WO2014030715A1 - ホウ素クラスター修飾peg脂質誘導体およびこれを用いた分子集合体 - Google Patents
ホウ素クラスター修飾peg脂質誘導体およびこれを用いた分子集合体 Download PDFInfo
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- WO2014030715A1 WO2014030715A1 PCT/JP2013/072458 JP2013072458W WO2014030715A1 WO 2014030715 A1 WO2014030715 A1 WO 2014030715A1 JP 2013072458 W JP2013072458 W JP 2013072458W WO 2014030715 A1 WO2014030715 A1 WO 2014030715A1
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- WIPO (PCT)
- Prior art keywords
- boron
- molecular assembly
- lipid
- peg
- lipid derivative
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- NRHMKIHPTBHXPF-TUJRSCDTSA-M sodium cholate Chemical compound [Na+].C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC([O-])=O)C)[C@@]2(C)[C@@H](O)C1 NRHMKIHPTBHXPF-TUJRSCDTSA-M 0.000 description 1
- 238000000527 sonication Methods 0.000 description 1
- 229940083466 soybean lecithin Drugs 0.000 description 1
- 239000012192 staining solution Substances 0.000 description 1
- 150000003432 sterols Chemical class 0.000 description 1
- 235000003702 sterols Nutrition 0.000 description 1
- 125000001424 substituent group Chemical group 0.000 description 1
- PXQLVRUNWNTZOS-UHFFFAOYSA-N sulfanyl Chemical class [SH] PXQLVRUNWNTZOS-UHFFFAOYSA-N 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 238000009210 therapy by ultrasound Methods 0.000 description 1
- 125000003396 thiol group Chemical group [H]S* 0.000 description 1
- 238000001269 time-of-flight mass spectrometry Methods 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000820 toxicity test Toxicity 0.000 description 1
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
- 230000036326 tumor accumulation Effects 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
- 238000002525 ultrasonication Methods 0.000 description 1
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Images
Classifications
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- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G65/00—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule
- C08G65/02—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring
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- C08G65/331—Polymers modified by chemical after-treatment with organic compounds containing oxygen
- C08G65/332—Polymers modified by chemical after-treatment with organic compounds containing oxygen containing carboxyl groups, or halides, or esters thereof
- C08G65/3322—Polymers modified by chemical after-treatment with organic compounds containing oxygen containing carboxyl groups, or halides, or esters thereof acyclic
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- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
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- C08G65/33324—Polymers modified by chemical after-treatment with organic compounds containing nitrogen containing carboxamide group acyclic
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- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
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- C08G65/02—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring
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- C08G65/335—Polymers modified by chemical after-treatment with organic compounds containing phosphorus
- C08G65/3353—Polymers modified by chemical after-treatment with organic compounds containing phosphorus containing oxygen in addition to phosphorus
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
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- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
- A61K47/6905—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a colloid or an emulsion
- A61K47/6911—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a colloid or an emulsion the form being a liposome
Definitions
- the present invention relates to a PEG lipid derivative useful as a drug carrier that recognizes tumor tissue and enables boron delivery, and a molecular assembly using the PEG lipid derivative.
- the boron neutron capture therapy for cancer (hereinafter also referred to as “BNCT”) is a 10 B nucleus previously incorporated into a tumor tissue and an ⁇ particle and a 7 Li particle produced by a thermal neutron capture reaction that does not substantially affect the living body. Is radiation therapy that damages tumor cells. Since the range of the particle beam generated as a result of this nuclear reaction is approximately the same as the cell diameter, if high concentrations of boron can be accumulated only in the tumor tissue beforehand, thermal neutrons can be irradiated on it to minimize damage to normal tissue. However, it is expected to have a therapeutic effect even in tumors with low radiosensitivity and invasive tumors by selectively damaging tumor tissues.
- Non-patent Document 1 Clinical studies have been conducted on malignant brain tumors (glioblastoma) that are clinically invasive and have a poor prognosis, and recurrent cases of head and neck cancer, and good results have been reported (Non-patent Document 1).
- the boron compound used clinically is a monomolecular compound of boron ion cluster called boronophenylalanine (BPA) and BSH. Although these compounds have been applied clinically, they are not sufficient from the viewpoint of tumor accumulation, and attempts to further improve the boron concentration ratio (T / N ratio) in tumors and normal tissues have been continued.
- BPA boronophenylalanine
- T / N ratio boron concentration ratio
- liposomes coated with polyethylene glycol (PEG) on the surface are mixed with BHA (10B enriched 4-Borono-L-phenylalanine, C 9 H 12 10 BNO 4 ) and BSH. (10B enriched Sodium mercaptododecaborate, BSH [ CAS No.12448-24-7], [10 B 12 H 11 SH] Na 2) have been used techniques for encapsulating hydrophilic boron compound such as (non-patent document 2) .
- BHA 4-Borono-L-phenylalanine
- BSH enriched Sodium mercaptododecaborate
- BSH [ CAS No.12448-24-7]
- [10 B 12 H 11 SH] Na 2 have been used techniques for encapsulating hydrophilic boron compound such as (non-patent document 2) .
- boron accumulation to an effective therapeutic area is difficult by these methods, and the achievement of the targeted T / N ratio cannot be achieved at the same time.
- Non-patent Document 3 Non-patent Document 3
- the general performance currently required for boron compounds for use in boron neutron capture therapy is as follows. i) A 10 B concentration of at least 20 ug / g or more can be achieved in the tumor. ii) On the other hand, in normal tissues, the 10 B concentration is as low as possible. iii) A Tumor / blood ratio of 5 to 10 or more is desirable. In order to achieve this concentration, unlike general pharmaceuticals, it is necessary to administer a large amount of a boron compound to a living body. Therefore, it is important that the toxicity of the boron compound is low for use in boron neutron capture therapy.
- the current compound BSH requires a rapid intravenous dose of 5 g per adult human and 15-25 g for BPA, which is 10 to 100 times the weight of common pharmaceuticals. Under such circumstances, there is an improvement in Drug Delivery System (DDS) efficiency that can be used in boron neutron capture therapy for cancer, provision of a new boron carrier compound that can achieve high concentration of boron in the tumor, and further low toxicity. It is desired.
