Astatine solution and method for producing same

Abstract

The present invention aims to provide an astatine solution that can be used in RI internal radiotherapy and the 5 like for the treatment of thyroid gland disease and the like, and a production method thereof. The present invention provides a method for producing a solution containing 211At- (astatide ion) at a high radiochemical purity by using 211At obtained by a nuclear 10 reaction as a starting material, including a step of adding a reducing agent to a solution containing an impurity derived from 211 At; and a solution containing 211At- (astatide ion) at a radiochemical purity of not less than 30%.20 24 26 69 68 28 N ov 2 02 4 A B S T R A C T 2 0 2 4 2 6 6 9 6 8 2 8 N o v 2 0 2 4

Description

Title of Invention: ASTATINE SOLUTION AND METHOD FOR PRODUCING SAME [Related Application] 5

[0001] This application is a divisional application of Australian application no. 2018395495, the entire disclosure of which is incorporated herein by reference. [Technical Field] 10

[0002] The present invention relates to a solution containing 211At_ (astatide ion) at a high radiochemical purity, and a production method thereof. [Background Art] 15

[0003] Radioisotope (RI) internal radiotherapy using sodium iodide (Na131!) is widely used for treating thyroid gland diseases such as thyroid cancer and hyperthyroidism (Graves' disease, etc.). The thyroid gland takes in iodide ions through 20 a sodium iodide symporter (hereinafter abbreviated as NIS). Thus, when Na131! is administered, iodide ion (131I_) accumulates specifically in the thyroid gland and thyroid cancer, and can treat by irradiating the affected area with [3 rays emitted from inside the body. However, internal radiotherapy using Na131! 25 requires hospitalization of patients, the patients are isolated in a special room for radiation control, and multiple administrations associated with recurrence and discontinuation of treatment due to the emergence of drug resistance pose problem. One of the factors of these problems is considered to 30 be the insufficient cell killing ability of |3-ray released by 131I. On the other hand, astatine-211 (211At) is an RI that releases a-ray with higher cell killing ability than |3-ray, and the development of RI internal radiotherapy using 211At-labeled 35 compound or astatide ion (211At_) is expected. 2024266968   28 Nov 2024 1005645380

[0004] Patent document 1 discloses a composition containing a halogen compound and recites astatine as one of the halogen compounds. However, it does not specifically disclose an 5 astatine solution or a production method thereof. Non-patent document 1 discloses experimental measurement results of distribution and radiation dose in the body of rats after 2nAt administration. It describes that 2nAt was produced by 209Bi (a, 2n) 211At reaction, but does not describe addition of a io reducing agent in the production step. Non-patent document 2 discloses experimental measurement results of in vitro cytotoxicity of astatide against glioma cells expressing the sodium iodide symporter. It is described that 211At was produced by 209Bi (a, 2n) 211At reaction, and after 15 isolation, Na2SOs was added. However, in this document, effects on radiochemical purity by the addition of reducing agents have not been investigated. Non-patent document 3 describes that expression of NIS is observed in stomach cancer, breast cancer, lung cancer, 20 prostate cancer, colon cancer, ovarian cancer and the like. [Document List] [Patent document]

[0005] patent document 1: National Publication of International Patent 25 Application No. 2017-507929 [non-patent documents]

[0006] non-patent document 1: Spetz J, et al., Cancer Biotherapy and Radiopharmaceuticals, Volume 28, Number 9, 2013, 657-664 30 non-patent document 2: Carlin S, et al., The Journal of Nuclear Medicine, Vol. 44, No. 11, November 2003, 1827-1838 non-patent document 3: Atsushi Shiozaki, et al., Functional analysis and clinical significance of sodium iodide symporter expression in gastric cancer, Gastric cancer, 35 https: / / doi.org / 10.1007 / sl0120-018-0874-2, published online: 06 2024266968   28 Nov 2024 1005645380 September 2018 [Summary of Invention] [Technical Problem]

[0007] 5       The present invention aims to provide an astatine solution that can be used in RI internal radiotherapy and the like for the treatment of thyroid gland disease and the like, and a production method thereof. [Solution to Problem] io

[0008] 2nAt can generally be produced by a nuclear reaction. For example, a production method in which helium is accelerated by an accelerator such as a cyclotron, irradiated to a target substance bismuth-209 (209Bi) , and 2nAt is produced in the 15 target substance by the nuclear reaction of 209Bi (a, 2n) 211At, the target substance is evaporated by heating to separate 211At, and 2nAt is dissolved in a solvent such as water to give a 2nAt solution is known. However, the present inventors have conducted intensive 20 studies and newly found that the 2nAt solution produced by the conventional method as described above has a low radiochemical purity (see Experimental Example 1 described later), and a 2nAt solution having such low radiochemical purity shows weak accumulation of radioactivity in the thyroid gland in an in 25 vivo experiment using rats (see Experimental Example 7 described later). The present inventors have conducted further studies based on the findings, and found that a 2nAt solution with a high radiochemical purity can be produced by adding a reducing 30 agent to a 2nAt solution, and further, that the 2nAt solution with a high radiochemical purity, which is produced by the method, shows enhanced accumulation of radioactivity in the thyroid gland after administration, as compared with one produced by the conventional method, and completed the present 35 invention. 2024266968   28 Nov 2024 1005645380

