KR101755295B1 - Iodine radioisotope-labeled azide compounds and compositions containing the same as an active ingredient for use Iodine radioisotope-labeling - Google Patents
Iodine radioisotope-labeled azide compounds and compositions containing the same as an active ingredient for use Iodine radioisotope-labeling Download PDFInfo
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Abstract
The present invention relates to a radioactive iodine-labeled azide compound and a radioactive iodine-labeled composition comprising the same as an active ingredient. More particularly, the present invention relates to a radioactive iodine-labeled azide compound represented by the following general formula (1) A radioactive iodine labeling composition and the like.
[Chemical Formula 1]
(Wherein n is an integer of 1-10 and X is radioactive iodine).
Description
The present invention relates to a radioactive iodine-labeled azide compound and a radioactive iodine-labeled composition comprising the same as an active ingredient. More particularly, the present invention relates to a radioactive iodine-labeled azide-tracer improved in uptake in blood, Can be applied in the field.
For decades, radioactive iodine has been used for radioactive labeling of biomolecules for diagnostic and therapeutic purposes. Specifically, 124 I is positron emission tomography (PET), 123 I and 125 I are radioiodine, which is mainly used for single photon emission computed tomography (SPECT), and 131 I It is used in the treatment of diseases such as thyroid cancer.
As described above, since radioactive iodine is widely used medically, various labeling methods have been developed for labeling biomolecules and small molecules of molecules with radioactive iodine. Among them, the electrophilic aromatic substitution reaction shows high efficiency by a method capable of directly labeling the radioactive isotope. However, most of the labeled compounds synthesized through the above labeling method are often unstable in the animal body and often fail to obtain desired image results, and the strong oxidizing agent used for labeling results in reducing the physiological activity of the biomaterial. A method of indirect labeling synthesis of radioactive iodine capable of solving the above problems has been studied and several prosthetic groups have been developed. However, the method of indirectly labeling radioactive iodine, which has been developed so far, has a problem in that a labeled compound is randomly generated due to lack of chemical selectivity for an active group, and an excessive amount of substrate is required for a high yield due to a slow reaction rate have.
Therefore, there is a need to develop a novel radioactive iodine labeling method that can be applied as an imaging diagnostic and therapeutic substance that is stable in living cells and animals and has less side effects from strong oxidants. Recently, a labeling method using a cyclooctyne compound through a copper-free click reaction has attracted attention as a radioisotope labeling method for nuclear medicine imaging due to its excellent specificity and rapid reaction rate.
Korean Patent Laid-Open Publication No. 2012-0101073 related to the related art discloses iodine-labeled homoglutamic acid and glutamate derivatives, and discloses a technique relating to 125 I-labeled homoglutamic acid and glutamate derivatives. However, the conventional technique has a problem that uptake in blood is not sufficient.
Accordingly, radioactive iodine can be labeled with a radioactive iodine-labeled biodegradable molecule or nanoparticles containing azaide and cyclooctyne at a high reaction rate and a high radiochemical yield through a capper-preclin reaction, If radioactive iodine-labeled azide-tracers are provided that have low uptake in the small intestine, large intestine, and rapid uptake into the kidney, and improved uptake in the blood, this can be used to detect radioiodine labeling of biomolecules or nanoparticles, It is expected to be useful for diagnosis.
Accordingly, one aspect of the present invention is to provide an azide compound labeled with radioactive iodine.
Another aspect of the present invention is to provide a method for labeling radioactive iodine using radioactive iodine-labeled azide compounds.
Another aspect of the present invention is to provide compounds labeled with radioactive iodine.
Another aspect of the present invention is to provide a radioactive iodine labeling composition comprising an azide compound labeled with radioactive iodine as an active ingredient.
Another aspect of the present invention is to provide a radioactive iodine labeling composition comprising a radioactive iodine-labeled compound as an active ingredient.
According to one aspect of the present invention, there is provided a radioactive iodine-labeled azide compound represented by the following formula (1).
[Chemical Formula 1]
(Wherein n is an integer of 1-10 and X is radioactive iodine).
According to another aspect of the present invention, there is provided a biomolecule comprising a radioactive iodine-labeled azide compound represented by Formula 1 and at least one moiety selected from the group consisting of dibenzocyclooctin (DBCO) and cyclooctyne , And a step of subjecting the fluorescent dye or the nanoparticle compound to a copper-free click reaction.
