KR100827293B1 - Gold Nanoparticle?Containing ?? Contrast Agent - Google Patents

Abstract

The present invention relates to a computed tomography (CT) contrast agent comprising gold nanoparticles (Au NPs). CT contrast agent of the present invention may exhibit contrast enhancement, such as iodine contrast agent in a relatively small amount compared to the conventional iodine contrast agent, Au NPs are discharged slowly because it has a much higher molecular weight than iodine contrast agent, which is lower than the iodine contrast agent Longer imaging times, which in turn greatly increase the utility of Au NPs as CT contrast agents. In addition, when applied in vivo, the CT contrast agent of the present invention exhibits contrast in a relatively short time, and the CT contrast agent of the present invention exhibits specificity for the liver or spleen. The CT contrast agent of the present invention can be particularly useful for diagnosing liver cancer.

CT, contrast agent, gold, nanoparticles, PVP, PEG, cancer

Description

CT contrast agent containing gold nanoparticles {Gold Nanoparticle-Containing CT Contrast Agent}

1A is an SEM image of PVP-coated gold nanoparticles (Au NPs) of the present invention having 20 nm (panel a) and 40 nm (panel b) sizes.

Figure 1b shows the results of Electrophoretic Light Scattering Spectrophotometer (ELS) analysis of the PVP-coated Au NPs of the present invention. When PVP-coated Au NPs of the present invention were prepared using 0.4 ml and 0.8 ml of citrate, Au NPs having spherical shapes of about 20 nm (panel a) and about 40 nm (panel b), respectively, I could see that.

Figure 2 is a photograph showing the results of in vitro HU value measurement for 40 nm size PVP-coated Au NPs of the present invention. White dots represent stock solutions of PVP-coated Au NPs.

Figure 3a is a CT image of the liver of the time zone when the mouse injected with 40 nm size PVP-coated Au NPs of the present invention.

Figure 3b is a CT image of the spleen of the time zone mice when the 40 nm size PVP-coated Au NPs of the present invention is injected into the mouse. The arrow points to the spleen.

FIG. 4 is a CT image taken at various time slots after intravenous injection (i.v) in a rat hepatoma model using 20 nm-sized PVP-coated Au NPs as a contrast agent. Panels (a)-(g) are CT images 5 minutes, 1.5 hours, 3 hours, 12 hours, 24 hours and 7 days after contrast agent injection, respectively, after contrast agent injection. The arrow points to hepatoma.

5 is a silver staining result showing the PVP-coated Au NPs Biodistribution. Panels (a)-(f) are results for normal liver tissue, spleen, hepatoma, kidney, lung and heart, respectively.

Figure 6 shows the results of size distribution analysis of PEG-coated Au NPs. Panel (a) is a size distribution graph before PEG coating and panel (b) after PEG coating.

FIG. 7 is a CT image taken at various time slots after injection of intravenous injection (i.v) into a rat hepatoma model using the PEG-coated Au NPs of the present invention as a contrast agent. Panels (a)-(h) are CT images 15 minutes, 1 hour, 2 hours, 4 hours, 12 hours, 24 hours, and 3 days after contrast injection, respectively, before contrast injection. The arrow points to hepatoma.

8 is a solid cancer CT image by PEG-coated Au NPs of the present invention. Panels (a)-(g) are CT images 10 minutes, 1 hour, 2 hours, 4 hours, 12 hours and 24 hours after contrast agent injection, respectively, after contrast agent injection. Arrows indicate solid cancer.

The present invention relates to novel computed tomography (CT) contrast agents.

CT (computed tomography) is a method in which a target area of the human body is irradiated from various directions to collect the transmitted X-rays with a detector, and a computer-based method of reconstructing the difference in absorption of the X-rays to the site using a mathematical technique Talk about shooting techniques. CT has excellent resolution and contrast in distinguishing blood, cerebrospinal fluid, white matter, gray matter, and tumors compared to conventional X-ray images, and can represent minute differences in absorption. .

