Amidite Monomers

Abstract

To provide a technique for incorporating a compound having a colchicine-like structure into a nucleic acid.SOLUTION: The invention provides a compound represented by a general formula (1) or a salt thereof, or their solvate. [In the formula: R1 are the same or different and each represent an alkoxy or phosphate group; R2 are the same or different and each represent an alkoxy or phosphate group; R3 represents -N= or -CH=; R4 represents =N- or =CH-; R5 represents a protecting group of a hydroxy group; R6 and R7 are the same or different and each represent an alkyl group; m represents an integer from 1 to 5; and n represents an integer from 1 to 4.]SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to an amidite monomer and the like. [Background technology]

[0002] Various anticancer drugs and their prodrugs with colchicine-like structures have been reported (e.g., Non-Patent Document 1). Like colchicine, these anticancer drugs specifically bind to tubulin and inhibit the formation of microtubules, thereby suppressing cell division of cancer cells. Therefore, there is a need for the development of drug delivery technologies, particularly those that can transport drugs specifically to cancer cells.

[0003] Generally, drug delivery technologies reported include liposomes, polymeric nanoparticles, inorganic nanoparticles, dendrimers, micelles, nanoemulsions, and polymer drug conjugates. However, some of these technologies lack uniformity in shape, have suboptimal biocompatibility, and often lack selectivity for diseased cells. Therefore, the development of new drug delivery technologies is desired. [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Borowiak et al., 2015, Cell, 162, 403-411. Summary of the Invention [Problem to be solved by the invention]

[0005] The present inventors have focused on nucleic acid structures such as DNA origami as a drug delivery technology. DNA origami has the inherent properties of biocompatibility, biodegradability, and the ability to control the site of drug uptake.

[0006] An objective of the present invention is to provide a technique for incorporating a compound having a colchicine-like structure into a nucleic acid. [Means for solving the problem]

[0007] The present inventors have conducted extensive research in light of the above-mentioned problems and have found that by using an amidite monomer represented by general formula (1), it is possible to obtain a nucleotide or polynucleotide to which a compound having a colchicine-like structure is attached and which has anticancer activity, as well as a nucleic acid construct containing the same. Based on this finding, the present inventors have conducted further research and have completed the present invention. That is, the present invention encompasses the following aspects.

[0008] Section 1. General formula (1):

[0009] [ka]

[0010] [In the formula: R 1 are the same or different and represent an alkoxy group or a phosphate group. 2 are the same or different and represent an alkoxy group or a phosphate group. 3 represents -N= or -CH=. 4 represents =N- or =CH-. 5 represents a protecting group for a hydroxy group. 6 and R 7 are the same or different and represent an alkyl group. m represents an integer of 1 to 5. n represents an integer of 1 to 4.] A compound represented by the formula (I), a salt thereof, or a solvate thereof.

[0011] Item 2. The compound represented by general formula (1A):

[0012] [ka]

[0013] [In the formula: R 1 , R 2 , R3 , R 4 , R 5 , R 6 , and R 7 is the same as above.] Item 2. The compound according to Item 1, or a salt thereof, or a solvate thereof, which is a compound represented by the formula:

[0014] Section 3.R 1 and R 2 Item 3. The compound or salt thereof, or a solvate thereof according to Item 1 or 2, wherein is an alkoxy group.

[0015] Section 4.R 5 is an alkyl group substituted with an electron-withdrawing group, and R 6 and R 7 Item 4. The compound or salt thereof, or a solvate of the compound or salt according to any one of Items 1 to 3, wherein is a branched alkyl group.

[0016] Item 5. The terminal phosphate group is represented by the general formula (2):

[0017] [ka]

[0018] [In the formula: R 1 are the same or different and represent an alkoxy group or a phosphate group. 2 are the same or different and represent an alkoxy group or a phosphate group. 3 represents -N= or -CH=. 4 represents =N- or =CH-. 8 represents O or S. R 9 O - or S - m represents an integer of 1 to 5. n represents an integer of 1 to 4.] A nucleotide or polynucleotide in which the nucleotide or polynucleotide is replaced with a group represented by the following formula:

[0019] Section 6. General formula (2A):

[0020] [ka]

[0021] [In the formula: R 1 , R 2 , R 3 , R 4 , R 8 , R 9 , m, and n are the same as above. 8A are the same or different and represent O or S. R 9A are the same or different, O - or S - Indicates R 10 R may be the same or different and represent a divalent group formed by removing two hydroxy groups from the sugar moiety of a nucleoside. 10A R may be the same or different and represent a divalent group formed by removing two hydroxy groups from the sugar moiety of a nucleoside. 11 represents a hydroxy group, a phosphate group, or a thiophosphate group, and p represents an integer of 0 or 1 or more. Item 6. The nucleotide or polynucleotide according to Item 5, which is a compound represented by the formula:

[0022] Item 7. A nucleic acid construct comprising the nucleotide or polynucleotide according to Item 5 or 6.

[0023] Item 8. The nucleic acid structure according to Item 7, which is a DNA origami.

[0024] Item 9. A pharmaceutical comprising at least one selected from the group consisting of the nucleotide or polynucleotide according to Item 5 or 6 and the nucleic acid construct according to Item 7 or 8.

[0025] Item 10. The pharmaceutical agent according to Item 9, which is an anticancer agent.

[0026] Item 11. A reagent comprising at least one selected from the group consisting of the nucleotide or polynucleotide according to Item 5 or 6 and the nucleic acid structure according to Item 7 or 8. [Effects of the Invention]

[0027] According to the present invention, it is possible to provide an amidite monomer of a compound having a colchicine-like structure, a nucleotide or polynucleotide obtained using the amidite monomer, and various acid structures containing the nucleotide or polynucleotide. [Brief explanation of the drawings]

