WO2022009788A1 - Cell killing method and scintillator particles - Google Patents

Cell killing method and scintillator particles Download PDF

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Publication number
WO2022009788A1
WO2022009788A1 PCT/JP2021/025048 JP2021025048W WO2022009788A1 WO 2022009788 A1 WO2022009788 A1 WO 2022009788A1 JP 2021025048 W JP2021025048 W JP 2021025048W WO 2022009788 A1 WO2022009788 A1 WO 2022009788A1
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scintillator
rays
cells
target
cell
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French (fr)
Japanese (ja)
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哲 石坂
拓司 相宮
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コニカミノルタ株式会社
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N13/00—Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues

Definitions

  • the present invention relates to a cell killing method and scintillator particles.
  • Patent Document 1 describes a method of guiding an ultraviolet ray generated by a xenon flash lamp to a vicinity of a target cell by an optical fiber and irradiating the target cell with the ultraviolet ray to damage the target cell.
  • Patent Document 2 describes a method of damaging a target cell by irradiating it with near-infrared rays in a state where the complex to which a dye and an antibody are bound is bound to the target cell.
  • Patent Document 1 if a shield exists between the optical fiber and the target cell, the ultraviolet ray is attenuated or the ultraviolet ray is shielded, so that sufficient ultraviolet ray does not reach the target cell.
  • the shield means a normal tissue other than a target cell such as a skin tissue in a living body, a container for controlling temperature and humidity, a member for blocking impurities and the like.
  • Patent Document 1 describes that the pulse irradiation of ultraviolet rays causes damage to tumor cells, but does not describe a mechanism that damages only tumor cells, and is reproducible and versatile. Was scarce.
  • the present invention is to provide a cell killing method and scintillator particles that can damage a target cell even if there is a shield between the light source and the target cell.
  • the method for killing cells is a step of irradiating a scintillator with X-rays to generate ultraviolet rays from the scintillator, and causing the generated ultraviolet rays to reach the target cells to kill the target cells.
  • the scintillator particles according to the embodiment of the present invention include particles containing a scintillator and a resin, and a target substance recognition substance that is supported on the surface of the particles and specifically binds to a target cell.
  • the target cell can be damaged even if there is a shield between the light source and the target cell.
  • FIG. 1A and 1B are schematic views of an experiment showing the effect of this embodiment.
  • FIG. 2 is another schematic diagram of an experiment showing the effect of this embodiment.
  • the cell killing method includes a step of irradiating the scintillator with X-rays to generate ultraviolet rays from the scintillator, and a step of causing the generated ultraviolet rays to reach the target cells and killing the target cells. .. Further, the cell killing method may further include a step of labeling the target cell with a scintillator having a target substance recognition substance that specifically binds to the target cell before irradiating the scintillator with X-rays.
  • a scintillator is a substance that emits fluorescence when excited by radiation (X-rays). Generally, the scintillator is widely used in medical X-ray diagnostic imaging equipment and the like. In this embodiment, a scintillator is used to kill the target cells.
  • the scintillator that can be used in the present embodiment is not particularly limited as long as it generates ultraviolet rays by being irradiated with radiation (X-rays).
  • the scintillator may be an organic scintillator or an inorganic scintillator. Examples of inorganic scintillators include single crystals of Cs 2 ZnCl 4.
  • Cs 2 ZnCl 4 generates ultraviolet rays having a spectral peak at 200 to 300 nm, which has a large cell killing effect, by irradiating with X-rays having an energy of 67.4 keV.
  • the wavelength of the ultraviolet light emitted by the scintillator can be adjusted by the material of the scintillator.
  • the shape of the scintillator is not particularly limited, but when the scintillator is used by adhering to cells as particles, the shape of the scintillator is preferably a spherical shape or a rectangular parallelepiped shape.
  • the diameter of the sphere is preferably 30 to 300 nm.
  • the shape of the scintillator is a rectangular parallelepiped shape, the length of one side of the rectangular parallelepiped is preferably 30 to 300 nm.
  • the diameter or maximum length of the scintillator is less than 30 nm or more than 300 nm, if it is less than the lower limit, the emission intensity of ultraviolet rays may be weakened and the killing effect may be insufficient. If the upper limit is exceeded, it becomes difficult for the reagent to reach the target cells, and there is a risk that the number of cells that cannot be killed will increase.
  • thermoplastic resins include polystyrene, polyacrylonitrile, polyfuran, or similar resins.
  • thermosetting resins include polyxylene, polylactic acid, glycidyl methacrylate, melamine resins, polyureas, polybenzoguanamines, polyamides, phenolic resins, polysaccharides and similar resins.
  • thermosetting resin is preferably a melamine resin from the viewpoint that the elution of the scintillator contained in the particles can be suppressed even by treatments such as dehydration, permeation, and encapsulation using an organic solvent such as xylene.
  • the target substance recognition substance is supported on the surface of the particles coated with the scintillator with resin or silicone.
  • the target substance recognition substance is, for example, a molecule that specifically binds to the target substance bound to the cell to be killed.
  • the cells to be killed are labeled by the scintillator particles in which the target substance recognition substance is carried on the surface of the particles.
  • the target substance is, for example, a membrane protein that is specifically present in the cell to be killed.
  • the target substance recognition substance is, for example, an antibody against a membrane protein specifically present in the cell to be killed.
  • the target substance and a combination of the target substance recognition substance that specifically binds to it can be interpreted as a generic term in the art of the present invention, association constant (K A) or dissociation constant which is the inverse (K D) Can be defined by.
  • Target substance recognition substance to be used in the present invention preferably binding constant between the target substance (K A) is in the range of 1 ⁇ 10 5 ⁇ 1 ⁇ 10 12. If binding constants (K A) is within the range, the target substance can be handled as a target substance recognition substance that specifically binds to the target substance.
  • X-rays are a type of electromagnetic wave, but because of their short wavelength and large properties as photons, the expression eV is often used as the energy of photons instead of wavelength.
  • the energy of X-rays irradiated to the scintillator is not particularly limited as long as ultraviolet rays having sufficient energy to kill cells are generated.
  • the energy of the X-rays applied to the scintillator is preferably 20 to 150 eV. If the energy of the X-rays applied to the scintillator is too low, it may not be possible to kill the cells. On the other hand, if the energy of the X-rays applied to the scintillator is too large, it may affect other tissues.
  • the ultraviolet rays generated from the scintillator preferably include ultraviolet rays having a wavelength of 200 to 300 nm.
  • Ultraviolet rays having a wavelength of 200 to 300 nm are effective in killing cells because they have a high ability to kill cells.
  • the method of irradiating X-rays is not particularly limited. For X-rays, a known X-ray irradiation device can be used.
  • FIG. 1A and 1B are schematic views of an experiment showing the effect of this embodiment.
  • 1A is a plan view for explaining the experiment
  • FIG. 1B is a cross-sectional view taken along the line AA shown in FIG. 1A.
  • the target cells 110 to be killed are cultured in a petri dish 100.
  • the cultured target cells 110 are partitioned into two regions A and B.
  • a scintillator plate 120 including the scintillator described above is arranged on the upper part of one region A, and nothing is arranged on the other region B.
  • the thickness of the scintillator plate 120 arranged in the region A is set to a thickness of several hundred ⁇ m, which is the same as the thickness of the scintillator plate 120 used in the medical field.
  • the scintillator plate 120 and the target cell 110 in the region A may be brought into contact with each other, or may be brought into close contact with each other via a thin optical member that transmits ultraviolet rays. In the present embodiment, the scintillator plate 120 and the target cell 110 in the region A are in contact with each other.
  • the arrangement of the scintillator and cells is not limited to the above example.