The basic principle of CT is that the X-ray tube is irradiated with X-ray beam as it travels around a section of the human body, and the detector collects each intensity and the computer calculates the absorption intensity of each part based on the data. Therefore, the image is reconstructed and displayed on the monitor.

On the other hand, Hounsfield unit (HU) is a unit indicating the degree of reduction of X-rays, and CT uses the Hounsfield unit. These hounsfield numbers represent the relative density of the tissue. In other words, a large HU value means that the density is large, so the absorption of X-rays is large and white on CT.

The main feature of CT is the use of X-rays. Short-wave X-rays are effective for imaging bone-dense structures, but the sharpness decreases for imaging soft tissues. That is, the X-ray alone has a limit in distinguishing the tissue from the surrounding area. Therefore, many researches and developments on contrast agents have been made to overcome these problems.

Structures with similar densities are difficult to distinguish by X-ray examination alone. Thus, contrast agents are needed to artificially provide contrast between the organ being examined and the surrounding tissue. Currently, iodine-based contrast agents such as ultravist are the most widely used in CT. The reason for using iodine is because it has a high density and has a high degree of absorption for X-rays, thereby showing an excellent contrast enhancement effect. That is, when X-rays come in contact with iodine in the contrast medium, the X-rays are absorbed and appear white on the CT, so the organ to be diagnosed looks bright. However, since the iodine contrast agent has a very small molecular weight, it has a disadvantage of having a short imaging time since it is removed through the kidney in a few minutes. In addition, instantaneously collected and discharged in the kidney has the disadvantage of having kidney toxicity sometimes.

Thus, there is a need in the art for the development of new contrast agents that can overcome the disadvantages of iodine-based contrast agents.

The present inventors earnestly researched to overcome the problems of the conventional iodine-based CT contrast agent, and confirmed that CT analysis of biological tissues using gold nanoparticles as contrast medium can obtain CT images of desired tissues with improved resolution. The present invention has been completed.

Accordingly, it is an object of the present invention to provide a computed tomography (CT) contrast agent.

Another object of the present invention is to provide a CT contrast agent for cancer imaging.

Other objects and advantages of the present invention will become apparent from the following detailed description, claims and drawings.

According to one aspect of the invention, the invention provides a computed tomography (CT) contrast agent comprising gold nanoparticles.

The present inventors have tried to overcome the problems of the conventional iodine-based CT contrast agent, it was confirmed that CT analysis of the biological tissue using gold nanoparticles as a contrast agent can obtain a CT image of the desired tissue with improved resolution.

The present invention proposes a novel use of gold nanoparticles as CT contrast agents to overcome the disadvantages of conventional iodine-based contrast agents.

Au is about 5 times denser than iodine (Au: 19.3, I: 4.94), and has a high absorption coefficient (X) for X-rays (Au: 5.16 cm ² g ^-1 , iodine: 1.94 cm ² g ^-1 ). Therefore, the inventors expected that gold nanoparticles (hereinafter referred to as "Au NPs") will have contrast enhancement such as iodine contrast agent in a relatively small amount. In addition, Au NPs have a much higher molecular weight than iodine contrast agents, so they are released slowly, which leads to longer imaging times than iodine contrast agents, which in turn increases the utility of Au NPs as CT contrast agents. These inventors' hypotheses were all demonstrated in the following examples, suggesting a novel use of Au NPs as CT contrast agents.

Examples of the use of Au NPs in general X-ray imaging have been reported, but their use as CT contrast agents is the first proposed by the present invention.

The gold nanoparticles used in the present invention can be prepared, for example, as follows: HAuCl ₄ is used as a gold source, and sodium citrate is used as a reducing agent to reduce HAuCl ₄ to prepare gold nanoparticles. In this case, the size of the gold nanoparticles can be adjusted by varying the amount of citrate added. That is, the size of the gold nanoparticles decreases as the amount of citrate increases, so that nucleation increases.