[0028] [Figure 1] The 1H NMR [CDCl3] spectrum and synthesis scheme of compound 1 are shown below. [Figure 2] The 1H NMR [CDCl3] spectrum and synthesis scheme of compound 2 are shown below. [Figure 3] The 1H NMR [CDCl3] spectrum and synthesis scheme of compound 3 are shown below. [Figure 4] The 1H NMR [CDCl3] spectrum and synthesis scheme of Photostatin are shown below. [Figure 5] 1 shows a mass spectrum of a PST-amidite monomer. [Figure 6] Mass spectra of PST-A, PST-G, PST-C, PST-T, and PST-T9 are shown. [Figure 7] The results of HPLC purification of PST-T and the mass spectrum of the collected peak are shown. [Figure 8] The results of HPLC purification of PST-T9 are shown. [Figure 9] The mass spectrum of the collected peak of PST-T9 is shown. [Figure 10] 1 shows the results of HPLC measurement evaluation of cis-trans isomerization in Example 3. [Figure 11] 1 shows the results of UV measurement evaluation of cis-trans isomerization in Example 4. [Figure 12] 1 shows the results of UV measurement (time course) evaluation of cis-trans isomerization in Example 5. [Figure 13] 1 shows the results of Tm measurement in Example 6. [Figure 14] 1 shows the results of thermal stability evaluation of Example 7. [Figure 15] 1 shows the results of the anticancer activity test in Example 8. [Figure 16] 1 shows an overall view of the DNA origami dendrimer prepared in Example 10. [Figure 17] 1 shows an enlarged view of a portion of the tip of the DNA origami dendrimer prepared in Example 10. [Figure 18] An enlarged view (left) of the tip of a portion of the DNA origami dendrimer prepared in Example 10 and an enlarged view (right) of the tip of a portion of the PST-modified DNA origami dendrimer are shown. DETAILED DESCRIPTION OF THE INVENTION

[0029] In this specification, the expressions "contain" and "comprise" include the concepts of "contain," "comprise," "consist essentially of," and "consist only of."

[0030] 1. Amidite Monomers In one aspect, the present invention provides a compound represented by general formula (1):

[0031] [ka]

[0032] or a salt thereof, or a solvate thereof (these may be collectively referred to as the "amidite monomer of the present invention" in this specification). This will be explained below.

[0033] R 1 are the same or different and represent an alkoxy group or a phosphate group. 1 is preferably an alkoxy group.

[0034] R 1The alkoxy group represented by the formula (I) includes both straight-chain and branched-chain (preferably straight-chain) ones. The number of carbon atoms in the alkoxy group is not particularly limited, but is, for example, 1 to 8, preferably 1 to 6, more preferably 1 to 4, even more preferably 1 to 2, and particularly preferably 1. Specific examples of the alkoxy group include a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an isobutoxy group, a sec-butoxy group, and a tert-butoxy group.

[0035] R 1 The phosphate group represented by is a group represented by H2PO4-.

[0036] m represents an integer of 1 to 5. m preferably represents an integer of 1 to 4, more preferably an integer of 2 to 4, still more preferably an integer of 2 to 3 or an integer of 3 to 4, and particularly preferably 3.

[0037] R 1 The position of R 3 For example, it can be in the para position, meta position, or ortho position relative to R 1 The position of R is preferably the para position or the meta position. 1 When there are multiple Rs, Rs at the para and / or meta positions 1 and preferably includes R 1 It is more preferred that the composition contains:

[0038] R 2 are the same or different and represent an alkoxy group or a phosphate group. 2 is preferably an alkoxy group.

[0039] R 2 The definition of the alkoxy group represented by R 1 The definition of the alkoxy group is the same as that of the alkoxy group represented by R 2 The definition of the phosphate group is R 1 This is the same as the definition of the phosphate group shown below.

[0040] n represents an integer of 1 to 4. n is preferably an integer of 1 to 3, more preferably an integer of 1 or 2, and even more preferably 1.

[0041] R 2 The position of R 4 For example, it can be in the para position, meta position, or ortho position relative to R 1 The position of is preferably the para position or the meta position, more preferably the para position.

[0042] In one preferred embodiment of the present invention, R 1 and R 2 can be an alkoxy group.

[0043] R 1 and R 2 Regarding the number (m, n) and position of, in one embodiment of the present invention, the compound represented by general formula (1) is particularly preferably represented by general formula (1A):

[0044] [ka]

[0045] The compound may be represented by the formula:

[0046] R 3 represents -N= or -CH=. 3 is preferably -N=.

[0047] R 4 represents =N- or =CH-. 4 is preferably =N-.

[0048] In one preferred embodiment of the present invention, R 3 is -N= and R 4 is =N-, or R 3 is -CH= and R 4 can be =CH-.

[0049] R 5indicates a protecting group for a hydroxy group.

[0050] R 5 As the protecting group for the hydroxy group represented by R, a wide range of known protecting groups used in amidite monomers can be used. 5 Examples of the alkyl group include an alkyl group, an alkenyl group, an alkynyl group, a cycloalkyl group, a haloalkyl group, an aryl group, a heteroaryl group, an arylalkyl group, a cycloalkenyl group, a cycloalkylalkyl group, a cyclylalkyl group, a hydroxyalkyl group, an aminoalkyl group, an alkoxyalkyl group, a heterocyclylalkenyl group, a heterocyclylalkyl group, a heteroarylalkyl group, a silyl group, a silyloxyalkyl group, a mono-, di-, or trialkylsilyl group, and a mono-, di-, or trialkylsilyloxyalkyl group, which may be substituted with an electron-withdrawing group.

[0051] R 5 R is preferably an alkyl group substituted with an electron-withdrawing group. Examples of the electron-withdrawing group include a cyano group, a nitro group, an alkylsulfonyl group, a halogen atom, an arylsulfonyl group, a trihalomethyl group, and a trialkylamino group, and is preferably a cyano group. 5 is particularly preferably —(CH 2 ) 2 —CN.

[0052] R 6 and R 7 are the same or different and represent alkyl groups.

[0053] R 6 and R 7 The alkyl group represented by may be either linear or branched, and is preferably an alkyl group having 1 to 12 carbon atoms, more preferably an alkyl group having 1 to 6 carbon atoms. Examples of the alkyl group include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, and hexyl. The alkyl group here also includes alkyl moieties such as alkoxy groups. R 6 and R 7may be bonded to each other to form a cyclic structure.

[0054] R 6 and R 7 and are particularly preferably both isopropyl groups.

[0055] The compound represented by general formula (1) is R 3 =R 4 That is, the compound represented by general formula (1) includes both a cis-isomer and a trans-isomer with respect to the double bond of general formula (11):

[0056] [ka]

[0057] or a compound represented by general formula (12):

[0058] [ka]

[0059] The compound may be represented by the formula:

[0060] The cis- and trans-isomers can be converted to the other isomer by irradiation with light. The conversion can be carried out according to known methods (for example, according to the method described in Non-Patent Document 1).

