Pharmaceutical composition for radiotherapy, and method for treating solid cancer using same

Abstract

Provided is a pharmaceutical composition to be used for radiotherapy that comprises a compound represented by formula (I) [wherein: R1 is an aryl group substituted by a substituent selected from an iodine atom, etc., wherein the aryl group may be optionally substituted by a substituent such as a hydroxy group; X is a bond, etc.; and ring A is a group represented by formula (A), wherein R2 is a non-aromatic heterocyclic group which may be optionally substituted by one or more C1-C4 alkyl groups] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. Also provided are nanoparticles and a method for treating solid cancer, each using the aforesaid compound.

Description

Pharmaceutical composition for radiation therapy and method for treating solid cancer using the same

[0001] The present invention relates to a pharmaceutical composition for radiation therapy, particularly X-ray therapy, and a method for treating solid cancers using the same. The present invention also relates to nanoparticles that can be used in radiation therapy.

[0002] Auger therapy is one of the currently under development radiation therapies. Auger therapy involves localizing high-Z atoms (atoms of high-Z elements) such as gadolinium and iodine within cells and then irradiating the high-Z atoms with X-rays. For example, a compound containing iodine atoms is incorporated into cancer cells and localized near the DNA. When X-rays of a specific energy are then irradiated, the X-ray energy is absorbed by the iodine atoms, resulting in a photoelectric effect, such as the release of K-shell electrons. As a result, Auger electrons, which have a strong DNA cleavage and cancer cell killing effect, are generated near the DNA, exerting a therapeutic effect. Auger electrons have a short flight distance (several hundred nanometers), but are highly effective in DNA damage and cell killing. Therefore, it is important to localize high-Z atoms near the DNA.

[0003] Previously, silica nanoparticles containing high Z atoms, which are suitable for use in Auger therapy, have been proposed by research groups including the present inventors (for example, Non-Patent Documents 1 and 2, Patent Document 1).

[0004] Auger therapy using iodine atoms is also known, and for example, a method using IUdR, a nucleotide analog, is known (see, for example, Non-Patent Document 3). However, this substance has problems such as being difficult to use because it is rapidly metabolized in the body and exhibits systemic toxicity.

[0005] International Publication No. 2021 / 060498

[0006] Scientific reports (2019), 9, 13275. Matsumoto et al. (https: / / doi.org / 10.1038 / s41598-019-49978-1)Scientific reports (2021), 11, 14192. Higashi et al. (https: / / doi.org / 10.1038 / s41598-021-93429-9)British Journal of Cancer (2004), 91, 544-551

[0007] The present invention has been made in view of the above circumstances, and aims to provide a pharmaceutical composition and nanoparticles that can be used in radiation therapy, particularly Auger therapy using X-rays, and a method for treating solid cancer using the same.

[0008] The present invention includes, but is not limited to, the embodiments listed below.

[0009] [1] Formula (I): [In the formula, R １ represents an iodine atom, a C substituted with one or more iodine atoms １ -C ４ an aryl group substituted with one or more substituents selected from the group consisting of an alkyl group, an aryl group substituted with one or more iodine atoms, a gadolinium atom-containing group, a gold atom-containing group, a silver atom-containing group, and a platinum atom-containing group, wherein the aryl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ X is optionally substituted with one or more substituents selected from the group consisting of an alkoxy group; ２ or C ４ and Ring A is an alkenylene group of formula (A): where R ２ optionally one or more C １ -C ４a non-aromatic heterocyclic group optionally substituted with an alkyl group, containing one or more heteroatoms selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms, and at least one of the heteroatoms being a nitrogen atom; or a pharmaceutically acceptable salt thereof (hereinafter also referred to as "the compound of the present invention"), and a pharmaceutically acceptable carrier, for use in radiation therapy (hereinafter also referred to as "the pharmaceutical composition of the present invention"). [2] R １ is an aryl group substituted with one or more iodine atoms, wherein the aryl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ optionally substituted with one or more substituents selected from the group consisting of alkoxy groups; and ２ optionally one or more C １ -C ４ [3] The pharmaceutical composition according to [1], wherein the compound of formula (I) is a compound of formula (II): [In the formula, R ３ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with 1 or 2 hydroxy groups; and R ４ is a hydrogen atom or C １ -C ４[4] The pharmaceutical composition according to any one of [1] to [3], wherein the radiation is X-rays. [5] The pharmaceutical composition according to [4], wherein the X-rays are X-rays capable of exciting K-shell electrons of an atom selected from the group consisting of iodine, gadolinium, gold, silver, and platinum in the compound of formula (I). [6] The pharmaceutical composition according to [4] or [5], wherein the X-rays are monochromatic X-rays or characteristic X-rays. [7] The pharmaceutical composition according to any one of [1] to [6], wherein the solid cancer is for treating or inhibiting the growth or proliferation of a solid cancer. [8] The pharmaceutical composition according to [7], wherein the solid cancer is brain tumor, lung cancer, ovarian cancer, digestive system cancer, osteosarcoma, or head and neck cancer. [9] The pharmaceutical composition according to [7] or [8], wherein the solid cancer is in a hypoxic state.

[10] Formula (I): [In the formula, R １ represents an iodine atom, a C substituted with one or more iodine atoms １ -C ４ an aryl group substituted with one or more substituents selected from the group consisting of an alkyl group, an aryl group substituted with one or more iodine atoms, a gadolinium atom-containing group, a gold atom-containing group, a silver atom-containing group, and a platinum atom-containing group, wherein the aryl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ X is optionally substituted with one or more substituents selected from the group consisting of an alkoxy group; ２ or C ４ and Ring A is an alkenylene group of formula (A): where R ２ optionally one or more C １ -C ４ a non-aromatic heterocyclic group optionally substituted with an alkyl group, containing one or more heteroatoms selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms, and at least one of the heteroatoms being a nitrogen atom; is a link to the X group.] or a pharmaceutically acceptable salt thereof; and a porous silica carrier, nanoparticles (hereinafter also referred to as "nanoparticles of the present invention"). [10-1] A nanoparticle of porous silica carrying the above-mentioned iodine atom-containing Hoechst compound (IH: Iodine-Hoechst).

[11] The nanoparticle according to

[10] , wherein the nanoparticle is a biodegradable mesoporous silica nanoparticle. [11-1] The nanoparticle according to

[10] or

[11] , for use in X-ray irradiation.

[12] A method for treating solid cancer or suppressing the growth or proliferation of solid cancer, comprising irradiating the compound of formula (I) or a pharmaceutically acceptable salt thereof according to [1], the pharmaceutical composition according to any one of [1] to [9], or the nanoparticle according to

[10] or

[11] , which has been taken into the body of a subject, with X-rays to destroy the cancer cells.

[13] The pharmaceutical composition according to any one of [4] to [9], the nanoparticle according to [11-1], or the method according to

[12] , wherein the compound of formula (I) contains an iodine atom and the X-rays are monochromatic X-rays having an energy of 33.2 keV.

[14] The compound of formula (I) according to [1] or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to any one of [1] to [9], or the nanoparticle according to

[10] ,

[11] , or [11-1], used for treating solid cancer or suppressing the growth or proliferation of solid cancer.

[15] Use of the compound of formula (I) according to [1] or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to any one of [1] to [9], or the nanoparticle according to

[10] ,

[11] , or [11-1], in the manufacture of a medicament for treating solid cancer or suppressing the growth or proliferation of solid cancer.

[0010] According to the present invention, highly effective radiation therapy can be performed using the Auger effect.

[0011] 1 shows the results (images and calculated values) of a docking simulation between iodine atom-containing Hoechst compounds (four types) and DNA, viewed along the helix axis. 2 shows the results (images) of a docking simulation between iodine atom-containing Hoechst compounds (four types) and DNA, viewed perpendicular to the helix axis. 3 shows images of OVCAR8 cancer cells observed under a microscope. A shows the fluorescent color (green) emitted by the OVCAR8 cancer cells, B shows the colored (blue) region produced by the Hoechst compounds functioning as a dye, and C shows a processed image obtained by superimposing A and B. 4 shows the results (images) of a tumor spheroid observed under a microscope. 5 shows an overview of the setup of a monochromatic X-ray irradiation device. 6 shows an overview of the tumor spheroid sample portion in the monochromatic X-ray irradiation device. 7 shows the results (images) of a tumor spheroid observed under a microscope after monochromatic X-ray irradiation (after incubation). 8 shows the results (images) of a tumor spheroid observed under a microscope after monochromatic X-ray irradiation (immediately after irradiation). Graph showing the results of a cytotoxicity test of an iodine atom-containing Hoechst compound. Images showing the results of microscopic observation of tumor spheroids in a hypoxic condition test. Graph showing the transition of fluorescent signal intensity of the hypoxia marker HIF-1α. Results of tumor spheroids after X-ray irradiation in a hypoxic condition test (photograph of the bottom of the tube). TEM image of an example of mesoporous silica nanoparticles. Absorption spectra of the solution and washing solution used in nanoparticle production, measured with a spectrophotometer. Results of nuclear staining using IH-BPMO. Results of a cancer spheroid destruction experiment using IH-BPMO.

[0012] Definitions To explain the present invention, the definitions of terms will be provided first. In this specification, an alkyl group means an aliphatic saturated hydrocarbon group composed of carbon atoms and hydrogen atoms. The alkyl group may be linear or branched. １ -C ４ The alkyl group represents an alkyl group having 1 to 4 carbon atoms. １ -C ４ Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, and tert-butyl groups.

[0013] In this specification, an alkoxy group refers to a group represented by (alkyl)-O-. The meaning of alkyl is as defined above. １ -C ４ The alkoxy group is an alkoxy group having 1 to 4 carbon atoms. １ -C ４ Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, and tert-butoxy groups.

[0014] In this specification, an aryl group refers to an aromatic hydrocarbon group. The aryl group preferably has 6 to 12 carbon atoms, more preferably 6 to 10 carbon atoms, and even more preferably 6 to 8 carbon atoms. Examples of the aryl group include, but are not limited to, a phenyl group, a naphthyl group, a phenanthryl group, and an anthracenyl group.