  • the scintillator may be formed into a particle shape and placed in the vicinity of the cell.
  • the cells do not have to have a planar distribution. Since X-rays basically pass through living tissues having atoms with small atomic numbers, cells may be on a mass.
  • the cell killing mechanism in this embodiment is the destruction of DNA in the cell nucleus by ultraviolet rays.
  • the advantage of the present embodiment is that even ultraviolet rays having a wavelength of 200 to 300 nm reach the cells to cause a killing effect.
  • the intensity of ultraviolet rays can be controlled by the intensity of the X-rays to be irradiated and the irradiation time of the X-rays.
  • FIG. 2 is another schematic diagram of an experiment showing the effect of this embodiment.
  • the scintillator 200 is shown in large size for the sake of explanation.
  • two identical cells a and b are partitioned by a light-shielding partition plate 210.
  • the cells a are attached with particles of a scintillator 200 that emits ultraviolet rays having a wavelength of 200 to 300 nm when irradiated with X-rays.
  • the size of the cells a and b is about 10 ⁇ m in diameter, and the diameter of the fine particles is about 100 nm.
  • the scintillator 200 emits ultraviolet rays in the cells a, so that the cells a in the vicinity are destroyed and die. Similar to the experiment shown in FIG. 1, the intensity of ultraviolet rays can be controlled by the intensity of X-rays to be irradiated and the irradiation time of X-rays.
  • the X-ray only permeates the cell b and does not interact with the cell b, so that the cell b does not die. Since the intensity of ultraviolet rays from the scintillator 200 attached to the cell a is rapidly attenuated by the inverse square law with respect to the distance from the scintillator 200, the killing effect on the cell b is extremely small even if the partition plate 210 is not present. it is conceivable that.
  • the wavelength of near-infrared light is known to be 660 to 740 nm.
  • the wavelength of X-rays is 1/1000 to 1/10000 of the wavelength of near-infrared light in the region of diagnostic X-rays. That is, the photon energy of X-rays is 1000 to 10,000 times larger than that of near-infrared photon energy.
  • the photon energy of X-rays is about 40,000 times larger than the photon energy of light (near infrared light) having a wavelength of 700 nm.
  • Ultraviolet rays directly interact with the bonds of the molecules that make up the living body, and can break the bonds and destroy the molecules.
  • the energy of near-infrared rays is only in the region of thermal vibration of the molecules constituting the living body, and the near-infrared rays cannot exert a direct destructive action on the molecules constituting the living body.
  • Ultraviolet light has a shorter wavelength than infrared light, but has a much longer wavelength than X-ray. Therefore, since ultraviolet rays can be generated from the photon energy of X-rays, a direct cell-killing action is exhibited.
  • X-rays are effective not only because the generator is easily available, but also because of their energy characteristics.
  • the transmittance of X-rays tends to decrease, and the influence of attenuation by the shielding material tends to increase.
  • it exceeds 150 keV the permeability becomes large and it becomes difficult to be absorbed by the scintillator, so that the emission intensity of ultraviolet rays becomes small and the bactericidal cell killing effect may be reduced.
  • Example 1 In the first embodiment, an example when there is a shield will be described.
  • the affected area is scraped with a grinding machine called a turbine and then covered with a metal or plastic crown (cover).
  • a metal or plastic crown cover
  • dental caries may recur internally after several years.
  • a scintillator that emits ultraviolet rays by irradiating the adhesive of the crown with X-rays of 200 to 300 nm is mixed to treat dental caries.
  • the scintillator in the adhesive is irradiated with X-rays to emit ultraviolet rays near the adhesive surface, and the remaining bacteria can be killed.
  • an X-ray generator for computer tomography (CT imaging) provided in many dentistry can be used.
  • the scintillator is made of a material that is harmless to the human body in consideration of leakage of the scintillator and accidental ingestion when the crown comes off. Alternatively, if there is a concern about safety, the scintillator is coated with a water-insoluble resin or silicon to prevent leakage.
  • Example 2 an example relating to the selective killing action will be described. Specifically, there is a case where there is a target cell such as a cancer cell to be selectively killed in the subject.
  • a target cell such as a cancer cell to be selectively killed in the subject.
  • Unique proteins (receptors) are often expressed on the cell membrane surface of cancer cells.
  • specific proteins expressed in cancer cells include human epidermal growth factor receptor type 2 (HER2) for breast cancer and epidermal growth factor receptor (EGFR) for colon cancer.
  • HER2 human epidermal growth factor receptor type 2
  • EGFR epidermal growth factor receptor
  • the scintillator is attached to the cell membrane by utilizing the antigen-antibody reaction.
  • scintillator particles obtained by surface-modifying a scintillator with a resin or silicone and binding to an antibody via polyethylene glycol (PEG), avidin, or biotin can be used as a reagent.
  • the antibody may be a combination of a primary antibody and a secondary antibody.
  • the selective killing action as in Example 2 can be applied in vitro or in vivo.
  • the scintillator is made of a material that is harmless to the human body when the selective killing action is used in vivo.
  • the scintillator is coated with a water-insoluble resin or silicon to prevent leakage.
  • the scintillator in vivo if the X-ray irradiation position is set to multiple ring-shaped positions centered on the irradiation target part as in CT, exposure to areas other than the killed target part can be minimized. can.
  • Example 2 the intensity of X-rays required for the cell-killing effect in application to tumor histiocytes in a subject as in Example 2 will be considered.
  • the photon energy of X-rays is about 10,000 times or more larger than the photon energy of ultraviolet rays. Specifically, theoretically, about 10,000 ultraviolet photons are generated for each X-ray photon. However, in reality, the conversion efficiency of ultraviolet photons generated from X-ray photons is about several percent due to the loss of heat energy and the like. For example, even if the conversion efficiency is underestimated to 1%, 1 to 100 ultraviolet photons of X-rays are generated. Given that even a single UV photon has the energy to destroy molecules in the cell nucleus, as long as the scintillator attached to the cell reacts with the X-ray photon, the attached cell and nearby cells will be killed. can.
  • the question is whether the X-ray photons and the scintillator can react.
  • the intensity of X-rays required for X-ray photons to react with the scintillator is difficult to calculate.
  • one X-ray photon is transmitted per projected area of one cell as a minimum requirement.
  • the calculated X-ray intensity (0.42 ⁇ Gy). Is a value that is four or more digits lower than this.
  • the X-ray intensity is low from the viewpoint of radiation exposure, and it can be said that there is no problem if it is the above.
  • the calculated value may fluctuate at the level of several digits.
  • the following may be considered as the possibility that it is necessary to irradiate a large amount of X-rays.
  • (2) Does the X-ray photon react with the scintillator attached to the cell? In order to reliably react the X-ray photons with the scintillator attached to the cells, it is necessary to irradiate a large number of X-rays in consideration of about 3 to 5 digits.
  • Tumors are thick in the depth direction and have a large number of cells present in the path of X-ray photons. In the case of a tumor with a thickness of 10 mm, 1000 cells will be present in the depth direction. Considering that the scintillator particles attached to any of these cells should react with the X-ray photons, the probability of the scintillator particles reacting with the X-ray photons increases by three orders of magnitude, so that the X-ray intensity can be reduced. .. (2) The X-ray irradiation is divided into a plurality of times.
  • X-rays are usually irradiated several times to a dozen times instead of once. If the same method as radiation therapy is adopted, the intensity of X-rays irradiated at one time can be reduced by about an order of magnitude.
  • the killing method according to the present embodiment can appropriately irradiate the target cells with ultraviolet rays, it is useful for appropriately sterilizing a narrow area and killing only the target cells in the living body.

