Conjugate as well as preparation method and application thereof

Abstract

The invention relates to a conjugate for inhibiting virus replication and a preparation method and application thereof. The present invention relates to a conjugate comprising a dimer or trimer of a moiety that inhibits influenza virus neuraminidase (e.g., zanamivir, peramivir, oseltamivir, or an analog thereof) coupled to an Fc domain monomer, an Fc domain, an Fc binding peptide, an albumin, or an albumin binding peptide. The conjugates are useful in the treatment of viral infections (e.g., influenza virus infections).

Description

[0001] Cross-reference

[0002] This application is based on an application with a CN application number of 202410111150.3 and a filing date of January 26, 2024, and claims its priority. The disclosure of this CN application is hereby incorporated herein by reference in its entirety. Technical Field

[0003] This application relates to conjugates for inhibiting virus replication, their preparation methods and uses. Background Art

[0004] In the medical field, there has been a continuous search for new and effective anti-influenza virus therapeutic drugs and strategies. Although most people can fully recover on their own within one to two weeks, some people will develop life-threatening complications, such as pneumonia. Therefore, for some susceptible populations, especially children, the elderly, or patients with chronic diseases, those with a weakened or compromised immune system, influenza can be fatal. For example, patients with advanced HIV or organ transplant patients (whose immune systems are medically suppressed to prevent rejection of the transplanted organ) are at a greater risk of influenza-related complications. Pregnant women and young children are also at high risk of complications.

[0005] The development of drugs for anti-influenza virus treatment has become an ongoing challenge. Several influenza antiviral preparations have been approved for clinical use. However, with long-term use, drug-resistant strains have now emerged against the most commonly used influenza inhibitors.

[0006] Anti-influenza virus drugs mainly target proteins on the surface of influenza virus particles. The envelope of the influenza virus contains two key glycoproteins, namely hemagglutinin (HA) and neuraminidase (NA), which play key roles in virus infection and spread. Neuraminidase is an exoglycosidase that cleaves sialic acid from the glycan structure on the surface of infected host cells, thereby releasing progeny viruses and spreading to uninfected surrounding cells. Therefore, neuraminidase has become a pharmacological target for antiviral drugs. There are already many small molecule drugs on the market for neuraminidase inhibitors to reduce virus spread, including oseltamivir (Tamiflu ,

[0007] , ,

[0006] , TM , TM , TM ), zanamivir (Relenza TM ), and peramivir (Rapivab TM ).

[0007] Targeted delivery of conjugated drugs is an emerging treatment method. Its basic principle is to link drugs with targeting molecules so that the drugs can selectively bind to disease-related cells or tissues. In cancer treatment, targeted delivery of conjugated drugs has achieved some encouraging results. For example, some antibody drugs have been successfully used to treat diseases such as breast cancer, colon cancer, and lymphoma. Currently, macromolecular conjugation technology has also been applied to the development of new antiviral drugs. For example, Chimerix Therapeutics has developed a human IgG Fc protein-conjugated dimer NA inhibitor (CN 113194983 A, WO 2021 / 046549A1), which has initially demonstrated therapeutic potential in the treatment of influenza.

[0008] However, due to the strong antigenic drift and antigenic shift of influenza viruses, new subtypes are constantly emerging, thus increasing the possibility of developing drug-resistant strains, and there is a need to develop new and more effective therapies for the treatment of influenza. Summary of the Invention

[0009] The present application relates to conjugates for inhibiting virus replication, their preparation methods, and uses. In particular, such conjugates couple an influenza virus neuraminidase inhibitor (e.g., zanamivir, peramivir, oseltamivir, or their analogs) to a biological macromolecule such as an antibody, antibody fragment, or albumin. The neuraminidase inhibitor in the conjugate targets the neuraminidase on the surface of virus particles. The Fc end or Fc protein of the antibody in the conjugate binds to FcγRs (such as FcRn, FcγRI, FcγrIa, FcγRIIc, FcγRIIIa, and FcγRIIIb) on immune cells (such as neutrophils), activating the phagocytic function of immune cells, such as antibody-dependent cell-mediated cytotoxicity (ADCC), thereby causing immune cells to phagocytose and destroy virus particles, further enhancing the antiviral activity of the conjugate. Albumin can extend the half-life of the conjugate. For example, albumin binds to FcRn to delay the metabolic loss of the conjugate.

[0010] Specifically, the conjugate contains a dimer or trimer of a moiety that inhibits influenza virus neuraminidase (e.g., zanamivir, peramivir, oseltamivir, or analogs thereof), which is conjugated to an Fc domain monomer, an Fc domain, an Fc-binding peptide, albumin, or an albumin-binding peptide. The neuraminidase inhibitor in the conjugate targets neuraminidase on the surface of virions. The Fc monomer or Fc domain in the conjugate binds to FcγRs (e.g., FcRn, FcγRI, FcγRIIa, FcγRIIc, FcγRIIIa, and FcγRIIIb) on immune cells (e.g., neutrophils) to activate phagocytosis and effector functions such as antibody-dependent cell-mediated cytotoxicity (ADCC), so that virions are engulfed and destroyed by immune cells and further enhance the antiviral activity of the conjugate. The albumin or albumin-binding peptide can prolong the half-life of the conjugate by binding to the recycling neonatal Fc receptor (FcRn). The conjugate can be used to inhibit virus growth and can also be used to treat virus infections (such as those caused by influenza A virus, influenza B virus, and influenza C virus).

[0011] In one aspect, the present application provides a conjugate of formula (I) or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof,

[0012]

[0013] wherein A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-XIV),

[0014]

[0015]

[0016] wherein L can be linked to any possible site of the compound of formula (A-XIV);

[0017] wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2, and -NHC(=NH)NHR6;

[0018] R2 and R3 are each independently selected from -H, -OH, -F, -Cl, and -Br;

[0019] R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3, and -C(=O)OCF3;

[0020] R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3;

[0021] X is selected from -O- and -S-;

[0022] Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)- and -NH(SO2)NR7-;

[0023] R’ is H or C1-C6 alkyl;

[0024] R6 is selected from

[0025]

[0026]

[0027] R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl;

[0028] Each E is a biological macromolecule; n is 1 or 2; T is a number between 1 and 20;

[0029] The wavy line connected to each E represents each covalent connection (e.g., by means of a covalent bond or a linker) to the N atom in each E (e.g., the N atom on the side-chain amino group of a lysine, arginine, asparagine or glutamine residue), and L is a linker covalently connected to each of the groups E, A1, A2 and A3.

[0030] In some embodiments, in the conjugate of formula (I), A1, A2 and A3 are each independently selected from formula (A-I) to formula (A-XIII),

[0031]

[0032] wherein the definitions of R1, R2, R3, R4, R5, R’, X and Y are as described in any embodiment of the present application.

[0033] In some embodiments, in the conjugate of formula (I), A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-IV), and formula (A-VII),

[0034]

[0035] wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0036] In some embodiments, each E in the conjugate of formula (I) comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence selected from SEQ ID NOs: 1-68), albumin (e.g., albumin having a sequence selected from SEQ ID NOs: 69-71), an albumin-binding peptide, or an Fc-binding peptide; n is 1 or 2, and when n is 2 and E is an Fc domain monomer, the two Es dimerize to form an Fc domain.

[0037] In some embodiments, in the conjugate of formula (I), A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-III), and formula (A-VII),

[0038]

[0039] wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0040] In some embodiments, in the conjugate of formula (I), A1, A2, and A3 are each independently selected from formula (A-I), formula (A-III), and formula (A-VII),

[0041]

[0042] wherein the definitions of R1, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0043] In some embodiments, E in the conjugate of formula (I) comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence selected from SEQ ID NOs: 1-68); n is 1 or 2, and when n is 2, the two Es dimerize to form an Fc domain.

[0044] In some embodiments, E in the conjugate of formula (I) is an Fc domain monomer, and the Fc domain monomer has an amino acid sequence selected from:

[0045] i) any one of the sequences shown in SEQ ID NOs: 1-68;

[0046] ii) a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to any of the sequences shown in SEQ ID NOs: 1-68; and

[0047] iii) a sequence having at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) to any of the sequences shown in SEQ ID NOs: 1-68.

[0048] In some embodiments, the substitutions described in ii) are conservative substitutions.

[0049] In some embodiments, E in the conjugate of formula (I) is a monomer of the Fc domain having at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) to the sequence of SEQ ID NO: 64.

[0050] In some embodiments, E in the conjugate of formula (I) is a monomer of the Fc domain having the sequence of SEQ ID NO: 64.

[0051] In some embodiments, E in the conjugate of formula (I) is a monomer of the Fc domain having the sequence of SEQ ID NO: 64, n is 2, and two Es dimerize to form the Fc domain.

[0052] In some embodiments, R1 in the conjugate of formula (I) is selected from -OH, -NH2, -NHC(=NH)NH2. In some embodiments, R1 in the conjugate of formula (I) is -NHC(=NH)NH2.

[0053] In some embodiments, R4 in the conjugate of formula (I) is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3. In some embodiments, R4 in the conjugate of formula (I) is selected from -C(=O)OH and -C(=O)OCH3.

[0054] In some embodiments, R5 in the conjugate of formula (I) is selected from -C(=O)CH3 and -C(=O)CF3. In some embodiments, R5 in the conjugate of formula (I) is -C(=O)CH3.

[0055] In some embodiments, R' in the conjugate of formula (I) is H or C1-C4 alkyl. In some embodiments, R' in the conjugate of formula (I) is H, methyl or ethyl. In some embodiments, R' in the conjugate of formula (I) is H or ethyl.

[0056] In some embodiments, X in the conjugate of formula (I) is -O-.

[0057] In some embodiments, Y in the conjugate of formula (I) is selected from -O-, -S-, -NH-. In some embodiments, Y in the conjugate of formula (I) is selected from -O- and -NH-.

[0058] In some embodiments, the conjugate of formula (I) has the structure shown in formula (I-II-A), formula (I-II-B) or formula (I-II-C),

[0059]

[0060]

[0061] wherein the definitions of R1, R4, R5, R', X, Y, E, n, L, T are as described in any embodiment of the present application.

[0062] In some embodiments, the conjugate of formula (I) has the structure shown in formula (I-II-A'), formula (I-II-B') or formula (I-II-C'),

[0063]

[0064] wherein the definitions of R1, R4, R5, R', X, Y, E, n, T are as described in any embodiment of the present application,

[0065] L1 is selected from:

[0066] Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20.

[0067] In some embodiments, in the conjugate of formula (I), L1 is selected from:

[0068] Wherein the definitions of V, p, m and Z are as described in any embodiment of the present application.

[0069] In some embodiments, in the conjugate of formula (I), L1 is Wherein the definitions of V, p, m and Z are as described in any embodiment of the present application.

[0070] In some embodiments, in the conjugate of formula (I), V is -CH2- or -O-. In some embodiments, in the conjugate of formula (I), V is -O-.

[0071] In some embodiments, in the conjugate of formula (I), Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)- or -C(=O)-. In some embodiments, in the conjugate of formula (I), Z is -NH-C(=O)- or -C(=O)-.

[0072] In some embodiments, in the conjugate of formula (I), p is an integer between 1 and 10. In some embodiments, in the conjugate of formula (I), p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6).

[0073] In some embodiments, in the conjugate of formula (I), m is an integer between 1 and 12. In some embodiments, in the conjugate of formula (I), m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0074] In some embodiments, the conjugate represented by formula (I) has the structure represented by formula (I-II-A'-1), formula (I-II-B'-1), formula (I-II-B'-2), or formula (I-II-C'-1);

[0075]

[0076] wherein the definitions of E, n, T, and L1 are as described in any embodiment of the present application.

[0077] In some embodiments, the conjugate represented by formula (I) has the structure represented by formula (I-II-A-2), formula (I-II-B-3), formula (I-II-B-4), or formula (I-II-C-2),

[0078]

[0079] wherein the definitions of E, n, T, and L1 are as described in any embodiment of the present application;

[0080] L2 is selected from:

[0081] wherein V is -CH2-, -O-, or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; Z1 and Z2 are independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; m and p are each independently an integer from 1 to 20;

[0082] L3 is wherein U is -C(=O)-NH-, -NH-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, -CH2-; W is -CH2-, -O-, or -S-; i and q are each independently an integer from 1 to 20, one end of L3 is covalently connected to E, and the other end is covalently connected to L2.

[0083] In some embodiments, in the conjugate of formula (I), L3 is covalently linked to E through an N atom, where the N atom is an N atom from E (e.g., the N atom on the side-chain amino group of a lysine, arginine, asparagine, or glutamine residue).

[0084] In some embodiments, in the conjugate of formula (I), L2 is selected from:

[0085] where V is -CH2-, -O-, or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; and m and p are each independently an integer from 1 to 20.

[0086] In some embodiments, in the conjugate of formula (I), L1 is where V, p, m, and Z are as defined in any embodiment of the present application. In some embodiments, V is -CH2- or -O-. In some embodiments, V is -O-. In some embodiments, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, or -C(=O)-. In some embodiments, Z is -NH-C(=O)- or -C(=O)-. In some embodiments, p is an integer between 1 and 10. In some embodiments, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6). In some embodiments, m is an integer between 1 and 12. In some embodiments, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0087] In some embodiments, in the conjugate of formula (I), L2 is Wherein the definitions of V, p, m, and Z are as described in any embodiment of the present application. In some embodiments, V is -CH2- or -O-. In some embodiments, V is -O-. In some embodiments, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-. In some embodiments, Z is -C(=O)-NH-. In some embodiments, p is an integer between 1 and 10. In some embodiments, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6). In some embodiments, m is an integer between 1 and 12. In some embodiments, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0088] In some embodiments, in the conjugate shown in formula (I), L3 is Wherein the definitions of U, i, and q are as described in any embodiment of the present application. In some embodiments, U is -NH-C(=O)-, -N(CH3)-C(=O)- or -N(CH2CH3)-C(=O)-. In some embodiments, U is -NH-C(=O)-. In some embodiments, the N atom in U is the N atom from E (e.g., the N atom on the side chain amino group of a lysine, arginine, asparagine, or glutamine residue). In some embodiments, i is an integer between 1 and 12. In some embodiments, i is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9). In some embodiments, q is an integer between 1 and 10. In some embodiments, q is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6).

[0089] In some embodiments, in the conjugate shown in formula (I), T is a number between 1 and 10. In some embodiments, in the conjugate shown in formula (I), T is a number between 1 and 9. In some embodiments, in the conjugate shown in formula (I), T is a number between 1 and 7. In some embodiments, in the conjugate shown in formula (I), T is a number between 1 and 5. In some embodiments, in the conjugate shown in formula (I), T is a number between 3 and 7. In some embodiments, in the conjugate shown in formula (I), T is a number between 3 and 5.5.

[0090] In some embodiments, the conjugate shown in formula (I) has the structure shown in formula (I-II-A-3), formula (I-II-B-5), formula (I-II-B-6), or formula (I-II-C-3),

[0091]

[0092]

[0093]

[0094] wherein the definitions of E, n, and T are as described in any embodiment of the present application; n1, n2, and n3 are each independently an integer between 1 and 20. In some embodiments, n1, n2, and n3 are each independently an integer between 1 and 12. In some embodiments, n1, n2, and n3 are each independently an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0095] In some embodiments, the conjugate represented by formula (I) is selected from:

[0096]

[0097]

[0098]

[0099]

[0100]

[0101] wherein the definitions of E, n, and T are as described in any embodiment of the present application.

[0102] On the other hand, the present application also provides a conjugate represented by formula (II) or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof,

[0103]

[0104] wherein A1 and A2 are each independently selected from formula (A-I) to formula (A-XIV),

[0105]

[0106]

[0107] wherein L can be connected to any possible site of the compound of formula (A-XIV);

[0108] wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2, and -NHC(=NH)NHR6;

[0109] R2 and R3 are each independently selected from -H, -OH, -F, -Cl, and -Br;

[0110] R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3 and -C(=O)OCF3;

[0111] R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3;

[0112] X is selected from -O- and -S-;

[0113] Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)- and -NH(SO2)NR7-;

[0114] R’ is H or C1-C6 alkyl;

[0115] R6 is selected from

[0116]

[0117]

[0118] R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl;

[0119] Each E is a biological macromolecule; n is 1 or 2; T is a number between 1 and 20;

[0120] The wavy line connected to each E indicates that each A1-L-A2 is covalently linked (e.g., by a covalent bond or a linker) to the N atom in each E (e.g., the N atom on the side chain amino group of a lysine, arginine, asparagine or glutamine residue), and L is a linker covalently linked to each of the groups E, A1, and A2.

[0121] In some embodiments, in the conjugate of formula (II), A1 and A2 are each independently selected from formula (A-I) to formula (A-XIII),

[0122]

[0123] wherein the definitions of R1, R2, R3, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0124] In some embodiments, in the conjugate represented by formula (II), A1 and A2 are each independently selected from formula (A-I) to formula (A-IV) and formula (A-VII),

[0125]

[0126] wherein the definitions of R1, R2, R3, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0127] In some embodiments, in the conjugate represented by formula (II), each E comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence selected from SEQ ID NO: 1-68), albumin (e.g., albumin having a sequence selected from SEQ ID NO: 69-71), an albumin-binding peptide, or an Fc-binding peptide; n is 1 or 2, and when n is 2 and E is an Fc domain monomer, the two Es dimerize to form an Fc domain.

[0128] In some embodiments, in the conjugate represented by formula (II), A1 and A2 are each independently selected from formula (A-I) to formula (A-III) and formula (A-VII),

[0129]

[0130] wherein the definitions of R1, R2, R3, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0131] In some embodiments, in the conjugate represented by formula (II), A1 and A2 are each independently selected from formula (A-I), formula (A-III), and formula (A-VII),

[0132]

[0133] wherein the definitions of R1, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0134] In some embodiments, in the conjugate represented by formula (II), E comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence selected from SEQ ID NO: 1-68); n is 1 or 2, and when n is 2, the two Es dimerize to form an Fc domain.

[0135] In some embodiments, in the conjugate represented by formula (II), E is an Fc domain monomer having an amino acid sequence selected from:

[0136] i) Any one of the sequences shown in SEQ ID NO: 1-68;

[0137] ii) A sequence having one or several amino acid substitutions, deletions or additions (such as 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared with any one of the sequences shown in SEQ ID NO: 1-68; and

[0138] iii) A sequence having at least 70% identity (such as 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) with any one of the sequences shown in SEQ ID NO: 1-68.

[0139] In some embodiments, the substitution described in ii) is a conservative substitution.

[0140] In some embodiments, in the conjugate shown in formula (II), E is a monomer of the Fc domain having at least 70% identity (such as 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) with the sequence of SEQ ID NO: 64.

[0141] In some embodiments, in the conjugate shown in formula (II), E is a monomer of the Fc domain having the sequence of SEQ ID NO: 64.

[0142] In some embodiments, in the conjugate shown in formula (II), E is a monomer of the Fc domain having the sequence of SEQ ID NO: 64, n is 2, and two Es dimerize to form the Fc domain.

[0143] In some embodiments, in the conjugate shown in formula (II), R1 is selected from -OH, -NH2, -NHC(=NH)NH2. In some embodiments, in the conjugate shown in formula (II), R1 is -NHC(=NH)NH2.

[0144] In some embodiments, in the conjugate shown in formula (II), R4 is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3. In some embodiments, in the conjugate shown in formula (II), R4 is selected from -C(=O)OH and -C(=O)OCH3.

[0145] In some embodiments, in the conjugate shown in formula (II), R5 is selected from -C(=O)CH3 and -C(=O)CF3. In some embodiments, in the conjugate shown in formula (II), R5 is -C(=O)CH3.

[0146] In some embodiments, R' in the conjugate represented by formula (II) is H or C1-C4 alkyl. In some embodiments, R' in the conjugate represented by formula (II) is H, methyl or ethyl. In some embodiments, R' in the conjugate represented by formula (II) is H or ethyl.

[0147] In some embodiments, X in the conjugate represented by formula (II) is -O-.

[0148] In some embodiments, Y in the conjugate represented by formula (II) is selected from -O-, -S-, -NH-. In some embodiments, Y in the conjugate represented by formula (II) is selected from -O- and -NH-.