[0061] The salt of the compound represented by general formula (1) is not particularly limited and includes, for example, salts with inorganic bases such as sodium salt, magnesium salt, potassium salt, calcium salt, and aluminum salt; salts with organic bases such as methylamine, ethylamine, and ethanolamine; salts with basic amino acids such as lysine, ornithine, and arginine; and ammonium salt. The salt may be an acid addition salt, and specific examples of such salts include acid addition salts with mineral acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, and phosphoric acid; organic acids such as formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, malic acid, tartaric acid, fumaric acid, succinic acid, lactic acid, maleic acid, citric acid, methanesulfonic acid, trifluoromethanesulfonic acid, and ethanesulfonic acid; and acidic amino acids such as aspartic acid and glutamic acid.

[0062] The solvates of the compound represented by general formula (1) or a salt thereof are not particularly limited, and examples thereof include solvates with solvents such as water, ethanol, glycerol, and acetic acid.

[0063] The amidite monomers of the present invention can be prepared in a variety of ways.

[0064] The compound represented by general formula (1) can be produced, for example, according to or in accordance with the following scheme.

[0065] [ka]

[0066] X is a halogen atom. Examples of the halogen atom represented by X include a fluorine atom, a chlorine atom, a bromine atom, and an iodine atom. Among these, a halogen atom is preferred.

[0067] As the compound represented by the general formula (101) and the compound represented by the general formula (102), commercially available compounds can be used as they are, or compounds synthesized according to or in accordance with known methods can be used as needed.

[0068] From the viewpoints of yield, ease of synthesis, etc., the amount of the compound represented by general formula (102) used is usually preferably 1 to 3 moles, more preferably 1.5 to 2.5 moles, per mole of the compound represented by general formula (101).

[0069] This reaction is usually carried out in the presence of a reaction solvent. The reaction solvent is not particularly limited, but examples thereof include acetonitrile, pyridine, dimethylformamide, dichloromethane, tetrahydrofuran, acetone, toluene, ethanol, etc., and preferably acetonitrile, etc. The solvent may be used alone or in combination.

[0070] This reaction is preferably carried out in the presence of a base (preferably a non-nucleophilic base) such as N,N-diisopropylethylamine. From the viewpoints of yield, ease of synthesis, etc., the amount of the base used is usually preferably 3 to 15 moles, more preferably 4 to 10 moles, per mole of the compound represented by general formula (101).

[0071] In addition to the above components, additives may also be used in this reaction as appropriate, provided that they do not significantly impair the progress of the reaction.

[0072] The reaction can be carried out under heating, at room temperature, or under cooling, and is usually preferably carried out at 10 to 100° C. The reaction time is not particularly limited, and can usually be 30 minutes to 30 hours.

[0073] The progress of the reaction can be monitored by conventional methods such as chromatography. After the reaction is complete, the solvent is removed by distillation, and the product can be isolated and purified by conventional methods such as chromatography and recrystallization. The structure of the product can also be determined by elemental analysis, MS (ESI-MS), IR analysis, and the like. 1 H-NMR, 13 It can be identified by C-NMR or the like.

[0074] 2. Nucleotides or polynucleotides In one aspect, the present invention provides a polymer having a terminal phosphate group represented by general formula (2):

[0075] [ka]

[0076] This is described below.

[0077] R 1 , R 2 , R 3 , R 4 The definitions of , m, and n are as explained above in "1. Amidite Monomer."

[0078] R 8 represents O or S. R 8 is preferably O.

[0079] R 9 O - or S - Indicates R 9 is preferably O - is.

[0080] The (poly)nucleotide of the present invention has a structure in which the terminal phosphate group of a nucleotide or polynucleotide is replaced with a group represented by general formula (2).

[0081] The terminal phosphate group may be either a 5'-terminal phosphate group or a 3'-terminal phosphate group, but is preferably a 5'-terminal phosphate group.

[0082] The nucleotides and polynucleotides are not particularly limited, and various building blocks of artificial nucleic acids, including natural nucleic acids, aTNA, and SNA, can be used. Specific examples of nucleic acids that can be used include DNA, RNA, and those that have been chemically modified as shown below. To prevent degradation by hydrolases such as nucleases, the phosphate residue of each nucleotide can be substituted with a chemically modified phosphate residue such as phosphorothioate (PS), methylphosphonate, or phosphorodithioate. Furthermore, the hydroxy group at the 2'-position of the sugar (ribose) of each ribonucleotide can be substituted with -OR (where R represents, for example, CH3(2'-O-Me), CH2CHOCH3(2'-O-MOE), CH2CH2NHC(NH)NH2, CH2CONHCH3, or CH2CH2CN). Furthermore, the base moiety (pyrimidine, purine) may be chemically modified, for example, by introducing a methyl group or a cationic functional group into the 5-position of the pyrimidine base, or by substituting a thiocarbonyl group for the carbonyl group at the 2-position. Further examples include, but are not limited to, those in which the phosphate moiety or hydroxyl moiety is modified with, for example, biotin, an amino group, a lower alkylamine group, or an acetyl group. Furthermore, BNA (LNA), in which the conformation of the sugar moiety of the nucleotide is fixed to the N-type by bridging the 2' oxygen and 4' carbon of the sugar moiety, may also be used.

[0083] The (poly)nucleotide of the present invention more particularly preferably has the general formula (2A):

[0084] [ka]

[0085] It is a compound represented by the formula:

[0086] R 8A are the same or different and represent O or S. R 8 A is preferably O.

[0087] R 9Aare the same or different, O - or S - Indicates R 9 A is preferably O - is.

[0088] R 10 are the same or different divalent groups formed by removing two hydroxy groups from the sugar moiety of a nucleoside (a nucleoside consisting of a sugar moiety such as ribose or deoxyribose and a nucleic acid base). Preferably, one hydroxy group is the 5' hydroxy group of ribose or deoxyribose or a corresponding hydroxy group in another nucleic acid backbone, and the other hydroxy group is the 3' hydroxy group of ribose or deoxyribose or a corresponding hydroxy group in another sugar moiety.

[0089] R 10A are the same or different divalent groups formed by removing two hydroxy groups from the sugar moiety of a nucleoside (a nucleoside consisting of a sugar moiety such as ribose or deoxyribose and a nucleic acid base). Preferably, one hydroxy group is the 5' hydroxy group of ribose or deoxyribose or a corresponding hydroxy group in another nucleic acid backbone, and the other hydroxy group is the 3' hydroxy group of ribose or deoxyribose or a corresponding hydroxy group in another sugar moiety.

