JP7312441B2 - photoacoustic imaging agent - Google Patents

Description

TECHNICAL FIELD The present invention relates to an environment-responsive photoacoustic imaging agent whose photoacoustic signal changes depending on the environment.

Photoacoustic imaging (PAI) is a method of detecting and imaging ultrasonic waves (photoacoustic waves) generated by thermoelastic expansion of a material that has absorbed light. The photoacoustic effect is a phenomenon in which photoacoustic waves are generated due to the thermoelastic expansion of tissues accompanying the absorption of light energy by fluorescent molecules and tissues. Since an acoustic wave is used as a detection signal, imaging of deep parts is possible. For example, sound waves generated by absorption of excitation laser light can be used to obtain functional images at depths of several centimeters. In addition, since it is possible to perform analysis by superimposing it on an ultrasonic echo image, it is expected that it will be widely used as a useful imaging method for diagnosing pathological conditions in clinical practice in the future. In fact, it has been reported that when a photoacoustic imaging agent targeting prostate-specific membrane antigen (PSMA) was used in prostate cancer model mice, tumors could be detected by superimposing and analyzing photoacoustic images on echo images (see Non-Patent Document 1).

In photoacoustic imaging, endogenous light absorbers such as hemoglobin in the living body can be used as light absorbers for generating photoacoustic waves. Photoacoustic imaging agents generally utilize the accumulation of the imaging agent in the target tissue or site to perform imaging.

In order to image deep parts of the body, it is preferable to use a light absorber having an absorption wavelength in the near-infrared region, which is highly permeable to the body, as a photoacoustic imaging agent. For example, indocyanine green (ICG) is a light absorber having an absorption wavelength in the near-infrared region, and is used to label various substances in vivo. For example, an ICG-fused antibody, in which a monoclonal antibody is bound directly to ICG having a carboxyl group or via a PEG chain, can be used to label a target substance in vivo (see, for example, Non-Patent Documents 2 to 4).

Zhang et al., Journal of Biophotonics, 2018, vol. 11(9), e201800021. Zhou et al., Bioconjugate Chemistry, 2014, vol.25, p.1801-1810. Aung et al., Molecular Imaging, 2016, vol.15, p.1-11. Sano et al., Bioconjugate Chemistry, 2013, vol.24, p.811-816.

A photoacoustic imaging agent always exhibits a photoacoustic signal upon absorption of light of a particular wavelength. Therefore, it is not easy to achieve high-contrast imaging due to non-specific accumulation and the influence of the photoacoustic imaging agent staying in the blood. Moreover, it is very difficult to target the organs involved in the metabolism and excretion of the photoacoustic imaging agent due to the large background.

An object of the present invention is to provide a compound useful as an active ingredient of an environmentally responsive photoacoustic imaging agent whose light absorption properties change under a specific environment, and a photoacoustic imaging agent containing the compound as an active ingredient.

The present inventors have found that a derivative in which a specific substituent is introduced near the center of the methine chain that connects two benzoindolenine rings of ICG changes its conformation only in an acidic pH environment, resulting in a change in light absorption properties, and completed the present invention.

Specifically, the present invention provides the following ICG derivatives, pharmaceutical compositions, and photoacoustic imaging agents.
[1] General formula (1) below

[In the formula, ring A is a cyclohexene ring or a cyclopentene ring, R ¹ is a group represented by -(CH ₂ ) _n1 -NR ³ R ⁴ , n1 is 0, 1, 2, or 3, and R ³ and R ⁴ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ² is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ¹ and R ² may together form a ring. Also, an asterisk means a bond. ]
An indocyanine green derivative having a structure represented by
[2] The structure represented by the general formula (1) is represented by the following general formulas (1-1) to (1-6)

[wherein ring A is a cyclohexene ring or a cyclopentene ring, n1 is 0, 1, 2 or 3, R ¹¹ and R ¹² are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, R ¹³ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, and R ¹⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. . Also, an asterisk means a bond. ]
The indocyanine green derivative of the above [1], which is a structure represented by any one of
[3] The structure represented by the general formula (1) is represented by the following general formulas (1-1-1) to (1-1-3), (1-4-1) and (1-4-2)