[0149] In some embodiments, the conjugate represented by formula (II) has a structure represented by formula (II-II-A), formula (II-II-B) or formula (II-II-C),

[0150]

[0151] wherein the definitions of R1, R4, R5, R', X, Y, E, n, L, T are as described in any embodiment of the present application.

[0152] In some embodiments, the conjugate represented by formula (II) has a structure represented by formula (II-II-A'), formula (II-II-B') or formula (II-II-C'),

[0153]

[0154] wherein the definitions of R1, R4, R5, R', X, Y, E, n, T are as described in any embodiment of the present application;

[0155] L1 is selected from:

[0156] wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1, Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m, p are each independently an integer from 1 to 20.

[0157] In some embodiments, L1 in the conjugate represented by formula (II) is selected from:

[0158] wherein V, p, m, and Z are defined as described in any embodiment of the present application. In some embodiments, L1 in the conjugate represented by formula (II) is wherein V, p, m, and Z are defined as described in any embodiment of the present application.

[0159] In some embodiments, V in the conjugate represented by formula (II) is -CH2- or -O-. In some embodiments, V in the conjugate represented by formula (II) is -O-.

[0160] In some embodiments, Z in the conjugate represented by formula (II) is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, or -C(=O)-. In some embodiments, Z in the conjugate represented by formula (II) is -NH-C(=O)- or -C(=O)-.

[0161] In some embodiments, p in the conjugate represented by formula (II) is an integer between 1 and 10. In some embodiments, p in the conjugate represented by formula (II) is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6).

[0162] In some embodiments, m in the conjugate represented by formula (II) is an integer between 1 and 12. In some embodiments, m in the conjugate represented by formula (II) is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0163] In some embodiments, the conjugate represented by formula (II) has a structure represented by formula (II-II-A-1), formula (II-II-B-1), formula (II-II-B-2), or formula (II-II-C-1),

[0164]

[0165]

[0166] wherein E, n, T, and L1 are defined as described in any embodiment of the present application.

[0167] In some embodiments, the conjugate of formula (II) has the structure of formula (II-II-A-2), formula (II-II-B-3), formula (II-II-B-4), or formula (II-II-C-2),

[0168]

[0169] wherein the definitions of E, n, T, and L1 are as described in any embodiment of the present application;

[0170] L2 is selected from:

[0171] wherein V is -CH2-, -O-, or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; Z1 and Z2 are independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; m and p are each independently an integer from 1 to 20;

[0172] L3 is wherein U is -C(=O)-NH-, -NH-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, -CH2-; W is -CH2-, -O-, or -S-; i and q are each independently an integer from 1 to 20, one end of L3 is covalently linked to E, and the other end is covalently linked to L2.

[0173] In some embodiments, in the conjugate of formula (II), L3 is covalently linked to E through an N atom, where the N atom is an N atom from E (for example, the N atom on the side chain amino group of a lysine, arginine, asparagine, or glutamine residue).

[0174] In some embodiments, L2 in the conjugate of formula (II) is selected from:

[0175] wherein V is -CH2-, -O-, or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; m and p are each independently an integer from 1 to 20.

[0176] In some embodiments, in the conjugate of formula (II), L1 is wherein V, p, m, and Z are as defined in any embodiment of the present application. In some embodiments, V is -CH2- or -O-. In some embodiments, V is -O-. In some embodiments, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, or -C(=O)-. In some embodiments, Z is -NH-C(=O)- or -C(=O)-. In some embodiments, p is an integer between 1 and 10. In some embodiments, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6). In some embodiments, m is an integer between 1 and 12. In some embodiments, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9);

[0177] In some embodiments, in the conjugate of formula (II), L2 is wherein V, p, m, and Z are as defined in any embodiment of the present application. In some embodiments, V is -CH2- or -O. In some embodiments, V is -O-. In some embodiments, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, or -N(CH2CH3)-C(=O)-. In some embodiments, Z is -C(=O)-NH-. In some embodiments, p is an integer between 1 and 10. In some embodiments, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6). In some embodiments, m is an integer between 1 and 12. In some embodiments, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0178] In some embodiments, in the conjugate of formula (II), L3 is Wherein the definitions of U, i, and q are as described in any embodiment of the present application. In some embodiments, U is -NH-C(=O)-, -N(CH3)-C(=O)-, or -N(CH2CH3)-C(=O)-. In some embodiments, U is -NH-C(=O)-. In some embodiments, the N atom in U is the N atom from E (e.g., the N atom on the side-chain amino group of a lysine, arginine, asparagine, or glutamine residue). In some embodiments, i is an integer between 1 and 12. In some embodiments, i is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9). In some embodiments, q is an integer between 1 and 10. In some embodiments, q is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6).

[0179] In some embodiments, in the conjugate shown in formula (II), T is a number between 1 and 10. In some embodiments, in the conjugate shown in formula (II), T is a number between 1 and 9. In some embodiments, in the conjugate shown in formula (II), T is a number between 1 and 7. In some embodiments, in the conjugate shown in formula (II), T is a number between 1 and 5. In some embodiments, in the conjugate shown in formula (II), T is a number between 3 and 7. In some embodiments, in the conjugate shown in formula (II), T is a number between 3 and 5.5.

[0180] In some embodiments, the conjugate shown in formula (II) has the structure shown in formula (II-II-A-3), formula (II-II-B-5), formula (II-II-B-6), or formula (II-II-C-3),

[0181]

[0182]

[0183] Wherein the definitions of E, n, and T are as described in any embodiment of the present application; n1, n2, and n3 are each independently an integer between 1 and 20. In some embodiments, n1, n2, and n3 are each independently an integer between 1 and 12. In some embodiments, n1, n2, and n3 are each independently an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0184] In some embodiments, the conjugate shown in formula (II) is selected from:

[0185]

[0186]

[0187]

[0188] wherein the definitions of E, n, and T are as described in any embodiment of the present application.

[0189] In another aspect, the present application also provides a compound represented by formula (IV) or a salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof,

[0190]

[0191] wherein A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-XIV),

[0192]

[0193]

[0194] wherein L can be connected to any possible site of the compound of formula (A-XIV);

[0195] wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2, and -NHC(=NH)NHR6;

[0196] R2 and R3 are each independently selected from -H, -OH, -F, -Cl, and -Br;

[0197] R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3, and -C(=O)OCF3;

[0198] R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3;

[0199] X is selected from -O- and -S-;

[0200] Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)-, and -NH(SO2)NR7-;

[0201] R' is H or a C1-C6 alkyl;

[0202] R6 is selected from

[0203]

[0204] R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl;

[0205] L’ is a linker covalently linked to each of the groups A1, A2, and A3.

[0206] In some embodiments, in the compounds of formula (IV), A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-XIII),

[0207]

[0208] wherein the definitions of R1, R2, R3, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0209] In some embodiments, in the compounds of formula (IV), A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-IV) and formula (A-VII),

[0210]

[0211] wherein the definitions of R1, R2, R3, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0212] In some embodiments, in the compounds of formula (IV), A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-III) and formula (A-VII),

[0213]

[0214] wherein the definitions of R1, R2, R3, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0215] In some embodiments, in the compounds of formula (IV), A1, A2, and A3 are each independently selected from formula (A-I), formula (A-III), and formula (A-VII),

[0216]

[0217] wherein the definitions of R1, R4, R5, R’, X, and Y are as described in any embodiment of the present application.

[0218] In some embodiments, R1 in the compound of formula (IV) is selected from -OH, -NH2, -NHC(=NH)NH2. In some embodiments, R1 in the compound of formula (IV) is -NHC(=NH)NH2.

[0219] In some embodiments, R4 in the compound of formula (IV) is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3. In some embodiments, R4 in the compound of formula (IV) is selected from -C(=O)OH and -C(=O)OCH3.

[0220] In some embodiments, R5 in the compound of formula (IV) is selected from -C(=O)CH3 and -C(=O)CF3. In some embodiments, R5 in the compound of formula (IV) is -C(=O)CH3.

[0221] In some embodiments, R' in the compound of formula (IV) is H or C1-C4 alkyl. In some embodiments, R' in the compound of formula (IV) is H, methyl or ethyl. In some embodiments, R' in the compound of formula (IV) is H or ethyl.

[0222] In some embodiments, X in the compound of formula (IV) is -O-.

[0223] In some embodiments, Y in the compound of formula (IV) is selected from -O-, -S-, -NH-. In some embodiments, Y in the compound of formula (IV) is selected from -O- and -NH-.

[0224] In some embodiments, the compound of formula (IV) has the structure of formula (IV-I),

[0225]

[0226] wherein the definitions of A1, A2 and A3 are as described in any embodiment of the present application,

[0227] L1 is selected from:

[0228]

[0229] L2' is selected from:

[0230]

[0231] wherein V is -CH2-, -O-, or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, -CH2- Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, or -CH2-; W’ is HC≡C-, -C(=O)OH, -NH2 or N3-; m and p are each independently an integer from 1 to 20.

[0232] In some embodiments, L1 in the compound of formula (IV) is selected from:

[0233] wherein V, p, m, and Z are as defined in any embodiment of the present application. In some embodiments, L1 in the compound of formula (IV) is wherein V, p, m, and Z are as defined in any embodiment of the present application.

[0234] In some embodiments, L2’ in the compound of formula (IV) is wherein W’, V, p, m, and Z are as defined in any embodiment of the present application.

[0235] In some embodiments, L2’ in the compound of formula (IV) is wherein W’, V, p, m, and Z are as defined in any embodiment of the present application.

[0236] In some embodiments, L2’ in the compound of formula (IV) is wherein W’, p, m, and Z are as defined in any embodiment of the present application.

[0237] In some embodiments, V in the compound of formula (IV) is -CH2- or -O-. In some embodiments, V in the compound of formula (IV) is -O-.

[0238] In some embodiments, in the compound of formula (IV), Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-.

[0239] In some embodiments, in the compound of formula (IV), Z is wherein the definitions of p, V, m, and Z1 are as described in any embodiment of the present application.

[0240] In some embodiments, in the compound of formula (IV), Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)- or -C(=O)-. In some embodiments, in the compound of formula (IV), Z is -NH-C(=O)- or -C(=O)-. In some embodiments, in the compound of formula (IV), Z is -NH-C(=O)-.

[0241] In some embodiments, in the compound of formula (IV), Z1 is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)- or -CH2CH2-C(=O)-. In some embodiments, in the compound of formula (IV), Z1 is -C(=O)-NH- or -CH2-C(=O)-NH-. In some embodiments, in the compound of formula (IV), Z1 is -C(=O)-NH-.

[0242] In some embodiments, in the compound of formula (IV), W' is HC≡C-, -C(=O)OH, -NH2 or N3-. In some embodiments, in the compound of formula (IV), W' is HC≡C-. In some embodiments, in the compound of formula (IV), W' is

[0243] In some embodiments, in the compound of formula (IV), p is an integer between 1 and 10. In some embodiments, in the compound of formula (IV), p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6).

[0244] In some embodiments, in the compound of formula (IV), m is an integer between 1 and 12. In some embodiments, in the compound of formula (IV), m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0245] In some embodiments, the compound represented by formula (IV) has the structures represented by formula (IV-I-1), formula (IV-I-2), and formula (IV-I-3):

[0246]

[0247]

[0248] wherein the definitions of R1, R4, R5, R’, X, Y, L1, and L2’ are as defined in any embodiment of the present application.

[0249] In some embodiments, the compound represented by formula (IV) has the structures represented by formula (IV-I-1-1), formula (IV-I-2-1), formula (IV-I-2-2), or formula (IV-I-3-1):

[0250]

[0251]

[0252] wherein the definitions of L1 and L2’ are as defined in any embodiment of the present application.

[0253] In some embodiments, the compound represented by formula (IV) is selected from:

[0254]

[0255]

[0256]

[0257]

[0258]

[0259] On the other hand, the present application also provides a compound represented by formula (V) or a salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof,

[0260]

[0261] wherein A1 and A2 are each independently selected from formula (A-I) to formula (A-XIV),

[0262]

[0263]

[0264] Among them, L can be connected to any possible site of the compound of formula (A-XIV); wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2, and -NHC(=NH)NHR6;

[0265] R2 and R3 are each independently selected from -H, -OH, -F, -Cl, and -Br;

[0266] R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3, and -C(=O)OCF3;

[0267] R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3;

[0268] X is selected from -O- and -S-;

[0269] Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)-, and -NH(SO2)NR7-;

[0270] R' is H or C1-C6 alkyl;

[0271] R6 is selected from

[0272]

[0273]

[0274] R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl, and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl;

[0275] L' is a linker covalently connected to each of the groups A1 and A2.

[0276] In some embodiments, in the compound of formula (V), A1 and A2 are each independently selected from formula (A-I) to formula (A-XIII),

[0277]

[0278] Wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0279] In some embodiments, in the compound of formula (V), A1 and A2 are each independently selected from formula (A-I) to formula (A-IV) and formula (A-VII),

[0280]

[0281] Wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0282] In some embodiments, in the compound of formula (V), A1 and A2 are each independently selected from formula (A-I) to formula (A-III) and formula (A-VII),

[0283]

[0284] Wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0285] In some embodiments, in the compound of formula (V), A1 and A2 are each independently selected from formula (A-I), formula (A-III), and formula (A-VII),

[0286]

[0287] Wherein the definitions of R1, R4, R5, R', X, and Y are as described in any embodiment of the present application.

[0288] In some embodiments, in the compound of formula (V), R1 is selected from -OH, -NH2, -NHC(=NH)NH2. In some embodiments, in the compound of formula (V), R1 is -NHC(=NH)NH2.

[0289] In some embodiments, in the compound of formula (V), R4 is selected from -C(=O)OH, -C(=O)OCH3, and -C(=O)OCF3. In some embodiments, in the compound of formula (V), R4 is selected from -C(=O)OH and -C(=O)OCH3.

[0290] In some embodiments, in the compound of formula (V), R5 is selected from -C(=O)CH3 and -C(=O)CF3. In some embodiments, in the compound of formula (V), R5 is -C(=O)CH3.

[0291] In some embodiments, R' in the compound of formula (V) is H or C1-C4 alkyl. In some embodiments, R' in the compound of formula (V) is H, methyl or ethyl. In some embodiments, R' in the compound of formula (V) is H or ethyl.

[0292] In some embodiments, X in the compound of formula (V) is -O-.

[0293] In some embodiments, Y in the compound of formula (V) is selected from -O-, -S-, -NH-. In some embodiments, Y in the compound of formula (V) is selected from -O- and -NH-.

[0294] In some embodiments, the compound of formula (V) has the structure shown in formula (V-I),

[0295]

[0296] wherein the definitions of A1 and A2 are as described in any embodiment of the present application,

[0297] L1 is selected from:

[0298]

[0299] L2' is selected from:

[0300]

[0301] wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; W' is HC≡C-, -C(=O)OH, -NH2 or N3-; m and p are each independently an integer from 1 to 20.

[0302] In some embodiments, L1 in the compound of formula (V) is selected from:

[0303]

[0304] Wherein the definitions of V, p, m, and Z are as described in any embodiment of the present application.

[0305] In some embodiments, in the compound of formula (V), L1 is Wherein the definitions of V, p, m, and Z are as described in any embodiment of the present application.

[0306] In some embodiments, in the compound of formula (V), L2’ is Wherein the definitions of W’, V, p, m, and Z are as described in any embodiment of the present application.

[0307] In some embodiments, in the compound of formula (V), L2’ is Wherein the definitions of W’, V, p, m, and Z are as described in any embodiment of the present application.

[0308] In some embodiments, in the compound of formula (V), V is -CH2- or -O-. In some embodiments, in the compound of formula (V), V is -O-.

[0309] In some embodiments, in the compound of formula (V), Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, or -C(=O)-. In some embodiments, in the compound of formula (V), Z is -NH-C(=O)- or -C(=O)-.

[0310] In some embodiments, in the compound of formula (V), W’ is HC≡C-, -C(=O)OH, -NH2, or N3-. In some embodiments, in the compound of formula (V), W’ is HC≡C-.

[0311] In some embodiments, in the compound of formula (V), p is an integer between 1 and 10. In some embodiments, in the compound of formula (V), p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6).

[0312] In some embodiments, in the compound of formula (V), m is an integer between 1 and 12. In some embodiments, in the compound of formula (V), m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

[0313] In some embodiments, the compound of formula (V) has the structures shown in formula (V-I-1), formula (V-I-2), and formula (V-I-3):

[0314]

[0315] Wherein the definitions of R1, R4, R5, R’, X, Y, L1, and L2’ are as described in any embodiment of the present application.

[0316] In some embodiments, the compound represented by formula (V) has a structure represented by formula (V-I-1-1), formula (V-I-2-1), formula (V-I-2-2), or formula (V-I-3-1):

[0317]

[0318] Wherein the definitions of L1 and L2’ are as described in any embodiment of the present application.

[0319] In some embodiments, the compound represented by formula (V) is selected from:

[0320]

[0321]

[0322]

[0323]

[0324] In another aspect, the present application further provides a pharmaceutical composition comprising the conjugate represented by formula (I) or formula (II) or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof, and a pharmaceutically acceptable carrier and / or excipient.

[0325] In another aspect, the present application further provides a pharmaceutical composition comprising the compound represented by formula (IV) or formula (V) or a salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof, and a pharmaceutically acceptable carrier and / or excipient.

[0326] In another aspect, the present application further provides the use of the conjugate represented by formula (I) or formula (II) or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof in the preparation of a drug for the treatment or prevention of viral infection.

[0327] In another aspect, the present application further provides the use of the compound represented by formula (IV) or formula (V) or a salt, geometric or optical isomer, hydrate, solvate, or polymorph thereof in the preparation of a drug for the treatment or prevention of viral infection.

[0328] In another aspect, the present application further provides the use of the pharmaceutical composition in the preparation of a drug for the treatment or prevention of viral infection.

[0329] In another aspect, the present application also provides a method for treating a subject having a viral infection or suspected of having a viral infection, the method comprising administering to the subject an effective amount of the conjugate represented by formula (I) or formula (II) of the present application or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof.

[0330] In another aspect, the present application also provides a method for treating a subject having a viral infection or suspected of having a viral infection, the method comprising administering to the subject an effective amount of the compound represented by formula (IV) or formula (V) of the present application or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof.

[0331] In another aspect, the present application also provides a method for prophylactically treating a viral infection in a subject in need thereof, the method comprising administering to the subject an effective amount of the conjugate represented by formula (I) or formula (II) of the present application or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof.

[0332] In some embodiments, the viral infection is an infection caused by an influenza virus or a parainfluenza virus. In some embodiments, the viral infection is an infection caused by an influenza A, B or C virus or a parainfluenza virus. In some embodiments, the viral infection is an infection caused by avian influenza.

[0333] In some embodiments, the subject may be further treated with an antiviral agent selected from oseltamivir, zanamivir, peramivir, laninamivir, amantadine, rimantadine or baloxavir marboxil.

[0334] In some embodiments, the subject is immunocompromised. In some embodiments, the subject has been diagnosed with a humoral immunodeficiency, a T cell deficiency, neutropenia, asplenia or a complement deficiency. In some embodiments, the subject is being treated or is about to be treated with an immunosuppressive therapy. In some embodiments, the subject has been diagnosed with a disease causing immunosuppression. In some embodiments, the disease is cancer or acquired immunodeficiency syndrome. In some embodiments, the cancer is leukemia, lymphoma or multiple myeloma. In some embodiments, the subject has undergone or is about to undergo a hematopoietic stem cell transplantation. In some embodiments, the subject has undergone or is about to undergo an organ transplantation.

[0335] In some embodiments, the conjugate represented by formula (I) or formula (II) described in the present application is capable of binding to one or more targets (e.g., antigens). In some embodiments, the target is a viral (e.g., influenza) protein, such as neuraminidase or hemagglutinin.

[0336] In some embodiments, the conjugate represented by formula (I) or formula (II), or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, or composition is administered intramuscularly, intravenously, intradermally, intraarterially, intraperitoneally, intralesionally, intracranially, intraarticularly, intraprostatically, intrathoracically, intratracheally, intranasally, intravitreally, intravaginally, rectally, transdermally, intratumorally, transperitoneally, subcutaneously, subconjunctivally, intracapsularly, transmucosally, intracardially, intraumbilically, intraocularly, orally, topically, by inhalation, by injection or by infusion.

