Bispecific antibodies for use in treatment of XIAP deficient or CDC42 mutated patients

Abstract

The present invention relates to a bivalent bispecific monoclonal antibody (bbmAb), or a variant thereof, for use in treating or alleviating symptoms of a self-inflammation or disease based on CDC42 mutation or XIAP deficiency in a subject in need thereof.

Description

Technical Field

[0001] This invention relates to the treatment of bivalent bispecific monoclonal antibodies for use in patients in need of treating autoinflammatory or inflammatory diseases based on XIAP deficiency or CDC42 mutations. This disclosure also relates to methods and treatment regimens for treating autoinflammatory or inflammatory diseases based on XIAP deficiency or CDC42 mutations using bispecific antibodies that simultaneously target both IL-1β and IL-18. In some cases, bbmAb (or a variant thereof) is used to treat subjects who carry mutations in the gene encoding the XIAP gene or in the CDC43 gene and suffer from, for example, hypogammaglobulinemia, cytopenia, inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, syndromic thrombocytopenia, megaloblastic thrombocytopenia, hematopoietic insufficiency, autoinflammatory disease, syndromic immunodeficiency or relapsing hemophagocytic lymphohistiocytosis (HLH), autoinflammatory disease with infantile enterocolitis (AIFEC) or macrophage activation syndrome (MAS). Background Technology

[0002] Inflammasomes are intracellular polyprotein complexes that typically form and activate in response to pathogens or risk-associated molecular patterns (PAMP / DAMP). Inflammasome pathologies are a group of mechanism-related diseases resulting from the overactivation of individual inflammasomes leading to differentiated clinical phenotypes (depending on the production of effector cytokines and tissue-specific expression). X-linked inhibitor of apoptosis (XIAP) deficiency (also known as X-linked lymphoproliferative syndrome type 2 (XLP2)) is an autoinflammatory or inflammasome pathology and adaptive immunodeficiency characterized by excessive inflammation and immune dysregulation (Geerlinks et al., Journal of Clinical Immunology (2022) 42:901-903). XIAP deficiency was first described in 2006. It is caused by pathogenic variants of the XIAP / BIRC4 gene (XIAP gene identification: HGNC: 592; NCBI gene: 331; Ensembl: ENSG00000101966; OMIM®: 300079; UniProtKB / Swiss-Prot: P98170). These pathogenic variants lead to a variety of clinical manifestations, most commonly hemophagocytic lymphohistiocytosis (HLH), inflammatory bowel disease, and splenomegaly.

[0003] Cell cycle 42 (CDC42) (CDC42 gene identification: HGNC: 1736; NCBI gene: 998; Ensembl: ENSG00000070831; OMIM®: 116952; UniProtKB / Swiss-Prot: P60953) is an intracellular member of Ras homologous (Rho) GTPases, which control cell polarity by regulating the assembly of actin cytoskeleton structures. From an immunological perspective, CDC42 regulates multiple cellular processes, such as migration, immune synapse formation, and polarized cytokine secretion. CDC42 also plays a crucial role in cell proliferation and hematopoiesis (Miyazawa H and Wada T (2022) Immune-mediated inflammatory diseases with chronic excess of serum interleukin-18. Front. Immunol. 13:930141).

[0004] Patients affected by XIAP deficiency or CDC42 mutation-related autoinflammatory diseases typically have significantly elevated IL-1β and / or IL-18 levels. These elevated effector cytokines drive the relevant clinical manifestations listed above, centered on neurodevelopment, hematopoiesis, and immune responses (Takenouchi et al. 2015, Asiri et al. 2021, Coppola et al. 2022). The inventors of this invention believe that only the combination of anti-IL-1 receptor (e.g., anakinin) and recombinant IL-18 binding protein (e.g., IL-18 BP) has been reported to be clinically effective in a limited number of infantile XIAP deficiency cases.

[0005] No approved therapeutics directly and specifically target the underlying IL-1β and IL-18-driven autoinflammatory processes to improve overall clinical outcomes in patients with XIAP-deficient or CDC42-mutant autoinflammatory or disease-related conditions. Therefore, there is a long-standing unmet need in the field for improved treatment of XIAP-deficient or CDC42-mutant autoinflammatory or disease-related conditions or inflammasome lesions. Summary of the Invention

[0006] Bispecific antibodies or functional fragments thereof that simultaneously target both IL-1β and IL-18 are described for use in the prevention or treatment of autoinflammatory diseases or inflammasome lesions based on XIAP deficiency or CDC42 mutation in subjects. In some cases, bispecific antibodies or functional fragments thereof that simultaneously target both IL-1β and IL-18 are used in the treatment of patients / methods for treating patients who are XIAP deficient or carry a CDC42 mutation and present with or have the following clinical symptoms: hypogammaglobulinemia, cytopenia, inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, syndromic thrombocytopenia, megaloblastic thrombocytopenia, hematopoietic insufficiency, autoinflammatory disease, syndromic immunodeficiency, or relapsing hemophagocytic lymphohistiocytosis (HLH). In some cases, methods are used to prevent or treat autoinflammatory diseases based on XIAP deficiency or CDC42 mutations in subjects who present with clinical symptoms of autoinflammatory disease accompanied by infantile enterocolitis (AIFEC) or macrophage activation syndrome by administering a therapeutically effective dose of a bispecific antibody that simultaneously targets both IL-1β and IL-18. This article also describes methods for preventing or treating autoinflammatory diseases based on XIAP deficiency or CDC42 mutations by administering a therapeutically effective dose of a bispecific antibody that simultaneously targets both IL-1β and IL-18 to subjects in need. In some cases, these methods involve treating autoinflammatory disease accompanied by infantile enterocolitis (AIFEC) or macrophage activation syndrome (MAS) in patients who are XIAP deficient or carry a CDC42 mutation and present with clinical symptoms of disease by administering a therapeutically effective dose of a bispecific antibody that simultaneously targets both IL-1β and IL-18.

[0007] This document further provides specific dosing regimens for the methods or uses of the bispecific antibody (e.g., bbmAb) that simultaneously targets both IL-1β and IL-18 as described herein. In one embodiment, in the method or therapeutic use of the bispecific antibody simultaneously targeting both IL-1β and IL-18 for the treatment of autoinflammatory or disease-related by XIAP deficiency or CDC42 mutation, the method or use includes administering about 1 mg / kg to about 35 mg / kg of the bispecific antibody to a subject in need. In another embodiment, about 10 mg / kg of the bispecific antibody is administered to the subject, wherein the bispecific antibody is administered intravenously or subcutaneously. In various embodiments, 10 mg / kg of the bispecific antibody simultaneously targeting both IL-1β and IL-18 is administered intravenously, optionally wherein the antibody is administered every other week. In another embodiment, the dose of the bispecific IL-1β and IL-18 antibody administered is about 50 mg to about 900 mg subcutaneously.

[0008] Further embodiments described herein relate to drug combinations and pharmaceutical compositions comprising a bispecific antibody (e.g., bbmAb) that simultaneously targets both IL-1β and IL-18, optionally with a pharmaceutically acceptable carrier, for use in the treatment or prevention of autoinflammatory or disease-associated with XIAP deficiency or CDC42 mutations, such as for the treatment of inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, autoinflammatory disease, recurrent hemophagocytic lymphohistiocytosis (HLH), AIFEC, or macrophage activation syndrome (MAS). Further features and advantages of the methods and uses described herein will become apparent from the detailed description that follows.

[0009] In a first aspect, this disclosure relates to a method for treating or preventing symptoms of an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, the method comprising administering to the subject a therapeutically effective amount of a bispecific antibody, wherein the antibody contains

[0010] a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and

[0011] b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18.

[0012] In a second aspect, this disclosure relates to a method for alleviating, preventing, or reducing the severity of symptoms of an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation in subjects in need, the method comprising administering to the subject a therapeutically effective amount of a bispecific antibody, wherein the antibody contains

[0013] a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and

[0014] b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18.

[0015] In a third aspect, this disclosure relates to bispecific antibodies that contain...

[0016] a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and

[0017] b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18 for use, such as in treating or preventing autoinflammatory or disease-related to XIAP deficiency or CDC42 mutation in subjects in need.

[0018] In a fourth aspect, this disclosure relates to the methods and treatments of the first, second, and third aspects, wherein the subject is administered a bispecific antibody that simultaneously targets both IL-1β and IL-18 at a dose of about 1 mg / kg to about 35 mg / kg. In a preferred embodiment of the fourth aspect, the subject receiving the treatment is administered a bispecific antibody at a dose of about 10 mg / kg.

