheses were analyzed to determine a comprehensive set of basic chemistry unit operations. Based around these operations, we created a software control system with a client–server architecture. In an attempt to maximize flexibility, the client software was designed to run on a variety of portable multi-touch devices. The software was used to create programs for the synthesis of several 18F-labeled probes on the ELIXYS radiosynthesizer, with [18F]FDG detailed here. To gauge the user-friendliness of the software, program lengths were compared to those from other systems. A small sample group with no prior radiosynthesizer experience was tasked with creating and running a simple protocol.ResultsThe software was successfully used to synthesize several 18F-labeled PET probes, including [18F]FDG, with synthesis times and yields comparable to literature reports. The resulting programs were significantly shorter and easier to debug than programs from other systems. The sample group of naive users created and ran a simple protocol within a couple of hours, revealing a very short learning curve. The client–server architecture provided reliability, enabling continuity of the synthesis run even if the computer running the client software failed. The architecture enabled a single user to control the hardware while others observed the run in progress or created programs for other probes.ConclusionsWe developed a novel unit operation-based software interface to control automated radiosynthesizers that reduced the program length and complexity and also exhibited a short learning curve. The client–server architecture provided robustness and flexibility.

Fully-automated synthesis of 16β-18F-fluoro-5α-dihydrotestosterone (FDHT) on the ELIXYS radiosynthesizer

Noninvasive in vivo imaging of androgen receptor (AR) levels with positron emission tomography (PET) is becoming the primary tool in prostate cancer detection and staging. Of the potential (18)F-labeled PET tracers, (18)F-FDHT has clinically shown to be of highest diagnostic value. We demonstrate the first automated synthesis of (18)F-FDHT by adapting the conventional manual synthesis onto the fully-automated ELIXYS radiosynthesizer. Clinically-relevant amounts of (18)F-FDHT were synthesized on ELIXYS in 90 min with decay-corrected radiochemical yield of 29±5% (n=7). The specific activity was 4.6 Ci/µmol (170 GBq/µmol) at end of formulation with a starting activity of 1.0 Ci (37 GBq). The formulated (18)F-FDHT yielded sufficient activity for multiple patient doses and passed all quality control tests required for routine clinical use.

On-chip product purification for complete microfluidic radiotracer synthesis

Solid phase extraction was incorporated into an electrowetting-on-dielectric chip for radiochemical purification of a positron emission tomography tracer that was radiolabeled on the same chip. The radiotracer droplet was mixed with alumina particles, and the alumina particles were filtered out from the droplet through a line of pillars, all by electrowetting droplet movement. Fluorination reaction and on-chip purification were analyzed with both Cerenkov imaging and off-chip radio-thin layer chromatography measurements. The measurements were compared to test the validity of the combined use of filtration on-chip and Cerenkov imaging as an alternative approach for monitoring reaction yield without the need to extract sample from the chip.

Performing radiosynthesis in microvolumes to maximize molar activity of tracers for positron emission tomography

Positron emission tomography (PET) is a molecular diagnostic imaging technology to quantitatively visualize biological processes in vivo. For many applications, including imaging of low-tissue density targets (e.g., neuroreceptors), imaging in small animals, and evaluation of novel tracers, the injected PET tracer must be produced with high molar activity to ensure low occupancy of biological targets and avoid pharmacologic effects. Additionally, high molar activity is essential for tracers with lengthy syntheses or tracers transported to distant imaging sites. Here we show that radiosynthesis of PET tracers in microliter volumes instead of conventional milliliter volumes results in substantially increased molar activity, and we identify the most relevant variables affecting this parameter. Furthermore, using the PET tracer [18F]fallypride, we illustrate that molar activity can have a significant impact on biodistribution. With full automation, microdroplet platforms could provide a means for radiochemists to routinely, conveniently, and safely produce PET tracers with high molar activity.For many applications, positron emission tomography tracers must be produced with high specific activity. Here the authors identify variables leading to increased specific activity when tracers are synthesized in microliter volumes, and show that specific activity can influence tracer biodistribution.

Production of diverse PET probes with limited resources: 24 18F-labeled compounds prepared with a single radiosynthesizer

Significance Molecular imaging with PET can provide a dynamic, whole-body picture of the rate of biological processes or distribution of biological targets by tracking the distribution of radiolabeled molecules or particles in the body over time. Continual efforts to develop new PET probes are expanding the variety of processes and targets that can be visualized, facilitating basic research, drug development, and patient care. However, access to these probes at all stages of their development is hindered by high costs arising, in large part, from the significant resources that are typically dedicated to production of a single probe. Emerging technologies with increased synthesis flexibility are allowing increased probe diversity with fewer resources and could significantly increase access to new molecular imaging agents. New radiolabeled probes for positron-emission tomography (PET) are providing an ever-increasing ability to answer diverse research and clinical questions and to facilitate the discovery, development, and clinical use of drugs in patient care. Despite the high equipment and facility costs to produce PET probes, many radiopharmacies and radiochemistry laboratories use a dedicated radiosynthesizer to produce each probe, even if the equipment is idle much of the time, to avoid the challenges of reconfiguring the system fluidics to switch from one probe to another. To meet growing demand, more cost-efficient approaches are being developed, such as radiosynthesizers based on disposable “cassettes,” that do not require reconfiguration to switch among probes. However, most cassette-based systems make sacrifices in synthesis complexity or tolerated reaction conditions, and some do not support custom programming, thereby limiting their generality. In contrast, the design of the ELIXYS FLEX/CHEM cassette-based synthesizer supports higher temperatures and pressures than other systems while also facilitating flexible synthesis development. In this paper, the syntheses of 24 known PET probes are adapted to this system to explore the possibility of using a single radiosynthesizer and hot cell for production of a diverse array of compounds with wide-ranging synthesis requirements, alongside synthesis development efforts. Most probes were produced with yields and synthesis times comparable to literature reports, and because hardware modification was unnecessary, it was convenient to frequently switch among probes based on demand. Although our facility supplies probes for preclinical imaging, the same workflow would be applicable in a clinical setting.