According to a preferred embodiment of the present invention, the gold nanoparticles are coated with a biocompatible polymer. By coating with such a polymer, gold nanoparticles can be stably present without aggregation, and safety is ensured even when injected in vivo. In addition, the coating by the polymer may prevent the proteins in the blood from adsorbed to the gold nanoparticles, to some extent to prevent the removal by the reticuloendothelial system (RES).

As the polymer used for the surface coating of the gold nanoparticles, any known as a biocompatible polymer can be used, and preferably polyethylene glycol (PEG), dextran, polyvinylpyrrolidone (PVP), albumin, polyvinyl Alcohol, Polyethylenimine, Chitosan, Hyaluronic Acid, Polylactic Glycolic Acid (PLGA), Polylactic Acid, Polylactide, Polyglycolic Acid, Polyamino Acid, Polyacetal, Polytherabolic Acid, Polycarbonate It is a biocompatible polymer selected from the group consisting of ton polyacrylic acid, polymethacrylic acid, polysaccharides, polyketals, polyethers, polyamides, polymaleic anhydrides, polymethylvinyl ethers and copolymers of the above polymers. Preferably PEG or PVP. When PEG is used, PEG-SH which has a -SH functional group with affinity for gold nanoparticles is especially preferable.

CT contrast agents of the present invention exhibit higher HU (hounsfield unit) values than any conventional CT contrast agent. Preferably, the CT contrast agent of the present invention has a 5-50 HU value in vitro , more preferably 10-30 HU value, even more preferably 20-30 HU value, most preferably on a 1 mg / ml basis Represents a 22-26 HU value.

The iodine-based contrast agent, a conventional CT contrast agent, has a concentration of 647 mg / ml for use as a contrast agent, and the HU value is 3000. When calculating the concentration when the HU value of the CT contrast agent of the present invention is 3000, preferably 100-300 mg / ml of gold nanoparticles, more preferably 100-150 mg / ml, most preferably 115-136 mg / ml concentration. Therefore, it can be seen that the contrast agent of the present invention exhibits an excellent contrast enhancement effect at a lower concentration than the conventional iodine-based contrast agent.

CT contrast agents of the present invention comprising gold nanoparticles have hepatic or splenic specificity. As used herein, the term "specificity" refers to a phenomenon in which the CT contrast agent of the present invention is accumulated (or targeted) in a relatively large amount relative to a specific organ or tissue in vivo. The CT contrast agent of the present invention is specifically accumulated in normal liver or spleen tissue, and is very useful for diagnosing a disease or condition with abnormalities in the liver or spleen (eg, abnormal growth of cells, that is, formation of tumor cells). Can be.

As illustrated in the examples below, the CT contrast agents of the present invention do not accumulate (or target) in the kidneys, lungs, and heart, but accumulate in normal liver and spleen.

According to a more preferred embodiment of the invention, the CT contrast agent of the invention scales to normal liver tissue but not to liver cancer tissue. Therefore, in the CT image, normal liver tissue appears bright but liver cancer tissue appears dark, so that liver cancer tissue can be easily diagnosed.

In the field of CT, no liver specific contrast agent has been reported so far, and thus the CT contrast agent of the present invention may be referred to as the first liver specific contrast agent.

According to a preferred embodiment of the present invention, the CT contrast agent of the present invention, when injected in vivo, contrasts at least twice the HU value for liver cancer tissues as compared to HU values of normal liver tissues within 10-90 minutes after injection. Demonstrate their ability The difference in HU values of 2 times is a difference that can sufficiently distinguish between normal liver tissue and liver cancer tissue.

According to a preferred embodiment of the invention, the gold nanoparticles or polymer coating-gold nanoparticles have a size of 1-500, more preferably 10-100 nm. If the size of the gold nanoparticles is less than 1 nm, there is a problem that the discharge through the kidney is too fast to obtain a suitable imaging time.

The targeting ligand may be additionally bound to the gold nanoparticles used in the CT contrast agent of the present invention. Examples of the targeting ligand include, but are not limited to, aptamers, proteins, peptides, antibodies, and sugars. Such targeting ligand may be directly bound to the gold nanoparticles, but may also be bound to a biocompatible polymer coated on the gold nanoparticles.

