JP2005145819A - Fluorescent contrast medium and method for in vitro fluorescence imaging - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a fluorescent contrast medium for diagnosing tumor or cancer having low toxicity and excellent water solubility and emitting fluorescence transmittable through biological tissues and providing specific imaging of the tumor, cancer and/or blood vessels, and to provide a method for in vitro fluorescence imaging using the fluorescent contrast medium. <P>SOLUTION: The fluorescent contrast medium for diagnosing the tumor or cancer comprises at least one kind of penta- or heptamethine cyanine dye having 3H-pyrrolopyridine with at least 2 acid groups in the molecule as at least one of parent nuclei. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

  The present invention relates to a near-infrared fluorescent contrast agent suitable for diagnosis of a tumor or cancer and a diagnosis of a tumor or cancer by an in vitro fluorescence contrast method using the near-infrared fluorescent contrast agent.

  When treating a disease, it is required to accurately, quickly and easily detect morphological changes caused in the living body by the disease at an early stage of the disease.

  Particularly in the case of treating cancer, the determination of the location and size of the initial small lesion is essential for early treatment. Known methods for this purpose include diagnostic imaging such as endoscopic biopsy, X-ray imaging, CT, MRI and ultrasound imaging.

  Biopsy is effective as a definitive diagnosis because it allows direct observation of the lesion, but at the same time it forces pain and distress on the subject. X-ray imaging and MRI expose the subject to radiation and magnetic fields that are harmful if excessive, and the exposure time increases in proportion to the tracking time when trying to track the time course. Equipment and devices are also large, and a great deal of labor and cost is required for their installation and maintenance.

  Near-infrared fluorescence contrast imaging is a diagnostic method that has been attracting attention in recent years as a diagnosis that can be miniaturized, reduce driving effort, and simplify use.

  It is said that near-infrared light can easily pass through a biological tissue composed of 70% or more of moisture, and can test and diagnose a thickness of 10 cm to 20 cm. Therefore, it has come to be developed as a diagnostic technique for near infrared CT while attracting attention in the field of clinical medicine.

  In this method, a compound that emits fluorescence when irradiated with excitation light having a near-infrared wavelength of 700 nm to 1000 nm as a contrast agent is irradiated into the living body and has a near-infrared wavelength from outside the body. The lesion is determined by irradiating excitation light and detecting fluorescence emitted from a compound administered in the body, that is, a so-called fluorescent contrast medium.

  As such fluorescent contrast agents, for example, porphyrin compounds and hematoporphyrin compounds that accumulate in tumors are known. These compounds are excited by near-infrared light irradiation to generate triplet oxygen that can oxidize the living body of the lesion, killing cells of the lesion such as cancer, and treating them. Although possible, there is a risk of destroying tissues other than the lesion.

  On the other hand, contrast methods using known fluorescent dyes such as fluorescein and fluoresamine are disclosed in US Pat. No. 4,945,239, but these fluorescent dyes are blue to green with very low light transmission in living organisms. Because it emits light, it cannot sufficiently detect lesions in the inner part of the body.

  Cyanine dyes that emit fluorescence in the near infrared region are expected as fluorescent contrast agents, and various cyanine compounds have been studied. Since the reporting of fluorescent contrast agents of cyanine compounds, the technology using various peripheral cyanine compounds as contrast agents has been disclosed in the following Patent Documents 1, 2, and 3 in order to change them to compounds with high hydrophilicity, molar extinction coefficient, and quantum yield. Is disclosed.

However, together with the ability to distinguish normal tissue from diseased tissue (contrast power / affinity with diseased tissue), it is necessary to be completely decomposed and harmless in the living body after imaging from the living body, or to be completely discharged (Non-accumulative), no safe contrast agent has been found.
JP 2000-95758 A JP-T-2002-526458 JP 2003-160558 A

  An object of the present invention is to diagnose a tumor or cancer that emits fluorescence that has low water toxicity, excellent water solubility, and can be transmitted through living tissue, and enables specific imaging of tumors, cancer and / or blood vessels. It is an object of the present invention to provide a fluorescent contrast agent for use in the present invention and to provide an extracorporeal fluorescent contrast method using the fluorescent contrast agent.