- DDS Drug Delivery System
- the inventors of the present invention have made it possible to increase the concentration of boron in the tumor and to reduce the normal cells by using DDS by modifying the lipid to construct the molecular assembly.
- the present inventors have found that there is a high possibility of realizing integration, and have completed the present invention.
- the present invention relates to the following boron cluster-modified PEG lipid derivative and a molecular assembly including the same.
- a molecular assembly comprising the PEG lipid derivative according to any one of [1] to [3].
- the molecular assembly according to [4] comprising a PEG lipid derivative in a molar ratio of 2 to 20% with respect to all constituent lipids of the molecular assembly.
- a novel boron carrier compound that can be used for boron neutron capture therapy for cancer can be provided.
- the molecular assembly containing the boron cluster-modified PEG lipid derivative of the present invention high concentration of boron in the tumor and low integration into normal cells by using DDS, Is likely to be realized.
- the present inventors have confirmed that there is no acute toxicity up to a boron concentration of 2000 ppm (215.6 mg / ml as a lipid concentration) and that there is no abnormality due to physical findings. Therefore, it can be said that the molecular assembly of the present invention is a boron carrier compound that has low toxicity and can be suitably used for boron neutron capture therapy.
- FIG. 2 is a mass spectrum of the compound obtained in Example 1.
- 2 is an enlarged view of a peak in a mass spectrum of the compound obtained in Example 1.
- FIG. 2 is a high performance liquid chromatograph of the compound obtained in Example 1.
- 4 is a graph showing an example of each fraction separated by gel exclusion chromatography obtained in Example 3.
- FIG. 4 is a graph showing the incorporation rate of boron cluster-modified PEG lipid derivatives in each liposome obtained in Example 3.
- FIG. It is a transmission electron micrograph (acceleration voltage: 100 kV) of PEG-Boron-lipid 5% liposome.
- FIG. 2 is a chart of 1 H-NMR spectrum of the compound obtained in Example 5.
- 6 is a chart of 10 B-NMR spectrum of the compound obtained in Example 5.
- 4 is a mass spectrum of the compound obtained in Example 5.
- 6 is an enlarged view of a peak in a mass spectrum of a compound obtained in Example 5.
- FIG. 2 is a high performance liquid chromatograph of the compound obtained in Example 5. It is the graph which showed the cytotoxic reaction when a thermal neutron beam was irradiated to the PEG-Boron-lipid modified liposome. It is the graph which showed the antitumor effect when a thermal neutron beam was irradiated to a PEG-Boron-lipid modified liposome.
- the boron cluster-modified PEG lipid derivative of the present invention has the formula (I): [Wherein, m and n are each independently an integer of 1 to 4, q is an integer of 1 to 280, and R 1 and R 2 are each independently an integer of 8 to 22 carbon atoms] It is a hydrocarbon group. ] It is a compound shown by these.
- the boron cluster-modified PEG lipid derivative of the present invention can be used as a constituent lipid of a molecular assembly such as a liposome because it has a structure in which a boron cluster ( 10 B 12 H 13 ) is modified to PEG lipid. It can be used as a boron carrier compound used for BNCT.
- m and n are each independently an integer of 1 to 4, preferably 1 or 2.
- M is preferably 2 and n is preferably 1 because raw materials are easily available.
- q is an integer of 1 to 280, preferably an integer of 10 to 114, and more preferably an integer of 44 to 66. Within this range, retention in blood is most easily obtained due to the thickness of the hydrated phase by PEG and steric hindrance due to movement.
- R 1 and R 2 are each independently a hydrocarbon group having 8 to 22 carbon atoms. The hydrocarbon group may be linear or branched, but is preferably linear.
- the hydrocarbon group may have a substituent selected from the group consisting of a carboxyl group, a hydroxyl group, an amino group, and a mercapto group.
- the number of carbon atoms of the hydrocarbon group is preferably 12-20, more preferably 14-18.
- the hydrocarbon group may have an unsaturated bond such as a double bond or a triple bond, in which case the number is preferably 1 to 4.
- R 1 and R 2 a linear or branched alkyl group having 12 to 22 carbon atoms is preferable, a linear alkyl group having 14 to 18 carbon atoms is more preferable, and a linear chain having 17 carbon atoms is preferable.
- the alkyl group is particularly preferred.
- the boron cluster-modified PEG lipid derivative of the present invention has the formula (II): [Wherein m, n, q, R 1 and R 2 have the same meaning as in formula (I). ] And a malechemi derivative represented by Katchem spol. sr. o. , Ltd., Ltd. 10B enriched Sodium mercapto dodecaborate (BSH) [CAS No. 12448-24-7] and [ 10 B 12 H 11 SH] Na 2 can be easily produced by reacting them in phosphate buffered saline. After the reaction, the desired compound can be obtained by freeze-drying, and then dissolving the crude product with ultrapure water, increasing the purity by dialysis, and freeze-drying.
- BSH Sodium mercapto dodecaborate
- the compound represented by the formula (II) can be produced, for example, by introducing a hydrophobic alkyl chain at the end of an amino acid to form a transmembrane type and bonding maleimide PEG to the end. Moreover, a commercial item can also be used as a compound shown by Formula (II).
- SUNBRIGHT registered trademark
- SERIES DSPE-020MA DSPE-050MA
- DSPE-PEG-MAL compound name: N-[(3-Maleimide-1-oxypropylidyl) aminopropylene) sold by NOF CORPORATION -Ethanolamine
- DEPE-PEG (2000) Maleimide Compound name: 1,2-distearoyl-sn-glycero-3-phosphoeethanolamine-N- [mal (polyethylene) glycol)-sold by Avanti Polar Lipids, Inc. am monium salt
- the boron cluster-modified PEG lipid derivative of the present invention can be used as a constituent lipid of a molecular assembly.
- the molecular assembly of the present invention is not particularly limited as long as it contains the boron cluster-modified PEG lipid derivative as a constituent lipid.
- the amount of the boron cluster-modified PEG lipid derivative used in the molecular assembly of the present invention is not particularly limited, but is preferably 2 to 20%, more preferably 3 to 15% in terms of molar ratio with respect to the total constituent lipid of the molecular assembly. Preferably, 5 to 10% is more preferable.
