Method for preserving intermediates of radiopharmaceutical compositions, method for preparing or preserving radiopharmaceutical compositions, intermediate compositions of radiopharmaceutical compositions, and pharmaceutical formulations.

Abstract

The storage method according to the present invention comprises storing an intermediate for a radiopharmaceutical composition in an acetate buffer. The intermediate is a complex between an anti-epidermal growth factor receptor antibody (anti-EGFR antibody) and 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA). The storage period is at least 24 hours.

Description

Technical Field

[0001] The present invention relates to a method for storing an intermediate of a radiopharmaceutical composition, a method for preparing or storing a radiopharmaceutical composition, an intermediate composition of a radiopharmaceutical composition, and a pharmaceutical preparation.

Background Art

[0002] Research and development of radioimmunopharmaceuticals obtained by labeling with a radionuclide using a bifunctional chelating agent such as 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) has been progressing. For example, Non-Patent Documents 1 to 5 describe radioimmunotherapy and diagnostic methods using radioimmunopharmaceuticals.

Prior Art Documents

Non-Patent Documents

[0003]

Non-Patent Document 1

Non-Patent Document 2

Non-Patent Document 3

Non-Patent Document 4

Non-Patent Document 5

Summary of the Invention

Problems to be Solved by the Invention

[0004] However, conventional technologies as described above do not guarantee the long-term storage of antibody ligand complexes (complexes of antibodies and chelating agents), which are intermediates of radioactive antibody drugs, making on-demand preparation essential. Furthermore, on-demand preparation requires aseptic techniques, skilled operation, and the risk of radiation exposure, making it difficult to stably supply radioactive antibody drugs. In addition, on-demand preparation of radioactive antibody drugs limits the amount that can be manufactured at one time, potentially increasing the manufacturing cost of radioactive antibody drugs. For these reasons, there is a need for technological development to manufacture radioactive antibody drugs at low cost and stably.

[0005] One aspect of the present invention aims to realize a technology for producing radioactive antibody drugs without requiring preparation before use. [Means for solving the problem]

[0006] As a result of diligent research, the inventors of the present invention discovered that intermediates for radioactive antibody drugs can be stored for a long period of time by using a storage solution of a specific composition. Furthermore, they found that the physical properties of radioactive antibody drugs prepared from intermediates stored in this storage solution for a certain period of time were equivalent to those of radioactive antibody drugs obtained by preparation immediately before use, thus completing the present invention.

[0007] A preservation method according to one aspect of the present invention comprises preserving an intermediate of a radiopharmaceutical composition in an acetate buffer, wherein the intermediate is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody), and the preservation time is 24 hours or more.

[0008] Furthermore, a preservation method according to one aspect of the present invention includes preserving a radiopharmaceutical composition in an acetate buffer, wherein the radiopharmaceutical composition is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody), and is labeled with a radionuclide that coordinates with PCTA.

[0009] Furthermore, an intermediate composition according to one aspect of the present invention is an intermediate composition used in the preparation of a radiopharmaceutical composition, comprising an intermediate of a radiopharmaceutical composition and a preservation solution, wherein the intermediate is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an antibody, and the preservation solution comprises at least one neutral amino acid, at least one polysorbate and an acetate buffer, and the intermediate is stored in the preservation solution for 24 hours or more.

[0010] Furthermore, a pharmaceutical preparation according to one aspect of the present invention comprises a radiopharmaceutical composition and a preservation solution, wherein the radiopharmaceutical composition is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an antibody, and is labeled with a radionuclide that coordinates with PCTA, and the preservation solution comprises at least one neutral amino acid, at least one polysorbate and an acetate buffer, and the radiopharmaceutical composition is obtained by labeling the complex, which has been stored in the preservation solution for 24 hours or more, with a radionuclide that coordinates with PCTA. [Effects of the Invention]

[0011] According to one aspect of the present invention, a radioactive antibody drug can be manufactured without preparing it for use. [Brief explanation of the drawing]