[0019] In one aspect of this disclosure, the subject was given an intravenous or subcutaneous administration of a bispecific antibody that simultaneously targets both IL-1β and IL-18.

[0020] In another preferred embodiment, a bispecific antibody that targets both IL-1β and IL-18 is administered intravenously to the treated subject at a dose of about 10 mg / kg.

[0021] In one embodiment, a single intravenous dose of 10 mg / kg is administered to the patient on day 1, simultaneously targeting bispecific antibodies against both IL-1β and IL-18. In another embodiment, a single intravenous dose of 10 mg / kg is administered to the patient on days 1 and 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, and / or 14, simultaneously targeting bispecific antibodies against both IL-1β and IL-18. In yet another embodiment, a single intravenous dose of 10 mg / kg is administered to the patient on days 1 and 14, simultaneously targeting bispecific antibodies against both IL-1β and IL-18. In yet another embodiment, a single intravenous dose of 10 mg / kg is administered to the patient every two weeks, simultaneously targeting bispecific antibodies against both IL-1β and IL-18. In one embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for a period of up to 28 weeks. In another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for a period of up to 24 weeks.

[0022] In one embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for a period of up to 3 years. In another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for 2 weeks. In yet another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for 8 weeks. In one embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for 14 weeks. In another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for 24 weeks. In yet another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient every two weeks for at least 2 weeks. In one embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient at a dose of 10 mg / kg every 2 weeks for at least 8 weeks. In another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient at a dose of 10 mg / kg every 2 weeks for at least 14 weeks. In yet another embodiment, a bispecific antibody targeting both IL-1β and IL-18 is administered intravenously to the patient at a dose of 10 mg / kg every 2 weeks for at least 24 weeks.

[0023] In a preferred embodiment, a bispecific antibody that targets both IL-1β and IL-18 is administered to the patient, for example, intravenously every two weeks at a dose of 10 mg / kg or about 10 mg / kg.

[0024] In another embodiment of the foregoing aspects of this disclosure, the bispecific antibody targeting both IL-1β and IL-18 is administered in combination with at least one additional therapeutic agent.

[0025] In a particular embodiment of any of the foregoing aspects disclosed herein, the first and second constant heavy chains of the bispecific antibody are human IgA, IgD, IgE, IgG, or IgM, preferably IgD, IgE, or IgG, such as human IgG1, IgG2, IgG3, or IgG4, preferably IgG1.

[0026] In another embodiment of any of the foregoing aspects disclosed herein, the first and second constant heavy chains of the bispecific antibody are IgG1, and

[0027] a. The first constant heavy chain has a point mutation that produces a pestle structure, and the second constant heavy chain has a point mutation that produces a mortar structure, or

[0028] b. The first constant heavy chain has a point mutation that produces a mortar structure, and the second constant heavy chain has a point mutation that produces a pestle structure, and optionally...

[0029] c. The first and second constant heavy chains have mutations that lead to disulfide bridges.

[0030] In particularly preferred embodiments of the first, second, and third aspects of the methods and uses described herein, the first immunoglobulin VH1 domain of the bispecific antibody comprises:

[0031] i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; or

[0032] ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 79, CDR2 has the amino acid sequence SEQ ID NO: 80, and CDR3 has the amino acid sequence SEQ ID NO: 81; and

[0033] The first immunoglobulin VL1 domain of the bispecific antibody includes:

[0034] iii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 93, and CDR3 has the amino acid sequence SEQ ID NO: 94 or...

[0035] iv. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 95, CDR2 has the amino acid sequence SEQ ID NO: 96, and CDR3 has the amino acid sequence SEQ ID NO: 97; and

[0036] The second immunoglobulin VH2 domain of the bispecific antibody includes:

[0037] v. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; or

[0038] vi. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 47, CDR2 has the amino acid sequence SEQ ID NO: 48, and CDR3 has the amino acid sequence SEQ ID NO: 49; and the second immunoglobulin VL2 domain of the bispecific antibody comprises:

[0039] vii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62 or...

[0040] viii. Hypervariable regions CDR1, CDR2 and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 63, CDR2 has the amino acid sequence SEQ ID NO: 64 and CDR3 has the amino acid sequence SEQ ID NO: 65.

[0041] In another preferred embodiment of this disclosure, the antibody used in the method / treatment according to any one of the foregoing aspects comprises:

[0042] a. The first immunoglobulin VH1 domain of the amino acid sequence SEQ ID NO: 85,

[0043] b. The first immunoglobulin VL1 domain of the amino acid sequence SEQ ID NO: 101,

[0044] c. The second immunoglobulin VH2 domain of the amino acid sequence SEQ ID NO: 53, and

[0045] d. The second immunoglobulin VL2 domain of the amino acid sequence SEQ ID NO: 69.

[0046] In another preferred embodiment of this disclosure, the antibody used in the method / treatment according to any one of the foregoing aspects comprises

[0047] e. The first immunoglobulin heavy chain, amino acid sequence SEQ ID NO: 87,

[0048] f. The first immunoglobulin light chain, amino acid sequence SEQ ID NO: 103,

[0049] g. The second immunoglobulin heavy chain with amino acid sequence SEQ ID NO: 55, and

[0050] h. Amino acid sequence SEQ ID NO: 71, second immunoglobulin light chain.

[0051] In another embodiment of the foregoing aspects of this disclosure, the treated subject suffers from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In another embodiment of the foregoing aspects of this disclosure, the treated subject has a loss-of-function mutation in the XIAP / BIRC4 gene encoding the XIAP protein (as disclosed below), a mutation in the CDC42 gene leading to aberrant palmitoylation of the CDC42 protein, or a missense mutation at the C-terminus of the CDC42 protein affecting CDC42 protein localization (as disclosed below).

[0052] In some cases, subjects requiring treatment for autoinflammatory or CDC42-mutant autoinflammatory or inflammatory diseases have excessively elevated IL-18 and IL-1β levels, or excessively elevated serum IL-18 and IL-1β levels, compared to a control group that does not exhibit XIAP-deficiency or CDC42-mutant autoinflammatory or inflammatory diseases. Excessively elevated IL-18 and / or IL-1β levels refer to a patient's IL-18 and / or IL-1β levels being significantly higher than the upper limit for healthy individuals. In some cases, subjects requiring treatment for XIAP-deficiency or CDC42-mutant autoinflammatory or inflammatory diseases have excessively elevated serum IL-18 and IL-1β levels compared to a control group that does not exhibit XIAP-deficiency or CDC42-mutant autoinflammatory or inflammatory diseases. In some cases, this excessively elevated serum IL-18 level refers to excessively elevated total serum IL-18 levels. In some cases, this excessively elevated serum IL-18 level refers to excessively elevated free serum IL-18 levels. In some cases, the subject has elevated serum C-reactive protein (CRP) levels compared to the control group. In some cases, the subject had high serum ferritin levels compared to the control group.

[0053] In a preferred embodiment, subjects requiring treatment for XIAP-deficiency or CDC42-mutant autoinflammatory or disease-related conditions have excessively elevated serum total IL-18 levels compared to a control group that does not exhibit XIAP-deficiency or CDC42-mutant autoinflammatory or disease-related conditions. In some cases, this excessively elevated serum total IL-18 level is greater than 1000 pg / mL. In some cases, this excessively elevated serum total IL-18 level is greater than 5000 pg / mL. In some cases, this excessively elevated serum total IL-18 level is greater than 10,000 pg / mL. In some cases, this excessively elevated serum total IL-18 level is between about 1000 pg / mL and about 20000 pg / mL. In some cases, this excessively elevated serum total IL-18 level is between about 5000 pg / mL and about 20000 pg / mL. In some cases, the excessively elevated serum total IL-18 level is between approximately 10,000 pg / mL and approximately 20,000 pg / mL. In some cases, the excessively elevated serum total IL-18 level is between approximately 1,000 pg / mL and approximately 25,000 pg / mL. In some cases, the excessively elevated serum total IL-18 level is between approximately 5,000 pg / mL and approximately 25,000 pg / mL. In some cases, the excessively elevated serum total IL-18 level is between approximately 10,000 pg / mL and approximately 25,000 pg / mL.

[0054] In some cases, subjects requiring treatment for autoinflammatory or CDC42-mutant autoinflammatory or ...