[In the formula, ring A is a cyclohexene ring or a cyclopentene ring, and R ²¹ , R ²² and R ²³ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ²⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. n2 is 0, 1, 2 or 3, and R ²⁵ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. Also, an asterisk means a bond. ]
The indocyanine green derivative of [2] above, which is a structure represented by any one of
[4] A photoacoustic imaging agent comprising the indocyanine green derivative according to any one of [1] to [3] as an active ingredient.
[5] A pharmaceutical composition comprising the indocyanine green derivative according to any one of [1] to [3] as an active ingredient.
[6] A method for producing a photoacoustic imaging image, comprising administering the photoacoustic imaging agent of [4] to an individual animal (excluding humans), irradiating near-infrared light from the outside, and detecting the generated photoacoustic waves to produce a photoacoustic imaging image.
[7] Furthermore, the animal individual is externally irradiated with ultrasonic waves to prepare an echo image,
The method for producing a photoacoustic imaging image according to [6], wherein the echo image and the photoacoustic imaging image are superimposed.

The ICG derivative according to the present invention has different light absorption characteristics in an acidic pH environment and in a neutral pH environment. Utilizing this change in light absorption characteristics, a photoacoustic imaging agent containing the ICG derivative as an active ingredient can increase the contrast of photoacoustic signals between the target site and other sites in an acidic environment, and the target site can be detected more accurately. Therefore, the ICG derivative according to the present invention is particularly useful as an active ingredient of a pharmaceutical composition used for obtaining photoacoustic imaging images for analysis of the internal state of an animal's body.

1 is the measured absorption spectrum of the ICG derivative MA-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative Mor-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative DMA-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative TMEDA-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative Pipe-Cy in Example 1. FIG. 2 is the measured absorption spectrum of the ICG derivative MP-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative BP-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative PBA-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative PDA-Cy in Example 1. FIG. 1 is the measured absorption spectrum of the ICG derivative TMPDA-Cy in Example 1. FIG. 1 shows the photoacoustic spectrum and absorption spectrum of TMEDA-Cy in Example 1. FIG. 1 shows the photoacoustic spectrum and absorption spectrum of TMPDA-Cy in Example 1. FIG.

ICG is a clinical diagnostic drug that has already been approved by the US FDA, and is a near-infrared light absorber known to have extremely low toxicity to living bodies. ICG has a high molar extinction coefficient and easily generates photoacoustic waves, but has a problem of low photostability. In addition, since ICG has no environmental response, when ICG is used as a photoacoustic imaging agent, photoacoustic waves are also generated from ICG present outside the target site, and the acquired photoacoustic imaging image tends to have high noise and insufficient accuracy.

In contrast, the ICG derivative according to the present invention changes its light absorption characteristics depending on pH. More specifically, as the pH becomes more acidic, the maximum absorption wavelength shifts to the longer wavelength side. By irradiating the ICG derivative with light, a photoacoustic signal corresponding to the absorption spectrum is detected. That is, the ICG derivative according to the present invention is suitable as an active ingredient of a pH-responsive photoacoustic imaging agent in which the detected photoacoustic signal changes depending on pH.

The ICG derivative according to the present invention is a compound having a structure represented by the following general formula (1). In general formula (1), an asterisk means a bond.

In general formula (1), ring A is a cyclohexene ring or a cyclopentene ring. By introducing a 6- or 5-membered alicyclic structure near the center of the methine chain that connects the two benzoindolenine rings of ICG, the ICG derivative according to the present invention has improved photostability compared to ICG. As for the ICG derivative according to the present invention, ring A is preferably a cyclohexene ring.

In general formula (1), R ¹ is a group represented by —(CH ₂ ) _n1 —NR ³ R ⁴ . where n1 is 0, 1, 2, or 3. By introducing a group in which two amino groups are linked by an alkylene group having 1 to 3 carbon atoms in the vicinity of the center of the methine chain linking two benzoindolenine rings of ICG, the conformation is probably changed in a pH-dependent manner, and the light absorption properties are changed. As for the ICG derivative according to the present invention, n1 is preferably 2 or 3, more preferably 2, because the maximum absorption wavelength can be shifted depending on pH.

R ³ and R ⁴ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. The alkyl group having 1 to 6 carbon atoms may be a linear group or a branched group. Examples of alkyl groups having 1 to 6 carbon atoms include methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, sec-butyl group, tert-butyl group, pentyl group, isopentyl group, neopentyl group, tert-pentyl group and hexyl group.

