Title of the invention: A compound having an affinity for an antibody, a cleaving moiety and a reactive group, or a salt thereof.
Technical field
[0001]
　The present invention relates to an affinity substance for an antibody, a compound having a cleaving moiety and a reactive group, or a salt thereof.
Background technology
[0002]
　In recent years, research and development of antibody-drug conjugates (ADCs) have been actively carried out. As the name implies, ADC is a drug in which a drug (eg, an anticancer drug) is conjugated to an antibody, and has direct cell-killing activity against cancer cells and the like. A typical ADC is T-DM1 (trade name: Kadcyla (registered trademark)) (Non-Patent Documents 1 to 3).
[0003]
　The non-uniformity of ADCs such as T-DM1 has been a problem from the beginning of development. That is, since a small molecule drug is randomly reacted with Lys residues of about 70 to 80 in the antibody, the drug antibody ratio (Drug / Antibody Ratio: DAR) and the conjugation position are not constant. It is known that usually, in such a random conjugation method, the DA is in the range of 0 to 8, and a plurality of drugs having different numbers of drug bindings are produced. In recent years, it has been reported that the pharmacokinetics, drug release rate, and effect change when the number and position of binding of a drug in ADC are changed. For these reasons, next-generation ADCs are required to control the number and position of conjugating drugs. It is considered that if the number and position are constant, the problems of efficacy, variation of conjugation drug, and lot difference, so-called regulation, as expected can be solved (Non-Patent Document 4).
[0004]
　Regioselective modifications of antibodies have been studied all over the world, most of which are genetically engineered or enzymatically modified. Regarding the genetic engineering modification method, although the regioselectivity and the number selectivity can be controlled, problems such as a decrease in the expression efficiency of the antibody itself (a decrease in the total yield when preparing the ADC) have been pointed out. In addition, it takes a long time to construct an antibody expression system (Non-Patent Documents 5 to 7).
[0005]
　In addition, a method of chemically modifying a protein in a congested environment such as inside a cell using a small molecule probe has been reported in recent years. This method is used for receptor identification in imaging and repositioning of small molecule drugs. Further, in the field of chemical biology, an organic chemical protein modification method using a synthetic small molecule probe has attracted attention (Non-Patent Documents 8 to 10).
[0006]
　Recently, the CCAP (Chemical Conjugation by Affinity Peptide) method has been developed. This method is carried out by reacting a peptide reagent in which an NHS activating ester and a drug are linked to an affinity peptide with an antibody (that is, a method for producing an ADC via a linker containing a peptide moiety). We have succeeded in position-selective modification of the antibody. This method was the first in the world to succeed in regioselectively modifying the antibody Fc region with a drug by a chemically synthesized method, and the results were also good in practice [reaction time 30 minutes, yield 70%]. (In the case of DAR 1), regioselectivity is 100%] has been confirmed. It has been demonstrated that the DA can be controlled by 2 by adding about 5 equivalents of the peptide reagent, and it is epoch-making in that the modification position can also be controlled (Patent Document 1).
Prior art literature
Patent documents
[0007]
Patent Document 1: International Publication No. 2016/186206
Non-patent literature
[0008]
Non-Patent Document 1: Reichert JM et al. , Nat Biotechnol 2005; 23: 1073-8
Non-Patent Document 2: Kubota T et al. , Cancer Science 2009; 100: 1566-72
Non-Patent Document 3: Wu AM et al. , Nat Biotechnol 2005; 23: 1137-46
Non-Patent Document 4: Juntura JR et al. , Nat Biotechnol 2008; 26: 925-32
Non-Patent Document 5: Shen BQ et al. , Nat Biotechnol 2012; 30: 184-9
Non-Patent Document 6: Hofer T et al. , Biochemistry 2009; 48: 12047-57
Non-Patent Document 7: Liu W et al. , Nat Methods 2007; 4: 239-44
Non-Patent Document 8: S.A. T. Laughlin et al. , Science 2008; 320,664
Non-Patent Document 9: A. E. Spaces et al. , ChemBioChem 2004; 5,41
Non-Patent Document 10: Y. et al . Takaoka et al. , Angew. Chem. Int. Ed. 2013; 52,4088
Outline of the invention
Problems to be solved by the invention
[0009]
　An object of the present invention is to develop a technique that enables modification of an antibody, particularly, regioselective modification of an antibody.
Means to solve problems
[0010]
　As a result of diligent studies, the present inventors have (1) an affinity substance for an antibody, (2) a reactive group for an amino acid residue constituting the antibody, and (3) the affinity substance and the reactive group. A structural unit containing a cleavable moiety between and (4) having a bioorthogonal functional group on the reactive group side by cleavage at the cleavable moiety (ie, a structural unit containing a bioorthogonal functional group and a reactive group ) Has been found to be useful for position-specific modification of antibodies (eg, FIG. 1-1, FIG.), A compound developed based on a novel and original design concept having the structural feature of being able to generate. 1-2, Fig. 1-3, Fig. 2). The present inventors also use such compounds to regioselectively have a functional substance (eg, drug) that does not contain a peptide moiety as a linker (eg, antibody drug conjugate (ADC)). It was found that it can be prepared. Avoidance of the use of linkers containing peptide moieties that have potential immunogenicity and are susceptible to hydrolysis in the blood is desirable in clinical applications of ADCs. That is, the method developed by the present inventors was the first in the world to succeed in regioselectively modifying the antibody Fc region with a drug by a chemically synthesized method and without using a linker containing a peptide moiety. be able to. The present inventors have also succeeded in developing various compounds having the structural characteristics as described in (1) to (4) above (eg, FIGS. 1-1, 1-2, 1-3). 2), the present invention has been completed.
[0011]
　That is, the present invention is as follows.
[0012]
　In a first embodiment, the invention provides a regioselective modifying reagent for an antibody comprising an affinity for the antibody, a compound having a cleaving moiety and a reactive group or a salt thereof, and the compound or a salt thereof. ..
[1] A compound having an affinity substance for an antibody, a cleaving moiety and a reactive group or a salt thereof,
　wherein the compound is the following formula (I):
ALBR (I)
[in the formula,
　A is
　a cleaving linker which is an affinity substance for an antibody, L is a divalent group containing a cleaving moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group, or (B) A divalent group that does not contain a bioorthogonal functional group, and
　R is a reactive group for the antibody. ], And the
　affinity substance for the antibody is the
formula 1-1: 1 (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-I-I-W- C- (X 0-3 ) b (SEQ ID NO
: 58) Equation 1-2: (X 0-3 ) a-Xaa1--C-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-I-V W-C-(X 0-3 ) b (SEQ ID NO: 59)
expression 1-3: (X 0-3 ) a -C- Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-VVWC- (X 0-3 ) b (SEQ ID NO
: 60) Equation 1-4: (X 0-3 ) a- C-Xaa1-Xaa2 -Xaa3-Xaa4-Xaa5-Xaa6-AV-WC- (X 0-3 ) b (SEQ ID NO
: 61) Equation 1-5: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3- Xaa4-Xaa5-Xaa6-L-L-WC- (X 0-3 ) b (SEQ ID NO
: 62) Equation 1-6: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-LIWC- (X 0-3 ) b (SEQ ID NO
: 63) Equation 1-7: (X 0-3 ) a- C -Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-VFC- (X 0-3 ) b (SEQ ID NO
: 64) Equation 1-8: (X 0-3 ) a- C-Xaa1- Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-QVWC- (X 0-3 ) b (SEQ ID NO
: 65) Equation 1-9: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3 -Xaa4-Xaa5-Xaa6-EVW-C- (X 0-3 ) b (SEQ ID NO: 66)
[In the formula,
　(X) 0-3 ) a is none, or 1 to 3 consecutive identical or different arbitrary amino acid residues (other than lysine and cysteine ​​residues), and
　(X 0-3 ) b is none, or 1 to 3 consecutive identical or different arbitrary amino acid residues (other than lysine residue and cysteine ​​residue),
　Xaa1 is an alanine residue, a glycine residue, a leucine residue, a proline residue, an arginine residue. , Valin residue, asparagine residue, glutamate residue, or phenylalanine residue,
　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue, and
　Xaa3 is a histidine residue, phenylalanine. Residues, tyrosine residues, tryptophan residues, arginine residues, or glycine residues,
　Xaa4 is a lysine residue,
　Xaa5 is a glycine residue, serine residue, asparagine residue, glutamine residue, With aspartic acid residue, glutamate residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue, alanine residue, valine residue, isoleucine residue, or arginine residue There,
　Xaa @ 6 is a glutamine residue, a glutamic acid residue, asparagine residue, aspartic acid residue, a proline residue, glycine residue, arginine residue, a phenylalanine residue or a histidine residue. ], Or
Equation 2-1: (X 0-3 ') a- C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-LVWC- (X 0-3 ') b (SEQ ID NO: 67)
[In the equation,
　(X 0-3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively.
　Xaa1', Xaa2', Xaa3', Xaa4', Xaa5', and Xaa6'are the same as Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different amino acid residues, (X 0-3 ') b.Except when 1 to 3 consecutive identical or different amino acid residues, Xaa3'is a histidine residue, and Xaa5'is a glycine residue. ], A compound or a salt thereof, which is a peptide containing any of the amino acid sequences of.
[2] (X 0-3 ) a is none, arginine residue-glycine residue-asparagin residue, asparaginic acid residue, or asparagine residue, and
　(X 0-3 ) b is none, threonine. Residue-tyrosine residue-histidine residue, or threonine residue
　,
　Xaa1 is an alanine residue, Xaa2 is a tyrosine residue, tryptophan residue, or histidine residue, and
　Xaa6 is a glutamine residue. A compound of [1] or a salt thereof, which is a group, a glutamic acid residue, an asparagine residue, or an aspartic acid residue.
[3] (X 0-3 ) a is a glycine residue-asparagin residue, a glycine residue, a glycine residue-glycine residue, or a glycine residue-glycine residue-glycine residue, and
　(X 0). -3 ) bIs a threonine residue-tyrosine residue, a glycine residue, a glycine residue-glycine residue, or a glycine residue-glycine residue-glycine residue, and
　Xaa1 is a glycine residue, a leucine residue, a proline residue. Group, arginine residue, valine residue, asparagine residue, glutamate residue, or phenylalanine residue,
　Xaa2 is a phenylalanine residue, and
　Xaa6 is a proline residue, glycine residue, arginine residue, phenylalanine. The compound of [1] or a salt thereof, which is a residue or a histidine residue.
[4] The compound of any of [1] to [3] or a salt thereof, wherein the affinity substance for the antibody is a peptide containing an amino acid sequence selected from the group consisting of the following:
(1) RGNCAYHKGQIIWCTYH (SEQ ID NO: 5). ;
(2) RGNCAYHKGQIVWCTYH (SEQ ID NO:
8); (3) RGNCAYHKGQVVWCTYH (SEQ ID NO:
9); (4) RGNCAYHKGQAVWCTYH (SEQ ID NO:
10); (5) RGNCAYHKGQLLWCTYH (SEQ ID NO:
11); (6) RGNCAYHKGQLIWCTYH (SEQ ID NO: 12) ;
(7) DCAYHKGQIVWCT (SEQ ID NO:
13); (8) DCAYHKGQVVWCT (SEQ ID NO:
14); (9) DCAYHKGQAVWCT (SEQ ID NO: 15);
(10) RGNCAYHKSQIIWCTYH (SEQ ID NO: 16);
(11) RGNCAYHKNQIIWCTYH (SEQ ID NO: 17);
(12) RGNCAYHKDQIIWCTYH (SEQ ID NO: 18);
(13) RGNCAYHKQQIIWCTYH (SEQ ID NO: 19);
(14)
(15) RGNCAYHKFQIIWCTYH (SEQ ID NO: 21);
(16) RGNCAYHKYQIIWCTYH (SEQ ID NO: 22);
(17) RGNCAYHKWQIIWCTYH (SEQ ID NO: 23);
(18) RGNCAYHKHQIIWCTYH (SEQ ID NO: 24
);
(20) RGNCAYHKLQIIWCTYH (SEQ ID NO: 26);
(21) CAYHKLQIVWC (SEQ ID NO: 27);
(22) CAYHKLQLIWC (SEQ ID NO: 28);
(23) CAYHKSQIVWC (SEQ ID NO: 29
);
(25) RGNCAYHKGQQVWCTYH (SEQ ID NO: 31);
(26) RGNCAYHKGQEVWCTYH (SEQ ID NO: 32);
(27) CAYHKGQLVWC (SEQ ID NO: 33);
(28) RGNCAYHKAQLVWCTYH (SEQ ID NO: 34);
(29) RGNCAYHKVQLVWCTYH (SEQ ID NO: 35);
(30) RGNCAYHKLQLVWCTYH (SEQ ID NO: 36);
(31)
(32) RGNCAYHKSQLVWCTYH (SEQ ID NO: 38);
(33) RGNCAYHKTQLVWCTYH (SEQ ID NO: 39);
(34) RGNCAYHKNQLVWCTYH (SEQ ID NO: 40);
(35) RGNCAYHKDQLVWCTYH (SEQ ID NO: 41
);
(37) RGNCAYHKEQLVWCTYH (SEQ ID NO: 43);
(38) RGNCAYHKFQLVWCTYH (SEQ ID NO: 44);
(39) RGNCAYHKRQLVWCTYH (SEQ ID NO: 45);
(40) RGNCAYHKHQLVWCTYH (SEQ ID NO: 46
);
(42) RGNCAYHKYQLVWCTYH (SEQ ID NO: 48);
(43) RGNCAYFKGQLVWCTYH (SEQ ID NO: 49);
(44) RGNCAYKYKGQLVWCTYH (SEQ ID NO: 50);
(45) RGNCAYWKGQLVWCTYH (SEQ ID NO: 51);
(46) RGNCAYRKGQLVWCTYH (SEQ ID NO: 52);
(47) RGNCAYGKGQLVWCTYH (SEQ ID NO: 53
);
(49) NCAYHKGQLVWC (SEQ ID NO: 55);
(50) CAYHKGQLVWCT (SEQ ID NO: 56);
(51) CAYHKSQLVWC (SEQ ID NO: 57);
(52) RGNCAWHKGQIIWCTYH (SEQ ID NO: 68);
(53) RGNCAFHKII
(54) RGNCAHHKGQIIWCTYH (SEQ ID NO: 70);
(55) RGNCGYHKGQIIWCTYH (SEQ ID NO: 71);
(56) RGNCLYHKGQIIWCTYH (SEQ ID NO: 72);
(57) RGNCPYHKGQIIWCTYH (SEQ ID NO: 73
);
(59) RGNCVYHKGQIIWCTYH (SEQ ID NO: 75);
(60) RGNCNYHKGQIIWCTYH (SEQ ID NO: 76);
(61) RGNCEYHKGQIIWCTYH (SEQ ID NO: 77);
(62) RGNCFYHKGQIIWCTYH (SEQ ID NO: 78);
(63) RGNCAYHKGEIIWCTYH (SEQ ID NO: 79);
(64) RGNCAYHKGNIIWCTYH (SEQ ID NO: 80);
(65)
(66) RGNCAYHKGGIIWCTYH (SEQ ID NO: 82);
(67) RGNCAYHKGDIIWCTYH (SEQ ID NO: 83);
(68) RGNCAYHKGRIIWCTYH (SEQ ID NO: 84);
(69) RGNCAYHKGFIIWCTYH (SEQ ID NO: 85);
(70) RGN
(71) DCAYHKGQIIWCT (SEQ ID NO: 87);
(72) NCAYHKGQIIWCT (SEQ ID NO: 88);
(73) GNCAYHKGQIIWCTY (SEQ ID NO: 89);
(74) GCAYHKGQIIWCG (SEQ ID NO: 90);
(75) And
(76) GGGCAYHKGQIIWCGGG (SEQ ID NO: 92).
[5] A cleavable linker in which L is a divalent group containing a cleavable moiety capable of producing a bioorthogonal functional group on the reactive group side by (i) cleavage, or bioorthogonal by (ii) cleavage. The compound or salt thereof according to any one of [1] to [4], which is a divalent group containing a cleaving moiety that does not have the ability to generate a sex functional group on the reactive group side.
[6] The compound of [5] or a salt thereof, wherein L is the cleaving linker of (i).
[7] The compound of [5] or [6] or a salt thereof, wherein L is the cleaving linker of (i) and B is the divalent group of (b).
[8] The compound of [5] or a salt thereof, wherein L is the cleaving linker of (ii) and B is the divalent group of (a).
[9] The compound according to any one of [1] to [8] or a salt thereof, wherein the peptide is a peptide that binds to the Fc region of a monoclonal antibody.
[10] The compound of [9] or a salt thereof, wherein the peptide is a peptide that binds to the Fc region of IgG.
[11] The affinity substance is an affinity substance for an antibody containing any one Fc region protein selected from the group consisting of the following (A) to (C) and having an antigen-binding ability [1]. ~ [10] Compound or salt thereof:
(A) Fc region protein containing the amino acid sequence of SEQ ID NO : 1;
(B) Insert one or several amino acid residues in the amino acid sequence of SEQ ID NO: 1. Fc region protein containing an added, deleted or substituted amino acid sequence; or
(C) An Fc region protein containing an amino acid sequence showing 90% or more identity with the amino acid sequence of SEQ ID NO: 1.
[12] A compound according to any one of [1] to [11] or a salt thereof, characterized in that the peptide can bind to human IgG.
[13] The cleavable part is (a) treated with one or more substances selected from the group consisting of acidic substances, basic substances, reducing agents, oxidizing agents, and enzymes, and (b) physics selected from the group consisting of light. Any of [1] to [12], which is a moiety that can be cleaved by either treatment with a chemical stimulus or (c) leaving it to stand when a cleaving linker containing a self-degradable cleaving moiety is used. A compound or a salt thereof.
[14] The cleaving moiety is a disulfide residue, an acetal residue, a ketal residue, an ester residue, a carbamoyl residue, an alkoxyalkyl residue, an imine residue, a tertiary alkyloxycarbamate residue, a silane residue, and the like. Hydrazone-containing residues, phosphoramidate residues, acetal residues, trityl residues, azo residues, vicinaldiol residues, selenium residues, aromatic ring-containing residues with electron-withdrawing groups, coumarin-containing residues , A compound according to any one of [1] to [13], or a salt thereof, selected from the group consisting of sulfone-containing residues, unsaturated bond-containing chain residues, and glycosyl residues.
[15] The cleaving moiety of (i) is selected from the group consisting of disulfide residues, ester residues, acetal residues, ketal residues, imine residues, and vicinaldiol residues [5] to [ 14] Any compound or salt thereof.
[16] The cleavable portion of (ii) is an ester residue, a carbamoyl residue, an alkoxyalkyl residue, an imine residue, a tertiary alkyloxycarbamate residue, a silane residue, a hydrazone-containing residue, or a phosphorami. Ester residues, aconityl residues, trityl residues, azo residues, vicinaldiol residues, selenium residues, aromatic ring-containing residues with electron-withdrawing groups, coumarin-containing residues, sulfone-containing residues, unsaturated The compound of any one of [5] to [14] or a salt thereof, which is selected from the group consisting of bond-containing chain residues and glycosyl residues.
[17] The cuttable part is as follows :
[Chemical

