Anti-Fas ligand (FasL) antibodies in the treatment of SJS / TEN disease

Abstract

The present invention relates to the use of human Fas ligand (FasL; also known as CD95L or Apo1L) antagonists, more particularly to the use of antibodies specifically directed against soluble FasL for the prevention and / or treatment of SJS and / or TEN skin diseases.

Description

Technical Field

[0001] The present invention relates to the use of antagonists of human Fas ligand (FasL; also referred to as CD95L or CD178 or Apo1L), particularly antagonists of soluble human Fas ligand (sFasL). More particularly, the present invention relates to the use of antibodies against FasL, particularly sFasL, for the prevention and / or treatment of Stevens-Johnson syndrome (SJS) and / or toxic epidermal necrolysis (TEN) skin diseases (hereinafter, "SJS / TEN").

Background Art

[0002] Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) are severe skin pathologies caused by adverse reactions to certain drugs or medical substances. Both skin pathologies are characterized by extensive erythema, epidermal necrosis, blister formation, and detachment of skin sections. Almost all SJS and TEN patients also have mucosal lesions in the eyes, mouth, and genitalia. These syndromes are considered the same disease with different severity spectra. SJS represents the lower severity disease spectrum and is defined by detachment of less than 10% of the body surface area. TEN represents a more severe disease spectrum with skin detachment involving more than 30% of the body surface area. The overlap of SJS / TEN is defined as 10 - 30% skin detachment.

[0003] The extensive skin detachment in SJS / TEN causes significant morbidity and mortality. The reported mortality rates of SJS / TEN patients are estimated to be 4.8% to 14.8% respectively. Therefore, SJS / TEN is a dermatological emergency, and early detection and successful treatment are considered life-saving.

[0004] SJS / TEN is a distressing condition, but there is no standard treatment or approved medication for these skin conditions. Patients are typically treated urgently with several medications, including cyclosporine, steroids, anti-TNF, IVIg, or plasmapheresis (Chang HH et al., Biomedicines, 2022). However, none of these medications have been shown to have a beneficial effect in controlled studies. Most patients receive only supportive care.

[0005] The clinical manifestation of skin peeling in SJS / TEN is primarily due to the widespread induction of keratinocyte apoptosis and necroptosis. However, the mechanism by which these processes are enhanced in SJS / TEN remains unclear.

[0006] Fas (or FasR) is a member of the TNF receptor superfamily and, upon binding to its ligand (FasL), induces apoptosis in many cell lines (Sharma et al, 2000). The Fas-FasL interaction is involved in the pathological mechanisms of several immunoinflammatory and infectious conditions. For example, the consequences of the Fas-FasL pathway have been shown to be a key mediator of keratinocyte apoptosis and squamous cell lysis in pemphigus (Lotti et al., 2018).

[0007] FasL exists in both membrane-bound (mFasL) and soluble (sFasL) forms. Soluble Fas ligand (sFasL) is generated and released when the extracellular domain of mFasL (mFasL), expressed on the plasma membrane, is hydrolytically cleaved by zinc-regulated matrix metalloproteinases (MMPs) and the metalloproteinase ADAM10. Soluble sFasL exhibits both apoptotic and non-apoptotic activity.

[0008] The Fas / Fas ligand system plays a crucial role in maintaining homeostasis of immune cells, preventing autoimmunity, and facilitating cancer progression. To maintain immune system homeostasis, membrane-bound mFasL expressed on cytotoxic lymphocytes binds to the Fas receptor (Fas), inducing the lysis of infected cells, hyperproliferative T lymphocytes, and cancer cells via apoptosis. Cell-cell interactions between activated immune cells possessing membrane-bound FasL and target cells possessing membrane-bound Fas are essential for maintaining immune system homeostasis.

[0009] In addition to their role in the physiological regulation of the immune system, Fas-FasL interactions and the resulting signaling pathways have been shown to be involved in the development and progression of various malignancies and immunoinflammatory and infectious conditions.

[0010] Furthermore, recent research suggests that the Fas / Fas ligand (FasL) system also plays a crucial role in the development of SJS / TEN, as evidenced by the following: (i) Serum FasL levels (i.e., soluble FasL, sFasL) in SJS / TEN patients are elevated, especially before the onset of skin peeling and / or mucosal lesions (Viard et al., 1998; Abe et al., 2003; Chang et al., 2004; Murata et al., 2008); (ii) Peripheral blood mononuclear cells (PBMCs) stimulated with serum from SJS / TEN patients and / or drugs that cause SJS / TEN disease release FasL (Abe et al. 2003); (iii) Serum derived from SJS / TEN patients induces keratinocyte cell death in culture (Abe et al.); and (iv) In SJS / TEN, FasL is released from keratinocytes, causing cell death in adjacent keratinocytes (Abe et al. 2015). This has been proven based on the following.

[0011] Therefore, the development of FasL-targeting inhibitors that do not affect the surveillance and homeostatic pathways controlled by the Fas / FasL system is extremely important.

[0012] Antagonists of the Fas receptor (or FasR) are known in the art. WO 2010 / 102792 A2 discloses binding members to human Fas (or FasR), particularly antibodies against human Fas, for use in inhibiting Fas-mediated apoptosis and treating Stevens-Johnson syndrome (SJS) and related diseases, including toxic epidermal necrolysis (TEN). However, these Fas inhibitors cannot distinguish between the sFasL and mFasL-mediated pathways.

[0013] Furthermore, Fas receptor antagonists can cause undesirable effects due to inhibition of the Fas / FasL pathway and its role in the physiological regulation of the immune system. This risk includes causing and / or exacerbating dysfunction of immunological homeostasis, which can lead to lymphoproliferative disorders, autoimmune disorders, and / or the development and / or progression of cancer. In addition, dysfunction of the Fas / FasL pathway due to inhibition and / or blockade of the Fas receptor can impair immune-privileged sites such as the eyes (ocular immune privilege), testes, placenta, and nervous system.

[0014] Viard et al. (Science, vol. 282, no. 5388, 1998, pp. 490-493) have disclosed an effective treatment of TEN with IVIG. IVIG exhibits Fas inhibitory activity due to the presence of naturally occurring anti-Fas antibodies contained in IVIG formulations. However, IVIG is a drug containing numerous antibodies with different specificities. Furthermore, the quality of IVIG is inconsistent from batch to batch. [Overview of the Initiative] [Problems that the invention aims to solve]

[0015] The object of the present invention is to provide therapeutic agents for the prevention and treatment of SJS / TEN. In light of the above evidence regarding the important role of FasL in SJS / TEN, the development of novel agents that block FasL, particularly selectively block sFasL in patient serum, could prevent keratinocyte apoptosis and subsequent cell exfoliation and squamous cell lysis, thereby preventing the formation of severe skin lesions in SJS / TEN.

[0016] A further object of the present invention is to provide a drug that can selectively inhibit the interaction between Fas and soluble FasL (sFasL) for use in the prevention and treatment of SJS / TEN by selectively inhibiting sFasL released into the serum of SJS / TEN patients. [Means for solving the problem]

[0017] Summary of the Invention This disclosure provides monoclonal antibodies or antigen-binding fragments specific to human Fas ligand protein (FasL), particularly sFasL, as activators for treating patients with Stevens-Johnson syndrome (SJS) and / or toxic epidermal necrolysis (TEN) (SJS / TEN). The antibodies are characterized by the amino acid sequences of the heavy chain variable region (VH) and the variable region (CDR) of at least one light chain variable region (VL), as defined in Sequence IDs 1-9 disclosed herein. The use of these antibodies is effective for treating SJS / TEN due to their high binding affinity to human FasL, particularly soluble human FasL.

[0018] The therapeutic utility of the antibodies or antigen-binding fragments disclosed herein is based on a combination of high binding affinity and selectivity to soluble human FasL. In particular, the antibodies or antigen-binding fragments disclosed herein specifically target sFasL, the pathological form in the patient's serum, while not effectively binding to mFasL, the homeostatic form.

[0019] Certain embodiments of the present invention relate to a monoclonal antibody or an antigen-binding fragment thereof comprising a VH region having complementarity-determining regions CDR H1, CDR H2 and CDR H3 shown in SEQ ID NOs: 1, 3 and 5, and a VL region having complementarity-determining regions CDR L1, CDR L2 and CDR L3 shown in SEQ ID NOs: 7-9. In a more preferred embodiment of the present invention, the antibody has an IgG heavy chain constant region, preferably an IgG1 or IgG4 heavy chain constant region.

[0020] A further aspect of the present invention relates to a nucleic acid molecule encoding the monoclonal antibody or antigen fragment thereof disclosed herein for use in the treatment of SJS / TEN.

[0021] Yet another aspect of the present invention relates to a pharmaceutical composition comprising the antibody or antigen-binding fragment or nucleic acid molecule disclosed herein for use in the treatment of SJS / TEN, together with one or more pharmaceutically acceptable carriers.