[0337] In another aspect, the present application also provides the use of the compound represented by formula (IV) or formula (V), or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, as an intermediate in the preparation of a conjugate. The compound represented by formula (IV) or formula (V) as an intermediate can be conjugated to an Fc domain or an Fc domain monomer (e.g., by means of a linker) by any suitable method known to those skilled in the art, including any method described or exemplified in the present application. In some embodiments, the conjugate (e.g., the conjugate represented by formula (I) or formula (II)) comprises E, where E is an Fc domain monomer (when n is 2, E forms an Fc domain as a dimer, and each Fc domain monomer independently has the sequence of any one of SEQ ID NOs: 1-68). In a preferred embodiment, one or more nitrogen atoms of one or more surface-exposed lysine residues of E are covalently conjugated to a linker (e.g., a PEG2-PEG20 linker). The linker conjugated to E can be functionalized so that it can react to form a covalent bond with any intermediate described herein (e.g., the compound represented by formula (IV) or formula (V)). In a preferred embodiment, E is conjugated to an azide-functionalized linker and the intermediate (e.g., the compound represented by formula (IV) or formula (V)) is alkynyl-functionalized. E is conjugated to the intermediate through the azide of the linker and the alkyne of the intermediate (e.g., by click chemistry) to form the conjugate of the present application, such as the conjugate represented by formula (I) or formula (II).

[0338] In some embodiments, in the conjugates of the present application, each E is an Fc domain monomer, including a hinge domain, a CH2 antibody constant domain, and a CH3 antibody constant domain. The Fc domain monomer can be an immunoglobulin antibody isotype IgG. The variant Fc domain monomer can also be any immunoglobulin antibody isotype (e.g., IgG1). The variant Fc domain monomer can be any immunoglobulin antibody allotype (e.g., IGHG1*01 (i.e., G1m(za)), IGHG1*07 (i.e., G1m(zax)), IGHG1*04 (i.e., G1m(zav)), IGHG1*03 (G1m(f)), IGHG1*08 (i.e., G1m(fa)), IGHG2*01, IGHG2*06, or IGHG2*02) (as described, for example, in Vidarsson et al. IgG subclasses and allotypes: from structure to effector function Frontiers in Immunology. 5(520):1-17(2014)). The variant Fc domain monomer can also be of any species, such as human, murine, or mouse. The dimer of the variant Fc domain monomer is a variant Fc domain that can bind to an Fc receptor, which is a receptor located on the surface of leukocytes.

[0339] In some embodiments, the Fc domain monomer includes the quadruple mutation C220S / M252Y / S254T / T256E relative to the wild-type Fc domain monomer. In some embodiments, the Fc domain monomer includes the quadruple mutation C220S / V309D / Q311H / N434S relative to the wild-type Fc domain monomer. In other embodiments, the Fc domain monomer includes the C220S mutation relative to the wild-type Fc domain monomer. The mutation is relative to the wild-type Fc monomer amino acid sequence, such as wild-type human IgG1.

[0340] In some embodiments, the Fc domain monomer in the conjugates of the present application can include one or more amino acid substitutions, additions, and / or deletions relative to the Fc domain monomer shown by the sequence of any one of SEQ ID NOs: 1-68. In some embodiments, the Fc domain monomer can include one or more amino acid substitutions, additions, and / or deletions relative to the Fc domain monomer shown by the sequence of any one of SEQ ID NOs: 1 to 29, 31 to 52, or 56 to 58. In some embodiments, Asn297 in the Fc domain monomer in the conjugates of the present application can be replaced by Ala to prevent N-linked glycosylation (see, for example, SEQ ID NO: 4, where the substitution of Asn297 to Ala is marked as (*)).

[0341] In some embodiments, the Fc domain monomer or Fc domain in the conjugate described in the present application is a non-glycosylated Fc domain monomer or Fc domain (e.g., an Fc domain monomer or Fc domain that maintains engagement with an Fc receptor (e.g., FcRn)). For example, the Fc domain is a non-glycosylated IgG1 variant that maintains engagement with an Fc receptor (e.g., IgG1 having amino acid substitutions at N297 and / or T299 of the glycosylation motif). Exemplary non-glycosylated Fc domains and methods for preparing non-glycosylated Fc domains are known in the art, for example, as described in Sazinsky S.L. et al., Aglycosylated immunoglobulin G1 variants productively engage activating Fc receptors, PNAS, 2008, 105(51):20167-20172.

[0342] In some embodiments, the C-terminal Lys447 of the Fc domain monomer in the conjugate described in the present application may be present or absent without affecting the structure or stability of the Fc region, such as any one of SEQ ID NOs: 1 to 29 and 31 to 52 that does not include a C-terminal Lys residue. In some embodiments, the N-terminal Asn of the Fc domain monomer in the conjugate described in the present application may be present or absent without affecting the structure or stability of the variant Fc domain monomer, such as any one of SEQ ID NOs: 1 to 29, 31 to 52, and 56 to 58 that does not include an N-terminal Asn residue.

[0343] In some embodiments, the Fc domain monomer in the conjugate described in the present application includes an additional moiety attached to the N or C terminus of the Fc domain monomer, such as a purification peptide (e.g., a hexahistidine peptide (HHHHHH (SEQ ID NO: 59))) or a signal sequence (e.g., the IL2 signal sequence MYRMQLLSCIALSLALVTNS (SEQ ID NO: 60)). In some embodiments, the Fc domain monomer in the conjugate does not contain any type of antibody variable region, such as VH, VL, complementarity-determining region (CDR), or hypervariable region (HVR).

[0344] In some embodiments, the Fc domain monomer in the conjugate described in the present application has a sequence with at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) to any one of SEQ ID NOs: 1-68 shown below. In some embodiments, the Fc domain monomer has the sequence of any one of SEQ ID NOs: 1-68 shown below.

[0345] SEQ ID NO:1: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), X1 is Asp or Glu, and X2 is Leu or Met, N-terminal Fab residue underlined, hinge residue italicized

[0346]

[0347] SEQ ID NO:2: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residue underlined, hinge residue italicized

[0348]

[0349] SEQ ID NO:3: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residue underlined, hinge residue italicized

[0350]

[0351] SEQ ID NO:4: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), Asn to Ala substitution (*), X1 is Asp or Glu, and X2 is Leu or Met, N-terminal Fab residue underlined, hinge residue italicized

[0352]

[0353] SEQ ID NO:5: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), Asn to Ala substitution (*), N-terminal Fab residue underlined, hinge residue italicized

[0354]

[0355] SEQ ID NO:6: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), Asn-to-Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0356]

[0357] SEQ ID NO:7: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), X6 is Asp or Glu, and X7 is Leu or Met, Z1 is Asn or absent, Z2 is Asn or Ala, Z3 is Lys or absent, N-terminal Fab residues underlined, hinge residues italicized

[0358]

[0359] SEQ ID NO:8: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0360]

[0361] SEQ ID NO:9: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0362]

[0363] SEQ ID NO:10: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0364]

[0365] SEQ ID NO:11: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0366]

[0367] SEQ ID NO:12: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0368]

[0369] SEQ ID NO:13: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0370]

[0371] SEQ ID NO:14: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), Asn-to-Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0372]

[0373] SEQ ID NO:15: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), Asn-to-Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0374]

[0375] SEQ ID NO:16: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), Asn-to-Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0376]

[0377] SEQ ID NO:17: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), Asn-to-Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0378]

[0379] SEQ ID NO:18: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(fa) (bold italic), Asn to Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0380]

[0381] SEQ ID NO:19: Adult IgG1 Fc, Cys to Ser substitution (#), YTE triple mutation (bold and underlined), allotype G1m(f) (bold italic), Asn to Ala substitution (*), N-terminal Fab residues underlined, hinge residues italicized

[0382]

[0383] SEQ ID NO:20: Adult IgG1 Fc, Cys to Ser substitution (#), X4 is Asp or Glu, X5 is Leu or Met; Z1 is Asn or absent, Z3 is Lys or absent, N-terminal Fab residues underlined, hinge residues italicized

[0384]

[0385] SEQ ID NO:21: Adult IgG1 Fc, Cys to Ser substitution (#), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0386]

[0387] SEQ ID NO:22: Adult IgG1 Fc, Cys to Ser substitution (#), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0388]

[0389] SEQ ID NO:23: Adult IgG1 Fc, Cys to Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, X6 is Asp or Glu, X7 is Leu or Met, Z1 is Asn or absent, Z2 is Asn or Ala, and Z3 is Lys or absent, N-terminal Fab residues underlined, hinge residues italicized

[0390]

[0391] SEQ ID NO:24: Adult IgG1 Fc, Cys-to-Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, X6 is Asp or Glu, X7 is Leu or Met, and Z2 is Asn or Ala, N-terminal Fab residues are underlined, hinge residues are italicized

[0392]

[0393] SEQ ID NO:25: Adult IgG1 Fc, Cys-to-Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, and Z2 is Asn or Ala, N-terminal Fab residues are underlined, hinge residues are italicized

[0394]

[0395] SEQ ID NO:26: Adult IgG1 Fc, Cys-to-Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, and Z2 is Asn or Ala, N-terminal Fab residues are underlined, hinge residues are italicized

[0396]

[0397] SEQ ID NO:27: Adult IgG1 Fc, Cys-to-Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, X6 is Asp or Glu, X7 is Leu or Met, and Z2 is Asn or Ala, N-terminal Fab residues are underlined, hinge residues are italicized

[0398]

[0399] SEQ ID NO:28: Adult IgG1 Fc, Cys-to-Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, and Z2 is Asn or Ala, N-terminal Fab residues are underlined, hinge residues are italicized

[0400]

[0401] SEQ ID NO:29: Adult IgG1 Fc, Cys-to-Ser substitution (#), X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, and Z2 is Asn or Ala, N-terminal Fab residues are underlined, hinge residues are italicized

[0402]

[0403] SEQ ID NO:30: Adult IgG1 Fc with murine heavy chain MigG Vh signal sequence (bold), Cys to Ser substitution (#), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0404]

[0405] SEQ ID NO:31: Adult Fc IgG1, where Z1 is Asn or absent, Z3 is Lys or absent, J1 is Cys or Ser, and where X1 is Met or Tyr, X2 is Ser or Thr, X3 is Thr or Glu, Z3 is Asn or Ala, X4 is Leu or Asp, X5 is Gln or His, X6 is Asp or Glu, and X7 is Leu or Met, X8 is Met or Leu, and X9 is Asn or Ser, Z2 is Asn or Ala, N-terminal Fab residues underlined, hinge residues italicized

[0406]

[0407] SEQ ID NO:32: Adult Fc IgG1, Cys to Ser substitution (#), where Z1 is Asn or absent, Z3 is Lys or absent, and where Z2 is Asn or Ala, X4 is Leu or Asp, X5 is Gln or His, X6 is Asp or Glu, X7 is Leu or Met, X8 is Met or Leu, and X9 is Asn or Ser, N-terminal Fab residues underlined, hinge residues italicized

[0408]

[0409] SEQ ID NO:33: Adult Fc IgG1, Cys to Ser substitution (#), DHS triple mutation (bold and underlined), where Z1 is Asn or absent, Z3 is Lys or absent, and where Z2 is Asn or Ala, X6 is Asp or Glu, and X7 is Leu or Met, X8 is Met or Leu, and X9 is Asn or Ser, N-terminal Fab residues underlined, hinge residues italicized

[0410]

[0411] SEQ ID NO:34: Adult Fc IgG1, Cys-to-Ser substitution (#), DHS triple mutation (bolded and underlined), where Z2 is Asn or Ala, X6 is Asp or Glu, and X7 is Leu or Met, N-terminal Fab residues underlined, hinge residues italicized

[0412]

[0413] SEQ ID NO:35: Adult Fc IgG1, Cys-to-Ser substitution (#), DHS triple mutation (bolded and underlined), where X6 is Asp or Glu and X7 is Leu or Met, N-terminal Fab residues underlined, hinge residues italicized

[0414]

[0415] SEQ ID NO:36: Adult Fc IgG1, Cys-to-Ser substitution (#), DHS triple mutation (bolded and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0416]

[0417] SEQ ID NO:37: Adult Fc IgG1, Cys-to-Ser substitution (#), DHS triple mutation (bolded and underlined

[0418]

[0419] SEQ ID NO:38: Adult Fc IgG1, Cys-to-Ser substitution (#), DHS triple mutation (bolded and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0420]

[0421] SEQ ID NO:39: Adult Fc IgG1, Cys-to-Ser substitution (#), DHS triple mutation (bolded and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0422]

[0423] SEQ ID NO:40: Adult Fc IgG1, Cys to Ser substitution (#), DHS triple mutations (bolded and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0424]

[0425] SEQ ID NO:41: Adult Fc IgG1, Cys to Ser substitution (#), DHS triple mutations (bolded and underlined)

[0426]

[0427] SEQ ID NO:42: Adult Fc IgG1, Cys to Ser substitution (#), DHS triple mutations (bolded and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0428]

[0429] SEQ ID NO:43: Adult Fc IgG1, Cys to Ser substitution (#), DHS triple mutations (bolded and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0430]

[0431] SEQ ID NO:44: Adult Fc IgG1, Cys to Ser substitution (#), Asn to Ala substitution (*), DHS triple mutations (bolded and underlined), where X6 is Asp or Glu and X7 is Leu or Met, N-terminal Fab residues underlined, hinge residues italicized

[0432]

[0433] SEQ ID NO:45: Adult Fc IgG1, Cys to Ser substitution (#), Asn to Ala substitution (*), DHS triple mutations (bolded and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0434]

[0435] SEQ ID NO:46: Adult Fc IgG1, Cys-to-Ser substitution (#), Asn-to-Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0436]

[0437] SEQ ID NO:47: Adult Fc IgG1, Cys-to-Ser substitution (#), Asn-to-Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0438]

[0439] SEQ ID NO:48: Adult Fc IgG1, Cys-to-Ser substitution (#), Asn-to-Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0440]

[0441] SEQ ID NO:49: Adult Fc IgG1, Cys-to-Ser substitution (#), Asn-to-Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0442]

[0443]

[0444] SEQ ID NO:50: Adult Fc IgG1, Cys-to-Ser substitution (#), Asn-to-Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0445]

[0446] SEQ ID NO:51: Adult Fc IgG1, Cys-to-Ser substitution (#), Asn-to-Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0447]

[0448] SEQ ID NO:52: Adult Fc IgG1, Cys to Ser substitution (#), Asn to Ala substitution (*), DHS triple mutation (bold and underlined), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italic

[0449]

[0450] SEQ ID NO:53: Adult Fc IgG1, added N-terminal ISAMVRS amino acid residues (italic), C-terminal G4S linker (italic), C-terminal myc tag (underlined), allotype G1m(f) (bold italic)

[0451]

[0452] SEQ ID NO:54: Adult Fc IgG1, added N-terminal ISAMVRS amino acid residues (italic), allotype G1m(fa) (bold italic)

[0453]

[0454] SEQ ID NO:55: Adult Fc IgG1, added N-terminal amino acid residues (italic), hinge residues (italic), allotype G1m(fa) (bold italic)

[0455]

[0456] SEQ ID NO:56: Adult IgG1 Fc, Cys to Ser substitution (#), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italic

[0457]

[0458] SEQ ID NO:57: Adult IgG1 Fc, Cys to Ser substitution (#), allotype G1m(f) (bold italic), N-terminal Fab residues underlined, hinge residues italic

[0459]

[0460] SEQ ID NO:58: Adult IgG1 Fc, Cys to Ser substitution (#), M428L, N434S (bold / underlined), allotype G1m(fa) (bold italic), N-terminal Fab residues underlined, hinge residues italicized

[0461]

[0462] SEQ ID NO:59: Adult IgG1 with murine heavy chain MigG1 signal sequence (bold), with Cys to Ser substitution (#), with M428L, N434S mutations (bold / underlined), with C-terminal G4S (italic) and with C-terminal IgA peptide (underlined), with allotype G1m(fa) (bold italic)

[0463]

[0464] SEQ ID NO:60: Adult Fc IgG1, where Z1 is Cys or Ser, and where X1 is Met or Trp, X2 is Ser or Thr, X3 is Thr or Glu, X4 is Asp or Glu, and X5 is Leu or Met, X6 is Met or Leu, and X7 is Asn or Ser

[0465]

[0466] SEQ ID NO:61: Adult Fc IgG1, Cys to Ser substitution (#), and where X1 is Met or Trp, X2 is Ser or Thr, X3 is Thr or Glu, X4 is Asp or Glu, and X5 is Leu or Met, X6 is Met or Leu, and X7 is Asn or Ser

[0467]

[0468] SEQ ID NO:62: Adult IgG1 Fc, Cys to Ser substitution (#), X4 is Asp or Glu, and X5 is Leu or Met

[0469]

[0470] SEQ ID NO:63: Adult IgG1 Fc, Cys to Ser substitution (#), allotype G1m(f) (bold italic)

[0471]

[0472] SEQ ID NO:64: Adult IgG1 Fc, Cys-to-Ser substitution (#), allotype G1m(fa) (bold italic)

[0473]

[0474] SEQ ID NO:65: Adult IgG1 Fc, Cys-to-Ser substitution (#), M428L, N434S mutations (bold / underlined), allotype G1m(fa) (bold italic)

[0475]

[0476] SEQ ID NO:66: Adult IgG1 Fc, Cys-to-Ser substitution (#), M428L, N434S mutations (bold / underlined), allotype G1m(f) (bold italic)

[0477]

[0478] SEQ ID NO:67: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutations (bold and underlined), allotype G1m(fa) (bold italic)

[0479]

[0480] SEQ ID NO:68: Adult IgG1 Fc, Cys-to-Ser substitution (#), YTE triple mutations (bold and underlined), allotype G1m(f) (bold italic)

[0481]

[0482] In some embodiments, in the conjugates of the present application, each E is a naturally occurring albumin or a variant thereof, such as an engineered variant of a naturally occurring albumin. The variants include polymorphisms, fragments such as domains and subdomains, and fusion proteins. The albumin can include the sequence of albumin obtained from any source. Preferably, the source is mammalian, such as human or bovine. Most preferably, the albumin is human serum albumin (HSA) or a variant thereof. Human serum albumin includes any albumin having the amino acid sequence naturally occurring in humans and variants thereof. The albumin coding sequence can be obtained by methods known to those skilled in the art for isolating the cDNA corresponding to the human gene and sequencing the cDNA. The albumin of the present application can include the amino acid sequence of human serum albumin (HSA) shown in SEQ ID NO: 69 or SEQ ID NO: 70, or the amino acid sequence of mouse serum albumin (MSA) shown in SEQ ID NO: 71, or a variant or fragment thereof, preferably a variant or fragment having its function. The fragment or variant may or may not be functional, or may retain the function of albumin to some extent. For example, the fragment or variant can retain the ability to bind to an albumin receptor, such as HSA or MSA, at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or 105% of the ability of the parental albumin (e.g., the parental albumin from which the fragment or variant is produced). The relative binding ability can be determined by methods known in the art, such as by surface plasmon resonance.

[0483] SEQ ID NO:69 (Human Serum Albumin (HSA), Variant 1)

[0484]

[0485] SEQ ID NO:70 (Human Serum Albumin (HSA), Variant 2)

[0486]

[0487]

[0488] SEQ ID NO:71 (Mouse Serum Albumin (MSA))

[0489]

[0490] In some embodiments of any aspect of the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 1. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 1.

[0491] In some embodiments of any aspect of the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 2. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 2.

[0492] In some embodiments of any aspect of the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 3. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 3.

[0493] In some embodiments of any aspect of the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 4. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 4.

[0494] In some embodiments of any aspect of the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 5. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 5.

[0495] In some embodiments of any aspect of the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 6. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 6.

[0496] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO:7. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO:7.

[0497] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO:8. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO:8.

[0498] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO:9. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO:9.

[0499] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO:10. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO:10.

[0500] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO:11. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO:11.

[0501] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO:12. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO:12.

[0502] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 13. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 13.

[0503] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 14. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 14.

[0504] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 15. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 15.

[0505] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 16. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 16.

[0506] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 17. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 17.

[0507] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 18. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 18.

[0508] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 19. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 19.

[0509] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 20. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 20.

[0510] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 21. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 21.

[0511] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 22. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 22.

[0512] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 23. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 23.

[0513] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 24. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 24.

[0514] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 25. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 25.