[0015] As used herein, an alkenylene group refers to a divalent unsaturated hydrocarbon group having at least one carbon-carbon double bond. ２ or C ４ The alkenylene group in the above means an alkenylene group having 2 or 4 carbon atoms, and specific examples include -C=C- and -C=C-C=C-.

[0016] In this specification, a non-aromatic heterocyclic group refers to a non-aromatic cyclic group containing one or more heteroatoms selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms among the atoms constituting the ring, with the remainder being carbon atoms. However, in the non-aromatic heterocyclic group in the compound of formula (I), at least one of the heteroatoms is a nitrogen atom. The non-aromatic heterocyclic group is preferably a 4- to 8-membered ring, more preferably a 5- or 6-membered ring, and even more preferably a 6-membered ring. The non-aromatic heterocyclic group may or may not have a double bond in the ring, as long as it maintains a non-aromatic system. The non-aromatic heterocyclic group may be a saturated cyclic group or an unsaturated cyclic group, with a saturated cyclic group being more preferred. Examples of non-aromatic heterocyclic groups include, but are not limited to, piperidinyl, piperazinyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, morpholinyl, thiomorpholinyl, oxazolidinyl, and thiazolidinyl groups.

[0017] In this specification, the term "bond" in the definition of group X means that the two moieties connected to X in the chemical structural formula are directly bonded (i.e., X is substantially absent). Specifically, for example, in the structure of ring A-X-benzene ring (see formula (I)), when X is a bond, it means ring A-benzene ring (i.e., ring A and the benzene ring are directly bonded).

[0018] In this specification, a hydroxy group refers to a group represented by —OH.

[0019] Compound The pharmaceutical composition of the present invention comprises a compound of formula (I): or a pharmaceutically acceptable salt thereof, wherein in the above formula (I), R １ represents an iodine atom, a C substituted with one or more iodine atoms １ -C ４an aryl group substituted with one or more substituents selected from the group consisting of an alkyl group, an aryl group substituted with one or more iodine atoms, a gadolinium atom-containing group, a gold atom-containing group, a silver atom-containing group, and a platinum atom-containing group, wherein the aryl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ X is optionally substituted with one or more substituents selected from the group consisting of an alkoxy group; ２ or C ４ and Ring A is an alkenylene group of formula (A): where R ２ optionally one or more C １ -C ４ a non-aromatic heterocyclic group optionally substituted with an alkyl group, containing one or more heteroatoms selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms, and at least one of the heteroatoms being a nitrogen atom; (ie, the wavy line) is the connection to the X group.

[0020] The compound of formula (I) generally comprises a benzimidazole ring structure having a group R attached to the imidazolyl group on the right side. １ is bonded to the phenyl group of the benzimidazole ring structure, and ring A (another benzimidazole ring structure located on the left side) is bonded to the phenyl group of the benzimidazole ring structure via an X group.

[0021] R １ represents an iodine atom, a C substituted with one or more iodine atoms １ -C ４ The R may be an alkyl group, an aryl group substituted with one or more substituents selected from the group consisting of an aryl group substituted with one or more iodine atoms, a gadolinium atom-containing group, a gold atom-containing group, a silver atom-containing group, and a platinum atom-containing group. １ has an aryl group as a basic skeleton, and this aryl group is bonded to the ring structure of benzimidazole. １The aryl group that forms the basic skeleton of is a group as defined above, and is preferably a phenyl group or a naphthyl group, although it is not limited thereto.

[0022] R １ represents an iodine atom, a C substituted with one or more iodine atoms １ -C ４ Preferably, R is an aryl group substituted with one or more substituents selected from the group consisting of an alkyl group and an aryl group substituted with one or more iodine atoms. １ In one preferred embodiment, R contains an iodine atom (I atom). １ The substituent of the aryl group of R may be one or more iodine atoms. The number of iodine atoms serving as the substituent may be one or more, and the number of iodine atoms may be, for example, 1 to 10, preferably 1 to 7, and more preferably 1 to 5. １ The substituents of the aryl group are C substituted with one or more iodine atoms. １ -C ４ It can be an alkyl group. １ -C ４ The alkyl group is a group as described in the definition above, and is preferably, but not limited to, a methyl group and an ethyl group (provided that R １ C which can be a substituent of the aryl group １ -C ４ The alkyl group is substituted with one or more iodine atoms. Specific examples of such a substituent include an iodomethyl group and an iodoethyl group. １ The substituent of the aryl group in R may be an aryl group substituted with one or more iodine atoms. The aryl group that can be a substituent is a group as defined above, and is not limited thereto, but is preferably a phenyl group or a naphthyl group (provided that R １ The aryl group that can be a substituent of the aryl group in R is substituted with one or more iodine atoms.) A specific example of such a substituent is an iodophenyl group. １is an aryl group substituted with one or more iodine atoms (i.e., R １ It is more preferred that the substituents of the aryl group are one or more iodine atoms.

[0023] In another embodiment, R １ may be an aryl group substituted with one or more substituents selected from the group consisting of a gadolinium atom-containing group, a gold atom-containing group, a silver atom-containing group, and a platinum atom-containing group. The gadolinium atom-containing group refers to a gadolinium atom (Gd) itself, or a group containing a gadolinium atom and an appropriate number and type of ligands (e.g., monodentate ligands such as halogen atoms (e.g., chlorine atom, fluorine atom), and phosphines (e.g., triphenylphosphine), bidentate ligands such as acetonates (e.g., acetylacetonate), and polydentate ligands such as ethylenediaminetetraacetic acid). The gadolinium atom-containing group is a group containing a gadolinium atom (Gd), and is bonded to R directly at the gadolinium atom or via a ligand to which the gadolinium atom is bonded. １ For example, the gadolinium atom-containing group may be a group that is bonded to the aryl group of R １ Here, DTPA means diethylenetriaminepentaacetic acid. In gadopentetic acid, Gd ３＋ The gadolinium atom-containing group is not limited to the above-mentioned embodiment.

[0024] The gold atom-containing group refers to a group containing a gold atom (Au) itself, or a gold atom and an appropriate number and type of ligands (for example, monodentate ligands such as halogen atoms (e.g., chlorine atom, fluorine atom), and phosphines (e.g., triphenylphosphine), bidentate ligands such as acetonates (e.g., acetylacetonate), and polydentate ligands such as ethylenediaminetetraacetic acid). The gold atom-containing group is a group containing a gold atom (Au), and is bonded to R directly at the gold atom or via a ligand to which the gold atom is bonded. １ The silver atom-containing group may be a group that bonds to an aryl group of the formula (I). The silver atom-containing group refers to a group that contains a silver atom (Ag) itself or a silver atom and an appropriate number and type of ligands (for example, monodentate ligands such as halogen atoms (e.g., chlorine atoms, fluorine atoms), and phosphines (e.g., triphenylphosphine), bidentate ligands such as acetonates (e.g., acetylacetonates), and polydentate ligands such as ethylenediaminetetraacetic acid). The silver atom-containing group is a group that contains a silver atom (Ag), and is bonded to R directly at the silver atom or via a ligand to which the silver atom is bonded. １ The platinum atom-containing group may be a group that bonds to an aryl group of the formula (I). The platinum atom-containing group refers to a platinum atom (Pt) itself, or a group that includes a platinum atom and an appropriate number and type of ligands (for example, monodentate ligands such as halogen atoms (e.g., chlorine atoms, fluorine atoms), and phosphines (e.g., triphenylphosphine), bidentate ligands such as acetonates (e.g., acetylacetonate), and polydentate ligands such as ethylenediaminetetraacetic acid). The platinum atom-containing group is a group that includes a platinum atom (Pt), and is bonded to R directly at the platinum atom or via a ligand to which the platinum atom is bonded. １ These metal atom (gold, silver, or platinum atom)-containing groups may be, for example, groups in which the metal atom is bonded to one or more oxygen atoms or ligands containing oxygen atoms, and the metal atom is bonded to the aryl group of R １ Of course, the metal atom (gold, silver or platinum atom)-containing group is not limited to the above-mentioned embodiments.

[0025] R １The substitution in the aryl group may be one substitution with one type of substituent, a plurality of substitutions with one type of substituent, or a plurality of substitutions with a plurality of types of substituents. For example, the substituents may be an iodine atom and a C substituted with one or more iodine atoms. １ -C ４ The substituent may be composed of both an iodine atom and an alkyl group. Alternatively, for example, the substituent may be composed of both an iodine atom and a gadolinium atom-containing group. Alternatively, for example, the substituent may be composed of two or more, three or more, or four or more iodine atoms.

[0026] As noted above, in the compounds of the present invention, R １ The substituent of the aryl group always contains one or more atoms from the group consisting of iodine atom, gadolinium atom, gold atom, silver atom, and platinum atom (hereinafter, sometimes referred to as "atoms to be irradiated with X-rays") as atoms to be irradiated with X-rays. As will be described later, these atoms are atoms that can easily emit Auger electrons when irradiated with X-rays (also referred to as high-Z atoms). Therefore, the presence of these atoms allows for effective radiation therapy utilizing the Auger effect.

[0027] R １ The aryl group that is the basic skeleton of the formula (I) may further optionally contain a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ The compound may be substituted with one or more substituents selected from the group consisting of alkoxy groups. The presence of such substituents can provide effects such as changing the properties of the compound itself (e.g., stability and safety) or adjusting the binding affinity with other compounds (e.g., increasing the binding affinity between the compound and DNA). １ -C ４ Alkyl groups, and C １ -C ４ The alkoxy group is as defined above. A hydroxy group is more preferred as the substituent, which can increase the binding affinity, for example.