When R ³ and R ⁴ are alkyl groups having 1 to 6 carbon atoms, the alkyl groups may have 1 to 3 substituents. The substituent is not particularly limited as long as it does not impair the effects of the present invention, and examples thereof include a phenyl group optionally having 1 to 3 substituents. Examples of substituents possessed by the phenyl group include alkyl groups having 1 to 6 carbon atoms, carboxyalkyl groups, and the like.

In general formula (1), R ² is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. As the alkyl group having 1 to 6 carbon atoms which may have a substituent, the same groups as those mentioned for R ³ or R ⁴ can be used.

In general formula (1), R ¹ and R ² may together form a ring. The ring formed by R ¹ and R ² is not particularly limited as long as it is a ring having a nitrogen atom at any one of positions 3 to 5 when the nitrogen atom connected to ring A is the 1st position. Examples of the ring include piperazine ring, tetrahydropyrimidine ring, hexahydropyrimidine ring, dihydroimidazole ring, tetrahydroimidazole ring and the like. Moreover, these rings may have a substituent. The substituent is not particularly limited as long as it does not impair the effects of the present invention, and examples thereof include an optionally substituted alkyl group having 1 to 6 carbon atoms.

The structure represented by general formula (1) possessed by the ICG derivative according to the present invention is preferably a structure represented by any one of the following general formulas (1-1) to (1-6). In general formulas (1-1) to (1-6), an asterisk means a bond.

In general formulas (1-1) to (1-6), ring A is a cyclohexene ring or a cyclopentene ring. In general formulas (1-1) to (1-6), ring A is preferably a cyclohexene ring.

In general formula (1-1), n1 is 0, 1, 2 or 3, R ¹¹ and R ¹² are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, and R ¹³ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. As the optionally substituted alkyl group having 1 to 6 carbon atoms in R ¹¹ , R ¹² and R ¹³ , the same groups as those mentioned for R ³ or R ⁴ can be used.

In general formulas (1-2), (1-4) and (1-5), R ¹⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. As the optionally substituted alkyl group having 1 to 6 carbon atoms in R ¹⁴ , the same groups as those mentioned for R ³ or R ⁴ can be used.

The structure represented by general formula (1-1) is preferably a structure represented by any one of the following general formulas (1-1-1) to (1-1-3). In general formulas (1-1-1) to (1-1-3), ring A is a cyclohexene ring or cyclopentene ring, and R ²¹ , R ²² and R ²³ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms.一般式（１－１－１）～（１－１－３）で表される構造としては、環Ａがシクロヘキセン環であり、Ｒ ^２１ 、Ｒ ^２２ 、及びＲ ^２３がそれぞれ独立して、水素原子、メチル基、エチル基、プロピル基、イソプロピル基、ブチル基、イソブチル基、ｓｅｃ－ブチル基、ｔｅｒｔ－ブチル基、ペンチル基、イソペンチル基、ネオペンチル基、ｔｅｒｔ－ペンチル基、又はヘキシル基である構造が好ましく、環Ａがシクロヘキセン環であり、Ｒ ^２１ 、Ｒ ^２２ 、及びＲ ^２３がそれぞれ独立して、水素原子、メチル基、エチル基、プロピル基、又はイソプロピル基である構造がより好ましく、環Ａがシクロヘキセン環であり、Ｒ ^２１ 、Ｒ ^２２ 、及びＲ ^２３がそれぞれ独立して、水素原子、メチル基、又はエチル基である構造がさらに好ましく、環Ａがシクロヘキセン環であり、Ｒ ^２１ 、Ｒ ^２２ 、及びＲ ^２３がそれぞれ独立して、水素原子又はメチル基である構造がよりさらに好ましい。

In the structure represented by general formula (1-2), ring A is a cyclohexene ring, and R¹⁴is a hydrogen atom, methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, sec-butyl group, tert-butyl group, pentyl group, isopentyl group, neopentyl group, tert-pentyl group, or hexyl group, ring A is a cyclohexene ring, and R¹⁴is a hydrogen atom, a methyl group, an ethyl group, a propyl group, or an isopropyl group, ring A is a cyclohexene ring, and R¹⁴is more preferably a hydrogen atom, a methyl group, or an ethyl group.

As the structure represented by general formula (1-3), a structure in which ring A is a cyclohexene ring is preferable.