[0009] That is, the present invention provides the following. [1] A method for producing a solution comprising 2nAt_ (astatide ion) at a high radiochemical purity by using 2nAt 5 obtained by a nuclear reaction as a starting material, the method comprising a step of adding a reducing agent to a solution comprising an impurity derived from 2nAt. [2] The method of the above-mentioned [1], wherein the reducing agent is a physiologically acceptable reducing agent. 10 [2-1] The method of the above-mentioned [2], wherein the reducing agent is an organic reducing agent or an inorganic reducing agent. [2-2] The method of the above-mentioned [2-1], wherein the inorganic reducing agent is iron (II) sulfate. 15 [3] The method of the above-mentioned [1], wherein the reducing agent is a physiologically acceptable organic reducing agent. [4] The method of the above-mentioned [1], wherein the reducing agent is selected from the group consisting of ascorbic acid, a salt with ascorbic acid, cysteine, glutathione and iron (II) 20 sulfate. [5] The method of the above-mentioned [1], wherein the reducing agent is ascorbic acid or sodium ascorbate. [6] The method of any of the above-mentioned [1] to [5], further comprising a step of adjusting to neutral or weakly 25 alkaline by adding a pH adjuster or buffer. [7] A solution comprising 211At" (astatide ion) at a radiochemical purity of not less than 30%. [8] A solution comprising 2nAt_ (astatide ion) at a high radiochemical purity, which solution is produced by the method 30 of any of the above-mentioned [1] to [6]. [Advantageous Effects of Invention]

[0010] According to the production method of the present invention, a solution containing 2nAt_ (astatide ion) at a high 35 radiochemical purity can be produced. 2024266968   28 Nov 2024 1005645380 A solution containing 211At" (astatide ion) at a high radiochemical purity produced by the production method of the present invention is useful in RI internal radiotherapy and the like for the treatment of thyroid gland diseases. 5        A solution containing 2nAt_ (astatide ion) at a high radiochemical purity which is produced by the production method of the present invention is useful in RI internal radiotherapy and the like for the treatment of stomach cancer, breast cancer, lung cancer, prostate cancer, colon cancer, ovarian cancer and io the like that show expression of NIS. [Brief Description

[0011] of Drawings] Fig. Fig. 1 2 shows shows the results the results of of Experimental Experimental Example 1. Example 2. 15 Fig. 3 shows the results of Experimental Example 3. Fig. 4 shows the results of Experimental Example 4. Fig. 5 shows the results of Experimental Example 5. Fig. 6 shows the results of Experimental Example 6. Fig. 7 shows the results of Experimental Example 8. 20 Fig. 8 shows the results of Experimental Example 9. Fig. 9 shows the results of Experimental Example 10. Fig. 10 shows ; the results of Experimental . Example 11 Fig. 11 shows ; the results of Experimental . Example 12 [Description of Embodiments] 25

[0012] The present invention is explained in detail in the following. The present invention relates to a production method of a solution containing 211At_ (astatide ion) at a high 30 radiochemical purity. The method of the present invention is characterized by a step of adding a reducing agent to a solution (e.g., aqueous solution, alcohol solution (e.g., ethanol solution), alcohol aqueous solution (e.g., ethanol aqueous solution)) containing 35 impurity derived from 2nAt (hereinafter to be referred to as 2024266968   28 Nov 2024 1005645380 211At stock solution) . In the present specification, the impurity derived from 2nAt refers to a compound derived from 211A which is other than the target compound 211At_ (astatide ion) . Examples of the 5 impurity derived from 2nAt include oxides of astatine (At+, AtO", AtO2-, At (OH), At (OH) 2"). In the present invention, the content of the impurity derived from 2nAt and contained in the 211At stock solution is not particularly limited. A solution in which the io radioactivity of the impurity derived from 211At with respect to the radioactivity of the whole solution is more than 0% and up to 100% can be used as the 2nAt stock solution of the present invention. A 2nAt solution (e.g., 2nAt aqueous solution) obtained by 15 a conventional method (e.g., the method described in the below-mentioned Reference Example 1) contains much impurity derived from 211At and can be used as the 2nAt stock solution in the method of the present invention. The impurity derived from 2nAt can be confirmed, for 20 example, by thin layer chromatography described in the below-mentioned Experimental Example 1, or a method analogous thereto.

[0013] The reducing agent to be used in the method of the present invention is preferably a physiologically acceptable 25 reducing agent. In the present invention, physiologically acceptable means being physiologically low toxic or nontoxic. Examples of the reducing agent to be used in the method of the present invention include organic reducing agents (e.g., ascorbic acid, salt with ascorbic acid (e.g., salts of ascorbic 30 acid with alkali metal or alkaline earth metal (e.g., sodium ascorbate, calcium ascorbate)), cysteine, glutathione, gentisic acid, glucose etc.), inorganic reducing agent (e.g., iron (II) sulfate), and ascorbic acid, salt with ascorbic acid (e.g., salts of ascorbic acid with alkali metal or alkaline earth 35 metals (e.g., sodium ascorbate, calcium ascorbate)), cysteine, 2024266968   28 Nov 2024 and glutathione are preferable, ascorbic acid, salts with ascorbic acid (e.g., salts of ascorbic acid with alkali metal or alkaline earth metals (e.g., sodium ascorbate, calcium ascorbate)) are more preferable, and ascorbic acid, sodium 5 ascorbate are particularly preferable.