[0012] [Figure 1]This figure shows the effect of the preservation solution on the labeling efficiency of 64Cu-PCTA-cetuximab. [Figure 2] This figure shows the effect of the preservation solution on the stability of labeled 64Cu-PCTA-cetuximab when PCTA-cetuximab is stored in the preservation solution for 3 months. [Figure 3] This figure shows the effect of the preservation solution on the stability of labeled 64Cu-PCTA-cetuximab when PCTA-cetuximab is stored in the preservation solution for 12 months. [Figure 4] This figure shows the effect of the preservation solution on the cell binding properties of 64Cu-PCTA-cetuximab. [Figure 5] This figure shows the effect of labeling-time radioactivity on the cell binding properties of 64Cu-PCTA-cetuximab. [Figure 6] This figure shows the effect of the preservation solution on the stability of 225Ac-PCTA-cetuximab after labeling. [Modes for carrying out the invention]

[0013] [Method for preserving intermediates of radioactive pharmaceutical compositions] The method for storing intermediates of a radiopharmaceutical composition according to this embodiment (hereinafter sometimes abbreviated as "the method for storing intermediates according to this embodiment") includes storing the intermediates of a radiopharmaceutical composition in a storage solution.

[0014] (Radiopharmaceutical compositions and their intermediates) In this specification, a radiopharmaceutical composition refers to a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA), labeled with a radionuclide, and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody). The radiopharmaceutical composition may be used for the diagnosis or treatment of diseases. In this specification, an intermediate of the radiopharmaceutical composition refers to the pharmaceutical composition before labeling with a radionuclide (i.e., the PCTA and anti-EGFR antibody complex that is not labeled with a radionuclide). Radionuclides will be described later.

[0015] (PCTA) PCTA is a bifunctional chelating agent. By introducing an isocyanate group or an N-hydroxysuccinimidyl group into PCTA and then reacting it with the side-chain amino group or N-terminal amino group of a lysine residue in an antibody, a chelation site is introduced into the antibody. Subsequently, by complexing a radionuclide with the chelation site, an antibody labeled with the radionuclide can be obtained.

[0016] (Anti-EGFR antibody) Examples of anti-EGFR antibodies include antibodies that specifically bind to cancer cells. Specific examples of anti-EGFR antibodies include cetuximab, panitumumab, and nesitumumab. Anti-EGFR antibodies may be mouse antibodies, chimeric antibodies, humanized antibodies, or human antibodies. Furthermore, anti-EGFR antibodies may be monoclonal or polyclonal antibodies. Commercially available anti-EGFR antibodies may be used, or anti-EGFR antibodies synthesized using known methods may be used.

[0017] The intermediate may be one synthesized using a known method. For example, as shown in the examples, the intermediate can be obtained by mixing a solution containing PCTA with a solution containing an anti-EGFR antibody and incubating it at 1 to 60°C for 1 to 24 hours.

[0018] (Preservation solution for intermediates) The preservation solution used in the intermediate preservation method according to this embodiment (hereinafter sometimes referred to as "intermediate preservation solution") is an acetate buffer. The pH of the acetate buffer is preferably 5 to 8. By using an acetate buffer as the preservation solution, protein aggregation is suppressed, and the intermediate can be preserved for a desired period of time.

[0019] The concentration of the intermediate in the above-mentioned preservation solution may be 0.1 mg / mL or higher, or 1 mg / mL or higher. It may also be 50 mg / mL or lower, or 5 mg / mL or lower.

[0020] The intermediate preservation solution preferably contains neutral amino acids, for example, because they have radical scavenging activity. Among neutral amino acids, those with high solubility are more preferable, for example, because they increase the stability of the antibody in the preservation solution. Examples of neutral amino acids with high solubility include asparagine, glutamine, serine, threonine, cysteine, alanine, or glycine. The preservation solution may contain one type of neutral amino acid or two or more types.

[0021] The concentration of neutral amino acids in the intermediate storage solution is preferably 1 μM or higher, more preferably 10 μM or higher, and even more preferably 50 μM or higher, as this increases the stability of the antibody in the storage solution. Furthermore, the concentration of neutral amino acids in the above storage solution is preferably 260 mM or lower, more preferably 200 mM or lower, and even more preferably 150 mM or lower.

[0022] The storage solution for the intermediate preferably contains polysorbate because it has radical scavenging activity and can stably store the intermediate by suppressing the aggregation of antibodies (proteins). Examples of polysorbate include polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65, and polysorbate 80. The storage solution may contain one type of polysorbate or two or more types.