[0090] The sugar moiety of a nucleoside is not particularly limited as long as it can constitute a nucleic acid, and examples include sugar moieties such as ribose, deoxyribose, and modified sugars (e.g., sugar moieties that constitute DNA, RNA, BNA, LNA, etc.).

[0091] As the nucleic acid base of nucleoside, any base constituting nucleic acid can be used without any particular limitation.The base constituting nucleic acid includes not only typical bases in natural nucleic acids such as RNA and DNA (adenine (A), thymine (T), uracil (U), guanine (G), cytosine (C), etc.), but also other bases such as hypoxanthine (I), modified bases, etc. Examples of modified bases include pseudouracil, 3-methyluracil, dihydrouracil, 5-alkylcytosine (e.g., 5-methylcytosine), 5-alkyluracil (e.g., 5-ethyluracil), 5-halouracil (5-bromouracil), 6-azapyrimidine, 6-alkylpyrimidine (6-methyluracil), 4-acetylcytosine, 5-(carboxyhydroxymethyl)uracil, 5'-carboxymethylaminomethyl-2-thiouracil, 5-carboxymethylaminomethyluracil, 1-methyladenine, 1-methylhypo Examples include xanthine, 2,2-dimethylguanine, 3-methylcytosine, 2-methyladenine, 2-methylguanine, N6-methyladenine, 7-methylguanine, 5-methoxyaminomethyl-2-thiouracil, 5-methylaminomethyluracil, 5-methylcarbonylmethyluracil, 5-methyloxyuracil, 5-methyl-2-thiouracil, 2-methylthio-N6-isopentenyladenine, uracil-5-oxyacetic acid, 2-thiocytosine, purine, 2-aminopurine, isoguanine, indole, imidazole, and xanthine.

[0092] R 11 represents a hydroxy group, a phosphate group, or a thiophosphate group.

[0093] p represents 0 or an integer of 1 or more. p is preferably 1 to 100,000, more preferably 5 to 100,000, even more preferably 10 to 100,000, still more preferably 15 to 100,000, and particularly preferably 20 to 100,000. The upper and / or lower limits of p can be, for example, 50, 100, 500, 1,000, 5,000, 10,000, or 50,000.

[0094] The (poly)nucleotide of the present invention can be in the form of a salt. The salt is not particularly limited, and examples include salts with inorganic bases such as sodium salts, magnesium salts, potassium salts, calcium salts, and aluminum salts; salts with organic bases such as methylamine, ethylamine, and ethanolamine; salts with basic amino acids such as lysine, ornithine, and arginine; and ammonium salts. The salt may also be an acid addition salt, and specific examples of such salts include acid addition salts with mineral acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, and phosphoric acid; organic acids such as formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, malic acid, tartaric acid, fumaric acid, succinic acid, lactic acid, maleic acid, citric acid, methanesulfonic acid, trifluoromethanesulfonic acid, and ethanesulfonic acid; and acidic amino acids such as aspartic acid and glutamic acid.

[0095] The (poly)nucleotide of the present invention may be in the form of a solvate. The solvate is not particularly limited, and examples thereof include solvates with solvents such as water, ethanol, glycerol, and acetic acid.

[0096] The (poly)nucleotide of the present invention can be produced by a method for producing a single-stranded polynucleotide by the phosphoramidite method, using the amidite monomer of the present invention.

[0097] The phosphoramidite method can be carried out according to known methods using, for example, a commercially available automated nucleic acid synthesizer. Specifically, the method includes the following steps: (A) deprotecting the hydroxyl group at the 5'-position (or a position corresponding thereto); (B) condensing the amidite monomer; (C) capping the hydroxyl group at the 5'-position (or a position corresponding thereto) of the unreacted compound; (D) converting the phosphite group to a phosphate group or a thiophosphate group; (E) cleaving the resulting compound from the solid support and deprotecting the phosphate moiety and nucleic acid base; and (F) deprotecting the hydroxyl group at the 5'-position (or a position corresponding thereto). Repeating steps (A) to (D) allows the production of a compound containing a polynucleotide backbone of a desired chain length. The (poly)nucleotide of the present invention can be produced by using the amidite monomer of the present invention in step (B).

[0098] In step (D), it is preferable to use a solution containing iodine / water as the oxidizing agent.

[0099] In the deprotection step (E), deprotection is carried out with a base such as aqueous ammonia. Although the bond between the phosphorus atom and the aromatic hydroxy group in the (poly)nucleotide of the present invention was expected to be unstable to bases, unexpectedly, cleavage of the above bond was suppressed during deprotection in the (poly)nucleotide of the present invention.

[0100] After step (E), it is preferable to perform chromatographic purification (e.g., reverse-phase chromatographic purification) before step (F), taking advantage of the hydrophobicity of the 5'-protecting group (hydrophobic group), which can further increase the purity of the target single-stranded polynucleotide.

[0101] The resulting single-stranded polynucleotide can be isolated and purified as necessary. It can usually be isolated by methods for precipitating, extracting, and purifying RNA. Specifically, methods include adding a solvent with low solubility for RNA, such as ethanol or isopropyl alcohol, to the reaction solution to precipitate the RNA, or adding a phenol / chloroform / isoamyl alcohol solution to the reaction solution to extract the RNA into the aqueous layer. The polynucleotide can then be isolated and purified by known high-performance liquid chromatography (HPLC) techniques, such as reverse-phase column chromatography, anion-exchange column chromatography, or affinity column chromatography.

[0102] 3.Nucleic acid construct In one aspect, the present invention relates to a nucleic acid construct (sometimes referred to herein as the "nucleic acid construct of the present invention") comprising the (poly)nucleotide of the present invention. This is described below.

[0103] The nucleic acid construct is not particularly limited as long as it is a construct mainly containing nucleic acids. The content of nucleic acids in the nucleic acid construct is, for example, 70% by mass or more, preferably 80% by mass or more, more preferably 90% by mass or more, even more preferably 95% by mass or more, even more preferably 98% by mass or more, particularly preferably 99% by mass or more, and particularly preferably 100% by mass. Substances other than nucleic acids in the nucleic acid construct are not particularly limited, and examples thereof include proteins, peptides, sugars, labeling substances, metal ions, etc.

[0104] The nucleic acid structure can be a three-dimensional structure, a two-dimensional structure (planar structure), or a structure that is a combination of these.

