KR20040098319A - Disulfide reduced mannosylated serum albumins and their radioisotope labeled compounds for lymphatic system image thereof - Google Patents

Abstract

PURPOSE: Provided are disulfide reduced mannosylated serum albumin and a radioisotope labeled compound containing the same for lymphaticsystem image. The radioisotope labeled compound containing disulfide reduced mannosylated serum albumin is labeled more easily than conventional isotope labeled compounds for lymphaticsystem image and has excellent absorption of lymphaticsystem. CONSTITUTION: The disulfide reduced mannosylated serum albumin is represented by the formula(1) of (Man-L)n-A-(SH)m, wherein Man is a mannosyl group; L is, as a linker, present or not and when it is not present, Man and A bind together; A is disulfide reduced serum albumin; SH is an thiol group; n is an integer of 1-42; and m is an integer of 2-34.

Description

Disulfide reduced mannosylated serum albumins and their radioisotope labeled compounds for lymphatic system image

The present invention relates to a disulfide reduced mannose-binding serum albumin and a method for preparing the same, and a radioisotope labeling compound and a kit for radiolabeling.

In nuclear medicine, lymphatic system imaging is important for imaging and diagnosing lymph node function. Particularly, in breast cancer or melanoma, if the tumor is injected with radioactive colloid and the lymph node, which is the closest lymph node that is ingested into the lymph node, is imaged and resected, the biopsy can be performed to determine the metastasis. It has a great effect on side effects and beauty after. Therefore, lymphatic system imaging, especially images of sentinel lymph nodes, will become increasingly important [G. Mariani, L. Moresco, G. Viale, G. Villa, M. Bagnasco, G. Canavese, J. Buscombe, HW Strauss, G. Paganelli. "Radioguided sentinel lymph node biopsy in breast cancer surgery." J. Nucl. Med. 2001; 42: 1198-1215; G. Mariani, M. Gipponi, L. Moresco, G. Villa, M. Bartolomei, G. Mazzarol, MC Bagnara, A. Romanini, F. Cafiero, G. Paganelli, HW Strauss. "Radioguided sentinel lymph node biopsy in malignant cutaneous melanoma" J. Nucl. Med. 2002; 43: 811-827.

The principle of lymph node simography is to image tissues that are absorbed into the lymphatic system, migrate to the lymph nodes, and accumulate when injected into the tissue. Most radiopharmaceuticals used for imaging lymph nodes are ^99m Tc-labeled colloidal strains, and the smaller the colloid particle size, the faster the lymph node absorption, which is an ideal radiopharmaceutical. The smallest colloid currently used is antimony sulfur colloid, which is about 10-20 nm in size, albumin nanocolloid is 50 nm, and sulfur colloid is 100-200 nm, so antimony sulfur colloid is the most ideal. . However, since the intercellular gap of the lymphatic system is about 5 nm, the antimony sulfur colloid is relatively smaller than other colloids, so that the intake is fast. However, there is a disadvantage in that the particles are too large. For patients who need to know, there is a difficult problem. In addition, it should be manufactured in strong acidity, but if neutralized after preparation, colloid may become unstable.

In order to avoid this disadvantage, ^99m Tc-phosphoral albumin with a molecular diameter of 6-8 nm may be used. However, the labeling time is short and lymph node migration speed is high. It is weak and has problems with intake in other lymph nodes as well as sentinel lymph nodes (WT Phillips, T. Andrews, H.-L. Liu, R. Klipper, AJ Landry Jr, R. Blumhardt, B. Goins. "Evaluation of [ ^99m Tc] liposomes as lymphoscintigraphic agents: comparison with [ ^99m Tc] sulfur colloid and [ ^99m Tc] human serum albumin " Nucl. Med. Biol. 2001; 28: 435-444).