[0023] The concentration of polysorbate contained in the intermediate storage solution is preferably 1 μM or higher, more preferably 10 μM or higher, and even more preferably 50 μM or higher, in order to suppress protein aggregation and stably store the intermediate. Furthermore, the concentration of polysorbate contained in the above storage solution is preferably 400 mM or lower, more preferably 200 mM or lower, and even more preferably 100 mM or lower.

[0024] The storage solution for the intermediate more preferably contains at least one neutral amino acid and at least one polysorbate. The presence of a neutral amino acid and polysorbate in the storage solution, which is an acetate buffer, allows for a synergistic effect between the acetate buffer, neutral amino acid, and polysorbate, further suppressing protein aggregation and enabling stable storage of the intermediate. Furthermore, this also has the effect of suppressing the radioactive degradation of the radiopharmaceutical composition prepared using the intermediate stored in the storage solution.

[0025] The storage time for the intermediate is 24 hours or more. The storage time for the intermediate may be 48 hours or more, or 1 week or more, or 1 month or more. Alternatively, the storage time for the intermediate may be 3 years or less, or 2 years or less, or 12 months or less.

[0026] The storage temperature of the intermediate is preferably 1°C or higher, and more preferably 3°C or higher, in terms of the stability of the intermediate. Furthermore, the storage temperature of the intermediate is preferably 8°C or lower, and more preferably 5°C or lower.

[0027] The method for preserving the intermediate according to this embodiment may include other steps. Examples of other steps include replacing the solvent of the synthesized intermediate with the preservation solution.

[0028] This embodiment also includes intermediates preserved by the intermediate preservation method according to this embodiment, and a preservation solution (intermediate composition) containing the intermediates after preservation by the said preservation method. Furthermore, a kit containing the intermediate composition and a radionuclide for labeling the intermediate is also included in this embodiment. As shown in the examples, the radiopharmaceutical composition prepared from intermediates preserved by the intermediate preservation method according to this embodiment has radiochemical purity suitable for use in clinical practice.

[0029] Conventional technology lacked a method for long-term storage of intermediates, requiring their preparation immediately before use. However, radiopharmaceutical compositions prepared from intermediates stored for 12 months using the intermediate storage method of this embodiment possess equivalent physical properties to those obtained through preparation immediately before use. Therefore, by using the intermediate storage method of this embodiment, the need for immediate preparation of intermediates is eliminated, reducing the cost of the raw materials for the intermediates, anti-EGFR antibody and PCTA. Furthermore, eliminating the need for immediate preparation of intermediates significantly reduces labor costs associated with intermediate preparation and quality testing. In addition, the intermediate storage method of this embodiment is compliant with pharmaceutical GMP (Good Manufacturing Practice).

[0030] [Method for preparing radiopharmaceutical compositions] The method for preparing the radiopharmaceutical composition according to this embodiment includes a labeling step of labeling the intermediate stored by the above storage method with a radionuclide that coordinates with PCTA.

[0031] (labeling process) The radionuclides used in the labeling process are radionuclides that coordinate with PCTA. Examples of such radionuclides include: 64 Cu and 225 Examples include Ac.

[0032] Labeling of the intermediate with a radionuclide can be done by known methods. For example, as shown in the examples, the intermediate can be labeled with a radionuclide by mixing a storage solution containing the intermediate with a solution containing the radionuclide and incubating it at 1 to 60°C for 5 minutes to 12 hours. The solution containing the radionuclide is preferably an acetate buffer, and more preferably an acetate buffer with the same pH as the storage solution.

[0033] The method for preparing the radiopharmaceutical composition according to this embodiment may include steps other than the labeling step described above. Examples of other steps include a step for measuring the radiochemical purity of the radiopharmaceutical composition.

[0034] [Method for preserving radioactive pharmaceutical compositions] The method for preserving the radiopharmaceutical composition according to this embodiment includes preserving the radiopharmaceutical composition in a preservation solution.

[0035] (Preservative solution for radiopharmaceutical compositions) The preservation solution used in the method for preserving the radiopharmaceutical composition according to this embodiment (hereinafter sometimes referred to as "preservation solution for the radiopharmaceutical composition") is an acetate buffer, similar to the preservation solution for the intermediate. By using an acetate buffer as the preservation solution, protein aggregation is suppressed, and the radiopharmaceutical composition can be preserved for a desired period of time.