formula 1] [Here, the wavy line orthogonal to the bond indicates the cut site, and the
　plurality of R 2a , the plurality of R 2b , and the plurality of R 2c are the same or different.
Te, (i) a hydrogen atom or a halogen
atom,; (ii) 1 monovalent hydrocarbon
group; (iii)
aralkyl; (iv) 1-valent heterocyclic
group; (v) R c -O-, R c -C (= O)-, R c - OC ( = O)-, or R c- C (= O) -O- (R c ) Indicates a hydrogen atom or a monovalent hydrocarbon group.
); (Vi) NR d R e -, NR d R e -C (= O) -, NR d R e -C (= O) -O-, or R d -C (= O) -NR e - (R d and Re represent the same or different hydrogen atoms or monovalent hydrocarbon groups.); Or (vi) consists of a nitro group, a sulfate group, a sulfonic acid group, a cyano group, or a carboxyl group. Selected from the group, 　J is -CH 2- , -O-, or -S-, 　r is any integer from 1 to 4, 　○ (white circle) indicates a bond to A, and ● ( Black circles) indicate the binding to B.

　If the chemical structure is asymmetric about the cleavage site, ● may indicate a bond to A and ◯ may indicate a bond to B. ], Which corresponds to any one of the chemical structures selected from the group consisting of [1] to [16], or a salt thereof.
[18] The cuttable portion of (i) above is as follows:
[Chemical

formula 2] [Here, the wavy line orthogonal to the bond indicates the cutting site,
　R 2a is the same as in [23], and
　○ (white circle). ) Indicates the bond to A, and ● (black circle) indicates the bond to B.
　If the chemical structure is asymmetric about the cleavage site, ● may indicate a bond to A and ◯ may indicate a bond to B. ], A compound according to any one of [5] to [13], [15], and [17], or a salt thereof, corresponding to any one of the chemical structures selected from the group consisting of.
[19] The cuttable portion of (ii) is as follows:
[Chemical

formula 3] [Here, the wavy line orthogonal to the bond indicates the cut site, and
　R 2b , R 2c , J, r are [23]. The same is true:
　○ (white circle) indicates a bond to A, and ● (black circle) indicates a bond to B.
　If the chemical structure is asymmetric about the cleavage site, ● may indicate a bond to A and ◯ may indicate a bond to B. ], A compound according to any one of [5] to [13], [16], and [17], or a salt thereof, corresponding to any one of the chemical structures selected from the group consisting of.
[20] L is the following formula (L1) to (L3):
La-C-Lb (L1)
La-C (L2)
　　　C-Lb (L3)
[In the formula,
　La and Lb are divalent, respectively. The base,
　C is the cuttable moiety. ], The compound according to any one of [1] to [19] or a salt thereof.
[21] The La and Lb are the following (La') and (Lb'), respectively:
[Chemical

formula 4] [In the formula,
　p and p'are the same or different, and are arbitrary integers from 0 to 10. ,
　Q and q'are the same or different, any integer from 0 to 10, and
　X and X'are the same or different, carbon atom, nitrogen atom, or single bond (where X is the nitrogen atom). If R 1b is absent and X'is a nitrogen atom, then R 1b'is absent. If X is a single bond, then R 1a and R 1b does not exist, and if X'is a single bond, then R 1a'and R 1b' do not exist), and
　R 1a , R 1b , R 1a'and R 1b'are the same or different. An atom or group selected from the group consisting of (i) to (vii). ], The compound of [20] or a salt thereof.
[22] Divalent groups including bioorthogonal functional groups are azide residue, aldehyde residue, thiol residue, alkin residue, alkene residue, tetrazine residue, nitron residue, hydroxylamine residue, nitrile. Residues, hydrazine residues, ketone residues, boronic acid residues, cyanobenzothiazole residues, allyl residues, phosphine residues, maleimide residues, disulfide residues, thioester residues, α-halocarbonyl residues, isonitriles The compound of any one of [1] to [21] or a salt thereof, which is a divalent group containing a bioorthogonal functional group selected from the group consisting of residues, sidonone residues, and selenium residues in the main chain.
[23] Divalent groups including bioorthogonal functional groups are azide residue, aldehyde residue, thiol residue, alkin residue, alkene residue, halogen residue, tetrazine residue, nitron residue, hydroxylamine. Residues, nitrile residues, hydrazine residues, ketone residues, boronic acid residues, cyanobenzothiazole residues, allyl residues, phosphine residues, maleimide residues, disulfide residues, α-halocarbonyl residues, isonitrile The compound of any one of [1] to [21] or a salt thereof, which is a divalent group containing a bioorthogonal functional group selected from the group consisting of residues, sidonone residues, and selenium residues in the side chain.
[24] The bioorthogonal functional groups are as follows:
[Chemical

formula 5] [In the formula,
　R 1f , single or multiple R 1 g and single or multiple R 1 h are the same or different, and the above (i) to It is an atom or group selected from the group consisting of (vii), or an electron-withdrawing group,
　and is a bonder. ), Which is any one of the compounds [1] to [23] or a salt thereof.
[25] The divalent group of (b) is an alkylene optionally substituted, a cycloalkylene optionally substituted, an aryl optionally substituted, or a divalent heterocycle optionally substituted. Group, -NR a- (R a)Indicates a hydrogen atom or a substituent), —O—, or a compound according to any one of [1] to [24] or a salt thereof selected from the group consisting of two or more combinations thereof.
[26] B is the following formula (B-1):
[Chemical

formula 6] [In the formula,
　Y is -NH-, -O-, -CH 2- , or the following formula (B-2):
[Chemical formula 7] ]

(wherein,
　V and V 'are the same or different, -NH -, - O -, - CH 2 -, or a single
　bond, V1 is a divalent group containing a bio-orthogonal functional group Yes,
　s is any integer from 0 to 10,
　and ○ and ● in equation (B-2) have the same orientation as ○ and ● in equation (B-1), respectively), and
　Z. Is an oxygen atom, a sulfur atom, or a hydrogen atom (when Z is a hydrogen atom, -C (= Z)-indicates -CH 2- ), and is a
　circle (white circle) in the formula (B-1). ) Indicates the bond to the L-side portion, and ● (black circle) indicates the bond to the R-side portion. ], The compound according to any one of [1] to [25] or a salt thereof.
[27] Any of [1] to [26], wherein the reactive group is a reactive group specific for any one side chain of a lysine residue, a tyrosine residue, or a tryptophan residue. Compound or salt thereof.
[28] The compound of [27] or a salt thereof, wherein the reactive group is a reactive group specific for the side chain of a lysine residue.
[29] The reactive group is as follows :
[Chemical

formula 8] [Here,
　R 5a and R 5c are atoms or groups selected from the group consisting of (i) to (vii), and
　R 5b is an electron. It is an attractive group,
　j is an arbitrary integer of 1 to 5, and
　k is an arbitrary integer of 1 to 4. ], Which corresponds to any one of the chemical structures selected from the group consisting of [1] to [28], or a salt thereof.
[30] A compound according to any one of [1] to [29] or a salt thereof, wherein the main chain connecting A and R has 4 to 20 atoms.
[31] The compound of any one of [1] to [30] or a salt thereof, wherein the main chain connecting A and R does not contain a ring structure.
[32] The compound according to any one of [1] to [31] or a salt thereof, wherein the partial structure represented by LB does not contain a peptide moiety.
[33] The compound represented by the formula (I) is the following (I'):
A-B2-L'-B1-R (I')
[In the formula,
　A and R are the same as those in the formula (I), and
　L'is a divalent group containing a cleaving moiety.
　B1 and B2 are sex linkers, which are the same or different, (a) a divalent group containing a bioorthogonal functional group, or (b) a divalent group not containing a bioorthogonal functional group, and
　B1. And B2 may have a symmetrical structure centered on L'. ], Which is any compound of [1] to [32] or a salt thereof.
[34] The compound represented by the formula (I') is the following (I''):
[Chemical

formula 9] [In the formula,
　A and R are the same as those of the formula (I) described in [1]. Yes,
　C is a cleavable moiety, and
　p, p', q, q', X, X', R 1a , R 1a' , R 1b , and R 1b' are the formulas (La) described in [19]. ') And (Lb'), and
　Y and Y'are the same as or different from Y in formula (B-1) described in [24].
　Z and Z'are the same as Z in the above formula (B-1), the same as or different from each other. ], The compound of [33] or a salt thereof.
[35] A position-selective modification reagent for an antibody, which has the following
　formula (I):
ALBR (I)
[In the formula,
　A is an affinity substance for the antibody, and
　L is a cleavage substance. It is a cleaving linker which is a divalent group containing a moiety, and
　B is a divalent group containing (a) a bioorthogonal functional group or (b) a divalent group containing no bioorthogonal functional group. Yes,
　R is a reactive group for the antibody. ], The substances having an affinity for an antibody, a cleaving moiety and a compound having a reactive group or a salt thereof, and having an affinity for an antibody are represented by the
　above formulas 1-1 to 1-9 and formula 2-. Peptide containing any one of the amino acid sequences of 1 [Preferably, a peptide containing an amino acid sequence selected from the group consisting of (1) to (76) above. The same applies hereinafter], reagents.
[36] A compound having an affinity substance, a cleaving moiety and a reactive group for an antibody or a salt thereof 　, wherein A is an antibody in the following
　formula (I):
ALBR (I)
[In the formula,
A is an antibody. for a affinity substance,
　L is a divalent cleavable linker is a group containing a cleavable moiety,
　B is (a) a divalent group containing a bioorthogonal functional group, or (b) a divalent group not containing a bioorthogonal functional group, and
　R is specific to the side chain of the lysine residue. Reactive group. ], And the
　substance having an affinity for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1. A compound or a salt thereof.
[37] A position-selective modification reagent for an antibody, which has the following
　formula (I):
ALBR (I)
[In the formula,
　A is an affinity substance for an antibody, and
　L is a cleavage substance. It is a cleavable linker which is a divalent group containing a moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group or (b) a divalent group not containing a bioorthogonal functional group. Yes,
　R is a reactive group specific for the side chain of the lysine residue. ], The substances having an affinity for an antibody, a cleaving moiety and a compound having a reactive group or a salt thereof, and having an affinity for an antibody are represented by the
　above formulas 1-1 to 1-9 and formula 2-. A reagent which is a peptide containing any of the amino acid sequences of 1.
[0013]
　Secondly, the present invention provides an affinity substance for an antibody, an antibody having a cleaving moiety or a salt thereof, and a method for producing the same.
(Antibody having an affinity for an antibody and an antibody having a cleaving moiety or a salt thereof)
[38] An antibody having an affinity for an antibody and a cleaving moiety or a salt thereof, wherein the following
　formula (II):
AL : -BR'-T (II)
[In the formula,
　A is an affinity substance for an antibody,
　L is a cleavable linker which is a divalent group containing a cleavable moiety, and
　B is (a). ) A divalent group containing a bioorthogonal functional group, or (b) a divalent group not containing a bioorthogonal functional group, where R'is the moiety
　produced by the reaction between the antibody and the reactive group. And
　T is an antibody. ], And the
　affinity substance for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1. The antibody or a salt thereof.
[39] The antibody of [38] or a salt thereof, wherein the antibody is a monoclonal antibody.
[40] An antibody of [38] or [39] or a salt thereof, wherein the antibody is an IgG antibody.
[41] The antibody or salt thereof according to any one of [38] to [40], wherein the antibody is of human origin.
[42] Any one of [38] to [41], wherein the antibody contains any one Fc region protein selected from the group consisting of the following (A) to (C) and has an antigen-binding ability. Antibody or salt thereof:
(A) Fc region protein containing the amino acid sequence of SEQ ID NO : 1;
(B) In the amino acid sequence of SEQ ID NO: 1, one or several amino acid residues are inserted, added, deleted or substituted. Fc region protein containing an amino acid sequence; or
(C) an Fc region protein containing an amino acid sequence showing 90% or more identity with the amino acid sequence of SEQ ID NO: 1.
[43] The antibody contains one or more specific amino acid residues in a target region consisting of 1 to 50 consecutive amino acid residues, and the specific amino acid residue in a non-target region other than the target region. The
　structural unit containing 5 or more groups and represented by ALBR'has a regioselectivity of 30% or more with respect to one or more specific amino acid residues contained in the target region. An antibody or salt thereof according to any one of [38] to [42] that is bound.
[44] The antibody of [43] or a salt thereof, wherein the target region is a region consisting of 1 to 10 consecutive amino acid residues.
[45] The antibody of [44] or a salt thereof, wherein the target region is a region consisting of 1 to 3 consecutive amino acid residues.
[46] The antibody or salt thereof of [45], wherein the target region is a region consisting of amino acid residues at positions 246 to 248 in the human IgG Fc region.
[47] The antibody or salt thereof according to any one of [43] to [46], wherein the regioselectivity is 50% or more.
[48] ​​The antibody or salt thereof of [47], which has 70% or more of the regioselectivity.
[49] The antibody or salt thereof of [48], which has 90% or more of the regioselectivity.
[50] The specific amino acid residue is a number with respect to the N-terminal side and the C-terminal side, respectively, centered on the specific amino acid existing at the specific position (where a is an arbitrary integer of 1 to 10). In the region up to the distant position of the amino acid residue of), the amino acid residue of the same type as the specific amino acid residue is not contained in addition to the specific amino acid residue existing at the specific position, [43] to [ 49] Any antibody or salt thereof.
[51] The antibody is an antibody containing a plurality of heavy chains, and
　T is a structure represented by ALBR'in a plurality of corresponding target regions in the plurality of heavy chains. The antibody or salt thereof according to any one of [38] to [50], wherein the antibody has a plurality of structural units represented by ALBR'as a result of having the unit.
[52] The antibody of [51] or a salt thereof, which has two heavy chains.
[53] The portion produced by the reaction between the antibody and the reactive group is any one of the lysine residue, tyrosine residue, or tryptophan residue with respect to the lysine residue, tyrosine residue, or tryptophan residue. The antibody or salt thereof according to any one of [38] to [52], which is a portion produced by the reaction of a reactive group specific to the side chain.
[54] The portion produced by the reaction between the antibody and the reactive group is the portion produced by the reaction between the lysine residue and the reactive group specific to the side chain of the lysine residue. , [38]-[53], or a salt thereof.
[55] The portion generated by the reaction is as follows:
[Chemical formula 10]

[Here, ● (black circle) indicates a bond to the T-side portion, and ○ (white circle) indicates a bond to the B-side portion. A straight line orthogonal to the bond indicates the bond formed by the reaction. ], Which corresponds to any one chemical structure selected from the group consisting of [38] to [54], or a salt thereof.
[56] The antibody or salt thereof according to any one of [38] to [55], wherein the main chain connecting A and R'has 4 to 20 atoms.
[57] The antibody or salt thereof according to any one of [38] to [56], wherein the main chain connecting A and R does not contain a ring structure.
[58] The antibody or salt thereof according to any one of [38] to [57], wherein the partial structure represented by LB does not contain a peptide moiety.
[59] The compound represented by the formula (II) is the following (II'):
A-B2-L'-B1-R'-T (II')
[In the formula,
　A, R'and T are Same as that of formula (II),
　L'is a cleavable linker that is a divalent group containing a cleavable moiety, and
　B1 and B2 are the same or different, (a) bioorthogonal functional groups. It is a divalent group containing, or (b) a divalent group not containing a bioorthogonal functional group, and
　B1 and B2 may have a symmetrical structure centered on L'. ], Which is the antibody of any one of [38] to [58] or a salt thereof.
[60] The compound represented by the formula (II') is described in the following (II''):
[Chemical formula 11].