[0055] In some cases, the excessively elevated serum IL-1β level is greater than 5 pg / mL. In some cases, the excessively elevated serum IL-1β level is greater than 10 pg / mL. In some cases, the excessively elevated serum IL-1β level is between approximately 5 pg / mL and approximately 25 pg / mL. In some cases, the excessively elevated serum IL-1β level is between approximately 10 pg / mL and approximately 25 pg / mL.

[0056] In some cases, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation is under 17 years of age and weighs at least 3 kg. In some cases, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation is under 10 years of age and weighs at least 3 kg. In some cases, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation is under 5 years of age and weighs at least 3 kg. In some cases, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation is an infant weighing at least 3 kg, and wherein the subject has both infantile enterocolitis and excessively elevated serum levels of total IL-18. In some cases, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation is an infant weighing at least 3 kg, and wherein the subject has both infantile enterocolitis and excessively elevated serum levels of free IL-18.

[0057] In a particularly preferred embodiment of the foregoing aspects of this disclosure, the subject receiving the treatment has AIFEC. In a further preferred embodiment, the subject receiving the treatment has AIFEC and excessively elevated serum levels of total IL-18. In another embodiment, the subject receiving the treatment has AIFEC and excessively elevated serum levels of free IL-18.

[0058] In a particularly preferred embodiment of the foregoing aspects of this disclosure, the treated subject's XIAP or CDC42 gene contains one or more point mutations or mutations including deletions (small or large exon deletions) and insertions, resulting in nonsense and missense mutations, intron or frameshift mutations. These mutations may result in the absence of functionally expressed XIAP and CDC42 proteins (which are either truncated or dysfunctional, respectively). A detailed overview of 90 pathogenic mutations in the XIAP gene is disclosed in Mudde et al., (2021) Evolution of Our Understanding of XIAP Deficiency. Front. Pediatr. 9:660520. Figure 1The content of this article is incorporated herein by reference. As disclosed by Martinelli et al. (The American Journal of Human Genetics, 102, 309-320, February 1, 2018), nine distinct gene mutations in the CDC42 gene have been identified (two amino acid substitutions affecting the N-terminal α-helix (residues Ile21 and Tyr23), three involving adjacent residues within the switch II motif (Tyr64, Arg66, and Arg68), two mapping to the fourth b-strand (Cys81 and Ser83), and the remaining two located near the C-terminus (Ala159 and Glu171)). Gernez et al. described the inflammatory and hematologic phenotypes of four patients with three variants affecting the C-terminus of CDC42 (p.R186C, p.C188Y, and p). 192Cext 24) is heterozygous (Gernez Y, de Jesus AA, Alsaleem H, Macaubas C, Roy A, Lovell D et al., Severe autoinflammation in 4 patients with C-terminal variants in celldivision control protein 42 homolog (CDC42) successfully treated with IL-1 inhibition. J Allergy Clin Immunol 2019;144:1122-5.e6).

[0059] In one embodiment, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation has a serum C-reactive protein (CRP) level higher than 20 mg / L. In another embodiment, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation has a serum ferritin level higher than 600 μg / L. In a further preferred embodiment, the subject requiring treatment for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation has a serum CRP level higher than 20 mg / L and a serum ferritin level higher than 600 μg / L.

[0060] In one embodiment of the foregoing aspects, treatment with the bispecific antibody targeting both IL-1β and IL-18 prolonged patient life compared to standard of care (SoC). In another embodiment, after 14 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced serum CRP and / or serum ferritin levels compared to SoC. In yet another embodiment, after 28 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced serum CRP and / or serum ferritin levels compared to SoC.

[0061] In another embodiment, after 7 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced the patient's serum CRP level compared to the standard of care. In another embodiment, after 14 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced the patient's serum CRP level compared to the standard of care. In another embodiment, after 28 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced the patient's serum CRP level compared to the standard of care. In another embodiment, after 7 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced the patient's serum ferritin level compared to the standard of care. In another embodiment, after 14 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced the patient's serum ferritin level compared to the standard of care. In another embodiment, after 28 days of treatment, treatment with the bispecific antibody targeting both IL-1β and IL-18 reduced patients’ serum ferritin levels compared to the standard of care.

[0062] In one embodiment, this document provides a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for use in subjects with autoinflammatory or disease-related by XIAP deficiency or CDC42 mutation to reduce serum C-reactive protein (CRP) levels. In some embodiments, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for manufacturing a medicament for reducing serum C-reactive protein (CRP) levels in subjects with autoinflammatory or disease-related by XIAP deficiency or CDC42 mutation. In some embodiments, the subject's serum CRP level is reduced by at least 1 mg / L, at least 2 mg / L, at least 3 mg / L, at least 4 mg / L, or at least 5 mg / L. In some embodiments, subjects with autoinflammatory or inflammatory diseases based on XIAP deficiency or CDC42 mutation who have received a bispecific antibody targeting both IL-1β and IL-18 (e.g., bbmAb) have serum CRP levels that are at least 10%, at least 20%, at least 30%, at least 40%, or at least 50% lower than baseline compared to patients who have not received the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction in serum CRP levels occurs 2, 3, 4, 5, 6, or 7 days after administration of the bispecific antibody targeting both IL-1β and IL-18 (e.g., bbmAb). In some embodiments, the reduction in serum CRP levels occurs 14 or 28 days after administration of the bispecific antibody targeting both IL-1β and IL-18 (e.g., bbmAb).

[0063] In one embodiment, this document provides a method for reducing serum ferritin levels in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, the method comprising administering to the subject a therapeutically effective amount of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18. In one embodiment, this document provides a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for use in reducing serum ferritin levels in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In some embodiments, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for manufacturing a medicament for reducing serum ferritin levels in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In some embodiments, the subject's serum ferritin level is reduced by at least 100 ng / L, at least 200 ng / L, at least 300 ng / L, at least 400 ng / L, or at least 500 ng / L. In some embodiments, subjects who received a bispecific antibody targeting both IL-1β and IL-18 (e.g., bbbmAb) had a reduction in serum ferritin levels of at least 10%, at least 20%, at least 30%, at least 40%, or at least 50% from baseline compared to patients who did not receive the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction in serum ferritin levels occurred 2, 3, 4, 5, 6, or 7 days after administration of the bispecific antibody targeting both IL-1β and IL-18 (e.g., bbbmAb). In some embodiments, the reduction in serum ferritin levels occurred 14 or 28 days after administration of the bispecific antibody targeting both IL-1β and IL-18 (e.g., bbbmAb).

[0064] In one embodiment, this document provides a method for reducing serum levels of a biomarker selected from the group consisting of CXCL9, CXCL10 (IP-10), IL-6, and sIL2R in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, the method comprising administering to the subject a therapeutically effective amount of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18. In another embodiment, this document provides a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for use in reducing serum levels of a biomarker selected from the group consisting of CXCL9, CXCL10 (IP-10), IL-6, and sIL2R in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In some embodiments, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the manufacture of a medicament intended to reduce serum levels of a group of biomarkers selected from CXCL9, CXCL10 (IP-10), IL-6, and sIL2R in subjects with an autoinflammatory or disease-associated with XIAP deficiency or CDC42 mutation. In some embodiments, subjects who have received a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 have serum levels of biomarkers selected from the group of CXCL9, CXCL10 (IP-10), IL-6, and sIL2R that are at least 10%, at least 20%, at least 30%, at least 40%, or at least 50% lower than baseline compared to patients who have not received the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction in serum levels of biomarkers selected from the group consisting of CXCL9, CXCL10 (IP-10), IL-6, and sIL2R occurs 2, 3, 4, 5, 6, or 7 days after administration of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18.

[0065] In an eighth aspect of this disclosure, a method is provided for preventing fever or reducing the incidence or severity of fever in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, the method comprising administering to the subject a therapeutically effective amount of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18. In one embodiment, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for preventing fever or reducing the incidence or severity of fever in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In some embodiments, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for manufacturing a medicament for preventing fever or reducing the incidence or severity of fever in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation.

[0066] In a ninth aspect of this disclosure, a method is provided for preventing diarrhea or reducing the incidence or severity of diarrhea in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, the method comprising administering to the subject a therapeutically effective amount of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18. In one embodiment, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for preventing diarrhea or reducing the incidence or severity of diarrhea in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In some embodiments, this document provides the use of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for manufacturing a medicament for preventing diarrhea or reducing the incidence or severity of diarrhea in a subject suffering from an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation.