Mode for Carrying Out the Invention

[0022] Embodiments of the present invention Hereinafter, specific embodiments of the present invention are disclosed as follows: 1. A monoclonal antibody or an antigen-binding fragment thereof specific for human Fas ligand protein (FasL) comprising at least one heavy chain variable (VH) region and at least one light chain variable (VL) region for use in a method for the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), wherein the antibody or antigen-binding fragment is as follows: (i) The following heavy chain CDR H1, CDR H2 and CDR H3: (a1) CDR H1: Arg His Gly Ile Thr (SEQ ID NO: 1) or (a2) CDR H1: Ser His Gly Ile Ser (SEQ ID NO: 2), (b1) CDR H2: Trp Ile Asn Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Val Gln Gly (SEQ ID NO: 3) or (b2) CDR H2: Trp Ile Asn Ala Tyr Ser Gly Asn Thr Asn Tyr Ala Gln Lys Leu Gln Gly (SEQ ID NO: 4), (c1) CDR H3: Glu Thr Met Val Arg Gly Val Pro Leu Asp Tyr (SEQ ID NO: 5), or (c2) CDR H3: Glu Thr Met Val Arg Gly Val Pro Cys Asp Tyr (SEQ ID NO: 6) having a VH region with complementarity determining regions (CDRs) of the following light chain CDR L1, CDR L2 and CDR L3: (a3) CDR L1: Arg Ala Ser Gln Ser Val Ser Tyr Leu Ala (SEQ ID NO: 7), (b3) CDR L2: Gly Ala Ser Ser Arg Ala Thr (SEQ ID NO: 8), (c3) CDR L3: Gln Gln Tyr Gly Ser Ser Pro Trp Thr (SEQ ID NO: 9) complementarity determining regions (CDRs) of an antibody or antigen-binding fragment thereof; or (ii) an antibody or antigen-binding fragment that competes with the antibody or antigen-binding fragment of (i) in binding to human Fas ligand protein (FasL); A monoclonal antibody or antigen-binding fragment thereof selected from 2. A monoclonal antibody or antigen-binding fragment thereof according to embodiment 1 for use according to embodiment 1, comprising at least one heavy chain variable (VH) region and at least one light chain variable (VL) region, wherein the antibody or antigen-binding fragment has the following heavy chain CDR H1, CDR H2 and CDR H3: (a1) CDR H1:Arg His Gly Ile Thr (Sequence ID 1), (b1) CDR H2: Trp Ile Asn Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Val Gln Gly (Sequence ID 3), (c1) CDR H3:Glu Thr Met Val Arg Gly Val Pro Leu Asp Tyr (SEQ ID NO: 5) VH region having complementarity determination region (CDR); and The following light chains CDR L1, CDR L2, and CDR L3: (a3) CDR L1: Arg Ala Ser Gln Ser Val Ser Tyr Leu Ala (SEQ ID NO: 7), (b3) CDR L2: Gly Ala Ser Ser Arg Ala Thr (Sequence ID 8), (c3) CDR L3: Gln Gln Tyr Gly Ser Pro Trp Thr (Sequence ID 9) Complementarity Determination Region (CDR); A monoclonal antibody or its antigen-binding fragment, selected from antibodies or antigen-binding fragments containing the above. 3. The VL region of the antibody has the following amino acid sequence: EIVLTQSPGTLSLSPGERATLSC RASQSVSSSYLA WYQQKPGQAPRLLIY GASSRAT GIPDRFSGSGSGTDFTLTISRLEPEDFAVYYC QQYGSSPWT FGQGTKVEIKRTVAAPSVFIFP(Sequence ID 10) including and The VH region of the antibody has the following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYIFI RHGIT WVRQAPGQGLEWMG WINAYNGNTNYAQKVQG RVTMTTDKSTSTAYMELRSLRSDDAAVYYCAR ETMVRGVPLDY WGQGTLVTVSSASTKGPSVFPLA (Sequence ID 11), or QVQLVQSGAEVKKPGASVKVSCKASGYIFI SHGIS WVRQAPGQGLEWMG WINAYSGNTNYAQKLQG RVTMTTDRSTSTAYMELRSLRSDDTAVYYCAR ETMVRGVPCDY WGQGTLVTVSSASTKGPSVFPLA(Sequence ID 12) An antibody or antigen-binding fragment thereof according to Embodiment 1 or 2 for use in Embodiment 1, including the antibody or antigen-binding fragment thereof. 4. The VL region of the antibody has the following amino acid sequence: EIVLTQSPGTLSLSPGERATLSC RASQSVSSSYLA WYQQKPGQAPRLLIY GASSRAT GIPDRFSGSGSGTDFTLTISRLEPEDFAVYYC QQYGSSPWT FGQGTKVEIK (Sequence ID 13), including and The VH region of the antibody has the following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYIFI RHGIT WVRQAPGQGLEWMG WINAYNGNTNYAQKVQG RVTMTTDKSTSTAYMELRSLRSDDAAVYYCAR ETMVRGVPLDY WGQGTLVTVSS (Sequence ID 14), or QVQLVQSGAEVKKPGASVKVSCKASGYIFI SHGIS WVRQAPGQGLEWMG WINAYSGNTNYAQKLQG RVTMTTDRSTSTAYMELRSLRSDDTAVYYCAR ETMVRGVPCDY WGQGTLVTVSS (Sequence ID 15) An antibody or antigen-binding fragment according to any one of Embodiments 1 to 3 for use in Embodiment 1, including the antibody or antigen-binding fragment according to Embodiment 1. 5. An antibody or antigen-binding fragment according to any one of Embodiments 1 to 4 for use in Embodiment 1, which recognizes the same epitope on human FasL as the antibody of Embodiment 1(i) or 2(i). 6. The antibody or antigen-binding fragment according to any one of Embodiments 1 to 5 for use in Embodiment 1, wherein the antibody is selected from a partially or fully human antibody, a chimeric antibody and / or a humanized antibody, and its antigen-binding fragment is selected from Fab, Fab' and / or F(ab')2 and / or a single-chain Fv fragment. 7. The antibody or antigen-binding fragment according to any one of Embodiments 1 to 6 for use in Embodiment 1, wherein the antibody has an IgG heavy chain constant region, preferably an IgG1 or IgG4 heavy chain constant region. 8. A nucleic acid molecule encoding a monoclonal antibody or its antigen fragment according to any one of Embodiments 1 to 7, for use in Embodiment 1. 9. A nucleic acid molecule according to Embodiment 8 for use in Embodiment 1, which is a DNA vector or RNA molecule. 10. An antibody or antigen-binding fragment thereof according to any one of Embodiments 1 to 7, or a nucleic acid molecule according to any one of Embodiments 8 to 9, for use in monotherapy in Embodiment 1. 11. For use in Embodiment 1, in combination with at least one further active ingredient effective against toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), an antibody or antigen-binding fragment thereof according to any one of Embodiments 1 to 7, or a nucleic acid molecule according to any one of Embodiments 8 to 9. 12. For use in Embodiment 11, an antibody or antigen-binding fragment thereof according to any one of Embodiments 1 to 7, or a nucleic acid molecule according to any one of Embodiments 8 to 9, wherein the further active ingredient is selected from at least one of a steroid, cyclosporine, IVIg, a TNF inhibitor, and / or plasmapheresis. 13. An antibody or antigen-binding fragment thereof according to any one of Embodiments 1 to 7, or a nucleic acid molecule according to any one of Embodiments 8 to 9, for use in human therapeutics. 14. A pharmaceutical composition comprising, together with one or more pharmaceutically acceptable carriers for use in Embodiment 1, an antibody or antigen-binding fragment thereof as described in any one of Embodiments 1 to 7, or a nucleic acid molecule as described in any one of Embodiments 8 to 9. 15. An antibody or antigen-binding fragment thereof according to any one of Embodiments 1 to 7, or a nucleic acid molecule according to any one of Embodiments 8 to 9, or a pharmaceutical composition according to Embodiment 14, which is administered systemically and / or locally.

[0023] Detailed explanation The present invention relates to the treatment of SJS / TEN disease associated with keratinocyte apoptosis by administration of a FasL antagonist. In a preferred embodiment, the present invention relates to the treatment of SJS / TEN disease in human subjects who additionally suffer from immunological homeostasis dysfunction and / or have an increased risk of developing immunological homeostasis dysfunction.

[0024] The term “immunological homeostasis” specifically refers to the proper and / or balanced functioning of the immune system being treated. The term “dysfunction” specifically refers to a disturbance in the immune system, for example, the functionality of immune cells with membrane-bound FasL, and more specifically, a disturbance to their ability to interact with target cells that have Fas receptors.

[0025] Generally, FasL antagonists can be selected from anti-FasL antibodies or their antigen-binding fragments, particularly humanized or human anti-FasL antibodies or their antigen-binding fragments, nucleic acid effector molecules of Fas expression such as antisense molecules or RNA interference molecules such as siRNA molecules, soluble Fas receptor molecules, antagonistic FasL muteins, and low molecular weight chemical compounds that inhibit Fas-FasL interactions. FasL antagonists prevent keratinocyte apoptosis and subsequent cell-cell detachment (acintholysis).

[0026] In a highly preferred embodiment, the anti-FasL antibody may be selected from an antibody or antigen-binding fragment that exhibits selective high-affinity binding to soluble FasL while substantially not inhibiting the binding of mFasL to Fas. Selective binding to sFasL allows for targeted inhibition of sFasL without substantially inhibiting mFasL. Thus, the interaction between mFasL and the Fas receptor remains substantially unaffected, thereby avoiding undesirable effects on immunological homeostasis. In certain embodiments, undesirable effects on immunological homeostasis include inhibition of the function of activated immune cells possessing mFasL.