[In the formula,
　A, R'and T are the same as those in formula (II) described in [36],
　C is a cutting part, and
　p and p'are the same or different, from 0 to. Any integer of 10,
　q and q'are the same or different, any integer from 0 to 10, and
　X and X'are the same or different, carbon atom, nitrogen atom, or single bond ( Here, when X is a nitrogen atom, R 1b does not exist , when X'is a nitrogen atom, R 1b' does not exist. When X is a single bond, R 1a and R 1b exist. If X'is a single bond, then R 1a' and R 1b' do not exist), and
　R 1a , R 1b , R 1a'and R 1b'are the same or different,
(i) hydrogen atom. , Or halogen atom;
(iii) monovalent hydrocarbon group;
(iii) aralkyl;
(Iv) monovalent heterocyclic group;
(v) R c- O-, R c- C (= O)-, R c - OC (= O)-, or R c- C (= O) -O- (R c is a hydrogen atom or a monovalent hydrocarbon group,.);
(vi) NR d R e -, NR d R e -C (= O) -, NR d R e -C (= O) -O-, or R d- C (= O) -NR e- (R d and Re represent the same or different hydrogen atoms or monovalent hydrocarbon groups); or (Vii) Selected from the group consisting of a nitro group, a sulfate group, a sulfonic acid group, a cyano group, or a carboxyl group .

　Y and Y'are the same or different from Y in the formula (B-1) described in [24], and
　Z and Z'are the same as or different from Z in the formula (B-1). It's the same. ], The antibody of [59] or a salt thereof.
[61] An antibody or a salt thereof having an affinity substance for the antibody and a cleaving moiety, wherein A is an antibody having the following
　formula (II):
ALBR'-T (II)
[In the formula,
　A is an antibody. for a affinity substance,
　L is a divalent cleavable linker is a group containing a cleavable moiety,
　B is (a) a divalent group containing a bio-orthogonal functional groups or (b) a biological, It is a divalent group free of orthogonal functional groups
　, where R'is the part produced by the reaction between the antibody and a reactive group specific for the side chain of the lysine residue, where
　T is. , An antibody. ] An
　antibody or a salt thereof, which is represented and the affinity substance for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1.
[0014]
(Method for producing an antibody having an affinity for an antibody and an antibody having a cleaving moiety or a salt thereof)
[62] A method for producing an antibody having an affinity for an antibody and an antibody having a cleaving moiety or a salt thereof, according to the following
　formula ( I):
ALBR (I)
[In the formula,
　A is an affinity for an antibody,
　L is a divalent group containing a cleaving moiety, and
　B is a cleaving linker. , (A) a divalent group containing a bioorthogonal functional group, or (b) a divalent group not containing a bioorthogonal functional group, where
　R is a reactive group for the antibody. ], A compound having an affinity substance for an antibody, a cleaving moiety and a reactive group or a salt thereof is reacted with the antibody to cause the following
　formula (II):
ALBR'-T ( II)
[In the formula,
　A, L, and B are the same as those in the above formula (I),
　R'is a portion produced by the reaction between the antibody and the reactive group, and
　T is It is an antibody. ], The affinity substance for the antibody, and
　the affinity substance for the antibody , which comprises producing an antibody having a cleavable moiety or a salt thereof, are the above formulas 1-1 to 1-9, and the formula 2 A method, the peptide comprising any of the amino acid sequences of -1.
[63] The method of [62], wherein the reactive group is a reactive group specific for the side chain of the lysine residue.
[0015]
　Thirdly, the present invention provides a complex having an affinity substance for an antibody, a cleaving moiety, a functional substance and an antibody or a salt thereof, and a method for producing the same.
(Complex having an affinity substance, a cleaving moiety, a functional substance and an antibody or a salt thereof)
[64] A complex having an affinity substance, a cleaving moiety, a functional substance and an antibody or a salt thereof. The following
　formula (III):
ALB'(-F) -R'-T (III)
[In the formula,
　A is an affinity substance for an antibody, and
　L contains a cleaving moiety. It is a cleaving linker that is a
　divalent group, B'is a trivalent group containing a moiety produced by a reaction between a functional substance and a bioorthogonal functional group, and
　F is a functional substance. Yes,
　R'is the part produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ], And the
　affinity substance for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1. The complex or a salt thereof.
[65] The complex of [64] or a salt thereof, wherein the antibody is a monoclonal antibody.
[66] A complex of [64] or [65] or a salt thereof, wherein the antibody is an IgG antibody.
[67] The complex of any of [64] to [66] or a salt thereof, wherein the antibody is of human origin.
[68] Any of [64] to [67], wherein the antibody contains any one Fc region protein selected from the group consisting of the following (A) to (C) and has an antigen-binding ability. Complex or salt thereof:
(A) Fc region protein containing the amino acid sequence of SEQ ID NO : 1;
(B) In the amino acid sequence of SEQ ID NO: 1, one or several amino acid residues are inserted, added, deleted or substituted. Fc region protein containing the amino acid sequence obtained; or
(C) Fc region protein containing an amino acid sequence showing 90% or more identity with the amino acid sequence of SEQ ID NO: 1.
[69] The antibody contains one or more specific amino acid residues in a target region consisting of 1 to 50 consecutive amino acid residues, and the specific amino acid residue in a non-target region other than the target region. The
　structural unit containing 5 or more groups and represented by ALB'(-F) -R'is 30% or more with respect to one or more specific amino acid residues contained in the target region. The complex of any of [64] to [68] or a salt thereof, which is bound by the regioselectivity of.
[70] The specific amino acid residue is a number with respect to the N-terminal side and the C-terminal side, respectively, centered on the specific amino acid existing at the specific position (where a is an arbitrary integer of 1 to 10). In the region up to the distant position of the amino acid residue of (), the complex of [69] does not contain an amino acid residue of the same type as the specific amino acid residue other than the specific amino acid residue existing at the specific position. Body or its salt.
[71] The antibody is an antibody containing a plurality of heavy chains.
　As a result of T having a structural unit represented by ALB'(-F) -R'in a plurality of corresponding target regions in a plurality of heavy chains, the antibody is ALB. The complex of any of [64] to [70] or a salt thereof, which has a plurality of structural units represented by'(-F) -R'.
[72] When the functional substance has a functional group that easily reacts with a bioorthogonal functional group, or is derivatized so as to have a functional group that easily reacts with a bioorthogonal functional group, the living body. Functional groups that easily react with orthogonal functional groups are azide residues, aldehyde residues, thiol residues, alkin residues, alken residues, halogen residues, tetrazine residues, nitron residues, hydroxylamine residues, and nitriles. Residues, hydrazine residues, ketone residues, boronic acid residues, cyanobenzothiazole residues, allyl residues, phosphine residues, maleimide residues, disulfide residues, thioester residues, α-halocarbonyl residues, isonitriles A complex of any of [64] to [71] or a salt thereof, which is a group selected from the group consisting of residues, sidonone residues, and selenium residues.
[73] When the functional substance has a functional group that easily reacts with a bioorthogonal functional group, or is derivatized so as to have a functional group that easily reacts with a bioorthogonal functional group, the living body. Functional groups that easily react with orthogonal functional groups are as follows:
[Derivatization 12]

[In the formula,
　R 1f , single or multiple R 1 g and single or multiple R 1 h.Is an atom or group or an electron-withdrawing group selected from the group consisting of the above (i) to (vii), which is the same or different, and: is a bond to a functional substance. A complex of any of [64] to [71] or a salt thereof, which is a group selected from the group consisting of the groups represented by).
[74] The moiety produced by the reaction between the antibody and the reactive group is one of a lysine residue, a tyrosine residue, or a tryptophan residue and a lysine residue, a tyrosine residue, or a tryptophan residue. The complex of any of [64] to [73] or a salt thereof, which is a portion formed by a reaction with a reactive group specific for a side chain.
[75] The portion produced by the reaction between the antibody and the reactive group is the portion produced by the reaction between the lysine residue and the reactive group specific to the side chain of the lysine residue. , [64] to [74], or a salt thereof.
[76] The part generated by the reaction is as follows:
[Chemical 13]

[Here, ● (black circle) indicates a bond to the T side part, and ○ (white circle) indicates a bond to the B side part. .. ], Which corresponds to any one chemical structure selected from the group consisting of [64] to [75], or a salt thereof.
[77] The complex of any of [64] to [76] or a salt thereof, wherein the functional substance is a drug or a labeling substance.
[78] The complex of any of [64] to [77] or a salt thereof, wherein the functional substance is a low molecular weight compound.
[79] A complex of [77] or [78] or a salt thereof, wherein the drug is an anticancer agent.
[80] The complex of any of [64] to [79] or a salt thereof, wherein the main chain connecting A and R'has 4 to 20 atoms.
[81] The complex of any of [64] to [80] or a salt thereof, wherein the main chain connecting A and R does not contain a ring structure.
[82] The complex of any of [64] to [81] or a salt thereof, wherein the partial structure represented by LB does not contain a peptide moiety.
[83] The compound represented by the formula (III) is the following formula (III'):
AB2'(-F2) -L'-B1'(-F1) -R'-T (III')
[ In the formula,
　A, R'and T are the same as those in formula (II) above,
　L'is a cleavable linker which is a divalent group containing a cleavable moiety,
　and B1'and B2' are cleavable linkers. , Identical or different, trivalent groups containing moieties produced by the reaction between the functional group and the bioorthogonal functional group,
　F1 and F2 are the same or different functional substances,
　B1 '(−F1) and B2'(−F2) may have a symmetrical structure centered on L'. ], The complex of any of [64] to [82] or a salt thereof.
[84] The compound represented by the formula (III') is the following (III''):
[Chemical