To address this problem, radiopharmaceuticals have been developed that bind to mannose receptors in lymphocyte immune cells. For example, a compound that binds amphoteric chelating agents such as DTPA or MAG _{3 to} dextran combined with mannose and labels ^99m Tc (DR Vera, AM Wallace, CK Hoh. "[ ^99m Tc] MAG _3- mannosyl-dextran: a receptor-binding radiopharmaceutical for sentinel node detection " Nucl. Med. Biol. 2001; 28: 493-498; DR Vera, AM Wallace, CK Hoh." A synthetic macromolecule for sentinel node detection: ^99m Tc-DTPA -mannosyl-dextran " J. Nucl. Med. 2001; 42: 951-959), ^99m Tc-labeled compound by binding DTPA to polylysine bound with mannose (DR Vera, ER Wisner, RC Stadalnik) "Sentinel node imaging via a nonparticulate receptor-binding radiotracer" J. Nucl. Med. 1997; 38: 530-535). However, in order to label ^99m Tc on a carrier such as dextran or polylysine, it is not a label on its own, so it is necessary to bind amphoteric chelates such as DTPA or MAG ₃ . As a result, it may cause problems such as change of isoelectric point or induction of immunoreactivity.

The present invention is to solve the above problems, it is easier to label the radioisotope than the radioisotope labeling compound for conventional lymphatic system imaging, has excellent stability, and also has a high lymphatic system accumulation rate has excellent characteristics in lymphatic system imaging It is to provide a disulfide reduced mannose binding serum albumin, a radioisotope labeling compound comprising the same, and a kit for radiolabeling.

The object of the present invention can be achieved using disulfide reduced mannose binding serum albumin which has a strong binding to mannose receptor, easy labeling for radioisotopes, and a molecular size that can be well taken into the lymphatic vessel.

Figure 1a is a schematic diagram of the disulfide reduced mannose binding serum albumin directly bonded to the mannosyl group in the present invention,

Figure 1b is a schematic diagram of the disulfide reduced mannose binding serum albumin in which the mannosyl group is bound through a linker in the present invention,

(At this time, Man represents a mannosyl group, L represents a linker and SH thiol group.)

Figure 2 is administered to the lymphatic system after administration of ^99m Tc-HSA (phosphoral albumin) and ^99m Tc-ASC (antimony sulfate colloid) and ^99m Tc-MSA (disulfide reduced mannose binding serum albumin) of the present invention to the rat This is the video.

In order to achieve the above object, the present invention provides a disulfide reduced mannose-binding serum albumin represented by the following formula (1) for the lymphatic system image and a method for preparing the same.

The present invention also provides a radioisotope labeling compound comprising the disulfide reduced mannose binding serum albumin.

In another aspect, the present invention provides a kit for lymphatic function imaging comprising the disulfide reduced mannose-binding serum albumin, weak chelate-based and stannous chloride, and additionally an antioxidant.

Hereinafter, the present invention will be described in detail.

The present invention includes disulfide reduced mannose binding serum albumin represented by the following formula (1) for lymphatic system function imaging.

(Man-L) _n -A- (SH) _m

(In the above formula, Man is a mannosyl group,

L is a linker, which may or may not exist, in which case Man and A directly bond,

A is disulfide reduced serum albumin,

SH is a thiol group, n is an integer of 1 to 42, and m is an integer of 2 to 34.)

1A and 1B are diagrams illustrating an example of the disulfide reduced mannose-binding serum albumin represented by Chemical Formula 1.

As shown in Formula 1 and Figure 1, the disulfide reduced mannose-binding serum albumin of the present invention is directly bonded to the terminal of the serum albumin (Fig. 1a), or bound using a linker (Fig. 1b) It contains a thiol group by reducing the disulfide in serum albumin.

The serum albumin is a human serum albumin, having a molecular weight of 66,462, a long axis of 8 nm, a short axis of 6 nm, and an isoelectric point (IEP) of 4.8. It also makes up 50% (4 g / dl) of plasma proteins and consists of a single polypeptide chain. In particular, the serum albumin has 17 disulfide bonds, so that 2 to 34 thiol groups are generated in serum albumin upon reduction using a suitable reducing agent.

The thiol group generated in the serum albumin acts as a portion that binds to the radioisotope, and is stable under anoxic conditions and increases in stability at low pH of less than 6 pH. In addition, rapid cooling to a liquid nitrogen temperature allows long-term storage without contamination or loss of thiol groups.

The mannose binding serum albumin of the present invention is bound directly or using a linker when binding mannose and serum albumin. The linker is a C ₁ ~ C ₁₀ alkyl group, C ₄ ~ C ₁₀ aryl group, monopeptide, dipeptide, oligopeptide, C ₄ ~ C ₁₀ cycloalkyl group, benzyl group, thioether, ether, amine, hydra One or more complexes selected from the group consisting of jides, pentose sugars, hexose sugars and alcohols.