[0105] The nucleic acid construct is preferably a structure formed by folding nucleic acid. From the viewpoint of suitability for drug delivery, the nucleic acid construct is preferably a structure (DNA origami) formed from a material containing a single-stranded circular nucleic acid and a staple nucleic acid containing a sequence complementary to the single-stranded circular nucleic acid.

[0106] The single-stranded circular nucleic acid that can constitute the nucleic acid construct is not particularly limited as long as it can be folded to form a DNA origami. Regardless of the base sequence of the single-stranded circular nucleic acid, DNA origami can be formed by designing the staple DNA sequence accordingly. The number of bases in the single-stranded circular nucleic acid is not particularly limited, but is, for example, 500 to 50,000, preferably 1,000 to 25,000, more preferably 2,000 to 15,000, even more preferably 3,000 to 12,000, and even more preferably 4,000 to 10,000. The base sequence of the single-stranded circular nucleic acid is not particularly limited, but examples include the M13 bacteriophage DNA sequence or a base sequence containing a sequence derived from the DNA sequence.

[0107] Staple nucleic acids that can constitute a nucleic acid construct are not particularly limited, as long as they contain a complementary sequence to a single-stranded circular nucleic acid and can fold the single-stranded circular nucleic acid to form a DNA origami through complementary base pairing with the single-stranded circular nucleic acid. The number of bases in the staple nucleic acid is not particularly limited, but is, for example, 10 to 500, preferably 20 to 250, and more preferably 30 to 100. In the staple nucleic acid, the number of bases in the sequence complementary to the single-stranded circular nucleic acid is not particularly limited, but is, for example, 10 to 500, preferably 20 to 250, and more preferably 30 to 100. One staple nucleic acid usually contains complementary sequences to each of the multiple strands (e.g., 2 to 5, preferably 2 to 4, and more preferably 2 to 3) that are adjacent to each other when the single-stranded circular nucleic acid is folded. Staple nucleic acids with such sequence characteristics can further stabilize the single-stranded circular nucleic acid in the folded state through complementary base pairing with the single-stranded circular nucleic acid. Various DNA origami methods have been reported, and the sequences of staple nucleic acids for forming nucleic acid constructs of desired shapes and structures can be designed according to known methods and information.

[0108] The size of the nucleic acid construct is not particularly limited and can be adjusted appropriately depending on the purpose. The nucleic acid construct is preferably nanosized (nanostructure). For example, the longest diameter (major axis) of the nucleic acid construct is 1000 nm or less. In one embodiment of the present invention, the upper and / or lower limits of the major axis of the nucleic acid construct can be, for example, 5 nm, 20 nm, 50 nm, 100 nm, 200 nm, 500 nm, or 800 nm. By making the nucleic acid construct of the present invention larger than a certain size, it can be selectively delivered to cancer tissues based on the EPR (enhanced permeability and retention) effect.

[0109] The nucleic acid construct of the present invention can be obtained by using the (poly)nucleotide of the present invention as a part of the nucleic acid constituting the nucleic acid construct. More specifically, the nucleic acid construct of the present invention can be obtained by using the (poly)nucleotide of the present invention, for example, as a nucleic acid that can hybridize to a part of a staple nucleic acid (a part that is not hybridized to a single-stranded circular nucleic acid), as a staple nucleic acid, or as part or all of a single-stranded circular nucleic acid. Alternatively, the nucleic acid construct of the present invention can be obtained by incorporating the (poly)nucleotide of the present invention into the nucleic acid construct of the present invention.

[0110] In order to effectively exert the anticancer activity derived from the (poly)nucleotide of the present invention, the nucleic acid construct of the present invention preferably has a plurality of (for example, 2 to 1000, 10 to 1000, 50 to 1000, or 100 to 500) overhanging structures, and each overhanging structure preferably contains the (poly)nucleotide of the present invention.

[0111] 4.Applications In one aspect, the present invention can be used as an active ingredient (active ingredient of the present invention) of a medicine, reagent, etc. (sometimes referred to as a "drug of the present invention" in this specification), which contains at least one selected from the group consisting of the (poly)nucleotide of the present invention and the nucleic acid construct of the present invention, more specifically, as an active ingredient of an anticancer agent, etc.

[0112] The pharmaceutical preparation of the present invention is not particularly limited as long as it contains the active ingredient of the present invention, and may further contain other ingredients as necessary. The other ingredients are not particularly limited as long as they are pharmaceutically acceptable. The other ingredients include not only ingredients with pharmacological effects but also additives. Examples of additives include bases, carriers, solvents, dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrants, lubricants, thickeners, moisturizers, colorants, fragrances, chelating agents, etc.

[0113] The mode of use of the agent of the present invention is not particularly limited, and an appropriate mode of use can be adopted depending on the type of agent. Depending on the intended use, the agent of the present invention can be used, for example, in vitro (e.g., added to the medium of cultured cells) or in vivo (e.g., administered to animals).

[0114] The agents of the present invention may be applied to, but are not limited to, mammals such as humans, monkeys, mice, rats, dogs, cats, rabbits, pigs, horses, cattle, sheep, goats, and deer. Examples of cells include animal cells. The types of cells are also not particularly limited, and examples include blood cells, hematopoietic stem cells and progenitor cells, gametes (sperm, eggs), fibroblasts, epithelial cells, vascular endothelial cells, nerve cells, hepatocytes, keratinocytes, muscle cells, epidermal cells, endocrine cells, ES cells, iPS cells, tissue stem cells, and cancer cells.

[0115] When the agent of the present invention is used as an anticancer agent or when used on cancer cells, the target cancer is not particularly limited and includes, for example, hepatocellular carcinoma, pancreatic cancer, kidney cancer, leukemia, esophageal cancer, stomach cancer, colon cancer, lung cancer, prostate cancer, skin cancer, breast cancer, cervical cancer, etc. Among these, solid cancers are preferred, and hepatocellular carcinoma is more preferred.

[0116] The agents of the present invention may take any dosage form, for example, oral preparations such as tablets (including orally disintegrating tablets, chewable tablets, effervescent tablets, troches, jelly drops, etc.), pills, granules, fine granules, powders, hard capsules, soft capsules, dry syrups, liquids (including drinks, suspensions, syrups), and jellies; and parenteral preparations such as injectable preparations (for example, drip injections (e.g., intravenous drip preparations), intravenous injections, intramuscular injections, subcutaneous injections, and intradermal injections), topical preparations (for example, ointments, poultices, and lotions), suppositories, inhalants, eye preparations, eye ointments, nasal drops, ear drops, and liposomes.