formula 14] [In the formula,
　A, R'and T are those of the formula (III) described in [62]. Is the same as
　C is the cuttable part,
　p and p'are any integers
　from 0 to 10 which are the same or different, and q and q'are any integers from 0 to 10 which are the same or different. ,
　X and X'are the same or different, carbon atom, nitrogen atom, or single bond (where R 1b does not exist if X is a nitrogen atom and R if X'is a nitrogen atom. 1b ' If no .X is a single bond, R 1a and R 1b is absent, X' when is a single bond, R 1a ' and R 1b' is does not
　exist), R 1a , R 1b , R 1a'and R 1b'are the same or different and are selected from the group consisting of the above (i) to (vii), and
　Y and Y'are the same or different and are the same or different from the above formula (B-1). It is a residue obtained by removing one hydrogen atom from Y of the above, and
　Z and Z'are the same as or different from Z of the above formula (B-1).
　F and F'are the same or different and are functional substances. ], The complex of [83] or a salt thereof.
[85] A complex having an affinity substance, a functional substance and an antibody, or a salt thereof, wherein the following
　formula (III):
ALB'(-F) -R'-T (III)
[in the formula ,
　A is an affinity substance for an antibody,
　L is a cleavable linker which is a divalent group containing a cleavable moiety,
　and B'is a reaction between a functional substance and a bioorthogonal functional group.
　Is a trivalent group containing a moiety produced by, F is a functional substance, and
　R'is a reaction between the antibody and a reactive group specific for the side chain of the lysine residue. The part to be produced, where
　T is an antibody. ], And the
　affinity substance for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1. The complex or a salt thereof.
[0016]
(Method for producing a complex having an affinity substance, a cleaving moiety, a functional substance and an antibody or a salt thereof)
[86] A complex having an affinity substance, a cleaving moiety, a functional substance and an antibody for an antibody or A method for producing the salt, which is the following
　formula (II):
ALBR'-T (II)
[In the formula,
　A is an affinity substance for an antibody, and
　L is a cleaving moiety. Cleaving linker which is a divalent group containing,
　B is (a) a divalent group containing a bioorthogonal functional group
　, and R'is produced by the reaction between the antibody and the reactive group. A portion, where
　T is an antibody. ], The antibody having an affinity for the antibody and the antibody having a cleavable moiety or a salt thereof is reacted with a functional substance, and the following
　formula (III):
ALB'(-F) -R '-T (III)
[In the formula,
　A, L, R'and T are the same as those in the above formula (II),
　and B'is the reaction between the functional substance and the bioorthogonal functional group. It is a trivalent group containing a portion produced by
　F, which is a functional substance. ], Which comprises producing a complex having an affinity substance, a cleavable moiety, a functional substance and an antibody, or a salt thereof, which is represented by.
　The method, wherein the affinity substance for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1.
[87] The method of [86], wherein the reactive group is a reactive group specific for the side chain of the lysine residue.
[88] affinity substance for the antibody, cleavage moiety, a complex or a salt thereof with a functional material and antibodies,
(A) the following formula (I):
A-L-B-R (I )
[In the formula,
　A is an affinity substance for an antibody,
　L is a cleavable linker which is a divalent group containing a cleavable moiety, and
　B is (a) a bioorthogonal functional group 2 It is a valence group, and
　R is a reactive group for the antibody. ] 　Is reacted with the antibody, and the following
　formula (II):
ALBR'-T (II)
[In the formula,
A, L, and B are the above formulas. Same as in (I),
　R'is the part produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ] To produce an antibody or a salt thereof having an affinity substance for an antibody and a cleaving moiety, and
(B) an antibody or a salt thereof having an affinity substance for the antibody and a cleaving moiety. React with functional substances,
　The following formula (III):
ALB'(-F) -R'-T (III)
[In the formula,
　A and L are the same as those in the formula (I),
　and R'and T are , The same as that of the above formula (II),
　B'is a trivalent group containing a moiety produced by the reaction between the functional substance and the bioorthogonal functional group, and
　F is the functional substance. Is. ], Which comprises producing a complex having an affinity substance for an antibody, a cleaving moiety, a functional substance and an antibody, or a salt thereof, and
　the affinity substance for the antibody is the above-mentioned formulas 1-1 to 1. -9, and a peptide comprising the amino acid sequence of any of formula 2-1.
[0017]
　Fourth, the present invention provides a method for producing an antibody having a bioorthogonal functional group.
[89] A method for producing an antibody having a bioorthogonal functional group or a salt thereof, wherein 　A is an affinity for the antibody according to the following
　formula (II):
ALBR'-T (II)
[In the formula,
A is an affinity for the antibody. A sex substance,
　L is a cleavable linker which is a divalent group containing a cleavable moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group or (b) bioorthogonality. It is a divalent group free of functional groups, where R'is the moiety
　produced by the reaction between the antibody and the reactive group and
　T is the antibody. ], The cleaving portion of the antibody or salt thereof having the affinity substance for the antibody and the cleaving moiety is cleaved, and the following
　formula (IV):
L1-BR'-T (IV)
[formula] Among them,
　B, R'and T are the same as those of the above formula (II), and
　L1 is a monovalent group containing (i') bioorthogonal functional group or (ii') bioorthogonal functional group. It is a monovalent group containing no group. ], The
　substance having an affinity for the antibody , which comprises producing an antibody having a bioorthogonal functional group or a salt thereof, is any one of the above formulas 1-1 to 1-9 and formula 2-1. A method, which is a peptide comprising the amino acid sequence of.
[90] L is a divalent group containing a cleavable moiety capable of producing a bioorthogonal functional group on the reactive group side by (i) cleavage, or a bioorthogonal by (ii) cleavage. The method of [89], wherein the cleaving linker is a divalent group containing a cleaving moiety that does not have the ability to generate a sex functional group on the reactive group side.
[91] L is the cleaving linker of (i),
　L1 is a monovalent group containing (i') a bioorthogonal functional group, and
　B is 2 of (a) or (b). The method of [89], which is the basis of the value.
[92] L is the cleaving linker of (i),
　L1 is a monovalent group containing (i') a bioorthogonal functional group, and
　B is a divalent group of (b). The method of [90] or [91].
[93] L is the cleaving linker of (ii),
　L1 is a monovalent group containing (i') a bioorthogonal functional group, and
　B is a divalent group of (a). , [90] method.
[94] The method of any of [89] to [93], wherein the reactive group is a reactive group specific for the side chain of the lysine residue.
[95]
A method for producing an antibody having a bioorthogonal functional group or a salt thereof, wherein (A) the following formula (I):
ALBR (I)
[In the formula,
　A is an affinity for an antibody. It is a sex substance
　L is a cleavable linker which is a divalent group containing a cleaving moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group or (b) not containing a bioorthogonal functional group. It is a divalent group, and
　R is a reactive group for the antibody. ] 　Is reacted with the antibody, and the following
　formula (II):
ALBR'-T (II)
[In the formula,
A, L, and B are the above formulas. Same as in (I),
　R'is the part produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ] To produce an antibody or a salt thereof having an affinity substance for an antibody and a cleaving moiety, and
(B) cleaving the antibody or a salt thereof having an affinity substance for the antibody and a cleaving moiety. By cutting off the sex part, the following
　formula (IV):
L1-BR'-T (IV)
[In the formula,
　B, R'and T are the same as those in the above formula (II), and
　L1 is , (I') a monovalent group containing a bioorthogonal functional group, or (ii') a monovalent group containing no bioorthogonal functional group. ], The
　substance having an affinity for the antibody , which comprises producing an antibody having a bioorthogonal functional group or a salt thereof, is any one of the above formulas 1-1 to 1-9 and formula 2-1. A method, which is a peptide comprising the amino acid sequence of.
[0018]
　Fifth, the present invention provides a method for producing an antibody having a functional substance or a salt thereof.
[96] A method for producing an antibody or a salt thereof having a functional substance, wherein the following
　formula (III):
ALB'(-F) -R'-T (III)
[In the formula,
　A is An affinity substance for an antibody,
　L is a cleavable linker which is a divalent group containing a cleavable moiety,
　and B'is a moiety produced by a reaction between a functional substance and a bioorthogonal functional group.
　Is a trivalent group containing, F is a functional substance,
　R'is a moiety produced by the reaction between an antibody and a reactive group, and
　T is an antibody. ], The cleaving part of the complex having an affinity substance, a cleaving part, a functional substance and an antibody for the antibody or a salt thereof is cleaved, and the following
　formula (V):
F- (L1-B) '-R'-T (V)
[In the formula,
　L1 is a monovalent group containing (i') bioorthogonal functional group or (ii') monovalent group not containing bioorthogonal functional group. Yes,
　B is (a) a divalent group containing a bioorthogonal functional group, or (b) a divalent group not containing a bioorthogonal functional group.
　The structural unit represented by (L1-B)'includes a moiety produced by a reaction between a functional substance and one or both of the bioorthogonal functional groups in (i') and (a). It is a divalent structural unit,
　F is a functional substance,
　R'is a part produced by the reaction between an antibody and a reactive group, and
　T is an antibody. ], Which comprises producing an antibody having a functional substance or a salt thereof, and
　the affinity substance for the antibody is an amino acid according to any one of the above formulas 1-1 to 1-9 and formula 2-1. A method that is a sequence-containing peptide.
[97] The following
　formula (V1):
L1-B'(-F) -is cleaved by cleaving the cleaving part of the complex having an affinity substance, a cleaving part, a functional substance and an antibody or a salt thereof. R'-T (V1)
[In the formula,
　L1 is (i') a monovalent group containing a bioorthogonal functional group, or (ii') a monovalent group containing no bioorthogonal functional group.
　B', F, R'and T are the same as those in the above formula (III). ], The method of [96], which comprises producing an antibody or a salt thereof having a functional substance.
[98] The antibody having a bioorthogonal functional group or a salt thereof is reacted with one or two kinds of functional substances, and the following
　formula (V2):
FL1'-BR'-T (V2) )
[In the formula,
　B, R'and T are the same as those of formula (IV) above, and
　L1'is due to the reaction between the functional material and (i') a monovalent group containing a bioorthogonal functional group. It is a divalent group containing the part to be produced, and
　F is a functional substance. ], Or the following
　formula (V3):
Fa-L1'-B'(-Fb) -R'-T (V3)
[In the formula, R'and
　T are the same as those in the formula (IV).
　L1'is the same as that of the above formula (V2)
　,
　B'is a trivalent group containing a moiety produced by the reaction between the functional substance and the bioorthogonal functional group, Fa and Fb. Are the same or different functional substances, respectively. ], The method of [96], which comprises producing an antibody or a salt thereof having a functional substance.
[99] The method according to any one of [96] to [98], wherein the reactive group is a reactive group specific for the side chain of the lysine residue.
[100]
A method for producing an antibody or a salt thereof having a functional substance, wherein (A) the following formula (II):
ALBR'-T (II)
[In the formula,
　A is for an antibody. An affinity substance,
　L is a cleaving linker that is a divalent group containing a cleaving moiety.
　B is (a) a divalent group containing a bioorthogonal functional group,
　R'is a portion produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ], The antibody having an affinity for the antibody and the antibody having a cleavable moiety or a salt thereof is reacted with a functional substance, and the following
　formula (III):
ALB'(-F) -R '-T (III)
[In the formula,
　A, L, R'and T are the same as those in the above formula (II),
　and B'is the reaction between the functional substance and the bioorthogonal functional group. It is a trivalent group containing a portion produced by
　F, which is a functional substance. ] To produce a complex having an affinity substance, a cleaving moiety, a functional substance and an antibody, or a salt thereof, and
(B) an affinity substance, a cleaving moiety, and functionality for the antibody. By cleaving the cleavable portion of the complex having the substance and antibody or a salt thereof, the following
　formula (V1):
L1-B'(-F) -R'-T (V1)
[In the formula,
　L1 is (i). ') A monovalent group containing a bioorthogonal functional group, or (ii') a monovalent group containing no bioorthogonal functional group, and
　B', F, R'and T are the above formula (III). It is the same as that of. ], Including producing an antibody having a functional substance or a salt thereof, and
　The method, wherein the affinity substance for the antibody is a peptide containing the amino acid sequence of any of the above formulas 1-1 to 1-9 and formula 2-1.
[101]
A method for producing an antibody or a salt thereof having a functional substance, wherein (A) the following formula (I):
ALBR (I)
[In the formula,
　A is an affinity substance for an antibody. in it,
　L is a divalent cleavable linker is a group containing a cleavable moiety,
　B is a divalent group comprising (a) a bio-orthogonal functional group,
　R is the response to the antibody It is a sex group. ], A compound having an affinity substance for an antibody, a cleaving moiety and a reactive group or a salt thereof is reacted with the antibody to cause the following
　formula (II):
ALBR'-T ( II)
[In the formula,
　A, L, and B are the same as those in the above formula (I),
　R'is a portion produced by the reaction between the antibody and the reactive group, and
　T is the antibody. Is. ] To produce an antibody having an affinity substance for an antibody and a cleavable moiety or a salt thereof,
(B) an antibody having an affinity substance for the antibody and a cleavable moiety or a salt thereof. Reacting with a sex substance, the following
　formula (III):
ALB'(-F) -R'-T (III)
[In the formula,
　A and L are the same as those of the formula (I),
　R'and T are the same as those of the formula (II),
　and B'is bioorthogonal with the functional substance. It is a trivalent group containing a moiety formed by a reaction with a functional group, and
　F is a functional substance. ] To produce a complex having an affinity substance, a cleaving moiety, a functional substance and an antibody, or a salt thereof, and
(C) an affinity substance, a cleaving moiety, and functionality for the antibody. By cleaving the cleavable portion of the complex having the substance and antibody or a salt thereof, the following
　formula (V1):
L1-B'(-F) -R'-T (V1)
[In the formula,
　L1 is (i). ') A monovalent group containing a bioorthogonal functional group, or (ii') a monovalent group containing no bioorthogonal functional group, and
　B', F, R'and T are the above formula (III). It is the same as that of. ], Which comprises producing an antibody having a functional substance or a salt thereof, and
　the affinity substance for the antibody is an amino acid according to any one of the above formulas 1-1 to 1-9 and formula 2-1. A method that is a sequence-containing peptide.
[102]
A method for producing an antibody or a salt thereof having a functional substance, wherein (A) the following formula (II):
ALBR'-T (II)
[in the formula,
　A is
　a cleaving linker which is an affinity substance for an antibody, L is a divalent group containing a cleaving moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group, or (B) A divalent group containing no bioorthogonal functional group, R'is
　a portion produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ], The cleaving portion of the antibody or salt thereof having the affinity substance for the antibody and the cleaving moiety is cleaved, and the following
　formula (IV):
L1-BR'-T (IV)
[formula] Among them,
　B, R'and T are the same as those of the above formula (II), and
　L1 is a monovalent group containing (i') bioorthogonal functional group or (ii') bioorthogonal functional group. It is a monovalent group containing no group. ] To produce an antibody having a bioorthogonal functional group or a salt thereof, and
(B) one or more functional substances of the antibody having a bioorthogonal functional group or a salt thereof. In response to the following
　formula (V2)
FL1'-BR'-T (V2)
[In the formula,
　B, R'and T are the same as those in the above formula (IV), and
　L1' Is a divalent group containing a moiety produced by the reaction between a functional group and (i') a monovalent group containing a bioorthogonal functional group.
　F is a functional substance. ], Or the following
　formula (V3):
Fa-L1'-B'(-Fb) -R'-T (V3)
[In the formula, R'and
　T are the same as those in the formula (IV).
　L1'is the same as that of the above formula (V2)
　,
　B'is a trivalent group containing a moiety formed by the reaction between the functional substance and the bioorthogonal functional group, Fa and Fb. Are the same or different functional substances, respectively. ], Which comprises producing an antibody having a functional substance or a salt thereof, and
　the affinity substance for the antibody is an amino acid according to any one of the above formulas 1-1 to 1-9 and formula 2-1. A method that is a sequence-containing peptide.
[103]
A method for producing an antibody or a salt thereof having a functional substance, wherein (A) the following formula (I):
ALBR (I)
[In the formula,
　A is an affinity substance for the antibody. in it,
　L is a divalent cleavable linker is a group containing a cleavable moiety,
　B is a divalent radical or (b) bio-orthogonal functional groups, comprising (a) a bio-orthogonal functional group
　Is a divalent group that does not contain, and R is a reactive group for the antibody. ] Is reacted with an antibody to the compound represented by the following
　formula (II):
ALBR'-T (II)
[In the formula,
　A, L, and B are the same as those in the above formula (I), and
　R'is the reaction between the antibody and the reactive group.
　Is the part produced by T, which is an antibody. ] To produce an antibody having an affinity substance for an antibody and a cleaving moiety or a salt thereof,
(B) a substance having an affinity for the antibody and a cleavage property of an antibody having a cleaving moiety or a salt thereof. By cutting the portion, the following
　formula (IV):
L1-BR'-T (IV)
[In the formula,
　B, R'and T are the same as those in the above formula (II), and
　L1 is (I') a monovalent group containing a bioorthogonal functional group, or (ii') a monovalent group containing no bioorthogonal functional group. ] To produce an antibody having a bioorthogonal functional group or a salt thereof, and
(C) one or more functional substances of the antibody having a bioorthogonal functional group or a salt thereof. In response to the following
　formula (V2)
FL1'-BR'-T (V2)
[In the formula,
　B, R'and T are the same as those in the above formula (IV), and
　L1' Is a divalent group containing a moiety produced by the reaction between a functional group and (i') a monovalent group containing a bioorthogonal functional group.
　F is a functional substance. ], Or the following
　formula (V3):
Fa-L1'-B'(-Fb) -R'-T (V3)
[In the formula, R'and
　T are the same as those in the formula (IV).
　L1'is the same as that of the above formula (V2)
　,
　B'is a trivalent group containing a moiety formed by the reaction between the functional substance and the bioorthogonal functional group, Fa and Fb. Are the same or different functional substances, respectively. ], Which comprises producing an antibody having a functional substance or a salt thereof, and
　the affinity substance for the antibody is an amino acid according to any one of the above formulas 1-1 to 1-9 and formula 2-1. A method that is a sequence-containing peptide.
Effect of the invention
[0019]
　(I) Compounds of the invention or salts thereof having affinity substances, cleaving moieties and reactive groups for antibodies are useful, for example, for regioselective modification of antibodies.
　(I) The compound of the present invention or a salt thereof, (II) an affinity substance for an antibody, and an antibody of the present invention or a salt thereof having a cleaving moiety (positionally), (III) an affinity substance for an antibody, cleavage. Complexes of the invention or salts thereof having (positionally selective) sex moieties, functional substances and antibodies are, for example, antibodies or salts thereof having bioorthogonal functional groups (positionally selective), and functional substances. It is useful as an intermediate for the preparation of antibodies or salts thereof (positionally selective). Antibodies or salts thereof having (regioselectively) functional substances are useful, for example, as pharmaceuticals or reagents (eg, diagnostic agents, research reagents). An antibody or salt thereof having a bioorthogonal functional group (regioselectively) is useful as an intermediate for the preparation of an antibody or salt thereof having a functional substance (regioselectively). Therefore, (I) the compound of the present invention or a salt thereof, (II) the antibody of the present invention or a salt thereof, and (III) the complex of the present invention or a salt thereof are useful as synthetic intermediates for, for example, pharmaceuticals or reagents. Is.
A brief description of the drawing
[0020]
[Fig. 1-1] Fig. 1-1 shows the position selection of an antibody by the compound of the present invention [compound having an affinity substance, a cleaving moiety and a reactive group for the antibody: ALBR (I)]. It is a schematic diagram (the 1) which shows the concept of a target modification. First, the compound of the present invention associates with the antibody (T) via the affinity substance (A) for the antibody. Next, the compound of the present invention contains a specific amino acid residue in a target region present in the vicinity of the association site between the affinity substance and the antibody via the reactive group (R) (activated ester in the figure). Reacts with the side chain of the lysine residue (in the figure, the side chain of the lysine residue) to selectively have a structural unit containing a conjugate of the compound of the present invention and an antibody [an affinity substance for the amino acid and a cleavage moiety Antibody: ALBR'-T (II)] is produced.
[Fig. 1-2] Fig. 1-2 is a schematic diagram (No. 2) showing the concept of regioselective modification of an antibody by the compound of the present invention. Cleavage of the cleaving moiety in the linker (L) produces an antibody position-specifically modified with a bioorthogonal functional group.
[Fig. 1-3] Fig. 1-3 is a schematic diagram (No. 3) showing the concept of regioselective modification of an antibody by a compound of the present invention. The reaction of a bioorthogonal functional group with a functional substance (eg, drug) produces an antibody that is position-specifically modified with the functional substance.
FIG. 2 is a diagram showing the relevance of each invention of the present invention (notation is omitted for salts). In the reaction (1), a compound having an affinity substance, a cleaving moiety and a reactive group for the antibody is reacted with the antibody to produce an antibody having an affinity substance for the antibody and a cleaving moiety. In the reaction (2), the antibody having an affinity substance for the antibody and the cleaving moiety is reacted with the functional substance to generate a complex having the affinity substance for the antibody, the cleaving moiety, the functional substance and the antibody. .. In the reaction (3), the cleaving part of the complex having the affinity substance, the cleaving part, the functional substance and the antibody to the antibody is cleaved to produce the antibody having the functional substance (at this time, the affinity substance). Is produced as a by-product). In the reaction (4), the cleaving portion of the antibody having an affinity substance for the antibody and the cleaving moiety is cleaved to produce an antibody having a bioorthogonal functional group (at this time, the portion containing the affinity substance is used. Produced as a by-product). In the reaction (5), an antibody having a bioorthogonal functional group is reacted with a functional substance to produce an antibody having the functional substance. Reactions (2) and (5) can be carried out in the same manner. Reactions (3) and (5) can also be carried out in the same manner.
FIG. 3 is a diagram showing the consensus amino acid sequences (SEQ ID NO: 1) of the Fc region in the heavy chain of trastuzumab and the IgG1 Fc region.
[Fig. 4] Fig. 4 shows (1) the amino acid sequence of the heavy chain of trusszumab (SEQ ID NO: 2), (2) the amino acid sequence of the IgG1 Fc region in which the sugar chain was cleaved with PNGase (SEQ ID NO: 3), and (3). It is a figure which shows the amino acid sequence (SEQ ID NO: 4) of the light chain of trastuzumab.
FIG. 5 is a diagram showing the results (detection wavelength: UV280 nm) of HIC (Hydrophobic Interaction Chromatography) -UPLC analysis of specific modification of trastuzumab (Example 1). The AU on the vertical axis indicates the absorbance (same in the following drawings).
FIG. 6 is a peptide consisting of 33 amino acid residues containing a site modified to a lysine residue by trypsin digestion of trussumab (a thiol-introduced product (+145.09Da) subjected to carbamidomylization with iodoacetamide), THTCPPCPAPELLLGGPSVFLFPPKPKDTLMISR (FIG. 6). It is a figure which shows the MS spectrum (measured value: m / z 1269.30717, theoretical value: 1269.30773, trivalent) of the peptide fragment of SEQ ID NO: 6) (Example 2).
FIG. 7-1 shows the modification of the lysine residue at position 248 of the human IgG heavy chain in EU numbering, m / z 603.29 corresponding to divalent y9 (theoretical value: 603. It is a figure which shows the CID spectrum of the product ion of 30) (Example 2).
[Fig. 7-2] Fig. 7-2 shows a peptide fragment containing a thiol-introduced product (+145.019 Da) modified to a lysine residue (Carbamidemethylized with iodoacetamide) from a tryptic digest of trastuzumab. It is a figure which shows the result of the search using (Thermo Fisher Scientific Co., Ltd.) (Example 2). The horizontal axis shows the identified lysine residue, and the vertical axis shows Peptide Spectrum Matches (PSMs). Regarding the residue number of the lysine residue, the number in the sequence for the heavy chain VH domain and the light chain (that is, the N-terminal amino acid is the first; the same applies hereinafter), and the heavy chain CH1, CH2, and CH3 domains. The above is described using EU numbering.
[Fig. 8] Fig. 8 shows the site of modification of trastuzumab to a lysine residue by digestion with Glu-C protease (thiol-introduced product (+145.09 Da) subjected to Carbamidometry with iodoacetamide. )) Is a diagram showing the MS spectrum (measured value: m / z 619.67299, theoretical value: 619.67112, trivalent) of the peptide fragment of LLGGPSVFLFPPKPKD (SEQ ID NO: 7), which is a peptide consisting of 16 amino acid residues. (Example 2).
[Fig. 9-1] Fig. 9-1 shows the modification of the lysine residue at the 246th or 248th position of the human IgG heavy chain in EU numbering, and m / z corresponding to monovalent y5. It is a figure which shows the CID spectrum of the product ion of 729.49 (theoretical value: 729.36) (Example 2).
[Fig. 9-2] Fig. 9-2 shows a peptide fragment containing a thiol-introduced product (+145.09Da) modified to a lysine residue (Carbamidemethylized with iodoacetamide) in the Glu-C protease digested product of trussumab. Is a diagram showing the results of searching using Proteome Discoverer (Thermofisher Scientific Co., Ltd.) (Example 2). The horizontal axis shows the identified lysine residue, and the vertical axis shows Peptide Spectram Matches (PSMs). Regarding the residue numbers of lysine residues, the numbers on the heavy chain VH domain and the light chain are shown in the sequence, and the numbers on the heavy chain CH1, CH2, and CH3 domains are shown using EU numbering
[Fig . ]. 10] FIG. 10 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of the specific modification of trusszumab (Example 3).
[Fig. 11] FIG. 11 is the HIC of the specific modification of trussumab. -The result of the UPLC analysis (detection wavelength: UV280 nm) is shown (Example 4).
[FIG. 12] FIG. 12 shows the result of the HIC-UPLC analysis (detection wavelength: UV280 nm) of the specific modification of trussumab. FIG
. 13 shows an amino acid containing a site of modification of trussumab to a lysine residue by trypsin digestion (a thiol-introduced product (+145.09 Da) that has undergone Carbamidemethylization with iodoacetamide). It is a figure which shows the MS spectrum (measured value: m / z 952.23170, theoretical value: 952.22900, tetravalent) of the peptide fragment of THTCPPCPAPELLLGGPSVFLFPPKPKDTLMISR (SEQ ID NO: 6) which is a peptide consisting of 33 residues (Example 6). ..
FIG. 14-1 shows modification of the lysine residue at position 246 or 248 of the human IgG heavy chain in EU numbering, m / z 1166.88 (theoretical value) corresponding to divalent y20. It is a figure which shows the CID spectrum of the product ion of: 1166.61) (Example 6).
[Fig. 14-2] Fig. 14-2 shows a peptide fragment containing a thiol-introduced product (+145.09 Da) modified to a lysine residue (Carbamidemethylized with iodoacetamide) from a tryptic digest of trastuzumab. It is a figure which shows the result of the search using (Thermo Fisher Scientific Co., Ltd.) (Example 6). The horizontal axis shows the identified lysine residue, and the vertical axis shows Peptide Spectrum Matches (PSMs). Regarding the residue numbers of lysine residues, the numbers on the heavy chain VH domain and the light chain are shown in the sequence, and the numbers on the heavy chain CH1, CH2, and CH3 domains are shown using EU numbering
[Fig. 15]. FIG. 15 is a diagram showing the results of HIC-UPLC analysis of specific modification of trastuzumab (detection wavelength: UV 280 nm) (Example 7).
[FIG. 16] FIG. 16 is a HIC-UPLC analysis of specific modification of trastuzumab. (Detection wavelength: UV280 nm).
[FIG. 17] FIG. 17 is a diagram showing the result of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab. (Example 9).
[FIG. 18] FIG. 18 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 10).
FIG. 19 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 11).
FIG. 20 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 12).
FIG. 21 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 13).
FIG. 22 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 14).
FIG. 23 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 15).
FIG. 24 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 16).
FIG. 25 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 17).
FIG. 26 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 18).