According to still another aspect of the present invention, there is provided a compound labeled with radioactive iodine represented by at least one of the following formulas (100) and (101).
(100)
(101)
Wherein Z and Z 'are each independently a biomolecule, a fluorescent dye or a nanoparticle, n is an integer of 1 to 10, and X is radioactive iodine.
According to another aspect of the present invention, there is provided a radioactive iodine labeling composition comprising the radioactive iodine-labeled azide compound represented by Formula 1 as an active ingredient.
According to another aspect of the present invention, there is provided a radioactive iodine labeling composition comprising a radioactive iodine-labeled compound represented by at least one of Chemical Formulas 100 and 101 as an active ingredient.
The radioactive iodine-labeled azide compound according to the present invention is capable of labeling radioactive iodine with a high reaction rate and a high radiochemical yield through a capper-free reaction to biomolecules or nanoparticles containing cyclooctyne It is stable in vivo, has low absorption in the liver, small intestine, large intestine, etc. and is quick in excretion into the kidney, while improving uptake in the blood, and is useful for radioiodine labeling or medical diagnosis of biomolecules or nanoparticles Lt; / RTI >
Figure 1 (a) shows the results of animal biodistribution of comparative compounds and Figure 1 (b) shows the results of animal biodistribution evaluation of the compound of Example (n = 5 mice per group).
Fig. 2 (a) shows SPECT / CT results after 1 hour of injection of the compound of the Example, and Fig. 2 (b) shows SPECT / CT results of 4 hours after injection of the compound of Example.
Hereinafter, preferred embodiments of the present invention will be described with reference to the accompanying drawings. However, the embodiments of the present invention can be modified into various other forms, and the scope of the present invention is not limited to the embodiments described below.
According to the present invention, there is provided a radioactive iodine-labeled azide follower having low absorption in the liver, small intestine, large intestine and the like and quick release into the kidney, while improving uptake in the blood, and said traceable substance is cyclooctyne ) Can be labeled with radioactive iodine at a rapid reaction rate and a high radiochemical yield through a capper-preclin reaction of biomolecules, fluorescent dyes or nanoparticles.
More specifically, according to the present invention, a radioactive iodine-labeled azide compound represented by the following general formula (1) is provided.
[Chemical Formula 1]
In Formula 1, n is an integer of 1 to 10; X is radioactive iodine. In this case, preferably, n is an integer of 3 to 5, and more preferably, n is 4. The exemplary structure of the radiolabeled iodide-labeled azide compound of the present invention is as follows:
The radioiodine X is of, for example 122 I, 123 I, 124 I , 125 I, 127 I, may be at least one radioactive iodine selected from the group consisting of 131 I and 132 I, preferably 125 I .
The compound represented by Formula 1 according to the present invention is a compound having a cyclooctyne group and / or a dibenzocyclooctyne (DBCO) group through an azide group (-N 3 ) (COCO) -free click reaction can be carried out at a high yield within a short period of time as long as it is a compound having cyclooctyne and / or dibenzocyclooctyne (DBCO) group, The radioactive iodine-labeled azide compound of the invention can be used to facilitate radioiodine labeling.
(I)
The radioactive iodine-labeled azide compound of the present invention can be synthesized by any known synthesizing method, and the synthesis method thereof is not particularly limited, but can be obtained, for example, by the same process as Scheme 1 below.
Scheme 1 below illustrates the radioactive iodine-labeled azide compound of the present invention represented by the general formula (1) wherein n is 4, but it is possible to substitute the starting material, which is a compound represented by the following general formula (4) It will be apparent to those skilled in the art to synthesize the radioactive iodine-labeled azide compounds of the present invention represented by formula (1).
[Scheme 1]
More specifically, a step (step 1) of reacting a compound represented by the formula (4) with a compound represented by the formula (a) 2-bromo-5-iodopyridine to obtain a compound represented by the formula (3) Reacting a compound represented by the formula (3) obtained in the above step 1 with a compound represented by the formula (b) Sn 2 Bu 6 to obtain a compound represented by the formula (2) (step 2); And a step (3) of reacting a compound represented by the formula (2) obtained in the above step 2 and [ 125 I] NaI with an oxidizing agent to prepare a compound represented by the formula (1) Compounds labeled with radioactive iodine can be prepared.