[0028] Ring A has a benzimidazole ring structure, and an X group is bonded to the imidazolyl group of this benzimidazole ring structure, and a R group is bonded to the phenyl group of the benzimidazole ring structure. ２ It has a chemical structure that bonds to

[0029] R ２ has a non-aromatic heterocyclic group as a basic skeleton containing one or more heteroatoms selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms, and at least one of the heteroatoms is a nitrogen atom. This non-aromatic heterocyclic group may optionally contain one or more C １ -C ４ The non-aromatic heterocyclic group may be unsubstituted. Preferably, the non-aromatic heterocyclic group is substituted with one or more C alkyl groups. １ -C ４ It is substituted with an alkyl group. ２ optionally one or more C １ -C ４ The non-aromatic heterocyclic group may be a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, optionally substituted with an alkyl group. The non-aromatic heterocyclic group is a group as defined above, and examples thereof include, but are not limited to, a piperidinyl group, a piperazinyl group, a pyrrolidinyl group, an imidazolidinyl group, a pyrazolidinyl group, a morpholinyl group, a thiomorpholinyl group, an oxazolidinyl group, and a thiazolidinyl group. The non-aromatic heterocyclic group is preferably a piperidinyl group, a piperazinyl group, a pyrrolidinyl group, an imidazolidinyl group, or a pyrazolidinyl group, more preferably a piperidinyl group or a piperazinyl group. The non-aromatic heterocyclic group may have a nitrogen atom bonded to the phenyl group in ring A, or a carbon atom bonded to the phenyl group in ring A. Possible substituents of C １ -C ４ Alkyl groups are as defined above, and include, for example, methyl and ethyl groups. １ -C ４When present, the alkyl group may be attached to a nitrogen atom or a carbon atom in the non-aromatic heterocyclic group. ２ However, the position on the phenyl group that bonds to the phenyl group of ring A is not particularly limited, and may be the position of a carbon atom adjacent to the carbon atom shared by the imidazole ring structure and the benzene ring structure in ring A, or may be the position of a carbon atom further adjacent to that carbon atom (the carbon atom adjacent to the carbon atom shared by the imidazole ring structure and the benzene ring structure), with the latter being more preferred.

[0030] X is a bond or C ２ an alkenylene group or C ４ It is an alkenylene group. When X is a bond, ring A is directly bonded to the phenyl group of the benzimidazole ring structure. In this case, a conjugated electron system is formed between the two benzimidazole ring structures. This allows the compound to have properties such as binding affinity with DNA. Also, when X is an alkenylene group, ring A is bonded to the phenyl group of the benzimidazole ring structure via the alkenylene group. C ２ Alkenylene group (—C═C—) and C ４ In the case of an alkenylene group (-C=C-C=C-), a conjugated electron system is formed between the two benzimidazole ring structures via the alkenylene group. This can provide the compound with properties such as binding affinity to DNA. X is more preferably a bond. The position on the phenyl group at which X is bonded to the phenyl group of the benzimidazole ring structure of formula (I) is not particularly limited, and may be the position of the carbon atom adjacent to the carbon atom shared by the imidazole ring structure and the benzene ring structure, or the carbon atom further adjacent to that carbon atom (the carbon atom adjacent to the carbon atom shared by the imidazole ring structure and the benzene ring structure), with the latter being more preferred.

[0031] In one preferred embodiment, R １ is an aryl group substituted with one or more iodine atoms, wherein the aryl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４and R ２ optionally one or more C １ -C ４ It is a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, which may be substituted with an alkyl group. Furthermore, X is more preferably a bond.

[0032] where R is an aryl group substituted with one or more iodine atoms. １ The maximum number of iodine atoms in the above formula (I) can be determined by taking into account the number of other optional substituents (the number of bonds of the aryl group). When the aryl group is a phenyl group, the phenyl group has five bonds available for substituents, excluding the bond to the benzimidazole ring structure, so the number of iodine atoms can be 1, 2, 3, 4, or 5 (maximum). However, in this case, the number of optional substituents is limited to 4 (maximum) or less, 3 or less, 2 or less, 1 or less, or 0, respectively (the total number of iodine atoms and the number of optional substituents is 5 or less). The greater the number of iodine atoms, the greater the Auger effect may be obtained.

[0033] In one preferred embodiment, the compound of formula (I) has formula (II): wherein, in the above formula, R ３ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with 1 or 2 hydroxy groups; and R ４ is a hydrogen atom or C １ -C ４ It is an alkyl group.

[0034] In comparison with the structure of formula (I), the structure of formula (II) is roughly the same as that of formula (I), in which X is a bond, the bonding positions of the two benzimidazole ring structures are specified, and the group R １ but a more limited group R ３ is replaced by the group R ２ is a group R ４ It has a chemical structure in which a piperazinyl group having as a substituent is substituted.

[0035] R３ is preferably a phenyl group substituted with one or two iodine atoms, which may be further optionally substituted with one hydroxy group, and more preferably a phenyl group substituted with one iodine atom, which may be further optionally substituted with one hydroxy group. ３ may be a phenyl group substituted with only one or two iodine atoms.

[0036] R ４ is C １ -C ４ An alkyl group is preferred, and a methyl group or an ethyl group is more preferred.

[0037] The compounds of formula (I) and formula (II) can be used in their original form (free form, not a salt). The compounds of formula (I) and formula (II) can also be used in pharmaceutical compositions in the form of their pharmaceutically acceptable salts. As used herein, "pharmaceutically acceptable" means that the compound is not toxic to a subject when administered (e.g., administered). The salt may be an acid addition salt or a base addition salt, but acid addition salts are preferred. This can be understood from the fact that multiple nitrogen atoms are contained in formula (I) and formula (II). Examples of acid addition salts include salts with inorganic acids such as hydrochloride, sulfate, and nitrate; and salts with organic acids such as acetate, sulfonate, and citrate. Acid addition salts also include salts with multiple acids (e.g., salts with multiple hydrochloric acids, specifically, dihydrochloride, trihydrochloride, etc.).

[0038] The compounds of formula (I) and formula (II) may contain isotopes of some atoms (e.g., deuterium ( ２ H), carbon 13 ( １３ C), nitrogen 15 ( １５ The compounds of formula (I) and formula (II) may be compounds substituted with N, etc. (isotopically labeled compounds), and such isotope-labeled compounds are also included in the compounds of the present invention. Furthermore, when the compounds of formula (I) and formula (II) have isomers (e.g., stereoisomers), the isomers (e.g., stereoisomers, for example, racemates, enantiomers, etc.) and mixtures thereof are also included in the compounds of the present invention.

[0039] Particularly preferred specific embodiments of the compounds of the present invention are shown below: Embodiment 1 A compound of formula (I) wherein R １ is an iodine atom, and C substituted with one or more iodine atoms １ -C ４ an aryl group substituted with one or more substituents selected from the group consisting of alkyl groups, wherein the aryl group is optionally substituted with one or more substituents selected from the group consisting of hydroxy groups, C １ -C ４ Alkyl groups, and C １ -C ４ alkoxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ A compound of formula (I) wherein R is a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, optionally substituted with an alkyl group. １ is an aryl group substituted with one or more iodine atoms, wherein the aryl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ alkoxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ A compound of formula (I) wherein R is a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, optionally substituted with an alkyl group. １ is an iodine atom, and C substituted with one or more iodine atoms １ -C ４ a phenyl group substituted with one or more substituents selected from the group consisting of alkyl groups, wherein the phenyl group is optionally substituted with one or more substituents selected from the group consisting of hydroxy groups, C １ -C ４ Alkyl groups, and C １ -C ４alkoxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ A compound of formula (I) wherein R is a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, optionally substituted with an alkyl group. １ is a phenyl group substituted with one or more iodine atoms, wherein the phenyl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ alkoxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ A compound of formula (I) wherein R is a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, optionally substituted with an alkyl group. １ is a phenyl group substituted with one or more iodine atoms, wherein the phenyl group optionally contains a hydroxy group, C １ -C ４ Alkyl groups, and C １ -C ４ alkoxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ A compound of formula (I) wherein R is a piperidinyl group or a piperazinyl group, optionally substituted with an alkyl group. １ is a phenyl group substituted with one or more iodine atoms, wherein the phenyl group may optionally be substituted with one or more hydroxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ Embodiment 7. A compound of formula (I) wherein R is a piperazinyl group, optionally substituted with an alkyl group. １is a phenyl group substituted with one or more iodine atoms, wherein the phenyl group may optionally be substituted with one or more hydroxy groups; X is a bond; and R ２ is a piperazinyl group optionally substituted with one or more methyl groups. １ is a phenyl group substituted with one or more iodine atoms, wherein the phenyl group may optionally be substituted with one or more hydroxy groups; X is a bond; and R ２ is a piperazinyl group substituted with a methyl group. １ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with 1 or 2 hydroxy groups; X is a bond; and R ２ is a piperazinyl group optionally substituted with one or more methyl groups. １ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with 1 or 2 hydroxy groups; X is a bond; and R ２ is a piperazinyl group substituted with a methyl group. １ is substituted with one or more iodine atoms １ -C ４ a phenyl group substituted with one or more alkyl groups, wherein the phenyl group may optionally be substituted with one or more hydroxy groups; X is a bond; and R ２ optionally one or more C １ -C ４ Embodiment 12. A compound of formula (I) wherein R is a piperazinyl group, optionally substituted with an alkyl group. １ is an iodine atom, and C substituted with one or more iodine atoms １ -C ４a phenyl group substituted with one or more substituents selected from the group consisting of alkyl groups, wherein the phenyl group is optionally substituted with one or more substituents selected from the group consisting of hydroxy groups, C １ -C ４ Alkyl groups, and C １ -C ４ and X is optionally substituted with one or more substituents selected from the group consisting of alkoxy groups; ２ or C ４ and R ２ optionally one or more C １ -C ４ A compound of formula (I) wherein R is a 6-membered non-aromatic heterocyclic group containing 1 to 4 nitrogen atoms, optionally substituted with an alkyl group. １ is a phenyl group substituted with one or more iodine atoms, wherein the phenyl group may optionally be substituted with one or more hydroxy groups; ２ or C ４ and R ２ optionally one or more C １ -C ４ A compound which is a piperazinyl group optionally substituted with an alkyl group.

[0040] Embodiment 14. A compound of formula (II) wherein R ３ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with 1 or 2 hydroxy groups; and R ４ is a hydrogen atom, a methyl group, an ethyl group, a propyl group, or an isopropyl group. ３ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with one hydroxy group; and R ４ is a hydrogen atom, a methyl group, or an ethyl group. ３ is a phenyl group substituted with 1 to 3 iodine atoms, wherein the phenyl group is optionally substituted with one hydroxy group; and R４ is a methyl group. ３ is a phenyl group substituted with one or two iodine atoms, wherein the phenyl group is optionally substituted with one hydroxy group; and R ４ is a methyl group. ３ is a phenyl group substituted with one iodine atom, wherein the phenyl group is optionally substituted with one hydroxy group; and R ４ is a methyl group. ３ is a phenyl group substituted with 1 to 3 iodine atoms; and R ４ is a methyl group. ３ is a phenyl group substituted with one iodine atom; and R ４ is a methyl group. ３ is a phenyl group substituted with 1 to 3 iodine atoms and one hydroxy group; and R ４ is a methyl group. ３ is a phenyl group substituted with one iodine atom and one hydroxy group; and R ４ is a methyl group.