In one embodiment, serum albumin to which mannose is bound directly or by using a linker is represented by the following Chemical Formulas 2-3. Formula 2 below shows mannose serum albumin in which mannose and serum albumin are directly bonded, and Formula 3 below is phenylmannosyl serum albumin using a phenyl group as a linker. If necessary, the mannose binding serum albumin may include mannose including 1 to 42 mannose or linker.

The disulfide reduced mannose-binding serum albumin binds strongly to the mannose receptor and accumulates in the lymph nodes so that it can be used as a lymphatic system image by labeling the radioisotope, and in particular, the size of the serum serum albumin is 6 to 8 Lymph node intake is faster because it is smaller in nm than the colloidal radiopharmaceuticals currently used. In addition, disulfide attached to serum albumin by a reducing agent does not use amphoteric chelates such as DTPA or MAG ₃ , thereby solving problems such as isoelectric point change or induction of immunoreactivity.

In another aspect, the present invention includes a method for producing a disulfide reduced mannose-binding serum albumin represented by the formula (1) for lymphatic system imaging.

Specifically, the step of binding mannose with serum albumin directly or using a linker to prepare mannose binding serum albumin (step 1); And,

Reducing the disulfide bond of the mannose-bound serum albumin using a reducing agent containing a thiol group (Step 2) comprises a manufacturing method.

Step 1 may be modified to have a bondable functional group, such as thiocyanate, ester, aldehyde, iminomethoxyalkyl, or the like, by using a conventional chemical reaction to bind the terminal portion of the mannose or the linker with serum albumin. Through the reaction, mannose binding serum albumin is prepared. Specifically, Scheme 1 below is 2-imino-2-methoxyethyl-1-thio-??-D-mannose represented by Chemical Formula 2 (2-imino-2-methoxyethyl-1-thio-β- D-mannose, hereinafter referred to as "IME-thiomannose", is used to show the preparation of mannosyl serum albumin.

As in Scheme 1, D-mannose is acetylated and brominated, thiourea is bound, and chloroacetonitrile is reacted to react cyanomethyl 2,3,4,6-tetra-O-acetyl-1-thio-. β-D-mannopyranoside is synthesized. This was reacted with sodium methoxide in methanol solution to synthesize IME-thiomannose, and the amino group of human serum albumin (HSA) (NH _2- ) and thiocarbamyl were prepared under pH 8-9. Combined in the form to prepare a mannosyl serum albumin represented by the formula (2).

In addition, Scheme 2 shows a process for preparing phenylmannosyl serum albumin represented by the formula (3) by combining phenylmanose and serum albumin.

As shown in Scheme 2, phenylmannose produces a stable phenylmannosyl serum albumin represented by the formula (2) by combining the SCN portion of the phenyl group terminal and the amino group of serum albumin.

Step 2 is to reduce the disulfide bond in the mannose-bound serum albumin with a reducing agent to generate a thiol group to prepare a disulfide reduced mannose-bound serum albumin of the formula (1). The reducing agent is mainly a reducing agent containing a thiol group, for example, 2-mercaptoethanol, dithiothreitol, thioglycolate, cysteine and glutathione are used. In addition, a reducing agent containing a thiol group used in this field Can be used.

In the reduction reaction of step 2, an additional chelating agent may be added to remove the remaining metal ions, thereby increasing the stability of the resulting compound and increasing the labeling efficiency of subsequent radioisotopes. The chelating agent may be used conventional chelate, preferably using EDTA.

The present invention also includes a radioisotope labeling compound containing the disulfide reduced mannose binding serum albumin of formula (1).

The radioisotope is preferably ^99m Tc, ¹⁸⁸ Re, ¹⁸⁶ Re, ⁶⁷ Cu, ⁶⁴ Cu, ²¹² Pb, ²¹² Bi or ¹⁰⁹ Pd, more preferably ^99m Tc or ¹⁸⁸ Re.

The radioisotope can be stably labeled by binding to a thiol group of the disulfide reduced mannose-binding serum albumin.