[0117] The route of administration of the drug of the present invention is not particularly limited as long as the desired effect can be obtained, and examples include enteral administration such as oral administration, tube feeding, and enema administration; and parenteral administration such as intravenous administration, intraarterial administration, intramuscular administration, intracardiac administration, subcutaneous administration, intradermal administration, and intraperitoneal administration.

[0118] The content of the active ingredient in the drug of the present invention depends on the mode of use, the subject to which it is applied, the condition of the subject to which it is applied, etc., and is not limited thereto, but can be, for example, 0.0001 to 100% by weight, preferably 0.001 to 50% by weight.

[0119] The dosage of the agent of the present invention when administered to an animal is not particularly limited as long as it is an effective amount that exerts a pharmacological effect, and is usually 0.1 to 1000 mg / kg body weight, preferably 0.5 to 500 mg / kg body weight per day in terms of the weight of the active ingredient, in the case of oral administration, and 0.01 to 100 mg / kg body weight, preferably 0.05 to 50 mg / kg body weight per day in the case of parenteral administration. The dosage can be increased or decreased as appropriate depending on the age, pathological condition, symptoms, etc. [Example]

[0120] The present invention will be described in detail below based on examples, but the present invention is not limited to these examples.

[0121] Example 1. Synthesis of PST-amidite monomers The PST-amidite monomer was synthesized according to the following scheme.

[0122] [ka]

[0123] Example 1-1. Synthesis of Compound 1 Cathechol (5.2 g, 47.2 mmol) and K2CO3 (6.3 g, 45.6 mmol) were weighed into a two-necked flask and dissolved in acetone (85 mL). A reflux apparatus was then attached and refluxing was initiated. Allyl bromide (4.0 mL, 46.3 mmol) was then slowly added dropwise via syringe, and the mixture was refluxed at 75°C for 4 hours. The color of the solution changed from white to reddish purple. The progress of the reaction was confirmed by TLC (hexane: ethyl acetate = 2:1). After filtration, the solvent was completely removed using an evaporator, and the product was purified by flash column chromatography using hexane: ethyl acetate (0 → 20%) as the mobile phase. A white solid (4.70 g) was obtained as the target product in a yield of 66.6%. Compound 1 1 The H NMR [CDCl3] spectrum and synthesis scheme are shown in Figure 1.

[0124] Example 1-2: Synthesis of Compound 2 3,4,5-Trimethoxyaniline (0.68 g, 3.7 mmol) was dissolved in ethanol (20 mL) and cooled on ice. Then, 3 M HCl (3.0 mL, 9.0 mmol) was slowly added via syringe, followed by the dropwise addition of 2.5 M NaNO2 (1.4 mL, 3.6 mmol). The solution changed color from reddish purple to black. After stirring for 45 min, compound 1 (0.60 g, 4.0 mmol) dissolved in 2 M NaOH (3.6 mL, 7.2 mmol) was added dropwise. The mixture was further stirred at room temperature for 2 h. The reaction was quenched by the addition of 3 M HCl (3.0 mL, 9.0 mmol). The progress of the reaction was monitored by TLC (hexane:ethyl acetate = 2:1). The mixture was filtered and the solvent was removed by evaporation. The residue was then dissolved in ethyl acetate, extracted twice with saturated aqueous NaHCO3, and washed once with saturated aqueous NaCl, and the organic layer was collected. Na2SO4 was added and the residue was dried by stirring for 30 minutes. The residue was then filtered and the solvent was removed using an evaporator. The residue was purified by flash column chromatography using hexane:ethyl acetate (0→30%) as the mobile phase. The target product was a yellow solid (0.71 g) in a yield of 57.7%. Compound 2 1 The H NMR [CDCl3] spectrum and synthesis scheme are shown in Figure 2.

[0125] <Example 1-3. Synthesis of Compound 3> Compound 2 (0.30 g, 0.87 mmol) was weighed into a two-neck flask and dissolved in DMF (50 mL). K2CO3 (0.26 g, 1.88 mmol) was then added, and the atmosphere was replaced with nitrogen three times. Mel (0.14 mL, 2.2 mmol) was then added dropwise via syringe, and the mixture was stirred at room temperature for 3 days. 2 M NaOH (30 mL) was added to quench the reaction. After confirming the progress of the reaction by TLC (hexane: ethyl acetate = 2:1), the mixture was filtered and the solvent was removed using an evaporator. The compound was then dissolved in ethyl acetate and saturated NaHCO 3aq Extracted twice with saturated NaClaq The organic layer was collected after washing once with Na2SO4. The mixture was dried by stirring for 30 minutes after addition of Na2SO4. After that, the mixture was filtered and the solvent was removed using an evaporator. The mixture was dissolved in ethyl acetate and purified by flash column chromatography using hexane:ethyl acetate (0→30%) as the mobile phase. The target product was a pale yellow solid (0.28 g) in a yield of 93.3%. 1 The H NMR [CDCl3] spectrum and synthesis scheme are shown in Figure 3.

[0126] Example 1-4: Synthesis of PST (Photostatin) Compound 3 (0.28 g, 0.45 mmol) was weighed into a two-neck flask and dissolved in a 5:3 MeOH:CHCl mixture (40 mL). The atmosphere was replaced with nitrogen, and Pd(PPh) (0.011 g, 0.0095 mmol) was added. After stirring at room temperature for 5 minutes, KCO (0.53 g, 3.8 mmol) was added and the mixture was stirred at room temperature for 90 minutes. The color of the solution changed from yellow to reddish-orange. After 90 minutes, the progress of the reaction was confirmed by TLC (hexane:ethyl acetate = 2:1), followed by filtration and removal of the solvent by evaporation. The compound was then dissolved in ethyl acetate and saturated NaHCO 3 . 3aq Extracted twice with saturated NaCl aq The organic layer was collected after washing once with Na2SO4. The mixture was dried by stirring for 30 minutes after adding Na2SO4. After that, the mixture was filtered and the solvent was removed using an evaporator. The residue was dissolved in ethyl acetate and purified by flash column chromatography using hexane:ethyl acetate (0→30%) as the mobile phase. The target product was a reddish-orange oily solid (0.17 g) in a yield of 70.8%. 1 The H NMR [CDCl3] spectrum and synthesis scheme are shown in Figure 4.