[0041] Further specific examples of the compounds of formula (I) and formula (II) are shown below.

[0042] The above compounds are known compounds, commercially available, and commercially available. As described above, they are commercially available as "Ortho-iodo Hoechst 33258" (MW=534.4), "Meta-iodo Hoechst 33258" (MW=534.4), "Para-iodo Hoechst 33258" (MW=534.4), and "Hoechst 33342 analog 2 trihydrochloride" (MW=659.8), respectively (MW means molecular weight). These compounds are derivatives or analogs of the commonly used Hoechst compounds (compounds used in Hoechst staining) shown below.

[0043] Hoechst staining is a well-known method for staining DNA in cells. Hoechst dyes (Hoechst compounds) have a high binding affinity with DNA, and DNA stained with them emits blue fluorescence. A particularly preferred embodiment (described above) of the compound of the present invention has a structure similar to that of a conventional Hoechst compound, except for the incorporation of an iodine atom, and can be considered an "iodine-atom-containing Hoechst compound" (or an iodinated Hoechst compound). Similar to conventional Hoechst compounds, the iodine-atom-containing Hoechst compound has a high binding affinity with DNA and excellent staining properties (blue fluorescence). This effect can also be understood from the structure of the compound's basic skeleton (a structure having two benzimidazole rings). Due to its binding affinity with DNA, the atom to be irradiated with X-rays can be positioned in close proximity to the DNA. The binding affinity of the iodine-atom-containing Hoechst compound to DNA has also been confirmed by molecular docking simulations (see the Examples below). Furthermore, the staining ability can clarify the location of target cancer cells and their DNA. Furthermore, compared to other DNA-binding compounds (DNA-binding dyes), such as YOYO-1 and propidium iodide, Hoechst compounds are safer and can be used to treat actual diseases (administerable to humans and non-human animals). Therefore, iodine-atom-containing Hoechst compounds can be used as binding substances that target DNA.

[0044] Similarly, compounds (compounds of formula (I) or formula (II)) having a structure modified from the structure of the Hoechst compound within the scope of the structure defined by formula (I) or formula (II) can also exhibit high binding affinity to DNA, excellent staining properties (blue fluorescence), and high safety. That is, the compounds of the present invention can bind to DNA to position the atom to be irradiated with X-rays in close proximity to the DNA, and can clarify the location of cancer cells and DNA through staining properties. Furthermore, they can be safe when administered to living organisms (humans and non-human animals). The compounds of the present invention can be used with the intention of utilizing such properties as DNA binding. Meanwhile, as described below, iodine atoms, gadolinium atoms, gold atoms, silver atoms, and platinum atoms are atoms that can easily emit Auger electrons upon X-ray irradiation. The present invention is characterized by utilizing the high binding affinity to DNA of the compounds of formula (I) or formula (II) and the Auger effect of iodine atoms, gadolinium atoms, etc., thereby enabling highly effective Auger therapy.

[0045] Specific examples of compounds (analogs or derivatives) having structures obtained by modifying the structure of Hoechst compounds are shown below.

[0046] Compounds of formula (I) or (II) other than the commercially available compounds described above can be synthesized by methods such as derivatization from the above-mentioned Hoechst compounds and iodine atom-containing Hoechst compounds. Alternatively, such compounds can be synthesized by modifying known synthesis methods for Hoechst compounds. As a method for derivatization of Hoechst compounds, for example, the methods described in the following documents can be appropriately used.・Shrivastava, N., Naim, M. J., Alam, M. J., Nawaz, F., Ahmed, S., & Alam, O. (2017). Benzimidazole Scaffold as Anticancer Agent: Synthetic Approaches and Structure-Activity Relationship. Archiv der Pharmazie, 350(6), 10.1002 / ardp.201700040. https: / / doi.org / 10.1002 / ardp.201700040 ・Di Gioia, M. L., Cassano, R., Costanzo, P., Herrera Cano, N., Maiuolo, L., Nardi, M., Nicoletta, F. P., Oliverio, M., & Procopio, A. (2019). Green Synthesis of Privileged Benzimidazole Scaffolds Using Active Deep Eutectic Solvent. Molecules (Basel, Switzerland), 24(16), 2885. https: / / doi.org / 10.3390 / molecules24162885 ・Balagurumoorthy, P., Xu, X., Wang, K., Adelstein, S. J., & Kassis, A. I. (2012). Effect of distance between decaying (125)I and DNA on Auger-electron induced double-strand break yield. International journal of radiation biology, 88(12), 998-1008. https: / / doi.org / 10.3109 / 09553002.2012.706360。

[0047] The present invention also relates to nanoparticles comprising the compound represented by formula (I) or a pharmaceutically acceptable salt thereof and a porous silica carrier. The present invention also relates to iodine-containing nanoparticles (IPOs). For details of IPOs, please also refer to the aforementioned Non-Patent Document 2.

[0048] Specifically, the nanoparticles of the present invention may be particles in which the compound represented by the above formula (I) or a pharmaceutically acceptable salt thereof is supported on a porous silica carrier.Preferred embodiments of the compound represented by formula (I) are as described above, and all of the descriptions also apply to the compound in the nanoparticles.For example, the compound represented by formula (I) is preferably a compound represented by formula (II), and more preferably the iodine atom-containing Hoechst compounds described above (particularly the four compounds).

[0049] The porous silica carrier is a carrier made of porous silica (a material capable of supporting a compound), and the following porous silica can be used: ２ ) as a main component and having many pores. Porous silica may be in the form of particles. Porous silica may constitute the main component (the component with the largest amount) of the components constituting nanoparticles. Porous silica has the property of increasing its specific surface area due to the pores. Porous silica may be in the form of nanoparticles. In this specification, nanoparticles refer to nano-sized particles. Porous silica nanoparticles can function as a support and / or substrate for holding the compound of the present invention. In this specification, nanosize generally refers to 10 nm or more and 500 nm or less, and a preferred nanosize is 40 nm or more and 400 nm or less.

[0050] In the nanoparticles of the present invention, the compound of the present invention may be present on the surface of the porous silica (the outer surface of the particle and the surface inside the pores). As a mode of support by porous silica, the compound of the present invention may be bonded to the porous silica. The bond between the compound and the porous silica may be a chemical bond or an electrical bond. Silica may have silanol groups (Si—OH) or groups derived from silanol groups on its surface. The compound may be bonded to the silica via the silanol groups. On the other hand, the compound of the present invention has a nitrogen atom, and the nitrogen atom may have an unshared electron pair. The compound may be bonded to the silica via the unshared electron pair. Alternatively, silica and the compound of the present invention may be electrically attracted to each other and bonded (attached) by electrostatic force. The mode of support of the compound by porous silica is not particularly limited, as long as it meets the purpose and intent of the present invention.

[0051] In a preferred embodiment, the porous silica is mesoporous silica. Mesoporous silica has a large number of pores, typically with a pore size (pore diameter) of 2 to 50 nm. Mesoporous silica has a larger specific surface area and can more efficiently retain the above-mentioned compounds. Furthermore, as will be described later, mesoporous silica has the advantage of being easily taken up by cells. Unless otherwise specified, all porous silicas mentioned in this specification can be replaced with mesoporous silica.

[0052] In a preferred specific embodiment, the nanoparticles (silica carriers) may be biodegradable mesoporous silica nanoparticles. An example of a method for synthesizing biodegradable mesoporous silica is described in Production Example 1 (paragraph

[0091] ) of the Examples section below. Biodegradable mesoporous silica can decompose over time in vivo. Examples of the mechanism of decomposition include enzymatic reactions. Biodegradable mesoporous silica allows the nanoparticles to be decomposed in the body and their components excreted, enabling safer treatments and the like. Biodegradable mesoporous silica can be obtained by using a silane compound having a biodegradable structure as a raw material. Examples of the biodegradable structure include bonds represented by S-S and / or S-S-S-S. For example, bis[3-(triethoxysilyl)propyl]tetrasulfide is a silane compound having an S-S-S-S bond between two Si atoms, and when this compound is incorporated into the structure of mesoporous silica, a structure containing an S-S-S-S bond between two Si atoms can be formed in the mesoporous silica. S-S and S-S-S-S bond strength is relatively weak, and therefore the mesoporous silica is easily biodegradable.

[0053] In the nanoparticles, the ratio of the compound of the present invention to the porous silica (compound of the present invention / porous silica) may vary depending on the type of atom to be irradiated with X-rays and is not particularly limited. For example, the ratio may be within a range of 0.0001 to 1 by weight, and this ratio is preferably 0.001 to 0.5, and more preferably 0.001 to 0.1. The ratio of the compound of the present invention to the porous silica may depend on the ratio of the atom to be irradiated with X-rays, such as an iodine atom, to the porous silica. The ratio of the atom to be irradiated with X-rays to the porous silica (atom to be irradiated with X-rays / porous silica) is not particularly limited. For example, the ratio may be within a range of 0.00001 to 1 by weight, and this ratio is preferably 0.0001 to 0.5, and more preferably 0.0001 to 0.1. It is particularly preferred that this ratio (atom to be irradiated with X-rays / porous silica) be 0.0005 or greater. Here, the weight of the atom to be irradiated with X in the nanoparticle can be obtained by analyzing the atom to be irradiated with X in the nanoparticle by inductively coupled plasma atomic emission spectroscopy (ICP-AES).

