A bicyclic peptide ligand specific to transferrin receptor 1 (TfR1)

Abstract

The present invention relates to polypeptides covalently attached to a molecular scaffold such that two or more peptide loops are interposed between the attachment points to the scaffold. In particular, the present invention describes peptides that bind to TfR1. The present invention also relates to multimeric binding complexes comprising at least two of the bicyclic peptide ligands. The present invention also includes pharmaceutical compositions comprising the peptide ligands and multimeric binding complexes, as well as the use of the peptide ligands, multimeric binding complexes, and pharmaceutical compositions in the prevention, suppression, or treatment of diseases or disorders through the delivery of therapeutic agents mediated by TfR1.

Description

[Technical Field]

[0001] (Field of invention) The present invention relates to polypeptides covalently bound to a molecular scaffold such that two or more peptide loops are embedded between attachment points to the scaffold. In particular, the present invention describes peptides that bind to TfR1. The present invention also relates to a multimer-binding complex comprising at least two of the bicyclic peptide ligands. The present invention also relates to a pharmaceutical composition comprising the peptide ligands and the multimer-binding complex, and the use of the peptide ligands, the multimer-binding complex and the pharmaceutical composition in the prevention, suppression or treatment of disease or disorder by delivery of a therapeutic agent mediated by TfR1. [Background technology]

[0002] (Background of the invention) Cyclic peptides can bind to protein targets with high affinity and specificity, and are therefore an attractive molecular class for the development of therapeutic drugs. In fact, several cyclic peptides have already been successfully used in clinical practice, such as the antimicrobial peptide vancomycin, the immunosuppressant cyclosporine, or the anticancer drug octreotide (Driggers et al. (2008), Nat Rev Drug Discov 7(7), 608-24). The excellent binding properties are due not only to the relatively large interaction surface formed between the peptide and the target, but also to the reduced conformational flexibility of the cyclic structure. Typically, macrocyclic molecules are used, for example, the cyclic peptide CXCR4 antagonist CVX15 (400Å). 2 ; Wu et al. (2007), Science 330, 1066-71), a cyclic peptide having an Arg-Gly-Asp motif that binds to integrin αVb3 (355Å 2 (Xiong et al. (2002), Science 296(5565), 151-5), or upain-1 (603Å), a cyclic peptide inhibitor that binds to urokinase-type plasminogen activator. 2As described in the literature by Zhao et al. (2007), J Struct Biol 160(1), 1-10), it binds to a surface area of ​​several hundred square angstroms.

[0003] Due to their cyclic configuration, macrocyclic peptide molecules are less flexible than linear peptides, resulting in less entropy loss upon binding to a target and consequently higher binding affinity. This reduced flexibility also leads to the fixation of target-specific conformations, increasing binding specificity compared to linear peptides. This effect is exemplified by the potent and selective inhibitory effects of matrix metalloproteinase 8 (MMP-8), which loses its selectivity for other MMPs when its ring is unfolded (Cherney et al. (1998), J Med Chem 41(11), 1749-51). The advantageous binding properties achieved by macrocyclization are even more pronounced in polycyclic peptides with multiple peptide rings, such as vancomycin, nisin, and actinomycin.

[0004] Various research teams have previously linked polypeptides containing cysteine ​​residues into synthetic molecular structures (Kemp and McNamara (1985), J. Org. Chem; Timmerman et al. (2005), ChemBioChem). Meloen and collaborators used tris(bromomethyl)benzene and related molecules for the rapid and quantitative cyclization of multiple peptide loops on synthetic scaffolds for structural mimicry of protein surfaces (Timmerman et al. (2005), ChemBioChem). Method for preparing candidate drug compounds (where the compound is prepared by linking a cysteine-containing polypeptide to a molecular scaffold such as 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (TATA)) (Heinis et al. (2014) Angewandte Chemie, International Edition 53(6) 1602-1606).

[0005] A phage display-based combinatorial approach has been developed for constructing and screening large libraries of bicyclic peptides targeting specific organisms (Heinis et al. (2009), Nat Chem Biol 5(7), 502-7 and WO 2009 / 098450). Briefly, a combinatorial library of linear peptides (Cys-(Xaa)6-Cys-(Xaa)6-Cys) containing three cysteine ​​residues and two random 6-amino acid regions was displayed on a phage, and the cysteine ​​side chains were covalently bonded to a small molecule scaffold to form a cyclization. [Overview of the project]

[0006] (Summary of the invention) According to a first aspect of the present invention, a peptide ligand specific to transferrin receptor 1 (TfR1) is provided, comprising a polypeptide having at least three reactive groups separated by at least two loop sequences, and a molecular scaffold that forms covalent bonds with the reactive groups of the polypeptide, thereby forming at least two polypeptide loops on the molecular scaffold.

[0007] A further aspect of the present invention provides a multimer-binding complex comprising a peptide ligand specific to transferrin receptor 1 (TfR1), wherein the peptide ligands are either the same or different, and each of these comprises a polypeptide having at least three reactive groups separated by at least two loop sequences, and a molecular scaffold that forms covalent bonds with the reactive groups of the polypeptide, resulting in at least two polypeptide loops being formed on the molecular scaffold.

[0008] A further aspect of the present invention provides a pharmaceutical composition comprising a peptide ligand or polymer-binding complex as defined herein in combination with one or more pharmaceutically acceptable excipients.

[0009] A further aspect of the present invention provides peptide ligands or polymer-bound complexes or pharmaceutical compositions as defined herein for use in preventing, suppressing, or treating a disease or disorder by delivery of a therapeutic agent mediated by TfR1. [Brief explanation of the drawing]

[0010] (Brief explanation of the drawing) [Figure 1] Figure 1: Results of a transcytosis assay using BCY17986 with primary cultures of human proximal tubular cells. [Figure 2] Figure 2: Results of a transcytosis assay using BCY17988 with primary cultures of human proximal tubular cells. [Figure 3] Figure 3: Results of a transcytosis assay using BCY17989 with primary cultures of human proximal tubular cells. [Figure 4] Figure 4: Results of a transcytosis assay using BCY17994 with primary cultures of human proximal tubular cells. [Modes for carrying out the invention]

[0011] (Detailed description of the invention) It will be understood that the present invention relates to both "monomer" bicyclic peptides, i.e., those containing a single (monomer) bicyclic peptide ligand, and "multimeric" bicyclic peptides, i.e., those containing multiple bicyclic peptides (e.g., two, three, or four) conjugated via one or more linkers.

[0012] (Monomer bicyclic peptide ligand) According to a first aspect of the present invention, a peptide ligand specific to transferrin receptor 1 (TfR1) is provided, comprising a polypeptide having at least three reactive groups separated by at least two loop sequences, and a molecular scaffold that forms covalent bonds with the reactive groups of the polypeptide, thereby forming at least two polypeptide loops on the molecular scaffold.

[0013] In one embodiment, the reactive group includes a cysteine ​​residue.

[0014] The term "TfR1-specific" will be understood to refer to the ability of a peptide ligand to bind to transferrin receptor 1 (TfR1). It will also be understood that the peptide ligand may have different effects on TfR1 depending on the exact epitope of binding. For example, the effect may be either inhibitory (i.e., the peptide ligand interferes with / inhibits the binding of transferrin to TfR1) or non-inhibitory (i.e., the peptide ligand does not interfere with / inhibit the binding of transferrin to TfR1).

[0015] (Inhibitory peptide ligand) In one embodiment, the peptide ligand is specific to TfR1 and binds to TfR1 in a manner that interferes with / inhibits the binding of transferrin to TfR1.

[0016] In a further embodiment, the loop sequence comprises 2, 3, 6, 8, or 9 amino acids.

[0017] In one embodiment, the loop sequence comprises three cysteine ​​residues separated by two loop sequences, the first of which consists of two amino acids, and the second of which consists of nine amino acids.

[0018] In one embodiment, the loop sequence comprises three cysteine ​​residues separated by two loop sequences, each consisting of six amino acids.

[0019] In one embodiment, the loop sequence comprises three cysteine ​​residues separated by two loop sequences, the first of which consists of three amino acids, and the second of which consists of eight amino acids.

[0020] In one embodiment, the peptide ligand is [Chemistry] (where C i 、C ii 、and C iii represent the first, second, and third cysteine residues, respectively) : comprises an amino acid sequence of or a pharmaceutically acceptable salt thereof.

[0021] In a further embodiment, the molecular scaffold is 1,1',1''-(1,3,5-triazinane-1,3,5-triyl)triprop-2-en-1-one (TATA), the peptide ligand comprises N- and / or C-terminal additions, and A-(SEQ ID NO: 1)-A (referred to herein as BCY12455); A-(SEQ ID NO: 1)-A-[Sar6]-[K-Fl] (referred to herein as BCY12652); A-(SEQ ID NO: 2)-A (referred to herein as BCY12452); A-(SEQ ID NO: 2)-A-[Sar6]-[K-Fl] (referred to herein as BCY12650); A-(SEQ ID NO: 3)-A (referred to herein as BCY12454); and A-(SEQ ID NO: 3)-A-[Sar6]-[K-Fl] (referred to herein as BCY12651) (where Sar represents sarcosine and Fl represents fluorescein) : is selected from.

[0022] For the purposes of this description, it is assumed that the inhibitory bicyclic peptide is cyclized with TATA to give a trisubstituted structure. However, as will be apparent from the description of the invention presented herein, cyclization can be carried out with any suitable molecular scaffold that forms a covalent bond with a reactive group of the polypeptide such that at least two polypeptide loops are formed. Cyclization occurs on C i 、C ii 、and C iii .

[0023] (Non-inhibitory peptide ligand) In one embodiment, the peptide ligand is specific to TfR1 and binds to TfR1 in a manner that does not interfere with or inhibit the binding of transferrin to TfR1.

[0024] In a further embodiment, the loop sequence comprises three or seven amino acids.

[0025] In one embodiment, the loop sequence comprises three cysteine ​​residues separated by two loop sequences, the first of which consists of seven amino acids, and the second of which consists of three amino acids.

[0026] In one embodiment, the peptide ligand is [ka] TIFF0007848202000003.tif248170TIFF0007848202000004.tif41170(where Abu represents aminobutyric acid, Aib represents aminoisobutyric acid, Aze represents azetidine, B-MeIle represents β-methylisoleucine, C5g represents cyclopentylglycine, Cba represents β-cyclobutylalanine, Cbg represents cyclobutylglycine, Chg represents cyclohexylglycine, Cpg represents cyclo [K(N3)] represents propylglycine, EPA represents 2-amino-3-ethylpentanoic acid, HyP represents trans-4-hydroxy-L-proline, [K(N3)] represents 6-azidridine, 1Nal represents 1-naphthylalanine, 2Nal represents 2-naphthylalanine, 4Pal represents 4-pyridylalanine, tBuAla represents t-butylalanine, tBuGly represents t-butylglycine, 3tBuTyr represents 3-t-butyltyrosine, and C i , C ii , and C iii (These represent the first, second, and third cysteine ​​residues, respectively.) Contains the amino acid sequence of : or a pharmaceutically acceptable salt thereof.

[0027] In a further embodiment, the peptide ligand is [ka] (Here, C i , C ii , and C iii (These represent the first, second, and third cysteine ​​residues, respectively.) Contains the amino acid sequence of : or a pharmaceutically acceptable salt thereof.

[0028] In a further embodiment, the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tris(2-bromoetanone)(TATB), and the peptide ligand includes N- and / or C-terminal additions, A-(Sequence ID 4)-A (referred to herein as BCY13983); A-(Sequence ID 4)-A-[Sar6]-[K-Fl] (referred to herein as BCY14474); A-(Sequence ID 5)-A (referred to herein as BCY13986); A-(Sequence ID 5)-A-[Sar6]-[K-Fl] (referred to herein as BCY14475); A-(Sequence ID 6)-A (referred to herein as BCY15466); Ac-(Sequence ID 6) (referred to as BCY15889 herein); A-(Sequence ID 7)-A (referred to herein as BCY15467); Ac-(Sequence ID 7) (referred to as BCY15890 herein); A-(Sequence ID 8)-A (referred to herein as BCY13989); A-(Sequence ID 8)-A-[Sar6]-[K-Fl] (referred to herein as BCY14476); A-(Sequence ID 9)-A (referred to herein as BCY15468); A-(Sequence ID 9)-A-[Sar6]-[K-Fl] (referred to herein as BCY15768); (Sequence ID 9)-[Sar6]-[K-Fl] (referred to herein as BCY15934); Ac-(Sequence ID 9)-A-[Sar6]-[K-Fl](referred to herein as BCY15937); Ac-(SEQ ID NO: 9)-[Sar6]-[K-Fl] (referred to herein as BCY15938); [Fl]G[Sar5]-A-(Sequence ID 9)-A (referred to herein as BCY15940); N[1Nal]N-(SEQ ID NO: 9) (referred to as BCY18030 in this specification); Ac-(Sequence ID 9)-E[Pip]W (referred to herein as BCY18039); Ac-(SEQ ID NO: 9)-EPW (referred to herein as BCY17994); NWN-(Sequence ID 9) (referred to as BCY18029 herein); NWN-(Sequence ID 9)-A (referred to herein as BCY17109); Ac-(Sequence ID 9)-E[Aze]W (referred to herein as BCY18037); Ac-NWN-(Sequence ID 9) (referred to herein as BCY17992); Ac-(Sequence ID 9)-E[dP]W (referred to herein as BCY18038); Ac-N[1Nal]N-(SEQ ID NO: 9) (referred to as BCY18034 herein); N[dW]N-(Sequence ID 9) (referred to as BCY18031 in this specification); Ac-N[dW]N-(SEQ ID NO: 9) (referred to as BCY18035 in this specification); HWM-(Sequence ID 9)-A (referred to herein as BCY17110); A-(Sequence ID 9)-PHP (referred to herein as BCY17115); A-(Sequence ID 9)-EPW (referred to herein as BCY17114); NEV-(Sequence ID 9)-A (referred to herein as BCY17112); A-(Sequence ID 9)-PIVH (referred to herein as BCY17120); Ac-(Sequence ID 9) (referred to as BCY15891 herein); HTS-(Sequence ID 9)-A (referred to herein as BCY17111); Ac-N[NMeTrp]N-(SEQ ID NO: 9) (referred to as BCY18036 in this specification); N[NMeTrp]N-(SEQ ID NO: 9) (referred to as BCY18032 in this specification); Ac-A-(Sequence ID 9)-A (referred to herein as BCY15939); A-(Sequence ID 9)-EHQE (referred to herein as BCY17119); ESF-(Sequence ID 9)-A (referred to herein as BCY17113); NWN-(Sequence ID 9)-[K(N3)] (referred to as BCY17870 in this specification); Ac-NWN-(Sequence ID 9)-[K(N3)] (referred to as BCY17871 herein); [AzPro]-NWN-(Sequence ID 9) (referred to as BCY17872 herein); Ac-(SEQ ID NO: 9)-EPW-[K(N3)] (referred to as BCY17873 herein); [AzPro]-(Sequence ID 9)-EPW (referred to herein as BCY17874); Ac-(Sequence ID 9)-[K(N3)] (referred to as BCY17868 herein); [AzPro]-(Sequence ID 9) (referred to as BCY17869 herein); Ac-N[dY]N-(SEQ ID NO: 9)-[K(N3)] (referred to as BCY17882 in this specification); Ac-(Sequence ID 9)-E-[dP]-W-[K(N3)] (referred to herein as BCY17890); Ac-(Sequence ID 9)-E-[Aze]-W-[K(N3)] (referred to herein as BCY17892); Ac-(Sequence ID 9)-E-[Pip]-W-[K(N3)] (referred to herein as BCY17894); Ac-(SEQ ID NO: 9)-[K(N3)(PYA-maleimide)](referred to herein as BCY17906); Ac-(Sequence ID 9)-EPW-[Peg 10 ]-[K(N3)] (referred to as BCY19405 in this specification); Ac-(Sequence ID 9)-EPW-[Peg 24 ]-[K(N3)] (referred to as BCY19406 in this specification); Ac-(SEQ ID NO: 9)-EPWGGSGGS-[K(N3)] (referred to herein as BCY19407); A-(Sequence ID 10)-A (referred to herein as BCY15469); Ac-(Sequence ID 10) (referred to as BCY15892 herein); A-(Sequence ID 11)-A (referred to herein as BCY15470); Ac-(Sequence ID 11) (referred to as BCY15893 herein); A-(Sequence ID 12)-A (referred to herein as BCY15471); Ac-(Sequence ID 12) (referred to as BCY15894 herein); Ac-(Sequence ID 13) (referred to as BCY17991 herein); Ac-(Sequence ID 13)-EPW (referred to herein as BCY17995); Ac-NWN-(Sequence ID 13) (referred to herein as BCY17993); NWN-(Sequence ID 13) (referred to as BCY18033 in this specification); A-(Sequence ID 13)-A (referred to herein as BCY16754); Ac-(Sequence ID 13)-[K(N3)] (referred to as BCY17896 herein); Ac-NWN-(Sequence ID 13)-[K(N3)] (referred to herein as BCY17899); Ac-(SEQ ID NO: 13)-EPW-[K(N3)] (referred to as BCY17901 in this specification); Ac-(Sequence ID 14) (referred to as BCY17990 herein); Ac-(SEQ ID NO: 14)-[K(N3)] (referred to as BCY17875 herein); [AzPro]-(Sequence ID 14) (referred to herein as BCY17876); Ac-(Sequence ID 15) (referred to as BCY17989 herein); A-(Sequence ID 15)-A (referred to herein as BCY16047); Ac-(Sequence ID 15)-[K(N3)] (referred to as BCY17877 herein); [AzPro]-(Sequence ID 15) (referred to as BCY17878 herein); A-(Sequence ID 16)-A (referred to herein as BCY16962); TYMN-(Sequence ID 17)-A (referred to as BCY17117 herein); A-(Sequence ID 17)-A (referred to herein as BCY16048); A-(Sequence ID 18)-A (referred to herein as BCY16963); Ac-(Sequence ID 19) (referred to as BCY17987 herein); A-(Sequence ID 20)-A (referred to herein as BCY16753); A-(Sequence ID 21)-A (referred to herein as BCY16046); A-(Sequence ID 22)-A (referred to herein as BCY16964); A-(Sequence ID 23)-A (referred to herein as BCY16965); Ac-(Sequence ID 24) (referred to herein as BCY17986); A-(Sequence ID 25)-A (referred to herein as BCY16550); A-(Sequence ID 26)-A (referred to herein as BCY16966); A-(Sequence ID 27)-A (referred to herein as BCY16051); IDSN-(Sequence ID 28)-A (referred to herein as BCY17118); WGKS-(Sequence ID 29)-A (referred to herein as BCY17116); A-(Sequence ID 30)-A (referred to herein as BCY16053); A-(Sequence ID 31)-A (referred to herein as BCY16557); A-(Sequence ID 32)-A (referred to herein as BCY16035); A-(Sequence ID 33)-A (referred to herein as BCY16043); A-(Sequence ID 34)-A-[Sar6]-[K-Fl](referred to herein as BCY15769); A-(Sequence ID 35)-A (referred to herein as BCY15648); A-(Sequence ID 36)-A (referred to herein as BCY16031); A-(Sequence ID 37)-A (referred to herein as BCY16079); A-(Sequence ID 38)-A (referred to herein as BCY16036); A-(Sequence ID 39)-A (referred to herein as BCY16029); A-(Sequence ID 40)-A (referred to herein as BCY16089); A-(Sequence ID 41)-A (referred to herein as BCY16088); A-(Sequence ID 42)-A (referred to herein as BCY16052); A-(Sequence ID 43)-A (referred to herein as BCY16033); A-(Sequence ID 44)-A (referred to herein as BCY16039); Ac-(Sequence ID 44) (referred to herein as BCY17988); Ac-(SEQ ID NO: 44)-[K(N3)] (referred to as BCY17879 herein); [AzPro]-(Sequence ID 44) (referred to herein as BCY17880); A-(Sequence ID 45)-A (referred to herein as BCY16038); A-(Sequence ID 46)-A (referred to herein as BCY16050); A-(Sequence ID 47)-A (referred to herein as BCY16034); A-(Sequence ID 48)-A (referred to herein as BCY16032); A-(Sequence ID 49)-A (referred to herein as BCY16049); A-(Sequence ID 50)-A (referred to herein as BCY16558); A-(Sequence ID 51)-A (referred to herein as BCY16041); A-(Sequence ID 52)-A (referred to herein as BCY16042); A-(Sequence ID 53)-A (referred to herein as BCY16045); A-(Sequence ID 54)-A (referred to herein as BCY16037); A-(Sequence ID 55)-A (referred to herein as BCY16044); A-(Sequence ID 56)-A (referred to herein as BCY16040); A-(Sequence ID 57)-A-[Sar6]-[K-Fl](referred to herein as BCY15771); A-(Sequence ID 58)-A-[Sar6]-[K-Fl](referred to herein as BCY15772); A-(Sequence ID 59)-A-[Sar6]-[K-Fl](referred to herein as BCY15773); A-(Sequence ID 60)-A-[Sar6]-[K-Fl](referred to herein as BCY15774); A-(Sequence ID 61)-A-[Sar6]-[K-Fl](referred to herein as BCY15775); A-(Sequence ID 62)-A-[Sar6]-[K-Fl](referred to herein as BCY15776); A-(Sequence ID 63)-A-[Sar6]-[K-Fl](referred to herein as BCY15777); A-(Sequence ID 64)-A-[Sar6]-[K-Fl](referred to herein as BCY15770); Ac-(Sequence ID 65) (referred to herein as BCY17903); Ac-(SEQ ID NO: 66) (referred to herein as BCY17904); and Ac-(Sequence ID 67) (referred to as BCY17905 herein); (Here, AzPro represents azidopropyl, Aze represents azetidine, 1Nal represents 1-naphthylalanine, NMeTrp represents N-methyltryptophan, [K(N3)] represents 6-azidridine, Peg represents polyethylene glycol, Pip represents pipecolic acid, Sar represents sarcosine, Fl represents fluorescein, and [K(N3)(PYA-maleimide)] has the following structure: [ka] (Represents modified lysine having) Selected from:

[0029] In a further embodiment, the molecular scaffold is a TATB, the peptide ligand includes N- and / or C-terminal additions, and A-(Sequence ID 4)-A (referred to herein as BCY13983); A-(Sequence ID 4)-A-[Sar6]-[K-Fl] (referred to herein as BCY14474); A-(Sequence ID 5)-A (referred to herein as BCY13986); A-(Sequence ID 5)-A-[Sar6]-[K-Fl] (referred to herein as BCY14475); A-(Sequence ID 6)-A (referred to herein as BCY15466); A-(Sequence ID 7)-A (referred to herein as BCY15467); A-(Sequence ID 8)-A (referred to herein as BCY13989); A-(Sequence ID 8)-A-[Sar6]-[K-Fl] (referred to herein as BCY14476); A-(Sequence ID 9)-A (referred to herein as BCY15468); A-(Sequence ID 9)-A-[Sar6]-[K-Fl] (referred to herein as BCY15768); (Sequence ID 9)-[Sar6]-[K-Fl] (referred to herein as BCY15934); Ac-(Sequence ID 9)-A-[Sar6]-[K-Fl](referred to herein as BCY15937); Ac-(SEQ ID NO: 9)-[Sar6]-[K-Fl] (referred to herein as BCY15938); [Fl]G[Sar5]-A-(Sequence ID 9)-A (referred to herein as BCY15940); A-(Sequence ID 10)-A (referred to herein as BCY15469); A-(Sequence ID 11)-A (referred to herein as BCY15470); and A-(Sequence ID 12)-A (referred to herein as BCY15471); (Here, Sar represents sarcosine and Fl represents fluorescein.) Selected from:

[0030] In an alternative embodiment, the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (TATA), and the peptide ligand includes N- and / or C-terminal additions, Ac-(Sequence ID 13) (referred to as BCY20546 herein) : is.

[0031] For the purposes of this explanation, it is assumed that a non-inhibitory bicyclic peptide is cyclized with TATA or TATB to produce a trisubstituted structure. However, as will be apparent from the description of the invention presented herein, cyclization can be carried out with any suitable molecular scaffold that forms covalent bonds with the reactant groups of the polypeptide so as to form at least two polypeptide loops. Cyclization is performed with C i , C ii , and C iii It happens above.

[0032] In further embodiments, the pharmaceutically acceptable salt is selected from free acids or sodium, potassium, calcium, or ammonium salts.

[0033] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by experts in the relevant fields, such as peptide chemistry, cell culture and phage display, nucleic acid chemistry, and biochemistry. Standard techniques are used in molecular biology, genetics, and biochemical methods (see Sambrook et al., *Molecular Cloning: A Laboratory Manual*, 3rd edition, 2001, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Ausubel et al., *Short Protocols in Molecular Biology* (1999), 4th edition, John Wiley & Sons, incorporated herein by reference).

[0034] (Multimeric bicyclic peptide ligand) A further aspect of the present invention provides a multimer-binding complex comprising a peptide ligand specific to transferrin receptor 1 (TfR1), wherein the peptide ligands are either the same or different, and each of these comprises a polypeptide having at least three reactive groups separated by at least two loop sequences, and a molecular scaffold that forms covalent bonds with the reactive groups of the polypeptide, resulting in at least two polypeptide loops being formed on the molecular scaffold.

[0035] Accordingly, in this embodiment of the present invention, the polymer-binding complex comprises at least two (i.e., 2, 3, or 4) monomeric bicyclic peptide ligands as defined herein.

[0036] This aspect of the present invention describes a series of polymerized bicyclic peptides having various chemical linkers and hinges of various lengths and stiffnesses, utilizing different binding sites within the bicyclic peptide that bind to and activate TfR1 with a wide range of strengths and potencies.

[0037] Those skilled in the art will understand that this aspect of the present invention presents a multi-configuration (multimeric) bicyclic peptide that offers synergistic benefits through the resulting properties of the multimer-binding complex compared to a corresponding monomer-binding complex containing a single bicyclic peptide. For example, the multimer-binding complex of this aspect of the present invention typically has a greater level of binding strength or binding force (measured herein by Kd value) than its monomeric counterpart. Furthermore, the multimer-binding complex of the present invention is designed to be small enough to be removed by the kidney.

[0038] While not constrained by theory, it is thought that multimerized bicyclic peptides can activate receptors by homocrosslinking multiple identical receptors. Therefore, in one embodiment, the bicyclic peptide ligand is specific to the same target within TfR1. In a further embodiment, the multimer-binding complex contains at least two identical bicyclic peptide ligands. "Identical" means bicyclic peptides having the same amino acid sequence, most strictly speaking, the same amino acid sequence refers to the binding portion of the bicyclic peptide (for example, the sequence may differ in its binding position). In this embodiment, each of the bicyclic peptides in the multimer-binding complex binds to the exact same epitope on the same target of TfR1—hence, the resulting target-binding complex generates a homodimer (if the multimer-binding complex contains two identical bicyclic peptides), a homotrimer (if the multimer-binding complex contains three identical bicyclic peptides), or a homotetramer (if the multimer-binding complex contains four identical bicyclic peptides), and so on.

[0039] In an alternative embodiment, the multimer-binding complex comprises at least two different bicyclic peptide ligands. By “different,” we mean bicyclic peptides having different amino acid sequences. In this embodiment, the different bicyclic peptide ligands within the multimer-binding complex bind to different epitopes on TfR1—and therefore the resulting target-binding complexes are biparatopic (if the multimer-binding complex contains two different bicyclic peptides), triparatopic (if the multimer-binding complex contains three different bicyclic peptides), or tetraparatopic (if the multimer-binding complex contains four different bicyclic peptides), and so on.

[0040] While not constrained by theory, it is thought that multimerized bicyclic peptides can activate receptors by heterocrosslinking different targets, for example, different target sites on TfR1. Therefore, in one embodiment, the bicyclic peptide ligand is specific to different targets on TfR1. In this embodiment, it will be understood that the multimer-binding complex contains at least two different bicyclic peptide ligands (i.e., bicyclic peptide ligands having different amino acid sequences). In this embodiment, each of the bicyclic peptides in the multimer-binding complex binds to a different epitope on TfR1—and thus the resulting target-binding complex generates a bispecific multimer-binding complex (when the multimer complex contains two different bicyclic peptides), a triplicate multimer-binding complex (when the multimer complex contains three different bicyclic peptides), or a quadruplicate multimer-binding complex (when the multimer complex contains four different bicyclic peptides), and so on.

[0041] It will be understood that the multimer-binding complex of the present invention can be designed to bind to various different targets on TfR1.

[0042] The bicyclic peptides within the polymer-binding complex of the present invention can be associated by several different options. For example, there may be a central hinge or branching portion having spacer or arm elements extending radially from the hinge or branching point, each containing a bicyclic peptide. Alternatively, it can be envisioned that a circular support member can hold several peptides protruding inward or outward.

[0043] In one embodiment, each bicyclic peptide ligand is connected to a central hinge portion by a spacer group.

[0044] It will be understood that the spacer group is linear and can connect a single bicyclic peptide to the central hinge portion. Therefore, in one embodiment, the polymer-binding complex is a compound of formula (I): [ka] (In the formula, CHM represents the central hinge portion; The term "bicyclic" represents a bicyclic peptide ligand as defined herein; and (m represents an integer selected from 2 to 10.) Includes.

[0045] In one embodiment, m represents an integer selected from 2, 3, or 4.

[0046] In a further embodiment, m represents 2.

[0047] If m represents 2, it will be understood that the central hinge portion requires two connection points. Thus, in one embodiment, m represents 2, and CHM is the motif of equation (A): [ka] .

[0048] (dimer) In one embodiment, a multimer-binding complex containing two identical bicyclic peptides includes the dimer-binding complexes listed in Table A below: Table A: Exemplary dimer-bound complexes of the present invention [Table 1]

[0049] (Numbering) When referring to the position of an amino acid residue within the peptide of the present invention, the cysteine ​​residue (C) i , C ii , and C iii Since ) are invariant, they are omitted from the numbering, and therefore the numbering of amino acid residues in the peptide of the present invention is referred to as follows: -C i -A1-L2-C ii -N3-D4-W5-T6-L7-P8-W9-H 10 -H 11 -C iii - (Sequence ID 1).

[0050] (Molecular format) N- or C-terminal extensions to a biring core sequence are added to the left or right side of the sequence, separated by a hyphen. For example, the N-terminal biotin-G-Sar5 tail is: [Biot]-G-[Sar5]-A-(Sequence ID X)

[0051] (Reverse peptide sequence) Considering the disclosure in Nair et al.'s paper (2003) J Immunol 170(3), 1362-1373, it is anticipated that the peptide sequences disclosed herein may also find usefulness in their retro-inverso forms. For example, the sequence may be reversed (i.e., the N-terminus becomes the C-terminus and the C-terminus becomes the N-terminus), and its stereochemistry may also be reversed (i.e., D-amino acids become L-amino acids and L-amino acids become D-amino acids).

[0052] (Definition of peptide ligand) As used herein, peptide ligands refer to peptides, peptidics, or peptimimetics covalently bonded to a molecular scaffold. Typically, such peptides, peptidics, or peptimimetics comprise a peptide having native or non-native amino acids, two or more reactive groups (i.e., cysteine ​​residues) capable of forming a covalent bond with the scaffold, and an inherent sequence between the reactive groups, which is called a loop sequence because it forms a loop when the peptide, peptidic, or peptimetic binds to the scaffold. In this case, the peptide, peptidic, or peptimetic comprises at least three cysteine ​​residues (as used herein, C i , C ii , and C iii It includes (called) and forms at least two loops on the scaffold.

[0053] (Advantages of peptide ligands) The specific bicyclic peptides of the present invention possess several advantageous properties that allow them to be considered suitable drug-like molecules for injection, inhalation, nasal, ocular, oral, or topical administration. Such advantageous properties include: - Cross-reactivity. This is a typical requirement for preclinical pharmacodynamic and pharmacokinetic evaluations; - Protease stability. Bicyclic peptide ligands should exhibit stability against plasma proteases, epithelial ("membrane-immobilized") proteases, gastrointestinal proteases, lung surface proteases, intracellular proteases, etc., in most situations. Protease stability should be maintained across different species so that bicyclic peptide lead candidates can be developed not only in animal models but also confidently administered to humans. - Desired solubility profile. This is a function of the ratio of charged residues and hydrophilic to hydrophobic residues, as well as intramolecular / intermolecular H-bonds, which are important for formulation and absorption purposes; and - Optimal plasma half-life in circulation. Depending on the clinical indication and treatment regimen, it may be necessary to develop bicyclic peptides with short or long in vivo exposure times for the management of either chronic or acute disease states. The optimal exposure time is determined by the requirement for sustained exposure (for maximum therapeutic efficiency) compared to the requirement for short exposure times to minimize toxicological effects resulting from sustained exposure of the drug.

[0054] (Salt that is acceptable as a medicine) The salt form is within the scope of the present invention, and it will be understood that any reference to a peptide ligand includes the salt form of said ligand.

[0055] The salts of the present invention can be synthesized from parent compounds containing a basic or acidic moiety by conventional chemical methods, for example, by the methods described in Pharmaceutical Salts: Properties, Selection, and Use, edited by P. Heinrich Stahl and Camille G. Wermuth, ISBN: 3-90639-026-8, Hardcover, 388 pages, August 2002. Typically, such salts can be prepared by reacting the free acidic or basic form of these compounds with a suitable base or acid in water or an organic solvent, or in a mixture thereof.