[0515] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 26. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 26.

[0516] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 27. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 27.

[0517] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 28. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 28.

[0518] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 29. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 29.

[0519] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO:30. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO:30.

[0520] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO:31. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO:31.

[0521] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO:32. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO:32.

[0522] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO:33. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO:33.

[0523] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO:34. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO:34.

[0524] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO:35. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO:35.

[0525] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 36. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 36.

[0526] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 37. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 37.

[0527] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 38. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 38.

[0528] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 39. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 39.

[0529] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 40. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 40.

[0530] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 41. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 41.

[0531] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 42. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 42.

[0532] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 43. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 43.

[0533] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 44. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 45.

[0534] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 46. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 46.

[0535] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 47. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 47.

[0536] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 48. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 48.

[0537] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 49. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 49.

[0538] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 50. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 50.

[0539] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 51. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 51.

[0540] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 52. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 52.

[0541] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 53. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 53.

[0542] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 54. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 54.

[0543] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 55. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 55.

[0544] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 56. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 56.

[0545] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 57. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 57.

[0546] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 58. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 58.

[0547] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 59. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 59.

[0548] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 60. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 60.

[0549] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 61. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 61.

[0550] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 62. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 62.

[0551] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 63. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 63.

[0552] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence set forth in SEQ ID NO: 64. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence set forth in SEQ ID NO: 64.

[0553] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 65. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 65.

[0554] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 66. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 66.

[0555] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 67. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 67.

[0556] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 68. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 68.

[0557] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 69. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 69.

[0558] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 70. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 70.

[0559] In some embodiments of any aspect described in the present application, E (e.g., each E) comprises the amino acid sequence shown in SEQ ID NO: 71. In some embodiments, E comprises an amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to the amino acid sequence shown in SEQ ID NO: 71.

[0560] Definitions

[0561] As used herein, the term "half maximal inhibitory concentration" refers to the concentration of a drug required to inhibit 50% in vitro.

[0562] As used herein, the term "viral infection" means the pathogenic growth of a virus (e.g., influenza virus) in a host organism (e.g., a mouse). A viral infection can be any situation in which the presence of a viral population is damaging to the host body. Thus, a subject "is experiencing" a viral infection when there is an excessive viral population in or on the body of the host organism, or when the presence of the viral population is damaging the cells or other tissues of the subject.

[0563] As used herein, the term "neuraminidase inhibitor" refers to a compound that reduces the activity of influenza virus neuraminidase (e.g., from influenza A, B or C virus). Viral neuraminidase inhibitors known to those skilled in the art include zanamivir, peramivir, oseltamivir and their analogs. The neuraminidase inhibitors described in the present application include zanamivir, peramivir, oseltamivir and their analogs, such as the neuraminidase inhibitors shown in formulas (A-I)-(A-XIII) in the conjugates.

[0564] As used herein, the term "Fc domain monomer" refers to a polypeptide chain that includes at least one hinge domain and second and third antibody constant domains (CH2 and CH3) or functional fragments thereof (e.g., fragments capable of (i) dimerizing with another Fc domain monomer to form an Fc domain, and (ii) binding to an Fc receptor). The Fc domain monomer can be any immunoglobulin antibody isotype, including IgG, IgE, IgM, IgA, or IgD (e.g., IgG). Additionally, the Fc domain monomer can be an IgG subtype including IgG1, IgG2a, IgG2b, IgG3, or IgG4 (e.g., IgG1). The Fc domain monomer does not include any portion of an immunoglobulin that can serve as an antigen-recognition region, such as a variable domain or complementarity-determining region (CDR). The Fc domain monomer in the conjugates described in the present application can contain one or more alterations (e.g., 1-10, 1-8, 1-6, 1-4 amino acid substitutions, additions, or deletions) relative to the wild-type Fc domain monomer sequence, which alter the interaction between the Fc domain and the Fc receptor. Examples of suitable alterations are known in the art. In certain embodiments, a human Fc domain monomer (e.g., an IgG heavy chain, such as IgG1) includes a region extending from any one of Asn208, Glu216, Asp221, Lys222, or Cys226 to the carboxyl terminus of the heavy chain at Lys447. The presence or absence of the C-terminal Lys447 of the Fc region does not affect the structure or stability of the Fc region. Unless otherwise specified herein, the numbering of amino acid residues in the IgG or Fc domain monomer is according to the EU numbering system for antibodies, which is also referred to as the Kabat EU index, as described, for example, in Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0565] As used herein, the term "Fc domain" refers to a dimer of two Fc domain monomers that is capable of binding to an Fc receptor. In a wild-type Fc domain, the two Fc domain monomers dimerize through the interaction between two CH3 antibody constant domains, and in some embodiments, one or more disulfide bonds are formed between the hinge domains of the two dimerized Fc domain monomers.

[0566] As used herein, the term "covalently linked" refers to two moieties in a conjugate that are linked to each other by a covalent bond formed between two atoms in the two moieties of the conjugate.

[0567] As used herein, the term "Fc-binding peptide" refers to a polypeptide having an amino acid sequence of 5 to 50 (e.g., 5 to 40, 5 to 30, 5 to 20, 5 to 15, 5 to 10, 10 to 50, 10 to 30, or 10 to 20) amino acid residues that has an affinity for an Fc domain (e.g., any Fc domain described in the present application) and has the function of binding to the Fc domain. The Fc-binding peptide can have different origins, such as synthetic, human, murine, or rat. The Fc-binding peptides of the present application include Fc-binding peptides that have been engineered to include one or more (e.g., two, three, four, or five) solvent-exposed lysine residues, which can provide sites for conjugation to the compounds of the present application (e.g., conjugation to a dimer or trimer of a neuraminidase inhibitor of formula (IV) or formula (V), e.g., by means of a linker). Most preferably, the Fc-binding peptide will contain a single solvent-exposed lysine, thus enabling site-specific conjugation of the compounds of the present application. The Fc-binding peptide can include only naturally occurring amino acid residues or can include one or more non-naturally occurring amino acid residues. Non-naturally occurring amino acid residues (e.g., side chains of non-naturally occurring amino acid residues) can be used as attachment points for the compounds of the present application (e.g., a dimer or trimer of a neuraminidase inhibitor of formula (IV) or formula (V), e.g., by means of a linker). The Fc-binding peptides of the present application can be linear or cyclic. The Fc-binding peptides of the present application include any Fc-binding peptides known to those skilled in the art.

[0568] As used herein, the term "albumin" refers to a polypeptide comprising amino acids corresponding to a naturally occurring albumin (e.g., human serum albumin) or a variant thereof, such as an engineered variant of a naturally occurring albumin. Variants of albumin include polymorphisms, fragments such as domains and subdomains, and fusion proteins (e.g., albumin having a C-terminal or N-terminal fusion with, for example, a polypeptide linker). Preferably, the albumin has the amino acid sequence of human serum albumin (HSA) or a variant or fragment thereof, and most preferably has the amino acid sequence of a functional variant or fragment thereof. The albumin of the present application includes a protein having at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to any of the sequences of SEQ ID NO: 69-71. The albumin of the present application includes albumin that has been engineered to include one or more (e.g., two, three, four or five) solvent-exposed lysine residues, which can provide a site for conjugation to a compound of the present application (e.g., conjugation to a neuraminidase inhibitor dimer or trimer represented by formula (IV) or formula (V), including via a linker). Most preferably, the albumin will contain a single solvent-exposed lysine, thus enabling site-specific conjugation of the compound of the present application (the neuraminidase inhibitor dimer or trimer represented by formula (IV) or formula (V)). The albumin may include only naturally occurring amino acid residues or may include one or more non-naturally occurring amino acid residues. The non-naturally occurring amino acid residues (e.g., the side chains of non-naturally occurring amino acid residues) can be used as attachment points for the compounds of the present application (e.g., the neuraminidase inhibitor dimer or trimer represented by formula (IV) or formula (V), including via a linker).

[0569] As used herein, the term "albumin-binding peptide" refers to a polypeptide having an amino acid sequence of 5 to 50 (e.g., 5 to 40, 5 to 30, 5 to 20, 5 to 15, 5 to 10, 10 to 50, 10 to 30, or 10 to 20) amino acid residues, which has an affinity for albumin (e.g., any albumin described in the present application) and has the function of binding to said albumin. Preferably, the albumin-binding peptide is conjugated to a naturally occurring serum albumin, most preferably human serum albumin. The albumin-binding peptide can have different origins, such as synthetic, human, murine, or rat. The albumin-binding peptides of the present application include albumin-binding peptides that have been engineered to include one or more (e.g., two, three, four, or five) solvent-exposed lysine residues, which can provide sites for conjugating to the compounds of the present application (e.g., conjugating to the neuraminidase inhibitor dimer or trimer represented by formula (IV) or formula (V), including by means of a linker). Most preferably, the albumin-binding peptide will contain a single solvent-exposed lysine, thus enabling site-specific conjugation of the compounds of the present application (the neuraminidase inhibitor dimer or trimer represented by formula (IV) or formula (V)). The albumin-binding peptide can include only naturally occurring amino acid residues or can include one or more non-naturally occurring amino acid residues. The non-naturally occurring amino acid residues (e.g., the side chains of non-naturally occurring amino acid residues) can be used as attachment points for the compounds of the present application (the neuraminidase inhibitor dimer or trimer represented by formula (IV) or formula (V)). The albumin-binding peptides of the present application can be linear or cyclic. The albumin-binding peptides of the present application include any albumin-binding peptides known to those skilled in the art.

[0570] As used herein, "surface-exposed amino acid" or "solvent-exposed amino acid", such as surface-exposed lysine, refers to an amino acid that is accessible to the solvent surrounding the protein. The surface-exposed amino acid can be a naturally occurring one or an engineered variant of the protein (e.g., a substitution or insertion). In some embodiments, the surface-exposed amino acid is an amino acid that, when substituted, does not substantially alter the three-dimensional structure of the protein.

[0571] As used herein, the terms "linker", "L'" refer to a covalent connection between two or more components in a compound (e.g., between two neuraminidase inhibitors in a conjugate described herein, between a neuraminidase inhibitor and an Fc domain monomer or Fc domain or albumin in a conjugate described herein, and between a multimer of two neuraminidase inhibitors and an Fc domain monomer or Fc domain or albumin in a compound described herein).

[0572] In some embodiments, the compounds described herein may contain a linker having a trivalent structure (e.g., a trivalent linker). The trivalent linker has four arms, each of which is covalently linked to a component of the compound (e.g., a first arm that binds to a first neuraminidase inhibitor (e.g., A1 in the conjugate shown in formula (I)), a second arm that binds to a second neuraminidase inhibitor (e.g., A2 in the conjugate shown in formula (I)), a third arm that binds to a third neuraminidase inhibitor (e.g., A3 in the conjugate shown in formula (I)), and a fourth arm that binds to an Fc protein or albumin).

[0573] As used herein, the term "amino acid" means naturally occurring amino acids and non-naturally occurring amino acids.

[0574] As used herein, the term "naturally occurring amino acid" means amino acids including Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, and Val.

[0575] As used herein, the term "T" refers to the average number of neuraminidase inhibitor dimers or neuraminidase inhibitor trimers conjugated to the Fc domain monomer, Fc domain, or albumin of the conjugate, which can be from 1 to 20 (e.g., T is from 1 to 2, 1 to 3, 1 to 4, 1 to 5, 5 to 10, 10 to 15, or 15 to 20).

[0576] As used herein, the term "pharmaceutical composition" refers to a pharmaceutical or medicinal formulation containing at least one active ingredient (e.g., any of the conjugates shown in formula (I) and formula (II), and the compounds shown in formula (IV) and formula (V)) and one or more carriers and / or excipients to render the active ingredient suitable for a method of administration.

[0577] As used herein, the term "about" can be understood to be within + / - 10%, + / - 9%, + / - 8%, + / - 7%, + / - 6%, + / - 5%, + / - 4%, + / - 3%, + / - 2%, + / - 1%, + / - 0.5%, + / - 0.4%, + / - 0.3%, + / - 0.2%, or + / - 0.1% of the value. For example, about 10% means 9% to 11%. Unless otherwise apparent from the context, all numerical values provided herein are modified by the term "about".

[0578] As used herein, the term "identity" is used to refer to the matching of sequences between two polypeptides or between two nucleic acids. When a position in each of two sequences being compared is occupied by the same base or amino acid monomer subunit (e.g., a position in each of two DNA molecules is occupied by adenine, or a position in each of two polypeptides is occupied by lysine), then the molecules are identical at that position. The "percent identity" between two sequences is a function of the number of matching positions shared by the two sequences divided by the number of positions compared × 100. For example, if 6 of 10 positions in two sequences match, then the two sequences have 60% identity. For example, the DNA sequences CTGACT and CAGGTT have 50% identity (3 of a total of 6 positions match). In general, comparison is made when the two sequences are aligned to yield maximum identity. Such alignment can be accomplished by using, for example, the method of Needleman et al. (1970) J. Mol. Biol. 48:443-453, which can be conveniently performed by computer programs such as the Align program (DNAstar, Inc.). The algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4:11-17 (1988)), incorporated into the ALIGN program (version 2.0), can also be used, with a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4 to determine the percent identity between two amino acid sequences. In addition, the algorithm of Needleman and Wunsch (J. Mol. Biol. 48:444-453 (1970)) incorporated into the GAP program in the GCG software package (available at www.gcg.com) can be used, with a Blossum 62 matrix or a PAM250 matrix and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6 to determine the percent identity between two amino acid sequences.

[0579] As used herein, the term "conservative substitution" means an amino acid substitution that does not adversely affect or alter the essential properties of a protein / polypeptide containing the amino acid sequence. For example, conservative substitutions can be introduced by standard techniques known in the art such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include substitutions of an amino acid residue with another amino acid residue having a similar side chain, such as a substitution with a residue that is physically or functionally similar to the corresponding amino acid residue (e.g., having a similar size, shape, charge, chemical properties, including the ability to form covalent or hydrogen bonds, etc.). Families of amino acid residues having similar side chains have been defined in the art. These families include amino acids having basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), β-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, a conservative substitution generally refers to the replacement of a corresponding amino acid residue with another amino acid residue from the same side chain family. Methods for identifying conservative amino acid substitutions are well known in the art (see, e.g., Brummell et al., Biochem. 32:1180-1187 (1993); Kobayashi et al. Protein Eng. 12(10):879-884 (1999); and Burks et al. Proc. Natl Acad. Set USA 94:412-417 (1997), which are incorporated herein by reference).

[0580] As used herein, the term "subject" can be a human, non-human primate, or other mammal, including but not limited to dogs, cats, horses, cows, pigs, turkeys, goats, fish, monkeys, chickens, rats, mice, and sheep.

[0581] As used herein, the term "effective amount" means an amount sufficient to achieve the desired therapeutic effect, e.g., an amount that achieves alleviation of symptoms associated with the disease to be treated. It should be further noted that the dosage and method of use of the conjugate or compound of the present application depend on many factors, including the age, weight, sex, natural health status, nutritional status, activity intensity of the compound, administration time, metabolic rate, severity of the disease, and subjective judgment of the treating physician. The preferred dosage is between 0.001 - 1000 mg / kg body weight / day.

[0582] As used herein, the term "pharmaceutically acceptable carrier and / or excipient" refers to a carrier and / or excipient that is pharmacologically and / or physiologically compatible with the subject and the active ingredient, which are well known in the art (see, e.g., Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995). Pharmaceutically acceptable carriers and / or excipients include, but are not limited to: pH regulators, surfactants, ionic strength enhancers, diluents, reagents for maintaining osmotic pressure, reagents for delaying absorption, preservatives, stabilizers. For example, pH regulators include, but are not limited to, phosphate buffer. Surfactants include, but are not limited to, cationic, anionic or nonionic surfactants, such as Tween-80. Ionic strength enhancers include, but are not limited to, sodium chloride. Preservatives include, but are not limited to, various antibacterial and antifungal reagents, such as parabens, chlorobutanol, phenol, sorbic acid, etc. Reagents for maintaining osmotic pressure include, but are not limited to, sugars, NaCl and its analogs. Reagents for delaying absorption include, but are not limited to, monostearate and gelatin. Diluents include, but are not limited to, water, aqueous buffers (such as buffered saline), alcohols and polyols (such as glycerol), etc. Preservatives include, but are not limited to, various antibacterial and antifungal reagents, such as thimerosal, 2-phenoxyethanol, parabens, chlorobutanol, phenol, sorbic acid, etc. Stabilizers have the meaning commonly understood by those skilled in the art, which can stabilize the desired activity of the active ingredient in the drug, including, but not limited to, sodium glutamate, gelatin, SPGA, sugars (such as sorbitol, mannitol, starch, sucrose, lactose, dextran, or glucose), amino acids (such as glutamic acid, glycine), proteins (such as dried whey, albumin or casein) or their degradation products (such as lactalbumin hydrolyzate), etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0583] Figure 1 and Figure 2 shows that the conjugate of the present application exhibits good NA enzyme activity inhibition ability;

[0584] Figure 3 and Figure 4 shows that the conjugate of the present application exhibits good NA enzyme activity inhibition ability against drug-resistant strain NA;

[0585] Figure 5 shows that the conjugate of the present application has a good plasma half-life and can remain above the effective drug concentration for a long time;

[0586] Figure 6 shows that the conjugate of the present application can maintain the same ADCC effect as Fc;

[0587] Figure 7 It shows that the conjugate of the present application can effectively inhibit virus replication after single administration at different virus challenge doses;

[0588] Figure 8 It shows that the conjugate of the present application can effectively alleviate the weight loss in the severe influenza DBA / 2 mouse model;

[0589] Figures 9 to 10 It shows that the negative control group and the small molecule drugs oseltamivir and zanamivir cannot protect the severe influenza DBA / 2 mouse model;

[0590] Figure 11 It shows the survival curve results of the conjugate of the present application on the severe influenza DBA / 2 mouse model, indicating that the conjugate of the present application has good protective effects on DBA / 2 mice suffering from severe influenza;

[0591] Figure 12 and Figure 13 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate I-1 prepared in an embodiment of the present application;

[0592] Figure 14 and Figure 15 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate I-5 prepared in an embodiment of the present application;

[0593] Figure 16 and Figure 17 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate I-6 prepared in an embodiment of the present application;

[0594] Figure 18 and Figure 19 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate II-1 prepared in an embodiment of the present application;

[0595] Figure 20 and Figure 21 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate II-6 prepared in an embodiment of the present application;

[0596] Figure 22 and Figure 23 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate I-9 prepared in an embodiment of the present application;

[0597] Figure 24 and Figure 25 respectively show the HPLC chromatogram and mass spectrum (MS) of conjugate I-6 prepared in another embodiment of the present application;

[0598] Figures 26 - 29The inhibitory curves of the conjugate of the present application against A / Puerto Rico / 8 / 1934 (H1N1-PR8), A / Victoria / 2570 / 2019 (H1N1), A / Darwin / 9 / 2021 (H3N2), and B / Austria / 1359417 / 2021 (BV) viruses are shown respectively;

[0599] Figure 30 It is shown that therapeutic administration of the conjugate of the present application can effectively alleviate the weight loss of the BALB / c mouse model infected with H1N1-PR8;

[0600] Figure 31 It is shown that therapeutic administration of the conjugate of the present application plays a protective role in the BALB / c mouse model infected with H1N1-PR8 and improves the survival rate of the BALB / c mouse model;

[0601] Figure 32 It is shown that therapeutic administration of the conjugate of the present application can effectively inhibit the replication of H1N1-PR8 virus in the BALB / c mouse model, and **** indicates a significance level of P<0.0001;

[0602] Figure 33 It is shown that prophylactic administration of the conjugate of the present application can effectively alleviate the weight loss of the BALB / c mouse model infected with H1N1-PR8;

[0603] Figure 34 It is shown that prophylactic administration of the conjugate of the present application plays a protective role in the BALB / c mouse model infected with H1N1-PR8 and improves the survival rate of the BALB / c mouse model;

[0604] Figure 35 It is shown that prophylactic administration of the conjugate of the present application can effectively inhibit the replication of H1N1-PR8 virus in the BALB / c mouse model, and **** indicates a significance level of P<0.0001. Detailed implementation mode

[0605] Example 1 Synthesis of divalent backbone

[0606]

[0607] 29) Preparation of compound 2:

[0608] Dissolve 2-amino-1,3-dihydroxypropane (5 g, 54.9 mmol) in 150 ml of anhydrous ethanol. Add di-tert-butyl dicarbonate Boc2O (dissolved in 100 ml of anhydrous ethanol, 12 g, 54.98 mmol), stir at room temperature, after reacting for 6 h, dry the reaction solution with a rotary evaporator, and recrystallize the product with n-heptane to obtain a white solid. Filter to obtain the solid and dry it under vacuum (7 mg, 90% yield). [M+H] + = 192.2

[0609] 2) Preparation of compound 3:

[0610] Under ice bath conditions, disperse KOH (7.87 g, 140.18 mmol) in 20 ml of ultra-dry DMF to obtain a DMF solution containing KOH. Dissolve compound 2 (5 g, 26.2 mmol) in 20 ml of THF / 8 ml of DMF, and then slowly add it to the DMF solution containing KOH. Then add 3-bromopropyne (12.46 g, 104.7 mmol) dropwise to the reaction solution, the solution turns brown, and react at 0 °C for 7 h. Extract with 100 ml of H2O and 140 ml of DCM respectively, repeat twice, combine the DCM extracts, wash with saturated brine, add anhydrous sodium sulfate and let stand for 8 h. After purification by column chromatography and vacuum drying, a brownish-yellow solid of 4 g, 50.2% yield, [M+H] + = 268.31

[0611] 3) Preparation of compound 4:

[0612] Dissolve compound 3 (2 g, 7.49 mmol) in 15 ml of HCl·EA, stir at room temperature, after 3 h, dry the reaction solution with a rotary evaporator to obtain a brown oily liquid. Since there are no other impurities in the reaction solution, the next step reaction is carried out directly.