As the structure represented by general formula (1-4), a structure represented by any one of the following general formulas (1-4-1) to (1-4-2) is preferable. In general formulas (1-4-1) to (1-4-2), ring A is a cyclohexene ring or a cyclopentene ring. In general formula (1-4-1), R ²⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. In general formula (1-4-2), n2 is 0, 1, 2, or 3, and R ²⁵ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms.

In the structure represented by general formula (1-4-1), ring A is a cyclohexene ring, and R²⁴is a hydrogen atom, methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, sec-butyl group, tert-butyl group, pentyl group, isopentyl group, neopentyl group, tert-pentyl group, or hexyl group, ring A is a cyclohexene ring, and R²⁴is a hydrogen atom, a methyl group, an ethyl group, a propyl group, or an isopropyl group, ring A is a cyclohexene ring, and R²⁴is more preferably a hydrogen atom, a methyl group, or an ethyl group.

The structure represented by general formula (1-4-2) is preferably a structure in which ring A is a cyclohexene ring, n2 is 0 or 1, R ²⁵ is a methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, sec-butyl group, tert-butyl group, carboxymethyl group, or carboxyethyl group, and ring A is a cyclohexene ring and n2 is 0, or ring A is a cyclohexene ring and n2 is 1 and R ²⁵ is a methyl group, an ethyl group, a carboxymethyl group, or a carboxyethyl group is more preferred.

In the structure represented by general formula (1-5), ring A is a cyclohexene ring, and R¹⁴is preferably a hydrogen atom, a methyl group, an ethyl group, a propyl group, an isopropyl group, or a benzyl group optionally having a substituent, ring A is a cyclohexene ring, and R¹⁴is more preferably a hydrogen atom, a methyl group, an ethyl group, a propyl group, an isopropyl group, a benzyl group, or a benzyl group substituted with a carboxyalkyl group, ring A is a cyclohexene ring, and R¹⁴is a hydrogen atom, a methyl group, an ethyl group, a propyl group, an isopropyl group, a benzyl group, or a benzyl group substituted with a carboxymethyl group.

As the structure represented by general formula (1-6), a structure in which ring A is a cyclohexene ring is preferable.

In the ICG derivative according to the present invention, the structural portion to which the structure represented by the general formula (1) is bonded is not particularly limited as long as it does not impair the effect of the structure represented by the general formula (1) as a photoacoustic imaging agent. Such structures include, for example, a sulfo group (--SO ₃ H) similar to ICG or a salt thereof, and a sulfo group or a salt thereof bound to an arbitrary linking group. Examples of the linking group include an alkylene group, an alkenylene group, a carbonyl group (-CO-), an ether bond (-O-), an ester bond (-COO-), an amide bond (-CONH-), and polyethylene glycol (PEG:-(C ₂ H ₄ O)n-), and these can be used in combination as appropriate.

When the ICG derivative according to the present invention is used as an active ingredient of a photoacoustic imaging agent, it can have a structure in which a molecule capable of binding to a target molecule to be detected by the photoacoustic imaging agent is bound to an arbitrary linking group. Examples of the linking group include those similar to those described above. A molecule that can bind to a target molecule may be a peptide or protein, a nucleic acid, a lipid, a sugar chain, or a low-molecular-weight compound. Such molecules include, for example, antibodies against target molecules.

In addition, tumor cells generally express more integrins. Therefore, when the target molecule is a tumor cell, an integrin-binding peptide containing an RGD sequence can be used as a molecule capable of binding to the target molecule. By using an ICG derivative in which an integrin-binding peptide and a structure represented by the general formula (1) are bonded directly or via an appropriate linking group as a photoacoustic imaging agent, a photoacoustic imaging image of tumor cells in the body of an animal can be obtained.

In the ICG derivative according to the present invention, the structural portion to which the structure represented by the general formula (1) is bound may have the same structure as a known ICG derivative. Examples of known ICG derivatives include ICG-Sulfo-OSu (code: I254, manufactured by Dojindo Laboratories), ICG-EG4-Sulfo-OSu (code: I289, manufactured by Dojindo Laboratories), and ICG-EG8-Sulfo-OSu (code: I290, manufactured by Dojindo Laboratories).