According to another aspect of the present invention, the present invention provides a CT contrast agent for cancer imaging comprising the computed tomography (CT) contrast agent of the present invention described above.

Since the CT contrast agent for cancer imaging of this invention contains the CT contrast agent of this invention mentioned above, the content common to both is abbreviate | omitted the description in order to avoid excessive complexity of this specification.

CT imaging agents for cancer imaging of the present invention can be applied to CT analysis of all cancer tissues. For example, it can be used for CT analysis of liver cancer, stomach cancer, lung cancer, breast cancer, ovarian cancer, bronchial cancer, nasopharyngeal cancer, laryngeal cancer, pancreatic cancer, bladder cancer, colon cancer, cervical cancer, brain cancer and prostate cancer. In particular, the CT contrast agent for cancer imaging of the present invention can be applied to solid cancer.

The mechanism by which the CT contrast agent for cancer imaging of the present invention contrasts normal and cancerous tissues is as follows: For example, PVP-coated Au NPs are uptake by a reticulorendothelial system (RES) in normal tissues, such as the liver or spleen. do. However, since cancer tissues (eg, liver cancer tissues) have RES disrupted, PVP-coated Au NPs cannot accumulate at cancer tissue sites. As a result, the contrast between contrast-enhanced tissues and cancerous tissues can easily occur because Au NPs will appear darker than normal liver tissue accumulated by RES.

Typically, RES is a problem to be solved in MRI or CT imaging. However, in the present invention, by using the RES mechanism, the innovative proposal can be made to contrast cancer tissue and normal tissue by Au NPs.

According to a preferred embodiment of the present invention, the CT contrast agent for cancer imaging of the present invention is a CT contrast agent for liver cancer imaging.

In summary, the features and advantages of the present invention are as follows:

(Iii) CT contrast agents of the present invention may exhibit contrast enhancement, such as iodine contrast agents, in a relatively small amount compared to conventional iodine contrast agents.

(Ii) Au NPs have a much higher molecular weight than iodine contrast agents, so they are released slowly, resulting in longer imaging times than iodine contrast agents, which in turn greatly increases the utility of Au NPs as CT contrast agents.

(Iii) The CT contrast agent of the present invention exhibits contrast in a relatively short time when applied in vivo.

(Iii) The CT contrast agent of the present invention shows specificity for the liver or spleen.

(Iii) The CT contrast agent of the present invention can be particularly useful for diagnosing liver cancer.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. .

Example

Experimental material

Poly (vinylpyrrolidone, poly (vinylpyrrolidone: PVP, Mw = 55,000), sodium citrate tribasic dehydrate, and hydrogen tetrachloroaurate (HAuCl ₄ ) were obtained from Sigma (Sigma Chemical Co., St. Louis). , Polyethylene glycol (Mw = 5,000) -SH was purchased from Nectar (Huntsville, AL) Lewis lung cancer cell line (LL) and rat hepatoma tumor cell line (N1S1). ) Were purchased from the American Type Culture Collection, Manassas, VA, USA The cells were obtained from DMEM containing 10% heat-inactivated fetal bovine serum (Gibco, Gramd Island, NY) and 1% penicillin and streptomycin. Incubated in Dulbecco's Modified Eagle Medium or IMDM (Iscoves' Modified Dulbecco's Medium; Gibco, Gramd Island, NY) at 37 ° C. under 5% CO ₂ , 95% air dust.

Experiment method

Synthesis of Au NPs (Gold Nanoparticles)

Au NPs were prepared by reducing HAuCl ₄ using sodium citrate as reducing agent. In addition, the size of the gold nanoparticles could be adjusted by varying the amount of citrate added. In other words, as the amount of citrate increased, the nucleation was increased, so the size of the gold nanoparticles decreased.

The synthesis process of Au NPs is represented by the following scheme 1.

The size change of gold nanoparticles according to the amount of citrate added is summarized in Table 1.