[0014] In the method of the present invention, the amount of the reducing agent to be used is an amount that achieves a final concentration of generally 0.1 - 30 wt% (w / v%), preferably 0.2 io - 10 wt%, more preferably 0.5-5 wt%, with respect to a solution containing impurity derived from 2nAt (that is, 2nAt stock solution). In the method of the present invention, the amount of the reducing agent to be used is generally 0.1 - 100 pmol, 15 preferably 0.2 - 50 pmol, more preferably 1-12 pmol, with respect to 1 MBq of 211At radioactivity of a solution containing impurity derived from 2nAt (that is, 2nAt stock solution). In the method of the present invention, a solution e.g., aqueous solution, alcohol solution (e.g., ethanol solution), 20 alcohol aqueous solution (e.g., ethanol aqueous solution)) containing the target 211At" (astatide ion) at a high radiochemical purity (e.g., not less than 30%) can be obtained by adding and mixing a reducing agent. The reducing agent may be dissolved in a solvent (e.g., water) in advance and added as 25 a reducing agent solution (e.g., aqueous solution). The mixing can be performed by a known method. When the solution is mixed while aerating with an inert gas such as nitrogen, argon, etc. to block the contact with air and oxygen, the effect of the reducing agent is highly exhibited. 30      The reducing agent is added and mixed at room temperature, specifically 0 - 40°C, preferably 15 - 30°C. The mixing time is, for example, 5 min - 2 hr, particularly, 10 - 30 min. Production of the target 2nAt_ (astatide ion) can be confirmed by thin layer chromatography. 2024266968   28 Nov 2024 The method of the present invention preferably further contains a step of adjusting to neutral or weakly alkaline by adding a pH adjuster or buffer. By adjusting to neutral or weakly alkaline, volatilization, transpiration of astatine, and 5 generation of oxide can be prevented. The step can be performed by a known method. The pH adjuster may be dissolved in a solvent (e.g., water) in advance and added as a pH adjuster solution (e.g., an aqueous solution). In the present specification, neutral or weakly alkaline io generally means pH 5 - pH 10, preferably pH 6 - pH 9, more preferably pH 6.5 - pH 8.5. When the reducing agent used makes the solution neutral or weakly alkaline, it is not necessary to add a pH adjuster or a buffer. 15

[0016] In the method of the present invention, a pH adjusting agent or buffer generally used in pharmaceutical preparations can be used as the pH adjuster or buffer. For example, pH adjusts such as sodium hydrogen carbonate, hydrochloric acid, 20 sodium hydroxide and the like, buffers such as phosphate buffer, acetate buffer, citrate buffer, borate buffer and the like can be mentioned, and sodium hydrogen carbonate is preferable. In the present invention, an amount capable of adjusting to neutral or weakly alkaline can be appropriately selected as 25 the amount of the pH adjuster or buffer.

[0017] In the method of the present invention, the order of the step of adding a reducing agent, and the step of adjusting to neutral or weakly alkaline by adding a pH adjuster or buffer is 30 not particularly limited. A reducing agent and a pH regulator or buffer may be added simultaneously. From the aspect of preventing the risk of volatilization of astatine, it is preferable to perform the step of adding a reducing agent after the step of adjusting to neutral or weakly alkaline by adding a 35 pH adjuster or buffer. 2024266968   28 Nov 2024 1005645380

[0018] The method of the present invention may include a step of adding additives (e.g., solvents such as water for injection, saline, alcohol and the like, isotonicity agent, surfactant, 5 sweetener, flavor, colorant, stabilizer, excipient) generally used for pharmaceutical preparations, in addition to the abovementioned reducing agent, pH adjuster, and buffer. These additives are used in amounts conventionally used for pharmaceutical preparations. io

[0019] Using the method of the present invention, a solution (e.g., aqueous solution, alcohol solution (e.g., ethanol solution), alcohol aqueous solution (e.g., ethanol aqueous solution) containing 2nAt_ (astatide ion) at a high 15 radiochemical purity (e.g., not less than 30%) can be produced.

[0020] In the present specification, the radiochemical purity is the ratio (%) of the radioactivity of 2nAt_ (astatide ion) to the total radioactivity of 2nAt in the solution. 20        The radiochemical purity of the solution containing 211At" (astatide ion) obtained by the method of the present invention is generally not less than 30%, preferably not less than 50%, more preferably not less than 80%, further preferably not less than 90%, further more preferably not less than 95%, 25 particularly preferably 100%. The radiochemical purity in the present invention can be confirmed by, for example, thin layer chromatography described in the below-mentioned Experimental Example 1, Experimental Example 8, or a method analogous thereto. 30

[0021] The embodiment of the method of the present invention is, for example, the following. (1) A reducing agent (e.g., ascorbic acid), a pH adjuster (e.g., sodium hydrogen carbonate), and, where necessary, water for 35 injection are added to a 2nAt solution (2nAt stock solution), 2024266968   28 Nov 2024 1005645380 and they are mixed to give solution containing 2nAt" (astatide ion) at a high radiochemical purity. (2) A reducing agent (e.g., sodium ascorbate), and, where necessary, water for injection are added to a 2nAt solution 5  (211At stock solution) , and they are mixed to give solution containing 2nAt_ (astatide ion) at a high radiochemical purity.