[0067] Various embodiments of the foregoing aspects may be beneficially combined with other treatments for autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutations in subjects in need. For example, such therapies may be any known treatment for a disease, disorder, symptom, or syndrome being treated. As a set of non-limiting examples, the at least one additional therapeutic agent may be selected from a list comprising: nonsteroidal anti-inflammatory drugs, cyclosporine, glucocorticoids, IL-18 binding protein (IL-18 BP), and combinations thereof.

[0068] In some cases, treatment with the bispecific antibody described herein reduces or eliminates the maintenance dose of glucocorticoids required for the treatment subject. In some cases, the treatment involves administering the bispecific antibody described herein and discontinuing the dose of glucocorticoids. In some cases, the treatment involves administering the bispecific antibody described herein and discontinuing the dose of cyclosporine.

[0069] In some cases, the treatment includes administration of the bispecific antibody described herein and a gradual reduction in the dose of an additional therapeutic agent. In some cases, the treatment includes administration of the bispecific antibody described herein and a gradual reduction in the dose of glucocorticoids. In some embodiments, the treatment includes reducing, gradually reducing, or discontinuing the administration of glucocorticoids in subjects receiving both glucocorticoids and bispecific antibodies. In some cases, the treatment includes reducing or gradually reducing the dose to less than or equal to a dose of prednisone equivalent to 0.2 mg / kg / day. In some cases, the treatment includes reducing or gradually reducing the dose to less than or equal to a dose of prednisone equivalent to 0.2 mg / kg / day. In some cases, after a gradual reduction to a dose of prednisone equivalent to 0.2 mg / kg / day, the subject receiving the treatment maintains at least a partial or complete response to the bispecific antibody for at least 2 weeks. In some embodiments, the treatment includes reducing, gradually reducing, or discontinuing the administration of glucocorticoids and discontinuing cyclosporine in subjects receiving bispecific antibodies, glucocorticoids, and cyclosporine.

[0070] In some embodiments, the subject in need has been or is currently receiving cyclosporine, anti-TNFα, corticosteroids, anti-IFNγ, anti-IL-1β, or anti-IL-18 therapy, or combinations thereof. In some embodiments, the subject in need has not achieved adequate control of XIAP deficiency- or CDC42 mutation-based autoinflammatory or disease by administration of cyclosporine, anti-TNFα, corticosteroids, anti-IFNγ, anti-IL-1β, or anti-IL-18 therapy, or combinations thereof. In some embodiments, the subject in need's XIAP deficiency- or CDC42 mutation-based autoinflammatory or disease is resistant to cyclosporine, anti-TNFα, corticosteroids, anti-IFNγ, anti-IL-1β therapy (as a single therapy), or combinations thereof. In some cases, the XIAP deficiency- or CDC42 mutation-based autoinflammatory or disease is unresponsive to cyclosporine, anti-TNFα, corticosteroids, anti-IFNγ, anti-IL-1β, or anti-IL-18 therapy, or combinations thereof. In some cases, resistance and / or inadequate control is indicated by failure to achieve a PPGA score of less than 2 as described herein. In some cases, resistance and / or inadequate control is indicated by failure to achieve a PPGA score of less than 1 as described herein. In some cases, non-response is indicated by failure to reduce the PPGA score as described herein.

[0071] In some embodiments, after 7, 14, 21, or 29 days of treatment, the treatment reduces or prevents the occurrence of disease flare-ups in subjects in need compared to the standard of care (SoC). In some embodiments, reducing or preventing the occurrence of disease flare-ups in subjects means reducing or preventing them for at least about one, two, or three weeks, or about four weeks.

[0072] In some embodiments, the treatment reduces or prevents the occurrence of one or more MAS characteristics in a subject, wherein these MAS characteristics are selected from the group consisting of fever, rash, tachycardia, cytopenia, liver dysfunction, and coagulopathy. In some cases, the treatment reduces or prevents enterocolitis in the subject. In some cases, the treatment reduces or prevents severe, refractory neonatal diarrhea in the subject. In some cases, the treatment reverses the patient's gastrointestinal pathology.

[0073] In some embodiments, the treatment increases the time to first onset, for example, by about (or at least about) one week, one month, two months, three months, six months, or one year. In some embodiments, the treatment further includes inducing serological remission in the patient. Serological remission can be indicated by adequately suppressing serum IL-18 to undetectable levels or to levels found in healthy individuals (e.g., below 500 pg / mL, below 1000 pg / mL, or below 5000 pg / mL).

[0074] In some embodiments, the patient has a XIAP mutation in the XIAP gene that results in a truncated or dysfunctional XIAP protein, wherein the mutation may be a point mutation, deletion (small or large exon deletion) or insertion, resulting in nonsense and missense mutations, intron or frameshift mutations.

[0075] In some embodiments, the patient has a CDC42 mutation in the CDC42 gene that results in a truncated or dysfunctional CDC42 protein, wherein the mutation can be a point mutation, deletion (small or large exon deletion), or insertion, resulting in nonsense and missense mutations, intron or frameshift mutations. Specifically, the mutation can be an amino acid substitution affecting adjacent residues (Tyr64, Arg66, and Arg68) within the N-terminal α-helix (residues Ile21 and Tyr23) or the switch II motif, or in the fourth β-chain (Cys81 and Ser83), or in the C-terminus (Ala159 and Glu171), or mutations such as p.R186C, p.C188Y, and p. disclosed in Gernez Y et al., J Allergy Clin Immunol [Journal of Allergy and Clinical Immunology] 2019;144:1122-5.e6. 192Cext twenty four.

[0076] In some embodiments, the patient is a newborn. In some embodiments, the patient is a child (< 18 years old). In some embodiments, the patient is less than 1 year old. In some embodiments, the patient is between 1 week and 1 year old. In some embodiments, the patient is between 1 month and 1 year old. In some embodiments, the patient is at least 1 week old. In some embodiments, the patient is at least 1 month old. Attached Figure Description

[0077] Figure 1 This is a schematic diagram of a treatment regimen used in clinical studies of treating autoinflammatory or disease-based conditions with bispecific antibodies (e.g., bbmAb) that target both IL-1β and IL-18.

[0078] Figure 2 is a schematic diagram of the treatment plans for Phase 1 (2A), Phase 2 (2B), and Phase 3 (2C).

[0079] 2A: Phase 1 is an open-label treatment phase designed to identify responders to bbmAb treatment and allow these patients to gradually reduce their glucocorticoid dose and / or discontinue cyclosporine treatment. Phase 1 is divided into three sub-parts (Phase 1a, 1b, and 1c).

[0080] 2B: Phase 2 consists of a 24-week placebo-controlled, double-blind, randomized withdrawal phase, primarily evaluating the efficacy of bbbab compared to placebo. At the start of Phase 2, bbbab responders (those who achieved a complete response to treatment at the end of open-label treatment in Phase 1) will be randomized 1:1 to either bbbab treatment (i.e., continued use of 10 mg / kg) or placebo. The first planned blinded dose after randomization in Phase 2 will be given 2 weeks after the last dose in Phase 1c, and doses will continue every 2 weeks until disease onset or 24 weeks have elapsed in Phase 2.

[0081] 2C: Phase 3 consists of a 3-year long-term safety profile, with treatment using open-label bbmAb (10 mg / kg). For Group 1, the first scheduled dose of Phase 3 will be administered 2 weeks after the last dose of Phase 1 or Phase 2, and will continue to be administered at intervals of at least 2 weeks. For Group 2, scheduled dosing will be initiated after completion of screening and baseline visits and confirmation that all group-specific eligibility criteria have been met. Dosing in Phase 3 will continue approximately every 2 weeks, and the protocol evaluation will have a reduced visit rate, as outlined in the evaluation schedule.

[0082] Figure 3 The curves depict the changes in serum bbmAb concentration over time from the first human healthy volunteer study, as predicted (line) and as measured (data points).