Citrate addition amount (ml) Gold nanoparticle size (nm) 1.0 16 0.75 25 0.5 41 0.3 72 0.21 98 0.16 147

Synthesis of PVP-Coated Au NPs

0.01 wt% HAuCl ₄ was added to a 250 ml round flask equipped with a condenser and heated. At the time of boiling of the solution, 0.8 ml or 4 ml of 1 wt% sodium citrate and 6.8 mg of PVP polymer were added rapidly. Over time, the color of the reaction solution changes from blue to red, indicating that Au NPs are formed. The PVP-coated Au NPs thus prepared were precipitated by centrifugation (Combi-514R, Hanil, Korea) for 30 minutes at 13000 rpm to finally collect 120 mg of PVP-coated Au NPs.

Synthesis of PEG-Coated Au NPs

0.01 wt% HAuCl ₄ was added to a 250 ml round flask equipped with a condenser and heated. At the time of solution boiling, 2 ml of 1 wt% sodium citrate was added quickly. Over time, the color of the reaction solution changes from blue to red, indicating that Au NPs are formed. Subsequently, after additional heat for 10 minutes, the temperature of the reaction solution was reduced to room temperature. Then, 1 mg of PEG-SH was added to the reaction solution, and stirred for 1 hour to obtain Au NPs coated with PEG. Since the -SH functional group in PEG has a high affinity for Au, PEG-SH is easily coated on Au NPs. The PEG-coated Au NPs thus prepared were precipitated by centrifugation (Combi-514R, Hanil, Korea) for 30 minutes at 13000 rpm to finally collect 120 mg of PEG-coated Au NPs.

Characterization of Au NPs

(a) Size analysis of Au NPs

The size of Au NPs was determined using ELS (Electrophoretic Light Scattering Spectrophotometer, Otsuka Electronics Co., Ltd, Japan).

(b) Types of Au NPs

The shape of Au NPs was confirmed using SEM (scanning electron microscopy, S4700, Hitachi Ltd, Japan). An appropriate amount of Au NPs suspension was dropped onto a silicon wafer, then dried and Pt coated for 40 seconds through a sputter, transferred to a holder of the SEM and a SEM image was obtained.

In Vitro Hound Value (HU) Measurement of PVP-Coated Au NPs

HU of PVP-coated AU NPs was measured by 64-channel computed tomography (CT).

Preparation of the animal model

(a) Rat Hepatoma Model

Sprague-Dawley rats (male, average body weight 300 g) were anesthetized by intraperitoneal injection of 0.5 ml ketamine and 0.05 ml rompun. Subsequently, after the rat was opened, 0.1 ml of rat hepatoma cell line N1S1 was inoculated into the liver in 10 ⁶ counts. There was no death of rats during and after surgery. The breeding process of rats was carried out according to the Institutional guidelines of the Institutional Animal Care and Use Committee (IACUC).

(b) LLC tumor mouse model

Lewis lung cancer (LLC) cells were injected into the back of C57BL / 6 mice (average body weight 25 g) by subcutaneous injection in 10 ⁶ counts. The breeding process of mice was carried out according to the Institutional guidelines of the Institutional Animal Care and Use Committee (IACUC).

Computed Tomography

400 μl of PVP-coated Au NPs (100 μg / μl) or PEG-coated Au NPs (100 μg / μl) were injected through the tail vein of the rat hepatoma model, and CT images were taken through 64 channels at various times after intravenous injection. Was taken.

150 μl of Au NPs (100 μg / μl) coated with PEG was injected through the tail vein into the LLC tumor mouse model. CT images were taken through 64 channels at various times after intravenous injection.