[0036] The preferred embodiment of the preservation solution for the radiopharmaceutical composition is the same as the preferred embodiment of the preservation solution for the intermediate. In particular, the inclusion of neutral amino acids and polysorbate in the preservation solution for the radiopharmaceutical composition, which is an acetate buffer, further suppresses protein aggregation, allowing for stable preservation of the radiopharmaceutical composition. This also has the effect of suppressing radioactive decomposition of the radiopharmaceutical composition.

[0037] The storage solution for the radiopharmaceutical composition and the storage solution for the intermediate may be the same or different. It is preferable that the storage solution for the radiopharmaceutical composition and the storage solution for the intermediate be the same, as this eliminates the need for solvent substitution, simplifies the preparation of the radiopharmaceutical composition, and reduces costs.

[0038] The concentration of the radiopharmaceutical composition contained in the above-mentioned preservation solution may be 5 MBq / mL or higher, or 25 MBq / mL or higher. It may also be 5 GBq / mL or lower, or 500 MBq / mL or lower.

[0039] The storage time for the radiopharmaceutical composition may be 30 minutes or more, 1 hour or more, or 3 hours or more. The storage time for the radiopharmaceutical composition may also be 24 hours or less, 18 hours or less, or 12 hours or less. The preferred storage temperature range for the radiopharmaceutical composition is preferably 1°C or higher, and more preferably 15°C or higher. The preferred storage temperature for the intermediate is preferably 30°C or lower, and more preferably 25°C or lower.

[0040] This embodiment also includes a radiopharmaceutical composition preserved by the radiopharmaceutical composition preservation method according to this embodiment, and a preservation solution (pharmaceutical preparation) containing the radiopharmaceutical composition after preservation by said preservation method. As shown in the examples, the radiopharmaceutical composition preserved by the radiopharmaceutical composition preservation method according to this embodiment has radiochemical purity suitable for use in clinical practice.

[0041] Conventional technology lacked a method for preserving radiopharmaceutical compositions, requiring their preparation immediately before use. However, the radiopharmaceutical composition preservation method according to this embodiment allows for stable storage of the radiopharmaceutical composition for 24 hours after preparation. Therefore, using the radiopharmaceutical composition preservation method according to this embodiment eliminates the need for immediate preparation of the radiopharmaceutical composition, enabling the transportation of the prepared radiopharmaceutical composition over a wide area. Furthermore, eliminating the need for immediate preparation of the radiopharmaceutical composition significantly reduces the labor costs required for the preparation and quality testing of the radiopharmaceutical composition. The radiopharmaceutical composition preservation method according to this embodiment is compliant with pharmaceutical GMP (Good Manufacturing Practice).

[0042] 〔summary〕 The method for preserving an intermediate of a radiopharmaceutical composition according to this embodiment includes preserving the intermediate in an acetate buffer, wherein the intermediate is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody), and the preservation time is 24 hours or more.

[0043] In the preservation method according to this embodiment, the acetate buffer may contain at least one neutral amino acid and at least one polysorbate.

[0044] In the storage method according to this embodiment, the anti-EGFR antibody may be cetuximab.

[0045] The method for preparing the radiopharmaceutical composition according to this embodiment includes a labeling step of labeling the intermediate stored by the above storage method with a radionuclide that coordinates with PCTA.

[0046] In the method for preparing a radiopharmaceutical composition according to this embodiment, the radionuclide is 64 Cu or 225 Ac is fine.

[0047] A method for preserving a radiopharmaceutical composition according to this embodiment includes preserving the radiopharmaceutical composition in an acetate buffer, wherein the radiopharmaceutical composition is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody), and is labeled with a radionuclide that coordinates with PCTA.

[0048] Regarding the method for storing the radiopharmaceutical composition according to this embodiment, the radiopharmaceutical composition may be stored in the acetate buffer for 30 minutes or more.

[0049] In the method for preserving a radiopharmaceutical composition according to this embodiment, the radionuclide is 64 Cu or 225 Ac is fine.

[0050] In the method for preserving the radiopharmaceutical composition according to this embodiment, the anti-EGFR antibody may be cetuximab.