FIG. 27 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 19).
FIG. 28 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 20).
FIG. 29 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 21).
FIG. 30 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 22).
FIG. 31 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 23).
FIG. 32 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 24).
FIG. 33 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 25).
FIG. 34 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 26).
FIG. 35 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 27).
FIG. 36 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 28).
FIG. 37 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 29).
FIG. 38 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 30).
FIG. 39 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 31).
FIG. 40 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 32).
FIG. 41 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 33).
FIG. 42 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 34).
FIG. 43 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 35).
FIG. 44 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 36).
FIG. 45 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 37).
FIG. 46 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 38).
FIG. 47 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 39).
FIG. 48 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 40).
FIG. 49 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 41).
FIG. 50 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 42).
FIG. 51 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 43).
FIG. 52 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 44).
FIG. 53 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 45).
FIG. 54 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 46).
FIG. 55 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 47).
FIG. 56 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 48).
FIG. 57 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 49).
FIG. 58 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 50).
FIG. 59 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 51).
FIG. 60 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 52).
FIG. 61 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 53).
FIG. 62 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 54).
FIG. 63 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 55).
FIG. 64 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 56).
FIG. 65 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 57).
FIG. 66 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 58).
FIG. 67 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 59).
FIG. 68 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 60).
FIG. 69 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 61).
FIG. 70 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 62).
FIG. 71 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 63).
FIG. 72 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 64).
FIG. 73 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 65).
FIG. 74 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 66).
FIG. 75 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 67).
FIG. 76 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 68).
FIG. 77 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 69).
FIG. 78 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 70).
FIG. 79 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 71).
FIG. 80 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 72).
FIG. 81 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 73).
FIG. 82 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 74).
FIG. 83 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 75).
FIG. 84 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 76).
FIG. 85 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 77).
FIG. 86 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 78).
FIG. 87 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 79).
FIG. 88 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under reducing conditions for a specific modifier of trastuzumab (Example 80).
FIG. 89 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 80).
FIG. 90 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under reduction conditions of a specific modified trastuzumab thiol-introduced product (Example 80).
FIG. 91 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 80).
FIG. 92 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under reducing conditions of a trastuzumab fluorescent label (Example 80).
FIG. 93 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under ADC mimic reduction conditions (Example 80).
FIG. 94 is a diagram showing a summary of the results of Example 80.
FIG. 95 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 81).
FIG. 96 is a peptide consisting of 33 amino acid residues containing a site modified to a lysine residue by trypsin digestion of trussumab (a thiol-introduced product (+145.09Da) subjected to carbamidomylization with iodoacetamide), THTCPPCPAPELLLGGPSVFLFPPKPKDTLMISR (FIG. 96). It is a figure which shows the MS spectrum (measured value: m / z 1269.30359, theoretical value: 1269.3273, trivalent) of the peptide fragment of SEQ ID NO: 6) (Example 81).
FIG. 97-1 shows modification of the lysine residue at position 246 or 248 of the human IgG heavy chain in EU numbering, m / z 1205.86 (theoretical value) corresponding to monovalent y9. It is a figure which shows the CID spectrum of the product ion of: 1205.60) (Example 81).
[Fig. 97-2] Fig. 97-2 shows a peptide fragment containing a thiol-introduced product (+145.019 Da) modified to a lysine residue (Carbamidemethylized with iodoacetamide) from a tryptic digest of trastuzumab. It is a figure which shows the result of the search using (Thermo Fisher Scientific Co., Ltd.) (Example 81). The horizontal axis shows the identified lysine residue, and the vertical axis shows Peptide Spectrum Matches (PSMs). Regarding the residue numbers of lysine residues, the numbers on the heavy chain VH domain and the light chain are shown in the sequence, and the numbers on the heavy chain CH1, CH2, and CH3 domains are shown using EU numbering
[Fig. 98]. FIG. 98 is a diagram showing the results of analysis of the antibody-nucleic acid conjugate by SDS-PAGE (Example 82).
[Fig. 99] Fig. 99 shows the weight of the antibody-nucleic acid conjugate by ESI-TOFMS analysis under the reducing conditions of the trastuzumab azide-introduced product. It is a figure which shows the confirmation of the chain selectivity (Example 83).
[FIG. 100] FIG. 100 is a figure which shows the confirmation of the heavy chain selectivity by ESI-TOFMS analysis under the reduction condition of ADC mimic (Example). 83).
[Fig. 101] Fig. 101 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under reduction conditions of a specific modifier of trastuzumab (Example 84)
[Fig. 102] Fig. 102. Is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of trastuzumab (Example 84).
FIG. 103 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under the reducing conditions of a specific modified trastuzumab azide-introduced product (Example 84).
FIG. 104 is a diagram showing the results of HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of a trastuzumab azide-introduced body (Example 84).
FIG. 105 shows a peptide of THTCPPCPAPELLLGGPSVFPFPPKPDTLMISS (SEQ ID NO: 6), a peptide consisting of 33 amino acid residues containing a site modified to a lysine residue by trypsin digestion of trussumab (azidocarboxylic acid introducer (+240.086)). It is a figure which shows the MS spectrum (measured value: m / z 1300.99241, theoretical value: 1300.99173, trivalent) of a fragment (Example 84).
FIG. 106-1 shows modification of the lysine residue at position 246 or 248 in EU numbering, m / z 1622.92 (theoretical value: 1622.87) corresponding to monovalent y12. It is a figure which shows the CID spectrum of the product ion of (Example 84).
[Fig. 106-2] Fig. 106-2 shows modifications to lysine residues (azido-introduced product (+255.097 Da), amine-introduced product (+229.106), azidocarboxylic acid-introduced product) for trypsin digested product of trussumab. (+240.086), an amine carboxylic acid introducer (+214.095)) is a diagram showing the results of searching for a peptide fragment using Protein Discoverer (Thermo Fisher Scientific Co., Ltd.) (Example 84). The horizontal axis shows the identified lysine residues, and the vertical axis shows the Peptide Spectrum Matches (PSMs). Regarding the residue number of the lysine residue, the numbers in the sequence are used for the heavy chain VH domain and the light chain, and EU numbering is used for the heavy chain CH1, CH2, and CH3 domains.
FIG. 107 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under ADC mimic reduction conditions (Example 85).
FIG. 108 is a diagram showing HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modifications of trastuzumab azide-introduced material (Example 85).
FIG. 109 is a peptide consisting of 33 amino acid residues containing a site modified to a lysine residue by trypsin digestion of trussumab (DBCO-Acid carboxylic acid introducer (+559.207)), THTCPPCPAPELLLGGPSVFLFPPKPKDTLMISS (SEQ ID NO: 6). It is a figure which shows the MS spectrum (measured value: m / z 1055.77631, theoretical value: 1055.77600, tetravalent) of the peptide fragment of (Example 85).
FIG. 110-1 shows modification of the lysine residue at position 246 or 248 in EU numbering, m / z 971.71 corresponding to divalent y12 (theoretical value: 971.50). It is a figure which shows the CID spectrum of the product ion of (Example 85).
[Fig. 110-2] Fig. 110-2 shows modifications to lysine residues (DBCO-Azide introducer (+574.218), DBCO-Acid carboxylic acid introducer (+559.207)) for tryptic digest of trussumab. , Azide-introduced body (+255.097), Azidecarboxylic acid-introduced body (+240.086)), and the results of searching for a peptide fragment using Protein Discoverer (Thermofisher Scientific Co., Ltd.) are shown (Example). 85). The horizontal axis shows the identified lysine residues, and the vertical axis shows the Peptide Spectrum Matches (PSMs). Regarding the residue number of the lysine residue, the numbers in the sequence are used for the heavy chain VH domain and the light chain, and EU numbering is used for the heavy chain CH1, CH2, and CH3 domains.
FIG. 111 is a diagram showing specific modification of the anti-TNF-α IgG1 antibody adalimumab and analysis by ESI-TOFMS (Example 86).
FIG. 112 is a diagram showing HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of adalimumab (Example 86).
FIG. 113 is a diagram showing linker cleavage of the adalimumab peptide complex (Example 86).
FIG. 114 is a diagram showing HIC-UPLC analysis (detection wavelength: UV 280 nm) of an adalimumab thiol-introduced product (Example 86).
FIG. 115 is a diagram showing a fluorescent label on an adalimumab thiol-introduced product (Example 86).
FIG. 116 is a diagram showing specific modification of the anti-RANKL IgG2 antibody denosumab and analysis by ESI-TOFMS (Example 87).
FIG. 117 is a diagram showing HIC-UPLC analysis (detection wavelength: UV280 nm) of specific modification of denosumab (Example 87).
FIG. 118 is a diagram showing linker cleavage of the denosumab peptide complex (Example 87).
FIG. 119 is a diagram showing HIC-UPLC analysis (detection wavelength: UV 280 nm) of a denosumab thiol-introduced body (Example 87).
FIG. 120 is a diagram showing a fluorescent label on a denosumab thiol-introduced body (Example 87).
FIG. 121 shows specific modification of the anti-IL-4 / 13 receptor IgG4 antibody dupilumab and analysis by ESI-TOFMS (Example 88).
FIG. 122 is a diagram showing HIC-UPLC analysis (detection wavelength: UV 280 nm) of specific modification of dupilumab (Example 88).
FIG. 123 is a diagram showing linker cleavage of the dupilumab peptide complex (Example 88).
FIG. 124 is a diagram showing HIC-UPLC analysis (detection wavelength: UV 280 nm) of a dupilumab thiol-introduced product (Example 88).
FIG. 125 is a diagram showing confirmation of heavy chain selectivity by ESI-TOFMS analysis under reducing conditions of a dupilumab fluorescent label (Example 88).
Mode for carrying out the invention
[0021]
1. 1. Compounds containing affinity substances, cleavage moieties and reactive groups for antibodies or salts thereof
1-1. Overview The
　present invention provides a compound or a salt thereof, which is represented by the formula (I) and contains an affinity substance for an antibody, a cleaving moiety and a reactive group.
ALBR (I)
[In the formula,
　A is an affinity substance for an antibody,
　L is a divalent group containing a cleaving moiety, and
　B is a (a). ) A divalent group containing a bioorthogonal functional group, or (b) a divalent group not containing a bioorthogonal functional group, where
　R is a reactive group for the antibody. ]
[0022]
　In formula (I) and other formulas presented in connection with the present invention,-(hyphen) indicates that the two units on either side of it are covalently bonded. Therefore, in formula (I), A is covalently bonded to L, L is covalently bonded to A and B, B is covalently bonded to L and R, and R is covalently bonded to B. Covalently bonded.
[0023]
1-2. Affinity substance for antibody (A) In
　formula (I), A is an affinity substance for antibody. Affinity substances are substances that have the ability to bind to antibodies by non-covalent bonds.
[0024]
　The affinity substances used in the present invention target antibodies. The antibody may be a protein modified with a biomolecule (eg, sugar) (eg, glycoprotein) or a protein unmodified with a biomolecule. As the antibody, any antibody against any component such as a biological component, a virus-derived component, and a component found in the environment can be used, but an antibody against a biological component or a virus-derived component is preferable. Examples of biological components include components derived from animals such as mammals and birds (eg, chickens), insects, microorganisms, plants, fungi, and fish (eg, proteins). Preferably, the biological component is a mammalian-derived component. Mammals include, for example, primates (eg humans, monkeys, chimpanzees), rodents (eg mice, rats, guinea pigs, hamsters, rabbits), pets (eg dogs, cats), domestic animals (eg dogs, cats) Examples include cows, pigs, goats) and working animals (eg horses, sheep). The biological component is more preferably a primate or rodent-derived component (eg, protein), and even more preferably a human-derived component (eg, protein) from the viewpoint of clinical application of the present invention. is there. Examples of the virus-derived component include components derived from influenza virus (eg, tri-influenza virus, swine flu virus), AIDS virus, Ebola virus, and phage virus (eg, protein).
[0025]
　The antibody is also a polyclonal antibody or a monoclonal antibody, preferably a monoclonal antibody. As the monoclonal antibody, for example, it is modified to have a sugar chain binding consensus sequence such as a chimeric antibody, a humanized antibody, a human antibody, and an antibody to which a predetermined sugar chain is added (eg, an N-type sugar chain binding consensus sequence). Antibodies), bispecific antibodies, scFv antibodies, Fab antibodies, F (ab') 2 antibodies, VHH antibodies, Fc region proteins, Fc fusion proteins. The antibody may also be a divalent antibody (eg, IgG, IgD, IgE) or a tetravalent or higher valent antibody (eg, IgA antibody, IgM antibody).
[0026]
　The antibody that is the target of the affinity substance may also be composed of any amino acid residue, but is preferably composed of 20 natural L-α-amino acid residues that normally constitute a protein. Examples of such amino acid residues include L-alanine (A), L-asparagine (N), L-cysteine ​​(C), L-glutamine (Q), L-isoleucine (I), and L-leucine ( L), L-methionine (M), L-phenylalanine (F), L-proline (P), L-serine (S), L-threonine (T), L-tryptophane (W), L-tyrosine (Y) ), L-valine (V), L-aspartic acid (D), L-glutamic acid (E), L-arginine (R), L-histidine (H), or L-lysine (K), and glycine (G). ) (Hereinafter, the notation of L is omitted). The antibody may be composed of, for example, 100 or more, preferably 120 or more, more preferably 150 or more, even more preferably 180 or more, and particularly preferably 200 or more amino acid residues. The number of antibodies may also be, for example, 1000 or less, preferably 900 or less, more preferably 800 or less, even more preferably 700 or less, and particularly preferably 600 or less. More specifically, the antibody has, for example, 100 to 1000 amino acids, preferably 120 to 900, more preferably 150 to 800, even more preferably 180 to 700 or more, and particularly preferably 200 to 600 amino acid residues. It may be composed of groups. When the antibody is an antibody (eg, a monoclonal antibody as described above), the above number of amino acid residues may correspond to the amino acid residues of the heavy chain of the antibody.
[0027]
　Antibodies that are the target of affinity substances also have a side chain or terminal (N-terminal and / or C-terminal), preferably a specific amino acid residue having a side chain, at one position to which a reactive group can react, as described below. Alternatively, it is a protein contained at a plurality of positions (preferably a plurality of positions). Examples of such specific amino acid residues include 14 types of amino acid residues as described later, preferably a group consisting of lysine residues, tyrosine residues, tryptophan residues, and cysteine ​​residues. Amino acid residues of choice. Considering that the compound of the present invention can regioselectively modify an antibody, an antibody containing such a specific amino acid residue at a plurality of positions is preferable. The plurality of positions are not particularly limited as long as they are two or more positions, but for example, three or more positions, preferably five or more positions, more preferably ten or more positions, and even more preferably 20 or more positions, particularly preferable. May be in more than 30 positions. The plurality of positions may also be, for example, 200 or less positions, preferably 180 or less positions, more preferably 150 or less positions, even more preferably 120 or less positions, and particularly preferably 100 or less positions. More specifically, the plurality of positions are, for example, 3 to 200 positions, preferably 5 to 180 positions, more preferably 10 to 150 positions, even more preferably 20 to 120 positions, and particularly preferably 30 to. It may be in the 100 position. Even in an antibody containing such a specific amino acid residue at a plurality of positions, the compound of the present invention can regioselectively modify a specific amino acid residue existing at a specific position. For example, the number of lysine residues in human IgG1 is generally said to be about 70 to 90, although it depends on the amino acid composition in the variable region. In the present invention, we have succeeded in regioselectively modifying a lysine residue existing at a specific position of such human IgG1.
[0028]
　More specifically, the present invention modifies amino acid residues present at specific positions in an antibody while preserving antibody function (ie, maintaining native folding without denaturing the antibody). From the viewpoint, regioselective modification of amino acid residues exposed on the surface of the antibody is preferable. For example, in human IgG such as human IgG1, the exposed lysine residue and the exposed tyrosine residue are present at the following positions (according to EU numbering; http: //www.imgt.org/IMGTScientificChart / Numbering / Hu_IGHGnber.html. reference). In the present application, for convenience of explanation, EU numbering is used for the heavy chain CH1, CH2, and CH3 domains, and the numbers in the sequence (that is, N) are used for the heavy chain VH domain and the light chain. The terminal amino acid may be expressed using the first).
(1) Exposed lysine residue
CH2 domain (246th, 248th, 274th, 288th, 290th, 317th, 320th, 322nd, 338th)
CH3 domain (360th, 414th, 439th)
( 2) Exposed tyrosine residue
CH2 domain (positions 278, 296, 300)
CH3 domain (position 436)
Therefore, when a human IgG such as human IgG1 is modified with a lysine residue or a tyrosine residue, at the above position. Modification is preferred.
[0029]
　Preferably, when human IgG such as human IgG1 is modified with a lysine residue or a tyrosine residue, it exists at the following positions with high surface exposure among the positions (1) and (2) above. Lysine residues or tyrosine residues may be modified.
(1') Exposed lysine residue
CH2 domain (246th, 248th, 274th, 288th, 290th, 317th, 320th, 322nd)
CH3 domain (360th, 414th, 439th)
(2' ) ) Exposed tyrosine residue
CH2 domain (positions 278, 296, 300)
CH3 domain (position 436)
Therefore, when a human IgG such as human IgG1 is modified with a lysine residue or a tyrosine residue, the modification at the above position. Is more preferable.
[0030]
　More preferably, when human IgG such as human IgG1 is modified with a lysine residue, a predetermined position (eg,) in the CH2 domain that can be efficiently modified in the present invention among the positions in (1) above. The lysine residues present at positions 246, 248, 288, 290, and 317) may be modified.
[0031]
　In certain embodiments, an antibody that is the target of an affinity substance is specified in a target region consisting of 1 to 50 consecutive amino acid residues when it contains a particular amino acid residue at multiple positions as described above. It may contain one or more amino acid residues of the above, and may contain five or more specific amino acid residues in a non-target region other than the target region. The target region is preferably 1 to 30, more preferably 1 to 20, even more preferably 1 to 10, 1 to 5, or 1 to 3 (ie, 1, 2, or 3). ) May consist of amino acid residues. Particularly preferably, the target region may be a region consisting of a specific amino acid residue present at a specific position. Such a specific position varies depending on the type of target protein and affinity substance, and is, for example, a specific position in a specific region (eg, CH1, CH2, CH3) in the constant region of an antibody. It may also be preferably the position in CH2 of the antibody. More specifically, the target region may be the following residues according to
Eunumbering in human IgG Fc: (1) Lys248 residues (hereinafter, also simply referred to as "Lys248" herein, human IgG CH2. The 16th residue of the human IgG CH2 region (SEQ ID NO: 1), which corresponds to the 18th residue of the region (SEQ ID NO: 1) or the Lys246 residue (hereinafter, also simply referred to as "Lys246" in the present specification).
equivalent) to; (2) Lys288 residues (hereinafter, simply herein denoted as "Lys288", human IgG CH2 region (SEQ ID NO: 1) corresponding to 58 th residues) or Lys290 residues ( Hereinafter, it is also simply referred to as “Lys290” in the present specification, and corresponds to the 60th residue of the human IgG CH2 region (SEQ ID NO: 1));
(3) Lys317 residue (hereinafter, also simply referred to as “Lys317” in the present specification, corresponds to the 87th residue of the human IgG CH2 region (SEQ ID NO: 1)).
[0032]
　According to the present invention, a specific amino acid residue in the target region can be modified highly regioselectively. Such regioselectivity is, for example, 30% or more, preferably 40% or more, more preferably 50% or more, even more preferably 60% or more, particularly preferably 70% or more, 80% or more, 90% or more, 95. It may be% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% or more.
[0033]
　The target region also has a specific amino acid residue present at a specific position with respect to the N-terminal side and the C-terminal side, respectively, centered on the specific position (where a is an arbitrary 1 to 10). In the region up to the remote position of the amino acid residue (which is an integer of), the amino acid residue of the same type as the specific amino acid residue is not contained other than the specific amino acid residue existing at the specific position. May be good. a is preferably an integer of 1 to 5, more preferably an integer of 1 to 3, even more preferably 1 or 2, and particularly preferably 1.
[0034]
　In a preferred embodiment, the antibody is a monoclonal antibody. Examples of antibody isotypes such as monoclonal antibodies include IgG (eg, IgG1, IgG2, IgG3, IgG4), IgM, IgA, IgD, IgE, and IgY. The monoclonal antibody is a full-length antibody or an antibody fragment (eg, F (ab') 2 , Fab', Fab, Fv, single-chain antibody), but a full-length antibody is preferable.
[0035]
　Antibodies are antibodies against any antigen. For example, such an antigen may be a component found in an organism or virus as described above. Such antigens also include, for example, proteins [oligopeptides, polypeptides. Proteins modified with biomolecules such as sugars (eg, glycoproteins)], sugar chains, nucleic acids, low molecular weight compounds can be mentioned.
[0036]
　Preferably, the antibody may be an antibody that uses a protein as an antigen. Examples of proteins include cell membrane receptors, cell membrane proteins other than cell membrane receptors (eg, extracellular substrate proteins), ligands, and soluble receptors.
[0037]
　More specifically, the protein that is the antigen of the antibody may be a disease target protein. Examples of disease target proteins include the following.
[0038]
(1) Cancer area
　PD-L1, GD2, PDGFRα (Plotlet-derived growth factor receptor), CD22, HER2, phosphatidylserine (PS), EpCAM, fibronectin, PD-1, VEGFR-2, CD33, HGF, gpNMB, CD27, DEC-205, Folic acid receptor, CD37, CD19, Trop2, CEACAM5, S1P, HER3, IGF-1R, DLL4, TNT-1 / B, CPAAs, PSMA, CD20, CD105 (endoglin), ICAM-1, CD30, CD16A, CD38, MUC1, EGFR, KIR2DL1,2, NKG2A, tenascin-C, IGF (Insulin-like growth factor), CTLA-4, meshothelin, CD138, c-Met, Ang2, VEGF-A, CD79b, ENPD3, folic acid receptor , TEM-1, GM2, Gripican 3, macrophage inhibitory factor, CD74, Notch1, Notch2, Notch3, CD37, TLR-2, CD3, CSF-1R, FGFR2b, HLA-DR, GM-CSF, EphA3, B7- CD123, gpA33, Frizzled7 receptor, DLL4, VEGF, RSPO, LIV-1, SLITRK6, Nectin-4, CD70, CD40, CD19, SEMA4D (CD100), CD25, MET, Tissue Factor, IL-8, EGFR, cMet, KIR3DL2, Bst1 (CD157), P-cadherin, CEA, GITR, TAM (tumor associated macrophage), CEA, DLL4, Ang2, CD73, FGFR2, CXCR4, LAG-3, GITR, FucosylG CSF-1R, FGFR3, OX40, BCMA, ErbB3, CD137 (4-1BB), PTK7, EFNA4, FAP, DR5, CEA, Ly6E, CA6, CEACAM5, LAMP1, thisset factor,EPHA2, DR5, B7-H3, FGFR4, FGFR2, α2-PI, A33, GDF15, CAIX, CD166, ROR1, GITR, BCMA, TBA, LAG-3, EphA2, TIM-3, CD-200, EGFRvIII, CD16A, CD32B, PIGF, Axl, MICA / B, Thomasen-Friendenrich, CD39, CD37, CD73, CLEC12A, Lgr3, transferrin receptor, TGFβ, IL-17, 5T4, RTK, Immunosuppressor Protein, NaPi2b, Lewis blood group A34, Lysil-Oxidase, DLK-1, TROP-2, α9 integrin, TAG-72 (CA72-4), CD70
[0039]
(2) Autoimmune diseases / inflammatory diseases
　IL-17, IL-6R, IL-17R, INF-α, IL-5R, IL-13, IL-23, IL-6, ActRIIB, β7-Integrin, IL- 4αR, HAS, Eotaxin-1, CD3, CD19, TNF-α, IL-15, CD3ε, Fibronectin, IL-1β, IL-1α, IL-17, TSLP (Thematic Structural Lymphopoietin), LAMP (Alpha4 Integrin) , IL-23, GM-CSFR, TSLP, CD28, CD40, TLR-3, BAFF-R, MAdCAM, IL-31R, IL-33, CD74, CD32B, CD79B, IgE (immunoglobulin E), IL-17A, IL-17F, C5, FcRn, CD28, TLR4, MCAM, B7RP1, CXCR1, Litens, IL-21, Cadherin-11, CX3CL1, CCL20, IL-36R, IL-10R, CD86, TNF-α, IL-7R , Kv1.3, α9 integrin, LIFEHT
[0040]
(3) Neurological diseases
　CGRP, CD20, β-amyloid, β-amyloid protofibrin, Calcitonin Gene-Related Peptide Receptor, LINGO (Ig Domain Contining1), α-synuclein, extracellular tau, CD52, insulin receptor Tau, CD52, insulin receptor t , SOD1, TauC3, JC virus
[0041]
(4) Infectious diseases
　Clostridium difficile toxin B, cytomegalovirus, RS virus, LPS, S. d. Aureus Alpha-toxin, M2e protein, Psl, PcrV, S. a. Aureus toxin, Influenza A, Alginate, Staphylococcus aureus, PD-L1, Influenza B, Acinetobacter, F-protein, Env, CD3, Pathogenic Escherichia coli, Klebsiella, Streptococcus pneumoniae
[0042]
(5) Hereditary / rare diseases
　Amyloid AL, SEMA4D (CD100), insulin receptor, ANGPTL3, IL4, IL13, FGF23, adrenocorticotropic hormone, transthyretin, huntingtin
[0043]
(6) Eye diseases
　Factor D, IGF-1R, PGDFR, Ang2, VEGF-A, CD-105 (Endoglin), IGF-1R, β-amyloid
[0044]
(7) Bone and Orthopedic Surgery Area
Sclerostin, Myostatin, Dickkopf-1, GDF8, RNAKL, HAS, SIGLEC-15
[0045]
(8) Blood diseases
　vWF, Factor IXa, Factor X, IFNγ, C5, BMP-6, Ferroportin, TFPI
[0046]
(9) Other diseases
　BAFF (B cell activating factor), IL-1β, PCSK9, NGF, CD45, TLR-2, GLP-1, TNFR1, C5, CD40, LPA, prolactin receptor, VEGFR-1, CB1, Endoglin, PTH1R, CXCL1, CXCL8, IL-1β, AT2-R, IAPP
[0047]
　In a more preferred embodiment, the affinity for the antibody is an affinity for the monoclonal antibody. The isotype of the monoclonal antibody is similar to that described above for the antibody, but IgG (eg, IgG1, IgG2, IgG3, IgG4) is preferred. Preferably, the monoclonal antibody is a full-length monoclonal antibody.
[0048]
　In an even more preferred embodiment, the affinity for the antibody is a chimeric antibody, a humanized antibody, or an affinity for a human antibody (eg, IgG such as IgG1, IgG2, IgG3, IgG4), which is a full-length monoclonal antibody.
[0049]
　In a particularly preferred embodiment, the amino acid-affinitive substance is an amino acid-affinitive substance containing any one Fc region protein selected from the group consisting of the following (A) to (C) and having an antigen-binding ability. :
(A) Fc region protein containing the amino acid sequence of SEQ ID NO : 1;
(B) Contains an amino acid sequence in which one or several amino acid residues are inserted, added, deleted or substituted in the amino acid sequence of SEQ ID NO: 1. Fc region protein; or
(C) an Fc region protein containing an amino acid sequence showing 90% or more identity with the amino acid sequence of SEQ ID NO: 1.
[0050]
　The amino acid sequence of SEQ ID NO: 1 is an Fc region protein. Such Fc region proteins are known to have secretory capacity. Therefore, the Fc region proteins (A) to (C) can have a secretory capacity. In addition, an antibody containing such an Fc region protein can have an antigen-binding ability. The amino acid residue at position 18 in SEQ ID NO: 1 is an arbitrary amino acid residue, but is preferably a neutral amino acid residue, and more preferably an amino acid residue having a non-polar side chain as described later. Even more preferably, it is leucine, isoleucine or alanine, and particularly preferably leucine or alanine. The amino acid residue at position 19 in SEQ ID NO: 1 is an arbitrary amino acid residue, preferably a neutral amino acid residue or an acidic amino acid residue, and more preferably an amino acid residue or an acidic amino acid having a non-polar side chain. It is an amino acid residue, and even more preferably leucine or glutamic acid. The amino acid residue at position 21 in SEQ ID NO: 1 is an arbitrary amino acid residue, but is preferably a neutral amino acid residue, more preferably an amino acid residue having a non-polar side chain, and even more preferably. Glycine or alanine. The amino acid residue at position 140 in SEQ ID NO: 1 is an arbitrary amino acid residue, but is preferably an acidic amino acid residue, and more preferably glutamic acid or aspartic acid. The amino acid residue at position 142 in SEQ ID NO: 1 is an arbitrary amino acid residue, but is preferably a neutral amino acid residue, more preferably an amino acid residue having a non-polar side chain, and even more preferably. It is methionine, leucine or isoleucine, and particularly preferably methionine or leucine. The amino acid residue at position 177 in SEQ ID NO: 1 is an arbitrary amino acid residue, but is preferably a neutral amino acid residue, and more preferably an amino acid residue having an uncharged polar side chain as described later. Amino acid residues with non-polar side chains, even more preferably threonine, alanine or glycine