The object of the present invention is achieved by the following configurations.
(Claim 1)
A fluorescent contrast agent for diagnosing a tumor or cancer comprising at least one penta- or heptamethine cyanine dye having 3H-pyrrolopyridine having at least two acid groups in the molecule as at least one mother nucleus.
(Claim 2)
2. The fluorescence for diagnosis of tumor or cancer according to claim 1, wherein the penta- or heptamethine cyanine dye is at least one kind represented by the following general formula (1), (2) or (3): Contrast agent.

[Wherein, R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ each represents an alkyl group, and the alkyl group represented by R ₁ and R ₄ may be substituted. Z ₁ and Z ₂ each represent a nonmetallic atom group necessary for forming a pyridine ring. Y ₁ represents a pyridine ring containing the following bond (1) in the ring, Y ₂ represents a dihydropyridine ring containing the following bond (2) in the ring, L represents a methine group, and X ⁻ represents an anion. m represents an integer of 3 or 4, and n represents an integer of 1 or 2. However, when m is 3, R ₁ and R ₄ are preferably substituted alkyl groups, and n is 1 when the dye forms an inner salt. ]

[R ₇ and R ₈ in bond (1) or bond (2) have the same meanings as R ₁ and R _{4 in} formula (1), (2) or (3). ]
(Claim 3)
The tumor according to claim 1 or 2, wherein at least two acid groups of the molecule of the penta or heptamethine cyanine dye are groups selected from a sulfonic acid group, a carboxylic acid group, and a phosphoric acid group. Fluorescent contrast agent for cancer diagnosis.
(Claim 4)
The at least 2 acid group which the molecule | numerator of the said penta or heptamethine cyanine dye has the group selected from a sulfonic acid group or a carboxylic acid group, and the sum total of an acid group is 2 or more, The fluorescent contrast agent for diagnosis of the tumor or cancer of any one of -3.
(Claim 5)
The tumor or cancer according to any one of claims 1 to 4, wherein at least two of the acid groups of the penta- or heptamethine cyanine dye molecule are sulfonic acid groups. Fluorescent contrast agent for diagnosis.
(Claim 6)
The fluorescence for diagnosis of tumor or cancer according to any one of claims 1 to 5, wherein at least two acid groups of the penta- or heptamethine cyanine dye molecule are sodium salts. Contrast agent.
(Claim 7)
The fluorescence imaging for diagnosis of a tumor or cancer according to any one of claims 1 to 6, wherein the tumor or cancer is at least one selected from the group consisting of sarcomas and melanomas. Agent.
(Claim 8)
Diagnosing a tumor or cancer by introducing the fluorescent contrast agent according to any one of claims 1 to 7 into a living body, irradiating the living body with excitation light, and detecting fluorescence from the contrast agent. An extracorporeal fluorescence imaging method characterized by the above.

  The near-infrared fluorescent contrast agent according to the present invention is highly permeable to biological tissues because the fluorescence excited and emitted by near-infrared light is excellent in the permeability of biological tissues. Therefore, it is possible to provide a fluorescent contrast agent and an extracorporeal fluorescent imaging method for diagnosing a tumor or cancer with high safety.

  Next, the best mode for carrying out the present invention will be described, but the present invention is not limited thereto.

  The above object of the present invention is achieved by a fluorescent contrast agent containing at least one penta- or heptamethine cyanine dye having 3H-pyrrolopyridine having at least two acid groups in the molecule as at least one mother nucleus. Is.

  Among the penta- or heptamethine cyanine dyes, preferred for use as a fluorescent contrast agent is at least one kind represented by the general formula (1), (2) or (3).

In the formula, R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ each represents an alkyl group, and the alkyl group represented by R ₁ and R ₄ may be substituted. Z ₁ and Z ₂ each represent a nonmetallic atom group necessary for forming a pyridine ring.

Y ₁ represents a pyridine ring containing the following bond (1) in the ring, Y ₂ represents a dihydropyridine ring containing the following bond (2) in the ring, L represents a methine group, and X ⁻ represents an anion. m represents an integer of 3 or 4, and n represents an integer of 1 or 2. However, when m is 3, R ₁ and R ₄ are preferably substituted alkyl groups, and n is 1 when the dye forms an inner salt.

