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KR-20260068077-A - Cysteine-manipulated antibodies and immune conjugates

KR20260068077AKR 20260068077 AKR20260068077 AKR 20260068077AKR-20260068077-A

Abstract

The present invention relates to antibodies engineered with reactive cysteine residues, and more specifically, to antibodies having therapeutic or diagnostic applications. The present invention also relates to immunoconjugates comprising engineered antibodies. The present invention further relates to pharmaceuticals or pharmaceutical compositions comprising antibodies or antibody-drug conjugates, and methods and uses for applying antibodies or immunoconjugates for the treatment of diseases such as tumors.

Inventors

  • 자오 멩후이
  • 주 첸첸
  • 가오 창수

Assignees

  • 이노벤트 바이오로직스 (쑤저우) 컴퍼니, 리미티드

Dates

Publication Date
20260513
Application Date
20240906
Priority Date
20230908

Claims (20)

  1. As a cysteine-engineered antibody or its antigen-binding fragment, (i) The antibody or its antigen-binding fragment comprises a modified Lambda light chain constant region, and the amino acid at position 160 of the modified Lambda light chain constant region is mutated to cysteine compared to the parent Lambda constant region; (ii) The antibody or its antigen-binding fragment comprises a modified lambda light chain constant region, wherein the amino acids at positions 160 and 166 of the modified lambda light chain constant region are mutated to cysteine compared to the parent lambda constant region; (iii) The antibody or its antigen-binding fragment comprises a modified lambda light chain constant region and a modified heavy chain constant region, wherein the amino acid at position 160 of the modified lambda light chain constant region is mutated to cysteine compared to the parent lambda constant region, and the amino acid at position 152 or 155 of the modified heavy chain constant region is mutated to cysteine compared to the parent heavy chain constant region; (iv) The antibody or its antigen-binding fragment comprises a modified lambda light chain constant region and a modified heavy chain constant region, wherein the amino acid at position 166 of the modified lambda light chain constant region is mutated to cysteine compared to the parent lambda constant region, and the amino acid at position 152 or 155 of the modified heavy chain constant region is mutated to cysteine compared to the parent heavy chain constant region; (v) The antibody or its antigen-binding fragment comprises a modified kappa light chain constant region and a modified heavy chain constant region, wherein the amino acid at position 165 of the modified kappa light chain constant region is mutated to cysteine compared to the parent kappa light chain constant region, and the amino acid at position 152 or 155 of the modified heavy chain constant region is mutated to cysteine compared to the parent heavy chain constant region; (vi) The antibody or its antigen-binding fragment comprises a modified kappa light chain constant region, wherein the amino acids at positions 160 and 166 of the modified kappa light chain constant region are mutated to cysteine, and the amino acid at position 178 is mutated to tyrosine compared to the parent kappa constant region; (vii) The antibody or its antigen-binding fragment comprises a modified kappa light chain constant region and a modified heavy chain constant region, wherein the amino acid at position 166 of the modified kappa light chain constant region is mutated to cysteine compared to the parent kappa constant region, and the amino acid at position 375 of the modified heavy chain constant region is mutated to cysteine compared to the parent heavy chain constant region; or (viii) An antibody or its antigen-binding fragment comprising a modified heavy chain constant region, wherein the amino acids at positions 239 and 375 of the modified heavy chain constant region are mutated to cysteine compared to the parent heavy chain constant region, and the antibody or its antigen-binding fragment.
  2. An antibody or an antigen-binding fragment thereof, wherein the parent heavy chain constant region is a human IgG1 heavy chain constant region and comprises, for example, an amino acid sequence described in SEQ ID NO. 3, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 3 and not containing mutations to cysteine.
  3. In claim 1 or 2, the parent lambda light chain invariant region is a human lambda light chain invariant region and comprises, for example, the amino acid sequence described in SEQ ID NO. 6, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 6 and not including mutations to cysteine; or An antibody or an antigen-binding fragment thereof comprising a human kappa light chain constant region, wherein the mokappa light chain constant region is a human kappa light chain constant region and comprises, for example, an amino acid sequence described in SEQ ID NO. 5, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 5 and not containing mutations to cysteine.