[0051] In the method for preserving a radiopharmaceutical composition according to this embodiment, the acetate buffer may contain at least one neutral amino acid and at least one polysorbate.

[0052] The intermediate composition according to this embodiment is an intermediate composition used in the preparation of a radiopharmaceutical composition, comprising an intermediate of the radiopharmaceutical composition and a preservation solution, wherein the intermediate is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an antibody, and the preservation solution comprises at least one neutral amino acid, at least one polysorbate and an acetate buffer, and the intermediate is stored in the preservation solution for 24 hours or more.

[0053] The pharmaceutical preparation according to this embodiment comprises a radiopharmaceutical composition and a preservation solution, wherein the radiopharmaceutical composition is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an antibody, and is labeled with a radionuclide that coordinates with PCTA, and the preservation solution comprises at least one neutral amino acid, at least one polysorbate and an acetate buffer, and the radiopharmaceutical composition is obtained by labeling the complex, which has been stored in the preservation solution for 24 hours or more, with a radionuclide that coordinates with PCTA.

[0054] The pharmaceutical preparation according to this embodiment contains the radioactive nuclide 64 Cu or 225 Ac is fine.

[0055] Examples are shown below to explain the embodiments of the present invention in more detail. Of course, the present invention is not limited to the following examples, and it is needless to say that various aspects are possible in terms of details. Furthermore, the present invention is not limited to the above-described embodiments, and various modifications are possible within the scope shown in the claims. Embodiments obtained by appropriately combining the disclosed technical means are also included in the technical scope of the present invention. Also, all the documents described in this specification are incorporated by reference.

Example

[0056] In the following examples, unless otherwise specified, % represents mass %.

[0057] 〔Materials and Methods〕 〔 64 Synthesis Method of Cu-PCTA-Cetuximab〕 Cetuximab, an EGFR antibody, was obtained by a known method. Specifically, the DNA strand encoding cetuximab, obtained using the gene sequence of cetuximab, was transfected into Chinese hamster ovary (CHO) cells, and the synthesized cetuximab was purified.

[0058] According to the description in Nucl Med Biol 43 (11):685-691, Cu was prepared and purified using a cyclotron. 64 Cu was prepared and purified. 64 For the purpose of coordinating with Cu and binding to cetuximab, 3,6,9,15-Tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-4-S-(4-isothiocyanatobenzyl)-3,6,9-triacetic acid (p-SCN-Bn-PCTA, Macrocyclics, Plano, TX, USA) was used. Sterile water of pharmaceutical grade was used, and other chemical substances of chemical grade were used.

[0059] Formation of the complex of cetuximab and PCTA and 64The Cu labeling was performed using the method described in the following literature, with some modifications: Cancer Sci 103 (3):600-605; PLoS One 8 (4):e61230; PLoS One 10 (4):e0123761. Briefly, a cetuximab solution in borate buffer (0.05 M, pH 8.5) (2 mg / mL) was prepared by buffer substitution using a Vivaspin ultrafiltration device (Sartorius, Aubagne, France). The cetuximab borate buffer was mixed with a PCTA dimethyl sulfoxide solution (0.42 mg / mL) in a 5:1 ratio (PCTA:cetuximab, molar ratio). The mixture was then reacted at 37°C for 16 hours to prepare PCTA-cetuximab. The prepared PCTA-cetuximab was diluted to 2 mg / mL using three preservation solutions shown in Table 1 via ultrafiltration centrifugation and concentration tubes.

[0060] [Table 1]

[0061] Each PCTA-cetuximab solution, which was replaced with the preservation solution listed in Table 1, was dissolved in acetate buffer (0.1 M, pH 6.0). 64 CuCl2 in a 3:1 ratio ( 64 CuCl2:PCTA-cetuximab was added and mixed in a vol:vol ratio. The mixture was then reacted at 40°C for 1 hour. 64 Cu-PCTA-cetuximab was prepared.