In the bond (1) and the bond (2), R ₇ and R ₈ have the same meanings as R ₁ and R _{4 in the} general formula (1), (2) or (3).

  The compound represented by the general formula (1), (2) or (3) has at least two acid groups in the molecule, and examples of the acid groups include sulfonic acid groups, carboxylic acid groups, phosphoric acid groups, Examples thereof include phosphonic acid groups, and each of these acid groups includes a salt thereof. Examples of the salt include alkali metal salts such as sodium and potassium, and organic ammonium salts such as ammonium, triethylammonium, and pyridinium.

  Further, in the penta- or heptamethine cyanine dye molecule used as the fluorescent contrast agent according to the present invention, particularly in the cyanine dye molecule represented by the general formula (1), (2) or (3), The at least two acid groups in the group are preferably acid groups selected from sulfonic acid groups, carboxylic acid groups, phosphoric acid groups and phosphonic acid groups.

  However, more preferably, in the general formula (1), (2) or (3), the acid group in the molecule contains a group selected from a sulfonic acid group or a carboxylic acid group, and This is a case where the total is 2 or more.

  More preferably, in the general formula (1), (2) or (3), at least two of the acid groups in the molecule are sulfonic acid groups.

  In the cyanine dye molecule represented by the general formula (1), (2) or (3), at least two acid groups in the molecule preferably form a salt, and the salt is a sodium salt. Is preferred.

The compound represented by the general formula (1), (2) or (3) may contain a —CH ₂ CH ₂ OR group. R represents a hydrogen atom or an alkyl group. The alkyl group represented by R is preferably a lower alkyl group having 4 or less carbon atoms.

Examples of the substituent containing a —CH ₂ CH ₂ OR group include a hydroxyethyl group, a hydroxyethoxyethyl group, a methoxyethoxyethyl group, a hydroxyethylcarbamoylmethyl group, a hydroxyethoxyethylcarbamoylmethyl group, and N, N-dihydroxyethylcarbamoylmethyl. Group, hydroxyethylsulfamoylethyl group, methoxyethoxyethoxycarbonylmethyl group and the like.

Z ₁ , Z ₂ , Y _1, and Y ₂ may have various substituents in addition to the acid group and —CH ₂ CH ₂ OR group described above. Groups, alkyl groups (eg, methyl, ethyl groups, etc.), alkoxy groups (eg, methoxy, ethoxy groups, etc.), aryl groups (eg, phenyl groups), halogen atoms (eg, fluorine, chlorine, bromine atoms, etc.), etc. It is done. The alkyl group represented by R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ is preferably a lower alkyl group having 1 to 8 carbon atoms (for example, methyl, ethyl, propyl, i-propyl, butyl). Group), and may have a substituent other than the above-described acid substituent or —CH ₂ CH ₂ OR group. The methine group represented by L may also have a substituent, and the substituent is a substituted or unsubstituted lower alkyl group having 1 to 5 carbon atoms (for example, methyl, ethyl, 3-hydroxypropyl, 2-sulfoethyl). Group), a halogen atom (eg, fluorine, chlorine, bromine atom, etc.), an aryl group (eg, phenyl group), an alkoxy group (eg, methoxy, ethoxy group, etc.) and the like. Further, substituents of the methine group may be bonded to each other to form a 6-membered ring containing three methine groups (for example, 4,4,4-dimethylcyclohexane ring).

The anion represented by X ⁻ is not particularly limited, but preferably a halogen ion, p-toluenesulfonic acid ion, ethylsulfuric acid ion, and the like.

  The cyanine dyes represented by the general formula (1), (2) or (3) of the present invention are m = 3 pentamethine cyanine dyes and m = 4 heptamethine cyanine dyes, preferably m = 4. Is a heptamethine cyanine dye.

  The dye represented by the general formula (1), (2) or (3) of the present invention is described in J. Chem. Soc., P. 189 (1933), US Patent. No. 2,895,955, Japanese Patent Laid-Open No. Sho 62-123454, Japanese Patent No. 2961549, and the like.

  Examples of the compound used as the mother nucleus of the dye represented by the general formula (1), (2) or (3) of the present invention include the following compounds A, B and C.