In the method for preparing the compound represented by Formula 1, the azide compound represented by Formula 4 and 2-bromo-5-iodopyridine represented by Formula a are reacted with potassium tert-butoxide (2- (2- (2- (2-azidethoxy) ethoxy) ethoxy) ethoxy) -5-iodopyridine compound represented by the formula (3).
Step 2 is a step of reacting the halogen group of the compound represented by the formula (3) obtained in the step 1 and the Sn 2 Bu 6 compound represented by the formula (b) under a catalyst of tetrakistriphenylphosphinephosphine palladium (Pd (Ph 3 P) 4 ) To obtain a radioactive iodine-labeled precursor compound.
The step 3 is a step of reacting the radioactive iodide metal salt with the tin (Sn, Tin) substituent of the compound represented by the formula 2 obtained in the step 2 as an oxidizing agent such as chloramine T, To obtain a labeled compound.
Here, the radioactive iodide metal salt may be a radioactive iodide alkali metal salt, and the alkali metal salt may be lithium (Li), sodium (Na), potassium (K), rubidium (Rb), cesium Is a radioactive iodide salt.
According to another aspect of the present invention, there is provided a radioactive iodine-labeled azide compound represented by the following formula (1) and a biomolecule comprising at least one moiety selected from the group consisting of dibenzocyclooctin (DBCO) and cyclooctyne , And a step of subjecting the fluorescent dye or the nanoparticle compound to a copper-free click reaction.
[Chemical Formula 1]
(Wherein n is an integer of 1-10 and X is radioactive iodine).
The radioactive iodine labeling method of the present invention can be applied to any object capable of causing a copper-free click reaction using the radioactive iodine-labeled azide compound of the present invention as described above. But not limited to, biomolecules, fluorescent dyes or nanoparticle compounds containing at least one moiety of, for example, dibenzocyclooctin (DBCO) and cyclooctyne.
According to another aspect of the present invention, the step of performing a copper-free click reaction comprises reacting a compound labeled with a radioactive iodine represented by the formula (1), at least one of the following chemical formulas (10) and (11) A fluorescent dye or a nanoparticle compound containing at least one moiety selected from the group consisting of dicyclohexylcarbodiimide (DBCO) and cyclooctyne is subjected to a copper-free click reaction, Wherein the radioactive iodine-labeled compound is labeled with at least one of the radioactive iodine-labeled compounds.
[Chemical Formula 10]
(In the
(100)
(101)
Wherein Z and Z 'are each independently a biomolecule, a fluorescent dye or a nanoparticle, n is an integer of 1 to 10, and X is radioactive iodine.
More specifically, the labeling method of radioactive iodine is a method of labeling a biomolecule, a fluorescent dye, or a nanoparticle compound by substituting at least one moiety of dibenzocyclooctene (DBCO) and cyclooctyne, 11 or a biomolecule, a fluorescent dye, or a nanoparticle compound represented by at least one of the following.
That is, the radioactive iodine labeling method of the present invention can be applied to an object containing at least one moiety of dibenzocyclooctin (DBCO) and cyclooctyne, and more specifically, A fluorescent dye or a nanoparticle compound, or a biomolecule, a fluorescent dye or a nanoparticle compound in which the moiety is substituted with a moiety as described above.
In this connection, in the present invention, Z and Z 'in formulas (10) and (11) are each independently defined as biomolecules, fluorescent dyes or nanoparticles. However, at least one of dibenzocyclooctene (DBCO) and cyclooctyne (10) and (11) themselves, including moieties, can also be referred to as biomolecules, fluorescent dyes or nanoparticle compounds, respectively, wherein at least one of the dibenzocyclooctene (DBCO) and cyclooctyne each of which may constitute a part of a biomolecule, a fluorescent dye or a nanoparticle compound, respectively.
The biomolecule is not particularly limited as long as it is a biomolecule including dibenzocyclooctin (DBCO) and / or cyclooctyne, but may be preferably a peptide, an affibody, an antibody, an oligonucleotide, or the like.
Examples of the peptide include RGD peptide, CGNSNPKSC peptide, VHSPNKK peptide, CTTHWGFTLC peptide, SGKGPRQITAL peptide, SGRSA peptide, FSRYLWS peptide and the like, preferably RGD peptide, as a tumor targeting peptide.
