Production of 211At and radiosynthesis of 211At-MABG
211At was produced through the 209Bi(α,2n)211At reaction using a Sumitomo multipurpose cyclotron (MP-30, Sumitomo Heavy Industries, Ltd. Japan) in the Advanced Clinical Research Center at Fukushima Medical University, Japan. A 30 MeV alpha-particle beam was degraded to 26.5 ± 0.9 MeV by inserting 70 μm of aluminum foil to prevent the production of 210At. The degraded beam was used to bombard the bismuth (99.999%, Goodfellow Cambridge Ltd., Huntingdon, England) layer on an aluminum backing for 110 min with 10.9 eμA. 211At was isolated from the irradiated target using the dry distillation procedure reported by Lindegren et al. [23] with slight modifications. Briefly, the target was inserted in a quartz tube placed in a tube furnace preheated to 700 °C. Under helium gas flow in a 700 °C oven, vaporized 211At was transported from the quartz tube to an externally connected PTFE tube that was immersed in dry ice/ethanol bath. The cooled 211At was trapped in the PTFE tube, eluted, and recovered using 0.5 mL of chloroform. The radioactivity of 211At was measured using a dose calibrator (CRC-25R, Capintec Inc., Ramsey, NJ, USA), which was previously calibrated by measuring a highly radioactive 211At source using a Ge detector (GEM30-70, ORTEC, Oak Ridge, TN, USA) and a dose calibrator. For the quantification of 211At radioactivity on a Ge detector, we selected 687.0 keV (gamma-ray intensity: Ir = 0.261%) of gamma-ray from 211At, and 569.65 keV (0.311%, against the decay of 211At) and 897.8 keV (0.321%, against the decay of 211At) from the daughter nuclide 211Po. Gamma-ray spectrometry was also performed using a Ge detector to assign the radionuclides produced in the target and the recovery solution of 211At. The radioactivity of 211At was 263.3 MBq at the end of the bombardment, and the radiochemical purity of 211At was more than 99.9% at the end of recovery; there was no contamination of 210At. Chloroform, the recovery solvent of 211At, was added to 50 μL of 0.1 M NaOH aqueous solution and then removed with nitrogen gas. The remaining 211At was redissolved in 3 mL of saline and administered to mice. In this study, 211At refers to “free astatine,” which likely consists not only of 211At-, but also, to some extent, other oxidation states [24].
211At-MABG was prepared in accordance with a slightly modified previously published method [25, 26]. Namely, 211At in chloroform and meta-trimethylsilylbenzylguanidine hemisulfate with N-chlorosuccinimide were dissolved in trifluoroacetic acid and heated at 70 °C for 10 min. Crude 211At-MABG was purified with a Sep-Pak tC18 Plus Light Cartridge (Waters, Milford, MA). After washing with water, 211At-MABG was eluted with 5% ethanol aqueous solution, providing a radiochemical yield of 36.3% (decay corrected). The eluate was diluted with saline, and sodium ascorbate was added at a final concentration of 2.5%. The radiochemical purity of 211At-MABG was determined using reverse-phase radio-high-performance liquid chromatography (radio-HPLC), and the value was > 98%.
Biodistribution study
The experimental procedures and care of animals were carried out with the approval of the Fukushima Medical University Institute of Animal Care and Use Committee. Normal male mice (C57BL/6 N, 9 weeks old) were administered 0.13 MBq of free 211At or 0.20 MBq of 211At-MABG by tail-vein injection. The mice were sacrificed at 5 min, and at 1, 3, 6, and 24 h after each tracer injection (n = 5 in each group). The radioactivities in organs (muscle, heart, lung, spleen, pancreas, white adipocyte, testis, stomach, small intestine inclusive of contents, large intestine inclusive of contents, kidneys, adrenal glands, liver, brown adipocyte, salivary gland, thyroid gland, bone, and brain) and blood were measured using a γ-counter (Wizard2®, Perkin Elmer, MA, USA). The activities in the abovementioned organs and blood were determined as the percentage of injected activity per mass (%IA/g), whereas that in the thyroid gland was determined as %IA because the gland could not be weighed accurately.
Radiation absorbed dose calculations
The data of biodistribution in the mice were used to estimate not only the mouse radiation absorbed doses but also the human radiation absorbed doses for both free 211At and 211At-MABG. The mean radioactivity in mouse organs at 5 min, and at 1, 3, 6, and 24 h (n = 5 in each group) was used to calculate the time-integrated activity coefficient (Bq-h/Bq) for each organ. The calculated %IA/organ in mouse and human organs was fitted with an exponential function and integrated to obtain the number of disintegrations (time-integrated activity coefficient) for source organs using the OLINDA/EXM version 1.1 software. One to three exponential terms can be selected for the modeling process [19]. An indirect blood-based method using patient-based red marrow-to-blood ratio (RMBLR) and bone marrow mass was used to determine bone marrow self-dose. The red marrow cumulated activity (ARM) is generally determined using the following equation (1) [27, 28]:
$$ {A}_{RM}=\left[{A}_{blood}\right]\times RMBLR\times {m}_{RM- phantom}, $$
(1)
where [Ablood] is the blood cumulated activity concentration obtained from serial whole-blood sampling and analysis of the resulting blood activity concentration-time curve, and mRM-phantom is the red marrow mass (kg) of the male human phantom. The RMBLR is a correction factor representing the marrow-to-blood activity concentration ratio. The RMBLR reported by previous studies was 0.36 [27, 28].
Using the percent kg/g method [29] with mass extrapolation of 73.0 kg (ICRP 89 adult male phantom), we extrapolated data to human dosimetry. In this method, the human %ID/organ is calculated using the following equation (2) [29, 30]:
$$ {\left(\% ID\right)}_{human}=\left[{\left(\frac{\% ID}{m_{organ}}\right)}_{animal}\times {\left({m}_{TB}\right)}_{animal}\right]\times {\left(\frac{m_{organ}}{m_{TB}}\right)}_{human}, $$
(2)
where morgan is the organ mass and mTB is the total body mass. The human body mass and organ masses were taken from OLINDA/EXM version 2.0 for adult male phantoms. The mouse body mass and organ masses were measured. For the thyroid, a mass of 14 mg for a 25 g mouse model installed in OLINDA/EXM version 2.0 was used for calculations because the gland could not be weighed accurately by the kg/g method. The 25 g mouse model is the closest to the average body mass (23.3 ± 1.2 g) of the mice used in this experiment.
OLINDA/EXM version 2.0 and IDAC-Dose 2.1 were used for human absorbed dose calculations. For animal absorbed dose assessment, only OLINDA/EXM version 2.0 was used. The absorbed dose contribution from the daughter nuclides can also be included in the absorbed dose calculations.