Since the ICG derivative according to the present invention has ICG as a basic skeleton, it can be expected to have low toxicity to living bodies like ICG. Moreover, the photoacoustic wave generated from the ICG derivative according to the present invention is a sound wave with high bio-permeability. Therefore, the ICG derivative is suitable as an active ingredient of a pharmaceutical composition to be administered to animals including humans.

A pharmaceutical composition containing an ICG derivative according to the present invention as an active ingredient can be formulated into oral solid formulations such as powders, granules, capsules, tablets, chewable formulations and sustained-release formulations, oral liquid formulations such as solutions and syrups, injections, enema formulations, sprays, patches, ointments, and the like, by conventional methods. Formulation can be carried out by conventional methods by blending excipients, binders, lubricants, disintegrants, fluidizing agents, solvents, solubilizers, buffers, suspending agents, emulsifiers, tonicity agents, stabilizers, preservatives, antioxidants, flavoring agents, coloring agents, etc., as necessary for the formulation.

The ICG derivative according to the present invention shifts the light absorption characteristics to the longer wavelength side as the pH becomes more acidic. Therefore, the ICG derivative according to the present invention is particularly suitable as an active ingredient of a photoacoustic imaging agent for detecting molecules and tissues that are expected to exist in an acidic environment.

For example, many extracellular substances are taken up into cells by endocytosis, but endosomes are generally acidic. Therefore, by using an ICG derivative in which the structure represented by the general formula (1) and a molecule that recognizes a specific cell (target cell) are linked by a linking group as a photoacoustic imaging agent, the photoacoustic signal from the target cell can be obtained in a state with less noise, and a more accurate photoacoustic imaging image can be obtained in real time with a high S/N ratio. In addition, the ICG derivative according to the present invention is preferable as an active ingredient of a photoacoustic imaging agent for obtaining a photoacoustic imaging image of pathological tissue associated with an acidic environment.

The administration route of the photoacoustic imaging agent and pharmaceutical composition containing the ICG derivative according to the present invention as an active ingredient is not particularly limited, and is appropriately determined according to target cells and tissues containing the target cells. For example, administration routes of the photoacoustic imaging agent containing the ICG derivative of the present invention as an active ingredient include oral administration, intravenous administration, intraperitoneal administration, enema administration and the like.

The animal to which the photoacoustic imaging agent and pharmaceutical composition containing the ICG derivative of the present invention as an active ingredient is administered is not particularly limited, and may be a human or a non-human animal. Non-human animals include mammals such as cows, pigs, horses, sheep, goats, monkeys, dogs, cats, rabbits, mice, rats, hamsters and guinea pigs, and birds such as chickens, quails and ducks.

The photoacoustic imaging agent containing the ICG derivative according to the present invention as an active ingredient can be used in the same manner as the photoacoustic imaging agents used in the preparation of conventional photoacoustic imaging images. Specifically, first, the photoacoustic imaging agent according to the present invention is administered to an individual animal. Next, the individual animal is irradiated with near-infrared light from the outside, and the generated photoacoustic wave signal is detected. Detection of the photoacoustic signal can be performed using an ultrasonic detector used in echo inspection or the like, and a photoacoustic imaging image can be produced from the detected photoacoustic signal by a conventional method.

The wavelength of the near-infrared light to be irradiated is not particularly limited as long as it is a wavelength at which the photoacoustic imaging agent in an acidic environment can generate a photoacoustic wave, but since a photoacoustic imaging image with less noise and a high S/N ratio can be produced, it is preferably near the maximum absorption wavelength of the photoacoustic imaging agent in an acidic environment and longer than the maximum absorption wavelength of the photoacoustic imaging agent in a neutral environment.

The photoacoustic imaging agent containing the ICG derivative according to the present invention as an active ingredient has a light absorption characteristic shifted to the longer wavelength side when taken into cells than when it is not taken into cells. By utilizing this difference in light absorption characteristics and irradiating light with a longer wavelength, it is possible to suppress the influence of noise on the photoacoustic signal generated and acquired from the ICG derivative in the state of being incorporated into cells.

An animal individual administered with a photoacoustic imaging agent can also be subjected to an echo examination. Specifically, the individual animal is externally irradiated with ultrasonic waves to create an echo image. By superimposing and analyzing the obtained echo image and photoacoustic imaging image, the target cell can be analyzed in more detail.