- the molecular assembly of the present invention may contain other constituent lipids other than the boron cluster-modified PEG lipid derivative.
- other constituent lipids include phosphatidylcholine (dimyristoylphosphatidylcholine (DMPC), L- ⁇ -phosphatidylcholine diasteroyl (DSPC), etc.), phosphatidylethanolamine, phosphatidylserine, phosphatidic acid, phosphatidylglycerol, phosphatidylinositol, cardiolipin,
- glycerophospholipids such as egg yolk lecithin, hydrogenated egg yolk lecithin, soybean lecithin, hydrogenated soybean lecithin; and sphingophospholipids such as sphingoemilin, ceramide phosphorylethanolamine, and ceramide phosphorylglycerol.
- phosphatidylcholine is preferable, and L- ⁇ -phosphatidylcholine diasteroyl (DSPC) is particularly preferable.
- the amount of phospholipid used in the molecular assembly of the present invention is preferably 40 to 95%, more preferably 40 to 70%, still more preferably 40 to 50% in terms of molar ratio to the total constituent lipid of the molecular assembly. .
- steroids are mentioned as another structural lipid.
- examples of sterols include ergosterol, cholesterol, triglyceride and the like. Of these, cholesterol is preferable.
- the amount of steroids used in the molecular assembly of the present invention is preferably 0 to 50%, more preferably 30 to 50%, still more preferably 40 to 50% in terms of a molar ratio with respect to all the constituent lipids of the molecular assembly. .
- the molecular assembly of the present invention includes glyceroglycolipids such as digalactosyl diglyceride and galactosyl diglyceride sulfate; galactosylceramide, galactosylceramide sulfate, lactosylceramide, ganglioside G7, ganglioside G6, ganglioside G4 and other sphingoglycolipids.
- Glycolipids such as: etc. may be included. Of these, ganglioside G4 is preferred.
- the amount of the glycolipid is preferably 4 to 24% in terms of a molar ratio with respect to all the constituent lipids of the molecular assembly.
- the form of the molecular assembly of the present invention is, for example, a polymer assembly, polymer micelle, emulsion, lipid microsphere, bilayer vesicle (liposome), other molecular assembly (tube, fiber, ribbon, sheet, etc.) ) And the like.
- the molecular assembly of the present invention is preferably in the form of liposomes.
- FIG. 1 is a conceptual diagram of a liposome containing a boron cluster-modified PEG lipid derivative according to an embodiment of the present invention.
- PEG surrounds the surface of the lipid bilayer vesicle formed by the constituent lipid, and boron clusters are further bound to the surface. It is done.
- the particle diameter of the liposome measured by a dynamic light scattering method is preferably 50 to 400 nm, more preferably 50 to 200 nm, and further preferably 100 to 200 nm.
- the molecular assembly of the present invention can be produced according to a known method.
- the liposome production method includes a probe method and a bath method in which an aqueous solvent is added to a single or mixed lipid thin film, followed by hydration / swell and ultrasonication.
- an agitation method vortex mixing, homogenizer
- an ethanol injection method or an ether injection method in which a single or mixed lipid is dissolved in an organic solvent and the solution is injected into an aqueous phase can be mentioned.
- a single or mixed lipid film can be dispersed in an aqueous phase with a nonionic surfactant such as sodium cholate, sodium dodecyl sulfate, Triton X, octyl glycoside or lauryl ether to form an emulsion and removed by dialysis.
- a nonionic surfactant such as sodium cholate, sodium dodecyl sulfate, Triton X, octyl glycoside or lauryl ether
- Liposomes can also be produced.
- an aqueous solvent is added to a thin film of single or mixed lipids, vigorously suspended, and the resulting solution is sonicated.
- a freeze-thaw method in which freezing and thawing with liquid nitrogen is then performed can be mentioned.
- single or mixed lipids are dissolved in an organic solvent, and a small amount of an aqueous solvent is added to the solution to give ultrasonic vibration.
- an aqueous solvent is added to the solution to give ultrasonic vibration.
- REV method reverse phase evaporation method
- an organic solvent such as ethanol or ether is removed by decompression or dialysis to produce liposomes.
- the particle size of the liposomes obtained above is adjusted by a high-pressure extrusion method, a French press method, or the like.
- the molecular assembly of the present invention containing a boron cluster-modified PEG lipid derivative as a constituent lipid can be produced.
- the molecular assembly of the present invention can efficiently incorporate 10 B nuclei contained in a boron cluster-modified PEG lipid derivative into tumor tissue by using DDS.
- the molecular assembly of the present invention can be suitably used for boron neutron capture therapy.
- the molecular assembly of the present invention can be administered, for example, orally, parenterally, intravenously, transdermally, via inhalation, or directly to the disease site.
- the dose may be within the range of the effective amount, and is usually about 1500 to about 7500 mg (lipid weight) per kg body weight, although it varies depending on the target disease, administration subject, administration method, symptoms and the like.
- the compound was identified by mass spectrum and high performance liquid chromatograph. The results are shown in FIGS. 2A, B and 3. 2A and B, the measured value of 3147.690 was confirmed with respect to 3146.07 which is the Exact mass of this compound. This has a molecular weight error (%) of 0.08%, and it can be said that this compound was successfully synthesized. Further, regarding the other peaks, there is a difference of 44 in molecular weight, which corresponds to 1mer (—CH 2 CH 2 O—) of polyethylene glycol (PEG), indicating that the synthesis of this compound was successful. I support it. Further, as shown in FIG. 3, the measured value with a peak area of 81.71% was confirmed. Therefore, it can be said that this compound was obtained with high purity by the above synthesis scheme.
- PEG polyethylene glycol
- the incorporation rate of the boron cluster-modified PEG lipid derivative into the liposome was calculated. As a result, it was found that the boron cluster-modified PEG lipid derivative was efficiently incorporated into each liposome obtained in Example 2.
- the incorporation rate of the boron cluster-modified PEG lipid derivative with respect to each liposome obtained in Example 2 is shown in Table 2 and FIG.