[0022] The present invention also relates to a solution (e.g., aqueous solution, alcohol solution (e.g., ethanol solution), io for example, alcohol aqueous solution (e.g., ethanol aqueous solution)) containing 2nAt_ (astatide ion) at a high radiochemical purity (e.g., not less than 30%), which is obtained by the above-mentioned production method of the present invention (hereinafter sometimes to be indicated as the 15 high purity 211At"-containing solution of the present invention). The definition and preferred range of the radiochemical purity are the same as above. The high purity 211At"-containing solution of the present invention may contain reducing agent, pH adjuster, buffer and 20 the like added in the production step, and can be administered to the subject without separation. In addition, the high purity 211At"-containing solution of the present invention may further contain additives generally used for pharmaceutical preparations . 25

[0023] The high purity 211At"-containing solution of the present invention can be used for the treatment and diagnosis (particularly RI internal radiotherapy) of, for example, thyroid gland diseases (e.g., thyroid cancer, hyperthyroidism 30 (Graves' disease) etc.), and thyroid cancer metastatic lesion in human and mammals (e.g., mouse, rat, rabbit, cat, dog, bovine, horse, monkey) . The high purity 211At_-containing solution of the present invention can also be used for the treatment and diagnosis (particularly RI internal radiotherapy) 35 of cancers (e.g., stomach cancer, breast cancer, lung cancer, 2024266968   28 Nov 2024 1005645380 prostate cancer, colon cancer, ovarian cancer etc.) which are other than thyroid cancer and in which NIS is markedly expressed. In the RI internal radiotherapy using the high purity 5 211At_-containing solution of the present invention, the affected part can be treated by radiation of a-ray released by 2nAt accumulated in the affected part after administration to patients . The high purity 211At_-containing solution of the present io invention can be safely administered orally or parenterally (intravenous injection, drip etc.).

[0024] The dose of the high purity 211At_-containing solution of the present invention varies depending on the subject of 15 administration, administration route, disease and the like. For example, when it is used for treatment of thyroid cancer in an adult, the dose of 211At" (astatide ion) is 1 MBq - 20 GBq. [Example]

[0025] 20       The present invention is explained in more detail in the following by referring to Examples and Experimental Examples, which are not to be construed as limitative. In Examples, % showing the concentration of components in the solution is wt% (w / v%). 25

[0026] Reference Example 1: Preparation of 2nAt aqueous solution Helium (28 MeV, 1-2 pA) accelerated by a cyclotron was irradiated to the target substance bismuth-209 (209Bi) , and astatine-211 (2nAt) was produced in the target substance by a 30 nuclear reaction of 209Bi (a, 2n) 211At. After irradiation, the target substance was dissolved by heating in an electric heating appliance to 800°C, and 211At evaporating from the target substance was dissolved in a small amount of water (0.1 - 2 mL) to prepare a 2nAt aqueous solution. The radioactivity 35 of this 2nAt aqueous solution on preparation was 5-25 MBq. 2024266968   28 Nov 2024 1005645380

[0027] Reference Example 2: Preparation of sodium hydrogen carbonate-added 2nAt aqueous solution To the 2nAt aqueous solution (0.1 mL, 14 MBq) prepared in 5 Reference Example 1 were successively added water for injection (0.5 mL) and 7% sodium hydrogen carbonate aqueous solution (0.8 mL), and the mixture was stirred at room temperature for 30 min to prepare a sodium hydrogen carbonate-added 2nAt aqueous solution (pH 7.5-9.0). io

[0028] Example 1: Preparation of 0.9% ascorbic acid-added 2nAt aqueous solution To the 2nAt aqueous solution (0.1 mL, 25 MBq) prepared in Reference Example 1 were successively added water for injection 15  (0.5 mL), 7% sodium hydrogen carbonate aqueous solution (0.8 mL), and 2% ascorbic acid aqueous solution (1.2 mL) , and the mixture was stirred at room temperature for 30 min to prepare a 0.9% ascorbic acid-added 211At aqueous solution (pH 7.5-9.0).

[0029] 20 Experimental Example 1: Analysis of radiochemical purity The radiochemical purity of the 0.9% ascorbic acid-added 2nAt aqueous solution prepared in Example 1, 2nAt aqueous solution prepared in Reference Example 1, sodium hydrogen carbonate-added 2nAt aqueous solution prepared in Reference 25 Example 2, and Na123! aqueous solution (manufactured by FUJIFILM RI Pharma Co., Ltd.) as a comparison control was analyzed by thin layer chromatography (TLC) according to the following method. Each sample (2 pL) was applied to the original line 30 (Origin) of a thin-layer plate (G60, manufactured by Merck & Co., Inc.) and developed with a mixed solution of acetonitrile : water : trifluoroacetic acid (67:33:0.5) as a developing solvent. After exposure to an imaging plate (BAS IP, GE Healthcare), the distribution ratio (%) of the radioactivity 35 on the thin-layer plate was measured by a bioimaging device 2024266968   28 Nov 2024 1005645380 (TYPHOON7000, manufactured by GE Healthcare). The results are shown in Fig. 1. In the 0.9% ascorbic acid-added 211At aqueous solution of Example 1 (Fig. 1, (a)), a radioactive spot corresponding to 5 211At-astatide ion (211At_) was detected at the position of Rf=0.82 with 100% radiochemical purity. The fact that this radioactive spot is 211At_ was considered appropriate by inferring from the radioactive spot of 123I-iodide ion (123I_) in the Na123I aqueous solution (Rf value of (d) = 0.86 in Fig. 1) obtained under the io same conditions. In contrast, in the 2nAt aqueous solution of Reference Example 1 (Fig. 1, (b)) and sodium hydrogen carbonate-added 211At aqueous solution of Reference Example 2 (Fig. 1, (c) ) , the radiochemical purity of 2nAt_ was not more than 2 0% in both, and most of the radioactivity was detected as a plurality of 15 radioactive impurities near the point of origin and near the solvent tip (Front) . From the above results, it was shown that highly pure 211At" is produced when ascorbic acid is added to the 2nAt aqueous solution.