[0056] Acid addition salts (mono or di salts) can be formed from a wide variety of acids, both inorganic and organic. Examples of acid addition salts include acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid, ascorbic acid (e.g., L-ascorbic acid), L-aspartic acid, benzenesulfonic acid, benzoic acid, 4-acetamidobenzoic acid, butanoic acid, (+)camphoric acid, camphorsulfonic acid, (+)-(1S)-camphor-10-sulfonic acid, capric acid, caproic acid, caprylic acid, cinnamic acid, citric acid, Cyclamic acid, dodecyl sulfate, ethane-1,2-disulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, formic acid, fumaric acid, mucoic acid, gentisic acid, glucoheptonic acid, D-gluconic acid, glucuronic acid (e.g., D-glucuronic acid), glutamic acid (e.g., L-glutamic acid), α-oxoglutaric acid, glycolic acid, hippuric acid, hydrohalic acid (e.g., hydrobromic acid, Hydrochloric acid, hydroiodic acid), isethionic acid, lactic acid (e.g., (+)-L-lactic acid, (±)-DL-lactic acid), lactobionic acid, maleic acid, malic acid, (-)-L-malic acid, malonic acid, (±)-DL-mandelic acid, methanesulfonic acid, naphthalene-2-sulfonic acid, naphthalene-1,5-disulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid, nitric acid, oleic acid, orotic acid, oxalic acid, palmitic acid Examples include monosalts or disalts formed from acids selected from the group consisting of tinic acid, pamoic acid, phosphoric acid, propionic acid, pyruvic acid, L-pyroglutamic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, sulfuric acid, tannic acid, (+)-L-tartaric acid, thiocyanic acid, p-toluenesulfonic acid, undecylenic acid, and valeric acid, as well as acylated amino acids and cation exchange resins.

[0057] One particular group of salts consists of salts formed from acetic acid, hydrochloric acid, hydroiodic acid, phosphoric acid, nitric acid, sulfuric acid, citric acid, lactic acid, succinic acid, maleic acid, malic acid, isethionic acid, fumaric acid, benzenesulfonic acid, toluenesulfonic acid, sulfuric acid, methanesulfonic acid (mesylic acid), ethanesulfonic acid, naphthalenesulfonic acid, valeric acid, propanoic acid, butanoic acid, malonic acid, glucuronic acid, and lactobionic acid. One particular salt is a hydrochloride salt. Another particular salt is an acetate salt.

[0058] The compound is anionic or has a functional group that can be anionic (for example, -COOH is -COO - If possible, a salt can be formed with an organic or inorganic base to generate a suitable cation. An example of a suitable inorganic cation is Li + kaNa + , and K + Alkali metal ions such as Ca 2+ and Mg 2+ Alkaline earth metal cations such as Al 3+ or Zn + Other cations include, but are not limited to, those listed above. A suitable example of an organic cation is the ammonium ion (i.e., NH4). + ) and substituted ammonium ions (e.g., NH3R + NH2R2 + NHR3 + NR4 + Examples of suitable substituted ammonium ions include, but are not limited to, those derived from methylamine, ethylamine, diethylamine, propylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids such as lysine and arginine. A common example of a quaternary ammonium ion is N(CH3)4 + That is the case.

[0059] If the peptides of the present invention contain amine functions, they can form quaternary ammonium salts by reaction with alkylating agents, for example, by methods well known to those skilled in the art. Such quaternary ammonium compounds are within the scope of the peptides of the present invention.

[0060] (Modified derivative) Modified derivatives of peptide ligands as defined herein will be understood to be within the scope of the present invention. Examples of such suitable modified derivatives include: N-terminal and / or C-terminal modifications; substitution of one or more amino acid residues with one or more unnatural amino acid residues (e.g., substitution of one or more polar amino acid residues with one or more isopartic or isoelectronic amino acids; substitution of one or more nonpolar amino acid residues with other unnatural isopartic or isoelectronic amino acids); addition of spacer groups; substitution of one or more oxidation-sensitive amino acid residues with one or more oxidation-resistant amino acid residues; substitution of one or more amino acid residues with one or more substituted amino acids, e.g., alanine; substitution of one or more L-amino acid residues with one or more D-amino acid residues; N-alkylation of one or more amide bonds in a bicyclic peptide ligand; substitution of one or more peptide bonds by substitute bonds; modification of peptide backbone length; Examples of modifications include substitution of a hydrogen atom on the α-carbon of one or more amino acid residues with another chemical group; modification of an amino acid such as cysteine, lysine, glutamic acid / aspartic acid, and tyrosine with a suitable amine, thiol, carboxylic acid, and phenol-reactive reagent to functionalize the amino acid; and introduction or substitution of an amino acid having an azide or alkyne group that introduces orthogonal reactivity suitable for functionalization, such as an alkyne or azide moiety.

[0061] In one embodiment, the modified derivative includes N-terminal and / or C-terminal modifications. In a further embodiment, the modified derivative includes N-terminal modifications using a preferred amino reaction chemistry and / or C-terminal modifications using a preferred carboxy reaction chemistry. In a further embodiment, the N-terminal or C-terminal modification includes, but is not limited to, the addition of an effector group, including a cytotoxic agent, a radiochelating agent, or a chromophore.

[0062] In a further embodiment, the modified derivative includes an N-terminal modification. In a further embodiment, the N-terminal modification includes an N-terminal acetyl group. In this embodiment, the N-terminal residue is capped with acetic anhydride or other suitable reagent during peptide synthesis, resulting in a molecule with an acetylated N-terminus. This embodiment offers the advantage of removing a potential recognition site for aminopeptidases and avoiding the possibility of degradation of the bicyclic peptide.

[0063] In an alternative embodiment, the N-terminal modification includes the addition of a molecular spacer group that facilitates the conjugation of the effector group and the retention of the bicyclic peptide's efficacy against its target.

[0064] In a further embodiment, the modified derivative includes a C-terminal modification. In a further embodiment, the C-terminal modification includes an amide group. In this embodiment, the C-terminal residue is synthesized as an amide during peptide synthesis, resulting in a molecule with an amidated C-terminus. This embodiment offers the advantage of removing a potential recognition site for carboxypeptidases, thereby reducing the potential for proteolysis of the bicyclic peptide.

[0065] In one embodiment, the modified derivative includes the substitution of one or more amino acid residues with one or more non-natural amino acid residues. In this embodiment, non-natural amino acids having isodistributed / isoelectronic side chains that are neither recognized by degradable proteases nor have any adverse effects on target efficacy may be selected.

[0066] Alternatively, non-natural amino acids having constrained amino acid side chains may be used so that proteolytic hydrolysis of nearby peptide bonds is sterically and sterically inhibited. In particular, these relate to cycloamino acids, which are proline analogs, bulky side chains, Cα-disubstituted derivatives (e.g., aminoisobutyric acid, Aib), and simple derivatives of amino-cyclopropylcarboxylic acids.

[0067] In one embodiment, the modified derivative includes the addition of a spacer group. In a further embodiment, the modified derivative is an N-terminal cysteine ​​(C i ) and / or C-terminal cysteine ​​(C iii This includes adding a spacer group to ).

[0068] In one embodiment, the modified derivative includes the substitution of one or more oxidation-sensitive amino acid residues with one or more oxidation-resistant amino acid residues. In a further embodiment, the modified derivative includes the substitution of a tryptophan residue with a naphthylalanine or alanine residue. This embodiment offers the advantage of improving the pharmaceutical stability profile of the resulting bicyclic peptide ligand.

[0069] In one embodiment, the modified derivative comprises the substitution of one or more charged amino acid residues with one or more hydrophobic amino acid residues. In an alternative embodiment, the modified derivative comprises the substitution of one or more hydrophobic amino acid residues with one or more charged amino acid residues. The correct balance of charged and hydrophobic amino acid residues is an important characteristic of bicyclic peptide ligands. For example, hydrophobic amino acid residues affect the degree of plasma protein binding and, therefore, the concentration of the available free fraction in plasma, while charged amino acid residues (in particular, arginine) may affect the interaction between the peptide and the phospholipid membrane on the cell surface. The combination of these two may affect the half-life, volume of distribution, and exposure of the peptide drug and can be adjusted according to the clinical endpoint. Furthermore, the correct combination and number of charged and hydrophobic amino acid residues can reduce irritation at the injection site (when the peptide drug is administered subcutaneously).

[0070] In one embodiment, the modified derivative includes the substitution of one or more L-amino acid residues with one or more D-amino acid residues. This embodiment is thought to enhance the stability of protein degradation due to steric hindrance and the tendency of D-amino acids to stabilize the β-turn stereostructure (Tugyi et al. (2005) PNAS, 102(2), 413-418).

[0071] In one embodiment, the modified derivative includes the removal of any amino acid residue and substitution with alanine such as D-alanine. This embodiment has the advantage of identifying key binding residues and eliminating potential proteolytic attack sites.

[0072] It should be noted that each of the modifications described above plays a role in intentionally improving the potency or stability of the peptide. Further potency enhancements based on modifications can be achieved through the following mechanisms: - To achieve higher affinity, incorporate hydrophobic moieties that utilize hydrophobic effects and result in lower dissociation rates; - Utilizing long-range ion interactions to achieve faster association rates and incorporate charged groups that provide higher affinity (see, for example, Schreiber et al., Rapid, electrostatically assisted association of proteins (1996), Nature Struct. Biol. 3, 427-31); and - For example, by correctly constraining the side chains of amino acids so that entropy loss is minimized at target binding, by constraining the torsion angle of the skeleton so that entropy loss is minimized at target binding, and by introducing further cyclization within the molecule for the same reasons, thereby incorporating further constraints into the peptide. (For review articles, see Gentilucci et al., Curr. Pharmaceutical Design, (2010), 16, 3185-203, and Nestor et al., Curr. Medicinal Chem (2009), 16, 4399-418).

[0073] (Isotope variations) The present invention includes all pharmaceutically acceptable (radioactive) isotope-labeled peptide ligands of the present invention, in which one or more atoms are replaced by atoms having the same atomic number but having an atomic mass or mass number different from that commonly found in nature; peptide ligands of the present invention (referred to as "effectors") to which a metal chelate group capable of holding the relevant (radioactive) isotope is attached; and peptide ligands of the present invention in which a specific functional group is covalently replaced by the relevant (radioactive) isotope or an isotope-labeled functional group.

[0074] Examples of isotopes suitable for inclusion in the peptide ligands of the present invention include hydrogen isotopes, for example, 2 H(D) and 3 H(T), an isotope of carbon, for example, 11 C, 13 C and 14 C, an isotope of chlorine, for example, 36 Cl, an isotope of fluorine, for example, 18 F, an isotope of iodine, for example, 123 I, 125 I, and 131 I. Isotopes of nitrogen, for example, 13 N and 15 N, an isotope of oxygen, for example, 15 O, 17 O, and 18 O, an isotope of phosphorus, for example, 32 P, an isotope of sulfur, for example; S, an isotope of copper, for example. 64 Isotopes of Cu and gallium, for example, 67 Ga or 68 Ga, an isotope of yttrium, for example, 90 Y, and lutetium isotopes, for example, 177 Lu, and bismuth isotopes, for example, 213 Includes Bi.

[0075] The specific isotope-labeled peptide ligands of the present invention, for example, those incorporating radioisotopes, are useful in studies of the tissue distribution of drugs and / or substrates, and for clinically assessing the presence and / or absence of targets on affected tissue. The peptide ligands of the present invention may further possess valuable diagnostic properties in that they can be used to detect or identify the formation of complexes between labeled compounds and other molecules, peptides, proteins, enzymes, or receptors. The detection or identification method may use compounds labeled with labeling agents such as radioisotopes, enzymes, fluorescent substances, or luminescent substances (e.g., luminol, luminol derivatives, luciferin, aequorin, and luciferase). For example, the radioisotope tritium, i.e., 3 H(T) and carbon-14, that is, 14 C is particularly useful for this purpose, given its ease of implementation and the availability of detection means.

[0076] Deuterium, that is, 2 Substitution with heavier isotopes, such as H(D), may result in certain therapeutic benefits, such as greater metabolic stability, an increased in vivo half-life, or a reduced required dosage, and therefore may be preferable in some situations.

[0077] 11 C, 18 F, 15 O, and 13 Substitution with positron-emitting isotopes such as 16N may be useful in positron emission topography (PET) studies to investigate target occupancy.

[0078] The isotope-labeled peptide ligand compounds of the present invention can typically be prepared by conventional techniques known to those skilled in the art, or by processes similar to those described in the attached examples, using appropriate isotope-labeling reagents instead of previously used unlabeled reagents.

[0079] (Molecular scaffold) In one embodiment, the molecular scaffold includes a non-aromatic molecular scaffold. The term "non-aromatic molecular scaffold" as used herein refers to any molecular scaffold as defined herein that does not contain aromatic (i.e., unsaturated) carbocyclic or heterocyclic ring systems.

[0080] A suitable example of a non-aromatic molecule scaffold is described in Heinis et al.'s (2014) Angewandte Chemie, International Edition 53(6) 1602-1606.

[0081] As described in the aforementioned document, the molecular scaffold may be made of low-molecular-weight molecules, such as low-organic molecules.

[0082] In one embodiment, the molecular scaffold may be a polymer. In one embodiment, the molecular scaffold is a polymer composed of amino acids, nucleotides, or carbohydrates.

[0083] In one embodiment, the molecular scaffold includes a reactive group that can react with the functional group of the polypeptide to form a covalent bond.

[0084] The molecular scaffold may contain chemical groups that form links with peptides, such as amines, thiols, alcohols, ketones, aldehydes, nitriles, carboxylic acids, esters, alkenes, alkynes, azides, anhydrides, succinimides, maleimides, alkyl halides, and acyl halides.

[0085] In one embodiment, the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (also known as triacryloylhexahydro-s-triazine (TATA)): [ka] That is the case.

[0086] Therefore, C i , C ii , and C iii After cyclization with the bicyclic peptide of the present invention on a cysteine ​​residue, the molecular scaffold has the following structure: [ka] (Here, * (This represents the binding site of three cysteine ​​residues.) It forms a trisubstituted 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropan-1-one derivative of TATA having the following properties.

[0087] In an alternative embodiment, the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tris(2-bromoetanone)(TATB).

[0088] Therefore, C i , C ii , and C iii After cyclization with the bicyclic peptide of the present invention on a cysteine ​​residue, the molecular scaffold has the following structure: [ka] This forms a trisubstituted derivative of TATB having the following properties.

[0089] (synthesis) The peptides of the present invention can be synthesized using standard techniques and then reacted in vitro with a molecular scaffold. Standard chemistry can be used when carrying this out. This allows for the rapid, large-scale preparation of soluble materials for further downstream experiments or validation. Such methods can be achieved using conventional chemistry, such as that disclosed in the literature by Timmerman et al. (above).

[0090] Accordingly, the present invention also relates to the production of a polypeptide or conjugate selected as described herein, wherein the production includes any further steps as described below. In one embodiment, these steps are carried out on a final product polypeptide / conjugate produced by chemical synthesis.

[0091] Optionally, amino acid residues in the target polypeptide may be substituted when producing the conjugate or complex.

[0092] By extending the peptide, for example, another loop can be incorporated, and therefore multiple specificities can be introduced.

[0093] To extend the peptide, it may simply be chemically extended at its N-terminus or C-terminus or within the loop using orthogonally protected lysine (and analogues) with standard solid-phase or liquid-phase chemistry. An activated or activatable N- or C-terminus may be introduced using standard (bio)conjugation techniques. Alternatively, the addition may be carried out enzymatically using a subtilis gase, for example, as described in (Dawson et al., 1994, Synthesis of Proteins by Native Chemical Ligation. Science 266:776-779), or using a subtilis gase, for example, as described in (Chang et al., Proc Natl Acad Sci US A. 1994 Dec 20; 91(26):12544-8 or Hikari et al., Bioorganic & Medicinal Chemistry Letters, Vol. 18, No. 22, November 15, 2008, pp. 6000-6003).

[0094] Alternatively, the peptide may be extended or modified by further conjugation via disulfide bonds. This has the additional advantage of allowing the first and second peptides to dissociate from each other in the reducing environment of the cell. In this case, a molecular scaffold (e.g., TATA or TATB) can be added during the chemosynthesis of the first peptide to react with three cysteine ​​groups; thereafter, an additional cysteine ​​or thiol can be added to the N or C-terminus of the first peptide, so that this cysteine ​​or thiol reacts only with the free cysteine ​​or thiol of the second peptide to form a disulfide-bonded bicyclic peptide-peptide conjugate.