In addition, the above-mentioned epitope bodies include, but are not limited to, Aβ peptides, Apolipo protein A1, CD25, CD28, c-Jun, EGFR, Factor VIII, Fibrinogen, Gp120, HER2, IgA, 7 KDa which can be bound to proteins such as IgE, IgM, IL-8, Insulin, RSV G protein, Taq polymerase, TNF-α, Transferrin, Transthyretin, It can be a small molecule.
Further, the antibody may be, but not limited to, anti-VEGFR, anti-ERBB2, anti-CD20, anti-CD19, anti-CD22, anti-CD33, anti-CD25, -CEA, Anti-MUC1, Anti-TAG 72, and the like.
The oligonucleotides may be DNA, RNA, siRNA, antisense oligonucleotides, and the like, without limitation.
In the radioactive iodine labeling method according to the present invention, the nanoparticles may be nanoparticles that may include dibenzocyclooctene (DBCO) and / or cyclooctyne, but may be metal nanoparticles, synthetic polymer nano- Particles, biopolymer nanoparticles, and the like.
The metal nanoparticles may include, but are not limited to, metals such as gold (Au), platinum (Pt), palladium (Pd), silver (Ag), and copper (Cu); Or metal oxides of metals such as cobalt (Co), manganese (Mn), iron (Fe), nickel (Ni), gadolinium (Gd), molybdenum (Mo), zinc (Zn) And preferably gold nanoparticles.
The synthetic polymer nanoparticles include, but are not limited to, poly (ethylene glycol), poly (vinyl alcohol), poly (acrylic acid), polyhydroxy But are not limited to, poly (hydroxyester), poly (ε-caprolactone), poly (orthoester), polymeric anhydride, polyphosphagene, polypropylene fumarate (propylenefumarate), poly (glycolic acid), poly (lactic acid), poly (lactic-co-glycolic acid) (Hydroxybutyrate-co-valerate), poly (urethane), polymethyl methacrylate (poly (hydroxybutyrate-co-valerate) methyl methacrylate)).
Further, the biopolymer nanoparticles may include, but are not limited to, chitin, chitosan, polylysine, hyaluronic acid, alginic acid, dextran, cellulose, And so on.
The nanoparticles are nanoparticles having a size of 1 nm to 1000 nm, preferably 10 nm to 500 nm, and more preferably 50 nm to 200 nm. When the size of the nanoparticles is less than 1 nm, it is difficult to produce the radioactive iodine-labeled nanoparticles according to the present invention. When the size of the nanoparticles is more than 1000 nm, the nanoparticle inherent characteristics may be lost.
Meanwhile, the fluorescent dye may be at least one selected from the group consisting of cyanine, fluorescein, and rhodamine.
The radioactive iodine labeling method according to the present invention can be used to label radioactive iodine with a biomolecule, a fluorescent dye, a nanoparticle compound or a combination thereof by using the compound represented by the general formula (1) And at the same time shows a high radiochemical yield value of 95% or more, and exhibits a high radiochemical yield value of 98% or more over a 24 hour period in an environment similar to a living body, so that it can be usefully used as a radioactive iodine labeling method.
According to another aspect of the present invention, there is provided a compound labeled with radioactive iodine represented by at least one of the following formulas (100) and (101).
(100)
(101)
In the above Formulas 100 and 101, Z and Z 'are each independently a biomolecule, a fluorescent dye or a nanoparticle; n is an integer from 1 to 10; And X is radioactive iodine.
The present invention also provides a radioactive iodine labeling composition comprising the radioactive iodine-labeled azide compound represented by Formula 1 as an active ingredient; And a radioactive iodine-labeled compound represented by at least one of the general formulas (100) and (101) as an active ingredient.
Further, the present invention may be provided in the form of a composition for medical diagnosis comprising a compound labeled with radioactive iodine represented by at least one of Chemical Formulas 100 and 101 as an active ingredient, wherein the medical diagnosis is not particularly limited, (PET), micro-PET, computed tomography (CT), magnetic resonance imaging (MRI), or radiological diagnostic imaging.
Particularly, the radioactive iodine-labeled azide compound according to the present invention has a rapid reaction rate (hereinafter referred to as " radioactive iodine ") through a capper-free reaction to biomolecules or nanoparticles containing dibenzocyclooctene (DBCO) and / or cyclooctyne And can be labeled with radioactive iodine with a high radiochemical yield, stable in vivo, low in absorption in the liver, small intestine, large intestine, etc., and quick in excretion into the kidney, while uptake in the blood is improved, Or may be useful for radioiodine labeling or medical diagnosis of nanoparticles.