[0030] 20 Examples 2-5: Preparation of 0.25 - 2% ascorbic acid-added 2nAt aqueous solution To the 2nAt aqueous solution (0.1 mL) prepared in Reference Example 1 were added an appropriate amount of water for injection, 7% sodium hydrogen carbonate aqueous solution, 25 and 4% ascorbic acid aqueous solution, and the mixture was stirred at room temperature for 30 min to respectively prepare ascorbic acid-added 211At aqueous solutions (pH 7.5-9.0) having a final concentration of 0.25% (Example 2), 0.5% (Example 3), 1% (Example 4) or 2% (Example 5). The final radioactivity 30 concentrations of 2nAt were all standardized to 10 MBq / mL.

[0031] Experimental Example 2: Study of effect of ascorbic acid concentration (0.25 - 2%) The radiochemical purity of the ascorbic acid-added 2nAt 35 aqueous solutions of Examples 2-5 was analyzed by TLC according 2024266968   28 Nov 2024 1005645380 to a method similar to that in Experimental Example 1. The results are shown in Fig. 2. When ascorbic acid was not added (2nAt aqueous solution of Experimental Example 1), the radiochemical purity of 2nAt_ 5 was 20%, whereas the radiochemical purity of 2nAt_ increased to 53.8% when the ascorbic acid concentration was 0.25%, and further, the radiochemical purity of 2nAt_ was not less than 95% when the ascorbic acid concentration was not less than 0.5%.

[0032] io Example 6: Preparation of 1% sodium ascorbate-added 2nAt aqueous solution To the 2nAt aqueous solution (0.1 mL, 2 MBq) prepared in Reference Example 1 was added 2% sodium ascorbate aqueous solution (0.1 mL) , and the mixture was stirred at room 15 temperature for 30 min to prepare a 1% ascorbate-added 2nAt aqueous solution (pH 5-7).

[0033] Experimental Example 3: Analysis of radiochemical purity The radiochemical purity of the 1% sodium ascorbate-added 20 2nAt aqueous solution of Example 6 was analyzed by TLC according to a method similar to that in Experimental Example 1. The results are shown in Fig. 3. The radiochemical purity of 211At" (Rf=0.93) was 92.4%. It was found that sodium ascorbate is effective for producing 25 211At" having high purity similar to that of ascorbic acid.

[0034] Example 7: Preparation of 1% cysteine-added 2nAt aqueous solution To the 2nAt aqueous solution (0.1 mL, 4 MBq) prepared in 30 Reference Example 1 were added water for injection (0.05 mL), 7% sodium hydrogen carbonate aqueous solution (0.1 mL), and 2% cysteine aqueous solution (0.25 mL), and the mixture was stirred at room temperature for 30 min to prepare a 1% cysteine-added 2nAt aqueous solution (pH 7-9) . 35

[0035] 2024266968   28 Nov 2024 1005645380 Experimental Example 4: Analysis of radiochemical purity The radiochemical purity of the 1% cysteine-added 2nAt aqueous solution of Example 7 was analyzed by TLC according to a method similar to that in Experimental Example 1. The 5 results are shown in Fig. 4. The radiochemical purity of 211At_ (Rf=0.89) was 100%. It was found that cysteine is effective for producing 2nAt_ having high purity similar to that of ascorbic acid.

[0036] io Example 8: Preparation of 1% glutathione addition 211At aqueous solution To the 2nAt aqueous solution (0.1 mL, 4 MBq) prepared in Reference Example 1 were added water for injection (0.05 mL), 7% sodium hydrogen carbonate aqueous solution (0.1 mL), and 2% 15 glutathione aqueous solution (0.25 mL), and the mixture was stirred at room temperature for 30 min to prepare a 1% glutathione-added 2nAt aqueous solution (pH 7-9) .

[0037] Experimental Example 5: Analysis of radiochemical purity 20       The radiochemical purity of the 1% glutathione-added 2nAt aqueous solution of Example 8 was analyzed by TLC according to a method similar to that in Experimental Example 1. The results are shown in Fig. 5. The radiochemical purity of 211At" (Rf=0.93) was 86.2%. 25 It was found that glutathione is effective for producing 211At" having high purity similar to that of ascorbic acid.