EXAMPLES Next, the present invention will be described in more detail with reference to examples, but the present invention is not limited to the following examples.

[Example 1]
An ICG derivative was synthesized and its light absorption spectrum was measured.

<Synthesis of ICG derivative>
Ten kinds of ICG derivatives were synthesized by the following synthetic reactions. It was confirmed by NMR and MS that the synthesized ICG derivative had the desired structure.

<Measurement of light absorption spectrum>
In order to examine the light absorption properties of the synthesized ICG derivatives, the absorption spectra were measured at a concentration of 1 μM in phosphate buffers (30% by volume DMSO and 70% by volume phosphate buffers) with dimethyl sulfoxide (DMSO) as a co-solvent, using an ultraviolet/visible spectrophotometer, and the pH dependence was investigated. Absorption spectra of each ICG derivative are shown in FIGS.

As a result, each ICG derivative exhibited different pH responsiveness depending on the structural features of the introduced amino group. MA-Cy (Fig. 1), DMA-Cy (Fig. 2), and Mor-Cy (Fig. 3) into which a monoamine group was introduced showed no change in absorption characteristics due to pH. On the other hand, in the ICG derivative into which a diamino group was introduced, the maximum absorption wavelength shifted to the longer wavelength side as the pH became more acidic. In particular, TMEDA-Cy (FIG. 4), Pipe-Cy (FIG. 5), MP-Cy (FIG. 6), BP-Cy (FIG. 7), and PBA-Cy (FIG. 8), which have two amino groups in the positional relationship of 1-position and 4-position, show an absorption maximum near 700 nm in a neutral environment, but the absorption near 800 nm increased in an acidic environment. These results indicated that the absorption spectral change in response to pH could be induced by protonation of the amine moiety of ICG.

<Measurement of photoacoustic spectrum>
For TMEDA-Cy and TMPDA-Cy, further photoacoustic signals were detected to examine the correlation between the photoacoustic spectrum and the absorption spectrum. A solution obtained by dissolving an ICG derivative in a phosphate buffer (pH 3.5 or 7.0) to 10 μM was used as a sample, and a wavelength-tunable OPO laser (pulse width: 6 to 8 ns, repetition frequency: 10 Hz, pulse energy: 20 to 50 mJ) was used to irradiate a short pulse laser beam, and the generated photoacoustic signal was measured. The excitation wavelength was measured in the range of 500 to 1000 nm in increments of 5 nm, and the irradiation energy was 100±50 μJ/pulse.

The photoacoustic spectrum and absorption spectrum of TMEDA-Cy are shown in FIG. 11, and the photoacoustic spectrum and absorption spectrum of TMPDA-Cy are shown in FIG. 12, respectively. As shown in the figure, photoacoustic signals corresponding to absorption spectra were observed for both compounds. From these results, the photoacoustic spectrum and the absorption spectrum are correlated, and therefore, TMEDA-Cy, Pipe-Cy, MP-Cy, BP-Cy, PBA-Cy, PDA-Cy, and TMPDA-Cy, which are ICG derivatives into which diamines are introduced, show a large change in the photoacoustic spectrum when the environment is changed from a neutral environment to an acidic environment, and the photoacoustic signal shifts to a longer wavelength in an acidic environment. It was shown that photoacoustic imaging can be performed according to the pH environment by using .

Claims (10)

General formula (1) below

[In the formula, ring A is a cyclohexene ring or a cyclopentene ring, R ¹ is a group represented by -(CH ₂ ) _n1 -NR ³ R ⁴ , n1 is 0, 1, 2, or 3, and R ³ and R ⁴ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ² is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ¹ and R ² may together form a ring. However, when n1 is 2 and R3 ^and R4 ^are pyridine-substituted methyl groups, R2 ^is not a hydrogen atom. Also, an asterisk means a bond. The bond is directly or via a linking group, and is bonded to a sulfo group or a salt thereof. The linking group is an alkylene group, an alkenylene group, a carbonyl group, an ether bond, an ester bond, an amide bond, polyethylene glycol, or a combination thereof. ]
An indocyanine green derivative having a structure represented by The structure represented by the general formula (1) is represented by the following general formulas (1-1) to (1-6)