Therefore, according to the present invention, it is expected that a radioactive iodine-labeled product can be effectively obtained for various target compounds including dibenzocyclooctin (DBCO) and / or cyclooctyne structure, for example, peptide. This result is expected to lead to the development of radiopharmaceuticals that can be used in clinical as well as future research.
In particular, when used as an efficient radioactive iodine labeling method for fluorescent wastes such as Cy5.5, which is widely used in in vivo and in vitro studies, a double observation can be used for diagnosis and treatment of diseases in the future (dual monitoring).
Hereinafter, the present invention will be described more specifically by way of specific examples. The following examples are provided to aid understanding of the present invention, and the scope of the present invention is not limited thereto.
Example
One. 125 I cover Azide Synthesis of compounds
Example
Scheme 1 below shows that the biomolecule and the nanomaterial including the dibenzocyclooctyne structure can be labeled with 125 I label which is a tracer compound of the present invention which is a tracer compound capable of efficiently labeling radioactive iodine ([≪ 125 > I] 1).
[Scheme 1]
step 1: 2 - (2- (2- (2- Ah Mapetoxi ) Ethoxy ) Ethoxy ) Ethoxy ) -5- Iodopyridine Produce
To dry DMF (dimethylformamide) (15 mL) of 2- (2- (2- (2-azidethoxy) ethoxy) ethoxy) ethan- Bromo-5-iodopyridine (771 mg, 2.72 mmol) and potassium tert-butoxide (605 mg, 2.72 mmol) were added at room temperature. The reaction mixture was stirred at 65 ° C for 12 hours and cooled at room temperature. The crude product was extracted with diethyl ether three times, and the organic layers were combined, dried over MgSO 4 (magnesium sulfate) and concentrated under reduced pressure. (2- (2- (2- (2-azidethoxy) ethoxy) ethoxy) ethoxy) -5-iodopyridine (100 mg) was obtained by silica gel column chromatography (hexane / ethyl acetate = 1: 426 mg, 63% yield) as a colorless oil.
1 H NMR (CDCl 3): δ 8.29 (d, 1H, J = 2.3), 7.76 (dd, 1H, J = 9.2, 2.3), 6.62 (d, 1H, J = 9.2), 4.42 (t, 2H, J = 4.6), 3.82 (t, 2H, J = 4.6), 3.63-3.72 (m, 10H), 3.37 (t, 2H, J = 5.0).
13 C NMR (CDCl 3 ): 163.01, 152.59, 146.39, 113.78, 82.3, 70.7, 70.1, 69.6, 65.5, 50.7.
HRMS ([M + H] + ) Calcd for C 13
step 2: 2 - (2- (2- (2- Ah Mapetoxi ) Ethoxy ) Ethoxy ) Ethoxy ) -5- ( Tributylstannyl ) Preparation of pyridine
Ethoxy) ethoxy) -5-iodopyridine (300 mg, 0.711 mmol) obtained in the above Step 1 was dissolved in 1, 2, Sn 2 Bu 6 (921 mg, 1.59 mmol) and tetrakistriphenylphosphine palladium (Pd (Ph 3 P) 4 ) (93 mg, 0.08 mmol) were added at room temperature to 4-dioxane (10 mL). The reaction solution was refluxed for 5 hours and cooled at room temperature. The crude product was extracted three times with ethyl acetate, and the organic layers were combined, dried over MgSO 4 and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (hexane / ethyl acetate = 1: 1) to give 2- (2- (2- (2- Butylstannyl) pyridine (198 mg, 48% yield) as a colorless oil.
1 H NMR (DMSO-d 6 ): δ 8.02 (dd, 1H, J = 10.7, 1.5), 7.63 (dd, 1H, J = 7.7, 1.5), 6.75 (d, 1H, J = 8.4), 4.30 ( 2H, J = 5.0), 3.69 (t, 2H, J = 5.0), 3.47-3.56 (m, (m, 6H), 0.99 (t, 6H, J = 8.4), 0.79 (t, 9H, J = 7.7).