[0038] Experimental Example 6: Planar imaging of normal rat Normal rats (Wistar rat, male, 3 months old) fed with a 30 low iodine diet for 2 weeks were used for the experiment. The rat was anesthetized with isoflurane, and the 0.9% ascorbic acid-added 2nAt aqueous solution (about 3 MBq / rat) prepared in Example 1 was administered into the tail vein (n=3). As a control group, similarly-treated rats were administered with 35 the 211At aqueous solution (about 5 MBq / rat) prepared in 2024266968   28 Nov 2024 Reference Example 1 (n=3). The rats were anesthetized 30 min, 3 hr, 6 hr, and 24 hr after administration, and planar images were taken with a gamma camera (E.CAM, manufactured by Siemens). Imaging time was 10 min in the case of 30 min to 6 hr after 5 administration, and 20 min in the case of 24 hr after administration. Fig. 6 shows the results of planar imaging. In Fig. 6, the ascorbic acid group (0.9% ascorbic acid-added 2nAt aqueous solution administration group) in the upper panel (a) showed strong accumulation of radioactivity in the io thyroid gland from the early stage (30 min point) after administration of the test solution as compared to the control group (211At aqueous solution administration group) in the lower panel (b), and the accumulation of radioactivity was enhanced with time until the 24 hr point. Other than the thyroid gland, 15 accumulation in the stomach and excretion into the urinary system (bladder) were shown. In the control group of (b), the accumulation of radioactivity in the thyroid gland was slightly weak, and retention of radioactivity in the stomach was shown. From the above results, it was suggested that administration of 20 a 211At" aqueous solution prepared to have a high purity by the addition of ascorbic acid to rats enhances the accumulation of radioactivity in the thyroid gland and, on the other hand, reduces the non-specific accumulation in the stomach.

[0039] 25 Experimental Example 7: Normal rat body distribution test After completion of the imaging at 24 hr point after administration described in Experimental Example 6, the rats in the ascorbic acid group (0.9% ascorbic acid-added 211At aqueous solution administration group) and the control group (211At 30 aqueous solution administration group) were dissected, and the thyroid gland, stomach, and other major organs were removed. The wet weight of each excised organ was measured with a microbalance, and the radioactivity was measured with a gamma counter (2480WIZARD2, manufactured by Perkin Elmer Co., Ltd.). 35 Table 1 shows the results of the body distribution test in 2024266968   28 Nov 2024 normal rats (radioactivity count per organ weight). In both groups, the radioactivity count of the thyroid gland was 10 times or higher than that of other organs. In the ascorbic acid group, the radioactivity count of the thyroid gland was 5 about 4 times higher than that of the control group. On the other hand, the radioactive count of the stomach, spleen and liver was higher in the control group than the ascorbic acid group. Table 2 shows the results of calculation of the io radioactivity count ratio per unit weight of the thyroid gland and other organs from the values in Table 1. In both the ascorbic acid group and the control group, the radioactivity count ratio of the thyroid gland and other major organs was as high as 19.3-fold to 2755.6-fold. The thyroid gland / stomach 15 ratio was 4.9 times higher in the ascorbic acid group than in the control group. From the above results, it was suggested that administration of a 211At" aqueous solution prepared to have a high purity by the addition of ascorbic acid to rats enhances the accumulation of radioactivity in the thyroid gland 20 and reduces the non-specific accumulation in the stomach and other organs. That is, it was suggested that an ascorbic acid-added 2nAt aqueous solution enhances the therapeutic effect on thyroid cancer, reduces exposure of other organs such as the stomach to radiation, and enables safer treatment. 25

[0040] [Table 1] Table 1. Normal rat body distribution test results (at 24 hr point from administration): radioactivity count (unit: counts / g) per weight of each organ test solution group organ ascorbic acid group (n=3) control group (n=3) thyroid gland 1309706 3283443 stomach 139652 169690 spleen 43518 57081 liver 4753 11729 2024266968   28 Nov 2024

[0041] [Table 2] Table 2. Rat body distribution test results (at 24 hr point 5 from administration): radioactivity count ratio per unit weight of thyroid gland and other organs (stomach, spleen, and liver) test solution group organ ascorbic acid group (n=3) control group (n=3) ascorbic acid group / control group thyroid gland / stomach 93.8 19.3 4.9 thyroid gland / spleen 300.9 57.5 5.2 thyroid gland / liver 2755.6 280.0 9.9

[0042] Examples 9-12: Preparation of reducing agent-added 2nAt io aqueous solution The 2nAt aqueous solution prepared in Reference Example 1 was diluted with water and to the prepared crudely purified 2nAt aqueous solution 0.1 mL (about 1 MBq) was added 2% reducing agent (ascorbic acid, cysteine, glutathione or 15 iron(II) sulfate) aqueous solution (0.1 mL) , and the mixture was stirred at room temperature for 1 hr to prepare ascorbic acid-added 2nAt aqueous solution (Example 9), cysteine-added 2nAt aqueous solution (Example 10), glutathione-added 211At aqueous solution (Example 11), and iron(II) sulfate-added 211At 20 aqueous solution (Example 12), each having a final concentration of the reducing agent of 1%.