  4. In any one of paragraphs 1 through 3, The lambda light chain invariant region modified in item (i) comprises the amino acid sequence described in SEQ ID NO. 7; The lambda light chain constant region modified in item (ii) comprises the amino acid sequence described in SEQ ID NO. 28; In item (iii), the modified lambda light chain constant region comprises the amino acid sequence described in SEQ ID NO. 7, and the modified heavy chain constant region comprises the amino acid sequence described in SEQ ID NO. 10 or 11; In item (iv), the modified lambda light chain constant region comprises the amino acid sequence described in SEQ ID NO. 8, and the modified heavy chain constant region comprises the amino acid sequence described in SEQ ID NO. 10 or 11; In item (v), the modified kappa light chain constant region comprises the amino acid sequence described in SEQ ID NO. 9, and the modified heavy chain constant region comprises the amino acid sequence described in SEQ ID NO. 10 or 11; The modified kappa light chain constant region in item (vi) comprises the amino acid sequence described in SEQ ID NO. 62; In item (vii), the modified kappa light chain constant region comprises the amino acid sequence described in SEQ ID NO. 66, and the modified heavy chain constant region comprises the amino acid sequence described in SEQ ID NO. 67; or The modified heavy chain constant region in item (viii) comprises the amino acid sequence described in SEQ ID NO. 67 and the amino acid sequence described in SEQ ID NO. 68, an antibody or its antigen-binding fragment.
  5. In any one of paragraphs 1 through 4, The modified light chain invariant region in item (i) comprises or consists of the following amino acid sequences: the amino acid sequence described in SEQ ID NO. 16, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 16 and having a substitution with cysteine at position 160, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 16 and including the amino acid sequence described in SEQ ID NO. 7; or The modified light chain invariant region in item (ii) comprises or is composed of the following amino acid sequences: the amino acid sequence described in SEQ ID NO. 18, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 18 and having a cysteine substitution at positions 160 and 166, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 18 and including the amino acid sequence described in SEQ ID NO. 28; or The modified light chain invariant region in item (iii) comprises or consists of the following amino acid sequences: the amino acid sequence described in SEQ ID NO. 16, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 16 and having a substitution with cysteine at position 160, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 16 and including the amino acid sequence described in SEQ ID NO. 7; The modified heavy chain invariant region is the following amino acid sequence: An amino acid sequence described in SEQ ID NO. 25, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 25 and having a substitution with cysteine at position 152, or an amino acid sequence comprising the amino acid sequence described in SEQ ID NO. 10 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 25; or or comprising or composed of: an amino acid sequence described in SEQ ID NO. 26; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 26 and having a substitution with cysteine at position 155; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 26 and including the amino acid sequence described in SEQ ID NO. 11; or The modified heavy chain invariant region contains the modified CH1, and the modified CH1 is the following amino acid sequence: An amino acid sequence described in SEQ ID NO. 22, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 22 and having a substitution with cysteine at position 152, or an amino acid sequence comprising the amino acid sequence described in SEQ ID NO. 10 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 22; or The amino acid sequence described in SEQ ID NO. 23, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 23 and having a substitution with cysteine at position 155, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 23 and including or composed of the amino acid sequence described in SEQ ID NO. 11; The modified light chain invariant region in item (iv) comprises or consists of the following amino acid sequences: the amino acid sequence described in SEQ ID NO. 17, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 17 and having a substitution with cysteine at position 166, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 17 and including the amino acid sequence described in SEQ ID NO. 8; The modified heavy chain invariant region is the following amino acid sequence: An amino acid sequence described in SEQ ID NO. 25, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 25 and having a substitution with cysteine at position 152, or an amino acid sequence comprising the amino acid sequence