- the liposome of the present invention (PEG-Boron-lipid 5% liposome or PEG-Boron-lipid 10% liposome) containing a boron cluster-modified PEG lipid derivative is minus like PEG-modified liposome (PEG liposome).
- a zeta potential of a value of Moreover, the particle diameter of the liposome of the present invention is about 100 nm to 200 nm or less, and is controlled to the particle diameter at which the EPR effect (enhanced permeability and retention effect) is most obtained.
- FIG. 6 is a transmission electron micrograph (acceleration voltage: 100 kV) of a PEG-Boron-lipid 5% liposome (indicated by an arrow in the figure). As shown in FIG. 6, the PEG-Boron-lipid modified liposome forms a bilamella and has a monolayer structure.
- Dialysis conditions External liquid: Ultrapure water External liquid exchange: 5 times / 2 hours Room temperature, light-shielded Reagents used in PBS preparation: Sodium hydrogen phosphate dodecahydrate Potassium dihydrogen phosphate Sodium chloride Potassium chloride All the above reagents were obtained from SIGMA Aldrich.
- the compound was identified by 1 H-NMR, 10 B-NMR, mass spectrum (time-of-flight mass spectrometry, TOF-MS) and high-performance liquid chromatograph.
- the results are shown in FIGS. 7, 8, 9 and 10, respectively.
- PEG-Boron-lipid observed boron peaks
- the antitumor effect by neutron irradiation was measured by the following method (in vivo).
- the tumor bearing model was prepared by seeding mouse colon cancer cells (CT26, 5 ⁇ 10 6 cells) on BALB / cA rice (female, 4 weeks old, weighting 16-20 g) on the right thigh, and having a tumor diameter of 6-8 mm. (About 8 days after injection).
- mice were treated without pain using an isoflurane inhalation anesthesia machine.
- Prior to conducting a series of animal experiments submit an animal experiment plan to the University of Tsukuba Animal Experiment Committee, comply with the laws and standards related to animal experiments, and take into account the reduction of animal suffering, etc., and obtain approval. It was. The same procedure was performed in the Kyoto University joint use application.
- PEG-Boron-lipid 5% liposome was intravenously injected at 10 mg 10 B / kg, and neutron irradiation was performed with KUR 24 hours later.
- the neutron dose was 4.5-7.0 ⁇ 10 12 neutron / cm 2 .
- Tumor diameter after irradiation was measured for tumor growth inhibitory effect over time until day 21 and compared with the control group.
- a BSH solution group (BSH, 6 cases), a neutron irradiation only group (irradiation only, 6 cases), and a group without medication / irradiation (no treatment, 12 cases) were compared and examined.
- PEG-Boron-lipid-modified liposomes showed significant tumor growth inhibition on day 10, 14, 17, and 21 compared to any group, and tumors increased especially until day 14 I never did.
- the boron cluster-modified PEG lipid derivative of the present invention is useful as a drug carrier that enables boron delivery.
- the molecular assembly of the present invention containing the PEG lipid derivative can realize high concentration of boron in the tumor and low integration into normal cells by using DDS. It is expected to be used as a boron carrier compound in boron neutron capture therapy for cancer.
Abstract
Description