[0613] 4) Preparation of compound 5:

[0614] Dissolve the brown oily liquid (compound 4) obtained in the previous step in 2 ml of MeOH, add 15 ml of NaHCO3, after reacting for 2 h, detect that the pH of the solution = 7. Extract with 50 ml of DCM and 30 ml of H2O respectively, repeat twice, combine the DCM extracts, wash with saturated NaCl, add anhydrous sodium sulfate and dry overnight. After purification by column chromatography and vacuum drying, a yellow oily substance of 1.26 g, 85% yield is obtained. 1 1H NMR (600 MHz, DMSO-d6) δ 4.13 (d, 4H), 3.42 (t, 2H), 3.38 (dd, 2H), 3.29 (dd, 2H), 2.97 (tt, 1H), 1.55 (s, 2H). [M+H] + = 168.01

[0615] Example 2 Synthesis of Trivalent Framework

[0616]

[0617] 29) Preparation of Compound 7:

[0618] Dissolve tris(hydroxymethyl)aminomethane (5 g, 41.27 mmol) in a mixture of 30 ml of methanol and 30 ml of tert-butanol to obtain a suspension solution. Dropwise add di-tert-butyl dicarbonate Boc2O (11.75 g, 53.84 mmol) dissolved in tert-butanol solution, stir at room temperature, and a white precipitate gradually forms in the reaction solution. After reacting for 8 h, dry the reaction solution with a rotary evaporator to obtain a white solid. Add ethyl acetate to the solid, cool to 0 °C, react for 2 h, filter to obtain a solid, and dry in vacuo (8.20 mg, 90% yield). [M+H] + = 222.80

[0619] 2) Preparation of Compound 8:

[0620] Under ice bath conditions, disperse KOH (7.87 g, 140.18 mmol) in 20 ml of ultra-dry DMF to obtain a DMF solution containing KOH. Dissolve BOC-amino tris(hydroxymethyl)methane (Compound 7) (5 g, 22.61 mmol) in 20 ml of THF / 8 ml of DMF, and then slowly add it to the DMF solution containing KOH. Then, dropwise add 3-bromopropyne (16.68 g, 140.18) to the reaction solution, the solution turns brown, and react at 0 °C for 6 h. Extract with 100 ml of H2O and 140 ml of DCM respectively, repeat twice, combine the DCM extracts, wash with saturated brine, add anhydrous sodium sulfate and let stand for 8 h. After purification by column chromatography and drying in vacuo, obtain 3.6 g of a brownish-yellow solid, 47.5% yield, [M+H] + = 336.29

[0621] 3) Preparation of Compound 9:

[0622] Dissolve Compound 8 (1 g, 2.98 mmol) in 15 ml of HCl·EA, stir at room temperature, and after 3 h, dry the reaction solution with a rotary evaporator to obtain a brown oily liquid. Since there are no other impurities in the reaction solution, directly proceed to the next step of the reaction.

[0623] 4) Preparation of Compound 10:

[0624] The brown oily liquid (Compound 9) obtained by drying the previous reaction suspension was dissolved in 2 ml of MeOH, 15 ml of NaHCO3 was added, and after reacting for 2 h, the pH of the solution was detected to be 7. It was extracted with 50 ml of DCM and 30 ml of H2O respectively, repeated twice, the DCM extracts were combined, washed with saturated NaCl, and dried over anhydrous sodium sulfate overnight. After purification by column chromatography and vacuum drying, 0.58 g of yellow oily substance was obtained, with a yield of 83%, 1 1H NMR (600 MHz, DMSO-d6) δ 4.14 (d, 6H), 3.42 (t, 3H), 3.32 (s, 6H), 1.47 (s, 2H). [M+H] + = 236.12

[0625] Example 3 Synthesis of Zanamivir Dimer Derivative

[0626]

[0627]

[0628] 29) Preparation of Compound R2:

[0629] Methyl 5-acetamido-7,8,9-O-triacetyl-2,6-anhydro-4-azido-3,4,5-trideoxy-D-glycero-D-galacto-non-2-enoate (Compound R1, 2.28 g, 5.0 mmol) was dissolved in anhydrous THF (15 mL), and the solution was cooled to about 13 °C. After 20 minutes, triphenylphosphine (1.58 g, 6 mmol) was added in portions. The resulting mixture was stirred at about 13 °C to room temperature for 2 hours, and then a solution of LiOH (12 mg, 0.5 mmol) in water (1 mL) was added dropwise. After stirring for 28 hours, N,N'-bis-boc-1-amidinopyrazole (1.63 g, 5.25 mmol) and 4-dimethylaminopyridine (122.2 mg, 1 mmol) were added to the reaction mixture. The reaction was stirred for 1.5 days. It was then diluted with a 1:1 mixture of ethyl acetate:hexane (100 mL) and extracted with water (30 mL). The aqueous layer was back-extracted with ethyl acetate (30 mL). The combined organic layers were concentrated by rotary evaporation. The residue was purified by C18 reverse-phase column chromatography (150 g, 25 to 70% acetonitrile and water). The collected eluates were concentrated by rotary evaporation at room temperature. A turbid aqueous solution was produced and most of the product deposited as a gel on the flask. The solution was then extracted with ethyl acetate (150 mL). The organic layer was used to redissolve the gel material. It was then dried over Na2SO4, concentrated by rotary evaporation, and further dried under high vacuum to give the title compound as a white foam. Yield 3 g, 90.2% yield. Ion found by LCMS: [M+H] + = 673.2.

[0630] 2) Preparation of Compound R3:

[0631] Compound R2 (3 g, 4.46 mmol) was cooled in an ice-water bath in anhydrous MeOH (15 mL), 0.5 M methanolic solution of sodium methoxide (13 mL, 6.5 mmol) was added slowly, stirred for 1 hour, and then the pH was carefully adjusted to 7 to 7.5 by dropwise addition of a 1,4-dioxane solution of hydrochloric acid (4 M, 2 mL, Innochem, catalog number A18256). The solvent was removed by rotary evaporation at a temperature not exceeding room temperature. The residue was diluted with a 2:1 mixture of ethyl acetate and hexane (150 mL), and the resulting solution was extracted with water (20 mL). The aqueous layer was back-extracted with ethyl acetate (30 mL). The combined organic layers were dried over anhydrous Na2SO4, concentrated by rotary evaporation, and dried in vacuo. The product was used in the subsequent step without further purification. Yield 2.7 g, 98.4% yield. Ion found by LCMS: [M+H] + = 547.26.

[0632] 3) Preparation of Compound R4:

[0633] Dissolve Compound R3 (1 g, 1.82 mmol) in anhydrous DCM (6 mL) and cool in an ice - water bath. Then add 4 - dimethylaminopyridine (221.26 mg, 1.82 mmol) and DIPEA (728 mg, 5.6 mmol). Next, add phenyl 4 - nitrophenyl carbonate (537.75 mg, 3.05 mmol) dropwise to the reaction solution. Then remove the ice - water bath, stir the reaction solution for 3 h and monitor by LCMS (add phenyl 4 - nitrophenyl carbonate again if necessary). After the reaction is completed, add water (10 mL) to end the reaction, separate the organic layer and concentrate it with a rotary evaporator, and then purify it by C18 reverse - phase chromatography (100 g, 20% to 70% acetonitrile and water). The collected eluate is evaporated to remove acetonitrile at room temperature with a rotary evaporator, and then extracted with a 1:1 mixture of ethyl acetate and hexane (120 mL). The aqueous layer is repeatedly extracted with ethyl acetate (30 mL). Combine the organic phases, add anhydrous Na2SO4 and dry for 8 h. After concentration with a rotary evaporator, dry under vacuum to obtain a white solid product. Yield 712 mg, 68.34% yield. 1 H NMR (600 MHz, DMSO - d6) δ11.39 (s, 1H), 8.26 (d, 2H), 8.15 (d, 1H), 5.80 (d, 1H), 5.71 (d, 1H), 4.98 (m, 1H), 4.88 (m, 1H), 4.59 (m, 1H), 4.55 (t, 1H), 4.17 (dd, 1H), 4.10 (m, 1H), 4.01 (m, 1H), 3.73 (s, 3H), 1.85 (s, 3H), 1.47 (s, 9H), 1.40 (s, 9H). [M + H] + = 573.24.

[0634] 4) Preparation of Compound R5:

[0635] Dissolve Compound R4 (700 mg, 1.22 mmol) in 5 ml of ultradry DCM, cool in an ice - water bath, then add phenyl 4 - nitrophenyl carbonate (294.6 mg, 1.46 mmol), then add dimethylaminopyridine (123.2 mg, 0.61 mmol), protect with nitrogen, remove the ice - water bath and continue stirring for 3 h. Add phenyl 4 - nitrophenyl carbonate (201.8 mg, 1 mmol) and continue the reaction for 1 h. After the reaction is completed, remove the solvent with a rotary evaporator and purify by column chromatography to obtain 540 mg of the product, 60% yield.

[0636] 5) Preparation of Compound R6:

[0637] Compound R5 (240 mg, 0.33 mmol) was dissolved in 5 ml of anhydrous DCM. Azido triethylene glycol amine (N3-PEG3-NH2, 2-(2-(2-(2-azidoethoxy)ethoxy)ethoxy)ethylamine, 71 mg, 0.33 mmol), 1-hydroxybenzotriazole (HOBT, 43.96 mg, 0.33 mmol), and N,N-diisopropylethylamine (DIPEA, 84.11 mg, 0.65 mmol) were added successively. Under nitrogen protection, the mixture was stirred at room temperature for 8 h, and then purified by column chromatography to obtain 140 mg of the product, with a 51% yield. 1 H NMR (600 MHz, DMSO-d6) δ 7.64 (m, 1H), 7.30 (m, 1H), 4.34 (t, 1H), 3.72 (d, 1H), 3.58 (m, 3H), 3.32 (m, 1H), 2.27 (t, 1H), 1.65 (m, 1H), 1.56 (m, 1H), 1.41 (t, 3H), 1.35 (m, 2H), 1.18 (s, 24H), 0.89 (t, 2H), 0.81 (t, 8H), 0.77 (t, 2H), [M+H] + = 817.34

[0638] Azido hexaethylene glycol amine (N3-PEG6-NH2) and azido nonaethylene glycol amine (N3-PEG9-NH2) were used to replace azido triethylene glycol amine (N3-PEG3-NH2) respectively, and compounds R6-1 and R6-2 were prepared by the same method.

[0639]

[0640] 6) Preparation of compound R7:

[0641] Compound R6 (140 mg, 0.168 mmol) and compound 5 (11.29 mg, 0.0672 mmol) were respectively dissolved in 1.5 ml of tetrahydrofuran (THF), and the mixture was stirred. An aqueous solution of CuSO4·5H2O (6.294 mg, 0.025 mmol) and sodium ascorbate (Na·VC, 9.98 mg, 0.05 mmol) was added successively to the reaction solution. After reacting at room temperature for 3 h, it was extracted with DCM, and finally purified by column chromatography to obtain 100 mg of the product, with a 33% yield. [M+H] + = 1780.3

[0642] Compound R6-1 and R6-2 were used to replace compound R6 respectively, and compounds R7-1 and R7-2 were prepared by the same method.

[0643]

[0644] 7) Preparation of Compound R8:

[0645] The raw material propargyl-PEG2-COOH (2-(2-(2-(propargyl-oxy)ethoxy)ethoxy)acetic acid, 14.68 mg, 0.168 mmol) was successively added to 3 ml of ultra-dry DMF along with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI, 28.31 mg, 0.145 mmol), HOBT (19.6 mg, 0.145 mmol), DMAP (2.96 mg, 0.024 mmol), and DIPEA (31.27 mg, 0.242 mmol). Under argon protection, the mixture was stirred at room temperature for 30 minutes. Then, Compound R7 (80 mg, 0.048 mmol) was added, and stirring was continued at room temperature under argon protection. After 12 h of reaction, the product was purified by column chromatography to obtain 60 mg with a 63% yield.

[0646] [M+H] + = 1984.6

[0647] Compound R7 was replaced with Compound R7-1, and Compound R8-1 was prepared using the same method.

[0648]

[0649] Propargyl-PEG3-COOH was used to replace propargyl-PEG2-COOH, and Compound R7 was replaced with Compound R7-2. Using the same method, Compound R8-2 was prepared.

[0650]

[0651] Propargyl-PEG6-COOH was used to replace propargyl-PEG2-COOH, and Compound R7 was replaced with Compound R7-2. Using the same method, Compound R8-3 was prepared.

[0652]

[0653] 8) Preparation of Compound R9:

[0654] Compound R8 (60 mg, 0.03 mmol) was dissolved in 2 ml of ultra-dry DMF, and then 2 ml of trifluoroacetic acid (TFA) was added. The mixture was stirred at room temperature for 4 h. TFA was removed using a rotary evaporator, and the product was purified by C18 reverse-phase chromatography to obtain 46 mg with a 97% yield. [M+H] + = 1584.5

[0655] The same method was used to prepare compounds R9-1 to R9-3.

[0656]

[0657]

[0658] 9) Preparation of Compound R10

[0659] Compound R9 (46 mg, 0.029 mmol) was dissolved in 1.5 ml of methanol, and 200 μl of double-distilled water was added. The pH was adjusted to 12-13 with 10 mol / L NaOH, and the mixture was stirred at room temperature for 4 h. The pH of the reaction system was adjusted to 4 with 4 M hydrochloric acid in 1,4-dioxane. Finally, the product was purified by C18 reverse-phase chromatography to obtain 40 mg of the product with a yield of 91.7%. [M+H] + =1503.9

[0660] Compounds R10-1 to R10-3 were prepared using the same method.

[0661]

[0662]

[0663] Example 4. Synthesis of Zanamivir Trimeric Derivatives

[0664]

[0665]

[0666]

[0667] 29) Preparation of compound R11:

[0668] Compound R6 (140 mg, 0.168 mmol) and compound 10 (12.34 mg, 0.0525 mmol) were separately dissolved in 1.5 ml of tetrahydrofuran (THF), mixed and stirred, and aqueous solutions of CuSO4·5H2O (6.294 mg, 0.025 mmol) and sodium ascorbate (Na·VC, 9.98 mg, 0.05 mmol) were added to the reaction solution in sequence. After reacting at room temperature for 3 h, the mixture was extracted with DCM and purified by column chromatography to obtain 106 mg of the product with a yield of 25.5%. 11H NMR (600 MHz, CDCl3) δ 11.38 (s, 2H), 8.43 (d, 3H), 7.88 (s, 2H), 5.97 (d, 2H), 5.68 (s, 2H), 5.62 (t, 2H), 5.17 (m, 3H), 5.12 (s, 2H), 4.66 (m, 12H), 4.54 (m, 8H), 4.18 (q, 3H), 3.89 (t, 6H), 3.78 (s, 8H), 3.68 (s, 3H), 3.57 (m, 32H), 3.31 (m, 2H), 3.23 (m, 3H), 3.31 (m, 2H), 3.23 (m, 3H), 1.91 (s, 7H), 1.47 (s, 54H), 1.29 (d, 4H), 1.26 (d, 6H), 0.87 (m, 3H). [M+2H] + = 1343.51

[0669] Replace compound R6 with compounds R6-1 and R6-2 respectively, and prepare compounds R11-1 and R11-2 by the same method.

[0670]

[0671] 2) Preparation of compound R12:

[0672] Add the raw material propargyl-PEG2-COOH (2-(2-(2-(propargyloxy)ethoxy)ethoxy)acetic acid, 9.83 mg, 0.0486 mmol), EDCI (18.96 mg, 0.0972 mmol), HOBT (13.127 mg, 0.0972 mmol), DMAP (1.979 mg, 0.0162 mmol), DIPEA (20.94 mg, 0.1621 mmol) into 3 ml of ultra-dry DMF in turn, under argon protection, stir at room temperature for 30 minutes, add compound R11 (80 mg, 0.03 mmol), continue to stir at room temperature under argon protection, after reacting for 12 h, purify by column chromatography to obtain 82 mg of the product, 95.3% yield. 11H NMR (600 MHz, CDCl3) δ 11.38 (s, 1H), 9.11 (s, 2H), 8.45 (d, 2H), 7.82 (dt, 2H), 7.79 (s, 1H), 7.67 (dt, 1H), 7.37 (m, 3H), 6.89 (m, 2H), 5.94 (d, 2H), 5.64 (t, 2H), 5.57 (m, 1H), 5.21 (t, 2H), 5.02 (m, 2H), 4.65 (m, 4H), 4.56 (m, 9H), 4.48 (dd, 2H), 4.17 (d, 1H), 4.14 (d, 1H), 3.88 (t, 6H), 3.78 (d, 11H), 3.66 (m, 2H), 3.63 (m, 3H), 3.59 (m, 21H), 3.52 (m, 3H), 3.18 (m, 4H), 3.34 (m, 3H), 3.26 (dd, 2H), 1.90 (s, 6H), 1.46 (t, 54H), 1.43 (d, 3H), 1.37 (s, 1H), 1.33 (s, 1H), 1.28 (d, 4H), 1.27 (s, 2H), 1.25 (s, 13H), 1.88 (t, 4H), 0.84 (m, 2H). [M + 3H] + = 957.42

[0673] Replace compound R11 with compound R11-1 and prepare compound R12-1 using the same method.

[0674]

[0675] Replace compound R11 with compound R11-2 and prepare compound R12-2 using the same method.

[0676]

[0677] Replace propargyl-1-yloxy-PEG2-COOH with propargyl-1-yloxy-PEG6-COOH and replace compound R11 with compound R11-2, and prepare compound R12-3 using the same method.

[0678]

[0679] 3) Preparation of compound R13:

[0680] Dissolve compound R12 (60 mg, 0.028 mmol) in 2 ml of ultradry DMF, add 2 ml of trifluoroacetic acid (TFA), stir at room temperature for 4 h, remove TFA using a rotary evaporator, and purify the product by C18 reverse-phase chromatography to obtain 41 mg of product, 64.5% yield. [M + H]+ = 2268.7

[0681] The compounds R13-1 to R13-3 were prepared by the same method.

[0682]

[0683]

[0684] 4) Preparation of compound R14

[0685] Compound R13 (41 mg, 0.018 mmol) was dissolved in 1.5 ml of methanol, 200 μl of double-distilled water was added, and the pH was adjusted to 12 - 13 with 10 mol / L NaOH. The mixture was stirred at room temperature for 4 h, and the pH of the reaction system was adjusted to 4 with a 1,4-dioxane solution of 4 M hydrochloric acid. Finally, the product (32 mg, 82.8% yield) was purified by C18 reverse-phase chromatography.