[0083] Figure 4 The graph depicts the predicted free IL-18 and IL-1β concentrations after administration of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 at 10 mg / kGi.v. relative to recombinant IL-18 BP and canagenzumab treatment (grey shading). The dashed line indicates the lower limit of quantitation (LLOQ). Detailed Implementation

[0084] X-linked apoptosis inhibitor protein deficiency (XIAP or XIAP deficiency) is a rare, inherited immunodeficiency that occurs almost exclusively in young men (Rigaud et al. 2006). XIAP exhibits pleiotropic functions in cell survival, innate immunity, and inflammation. In particular, in addition to its role in regulating caspase activity, XIAP is considered an essential regulator of NLRP3 inflammasome activation (Miyazawa and Wado 2022). Key features of XIAP deficiency include inflammatory bowel disease, typically presenting as abdominal pain and diarrhea, relapsing fever, splenomegaly, and hemophagocytic lymphohistiocytosis (HLH). The latter manifestation is usually triggered by infection with Epstein-Barr virus (EBV). Once diagnosed, the initial goal of treatment is to suppress inflammation, typically using corticosteroids and biologics. Inflammatory bowel disease can be treated with standard immunosuppressive drugs, although response rates are generally low. EBV infection can be treated with specific antiviral therapy, and in some cases, immunoglobulin therapy. Currently, the only potentially curative treatment for XIAP deficiency is hematopoietic stem cell transplantation (HSCT). However, due to the characteristics of the underlying disease, low-intensity conditioning regimens are required, and HSCT results are generally worse than normal for patients of this age. Despite an increase in life expectancy in patients with XIAP over the past decade, the condition still presents with significant morbidity and mortality (Mudde et al. 2021). Loss of XIAP leads to dysregulation of inflammasome activity, excessive production of inflammasome-activated cytokines (including IL-18), and cell death (Wada et al. 2014). In XIAP-deficient patients, IL-18 levels rise sharply during HLH episodes, exceeding the blocking capacity of IL-18BP, and free IL-18 levels are detected in peripheral blood. In one published case, a combined approach of IL-1 and IL-18 blockade has been used to alleviate clinical disease activity, including HLH and gastrointestinal symptoms (Geerlinks et al. 2022). An 8-year-old male presented with relapsing fever from infancy, followed by abdominal pain and diarrhea associated with elevated transaminases (transaminitis). The patient exhibited hypersensitivity to anakinase and was unable to discontinue steroids despite multiple immunotherapies, including canagencinumab, tocilizumab, colchicine, etanercept, infliximab, mesalazine, rituximab, and vedozhumab. After treatment with recombinant IL-18 BP tadalafil-α, the patient was able to discontinue oral steroids and experienced almost complete symptom remission over more than two years of treatment, except for one episode of fever and diarrhea (Geerlinks et al. 2022).

[0085] Cell division control protein 42 (CDC42) belongs to the Rho family of small monomeric GTPases (Heasman et al. 2008). It influences multiple cellular processes, including cell division and migration, and regulates the nervous, immune, and hematopoietic systems (Melendez et al. 2011). Pathogenic mutations in CDC42 can have a variety of effects on protein function, leading to increasingly prominent and diverse disease phenotypes centered on neurodevelopment, hematopoiesis, and immune responses (Takenouchi et al. 2015, Asiri et al. 2021, Coppola et al. 2022). In particular, aberrant palmitoylation of CDC42R186C has been shown to trap CDC42 within the Golgi apparatus, which may induce excessive activation of the heat protein inflammasome and significant increases in IL-1β and IL-18 (Coppola et al. 2022). In four pediatric patients, a neonatal missense mutation at the C-terminus (p.R186C) has been demonstrated to affect CDC42 localization and cause a specific set of neonatal episodic cytopenia, autoinflammation, and recurrent HLH (Lam et al. 2019). The presence of chronically elevated serum IL-18 (even after a clinical response to anti-IL-1 therapy) suggests that IL-18 is a promising therapeutic target for CDC42 C-terminal disease (Lam et al. 2019). In a single surviving patient in a case series of four patients identified with the CDC42R186C mutation, ongoing HLH was associated with significant and persistent elevations in IL18. Pharmacological interventions with pulsed glucocorticoids, cyclosporine A, and high-dose anakinin (10 mg / kg / day) failed to prevent ongoing HLH, and the attacks responded only to imarumab (a monoclonal antibody targeting IFN-γ, a downstream mediator of IL18 activity). The patient eventually underwent a curative hematopoietic stem cell transplant, after which IL18 levels normalized and the patient did not experience any further HLH episodes (Lam et al. 2019).

[0086] As used in this article, the general term "autoinflammatory or disease based on XIAP deficiency or CDC42 mutation" will be used because it best reflects the underlying cause of the disease.

[0087] Autoinflammatory diseases based on XIAP deficiency or CDC42 mutations refer to / can cause the following clinical symptoms: hypogammaglobulinemia, cytopenia, inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, syndromic thrombocytopenia, megaloblastic thrombocytopenia, hematopoietic insufficiency, autoinflammation, syndromic immunodeficiency, recurrent hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome, or AIFEC.

[0088] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is hypogammaglobulinemia.

[0089] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is cytopenia.

[0090] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is inflammatory bowel disease.

[0091] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptoms are abdominal pain and diarrhea.

[0092] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is relapsing fever.

[0093] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is splenomegaly.

[0094] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is hemophagocytic lymphohistiocytosis.

[0095] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is neonatal episodic cytopenia.

[0096] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is syndromic thrombocytopenia.

[0097] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is megaloblastic thrombocytopenia.

[0098] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is hematopoietic insufficiency.

[0099] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of autoinflammatory or disease-based on XIAP deficiency, wherein the clinical symptom is autoinflammatory.

[0100] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is a syndromic immunodeficiency.

[0101] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is recurrent hemophagocytic lymphohistiocytosis (HLH).

[0102] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is macrophage activation syndrome (MAS).

[0103] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on XIAP deficiency, wherein the clinical symptom is AIFEC.

[0104] Autoinflammatory diseases or conditions based on CDC42 mutations refer to / can cause the following clinical symptoms: hypogammaglobulinemia, cytopenia, inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, syndromic thrombocytopenia, megaloblastic thrombocytopenia, hematopoietic insufficiency, autoinflammation, syndromic immunodeficiency, or relapsing hemophagocytic lymphohistiocytosis (HLH). In one embodiment, this disclosure relates to the use or method of using a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 to treat autoinflammatory diseases or conditions based on CDC42 mutations, wherein the clinical symptom is hypogammaglobulinemia.

[0105] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is a decrease in blood cells.

[0106] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of autoinflammatory or disease-based on CDC42 mutations, wherein the clinical symptom is inflammatory bowel disease.

[0107] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptoms are abdominal pain and diarrhea.

[0108] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is relapsing fever.

[0109] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is splenomegaly.

[0110] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is hemophagocytic lymphohistiocytosis.

[0111] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is neonatal episodic cytopenia.

[0112] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is syndromic thrombocytopenia.

[0113] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is megaloblastic thrombocytopenia.

[0114] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is hematopoietic insufficiency.

[0115] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of autoinflammatory or disease-based on CDC42 mutations, wherein the clinical symptom is autoinflammatory.

[0116] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of autoinflammatory or disease-based on CDC42 mutations, wherein the clinical symptom is syndromic immunodeficiency.

[0117] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is recurrent hemophagocytic lymphohistiocytosis (HLH).

[0118] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of autoinflammatory or disease-associated with a CDC42 mutation, wherein the clinical symptom is macrophage activation syndrome (MAS).

[0119] In one embodiment, this disclosure relates to the use or method of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 for the treatment of an autoinflammatory or disease based on a CDC42 mutation, wherein the clinical symptom is AIFEC.

[0120] In subjects (e.g., infants), it is hypothesized that early treatment of MAS-like features and enterocolitis can prevent disease progression to irreversible end-organ damage, which typically leads to fatal outcomes (Romberg et al. 2014, Moghaddas et al. 2018). In critically ill children, the disease has been found to be resistant to combination or single cyclosporine, anti-TNFα therapy, systemic glucocorticoids, and anti-IL-1β therapies. Currently, apart from supportive medical care and immunosuppression with limited benefit in this population, there are no approved therapeutics that directly and specifically target the underlying IL-1β and IL-18-driven autoinflammatory processes to improve overall clinical outcomes in patients with autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutations.

[0121] This article describes a method for treating or preventing autoinflammatory or disease-related to XIAP deficiency or CDC42 mutation by administering an effective amount of a bispecific antibody (e.g., bbmAb) or a functional fragment thereof that simultaneously targets both IL-1β and IL-18 to a subject in need. Therefore, in one aspect, a method for preventing or treating AIFEC is provided, comprising administering an effective amount of a bispecific antibody (e.g., bbmAb) that simultaneously targets both IL-1β and IL-18 to a subject in need.