The radioisotope labeling compound is prepared by adding a disulfide reduced mannose-binding serum albumin of Formula 1 and a radioisotope. These labeling reactions can be injected into the human body immediately after labeling in an sterile, pyrogen-free aqueous solution for administration to the human body.

The labeled radioisotope labeling compound can be parenterally injected into the body, accumulated in the lymphatic system, and used for imaging lymphatic system function through conventional nuclear medicine equipment.

In addition, the present invention is 0.1 to 100 mg / unit dose of disulfide reduced mannose-binding serum albumin represented by the formula (1), 0.1 to 500 mg / unit dose of the weak chelating agent and 0.01 μg / unit dose of stannous chloride Kit for lymphatic system functional imaging in a pharmaceutically acceptable nonpyrogenic sterile form comprising -100 mg / unit dose, and additionally 0-500 mg / unit dose of antioxidant.

The weak chelating agent plays a role of preventing the phenomenon of labeling at the weak binding site of mannose binding albumin and the formation of colloids in the preparation of the radioisotope labeling compound using the liver function imaging kit. , Phosphonate, glucoheptonate, gluconate, glucarate, tartarate, succinate and citric acid, consisting of one or two or more, and the kit of the present invention is preferably 0.1 to 500 mg per unit dose. .

As for stannous chloride, 0.01 microgram-100 mg are preferable.

In addition, the lymphatic system imaging kit of the present invention may further include an antioxidant. The antioxidant is to prevent the formation of a polymer that can be produced by the oxidation of the thiol group of the disulfide reduced galactose-binding serum albumin is bonded to each other, preferably vitamin C or gentic acid, etc., in the kit of the present invention 0 500 mg is preferable.

The lymphatic imaging kit is frozen or lyophilized in a sterile container and in an inert gas atmosphere, preferably cooled in liquid nitrogen and slowly dissolved immediately before use. The kit may supplement buffer sterile vials, saline, syringes, filters, columns, and other auxiliary devices to prepare injection preparations for use by clinical pathologists or technicians. It is well known to those of ordinary skill in the art that the kits can be changed and modified in accordance with the patient's individual needs or diet, as well as in the form in which radioisotopes can be provided or obtained. .

The kit for lymphatic imaging is a radioisotope labeling compound prepared by adding 0.1-500 mCi of radioisotope per mg of disulfide reduced mannose-binding serum albumin at room temperature immediately before use and reacting for 0.1-30 minutes.

As an example, pertechnic acid salt ( ^99m TcO ₄ ⁻ ) prepared by a generator was added to a lymphatic imaging kit and reacted at room temperature for 1 to 30 minutes to prepare a technetium labeled compound with an efficiency of 98 to 100%.

According to an embodiment of the present invention, when the labeling efficiency of the radioisotope labeling compound was measured, ^99m Tc-mannosyl serum albumin was able to obtain a labeling efficiency of 99% or more, and the stability of the radioisotope labeling compound was measured. As shown in Table 2 , the radiochemical purity was found to be stable for 24 hours. In addition, after measuring the femoral lymph node migration of radioisotope labeled compounds after administration to the paw of the mouse, it can be seen that ^99m Tc-mannosyl serum albumin moves more rapidly than other radioisotope labeled compounds as shown in Table 3 This was also confirmed in Figure 2 obtained gamma image after administration to the sole of the rat.

An embodiment of the present invention will be described below.

However, the present invention is not limited by the examples.