Abstract

The killing method according to the present invention comprises: a step for emitting X-rays to a scintillator to generate ultraviolet rays from the scintillator; and a step for causing the generated ultraviolet rays to reach target cells to kill the target cells.

Description

細胞の殺傷方法、シンチレーター粒子How to kill cells, scintillator particles
　本発明は、細胞の殺傷方法と、シンチレーター粒子とに関する。 The present invention relates to a cell killing method and scintillator particles.
　従来、紫外線を照射することで、殺菌対象物を殺菌処理したり、目的細胞を殺傷したりする方法が知られている。 Conventionally, a method of sterilizing an object to be sterilized or killing a target cell by irradiating with ultraviolet rays has been known.
　特許文献１には、キセノンフラッシュランプで発生させた紫外線を光ファイバーで目的細胞の近くまで導き、目的細胞に向けて紫外線を照射することで、目的細胞に損傷を与える方法が記載されている。 Patent Document 1 describes a method of guiding an ultraviolet ray generated by a xenon flash lamp to a vicinity of a target cell by an optical fiber and irradiating the target cell with the ultraviolet ray to damage the target cell.
　また、特許文献２には、色素および抗体を結合させた複合体を目的細胞に結合させた状態で、近赤外線を照射することで、目的細胞に損傷を与える方法が記載されている。 Further, Patent Document 2 describes a method of damaging a target cell by irradiating it with near-infrared rays in a state where the complex to which a dye and an antibody are bound is bound to the target cell.
特開２０１１－０７８７５０号公報Japanese Unexamined Patent Publication No. 2011-078750 特表２０１４－５２３９０７号公報Special Table 2014-523907 Gazette
　しかしながら、特許文献１に記載の方法では、光ファイバーと、目的細胞との間に遮蔽物が存在すると、紫外線が減衰されるか、紫外線が遮蔽されるため、目的細胞に十分な紫外線が到達しないことがある。ここで、遮蔽物とは、生体における皮膚組織などの目的細胞以外の正常組織、温度および湿度を管理するための容器、不純物などを遮断するための部材を意味する。また、特許文献１には、紫外線をパルス照射することで、腫瘍細胞に損傷を与えることが記載されているが、腫瘍細胞にのみ損傷を与える機構が記載されておらず、再現性および汎用性が乏しかった。 However, in the method described in Patent Document 1, if a shield exists between the optical fiber and the target cell, the ultraviolet ray is attenuated or the ultraviolet ray is shielded, so that sufficient ultraviolet ray does not reach the target cell. There is. Here, the shield means a normal tissue other than a target cell such as a skin tissue in a living body, a container for controlling temperature and humidity, a member for blocking impurities and the like. Further, Patent Document 1 describes that the pulse irradiation of ultraviolet rays causes damage to tumor cells, but does not describe a mechanism that damages only tumor cells, and is reproducible and versatile. Was scarce.
　また、特許文献２に記載の方法では、特許文献１と同様に、遮蔽物の存在により近赤外光が減衰されてしまうか、近赤外光が遮蔽されることが考えられる。近赤外光は、生体組織に対する透過率が高いことが知られている。しかしながら、生体組織の厚みが厚い場合には、近赤外光が生体組織に吸収されたり、散乱することで、目的細胞に到達する近赤外光の強度が低くなってしまう問題があった。さらに、エネルギーの低い近赤外光を使用するため、効果が得られる条件が限られたり、長時間の近赤外線照射が必要な場合がある。 Further, in the method described in Patent Document 2, it is conceivable that the near-infrared light is attenuated or the near-infrared light is shielded due to the presence of the shield, as in Patent Document 1. Near-infrared light is known to have high transmittance for living tissues. However, when the living tissue is thick, there is a problem that the intensity of the near-infrared light reaching the target cell is lowered due to the near-infrared light being absorbed or scattered by the living tissue. Furthermore, since near-infrared light with low energy is used, the conditions under which the effect can be obtained may be limited, or long-term near-infrared irradiation may be required.
　本発明は、光源および目的細胞の間に遮蔽物があった場合でも、目的細胞に損傷を与えることができる細胞の殺傷方法およびシンチレーター粒子を提供することである。 The present invention is to provide a cell killing method and scintillator particles that can damage a target cell even if there is a shield between the light source and the target cell.
　本発明の一実施の形態に係る細胞の殺傷方法は、シンチレーターにＸ線を照射して、前記シンチレーターから紫外線を発生させる工程と、発生した紫外線を目的細胞に到達させて、前記目的細胞を殺傷する工程と、を含む。 The method for killing cells according to an embodiment of the present invention is a step of irradiating a scintillator with X-rays to generate ultraviolet rays from the scintillator, and causing the generated ultraviolet rays to reach the target cells to kill the target cells. Including the process of
　本発明の一実施の形態に係るシンチレーター粒子は、シンチレーターおよび樹脂を含む粒子と、前記粒子の表面に担持された、目的細胞に特異的に結合する標的物質認識物質と、を有する。 The scintillator particles according to the embodiment of the present invention include particles containing a scintillator and a resin, and a target substance recognition substance that is supported on the surface of the particles and specifically binds to a target cell.
　本発明によれば、光源および目的細胞の間に遮蔽物があった場合でも、目的細胞に損傷を与えることができる。 According to the present invention, even if there is a shield between the light source and the target cell, the target cell can be damaged.
図１Ａ、Ｂは、本実施の形態の効果を示す実験の模式図である。1A and 1B are schematic views of an experiment showing the effect of this embodiment. 図２は、本実施の形態の効果を示す実験の他の模式図である。FIG. 2 is another schematic diagram of an experiment showing the effect of this embodiment.
　以下、本発明の一実施の形態に係る細胞の殺傷方法について説明する。 Hereinafter, a method for killing cells according to an embodiment of the present invention will be described.
　本実施の形態に係る細胞の殺傷方法は、シンチレーターにＸ線を照射して、シンチレーターから紫外線を発生させる工程と、発生した紫外線を目的細胞に到達させて、目的細胞を殺傷する工程とを含む。また、細胞の殺傷方法は、シンチレーターにＸ線を照射する前に、目的細胞に特異的に結合する標的物質認識物質を有するシンチレーターで目的細胞を標識する工程をさらに有してもよい。 The cell killing method according to the present embodiment includes a step of irradiating the scintillator with X-rays to generate ultraviolet rays from the scintillator, and a step of causing the generated ultraviolet rays to reach the target cells and killing the target cells. .. Further, the cell killing method may further include a step of labeling the target cell with a scintillator having a target substance recognition substance that specifically binds to the target cell before irradiating the scintillator with X-rays.
　シンチレーターとは、放射線（Ｘ線）による励起で蛍光を発生する物質である。一般に、シンチレーターは、医療用のＸ線画像診断装置などに広く使用されている。本実施の形態では、目的細胞を殺傷するためにシンチレーターを使用している。本実施の形態で使用できるシンチレーターは、放射線（Ｘ線）が照射されることで、紫外線を発生させるものであれば特に限定されない。シンチレーターは、有機シンチレーターでもよいし、無機シンチレーターでもよい。無機シンチレーターの例には、Ｃｓ２ＺｎＣｌ４の単結晶が含まれる。例えば、Ｃｓ２ＺｎＣｌ４は、６７．４ｋｅＶのエネルギーのＸ線を照射することで、細胞殺傷効果の大きい２００～３００ｎｍにスペクトルピークを有する紫外線を発生させる。シンチレーターが発光する紫外線の波長は、シンチレーターの材料により調整できる。 A scintillator is a substance that emits fluorescence when excited by radiation (X-rays). Generally, the scintillator is widely used in medical X-ray diagnostic imaging equipment and the like. In this embodiment, a scintillator is used to kill the target cells. The scintillator that can be used in the present embodiment is not particularly limited as long as it generates ultraviolet rays by being irradiated with radiation (X-rays). The scintillator may be an organic scintillator or an inorganic scintillator. Examples of inorganic scintillators include single crystals of Cs 2 ZnCl 4. For example, Cs 2 ZnCl 4 generates ultraviolet rays having a spectral peak at 200 to 300 nm, which has a large cell killing effect, by irradiating with X-rays having an energy of 67.4 keV. The wavelength of the ultraviolet light emitted by the scintillator can be adjusted by the material of the scintillator.