[0122] This disclosure relates to bispecific monoclonal antibodies (bbmAbs) or variants thereof (e.g., bispecific antibodies that simultaneously target both IL-1β and IL-18, e.g., bbmAbs) for use in treating patients with XIAP-deficient or CDC42-mutant autoinflammatory or disease-related with excessively elevated IL-1β and / or IL-18. This disclosure also relates to methods, treatment regimens, uses, kits, and therapies for treating XIAP-deficient or CDC42-mutant autoinflammatory or disease-related with XIAP-deficient or CDC42-mutant autoinflammatory or disease-related by employing bispecific antibodies that simultaneously target both IL-1β and IL-18.

[0123] The data described herein indicate that the combination of simultaneous neutralization of IL-1β and IL-18 more potently attenuates the production of IFN-γ (and other pro-inflammatory cytokines) compared to the individual neutralization of IL-1β or IL-18 by anti-IL-1 or anti-IL-18 mAb. Therefore, this disclosure is based in particular on the unexpected finding that certain antibodies that simultaneously neutralize IL-1β and IL-18 more potently attenuate the production of IFN-γ (and other pro-inflammatory cytokines) compared to the individual neutralization of IL-1β or IL-18, which the inventors believe is an effective treatment for patients with (i) autoinflammatory or disease based on XIAP deficiency, or (ii) autoinflammatory or disease based on CDC42 mutation, or (iii) AIFEC.

[0124] Furthermore, the inventors of this invention hypothesize that treatment with an antibody that simultaneously neutralizes IL-1β and IL-18 (e.g., bbmAb) could allow for a significantly reduced dosing frequency in patients with autoinflammatory or disease-based conditions such as XIAP deficiency or CDC42 mutations, compared to more complex investigational combinations requiring anti-IL-1β (canagenumab every two weeks or anaphylaxis once daily) and possibly glucocorticoids, cyclosporine, and IL-18BP (every two days).

[0125] 1. Definition

[0126] For the purposes of interpreting this specification, the following definitions will apply, and where appropriate, terms used in the singular will also include the plural form, and vice versa. Additional definitions are set forth throughout the specific embodiments. All references, publications, patents, and database accessions, including those from GenBank and OMIM and their contents, are incorporated herein by reference in their entirety and for all purposes.

[0127] In the context of treating autoinflammatory or other diseases based on XIAP deficiency or CDC42 mutations, the term "episode" refers to:

[0128] • As described in this article, in the physician's overall assessment of disease activity, the increase in disease activity ranges from none or very slight to greater than very slight;

[0129] • In treated patients, serum ferritin and / or C-reactive protein (CRP) were increased by 60% from normalization levels, where normalization levels indicate very mild or absent disease activity of autoinflammation or disease based on XIAP deficiency or CDC42 mutation (e.g., CRP < 20 mg / L; ferritin < 600 ng / L); or

[0130] • Increased serum ferritin levels > 2500 ng / mL and / or elevated CRP levels > 20 mg / mL.

[0131] The term "IL-18" is a synonym for IL-18 polypeptide, interleukin-18 polypeptide, IFN-γ inducible factor, or interferon-γ inducible factor or INF-γ inducible factor. Unless otherwise stated, the term "IL-18" refers to human IL-18. IL-18 is well known to those skilled in the art and is available, for example, from MBL® International Corporation under product number #B001-5. Throughout this specification, the term IL-18 is used interchangeably to refer to pre-IL-18 (the precursor of mature IL-18 before protease cleavage) and mature IL-18 (after protease cleavage), unless specifically stated to mean either pre- or mature form.

[0132] The terms “IL-1β” or “IL-1b” are synonyms for IL-1β polypeptide and interleukin-1β polypeptide. Unless otherwise stated, the term “IL-1β” refers to human IL-1β. IL-1β is well known to those skilled in the art and is available, for example, from Sino Biological under product number #10139-HNAE-5.

[0133] The term "antibody" refers to a complete immunoglobulin or a functional fragment thereof. Naturally occurring antibodies typically consist of tetramers, which are usually composed of at least two heavy (H) chains and at least two light (L) chains. Each heavy chain consists of a heavy chain variable region (abbreviated as VH) and a heavy chain constant region (usually composed of three domains (CH1, CH2, and CH3)). The heavy chain can belong to any isotype, including IgG (IgG1, IgG2, IgG3, and IgG4 subtypes), IgA (IgA1 and IgA2 subtypes), IgM, and IgE. Each light chain consists of a light chain variable region (abbreviated as VL) and a light chain constant region (CL). The light chains include κ (kappa) chains and λ (lambda) chains. The heavy and light chain variable regions are typically responsible for antigen recognition, while the heavy and light chain constant regions mediate the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system. The VH and VL regions can be further subdivided into hypervariable regions called complementarity-determining regions (CDRs), interspersed with more conserved regions called framework regions (FRs). Each VH and VL consists of three CDRs and four FRs arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with the antigen.

[0134] As used herein, the term “antigen-binding portion” (or simply “antigen portion”) of an antibody refers to a full-length antibody or one or more fragments of an antibody that retain the ability to specifically bind to the IL-18 or IL-1β antigen. Fragments of full-length antibodies have been shown to perform the antigen-binding function of the antibody. Examples of binding fragments encompassed within the term “antigen-binding portion” of an antibody include Fab fragments, which are monovalent fragments consisting of VL, VH, CL, and CH1 domains; F(ab)2 fragments, which are fragments containing two monovalent Fab fragments connected by a disulfide bridge in the hinge region; Fd fragments consisting of VH and CH1 domains; Fv fragments consisting of the VL and VH domains of a single arm of the antibody; dAb fragments (Ward et al., (1989) Nature; 341:544-546), which consist of a VH domain; and separate complementarity-determining regions (CDRs).

[0135] Furthermore, although the two domains VL and VH of the Fv fragment are encoded by separate genes, these two domains can be linked using recombination methods via flexible linkers that enable them to form a single protein chain, where the VL and VH regions pair to form a monovalent molecule (referred to as a single-chain Fv (scFv); see, for example, Bird et al., (1988) Science 242:423-426; and Huston et al., (1988) Proc Natl Acad Sc. 85:5879-5883). Such single-chain antibodies are also intended to be encompassed within the term "antigen-binding portion" of antibody. These antibody fragments are obtained using conventional techniques known to those skilled in the art and are screened for efficacy in the same manner as intact antibodies.

[0136] Throughout the instructions, the term "isolated" means that the immunoglobulins, antibodies, or polynucleotides (as the case may be) exist in a physical environment different from their natural environment.

[0137] Throughout this specification, the complementary determination region (“CDR”) is defined according to the Kabat definition, unless otherwise specified that the CDR is defined according to another definition. The precise amino acid sequence boundaries of a given CDR can be determined using any of many well-known schemes, including those described in the following literature: Kabat et al. (1991), “Sequences of Proteins of Immunological Interest”, 5th edition, National Institutes of Health, Public Health Service, Bethesda, MD (“Kabat” numbering scheme); Al-Lazikani et al. (1997) JMB 273, 927-948 (“Chothia” numbering scheme); and ImMunoGenTics (IMGT) numbering (Lefranc, M.-P., The Immunologist, 7, 132-136 (1999); Lefranc, M.-P. et al., Dev. Comp. Immunol). [Developmental Immunology and Comparative Immunology], 27, 55-77 (2003) (“IMGT” numbering scheme). For example, for the classical form, according to Kabart, the CDR amino acid residues in the heavy chain variable domain (VH) are numbered 31-35 (HCDR1), 50-65 (HCDR2), and 95-102 (HCDR3); and the CDR amino acid residues in the light chain variable domain (VL) are numbered 24-34 (LCDR1), 50-56 (LCDR2), and 89-97 (LCDR3). According to Josiah, the CDR amino acids in VH are numbered 26-32 (HCDR1), 52-65 (HCDR2), 50-65 (HCDR2), and 95-102 (HCDR3). -56 (HCDR2) and 95-102 (HCDR3); and number the amino acid residues in VL as 26-32 (LCDR1), 50-52 (LCDR2) and 91-96 (LCDR3). By combining the CDR definition of Cabat and Josiah, the CDR consists of amino acid residues 26-35 (HCDR1), 50-65 (HCDR2) and 95-102 (HCDR3) in human VH and amino acid residues 24-34 (LCDR1), 50-56 (LCDR2) and 89-97 (LCDR3) in human VL.According to IMGT, the CDR amino acid residues in VH are numbered approximately 26-35 (CDR1), 51-57 (CDR2), and 93-102 (CDR3), and the CDR amino acid residues in VL are numbered approximately 27-32 (CDR1), 50-52 (CDR2), and 89-97 (CDR3) (according to "Carbart" numbering). The CDR regions of the antibody can be determined using the IMGT / DomainGapAlign procedure according to IMGT.