Example 1 Preparation of Disulfide Reduced Mannosyl Serum Albumin

Step 1: Preparation of Mannosyl Serum Albumin

As in Scheme 1, 100 g of D-mannose was added to 400 ml of acetic anhydride and 3 ml of perchloric acid while slowly stirring while maintaining the temperature at 30 to 40 ° C. After 30 g of amorphous phosphorus was added to the mixed solution containing D-mannose, the reaction vessel was cooled with ice. 180 g of bromine was slowly added while maintaining the temperature of the liquid mixture at 20 ° C. or lower, and 36 ml (0.9 equivalent) of distilled water was slowly added over 30 minutes while preventing the temperature from rising. After the reaction vessel was blocked and reacted for another 2 hours at room temperature, 300 ml of chloroform was added, and the mixture was poured into a separatory funnel containing 800 ml of ice water. The chloroform layer was collected, filtered to remove phosphorus, and then washed twice with the same amount of ice water. The solution was washed once more with an aqueous sodium bicarbonate solution to remove the remaining acid, dehydrated with calcium chloride, dried under reduced pressure, dissolved in diethyl ether and recrystallized to obtain a crystal having a melting point of 87 ° C. 9.74 g (20 mmol) of 2-S- (2,3,4,6-tetra-O-acetyl-β-D-mannopyranosyl) -2-thioshudourea hydrobromide was added to 40 ml of water and acetone 1: 1 Dissolved in a mixed solution, and 5 ml (79 mmol) of chloroacetonitrile was added to dissolve completely. 3.2 g (23.2 mmol) of potassium carbonate and 4.0 g (40.4 mmol) of sodium bisulfite were added and stirred well at room temperature for 30 minutes, and then the reaction mixture was added to 160 ml of ice water and further stirred for 2 hours. The resulting precipitate was collected by filtration and washed with cold water. The precipitate, dried in air, was dissolved in boiling methanol, filtered to remove impurities and stored in a refrigerator to obtain crystals: the recovery was 72% and the melting point was 95-97 ° C.

40 mg of the recovered crystals were dissolved in 1.5 ml of anhydrous methanol. At this time, it was heated to about 40 ℃ to make it melt well. 0.8 mg of sodium methoxide was added thereto, mixed well, and reacted at room temperature for 48 hours to synthesize 22 mg of IME-thiomannose: yield 55%.

Next, 100 mg of serum albumin was dissolved in 1 ml of 0.2 M boric acid buffer solution (pH = 8.5) and mixed vigorously with 22 mg IME-thiomannose. The reaction was carried out at 37 ° C. for 1.5 hours and frozen at −70 ° C.

Step 2: Preparation of Disulfide Reduced Mannosyl Serum Albumin

40 μl of 0.3 M EDTA (pH 8.0), 40 μl of 1 M sodium bicarbonate and 50 μl of 1.5 M β-mercaptoethanol were added to 1 ml mannosyl serum albumin (13.6 mg / ml) prepared in Step 1, and then 37 ° C. The reaction was carried out for 1 hour at. After the reaction, disulfide reduced mannosyl serum albumin was prepared using a Sephadex G-25 column.

Example 2 Disulfide Reduced Phenylmannosyl Serum Albumin

Step 1: Preparation of Phenylmannosyl Serum Albumin

As in Scheme 2, after dissolving 268 mg serum albumin in 25 ml of 0.1 M carbonic acid buffer solution (pH 9.5), 25 mg α-L-mannopyranosylphenylisothiocyanate was added and then stirred well. The reaction was carried out for 20 hours. The reaction solution was stored frozen at -70 ° C.

Step 2: Preparation of Disulfide Reduced Phenylmannosyl Serum Albumin

To 40 ml of 0.3 M EDTA (pH 8.0), 40 µl of 1 M sodium bicarbonate and 50 µl of 1.5 M β-mercaptoethanol were added to 1 ml phenylmannosyl serum albumin (13.6 mg / ml) prepared in Step 1, 37 ° C. Reaction was carried out for 1 hour. After the reaction, disulfide reduced phenylmannosyl serum albumin was separated using a Sephadex G-25 column, followed by addition of 0.3 ml of glucarate solution (2 mg / ml, pH 7.0, including 6 μg SnCl ₂ · H ₂ O). The aliquots were dispensed into vials at an amount corresponding to 3 mg of protein, and then stored at -70 ° C for freezing or lyophilization.

Example 3 Preparation of Lymphatic System Imaging Kit

1 ml of disulfide-reduced mannosyl serum albumin (13.6 mg / ml), 0.3 ml of glutarate solution (2 mg / ml, pH 7.0, 6 μg SnCl ₂ · H ₂ O) was added to 1 mg of protein The aliquots were dispensed into vials and then frozen or lyophilized at -70 ° C and stored.

Example 4 Preparation of Technicium Labeling Compound Using Lymphatic System Imaging Kit

To the kit of Example 3, 2 ml or 5 ml of a physiological saline solution of pertechnic acid salt ( ^99m TcO ₄ ⁻ ) prepared with ⁹⁹ Mo / ^99m Tc-generator (Du Pont) was added for 1 to 30 minutes at room temperature. Reaction.