13 C NMR (DMSO-d 6 ): 163.8, 153.2, 146.7, 127.1, 111.7, 70.4, 70.2, 69.8, 69.3, 64.9, 50.5, 29.1, 27.2, 14.0, 9.7
HRMS ([M + H] + ) Calcd for C 25 H 47 N 4 O 4 Sn + : 587.2614; Found 587.2625.
step 3: 125 I The labeled 2- (2- (2- (2- (2- Ah Mapetoxi ) Ethoxy ) Ethoxy ) Ethoxy ) -5- Iodopyridine Produce
To a solution of 2- (2- (2- (2-azidethoxy) ethoxy) ethoxy) ethoxy) -5- (tributylstannyl) pyridine (1 mg) obtained in Step 2 in DMSO (Chloramine T) solution (20 μL of 1 mg of 1 X PBS) and acetic acid (20 μL) were added to 100 μL of dimethyl sulfoxide (100 μL). 0.1 M NaOH (50 [mu] L) of [ 125 I] NaI (407 MBq (megabecquerel)) was added to the reaction mixture. The reaction was stirred at room temperature for 10 minutes and quenched with aqueous sodium metabisulfite solution. The crude product was purified by prep HPLC (eluent change: 40% solvent B 0-2 min; 40-100% solvent B 2-22 min; 100% solvent B 23-28 min, retention time: 14.0 min) To give Example compound [ 125 I] 1 (75% radiochemical yield) labeled with 125 I (305.25 MBq).
Radiochemical purity: 99% (analytical HPLC), specific radioactivity; 15.17 GBq / [mu] mol.
The case of the 125 I labeled azide compound ([125 I] 1) by utilizing a simple molecular structure of DBCO (dibenzocyclooctyne) -NH 2 with the response obtained in the [125 I] 1 and 30 minutes a molecule having a structure DBCO The reaction was carried out at a very high yield of 95% or more in a short period of time.
Comparative Example
Scheme 2 below shows a process for synthesizing a 125 I labeled azide compound ([ 125 I] 1) having a structure different from that of the tracer compound of the present invention. The final synthesized compound is [ 125 I] 1 '.
[Scheme 2]
Step 1: N- (2- (2- (2- (2- Ah Mapetoxi ) Ethoxy ) Ethoxy ) Ethyl) -4- Iodobenzamide Produce
To dry DMF (dimethylformamide) (15 mL) of 2- (2- (2- (2-azidethoxy) ethoxy) ethoxy) ethan- 1- (1240 mg, 5 mmol), HBTU (N, N, N ', N "-tetramethyl-O- (lH-benzotriazol- l-yl) uronium hexafluorophosphate) 2000 mg, 5.26 mmol) and DIPEA (diisopropylethylamine) (1.74 mL) at room temperature. The reaction mixture was stirred at room temperature for 3 hours. After the reaction, a 0.2 M aqueous hydrochloric acid solution (50 mL) was added to the reaction solution. The crude product was extracted with ethyl acetate three times, and the organic layers were combined, dried over MgSO 4 (magnesium sulfate) and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (hexane / ethyl acetate = 1: 3) to give N- (2- (2- (2- Amide (2017 mg, 90% yield) as a colorless oil.
1 H NMR (CDCl 3): 7.77 (d, 2H, J = 8.4), 7.52 (d, 2H, J = 8.4), 6.81 (br s, 1H), 3.60-3.66 (m, 14H), 3.35 (t , 2H, J = 5.4).
13 C NMR (CDCl 3 ): 166.94, 137.71, 133.96, 128.85, 98.47, 70.67, 70.65, 70.56, 70.26, 70.08, 69.69, 50.67, 39.94.
HRMS ([M + H] + ) Calcd for C 15 H 22 IN 4 O 4 +: 449.0680; Found 449.0685.
Step 2: N- (2- (2- (2- (2- Ah Mapetoxi ) Ethoxy ) Ethoxy ) Ethyl) -4- ( Tributylstannyl ) Benzamide Produce
Ethyl) -4-iodobenzamide (224 mg, 0.5 mmol) obtained in the above Step 1 was dissolved in 1, 2, To the 4-dioxane (10 mL) was added Sn 2 Bu 6 (580 mg, 1 mmol) and tetrakistriphenylphosphine palladium (Pd (Ph 3 P) 4 ) (58 mg, 0.05 mmol) at room temperature. The reaction solution was refluxed for 5 hours and cooled at room temperature. The crude product was extracted three times with ethyl acetate, and the organic layers were combined, dried over MgSO 4 and concentrated using a rotary evaporator. The residue was purified by silica gel column chromatography (hexane / ethyl acetate = 1: 1) to give N- (2- (2- (2- Stanyl) benzamide (198 mg, 65% yield) as a colorless oil.