[0095] Furthermore, the addition of other functional groups or effector groups may be achieved in the same manner by coupling at the N- or C-terminus or via the side chain using appropriate chemistry. In one embodiment, the coupling is carried out in such a manner that it does not block the activity of any of the entities.

[0096] (Pharmaceutical composition) A further aspect of the present invention provides a pharmaceutical composition comprising a peptide ligand as defined herein in combination with one or more pharmaceutically acceptable excipients.

[0097] Typically, the peptide ligand is used in a purified form with a pharmacologically appropriate excipient or carrier. These excipients or carriers typically include aqueous or alcohol / aqueous solutions, emulsions, or suspensions containing physiological saline and / or a buffering medium. Parenteral vehicles include sodium chloride solution, Ringer's dextrose, dextrose, and sodium chloride, as well as lactated Ringer's dextrose. A physiologically acceptable adjuvant may be selected from thickeners such as carboxymethylcellulose, polyvinylpyrrolidone, gelatin, and alginates, if necessary to maintain the polypeptide complex in suspension.

[0098] Intravenous vehicles include fluids, nutritional supplements, and electrolyte supplements, such as those based on Ringer dextrose. Preservatives and other additives, such as antimicrobial agents, antioxidants, chelating agents, and inert gases, may also be present (Mack (1982), Remington's Pharmaceutical Sciences, 16th edition).

[0099] The peptide ligands of the present invention may be administered separately as a composition or in combination with other agents. These include antibodies, antibody fragments, and various immunotherapeutic agents, such as cyclosporine, methotrexate, adriamycin, or cisplatin, and immunotoxins. Further examples of other agents that may be administered separately or in combination with the peptide ligands of the present invention include cytokines, lymphokines, other hematopoietic factors, thrombolytic factors, and antithrombotic factors. The pharmaceutical composition may include a "cocktail" of various cytotoxic agents or other agents in combination with the protein ligands of the present invention, or even a combination of selected polypeptides according to the present invention having different specificities, such as polypeptides selected using different target ligands, whether pooled or not before administration.

[0100] The route of administration of the pharmaceutical composition according to the present invention may be any of those commonly known to those skilled in the art. For therapeutic purposes, the peptide ligand of the present invention can be administered to any patient according to standard techniques. Administration may be by any suitable mode, including parenteral, intravenous, intramuscular, intraperitoneal, percutaneous, pulmonary routes, or, as appropriate, direct infusion using a catheter. Preferably, the pharmaceutical composition according to the present invention is administered intravenously. The dosage and frequency of administration will depend on the patient's age, sex, and condition, concurrent administration of other drugs, contraindications, and other parameters considered by the clinician.

[0101] The peptide ligands of the present invention can be lyophilized for storage and reconstituted in a suitable carrier before use. This technique has been shown to be effective, and lyophilization and reconstitution techniques known in the art can be utilized. It will be understood by those skilled in the art that lyophilization and reconstitution may result in varying degrees of activity loss, and that it may be necessary to adjust the levels upward to compensate for this.

[0102] Compositions containing the peptide ligand or cocktail thereof of the present invention may be administered for prophylactic and / or therapeutic purposes. In a particular therapeutic use, an amount sufficient to achieve at least partial inhibition, suppression, regulation, death, or any other measurable parameter of a selected population of cells is defined as the “therapeutic effective dose.” The amount required to achieve this dose depends on the severity of the disease, but is generally in the range of 0.005 to 5.0 mg of the selected peptide ligand per kilogram of body weight, with doses of 0.05 to 2.0 mg / kg being more commonly used. For prophylactic purposes, compositions containing the peptide ligand or cocktail thereof may also be administered in similar or slightly lower doses.

[0103] Compositions containing peptide ligands according to the present invention can be used in prophylactic and therapeutic settings to assist in altering, inactivating, killing, or removing selective target cell populations in mammals. Furthermore, the peptide ligands described herein can be selectively used in vitro or in vitro to kill, deplete, or otherwise effectively remove target cell populations derived from heterogeneous cell aggregates. Mammalian blood can be combined with selected peptide ligands in vitro to kill or otherwise remove unwanted cells from the blood for return to the mammal according to standard techniques.

[0104] (therapeutic use) The bicyclic peptide of the present invention has specific utility as a transferrin receptor 1 (TfR1) conjugate. Further aspects of the present invention provide peptide ligands or pharmaceutical compositions as defined herein for use in preventing, suppressing, or treating diseases or disorders by the delivery of therapeutic agents mediated by TfR1.

[0105] Transferrin is a glycoprotein found in vertebrates that binds to iron (Fe) and, as a result, mediates the transport of iron (Fe) from blood plasma. It is produced in the liver and contains two Fe molecules. 3+ It contains atomic bonding sites. Human transferrin is encoded by the TF gene and produced as a 76 kDa glycoprotein.

[0106] Transferrin glycoprotein binds to iron firmly but reversibly. While the iron bound to transferrin accounts for less than 0.1% (4 mg) of the body's total iron, it forms the most vital iron pool, possessing the highest turnover rate (25 mg / 24 hours). Transferrin has a molecular weight of approximately 80 kDa and contains two specific high-affinity Fe(III) binding sites. Transferrin has extremely high affinity for Fe(III) (association constant is 10 at pH 7.4). 20 M -1 Transferrin, however, decreases the pH below neutral and gradually decreases. Transferrin is not limited to binding to iron, but also to various metal ions. These glycoproteins are found in various body fluids of vertebrates. When not bound to iron, transferrin is known as "apotransferrin."

[0107] In one embodiment, transferrin is mammalian transferrin. In a further embodiment, mammalian transferrin is human transferrin. In one embodiment, human transferrin is human transferrin receptor 1 (TfR1; also known as CD71).

[0108] It will be understood that TfR1-binding peptides may be useful in the treatment of neurological disorders. Examples of such neurological disorders include, but are not limited to, neuropathies, neurodegenerative diseases, cancer, ocular disorders, paroxysmal disorders, lysosomal storage disorders, amyloidosis, viral or microbial diseases, ischemia, behavioral disorders, and CNS inflammation.

[0109] In one embodiment, the neurological impairment is in a human subject. It will be understood that the dosage and / or frequency of administration are adjusted to reduce the concentration of the peptide ligand to which the red blood cells are exposed. In a further embodiment, the treatment further includes the step of monitoring the human subject for red blood cell removal.

[0110] In this specification, the term “prevention” includes the administration of a protective composition before the induction of disease. “Suppression” refers to the administration of a composition after an inducible event but before the clinical manifestation of the disease. “Treatment” includes the administration of a protective composition after disease symptoms have become apparent.

[0111] Animal model systems are available that can be used to screen the efficacy of peptide ligands in the protection against or treatment of disease. The use of animal model systems is facilitated by the present invention, thereby enabling the development of polypeptide ligands that can cross-react with human and animal targets, and enabling the use of animal models.

[0112] Transferrin receptor 1 (TfR1) is a widely studied model receptor-ligand system that provides considerable insight into cellular properties and the mechanisms of nutrient / scavenger receptor cargo internalization and endocytosis sorting (Qian et al. (2002) Pharmacological Reviews 54(4), 561-587). TfR1 is known to undergo constitutive endocytosis and recycling to the plasma membrane and possesses pH-dependent ligand binding to enable proper sorting of endocytized cargo. While anti-TfR1 antibodies have been considered the primary agents for TfR1 targeting with oligonucleotide therapies, the TfR1-binding peptide ligand of the present invention may exhibit efficient and significant knockdown of gene expression in skeletal and cardiac muscle via systemic delivery of the TfR1-bicyclic peptide-siRNA conjugate.

[0113] Therefore, considering this mechanism, the peptide ligand of the present invention is thought to be useful as a tissue delivery complex, for example, for the delivery of the Tfr1-peptide ligand-payload (i.e., siRNA) complex to tissue cells, particularly muscle cells.

[0114] Therefore, according to a further aspect of the present invention, a tissue delivery complex is provided comprising the peptide ligand of the present invention bound to TfR1 in combination with another peptide, small molecule drug, or oligonucleotide, in particular, a payload such as siRNA.