　本実施の形態においてシンチレーターの形状は、特に限定されないが、粒子状として細胞に付着させて用いる場合は、シンチレーターの形状は、球形状または直方体形状が好ましい。シンチレーターの形状が球形状の場合、球の直径は、３０～３００ｎｍが好ましい。シンチレーターの形状が直方体形状の場合、直方体の一辺の長さは、３０～３００ｎｍが好ましい。シンチレーターの直径または最大長さが３０ｎｍ未満または３００ｎｍ超の場合、下限を下回ると、紫外線の発光強度が弱くなり殺傷効果が不十分となるおそれがある。また上限を超えると、目的とする細胞に試薬が届きにくくなり、殺傷できない細胞が増えてくるおそれがある。 In the present embodiment, the shape of the scintillator is not particularly limited, but when the scintillator is used by adhering to cells as particles, the shape of the scintillator is preferably a spherical shape or a rectangular parallelepiped shape. When the shape of the scintillator is spherical, the diameter of the sphere is preferably 30 to 300 nm. When the shape of the scintillator is a rectangular parallelepiped shape, the length of one side of the rectangular parallelepiped is preferably 30 to 300 nm. When the diameter or maximum length of the scintillator is less than 30 nm or more than 300 nm, if it is less than the lower limit, the emission intensity of ultraviolet rays may be weakened and the killing effect may be insufficient. If the upper limit is exceeded, it becomes difficult for the reagent to reach the target cells, and there is a risk that the number of cells that cannot be killed will increase.
　シンチレーターは、その表面の少なくとも一部が樹脂またはシリコーンで修飾されていることが好ましい。シンチレーターを被覆する樹脂の例には、熱可塑性樹脂または熱硬化性樹脂が含まれる。熱可塑性樹脂の例には、ポリスチレン、ポリアクリロニトリル、ポリフラン、または、これに類する樹脂が含まれる。熱硬化性樹脂の例には、ポリキシレン、ポリ乳酸、グリシジルメタクリレート、メラミン樹脂、ポリウレア、ポリベンゾグアナミン、ポリアミド、フェノール樹脂、多糖類またはこれに類する樹脂が含まれる。熱硬化性樹脂は、キシレンなどの有機溶媒を用いる脱水、透徹、封入などの処理によっても、粒子に内包させたシンチレーターの溶出を抑制できる観点から、メラミン樹脂が好ましい。 It is preferable that at least a part of the surface of the scintillator is modified with resin or silicone. Examples of resins that coat the scintillator include thermoplastic or thermosetting resins. Examples of thermoplastic resins include polystyrene, polyacrylonitrile, polyfuran, or similar resins. Examples of thermosetting resins include polyxylene, polylactic acid, glycidyl methacrylate, melamine resins, polyureas, polybenzoguanamines, polyamides, phenolic resins, polysaccharides and similar resins. The thermosetting resin is preferably a melamine resin from the viewpoint that the elution of the scintillator contained in the particles can be suppressed even by treatments such as dehydration, permeation, and encapsulation using an organic solvent such as xylene.
　シンチレーターを樹脂またはシリコーンで被覆した粒子の表面には、標的物質認識物質が担持されていることが好ましい。 It is preferable that the target substance recognition substance is supported on the surface of the particles coated with the scintillator with resin or silicone.
　標的物質認識物質は、例えば殺傷対象の細胞に結合した標的物質に特異的に結合する分子である。標的物質認識物質が標的物質に結合することで、粒子の表面に標的物質認識物質が担持されたシンチレーター粒子によって殺傷対象の細胞が標識されることになる。標的物質は、例えば殺傷対象の細胞に特異的に存在する膜タンパク質である。標的物質認識物質は、例えば殺傷対象の細胞に特異的に存在する膜タンパク質に対する抗体である。標的物質およびそれに特異的に結合する標的物質認識物質の組み合わせは、本発明の属する技術分野における一般的な用語として解釈できるが、結合定数（ＫＡ）またはその逆数である解離定数（ＫＤ）によって定義できる。 The target substance recognition substance is, for example, a molecule that specifically binds to the target substance bound to the cell to be killed. When the target substance recognition substance binds to the target substance, the cells to be killed are labeled by the scintillator particles in which the target substance recognition substance is carried on the surface of the particles. The target substance is, for example, a membrane protein that is specifically present in the cell to be killed. The target substance recognition substance is, for example, an antibody against a membrane protein specifically present in the cell to be killed. The target substance and a combination of the target substance recognition substance that specifically binds to it, can be interpreted as a generic term in the art of the present invention, association constant (K A) or dissociation constant which is the inverse (K D) Can be defined by.
　本発明で用いられる標的物質認識物質は、標的物質との間の結合定数（ＫＡ）が１×１０５～１×１０１２の範囲にあることが好ましい。結合定数（ＫＡ）が当該範囲内にある場合は、その標的物質は標的物質と特異的に結合する標的物質認識物質として取り扱うことができる。 Target substance recognition substance to be used in the present invention preferably binding constant between the target substance (K A) is in the range of 1 × 10 5 ~ 1 × 10 12. If binding constants (K A) is within the range, the target substance can be handled as a target substance recognition substance that specifically binds to the target substance.
　Ｘ線は、電磁波の一種であるが、波長が短く光子としての性質が大きいことから、波長に代わって、光子のエネルギーとしてｅＶの表現を採用することが多い。波長（λ）と、エネルギー（Ｐ）との関係は、Ｐ（ｋｅＶ）＝１２．４／λ（Å）で表される。 X-rays are a type of electromagnetic wave, but because of their short wavelength and large properties as photons, the expression eV is often used as the energy of photons instead of wavelength. The relationship between the wavelength (λ) and the energy (P) is represented by P (keV) = 12.4 / λ (Å).
　シンチレーターに照射されるＸ線のエネルギーは、細胞を殺傷するのに十分なエネルギーの紫外線が発生されれば特に限定されない。シンチレーターに照射されるＸ線のエネルギーは、２０～１５０ｅＶが好ましい。シンチレーターに照射されるＸ線のエネルギーが小さすぎる場合、細胞を殺傷できないおそれがある。一方、シンチレーターに照射されるＸ線のエネルギーが大きすぎる場合、他の組織に影響をおよぼしてしまうことがある。 The energy of X-rays irradiated to the scintillator is not particularly limited as long as ultraviolet rays having sufficient energy to kill cells are generated. The energy of the X-rays applied to the scintillator is preferably 20 to 150 eV. If the energy of the X-rays applied to the scintillator is too low, it may not be possible to kill the cells. On the other hand, if the energy of the X-rays applied to the scintillator is too large, it may affect other tissues.
　シンチレーターから発生する紫外線は、波長が２００～３００ｎｍの紫外線を含むことが好ましい。波長が２００～３００ｎｍの紫外線は、細胞を殺傷する能力が高いため、細胞の殺傷に効果的である。Ｘ線を照射する方法は、特に限定されない。Ｘ線は、公知のＸ線照射装置を使用できる。 The ultraviolet rays generated from the scintillator preferably include ultraviolet rays having a wavelength of 200 to 300 nm. Ultraviolet rays having a wavelength of 200 to 300 nm are effective in killing cells because they have a high ability to kill cells. The method of irradiating X-rays is not particularly limited. For X-rays, a known X-ray irradiation device can be used.