[0138] By convention, the CDR regions in the heavy chain are usually referred to as H-CDR1, H-CDR2, and H-CDR3, while the CDR regions in the light chain are usually referred to as L-CDR1, LCDR2, and L-CDR3. They are numbered sequentially from the amino terminus to the carboxyl terminus.

[0139] As used herein, the terms "monoclonal antibody" or "monoclonal antibody composition" refer to formulations of antibody molecules that are single-molecule compositions. Monoclonal antibody compositions exhibit single binding specificity and affinity for a specific epitope.

[0140] As used herein, the term "human antibody" is intended to include antibodies with variable regions having both the frame region and the CDR region derived from human-derived sequences. Furthermore, if the antibody contains a constant region, the constant region is also derived from human sequences such as germline human sequences or mutant forms of germline human sequences, or antibodies containing a common frame sequence derived from human frame sequence analysis, for example, as described in Knappik et al., (2000) J Mol Biol [Journal of Molecular Biology]; 296:57-86).

[0141] Human antibodies of the present invention may include amino acid residues not encoded by human sequences (e.g., mutations introduced by random mutagenesis or site-specific mutagenesis in vitro or by somatic mutation in vivo). However, as used herein, the term "human antibody" is not intended to include antibodies in which CDR sequences derived from another mammalian species (such as mice) have been grafted onto human frame sequences.

[0142] The term "human monoclonal antibody" refers to an antibody with a variable region that exhibits single binding specificity, where both the frame region and the CDR region are derived from human sequences.

[0143] As used herein, the term "recombinant human antibody" includes all human antibodies prepared, expressed, generated, or isolated in a recombinant manner, such as antibodies isolated from transgenic or transchromosomally modified animals (e.g., mice) or hybridomas prepared therefrom, antibodies isolated from host cells transformed to express human antibodies (e.g., from transfected tumors), antibodies isolated from a library of recombinant human antibodies, and antibodies prepared, expressed, generated, or isolated by any other means involving splicing of all or part of the human immunoglobulin gene. Such recombinant human antibodies have variable regions, wherein both the framework region and the CDR region are derived from human germline immunoglobulin sequences. However, in some embodiments, such recombinant human antibodies may be mutagenized in vitro (or, when using animals with transgenic human Ig sequences, in vivo somatic cell mutagenesis), and thus the amino acid sequences of the VH and VL regions of the recombinant antibody are derived from and associated with human germline VH and VL sequences that may not be naturally present in a human antibody germline library in vivo.

[0144] The phrases “antibody that recognizes an antigen” and “antibody that is specific to an antigen” are used interchangeably with the term “antibody that binds specifically to an antigen” in this document.

[0145] As used in this article, the term "specifically binding to IL-18" refers to a binding molecule with a K+ concentration of 100 nM or less, 10 nM or less, or 1 nM or less. D A binding molecule that binds to human IL-18.

[0146] As used in this article, the term "specifically binding IL-1β" refers to a binding molecule with a K+ concentration of 100 nM or less, 10 nM or less, or 1 nM or less. D A binding molecule that binds to human IL-1β.

[0147] As used herein, the term "antagonist" is intended to refer to a binding molecule that inhibits signal transduction activity in the presence of an activating compound. For example, in the case of IL-18, an IL-18 antagonist would be a binding molecule that inhibits signal transduction activity in human blood cells in the presence of IL-18 in a human cell assay (such as an IL-18-dependent interferon-γ (IFN-γ) production assay). Examples of IL-18-dependent IFN-γ production assays in human blood cells are described in more detail in the following examples.

[0148] The term bivalent bispecific antibody or multiple bivalent bispecific antibodies refers to antibodies that bind to two different targets (such as IL-18 and IL-1β). Typically, the bivalent bispecific antibody binds to each target monovalently.

[0149] Bispecific antibodies are “heterodimers,” meaning that one part comes from a first antibody that is specific to a first target, and the other part comes from a second antibody that is specific to a second target. “Heterodimerization modification” is a modification of one or both parts of the antibody that forms a heterodimeric bispecific antibody, designed to promote such formation. Examples of heterodimerization modifications of the Fc domains of the two IgG1 moieties designed to form bispecific antibodies are “mortars” with large amino acid (aa) side chains (S354C, T366W) in the first heavy chain and “mortars” with small amino acid side chains (Y349C, T366S, L368A, Y407V) introduced in the second heavy chain, as well as additional disulfide bridges connecting the two heavy chains in the CH3 region (Merchant et al., Nat. Biotechnol. [Nature Biotechnology], 16:677-681 (1998), p. 678, Table 1).

[0150] As used in this article, the term "K" D "This refers to the dissociation constant, which is obtained from K..." d With K a The ratio (i.e., K) d / K a The Kc of the antibody is expressed as a molar concentration (M). The Kc of the antibody can be determined using methods well-established in the art. D Value. K value used to determine the antibody. D The method involves using surface plasmon resonance, such as the Biacore® system.

[0151] As used in this article, the term "affinity" refers to the strength of the interaction between the binding molecule and the antigen at a single antigenic site.

[0152] As used in this article, the term "high affinity" for antibodies refers to antibodies with a KD of 1 nM or less against the target antigen.

[0153] As used herein, the term “subject” includes any subject who receives a bispecific antibody as described herein. The term “subject” may additionally or alternatively include any subject who has, for example, symptoms of autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, or inflammasome lesions based on XIAP deficiency or CDC42 mutation, or AIFEC as defined above, or who is at risk of, for example, autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, or inflammasome lesions based on XIAP deficiency or CDC42 mutation, or AIFEC as defined above.

[0154] The term "immune response" refers to the action of, for example, lymphocytes, antigen-presenting cells, phagocytes, granulocytes, and soluble macromolecules (including antibodies, cytokines, and complement) produced by the aforementioned cells or liver, resulting in selective damage, destruction, or elimination from the body of invading pathogens, pathogen-infected cells or tissues, cancer cells (or, in the case of autoimmunity or pathological inflammation, normal human cells or tissues).

[0155] Nucleotides in "polynucleotides" or "nucleic acids" can contain modifications, including base modifications such as bromouridine and inosine derivatives; and ribose modifications such as thiophosphates, dithiophosphates, selenophosphates, diselenophosphates, aniline thiophosphates, acylaniline phosphates, and aminophosphates.

[0156] The terms "treat," "treating," and "prevent" encompass therapeutic treatment, preventative treatment, and applications that reduce a subject's risk of developing a disorder or other risk factors. Treatment does not require a complete cure for the disorder and covers alleviating symptoms or underlying risk factors.

[0157] The term "treatment or prevention" includes the administration of a compound, such as a bispecific antibody targeting both IL-1β and IL-18 (e.g., bbmAb), optionally in combination with at least one additional therapeutic agent, to prevent or delay the onset of symptoms, complications, or biochemical markers of a disease, condition, disorder, or syndrome (e.g., autoinflammatory or disease based on XIAP deficiency or CDC42 mutation, AIFEC), to alleviate symptoms, or to prevent or inhibit the further development of the disease, condition, disorder, or syndrome. Treatment can be preventative (to prevent or delay the onset of a disease, condition, disorder, or syndrome, or to prevent the manifestation of its clinical or subclinical symptoms) or therapeutic suppression or relief of symptoms after the onset of a disease, condition, disorder, or syndrome.

[0158] As used herein, the term “prevent, preventing, or prevention” in relation to a disease, condition, disorder, or syndrome refers to preventive treatment of a subject who is at risk of developing a condition (e.g., a specific disease, condition, disorder, or syndrome or its clinical symptoms, such as an autoinflammatory or disease based on XIAP deficiency or CDC42 mutation or AIFEC) in order to reduce the likelihood that the subject will develop the condition.

[0159] The term "treatment" refers to therapeutic treatment and preventive or preventative measures in which the aim is to improve a disease, symptom, disorder, or syndrome (i.e., to slow, stop, or reduce the development of the disease or at least one of its clinical symptoms) by reducing or improving at least one physical parameter (including those that the patient may not be able to discern). The term "treatment" also refers to regulating a disease or disorder physically (e.g., stabilization of identifiable symptoms), physiologically (e.g., stabilization of physical parameters), or in both aspects, and / or preventing or delaying the occurrence, development, or progression of a disease or disorder.