[0054] Synthesized porous silica can be used. An example of a method for synthesizing porous silica is described in Production Example 1 (paragraph

[0091] ) of the Examples section below. The method for synthesizing porous silica is not particularly limited, and known methods can be used. For example, a precursor substance for forming porous silica (e.g., an organosilane compound, specifically, an alkylalkoxysilane, etc.) can be condensed in the presence of a template compound for forming pores (e.g., a tetraalkoxysilane, specifically, tetraethoxysilane, etc.), and the template compound can be removed from the silica particles produced by the condensation to obtain porous silica particles (see, for example, International Publication No. 2021 / 060498). For example, a specific example of a precursor substance for porous silica is 1,2-bis(triethoxysilyl)ethane. Furthermore, a specific example of a template compound is cetyltrimethylammonium bromide (CTAB).

[0055] The compound of the present invention can be supported on porous silica by, but not limited to, mixing the compound of the present invention with porous silica in an appropriate solvent (e.g., adding porous silica particles to a solution containing the compound and suspending them). Upon contact between the compound and porous silica, the compound is supported on the porous silica via the aforementioned bonding mechanism (e.g., chemical or electrical bonding). Mixing can be performed using a rotary mixer. After mixing, the mixture can be centrifuged, the supernatant removed, and the mixture washed several times with a washing solution (e.g., water) to remove excess compound, thereby obtaining nanoparticles of the present invention in which the compound of the present invention is supported on porous silica. Iodine-containing compounds other than IH (e.g., propidium iodide) can also be supported by the above method and used for Auger therapy.

[0056] Here, the nanoparticles of the present invention are characterized by their easy uptake by cells, particularly cancer cells. It has been confirmed that when nanoparticles are brought into contact with cells, they enter the cells. It is hypothesized that cellular uptake occurs via the endocytic mechanism involving endosomal vesicles, and that this vesicular transport can deliver nanoparticles to lysosomes located adjacent to the cell nucleus. Of course, the present invention is not limited by this hypothesis. Therefore, the nanoparticles of the present invention can deliver the compounds of the present invention to the vicinity of the cell nucleus.

[0057] The nanoparticles of the present invention are also useful for targeting solid tumors, such as tumors. When nanoparticles are administered to humans and animals, they may accumulate in solid tumors. Therefore, nanoparticles have at least two advantages: delivery to solid tumors and uptake by cancer cells. Furthermore, after uptake by cancer cells, the compounds of the present invention (compounds of formula (I), specifically, for example, iodine-containing Hoechst compounds) are released from the nanoparticles under conditions such as the low pH of intracellular endosomes, and then translocate to the cell nucleus and bind to DNA within the nucleus. In other words, the nanoparticles of the present invention have the ability to accumulate in tumors and deliver X-ray-irradiated atoms, such as iodine atoms, to DNA in cancer cells. Furthermore, such targeting can prevent side effects from systemic administration (see Figure 15).

[0058] As described above, the compound of the present invention has a chemical structure similar to that of the dye in Hoechst staining, has excellent binding ability to cell nuclei, and also contains atoms that emit Auger electrons when irradiated with X-rays (atoms that are subject to X-ray irradiation), such as iodine atoms. Therefore, the compound of the present invention is placed near cancer cells by the nanoparticles and is efficiently taken up into the cell nuclei, allowing for extremely effective Auger therapy.

[0059] Pharmaceutical Compositions According to one aspect of the present invention, the present invention relates to pharmaceutical compositions for use in radiation therapy, comprising the compound of the present invention and / or the nanoparticles of the present invention. The pharmaceutical composition comprises the compound of the present invention described above and a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier may be liquid or solid. The carrier may be an excipient, diluent, adjuvant, etc. Liquid carriers include, for example, water and organic solvents. Organic solvents include, but are not limited to, alcoholic solvents such as methanol and ethanol, ketone solvents such as acetone, ether solvents such as diethyl ether, and ester solvents such as ethyl acetate. Solid carriers include, for example, lactose, crystalline cellulose, and starch. Note that the carriers described here are merely examples, and known carriers can be used appropriately in pharmaceutical compositions.

[0060] In certain embodiments, the pharmaceutical composition comprises the nanoparticles of the present invention and a pharmaceutically acceptable carrier, the nanoparticles carrying the compound of the present invention, and the pharmaceutical composition comprises the compound of the present invention via the nanoparticles carrying the compound of the present invention.

[0061] The pharmaceutical composition can be irradiated with X-rays as the radiation for irradiation. As described above, X-ray irradiation causes the atom (e.g., iodine atom) in the compound of the present invention to emit electrons due to the Auger effect. The pharmaceutical composition of the present invention makes it easier for the compound to reach the target site.

[0062] The pharmaceutical composition can be used to treat solid cancers or to suppress the growth or proliferation of solid cancers. As described above, Auger electrons released from atoms (such as iodine atoms) irradiated with X-rays can destroy cancer cells. Therefore, the pharmaceutical composition is useful for treating solid cancers or suppressing the growth or proliferation of solid cancers.

[0063] Solid cancers include, but are not limited to, brain cancer, lung cancer, ovarian cancer, gastrointestinal cancer, osteosarcoma, or head and neck cancer.

[0064] Solid cancers may be in a hypoxic state. The present invention makes it possible to efficiently treat solid cancers in a hypoxic state. While many existing X-ray irradiation methods utilize reactive oxygen, under hypoxic conditions, sufficient therapeutic effects may not be achieved because reactive oxygen is not generated sufficiently. However, the radiotherapy according to the present invention does not rely on reactive oxygen but utilizes the Auger effect, and has been confirmed to be able to effectively treat cancer even in a hypoxic state (see the Examples below). Therefore, the present invention makes it possible to effectively treat cancer even under hypoxic conditions where existing radiotherapy is difficult to achieve.

[0065] The pharmaceutical composition can be administered by an appropriate administration method. The administration method may be oral administration or parenteral administration. Examples of parenteral administration include injection (intravenous injection, subcutaneous injection, intramuscular injection, etc.), suppository administration, and topical application (cutaneous application, mucosal application), etc. The dose of the pharmaceutical composition for radiotherapy is not particularly limited, but is preferably an amount that can produce the Auger effect when the compound of the present invention is irradiated with X-rays.

[0066] Radiation Therapy (X-ray Irradiation) and Cancer Cell Destruction Radiation therapy (X-ray irradiation) according to the present invention is described below. This description is applicable to the compound, nanoparticles, and pharmaceutical composition of the present invention described above. According to one aspect of the present invention, the present invention relates to X-ray irradiation of the compound of the present invention (including when supported on nanoparticles) and the resulting cancer cell destruction. Specifically, X-rays can be irradiated targeting an atom to be irradiated with X-rays (specifically, an iodine atom, a gadolinium atom, a gold atom, a silver atom, or a platinum atom). By irradiating with X-rays capable of exciting the K-shell electrons of the atom to be irradiated with X-rays, Auger electrons can be emitted from the atom to be irradiated with X-rays (e.g., an iodine atom).

[0067] In general, Auger electrons can damage DNA and other cellular components. The atoms irradiated with X-rays are suitable for emitting Auger electrons. However, the range of Auger electrons is limited, and previous studies have not adequately demonstrated the cell-destructive effect of Auger electrons. The present invention has the advantage of using a compound that combines a specific atom (the atom irradiated with X-rays) with a Hoechst-like structure, positioning the atom irradiated with X-rays near the nucleus (especially DNA), and efficiently destroying cancer cells with Auger electrons.

[0068] When an atom to be irradiated, placed near a cell nucleus, is irradiated with X-rays capable of exciting the K-shell electrons of the atom, Auger electrons are emitted from the atom, and these electrons can damage the cell. The cell nucleus and its surroundings contain important cellular functions, including organelles, and the Auger electrons can damage these, damaging the cell efficiently and effectively. This cellular damage can destroy or kill cancer cells.

[0069] The X-rays capable of exciting the K-shell electrons of atoms irradiated with X-rays vary depending on the atom, and each atom irradiated with X-rays has its own specific energy level and / or wavelength. The X-rays may have an energy capable of exciting K-shell electrons. The X-rays may also have a wavelength capable of exciting K-shell electrons. Based on "International Tables for Crystallography C, Table 4.2.2.4 Theoretical Calculations" of the International Union of Crystallography (IUCr), the K-shell electron excitation wavelengths (corresponding X-ray wavelengths) and K-shell electron excitation energies of I, Gd, Au, Ag, and Pt are as shown in Table 1 below. In iodine atoms, X-rays with an energy of 33.2 keV are suitable for exciting K-shell electrons. This is because the K-shell electron excitation energy of iodine atoms is 33.2 keV (more precisely, 33.17 keV). However, even if the energy level is not optimal, K-shell electrons may be excited, and it has been confirmed that X-rays with an energy of 33.4 keV are also effective in iodine atoms. Therefore, by irradiating iodine atoms with X-rays with an energy of 33.2 keV or 33.4 keV, Auger electrons can be emitted from the iodine atoms. The wavelength of X-rays corresponding to 33.17 keV, which can excite the K-shell electrons of iodine atoms, is 0.03737 nm. Similarly, for gadolinium atoms, gold atoms, silver atoms, and platinum atoms, the energy (or wavelength) of X-rays suitable for exciting K-shell electrons is 50.25 keV for gadolinium atoms, 80.73 keV for gold atoms, 25.52 keV for silver atoms, and 78.4 keV for platinum atoms. Therefore, by irradiating these atoms with X-rays of the respective energies (or wavelengths), Auger electrons can be emitted. Note that the K-shell electron excitation energy is also referred to as the K-shell absorption edge energy, and the K-shell electron excitation wavelength is also referred to as the K-shell absorption edge wavelength.