described in SEQ ID NO. 10 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 25; or or comprising or composed of: an amino acid sequence described in SEQ ID NO. 26; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 26 and having a substitution with cysteine at position 155; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 26 and including the amino acid sequence described in SEQ ID NO. 11; or The modified heavy chain invariant region contains the modified CH1, and the modified CH1 is the following amino acid sequence: An amino acid sequence described in SEQ ID NO. 22, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 22 and having a substitution with cysteine at position 152, or an amino acid sequence comprising the amino acid sequence described in SEQ ID NO. 10 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 22; or or comprising or composed of: an amino acid sequence described in SEQ ID NO. 23; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 23 and having a substitution with cysteine at position 155; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 23 and including the amino acid sequence described in SEQ ID NO. 11; or The modified light chain invariant region in item (v) comprises or consists of the following amino acid sequence: the amino acid sequence described in SEQ ID NO. 20, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 20 and having a substitution with cysteine at position 165, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 20 and including the amino acid sequence described in SEQ ID NO. 9; The modified heavy chain invariant region is the following amino acid sequence: An amino acid sequence described in SEQ ID NO. 25, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 25 and having a substitution with cysteine at position 152, or an amino acid sequence comprising the amino acid sequence described in SEQ ID NO. 10 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 25; or or comprising or composed of: an amino acid sequence described in SEQ ID NO. 26; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 26 and having a substitution with cysteine at position 155; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 26 and including the amino acid sequence described in SEQ ID NO. 11; or The modified heavy chain invariant region contains the modified CH1, and the modified CH1 is the following amino acid sequence: An amino acid sequence described in SEQ ID NO. 22, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 22 and having a substitution with cysteine at position 152, or an amino acid sequence comprising the amino acid sequence described in SEQ ID NO. 10 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 22; or The amino acid sequence described in SEQ ID NO. 23, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 23 and having a substitution with cysteine at position 155, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 23 and including or composed of the amino acid sequence described in SEQ ID NO. 11; The modified light chain constant region in item (vi) comprises or consists of the following amino acid sequences: the amino acid sequence described in SEQ ID NO. 60, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 60, having substitutions with cysteine at positions 160 and 166 and substitution with tyrosine at position 178, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 60, including the amino acid sequence described in SEQ ID NO. 62; or The modified light chain invariant region in item (vii) comprises or consists of the following amino acid sequences: the amino acid sequence described in SEQ ID NO. 44, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 44 and having a substitution with cysteine at position 166, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 44 and including the amino acid sequence described in SEQ ID NO. 66; The modified heavy chain invariant region is the following amino acid sequence: or comprising or composed of: an amino acid sequence described in SEQ ID NO. 61; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 61 and having a substitution with cysteine at position 375; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 61 and including the amino acid sequence described in SEQ ID NO. 67; or The modified heavy chain invariant region contains the modified Fc region, and the modified Fc region is the following amino acid sequence: The amino acid sequence described in SEQ ID NO. 63, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 63 and having a substitution with cysteine at position 375, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 63 and including or composed of the amino acid sequence described in SEQ ID NO. 67; The modified heavy chain invariant region in item (viii) is the following amino acid sequence: or comprising or composed of: an amino acid sequence described in SEQ ID NO. 64; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 64 and having cysteine substitutions at positions 239 and 375; or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 64 and including the amino acid sequences described in SEQ ID NO. 67 and SEQ ID NO. 68; or The modified heavy chain invariant region contains the modified Fc region, and the modified Fc region is the following amino acid sequence: An antibody or an antigen-binding fragment thereof comprising or composed of the amino acid sequence described in SEQ ID NO. 65, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 65 and having a substitution with cysteine at position 375, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence described in SEQ ID NO. 65 and including the amino acid sequences described in SEQ ID NO. 67 and SEQ ID NO. 68.