[0027] Accordingly, the present invention relates to the use of at least one anti-FasL compound that can inhibit the biological effects of FasL, particularly sFasL. As used herein, the expressions “inhibit the biological effects of FasL” or “inhibit the biological effects of sFasL” relate to a compound that can completely or at least substantially inhibit or neutralize the biological effects of FasL or sFasL, particularly sFasL. For example, the inhibitory or neutralizing effect may be based on suppressing the binding of FasL, particularly sFasL, to its native receptor and thereby suppressing the signaling it causes. This can be achieved, for example, by using an antibody that binds to FasL / sFasL itself, or a soluble receptor that mimics Fas / sFasL, or an antagonistic FasL / sFasL mutein, thus blocking the binding of FasL / sFasL to its cellular receptor. Alternatively, the Fas / FasL system can be similarly blocked by interfering with the expression of Fas or FasL / sFasL by siRNA.

[0028] One aspect of the present invention relates to the use of an anti-FasL antibody or its active fragment as a therapeutic agent in the prevention and / or treatment of SJS / TEN. In a preferred embodiment, the present invention relates to the use of an anti-sFasL antibody or its active fragment as a therapeutic agent in the prevention and / or treatment of SJS / TEN.

[0029] In the context of the present invention, the term "prevention of SJS / TEN" means treating the disease at the time of diagnosis, for example, at an early stage, in order to prevent progression of the disease to a more severe severity spectrum. The term "treatment of SJS / TEN" means managing the disease, preferably to the point of curing it, in order to alleviate and / or improve the symptoms of SJS / TEN disease.

[0030] The development of active agents, particularly antibodies, that specifically inhibit sFasL will enable targeted therapeutic approaches to SJS and TEN. Furthermore, this therapeutic approach minimizes the risk of inducing or exacerbating dysfunction of immunological homeostasis related to the Fas / mFasL system.

[0031] The antibody is preferably a chimeric antibody, a humanized antibody, or a human anti-FasL antibody, preferably an anti-sFasL antibody. Furthermore, the antibody may be monovalent or polyvalent and may include modifications such as different glycosylation patterns or modifications of the Fc region to alter antibody-dependent cytotoxicity (ADCC) or complement-dependent cytotoxicity (CDC). The antigen-binding fragment or derivative of the anti-FasL antibody, preferably an anti-sFasL antibody, of the present invention may be a recombinant single-chain antibody or a single-chain variable fragment. In a very preferred embodiment of the present invention, the antibody is a human FasL antibody, more preferably a human anti-sFasL antibody. Optionally, the antibody may be conjugated to an effector molecule, such as a cell proliferation inhibitor, cytotoxic, and / or radioactive compound.

[0032] In the first aspect, the present invention relates to a soluble human Fas ligand protein (sFasL) specific monoclonal antibody or antigen-binding fragment, preferably comprising at least one heavy chain variable (VH) region and at least one light chain variable (VL) region, for use in methods for the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), wherein the antibody or antigen-binding fragment comprises the following heavy chains CDR H1, CDR H2 and CDR H3: (a1) CDR H1:Arg His Gly Ile Thr (Sequence ID 1) or (a2) CDR H1:Ser His Gly Ile Ser (Sequence ID 2), (b1) CDR H2: Trp Ile Asn Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Val Gln Gly (SEQ ID NO: 3), or (b2) CDR H2:Trp Ile Asn Ala Tyr Ser Gly Asn Thr Asn Tyr Ala Gln Lys Leu Gln Gly (SEQ ID NO: 4), and (c1) CDR H3:Glu Thr Met Val Arg Gly Val Pro Leu Asp Tyr (SEQ ID NO: 5), or (c2) CDR H3:Glu Thr Met Val Arg Gly Val Pro Cys Asp Tyr (SEQ ID NO: 6) VH region having complementarity determination region (CDR), and The following light chains CDR L1, CDR L2, and CDR L3: (a3) CDR L1: Arg Ala Ser Gln Ser Val Ser Tyr Leu Ala (SEQ ID NO: 7), (b3) CDR L2:Gly Ala Ser Ser Arg Ala Thr (SEQ ID NO: 8), and (c3) CDR L3: Gln Gln Tyr Gly Ser Ser Pro Trp Thr (Sequence ID 9) It includes the complementarity determination region (CDR).

[0033] In a preferred embodiment, the present invention relates to the following heavy chain CDR H1, CDR H2, and CDR H3: (a1) CDR H1:Arg His Gly Ile Thr (Sequence ID 1), (b1) CDR H2: Trp Ile Asn Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Val Gln Gly (Sequence ID 3), (c1) CDR H3:Glu Thr Met Val Arg Gly Val Pro Leu Asp Tyr (SEQ ID NO: 5) At least one heavy chain variable (VH) region having a complementarity determination region (CDR), and The following light chains CDR L1, CDR L2, and CDR L3: (a3) CDR L1: Arg Ala Ser Gln Ser Val Ser Tyr Leu Ala (SEQ ID NO: 7), (b3) CDR L2:Gly Ala Ser Ser Arg Ala Thr (SEQ ID NO: 8), and (c3) CDR L3: Gln Gln Tyr Gly Ser Ser Pro Trp Thr (Sequence ID 9) This relates to a monoclonal antibody or antigen-binding fragment specific to human Fas ligand protein (FasL) for use in methods for the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), comprising at least one light chain variable (VL) region having a complementarity-determining region (CDR).

[0034] Furthermore, the present invention relates to a monoclonal antibody or antigen-binding fragment that competes with the monoclonal antibodies or antigen-binding fragments disclosed above for selective inhibition of binding to human Fas ligand protein (FasL), particularly soluble human Fas ligand protein (sFasL). Preferably, the antibody competes with a monoclonal antibody having heavy chain CDR sequences H1-H3 containing the amino acid sequences of SEQ ID NOs. 1-5 and light chain CDR sequences L1-L3 containing the amino acid sequences of SEQ ID NOs. 7-9. In a very preferred embodiment, the antibody competes with a monoclonal antibody having heavy chain CDR sequences H1-H3 of SEQ ID NOs. 1, 3, and 5 and light chain CDR sequences L1-L3 of SEQ ID NOs. 7-9.

[0035] In certain embodiments, the competing antibody of the present invention binds to the same or overlapping epitopes on human FasL, particularly human sFasL, as the monoclonal antibody having a CDR as defined in SEQ ID NOs. Competition can be determined by standard assays in the art that can quantify the binding affinity of the antibody to the binding protein, particularly human FasL, relative and absolute to a given reference antibody.

[0036] The quantitative binding of FasL antibodies to FasL proteins can be determined using one of several surface plasmon resonance (SPR) measurement platforms, such as Biacore and Forte Octet. The recombinant target protein, FasL (e.g., Acro Bio FAL-H5241), is immobilized on a capture chip, and the FasL antibody is flowed over the chip while changes in intermolecular interactions are recorded in real time. The on-rate (ka; M) is then measured. -1 s-1 ), off-rate (kd; s -1 The ) can be calculated by adding or removing FasL from the flow. The binding affinity index (KD, μM) can be obtained by dividing the off-rate by the on-rate. A KD value of less than 10 μM is usually required for therapeutic use. KD values ​​between 10 nM and 1 pM are optimally used.

[0037] In certain embodiments of the present invention, at least one amino acid in the above-mentioned CDR1, CDR2, or CDR3 amino acid sequence of the VH chain and / or VL chain is replaced with another amino acid while maintaining the structural integrity and epitope binding ability of the antibody. These replacements may be conservative (i.e., by similar amino acids) or non-conservative.

[0038] In particular, at least one amino acid in the CDR1, CDR2, or CDR3 sequences of VH and VL may be a conserved amino acid substitution, i.e., a substitution of an amino acid with another amino acid having similar biochemical properties, for example, an aliphatic amino acid, such as Gly, Ala, Val, Leu, or Ile; a basic amino acid, such as His, Lys, or Arg, with another basic amino acid or Met; an acidic amino acid or its amide, such as Asp, Glu, Asn, or Gln, with another acidic amino acid or its amide; or an aromatic amino acid, such as Phe, Tyr, or Trp, with another aromatic amino acid. In certain preferred embodiments of the present invention, amino acids 1, 2, 3, 4, or 5 of SEQ ID NOs: 1-9 are substituted with another conserved or non-conserved amino acid.

[0039] In a preferred embodiment, the present invention relates to the use of an anti-FasL human antibody or its antigen-binding moiety comprising a light chain variable region and / or a heavy chain variable region (SEQ ID NOs. 2 and SEQ ID NOs. 10 or 18, respectively, of US 7,262,277, the contents of which are incorporated herein by reference. Surprisingly, these antibodies disclosed in US 7,262,277 have been found to have high target affinity and specificity for soluble FasL, but do not bind to or substantially bind to membrane-bound FasL (mFasL).