  Compound (A) is described in J. Org. Chem. Soc. 3202 (1959) or the method described in British Patent 870,753. Compound (B) is described in J. Org. Chem. Soc. , 584 (1961). Compound (C) can be synthesized by the method described in British Patent 841,588.

  These mother nuclei can be used for quaternization, sulfonation and the like as required. Chem. Soc. 3202 (1959) and J.A. Chem. Soc. , 584 (1961), N-alkyl-N-pyridylhydrazine was synthesized, cyclized through hydrazone, and treated with acid as necessary to give 1-alkyl-substituted 3H-pyrrolopyridine. Derivatives can also be obtained and used as starting materials.

  The compound of the present invention can be easily obtained by reacting an appropriate methine chain supplier using these quaternized and optionally sulfonated mother nucleus compounds.

  If glutaconaldehyde dianil hydrochloride is used as a methine chain supplier, a heptamethine dye can be obtained.

  The compounds of the present invention are listed below, but the present invention is not limited thereto.

  It is important that the fluorescent contrast agent to be used in the living body is not accumulated in the body but is quickly discharged out of the body, and is basically a water-soluble condition. As a means for improving water solubility, it is preferable to have an anionic carboxylic acid or sulfonic acid in the molecule, and these carboxylic acid and sulfonic acid preferably form a salt, such as sodium or potassium. And organic ammonium salts such as ammonium, triethylammonium and pyridinium.

  The near-infrared fluorescent contrast agent of the present invention preferably contains two or more sulfonic acid groups among the above acid groups in the above compound, and among them, the introduction of three or more sulfonic acid groups. As a result, water solubility is remarkably improved. In order to obtain excellent water solubility, the number of sulfonic acid groups is preferably 4 or more. In order to facilitate the synthesis, the number of sulfonic acid groups is 10 or less, preferably 8 or less.

  A measure of water solubility can be determined by measuring the partition coefficient of each compound, for example, by measuring the partition coefficient in a two-phase system of an aliphatic alcohol such as butanol and water.

  For example, by introducing 3 or more sulfonic acid groups, the distribution coefficient logPo / w of n-butanol / water becomes −1.00 or less.

  A method for determining water solubility in a living body is that it dissolves in physiological saline and does not precipitate or precipitate after a lapse of time at 36 ° C. The salt that is tolerated when administered in vivo may be any salt that forms a non-toxic salt with the compound of general formula (1), (2) or (3). Examples thereof include alkali metal salts such as sodium salts and potassium salts; alkaline earth metal salts such as magnesium salts and calcium salts, tryptophan, methionine, lysine, phenylalanine, leucine, isoleucine, valine, threonine and arginine. And amino acid salts such as salts thereof. Particularly preferred are sodium salts that have low toxicity in vivo.

  The compound according to the present invention to be contained in the fluorescent contrast agent of the present invention exhibits absorption and fluorescence in the near infrared light region of 700 to 1300 nm, particularly about 700 to 900 nm, and has a molar extinction coefficient of 100,000 or more.

  The fluorescent contrast agent of the present invention is an in vitro fluorescent contrast imaging method for diagnosing a tumor or cancer by introducing a fluorescent contrast agent into a living body and then irradiating the living body with excitation light and detecting fluorescence from the contrast agent. It is used.

  Tumors or cancers diagnosed by extracorporeal fluorescence imaging include brain, breast, breast, prostate, colon, lung, liver, pancreas, stomach, lymph nodes, uterine body, cervix, and upper and lower limbs. It is selected from the group consisting of sarcomas and melanomas.

  Specifically, the fluorescent contrast agent used in the extracorporeal fluorescence contrast method according to the present invention is prepared by using at least one of the penta- or heptamethine cyanine dyes according to the present invention such as distilled water for injection, physiological saline, Ringer's solution, or the like. It is suspended or dissolved in a solvent, and pharmacologically acceptable additives such as carriers, excipients and the like may be added as necessary. These additives include substances such as pharmacologically acceptable electrolytes, buffers, detergents, and substances for adjusting osmotic pressure to improve stability and solubility (eg, cyclodextrins, liposomes, etc.). . Various additives commonly used in related fields can be used. The fluorescent contrast medium according to the present invention is preferably manufactured through sterilization treatment for medical use. The fluorescent contrast agent can be administered to the living body site by injection, injection or application, intravascular (vein, artery), oral, intraperitoneal, transdermal, subcutaneous, intravesical or intrabronchial administration. Preferably, the contrast agent is administered into the blood vessel in the form of a solution, emulsion or suspension.