  6. In any one of claims 1 to 5, the antigen-binding fragment is an antibody or an antigen-binding fragment thereof selected from Fab, Fab', F(ab')2 or diabody.
  7. In any one of claims 1 to 6, the antibody or its antigen-binding fragment is an antibody or its antigen-binding fragment that specifically binds to an antigen, such as an immune checkpoint molecule like HER2, or a tumor-associated antigen.
  8. In claim 7, the antibody or its antigen-binding fragment that specifically binds to HER2 comprises three heavy chain variable region CDRs and three light chain variable region CDRs of a known antibody that specifically binds to HER2, such as trastuzumab; or the antibody or its antigen-binding fragment comprising a heavy chain variable region and a light chain variable region of a known antibody that specifically binds to HER2, such as trastuzumab.
  9. In paragraph 7, the parent antibody or its antigen-binding fragment is an antibody or its antigen-binding fragment that is trastuzumab or its antigen-binding fragment.
  10. An isolated nucleic acid molecule encoding an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 9.
  11. An expression vector comprising a nucleic acid molecule according to paragraph 10.
  12. A host cell comprising an isolated nucleic acid molecule according to paragraph 10 or an expression vector according to paragraph 11.
  13. A method for producing a cysteine-engineered antibody or an antigen-binding fragment thereof, A step of introducing into a host cell an antibody according to any one of claims 1 to 9, a nucleic acid encoding each chain of the antigen-binding fragment thereof, or an expression vector comprising said nucleic acid(s); and The method includes the step of expressing and assembling an antibody or its antigen-binding fragment in a host cell in vivo; Optionally, the antibody or its antigen-binding fragment is purified, for example, by protein A, in a method.
  14. An immunoconjugate comprising an antibody or an antigen-binding fragment thereof and a payload according to any one of claims 1 to 9.
  15. In paragraph 14, the payload is an immunoconjugate selected from drugs such as small molecule drugs, radionuclides, DNA, RNA, enzymes, or polypeptides; for example, cytotoxic agents, chemotherapy agents, innate immune agents (e.g., Toll-like receptor agonist (TLR) ISAC drugs SBT6050, SBT6290, BDC-1001; STING agonist ISAC drug XMT-2056, Treg cell modulating ISAC drug ADCT-301, etc.), immunomodulators, therapeutic oligonucleotides (siRNA, PMO, etc.), or radionuclides.
  16. In paragraph 14, the immunoconjugate is an immunoconjugate selected from an antibody-drug conjugate (ADC), an immunostimulating antibody conjugate (ISAC), an antibody-oligonucleotide conjugate (AOC), an antibody-polypeptide conjugate (APC), an antibody-radionuclide drug conjugate (RDC), or an antibody degraducer conjugate (ADeC).
  17. In paragraph 16, the immunoconjugate is an antibody-drug conjugate (ADC), and the payload is a topoisomerase I inhibitor such as a camptothecin derivative or camptothecin analog (e.g., DXd); an anti-tubulin agent such as auristatin (MMAF, MMAE); a taxane analog (e.g., epotilone A and B); maytansinoid (DM1, DM4); tubulicin and analogs thereof; a DNA-acting drug such as PBD, etc., the immunoconjugate is a drug moiety selected from the group consisting of these.
  18. In paragraph 16 or 17, the immunoconjugate is an antibody-drug conjugate, and the ADC is an immunoconjugate having an average DAR value of 1-4.
  19. An immunoconjugate in any one of claims 14 to 18, wherein the antibody or its antigen-binding fragment is connected to the payload, for example with or without a linker, by cysteine(s) introduced by the mutation(s).
  20. In claim 19, the linker comprises a functional group capable of reacting with a thiol group of a cysteine residue present in an antibody or its antigen-binding fragment to form a covalent bond, wherein the functional group is selected from, for example, maleimide, iodoacetamide, bromoacetamide, vinylpyridine, disulfide, pyridyl disulfide, haloacetamide, α-haloacetyl, succinimidyl ester, 4-nitrophenyl ester, pentafluorophenyl ester, tetrafluorophenyl ester, active esters, anhydrides, acyl chlorides, sulfonyl chlorides, isocyanates, and isothiocyanates, and preferably maleimide, an immunoconjugate.