[0115] Therefore, the tissue delivery complex finds usefulness in the treatment of musculoskeletal disorders. Suitable examples of musculoskeletal disorders include: 12q14 microdeletion syndrome 2q37 deletion syndrome 3M syndrome Absence of the tibia Tibial deficiency accompanied by polydactyly Patella absence Anomalism Achonzodysplasia type 1A - See Achonzodysplasia Achonzodysplasia type 1B - See Achonzodysplasia Achonzodysplasia type 2 - See Achonzodysplasia achondroplasia Acute, thoracic, and renal field defects Acromiocorpus callosum syndrome, Schinzel type Apical femoral head dysplasia acral polydactyly acrodysostosis Acropora dysplastic scoliosis Acropacial dysostosis, Catania type Acropacial dysostosis, paragonia type Acropacial dysostosis, Rodriguez type Acroporafrontofacial rhinoplasty Acroporasal dysostosis Acropora and midline anterior dysplasia Acropora and midline anterior dysplasia, Hunter-Thompson type Acropora and midline anterior dysplasia, Maroto type acrobrachydysplasia Acromiolysis, dominant type acrothoracic syndrome Apical thoracic spondylodysplasia, type F Acute febrile neutrophilic dermatosis Unilateral adactyly Adams-Oliver syndrome Adenosine deaminase 2 deficiency ADULT syndrome Adult-onset Still's disease Ecardi-Goutier syndrome Alghazari Sabrina Tannile Syndrome Alain Babin De Marquez Syndrome α-mannosidosis Scapulofibular neurogenic muscular atrophy, New England type ahypertrophic dysplasia Angeloid phalangeal epiphyseal dysplasia Eyelid adhesions, ectodermal defects, cleft lip and palate syndrome Ankylosing spondylitis - Not a rare disease Hyperossification of the ankylosing spondylosis with calluses Onychomycosis / onychoplasia with hypoplasia or absence of distal phalanges Antley-Bixler syndrome Apert syndrome Congenital multiple arthral contractures Arts syndrome Aspartylglucosamineuria Osteogenesis imperfecta type 1 Osteogenesis imperfecta type 2 Osteogenesis imperfecta type 3 Otocephalic syndactyly auricular condylar syndrome Auricular dysplasia Autosomal dominant late-onset spondyloepiphyseal dysplasia Autosomal recessive early-onset inflammatory bowel disease Autosomal recessive protein C deficiency axial osteomalacia Axial spondylometaphyseal dysplasia Baby rattle pelvic dysplasia Bohler-Gerrold syndrome Banki syndrome Behle-Stevenson gyrus of the scalp Behcet's disease Benaleg-Lacete syndrome Vethlemmyopathy Bueks familial hip dysplasia Blau syndrome Blount's disease BOD syndrome Osteodysplasia, Azuz type Osteodysplasia, lethal Holmgren type Boomerang dysplasia Forward curvature of the legs associated with dwarfism Brachyfrontal nasal dysplasia Brachydactyly dwarfism, Museleni type Brachydactyly (shoulder, elbow, and wrist) Brachydactyly, long thumb type Brachydactyly, mononen type Brachydactyly type A1 Brachydactyly type A2 Brachydactyly type A4 Brachydactyly type A5 Brachydactyly type A6 Brachydactyly type A7 Brachydactyly type B Brachydactyly type C Brachydactyly type E Brachydactyly B and E mixed type Brachytaxial type 3 Branchial arch syndrome, X-linked Brody Myopathy Brooks syndrome 1 Bushke-Olendorf syndrome C syndrome Caffe's disease Limb flexion disorder, cumming type flexural limb dysplasia brachydactyly Flexor flexor, arthropathy, coxa vara, pericarditis syndrome Finger flexion syndrome, Guadalajara type 2 Dermatitis, hypertension, and hearing loss syndrome Kamrathi-Engelmann disease Cantu syndrome Carpenter syndrome carpal tarsal osteochondromatosis cartilage hypoplasia Cater Manzquet syndrome Cerebellar hypoplasia with endosteal sclerosis Brain-rib-mandibular syndrome Cervical dystonia Charlie M syndrome Cherubimic disease CHILD syndrome Childhood hypophosphatasia Chondrocalcinosis 2 Chondrodysplasia, Blomstrand type Chondrodysplasia punctata 1, X-linked recessive Chondrodysplasia punctata, Sheffield type Chondrodysplasia with joint dislocation, GPAPP type Chondrodysplasia, Glebe type Chondrosarcoma Chordoma Chronic atypical neutrophilic dermatosis with lipodystrophy and elevated body temperature Chronic relapsing multifocal osteomyelitis Tibial bone defect, split hand cleidocranial dysplasia Obstecranial dysplasia, recessive type Clavicular limb root syndrome CLOVES syndrome Coccyx pain CODAS syndrome Coffin-Siris syndrome COG1-CDG (CDG-IIg) Cole Carpenter Syndrome Collagen disorder type 2α1 Condensed osteitis of the clavicle Congenital adrenal hyperplasia due to cytochrome P450 oxidoreductase deficiency Congenital contracture of the arachnodactyly Congenital femoral deficiency Congenital primary aphakia Congenital radioulnar synostosis Cornelia de Lange syndrome Cousin Syndrome Craniotruncal dysplasia Extracephalic dysplasia Craniofacial dysostosis with diaphysis hyperplasia Craniofacial dysfunction Cranioporatonar dysplasia Craniometrial dysplasia, autosomal dominant type Craniometrial dysplasia, autosomal recessive type Craniosynostosis, anal abnormalities, and porokeratosis Craniotelenial dysplasia Crouzon syndrome Collar-Jones syndrome Clarino Tricolor Currie-Jones syndrome Czech dysplasia, metatarsal type Dandy-Walker malformation with postaxial polydactyly Dandy-Walker malformation with sagittal craniosynostosis and hydrocephalus Deficiency of interleukin-1 receptor antagonists Late-onset membranous ossification Dentate nucleobyl pallidolus atrophy Debuqueror Syndrome Desmosterol disease Diaphyseal medulla stenosis associated with malignant fibrous histiocytoma Torsional osteodysplasia Dihydropyrimidine dehydrogenase deficiency - not a rare disease Digbe-Melchior-Clausen syndrome Chondroosteodysplastic nephritis Disferlinopathy Heterotopic osteosclerosis Unilateral epiphyseal dysplasia Segmental dysplasia, Roland-Desbuquois type Segmental dysplasia, Silverman-Handmaker type DYT-GNAL EEC syndrome EEM syndrome Ehlers-Van Creveld syndrome Juvenile idiopathic arthritis associated with enthesitis Epidermolysis bullosa simplex with muscular dystrophy Multiple epiphyseal dysplasia associated with early-onset diabetes Erdheim-Chester disease Ewing sarcoma Familial avascular necrosis of the femoral head Familial cold autoinflammatory syndrome Familial hypocalciuric hypercalcemia type 1 Familial hypocalciuric hypercalcemia type 2 Familial hypocalciuric hypercalcemia type 3 Familial Mediterranean fever Familial dyschondrosteosis Familial tumoral calcinosis Fanconi anemia Finegold syndrome Felty syndrome Femoral-facial syndrome Bifid femur with monodactyly Femoral-fibular-ulnar syndrome Fetal thalidomide syndrome Fibrochondrogenesis Progressive fibrodysplasia ossificans Fibular aplasia with ectrodactyly Fibular aplasia, tibial bowing, and syndactyly syndrome Fibular agenesis Fibular hypoplasia and complex brachydactyly Filippi syndrome Fitzsimons-Gilbert syndrome Nodular segmental glomerulosclerosis Frank Ter Haar syndrome Freiberg disease Frontofacial rhinoplasia Frontal metaphyseal dysplasia frontonasal dysplasia Frontorhinostomy with alopecia and genital abnormalities - See Frontorhinostomy Frontorhinoplasia, Severe Microphthalmia, Severe Facial Cleft Syndrome - See Frontorhinoplasia See Frontorhiny - Frontornasal Dysplasia Flynn-Hoffkens-Fabry syndrome Fucosidosis Foolman syndrome Galactosialidosis Gaucher disease type 1 Gaucher disease type 3 Ecstatic Dwarfism Genital patellar syndrome Genoa syndrome Hereditary chondromatosis osteodysplastic senile dermatosis Gosal hematodiaphyseal dysplasia syndrome giant cell tumor of bone GM1 gangliosidosis type 1 GM1 gangliosidosis type 2 GM1 gangliosidosis type 3 Goldenhar disease Gorham's disease thin bone dysplasia Grant syndrome Greenberg dysplasia Craig's polysyndactyly syndrome Gurieri syndrome Harrahman-Strife syndrome Hand-foot-uterine syndrome Hanhart syndrome Slovenian type of cardiac-hand syndrome Hand syndrome, Spanish type Hemifacial dwarfism Hemifacial hyperplasia Hereditary antithrombin deficiency Hereditary multiple osteochondroma Holt-Oram syndrome Hunter-MacAlpine syndrome Hurler syndrome Hurler-Scheie syndrome Mucopolysaccharidosis syndrome Hyper IgD syndrome Generalized cortical hyperostosis Hyperphosphatemic familial tumoral calcinosis Chondrodysplasia Hypophosphatasia Hypophosphatemic rickets I-cell disease IMAGe syndrome Imperforate pharynx and costovertebral anomalies Inclusion body myopathy 3 Early-onset Paget's disease and inclusion body myopathy with frontotemporal dementia Inclusion body myositis Intellectual disability, contractures, and ectodactyly syndrome Iris-corneal angle dysgenesis type 1 IVIC syndrome Jackson-Weiss syndrome Jansen type metaphyseal chondrodysplasia June syndrome Johnson-Manson syndrome Juvenile dermatomyositis Juvenile osteoporosis Juvenile Paget's disease Kaplan-Prautchett-Fitch syndrome Kenny-Caffey syndrome type 1 Kenny-Caffey syndrome type 2 Keutel syndrome Kienböck disease Klippel-Hommes syndrome Klippel-Feil syndrome Klippel-Trenaunay syndrome Kniest dysplasia Kniest-like dysplasia, lethal Köhler disease Posteriorly angulated limb dysplasia Lacrimal-ear-tooth-finger syndrome Lambda type synostosis Lambert-Eaton myasthenic syndrome Langer mesomelic shortening dysplasia Larsen syndrome Lateral meningocele syndrome Laurin Sandrow syndrome Legg-Calvé-Perthes disease Lenz-Maszieski hyperosteal dwarfism Reri hyperossification Leri-Weil chondrodysplasia Fatal chondrodysplasia, Moerman type Fatal chondrodysplasia, Seller type Levator muscle syndrome Limb-girdle muscular dystrophy type 1A Limb-girdle muscular dystrophy type 2A Limb-girdle muscular dystrophy type 2B Limb-girdle muscular dystrophy type 2E Limb-girdle muscular dystrophy type 2F Limb-girdle muscular dystrophy type 2H Limb-girdle muscular dystrophy, type 2C Limb-girdle muscular dystrophy, type 2D limb breast syndrome Loeys-Dietz syndrome Laurie MacLean Syndrome Lorey Wood Syndrome Macrophage fasciitis Muffucci syndrome MAGIC syndrome Magid syndrome Mandibular dysplasia with type A adipose dysplasia Mandibular dysplasia with type B adipose dysplasia Mandibular and facial dysostosis with microcephaly Mannosidosis, βA, lysosomal type Marshall syndrome Marshall-Smith syndrome McCune-Albright syndrome Meckel syndrome Median cleft upper lip accompanied by polyps of the facial skin and nasal mucosa. Mayer-Gorlin syndrome Melnick Needles syndrome waxy osteosis Familial osteopathy with osteoporosis Mid-columbar shortening and osteofusion syndrome Mid-columnar dwarfism, cleft palate, and flexed fingers Cantaputra type of mid-septal anterior dysplasia Midcostomy dysplasia, Savalayan type Fourth and fifth metacarpal synostosis Chondromatosis Metaphyseal epiphyseal dysplasia Schmidt type metaphyseal chondrodysplasia Chondrodysplasia of the epiphyseal region, spur type Metaphyseal chondrodysplasia, intellectual disability, conductive hearing loss syndrome Metaphyseal dysplasia / micromaxilla / brachydactyly Metaphyseal dysplasia without trichothyroidism Degenerative dysplasia of bone Mevalonic aciduria Microcephalic osteodysplastic primitive dwarfism type 1 Microcephalic osteodysplastic primitive dwarfism type 2 Microcephalic primitive dwarfism, Torriero type Hemifacial dwarfism and radial agenesis Mirror syndrome Minicoar myopathy with extraocular muscle paralysis Monomer muscular atrophy Mackle-Wells syndrome Mucolipidosis IIIα / β Mucolipidosis type 4 Mucopolysaccharidosis type III Mucopolysaccharidosis type IIIA Mucopolysaccharidosis type IIIB Mucopolysaccharidosis type IIIC Mucopolysaccharidosis type IIID Mucopolysaccharidosis type IV Mucopolysaccharidosis type IVA Mucopolysaccharidosis type VII Muwenke syndrome Multicentric carpotarsal osteolysis syndrome Multiple epiphyseal dysplasia Multiple epiphyseal dysplasia 2 Multiple sulfatase deficiency Multiple osteosynostosis syndrome 1 multiple system atrophy Muscular dystrophy Muscular dystrophy, congenital, megaconus type MYH7-related scapulofibular myopathy Mayer syndrome Myosinopathy Myostatin-related muscle hypertrophy Myotonic dystrophy Myotonic dystrophy type 2 Najjer acrofacial dysostosis Nail-patella syndrome Nakajo-Nishimura Syndrome Neonatal-onset multisystem inflammatory disease Severe hyperparathyroidism in newborns Nestor Guillermo syndrome Neurofibromatosis type 1 Nibelgaard syndrome Familial tumor-like calcification with normal phosphate occipital horn syndrome oculofrontal-nasal syndrome Oculodontodigital dysplasia Oculofacial Dysostosis Oculopharyngeal muscular dystrophy Oliver syndrome Olier's disease Skeletal dysplasia 1 Skeletal dysplasia 2 Delayed Maturation Osteodysplasia Orofacial Finger Syndrome 1 Orofacial Finger Syndrome 10 Orofacial Finger Syndrome 11 Orofacial Finger Syndrome 2 Orofacial Finger Syndrome 3 Orofacial Finger Syndrome 4 Orofacial Finger Syndrome 5 Orofacial Finger Syndrome 6 Orofacial Finger Syndrome 8 Orofacial Finger Syndrome 9 Osram syndrome OSMED syndrome Ossification of the posterior longitudinal ligament of the spine - not a rare disease Familial phalangeal arthropathy osteochondrosis dissecans Familial dysplasia, Anderson type Danks-Maine and Kozlovsky's precocious dysplasia osteofibrous dysplasia Osteogenesis imperfecta type 1 Osteogenesis imperfecta type II Osteogenesis imperfecta type III Osteogenesis imperfecta type IV Osteogenesis imperfecta type V Osteogenesis imperfecta type VI Cavity osteodysplasia Intermediate bone density Linear osteopathy with craniosclerosis Osteopenia and hair loss Osteopetrosis, autosomal dominant type 1 Osteopetrosis, autosomal dominant type 2 Osteopetrosis, autosomal recessive type 3 Osteopetrosis, autosomal recessive type 4 Osteopetrosis, autosomal recessive 7 Osteoporosis and dacryocystitis Osteoporosis and oculocutaneous hypopigmentation syndrome Osteoporosis / Pseudoglioma Syndrome Osteosarcoma Ear-palate-finger syndrome type 1 Ear-palate-finger syndrome type 2 Hypertrophic cutaneous periosteal disease Pacman dysplasia Pallister Hall syndrome Congenital paramyotonia Parastremmatic dwarfism PARC syndrome Parkes-Weber syndrome Patterson-Stevenson-Fontaine syndrome Pelvic dysplasia and lower limb joint contractures Periodic fever, aphthous stomatitis, pharyngitis, and adenitis Pfeiffer's type of cardiocranial syndrome Pheasant limb syndrome, external digitia, hearing loss, sinus arrhythmia Pigmented trophoblastic nodular synovitis Piriformis syndrome Lethal skeletal dysplasia of flat vertebrae, Torrance type Polyconus myopathy accompanied by salt cravings Poland Syndrome Polycystic bone disease Polycystic lipomegaly dysplasia with sclerosing leukoencephalopathy Polydactyly / myopia syndrome Polyostotic osteolytic dysplasia, hereditary expansive Potassium-induced myotonia Preaxial deficiency, postaxial polydactyly, and hypospadias Preaxial polydactyly type 1 Preaxial polydactyly type 2 Preaxial polydactyly type 3 Preaxial polydactyly type 4 progeria progressive bone formation Progressive pseudorheumatic dysplasia Protein C deficiency - not a rare disease Proteus syndrome Proximal phalangeal fusion pseudoachondroplasia Pseudoaminopterin syndrome Pseudovolvulus dysplasia Pseudohypoparathyroidism type 1A Pseudohypoparathyroidism type 1C pseudopseudohypoparathyroidism Psoriatic juvenile idiopathic arthritis dysostosis pyknosis pyknotic achondroplasia Pile disease Pyoderma gangrene Septic arthritis, pyoderma gangrenosum, and acne See Radioulnar Synostosis Type 1 - Congenital Radioulnar Synostosis. See Radioulnar Synostosis Type 2 - Congenital Radioulnar Synostosis. Radioulnar synostosis, microcephaly, scoliosis syndrome Rains syndrome Ramon syndrome Lapadelino syndrome Reactive arthritis Renal dysplasia, retinitis pigmentosa, cerebellar ataxia, and skeletal dysplasia Retinal vascular disease with systemic symptoms of cerebral white matter dystrophy Limb root punctate chondrodysplasia type 1 Limb root dysplasia, Patterson-Lowry type extremity root syndrome Ricchieri-Costa da Silva syndrome ankylosing spine syndrome Roberts syndrome Seetzle-Kotzen syndrome See Salah's disease - Free sialic acid storage. SAPHO syndrome Sarcoidosis - Not a rare disease Say Meyer syndrome Sayfield-Caldwell syndrome Scalp defects and retroaxial polydactyly SCARF syndrome Chaillet syndrome Scheuermann's disease Schimke immunoosteodysplasia Schinzel-Gideon syndrome Schinzel's phoenix limb disease Cochlear pelvic dysplasia Schnitzler syndrome Schwarz-Jampel syndrome Sclerosing ossification Seckel syndrome Sepiapterin reductase deficiency Short rib polydactyly syndrome type 3 Short rib polydactyly syndrome type 1 Short rib polydactyly syndrome type 4 Short rib polydactyly syndrome, Majooski type Short stature syndrome, Brussels type Sprinzen-Goldberg craniosynostosis syndrome Schwachmann-Diamond Syndrome Sickle-shaped β-thalassemia Sickle cell anemia Silens syndrome Singleton-Merten syndrome Slipped capital femoral epiphysis - not a rare disease lesser patella syndrome Smith-McCourt dysplasia Smith-Lemle-Oppitz syndrome Sotos syndrome Spheroid body myopathy Spinal muscular atrophy, Ryukyuan type Spinal muscular atrophy type 1 with congenital fractures Spinal muscular atrophy type 3 Spinal muscular atrophy type 4 Spinal muscular atrophy type 1 with dyspnea Splenic gland fusion, limb defects, micrognathia Syndactyly Syndactyly, nystagmus Spinal digit flexion Spinal carpal tarsal coalition syndrome Spinal and costal heterotosis 1 - See spinal and costal heterotosis Spinal and costal heterotosis 2 - See spinal and costal heterotosis Spinal and costal heterotosis 3 - See spinal and costal heterotosis Spinal and costal heterotosis 4 - See spinal and costal heterotosis Spinal and costal heterotosis 5 - See spinal and costal heterotosis Spinal and costal heterotosis 6 - See spinal and costal heterotosis Spinal dysplastic Ehlers-Danlos syndrome Intraspinal chondrodysplasia with immunodeficiency Spinal epiphyseal dysplasia, Juvinove type Spinal epiphyseal dysplasia, joint laxity Spinal epiphyseal dysplasia, matrilin-3 related Spinal epiphyseal dysplasia, Missouri type Spinal epiphyseal dysplasia, Shohat type Spinal epiphyseal dysplasia, Spornastrim type Spinal epiphyseal dysplasia, Strudwick type Spinal epiphyseal dysplasia with hypotrichosis Spinal epiphyseal dysplasia with multiple dislocations Spinal epiphyseal dysplasia, X-linked type Spinal epiphyseal dysplasia, aggrecan type Congenital spinal epiphyseal dysplasia Spinal epiphyseal dysplasia, Maroteaux type Late-onset spinal epiphyseal dysplasia, X-linked type Spinal epiphyseal dysplasia, brachydactyly and characteristic speech Spinal epiphyseal dysplasia, short limb, hand type Spinal metaphyseal dysplasia, Algerian type Spinal metaphyseal dysplasia, angular fracture type Spinal metaphyseal dysplasia, Sedaghatian type Spinal metaphyseal dysplasia, A4 type Spinal metaphysical dysplasia with pyramidal-rod dystrophy Vertebrophyseal dysplasia with dentinogenesis imperfecta Spondylometaphyseal dysplasia, X-linked type Spondylometaphyseal dysplasia, Kozlowski type Perisvertebral dysplasia Spondylothoracic dysostosis Sprengel deformation STAR syndrome Stiff Person Syndrome Stub-Wiedemann syndrome Dactylosynostosis with multiple abnormalities in the hands and feet Syndactyly, Sennani-Lenz type Syndactyly type 3 Syndactyly type 5 Syndactyly type 9 Syndactyly, Polydactyly, and Earlobe Syndrome Jaw fusion / multiple abnormalities synovial chondroma Systemic onset juvenile idiopathic arthritis TAR syndrome TARP syndrome Tarsal-carpal syndrome tarsal tunnel syndrome Tetra-amelia syndrome Tetra-amelia syndrome (multiple malformations) Monodactyly (four-limb monodactyly) Fatal osteodysplasia type 1 Fatal osteodysplasia type 2 Thoracic dysplasia / hydrocephalus syndrome Thoracic, laryngeal, and pelvic dysplasia Tibial agenesis, polydactyly, arachnoid cyst Tietze syndrome TMEM165-CDG(CDG-IIk) Townes-Brocks syndrome Treacher Collins syndrome Hair, Teeth, and Bone Syndrome Hair-related liver-collapse syndrome Tricho-nasopharyngeal syndrome type 1 Trichonasophalangeal syndrome type 2 Trichonasophalangeal syndrome type 3 Trigonobrachial syndrome, cleft nasal bulb, micrognathia, and limb abnormalities. Triphalangeal thumb / brachydactyly Humeral trochlear aplasia Trochlear dysplasia Troyer syndrome Tubular aggregate myopathy Tumor necrosis factor-associated periodic fever syndrome Hypoplasia of the ulna and fibula Ulnar hypoplasia and intellectual disability syndrome Ulnar metaphyseal dysplasia syndrome Ulnar hypoplasia / Foot deformity (shrimp claw-like shape) Ulnar-breast syndrome Undifferentiated pleomorphic sarcoma Apington's disease Verlo-Bourguignon syndrome Viljoen-Karis-Vosges syndrome Warman-Mullichen-Hayward Syndrome Weaver syndrome Weil-Marchesani syndrome Weissenbacher-Zweimüller syndrome Weyers type acrofacial dysostosis Wilderwanck syndrome Wirth type autosomal dominant osteosclerosis Wrinkled skin syndrome X-linked dominant chondrodysplasia punctata 2 X-linked dominant scapulofibular myopathy X-linked hypophosphatemia X-linked intellectual disability-plagiocephaly syndrome X-linked skeletal dysplasia - intellectual disability syndrome Eunice Baron syndrome These are some examples, but are not limited to them.

[0116] The present invention will be further described below with reference to the following examples. [Examples]

[0117] (Examples) (Materials and Methods) (Preparation of bicyclic peptide ligands (general method)) Bicyclic peptides were synthesized on Rink amide resin using standard Fmoc (9-fluorenylmethyloxycarbonyl) solid-phase peptide synthesis, either by manual coupling (for large-scale studies) or using a Biotage SyroII automated peptide synthesizer (for small-scale studies). After cleavage from the resin based on TFA, the peptides were precipitated with diethyl ether and dissolved in 50:50 acetonitrile / water. The crude peptides (~1 mM concentration) were then cyclized using ammonium bicarbonate (100 mM) as a base with a 1.3 equivalent scaffold. The completion of cyclization was determined by matrix-assisted laser desorption / ionization time-of-flight (MALDI-TOF) or LC-MS. After completion, the cyclization reaction was quenched with N-acetylcysteine ​​(10 equivalents relative to the peptide), and the solution was lyophilized. The residue was dissolved in a suitable solvent and purified by RP-HPLC. Peptide fractions of sufficient purity and accurate molecular weight (confirmed by MALDI-TOF and either HPLC or LC-MS) were pooled and lyophilized. Concentrations were determined by UV absorption using a decay coefficient at 280 nm based on the Trp / Tyr content.

[0118] Unless otherwise specified, all amino acids were used in their L-stereoconfiguration.