[0686] [M + H] + = 2148.9

[0687] The compounds R14-1 to R14-3 were prepared by the same method.

[0688]

[0689] Example 5. Synthesis of peramivir dimeric derivatives

[0690]

[0691]

[0692] 29) Preparation of compound P2:

[0693] Compound P1 (5 g, 16.7 mmol) was added to DCM (100 ml), N,N'-bis(tert-butoxycarbonyl)-1H-pyrazole-1-carboxamidine (7.77 g, 25 mmol) was added, diisopropylethylamine DIEA (4.3 g, 33.3 mmol) and dimethylaminopyridine DMAP (0.1 g) were added. The mixture was stirred at room temperature for 24 h, and monitored by TLC (DCM:MeOH = 10:1). When the reaction of compound P1 was complete, water (100 ml) was added to the reaction solution, and the mixture was stirred for 10 min, then allowed to stand. The aqueous phase was separated, and the organic phase was washed with 100 ml of water again. The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and the filtrate was collected and concentrated to dryness. It was purified by column chromatography (eluent: DCM / MeOH = 50:1), and the target product was collected to obtain 5.8 g of compound P2, with a yield of 66%.

[0694] 2) Preparation of compound P3:

[0695] Compound P2 (100 mg, 0.184 mmol) was dissolved in 3 ml of anhydrous DCM, stirred in an ice bath, phenyl chloroformate p-nitro (55.7 mg, 0.276 mmol) was added successively, and then dimethylaminopyridine DMAP (45.03 mg, 0.368 mmol) was added. Under nitrogen protection, after removing the ice bath, stirring was continued for 6 hours. Phenyl chloroformate p-nitro (27.5 mg, 0.184 mmol) was added additionally, and the reaction was continued for 6 hours. After the reaction was completed, the solvent was removed by a rotary evaporator, and the product was purified by column chromatography to obtain 98 mg, 75.3% yield.

[0696] 3) Preparation of compound P4:

[0697] Compound P3 (60 mg, 0.0848 mmol) was dissolved in 5 ml of anhydrous DCM, and azido triethylene glycol amine (N3-PEG3-NH2, 2-(2-(2-(2-azidoethoxy)ethoxy)ethoxy)ethylamine, 27.783 mg, 0.127 mmol), HOBT (11.47, 0.0848 mmol), and DIPEA (21.94 mg, 0.1697 mmol) were added successively. Under nitrogen protection, stirring was carried out at room temperature. After 8 hours of reaction, the product was purified by column chromatography to obtain 62 mg, 93.0% yield, [M+H] + = 787.3

[0698] Using the same method, azido triethylene glycol amine (N3-PEG3-NH2) was replaced with azido hexaethylene glycol amine (N3-PEG6-NH2) or azido nonaethylene glycol amine (N3-PEG9-NH2) to prepare compound P4-1 or P4-2.

[0699]

[0700] 4) Preparation of compound P5:

[0701] Compound P4 (60 mg, 0.076 mmol) and compound 5 (5.77 mg, 0.0345 mmol) were respectively dissolved in 1.5 ml of tetrahydrofuran (THF), mixed and stirred. An aqueous solution of CuSO4·5H2O (9.57 mg, 0.038 mmol) and sodium ascorbate (Na·VC, 15.12 mg, 0.076 mmol) was added successively to the reaction solution. After reacting at room temperature for 3 hours, extraction was carried out with DCM, and finally the product was purified by column chromatography to obtain 54 mg, 40.8% yield. 11H NMR (600 MHz, CDCl3) δ 11.43 (s, 2H), 8.56 (d, 2H), 8.24 (m, 2H), 7.97 (s, 1H), 6.14 (t, 2H), 5.18 (dd, 2H), 4.68 (s, 3H), 4.56 (t, 4H), 4.39 (m, 2H), 4.24 (m, 2H), 3.89 (t, 4H), 3.80 (s, 2H), 3.72 (s, 6H), 3.61 (m, 17H), 3.54 (t, 5H), 3.33 (m, 5H), 2.96 (m, 2H), 2.50 (m, 2H), 2.36 (q, 2H), 2.05 (d, 6H), 1.83 (m, 2H), 1.48 (d, 36H), 1.41 (d, 3H), 1.28 (s, 2H), 1.25 (s, 4H), 1.11 (m, 3H), 1.05 (m, 2H), 0.85 (m, 14H), [M+H] + = 1741.0540

[0702] Replace compound P4 with compounds P4-1 and P4-2 respectively, and prepare compounds P5-1 and P5-2 by the same method.

[0703]

[0704] 5) Preparation of compound P6:

[0705] Add raw material propargyl-PEG2-COOH (2-(2-(2-(propargyloxy)ethoxy)ethoxy)acetic acid, 4.68 mg, 0.023 mmol), EDCI (8.868 mg, 0.046 mmol), HOBT (6.245 mg, 0.046 mmol), DMAP (0.942 mg, 0.008 mmol), DIPEA (9.96 mg, 0.077 mmol) into 3 ml of ultra-dry DMF in turn under argon protection, stir at room temperature for 30 minutes, add compound P5 (40 mg, 0.023 mmol), continue to stir at room temperature under argon protection, after reacting for 12 h, purify by column chromatography to obtain 43 mg of product, 97.2% yield. [M+H] + = 1925.4

[0706] Replace compound P5 with compound P5-1, and replace propargyl-PEG2-COOH with propargyl-PEG6-COOH, and prepare compound P6-1 by the same method.

[0707]

[0708] Replace compound P5 with compound P5-2 and prepare compound P6-2 using the same method.

[0709]

[0710] 6) Preparation of compound P7:

[0711] Dissolve compound P6 (40 mg, 0.021 mmol) in 2 ml of ultradry DMF, add 2 ml of trifluoroacetic acid (TFA), stir at room temperature for 4 h, remove TFA by rotary evaporation, and purify by C18 reverse-phase chromatography to obtain 29 mg of the product, 92.0% yield. [M+H] + = 1524.6

[0712] Prepare compounds P7-1 to P7-2 using the same method.

[0713]

[0714]

[0715] 7) Preparation of compound P8

[0716] Dissolve compound P7 (29 mg, 0.019 mmol) in 1.5 ml of methanol, add 200 μl of double-distilled water, adjust the pH to 12 - 13 with 10 mol / L NaOH, stir at room temperature for 4 h, adjust the pH of the reaction system to 4 with a 4 M hydrochloric acid solution in 1,4-dioxane, and finally purify by C18 reverse-phase chromatography to obtain 20 mg of the product, 70.4% yield. [M+H] + = 1496.3

[0717] Prepare compounds P8-1 to P8-2 using the same method.

[0718]

[0719] Example 6. Synthesis of peramivir trimer derivatives

[0720]

[0721]

[0722]

[0723] 29) Preparation of compound P9:

[0724] Compound P4 (300 mg, 0.382 mmol) and compound 10 (28.06 mg, 0.119 mmol) were separately dissolved in 1.5 ml of tetrahydrofuran (THF), mixed and stirred. An aqueous solution of CuSO4·5H2O (14.29 mg, 0.057 mmol) and sodium ascorbate (Na·VC, 22.68 mg, 0.115 mmol) was successively added to the reaction solution. After reacting at room temperature for 3 h, it was extracted with DCM, and finally purified by column chromatography to obtain 190 mg of the product, with a yield of 57.5%. 1 H NMR (600 MHz, CDCl3) δ 11.43 (s, 3H), 8.55 (d, 3H), 8.20 (d, 2H), 7.85 (s, 2H), 5.91 (t, 2H), 5.15 (dd, 3H), 4.62 (s, 4H), 4.55 (t, 6H), 4.41 (m, 3H), 4.22 (m, 3H), 3.89 (t, 6H), 3.72 (s, 9H), 3.62 (d, 28H), 3.54 (m, 6H), 3.32 (q, 6H), 2.97 (m, 3H), 2.49 (m, 3H), 2.35 (m, 3H), 2.06 (s, 8H), 1.82 (m, 3H), 1.48 (d, 54H), 1.42 (s, 2H), 1.38 (m, 4H), 1.28 (s, 3H), 1.25 (s, 8H), 1.13 (m, 3H), 1.04 (m, 3H), 0.84 (m, 20H), [M + 2H] + = 1298.2681

[0725] Compound P4 was replaced with compound P4-1 and P4-2 respectively, and compounds P9-1 and P9-2 were prepared by the same method.

[0726]

[0727] 2) Preparation of compound P10:

[0728] The raw material 2-(2-(2-(prop-2-yn-1-yloxy)ethoxy)ethoxy)acetic acid (18.72 mg, 0.093 mmol), EDCI (35.452 mg, 0.185 mmol), HOBT (24.98 mg, 0.185 mmol), DMAP (3.768 mg, 0.032 mmol), and DIPEA (39.8 mg, 0.308 mmol) were successively added to 3 ml of ultra-dry DMF under argon protection, stirred at room temperature for 30 minutes, compound P9 (160 mg, 0.062 mmol) was added, and stirring was continued at room temperature under argon protection. After reacting for 12 h, the product was purified by column chromatography to obtain 126 mg of the product, with a yield of 73.1%.1 1H NMR (600 MHz, CDCl3) δ 11.43 (s, 2H), 8.55 (d, 2H), 8.20 (d, 2H), 7.71 (d, 2H), 7.36 (m, 1H), 7.31 (t, 1H), 5.79 (m, 2H), 5.30 (s, 3H), 5.15 (m, 3H), 4.58 (s, 4H), 4.54 (t, 4H), 4.41 (m, 3H), 4.21 (m, 2H), 4.17 (d, 1H), 3.88 (m, 6H), 3.80 (s, 4H), 3.72 (s, 6H), 3.61 (d, 20H), 3.52 (m, 4H), 3.41 (m, 2H), 3.32 (m, 6H), 2.96 (m, 3H), 2.48 (m, 3H), 2.35 (q, 3H), 2.31 (t, 1H), 2.05 (s, 6H), 1.82 (m, 3H), 1.48 (d, 54H), 1.42 (s, 3H), 1.37 (s, 3H), 1.28 (s, 5H), 1.25 (s, 18H), 1.13 (m, 4H), 1.04 (m, 3H), 0.84 (m, 26H), [M+H] + = 2780.5539

[0729] Replace compound P9 with compound P9-1, and replace propargyl-1-yloxy-PEG2-COOH with propargyl-1-yloxy-PEG6-COOH. Using the same method, prepare compound P10-1.

[0730]

[0731] Replace compound P9 with compound P9-2. Using the same method, prepare compound P10-2.

[0732]

[0733] 3) Preparation of compound P11:

[0734] Dissolve compound P10 (120 mg, 0.043 mmol) in 2 ml of ultradry DMF, then add 2 ml of trifluoroacetic acid TFA. Stir at room temperature for 4 h. Remove TFA using a rotary evaporator and purify by C18 reverse-phase chromatography to obtain 81 mg of the product, 86.5% yield. [M+H] + = 2179.3

[0735] Prepare compounds P11-1 and P11-2 using the same method.

[0736]

[0737]

[0738] 4) Preparation of Compound P12

[0739] Dissolve compound P11 (80 mg, 0.036 mmol) in 1.5 ml of methanol, add 200 μl of double-distilled water, adjust the pH to 12 - 13 with 10 N NaOH, stir at room temperature for 4 h, adjust the reaction pH to 4 with dioxane·4 N HCl, and finally purify by C18 reverse-phase chromatography to obtain 36 mg of the product, 46.8% yield. [M+H] + = 2137.4

[0740] Compounds P12-1 and P12-2 were prepared using the same method.

[0741]

[0742]

[0743] Example 7 Synthesis of Oseltamivir Dimer Derivatives

[0744]

[0745] 29) Preparation of Compound O2

[0746] Add the raw material N3-PEG2-(CH2)2-COOH (1.084 g, 5.34 mmol), EDCI (1.53 g, 8.01 mmol), HOBT (1.81 g, 8.01 mmol), and DIPEA (1.378 g, 10.68 mmol) to 10 ml of ultra-dry DMF in sequence, under argon protection, stir at room temperature for 30 minutes, add compound O1 (2 g, 6.4 mmol), continue to stir at room temperature under argon protection, after reacting for 12 h, purify by column chromatography to obtain 1.76 g of the product, 60.65% yield. [M+H] + = 498.53

[0747] Using the same method, replace N3-PEG2-(CH2)2-COOH with N3-PEG6-(CH2)2-COOH to prepare compound O2-1.

[0748]

[0749] 2) Preparation of Compound O3

[0750] Compound O2 (430 mg, 0.865 mmol) and compound 5 (57.76 mg, 0.345 mmol) were separately dissolved in 2 ml of tetrahydrofuran (THF), mixed and stirred. An aqueous solution of CuSO4·5H2O (86 mg, 0.345 mmol) and sodium ascorbate (Na·VC, 136.7 mg, 0.69 mmol) was successively added to the reaction solution. After reacting at room temperature for 3 h, it was extracted with DCM, and finally purified by column chromatography to obtain 160 mg of the product, 79.69% yield. [M+H] + = 1162.48

[0751] Compound O2 was replaced with compound O2-1, and compound O3-1 was prepared by the same method.

[0752]

[0753] 3) Preparation of compound O4:

[0754] Propargyl-PEG2-COOH (2-(2-(2-(prop-2-ynyloxy)ethoxy)ethoxy)acetic acid, 41.8 mg, 0.2067 mmol), EDCI (80.62 mg, 0.4134 mmol), HOBT (55.81 mg, 0.4134 mmol), DMAP (8.42 mg, 0.0689 mmol), and DIPEA (89.05 mg, 0.6891 mmol) were successively added to 3 ml of ultra-dry DMF under argon protection and stirred at room temperature for 30 minutes. Compound O3 (160 mg, 0.1378 mmol) was added, and stirring was continued at room temperature under argon protection. After reacting for 12 h, it was purified by column chromatography to obtain 170 mg of the product, 91.72% yield. [M+H] + = 1346.22

[0755] Compound O3 was replaced with compound O3-1, and compound O4-1 was prepared by the same method.

[0756]

[0757] 4) Preparation of compound O5:

[0758] Compound O4 (170 mg, 0.1264 mmol) was completely dissolved in 1 ml of methanol solution, and 5% aqueous NaOH solution was slowly added dropwise. A precipitate formed, and then the solution gradually became clear. The reaction was carried out for 1.5 h. Under ice bath conditions, the reaction pH was adjusted to 4 with 1 N dilute hydrochloric acid, and the reaction solution was extracted with n-butanol. After purification by column chromatography, 95 mg of the product was obtained, 62.58% yield, [M+H] += 1202.03

[0759] Replace compound O4 with compound O4-1 and prepare compound O5-1 using the same method.

[0760]

[0761] Example 8. Synthesis of oseltamivir trimer derivative

[0762]

[0763]

[0764] 29) Preparation of compound O6

[0765] Dissolve compound O2 (1.76 g, 3.541 mmol) and compound 10 (250 mg, 1.063 mmol) separately in 1.5 ml of tetrahydrofuran (THF), mix and stir. Add aqueous solutions of CuSO4·5H2O (132.66 mg, 0.53 mmol) and sodium ascorbate (Na·VC, 210.60 mg, 1.06 mmol) to the reaction solution in sequence. After reacting at room temperature for 3 h, extract with DCM and finally purify by column chromatography to obtain 1.28 g of the product, 62.8% yield. [M+H] + = 1727.34

[0766] Replace compound O2 with compound O2-1 and prepare compound O6-1 using the same method.

[0767]

[0768] 2) Preparation of compound O7:

[0769] Add the raw material propargyl-PEG2-COOH (2-(2-(2-(propargyloxy)ethoxy)ethoxy)acetic acid, 57.99 mg, 0.2868 mmol), EDCI (111.85 mg, 0.5736 mmol), HOBT (77.436 mg, 0.5736 mmol), DMAP (11.68 mg, 0.0956 mmol), and DIPEA (123.55 mg, 0.956 mmol) to 3 ml of ultra-dry DMF in sequence under argon protection. Stir at room temperature for 30 minutes, add compound O6 (330 mg, 0.1912 mmol), continue stirring at room temperature under argon protection. After reacting for 12 h, purify by column chromatography to obtain 305 mg of the product, 89.72% yield.

[0770] [M+H] += 1779.4

[0771] Replace compound O6 with compound O6-1 and prepare compound O7-1 using the same method.

[0772]

[0773] 3) Preparation of compound O8:

[0774] Completely dissolve compound O7 (150 mg, 0.078 mmol) in 1 ml of methanol solution, slowly add dropwise 5% aqueous NaOH solution, precipitate forms, and then the solution gradually becomes clear. React for 1.5 h, adjust the reaction pH to 4 with 1 N dilute hydrochloric acid under ice bath conditions, and then extract the reaction solution with n-butanol. The product is purified by column chromatography to obtain 95 mg, 71.89% yield, [M+H] + = 1695.79

[0775] Replace compound O7 with compound O7-1 and prepare compound O8-1 using the same method.

[0776]

[0777] Example 9 Synthesis of Conjugate I

[0778] Compound R14 is conjugated with Fc protein (sequence SEQ ID NO: 64) to synthesize conjugate I-1

[0779]

[0780] Replace Fc protein (Fc domain, whose sequence is shown in SEQ ID NO: 64) into PBS solution to prepare a DMSO stock solution of 5 mg / ml N3-PEG4-TFP (customized by Hunan Huateng Pharmaceutical Co., Ltd.). Take 5 mg of Fc protein, add N3-PEG4-TFP (0.278 mg, 0.633 μmol), and react overnight at room temperature. Fc protein is conjugated to the site # of N3-PEG4-TFP through its N atom. The product is purified by PD-10 desalting chromatography column and then ultrafiltered and concentrated to obtain 4.6 mg of Fc-PEG4-N3. Then add compound R14 (R14:Fc = 20 eq:1 eq), and successively add CuSO4·5H2O (0.95 mg, 3.81 μmol), tris(3-hydroxypropyltriazylmethyl)amine (THPTA, 1.66 mg, 3.81 μmol), sodium ascorbate (Na·VC, 2.7 mg, 13.6 μmol), and react at room temperature for 4 h. After purification by PD-10 desalting chromatography column and ultrafiltration and concentration, conjugate I-1 is obtained. Its HPLC chromatogram is as Figure 12 shown, with a purity of 95.57%; the mass spectrum (MS) is asFigure 13 As shown. In conjugate I-1, E represents a monomer of the Fc domain (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0781] In the same way, compound R14-1 and Fc-PEG4-N3 were subjected to a click reaction to obtain conjugate I-2, wherein the monomer of the Fc domain (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0782]

[0783] In the same way, compound R14-2 and Fc-PEG5-N3 were subjected to a click reaction to obtain conjugate I-3, wherein the monomer of the Fc domain (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0784]

[0785] In the same way, compound R14-3 and Fc-PEG6-N3 were subjected to a click reaction to obtain conjugate I-4, wherein the monomer of the Fc domain (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0786]

[0787] In the same way, compound P11 and Fc-PEG4-N3 were subjected to a click reaction to obtain conjugate I-5, the HPLC chromatogram of which is as Figure 14 shown, with a purity of: 95.49%; the mass spectrometry (MS) is as Figure 15 shown. In conjugate I-5, the monomer of the Fc domain (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0788]

[0789] In the same way, compound P12 and Fc-PEG4-N3 were subjected to a click reaction to obtain conjugate I-6, the HPLC chromatogram of which is as Figure 16 shown, with a purity of: 97.41%; the mass spectrometry (MS) is as Figure 17 shown. In conjugate I-6, the monomer of the Fc domain (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.5.

[0790]

[0791] In the same way, the compound P12-1 and Fc-PEG6-N3 are subjected to a click reaction to obtain the conjugate I-7, wherein the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is about 3.5.

[0792]

[0793] In the same way, the compound P12-2 and Fc-PEG4-N3 are subjected to a click reaction to obtain the conjugate I-8, wherein the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is about 3.5.

[0794]

[0795] In the same way, the compound O8 and Fc-PEG4-N3 are subjected to a click reaction to obtain the conjugate I-9, and its HPLC chromatogram is as Figure 22 shown, with a purity of: 93.22%; the mass spectrum (MS) is as Figure 23 shown. In the conjugate I-9, E is the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is about 3.2.