  The dose of the fluorescent contrast agent of the present invention is not particularly limited as long as the site to be finally diagnosed can be detected by the dose, and the type, age, and weight of the compound emitting near-infrared fluorescence to be used And appropriately adjusted according to the target organ to be administered. Typically, the dosage is 0.1-100 mg / kg body weight, preferably 0.5-20 mg / kg body weight as the amount of the compound. The contrast agent of the present invention can be appropriately used for various animals other than humans. The form, route and dose of administration are appropriately determined according to the weight and condition of the subject animal.

  In the present invention, when the penta or heptamethine cyanine dye is accumulated in the tumor tissue, the tumor tissue can be specifically imaged using the fluorescent contrast agent of the present invention. Furthermore, a series of the compounds can stay in the blood vessels for a long time and is expected to function well as an angiographic agent.

  The extracorporeal fluorescence imaging method of the present invention is characterized by the use of the fluorescent contrast agent of the present invention. This method is performed according to a known method, and parameters such as the excitation wavelength and the fluorescence wavelength to be detected are appropriately determined so as to be optimal depending on the type of the near-infrared fluorescent contrast agent to be administered and the administration target. The The time taken from the administration of the fluorescent contrast agent of the present invention to the measurement object to the start of measurement by the fluorescence imaging method of the present invention varies depending on the type of near-infrared fluorescent contrast agent used and the administration object. For example, for tumor imaging purposes, the disappearance time is considered to be approximately 4-120 hours after administration. If the disappearance time is too short, the fluorescence is too strong to clearly separate the target site from other sites. If the length is too long, the contrast medium may be excreted from the body. When angiography is required, detection is performed immediately after administration or within about 30 minutes.

  In this method, the fluorescent contrast agent is administered to a detection target, and the detection target is exposed to excitation light from an excitation light source. Next, the fluorescence generated by the excitation light from the near-infrared fluorescent contrast agent is detected by a fluorescence detector. The excitation wavelength varies depending on the near-infrared fluorescent contrast agent used, but is not limited as long as the compound emits fluorescence effectively in the near-infrared region. Preferably, near-infrared light having excellent biological permeability is used.

  The wavelength of near-infrared fluorescence to be detected also varies depending on the contrast agent used. Generally speaking, near-infrared fluorescence in the wavelength region of 700-1000 nm, preferably 750-900 nm is detected using excitation light having a wavelength of 600-1000 nm, preferably 700-850 nm. In this case, the excitation light source may be a normal excitation light source such as various lasers (for example, an ion laser, a dye laser, and a semiconductor laser), a halogen light source, a xenon light source, or the like. If necessary, various optical filters can be used to obtain the optimum excitation wavelength. Similarly, fluorescence can be detected by capturing only fluorescence from the fluorescent contrast agent using various optical filters.

  The detected fluorescence is data-processed as fluorescence information and used to generate a recordable fluorescence image. The fluorescence image is generated by irradiating a wide area including the target tissue, detecting fluorescence with a CCD camera, and processing the obtained fluorescence information. Alternatively, an optical CT apparatus, an endoscope, or a fundus camera may be used. The fluorescence imaging method of the present invention can visualize systemic diseases, tumors, blood vessels and the like without damaging the living body.

  EXAMPLES Hereinafter, although an Example demonstrates this invention, this invention is not limited to these.

Example 1
(Synthesis of Exemplified Compound 31)
The compound (B) synthesized according to the above-mentioned literature is quaternized with propane sultone according to a conventional method, and then 5.0 g of this quaternized product is dissolved by heating in 40 ml of acetic acid to give 1.0 g of glutaconaldehyde dianyl hydrochloride. Then, 9 ml of acetic anhydride and 13 ml of pyridine were added, and the mixture was heated and stirred at a bath temperature of 80 to 85 ° C. for 30 minutes. After allowing to cool, isopropyl ether was added and decanted. Further, methynol, acetone and isopropyl ether were added and decanted for purification, and further 100 ml of saturated saline was added for purification to obtain compound (31). [lambda] max (MeOH) 739 nm.