Comparative Example 1 ^99m Tc-antimony sulfur colloid ( ^99m Tc-antimony sulfur colloid: ^99m Tc-ASC)

Dissolved in the ASC kit vial prepared by the Korea Atomic Energy Research Institute, ^99m TcO ₄ ^- was mixed and incubated for 30 minutes at 50 ℃ was labeled ( ^99m Tc-DTPA-GSA-Gal).

Comparative Example 2 ^99m Tc-tin colloid ( ^99m Tc-tin colloid)

^99m TcO ₄ ^- was mixed in a tin colloid kit vial prepared by the Korea Atomic Energy Research Institute and incubated at room temperature for 10 minutes for labeling.

Comparative Example 3 ^99m Tc-serum albumin ( ^99m Tc-human serum albumin; HSA)

^99m TcO ₄ ^- was mixed in a HSA kit vial made by Daiichi, and incubated at room temperature for 10 minutes for labeling.

Experimental Example 1 Measurement of Radioisotope Labeling Efficiency

The radioisotope labeling efficiency of ^99m Tc-ASC, ^99m Tc-tin colloid, ^99m Tc-HSA prepared in Comparative Examples 1 to 3 and mannosyl serum albumin of Example 4 was determined by instant thin layer chromatography. The reaction was determined by taking small amounts of the reactants in) and developing with physiological saline and then measuring the radioactivity distribution with a TLC-scanner. The labeled technics remained at the origin and all other technics moved along the solvent shear. The results are shown in Table 1 below.

Labeling Efficiency of Mannosyl Serum Albumin Radiopharmaceutical Cover efficiency (%) ^99m Tc-MSA (Example 4) 99.1 ^99m Tc-ASC (Comparative Example 1) 96.5 ^99m Tc-tin colloid (Comparative Example 2) 99.9 ^99m Tc-HSA (Comparative Example 3) 94.7

As shown in Table 1 , as a result of measuring the labeling efficiency of each radiopharmaceutical, it was confirmed that disulfide-reduced mannosyl serum albumin (MSA) was labeled with an efficiency of 99% or more, and compared with other radiopharmaceuticals of the comparative example. Showed efficiency. Therefore, it was found that practical use is possible.

Experimental Example 2 Stability Experiment of Radioisotope Labeling Compound

Stability was measured by radiochemical purity when the ^99m Tc-MSA labeled in Example 4 was left at room temperature and mixed with phosphorous serum and incubated at 37 ° C. The results are shown in Table 2 below.

Radiochemical Purity over Time (%) Hours (hr) water(%) 37 ℃ Room temperature 0.5 96.0 94.5 1.0 92.2 96.4 2.0 96.0 91.1 3.0 94.4 94.0 6.0 97.4 N.D. 20.0 90.5 91.0 24.0 88.7 95.3

As shown in Table 2 above, when left at room temperature and incubated at 37 ° C. in serum, the purity was maintained at 90% or more for 20 hours, and when incubated at 37 ° C. in serum, it did not fall to 88.7% at 24 hours. Only the actual nuclear medical images were mostly injected within 1 hour after the labeling, so it was confirmed that they are sufficiently practical and stable.

Experimental Example 3 Confirmation of Lymph Node Intake by In Vivo Distribution Experiment

Mice (20 g body weight, male, ICR) were used to measure lymph node uptake by in vivo distribution experiments. 0.5 μCi / μl of the technetium labeling material prepared in Example 4, Comparative Examples 1, 2, and 3 was administered to the sole of the mouse. After 10 minutes and 1 hour, the mice were sacrificed to dissect, each organ was removed, and the radioactivity distributed in each tissue was measured by a gamma counter (CobraIII, Packard, USA), weighed using a scale, and injected per unit g. In vivo distribution was measured by standard methods of determining the percentage for. The results are shown in Table 3 below.

Percent Radioactivity According to Biological Tissues after Administration of Example 4, Comparative Examples 1, 2, and 3 (% ID / g)

Table 3 shows the percentage of radioactivity distributed in each tissue after 10 and 60 minutes. Foot is the injection site and leg is the site of lymph nodes. After 10 minutes, ^99m Tc-MSA migrated to the lymph node most frequently, moving 16.1%, followed by ^99m Tc-ASC 9.9%, ^99m Tc-HSA 2.6%, and finally ^99m Tc-tin. colloid is 0.1% net.