Silver dyed

Liver cancer tissue, normal liver tissue, kidney tissue, lung tissue, kidney tissue, and intestinal tissue were extracted, fixed on slide glass, and oxidized through 10% chromic acid at room temperature for 5 minutes. Washed three times with distilled water, reduced for 2 minutes with 1% sodium bisulfate, and washed for 5-10 minutes with distilled water. Subsequently, the slide was reacted with (methenamine silver working solution) (40-50 minutes at 60 ℃), and washed with distilled water at room temperature. Then, treated with 2% gold chloride for 3-5 minutes and washed with distilled water. Then treated with 2% sodium thiosulfate for 2-4 minutes and washed with tap water for 10 minutes. Counter staining was performed with 0.4% light green for 10 seconds. Finally, the mounting medium was dehydrated.

Experiment result

SEM image of 40 nm PVP-coated Au NPs

SEM and ELS analysis were performed on PVP-coated Au NPs coated with biocompatible polymer PVP as the first contrast agent for cancer imaging and prepared by adding 0.4 ml or 0.8 ml of citrate as reducing agent. As can be seen in Figures 1a and 1b, when the PVP-coated Au NPs of the present invention was prepared using 0.4 ml and 0.8 ml of citrate, Au having a spherical shape of about 20 nm and about 40 nm, respectively It can be seen that it is NPs.

HU measurement of PVP-coated Au NPs

Was collected for the 40 nm Au NPs size of PVP- coating prepared in this example is 34 mg / ml concentration, it was measured Hounsfield value (HU) in vitro for it (Fig. 2).

In FIG. 2, the bright white spot at the bottom left of the right panel is a CT image of a tube containing 34 mg / ml PVP-coated NPs, showing an 800 HU value in vitro . In the case of iodine-based contrast agent, which is currently used CT contrast agent, the concentration to be used as contrast agent is 647 mg / ml, and the HU value is 3000. Based on the above in vitro results, the concentration of Au NPs at HU of 3000 was calculated to be 127.5 mg / ml, which shows excellent contrast enhancement effect with less farming than iodine-based contrast agent.

In vivo mouse model data

In order to determine how Au NPs coated with the biocompatible polymer PVP and having a size of 40 nm exhibited a contrast agent in vivo , an experiment was conducted using a mouse model. Based on the results of the in vitro experiment, PVP-coated Au NPs at a concentration of 68 mg / ml were prepared, and 200 μl was injected into the C57Bl / 6 mouse through the tail vein (intravenous injection iv), followed by CT scan. 3A and 3B are CT images of liver and spleen of mice at various time zones after injection of 40 nm PVP-coated Au NPs. The numbers above the image represent the time and Hounsfield values of the CT taken after the injection. "Pre" is a CT image of the liver and spleen of mice before injecting PVP-coated Au NPs.

As shown in Figures 3a and 3b, immediately after injection of 40 nm PVP-coated Au NPs it was observed that the liver and spleen of the mouse rapidly whitened. Thus, it can be seen that PVP-coated Au NPs were uptaken by the reticulorendothelial system (RES) in the liver or spleen.

The experimental results show that the PVP-coated Au NPs of the present invention can be used as a liver specific contrast agent. Because liver cancer tissue has RES destroyed, PVP-coated Au NPs cannot accumulate in the liver cancer tissue region. This is because contrast enhancement occurs because Au NPs will appear darker than normal liver tissue accumulated by RES, and liver cancer tissue can be easily distinguished from normal liver tissue.

Typically, RES is a problem to be solved in MRI or CT imaging. However, the results of this experiment suggest that the RES mechanism can be used to image liver cancer tissue and normal liver tissue by Au NPs.

On the other hand, as can be seen in Figures 3a and 3b, even after 5 days after injection, the liver and spleen show a white CT image, 40 nm PVP-coated Au NPs are not erased from the liver and spleen even after 5 days Can be confirmed. This result may be limited to the application of the PVP-coated Au NPs of the present invention as a contrast agent. Therefore, in order to overcome this problem, PVP-coated Au NPs having a size of about 20 nm smaller than 40 nm were fabricated, and further experiments were performed. The experiment was conducted.