　本実施の形態における効果は、例えば以下のような実験で確認できる。図１Ａ、Ｂは、本実施の形態の効果を示す実験の模式図である。図１Ａは、実験を説明するための平面図であり、図１Ｂは、図１Ａに示されるＡ－Ａ線の断面図である。 The effect of this embodiment can be confirmed by the following experiment, for example. 1A and 1B are schematic views of an experiment showing the effect of this embodiment. 1A is a plan view for explaining the experiment, and FIG. 1B is a cross-sectional view taken along the line AA shown in FIG. 1A.
　図１Ａ、Ｂに示されるように、シャーレ１００に殺傷対象となる目的細胞１１０を培養する。培養した目的細胞１１０を２つの領域Ａおよび領域Ｂに区画する。一方の領域Ａの上部には、前述したシンチレーターを含むシンチレータープレート１２０を配置し、他方の領域Ｂには、何も配置しない。領域Ａに配置したシンチレータープレート１２０の厚みは、医療の分野で使用されるシンチレータープレート１２０の厚みと同じ数百μｍの厚さとする。シンチレータープレート１２０と、領域Ａの目的細胞１１０とは、接触させてもよいが、紫外線を透過する薄い光学部材を介して近接させてもよい。本実施の形態では、シンチレータープレート１２０と、領域Ａの目的細胞１１０とは、接触させている。 As shown in FIGS. 1A and 1B, the target cells 110 to be killed are cultured in a petri dish 100. The cultured target cells 110 are partitioned into two regions A and B. A scintillator plate 120 including the scintillator described above is arranged on the upper part of one region A, and nothing is arranged on the other region B. The thickness of the scintillator plate 120 arranged in the region A is set to a thickness of several hundred μm, which is the same as the thickness of the scintillator plate 120 used in the medical field. The scintillator plate 120 and the target cell 110 in the region A may be brought into contact with each other, or may be brought into close contact with each other via a thin optical member that transmits ultraviolet rays. In the present embodiment, the scintillator plate 120 and the target cell 110 in the region A are in contact with each other.
　この状態で、目的細胞１１０の上方からシンチレータープレート１２０を介してＸ線を照射する。シンチレータープレート１２０が配置された領域Ａでは波長２００～３００ｎｍの紫外線が発生する。発生した紫外線により、領域Ａの目的細胞１１０は殺傷する。紫外線の強度は、照射するＸ線の強度および照射時間で調整できる。一方、シンチレータープレート１２０が配置されていない領域ＢではＸ線が透過するのみであり、目的細胞１１０と相互作用しないため、目的細胞１１０は殺傷されない。以上の工程により、本実施の形態の効果を実証できる。 In this state, X-rays are irradiated from above the target cells 110 via the scintillator plate 120. In the region A where the scintillator plate 120 is arranged, ultraviolet rays having a wavelength of 200 to 300 nm are generated. The generated ultraviolet rays kill the target cells 110 in region A. The intensity of ultraviolet rays can be adjusted by the intensity of X-rays to be irradiated and the irradiation time. On the other hand, in the region B where the scintillator plate 120 is not arranged, only X-rays are transmitted and the target cells 110 do not interact with each other, so that the target cells 110 are not killed. Through the above steps, the effect of this embodiment can be demonstrated.
　なお、シンチレーターおよび細胞の配置は、上記の例に限定されない。例えば、シンチレーターを粒子形状にして細胞の近傍に配置してもよい。また、細胞も平面的な分布でなくてもよい。Ｘ線は、基本的に原子番号の小さい原子を有する生体組織を透過するため、細胞は塊上でもよい。本実施の形態における細胞殺傷機序は、紫外線による細胞核内のＤＮＡの破壊である。このように、本実施の形態の利点は、波長が２００～３００ｎｍの紫外線さえ細胞に届けば、殺傷作用が生じる点である。すなわち、紫外線を遮蔽物の存在下で確実に細胞に照射させるには、遮蔽物を透過するＸ線と、様々な形態で細胞の近傍に配置できるシンチレーターを組み合わせて用いることが極めて有効である。なお、紫外線の強度は、照射するＸ線の強度およびＸ線の照射時間で制御できる。 The arrangement of the scintillator and cells is not limited to the above example. For example, the scintillator may be formed into a particle shape and placed in the vicinity of the cell. Also, the cells do not have to have a planar distribution. Since X-rays basically pass through living tissues having atoms with small atomic numbers, cells may be on a mass. The cell killing mechanism in this embodiment is the destruction of DNA in the cell nucleus by ultraviolet rays. As described above, the advantage of the present embodiment is that even ultraviolet rays having a wavelength of 200 to 300 nm reach the cells to cause a killing effect. That is, in order to reliably irradiate cells with ultraviolet rays in the presence of a shield, it is extremely effective to use a combination of X-rays that pass through the shield and a scintillator that can be placed in the vicinity of the cells in various forms. The intensity of ultraviolet rays can be controlled by the intensity of the X-rays to be irradiated and the irradiation time of the X-rays.
　本実施の形態における細胞の選択的殺傷作用は、例えば以下のような実験も確認できる。図２は、本実施の形態の効果を示す実験の他の模式図である。図２では、説明のためにシンチレーター２００を大きく示している。 For example, the following experiments can be confirmed for the selective killing action of cells in this embodiment. FIG. 2 is another schematic diagram of an experiment showing the effect of this embodiment. In FIG. 2, the scintillator 200 is shown in large size for the sake of explanation.
　図２に示されるように、２つの同じ細胞ａ、ｂを遮光性の仕切り板２１０で仕切る。細胞ａには、Ｘ線を照射されることで波長が２００～３００ｎｍの紫外線を発光するシンチレーター２００の粒子を付着させる。細胞ａ、ｂの大きさは直径１０μｍ程度であり、微粒子の直径は１００ｎｍ程度である。細胞ａ、ｂに対してＸ線を照射すると、細胞ａでは、シンチレーター２００が紫外線を発するため、近接している細胞ａは細胞核を破壊され死滅する。図１に示した実験と同様に、紫外線の強度は、照射するＸ線の強度およびＸ線の照射時間で制御できる。 As shown in FIG. 2, two identical cells a and b are partitioned by a light-shielding partition plate 210. The cells a are attached with particles of a scintillator 200 that emits ultraviolet rays having a wavelength of 200 to 300 nm when irradiated with X-rays. The size of the cells a and b is about 10 μm in diameter, and the diameter of the fine particles is about 100 nm. When the cells a and b are irradiated with X-rays, the scintillator 200 emits ultraviolet rays in the cells a, so that the cells a in the vicinity are destroyed and die. Similar to the experiment shown in FIG. 1, the intensity of ultraviolet rays can be controlled by the intensity of X-rays to be irradiated and the irradiation time of X-rays.
　一方、細胞ｂでは、Ｘ線が細胞ｂを透過するのみで相互作用をしないため、細胞ｂは死滅しない。細胞ａに付着したシンチレーター２００からの紫外線の強さは、シンチレーター２００からの距離に対する逆２乗則で急激に減衰するため、仮に仕切り板２１０がなくても、細胞ｂへの殺傷作用は極めて小さいと考えられる。 On the other hand, in the cell b, the X-ray only permeates the cell b and does not interact with the cell b, so that the cell b does not die. Since the intensity of ultraviolet rays from the scintillator 200 attached to the cell a is rapidly attenuated by the inverse square law with respect to the distance from the scintillator 200, the killing effect on the cell b is extremely small even if the partition plate 210 is not present. it is conceivable that.