臨床で用いられているホウ素化合物は、ボロノフェニルアラニン(BPA)および、BSHとよばれるホウ素イオンクラスターの単分子化合物である。これら化合物は、臨床応用されているものの、腫瘍集積の観点からは十分とはいえず、腫瘍・正常組織におけるホウ素濃度比(T/N比)をさらに向上させる試みが続けられている。さらに腫瘍組織のみへ高濃度でホウ素を集積させる技術として、表面をポリエチレングリコール(PEG)で被覆したリポソームに、BHA(10B enriched 4-Borono-L-phenylalanine、C9H12 10BNO4)およびBSH(10B enriched Sodium mercaptododecaborate、BSH[CAS No.12448-24-7]、[10B12H11SH]Na2)などの親水性ホウ素化合物を封入する手法が用いられている(非特許文献2)。
しかしながら、これらの方法では有効な治療域までのホウ素集積が難しいこと、また、目標とされるT/N比の達成との両立ができていないため、正常組織でのホウ素による正常組織への放射線障害や治療効果の低減が大きな問題となっている。
脂質と水性ホウ素化合物を共有結合によって結合させ、リン脂質二重膜内にホウ素化合物を取り込ませる技術も開発されているが、多段階かつ複雑な合成を必要とするため、効率やコスト面から問題が多い。また動物実験において急性毒性が数例報告されており、安全面においても問題がある(非特許文献3)。
i)腫瘍内に少なくとも20ug/g以上の10B濃度を達成できる。ii)一方で正常組織において10B濃度は可能な限り低濃度である。iii)Tumor/blood比で5~10以上が望ましい。
この濃度を達成するためには、一般の医薬品と異なり、生体へのホウ素化合物の大量投与が必要である。そのため、ホウ素中性子捕捉療法に用いるためには、ホウ素化合物の毒性が低いことが重要となる。現在の化合物BSHではヒト成体あたり5g、BPAでは15~25gの急速静注投与を要しており、これは一般の医薬品の10~100倍の重量である。
このような状況下、癌のホウ素中性子捕捉療法において利用可能なDrug Delivery System(DDS)効率の向上と腫瘍内のホウ素の高濃度化、さらには低毒性を実現しうる新規ホウ素担体化合物の提供が望まれている。
[1]式(I):
[式中、mおよびnは、それぞれ独立して、1~4の整数であり、qは、1~280の整数であり、R1およびR2は、それぞれ独立して、炭素数8~22の炭化水素基である。]
で示されるホウ素クラスター修飾PEG脂質誘導体。
[2]R1およびR2が、それぞれ独立して、炭素数12~22の直鎖状または分岐状のアルキル基である、[1]記載のPEG脂質誘導体。
[3]mが2であり、かつ、nが1である、[1]または[2]記載のPEG脂質誘導体。
[4][1]から[3]の何れか1項記載のPEG脂質誘導体を含む分子集合体。
[5]PEG脂質誘導体を分子集合体の全構成脂質に対してモル比で2~20%含む、[4]記載の分子集合体。
[6]L-α-ホスファチジルコリンジアステロイルおよびコレステロールをさらに含む、[4]または[5]記載の分子集合体。
[7]リポソームである、[4]から[6]の何れか1項記載の分子集合体。
[8]動的光散乱法により測定したリポソームの粒子径が50~400nmである、[7]記載の分子集合体。
本発明のホウ素クラスター修飾PEG脂質誘導体は、式(I):
[式中、mおよびnは、それぞれ独立して、1~4の整数であり、qは、1~280の整数であり、R1およびR2は、それぞれ独立して、炭素数8~22の炭化水素基である。]
で示される化合物である。
本発明のホウ素クラスター修飾PEG脂質誘導体は、PEG脂質に、ホウ素クラスター(10B12H13)が修飾された構造を有していることにより、リポソームなどの分子集合体の構成脂質として用いることができ、BNCTに用いるホウ素担体化合物として利用可能である。
式中、mおよびnは、それぞれ独立して、1~4の整数であり、好ましくは、1または2である。原料を入手しやすいことから、mは2であることが好ましく、nは1であることが好ましい。
qは、1~280の整数であり、好ましくは、10~114の整数であり、より好ましくは、44~66の整数である。この範囲であると、PEGによる水和相の厚さおよび運動による立体障害性から最も血中滞留性を得やすい。
R1およびR2は、それぞれ独立して、炭素数8~22の炭化水素基である。炭化水素基は、直鎖状でも分岐状でもよいが、直鎖状であることが好ましい。また、炭化水素基は、カルボキシル基、水酸基、アミノ基およびメルカプト基からなる群から選択される置換基を有していてもよい。炭化水素基の炭素数は、好ましくは12~20であり、より好ましくは14~18である。炭化水素基は、二重結合や三重結合などの不飽和結合を有していてもよく、その場合にその数は1~4であることが好ましい。中でも、R1およびR2としては、炭素数12~22の直鎖状または分岐状のアルキル基が好ましく、炭素数14~18の直鎖状のアルキル基がより好ましく、炭素数17の直鎖状のアルキル基が特に好ましい。
[式中のm、n、q、R1およびR2は、式(I)におけるものと同義である。]
で示されるマレイミド誘導体と、Katchem spol.sr.o., Ltd.より販売されている、10B enriched Sodium mercaptododecaborate(BSH)[CAS No.12448-24-7]、[10B12H11SH]Na2とを、リン酸緩衝生理食塩水中で反応させることで簡便に製造することができる。
反応後は、凍結乾燥し、その後、粗生成物を超純水で溶解後、透析により純度を上げ、凍結乾燥することで目的の化合物を得ることができる。
式(II)で示される化合物は、例えばアミノ酸の末端に疎水的なアルキル鎖を導入して、膜貫通型とし、その末端にマレイミドPEGを結合することで製造することができる。また、式(II)で示される化合物として市販品を用いることもできる。例えば、NOF CORPORATIONより販売されているSUNBRIGHT(登録商標)SERIES DSPE-020MA、DSPE-050MA、DSPE-PEG-MAL(化合物名:N-[(3-Maleimide-1-oxopropyl)aminopropyl polyethyleneglycol-carbamyl] distearoylphosphatidyl-ethanolamine)、Avanti Polar Lipids社より販売されているDEPE-PEG(2000)Maleimide(化合物名:1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[mal(polyethylene glycol)-2000](ammonium salt))などが挙げられる。
本発明のホウ素クラスター修飾PEG脂質誘導体は、分子集合体の構成脂質として用いることができる。
本発明の分子集合体は、構成脂質として、前記ホウ素クラスター修飾PEG脂質誘導体を含むものであれば特に制限されない。
本発明の分子集合体に用いられるホウ素クラスター修飾PEG脂質誘導体の量は、特に制限されないが、分子集合体の全構成脂質に対してモル比で2~20%が好ましく、3~15%がより好ましく、5~10%がさらに好ましい。ホウ素クラスター修飾PEG脂質誘導体の量を適宜調整することで、目的に応じて腫瘍内のホウ素の濃度を制御することが可能である。
他の構成脂質としては、例えば、ホスファチジルコリン(ジミリストイルホスファチジルコリン(DMPC)、L-α-ホスファチジルコリンジアステロイル(DSPC)など)、ホスファチジルエタノールアミン、ホスファチジルセリン、ホスファチジン酸、ホスファチジルグリセロール、ホスファチジルイノシトール、カルジオリピン、卵黄レシチン、水添卵黄レシチン、大豆レシチン、水添大豆レシチン等のグリセロリン脂質類;スフィンゴエミリン、セラミドホスホリルエタノールアミン、セラミドホスホリルグリセロールなどのスフィンゴリン脂質類;などのリン脂質が挙げられる。中でも、ホスファチジルコリンが好ましく、L-α-ホスファチジルコリンジアステロイル(DSPC)が特に好ましい。