[0062] [ 64 [Analytical method for the radiochemical purity of Cu-PCTA-cetuximab] Prepared 64The radiochemical purity of Cu-PCTA-cetuximab was analyzed by RadioTLC (radio-thin layer chromatography). Silica gel (silica gel 60 plate, Merck, Whitehouse Station, NJ, USA) was used as the support, and 80% methanol (methanol:water = 80:20 (vol / vol)) was used as the mobile phase. Free particles that do not coordinate with PCTA-cetuximab were analyzed. 64 To detect Cu, 64 5 μL of 25 mM EDTA was added to a 5 μL sample of Cu-PCTA-cetuximab and analyzed by radioTLC. For comparison, 64 A 0.1M Cu acetate buffer (pH 6.0) solution is treated similarly. 64 Cu was used as a standard sample. These analytical samples were dropped onto thin silica gel plates in 1 μL increments and developed using a mobile phase. The radiochemical purity of the TLC plates was then detected using an imaging analyzer (FLA7000, GE Healthcare Life Science, Marlborough, MA, USA). 64 Cu-PCTA-cetuximab and 64 The proportion of Cu was calculated. Relative radioactivity is the intact radioactivity relative to total radioactivity. 64 The percentage was calculated as Cu-PCTA-cetuximab. Radiochemical purity of 95% or higher was used. 64 This was used as the target specification standard for Cu-PCTA-cetuximab.

[0063] [To cetuximab-expressing cells] 64 [Method for evaluating the binding affinity of Cu-PCTA-cetuximab] 64 HCT116 cells (human colorectal cancer cells; CCL-247; American Type Cell Collection) were used to evaluate the cell binding affinity of Cu-PCTA-cetuximab. HCT116 cells were diluted in phosphate buffer containing 1% bovine serum albumin (Sigma-Ace) and incubated on ice for 1 hour. 64The cells were reacted with Cu-PCTA-cetuximab. After washing the cells, the radioactivity was measured using a gamma counter (1480 Automatic gamma counter Wizard 3, PerkinElmer). 64 The cell binding activity of Cu-PCTA-cetuximab was evaluated. Immunobinding activity was determined by the method of Lindmo et al. (J Immunol Methods. 1984; 72:77-89). In addition, HCT116 cells were subjected to a competitive inhibition assay. 64 Cu-PCTA-cetuximab and unlabeled cetuximab at various concentrations were incubated on ice for 1 hour. After washing, as described above, 64 The cell binding affinity of Cu-PCTA-cetuximab was evaluated. The dissociation constant was determined based on the results of a competitive inhibition assay.

[0064] [Statistical analysis] Data regarding cell connectivity were expressed as mean values ​​with corresponding standard deviations. P-values ​​were calculated using analysis of variance and compared across multiple groups. A P-value less than 0.05 was considered statistically significant.

[0065] [Experiment 1.] 64 Effect of preservation solution on the labeling efficiency of Cu-PCTA-cetuximab Using PCTA-cetuximab prepared with the three preservation solutions listed in Table 1, samples were collected immediately after preparation (0 months) and after refrigeration at 4°C for 3, 6, 8, 9, and 12 months. 64 Cu-PCTA-cetuximab was prepared. 64 The stabilizing effect of the three storage solutions on PCTA-cetuximab was evaluated by comparing and examining the radiochemical purity (labeling efficiency) immediately after labeling. The evaluation results are shown in Figure 1.

[0066] In Figure 1, the vertical axis represents radiochemical purity (labeling efficiency, %). The horizontal axis represents the storage time (months) of PCTA-cetuximab after preparation with the preservation solution. "*" indicates results where the radiochemical purity was less than 95%.

[0067] As shown in Figure 1, PCTA-cetuximab stored in the comparative example's storage solution (triangle in Figure 1) showed a decrease in labeling efficacy after long-term storage, falling below the target specification standard of 95%. On the other hand, the decrease in labeling efficacy was suppressed for PCTA-cetuximab stored in the storage solution of Example 1 (square in Figure 1) and the storage solution of Example 2 (circle in Figure 1). In particular, the PCTA-cetuximab stored in the storage solution of Example 2 showed a high level of maintenance of labeling efficacy.

[0068] [Experiment 2.] 64 [Effect of preservation solution on the stability of Cu-PCTA-cetuximab after labeling] PCTA-cetuximab was prepared using the three preservation solutions listed in Table 1 above, and samples were stored refrigerated at 4°C for 3 months or 12 months. 64 Cu-PCTA-cetuximab was prepared. 64 (Cu, 37 MBq, 20 μg PCTA-cetuximab, total volume 40 μL). By comparing and examining the radiochemical purity (stability) of the three storage solutions immediately after labeling, and at 0.5, 1, 3, and 24 hours after labeling, the three storage solutions were evaluated. 64 The stabilizing effect on Cu-PCTA-cetuximab was evaluated. The evaluation results are shown in Figures 2 and 3.