[0070] The X-rays may be X-rays having a spectral peak preferably at E -0.5 keV or higher, more preferably E -0.3 keV or higher, and even more preferably E -0.1 keV or higher, relative to the K-shell electron excitation energy E of the atom to be irradiated with X-rays. The X-rays may be X-rays having a spectral peak preferably at E +0.8 keV or lower, more preferably E +0.7 keV or lower, even more preferably E +0.6 keV or lower, and even more preferably E +0.5 keV or lower, relative to the K-shell electron excitation energy E of the atom to be irradiated with X-rays. For example, the X-rays may have a spectral peak at any one of E -0.5 keV or higher, E -0.3 keV or higher, E -0.1 keV or higher, or E -0.05 keV or higher, and also have a spectral peak at any one of E +0.8 keV or lower, E +0.7 keV or lower, E +0.6 keV or lower, or E +0.5 keV or lower. In particular, the X-rays are preferably X-rays having a spectral peak in the range of E - 0.1 keV or more and E + 0.5 keV or less, where E is the K-shell electron excitation energy of the atom to be irradiated with X-rays. As described above, E differs for each atom to be irradiated with X-rays, and Auger electrons can be efficiently emitted by irradiating each atom with X-rays at an energy corresponding to the atom to be irradiated with X-rays. Furthermore, the Auger effect may be obtained even at energies close to the K-shell electron excitation energy E of the atom to be irradiated with X-rays. In particular, the Auger effect may be obtained at energies slightly higher than the K-shell electron excitation energy E, and therefore, the above-mentioned energy of E + 0.5 keV or less is preferred. On the other hand, the Auger effect is difficult to obtain at energies lower than the K-shell electron excitation energy E, and therefore, the above-mentioned energy of E - 0.1 keV or more is preferred. From the above, it can be understood that, in the case of iodine atoms (E = 33.2 keV), an energy of 33.1 to 33.7 keV is preferred.

[0071] The X-rays are preferably monochromatic X-rays or characteristic X-rays. Furthermore, the X-rays are more preferably monochromatic X-rays. Monochromatic X-rays refer to X-rays with an extremely narrow range of energy. In this specification, the energy E １ When we say monochromatic X-rays, the spectral peak of the monochromatic X-rays has energy E １For example, １ Energy below -ΔE keV and E １ It does not include X-rays with energies of +ΔE keV or more, where ΔE≦E×10 －３ Characteristic X-rays are emitted as X-rays due to the excess energy generated when an electron transitions from the outer shell to a vacancy created by excitation of the inner core. The energy of characteristic X-rays is determined by the energy difference between the inner and outer shell levels and is a material-specific value. Therefore, it is difficult to extract monochromatic X-rays of any energy. Unlike continuous X-rays (or white X-rays), monochromatic X-rays and characteristic X-rays have a narrow energy range, which effectively reduces damage to normal cells and tissues caused by X-ray irradiation. Monochromatic X-rays are particularly effective in this regard. Monochromatic X-rays can be extracted by monochromatizing white X-rays generated by a synchrotron radiation facility or white X-rays generated by an X-ray generator using a spectroscope. Characteristic X-rays can be extracted by monochromatizing only the characteristic X-rays contained in white X-rays generated by an X-ray generator using a spectroscope. However, the method of generating monochromatic X-rays and characteristic X-rays is not limited to this.

[0072] As shown in the examples below, when iodine atoms were used as the target of X-ray irradiation, cancer cells were effectively destroyed when irradiated with X-rays of 33.2 keV energy. Furthermore, when irradiated with X-rays of 33.4 keV energy, cancer cells were still destroyed, although the level was lower than that of 33.2 keV. On the other hand, when irradiated with X-rays of 33.0 keV energy, there was almost no destruction of cancer cells. This dramatic difference in the effects of 33.0 keV and 33.2 keV X-rays suggests the idea that Auger electrons exert a cell-destructive effect. X-ray sensitization was observed when X-rays obtained from commercially available X-ray irradiation equipment for experimental and research use were used.

[0073] According to one aspect of the present invention, the present invention relates to a method for treating solid cancer or inhibiting the growth or proliferation of solid cancer. This method comprises irradiating the above-mentioned compound, nanoparticles, or pharmaceutical composition taken into the body of a subject with X-rays to destroy cancer cells. X-rays that can excite K-shell electrons in the subject can be used. The mechanism of cancer cell destruction is as explained above. Subjects include patients. While the method is applicable to humans, it can also be applied to non-human animals.

[0074] The X-ray irradiation time may vary depending on the severity of the disease to be treated, the patient's tolerance to X-ray irradiation, etc., but is not particularly limited and can be, for example, 1 minute or more, 2 minutes or more, 3 minutes or more, or 5 minutes or more. The X-ray irradiation time is also not particularly limited and can be, for example, 240 minutes or less, 180 minutes or less, 150 minutes or less, or 120 minutes or less. For example, the X-ray irradiation time may be 10 minutes, 30 minutes, 60 minutes, 90 minutes, etc.

[0075] As shown in the examples below, the compounds of the present invention have excellent targeting properties for solid cancers and can easily penetrate the nuclei of cancer cells. Furthermore, by irradiating the compounds of the present invention with X-rays, Auger electrons can be used to destroy cancer cells. Therefore, this method can effectively treat solid cancers or effectively inhibit the growth or proliferation of solid cancers.

[0076] The present invention will be further explained below with reference to examples, but it goes without saying that the present invention is not limited to these examples.

[0077] Materials (Compounds) The following commercially available iodine-containing Hoechst compounds were obtained as iodine-containing Hoechst compounds: Hoechst 33342 analog 2 trihydrochloride (manufactured by MedChemExpress), meta-iodo Hoechst 33258 (manufactured by MedChemExpress), para-iodo Hoechst 33258 (manufactured by MedChemExpress), ortho-iodo Hoechst 33258 (manufactured by MedChemExpress).

[0078] Test Example 1 Docking Simulation of Iodine-Containing Hoechst Compounds with DNA A docking simulation of iodine-containing Hoechst compounds with DNA was performed using a molecular docking technique. Four iodine-containing Hoechst compounds were used: "Hoechst 33342 analog 2," "meta-iodo Hoechst 33258," "para-iodo Hoechst 33258," and "ortho-iodo Hoechst 33258." To obtain DNA for docking, a typical B-form DNA was constructed using the software "3DNA" (see Lu XJ, Olson WK. 3DNA: a software package for the analysis, rebuilding, and visualization of three-dimensional nucleic acid structures. Nucleic Acids Res. 2003 Sep 1;31(17):5108-21. doi: 10.1093 / nar / gkg680). The docking simulation software used was AutoDock4 (see Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS, Olson AJ. AutoDock4 and AutoDockTools4: Automated docking with selective receptor flexibility. J Comput Chem. 2009 Dec;30(16):2785-91. doi: 10.1002 / jcc.21256). The iodine-containing Hoechst compound ligand was docked to DNA with full flexibility, and from 20 predicted binding states, the one that bound to the target binding site and had the lowest estimated binding free energy (ΔG) was selected.

[0079] Figures 1 and 2 and Table 2 show the results of the docking simulation (images and calculated values).

[0080] FIG. 1 is a diagram viewed along the helix axis (top view), and FIG. 2 is a diagram viewed perpendicular to the helix axis (side view). As shown in these figures and tables, molecular docking simulations confirmed that all four of the above-mentioned iodine-atom-containing Hoechst compounds have high binding affinity to DNA. This suggests that the iodine-atom-containing Hoechst compounds can position the atom to be irradiated with X-rays (iodine atom) in close proximity to DNA. Of the four iodine-atom-containing Hoechst compounds, "Hoechst 33342 analog 2" had the smallest estimated binding free energy (ΔG) (-13.56 kcal / mol), indicating high binding affinity with DNA, and that the iodine was close to the center of DNA (5 Å). Therefore, of the four compounds, "Hoechst 33342 analog 2" is presumed to be the most advantageous. Therefore, in the following experiments, "Hoechst 33342 analog 2" (hereinafter also referred to as "I-Hoechst") was mainly investigated as an iodine atom-containing Hoechst compound. However, when using "Hoechst 33342 analog 2", its trihydrochloride salt, "Hoechst 33342 analog 2 trihydrochloride" (hereinafter also referred to as "I-Hoechst.3HCl") was used due to its easy availability.

[0081] Test Example 2: Uptake of Iodine-Containing Hoechst Compound into Cancer Cells Human ovarian cancer cells, OVCAR8, which express green fluorescent protein (GFP), were used as cancer cells. OVCAR8 cancer cells were cultured on a 100 mm diameter culture dish in RPMI 1640 medium supplemented with 10% inactivated FBS and 1% penicillin / streptomycin. Furthermore, an iodine-containing Hoechst compound (I-Hoechst·3HCl) (0.1 mg, 151.57 nmol) was dissolved in water (0.15 mL) to prepare a solution containing the iodine-containing Hoechst compound (amount of Hoechst compound: 1 μmol / mL). This solution was added to the culture medium in an amount such that the amount of Hoechst compound in the culture medium became 50 nmol / mL. The cells were then incubated for 1 hour at 25°C for 1 hour at 25°C. ２The cells were incubated in an incubator at 37°C for 24 hours. Then, the medium was removed and the cells were washed. The cells were observed under a confocal microscope. Cell nuclei can be detected by Hoechst compound dye staining (blue coloring) and GFP expression (green fluorescence).

[0082] Figure 3 shows images of microscopic observation. This figure shows a representative cell. In this figure, A represents the fluorescent color (green) emitted by the cancer cell OVCAR8, B represents the stained area (blue) where the Hoechst compound functions as a dye, and C represents a processed image obtained by superimposing A and B. Figures 3A to 3C confirm that the Hoechst-stained area is located inside the cancer cell, and it is clear that the Hoechst compound is taken up into the cell, transported into the cell nucleus, and localized in the DNA.

[0083] Test Example 3: Incorporation of Hoechst Compound into Tumor Spheroids Human ovarian cancer cells OVCAR8 expressing green fluorescent protein (GFP) were used as cancer cells, and tumor spheroids (hereinafter also referred to as "spheroids") were formed from these cancer cells. OVCAR8 cancer cells were cultured on a 100 mm diameter culture dish in RPMI 1640 medium supplemented with 10% inactivated FBS and 1% penicillin / streptomycin. 5.0 × 10 cells were used to form spheroids. ３ OVCAR8 cells were seeded onto a PrimeSurface 96U culture plate (MS-9096U, Sumitomo Bakelite Co., Ltd.). ２ The cells were cultured in an incubator at 37°C for 7 days. Here, because the cells could not adhere to the hydrophilic plate surface, they gathered at the bottom of the well, where three-dimensional spheroids were formed. This resulted in spheroids with a diameter of approximately 500 μm (estimated cell count of approximately 100,000 cells). Next, an iodine-containing Hoechst compound (I-Hoechst·3HCl) (0.1 mg, 151.57 nmol) was dissolved in water (1.5 mL) to prepare a solution containing the iodine-containing Hoechst compound (amount of Hoechst compound: 100 nmol / mL). This solution was added to the spheroids in an amount such that the amount of Hoechst compound was 5 nmol / mL, and CO２ The cells were incubated in an incubator at 37°C for a predetermined period of time. After incubation, the spheroids were collected in an Eppendorf tube. The spheroids were washed with ice-cold PBS and fixed with 4% paraformaldehyde at 4°C overnight. The spheroids were washed with ice-cold PBS and treated with 99.8% methanol at -80°C for 30 minutes. The spheroid samples obtained as described above were observed under a confocal microscope.