本発明の分子集合体に用いられるリン脂質の量は、分子集合体の全構成脂質に対してモル比で40~95%が好ましく、40~70%がより好ましく、40~50%がさらに好ましい。
本発明の分子集合体に用いられるステロイド類の量は、分子集合体の全構成脂質に対してモル比で0~50%が好ましく、30~50%がより好ましく、40~50%がさらに好ましい。
糖脂質の量は、分子集合体の全構成脂質に対してモル比で4~24%が好ましい。
例えば、リポソームの製造方法としては、単独もしくは混合脂質の薄膜に水系溶媒を加え、水和・膨潤させ超音波処理する、プローブ法、浴槽法が挙げられる。また、単独もしくは混合脂質の薄膜に水系溶媒を加えてからボルテックス処理し、その後超音波処理する撹拌(ボルテックスミキシング、ホモジナイザー)法が挙げられる。
あるいは、単独または混合脂質を有機溶媒に溶解させ、その溶液を水相の中に注入するエタノール注入法、エーテル注入法が挙げられる。
あるいは、単独もしくは混合脂質の薄膜にコール酸ナトリウム、ドデシル硫酸ナトリウム、Triton X、オクチルグリコシドまたはラウリルエーテルなどの非イオン性界面活性剤と共に水相に分散させてエマルジョンを形成させ、透析によって除去してリポソームを製造することもできる。
あるいは単独もしくは混合脂質の薄膜に水系溶媒を加え、激しく懸濁し、得られた溶液を超音波処理する。その後液体窒素による凍結・融解をおこなう凍結融解法が挙げられる。
あるいは、単独または混合脂質を有機溶媒に溶解させ、その溶液に水系溶媒を少量加え超音波振動を与える。そしてエタノール、エーテルなどの有機溶媒を減圧または透析などにより除去してリポソームを製造する逆相蒸発法(REV法)が挙げられる。
上記で得られたリポソームを高圧押出(エクストルージョン)法、フレンチプレス法などにより粒子径を調節する。
本発明の好ましい態様によれば、本発明の分子集合体は、DDSを用いることにより、ホウ素クラスター修飾PEG脂質誘導体に含まれる10B核を効率よく腫瘍組織に取り込ませることができる。本発明の好ましい態様によれば、本発明の分子集合体はホウ素中性子捕捉療法に好適に用いることができる。
本発明の分子集合体は、例えば、経口、非経口、静脈、経皮、吸入経由さらには疾患部位に対して直接投与することができる。投与量は、有効量の範囲内であれば良く、対象疾患、投与対象、投与方法、症状などによっても異なるが、通常、体重1kg当たり、約1500~約7500mg(脂質重量)である。
SUNBRIGHT(登録商標)SERIES DSPE-020MA(NOF CORPORATION)120mgおよび10B enriched Sodium mercaptododecaborate(BSH)(Katchem spol.sr.o., Ltd.)40mg、PBS(6mL、pH7.0)を0.1M NaOH水溶液にてpH8.0に調製し、室温で、3時間反応させた。反応は、高速液体クロマトグラフ(HPLC)で進行度合いを確認しながら行った。
その後、透析(MWCO:500~1000)により粗生成物の純度を上げ、凍結乾燥を行って次式で示される目的物を得た。目的物の純度は高速液体クロマトグラフの結果(図3)から81.7%(重量基準)、収量は115.4mgであった。
透析の条件:
外液:超純水
外液交換:5回/2時間
室温
PBS調製時に用いた各試薬:
リン酸水素ナトリウム12水和物
リン酸二水素カリウム
塩化ナトリウム
塩化カリウム
上記試薬は、いずれも、SIGMA Aldrichから入手した。
図2AおよびBに示されるとおり、本化合物のExact massである3146.07に対し、3147.690の測定値を確認できた。これは分子量誤差(%)が0.08%であり、本化合物の合成に成功したといえる。また、他のピークに関して、分子量44ずつの差がみられているが、これはポリエチレングリコール(PEG)の1mer(-CH2CH2O-)に相当し、本化合物の合成が成功したことを裏付けている。
また、図3に示されるとおり、ピーク面積81.71%の測定値を確認できた。ゆえに本化合物は上記合成スキームにより高純度で得られたといえる。
実施例2で調製した各リポソームについて、ゲルサイズ排除クロマトグラフィーにより、ホウ素クラスター修飾PEG脂質誘導体のリポソームへの組込率を調べた。
まず、実施例2で得られたリポソーム混合溶液を、ゲルサイズ排除クロマトグラフィーにより、リポソーム画分、ミセル画分、単分子画分に分離した。各分画の一例を図4に示す。
次に、リポソーム分画に含まれるホウ素原子の濃度(ppm)を誘導結合プラズマ発光分光分析装置(ICP-AES)(株式会社島津製作所製、「ICPS-8100」)により測定し、その測定値から、ホウ素クラスター修飾PEG脂質誘導体のリポソームへの組込率を算出した。
その結果、実施例2で得られた各リポソームにおいて、ホウ素クラスター修飾PEG脂質誘導体は効率よく組み込まれていることが分かった。実施例2で得られた各リポソームに対するホウ素クラスター修飾PEG脂質誘導体の組込率を表2および図5に示す。
動的光散乱法(Zetasizer Nano ZS、Malvern Instruments Ltd.社製)により、L-α-ホスファチジルコリンジアステロイル(DSPC)、コレステロール(Chol)、L-α-ジステアロイル-ホスファチジルエタノールアミン(DSPE)-PEG2000(DSPE-PEG2000)および実施例1で製造したホウ素クラスター修飾PEG脂質誘導体の2種以上で構成される単分散の各リポソームの粒子径およびゼータ電位を測定し、比較した。各リポソームは、実施例2と同様に、脂質薄膜法、続いて押出法により調製した。結果を表3に示す。
表3に示すとおり、ホウ素クラスター修飾PEG脂質誘導体を含む本発明のリポソーム(PEG-Boron-lipid 5%リポソームあるいはPEG-Boron-lipid 10%リポソーム)は、PEG修飾リポソーム(PEGリポソーム)と同様、マイナスの値のゼータ電位を示した。また、本発明のリポソームの粒子径は、およそ100nmから200nm以下であり、最もEPR効果(enhanced permeability and retention effect)が得られる粒子径にコントロールされている。
図6は、PEG-Boron-lipid 5%リポソームの透過型電子顕微鏡写真(加速電圧:100kV)である(図中、矢印で示す。)。図6に示されるとおり、PEG-Boron-lipid修飾リポソームは二重ラメラを形成し、単層構造である。
SUNBRIGHT(登録商標)SERIES DSPE-020MA(NOF CORPORATION)120mgおよび10B enriched Sodium mercaptododecaborate(BSH)(Katchem spol.sr.o., Ltd.)40mg、PBS(6mL、pH7.0)を0.1M NaOH水溶液にてpH8.0に調製し、遮光、室温で、3時間反応させた。反応は、高速液体クロマトグラフ(HPLC)で進行度合いを確認しながら行った。
その後、透析(MWCO:500~1000)により粗生成物の純度を上げ、凍結乾燥を行って次式で示される目的物を得た。目的物の純度は、89.05%(重量基準)、収量は135.0mgであった。
透析の条件:
外液:超純水
外液交換:5回/2時間
室温、遮光
PBS調製時に用いた各試薬:
リン酸水素ナトリウム12水和物
リン酸二水素カリウム
塩化ナトリウム
塩化カリウム
上記試薬は、いずれも、SIGMA Aldrichから入手した。