1 H NMR (DMSO-d 6 ) 8.42 (t, 1H), 7.72 (d, 2H, J = 8.4), 7.48 (d, 2H, J = 8.4), 3.48-3.54 (m, 12H), 3.32-3.39 (m, 4H), 1.43-1.50 (m, 6H), 1.21-1.29 (m, 6H), 1.02 (t, 6H, J = 8.0), 0.81 (t, 9H, J = 7.3).
13 C NMR (DMSO-d 6 ): 167.02, 146.23, 136.54, 134.64, 126.90, 70.33, 70.30, 70.21, 70.15, 69.76, 69.42, 50.49, 29.11, 27.20, 14.07, 9.71.
HRMS ([M + H] + ) Calcd for C 27 H 49 N 4 O 4 Sn + : 613.2770; Found 613.2781.
step 3: 125 I The labeled N- (2- (2- (2- (2- Ah Mapetoxi ) Ethoxy ) Ethoxy ) Ethyl) -4- Iodobenzamide Produce
To a solution of the N- (2- (2- (2- (2-azidethoxy) ethoxy) ethoxy) ethyl) -4- (tributylstannyl) benzamide (1 mg) 200 μL) and chloramine T (40 μL, 2 mg) were added to 1 × PBS. 0.1 M NaOH (100 [mu] L) of [ 125 I] NaI (133 MBq (megabecquerel)) was added to the reaction mixture. The reaction was stirred at room temperature for 15 minutes and quenched with aqueous sodium metabisulfite solution. The crude product was purified by prep HPLC (eluent change: 20% solvent B 0-2 min; 20-80% solvent B 2-22 min; 80-100% solvent B 22-23 min; 100% solvent B 23-28, retention time (retention time: 18.3 minutes) to obtain a comparative compound [ 125 I] 1 '(85% radiochemical yield) labeled with 125 I (113 MBq).
Radiochemical purity: 99% (analytical HPLC), specific radioactivity; 40.7 GBq / μmol.
2. Cooper - free Through a copper-free click reaction 125 I cover cRGD Of peptide Produce
The following Scheme 3 illustrates the use of the tracer compound [ 125 I] 1 of the present invention synthesized in Scheme 1 above as a cancer targeting peptide as a radioactive iodine-labeled cyclic RGD peptide, which is widely used in biological research and clinical studies .
[Scheme 3]
Specific steps of the scheme 3 are as follows.
Step a: DBCO-NHS ester, DIPEA (diisopropylethylamine) and DMF (dimethylformamide) were added to the substance of the fourth step at room temperature. Stirring at room temperature under basic conditions yielded
Step b: 2 nmol of
Using this method, it is expected that radioactive iodine-labeled products can be obtained using [ 125 I] 1 for various peptides containing the cyclooctane structure.
3. Cooper - free Through a copper-free click reaction 125 I cover DBCO - Cy5 .5 manufacturing
Scheme 4 below is a procedure for labeling Cy5.5 containing dibenzocyclooctane structure with radioactive iodine utilizing [ 125 I] 1.
More specifically, step a of Scheme 4 is as follows.
20 nmol of substance No. 7 containing a commercially available dibenzocyclooctane structure was reacted with [ 125 I] 1 (0.74 MBq (megabecquerel)) at 37 ° C. The radiochemical yield was determined by analytical radio-HPLC (eluent change: 40% solvent D 0-2 min; 40-100% solvent D 2-22 min; 100% solvent D 23-28 min, retention time: ). [ 125 I] 6 could be synthesized with a labeling efficiency of 82% within 30 minutes and 95% within 1 hour.
[Scheme 4]
As a result of this experiment, the tracer compound [ 125 I] 1 of the present invention can be used as an efficient radioactive iodine labeling method for a fluorescent substance such as Cy5.5, which is widely used in in vivo and in vitro studies It is expected to be.
In addition, since Cy5.5 itself is used for the imaging of tumor and joint inflammation such as U87MG, the present radioactive iodine-labeled substance can be applied to dual monitoring that can be used for the diagnosis and treatment of disease in the future, Furthermore, it can be developed as a radiopharmaceutical.