CT image of rat hepatoma using 20 nm PVP-coated Au NPs

The basic concept of this experiment is as follows: PVP-coated Au NPs as CT contrast agent are injected into rat hepatoma model with liver cancer through the tail vein of the rat. PVP-coated Au NPs cannot accumulate in liver cancer tissues where RES is destroyed, whereas Au NPs accumulate in RES in normal tissues. When X-rays are irradiated, normal liver tissue accumulating Au NPs is weakened as X-rays pass through Au NPs, which makes them appear white on CT images. X-rays are weak in liver cancer tissues without Au NPs accumulated. Because it passes through without being supported, it appears darker than normal liver tissue on the CT image. Therefore, liver cancer tissues can be easily distinguished by using PVP-coated Au NPs of the present invention as liver-specific contrast agents.

Rat hepatoma models were tested to see whether PVP-coated Au NPs could substantially differentiate liver and liver cancer tissues by acting as a liver-specific contrast agent. In addition, PVP-coated Au NPs with a size of 20 nm smaller than 40 nm were used as contrast medium for adequate time emission (about 2-3 days).

FIG. 4 is a CT image taken at various time slots after intravenous injection (i.v) in a rat hepatoma model using 20 nm-sized PVP-coated Au NPs as a contrast agent. The number at the top right of the image represents the HU value for each time zone, the number on the left represents the HU value of liver cancer tissue, and the number on the right represents the HU value of normal liver tissue. White arrows also indicate liver cancer tissue. Panel (a) is a CT image of liver before Au NPs were used as contrast medium, and it is not possible to clearly distinguish between liver cancer tissue and normal liver tissue. However, after the injection of PVP-coated Au NPs, the Au NPs accumulate in normal liver tissues, and they appear brighter on CT images, while liver cancer tissues without Au NPs accumulate appear darker, making it easier to distinguish between liver cancer tissues and normal liver tissues. can do. However, as can be seen in panel (g), in the CT image of liver 7 days after injection, the HU value of normal tissue decreased to 110, a little below the maximum HU value of 125, ie slightly discharged but still Au NPs remained in normal liver tissue for a long time. It can be seen that it is not discharged while Therefore, in order to reduce the efficient emission of Au NPs and uptake of RES, experiments were carried out by coating PEG with Au NPs, a biocompatible polymer having stealth function stronger than PVP.

Biodistribution of PVP-Coated Au NPs

It was confirmed by silver staining to see what tissue accumulated 20 nm PVP-coated Au NPs injected in vivo (Fig. 5). If Au NPs are present in any tissue, Au NPs will act as a nucleation site of silver during silver staining and the tissue will look black as the particles grow.

Panel a of FIG. 5 shows an experimental result of normal liver tissue, in which Au NPs are present in cooper cells and RES cells. Panel b shows the presence of Au NPs as a result of the experiment on the spleen. However, it was confirmed by silver staining that Au NPs were not present in liver cancer tissue, kidney, lung and heart.

Size Distribution Analysis of PEG-Coated Au NPs

The size of Au NPs before and after coating with PEG was measured by ELS. As can be seen in Figure 6, the size of the Au NP before the PEG coating is about 20-25 nm, the size after the coating is about 40-45 nm, it was confirmed that the well coated with PEG.

Hepatoma CT Images by PEG-Coated Au NPs

Experimental experiments were conducted using a rat hepatoma model to show how the Au NPs (size 20-25 nm before coating and size 40-45 nm after coating) coated with PEG, a stealth polymer stronger than PVP, appeared as contrast agents. FIG. 7 is a CT image of various time periods after injection of PEG-coated Au NPs into rats in the same amount as the PVP-coated Au NPs experiment.

In FIG. 7, the number in the upper right of the image represents the HU value for each time zone, the left number represents the HU value of liver cancer tissue, and the right number represents the HU value of normal liver tissue. White arrows also indicate liver cancer tissue. Panel (a) is a CT image of liver before Au NPs were used as a contrast agent, and there is no clear distinction between liver cancer and normal liver tissue. On the other hand, PGE-coated Au NPs were reduced uptake by RES of normal liver tissues compared to PVP-coated Au NPs (maximum HU value of PVP-coated Au NPs in normal liver tissues 125, PEG-coated Au NPs The maximum HU value 108), accumulated in normal liver tissue to distinguish it from liver cancer tissue, showed an contrast enhancement effect.