図7に示されるとおり、PEG-Boron-lipidはアルキル鎖末端のメチル基(2CH2CH3)のピーク δ=0.89[t、6H]を観測し、合成物にアルキル鎖が2本あることを確認した。
また、図8に示されるとおり、PEG-Boron-lipidはホウ素ピークδ=-11からδ=-16を観測し、合成物はホウ素化合物であることを確認した。
また、図9A及びBに示されるとおり、本化合物のExact massである3146.07に対し、3145.520の測定値を確認できた。これは分子量誤差(%)が0.017%であり、本化合物の合成に成功したといえる。また、他のピークに関して、分子量44ずつの差がみられているが、これはポリエチレングリコール(PEG)の1mer(-CH2CH2O-)に相当し、本化合物の合成が成功したことを裏付けている。
また、図10に示されるとおり、ピーク面積89.05%の測定値を確認できた。ゆえに本化合物は上記合成スキームにより高純度で得られたといえる。
以下に示す方法で、中性子線照射による正常細胞への影響 (in vitro)を検討した。試験に用いた細胞腫はV79 379A(classification;normal)である。
<方法>
脂質組成比(DSPC:Chol:PEG-Boron-lipid=1:1:0.12)、脂質濃度50mg/mL、超音波処理法で調製したリポソーム懸濁液を超遠心分離(100,000×g、2時間、4℃)し、得られたPEG-Boron-lipid 5%リポソームを用いた。中性子線照射はKURで行った。2.65×1012thermal neutron/cm2を各時間(15分、30分、5分)照射した。照射後、ギムザ染色液を用いて、コロニーフォメーションアッセイを行った。照射時に用いたサンプルは次の2つである。
(1)wash
V79 379A細胞(4×105cells)を25Tflaskに播種し、37℃、5%CO2条件下6時間培養した。6時間後、培地を交換し、新鮮培養液4mLにPEG-Boron-lipid 5%リポソーム溶液を1mL添加し、37℃、5%CO2条件下2時間培養した。2時間後、培地を除去し、PBSで洗浄後、トリプシンを加えて細胞をはがし、細胞を遠心分離で沈殿させ、上清を除き、新鮮培養液2mLを加えて細胞懸濁液とした。細胞懸濁液2mLから500μLをクライオチューブに分取し、1サンプルとした。
(2)non wash
V79 379A 細胞(4×105cells)を25Tflaskに播種し、37℃、5%CO2条件下6時間培養した。6時間後、培地を除去し、PBSで洗浄後トリプシンを加えて細胞をはがし、細胞数をカウントした。細胞を遠心分離で沈殿させ、上清を除き、新鮮培養液1mLを加えて細胞懸濁液とした。クライオチューブにV79 379A細胞 250μl(1×105cells)を分取し、PEG-Boron-lipid 5%リポソーム溶液250μLを添加し、2時間培養した細胞懸濁液を1サンプルとした。
<結果>
図11に示されるとおり、PEG-Boron-lipid 5%リポソームをV79 379A細胞に添加後、1washの操作を行った群(PBL(wash))は、中性子線照射のみを行った群(irradiation only)と比較して、中性子線照射のみを行った群と同等の殺細胞効果しか観察されず、PEG-Boron-lipid 5%修飾リポソーム添加による有意差がないことを確認した。検定にはスチューデントのt検定を用いた。また、2non washの操作を行った群(PBL(non wash))においては、中性子線照射により殺細胞効果が観察され、PEG-Boron-lipid 5%修飾リポソームが10B-enrichのホウ素原子団を含有していることを確認した。
V79 379Aは、Classification normalであり、正常細胞との毒性実験として行った。その結果、PEG-Boron-lipidは中性子照射によってのみ殺細胞効果を得ると考えられる。
以下に示す方法で中性子線照射による抗腫瘍効果を測定した(in vivo)。
<方法>
実施例6と同様にして作製したPEG-Boron-lipid 5%リポソームを担がんマウスに投与し、そのBNCT治療効果を検討した。担がんモデルは、BALB/cA mice(female、4weeks old,weighting 16-20g)にマウス大腸がん細胞(CT26、5×106cells)を右大腿部に播種し、腫瘍直径6-8mmとなるように作製した(injection後、約8日)。実験の際には、マウスはイソフルレンの吸入麻酔器を用いて、苦痛なく処置を行った。一連の動物実験にあたっては、あらかじめ、動物実験計画書を筑波大学動物実験委員会に提出し、動物実験に関する法規、基準に準拠し、動物に対する苦痛の軽減等についても考慮した計画とし、認可を得た。また、同様の手続きを京都大学共同利用申請において行った。
作製した担がんマウスにPEG-Boron-lipid 5%リポソームを10mg10B/kgで尾静注し、24時間後に中性子線照射をKURで行った。中性子線量は、4.5-7.0×1012neutron/cm2とした。
腫瘍増殖抑制効果を照射後の腫瘍径を経時的に21日目まで測定し、コントロール群と比較した。コントロールには、BSH溶液群(BSH、6例)、中性子線照射のみ群(irradiation only、6例)、投薬・照射なし群(no treatment、12例)を用いて、比較検討した。腫瘍サイズの測定は下記の計算式を用いた。
(長径(mm))×(短径(mm))2 = 腫瘍サイズ(mm3)
<結果>
図12に示されるとおり、PEG-Boron-lipid 修飾リポソームは10、14、17、21日目において、いずれの群とも比較して有意に腫瘍増殖抑制が見られ、特に14日までは腫瘍が増大することはなかった。また、PEG-Boron-lipid 5%リポソーム投与群6例のうち1例に腫瘍の完全消失を確認した。
腫瘍増殖抑制効果は図12からはBSHとの比較によってのみの結果である。しかし、臨床例として用いられているBSHよりも良好な結果を得たことから、その効果は高いといえる。
Claims (8)
- R1およびR2が、それぞれ独立して、炭素数12~22の直鎖状または分岐状のアルキル基である、請求項1記載のPEG脂質誘導体。
- mが2であり、かつ、nが1である、請求項1または2記載のPEG脂質誘導体。
- 請求項1から3の何れか1項記載のPEG脂質誘導体を含む分子集合体。
- PEG脂質誘導体を分子集合体の全構成脂質に対してモル比で2~20%含む、請求項4記載の分子集合体。
- L-α-ホスファチジルコリンジアステロイルおよびコレステロールをさらに含む、請求項4または5記載の分子集合体。
- リポソームである、請求項4から6の何れか1項記載の分子集合体。
- 動的光散乱法により測定したリポソームの粒子径が50~400nmである、請求項7記載の分子集合体。
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EP13831030.5A EP2889302B1 (en) | 2012-08-23 | 2013-08-22 | Boron cluster-modified peg lipid derivative, and molecular assembly using same |
JP2014531673A JP6206925B2 (ja) | 2012-08-23 | 2013-08-22 | ホウ素クラスター修飾peg脂質誘導体およびこれを用いた分子集合体 |
US14/423,049 US9468688B2 (en) | 2012-08-23 | 2013-08-22 | Boron cluster-modified PEG lipid derivative, and molecular assembly using same |