4. 125 Animal vivo distribution of I-labeled azide compounds ( Biodistribution ) evaluation
Comparative Experimental Example
In order to evaluate the in vivo distribution of the compounds synthesized in the comparative examples, the following experiment was conducted.
Comparative Example Compound [ 125 I] 1 '(1 μCi / 100 μL per mouse) was intravenously administered to 20 ICR male mice. Five mice were sacrificed at 0.25, 1, 2, and 4 hours after anesthesia to remove heart, lung, liver, spleen, kidney, stomach, thyroid, and enteric blood. The organ and blood weights and radioactivity were measured and analyzed. The radioactivity was measured with a gamma-counter. The distribution data is expressed as a percentage injected dose per gram tissue value, and the results are shown graphically in Fig. 1 (a). FIG. 1 (a) is a graph showing the biodistribution in the organ and blood according to time after intravenous administration of the track of Comparative Example to ICR mice.
As shown in Fig. 1 (a), the comparative compound was absorbed into the liver and kidney at a rapid rate within 15 minutes and accumulated at a high concentration, falling to a concentration of 20% or less within 1 hour, , It was found that 90% or more was excreted into the body within 2 hours.
In other words, since the pendulum synthesized in the comparative example has a high integration speed and is quick in time to discharge, the diagnosis can be speeded up at the time of medical diagnosis and the radioactive material remains in the body since it is discharged outside the body after diagnosis And can be usefully used for medical diagnosis.
However, since the uptake in the blood is low in the tracer synthesized in the comparative example, there is a problem that the rate of the copper-free click reaction in vivo is very low.
In addition, the traces synthesized in the comparative examples show high absorption in the small intestine and large intestine, because the body excretion is relatively slow. Therefore, there is a lot of room to act as a background signal in image research.
Experimental Example
In order to evaluate the in vivo organs distribution and in vivo behavior of the 125 I labeled azide of the present invention synthesized in the examples, 6-week-old ICR mice were intravenously injected, and after 15 minutes, 1 hour, 2 hours and 4 hours, The liver, spleen, stomach, small intestine, kidney, heart, lung, muscle, bone and brain were sacrificed and the weight and radioactivity of the sample were measured and analyzed.
The 125 I-labeled azide of the present invention synthesized in the Examples was rapidly observed to be excreted in the kidney and blood through the urinary tract in vivo, and as shown in Fig. 2, the result was consistent with the image result using SPECT / CT Respectively. 2 (a) shows the results after 1 hour after the injection, and FIG. 2 (b) shows the results after 4 hours after the injection.
These results are consistent with a variety of imaging and therapeutic applications in the future and have lower absorption in the liver, small intestine, and large intestine than in the follower compounds synthesized in the above Comparative Examples, . In addition, it was confirmed that the 125 I-labeled azide of the present invention had a particularly high uptake in the undiluted solution as compared to the tracked compound synthesized in the above Comparative Example.
Based on these results, it can be concluded that the tracer compound of the present invention has excellent properties that can be used for pre-targeting imaging in vivo.
While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments, but, on the contrary, It will be obvious to those of ordinary skill in the art.
Claims (16)
[Chemical Formula 1]
(Wherein n is an integer of 1-10 and X is radioactive iodine).
[Chemical Formula 1]
(Wherein n is an integer of 1-10 and X is radioactive iodine).
[Chemical Formula 10]
(In the above formulas 10 and 11, Z and Z 'are each independently a biomolecule, a fluorescent dye or a nanoparticle)
A method of labeling radioactive iodine, which is carried out by obtaining a radioactive iodine-labeled compound represented by at least one of the following formulas (100) and (101)
(100)
(101)
Wherein Z and Z 'are each independently a biomolecule, a fluorescent dye or a nanoparticle, n is an integer of 1 to 10, and X is radioactive iodine.
(100)
(101)
Wherein Z and Z 'are each independently a biomolecule, a fluorescent dye or a nanoparticle, n is an integer of 1 to 10, and X is radioactive iodine.
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| US11298431B2 (en) | 2018-07-17 | 2022-04-12 | Korea Atomic Energy Research Institute | Method for labeling radioisotope radiolabeling compound and kit comprising the same for labeling radioisotope |
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| KR102133823B1 (en) * | 2019-06-12 | 2020-07-15 | 한국원자력연구원 | Method for labeling radioisotope, radiolabeled compounds using quinine compound and kit comprising the same for labeling radioisotope |
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