[0084] Figure 4 shows the results (images) of microscopic observation of spheroids. In the horizontal columns, the image labeled "GFP" shows the results of fluorescent coloring (green), and the image labeled "I-Hoechst" shows the results of dye staining with the Hoechst compound (blue). In the vertical columns, "No Loading" indicates no treatment with the Hoechst compound solution (i.e., control), and 1, 24, and 96 indicate the incubation times with the Hoechst compound (1 hour, 24 hours, and 96 hours, respectively). As shown in this figure, the Hoechst compound was efficiently incorporated into the spheroids and was evenly distributed throughout the spheroids after 24 hours. Interestingly, by operating the confocal microscope, it was confirmed that the Hoechst compound reached the interior of the spheroids. In other words, it was confirmed that the Hoechst compound reached not only the cells near the surface of the spheroids, but also the center of the spheroids. This is contrary to initial expectations. Such high permeability is considered to be an advantageous feature of Hoechst compounds.

[0085] Experimental Example 4: Irradiation of Iodine-Containing Hoechst Compounds with Monochromatic X-Rays. Monochromatic X-Ray Irradiation System Setup. Monochromatic X-ray irradiation was performed at beamline BL14B1 of the SPring-8 synchrotron radiation facility in Sayo-cho, Sayo-gun, Hyogo Prefecture, Japan. Figure 5 shows an overview of the irradiation system setup. First, white X-rays generated by the SPring-8 bending magnet were guided into a fixed-exit position double-crystal monochromator with a silicon 311 crystal to generate a monoenergetic X-ray beam (monochromatic X-rays). The SPring-8 storage ring was operated in top-up mode with a constant storage current of 100 mA, allowing for negligible fluctuations in X-ray intensity over time. The X-ray beam shape was adjusted using horizontal and vertical transport channel (TC) slits. The X-ray beam size at the sample position was 0.7 mm high x 1.4 mm wide. This is sufficient to cover a spheroid measuring 0.5 mm x 0.5 mm. During the experiment, the intensity of the X-ray beam was monitored by two ion chambers placed on the optical axis. The transmitted X-rays were monitored by a CCD camera to adjust the sample position. Figure 6 shows an overview of the spheroid sample area. The spheroid was placed at the bottom of a tube placed on a sample rack.

[0086] Test Example 5: Destruction of Spheroids by Monochromatic X-ray Irradiation. Spheroids were incubated with an iodine-containing Hoechst compound (I-Hoechst) (24 hours) using the same method as in Test Example 3 to obtain spheroids incorporating the Hoechst compound. These spheroids were placed in tubes, which were then placed in the sample rack of an X-ray irradiator. The sample rack was set up to be movable on an XYZ stage, allowing the experimenter to move the sample along the optical axis and irradiate it with X-rays without entering the experimental hutch. Furthermore, upon completion of X-ray irradiation of one sample, the sample rack moved and the irradiation position automatically moved to the next sample, enabling a series of X-ray irradiations to be performed automatically (see Figure 6). The sample position was confirmed using an optical microscope and laser before irradiation. Furthermore, X-ray irradiation was monitored using a CCD camera. Due to the high energy of the X-rays, it was not possible to observe the absorption contrast of the spheroids or tubes using the CCD camera. The sample position was monitored by obtaining refraction-enhanced X-ray images of the tubes. The photon flux at the sample is 3.9 × 10 using the SPECTRA code. ８ The spheroids prepared as described above were irradiated with monochromatic X-rays (energy: 33.0 keV, 33.2 keV, or 33.4 keV) for 30 minutes. After X-ray irradiation, the spheroids were heated in a CO ２ After incubation at 37°C for 3 days, the spheroids were observed under a confocal microscope (visible and fluorescent). For comparison, a non-iodine-containing Hoechst compound (Hoechst 33258, Dojindo Laboratories) was used, and the same treatments (incubation with spheroids, monochromatic X-ray irradiation, and post-irradiation incubation) were performed.

[0087] Figure 7 shows the results (images) of microscopic observation of spheroids after incubation following monochromatic X-ray irradiation. In the horizontal columns, the images labeled "BF" represent bright-field image results, while the images labeled "GFP" represent fluorescent (green) color observation results. The vertical columns represent the energy values ​​of monochromatic X-rays. As shown in this figure, when spheroids incorporating an iodine-atom-containing Hoechst compound were irradiated with monochromatic X-rays with an energy of 33.2 keV, the presence of spheroids was not confirmed in either the bright-field image (BF) or the GFP image. Therefore, it was demonstrated that spheroids incorporating an iodine-atom-containing Hoechst compound were destroyed and broken apart when irradiated with monochromatic X-rays with an energy of 33.2 keV for 30 minutes. Furthermore, a reduction in spheroids was observed with 33.4 keV monochromatic X-rays, and although to a lesser extent than with 33.2 keV, the spheroid destruction effect was observed. On the other hand, when irradiated with monochromatic X-rays at 33.0 keV, the spheroids did not decrease in size, and no spheroid destruction effect was observed. Furthermore, when spheroids that incorporated Hoechst compounds that did not contain iodine were irradiated with monochromatic X-rays at either energy, no spheroid destruction was observed. The above energy corresponds to the K-shell electron excitation energy of iodine atoms, suggesting that the cells were destroyed by the Auger effect.

[0088] To investigate the mechanism of spheroid destruction described above, we performed a γH2AX assay on spheroids immediately after 30 minutes of monochromatic X-ray irradiation (before incubation) to confirm the state of the DNA double strand. In the γH2AX assay, red fluorescence is emitted when double-strand breaks occur. The γH2AX assay was performed using a commercially available kit according to the kit's instructions. Figure 8 shows the results (images) of microscopic observation of spheroids immediately after monochromatic X-ray irradiation. In the horizontal columns, the image labeled "BF" represents the bright-field image, the image labeled "γH2AX" represents the fluorescent coloration (red) due to γH2AX, and the image labeled "GFP" represents the fluorescent coloration (green). The vertical columns represent the energy values ​​of the monochromatic X-rays (where "0" means no irradiation). As shown in this figure, when spheroids incorporating an iodine-containing Hoechst compound were irradiated with monochromatic X-rays at 33.2 keV, red fluorescence was observed, indicating double-strand breaks in the DNA (note that green fluorescence (GFP) from cancer cells was observed before incubation). Furthermore, 33.4 keV monochromatic X-rays also induced double-strand breaks in the DNA, although to a lesser extent than 33.2 keV. On the other hand, no double-strand breaks in the DNA were observed with 33.0 keV monochromatic X-rays or without monochromatic X-ray irradiation. This result indicates that Auger electrons generated on the DNA can cause double-strand breaks in the DNA, destroying the spheroids. Taking the results of Figures 7 and 8 together, it is believed that monochromatic X-ray irradiation causes double-strand breaks in the DNA, and that these DNA breaks ultimately lead to the destruction of the spheroids. As described above, the effect of X-ray irradiation on spheroid destruction was most pronounced at 33.2 keV, the K-edge energy of iodine atoms, but not at the slightly lower energy level of 33.0 keV. It was also found that the slightly higher energy level of 33.4 keV was effective, but less so than 33.2 keV. This demonstrates that the use of monochromatic X-rays with energies near the K-edge energy of iodine atoms is most effective.

[0089] Test Example 6: Safety of Iodine-Containing Hoechst Compounds Hoechst compounds (ordinary compounds not containing iodine atoms) are generally considered to have low toxicity. The low toxicity of iodine-containing Hoechst compounds was also confirmed by the following test. Ovarian cancer OVCAR8 cells were incubated with various amounts of iodine-containing Hoechst compounds (I-Hoechst·3HCl). Cytotoxicity was examined by the LDH (lactate dehydrogenase) method using a commercially available kit. Figure 9 shows the results of the cytotoxicity test. As shown in this figure, the iodine-containing Hoechst compounds showed no toxicity at concentrations of 20 μM or less.

[0090] Test Example 7: Destruction of Hypoxic Spheroids The effect of spheroid destruction under hypoxic conditions was examined by the following test. Spheroids were incubated with an iodine-containing Hoechst compound (I-Hoechst) for 24 hours in the same manner as in Test Example 3, yielding spheroids incorporating the Hoechst compound. These spheroids were placed in tubes and incubated for an additional 18 hours under hypoxic conditions (0% oxygen concentration). As a control, spheroids were also incubated for 18 hours under normal oxygen conditions (20% oxygen concentration), not hypoxic conditions. After incubation under hypoxic conditions (or normal oxygen conditions), monochromatic X-ray irradiation was performed in the same manner as in Test Example 5. In this test, hypoxia in the spheroids was confirmed by the induction of expression of HIF-1α, a hypoxia marker. HIF-1α is a type of hypoxia-inducible factor and can be measured using a commercially available kit. Figure 10 shows the results (images) of microscopic observation of spheroids after incubation under hypoxic or normoxic conditions. Figure 11 shows a graph of the transition in HIF-1α fluorescent signal intensity. As shown in these figures, incubation under hypoxic conditions confirmed the expression of HIF-1α, which is not observed under normoxic conditions, confirming the formation of hypoxic spheroids. As shown in this figure, although there was a time lag (5 hours) before the actual irradiation time due to transportation to the X-ray irradiation device, it was confirmed that the hypoxic state was maintained even before irradiation. Figure 12 shows the results of X-ray-irradiated spheroids (photographs of the bottom of the tube). As shown in this figure, the destruction effect was confirmed not only in spheroids under normoxic conditions but also in hypoxic spheroids. This suggests that the method of the present invention can effectively treat cancer even under hypoxic conditions, where existing radiation therapies are not sufficiently effective.