[wherein ring A is a cyclohexene ring or a cyclopentene ring, n1 is 0, 1, 2 or 3, R ¹¹ and R ¹² are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, R ¹³ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, and R ¹⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. . However, when n1 is 2 and R11 ^and R12 ^are pyridine-substituted methyl groups, R13 ^is not a hydrogen atom. Also, an asterisk means a bond. The bond is directly or via a linking group, and is bonded to a sulfo group or a salt thereof. The linking group is an alkylene group, an alkenylene group, a carbonyl group, an ether bond, an ester bond, an amide bond, polyethylene glycol, or a combination thereof. ]
2. The indocyanine green derivative according to claim 1, which has a structure represented by any one of: The structure represented by the general formula (1) is represented by the following general formulas (1-1-1) to (1-1-3), (1-4-1) and (1-4-2)

[In the formula, ring A is a cyclohexene ring or a cyclopentene ring, and R ²¹ , R ²² and R ²³ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ²⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. n2 is 0, 1, 2 or 3, and R ²⁵ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. Also, an asterisk means a bond. The bond is directly or via a linking group, and is bonded to a sulfo group or a salt thereof. The linking group is an alkylene group, an alkenylene group, a carbonyl group, an ether bond, an ester bond, an amide bond, polyethylene glycol, or a combination thereof. ]
2. The indocyanine green derivative according to claim 1, which has a structure represented by any one of: A photoacoustic imaging agent comprising the indocyanine green derivative according to any one of claims 1 to 3 as an active ingredient. General formula (1) below

[In the formula, ring A is a cyclohexene ring or a cyclopentene ring, R ¹ is a group represented by -(CH ₂ ) _n1 -NR ³ R ⁴ , n1 is 0, 1, 2, or 3, and R ³ and R ⁴ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ² is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ¹ and R ² may together form a ring. However, when n1 is 2 and R3 ^and R4 ^are pyridine-substituted methyl groups, R2 ^is not a hydrogen atom. Also, an asterisk means a bond. The bond binds to a molecule capable of binding to a target molecule via a linking group. The linking group is an alkylene group, an alkenylene group, a carbonyl group, an ether bond, an ester bond, an amide bond, polyethylene glycol, or a combination thereof. ]
A photoacoustic imaging agent having a structure represented by The structure represented by the general formula (1) is represented by the following general formulas (1-1) to (1-6)

[wherein ring A is a cyclohexene ring or a cyclopentene ring, n1 is 0, 1, 2 or 3, R ¹¹ and R ¹² are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, R ¹³ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms, and R ¹⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. . However, when n1 is 2 and R11 ^and R12 ^are pyridine-substituted methyl groups, R13 ^is not a hydrogen atom. Also, an asterisk means a bond. The bond binds to a molecule capable of binding to a target molecule via a linking group. The linking group is an alkylene group, an alkenylene group, a carbonyl group, an ether bond, an ester bond, an amide bond, polyethylene glycol, or a combination thereof. ]
The photoacoustic imaging agent according to claim 5, which is a structure represented by any one of. The structure represented by the general formula (1) is represented by the following general formulas (1-1-1) to (1-1-3), (1-4-1) and (1-4-2)

[In the formula, ring A is a cyclohexene ring or a cyclopentene ring, and R ²¹ , R ²² and R ²³ are each independently a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. R ²⁴ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. n2 is 0, 1, 2 or 3, and R ²⁵ is a hydrogen atom or an optionally substituted alkyl group having 1 to 6 carbon atoms. Also, an asterisk means a bond. The bond binds to a molecule capable of binding to a target molecule via a linking group. The linking group is an alkylene group, an alkenylene group, a carbonyl group, an ether bond, an ester bond, an amide bond, polyethylene glycol, or a combination thereof. ]
The photoacoustic imaging agent according to claim 5, which is a structure represented by any one of. A pharmaceutical composition comprising the indocyanine green derivative according to any one of claims 1 to 3 or the photoacoustic imaging agent according to any one of claims 4 to 7 as an active ingredient. A method for producing a photoacoustic imaging image, comprising administering the photoacoustic imaging agent according to any one of claims 4 to 7 to an animal individual (excluding humans), irradiating near-infrared light from the outside, and detecting the generated photoacoustic waves to produce a photoacoustic imaging image. Furthermore, the animal individual is externally irradiated with ultrasonic waves to create an echo image,
The method for producing a photoacoustic imaging image according to claim 9 , wherein the echo image and the photoacoustic imaging image are superimposed.

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