[0069] Figure 2 shows PCTA-cetuximab stored in preservation solution for 3 months. 64 Figure 3 shows the results of a comparison of radiochemical purity (stability) after Cu labeling. 64 This shows the comparative results of radiochemical purity (stability) after Cu labeling. The vertical axis in Figures 2 and 3 shows radiochemical purity (labeling efficiency, %). The horizontal axis shows: 64 This shows the elapsed time after Cu labeling. "*" and "#" indicate results where the radiochemical purity was less than 95%.

[0070] As shown in Figures 2 and 3, PCTA-cetuximab stored in the comparative example's storage solution (triangle in Figures 2 and 3) showed a radiochemical purity below the target specification standard of 95% immediately after labeling. On the other hand, PCTA-cetuximab stored in the storage solution of Example 1 (square in Figures 2 and 3) and the storage solution of Example 2 (circle in Figures 2 and 3) showed a radiochemical purity above the target specification standard of 95% immediately after labeling.

[0071] As shown in Figure 2, the radiochemical purity of PCTA-cetuximab stored in the preservation solution of Example 1 fell below 95% after 3 hours from labeling. On the other hand, as shown in Figures 2 and 3, the radiochemical purity of PCTA-cetuximab stored in the preservation solution of Example 2 was maintained even after 3 months or 12 months of storage, up to 24 hours after labeling.

[0072] The results of Experiments 1 and 2 suggest that, for example, it is possible to transport the intermediate stored in the preservation solution of the Examples and prepare a radiopharmaceutical composition from the intermediate at the destination. Furthermore, it was found that the radiopharmaceutical composition can be stored stably for 24 hours by storing it in the preservation solution of the Examples after preparation.

[0073] [Experiment 3.] 64 [Effect of preservation solution on the cell binding properties of Cu-PCTA-cetuximab] PCTA-cetuximab prepared in the three preservation solutions listed in Table 1 above and stored at 4°C for 12 months was used. 64 Cu-PCTA-cetuximab was prepared. 64 Cu, 37 MBq, 20 μg PCTA-cetuximab (total volume 40 μL). HCT116 cells (6.25 × 10⁶) 5 The individual was diluted in 1 mL of phosphate buffer containing 1% bovine serum albumin (BSA). 64 Add Cu-PCTA-cetuximab and incubate on ice for 1 hour. 64The cell binding properties of Cu-PCTA-cetuximab were compared. Following the description in Non-Patent Document 3, a conjugate of commercially available cetuximab (trade name Erbitux, Merck) and PCTA was used. 64 The solution labeled with Cu was used as a positive control. The biochemical evaluation of the stabilizing effect of the three preservation solutions on PCTA-cetuximab is shown in Figure 4.

[0074] The vertical axis in Figure 4 shows the percentage of cell binding compared to the positive control, where the cell binding result is set to 100%. "NS" in Figure 4 indicates that there was no significant difference compared to the positive control's cell binding result.

[0075] As shown in Figure 4, under any storage conditions 64 The cell binding affinity of cu-PCTA-cetuximab was maintained at a level comparable to that of the positive control.

[0076] [Experiment 4.] 64 [Effect of labeling radioactivity on the cell binding affinity of Cu-PCTA-cetuximab] The effect of radioactivity concentration at the time of labeling on cell binding was investigated using PCTA-cetuximab stored in the preservation solution of Example 2. 64 Mix Cu with 0.1M acetate buffer (pH 6.0) in a 3:1 ratio. 64 Cu:PCTA-cetuximab (v / v) was mixed. 64 Three solutions with radioactivity concentrations of 520 MBq, 370 MBq, and 130 MBq were tested as 0.1 M Cu acetate buffer. The reaction was carried out at 40°C for 1 hour. 64 Cu-PCTA-cetuximab was prepared (PCTA-cetuximab 90 μg, total volume 5 mL). Then, under the same conditions as in Experiment 3, 64 The cell binding affinity of Cu-PCTA-cetuximab was evaluated. This was evaluated in Experiment 3. 64 Cu-PCTA-cetuximab ( 64 A comparison was made using a 20 μg dose of PCTA-cetuximab (37 MBq of Cu, total volume 40 μL). The comparison results are shown in Figure 5.