[0040] This finding provides novel therapeutic applications for these antibodies in more targeted therapies for SJS and / or TEN. In preferred embodiments, the antibodies are particularly useful for administration to subjects with SJS / TEN who have further immunological homeostasis dysfunction and / or an increased risk of developing immunological homeostasis dysfunction. In more specific embodiments, the disclosure provides the use of the antibodies in subjects with SJS / TEN who have further immune system-related dysfunction or an increased risk of developing immune system-related dysfunction, such as immunoinflammatory disorders, autoimmune disorders and / or cancer.

[0041] Therefore, in a more preferred embodiment of the present invention, an anti-FasL antibody, particularly an anti-sFasL antibody, for use in the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS) has the following amino acid sequence: EIVLTQSPGTLSLSPGERATLSC RASQSVSSSYLA WYQQKPGQAPRLLIY GASSRAT GIPDRFSGSGSGTDFTLTISRLEPEDFAVYYC QQYGSSPWT FGQGTKVEIKRTVAAPSVFIFP(Sequence ID 10) A light chain variable region (VL) comprising a polypeptide having, and The following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYIFI RHGIT WVRQAPGQGLEWMG WINAYNGNTNYAQKVQG RVTMTTDKSTSTAYMELRSLRSDDAAVYYCAR ETMVRGVPLDY WGQGTLVTVSSASTKGPSVFPLA (Sequence ID 11), or QVQLVQSGAEVKKPGASVKVSCKASGYIFI SHGIS WVRQAPGQGLEWMG WINAYSGNTNYAQKLQG RVTMTTDRSTSTAYMELRSLRSDDTAVYYCAR ETMVRGVPCDY WGQGTLVTVSSASTKGPSVFPLA(Sequence ID 12) It contains a heavy chain variable region (VH) comprising a polypeptide having [a specific characteristic].

[0042] In a more preferred embodiment, the present disclosure relates to an anti-FasL antibody, particularly an anti-sFasL antibody or an antibody-conjugated fragment thereof, for use in the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), wherein the light chain variable region (VL) of the antibody or the antibody-conjugated fragment comprises the following amino acid sequence: EIVLTQSPGTLSLSPGERATLSC RASQSVSSSYLA WYQQKPGQAPRLLIY GASSRAT GIPDRFSGSGSGTDFTLTISRLEPEDFAVYYC QQYGSSPWT FGQGTKVEIK(Sequence ID 13) comprising a polypeptide having, The heavy chain variable region (VH) of the antibody or its antibody-binding fragment has the following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYIFI RHGIT WVRQAPGQGLEWMG WINAYNGNTNYAQKVQG RVTMTTDKSTSTAYMELRSLRSDDAAVYYCAR ETMVRGVPLDY WGQGTLVTVSS (Sequence ID 14), or QVQLVQSGAEVKKPGASVKVSCKASGYIFI SHGIS WVRQAPGQGLEWMG WINAYSGNTNYAQKLQG RVTMTTDRSTSTAYMELRSLRSDDTAVYYCAR ETMVRGVPCDY WGQGTLVTVSS (Sequence ID 15) Contains polypeptides having

[0043] The amino acid sequences of SEQ ID NOs: 13, 14, and 15 differ from those of SEQ ID NOs: 10, 11, and 12 only in that the last 12 amino acids of the latter are no longer assigned to the VH or VL region, but are instead assigned to their respective constant regions.

[0044] In a particularly preferred embodiment, the anti-FasL antibody of the present disclosure, in particular the anti-sFasL antibody, comprises a light chain variable region (VL) containing a polypeptide having the amino acid sequence shown in SEQ ID NO: 10 or SEQ ID NO: 13, and a heavy chain variable region (VH) containing a polypeptide having the amino acid sequence shown in SEQ ID NO: 11 or SEQ ID NO: 14.

[0045] In a preferred embodiment, the anti-FasL antibody, particularly the anti-sFasL antibody, or its antigen-binding fragment may include a VL amino acid sequence having at least 85%, at least 90%, at least 95%, or at least 99% identity with SEQ ID NO: 10 or SEQ ID NO: 13, and a VH amino acid sequence having at least 85%, at least 90%, at least 95%, or at least 99% identity with SEQ ID NO: 11 or 12, or with SEQ ID NO: 14 or 15, respectively, thereby maintaining high binding affinity and selectivity for FasL, particularly sFasL.

[0046] "Percent (%) amino acid sequence identity" for a peptide or polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical to amino acid residues in a particular peptide or polypeptide sequence, after the sequences have been aligned and gaps introduced as necessary to maximize the percentage sequence identity. Alignment for determining percentage amino acid sequence identity can be achieved in various ways by those skilled in the art, such as using readily available computer software like BLAST.

[0047] The antibodies of the present invention for use in the prevention and / or treatment of SJS / TEN can be selected from partially or completely human antibodies, chimeric antibodies and / or humanized antibodies. Preferably, the antibody is a human antibody. Furthermore, the antibodies of the present invention may be monospecific and / or bispecific. Alternatively, the antibody may be polyvalent and multispecific. Furthermore, the antigen-binding fragment of the antibody of the present invention may be selected from Fab, Fab' and / or F(ab')2 and / or single-chain Fv fragments.

[0048] The antibody may be of any suitable class. The term “class” refers to the type of constant domain or constant region in its heavy chain. In this specification, “constant domain” or “constant region” refers to the sum of the domains of an antibody other than the variable region. The constant region does not directly participate in antigen binding but exhibits various effector functions. The antibodies of this disclosure may be of any of the five major classes of antibodies: IgA, IgD, IgE, IgG, and IgM, or their subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant domains corresponding to different classes of immunoglobulin are called α, δ, ε, γ, and μ, respectively. According to this disclosure, antibodies or fragments of the IgG, IgA, and IgM classes are particularly preferred.

[0049] In a very preferred embodiment of the present invention, the antibody has an IgG heavy chain constant region, preferably an IgG1 or IgG4 heavy chain constant region. In a further preferred embodiment of the present invention, the antibody has a kappa light chain. In the most preferred embodiment of the present invention, the antibody has an IgG4 heavy chain constant region.

[0050] Therefore, the most preferred antibody of the present invention is a monoclonal IgG4 anti-FasL antibody, preferably a monoclonal IgG4 anti-sFasL antibody or its antigen-binding fragment comprising a VH region having complementarity-determining regions CDR H1, CDR H2, and CDR H3 as shown in SEQ ID NOs. 1, 3, and 5 above, and a VL region having complementarity-determining regions CDR L1, CDR L2, and CDR L3 as shown in SEQ ID NOs. 7 to 9 above. In a more preferred embodiment of the present invention, the antibody comprises a light chain variable region (VL) containing a polypeptide having the sequence shown in SEQ ID NOs. 10 or 13 above, and a heavy chain variable region (VH) containing a polypeptide having the sequence shown in SEQ ID NOs. 11 or 14 above.

[0051] In a more preferred embodiment of the present invention, the above-mentioned anti-FasL antibody, preferably anti-sFasL antibody, has the following amino acid sequence: PSDEQLKS GTASVVCLLN NFYPREAKVQ WKVDNALQSG NSQESVTEQD SKDSTYSLSS TLTLSKADYE KHKVYACEVT HQGLSSPVTK SFNRGEC (Sequence ID 16) It includes an IgG4 heavy chain constant region containing a light chain constant region (LC) having a polypeptide having, or The following amino acid sequence: RTVAAPSVFI FPPSDEQLKS GTASVVCLLN NFYPREAKVQ WKVDNALQSG NSQESVTEQD SKDSTYSLSS TLTLSKADYE KHKVYACEVT HQGLSSPVTK SFNRGEC (Sequence ID 18) It comprises a light chain constant region (LC) having a polypeptide having and The following amino acid sequence: PCSRSTSE STAALGCLVK DYFPEPVTVS WNSGALTSGV HTFPAVLQSS GLYSLSSVVT VPSSSLGTKT YTCNVDHKPS NTKVDKRVES KYGPPCPPCP APEFLGGPSV FLFPPKPKDT LMISRTPEVT CVVVDVSQED PEVQFNWYVD GVEVHNAKTK PREEQFNSTY RVVSVLTVLH QDWLNGKEYK CKVSNKGLPS SIEKTISKAK GQPREPQVYT LPPSQEEMTK NQVSLTCLVK GFYPSDIAVE WESNGQPENN YKTTPPVLDS DGSFFLYSRL TVDKSRWQEG NVFSCSVMHE ALHNHYTQKS LSLSLGK (Sequence number 17) A heavy chain constant region (HC) containing a polypeptide having, or The following amino acid sequence: ASTKGPSVFP LAPCSRSTSE STAALGCLVK DYFPEPVTVS WNSGALTSGV HTFPAVLQSS GLYSLSSVVT VPSSSLGTKT YTCNVDHKPS NTKVDKRVES KYGPPCPPCP APEFLGGPSV FLFPPKPKDT LMISRTPEVT CVVVDVSQED PEVQFNWYVD GVEVHNAKTK PREEQFNSTY RVVSVLTVLH QDWLNGKEYK CKVSNKGLPS SIEKTISKAK GQPREPQVYT LPPSQEEMTK NQVSLTCLVK GFYPSDIAVE WESNGQPENN YKTTPPVLDS DGSFFLYSRL TVDKSRWQEG NVFSCSVMHE ALHNHYTQKS LSLSLGK (Sequence number 19) It contains a heavy chain constant region (HC) comprising a polypeptide having [a specific characteristic].