　ここで、細胞殺傷作用のトリガーとしてＸ線を使用するメリットについて、光子エネルギーの観点から説明する。近赤外光の波長は、６６０～７４０ｎｍであることが知られている。一方、Ｘ線の波長は、診断用Ｘ線の領域では、近赤外光の波長に対して１／１０００～１／１００００の長さである。すなわちＸ線の光子エネルギーは、近赤外線の光子エンルギーよりも１０００～１００００倍大きい。例えば、Ｘ線の波長を０．１８４Å（１．８４ｎｍ）とすると、Ｘ線の光子エネルギーは、波長７００ｎｍの光（近赤外光）の光子エネルギーより、約４００００倍大きいことになる。 Here, the merit of using X-rays as a trigger for cell killing action will be explained from the viewpoint of photon energy. The wavelength of near-infrared light is known to be 660 to 740 nm. On the other hand, the wavelength of X-rays is 1/1000 to 1/10000 of the wavelength of near-infrared light in the region of diagnostic X-rays. That is, the photon energy of X-rays is 1000 to 10,000 times larger than that of near-infrared photon energy. For example, assuming that the wavelength of X-rays is 0.184 Å (1.84 nm), the photon energy of X-rays is about 40,000 times larger than the photon energy of light (near infrared light) having a wavelength of 700 nm.
　シンチレーターのような蛍光体を発光させる場合、原理的に励起光より光子エネルギーの高い（波長の短い）光を発生させることはできない。紫外線は、生体を構成する分子の結合に直接相互作用し、結合を切断して分子を破壊できる。一方、近赤外線のエネルギーは生体を構成する分子の熱振動の領域でしかなく、近赤外線は生体を構成する分子に対して直接の破壊作用を発揮できない。紫外線は、赤外線より短波長であるが、Ｘ線に対しては遥かに長波長である。よって、Ｘ線の光子エネルギーから紫外線を発生させることができるため、直接的な細胞殺傷作用が発現する。 When emitting a phosphor such as a scintillator, in principle, it is not possible to generate light with higher photon energy (shorter wavelength) than excitation light. Ultraviolet rays directly interact with the bonds of the molecules that make up the living body, and can break the bonds and destroy the molecules. On the other hand, the energy of near-infrared rays is only in the region of thermal vibration of the molecules constituting the living body, and the near-infrared rays cannot exert a direct destructive action on the molecules constituting the living body. Ultraviolet light has a shorter wavelength than infrared light, but has a much longer wavelength than X-ray. Therefore, since ultraviolet rays can be generated from the photon energy of X-rays, a direct cell-killing action is exhibited.
　Ｘ線は、発生装置の入手が容易であるという他に、エネルギー特性の点からも有効である。線のエネルギーの下限が２０ｋｅＶより低くなると、Ｘ線といえども透過率が下がり、遮蔽物質による減衰の影響が大きくなる傾向がある。また１５０ｋｅＶを超えると、今度は透過性が大きくなりシンチレーターに吸収されにくくなるため、紫外線の発光強度が小さくなり殺菌殺細胞効果が低減してしまうおそれがある。 X-rays are effective not only because the generator is easily available, but also because of their energy characteristics. When the lower limit of the energy of the line is lower than 20 keV, the transmittance of X-rays tends to decrease, and the influence of attenuation by the shielding material tends to increase. On the other hand, if it exceeds 150 keV, the permeability becomes large and it becomes difficult to be absorbed by the scintillator, so that the emission intensity of ultraviolet rays becomes small and the bactericidal cell killing effect may be reduced.
　以下、実施例により本発明をさらに詳細に述べるが、本発明はこれらの実施例に限定されない。 Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited to these Examples.
　（実施例１）
　実施例１では、遮蔽物がある場合の例を記す。歯科診療では、虫歯を治療する際、タービンと呼ばれる研削機で患部を削った後、金属製やプラスチック製のクラウン（被せ物）で被覆する治療が行われている。しかしながら、治療の際に僅かな細菌が混入すると、数年経過後に内部で虫歯が再発することがある。 (Example 1)
In the first embodiment, an example when there is a shield will be described. In dental practice, when treating dental caries, the affected area is scraped with a grinding machine called a turbine and then covered with a metal or plastic crown (cover). However, if a small amount of bacteria is mixed in during treatment, dental caries may recur internally after several years.
　従来、歯を一度被覆してしまうと、歯の内部を殺菌処置できないが、上述した方法を採用すれば被覆後でも殺菌処置できる。具体的には、クラウンの接着剤に２００～３００ｎｍのＸ線を照射することで紫外線を発光するシンチレーターを混入させて虫歯を治療する。虫歯が再発したら、接着剤中のシンチレーターにＸ線を照射することで接着面付近で紫外線が発光し、残存する細菌を死滅できる。Ｘ線の照射には、多くの歯科に備えられているコンピュータ断層撮影（ＣＴ撮影）用のＸ線発生装置を使用できる。 Conventionally, once a tooth is covered, the inside of the tooth cannot be sterilized, but if the above method is adopted, sterilization can be performed even after covering. Specifically, a scintillator that emits ultraviolet rays by irradiating the adhesive of the crown with X-rays of 200 to 300 nm is mixed to treat dental caries. When the tooth decay recurs, the scintillator in the adhesive is irradiated with X-rays to emit ultraviolet rays near the adhesive surface, and the remaining bacteria can be killed. For X-ray irradiation, an X-ray generator for computer tomography (CT imaging) provided in many dentistry can be used.
　この際、歯とクラウンとの接合部を中心にＸ線発生装置のアームを回転させれば、照射対象部分以外の被曝を最小に抑えることができる。なお、シンチレーターの漏れやクラウンが外れた場合の誤飲を考慮し、シンチレーターは人体に無害な材料で構成されている。あるいは安全性に懸念がある場合は、シンチレータを非水溶性の樹脂やシリコンで被覆し、漏れ出しを防止する構成とする。 At this time, if the arm of the X-ray generator is rotated around the joint between the tooth and the crown, the exposure other than the irradiation target portion can be minimized. The scintillator is made of a material that is harmless to the human body in consideration of leakage of the scintillator and accidental ingestion when the crown comes off. Alternatively, if there is a concern about safety, the scintillator is coated with a water-insoluble resin or silicon to prevent leakage.
　（実施例２）
　実施例２では、選択的殺傷作用に関する例を記す。具体的には、被検体中にがん細胞などの選択的に殺傷したい目的細胞がある場合である。がん細胞の細胞膜表面には、特有のタンパク質（受容体）が発現する場合が多い。例えば、がん細胞に発現する特有のタンパク質の例には、乳がんのヒト上皮成長因子受容体２型（ＨＥＲ２）、大腸がんの上皮成長因子受容体（ＥＧＦＲ）が含まれる。これらのタンパク質を細胞膜上の標的分子（抗原）として、抗原抗体反応を利用して、シンチレーターを細胞膜に付着させる。例えば、シンチレーターを樹脂またはシリコーンで表面修飾し、ポリエチレングリコール（ＰＥＧ）、アビジン、ビオチンを介して抗体と結合させ、試薬としたシンチレーター粒子を使用できる。抗体は、一次抗体と二次抗体の組み合わせでもよい。 (Example 2)
In Example 2, an example relating to the selective killing action will be described. Specifically, there is a case where there is a target cell such as a cancer cell to be selectively killed in the subject. Unique proteins (receptors) are often expressed on the cell membrane surface of cancer cells. For example, examples of specific proteins expressed in cancer cells include human epidermal growth factor receptor type 2 (HER2) for breast cancer and epidermal growth factor receptor (EGFR) for colon cancer. Using these proteins as target molecules (antigens) on the cell membrane, the scintillator is attached to the cell membrane by utilizing the antigen-antibody reaction. For example, scintillator particles obtained by surface-modifying a scintillator with a resin or silicone and binding to an antibody via polyethylene glycol (PEG), avidin, or biotin can be used as a reagent. The antibody may be a combination of a primary antibody and a secondary antibody.