The scope of the claims
[Claim 1]
　A compound having an affinity substance, a cleaving moiety and a reactive group for an antibody or a salt thereof,
　wherein the compound is the following formula (I):
ALBR (I)
[In the formula,
　A is An affinity substance for an antibody,
　L is a cleavable linker which is a divalent group containing a cleavable moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group, or (b). It is a divalent group containing no bioorthogonal functional group, and
　R is a reactive group for the antibody. ], And the
　affinity substance for the antibody is the
formula 1-1: 1 (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-I-I-W- C- (X 0-3 ) b (SEQ ID NO
: 58) Equation 1-2: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-IVW-C- ( X 0-3 ) b (SEQ ID NO: 59)
Equation 1-3: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-VVW-C- (X 0-3 ) b (SEQ ID NO: 60)
Equation 1- 4: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-A-V-WC- (X 0-3 ) b (SEQ ID NO
: 61) Equation 1-5 :( X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-L-W-C- (X 0-3 ) b (SEQ ID NO
: 62) Equation 1-6: (X 0-) 3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-IWC- (X 0-3 ) b(SEQ ID NO
: 63) Equation 1-7: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-LVFC- (X 0-3 ) b (SEQ ID NO : 63) 64)
formula 1-8: (X 0-3 ) a -C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-Q-V-W-C-(X 0-3 ) b (SEQ ID NO: 65)
expression 1-9: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-EVW-C- (X 0-3 ) b (SEQ ID NO: 66)
[In the formula,
　(X 0-3 ) a is none, or 1 to 3 consecutive identical or different arbitrary amino acid residues (other than lysine and cysteine ​​residues).
　(X 0-3 ) b is none, or 1 to 3 consecutive identical or different amino acid residues (other than lysine and cysteine ​​residues), and
　Xaa1 is an alanine residue, glycine residue. , Leucine residue, proline residue, arginine residue, valine residue, asparagine residue, glutamate residue, or phenylalanine residue, and
　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue. a
　group, Xaa3 is histidine residue, a phenylalanine residue, a tyrosine residue, a tryptophan residue, an arginine residue or glycine
　residue,, Xaa4 is a lysine
　residue, Xaa @ 5 is a glycine residue, Serine residue, asparagine residue, glutamine residue, aspartic acid residue, glutamic acid residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue, alanine residue, valine A residue, isoleucine residue, or arginine residue,
　Xaa6 is a glutamine residue, glutamic acid residue, asparagine residue, aspartic acid residue, proline residue, glycine residue, arginine residue, phenylalanine residue, Or a histidine residue. ], Or
Equation 2-1: (X 0-3 ') a-C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-LVWC- (X 0-3 ') b (SEQ ID NO: 67)
[In the formula,
　(X 0- 3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively, and are
　Xaa1', Xaa2', Xaa3', Xaa4', Xaa5. ', Xaa6' are the same as the Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different amino acid residues, (X 0-3 ') b.Except when 1 to 3 consecutive identical or different arbitrary amino acid residues, Xaa3'is a histidine residue, and Xaa5'is a glycine residue. ], A compound or a salt thereof, which is a peptide containing any of the amino acid sequences of.
[Claim 2]
　(X 0-3 ) a is none, arginine residue-glycine residue-aspartic acid residue, or aspartic acid residue, and
　(X 0-3 ) b is none, threonine residue- Tyrosine residue-histidine residue, or threonine residue
　,
　Xaa1 is an alanine residue, Xaa2 is a tyrosine residue, tryptophan residue, or histidine residue, and
　Xaa6 is a glutamine residue, glutamic acid. The compound according to claim 1, or a salt thereof, which is a residue, an aspartic residue, or an aspartic acid residue.
[Claim 3]
　(X 0-3 ) a is a glycine residue-asparagin residue, a glycine residue, a glycine residue-glycine residue, or a glycine residue-glycine residue-glycine residue, and
　(X 0-3 ). b is threonine residue-tyrosine residue, glycine residue, glycine residue-glycine residue, or glycine residue-glycine residue-glycine residue, and
　Xaa1 is glycine residue, leucine residue, proline. Residues, arginine residues, valine residues, asparagine residues, glutamate residues, or phenylalanine residues,
　Xaa2 is a phenylalanine residue,
　Xaa6 is a proline residue, glycine residue, arginine residue, The compound according to claim 1, or a salt thereof, which is a phenylalanine residue or a histidine residue.
[Claim 4]
　Affinity substance to the antibody is a peptide comprising an amino acid sequence selected from the group consisting of: The compound of any one of claims 1 to 3 or a salt
thereof: (1) RGNCAYHKGQIIWCTYH (SEQ ID NO:
5); ( 2) RGNCAYHKGQIVWCTYH (SEQ ID NO:
8); (3) RGNCAYHKGQVVWCTYH (SEQ ID NO:
9); (4) RGNCAYHKGQAVWCTYH (SEQ ID NO:
10); (5) RGNCAYHKGQLLWCTYH (SEQ ID NO:
11); (6) RGNCAYHKGQLIWCTYH (SEQ ID NO:
12); ( 7) DCAYHKGQIVWCT (SEQ ID NO:
13); (8) DCAYHKGQVVWCT (SEQ ID NO:
14); (9) DCAYHKGQAVWCT (SEQ ID NO:
15); (10) RGNCAYHKSQIIWCTYH (SEQ ID NO:
16); (11) RGNCAYHKNQIIWCTYH (SEQ ID NO:
17); ( 12) RGNCAYHKDQIIWCTYH (SEQ ID NO: 18);
(13) RGNCAYHKQQIIWCTYH (SEQ ID NO: 19);
(14) RGNCAYHKEQIIWCTYH (SEQ ID NO: 20);
(15) RGNCAYHKFQIIWCTYH (SEQ ID NO: 21);
(16) RGNCAYHKYQIIWCTYH (SEQ ID NO: 22);
(17) RGNCAYHKWQIIWCTYH (SEQ ID NO: 23);
(18) RGNCAYHKHQIIWCTYH (SEQ ID NO: 24);
(19) RGNCAYHKTQIIWCTYH (SEQ ID NO: 25);
(20)
(21) CAYHKLQIVWC (SEQ ID NO: 27);
(22) CAYHKLQLIWC (SEQ ID NO: 28);
(23) CAYHKSQIVWC (SEQ ID NO: 29);
(24) RGNCAYHKGQLVFCTYH (SEQ ID NO: 30);
(25) RGNCAYHKGQV
(26) RGNCAYHKGQEVWCTYH (SEQ ID NO: 32);
(27) CAYHKGQLVWC (SEQ ID NO: 33);
(28) RGNCAYHKAQLVWCTYH (SEQ ID NO: 34);
(29) RGNCAYHKVQLVWCTYH (SEQ ID NO: 35);
(30)
(31) RGNCAYHKIQLVWCTYH (SEQ ID NO: 37);
(32) RGNCAYHKSQLVWCTYH (SEQ ID NO: 38);
(33) RGNCAYHKTQLVWCTYH (SEQ ID NO: 39);
(34) RGNCAYHKNQLVWCTYH (SEQ ID NO: 40);
(35) RGNCAYHKDQLVWCTYH (SEQ ID NO: 41);
(36) RGNCAYHKQQLVWCTYH (SEQ ID NO: 42
);
(38) RGNCAYHKFQLVWCTYH (SEQ ID NO: 44);
(39) RGNCAYHKRQLVWCTYH (SEQ ID NO: 45);
(40) RGNCAYHKHQLVWCTYH (SEQ ID NO: 46);
(41) RGNCAYHKWQLVWCTYH (SEQ ID NO: 47
);
(43) RGNCAYFKGQLVWCTYH (SEQ ID NO:
49); (44) RGNCAYYKGQLVWCTYH (SEQ ID NO:
50); (45) RGNCAYWKGQLVWCTYH (SEQ ID NO:
51); (46) RGNCAYRKGQLVWCTYH (SEQ ID NO:
52); (47) RGNCAYGKGQLVWCTYH (SEQ ID NO: 53);
(48) DCAYHKGQLVWC (SEQ ID NO: 54);
(49) NCAYHKGQLVWC (SEQ ID NO: 55);
(50) CAYHKGQLVWCT (SEQ ID NO: 56);
(51) CAYHKSQLVWC (SEQ ID NO: 57);
(52) RGNCAWHKGQIIWCTYH (SEQ ID NO: 68);
(53) RGNCAFHKGQIIWCTYH (SEQ ID NO: 69);
(54) RGNCAHKH
(55) RGNCGYHKGQIIWCTYH (SEQ ID NO: 71);
(56) RGNCLYHKGQIIWCTYH (SEQ ID NO: 72);
(57) RGNCPYHKGQIIWCTYH (SEQ ID NO: 73);
(58) RGNCRYHKGQIIWCTYH (SEQ ID NO: 74
);
(60) RGNCNYHKGQIIWCTYH (SEQ ID NO: 76);
(61) RGNCEYHKGQIIWCTYH (SEQ ID NO: 77);
(62) RGNCFYHKGQIIWCTYH (SEQ ID NO: 78);
(63) RGNCAYHKGEIIWCTYH (SEQ ID NO: 79);
(64)
(65) RGNCAYHKGPIIWCTYH (SEQ ID NO: 81);
(66) RGNCAYHKGGIIWCTYH (SEQ ID NO: 82);
(67) RGNCAYHKGDIIWCTYH (SEQ ID NO: 83);
(68) RGNCAYHKGRIIWCTYH (SEQ ID NO: 84);
(69) RGNCAYHKGFIIWCTYH (SEQ ID NO: 85);
(70) RGNCAYHKGHIIWCTYH (SEQ ID NO: 86);
(71)
(72) NCAYHKGQIIWCT (SEQ ID NO: 88);
(73) GNCAYHKGQIIWCTY (SEQ ID NO: 89);
(74) GCAYHKGQIIWCG (SEQ ID NO: 90);
(75) GGCAYHKGQIIWCGG (SEQ ID NO: 91); and
(76) ..
[Claim 5]
　A cleavage linker in which L is a divalent group containing a cleavable moiety capable of producing a bioorthogonal functional group on the reactive group side by (i) cleavage, or a bioorthogonal functional group by (ii) cleavage. The compound according to any one of claims 1 to 4, or a salt thereof, which is a cleaving linker which is a divalent group containing a cleaving moiety which does not have the ability to form a reactive group side.
[Claim 6]
　The cleavable moiety is treated with one or more substances selected from the group consisting of (a) acidic substances, basic substances, reducing agents, oxidizing agents, and enzymes, and (b) physicochemically selected from the group consisting of light. The compound according to any one of claims 1 to 5, which is a moiety that can be cleaved by treatment with a stimulus or (c) a moiety that can be cleaved by leaving it to stand when using a cleaving linker containing a self-degradable cleaving moiety. Or its salt.
[Claim 7]
　The cleaving moiety contains a disulfide residue, an acetal residue, a ketal residue, an ester residue, a carbamoyl residue, an alkoxyalkyl residue, an imine residue, a tertiary alkyloxycarbamate residue, a silane residue, and a hydrazone-containing residue. Group, phosphoramidate residue, acetal residue, trityl residue, azo residue, vicinaldiol residue, selenium residue, aromatic ring-containing residue with electron-withdrawing group, coumarin-containing residue, sulfone-containing The compound according to any one of claims 1 to 6, or a salt thereof, selected from the group consisting of residues, unsaturated bond-containing chain residues, and glycosyl residues.
[Claim 8]
　Any of claims 5 to 7, wherein the cleaving moiety of (i) is selected from the group consisting of disulfide residues, ester residues, acetal residues, ketal residues, imine residues, and vicinaldiol residues. The compound according to one item or a salt thereof.
[Claim 9]
　The cleaving portion of (ii) is an ester residue, a carbamoyl residue, an alkoxyalkyl residue, an imine residue, a tertiary alkyloxycarbamate residue, a silane residue, a hydrazone-containing residue, or a phosphoramidate residue. , Aconityl residue, Trityl residue, Azo residue, Vicinaldiol residue, Selenium residue, Aromatic ring-containing residue with electron-withdrawing group, Kumarin-containing residue, Sulfone-containing residue, Unsaturated bond-containing chain The compound according to any one of claims 5 to 7, or a salt thereof, which is selected from the group consisting of residues and glycosyl residues.
[Claim 10]
　Bioorthogonal functional groups are azide residues, aldehyde residues, thiol residues, alkin residues, alkene residues, halogen residues, tetrazine residues, nitron residues, hydroxylamine residues, nitrile residues, hydrazine residues Group, ketone residue, boronic acid residue, cyanobenzothiazole residue, allyl residue, phosphine residue, maleimide residue, disulfide residue, thioester residue, α-halocarbonyl residue, isonitrile residue, sidonone residue The compound according to any one of claims 1 to 9, or a salt thereof, selected from the group consisting of a group and a selenium residue.
[Claim 11]
　The compound according to any one of claims 1 to 10, wherein the reactive group is a reactive group specific for any one side chain of a lysine residue, a tyrosine residue, or a tryptophan residue. That salt.
[Claim 12]
　The compound according to claim 11, or a salt thereof, wherein the reactive group is a reactive group specific for the side chain of the lysine residue.
[Claim 13]
　The compound according to any one of claims 1 to 12 or a salt thereof, which has one or more characteristics selected from the following:
(a) The number of atoms in the main chain connecting A and R is 4 to 20;
(B) The main chain connecting A and R does not contain a ring structure;
(c) the partial structure represented by LB does not contain a peptide moiety.
[Claim 14]
　A position-selective modification reagent for an antibody, which has the following
　formula (I):
ALBR (I)
[In the formula,
　A is an affinity substance for the antibody, and
　L contains a cleaving moiety. It is a cleaving linker which is a divalent group, and
　B is (a) a divalent group containing a bioorthogonal functional group or (b) a divalent group containing no bioorthogonal functional group, and
　R Is a reactive group for the antibody. ], A compound having an affinity for an antibody, a cleavage moiety and a compound having a reactive group or a salt thereof, and
　an affinity for an antibody is represented by the
formula 1-1: 1 (X 0-3 ) a− C -Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-I-IWC- (X 0-3 ) b (SEQ ID NO
: 58) Equation 1-2: (X 0-3 ) a- C-Xaa1- Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-IVWC- (X 0-3 ) b (SEQ ID NO: 59)
Equation 1-3: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-VVW-C- (X 0-3 ) b (SEQ ID NO: 60)
Equation 1- 4: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-A-V-WC- (X 0-3 ) b (SEQ ID NO
: 61) Equation 1-5 :( X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-L-W-C- (X 0-3 ) b (SEQ ID NO
: 62) Equation 1-6: (X 0-) 3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-IWC- (X 0-3 ) b(SEQ ID NO
: 63) Equation 1-7: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-LVFC- (X 0-3 ) b (SEQ ID NO : 63) 64)
formula 1-8: (X 0-3 ) a -C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-Q-V-W-C-(X 0-3 ) b (SEQ ID NO: 65)
expression 1-9: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-EVW-C- (X 0-3 ) b (SEQ ID NO: 66)
[In the formula,
　(X 0-3 ) a is none, or 1 to 3 consecutive identical or different arbitrary amino acid residues (other than lysine and cysteine ​​residues).
　(X 0-3 ) b is none, or 1 to 3 consecutive identical or different amino acid residues (other than lysine and cysteine ​​residues), and
　Xaa1 is an alanine residue, glycine residue. , Leucine residue, proline residue, arginine residue, valine residue, asparagine residue, glutamate residue, or phenylalanine residue, and
　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue. a
　group, Xaa3 is histidine residue, a phenylalanine residue, a tyrosine residue, a tryptophan residue, an arginine residue or glycine
　residue,, Xaa4 is a lysine
　residue, Xaa @ 5 is a glycine residue, Serine residue, asparagine residue, glutamine residue, aspartic acid residue, glutamic acid residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue, alanine residue, valine A residue, isoleucine residue, or arginine residue,
　Xaa6 is a glutamine residue, glutamic acid residue, asparagine residue, aspartic acid residue, proline residue, glycine residue, arginine residue, phenylalanine residue, Or a histidine residue. ], Or
Equation 2-1: (X 0-3 ') a-C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-LVWC- (X 0-3 ') b (SEQ ID NO: 67)
[In the formula,
　(X 0- 3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively, and are
　Xaa1', Xaa2', Xaa3', Xaa4', Xaa5. ', Xaa6' are the same as the Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different amino acid residues, (X 0-3 ') b.Except when 1 to 3 consecutive identical or different arbitrary amino acid residues, Xaa3'is a histidine residue, and Xaa5'is a glycine residue. ], A reagent that is a peptide containing any of the amino acid sequences of.
[Claim 15]
　An antibody having an affinity substance for an antibody and a cleaving moiety or a salt thereof, wherein the following
　formula (II):
ALBR'-T (II)
[In the formula,
　A is an affinity for an antibody. A substance,
　L is a cleavable linker which is a divalent group containing a cleavable moiety, and
　B is (a) a divalent group containing a bioorthogonal functional group or (b) a bioorthogonal functional group. It is a group-free divalent group, where
　R'is the moiety produced by the reaction between the antibody and the reactive group and
　T is the antibody. ], And the
　affinity substance for the antibody is the
formula 1-1: 1 (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-I-I-W- C- (X 0-3 ) b (SEQ ID NO
: 58) Equation 1-2: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-IVW-C- ( X 0-3 ) b(SEQ ID NO
: 59) Equation 1-3: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-VVW-C- (X 0-3 ) b (SEQ ID NO : 59) 60)
formula 1-4: (X 0-3 ) a -C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-A-V-W-C-(X 0-3 ) b (SEQ ID NO: 61)
expression 1-5: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-L-WC- (X 0-3 ) b (SEQ ID NO: 62)
Equation 1-6 : (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-I-WC- (X 0-3 ) b (SEQ ID NO: 63)
expression 1-7: (X 0-3 ) a -C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-L-V-F-C-(X 0-3 ) b (SEQ No. 64)
Equation 1-8: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Q-VWC- (X 0-3 ) b (SEQ ID NO: 65)
Formula 1-9: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-EVW-C- (X 0-3 ) b (SEQ ID NO: 66)
[in the formula ,
　(X 0-3 ) aIs none, or any one to three consecutive identical or different amino acid residues (other than lysine and cysteine ​​residues), and
　(X 0-3 ) b is none, or one to three. Any consecutive identical or different amino acid residue (other than lysine residue and cysteine ​​residue),
　Xaa1 is an alanine residue, glycine residue, leucine residue, proline residue, arginine residue, valine residue, Asparagine residue, glutamate residue, or phenylalanine residue,
　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue, and
　Xaa3 is a histidine residue, phenylalanine residue, tyrosine residue. A group, tryptophan residue, arginine residue, or glycine residue,
　Xaa4 is a lysine residue,
　Xaa5 is a glycine residue, serine residue, asparagine residue, glutamine residue, aspartic acid residue, Glutamic acid residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue, alanine residue, valine residue, isoleucine residue, or arginine residue,
　Xaa6 is Glutamine residue, glutamic acid residue, asparagine residue, aspartic acid residue, proline residue, glycine residue, arginine residue, phenylalanine residue, or histidine residue. ], Or
Equation 2-1: (X 0-3)') a- C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-LVWC- (X 0-3 ') b (SEQ ID NO: 67)
[In the formula,
　( X 0-3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively, and are
　Xaa1', Xaa2', Xaa3', and Xaa4. ', Xaa5', and Xaa6'are the same as Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different amino acid residues, (X 0-3 ') b.Except when 1 to 3 consecutive identical or different arbitrary amino acid residues, Xaa3'is a histidine residue, and Xaa5'is a glycine residue. ], Which is a peptide containing any of the amino acid sequences of an antibody or a salt thereof.
[Claim 16]
　The antibody according to claim 15, or a salt thereof, wherein the antibody is a monoclonal antibody.
[Claim 17]
　The antibody or salt thereof according to claim 15 or 16, wherein the antibody is an IgG antibody.
[Claim 18]
　The antibody or salt thereof according to any one of claims 15 to 17, wherein the antibody is of human origin.
[Claim 19]
　The antibody or salt thereof according to any one of claims 15 to 18, wherein the antibody contains any one Fc region protein selected from the group consisting of the following (A) to (C) and has an antigen-binding ability. :
(A) Fc region protein containing the amino acid sequence of SEQ ID NO : 1;
(B) Contains an amino acid sequence in which one or several amino acid residues are inserted, added, deleted or substituted in the amino acid sequence of SEQ ID NO: 1. Fc region protein; or
(C) an Fc region protein containing an amino acid sequence showing 90% or more identity with the amino acid sequence of SEQ ID NO: 1.
[Claim 20]
　The antibody contains one or more specific amino acid residues in a target region consisting of 1 to 50 consecutive amino acid residues, and 5 of the specific amino acid residues in a non-target region other than the target region. wherein FOB,
　a-L-B-R 'represented by the structural units, combined with 30% or more positions selectivity for one or more specific amino acid residues contained in said target area The antibody or salt thereof according to any one of claims 15 to 19.
[Claim 21]
　The antibody or salt thereof according to claim 20, wherein the target region is a region consisting of 1 to 10 consecutive amino acid residues.
[Claim 22]
　The antibody or salt thereof according to claim 21, wherein the target region is a region consisting of 1 to 3 consecutive amino acid residues.
[Claim 23]
　The antibody or salt thereof according to claim 22, wherein the target region is a region consisting of amino acid residues at positions 246 to 248 in the human IgG Fc region.
[Claim 24]
　The antibody or salt thereof according to any one of claims 20 to 23, wherein the regioselectivity is 50% or more.
[Claim 25]
　The antibody or salt thereof according to claim 24, wherein the regioselectivity is 70% or more.
[Claim 26]
　The antibody or salt thereof according to claim 25, wherein the regioselectivity is 90% or more.
[Claim 27]
　The portion produced by the reaction between the antibody and the reactive group is on the side chain of any one of the lysine residue, tyrosine residue, or tryptophan residue with respect to the lysine residue, tyrosine residue, or tryptophan residue. The antibody or salt thereof according to any one of claims 15 to 26, which is a portion produced by the reaction of a specific reactive group.
[Claim 28]
　Claim that the moiety produced by the reaction between the antibody and the reactive group is the moiety produced by the reaction between the lysine residue and the reactive group specific for the side chain of the lysine residue. The antibody or salt thereof according to any one of 15 to 27.
[Claim 29]
　A method for producing an antibody having a bioorthogonal functional group or a salt thereof, wherein the following
　formula (II):
ALBR'-T (II)
[In the formula,
　A is an affinity substance for the antibody. Yes,
　L is a cleavable linker which is a divalent group containing a cleavable moiety, and
　B is a (a) divalent group containing a bioorthogonal functional group or (b) a bioorthogonal functional group. It is a divalent group that does not contain
　, R'is the part produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ], The cleaving portion of the antibody or salt thereof having the affinity substance for the antibody and the cleaving moiety is cleaved, and the following
　formula (IV):
L1-BR'-T (IV)
[formula] Among them,
　B, R'and T are the same as those of the above formula (II), and
　L1 is a monovalent group containing (i') bioorthogonal functional group or (ii') bioorthogonal functional group. It is a monovalent group containing no group. ], Which comprises producing an antibody having a bioorthogonal functional group or a salt thereof, and
　an affinity substance for the antibody is represented by the
formula 1-1: 1 (X 0-3 ) a.-Xaa1--C-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6 I-I-W-C-(X 0-3 ) b (SEQ ID NO: 58)
expression 1-2: (X 0-3 ) a -C- Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-IVWC- (X 0-3 ) b (SEQ ID NO
: 59) Equation 1-3: (X 0-3 ) a- C-Xaa1-Xaa2 -Xaa3-Xaa4-Xaa5-Xaa6-VVVWC- (X 0-3 ) b (SEQ ID NO
: 60) Equation 1-4: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3- Xaa4-Xaa5-Xaa6-AVWC- (X 0-3 ) b (SEQ ID NO
: 61) Equation 1-5: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-L-WC- (X 0-3 ) b (SEQ ID NO
: 62) Equation 1-6: (X 0-3 ) a- C -Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-IWC- (X 0-3 ) b (SEQ ID NO
: 63) Equation 1-7: (X 0-3 ) a- C-Xaa1- Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-LVFC- (X 0-3 ) b (SEQ ID NO
: 64) Equation 1-8: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3 -Xaa4-Xaa5-Xaa6-Q-V-WC- (X 0-3 ) b (SEQ ID NO
: 65) Equation 1-9: (X 0-3)) A -C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-E-V-W-C-(X 0-3 ) b (SEQ ID NO: 66) 　wherein,
(X 0-3 ) a is None, or any 1 to 3 consecutive identical or different amino acid residues (other than lysine and cysteine ​​residues), where 　(X 0-3 ) b is none or 1 to 3 consecutive Any amino acid residue (other than lysine residue and cysteine ​​residue) that is the same or different, 　Xaa1 is an alanine residue, glycine residue, leucine residue, proline residue, arginine residue, valine residue, asparagine residue. Group, glutamic acid residue, or phenylalanine residue, 　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue, and 　Xaa3 is a histidine residue, phenylalanine residue, tyrosine residue, A tryptophan residue, an arginine residue, or a glycine residue, 　Xaa4 is a lysine residue,