[0052] In further embodiments, the present invention relates to the use of anti-hFas ligand human antibodies or their antigen-binding fragments produced by hybridoma cells of accession number ATCC PTA-4017, described as antibodies 3E1 and 4G11 in US 7,262,277, respectively, and / or the use of anti-hFas ligand human antibodies or their antigen-binding fragments produced by hybridoma cells of accession number ATCC PTA-4018, for the prevention and / or treatment of SJS / TEN. In the most preferred embodiments of the present invention, the use of anti-hFas ligand human antibody 3E1 (produced by hybridoma cells of accession number ATCC PTA-4017) or its antigen-binding fragment for the prevention and / or treatment of SJS / TEN.

[0053] The hybridoma cells with accession numbers ATCC PTA-4017 and ATCC PTA-4018 were deposited on January 29, 2002, at the American Type Culture Collection, 10801 University Boulevard, Manassas, Virginia 20110-2209 (USA).

[0054] In a more ultimately preferred aspect, the present invention refers to the use of a monoclonal antibody or its antigen-binding fragment that recognizes the same human FasL epitope as the antibody described above, preferably the same human sFasL epitope.

[0055] The antibodies or antigen-binding fragments of this disclosure bind specifically and with high affinity to soluble human FasL (sFasL). Furthermore, the antibodies or antigen-binding fragments of this disclosure do not effectively bind to membrane-bound FasL. The binding properties of these antibodies enable selective targeting of sFasL without the undesirable side effects caused by binding to mFasL. In particular, administration of the antibodies of this invention may lead to a reduction in the risk of inducing physiological and / or pathological processes mediated and / or induced by inhibition of the mFasL pathway.

[0056] As used herein, the terms “binding” and “specific binding” preferably refer to the selective binding of the antibody or fragment of the present invention to human sFasL. The measure of the strength of antibody binding is called affinity. Methods for determining such binding and / or affinity using in vitro assays are known to those skilled in the art. According to this disclosure, detection by flow cytometry, immunohistochemistry and / or fluorescence is described and is particularly preferred herein.

[0057] A more preferred embodiment of the present invention refers to a nucleic acid molecule encoding the monoclonal antibody or its antigen fragment disclosed above for use in the prevention and / or treatment of SJS / TEN. The nucleic acid molecule may be a DNA vector or an RNA molecule formulated typically as a lipid nanoparticle encapsulation. When injected into a patient, the nucleic acid molecule encoding the antibody of the present invention is taken up by antibody-expressing cells and expresses the antibody in circulation. The use of DNA and RNA delivery technologies in antibody therapy, particularly the use of mRNA-encoded therapeutic antibodies, is a well-known therapeutic approach in the art (e.g., Van Hoecke and Roose, J Trans Med (2019) 17:54 and Deal at al., Vaccines (2021), 9, 2018).

[0058] In therapeutic applications, the anti-FasL antibody or its antibody fragment of the present invention is administered in an effective dose to a subject in need, particularly a human subject. The dose depends on the specific type of antibody, the severity and stage of the disease, and the route of administration.

[0059] In a preferred embodiment of the present invention, subjects suffering from SJS / TEN who require the anti-FasL antibody of the present invention are those whose serum FasL concentration levels are elevated compared to healthy subjects.

[0060] Typically, the anti-FasL antibody or antibody fragment of the present invention is administered as a pharmaceutical composition comprising an activator and a pharmaceutically acceptable carrier or excipient. Examples of carriers and excipients suitable for formulating antibodies or antibody fragments are well known in the art. The effective dose of the pharmaceutical of the present invention can range from 0.1 μg to 100 mg, with a maximum total dose of approximately 1 g, depending on the route of administration.

[0061] Accordingly, a further aspect of the present invention is a pharmaceutical composition comprising the above-mentioned anti-FasL antibody or its antigen-binding fragment together with one or more pharmaceutically acceptable carriers for use in methods for the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS). More preferably, the pharmaceutical composition comprises the above-mentioned anti-sFasL antibody or its antigen-binding fragment together with one or more pharmaceutically acceptable carriers for use in methods for the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS).

[0062] Depending on the stage and severity of the disorder, the pharmaceutical composition may be administered once or several times during the course of the disorder. For example, it may be administered once or several times a day, every other day, twice a week, or once a week for an appropriate period of time. The pharmaceutical composition may be administered in a single treatment cycle consisting of one or several doses, or in multiple treatment cycles, each consisting of one or several doses. Each treatment cycle may last from one day to several weeks, several months, or several years.

[0063] In certain embodiments, the pharmaceutical composition is administered parenterally, for example, by subcutaneous, intramuscular, or intravenous injection, or by infusion. In further embodiments, the pharmaceutical composition is administered topically, for example, topically, orally, nasally, or intrapulmonaryly, for example, by inhalation as an aerosol. Preferably, the composition is administered systemically.

[0064] Anti-FasL antibodies, preferably anti-sFasL antibodies or their antibody fragments, can be used as monotherapy or in combination therapy. Therefore, anti-FasL antibodies or their antibody fragments may be administered alone or together with at least one additional activator effective in treating SJS and / or TEN. In particular, the anti-FasL antibodies of the present invention can be used in combination with other agents selected from steroids, cyclosporine, IVIg, TNF inhibitors, or plasmapheresis.

[0065] Furthermore, the present invention will be described in more detail with reference to the following drawings and embodiments. [Brief explanation of the drawing]

[0066] [Figure 1] FasL concentrations in SJS / TEN serum measured by ELISA. S01 to S08 are all serum samples from SJS / TEN patients. The dotted line indicates a normal range; ≤124 pg / ml. [Figure 2] Viability of HaCaT cells after serum treatment. H0A is healthy serum, and S0A-S0B are SJS / TEN serum. Serum concentrations were 1%, 5%, and 10%. [Figure 3-3a] Cell death inhibition effect of PC111 antibody (including statistical analysis). PC111 antibody showed a concentration-dependent inhibitory effect on cell death. NS = No serum; PC111 antibody dose is expressed in μg / mL; "PC111 0" dose is SJS / TEN serum only. [Figure 4-4a] Cell death inhibition effect of zVAD (including statistical analysis) (Z-VAD-FMK, pancaspase inhibitor), μM. NS = no serum. "zVAD 0" dose is SJS / TEN serum only. [Figure 5-5a] No induction of HaCaT cell death by serum from healthy patients was observed (including statistical analysis). [Figure 6a] Overview of the experimental design and schedule for the in vivo study. An acetaminophen-induced SJS / TEN mouse model was constructed using patient-derived PBMCs and the causative drug acetaminophen. The effect of the PC111 antibody was compared between a group that received PC111 injection and a group that did not. [Figure 6b] Schematic diagram of the experimental design for the in vivo study. An acetaminophen-induced SJS / TEN mouse model was constructed using PBMCs (intravenous, iv) derived from one patient and the causative drug acetaminophen (oral administration (po) once daily for 14 days (po)) (vehicle group, n=6 mice). Another group was treated similarly, but PC111 was administered intravenously (iv) every two days (PC111 group, n=6 mice). Body weight, eye and skin appearance / changes were analyzed daily. [Figure 7a] Ocular symptoms on D14. The degree of conjunctival redness and edema was evaluated and scored on a 0-3 scale. N / A = Not available. [Figure 7b] Cases of ocular symptoms on D14. The degree of conjunctival hyperemia and edema was evaluated and scored on a 0-3 scale. A third group of age-matched, normal, untreated animals was used as a negative control (n=4). [Figure 8] Histopathological findings of each mouse group tested. Scale bars: 200 μm (top photo), 50 μm (bottom photo). [Figure 9] TUNEL staining for counting dead epithelial cells. The TUNEL assay detected a significantly higher number of dead epithelial cells in the vehicle group compared to the PC111-treated group and the control (normal) group. Scale bars: 200 μm (top photo), 50 μm (bottom photo). [Figure 10] The percentage of TUNEL-positive cells relative to the total number of conjunctival cells. The proportion of dead cells was significantly reduced in the PC111-treated group compared to the untreated control group. Significant differences were assessed using one-way ANOVA, followed by Tukey's multiple comparison test (***, p<0.001; ****, p<0.0001). Mean differences were analyzed using Dunnett's multiple comparison test. [Figure 11] Weight analysis over time. (A) Weight changes over time for the three animal groups. (B) Weight changes over time relative to baseline (day 1). (C) Area under the curve (AUC) analysis of weight curves, significance was evaluated by one-way ANOVA, followed by Tukey's multiple comparison test (**, p<0.01). (D) Analysis of mean AUC differences using Tukey's multiple comparison test. [Figure 12] The coupling curve (A) of FasL (sFasL) titrated against a constant concentration of 15 nM PC111, with data points related to higher-order oligomers shown in orange. Panel A shows the coupling isotherm with the three points marked in orange removed to allow for fitting (B). [Figure 13] An overview of membrane proteome arrays. [Figure 14] Membrane proteome array (MPA): Optimization of ligand concentrations for screening. Serial dilutions of each test ligand were incubated with known targets and assayed, and target binding was measured by flow cytometry. The optimal ligand concentration for screening was selected based on a joint evaluation of binding strength, background signal (left), and the incidence of high-background events (right). [Figure 15] MPA screening results. Each ligand was tested for binding to MPA at a pre-determined optimal concentration using flow cytometry. Binding interactions confirmed in downstream validation tests are shown, and proteins that failed validation were excluded. [Examples]

[0067] In vitro and in vivo experiments were conducted to evaluate the therapeutic effect of FasL antibodies against SJS / TEN.