[0119] (Biological data) (1. Direct TfR1 binding assay) The affinity (Kd) of the peptides of the present invention for human or cynomolgus monkey TfR1 was determined using a fluorescence polarization assay according to the following method. The peptides of the present invention were labeled with a fluorescent tag (fluorescein) and diluted to 2.5 nM in 25 mM HEPES, pH 7.4 containing 100 mM NaCl, 4 mM CaCl2, and 0.005% P20. TfR1 protein (human: R&D Systems, 2474-TR or Acro Biosystems, CD1-H5243; cynomolgus monkey: Acro Biosystems, TFR-C524a) was titrated starting from 1 - 5 μM in the same assay buffer as the peptide, and 1 nM peptide was assayed in a total volume of 25 μL in a black-wall and black-bottom low-binding low-volume 384-well plate. This assay was typically prepared by adding 5 μL of assay buffer, 10 μL of TfR1 protein, and then 10 μL of fluorescent peptide. The concentration of TfR1 protein was serially diluted by half to generate 12 different concentrations starting from 1 - 5 μM. Measurements were performed at 25 °C on a BMG PHERAstar FS equipped with an FP 485 520 520 light module, with 200 flashes per well and a positioning delay of 0.1 s. Each well was measured every 5 minutes for 60 minutes. The gain used for analysis was determined for each tracer at the end of 60 minutes when no protein was present in the well. mP was fitted to a standard 1:1 binding model using a quadratic equation to obtain the Kd value. The selected peptides of the present invention were tested in the above assay. The results are shown in Table 1: Table 1: FP direct binding of selected peptide ligands of the present invention

Table 2

[0120] (2. TfR1 SPR binding assay) Biacore experiments were performed to determine the k a (M -1 s -1 )、k d (s -1 )、KD The (nM) value was determined.

[0121] Recombinant human and cynomolgus monkey TfR1 cells were received from two sources as His6-tagged TfR1 cells (aa89~760) (ACRO Biosystems, CD1-H5243 and TFR-C524a).

[0122] For the analysis of TfR1 peptide bonds, a capture / coupling approach using a Cytiva NTA tip was employed with a Biacore T200 or S200 instrument, using 25 mM HEPES, 0.1 M NaCl, and 0.05% Tween 20 pH 7.4 as electrophoresis buffers at 25°C. Immobilization was performed as follows: The tip was pre-equilibrated by injecting 500 mM EDTA (pH 8) before activation with 5 mM NiSO4. The surface was then activated using standard amine coupling chemistry. Briefly, the carboxymethyl dextran surface was activated with a 1:1 ratio of 0.4 M 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) / 0.1 M N-hydroxysuccinimide (NHS). These were then added to electrophoresis buffers and diluted to 200 nM and 250 nM, respectively, before TfR1 protein (human or cynomolgus monkey) was captured on the activated surface. The remaining active groups were blocked by injection of 1M ethanolamine (pH 8.5):HBS-N (1:1) for 7 minutes. When the reference surface was activated and blocked as described above, there was no TfR1 protein capture. Capture levels ranged from 1,500 to 5,000 RU by individual tests. The buffer was replaced with 25 mM HEPES, 0.1 M NaCl, 0.05% Tween 20 pH 7.4, and 1% DMSO.

[0123] A dilution series of the test peptide was prepared in this buffer using a maximum peptide concentration of 5 μM and six further 2-fold dilutions. SPR analysis was performed at 25°C at a flow rate of 30 μl / min with association for 160 seconds and dissociation for 700–800 seconds. Data were corrected for DMSO exclusion volume effect. All data were double-referenced to a blank injection and reference plane using standard processing procedures, and data processing and dynamic fitting were performed using Scrubber software, version 2.0c (BioLogic Software). Data were fitted using a simple 1:1 binding model, taking mass transfer effects into consideration as appropriate.

[0124] The selected peptides of the present invention were tested using the assay described above. The results are shown in Table 2: Table 2: SPR binding of selected peptide ligands of the present invention [Table 3]

[0125] Further selected peptides of the present invention were tested using the assay described above. The results are shown in Table 3: Table 3: SPR binding of selected peptide ligands of the present invention [Table 4] TIFF0007848202000016.tif243170TIFF0007848202000017.tif208170nd=Undecided NB=Not bonded

[0126] (3. TfR1 Inhibition Assay) TfR1 inhibitory activity of the peptide of the present invention (IC 50The following method was used to determine the concentration of the protein, peptide, and alpha reagent: Proteins, peptides, and alpha reagents were prepared to 5× concentrations, and 5 μl of each reagent was added to a white 384-well Optiplate up to a total volume of 25 μl to obtain a 1× final concentration. Fluorescently labeled human transferrin (Invitrogen, T2871) was diluted to 2.5 nM in 25 mM HEPES, pH 7.4, containing 100 mM NaCl, 4 mM CaCl2, 0.5% BSA, and 0.05% P2O. Human or cynomolgus monkey TfR1 protein was diluted to 50 nM, and unlabeled human transferrin (R&D Systems, 2914-HT) was diluted to 500 nM in the same buffer assay. Unlabeled peptides from DMSO stock were diluted 20-fold in the same buffer assay, and then serially diluted by 1 / 3 in a buffer assay containing 5% DMSO to obtain 11 different concentrations. 5 μl of fluorescently labeled transferrin, 5 μl of human or cynomolgus monkey TfR1, and 5 μl of unlabeled peptide or unlabeled human transferrin (R&D Systems, 2914-HT) were added to a white 384-well Optiplate and incubated for 30 minutes. 5 μl of anti-FITC acceptor (PerkinElmer, AL127), diluted 50-fold in assay buffer, was added to the assay plate and incubated for 30 minutes. 5 μl of nickel chelate donor (PerkinElmer, AS101), diluted 50-fold in assay buffer, was added to the assay plate and incubated for 180 minutes. Emission measurements were performed at 25°C using a BMG PHERAstar FS or FSX with an AlphaScreen 520-620 module after excitation at 680 nm. Raw data were normalized to 100 nM unlabeled transferrin and buffer. The data were standardized against 100 nM unlabeled transferrin and buffer controls, and IC50 values ​​were obtained by fitting them to a standard four-parameter fit.

[0127] The selected peptides of the present invention were tested using the assay described above. The results are shown in Table 4: Table 4: Transferrin inhibition assay of selected peptide ligands of the present invention [Table 5] nd=undecided

[0128] (4. TfR1 competitive binding assay) Peptides without a fluorescent tag were tested in competition with a 1 nM peptide (BCY15768) that had a fluorescent tag and a known Kd. The peptides were first diluted in 100% DMSO, then diluted to an appropriate concentration in assay buffer with up to 2.5% DMSO as described in the direct binding assay, and then serially diluted by half. 10 μL of the diluted peptide was added to a plate, and then 10 μL of human TfR1 was added at a constant concentration (200 nM) as described in the direct binding assay. Subsequently, 5 μL of the fluorescent peptide was added. Measurements were performed in the same manner as in the direct binding assay, but the gain was determined before the first measurement. Data analysis was performed using Dotmatics, applying the Cheng-Prusoff equation.

[0129] The selected peptides of the present invention were tested using the assay described above. The results are shown in Table 5: Table 5: TfR1 competitive binding assay of selected peptide ligands according to the present invention [Table 6] TIFF0007848202000020.tif242170TIFF0007848202000021.tif174170

[0130] The selected peptides of the present invention were tested in humans and / or cynomolgus monkeys (TfR1) using the assay described above. The results are shown in Table 6: Table 6: TfR1 competitive binding assay of selected peptide ligands according to the present invention [Table 7] TIFF0007848202000023.tif243170TIFF0007848202000024.tif243170TIFF0007848202000025.tif84170nd=Undecided

[0131] (5. Transcytosis assay using TfR1-linked bicyclic peptide in primary cultures of human proximal tubular cells) To understand the processing of TfR1-linked bicyclic peptides, transepithelial fluxes were measured across the polarization monolayer of human proximal tubular cells. Two fluxes, JAB (absorption-oriented flux) and JBA (secretion-oriented flux), were measured over a 180-minute flux period. From these fluxes, the net direction (absorption or secretion) and magnitude of the TA flux were determined. The experimental details are outlined below: The compound was applied to either the apical or basal side of a confluent monolayer, and the time-resolved distribution of the substrate between these two compartments was monitored to determine the absorption flux (JAB) and secretion flux (JBA), which are the fluxes of TA. From these, the net flux (Jnet) was calculated. The bicyclic peptide was tested at three concentrations: 0.1, 1, and 10 μM. Confluent monolayers were paired so that the monolayers used for measuring absorption flux (JAB) and secretion flux (JBA) had similar TEER values. The culture medium was aspirated from the insert wells before successively transferring the insert into three beakers of approximately 100 ml of warm, modified Krebs buffer. Inserts containing a monolayer of human proximal tubular cells were placed in new 24-well plates, each containing 800 μl of warm modified Krebs buffer at pH 7.4. 200 μl of modified Krebs buffer at pH 7.4 was added to the upper chamber (apical chamber) of the insert. The experimental temperature was maintained at 37°C. • Before starting the fluxing of the test sample, the monolayer was pre-incubated with Krebs buffer alone or with Krebs buffer plus vehicle. The monolayer was incubated with Krebs at pH 7.4 in either the apical or basal membrane, as appropriate. Flux was initiated when the modified Krebs buffer was aspirated from the apical or bottom chamber and replaced with an equivalent volume of the required test concentration of bicyclic peptide at the appropriate pH. This chamber is called the donor chamber. In addition to the bicyclic peptide, Lucifer Yellow containing the same concentration of bicyclic peptide was co-administered to determine the paracellular flux. Subsequently, a 50 μl sample was collected from the opposite chamber (called the receiver chamber) at a predetermined point after the start of the experiment. After gently pipetting twice to mix the buffer, the sample was collected. After each sample was taken, an equal volume of fresh Krebs buffer with the appropriate pH and substrate was replaced. At the time of the final sample, the reaction was terminated by successively transferring the insert to three beakers of ice-cold Krebs buffer and allowed to dry. 50 μl of the sample was stored in a 96-well PCR plate, and 5.6 μl of 0.1% trifluoroacetic acid (TFA) was added to obtain a final concentration of 0.01% TFA. The plate was then flash-frozen in dry ice for storage. The monolayer was dissolved in 50 μl of 0.01% TFA to determine the intracellular volume of the bicyclic peptide, and then flash-frozen as described above. All samples were stored at -80°C. The samples were subjected to LC-MS / MS determination of the bicyclic peptide concentration.

[0132] (6. Detection of bicyclic peptides by LC-MS / MS) A total of 648 samples were received for LC-MS / MS analysis.

[0133] BCY17986, BCY17988, BCY17989, and BCY17994 were provided individually as 1 mg / mL solutions in DMSO. These were further diluted in acetonitrile / DMSO (50 / 50, v / v) to prepare working solutions.

[0134] Bulk calibration standards for BCY17986, BCY17988, BCY17989, and BCY17994 in transporter medium (modified Krebs buffer) with matrix concentrations in the range of 1.00 to 1000 nmol / L were prepared by fortifying the transporter medium with appropriate amounts of BCY17986, BCY17988, BCY17989, and BCY17994 working solutions.

[0135] The donor chamber, receiver chamber, and lysed kidney cell samples were all quantified using bulk calibration standards, and QC samples were prepared in transporter medium. Any samples expected to exceed ULOQ in the initial analysis were diluted 20-fold before re-analysis. After dose administration, BCY17986, BCY17988, BCY17989, and BCY17994 were detected in transporter medium and in lysed kidney cell samples derived from all in vitro kidney monolayers administered with the test samples.

[0136] The overall bicyclic peptide content in each chamber was calculated from the analytical concentrations, and the true net flux at each bicyclic peptide concentration in each direction was derived by correcting for paracellular leakage using the leakage rate of Lucifer Yellow. The net flux was expressed as pmol / cm³. 2 The data was represented as follows and plotted against time in the direction from the apex to the base (AB) and from the base to the apex (BA).