　Xaa5 contains glycine residue, serine residue, asparagine residue, glutamine residue, aspartic acid residue, glutamic acid residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue. A group, alanine residue, valine residue, isoleucine residue, or arginine residue,
　Xaa6 is glutamine residue, glutamic acid residue, asparagine residue, aspartic acid residue, proline residue, glycine residue, arginine. A residue, a phenylalanine residue, or a histidine residue. ], Or
formula 2-1: (X 0-3 ') a- C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-L-VWC- (X 0-3) ') b (SEQ ID NO: 67)
[In the equation,
　(X 0-3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively. And
　Xaa1', Xaa2', Xaa3', Xaa4', Xaa5', and Xaa6'are the same as Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different arbitrary amino acid residues, and (X 0-3 ') b is 1 to 3 consecutive identical or different arbitrary amino acid residues. Except when the group, Xaa3'is a histidine residue and Xaa5' is a glycine residue. ] Is a peptide comprising any of the amino acid sequences of.
[Claim 30]
　A method for producing an antibody having a functional substance or a salt thereof, wherein the following
　formula (III):
ALB'(-F) -R'-T (III)
[In the formula,
　A is an affinity for an antibody. It is a sex substance,
　L is a cleavable linker which is a divalent group containing a cleavable moiety,
　and B'contains a moiety produced by a reaction between a functional substance and a bioorthogonal functional group 3 The valence group,
　F is the functional substance,
　R'is the part produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ], The cleaving part of the complex having an affinity substance, a cleaving part, a functional substance and an antibody for the antibody or a salt thereof is cleaved, and the following
　formula (V1):
L1-B'(-F) ) -R'-T (V1)
[In the formula,
　L1 is a monovalent group containing (i') bioorthogonal functional group or (ii') monovalent group not containing bioorthogonal functional group. Yes,
　B', F, R'and T are the same as those in formula (III) above. ], Which comprises producing an antibody having a functional substance or a salt thereof, and
　an affinity substance for the antibody is represented by the
formula 1-1: 1 (X 0-3 ) a.-Xaa1--C-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6 I-I-W-C-(X 0-3 ) b (SEQ ID NO: 58)
expression 1-2: (X 0-3 ) a -C- Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-IVWC- (X 0-3 ) b (SEQ ID NO
: 59) Equation 1-3: (X 0-3 ) a- C-Xaa1-Xaa2 -Xaa3-Xaa4-Xaa5-Xaa6-VVVWC- (X 0-3 ) b (SEQ ID NO
: 60) Equation 1-4: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3- Xaa4-Xaa5-Xaa6-AVWC- (X 0-3 ) b (SEQ ID NO
: 61) Equation 1-5: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-L-WC- (X 0-3 ) b (SEQ ID NO
: 62) Equation 1-6: (X 0-3 ) a- C -Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-IWC- (X 0-3 ) b (SEQ ID NO
: 63) Equation 1-7: (X 0-3 ) a- C-Xaa1- Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-LVFC- (X 0-3 ) b (SEQ ID NO
: 64) Equation 1-8: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3 -Xaa4-Xaa5-Xaa6-Q-V-WC- (X 0-3 ) b (SEQ ID NO
: 65) Equation 1-9: (X 0-3)) A -C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-E-V-W-C-(X 0-3 ) b (SEQ ID NO: 66) 　wherein,
(X 0-3 ) a is None, or any 1 to 3 consecutive identical or different amino acid residues (other than lysine and cysteine ​​residues), where 　(X 0-3 ) b is none or 1 to 3 consecutive Any amino acid residue (other than lysine residue and cysteine ​​residue) that is the same or different, 　Xaa1 is an alanine residue, glycine residue, leucine residue, proline residue, arginine residue, valine residue, asparagine residue. Group, glutamic acid residue, or phenylalanine residue, 　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue, and 　Xaa3 is a histidine residue, phenylalanine residue, tyrosine residue, A tryptophan residue, an arginine residue, or a glycine residue, 　Xaa4 is a lysine residue,