[0068] An anti-FasL human IgG4,κ, monospecific bivalent antibody was used, which included heavy chain variable (VH) regions having complementarity-determining regions CDR H1, CDR H2, and CDR H3 of SEQ ID NOs: 1, 3, and 5, and light chain variable (VL) regions having complementarity-determining regions CDR L1, CDR L2, and CDR L3 of SEQ ID NOs: 7-9. This antibody is internally referred to as "antibody PC111" below.

[0069] 1. In-vitro testing 1.1 Materials and Methods 1.1.1 Cell culture HaCaT cells (spontaneously immortalized adult keratinocyte cell line) were purchased from CosmoBio (Tokyo, Japan) and cultured in an incubator at 37°C and 5% CO2 using CnT-PR (CELLnTEC, Stauffacherstrasse, Switzerland).

[0070] 1.1.2 Creation of SJS / TEN model cells To create SJS / TEN model cells, HaCaT cells cultured in CnT-PR were treated with 1%, 5%, and 10% SJS / TEN serum for 24 hours. PC111 antibody was added simultaneously with the serum. SJS / TEN patient serum was obtained from Niigata University Hospital.

[0071] 1.1.3 Cytotoxicity Test Cell viability or toxicity was evaluated using the Live / Dead Cell Staining Kit II (PromoCell, Sickingenstr, Germany). HaCaT cells were treated in 96-well plates and then cultured in a 37°C, 5% CO2 incubator. Images were acquired using a Keyence BZ-X710 all-in-one fluorescence microscope (Keyence, Osaka, Japan). The number of viable and dead cells was counted using ImageJ. Cytotoxicity was calculated as (number of dead cells) / (number of dead and viable cells). FasL levels in SJS / TEN and healthy serum were measured using an enzyme-linked immunosorbent assay (ELISA) kit (R&D systems, Minneapolis, MN).

[0072] 1.2 Results 1.2.1 FasL concentrations were high in the serum of SJS / TEN patients. Before performing in vitro assays using serum samples from SJS / TEN patients, FasL levels in SJS / TEN serum were measured by enzyme-linked immunosorbent assay (ELISA). FasL levels were elevated in the serum of 4 out of 8 SJS / TEN patients (normal range: <124 pg / ml) (Figure 1). This experiment was performed only once.

[0073] 1.2.2 Anti-FasL antibodies suppress cell death in SJS / TEN serum. SJS / TEN patient serum and serum from healthy control subjects were tested at concentrations of 1%, 5%, and 10% for 24 hours, and a decrease in cell viability was observed. SJS / TEN serum was confirmed to induce cell death in HaCaT cells. Compared to healthy serum, SJS / TEN serum significantly induced cell death even at low concentrations of SJS / TEN serum (1%) (S0A 1%; p=0.0027, S0B 1%; p=0.0024) (Figure 2). Survival rates decreased from 80-90% in normal serum to 50-60% in patient serum. It was found that cell death can be sufficiently induced even at a serum concentration of 1%.

[0074] Next, the efficacy of the PC111 antibody was confirmed using serum S08 with the highest FasL concentration, i.e., in the range of 90–200 pg / mL (analyzed in Figure 1) (see Figure 1). Then, the serum with the highest FasL concentration was used in the PC111 titration experiment. As a result, it was found that SJS / TEN serum S08 induced HaCaT cell death, and the PC111 antibody suppressed SJS / TEN serum S08-induced cell death in a dose-dependent manner (Figures 3 and 3a). Cell death was significantly suppressed when the PC111 antibody concentration was 10–100 μg / ml (PC111 10 μg / ml; p=0.0025, 100 μg / ml; p=0.0159, respectively). This experiment was repeated three times, and similar results were obtained (see statistical analysis in Figure 3a).

[0075] Furthermore, experiments using the pancaspase inhibitor zVAD were conducted as a control for pathway comparison of the PC111 antibody, and it was confirmed that zVAD has a significant effect in suppressing apoptosis (zVAD 50 μM; p=0.0025, 100 μM; p=1.57 x 10⁻¹⁰). -5 (Figures 4 and 4a). Healthy serum did not induce HaCaT cell death (Figures 5 and 5a). This experiment was repeated three times, and the results were the same (see statistical analysis in Figures 4a and 5a).

[0076] Since cell death is suppressed by zVAD, it is thought that apoptosis, a caspase-dependent form of cell death, was primarily observed in this experimental system using serum from SJS / TEN patients. Furthermore, the experimental results suggest that the PC111 antibody has a therapeutic effect against SJS / TEN through the suppression of keratinocyte apoptosis via Fas-FasL interaction.

[0077] In short, in vitro studies showed that SJS / TEN serum induces cell death in HaCaT cells in vitro, and that the PC111 antibody can suppress apoptosis in HaCaT cells in a dose-dependent manner.

[0078] 2. In vivo testing To evaluate the in vivo efficacy of the anti-FasL antibody PC111, we used a validated SJS / TEN mouse model (Saito et al. 2013) transplanted with PBMCs derived from SJS / TEN patients. As reported below, in vivo studies confirmed that the PC111 antibody of the present invention is an effective therapeutic agent for use in the treatment of SJS / TEN.

[0079] 2.1 Method 2.1.1 Mouse Six-week-old immunodeficient NOD / Shi-scid, IL-2Rγ null (NOG) mice were purchased from InVivoScience, Inc. (Tokyo, Japan). Six mice were included in the PC111 treatment group and vehicle group, and four mice were included in the control or normal group. All animal experiments were conducted with the approval of the Animal Experiment Ethics Committee of Niigata University.

[0080] 2.1.2 SJS / TEN mouse model using patient PBMCs The experimental design and schedule are outlined in Figures 6a and 6b. Peripheral blood mononuclear cells (PBMCs) were obtained from patients who had recovered from SJS / TEN. Patients did not receive systemic glucocorticoids at the time of PBMC collection. PBMCs (2 × 10⁻⁶) 6NOD-scid IL2r gamma (null)(NOG) mice were intravenously injected with ) followed by oral administration of the causative agent (acetaminophen, 1.5 mg / 100 μl).

[0081] The dosage used in this model was calculated from the usual adult dosage in milligrams per kg of body weight. Mice were administered the drug once daily. The dosage was also confirmed to be below the median lethal dose for mice. The drug dosage was estimated by dose conversion based on body weight. Specifically, PC111 antibody (100 μg / 100 μl) or PBS (100 μl) as a control was administered intravenously every two days starting from day 1 (D1, 100 μl). Mice were observed for 14 days. Any changes in the skin, eyes, and mucous membranes (skin color, mucosal bleeding, etc.) were checked daily. Body weight was recorded daily. Endpoints were collected on day 14 (D14). On day 14, general anesthesia was administered, the conjunctiva of the eyeball was dissected, and findings of hyperemia and edema were evaluated in detail. The mice were then sacrificed, and the eyeballs were collected as specimens. Ocular lesions were examined by histopathological examination and immunohistochemical staining.

[0082] 2.1.3 Immunohistochemical staining Terminal deoxyribonucleotide transferase-mediated dUTP nick end labeling (TUNEL) is a method for detecting apoptotic cells accompanied by DNA fragmentation by labeling the ends of nucleic acids. The TUNEL assay was performed according to the manufacturer's protocol (Takara Bio, Shiga, Japan). The conjunctiva of the eyeball was observed macroscopically, and the number of dead cells was macroscopically counted for all animals in each group. Significant differences in the proportion of dead cells between groups were analyzed by one-way analysis of variance (ANOVA), followed by Tukey's multiple comparison test. A p-value of < 0.05 was defined as statistically significant.

[0083] 2.2 Results The condition of the skin was observed daily throughout the experiment. No significant changes in the skin were observed until day 14. One mouse in the vehicle group died on day 12 (day 12) for unknown reasons.

[0084] Before sacrificing the mice on D14, the eyeballs were dislocated and the conjunctiva was examined in detail (Figures 7a and 7b). In the control vehicle group (n=2), conjunctival hyperemia was prominent (score 2 on a 0-3 scale), and mild edema (score 1 on a 0-3 scale) was also observed. In the PC111 treatment group (n=3), only one mouse showed mild conjunctival hyperemia (score 1 on a 0-3 scale) and no edema, while the other two PC111-treated mice showed neither hyperemia nor edema.

[0085] In a further step, the retrieved eyeballs were histologically evaluated (Figure 8). Hematoxylin-eosin staining revealed mild conjunctival edema in both PBMC-treated groups. Epithelial cell keratinization abnormalities, similar to those seen in SJS / TEN patients, were observed in the vehicle group. Therefore, TUNEL staining was performed to objectively evaluate the number of dead cells (Figure 9). As a result, a large number of dead epithelial cells were observed in the vehicle group, while only a few were found in the PC111 treatment group. The ratio of TUNEL-positive cells to the total number of conjunctival cells was calculated for all mice, and after one-way analysis of variance (ANOVA), Dunnett's multiple comparison test was performed to evaluate the significance of the difference (Figure 10). As a result, the PC111 treatment group had a significantly lower ratio of TUNEL-positive cells than the vehicle treatment group (p=0.0003), and there was no difference compared to the untreated control group (p=0.195). In this experiment, conjunctival hyperemia and edema were clearly suppressed, and TUNEL staining showed a significant decrease in TUNEL-positive cells in the PC111 treatment group.