[0137] The results of the analysis in Sections 5 and 6 above are shown in Figures 1-4, in which it can be seen that all four bicyclic peptides tested exhibited concentration- and time-dependent transcytosis in both the A-B and B-A directions. This is consistent with concurrent tests showing binding of both FITC-transferrin and TfR1 localized on both membranes. Generally, the flux from the basolateral to the apical side was greater than that from the apical to the basolateral side. Previous studies have shown internalization of these bicyclic peptides. This data demonstrates transcytosis of TfR1-binding bicyclic peptides in primary human cultures expressing TfR1, and the crossing of polarized cells indicates the potential for transport across endothelial cells of the peripheral and cerebral vascular systems. This application provides the invention in the following embodiments. (Aspect 1) A peptide ligand specific to transferrin receptor 1 (TfR1), comprising a polypeptide having at least three reactive groups separated by at least two loop sequences, and a molecular scaffold that forms covalent bonds with the reactive groups of the polypeptide, thereby forming at least two polypeptide loops on the molecular scaffold. (Aspect 2) The peptide ligand according to embodiment 1, wherein the reactive group comprises a cysteine ​​residue. (Aspect 3) The peptide ligand according to embodiment 1 or embodiment 2, wherein the peptide ligand inhibits the binding of transferrin to TfR1. (Aspect 4) The peptide ligand according to any one of embodiments 1 to 3, wherein the loop sequence comprises 2, 3, 6, 8, or 9 amino acids. (Appendix 5) The peptide ligand according to embodiment 4, wherein the loop sequence comprises three cysteine ​​residues separated by two loop sequences, the first of which consists of two amino acids and the second of which consists of nine amino acids. (Aspect 6) The peptide ligand according to embodiment 4, wherein the loop sequence comprises three cysteine ​​residues separated by two loop sequences, each consisting of six amino acids. (Aspect 7) The peptide ligand according to embodiment 4, wherein the loop sequence comprises three cysteine ​​residues separated by two loop sequences, the first of which consists of three amino acids and the second of which consists of eight amino acids. (Pattern 8) The peptide ligand, (chemical 1) TIFF0007848202000026.tif19170 (Here, C i 、C ii , and C iii (These represent the first, second, and third cysteine ​​residues, respectively.) : comprises the amino acid sequence of or a pharmaceutically acceptable salt thereof, for example, wherein the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (TATA), and the peptide ligand comprises N- and / or C-terminal additions, A-(Sequence ID 1)-A (referred to herein as BCY12455); A-(Sequence ID 1)-A-[Sar 6 ]-[K-Fl] (referred to as BCY12652 in this specification); A-(Sequence ID 2)-A (referred to herein as BCY12452); A-(Sequence No. 2)-A-[Sar 6 ]-[K-Fl] (referred to as BCY12650 in this specification); A-(Sequence ID 3)-A (referred to herein as BCY12454); and A-(Sequence No. 3)-A-[Sar 6 ]-[K-Fl] (referred to as BCY12651 in this specification). (Here, Sar represents sarcosine and Fl represents fluorescein.) A peptide ligand according to embodiment 4, selected from: (Aspect 9) The peptide ligand according to embodiment 1 or embodiment 2, wherein the peptide ligand does not inhibit the binding of transferrin to TfR1. (Aspect 10) The peptide ligand according to embodiment 9, wherein the loop sequence comprises three or seven amino acids. (Aspect 11) The peptide ligand according to embodiment 10, wherein the loop sequence comprises three cysteine ​​residues separated by two loop sequences, the first of which consists of seven amino acids and the second of which consists of three amino acids. (Aspect 12) The peptide ligand, (Case 2) TIFF0007848202000027.tif121170TIFF0007848202000028.tif248170TIFF0007848202000029.tif62170 (Here, Abu represents aminobutyric acid, Aib represents aminoisobutyric acid, Aze represents azetidine, B-MeIle represents β-methylisoleucine, C5g represents cyclopentylglycine, Cba represents β-cyclobutylalanine, Cbg represents cyclobutylglycine, Chg represents cyclohexylglycine, Cpg represents cyclopropylglycine, EPA represents 2-amino-3-ethylpentanoic acid, HyP represents trans-4-hydroxy-L-proline, [K(N 3 ) represents 6-azidryzine, 1Nal represents 1-naphthylalanine, 2Nal represents 2-naphthylalanine, 4Pal represents 4-pyridylalanine, tBuAla represents t-butylalanine, tBuGly represents t-butylglycine, 3tBuTyr represents 3-t-butyltyrosine, and C i 、C ii , and C iii (These represent the first, second, and third cysteine ​​residues, respectively.) : Containing the amino acid sequence of : or a pharmaceutically acceptable salt thereof, For example, here the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tris(2-bromoetanone)(TATB), and the peptide ligand includes N- and / or C-terminal additions, A-(Sequence ID 4)-A (referred to herein as BCY13983); A-(Sequence No. 4)-A-[Sar 6 ]-[K-Fl] (referred to as BCY14474 in this specification); A-(Sequence ID 5)-A (referred to herein as BCY13986); A-(Sequence No. 5)-A-[Sar 6 ]-[K-Fl] (referred to as BCY14475 in this specification); A-(Sequence ID 6)-A (referred to herein as BCY15466); Ac-(Sequence ID 6) (referred to as BCY15889 herein); A-(Sequence ID 7)-A (referred to herein as BCY15467); Ac-(Sequence ID 7) (referred to as BCY15890 herein); A-(Sequence ID 8)-A (referred to herein as BCY13989); A-(Sequence No. 8)-A-[Sar 6 ]-[K-Fl] (referred to as BCY14476 in this specification); A-(Sequence ID 9)-A (referred to herein as BCY15468); A-(Sequence No. 9)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15768 in this specification); (Sequence number 9)-[Sar 6 ]-[K-Fl] (referred to as BCY15934 in this specification); Ac-(Sequence No. 9)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15937 in this specification); Ac-(Sequence No. 9)-[Sar 6 ]-[K-Fl] (referred to as BCY15938 in this specification); [Fl]G[Sar 5 ]-A-(Sequence ID 9)-A (referred to herein as BCY15940); N[1Nal]N-(SEQ ID NO: 9) (referred to as BCY18030 in this specification); Ac-(Sequence ID 9)-E[Pip]W (referred to herein as BCY18039); Ac-(SEQ ID NO: 9)-EPW (referred to herein as BCY17994); NWN-(Sequence ID 9) (referred to as BCY18029 herein); NWN-(Sequence ID 9)-A (referred to herein as BCY17109); Ac-(Sequence ID 9)-E[Aze]W (referred to herein as BCY18037); Ac-NWN-(Sequence ID 9) (referred to herein as BCY17992); Ac-(Sequence ID 9)-E[dP]W (referred to herein as BCY18038); Ac-N[1Nal]N-(SEQ ID NO: 9) (referred to as BCY18034 herein); N[dW]N-(Sequence ID 9) (referred to as BCY18031 in this specification); Ac-N[dW]N-(SEQ ID NO: 9) (referred to as BCY18035 in this specification); HWM-(Sequence ID 9)-A (referred to herein as BCY17110); A-(Sequence ID 9)-PHP (referred to herein as BCY17115); A-(Sequence ID 9)-EPW (referred to herein as BCY17114); NEV-(Sequence ID 9)-A (referred to herein as BCY17112); A-(Sequence ID 9)-PIVH (referred to herein as BCY17120); Ac-(Sequence ID 9) (referred to as BCY15891 herein); HTS-(Sequence ID 9)-A (referred to herein as BCY17111); Ac-N[NMeTrp]N-(SEQ ID NO: 9) (referred to as BCY18036 in this specification); N[NMeTrp]N-(SEQ ID NO: 9) (referred to as BCY18032 in this specification); Ac-A-(Sequence ID 9)-A (referred to herein as BCY15939); A-(Sequence ID 9)-EHQE (referred to herein as BCY17119); ESF-(Sequence ID 9)-A (referred to herein as BCY17113); NWN-(Sequence ID 9)-[K(N 3 )] (referred to as BCY17870 in this specification); Ac-NWN-(Sequence ID 9)-[K(N 3 )] (referred to as BCY17871 in this specification); [AzPro]-NWN-(Sequence ID 9) (referred to as BCY17872 herein); Ac-(Sequence ID 9)-EPW-[K(N 3 )] (referred to as BCY17873 in this specification); [AzPro]-(Sequence ID 9)-EPW (referred to herein as BCY17874); Ac-(Sequence No. 9)-[K(N 3 )] (referred to as BCY17868 in this specification); [AzPro]-(Sequence ID 9) (referred to as BCY17869 herein); Ac-N[dY]N-(Sequence ID 9)-[K(N 3 )] (referred to as BCY17882 in this specification); Ac-(Sequence ID 9)-E-[dP]-W-[K(N 3 )] (referred to as BCY17890 in this specification); Ac-(Sequence No. 9)-E-[Aze]-W-[K(N 3 )] (referred to as BCY17892 herein); Ac-(Sequence No. 9)-E-[Pip]-W-[K(N 3 )] (referred to as BCY17894 in this specification); Ac-(Sequence No. 9)-[K(N 3 (PYA-maleimide) (referred to as BCY17906 in this specification); Ac-(Sequence ID 9)-EPW-[Peg 10 ]-[K(N 3 )] (referred to as BCY19405 in this specification); Ac-(Sequence ID 9)-EPW-[Peg 24 ]-[K(N 3 )] (referred to as BCY19406 in this specification); Ac-(Sequence No. 9)-EPWGGSGGS-[K(N 3 )] (referred to as BCY19407 in this specification); A-(Sequence ID 10)-A (referred to herein as BCY15469); Ac-(Sequence ID 10) (referred to as BCY15892 herein); A-(Sequence ID 11)-A (referred to herein as BCY15470); Ac-(Sequence ID 11) (referred to as BCY15893 herein); A-(Sequence ID 12)-A (referred to herein as BCY15471); Ac-(Sequence ID 12) (referred to as BCY15894 herein); Ac-(Sequence ID 13) (referred to as BCY17991 herein); Ac-(Sequence ID 13)-EPW (referred to herein as BCY17995); Ac-NWN-(Sequence ID 13) (referred to herein as BCY17993); NWN-(Sequence ID 13) (referred to as BCY18033 in this specification); A-(Sequence ID 13)-A (referred to herein as BCY16754); Ac-(Sequence No. 13)-[K(N 3 )] (referred to as BCY17896 in this specification); Ac-NWN-(Sequence ID 13)-[K(N 3 )] (referred to as BCY17899 in this specification); Ac-(Sequence No. 13)-EPW-[K(N 3 )] (referred to as BCY17901 in this specification); Ac-(Sequence ID 14) (referred to as BCY17990 herein); Ac-(Sequence No. 14)-[K(N 3 )] (referred to as BCY17875 in this specification); [AzPro]-(Sequence ID 14) (referred to herein as BCY17876); Ac-(Sequence ID 15) (referred to as BCY17989 herein); A-(Sequence ID 15)-A (referred to herein as BCY16047); Ac-(Sequence No. 15)-[K(N 3 )] (referred to as BCY17877 in this specification); [AzPro]-(Sequence ID 15) (referred to as BCY17878 herein); A-(Sequence ID 16)-A (referred to herein as BCY16962); TYMN-(Sequence ID 17)-A (referred to as BCY17117 herein); A-(Sequence ID 17)-A (referred to herein as BCY16048); A-(Sequence ID 18)-A (referred to herein as BCY16963); Ac-(Sequence ID 19) (referred to as BCY17987 herein); A-(Sequence ID 20)-A (referred to herein as BCY16753); A-(Sequence ID 21)-A (referred to herein as BCY16046); A-(Sequence ID 22)-A (referred to herein as BCY16964); A-(Sequence ID 23)-A (referred to herein as BCY16965); Ac-(Sequence ID 24) (referred to herein as BCY17986); A-(Sequence ID 25)-A (referred to herein as BCY16550); A-(Sequence ID 26)-A (referred to herein as BCY16966); A-(Sequence ID 27)-A (referred to herein as BCY16051); IDSN-(Sequence ID 28)-A (referred to herein as BCY17118); WGKS-(Sequence ID 29)-A (referred to herein as BCY17116); A-(Sequence ID 30)-A (referred to herein as BCY16053); A-(Sequence ID 31)-A (referred to herein as BCY16557); A-(Sequence ID 32)-A (referred to herein as BCY16035); A-(Sequence ID 33)-A (referred to herein as BCY16043); A-(Sequence No. 34)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15769 in this specification); A-(Sequence ID 35)-A (referred to herein as BCY15648); A-(Sequence ID 36)-A (referred to herein as BCY16031); A-(Sequence ID 37)-A (referred to herein as BCY16079); A-(Sequence ID 38)-A (referred to herein as BCY16036); A-(Sequence ID 39)-A (referred to herein as BCY16029); A-(Sequence ID 40)-A (referred to herein as BCY16089); A-(Sequence ID 41)-A (referred to herein as BCY16088); A-(Sequence ID 42)-A (referred to herein as BCY16052); A-(Sequence ID 43)-A (referred to herein as BCY16033); A-(Sequence ID 44)-A (referred to herein as BCY16039); Ac-(Sequence ID 44) (referred to herein as BCY17988); Ac-(Sequence No. 44)-[K(N 3 )] (referred to as BCY17879 in this specification); [AzPro]-(Sequence ID 44) (referred to herein as BCY17880); A-(Sequence ID 45)-A (referred to herein as BCY16038); A-(Sequence ID 46)-A (referred to herein as BCY16050); A-(Sequence ID 47)-A (referred to herein as BCY16034); A-(Sequence ID 48)-A (referred to herein as BCY16032); A-(Sequence ID 49)-A (referred to herein as BCY16049); A-(Sequence ID 50)-A (referred to herein as BCY16558); A-(Sequence ID 51)-A (referred to herein as BCY16041); A-(Sequence ID 52)-A (referred to herein as BCY16042); A-(Sequence ID 53)-A (referred to herein as BCY16045); A-(Sequence ID 54)-A (referred to herein as BCY16037); A-(Sequence ID 55)-A (referred to herein as BCY16044); A-(Sequence ID 56)-A (referred to herein as BCY16040); A-(Sequence No. 57)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15771 in this specification); A-(Sequence No. 58)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15772 in this specification); A-(Sequence No. 59)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15773 in this specification); A-(Sequence No. 60)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15774 in this specification); A-(Sequence No. 61)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15775 in this specification); A-(Sequence No. 62)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15776 in this specification); A-(Sequence No. 63)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15777 in this specification); A-(Sequence No. 64)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15770 in this specification); Ac-(Sequence ID 65) (referred to herein as BCY17903); Ac-(SEQ ID NO: 66) (referred to herein as BCY17904); and Ac-(Sequence ID 67) (referred to as BCY17905 herein); (Here, AzPro represents azidopropyl, Aze represents azetidine, 1Nal represents 1-naphthylalanine, NMeTrp represents N-methyltryptophan, [K(N 3 )] represents 6-azidridine, Peg represents polyethylene glycol, Pip represents pipecolic acid, Sar represents sarcosine, Fl represents fluorescein, [K(N 3 (PYA-maleimide) has the following structure: (3) TIFF0007848202000030.tif27170 (Represents modified lysine having) : selected from or wherein the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (TATA), and the peptide ligand includes N- and / or C-terminal additions, Ac-(Sequence ID 13) (referred to as BCY20546 herein) A peptide ligand according to any one of embodiments 9 to 11. (Aspect 13) The peptide ligand according to any one of embodiments 1 to 12, wherein the pharmaceutically acceptable salt is selected from free acids or sodium, potassium, calcium, or ammonium salts. (Aspect 14) A multimer-binding complex comprising at least two peptide ligands described in any one of embodiments 1 to 13. (Aspect 15) Compound of formula (I): (C4) TIFF0007848202000031.tif45170 (In the formula, CHM represents the central hinge portion; The two rings represent a bicyclic peptide ligand as described in any one of embodiments 1 to 13; and m represents an integer selected from 2 to 10, for example, 2, 3, or 4. A polymer-binding complex according to embodiment 14, comprising: (Aspect 16) m represents 2, and CHM is given by equation (A): (C5) TIFF0007848202000032.tif49170 For example, BCY19409 The polymer-binding complex described in embodiment 15 is the motif of the above. (Aspect 17) A pharmaceutical composition comprising a peptide ligand according to any one of embodiments 1 to 13 or a polymer-binding complex according to any one of embodiments 14 to 16, in combination with one or more pharmaceutically acceptable excipients. (Aspect 18) A peptide ligand according to any one of embodiments 1 to 13, a polymer-bound complex according to any one of embodiments 14 to 16, or a pharmaceutical composition according to embodiment 17, for use in preventing, suppressing, or treating a disease or disorder by delivery of a therapeutic agent mediated by TfR1. (Aspect 19) A tissue delivery complex comprising a peptide ligand according to any one of embodiments 1 to 13 or a multimer-binding complex according to any one of embodiments 14 to 16, bound to Tfr1, in combination with a payload, for example, an oligonucleotide, particularly siRNA. (Aspect 20) The tissue delivery complex according to embodiment 19, which is a muscle tissue delivery complex. (Aspect 21) A tissue delivery complex according to embodiment 19 or embodiment 20 for use in the treatment of musculoskeletal disorders.

Claims

1. A peptide ligand specific to transferrin receptor 1 (TfR1) or a pharmaceutically acceptable salt thereof, comprising a polypeptide having at least three reactive groups separated by at least two loop sequences, and a molecular scaffold that forms covalent bonds with the reactive groups of the polypeptide, wherein at least two polypeptide loops are formed on the molecular scaffold, The reactive group contains a cysteine ​​residue, The molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)trippropa-2-en-1-one (TATA) or 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tris(2-bromoetanone) (TATB), and The peptide ligand is: C i ALC ii NDWTLPWHHC iii (Sequence ID: 1); C i REFFDTC ii GLAFIEC iii (Sequence ID: 2); C i LEAC ii YDGVYWYSC iii (Sequence ID: 3); C i SADDWLGC ii ISWC iii (Sequence ID: 4); C i SSDAYLGC ii ISWC iii (Sequence ID: 5); C i PPDAHLGC ii ISWC iii (Sequence ID: 6); C i PQDAYLGC ii ISWC iii (Sequence ID: 7); C i PPDSWQGC ii ISYC iii (Sequence ID: 8); C i SPDAHLGC ii ISYC iii (Sequence ID: 9) (referred to as BCY15935 herein); C i PGDAHLGC ii ISYC iii (Sequence ID: 10); C i PPDSHLGC ii ISYC iii (Sequence ID: 11); C i SADDWLGC ii ISYC iii (Sequence ID: 12); C i P[HyP]DAYLGC ii [tBuGly]SYC iii (Sequence ID: 13); C i P[HyP]DAYLGC ii ISYC iii (Sequence ID: 14); C i S[HyP]DAHLGC ii ISYC iii (Sequence ID: 15); C i P[Aib]DAHLGC ii [tBuGly]SYC iii (Sequence ID: 16); C i PPDAHLGC ii ISYC iii (Sequence ID: 17); C i P[Aib]DAYLGC ii [tBuGly]SYC iii (Sequence ID: 18); C i SADAHLGC ii ISYC iii (Sequence ID: 19); C i S[Aib]DAHLGC ii [tBuGly]SYC iii (Sequence ID: 20); C i SPDAHLGC ii [EPA]SYC iii (Sequence ID: 21); C i PPDAYLGC ii [tBuGly]SYC iii (Sequence ID: 22); C i S[Aib]DAYLGC ii [tBuGly]SYC iii (Sequence ID: 23); C i APDAHLGC ii ISYC iii (Sequence ID: 24); C i P[Aib]DAHLGC ii ISYC iii (Sequence ID: 25); C i SPDAYLGC ii [tBuGly]SYC iii (Sequence ID: 26); C i SPDAHLGC ii [tBuGly]SYC iii (Sequence ID: 27); C i PNDAHLGC ii ISYC iii (Sequence ID: 28); C i PIDAHLGC ii ISYC iii (Sequence ID: 29); C i SPDAYLGC ii ISYC iii (Sequence ID: 30); C i PPDAYLGC ii ISYC iii (Sequence ID: 31); C i S[Aib]DAHLGC ii ISYC iii (Sequence ID: 32); C i SPDAHLGC ii [Chg]SYC iii (Sequence ID: 33); C i APDAHLGC ii ISYC iii (Sequence ID: 34); C i YLPDW[tBuAla]C ii GDEYC iii (Sequence ID: 35); C i SPDAHLGC ii IS[2Nal]C iii (Sequence ID: 36); C i SPDAHLGC ii IS[3tBuTyr]C iii (Sequence ID: 37); C i SPD[Aib]HLGC ii ISYC iii (Sequence ID: 38); C i SPDAHLGC ii IS[1Nal]C iii (Sequence ID: 39); C i SPDAH[tBuAla]GC ii ISYC iii (Sequence ID: 40); C i SPDAH[Cba]GC ii ISYC iii (Sequence ID: 41); C i SPDAHLGC ii ISWC iii (Sequence ID: 42); C i SPD[Abu]HLGC ii ISYC iii (Sequence ID: 43); C i S[Aze]DAHLGC ii ISYC iii (Sequence ID: 44); C i SPDDHLGC ii ISYC iii (Sequence ID: 45); C i SPDSHLGC ii ISYC iii (Sequence ID: 46); C i SPDAH[Abu]GC ii ISYC iii (Sequence ID: 47); C i SPDAHLGC ii IS[4Pal]C iii (Sequence ID: 48); C i P[dA]DAHLGC ii ISYC iii (Sequence ID: 49); C i SPDAYLGC ii [tBuAla]SYC iii (Sequence ID: 50); C i SPDAHLGC ii [C5g]SYC iii (Sequence ID: 51); C i SPDAHLGC ii [Cbg]SYC iii (Sequence ID: 52); C i SPDAHL[dA]C ii ISYC iii (Sequence ID: 53); C i SPDAH[Aib]GC ii ISYC iii (Sequence ID: 54); C i SPDAHLGC ii [Cpg]SYC iii (Sequence ID: 55); C i SPDAHLGC ii [B-MeIle]SYC iii (Sequence ID: 56); C i SADAHLGC ii ISYC iii (Sequence ID: 57); C i SPAAHLGC ii ISYC iii (Sequence ID: 58); C i SPDAALGC ii ISYC iii (Sequence ID: 59); C i SPDAHAGC ii ISYC iii (Sequence ID: 60); C i SPDAHLAC ii ISYC iii (Sequence ID: 61); C i SPDAHLGC ii ASYC iii (Sequence ID: 62); C i SPDAHLGC ii IAYC iii (Sequence ID: 63); C i SPDAHLGC ii ISAC iii (Sequence ID: 64); C i [K(N 3)]PDAHLGC ii ISYC iii (Sequence ID: 65); C i S[K(N 3 )]DAHLGC ii ISYC iii (Sequence ID: 66); or C i SPD[K(N 3)]HLGC ii ISYC iii (Sequence ID: 67); (Here, Abu represents aminobutyric acid, Aib represents aminoisobutyric acid, Aze represents azetidine, B-MeIle represents β-methylisoleucine, C5g represents cyclopentylglycine, Cba represents β-cyclobutylalanine, Cbg represents cyclobutylglycine, Chg represents cyclohexylglycine, Cpg represents cyclopropylglycine, EPA represents 2-amino-3-ethylpentanoic acid, HyP represents trans-4-hydroxy-L-proline, [K(N 3 ) represents 6-azidridine, 1Nal represents 1-naphthylalanine, 2Nal represents 2-naphthylalanine, 4Pal represents 4-pyridylalanine, tBuAla represents t-butylalanine, tBuGly represents t-butylglycine, 3tBuTyr represents 3-t-butyltyrosine, and C i, C ii, and C iii represent the first, second, and third cysteine ​​residues, respectively. A peptide ligand or a pharmaceutically acceptable salt thereof, comprising a polypeptide or a pharmaceutically acceptable salt thereof.