[0127] <Example 1-5. Synthesis of PST-amidite monomer> Photostatin (0.08 g, 0.25 mmol) was dissolved in acetonitrile (5 mL) and transferred to a two-neck flask. Under nitrogen, the solution was azeotropically dried three times with dry acetonitrile (2 mL). Then, acetonitrile (5 mL) was added dropwise. Next, DIPEA (348 μL, 2.0 mmol, 8 eq) and 2-cyanoethyldiisopropylchlorophosphoramidite (111 μL, 0.50 mmol, 2 eq) were slowly added dropwise via pipette while cooling on ice. This procedure was carried out under nitrogen. The mixture was stirred at room temperature for 90 min. The color of the solution changed from reddish-orange to bright orange. After 90 min, the progress of the reaction was confirmed by TLC (hexane: ethyl acetate / triethylamine = 60:40:3). The organic layer was extracted twice with saturated NH4Cl2 and washed once with saturated NaCl solution. The mixture was dried by adding Na2SO4 and stirring for 30 minutes. It was then filtered and the solvent was removed using an evaporator. The evaporation was performed without immersing the mixture in a hot water bath. The residue was dissolved in ethyl acetate and purified by flash column chromatography using hexane with triethylamine: ethyl acetate: (0→40%) as the mobile phase. The target product, an orange oily solid (0.12 g), was obtained in 75.2% yield.

[0128] The obtained substance was analyzed by TLC (eluent: hexane: ethyl acetate: triethylamine = 50:50:3). Before the reaction, only a spot with an Rf value of 0.57 was observed. After the reaction, a spot with an Rf value of 0.65 was also observed in addition to the spot with an Rf value of 0.57 (these were the only two spots). The obtained substance was then purified by HPLC and analyzed by mass spectrometry. The mass spectrum of the mass spectrometry is shown in Figure 5. It was confirmed that the PST-amidite monomer had been obtained.

[0129] Example 2. Synthesis of PST-modified nucleic acids Photostatin was subjected to five types of DNA modifications: T9, A, T, G, and C. The structural formula of the target product is shown below.

[0130] [ka]

[0131] The PST-amidite monomer was azeotropically dried three times with dry acetonitrile (2 mL). It was then dissolved in dry acetonitrile and transferred to a brown bottle with a syringe, which was then loaded onto an automated DNA synthesizer. After DNA synthesis, the mixture was transferred to a screw tube, and 500 μL of aqueous ammonia was added. The protecting groups at the nucleobase and phosphate groups of T and T9 were deprotected by leaving them at room temperature for 1 hour; A and C by leaving them at 37°C for 8 hours; and G by leaving them at 65°C for 8 hours. Mass spectrometry was then performed using MALDI-TOF / MS (Figure 6). Furthermore, PST-T9 and PST-T were purified by HPLC before mass spectrometry (Figures 7–9).

[0132] Example 3. Evaluation of cis-trans isomerization Evaluation was carried out by HPLC. Purified PST-T9 was prepared by adding sterile water to a final concentration of 5 μM. Samples were prepared by irradiating visible light at 50% for 30 seconds and ultraviolet light at 50% for 30 seconds. 10 μL of each was injected into an HPLC injector using a syringe, and changes in the peaks were examined. The results are shown in Figure 10. UV irradiation increased the proportion of peaks derived from cis isomers. This demonstrates that UV irradiation can also control the structure of PST-modified nucleic acids.

[0133] Example 4. UV measurement We investigated how the absorption wavelength changes between the cis and trans forms of Photostatin. Purified PST-T9 was adjusted to a final concentration of 5 μM with sterile water and 10x PBS. First, UV measurement of the trans form was performed by irradiating it with 50% visible light for 30 seconds. Then, UV measurement of the cis form was performed by irradiating it with 50% ultra violet light for 30 seconds in a dark place. The results are shown in Figure 11. It was found that cis-trans isomerization occurs.

[0134] Example 5. Time-varying measurements Time-dependent changes were measured to investigate reversibility over time. A similar sample was irradiated with 50% ultra violet light for 30 seconds in the dark, converting it to the cis form. The system was set to automatically measure every 10 minutes, and the presence or absence of reversibility was investigated from the absorption wavelength and absorbance. The results are shown in Figure 12. It was found that the sample exhibited cis-trans reversibility.

[0135] Example 6. Tm measurement PST-T9 was mixed with a complementary single-stranded construct (A9) to form a double-stranded construct, and its melting point was measured. PST-T9 and A9, each adjusted to 10 μM, were mixed and the resulting sample was prepared in 10x PBS to a final concentration of 5 μM. First, the internal temperature of the apparatus was raised to 80°C. The temperature was then lowered to 10°C (-1°C / min) and held at that temperature for 5 minutes. The temperature was then raised from 10°C to 80°C (1°C / min). Finally, the construct was converted to the cis form by irradiating it with 50% ultra violet light for 30 seconds in the dark, and measurements were performed at a rate of 10°C to 60°C (1°C / min). This procedure was performed under a nitrogen gas flow to minimize the effects of condensation. The results are shown in Figure 13. The Tm slightly decreased when the construct was converted to the cis form.

[0136] Example 7. Evaluation of thermal stability The structure of PST-T9 at high temperatures was confirmed by HPLC. Purified PST-T9 was prepared as a sample with sterile water and 10x PBS to a final concentration of 5 μM. The sample was heated to 80°C in a thermal cycler and held at that temperature for 10 minutes. The heated sample was then injected into an HPLC injector using a syringe, and the elution peaks were examined. The results are shown in Figure 14. As the temperature increased, only the trans-isomer peaks were observed.

[0137] Example 8. Anticancer activity test The anticancer activity of PST-T against HeLa cells was examined. Purified PST-T was diluted with sterile water to a final concentration of 20 μM for sample preparation. HeLa cells were seeded in a 96-well plate and pre-cultured for two days. After two days, the medium was removed. The PST-T solution was filter-sterilized and then dispensed into two Eppendorf tubes (50 μL each). One tube was then converted to cis isomers by irradiating with UV light at 50% for 30 seconds, and the other tube was converted to trans isomers by irradiating with viscous light at 50% for 30 seconds. Each tube was then added to a 96-well plate with 2× E-MEM (50 μL) to a final concentration of 1× E-MEM and 10 μM for the sample, and incubated for 8 days. After 8 days, the solution was removed, Live / Dead reagent was added, and the cells were incubated at 37°C. After 30 minutes, the cells were observed using a confocal laser microscope. A control well containing 1× E-MEM alone was also prepared. The results are shown in Figure 15. By converting it into a cis-isomer, stronger anti-cancer activity was exhibited.