[0086] The inventors found that the average body weight on day 1 was similar in the vehicle treatment group and the PC111 treatment group, but PC111 treatment caused a rapid recovery in body weight to a level comparable to that of the normal control group (Figure 11A). Furthermore, the AUC of the PC111 treatment group was very similar to that of the control group. Conversely, the vehicle treatment group showed a generally lower AUC. Although no statistically significant difference was observed between the vehicle and PC111 groups, this is likely due to the smaller number of animals, and the trend of improvement in AUC was clear (Figure 11C).

[0087] In summary, the data shown in Figure 11 provide valuable therapeutic conclusions, particularly considering the rapid recovery in the PC111-treated group compared to the vehicle-treated group. The evidence of AUC similarity between PC111 and the control group has high therapeutic value. Furthermore, these data indicate that blocking soluble FasL does not have an acute toxic effect, and in fact, the PC111 group is nearly identical to the untreated control group.

[0088] Thus, the combined results of the in vivo model studies provide evidence that the PC111 antibody is effective in reducing inflammatory symptoms and inhibiting apoptotic death of epithelial cells in the posterior conjunctival epithelium in a mouse model of SJS / TEN.

[0089] 3. Binding affinity and specificity of PC111 3.1 Surface Plasmon Resonance (SPR) Analysis of the Binding Affinity of PC111 to sFasL The binding affinity of PC111 to sFasL was measured for three different batches of PC111 using sFasL from different sources and different SPR instruments. The average binding affinity (equilibrium dissociation constant; KD) for the six measurements shown in Table 1 was 238 pM, with a range from 650 pM to 93 pM.

[0090] [Table 1]

[0091] 3.2 Flow-induced analysis of variance In the following experiments, the antibody PC111 was tested against the FasL protein for binding. Flow-induced variance analysis (FIDA) was used in these tests. This analysis utilizes dispersion phenomena in pressure-driven flow to accurately assess molecular diffusivity and hydrodynamic radius (Rh) after signal analysis. Changes in the hydrodynamic radius of the binder reflect the strength of the interaction.

[0092] 3.2.1 Methods and Materials Capillary: Coated capillary Detector: 480nm fluorescence detection of riboflavin Indicator (labeled molecule) consumption per measurement: 39 nL Analyte consumption per measurement: 12 μL Analysis time: 5.5 minutes per measurement PC111-Alexa concentration: 100nM FasL concentration: 0.01-320nM Assay buffer: PBS pH 7.45, BSA 0.1% (1 mg / ml) Mixing principle: Premix (>10 minutes) Temperature: 25℃

[0093] 3.2.2 Results PC111 was used as an indicator at a constant concentration of 15 nM, and FasL was titrated in the range of 0.01 to 320 nM. The Rh value of PC111 alone was measured at 4.8 nm, and in combination with FasL at 6.8 nm. Since FasL is a homotrimer, PC111 can theoretically bind to FasL in a stoichiometric ratio of 1 FasL:3 PC111 depending on steric hindrance. Multiple stoichiometric combinations are possible.

[0094] PC111 was labeled, and its Rh increase was detected by adding sFasL (Figure 12A). Higher Rh levels were detected when the sFasL concentration reached the range of 10–80 nM. The orange square dots in Figure 12A strongly suggest the presence of higher-order oligomers. When sFasL exceeded 100 nM, the Rh of PC11 returned to 6.8 nm, suggesting a 1:1 stoichiometry. Fitting models including higher-order oligomers have not yet been developed, as they may differ from system to system. Therefore, the three points shown in orange in Figure 12A were removed, and a standard 1:1 Excess Indicator fitting model was applied (Figure 12B). The dissociation constant between PC111 and sFasL in solution KD was 340 pM.

[0095] 3.3 Membrane Proteome Arrays To investigate the target specificity of the antibody and the possibility of off-target binding, we screened PC111 binding targets for the human membrane proteome using membrane proteome array (MPA) technology. This technology uses flow cytometry to directly detect antibody binding to membrane proteins expressed in unfixed cells (Figure 13). All target proteins possess native conformations and appropriate post-translational modifications.

[0096] 3.3.1 Determination of Assay Screening Conditions To optimize ligand concentrations and cell lines for screening, HEK-293T cells (ATCC CRL-3216) and QT6 cells (ATCC CRL-1708) were transfected in 384-well cell culture plates at a density of 18,000 cells / well with plasmids encoding known ligand targets, protein A (bound to antibody Fc; positive control), or vector alone (pUC; negative control). Transfected cells were incubated for 36 hours at 37°C and 5% CO2 in a medium consisting of Corning DMEM, 10% FBS, 2 mM L-alanyl-L-glutamine, Pen Strep, MEM NEAA, and 10 mM HEPES. After incubation, a 4-fold dilution series of each ligand, starting at 20 μg / ml, was added to the transfected cells in four fractional doses. Binding ligands were detected by high-throughput immunofluorescence flow cytometry using a single dilution of a fluorescently labeled secondary antibody.

[0097] Mean fluorescence intensity (MFI) values ​​were determined for each test ligand dilution in each cell line using ForeCyt Software (Intellicyt) and plotted using Excel (Microsoft). The high background rate for each assay condition was calculated as the percentage of positive events above the defined fluorescence threshold in cells transfected with the negative control. The optimal screening concentration and cell line for each test ligand (Table 2 below) were determined by background signal (MFI) and the high background rate in the vector control (Figure 14). Molecules were preferentially screened in HEK-293T cells at the highest concentration that yielded an acceptable low background (<50,000 MFI) and the minimum high background rate (<1%). If an acceptable screening condition was not identified in HEK-293T cells, the molecule was screened in QT6 cells at the highest acceptable concentration.

[0098] [Table 2]

[0099] 3.3.2. Membrane Proteome Array Screen Plasmids containing cDNA clones of approximately 6,000 membrane proteins (corresponding to over 94% of the human membrane proteome) were transfected into HEK-293T cells (18,000 cells / well) in 384-well cell culture plates (one unique cDNA-containing plasmid per well). The cells were incubated at 37°C and 5% CO2 in a medium consisting of Corning DMEM, 10% FBS, 2 mM L-alanyl-L-glutamine, Penn / strep, MEM NEAA, and 10 mM HEPES. Each 384-well plate also contained wells independently transfected with plasmids encoding either GFP or a membrane-bound protein A construct, serving as controls for transfection efficiency and binding of fluorescently labeled detection antibodies. After 36 hours of incubation, cells were pulled up using a CellStripper and reformatted into a two-dimensional matrix of rows and columns in new 384-well plates using a JANUS Automated Workstation. Each well in the matrix plate contains 48 overexpression protein components, and each protein is represented by a unique combination of two different wells in the matrix plate, as it is contained in both a "row" pool and a "column" pool. Therefore, all proteins in the MPA are tested twice for reactivity. Test ligands were added to the Membrane Proteome Array matrix plate at predetermined concentrations (Table 2 above), washed with 1×PBS, and detected by flow cytometry using fluorescently labeled antibodies. All flow cytometry data were obtained using ForeCyt Software (Intellicyt).

[0100] Next, the test ligand targets were identified by detecting their binding to overlapping pool matrix wells obtained from the same transfection plate. This allowed for specific deconvolution. Individual targets showing binding at least 3 standard deviations higher than the background in both wells were selected for downstream validation experiments.

[0101] To represent the binding of test ligands to each protein in the Membrane Proteome Array with a single value, two-dimensional binding data was transformed using a standard matrix deconvolution method. In short, each point (representing individual overexpressed proteins) was converted to radians, then transformed using the formula r.sin(2θ), and plotted as target binding (Figure 15). Non-specific fluorescence was defined as a value below three standard deviations of the transformed value of the calculated background fluorescence. All data and analyses were performed in Excel (Microsoft).

[0102] 3.3.3 Target Verification To validate the identified off-target interactions, cells were transfected in a 384-well format with plasmids encoding the identified target, protein A, or the vector alone. After incubation at 37°C and 5% CO2 for 36 hours, each test ligand, diluted 4-fold from 20 μg / ml, was added to the transfected cells, and ligand binding was detected using a high-throughput immunofluorescence flow cytometry assay (under the same conditions as described in Table 2 above).

[0103] If the known target of the molecule was the only protein validated by the MPA screening, validation is not shown because the Assay Setup results have already demonstrated specific reactivity in a screening equivalent to the validation screening.

[0104] 3.3.4 Results During the MPA trial, binding of PC111 to Fc receptors FCGR1A and FCGR2B, as well as to the Fc binding control protein A, was confirmed. Binding of PC111 to membrane-bound FASLG was not detected. No off-target binding of PC111 was identified.