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WO2020040296A1 (ja) * | 2018-08-23 | 2020-02-27 | 森田薬品工業株式会社 | Peg化ホウ素クラスター化合物、およびpeg化ホウ素クラスター化合物を含む抗腫瘍剤、およびpeg化ホウ素クラスターを含む増感剤 |
CN111281975A (zh) * | 2020-03-13 | 2020-06-16 | 四川瑶天纳米科技有限责任公司 | 一种含硼纳米靶向药物的制备方法 |
Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2005343858A (ja) * | 2004-06-07 | 2005-12-15 | Mebiopharm Co Ltd | ホウ素イオンクラスター脂質及びこれを用いたリポソーム |
JP2008013498A (ja) * | 2006-07-06 | 2008-01-24 | Stella Chemifa Corp | ホウ素含有化合物およびこれを用いたリポソーム |
JP2008094730A (ja) * | 2006-10-06 | 2008-04-24 | Hiroyuki Nakamura | ホウ素含有化合物およびこれを用いたリポソーム |
-
2013
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Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2005343858A (ja) * | 2004-06-07 | 2005-12-15 | Mebiopharm Co Ltd | ホウ素イオンクラスター脂質及びこれを用いたリポソーム |
JP2008013498A (ja) * | 2006-07-06 | 2008-01-24 | Stella Chemifa Corp | ホウ素含有化合物およびこれを用いたリポソーム |
JP2008094730A (ja) * | 2006-10-06 | 2008-04-24 | Hiroyuki Nakamura | ホウ素含有化合物およびこれを用いたリポソーム |
Non-Patent Citations (6)
Title |
---|
HIROYUKI NAKAMURA: "Minimally Invasive Cytoselective Radiation Therapy Using Boron Neutron Capture Reaction", YAKUGAKU ZASSHI, vol. 130, no. 12, 2010, pages 1687 - 1694, XP055197822 * |
LEE JD; UENO M; MIYAJIMA Y; NAKAMURA H.: "Synthesis of boron cluster lipids: closo-dodecaborate as an alternative hydrophilic function of boronated liposomesfor neutron capture therapy", ORG LETT., vol. 9, no. 2, 18 January 2007 (2007-01-18), pages 323 - 6 |
MANABU UENO: "Dodecaborate lipid liposomes as new vehicles for boron delivery system of neutron capture therapy", BIOORGANIC & MEDICINAL CHEMISTRY, vol. 18, no. 9, 2010, pages 3059 - 3065, XP027030036 * |
MARUYAMA K; ISHIDA O; KASAOKA S; TAKIZAWA T; UTOGUCHI N; SHINOHARA A; CHIBA M; KOBAYASHI H; ERIGUCHI M; YANAGIE H.: "Intracellular targeting of sodium mercaptoundecahydrocloctecabonate (BSH) to solid tumors by transferrin-PEG liposomes, for boron neutron-capture therapy (BNCT)", J CONTROL RELEASE., vol. 98, no. 2, 11 August 2004 (2004-08-11), pages 195 - 207, XP004521710, DOI: doi:10.1016/j.jconrel.2004.04.018 |
See also references of EP2889302A4 |
YAMAMOTO T; NAKAI K; KAGEJI T; KUMADA H; ENDO K; MATSUDA M; SHIBATA Y; MATSUMURA A.: "Boron neutron capture therapy for newly diagnosed glioblastoma", RADIOTHER ONCOL., vol. 91, no. 1, 11 March 2009 (2009-03-11), pages 80 - 4, XP026026796, DOI: doi:10.1016/j.radonc.2009.02.009 |
Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
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WO2020040296A1 (ja) * | 2018-08-23 | 2020-02-27 | 森田薬品工業株式会社 | Peg化ホウ素クラスター化合物、およびpeg化ホウ素クラスター化合物を含む抗腫瘍剤、およびpeg化ホウ素クラスターを含む増感剤 |
JPWO2020040296A1 (ja) * | 2018-08-23 | 2021-08-26 | 森田薬品工業株式会社 | Peg化ホウ素クラスター化合物、およびpeg化ホウ素クラスター化合物を含む抗腫瘍剤、およびpeg化ホウ素クラスターを含む増感剤 |
JP7337342B2 (ja) | 2018-08-23 | 2023-09-04 | 森田薬品工業株式会社 | Peg化ホウ素クラスター化合物、およびpeg化ホウ素クラスター化合物を含む抗腫瘍剤、およびpeg化ホウ素クラスターを含む増感剤 |
CN111281975A (zh) * | 2020-03-13 | 2020-06-16 | 四川瑶天纳米科技有限责任公司 | 一种含硼纳米靶向药物的制备方法 |
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US20150238622A1 (en) | 2015-08-27 |
EP2889302A1 (en) | 2015-07-01 |
JPWO2014030715A1 (ja) | 2016-07-28 |
EP2889302A4 (en) | 2016-05-18 |
US9468688B2 (en) | 2016-10-18 |
EP2889302B1 (en) | 2017-05-17 |
JP6206925B2 (ja) | 2017-10-04 |
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