2. The peptide ligand according to claim 1, or a pharmaceutically acceptable salt thereof, wherein the peptide ligand inhibits the binding of transferrin to TfR1.

3. The molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (TATA), and the peptide ligand includes N- and / or C-terminal additions, A-(Sequence ID 1)-A (referred to herein as BCY12455); A-(Sequence ID 1)-A-[Sar 6 ]-[K-Fl] (referred to as BCY12652 in this specification); A-(Sequence ID 2)-A (referred to herein as BCY12452); A-(Sequence No. 2)-A-[Sar 6 ]-[K-Fl] (referred to as BCY12650 in this specification); A-(Sequence ID 3)-A (referred to herein as BCY12454); or A-(Sequence No. 3)-A-[Sar 6 ]-[K-Fl] (referred to as BCY12651 in this specification) or a salt that is acceptable as a medicine (Here, Sar represents sarcosine and Fl represents fluorescein.) A peptide ligand according to claim 1 or a pharmaceutically acceptable salt thereof, selected from:

4. The peptide ligand according to claim 1, or a pharmaceutically acceptable salt thereof, wherein the peptide ligand does not inhibit the binding of transferrin to TfR1.

5. The molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tris(2-bromoetanone)(TATB), and the peptide ligand or a pharmaceutically acceptable salt thereof includes N- and / or C-terminal additions, A-(Sequence ID 4)-A (referred to herein as BCY13983); A-(Sequence No. 4)-A-[Sar 6 ]-[K-Fl] (referred to as BCY14474 in this specification); A-(Sequence ID 5)-A (referred to herein as BCY13986); A-(Sequence No. 5)-A-[Sar 6 ]-[K-Fl] (referred to as BCY14475 in this specification); A-(Sequence ID 6)-A (referred to herein as BCY15466); Ac-(Sequence ID 6) (referred to as BCY15889 herein); A-(Sequence ID 7)-A (referred to herein as BCY15467); Ac-(Sequence ID 7) (referred to as BCY15890 herein); A-(Sequence ID 8)-A (referred to herein as BCY13989); A-(Sequence No. 8)-A-[Sar 6 ]-[K-Fl] (referred to as BCY14476 in this specification); A-(Sequence ID 9)-A (referred to herein as BCY15468); A-(Sequence No. 9)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15768 in this specification); (Sequence number 9)-[Sar 6 ]-[K-Fl] (referred to as BCY15934 in this specification); Ac-(SEQ ID NO:9)-A-[Sar 6 -[K-Fl](referred to herein as BCY15937); Ac-(Sequence No. 9)-[Sar 6 ]-[K-Fl] (referred to as BCY15938 in this specification); [Fl]G[Sar 5 ]-A-(Sequence ID 9)-A (referred to herein as BCY15940); N[1Nal]N-(SEQ ID NO: 9) (referred to as BCY18030 in this specification); Ac-(Sequence ID 9)-E[Pip]W (referred to herein as BCY18039); Ac-(SEQ ID NO: 9)-EPW (referred to herein as BCY17994); NWN-(Sequence ID 9) (referred to as BCY18029 herein); NWN-(Sequence ID 9)-A (referred to herein as BCY17109); Ac-(Sequence ID 9)-E[Aze]W (referred to herein as BCY18037); Ac-NWN-(Sequence ID 9) (referred to herein as BCY17992); Ac-(Sequence ID 9)-E[dP]W (referred to herein as BCY18038); Ac-N[1Nal]N-(SEQ ID NO: 9) (referred to as BCY18034 herein); N[dW]N-(Sequence ID 9) (referred to as BCY18031 in this specification); Ac-N[dW]N-(SEQ ID NO: 9) (referred to as BCY18035 in this specification); HWM-(Sequence ID 9)-A (referred to herein as BCY17110); A-(Sequence ID 9)-PHP (referred to herein as BCY17115); A-(Sequence ID 9)-EPW (referred to herein as BCY17114); NEV-(Sequence ID 9)-A (referred to herein as BCY17112); A-(Sequence ID 9)-PIVH (referred to herein as BCY17120); Ac-(Sequence ID 9) (referred to as BCY15891 herein); HTS-(Sequence ID 9)-A (referred to herein as BCY17111); Ac-N[NMeTrp]N-(SEQ ID NO: 9) (referred to as BCY18036 in this specification); N[NMeTrp]N-(SEQ ID NO: 9) (referred to as BCY18032 in this specification); Ac-A-(Sequence ID 9)-A (referred to herein as BCY15939); A-(Sequence ID 9)-EHQE (referred to herein as BCY17119); ESF-(Sequence ID 9)-A (referred to herein as BCY17113); NWN-(Sequence ID 9)-[K(N 3 )] (referred to as BCY17870 in this specification); Ac-NWN-(Sequence ID 9)-[K(N 3 )] (referred to as BCY17871 in this specification); [AzPro]-NWN-(Sequence ID 9) (referred to as BCY17872 herein); Ac-(Sequence ID 9)-EPW-[K(N 3 )] (referred to as BCY17873 in this specification); [AzPro]-(Sequence ID 9)-EPW (referred to herein as BCY17874); Ac-(Sequence No. 9)-[K(N 3 )] (referred to as BCY17868 in this specification); [AzPro]-(Sequence ID 9) (referred to as BCY17869 herein); Ac-N[dY]N-(Sequence ID 9)-[K(N 3 )] (referred to as BCY17882 in this specification); Ac-(Sequence ID 9)-E-[dP]-W-[K(N 3 )] (referred to as BCY17890 in this specification); Ac-(Sequence No. 9)-E-[Aze]-W-[K(N 3 )] (referred to as BCY17892 herein); Ac-(Sequence No. 9)-E-[Pip]-W-[K(N 3 )] (referred to as BCY17894 in this specification); Ac-(Sequence No. 9)-[K(N 3 (PYA-maleimide) (referred to as BCY17906 in this specification); Ac-(Sequence ID 9)-EPW-[Peg 10 ]-[K(N 3 )] (referred to as BCY19405 in this specification); Ac-(Sequence ID 9)-EPW-[Peg 24 ]-[K(N 3 )] (referred to as BCY19406 in this specification); Ac-(Sequence No. 9)-EPWGGSGGS-[K(N 3 )] (referred to as BCY19407 in this specification); A-(Sequence ID 10)-A (referred to herein as BCY15469); Ac-(Sequence ID 10) (referred to as BCY15892 herein); A-(Sequence ID 11)-A (referred to herein as BCY15470); Ac-(Sequence ID 11) (referred to as BCY15893 herein); A-(Sequence ID 12)-A (referred to herein as BCY15471); Ac-(Sequence ID 12) (referred to as BCY15894 herein); Ac-(Sequence ID 13) (referred to as BCY17991 herein); Ac-(Sequence ID 13)-EPW (referred to herein as BCY17995); Ac-NWN-(Sequence ID 13) (referred to herein as BCY17993); NWN-(Sequence ID 13) (referred to as BCY18033 in this specification); A-(Sequence ID 13)-A (referred to herein as BCY16754); Ac-(Sequence No. 13)-[K(N 3 )] (referred to as BCY17896 in this specification); Ac-NWN-(Sequence ID 13)-[K(N 3 )] (referred to as BCY17899 in this specification); Ac-(Sequence No. 13)-EPW-[K(N 3 )] (referred to as BCY17901 in this specification); Ac-(Sequence ID 14) (referred to as BCY17990 herein); Ac-(Sequence No. 14)-[K(N 3 )] (referred to as BCY17875 in this specification); [AzPro]-(Sequence ID 14) (referred to herein as BCY17876); Ac-(Sequence ID 15) (referred to as BCY17989 herein); A-(Sequence ID 15)-A (referred to herein as BCY16047); Ac-(Sequence No. 15)-[K(N 3 )] (referred to as BCY17877 in this specification); [AzPro]-(Sequence ID 15) (referred to as BCY17878 herein); A-(Sequence ID 16)-A (referred to herein as BCY16962); TYMN-(Sequence ID 17)-A (referred to as BCY17117 herein); A-(Sequence ID 17)-A (referred to herein as BCY16048); A-(Sequence ID 18)-A (referred to herein as BCY16963); Ac-(Sequence ID 19) (referred to as BCY17987 herein); A-(Sequence ID 20)-A (referred to herein as BCY16753); A-(Sequence ID 21)-A (referred to herein as BCY16046); A-(Sequence ID 22)-A (referred to herein as BCY16964); A-(Sequence ID 23)-A (referred to herein as BCY16965); Ac-(Sequence ID 24) (referred to herein as BCY17986); A-(Sequence ID 25)-A (referred to herein as BCY16550); A-(Sequence ID 26)-A (referred to herein as BCY16966); A-(Sequence ID 27)-A (referred to herein as BCY16051); IDSN-(Sequence ID 28)-A (referred to herein as BCY17118); WGKS-(Sequence ID 29)-A (referred to herein as BCY17116); A-(Sequence ID 30)-A (referred to herein as BCY16053); A-(Sequence ID 31)-A (referred to herein as BCY16557); A-(Sequence ID 32)-A (referred to herein as BCY16035); A-(Sequence ID 33)-A (referred to herein as BCY16043); A-(Sequence No. 34)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15769 in this specification); A-(Sequence ID 35)-A (referred to herein as BCY15648); A-(Sequence ID 36)-A (referred to herein as BCY16031); A-(Sequence ID 37)-A (referred to herein as BCY16079); A-(Sequence ID 38)-A (referred to herein as BCY16036); A-(Sequence ID 39)-A (referred to herein as BCY16029); A-(Sequence ID 40)-A (referred to herein as BCY16089); A-(Sequence ID 41)-A (referred to herein as BCY16088); A-(Sequence ID 42)-A (referred to herein as BCY16052); A-(Sequence ID 43)-A (referred to herein as BCY16033); A-(Sequence ID 44)-A (referred to herein as BCY16039); Ac-(Sequence ID 44) (referred to herein as BCY17988); Ac-(Sequence No. 44)-[K(N 3 )] (referred to as BCY17879 in this specification); [AzPro]-(Sequence ID 44) (referred to herein as BCY17880); A-(Sequence ID 45)-A (referred to herein as BCY16038); A-(Sequence ID 46)-A (referred to herein as BCY16050); A-(Sequence ID 47)-A (referred to herein as BCY16034); A-(Sequence ID 48)-A (referred to herein as BCY16032); A-(Sequence ID 49)-A (referred to herein as BCY16049); A-(Sequence ID 50)-A (referred to herein as BCY16558); A-(Sequence ID 51)-A (referred to herein as BCY16041); A-(Sequence ID 52)-A (referred to herein as BCY16042); A-(Sequence ID 53)-A (referred to herein as BCY16045); A-(Sequence ID 54)-A (referred to herein as BCY16037); A-(Sequence ID 55)-A (referred to herein as BCY16044); A-(Sequence ID 56)-A (referred to herein as BCY16040); A-(Sequence No. 57)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15771 in this specification); A-(Sequence No. 58)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15772 in this specification); A-(Sequence No. 59)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15773 in this specification); A-(Sequence No. 60)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15774 in this specification); A-(Sequence No. 61)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15775 in this specification); A-(Sequence No. 62)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15776 in this specification); A-(Sequence No. 63)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15777 in this specification); A-(Sequence No. 64)-A-[Sar 6 ]-[K-Fl] (referred to as BCY15770 in this specification); Ac-(Sequence ID 65) (referred to herein as BCY17903); Ac-(Sequence ID 66) (referred to herein as BCY17904); or Ac-(Sequence ID 67) (referred to as BCY17905 herein); or a salt that is acceptable as a medicine (Here, AzPro represents azidopropyl, Aze represents azetidine, 1Nal represents 1-naphthylalanine, NMeTrp represents N-methyltryptophan, [K(N 3 )] represents 6-azidridine, Peg represents polyethylene glycol, Pip represents pipecolic acid, Sar represents sarcosine, Fl represents fluorescein, [K(N 3 (PYA-maleimide) has the following structure: 【Chemistry 1】 (Represents modified lysine having) : selected from or the molecular scaffold is 1,1',1''-(1,3,5-triazinan-1,3,5-triyl)tripropa-2-en-1-one (TATA) and the peptide ligand includes N- and / or C-terminal additions, Ac-(Sequence ID 13) (referred to as BCY20546 herein) or a salt that is acceptable as a medicine A peptide ligand or a pharmaceutically acceptable salt thereof according to claim 1 or 4.

6. The peptide ligand according to any one of claims 1 to 5, wherein the peptide ligand comprises a C-terminal amide group, or a pharmaceutically acceptable salt thereof.

7. The peptide ligand or pharmaceutically acceptable salt thereof according to any one of claims 1 to 6, wherein the pharmaceutically acceptable salt is selected from a free acid or a sodium, potassium, calcium, or ammonium salt.

8. A polymer-binding complex comprising at least two peptide ligands or pharmaceutically acceptable salts thereof as described in any one of claims 1 to 7.

9. Compound of formula (I): 【Chemistry 2】 (In the formula, CHM represents the central hinge portion; The two rings represent the peptide ligand described in any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof; and (m represents an integer selected from 2 to 10.) The polymer-bound complex according to claim 8, comprising:

10. The polymer-bound complex according to claim 9, wherein m represents an integer selected from 2, 3, or 4.

11. m represents 2, and CHM is given by equation (A): 【Transformation 3】 A polymer-bound complex according to claim 9 or 10, which is the motif of the above.

12. The polymer-binding complex according to claim 11, wherein the polymer-binding complex is BCY19409.

13. A pharmaceutical composition comprising a peptide ligand according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof, or a polymer-bound complex according to any one of claims 8 to 12.

14. A pharmaceutical for preventing, suppressing, or treating a disease or disorder by delivery of a therapeutic agent mediated by TfR1, comprising as an active ingredient a peptide ligand according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof, or a polymer-bound complex according to any one of claims 8 to 12, or a pharmaceutical composition according to claim 13.

15. A tissue delivery complex comprising, in combination with a payload, a peptide ligand according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof, or a polymer-bound complex according to any one of claims 8 to 12, conjugated to Tfr1.

16. The tissue delivery complex according to claim 15, which is a muscle tissue delivery complex.

17. A pharmaceutical agent for treating musculoskeletal disorders, comprising a tissue delivery complex according to claim 15 or claim 16 as an active ingredient.

18. Use of a peptide ligand according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof, a polymer-bound complex according to any one of claims 8 to 12, or a pharmaceutical composition according to claim 13 in the manufacture of a pharmaceutical for treating a disease or disorder by delivery of a therapeutic agent mediated by TfR1.

19. Use of the tissue delivery complex according to claim 15 or 16 in the manufacture of a pharmaceutical product for treating musculoskeletal disorders.

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