In addition, PEG-coated Au NPs were released through the kidneys for the first time after injection compared to PVP-coated Au NPs, but as shown in panel (h), HU of normal liver tissue 108 shows PEG-coated Au NPs accumulated in normal liver tissue.

Cancer CT Images by PEG-Coated Au NPs

Whether PEG-coated Au NPs (size 20-25 nm before coating, size 40-45 nm after coating) can be used to image common solid cancers, not liver cancer, the study was performed using a mouse model of tumor-induced LLS tumors .

In Figure 8, the upper right number of the image represents the HU value according to time zone, the left number represents the HU value of the muscle tissue, the right number represents the solid cancer HU value. White arrows also indicate solid cancer on the dorsal side of the mouse.

As shown in Figure 8, 10 minutes after the PEG-coated Au NPs injection HU value of the solid cancer increases to 71, it can be seen that the PEG-coated Au NPs accumulate in the solid cancer. PEG-coated Au NPs of the present invention are coated with a biocompatible polymer that avoids RES and can accumulate around cancer by an enhanced permeation and retention (EPR) effect. When PEG-coated Au NPs accumulate around solid cancers, X-rays are absorbed by the Au NPs and weaken and appear white on the CT image.

As described in detail above, the CT contrast agent of the present invention may exhibit contrast enhancement, such as iodine contrast agent, in a relatively small amount compared to conventional iodine contrast agents, and Au NPs have a much higher molecular weight than iodine contrast agents. This results in longer imaging times than iodine contrast agents, which in turn greatly increases the utility of Au NPs as CT contrast agents. In addition, when applied in vivo, the CT contrast agent of the present invention exhibits contrast in a relatively short time, and the CT contrast agent of the present invention exhibits specificity for the liver or spleen. The CT contrast agent of the present invention can be particularly useful for diagnosing liver cancer.

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that the specific technology is merely a preferred embodiment, and the scope of the present invention is not limited thereto. Thus, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

Claims (11)

Computed tomography (CT) contrast agent comprising gold nanoparticles and having liver or spleen specificity. The method of claim 1, wherein the gold nanoparticles are polyethylene glycol (PEG), dextran, albumin, polyvinyl alcohol, polyethyleneimine, polylactic glycolic acid (PLGA), polylactic acid, polylactide, polyglycolic Acids, polyamino acids, polyacetals, polyacetalates, polycarbonates, polycarbolactone polyacrylic acids, polymethacrylic acids, polysaccharides, polyketals, polyethers, polyamides, polymaleic anhydrides, polymethylvinyl CT contrast agent characterized in that the surface-coated with a biocompatible polymer selected from the group consisting of ethers and copolymers of the polymer. The CT contrast agent according to claim 1, wherein the gold nanoparticles are surface-coated with polyethylene glycol (PEG). The CT contrast agent according to claim 1, wherein the CT contrast agent has a value of 5-50 hunsfield units (HU) in vitro on a 1 mg / ml basis. The CT contrast agent according to claim 4, wherein the CT contrast agent has a value of 20-30 HU (hounsfield unit) in vitro based on 1 mg / ml. delete The CT contrast agent according to claim 1, wherein the CT contrast agent is accumulated in normal liver tissue but not in liver cancer tissue. The CT contrast agent according to claim 1, wherein the CT contrast agent exhibits at least twice the HU value for liver cancer tissues as compared to the HU values of normal liver tissues within 10 to 90 minutes after injection. . The CT contrast agent according to claim 1, wherein the gold nanoparticles have a size of 1-500 nm. A CT contrast agent for cancer imaging, comprising the computed tomography (CT) contrast agent of any one of claims 1 to 5 and 7 to 9. 11. The CT contrast agent for cancer imaging according to claim 10, wherein the cancer is liver cancer.

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