(Synthesis of Compound Example 32)
The compound (A) synthesized according to the above literature was quaternized with propane sultone according to a conventional method, reacted under the same reaction conditions as compound (31), and then purified to obtain compound (32). [lambda] max (MeOH) 731 nm.

(Synthesis of Compound Example 20)
The compound (A) was quaternized with propane sultone and then sulfonated with concentrated sulfuric acid and fuming sulfuric acid. 2.6 g of the sulfonated product was dissolved in 10 ml of N, N-dimethylformamide, 1.0 g of glutaconaldehyde dianil hydrochloride, 2 ml of pyridine and 2 ml of acetic anhydride were added, and the mixture was heated and stirred at a bath temperature of about 100 ° C. for 20 minutes. Isopropyl ether was added to the reaction mixture, followed by decantation and purification to obtain compound (20). [lambda] max (MeOH) 737 nm.

(Synthesis of Compound Example 33)
According to the above literature, 2,3,3-trimethyl-5-iodo-3H-pyrrolo [3,2-b] pyridine was synthesized. This was quaternized with propane sultone according to a conventional method, 0.41 g of the quaternized product was dissolved in 3 ml of acetic acid, 0.19 g of glutaconaldehyde dianyl hydrochloride, 0.7 ml of acetic anhydride and 1 ml of pyridine were added, and the bath temperature was increased. The mixture was heated and stirred at 90 to 105 ° C. for 1 hour. After allowing to cool, 40 ml of ethyl acetate was added to the reaction solution and stirred, followed by filtration to obtain 0.36 g of compound (33) as a yellowish green solid. [lambda] max (MeOH) 747 nm.

(Synthesis of Compound Example 34)
According to the above literature, 2,3,3-trimethyl-5-hydroxyethoxy-3H-pyrrolo [3,2-b] pyridine was synthesized. This was quaternized in the same manner as the synthesis of compound (33) and then condensed to obtain compound (34). [lambda] max (MeOH) 767 nm.

Example 2
(Creation of a breast cancer carcinogenesis model mouse)
Senescence accelerated mouse In order to develop breast cancer in SAMP6 / Ta mouse, which is one of the SAM strains, carcinogen 7,12-dimethylbenz [a] anthracene (DMBA) is administered, and a breast cancer model mouse is treated. Created.

The method for carcinogenesis in mice was performed according to Japanese Patent Application Laid-Open No. 2003-33125. 20 SAMP6 / Ta mice were administered to DMBA at a dose of 0.5 mg / mouse / week for a total of 6 times. As feed, high protein and high calorie CA-1 solid (manufactured by Claire Japan) was given. After the sixth administration of the carcinogen, the period from the first administration up to the 20th week was defined as a drug holiday. Breast cancer and lung metastases of breast cancer were searched histopathologically. DMBA was administered 6 times at 1-week intervals, then the drug was withdrawn from the start of the subsequent administration until the 20th week, and mice with breast cancer (75% breast cancer incidence) were used.
(Fluorescence contrast test)
A tumor tissue fragment (2 mm × 2 mm square) of mouse breast cancer was transplanted subcutaneously into the breast of the left breast of a BALB / c nude mouse (5 weeks old, Claire Japan). Ten days later, when the tumor grew to a diameter of about 5 mm, the mouse was subjected to the test. A titanium sapphire laser was used as the fluorescence excitation light source. Using a ring-type light guide (Sumita Optical Glass Co., Ltd.), the test mice were uniformly irradiated with laser light so that the dispersion of irradiation was within 2%. The irradiation output was adjusted to be about 36 μW / cm ² near the skin surface of the mouse. Fluorescence was excited at the maximum excitation wavelength of each compound, and fluorescence emission from the mouse was detected and imaged through a short wavelength cut-off filter using a CCD camera (C4880, Hamamatsu Photonics). The cut-off filter was selected to match the compound excitation wavelength (800 nm to 900 nm). The irradiation time was adjusted according to the fluorescence intensity of each compound.