This suggests that the disulfide reduced mannose-binding serum albumin (MSA) -labeled technetium labeled compound of the present invention can be sufficiently used for lymphatic imaging.

Experimental Example 4 Lymph Node Intake Confirmation by Gamma Camera Image

Rats (weight average 200 g, male, Sprague-Dawley) were used to measure lymph node uptake by gamma camera images. 500 μCi / 10 μl of the technique labeled substance prepared in Example 4, Comparative Examples 1, 2, and 3 was administered to the sole of the rat. Images were obtained with a 1 hour gamma ray camera (Sigma 410, Ohio-Nuclear, USA). The results are shown in Fig.

2 , it can be seen that the ^99m Tc-MSA of the present invention has higher lymph node intake than the ^99m Tc-HSA and ^99m Tc-ASC of the comparative example.

As described above, the disulfide reduced mannose-binding serum albumin of the present invention and the radioisotope labeling compound including the same are easier to label the radioisotope than the radioisotope labeling compound for lymphoid imaging, and have superior stability. In addition, the lymphatic system has a high accumulation rate and has excellent characteristics in lymphatic imaging.

Claims (14)

Disulfide reduced mannose binding serum albumin represented by the following formula (1). Formula 1 (Man-L) _n -A- (SH) _m (In the above formula, Man is a mannosyl group, L is a linker, which may or may not exist, in which case Man and A directly bond, A is disulfide reduced serum albumin, SH is a thiol group, n is an integer of 1 to 42, and m is an integer of 2 to 34.) The method according to claim 1, wherein the linker (L) is C _{1 ~} C ₁₀ alkyl group, C _{4 ~} C ₁₀ aryl group, monopeptide, dipeptide, oligopeptide, C ₄ ~ C ₁₀ cycloalkyl group, benzyl group, Disulfide-reduced mannose-binding serum albumin, characterized in that it is a complex of one or more selected from the group consisting of thioether, ether, amine, hydrazide, pentose, hexose and alcohol. The disulfide reduced mannose-binding serum albumin according to claim 1, wherein the Man-L is a mannosyl group or a phenylmannosyl group. Binding mannose to serum albumin directly or using a linker to produce mannose binding serum albumin (step 1); And, A method for producing the disulfide reduced mannose-binding serum albumin of claim 1, comprising reducing the disulfide bond of the mannose-binding serum albumin using a reducing agent containing a thiol group (step 2). The method of claim 4, wherein the reducing agent containing a thiol group is 2-mercaptoethanol, 1,4-dithiodreitol, 2-aminoethanethiol, thioglycolate, cysteine or glutathione. The method of claim 4, wherein the reduction further comprises a chelating agent. 7. A process according to claim 6, wherein said chelating agent is EDTA. A radioisotope labeling compound for lymphatic imaging comprising a disulfide reduced mannose binding serum albumin of claim 1 and a metallic radioisotope. 9. The radioisotope labeling compound for lymphatic imaging according to claim 8, wherein the metallic radioisotope is ^99m Tc, ¹⁸⁸ Re, ¹⁸⁶ Re, ⁶⁷ Cu, ⁶⁴ Cu, ²¹² Pb, ²¹² Bi or ¹⁰⁹ Pd. Kit for lymphoid imaging in a pharmaceutically acceptable nonpyrogenic sterile form comprising the disulfide reduced mannose binding serum albumin, weak chelating agent and stannous chloride. The method of claim 10, wherein the mixing ratio of the lymphatic imaging kit is 0.1 to 100 mg / unit dose of disulfide reduced mannose-binding serum albumin, 0.1 to 500 mg / unit dose of the chelating agent, and 0.1 μg of stannous chloride. Lymphatic system imaging kit, characterized in that the unit dose-100 mg / unit dose. The method of claim 10, wherein the weak chelating agent is for lymphatic system imaging, characterized in that consisting of one or more selected from the group consisting of phosphonate, glucoheptonate, gluconate, glucrate, tartarate, succinate and citric acid Kit. 11. The lymphatic imaging kit according to claim 10, further comprising an antioxidant dose of 0 to 500 mg / unit. The kit for imaging a lymphatic system according to claim 13, wherein the antioxidant is vitamin C or gentic acid.