[0138] Example 9. Synthesis of CA4-amidite monomer

[0139] [ka]

[0140] CA4 (0.10 g, 0.31 mmol) was dissolved in acetonitrile (5 mL) and transferred to a two-neck flask. Under nitrogen, the solution was azeotropically dried three times with dry acetonitrile (2 mL). Then, acetonitrile (5 mL) was added dropwise. Next, DIPEA (431 μL, 2.48 mmol, 8 eq) and 2-cyanoethyldiisopropylchlorophosphoramidite (137 μL, 0.62 mmol, 2 eq) were slowly added dropwise via pipette while cooling on ice. This procedure was carried out under nitrogen. The mixture was stirred at room temperature for 90 min. The color of the solution changed from white to pale yellow. After 90 min, the progress of the reaction was confirmed by TLC (hexane: ethyl acetate / triethylamine = 60:40:3). The organic layer was then extracted twice with saturated NH4Cl2 and washed once with saturated NaCl solution. The mixture was dried by adding Na2SO4 and stirring for 30 minutes. The mixture was then filtered and the solvent removed using an evaporator. The evaporation was performed without immersing in a hot water bath. The product was dissolved in ethyl acetate and purified by flash column chromatography using hexane with triethylamine: ethyl acetate (0% to 30%) as the mobile phase. The target product, a white solid (0.10 g), was obtained in 63.2% yield. TLC analysis of the resulting product (eluent: hexane: ethyl acetate: triethylamine = 50:50:3) revealed that before the reaction, only a spot with an Rf value of 0.34 was observed. After the reaction, a spot with an Rf value of 0.47 was also observed (these two spots were the only ones present). The resulting product was then purified by HPLC, confirming the formation of the CA4-amidite monomer.

[0141] Example 10. Preparation of PST-modified DNA origami dendrimers PST-modified DNA origami dendrimers were prepared as follows.

[0142] A staple DNA mix for forming a DNA origami dendrimer (overall view: Figure 16, enlarged view of a part of the tip of the dendrimer: Figure 17) was prepared. This was mixed with M13mp18 ssDNA and 10× TAE / Mg 2+ The mixture was diluted with sterile water to a final concentration of 20 nM staple DNA and 4 nM M13mp18 ssDNA. Annealing was then performed using a thermal cycler (90°C for 10 min, 90°C → 25°C, -1°C / min) to form a DNA origami dendrimer. As shown in the left side of Figure 18, the staple DNA at the end has a portion (free strand) that does not anneal with the M13mp18 ssDNA. Staple DNA (PST-staple DNA 1 and PST-staple DNA 2), which has a complementary sequence to the free strand and whose 5' end is modified with PST, can be annealed to the free strand (right side of Figure 18) to obtain a PST-modified DNA origami dendrimer.

[0143] [Table 1]

[0144] After the DNA origami dendrimer was formed, 1.1 μL and 1.3 μL of 67.3 μM PST-staple DNA 1 and 49.6 μM PST-staple DNA 2 were added, respectively, and the mixture was left to stand overnight in a refrigerator. Excess staple DNA was then removed by ultrafiltration (Amicon Ultra 50K). After purification, the solution was concentrated 50-fold and UV absorption measurements were performed using a Nanodrop, confirming the presence of PST absorption (375 nm).

Claims

1. General formula (1): 【Chemistry 1】 [In the formula: R 1 are the same or different and represent an alkoxy group or a phosphate group. 2 are the same or different and represent an alkoxy group or a phosphate group. 3 indicates -N=. R 4 indicates =N-. R 5 represents a protecting group for a hydroxy group. 6 and R 7 are the same or different and represent an alkyl group; m represents an integer of 1 to 5; and n represents an integer of 1 to 4. A compound represented by the formula (I), a salt thereof, or a solvate thereof.

2. The compound represented by general formula (12): 【Chemistry 2】 [wherein R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , m and n are the same as defined above.] The compound according to claim 1, or a salt thereof, or a solvate thereof, which is a compound represented by the formula:

3. The compound has the general formula (1A): 【Transformation 3】 [In the formula: R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , and R 7 is the same as above.] The compound according to claim 1, or a salt thereof, or a solvate thereof, which is a compound represented by the formula:

4. R 1 and R 2 The compound according to claim 1, or a salt thereof, or a solvate thereof, wherein is an alkoxy group.

5. R 5 is an alkyl group substituted with a cyano group, a nitro group, an alkylsulfonyl group, a halogen atom, an arylsulfonyl group, a trihalomethyl group, or a trialkylamino group, and R 6 and R 7 The compound or salt thereof, or a solvate of the compound or salt according to any one of claims 1 to 4, wherein is a branched alkyl group.

6. The terminal phosphate group has the general formula (2): 【Chemistry 4】 [In the formula: R 1 are the same or different and represent an alkoxy group or a phosphate group. 2 are the same or different and represent an alkoxy group or a phosphate group. 3 indicates -N=. R 4 indicates =N-. R 8 represents O or S. R 9 O - or S - m represents an integer of 1 to 5, and n represents an integer of 1 to 4. A nucleotide or polynucleotide in which the nucleotide or polynucleotide is replaced with a group represented by the following formula:

7. General formula (2A): 【Transformation 5】 [In the formula: R 1 , R 2 , R 3 , R 4 , R 8 , R 9 , m, and n are the same as above. 8A are the same or different and represent O or S. R 9A are the same or different, O - or S - Indicates R 10 R may be the same or different and represent a divalent group formed by removing two hydroxy groups from the sugar moiety of a nucleoside. 10A R may be the same or different and represent a divalent group formed by removing two hydroxy groups from the sugar moiety of a nucleoside. 11 represents a hydroxy group, a phosphate group, or a thiophosphate group, and p represents an integer of 0 or 1 or more. The nucleotide or polynucleotide according to claim 6, which is a compound represented by the formula:

8. A nucleic acid construct comprising the nucleotide or polynucleotide of claim 6.

9. The nucleic acid structure of claim 8, which is a DNA origami.

10. A pharmaceutical comprising at least one selected from the group consisting of the nucleotide or polynucleotide according to claim 6 or 7 and the nucleic acid structure according to claim 8 or 9.

11. The pharmaceutical composition according to claim 10, which is an anticancer agent.

12. A reagent comprising at least one selected from the group consisting of the nucleotide or polynucleotide according to claim 6 or 7 and the nucleic acid structure according to claim 8 or 9.

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