  As a fluorescent contrast agent, each test compound was dissolved in distilled water (0.5 mg / ml) and administered to the mice from the milk ducts and the filaments. The dose was 1 mg / kg each. Twelve minutes after administration of the compound, the mouse was anesthetized with diethyl ether, and a fluorescence image of the whole body of the mouse was imaged (experiment numbers 101 to 111). As a sensor, a photodiode (wavelength range: 220 to 1100 nm) of a fluorometer (JASCO FP-6600) was used.

  The fluorescence sensitivity was represented by the logarithm of the amount of fluorescence generated with respect to the reference, and the relative sensitivity with the experiment number 101 (comparison) as 100.

  From the liquid chromatograph 2010A (manufactured by Shimadzu Corporation), the amount of the contrast medium discharged from the body is 100% of the amount of the compound after contrast medium administration, and the cumulative amount discharged by the catheter from the bladder and kidney after 3 weeks. Extracorporeal discharge was calculated and the discharge rate relative to the initial concentration was determined.

  In addition, the compound described in Unexamined-Japanese-Patent No. 2003-160558 was used as a comparative compound.

  In the water solubility test of the compound, 0.5 mg was dissolved in 1 ml of 0.9% physiological saline and left standing at 36 ° C. for 2 weeks to confirm precipitation and precipitation. No level at all, ◎, slightly haze applied, but disappeared by stirring ○, level that haze is applied but does not disappear by stirring △, level that precipitates × did.

  These results are summarized in Table 1.

  From Table 1, the fluorescence contrast agent for the diagnosis of tumor or cancer containing the cyanine dye represented by the general formula (1), (2) or (3) of the present invention is compared with the comparative fluorescence contrast agent. It turns out that it is excellent in water solubility, fluorescence sensitivity, and extracorporeal excretion rate.

Claims (8)

A fluorescent contrast agent for diagnosing a tumor or cancer comprising at least one penta- or heptamethine cyanine dye having 3H-pyrrolopyridine having at least two acid groups in the molecule as at least one mother nucleus. 2. The fluorescence for diagnosis of tumor or cancer according to claim 1, wherein the penta- or heptamethine cyanine dye is at least one kind represented by the following general formula (1), (2) or (3): Contrast agent.

[Wherein, R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ each represents an alkyl group, and the alkyl group represented by R ₁ and R ₄ may be substituted. Z ₁ and Z ₂ each represent a nonmetallic atom group necessary for forming a pyridine ring. Y ₁ represents a pyridine ring containing the following bond (1) in the ring, Y ₂ represents a dihydropyridine ring containing the following bond (2) in the ring, L represents a methine group, and X ⁻ represents an anion. m represents an integer of 3 or 4, and n represents an integer of 1 or 2. However, when m is 3, R ₁ and R ₄ are preferably substituted alkyl groups, and n is 1 when the dye forms an inner salt. ]

[R ₇ and R ₈ in bond (1) or bond (2) have the same meanings as R ₁ and R _{4 in} formula (1), (2) or (3). ] The tumor according to claim 1 or 2, wherein at least two acid groups of the molecule of the penta or heptamethine cyanine dye are groups selected from a sulfonic acid group, a carboxylic acid group, and a phosphoric acid group. Fluorescent contrast agent for cancer diagnosis. The at least 2 acid group which the molecule | numerator of the said penta or heptamethine cyanine dye has the group selected from a sulfonic acid group or a carboxylic acid group, and the sum total of an acid group is 2 or more, The fluorescent contrast agent for diagnosis of the tumor or cancer of any one of -3. The tumor or cancer according to any one of claims 1 to 4, wherein at least two of the acid groups of the penta- or heptamethine cyanine dye molecule are sulfonic acid groups. Fluorescent contrast agent for diagnosis. The fluorescence for diagnosis of tumor or cancer according to any one of claims 1 to 5, wherein at least two acid groups of the penta- or heptamethine cyanine dye molecule are sodium salts. Contrast agent. The fluorescence imaging for diagnosis of a tumor or cancer according to any one of claims 1 to 6, wherein the tumor or cancer is at least one selected from the group consisting of sarcomas and melanomas. Agent. Diagnosing a tumor or cancer by introducing the fluorescent contrast agent according to any one of claims 1 to 7 into a living body, irradiating the living body with excitation light, and detecting fluorescence from the contrast agent. An extracorporeal fluorescence imaging method characterized by the above.