[0796]

[0797] In the same way, the compound O8-1 and Fc-PEG4-N3 are subjected to a click reaction to obtain the conjugate I-10, wherein E is the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is about 3.2.

[0798]

[0799] In the same way, the compound R10 and Fc-PEG4-N3 are subjected to a click reaction to obtain the conjugate II-1, and its HPLC chromatogram is as Figure 18 shown, with a purity of: 93.27%; the mass spectrum (MS) is as Figure 19 shown. In the conjugate II-1, the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is about 3.7.

[0800]

[0801] Compound R10-1 and Fc-PEG4-N3 were reacted via click chemistry to obtain conjugate II-2 in the same manner, where the Fc domain monomer (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0802]

[0803] Compound R10-2 and Fc-PEG5-N3 were reacted via click chemistry to obtain conjugate II-3 in the same manner, where the Fc domain monomer (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0804]

[0805] Compound R10-3 and Fc-PEG6-N3 were reacted via click chemistry to obtain conjugate II-4 in the same manner, where the Fc domain monomer (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.7.

[0806]

[0807] Compound P8 and Fc-PEG4-N3 were reacted via click chemistry to obtain conjugate II-6 in the same manner, and its HPLC chromatogram is as Figure 20 shown, with a purity of: 100.00%; the mass spectrometry (MS) is as Figure 21 shown. In conjugate II-6, the Fc domain monomer (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.5.

[0808]

[0809] Compound P7 and Fc-PEG4-N3 were reacted via click chemistry to obtain conjugate II-5 in the same manner, where the Fc domain monomer (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.5.

[0810]

[0811] Compound P8-1 and Fc-PEG6-N3 were reacted via click chemistry to obtain conjugate II-7 in the same manner, where the Fc domain monomer (the amino acid sequence of which is shown in SEQ ID NO: 64), I2 represents the Fc protein, and T is approximately 3.5.

[0812]

[0813] Compound P8-2 and Fc-PEG4-N3 were subjected to click reaction in the same manner to obtain conjugate II-8, where the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents Fc protein, and T is about 3.5.

[0814]

[0815] Compound O5 and Fc-PEG4-N3 were subjected to click reaction in the same manner to obtain conjugate II-9, where the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents Fc protein, and T is about 3.2.

[0816]

[0817] Compound O5-1 and Fc-PEG4-N3 were subjected to click reaction in the same manner to obtain conjugate II-10, where the Fc domain monomer (the amino acid sequence thereof is as shown in SEQ ID NO: 64), I2 represents Fc protein, and T is about 3.2.

[0818]

[0819] Example 9-1 Synthesis of Conjugates II

[0820] 1. Preparation of Compound P12-A

[0821]

[0822]

[0823] At room temperature, in N,N-dimethylformamide (2 mL), dimethyl sulfoxide (2 mL) and water (4 mL) dissolved with compound P12 (300 mg, 0.140 mmol) and 2,3,5,6-tetrafluorophenyl 1-azido-3,6,9,12-tetraoxapentadecane-15-carboxylate (93 mg, 0.212 mmol), an aqueous solution of copper(II) sulfate pentahydrate (18 mg, 0.072 mmol) and sodium ascorbate (28 mg, 0.141 mmol) was added successively. After addition, the whole mixture was stirred at room temperature for 30 min. The reaction solution was directly separated by preparative HPLC without treatment. Preparation conditions: Shim-pack GIST C18 (20 mm×250 mm, 5 μm); A: 0.1% TFA in water, B: MeCN; 40 mL / min; 0 - 3 min, 10% B; 3 - 3.5 min, 10% - 25% B; 3.5 - 14 min, 25% - 60% B; 14 - 14.5 min, 60% - 100% B; detector, UV 254 nm. The product P12-A as a white solid was obtained (98.28 mg, 0.034 mmol, 24.03%). 1 1H NMR (500 MHz, DMSO-d6) δ ppm 12.64 (br s, 3H), 8.01 (s, 3H), 7.63 (d, J = 7.6 Hz, 3H), 7.29 (br s, 4H), 7.01 (t, J = 6.3 Hz, 3H), 6.82 - 6.88 (m, 3H), 6.70 (br s, 4H), 5.14 - 5.20 (m, 3H), 4.45 - 4.54 (m, 15H), 4.34 (t, J = 10.4 Hz, 3H), 3.75 - 3.85 (m, 14H), 3.66 (s, 6H), 3.35 - 3.56 (m, 44H), 3.06 - 3.18 (m, 6H), 3.01 (t, J = 6.0 Hz, 2H), 2.57 - 2.69 (m, 7H), 2.26 - 2.34 (m, 3H), 1.96 - 2.04 (m, 2H), 1.78 (s, 9H), 1.52 - 1.60 (m, 3H), 1.40 - 1.48 (m, 5H), 1.20 - 1.32 (m, 12H), 1.01 - 1.08 (m, 4H), 0.78 - 0.91 (m, 21H); ESI-MS m / z = 1289.3 [M / 2 + H] + ; Calcu. = 2575.3。

[0824] 2. Synthesis of conjugate I-6

[0825] Prepare a stock solution of P12-A at 50 mg / mL using DMSO. Take Fc protein (10 mg / mL, 37.32 mg, 0.67 μmol), displace it into 50 mM PBS (pH 7.4) solution, add 11 eq of the stock solution of P12-A small molecule (50 mg / mL, 19.05 mg, 7.4 μmol), with DMSO accounting for 10% of the entire system. Shake at room temperature overnight, and use a 30 kD ultrafiltration centrifugal tube (Millipore) to displace the sample into 20 mM histidine buffer (pH 5.0) to obtain a total of 12.5 mg of conjugate I-6. Its SEC-HPLC chromatogram is as shown in Figure 1 shown, with a purity of 99.6%; the mass spectrometry (MS) is as shown in Figure 2 shown, and the DAR value is 3.8.

[0826] Conjugates I-1 to I-5, I-7 to I-10, and conjugates II-1 to II-10 can also be synthesized and prepared using the same method.

[0827] Example 10 Evaluation of NA Enzyme Activity Inhibition

[0828] To verify the activity of inhibiting influenza virus at the enzyme level, the poly-small molecule derivatives, conjugates, and small molecule drugs (zanamivir (purchased from Adamas-beta, Catalog# C63-102157A), peramivir (Bidepharm, Catalog# BD317210), oseltamivir phosphate (purchased from InnoChem, Catalog# A90108)) provided in the examples of this application were tested for the half-maximal inhibitory concentration (IC 50 ) against NA (A / California / 04 / 2009, H1N1, purchased from Sino Biological, Catalog# 11058-VNAHC). Mix the serially diluted drugs with NA protein and incubate together at room temperature for 30 minutes, then add the MUNANA substrate (purchased from Sigma, Catalog# M8639), detect its fluorescence intensity on an enzyme-linked immunosorbent assay (ELISA) reader (Molecular Devices, SpectraMax Gemini EM), and finally calculate the IC 50 , as shown in Figure 1 and Figure 2 shown, and the conjugates all showed good NA enzyme activity inhibition ability. [[ID=~23]]

[0829] Example 11 Evaluation of NA Enzyme Activity Inhibition of Drug-Resistant Strains

[0830] To verify the activity of inhibiting influenza virus at the enzyme level, the poly-small molecule derivatives and conjugates provided in the examples of this application, as well as small molecule drugs (zanamivir, peramivir, oseltamivir phosphate), were tested for their half-maximal inhibitory concentration (IC 50 ) against NA (A / California / 04 / 2009, H275Y, H1N1, purchased from Sino Biological, Catalog#11058-VNAHC). The gradient-diluted drugs were mixed with NA protein and co-incubated at room temperature for 30 minutes, then MUNANA substrate was added, and its fluorescence intensity was detected on a microplate reader. Finally, the IC 50 was calculated. As shown in Figure 3 and Figure 4 , the conjugates showed good inhibitory ability against the NA enzyme activity of drug-resistant strains.

[0831] Example 12 Evaluation of Anti-Influenza Virus Activity in Vitro

[0832] To verify the activity of inhibiting influenza virus at the cellular level, its antiviral activity was tested in a model of MDCK cells (kindly provided by Xiamen University) infected with nine strains of viruses: A / Victoria / 2570 / 2009 (H1N1) (kindly provided by Xiamen University), A / California / 04 / 2009 (H1N1) (kindly provided by Xiamen University), A / Vietnam / 2004 (H5N1), A / Shanghai / 02 / 2013 (H7N9) (kindly provided by Xiamen University), B / Phuket / 3073 / 2013 (BY) (kindly provided by Xiamen University), A / Puerto Rico / 8 / 1934 (H1N1-PR8) (purchased from NIBSC, NIBSC16 / 108), A / Victoria / 2570 / 2019 (H1N1) (purchased from NIBSC, NIBSC21 / 346), A / Darwin / 9 / 2021 (H3N2) (purchased from NIBSC, NIBSC21 / 214), and B / Austria / 1359417 / 2021 (BV) (purchased from NIBSC, NIBSC21 / 224). Different concentrations of the poly-small molecule derivatives or conjugates or small molecule drugs (zanamivir, peramivir, oseltamic acid) provided in the examples of this application were mixed with the virus. After incubating at 37 °C for one hour, the mixture was added to monolayer MDCK cells at 80 - 90% confluence. After culturing for 90 minutes, the cells were washed. The monolayer cells were then covered with carboxymethyl cellulose (purchased from Energy Chemical, A05925) to minimize virus spread and allow the incubation to continue for two days. After two days of culture, the effective drug concentration (EC 50) The results are shown in Tables 1 and 2. The inhibition curves of the drug against A / Puerto Rico / 8 / 1934 (H1N1-PR8), A / Victoria / 2570 / 2019 (H1N1), A / Darwin / 9 / 2021 (H3N2), and B / Austria / 1359417 / 2021 (BV) viruses are as Figures 26 - 29 shown.

[0833] Table 1

[0834]

[0835] Table 2

[0836]

[0837] Example 13 Pharmacokinetic Analysis

[0838] To verify whether the conjugate drug provided in the examples of this application has a long-lasting protective effect, the pharmacokinetics of the conjugate drug in mice were analyzed. Four healthy BALB / c mice (purchased from Vital River) were selected for each group and intravenously injected with 10 mg / kg of I-1, 1-6, or Fc protein (purchased from Sino Biological, customized, JS95A). At different time points, 20 μL of blood samples were taken from the mice to detect the drug concentration. Blood samples were taken at different intervals. The collected blood samples were immediately added with anticoagulant (saturated EDTA-Na2), and the supernatant was extracted after centrifugation for standby. A 96-well plate was coated with NA protein (purchased from Sino Biological, 11058-V08B-(MF14DE2511)), and incubated with different concentrations of I-1 and I-6 at room temperature for 1 hour. The drug concentration in the serum samples was detected by sandwich ELISA using Goat anti-human IgG (Fc) Antibody, Peroxidase-Labeled (SeraCare, CAT: 5220-0279). The results are as Figure 5 shown, indicating that the conjugate has a good plasma half-life and can maintain above the effective drug concentration for a long time.

[0839] Example 14 ADCC Effect Analysis

[0840] Fc has certain biological functions and may activate the body's ADCC and ADCP effects to enhance antiviral activity. To verify whether the conjugate has this function, a virus-infected cell model was constructed and cultured at 37°C and 5% CO2 for 18 - 24 hours. Then, an ADCC reporter cell line that stably expresses the FcγRIIIa receptor was added. Finally, drugs to be tested at different concentrations were added. After incubating for 15 minutes at 37°C and 5% CO2, the fluorescence intensity emitted by the ADCC reporter cell line was detected, and then the intensity of the ADCC effect caused by the conjugate was detected. As Figure 6 shown, the conjugate provided in the embodiments of the present application can maintain the same ADCC effect as Fc.

[0841] Example 15 Evaluation of the Conjugate's Inhibition of Virus Replication in Mice

[0842] 1. To study whether the conjugate provided in the embodiments of the present application can inhibit virus replication in the lungs and nasal turbinates of mice, approximately 1×10 6 PFU or approximately 1×10 7 PFU of H1N1 influenza virus (A / California / 07 / 2009, generously provided by Xiamen University) was instilled into the nasal cavities of 6-week-old BALB / c mice (purchased from Vital River). Drugs were administered 2 hours after virus challenge. The conjugate (3, 9 mg / kg), PBS, or negative control Fc protein (purchased from Sino Biological, custom-made, JS95A, 3 mg / kg) was intravenously injected as a single dose. The positive control zanamivir (5 mg / kg) was intravenously injected 2 hours after virus challenge, once a day for 3 consecutive days. Oseltamivir phosphate (20 mg / kg) was orally administered 2 hours after virus challenge, twice a day for 3 consecutive days. All mice were euthanized on the fourth day after virus challenge, and the viral loads in the lungs and nasal turbinates were detected. The results are as Figure 7 shown, indicating that the conjugate provided in the embodiments of the present application can effectively inhibit virus replication at different virus challenge doses.

[0843] 2. BALB / c mice were inoculated with 10×LD 50 of influenza virus (A / Puerto Rico / 8 / 1934 (H1N1-PR8)). Drugs were administered 2 hours after virus challenge. The conjugate (3 mg / kg) was intravenously injected as a single dose, and the positive controls oseltamivir (10 mg / kg) and baloxavir marboxil (5 mg / kg) were orally administered. The virus content in the bronchoalveolar lavage fluid was detected on the 6th day after drug administration, and the body weight and survival rate were monitored daily. The results are as Figures 30 - 32 shown, indicating that therapeutic administration of the conjugate provided in the embodiments of the present application, such as I-6, I-1, II-6, can effectively inhibit virus replication, has low toxicity and side effects, the body weight of mice can increase normally, and the survival rate of mice is high.

[0844] 3. BALB / c mice were intravenously injected with the conjugate at a single dose of 3 mg / kg, and the positive controls oseltamivir (10 mg / kg) and baloxavir marboxil (5 mg / kg) were orally administered. After 15 days of administration, the mice were challenged with 50 influenza virus (A / Puerto Rico / 8 / 1934 (H1N1-PR8)) at a dose of 10×LD Figures 33 - 35 . On the 6th day after inoculation, the virus content in the bronchoalveolar lavage fluid was detected, and the body weight and survival rate were monitored daily. The results are as

[0845] shown, indicating that prophylactic administration of the conjugate provided in the examples of the present application, such as I-6, I-1, and II-6, can effectively inhibit virus replication, with low toxicity and side effects, and the body weight of the mice can increase normally, and the survival rate of the mice is high.

[0846] In order to significantly compare the antiviral activities of the conjugates provided in the examples of the present application, a mouse model of influenza-susceptible DBA / 2 (purchased from Vital River) was selected. Compared with the BALB / c mouse model, this model can develop severe influenza symptoms at a lower challenge dose. Six-week-old female DBA / 2 mice (10 mice per group) were challenged by nasal drip with a dose of 1×10 4 PFU of A / California / 07 / 2009. Two hours after challenge, the conjugates (I-1, I-5) (0.3, 3 mg / kg), PBS, or the negative control Fc protein (3 mg / kg, 9 mg / kg, purchased from Sino Biological, customized, JS95A) were injected via the tail vein once. The positive control zanamivir (5 mg / kg) was intravenously injected two hours after challenge, once a day for 5 consecutive days, and oseltamivir phosphate (20 mg / kg) was orally administered two hours after challenge, twice a day for 5 consecutive days. The body weight and survival rate were monitored daily (a 25% decrease in body weight was considered death), and the observation continued for 14 days. The results are as Figures 9 - 12 shown. In the DBA / 2 mouse model, as Figure 8 can be seen, the body weight dropped to the lowest on the 7th day after challenge. Among them, I-1 (3 mg / kg) could significantly relieve the body weight loss of the mice. While the PBS group and the Fc protein control group all died on the fifth day ( Figure 9 ), and the positive control drugs oseltamivir phosphate and zanamivir also all died in the mice after high-dose and multiple-frequency administration ( Figure 10 ), while I-1 and I-5 still achieved a good protection rate with a single dose as low as 3 mg / kg ( Figure 11 ), and I-5 could achieve a protection rate as high as 90%. The above results reflect the significant antiviral activity of the single-dose conjugate.

Claims

1. The conjugate of formula (I) or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1, A2 and A3 are each independently selected from formulae (A-I) to (A-XIV), Among them, L can be attached to any possible site of the compound of formula (A-XIV); wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2 and -NHC(=NH)NHR6; R2 and R3 are each independently selected from -H, -OH, -F, -Cl and -Br; R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3 and -C(=O)OCF3; R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3; X is selected from -O- and -S-; Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)- and -NH(SO2)NR7-; R' is H or C1-C6 alkyl; R6 is selected from R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl; each E is a biological macromolecule; n is 1 or 2; T is a number between 1 and 20; The wavy lines connecting to each E represent each covalently linked (e.g., by a covalent bond or a linker) to an N atom in each E (e.g., the N atom on the side-chain amino group of a lysine, arginine, asparagine, or glutamine residue), and L is a linker covalently linked to each of the groups E, A1, A2, and A3.

2. The conjugate of claim 1 or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1, A2 and A3 are each independently selected from formulae (A-I) to (A-XIII), Preferably, A1, A2 and A3 are each independently selected from formulae (A-I) to (A-IV) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X and Y are as described in claim 1; each E comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence of any one of SEQ ID NOs: 1-68), albumin (e.g., albumin having a sequence of any one of SEQ ID NOs: 69-71), an albumin-binding peptide or an Fc-binding peptide; n is 1 or 2, and when n is 2 and E is an Fc domain monomer, two Es dimerize to form an Fc domain; L is a linker covalently attached to each of the groups E, A1, A2 and A3; Preferably, A1, A2 and A3 are each independently selected from formulae (A-I) to (A-III) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X and Y are as above; Preferably, A1, A2 and A3 are each independently selected from formula (A-I), formula (A-III) and formula (A-VII), wherein the definitions of R1, R4, R5, R', X and Y are as above.

3. The conjugate of claim 1 or 2, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein E comprises a monomer of the Fc domain (e.g., a monomer of the Fc domain having a sequence selected from SEQ ID NOs: 1-68); n is 1 or 2, and when n is 2, two Es dimerize to form the Fc domain; L is a linker covalently linked to each of the groups E, A1, A2 and A3; Preferably, the monomer of the Fc domain has an amino acid sequence selected from: i) Any one of the sequences shown in SEQ ID NOs: 1-68; ii) A sequence having one or several amino acid substitutions, deletions or additions (e.g., substitution, deletion or addition of 1, 2, 3, 4 or 5 amino acids) as compared with any one of the sequences shown in SEQ ID NOs: 1-68; and iii) A sequence having at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) with any one of the sequences shown in SEQ ID NOs: 1-68; Preferably, the substitution in ii) is a conservative substitution; Preferably, E is a monomer of the Fc domain having a sequence with at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) with the sequence of SEQ ID NO: 64; Preferably, E is a monomer of the Fc domain having the sequence of SEQ ID NO: 64; Preferably, R1 is selected from -OH, -NH2, -NHC(=NH)NH2; more preferably, R1 is -NHC(=NH)NH2; Preferably, R4 is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3; more preferably, R4 is selected from -C(=O)OH and -C(=O)OCH3; Preferably, R5 is selected from -C(=O)CH3 and -C(=O)CF3; more preferably, R5 is -C(=O)CH3; Preferably, R' is H or C1-C4 alkyl; more preferably, R' is H, methyl or ethyl; more preferably, R' is H or ethyl; Preferably, X is -O-; Preferably, Y is selected from -O-, -S-, -NH-; more preferably, Y is selected from -O- and -NH-.

4. The conjugate of any one of claims 1-3, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has a structure shown in formula (I-II-A), formula (I-II-B) or formula (I-II-C), wherein the definitions of R1, R4, R5, R', X, Y, E, n, L, T are as described in any one of claims 1-3.