Description

Cysteine-manipulated antibodies and immune conjugates The present invention relates to antibodies engineered with reactive cysteine residues, and more specifically, to antibodies having therapeutic or diagnostic applications. The present invention also relates to immunoconjugates comprising engineered antibodies. The present invention further relates to pharmaceutical agents or pharmaceutical compositions comprising antibodies or immunoconjugates, and methods and uses for applying antibodies or immunoconjugates for the treatment of diseases such as tumors. Despite the clinical success of therapeutic antibodies, naked antibodies targeting cell-surface tumor antigens rarely provide sufficient efficacy on their own. To increase the low activity of antibodies, new strategies are focusing on conjugating antibodies to toxic molecules. Plant and bacterial toxins, as well as chemotherapy small molecules, can be excellent candidates because they are highly effective and active even in very small amounts. Therefore, conjugating antibodies to other components to form novel immunoconjugates, such as antibody-drug conjugates (ADCs), has become a new strategy for the development of oncology therapies. Currently, most methods for ADC conjugation are still random conjugations, including cysteine and lysine conjugations, which lead to heterogeneous distribution of ADC products. For example, a cysteine-conjugated ADC with an average DAR value of 4 contains multiple components ranging from DAR0 to DAR8. Even among components with the same DAR value, differences in conjugation at different sites still exist. Meanwhile, these components tend to have different characteristics. For instance, the DAR0 component binds competitively to the target, while the DAR8 component is prone to aggregation due to the presence of multiple conjugated hydrophobic drugs, leading to easier clearance from the body. The different pharmacodynamic and pharmacokinetic properties corresponding to each of these components make PK/PD analysis of heterogeneous ADC mixtures more difficult and require higher manufacturing processes to produce relatively stable batches of ADC products. To obtain a homogeneous ADC product, Genentech attempted to mutate a region of an antibody to cysteine in 2006 (WO2006034488A2) so that the drug could be homogeneously conjugated to this mutated site. The cysteine introduced into the antibody screened by this patent has higher thiol reactivity, and the mutation to cysteine does not affect the antibody's ability to bind to the antigen; furthermore, the ADC has potenter efficacy in vivo than a randomly conjugated ADC. However, this technology still has several limitations. As recent research has shown that conjugating only two drugs is insufficient to achieve the expected efficacy for some diseases or targets, technology for obtaining homogeneous products with higher DAR is also important. Strategies for screening engineered sites disclosed in the prior art are relatively limited. To screen sites with higher thiol reactivity and obtain ADCs with higher DAR, conventional site selection is biased toward sites exposed on the antibody surface that possess higher solvent accessibility. However, a series of other negative effects caused by sites with high solvent accessibility, such as ADC stability after drug conjugation, hydrophilicity and hydrophobicity, PK behavior, and in vivo efficacy, are not considered. Therefore, there is still a need in the industry for new cysteine-manipulation technologies for antibodies, and antibodies obtained based on this can be used to construct ADCs with superior stability and efficacy. The present invention relates to a modified antibody or an antigen-binding fragment thereof having a mutation to cysteine in a hidden site (cryptic site) of an invariant region, thereby imparting greater stability and/or hydrophilicity to an immunoconjugate containing the same. In one embodiment, the cysteine-modified antibody of the present invention generates two or four cysteine-modified antibody molecules due to the dimerization properties of IgG antibodies after one or two amino acids in the light or heavy chain are mutated to cysteine. Thiol groups on cysteine can undergo a nucleophilic reaction with a toxin small molecule having a maleimide linker, thereby enabling the preparation of site-specifically conjugated ADC molecules by conjugating the toxin small molecule to four cystes. The selection of the mutation site is a key aspect of the present invention, and the quality of the site directly determines the characteristics of the ADC. After a small molecule drug is linked to a site with high solvent accessibility, the potential for thiol exchange between the small molecule and the protein in plasma increases due to the relatively exposed environment of the small molecule drug, thereby weakening efficacy and making it more susceptible to toxicity and side effects. Furthermore, the more exposed small molecule also