[0091] Preparation Example 1: Preparation of Nanoparticles Supported by Iodine-Containing Hoechst Compound Synthesis of Biodegradable Mesoporous Silica Nanoparticles Biodegradable mesoporous silica nanoparticles (BPMO) were synthesized as a porous silica support by the following method. A mixture of cetyltrimethylammonium bromide (CTAB, 98%, Sigma-Aldrich) (250 mg), 8 M aqueous NaOH solution (219 μL), and water (120 mL) was vigorously stirred at 80°C to prepare a CTAB solution. Separately, rhodamine B isothiocyanate (RITC, Sigma-Aldrich) (2.5 mg) was dissolved in ethanol (5 mL), and 3-aminopropyltriethoxysilane (APTS, 99%, Wako) (6 μL) was added. The mixture was stirred at room temperature for 30 minutes to prepare a RITC-APTS solution. Next, 1,2-bis(triethoxysilyl)ethane (300 μL, Fluorochem) was mixed with the above RITC-APTS solution, and this mixture was added dropwise to the CTAB solution. Then, bis[3-(triethoxysilyl)propyl]tetrasulfide (100 μL, Fluorochem) was immediately added. The mixture was stirred for 15 minutes. Then, 315 μL of 50% aqueous 3-(trihydroxysilyl)propylmethylphosphonate monosodium solution was added to the mixture and stirred. The resulting solid product was collected by centrifugation and washed twice with ethanol. The solid product was refluxed in a mixed solution of ammonium nitrate (0.3 g) and ethanol (50 mL). This removed the template CTAB. The solid product was then centrifuged, washed three times with ethanol, and dried overnight. This yielded biodegradable mesoporous silica nanoparticles (BPMO). Figure 13 shows a TEM image (image taken by a transmission electron microscope) of an example of mesoporous silica nanoparticles, which confirms the mesoporous nature of the nanoparticles.

[0092] Loading of iodine-containing Hoechst compound onto nanoparticles The iodine-containing Hoechst compound described above, "Hoechst 33342 analog 2 trihydrochloride," was dissolved in water to prepare a 0.8 mg / mL solution. 110 μL of this solution was added to an Eppendorf tube containing 1 mg of the silica nanoparticles (BPMO). The mixture was stirred and mixed using a rotary mixer (80 rpm) at 4°C for 24 hours. The mixture was centrifuged (14,000 rpm, room temperature, 10 minutes), and the supernatant was removed. The nanoparticles were washed three times with 110 μL of water. This yielded nanoparticles loaded with the iodine-containing Hoechst compound.

[0093] Figure 14 shows the absorption spectra of the solution and washings used in nanoparticle production, measured using a spectrophotometer. The absorption spectra are shown for the solution of the iodine-containing Hoechst compound (before loading), the supernatant after loading, the first washing, the second washing, and the third washing. The absorbance was measured using a NanoDrop ultra-microspectrophotometer. Table 3 also shows the absorbance values ​​at a wavelength of 346 nm and the NanoDrop dilution ratio (the sample is diluted in the NanoDrop method). As shown above, the absorption due to the Hoechst compound decreases with increasing number of washings, and after the third washing, the absorption almost disappears. This indicates that the Hoechst compound is firmly supported on the silica nanoparticles. Furthermore, in this experiment, when 1 mg of silica nanoparticles was reacted with 0.088 mg of the iodine-containing Hoechst compound, it was confirmed that about 0.005 mg of the iodine-containing Hoechst compound was supported.

[0094] Test Example 8: Uptake of nanoparticles carrying an iodine atom-containing Hoechst compound into cancer cells. A test for uptake into cancer cells was conducted using the same method as in Test Example 2, except that nanoparticles carrying an iodine atom-containing Hoechst compound (the same amount of Hoechst compound) were used instead of the iodine atom-containing Hoechst compound in Test Example 2. This test confirmed that the iodine atom-containing Hoechst compound-carrying nanoparticles were efficiently taken up into cancer cells. Furthermore, a test for uptake into spheroids was conducted using the same method as in Test Example 3, except that nanoparticles carrying an iodine atom-containing Hoechst compound (the same amount of Hoechst compound) were used instead of the iodine atom-containing Hoechst compound in Test Example 3. This test demonstrated that the nanoparticles were evenly distributed within the spheroids, demonstrating excellent nanoparticle penetration into the spheroids. Furthermore, the safety of the iodine atom-containing Hoechst compound-carrying nanoparticles was confirmed using the same method as in Test Example 6.

[0095] Test Example 9 Irradiation of Monochromatic X-Rays onto Iodine-Atom-Containing Hoechst Compound-Supported Nanoparticle-Spheroids The spheroids (iodine-atom-containing Hoechst compound-supported nanoparticle-spheroids) prepared in Test Example 8 above were irradiated with monochromatic X-rays in the same manner as in Test Example 5. This confirmed the destruction of the spheroids after X-ray irradiation. That is, it was confirmed that the spheroids were completely destroyed when irradiated with 33.2 keV X-rays. Furthermore, the spheroids (iodine-atom-containing Hoechst compound-supported nanoparticle-spheroids) prepared in Test Example 8 above were irradiated with monochromatic X-rays in the same manner as in Test Example 7 to form hypoxic spheroids. This confirmed the destruction of hypoxic spheroids after X-ray irradiation. That is, it was confirmed that the hypoxic spheroids were completely destroyed when irradiated with 33.2 keV X-rays.

[0096] Test Example 10: In accordance with Test Example 8, nanoparticles (IH-BPMO) loaded with iodine-containing Hoechst (I-Hoechst) were added to a culture medium containing cancer cells, and the cells were cultured for 24 hours, followed by observation of fluorescence under a microscope. The results are shown in Figure 15. I-Hoechst fluorescence was observed in the nucleus, confirming nuclear staining as a result of delivery of IH by IH-BPMO.

[0097] Test Example 11: In accordance with Test Example 9, nanoparticles (IH-BPMO) loaded with iodine-containing Hoechst (I-Hoechst) were added to a culture medium containing cancer spheroids, incubated, and then irradiated with monochromatic X-rays. The results are shown in Figure 16. Destruction of the cancer spheroids was confirmed.

[0098] The pharmaceutical composition for use in radiotherapy according to the present invention enables highly effective radiotherapy.

Claims

1. Equation (I): 【Chemistry 1】 [In the formula, R １ C is an iodine atom, or a carbon atom substituted with one or more iodine atoms. １ -C ４ An aryl group substituted with one or more substituents selected from the group consisting of alkyl groups, aryl groups substituted with one or more iodine atoms, gadolinium atom-containing groups, gold atom-containing groups, silver atom-containing groups, and platinum atom-containing groups, wherein the aryl group may optionally contain a hydroxyl group, C １ -C ４ Alkyl alkyl groups, and C １ -C ４ They may be substituted with one or more substituents selected from the group consisting of alkoxy groups; X is a linking or C ２ or C ４ alkenylene group; and Ring A is given by equation (A): 【Chemistry 2】 It is a base represented by, where, R ２ is any one or more C １ -C ４ A non-aromatic heterocyclic group containing one or more heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur atoms, which may be substituted with alkyl groups, and at least one of the heteroatoms being a nitrogen atom; 【Transformation 3】 This is a linkage to the X group. A pharmaceutical composition for use in radiotherapy, comprising a compound represented by or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

2. R １ However, it is an aryl group substituted with one or more iodine atoms, where the aryl group may optionally be a hydroxyl group, C １ -C ４ Alkyl alkyl groups, and C １ -C ４ They may be substituted with one or more substituents selected from the group consisting of alkoxy groups; and, R ２ However, any one or more C １ -C ４ A six-membered non-aromatic heterocyclic group containing one to four nitrogen atoms, which may be substituted with an alkyl group. The pharmaceutical composition according to claim 1.

3. The compound of formula (I) is, formula (II): 【Chemistry 4】 [In the formula, R ３ is a phenyl group substituted with 1 to 3 iodine atoms, where the phenyl group may optionally be substituted with 1 or 2 hydroxyl groups; and, R ４ is a hydrogen atom, or C １ -C ４ It is an alkyl group. The pharmaceutical composition according to claim 1, wherein the compound is represented by [the compound shown].

4. The pharmaceutical composition according to claim 1, wherein the radiation is X-rays.

5. The pharmaceutical composition according to claim 4, wherein the X-rays are capable of exciting the K-shell electrons of atoms selected from the group consisting of iodine, gadolinium, gold, silver, and platinum atoms in the compound of formula (I).

6. The pharmaceutical composition according to claim 4, wherein the X-rays are monochromatic X-rays or characteristic X-rays.

7. A pharmaceutical composition according to any one of claims 1 to 6, for treating solid tumors or for suppressing the growth or proliferation of solid tumors.

8. The pharmaceutical composition according to claim 7, wherein the solid tumor is a brain tumor, lung cancer, ovarian cancer, digestive system cancer, osteosarcoma, or head and neck cancer.

9. The pharmaceutical composition according to claim 7, wherein the solid tumor is in a hypoxic state.

10. Equation (I): 【Transformation 5】 [In the formula, R １ C is an iodine atom, or a carbon atom substituted with one or more iodine atoms. １ -C ４ An aryl group substituted with one or more substituents selected from the group consisting of alkyl groups, aryl groups substituted with one or more iodine atoms, gadolinium atom-containing groups, gold atom-containing groups, silver atom-containing groups, and platinum atom-containing groups, wherein the aryl group may optionally contain a hydroxyl group, C １ -C ４ Alkyl alkyl groups, and C １ -C ４ They may be substituted with one or more substituents selected from the group consisting of alkoxy groups; X is a combination, or C ２ or C ４ It is an alkenylene group; and, Ring A is given by equation (A): 【Transformation 6】 It is a base represented by, where, R ２ is any one or more C １ -C ４ A non-aromatic heterocyclic group containing one or more heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur atoms, which may be substituted with alkyl groups, and at least one of the heteroatoms being a nitrogen atom; 【Transformation 7】 This is a linkage to the X group. Compounds represented by or pharmaceutically acceptable salts thereof; and, Porous silica support, Nanoparticles consisting of these nanoparticles.

11. The nanoparticles according to claim 10, wherein the nanoparticles are biodegradable mesoporous silica nanoparticles.

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