5. The conjugate of any one of claims 1-4 or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has a structure represented by formula (I-II-A'), formula (I-II-B') or formula (I-II-C'), wherein the definitions of R1, R4, R5, R', X, Y, E, n, and T are as described in any one of claims 1-4. L1 is selected from: Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20; Preferably, L1 is selected from: wherein V, p, m, and Z are defined as described above; Preferably, L1 is wherein V, p, m, and Z are defined as described above; Preferably, V is -CH2- or -O-; more preferably, V is -O-. Preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)- or -C(=O)-; more preferably, Z is -NH-C(=O)- or -C(=O)-. Preferably, p is an integer between 1 and 10; more preferably, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6). Preferably, m is an integer between 1 and 12; more preferably, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

6. The conjugate of any one of claims 1-5 or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has a structure represented by formula (I-II-A'-1), formula (I-II-B'-1), formula (I-II-B'-2) or formula (I-II-C'-1); wherein the definitions of E, n, and T are as described in any one of claims 1-4, and the definition of L1 is as described in claim 5.

7. The conjugate of any one of claims 1-6 or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has a structure represented by formula (I-II-A-2), formula (I-II-B-3), formula (I-II-B-4) or formula (I-II-C-2), wherein the definitions of E, n, and T are as described in any one of claims 1-4, and the definition of L1 is as described in claim 5; L2 is selected from: Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20; Preferably, L2 is selected from: Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20; L3 is wherein U is -C(=O)-NH-、 -NH-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, -CH2-; W is -CH2-, -O- or -S-; i and q are each independently an integer from 1 to 20, one end of L3 is covalently linked to E, and the other end is covalently linked to L2; Preferably, L1 is wherein V, p, m, and Z are defined as described above; preferably, V is -CH2- or -O-; more preferably, V is -O-; preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, or -C(=O)-; more preferably, Z is -NH-C(=O)- or -C(=O)-; preferably, p is an integer between 1 and 10; more preferably, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6); preferably, m is an integer between 1 and 12; more preferably, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9); Preferably, L2 is wherein V, p, m, and Z are defined as above; preferably, V is -CH2- or -O-; more preferably, V is -O-; preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-; more preferably, Z is -C(=O)-NH-; preferably, p is an integer between 1 and 10; more preferably, p is an integer between 1 and 6 (such as 1, 2, 3, 4, 5, 6); preferably, m is an integer between 1 and 12; more preferably, m is an integer between 1 and 10 (such as 1, 2, 3, 4, 5, 6, 7, 8, 9); Preferably, L3 is wherein U, i, and q are defined as described above; preferably, U is -NH-C(=O)-, -N(CH3)-C(=O)-, or -N(CH2CH3)-C(=O)-; more preferably, U is -NH-C(=O)-; preferably, i is an integer between 1 and 12; more preferably, i is an integer between 1 and 10 (such as 1, 2, 3, 4, 5, 6, 7, 8, 9); preferably, q is an integer between 1 and 10; more preferably, q is an integer between 1 and 6 (such as 1, 2, 3, 4, 5, 6); Preferably, T is a number between 1 and 10; more preferably, T is a number between 1 and 9; more preferably, T is a number between 1 and 7; more preferably, T is a number between 1 and 5, more preferably, T is a number between 3 and 7, more preferably, T is a number between 3 and 5.

5.

8. The conjugate according to any one of claims 1-7, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has the structure shown in formula (I-II-A-3), formula (I-II-B-5), formula (I-II-B-6) or formula (I-II-C-3), wherein E, n, T are defined as in any one of claims 1-7; n1, n2, n3 are each independently an integer between 1 and 20; Preferably, n1, n2, n3 are each independently an integer between 1 and 12; More preferably, n1, n2, n3 are each independently an integer between 1 and 10 (such as 1, 2, 3, 4, 5, 6, 7, 8, 9).

9. The conjugate according to any one of claims 1-8, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate is selected from: wherein E, n, T are defined as in any one of claims 1-8.

10. The conjugate shown in formula (II), or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1, A2 are each independently selected from formula (A-I) to formula (A-XIV), Among them, L can be connected to any possible site of the compound of formula (A-XIV); wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2 and -NHC(=NH)NHR6; R2 and R3 are each independently selected from -H, -OH, -F, -Cl and -Br; R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3 and -C(=O)OCF3; R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3; X is selected from -O- and -S-; Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)- and -NH(SO2)NR7-; R' is H or C1-C6 alkyl; R6 is selected from R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl; Each E is a biological macromolecule; n is 1 or 2; T is a number between 1 and 20; The wavy line connected to each E indicates that each A1-L-A2 is covalently connected (for example, by means of a covalent bond or a linker) to the N atom in each E (for example, the N atom on the side chain amino group of a lysine, arginine, asparagine or glutamine residue), and L is a linker covalently connected to each of the groups E, A1, A2.

11. The conjugate of claim 10 or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1 and A2 are each independently selected from formula (A-I) to formula (A-XIII), Preferably, A1 and A2 are each independently selected from formula (A-I) to formula (A-IV) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in claim 10; Each E comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence of any one of SEQ ID NOs: 1-68), albumin (e.g., albumin having a sequence of any one of SEQ ID NOs: 69-71), an albumin-binding peptide or an Fc-binding peptide; n is 1 or 2, and when n is 2 and E is an Fc domain monomer, two Es dimerize to form an Fc domain; Preferably, A1 and A2 are each independently selected from formula (A-I) to formula (A-III) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are the same as above; Preferably, A1 and A2 are each independently selected from formula (A-I), formula (A-III), and formula (A-VII), wherein the definitions of R1, R4, R5, R', X, and Y are the same as above.

12. The conjugate of claim 10 or 11 or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein E comprises an Fc domain monomer (e.g., an Fc domain monomer having a sequence of any one of SEQ ID NOs: 1-68); n is 1 or 2, and when n is 2, two Es dimerize to form an Fc domain; Preferably, the Fc domain monomer has an amino acid sequence selected from the following: i) Any one of the sequences shown in SEQ ID NOs: 1-68; ii) A sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to any one of the sequences shown in SEQ ID NOs: 1-68; and iii) A sequence having at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) with any one of the sequences shown in SEQ ID NOs: 1-68; Preferably, the substitution in ii) is a conservative substitution; Preferably, E is an Fc domain monomer having a sequence with at least 70% identity (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity) with the sequence of SEQ ID NO: 64; Preferably, E is an Fc domain monomer having the sequence of SEQ ID NO: 64; Preferably, R1 is selected from -OH, -NH2, -NHC(=NH)NH2; more preferably, R1 is -NHC(=NH)NH2; Preferably, R4 is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3; More preferably, R4 is selected from -C(=O)OH and -C(=O)OCH3; Preferably, R5 is selected from -C(=O)CH3 and -C(=O)CF3; More preferably, R5 is -C(=O)CH3; Preferably, R' is H or a C1-C4 alkyl group; More preferably, R' is H, methyl or ethyl; Even more preferably, R' is H or ethyl; Preferably, X is -O-; Preferably, Y is selected from -O-, -S-, -NH-; More preferably, Y is selected from -O- and -NH-.

13. The conjugate according to any one of claims 10-12, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has the structure shown in formula (II-II-A), formula (II-II-B) or formula (II-II-C), wherein the definitions of R1, R4, R5, R', X, Y, E, n, L, T are as described in any one of claims 10-12.

14. The conjugate according to any one of claims 10-13, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has the structure shown in formula (II-II-A'), formula (II-II-B') or formula (II-II-C'), wherein the definitions of R1, R4, R5, R', X, Y, E, n, T are as described in any one of claims 10-12; L1 is selected from: Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20; Preferably, L1 is selected from: wherein the definitions of V, p, m, and Z are as described above; Preferably, L1 is wherein V, p, m, and Z are defined as described above; Preferably, V is -CH2- or -O-; More preferably, V is -O-; Preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)- or -C(=O)-; More preferably, Z is -NH-C(=O)- or -C(=O)-; Preferably, p is an integer between 1 and 10; More preferably, p is an integer between 1 and 6 (such as 1, 2, 3, 4, 5, 6); Preferably, m is an integer between 1 and 12; More preferably, m is an integer between 1 and 10 (such as 1, 2, 3, 4, 5, 6, 7, 8, 9).

15. The conjugate according to any one of claims 10-14, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has the structure shown in formula (II-II-A-1), formula (II-II-B-1), formula (II-II-B-2) or formula (II-II-C-1), wherein the definitions of E, n, T are as described in any one of claims 10-13, and the definition of L1 is as described in claim 14.

16. The conjugate of any one of claims 10 - 15, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has the structure shown in formula (II-II-A-2), formula (II-II-B-3), formula (II-II-B-4) or formula (II-II-C-2), wherein E, n, and T are defined as in any one of claims 10 - 13, and L1 is defined as in claim 14; L2 is selected from: Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20; Preferably, L2 is selected from: wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; m and p are each independently an integer from 1 to 20; L3 is wherein U is -C(=O)-NH-、 -NH-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)-, -CH2-; W is -CH2-, -O- or -S-; i and q are each independently an integer from 1 to 20, one end of L3 is covalently linked to E, and the other end is covalently linked to L2; Preferably, L1 is wherein V, p, m, and Z are defined as above; preferably, V is -CH2- or -O-; more preferably, V is -O-; preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, or -C(=O)-; more preferably, Z is -NH-C(=O)- or -C(=O)-; preferably, p is an integer between 1 and 10; more preferably, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6); preferably, m is an integer between 1 and 12; more preferably, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9); Preferably, L2 is wherein V, p, m, and Z are defined as described above; preferably, V is -CH2- or -O-; more preferably, V is -O-; preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-; more preferably, Z is -C(=O)-NH-; preferably, p is an integer between 1 and 10; more preferably, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6); preferably, m is an integer between 1 and 12; more preferably, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9); Preferably, L3 is wherein U, i, and q are defined as described above; preferably, U is -NH-C(=O)-, -N(CH3)-C(=O)-, or -N(CH2CH3)-C(=O)-; more preferably, U is -NH-C(=O)-; preferably, i is an integer between 1 and 12; more preferably, i is an integer between 1 and 10 (such as 1, 2, 3, 4, 5, 6, 7, 8, 9); preferably, q is an integer between 1 and 10; more preferably, q is an integer between 1 and 6 (such as 1, 2, 3, 4, 5, 6). Preferably, T is a number between 1 and 10; more preferably, T is a number between 1 and 9; more preferably, T is a number between 1 and 7; more preferably, T is a number between 1 and 5, more preferably, T is a number between 3 and 7, more preferably, T is a number between 3 and 5.

5.

17. The conjugate of any one of claims 10 - 16, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate has the structure shown in formula (II-II-A-3), formula (II-II-B-5), formula (II-II-B-6) or formula (II-II-C-3), wherein E, n, and T are defined as in any one of claims 10 - 16; n1, n2, and n3 are each independently an integer between 1 and 20; Preferably, n1, n2, and n3 are each independently an integer between 1 and 12; more preferably, n1, n2, and n3 are each independently an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

18. The conjugate of any one of claims 10 - 17, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the conjugate is selected from: wherein E, n, and T are defined as in any one of claims 10 - 17.

19. A compound of formula (IV) or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1, A2, and A3 are each independently selected from formula (A-I) to formula (A-XIV), Among them, L can be linked to any possible site of the compound of formula (A-XIV); wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2, and -NHC(=NH)NHR6; R2 and R3 are each independently selected from -H, -OH, -F, -Cl, and -Br; R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3, and -C(=O)OCF3; R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3; X is selected from -O- and -S-; Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)- and -NH(SO2)NR7-; R' is H or C1-C6 alkyl; R6 is selected from R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl; L' is a linker covalently linked to each of the groups A1, A2 and A3.

20. The compound of claim 19 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1, A2 and A3 are each independently selected from formula (A-I) to formula (A-XIII), Preferably, A1, A2, A3 are each independently selected from formula (A-I) to formula (A-IV) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X, Y are as described in claim 19; Preferably, A, A2 and A3 are each independently selected from formula (A-I) to formula (A-III) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X, Y are the same as above; Preferably, A1, A2 and A3 are each independently selected from formula (A-I), formula (A-III) and formula (A-VII), wherein the definitions of R1, R4, R5, R', X, Y are the same as above.

21. The compound of claim 19 or 20 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2; more preferably, R1 is -NHC(=NH)NH2; Preferably, R4 is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3; more preferably, R4 is selected from -C(=O)OH and -C(=O)OCH3; Preferably, R5 is selected from -C(=O)CH3 and -C(=O)CF3; more preferably, R5 is -C(=O)CH3; Preferably, R' is H or C1-C4 alkyl; more preferably, R' is H, methyl or ethyl; more preferably, R' is H or ethyl; Preferably, X is -O-; Preferably, Y is selected from -O-, -S-, -NH-; more preferably, Y is selected from -O- and -NH-.

22. The compound of any one of claims 19-21 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound has the structure shown in formula (IV-I), wherein the definitions of A1, A2 and A3 are as described in any one of claims 19-21, L1 is selected from: L2' is selected from: wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; or Z is Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; W' is HC≡C-, -C(=O)OH, -NH2 or N3-; m and p are each independently an integer from 1 to 20; Preferably, L1 is selected from: wherein V, p, m, and Z are defined as described above; Preferably, L1 is wherein V, p, m, and Z are defined as described above; Preferably, L2’ is wherein W’, V, p, m, and Z are defined as described above; Preferably, L2’ is wherein W’, V, p, m, and Z are defined as described above; Preferably, L2’ is wherein W’, p, m, and Z are defined as described above; Preferably, V is -CH2- or -O-; more preferably, V is -O-; Preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)- or -C(=O)-; More preferably, Z is -NH-C(=O)- or -C(=O)-; Preferably, W' is HC≡C-, -C(=O)OH, -NH2 or N3-; More preferably, W' is HC≡C-; Preferably, W’ is Preferably, Z is Preferably, Z1 is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)- or -CH2CH2-C(=O)-; More preferably, Z1 is -C(=O)-NH- or -CH2-C(=O)-NH-; Even more preferably, Z1 is -C(=O)-NH-; Preferably, p is an integer between 1 and 10; More preferably, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6); Preferably, m is an integer between 1 and 12; More preferably, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

23. The compound according to any one of claims 19 - 22, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound has the structures shown in formula (IV-I-1), formula (IV-I-2), formula (IV-I-3): wherein the definitions of R1, R4, R5, R', X, Y are as described in any one of claims 19 - 21, and the definitions of L1, L2' are as described in claim 22.

24. The compound according to any one of claims 19 - 23, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound has the structures shown in formula (IV-I-1-1), formula (IV-I-2-1), formula (IV-I-2-2) or formula (IV-I-3-1): wherein the definitions of L1, L2' are as described in claim 22.

25. The compound according to any one of claims 19 - 24, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound is selected from:

26. The compound shown in formula (V) or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1 and A2 are each independently selected from formula (A-I) to formula (A-XIV), Among them, L can be connected to any possible site of the compound of formula (A-XIV); wherein R1 is selected from -OH, -NH2, -NHC(=NH)NH2 and -NHC(=NH)NHR6; R2 and R3 are each independently selected from -H, -OH, -F, -Cl and -Br; R4 is selected from -C(=O)OH, -P(=O)(OH)2, -SO3H, -C(=O)OCH3 and -C(=O)OCF3; R5 is selected from -C(=O)CH3, -C(=O)CF3, -SO2CH3; X is selected from -O- and -S-; Y is selected from -O-, -S-, -NR7-, -OC(=O)NR7-, -OC(=S)NR7-, -OC(=O)O-, -OC(=O)-, -NHC(=O)O-, -NHC(=O)-, -NHC(=NH)-, -NHC(=O)NR7-, -NHC(=NH)NR7-, -NHC(=S)NR7-, -NHC(=S)-, -OCH2C(=O)NR7-, -NH(SO2)- and -NH(SO2)NR7-; R' is H or C1-C6 alkyl; R6 is selected from R7 is selected from H, C1-C 20 alkyl, C3-C 20 cycloalkyl, C3-C 20 heterocycloalkyl; C5-C 15 aryl and C5-C 15 heteroaryl; preferably, R7 is H; preferably, C1-C6 alkyl; L' is a linker covalently linked to each of the groups A1 and A2.

27. The compound of claim 26 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein A1 and A2 are each independently selected from formula (A-I) to formula (A-XIII), Preferably, A1 and A2 are each independently selected from formula (A-I) to formula (A-IV) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as described in claim 21; Preferably, A1 and A2 are each independently selected from formula (A-I) to formula (A-III) and formula (A-VII), wherein the definitions of R1, R2, R3, R4, R5, R', X, and Y are as above; Preferably, A1 and A2 are each independently selected from formula (A-I), formula (A-III) and formula (A-VII), wherein the definitions of R1, R4, R5, R', X, and Y are as above.

28. The compound of claim 26 or 27 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein Preferably, R1 is selected from -OH, -NH2, -NHC(=NH)NH2; more preferably, R1 is -NHC(=NH)NH2; Preferably, R4 is selected from -C(=O)OH, -C(=O)OCH3 and -C(=O)OCF3; more preferably, R4 is selected from -C(=O)OH and -C(=O)OCH3; Preferably, R5 is selected from -C(=O)CH3 and -C(=O)CF3; more preferably, R5 is -C(=O)CH3; Preferably, R' is H or C1-C4 alkyl; more preferably, R' is H, methyl or ethyl; more preferably, R' is H or ethyl; Preferably, X is -O-; Preferably, Y is selected from -O-, -S-, -NH-; more preferably, Y is selected from -O- and -NH-.

29. The compound of any one of claims 26-28 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound has the structure shown in formula (V-I), wherein the definitions of A1 and A2 are as described in any one of claims 26-28, L1 is selected from: L2' is selected from: Wherein V is -CH2-, -O- or -S-; Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; Z1 and Z2 are each independently selected from -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)-, -O-C(=O)-, -C(=O)- or -CH2-; W’ is HC≡C-, -C(=O)OH, -NH2 or N3-; m and p are each independently an integer from 1 to 20; Preferably, L1 is selected from: wherein the definitions of V, p, m, and Z are as described above; Preferably, L1 is wherein V, p, m, and Z are defined as described above; Preferably, L2' is wherein W', V, p, m, and Z are defined as described above; Preferably, L2’ is wherein W’, V, p, m, and Z are defined as described above; Preferably, V is -CH2- or -O-; more preferably, V is -O-; Preferably, Z is -C(=O)-NH-, -CH2-C(=O)-NH-, -NH-C(=O)-, -CH2CH2-C(=O)-, -N(CH3)-C(=O)-, -N(CH2CH3)-C(=O)- or -C(=O)-; More preferably, Z is -NH-C(=O)- or -C(=O)-; Preferably, W' is HC≡C-, -C(=O)OH, -NH2 or N3-; More preferably, W' is HC≡C-; Preferably, p is an integer between 1 and 10; More preferably, p is an integer between 1 and 6 (e.g., 1, 2, 3, 4, 5, 6); Preferably, m is an integer between 1 and 12; More preferably, m is an integer between 1 and 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9).

30. A compound according to any one of claims 26-29, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound has a structure represented by formula (V-I-1), formula (V-I-2) or formula (V-I-3): wherein the definitions of R1, R4, R5, R', X and Y are as described in any one of claims 26-28, and the definitions of L1 and L2' are as described in claim 29.

31. A compound according to any one of claims 26-30, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound has a structure represented by formula (V-I-1-1), formula (V-I-2-1), formula (V-I-2-2) or formula (V-I-3-1): wherein the definitions of L1 and L2' are as described in claim 29.

32. A compound according to any one of claims 26-31, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, wherein the compound is selected from:

33. A pharmaceutical composition comprising the conjugate according to any one of claims 1 to 18, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, or a compound according to any one of claims 19-32, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, and a pharmaceutically acceptable carrier and / or excipient.

34. Use of the conjugate according to any one of claims 1 to 18, or a pharmaceutically acceptable salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, or a compound according to any one of claims 19-32, or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof, or the pharmaceutical composition according to claim 33 in the preparation of a drug for the treatment or prevention of viral infections, Preferably, the viral infection is an infection caused by influenza virus or parainfluenza virus; Preferably, the viral infection is an infection caused by influenza virus type A, B or C or parainfluenza virus; Preferably, the viral infection is an infection caused by avian influenza.

35. The use according to claim 34, wherein the subject can be further treated with an antiviral agent selected from oseltamivir, zanamivir, peramivir, laninamivir, amantadine, rimantadine or baloxavir marboxil.

36. Use of a compound according to any one of claims 19-32 or a salt, geometric or optical isomer, hydrate, solvate or polymorph thereof as an intermediate in the preparation of a conjugate.

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