　がん細胞への選択的な付着方法としては、上記抗原抗体反応を利用したものの他に、ＥＰＲ（ｅｎｈａｎｃｅｄ　ｐｅｒｍｅａｂｉｌｉｔｙ　ａｎｄ　ｒｅｔｅｎｔｉｏｎ）効果によるものも知られており、目的細胞の位置や種類により使い分ければよい。 As a method of selectively adhering to cancer cells, in addition to the method using the above-mentioned antigen-antibody reaction, the method by EPR (enhanced permeability and retention) effect is also known. good.
　実施例２のような選択的殺傷作用は、ｉｎ　ｖｉｔｒｏでも適用できるし、ｉｎ　ｖｉｖｏでも適用できる。シンチレーターは、選択的殺傷作用をｉｎ　ｖｉｖｏで使用する場合、人体に無害な材質で構成されている。シンチレーターは、安全性に懸念がある場合、非水溶性の樹脂やシリコンで被覆し、漏れ出しを防止する構成である。シンチレーターをｉｎ　ｖｉｖｏで使用する場合は、Ｘ線の照射位置は、ＣＴのように照射対象部分を中心としたリング状の複数位置にすれば、殺傷対象部分以外への被曝を最小に抑えることができる。 The selective killing action as in Example 2 can be applied in vitro or in vivo. The scintillator is made of a material that is harmless to the human body when the selective killing action is used in vivo. When there is a concern about safety, the scintillator is coated with a water-insoluble resin or silicon to prevent leakage. When using the scintillator in vivo, if the X-ray irradiation position is set to multiple ring-shaped positions centered on the irradiation target part as in CT, exposure to areas other than the killed target part can be minimized. can.
　ここで、実施例２のような被検体内の腫瘍組織細胞への適用において、細胞殺傷効果に必要なＸ線の強度について考察する。 Here, the intensity of X-rays required for the cell-killing effect in application to tumor histiocytes in a subject as in Example 2 will be considered.
　前述したように、Ｘ線の光子エネルギーは、紫外線の光子エネルギーより約１００００倍以上大きい。具体的には、理論上、Ｘ線の光子１個につき約１００００個の紫外線の光子が発生する。しかし、実際には、Ｘ線の光子から発生する紫外線の光子の変換効率は、熱エネルギーとしての損失などがあるため、数％程度である。例えば、変換効率を低く見積もって１％としても、Ｘ線の光子１個から１００個の紫外線の光子が発生する。紫外線の光子が１個でも細胞核内の分子を破壊するエネルギーを有することを考えると、細胞に付着させたシンチレーターとＸ線光子とが反応しさえすれば、付着された細胞および近接する細胞を殺傷できる。 As mentioned above, the photon energy of X-rays is about 10,000 times or more larger than the photon energy of ultraviolet rays. Specifically, theoretically, about 10,000 ultraviolet photons are generated for each X-ray photon. However, in reality, the conversion efficiency of ultraviolet photons generated from X-ray photons is about several percent due to the loss of heat energy and the like. For example, even if the conversion efficiency is underestimated to 1%, 1 to 100 ultraviolet photons of X-rays are generated. Given that even a single UV photon has the energy to destroy molecules in the cell nucleus, as long as the scintillator attached to the cell reacts with the X-ray photon, the attached cell and nearby cells will be killed. can.
　よって、Ｘ線の光子とシンチレーターとが反応できるかが問題となる。Ｘ線の光子がシンチレーターと反応するために必要なＸ線の強度は、計算が困難である。ここでは、１細胞の投影面積あたりに１個のＸ線の光子が透過することを最低限の必要条件として仮定する。細胞の大きさを半径ｒｃとすると、細胞１個あたりの投影面積（Ｓｃ）は、Ｓｃ＝π・ｒｃ ２で表すことができる。 Therefore, the question is whether the X-ray photons and the scintillator can react. The intensity of X-rays required for X-ray photons to react with the scintillator is difficult to calculate. Here, it is assumed that one X-ray photon is transmitted per projected area of one cell as a minimum requirement. Assuming that the size of the cell is a radius r c , the projected area (S c ) per cell can be expressed by S c = π · r c 2.
　一方、Ｘ線の光子数（Ｎ）は、診断領域の代表的な線質としてＲＱＡ５（ＩＥＣ６２２２０－１などに示されるように、管電圧７０ｋＶで人体透過後を想定しアルミフィルターを２１ｍｍ付加したときの線質）を仮定すると、１μＧｙあたりＮ＝３．０×１０４個／ｍｍ２となる。よって投影面積（Ｓｃ）あたりＸ線の光子が１個通過するためのＸ線の強度Ｐ（μＧｙ）は、Ｐ・Ｓｃ・Ｎ＝１により算出される。ここで、細胞の半径を５μｍと仮定し、長さの単位をｍｍに統一すると、ｒｃ＝５μｍ＝５×１０－３ｍｍ。これらより、Ｐ＝１／（Ｐ・Ｓｃ）＝１／（Ｎ・π・ｒｃ ２）＝０．４２（μＧｙ）と算出される。 On the other hand, the number of photons (N) of X-rays is when an aluminum filter is added by 21 mm assuming that the tube voltage is 70 kV and the tube voltage is 70 kV, as shown in RQA5 (IEC6220-1) as a typical radiation quality in the diagnostic region. Assuming the quality of the line), N = 3.0 × 10 4 pieces / mm 2 per 1 μGy. Thus the projected area (S c) the intensity of X-rays for photon per X-rays pass through one P (μGy) is calculated by P · S c · N = 1 . Here, if the radius of the cell assuming 5 [mu] m, to unify the unit of length in mm, r c = 5μm = 5 × 10 -3 mm. From these, it is calculated that P = 1 / (P · S c ) = 1 / (N · π · r c 2 ) = 0.42 (μGy).
　例えば、ヒトのＸ線検査において、腰椎側面を撮影する際のＸ線強度が１５ｍＧｙ以下（日本診療放射線技師会ガイドライン参照）であることを考慮すると、算出されたＸ線の強度（０．４２μＧｙ）は、これより４桁以上低い値ということになる。細胞が生体を含むことを考えると、被曝の観点から、Ｘ線強度は低い方が望ましく、上記であれば問題ないと言える。 For example, considering that the X-ray intensity when photographing the lateral surface of the lumbar spine is 15 mGy or less (see the guidelines of the Japan Association of Radiological Technologists) in human X-ray examination, the calculated X-ray intensity (0.42 μGy). Is a value that is four or more digits lower than this. Considering that cells contain a living body, it is desirable that the X-ray intensity is low from the viewpoint of radiation exposure, and it can be said that there is no problem if it is the above.
　ただし、前述の算出プロセスでは、数桁レベルで算出値が振れる可能性がある。多くのＸ線を照射する必要が生じる可能性として、下記事項が考えられる。
　（１）細胞が生体のどの位置に存在するか。例えば、Ｘ線を照射する側から見たときに、細胞が奥側に存在する場合は、生体によるＸ線の減衰を１桁程度考慮して多くのＸ線を照射する必要がある。
　（２）Ｘ線の光子が細胞に付着したシンチレーターと反応するか。細胞に付着したシンチレーターにＸ線の光子を確実に反応させるために、３～５桁程度考慮して多くのＸ線を照射する必要がある。 However, in the above-mentioned calculation process, the calculated value may fluctuate at the level of several digits. The following may be considered as the possibility that it is necessary to irradiate a large amount of X-rays.