　Xaa5 contains glycine residue, serine residue, asparagine residue, glutamine residue, aspartic acid residue, glutamic acid residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue. A group, alanine residue, valine residue, isoleucine residue, or arginine residue,
　Xaa6 is glutamine residue, glutamic acid residue, asparagine residue, aspartic acid residue, proline residue, glycine residue, arginine. A residue, a phenylalanine residue, or a histidine residue. ], Or
formula 2-1: (X 0-3 ') a- C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-L-VWC- (X 0-3) ') b (SEQ ID NO: 67)
[In the equation,
　(X 0-3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively. And
　Xaa1', Xaa2', Xaa3', Xaa4', Xaa5', and Xaa6'are the same as Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different arbitrary amino acid residues, and (X 0-3 ') b is 1 to 3 consecutive identical or different arbitrary amino acid residues. Except when the group, Xaa3'is a histidine residue and Xaa5' is a glycine residue. ] Is a peptide comprising any of the amino acid sequences of.
[Claim 31]

A 　method for producing an antibody or a salt thereof having a functional substance, wherein (A) the following formula (II):
ALBR'-T (II)
[In the formula,
　A is an affinity substance for the antibody. in it,
　L is a divalent cleavable linker is a group containing a cleavable moiety,
　B is a divalent radical or (b) bio-orthogonal functional groups, comprising (a) a bio-orthogonal functional group
　Is a divalent group that does not contain , R'is the moiety produced by the reaction between the antibody and the reactive group, and
　T is the antibody. ], The cleaving portion of the antibody or salt thereof having the affinity substance for the antibody and the cleaving moiety is cleaved, and the following
　formula (IV):
L1-BR'-T (IV)
[formula] Among them,
　B, R'and T are the same as those of the above formula (II), and
　L1 is a monovalent group containing (i') bioorthogonal functional group or (ii') bioorthogonal functional group. It is a monovalent group containing no group. ] To produce an antibody having a bioorthogonal functional group or a salt thereof, and
(B) one or more functional substances of the antibody having a bioorthogonal functional group or a salt thereof. In response to the following
　formula (V2)
FL1'-BR'-T (V2)
[in the formula,
　B, R'and T are the same as those of formula (IV) above, and
　L1'is due to the reaction between the functional material and (i') a monovalent group containing a bioorthogonal functional group. It is a divalent group containing the part to be produced, and
　F is a functional substance. ], Or the following
　formula (V3):
Fa-L1'-B'(-Fb) -R'-T (V3)
[In the formula, R'and
　T are the same as those in the formula (IV).
　L1'is the same as that of the above formula (V2)
　,
　B'is a trivalent group containing a moiety formed by the reaction between the functional substance and the bioorthogonal functional group, Fa and Fb. Are the same or different functional substances, respectively. ], The substance having
　an affinity for the antibody , which comprises producing an antibody having a functional substance or a salt thereof, is represented by the
formula 1-1: 1 (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3. -Xaa4-Xaa5-Xaa6-I-IWC- (X 0-3 ) b (SEQ ID NO
: 58) Equation 1-2: (X 0-3 ) a-Xaa1--C-Xaa2-Xaa3-Xaa4-Xaa @ 5-Xaa @ 6-I-V W-C-(X 0-3 ) b (SEQ ID NO: 59)
expression 1-3: (X 0-3 ) a -C- Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-VVWC- (X 0-3 ) b (SEQ ID NO
: 60) Equation 1-4: (X 0-3 ) a- C-Xaa1-Xaa2 -Xaa3-Xaa4-Xaa5-Xaa6-AV-WC- (X 0-3 ) b (SEQ ID NO
: 61) Equation 1-5: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3- Xaa4-Xaa5-Xaa6-L-L-WC- (X 0-3 ) b (SEQ ID NO
: 62) Equation 1-6: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-LIWC- (X 0-3 ) b (SEQ ID NO
: 63) Equation 1-7: (X 0-3 ) a- C -Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-L-VFC- (X 0-3 ) b (SEQ ID NO
: 64) Equation 1-8: (X 0-3 ) a- C-Xaa1- Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-QVWC- (X 0-3 ) b (SEQ ID NO
: 65) Equation 1-9: (X 0-3 ) a- C-Xaa1-Xaa2-Xaa3 -Xaa4-Xaa5-Xaa6-EVW-C- (X 0-3 ) b (SEQ ID NO: 66)
[In the formula,
　(X) 0-3 ) a is none, or 1 to 3 consecutive identical or different arbitrary amino acid residues (other than lysine and cysteine ​​residues), and
　(X 0-3 ) b is none, or 1 to 3 consecutive identical or different arbitrary amino acid residues (other than lysine residue and cysteine ​​residue),
　Xaa1 is an alanine residue, a glycine residue, a leucine residue, a proline residue, an arginine residue. , Valin residue, asparagine residue, glutamate residue, or phenylalanine residue,
　Xaa2 is a tyrosine residue, tryptophan residue, histidine residue, or phenylalanine residue, and
　Xaa3 is a histidine residue, phenylalanine. Residues, tyrosine residues, tryptophan residues, arginine residues, or glycine residues,
　Xaa4 is a lysine residue,
　Xaa5 is a glycine residue, serine residue, asparagine residue, glutamine residue, With aspartic acid residue, glutamate residue, phenylalanine residue, tyrosine residue, tryptophan residue, histidine residue, threonine residue, leucine residue, alanine residue, valine residue, isoleucine residue, or arginine residue There,
　Xaa @ 6 is a glutamine residue, a glutamic acid residue, asparagine residue, aspartic acid residue, a proline residue, glycine residue, arginine residue, a phenylalanine residue or a histidine residue. ], Or
Equation 2-1: (X 0-3 ') a- C-Xaa1'-Xaa2'-Xaa3'-Xaa4'-Xaa5'-Xaa6'-LVWC- (X 0-3 ') b (SEQ ID NO: 67)
[In the equation,
　(X 0-3 ') a and (X 0-3 ') b are the same as (X 0-3 ) a and (X 0-3 ) b , respectively.
　Xaa1', Xaa2', Xaa3', Xaa4', Xaa5', and Xaa6'are the same as Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, and Xaa6, respectively.
　However, (X 0-3 ') a is 1 to 3 consecutive identical or different amino acid residues, (X 0-3 ') b.Except when 1 to 3 consecutive identical or different arbitrary amino acid residues, Xaa3'is a histidine residue, and Xaa5'is a glycine residue. ] Is a peptide comprising any of the amino acid sequences of.