[0105] References 1) Viard I, et al. Inhibition of toxic epidermal necrolysis by blockade of CD95 with human intravenous immunoglobulin. Science 1998; 282:490-3. 2) Abe R, et al. Toxic epidermal necrolysis and Stevens-Johnson syndrome are induced by soluble Fas ligand. Am J Pathol 2003; 162: 1515-20. 3) Chang HY, et al. Kinetics and specificity of fas ligand induction in toxic epidermal necrolysis. Arch Dermatol 2004; 140: 242-4. 4) Abe R, et al. Toxic epidermal necrolysis, and Stevens-Johnson syndrome: soluble Fas ligand involvement in the pathomechanisms of these diseases. J Dermatol Sci 2008; 52: 151-9. 5) Abe R, et al. Immunological response in Stevens-Johnson syndrome and toxic epidermal necrolysis. J Dermatol 2015; 42: 42-8. 6) Puviani M, et al. Fas ligand in pemphigus sera induces keratinocyte apoptosis through the activation of caspase-8. J Invest Dermatol 2003; 120: 164-7. 7) Lotti R, et al. Soluble Fas Ligand Is Essential for Blister Formation in Pemphigus. Front Immunol. 2018; 9: 370. 8) Chang HH. et al, A Review of the Systemic Treatment of Stevens-Johnson Syndrome and Toxic Epidermal Necrolysis, Biomedicines, 2022, 10, 2105 9) Sharma K et al., Death the Fas way: regulation and pathophysiology of Fas and its ligand. Pharmacol Ther. 88:333-347, 2000 10) Saito N, et al. Stevens-Johnson syndrome / toxic epidermal necrolysis mouse model generated by using PBMCs and the skin of patients. J Allergy Clin Immunol 2013; 131: 434-41. 11) Saito N, et al. An annexin A1-FPR1 interaction contributes to necroptosis of keratinocytes in severe cutaneous adverse drug reactions. Sci Transl Med 2014; 6: 245ra95. 12) Zhang S, et al. Biologic TNF-alpha inhibitors in the treatment of Stevens-Johnson syndrome and toxic epidermal necrolysis: a systemic review. J Dermatolog Treat. 2020; 31: 66-73. 13) Van Hoecke and Roose, “How mRNA therapeutics are entering the monoclonal antibody filed”, J Trans Med (2019) 17:54 14) Deal at al., “Advancements in mRNA Encoded Antibodies for Passive Immunotherapy”, Vaccines (2021), 9, 2018

Claims

1. A monoclonal antibody or antigen-binding fragment specific to human Fas ligand protein (FasL), comprising at least one heavy chain variable (VH) region and at least one light chain variable (VL) region, for use in methods for the prevention and / or treatment of toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), wherein the antibody or antigen-binding fragment is: (i) The following heavy chain CDR H1, CDR H2, and CDR H3: (a 1 ) CDR H1: Arg His Gly Ile Thr (Sequence ID 1) or (a 2 ) CDR H1: Ser His Gly Ile Ser (Sequence No. 2), (b 1 ) CDR H2: Trp Ile Asn Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Val Gln Gly (Sequence ID 3) or (b) 2 ) CDR H2: Trp Ile Asn Ala Tyr Ser Gly Asn Thr Asn Tyr Ala Gln Lys Leu Gln Gly (sequence number: 4), (c 1 ) CDR H3: Glu Thr Met Val Arg Gly Val Pro Leu Asp Tyr (SEQ ID NO: 5), or (c 2 CDR H3: Glu Thr Met Val Arg Gly Val Pro Cys Asp Tyr (Layout No. 6) VH region having complementarity determination region (CDR), and The following light chains CDR L1, CDR L2, and CDR L3: (a 3 ) CDR L1: Arg Ala Ser Gln Ser Val Ser Tyr Leu Ala (SEQ ID NO: 7), (b 3 ) CDR L2: Gly Ala Ser Ser Arg Ala Thr (Sequence No. 8), (c 3 ) CDR L3: Gln Gln Tyr Gly Ser Ser Pro Trp Thr (SEQ ID NO: 9) Complementarity Determination Region (CDR) Antibodies or antigen-binding fragments containing; or (ii) An antibody or antigen-binding fragment that competes with the antibody or antigen-binding fragment of (i) for binding to human Fas ligand protein (FasL); A monoclonal antibody or its antigen-binding fragment, selected from the following.

2. A monoclonal antibody or antigen-binding fragment according to claim 1 for use of claim 1, comprising at least one heavy chain variable (VH) region and at least one light chain variable (VL) region, wherein the antibody or antigen-binding fragment comprises the following heavy chains CDR H1, CDR H2, and CDR H3: (a 1 ) CDR H1: Arg His Gly Ile Thr (Sequence ID 1), (b) 1 ) CDR H2: Trp Ile Asn Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Val Gln Gly (sequence number 3), (c 1 ) CDR H3: Glu Thr Met Val Arg Gly Val Pro Leu Asp Tyr (Allocation No. 5) VH region having a complementarity determination region (CDR); and The following light chains CDR L1, CDR L2, and CDR L3: (a 3 ) CDR L1: Arg Ala Ser Gln Ser Val Ser Tyr Leu Ala (SEQ ID NO: 7), (b 3 ) CDR L2: Gly Ala Ser Ser Arg Ala Thr (Sequence No. 8), (c 3 ) CDR L3: Gln Gln Tyr Gly Ser Pro Trp Thr (SEQ ID NO: 9) Complementarity Determination Region (CDR); A monoclonal antibody or its antigen-binding fragment, selected from antibodies or antigen-binding fragments containing the above.

3. The VL region of the antibody has the following amino acid sequence: EIVLTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQYGSSPWTFGQGTKVEIKRTVAAPSVFIFP (Sequence ID 10) including and The VH region of the antibody has the following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYIFIRHGITWVRQAPGQGLEWMGWINAYNGNTNYAQKVQGRVTMTTDKSTSTAYMELRSLRSDDAAVYYCARETMVRGVPLDYWGQGTLVTVSSASTKGPSVFPLA (Sequence ID 11), or QVQLVQSGAEVKKPGASVKVSCKASGYIFISHGISWVRQAPGQGLEWMGWINAYSGNTNYAQKLQGRVTMTTDRSTSTAYMELRSLRSDDTAVYYCARETMVRGVPCDYWGQGTLVTVSSASTKGPSVFPLA (Sequence ID 12) An antibody or antigen-binding fragment according to claim 1 or 2 for use in claim 1, comprising:

4. The VL region of the antibody has the following amino acid sequence: EIVLTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQYGSSPWTFGQGTKVEIK (Sequence ID 13), including and The VH region of the antibody has the following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYIFIRHGITWVRQAPGQGLEWMGWINAYNGNTNYAQKVQGRVTMTTDKSTSTAYMELRSLRSDDAAVYYCARETMVRGVPLDYWGQGTLVTVSS (Sequence ID 14), or QVQLVQSGAEVKKPGASVKVSCKASGYIFISHGISWVRQAPGQGLEWMGWINAYSGNTNYAQKLQGRVTMTTDRSTSTAYMELRSLRSDDTAVYYCARETMVRGVPCDYWGQGTLVTVSS (Sequence ID 15) An antibody or antigen-binding fragment according to any one of claims 1 to 3 for use in claim 1, comprising:

5. An antibody or antigen-binding fragment according to any one of claims 1 to 4 for use in claim 1, which recognizes the same epitope on human FasL as the antibody of claim 1(i) or 2(i).

6. The antibody or antigen-binding fragment according to any one of claims 1 to 5 for use in claim 1, wherein the antibody is selected from a partially or fully human antibody, a chimeric antibody and / or a humanized antibody, and the antigen-binding fragment thereof is selected from Fab, Fab' and / or F(ab')2 and / or a single-chain Fv fragment.

7. The antibody or antigen-binding fragment according to any one of claims 1 to 6 for use in claim 1, wherein the antibody has an IgG heavy chain constant region, preferably an IgG1 or IgG4 heavy chain constant region.

8. A nucleic acid molecule encoding a monoclonal antibody or its antigen fragment according to any one of claims 1 to 7, for use in claim 1.

9. A nucleic acid molecule according to claim 8 for use in claim 1, which is a DNA vector or an RNA molecule.

10. An antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, or a nucleic acid molecule according to any one of claims 8 to 9, for use in monotherapy as described in claim 1.

11. For use in claim 1, in combination with at least one further active ingredient effective against toxic epidermal necrolysis (TEN) and / or Stevens-Johnson syndrome (SJS), an antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, or a nucleic acid molecule according to any one of claims 8 to 9.

12. An antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, or a nucleic acid molecule according to any one of claims 8 to 9, for use in claim 11, wherein the further active ingredient is selected from at least one of a steroid, cyclosporine, IVIg, a TNF inhibitor, and / or plasmapheresis.

13. An antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, or a nucleic acid molecule according to any one of claims 8 to 9, for use in human therapeutics.

14. A pharmaceutical composition comprising, together with one or more pharmaceutically acceptable carriers for use in claim 1, an antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, or a nucleic acid molecule according to any one of claims 8 to 9.

15. An antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, or a nucleic acid molecule according to any one of claims 8 to 9, or a pharmaceutical composition according to claim 14, which is administered systemically and / or locally.

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