(1) Where are the cells in the living body? For example, when cells are present on the back side when viewed from the side to be irradiated with X-rays, it is necessary to irradiate a large amount of X-rays in consideration of the attenuation of X-rays by a living body by about an order of magnitude.
(2) Does the X-ray photon react with the scintillator attached to the cell? In order to reliably react the X-ray photons with the scintillator attached to the cells, it is necessary to irradiate a large number of X-rays in consideration of about 3 to 5 digits.
　一方、少ないＸ線の強度でよい可能性として、下記事項が考えられる。
　（１）腫瘍では、奥行き方向に厚みがあり、Ｘ線の光子の通り道に存在する細胞数が多い。厚み１０ｍｍの腫瘍であれば、奥行き方向に１０００個の細胞が存在することになる。このいずれかの細胞に付着したシンチレーター粒子とＸ線の光子とが反応すればよいと考えると、シンチレーター粒子とＸ線光子とが反応する確率は３桁増加するため、Ｘ線の強度を低くできる。
　（２）Ｘ線照射を複数回に分割する。現在、がん治療で行われている放射線治療では、通常、１回ではなく数回から十数回に日を分けてＸ線を照射している。放射線治療と同様の手法を採用すれば、１回に照射するＸ線の強度は、１桁程度減らすことができる。
　（３）細胞に対するシンチレーター粒子の付着条件を検討すること。付着条件を最適化することにより、細胞に対するシンチレーター粒子の付着数は、２～３桁個分増やすことができるため、照射するＸ線の強度を低くできる。 On the other hand, the following items can be considered as the possibility that a small amount of X-ray intensity is sufficient.
(1) Tumors are thick in the depth direction and have a large number of cells present in the path of X-ray photons. In the case of a tumor with a thickness of 10 mm, 1000 cells will be present in the depth direction. Considering that the scintillator particles attached to any of these cells should react with the X-ray photons, the probability of the scintillator particles reacting with the X-ray photons increases by three orders of magnitude, so that the X-ray intensity can be reduced. ..
(2) The X-ray irradiation is divided into a plurality of times. Currently, in radiation therapy performed in cancer treatment, X-rays are usually irradiated several times to a dozen times instead of once. If the same method as radiation therapy is adopted, the intensity of X-rays irradiated at one time can be reduced by about an order of magnitude.
(3) Examine the conditions for attachment of scintillator particles to cells. By optimizing the adhesion conditions, the number of scintillator particles attached to the cells can be increased by 2 to 3 orders of magnitude, so that the intensity of the irradiated X-rays can be reduced.
　このように、見積もりの振れ幅は大きいが、総合的に現在人へのＸ線検査において使用されるＸ線の強度に対し、極端に強いＸ線の強度が必要となる可能性は低いと考えられる。 In this way, although the estimated fluctuation range is large, it is unlikely that an extremely strong X-ray intensity will be required for the X-ray intensity currently used in X-ray inspections for humans. Be done.
　本出願は、２０２０年７月８日出願の特願２０２０－１１７７４７に基づく優先権を主張する。当該出願明細書および図面に記載された内容は、すべて本願明細書に援用される。 This application claims priority based on Japanese Patent Application No. 2020-117747 filed on July 8, 2020. All the contents described in the application specification and drawings are incorporated herein by reference.
　本実施の形態に係る殺傷方法は、目的細胞に対して適切に紫外線を照射できるため、狭い領域を適切に殺菌することや、生体内における目的細胞のみの殺傷に有用である。 Since the killing method according to the present embodiment can appropriately irradiate the target cells with ultraviolet rays, it is useful for appropriately sterilizing a narrow area and killing only the target cells in the living body.
　１００　シャーレ
　１１０　目的細胞
　１２０　シンチレータープレート
　２００　シンチレーター
　２１０　仕切り板
　ａ、ｂ　細胞 100 Petri dish 110 Target cell 120 Scintillator plate 200 Scintillator 210 Divider a, b cells

Claims (7)

  1. 　シンチレーターにＸ線を照射して、前記シンチレーターから紫外線を発生させる工程と、
    　発生した紫外線を目的細胞に到達させて、前記目的細胞を殺傷する工程と、
    　を含む、細胞の殺傷方法。     The process of irradiating the scintillator with X-rays to generate ultraviolet rays from the scintillator,
    The process of causing the generated ultraviolet rays to reach the target cells and killing the target cells,
    How to kill cells, including.
  2. 　前記シンチレーターにＸ線を照射する前に、前記目的細胞に特異的に結合する標的物質認識物質を有する前記シンチレーターで前記目的細胞を標識する工程をさらに有する、請求項１に記載の細胞の殺傷方法。 The method for killing cells according to claim 1, further comprising a step of labeling the target cells with the scintillator having a target substance recognition substance that specifically binds to the target cells before irradiating the scintillator with X-rays. ..
  3. 　前記シンチレーターから発生する紫外線は、波長が２００～３００ｎｍの紫外線を含む、請求項１または請求項２に記載の細胞の殺傷方法。 The method for killing cells according to claim 1 or 2, wherein the ultraviolet rays generated from the scintillator include ultraviolet rays having a wavelength of 200 to 300 nm.
  4. 　前記シンチレーターに照射されるＸ線のエネルギーは、２０～１５０ｅＶである、請求項１～３のいずれか一項に記載の細胞の殺傷方法。 The method for killing cells according to any one of claims 1 to 3, wherein the energy of the X-ray irradiated to the scintillator is 20 to 150 eV.
  5. 　前記シンチレーターは、直径が３０～３００ｎｍの球形状または一辺が３０～３００ｎｍの直方体形状である、請求項１～４のいずれか一項に記載の細胞の殺傷方法。 The method for killing cells according to any one of claims 1 to 4, wherein the scintillator has a spherical shape having a diameter of 30 to 300 nm or a rectangular parallelepiped shape having a side of 30 to 300 nm.
  6. 　前記シンチレーターは、その表面が樹脂またはシリコーンで修飾されている、請求項１～５のいずれか一項に記載の細胞の殺傷方法。 The method for killing cells according to any one of claims 1 to 5, wherein the surface of the scintillator is modified with resin or silicone.
  7. 　シンチレーターおよび樹脂を含む粒子と、
    　前記粒子の表面に担持された、目的細胞に特異的に結合する標的物質認識物質と、
    　を有する、シンチレーター粒子。     With particles containing scintillators and resins,
    A target substance recognition substance that is supported on the surface of the particles and specifically binds to the target cell,
    Has scintillator particles.
PCT/JP2021/025048 2020-07-08 2021-07-01 Cell killing method and scintillator particles WO2022009788A1 (en)

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JP2019523757A (en) * 2016-05-20 2019-08-29 ザ ユニバーシティ オブ シカゴＴｈｅ Ｕｎｉｖｅｒｓｉｔｙ Ｏｆ Ｃｈｉｃａｇｏ Nanoparticles for chemotherapy, targeted therapy, photodynamic therapy, immunotherapy and any combination thereof
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Publication number Priority date Publication date Assignee Title
JP2011078750A (en) * 2009-09-09 2011-04-21 Tokai Univ Method for killing tumor cells selectively and apparatus for the method
JP2017532340A (en) * 2014-10-14 2017-11-02 ザ ユニバーシティ オブ シカゴＴｈｅ Ｕｎｉｖｅｒｓｉｔｙ Ｏｆ Ｃｈｉｃａｇｏ Nanoparticles for photodynamic therapy, X-ray induced photodynamic therapy, radiation therapy, chemotherapy, immunotherapy, and any combination thereof
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