Microliter-scale reaction arrays for economical high-throughput experimentation in radiochemistry

Abstract

The increasing number of positron-emission tomography (PET) tracers being developed to aid drug development and create new diagnostics has led to an increased need for radiosynthesis development and optimization. Current radiosynthesis instruments are designed to produce large-scale clinical batches and are often limited to performing a single synthesis before they must be decontaminated by waiting for radionuclide decay, followed by thorough cleaning or disposal of synthesizer components. Though with some radiosynthesizers it is possible to perform a few sequential radiosyntheses in a day, none allow for parallel radiosyntheses. Throughput of one or a few experiments per day is not well suited for rapid optimization experiments. To combat these limitations, we leverage the advantages of droplet-radiochemistry to create a new platform for high-throughput experimentation in radiochemistry. This system contains an array of 4 heaters, each used to heat a set of 16 reactions on a small chip, enabling 64 parallel reactions for the rapid optimization of conditions in any stage of a multi-step radiosynthesis process. As examples, we study the syntheses of several 18F-labeled radiopharmaceuticals ([18F]Flumazenil, [18F]PBR06, [18F]Fallypride, and [18F]FEPPA), performing > 800 experiments to explore the influence of parameters including base type, base amount, precursor amount, solvent, reaction temperature, and reaction time. The experiments were carried out within only 15 experiment days, and the small volume (~ 10 μL compared to the ~ 1 mL scale of conventional instruments) consumed ~ 100 × less precursor per datapoint. This new method paves the way for more comprehensive optimization studies in radiochemistry and substantially shortening PET tracer development timelines.

Rapid Purification and Formulation of Radiopharmaceuticals via Thin-Layer Chromatography

“…Compared to our prior reports of droplet radiosyntheses that used analytical-scale HPLC purification (with purification efficiency of ~80%), the efficiency of the TLC purification and formulation process was significantly higher (nearly quantitative), leading to higher overall radiochemical yield. In particular, a prior report of droplet-based [ 18 F]PBR-06 production showed high crude RCY (94 ± 2%, n = 4), but due to losses during HPLC purification, the isolated RCY was only 76% ( n = 1) [ 27 ], and further losses would have been expected during downstream formulation, which was not performed in that study. Similarly, a prior report of droplet-based [ 18 F]Fallypride production exhibited high crude RCY (96 ± 2%, n = 4), but, due to losses during HPLC purification, the isolated yield was 78% ( n = 1) [ 26 ].…”

“…[ 18 F]PBR-06 and [ 18 F]Fallypride were prepared using droplet radiochemistry methods on Teflon-coated silicon surface tension trap chips [ 26 ]. Detailed protocols for preparing these radiotracers have been previously reported [ 27 ]. Stock solutions of reference standards were prepared at 20 mM concentrations: 5 mg of Fallypride was added to 685 µL of MeOH, and 5 mg of PBR-06 was added to 632 µL of MeOH.…”

“…Recently, our group and others have shown that microscale synthesis methods enable efficient reactions while enabling a vast reduction of reagent masses [ 25 , 26 , 27 , 28 , 29 ]. Consequently, the quantity of impurities is drastically reduced, and it appears in some cases that the number of different impurities may also be reduced [ 28 , 30 ].…”

Scalable droplet-based radiosynthesis of [18F]fluorobenzyltriphenylphosphonium cation ([18F]FBnTP) via a “numbering up” approach

“…A further advantage of droplet radiochemistry is the ability to perform high-throughput optimization via arrays of droplet reactions performed in parallel. 21,22 Using a newly developed robotic platform, we used this technique to develop a preliminary droplet-based radiosynthesis of [ 18 F]FBnTP, 23 resulting in substantial reduction in reagent usage and enhancement in radiosynthesis performance (RCC: 89 ± 1%, n = 4). Following purification and formulation, [ 18 F]FBnTP was produced with high isolated radiochemical yield (RCY, 66 ± 6%, n = 3) within 42 min, corresponding to an activity yield of 49 ± 3% ( n = 3).…”

“…In addition, a modest drop in RCY was observed, potentially due to the increased amount of impurities present when using large volumes of radioisotope source solution. 22 Conversely, the second approach effectively worked with much larger volumes and avoids the build-up of impurities, but required a more complex setup. Furthermore, this approach requires optimization of the [ 18 F]fluoride elution protocol for each radiotracer because the type and amount of phase transfer catalyst (PTC) and base needed for efficient elution can impact the subsequent radiotracer synthesis.…”