[0077] The vertical axis in Figure 5 represents the results evaluated in Experiment 3. 64 This shows the percentage of cell binding when the cell binding result of Cu-PCTA-cetuximab is set to 100%.

[0078] As shown in Figure 5, no effect on cell binding was observed even at a radioactivity concentration of 520 MBq (PCTA-cetuximab 90 μg, total volume 5 mL). Based on these results, at amounts and concentrations usable for diagnosis in clinical practice, 64 It was confirmed that Cu-PCTA-cetuximab can be prepared stably.

[0079] [Experiment 5.] 225 [Effect of preservation solution on the stability of Ac-PCTA-cetuximab after labeling] Using PCTA-cetuximab preserved in the preservation solution of Example 2, 225 Labeling with Ac and the stability after labeling were investigated. 225 Mix 0.1M Acetate buffer (pH 6.0) with Acetate in a 3:1 ratio. 225 Ac:PCTA-cetuximab (v / v) was added and mixed. The mixture was reacted at 40°C for 1 hour. 225 Ac-PCTA-cetuximab was prepared. 225 The evaluation results for Ac-PCTA-cetuximab are shown in Figure 6.

[0080] The bar graph on the left side of Figure 6 is, 225 This shows the radiochemical purity immediately after labeling with Ac. The bar graph on the right shows: 225 This shows the radiochemical purity 24 hours after labeling with Ac.

[0081] As shown in Figure 6, 225 The radiochemical purity immediately after Ac labeling exceeded the target standard of 95%. 225 It was found that the radiochemical purity was maintained up to 24 hours after Ac labeling. From these results, the preservation solution of the present invention is 64 Other than Cu 225 It was found that this method can also be applied to other metallic radionuclides such as Ac.

[0082] Cetuximab 225 When labeling Ac, DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) is widely used as a ligand. 225 High pH and high temperature are required to coordinate Ac to DOTA.

[0083] on the other hand, 225 When preparing Ac-PCTA-cetuximab, under mild conditions (40°C, pH 6.0) 225 Ac was efficiently coordinated to PCTA. Furthermore, despite being coordinated under mild conditions, the radiochemical purity was high, exceeding 95%. [Industrial applicability]

[0084] This invention can be used for intermediates of radiopharmaceutical compositions and for the long-term storage of radiopharmaceutical compositions. Therefore, this invention enables the stable and efficient production of radiopharmaceutical compositions.

Claims

1. This includes storing an intermediate of a radiopharmaceutical composition in an acetate buffer, The intermediate is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody). A storage method that allows for storage for 24 hours or more.

2. The storage method according to claim 1, wherein the acetate buffer comprises at least one neutral amino acid and at least one polysorbate.

3. The storage method according to claim 1 or 2, wherein the anti-EGFR antibody is cetuximab.

4. A method for preparing a radiopharmaceutical composition, comprising a storage step of storing the intermediate by the storage method described in any one of claims 1 to 3, and a labeling step of labeling with a radionuclide that coordinates with PCTA.

5. The aforementioned radioactive nuclide ６４ Cu or ２２５ The preparation method according to claim 4, wherein the result is Ac.

6. The method includes storing the radiopharmaceutical composition in an acetate buffer, The radiopharmaceutical composition is a complex of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid (PCTA) and an anti-epidermal growth factor receptor antibody (anti-EGFR antibody), wherein the complex, which has been stored in acetate buffer for 24 hours or more, is labeled with a radionuclide that coordinates with PCTA. A method for preserving the radiopharmaceutical composition, comprising preserving it in the acetate buffer for one hour or more.

7. The aforementioned radioactive nuclide ６４ Cu or ２２５ The storage method according to claim 6, wherein it is A.

8. The storage method according to claim 6 or 7, wherein the anti-EGFR antibody is cetuximab.

9. The storage method according to any one of claims 6 to 8, wherein the acetate buffer comprises at least one neutral amino acid and at least one polysorbate.

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