[0043] Experimental Example 8: Analysis of radiochemical purity: effect of addition of various reducing agents 25       A crudely-purified 2nAt aqueous solution, and the reducing agent-added 2nAt aqueous solutions prepared in Examples 9-12 were analyzed by TLC (thin-layer plate: silica gel G6OF254 (manufactured by Merck), solvent: acetonitrile-water-trifluoroacetic acid (67 / 33 / 0.1)) (Fig. 7). The 30 radiochemical purity of 211At" (astatide ion) was the highest 2024266968   28 Nov 2024 1005645380 (95.7%) when ascorbic acid was used, and then in the order of cysteine, glutathione, and iron(II) sulfate.

[0044] Experimental Example 9: Comparison of intracellular uptake of 5 2nAt by KI cell (human thyroid cancer cell) and Kl-NIS cell (NIS expression KI cell) KI cells (human thyroid cancer cell) and Kl-NIS cells (NIS expression KI cells), each IxlO5 cells, were seeded on a 96-well plate, and 10 pL (about 10 kBq) of 211At aqueous io solution was added. The cells were cultured at 37°C for 5, 30, and 60 min, lysed with 0. IN NaOH, and the intracellular radioactivity was measured with a gamma counter (2480 Wizard2, Perkin Elmer). The amount of protein was measured with a BCA protein quantification kit (FUJIFILM Corporation) and a plate 15 reader (Thermo Fisher) . As shown in Fig. 8, 211At was taken up by Kl-NIS cells but was not taken up by KI cells. It was shown that 211At is specifically taken up intracellularly via NIS.

[0045] Experimental Example 10: changes in time-course uptake amount 20 of 211At-AA( + ) and 211At-AA(-) by thyroid cancer cell (Kl-NIS) The 2nAt aqueous solution prepared in Reference Example 1 was diluted with water to prepare astatine aqueous solution (211At-AA (-) , 1 MBq / mL) without ascorbic acid. In addition, the 2nAt aqueous solution prepared in Reference Example 1 was 25 diluted with water, and ascorbic acid was further added to a final concentration of 1% to prepare ascorbic acid-added astatine aqueous solution (211At-AA ( + ) , 1 MBq / mL). Kl-NIS cells (NIS expressing KI cells) were seeded on a 96-well plate by IxlO5 cells, and 10 pL (about 10 kBq) of 30 ascorbic acid-added astatine aqueous solution (2nAt-AA(+)) or astatine aqueous solution (211At-AA (-) ) without ascorbic acid was added. The cells were cultured at 37°C for 5, 30, or 60 min, lysed with 0. IN NaOH, and the intracellular radioactivity was measured with a gamma counter (2480 Wizard2, Perkin Elmer). 35 The amount of protein was measured with a BCA protein 2024266968   28 Nov 2024 quantification kit (FUJIFILM Corporation) and a plate reader (Thermo Fisher). As shown in Fig. 9, the uptake amount of 211At-AA( + ) by Kl-NIS cells increased over time and reached about 2 times that of AA(-) (60 min later). It was shown that 5 addition of ascorbic acid to the 2nAt aqueous solution increases 2nAt uptake amount of the cell.

[0046] Experimental Example 11: SPECT imaging of mice transplanted with thyroid cancer cell (Kl-NIS) by 211At-AA( + ) io       The 2nAt aqueous solution prepared in Reference Example 1 was diluted with water, and ascorbic acid was further added to a final concentration of 1% to prepare 211At-AA(+) aqueous solution. A thyroid cancer-transplanted mouse was prepared by 15 injecting IxlO7 Kl-NIS cells subcutaneously into SCID mouse. A 211At-AA(+) aqueous solution (1 MBq) was intravenously injected to this mouse, and images were taken with a SPECT camera (E-cam, Siemens) after 3 hr and 24 hr. The tumor site was clearly depicted at any time point (Fig. 10) . The radioactivity 20 accumulation rate at the tumor site was 22.5110.4% (3 hr later) and 12.916.8% (24 hr later). In addition, weak physiological accumulation of radioactivity was also observed in the stomach. This result suggests that 2nAt-AA(+) is useful for diagnosis and treatment of NIS-expressing tumors including thyroid cancer. 25

[0047] Experimental Example 12: Treatment of thyroid cancer cell (Kl-NIS) -transplanted mouse by 211At-AA( + ) aqueous solution: Changes in tumor size and body weight over time after single intravenous injection of 0.1 MBq, 0.4 MBq and 1 MBq 30       The 2nAt aqueous solution prepared in Reference Example 1 was diluted with water, and ascorbic acid was further added to a final concentration of 1% to prepare 211At-AA(+) aqueous solution. A thyroid cancer-transplanted mouse was prepared by 35 injecting IxlO7 Kl-NIS cells subcutaneously into SCID mouse. A 2024266968   28 Nov 2024 1005645380 211At-AA( + ) aqueous solution (1 MBq, 0.4 MBq, and 1 MBq) was intravenously injected once to this mouse, and changes in the tumor size and body weight were measured for 60 days (n=6). As shown in Fig. Ila (Fig. 11, left figure) , the tumor size 5 increased over time in the non-treated group (control), but tumor reduction or growth suppressive effect was observed for about 40 days in the 2nAt 1 MBq intravenous injection group. The tumor growth suppressive effect was dependent on the amount of radioactivity of 211At. As shown in Fig. 11b (Fig. 11, right io figure.), transient weight loss was initially observed, but it was recovered later in the 1 MBq intravenous injection group. The body weight of the 0.1 MBq group and the 0.4 MBq group did not differ much as compared to that of the control group. [Industrial Applicability] 15

[0048] The present invention can provide a solution containing 211At" (astatide ion) at a high radiochemical purity, that is useful for RI internal radiotherapy and the like for the treatment of thyroid gland disease and the like. 20

[0049] This application is based on patent application No. 2017255109 filed in Japan, the contents of which are encompassed in full herein.

[0050] 25        In the claims which follow and in the preceding description of the invention, except where the context requires otherwise due to express language or necessary implication, the word "comprise" or "contain" or variations such as "comprises", "comprising", "contains" or "containing" is used in an 30 inclusive sense, i.e. to specify the presence of the stated features but not to preclude the presence or addition of further features in various embodiments of the invention.

[0051] Reference to any prior art in the specification is not 35 an acknowledgement or suggestion that this prior art forms part 2024266968   28 Nov 2024 1005645380 of the common general knowledge in any jurisdiction or that this prior art could reasonably be expected to be combined with any other piece of prior art by a skilled person in the art.

Claims

1. A method for treating or diagnosing a thyroid cancer or a thyroid cancer metastatic lesion, comprising administering an effective5 amount of a composition comprising 211At" (astatide ion) as an active ingredient.

2. The method according to claim 1, wherein the composition further comprises a reducing agent selected from the group consisting of10 ascorbic acid, a salt of ascorbic acid, cysteine, glutathione and iron(II) sulfate.

3. The method according to claim 1 or 2, wherein the reducing agent is ascorbic acid or a salt of ascorbic acid.

154. The method according to claim 3, wherein the reducing agent is ascorbic acid or sodium ascorbate.

5. The method according to claim 3, wherein the reducing agent is20 sodium ascorbate.

6. The method according to any one of claims 3 to 5, wherein aconcentration of the reducing agent is 0.5 wt% to 5 wt% .25 7. The method according to any one (astatide ion) has a radiochemical8. The method according to any one (astatide ion) has a radiochemical 309. The method according to any one (astatide ion) has a radiochemicalof claims 1 to 6, wherein the 2nAt purity of not less than 30%.of claims 1 to 6, wherein the 2nAt purity of not less than 80%.of claims 1 to 6, wherein the 2nAt purity of not less than 90%.

10. The method according to any one of claims 1 to 6, wherein the352nAt_ (astatide ion) has a radiochemical purity of not less than 95%.

11. The method according to any one of claims 1 to 6, wherein the2nAf(astatide ion)has a radiochemical purity of not less than 100%.2024266968   28 Nov 2024100564538012. The method according to any one of claims 1 to 11, wherein the thyroid cancer expresses NIS and the thyroid cancer metastatic lesion expresses NIS.5 13. The method according to any one of claims 1 to 12, wherein the composition is administered by intravenous injection.

14. The method according to any one of claims 1 to 13, wherein a dose of the composition is 1 MBq to 20 GBq.io15. The method according to any one of claims 1 to 14, the composition further comprises a pH adjuster, wherein the pH adjuster is sodium hydrogen carbonate.15 16. The method according to any one of claims 1 to 15, wherein the composition has pH 6 to pH 9.

17. A composition comprising (1) 211At" (astatide ion) as an active ingredient, and (2) a reducing agent, wherein the reducing agent is 20 ascorbic acid or a salt of ascorbic acid, andthe 211At_ (astatide ion) has a radiochemical purity of not less than 30%.

18. The composition according to claim 17, whereinthe reducing agent25 is ascorbic acid or sodium ascorbate.

19. The composition according to claim 18, wherein the reducing agentis sodium ascorbate.30 20. The composition according to any one of claims 17 to 19, whereina concentration of the reducing agent is 0.5 wt% to 5 wt% .

21. The composition according to any one of claims17 to 20, whereinthe 211At_ (astatide ion) has a radiochemical purity of not less than35 80%.

22. The composition according to any one of claims 17 to 20, whereinthe 211At_ (astatide ion) has a radiochemical purity of not less than2024266968   28 Nov 202423.The composition according to any one of claims 17 to 20,whereinthe 211At_ (astatide ion) has a radiochemical purity of not less than95%.

24. The composition according to any one of claims 17 to 20, whereinthe 211At_ (astatide ion) has a radiochemical purity of not less than100%.io 25. The composition according to any one of claims 17 to 24, whereinthe composition is administered by intravenous injection.

26. The composition according to any one of claims 17 to 25, whereina dose of the composition is 1 MBq to 20 GBq.1527. The composition according to any one of claims 17 to 26, further comprising a pH adjuster, wherein the pH adjuster is sodium hydrogen carbonate.20 28. The composition according to any one of claims 17 to 27, whereinthe composition has pH 6 to pH 9.2 9. Use of 211At_ (astatide ion) in the production of a composition forthe treatment or diagnosis of a thyroid cancer or a thyroid cancer25 metastatic lesion.

30. Use of 211At" (astatide ion) and a reducing agent in the production of a composition for the treatment or diagnosis of a thyroid cancer or a thyroid cancer metastatic lesion, wherein the reducing agent is30 selected from the group consisting of ascorbic acid, a salt of ascorbic acid, cysteine, glutathione and iron (II) sulfate.

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