[0160] For example, "treatment of autoinflammatory or disease-associated with XIAP deficiency or CDC42 mutation" can refer to improving, alleviating, or modulating at least one of the symptoms or pathological features associated with autoinflammatory or disease-associated with XIAP deficiency or CDC42 mutation; for example, elevated serum inflammatory markers (such as one or more of serum CRP, serum ferritin, serum IL-18, serum total IL-18, serum free IL-18, serum IL-1β, serum total IL-1β, and serum free IL-1β), fever, diarrhea, rash, tachycardia, cytopenia, liver dysfunction, and / or coagulopathy; for example, it can refer to slowing the progression, reducing, or stopping the progression of autoinflammatory or disease-associated with XIAP deficiency or CDC42 mutation. 42. At least one of the following symptoms or pathological features associated with autoinflammatory or disease-related disorders: elevated serum inflammatory markers (such as one or more of serum CRP, serum ferritin, and serum IL-18), fever, diarrhea, rash, tachycardia, cytopenia, liver dysfunction and / or coagulopathy or hypogammaglobulinemia, cytopenia, inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, syndromic thrombocytopenia, megaloblastic thrombocytopenia, hematopoietic insufficiency, autoinflammatory disease, syndromic immunodeficiency, or relapsing hemophagocytic lymphohistiocytosis (HLH), MAS, or AIFEC. It may also refer to the prevention or delay of one or more of the aforementioned symptoms, for example, slowing the progression of the disease, condition, disorder, or syndrome, stopping or reversing the progression of the disease, condition, disorder, or syndrome, and improving clinical outcomes (e.g., preventing lethal progression of AIFEC and improving survival).

[0161] Total IL-18 in serum can be measured by conjugating anti-human IL-18 antibodies (e.g., clone 125-2H, MBL International) with Bio-plex COOH magnetic beads (Bio-Rad, Inc.), detected using biotinylated anti-human IL-18 (clone 159-12B, MBL), and the concentration calculated using the IL-18 contained in the Group II cytokine standard curve (Bio-Rad). Free IL-18 can be measured as described in Girard et al., Rheumatology (Oxford University Press). Dec. 2016;55(12):2237-2247.

[0162] Serum IL-1β can be measured using a commercially available ELISA kit (88-7261-88, eBioscience) (using the manufacturer’s instructions).

[0163] In some embodiments, the reduction of one or more elevated serum inflammatory markers by administration of a bispecific antibody or functional fragment thereof that simultaneously targets both IL-1β and IL-18 may be at least 10% lower than baseline compared to patients who did not receive the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction of one or more elevated serum inflammatory markers by administration of a bispecific antibody or functional fragment thereof that simultaneously targets both IL-1β and IL-18 may be at least 20% lower than baseline compared to patients who did not receive the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction of one or more elevated serum inflammatory markers by administration of a bispecific antibody or functional fragment thereof that simultaneously targets both IL-1β and IL-18 may be at least 30% lower than baseline compared to patients who did not receive the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction of one or more elevated serum inflammatory markers by administration of a bispecific antibody or functional fragment thereof that simultaneously targets both IL-1β and IL-18 may be at least 40% lower than baseline compared to patients who did not receive the same treatment (e.g., patients receiving standard of care (SOC)). In some embodiments, the reduction of one or more elevated serum inflammatory markers by administration of a bispecific antibody or a functional fragment thereof that simultaneously targets both IL-1β and IL-18 can be at least 50% lower than baseline compared to patients who have not received the same treatment (e.g., patients who have received standard of care (SOC)).

[0164] As an example, autoinflammatory or inflammatory diseases based on XIAP deficiency or CDC42 mutations that are suitable for treatment with the compositions and methods described herein include those caused by or related to point mutations, deletions (small or large exon deletions) or insertions that result in nonsense and missense mutations, intron or frameshift mutations.

[0165] In some embodiments, suitable or desired subjects have one of the 90 mutations disclosed in the XIAP gene by Mudde et al. ((2021) Evolution of Our Understanding of XIAP Deficiency. Front. Pediatr. 9:660520), or a mutation in the CDC42 gene that results in truncated or dysfunctional CDC42 protein, wherein the mutation can be a point mutation, deletion (small or large exon deletion) or insertion, resulting in nonsense and missense mutations, intron or frameshift mutations, particularly mutations in the N-terminal α-helix (e.g., residues Ile21 and Tyr23) or switch II motifs (e.g., Tyr64, Arg66, and Arg68), or in the fourth β chain (e.g., Cys81 and Ser83) or in the C-terminus (e.g., Ala159 and Glu171, or deletion), or p.R186C, p.C188Y, and p. 192Cext Mutations in 24 (as disclosed in Gernez Y et al., J Allergy Clin Immunol 2019;144:1122-5.e6).

[0166] "Treatment" can also refer to slowing, stopping, or reversing the progression of a disease, symptom, disorder, or syndrome, and improving clinical outcomes, such as moving from a higher number to a lower number on the following five ordinal scales:

[0167]

[0168] As used herein, the term "therapeuticly effective amount" of a compound refers to an amount of a compound that will elicit a biological or medical response in a subject (e.g., improvement of symptoms, reduction of symptom, slowing or delaying disease progression, or prevention of disease, symptom, disorder, or syndrome, etc.). In one non-limiting embodiment, the term "therapeuticly effective amount" refers to an amount of a compound described herein that, when administered to a subject, effectively at least partially reduces, inhibits, prevents, and / or improves autoinflammatory or disease based on XIAP deficiency or CDC42 mutation. In one non-limiting embodiment, the term "therapeuticly effective amount" refers to an amount of a compound described herein that, when administered to a subject, effectively at least partially reduces, inhibits, prevents, and / or improves autoinflammatory or disease based on XIAP deficiency or CDC42 mutation presenting with elevated IL-18 and IL-1β levels. In one non-limiting embodiment, the term "therapeuticly effective amount" refers to an amount of a compound described herein that, when administered to a subject, effectively at least partially reduces, inhibits, prevents, and / or improves AIFEC.

[0169] As used herein, a human antibody or fragment thereof contains a variable region or full-length heavy or light chain that is either “a product” or “derived from” a specific germline sequence if the variable region or full-length chain of the antibody is obtained from a system using a human germline immunoglobulin gene. Such systems include immunizing transgenic mice carrying human immunoglobulin genes with a target antigen or screening a library of human immunoglobulin genes displayed on a bacteriophage with a target antigen. A human antibody or fragment thereof that is either “a product” or “derived from” a human germline immunoglobulin sequence can be identified in this way by comparing the amino acid sequence of the human antibody with the amino acid sequence of a human germline immunoglobulin, and selecting the human germline immunoglobulin sequence that is closest to the human antibody sequence (i.e., has the highest identity percentage). A human antibody that is either “a product” or “derived from” a specific human germline immunoglobulin sequence may contain amino acid differences compared to the germline sequence due to, for example, naturally occurring somatic mutations or intentionally introduced site-directed mutations. However, the selected human antibody typically has an amino acid sequence that is at least 90% identical to the amino acid sequence encoded by the human germline immunoglobulin gene, and contains amino acid residues that identify the human antibody as belonging to humans when compared with germline immunoglobulin amino acid sequences of other species (e.g., mouse germline sequences). In some cases, the amino acid sequence of the human antibody may be at least 60%, 70%, 80%, 90%, or at least 95%, or even at least 96%, 97%, 98%, or 99% identical to the amino acid sequence encoded by the germline immunoglobulin gene. Typically, human antibodies derived from a specific human germline sequence will show a difference of no more than 10 amino acids from the amino acid sequence encoded by the human germline immunoglobulin gene. In some cases, the human antibody may show a difference of no more than 5 amino acids, or even no more than 4, 3, 2, or 1 amino acid from the amino acid sequence encoded by the germline immunoglobulin gene.

[0170] 2. IL-18 antibody

[0171] The particularly preferred IL-18 antibody or its antigen-binding fragment used in the disclosed method is a human antibody.

[0172] For ease of reference, Table 1 below provides the hypervariable regions of specific IL-18 antibodies (referred to as mAb1) based on the Cabart and Josiah definitions, as well as V... L and V H The structural domains and the complete amino acid sequences of the heavy and light chains.

[0173] Table 1. Hypervariable region (CDR), variable domains (VH and VL), and full-strand amino acid sequence of mAb1. The DNA encoding the VL of mAb1 is listed in SEQ ID NO: 18. The DNA encoding the VH of mAb1 is listed in SEQ ID NO: 8.

[0174]

[0175] In one embodiment, the IL-18 antibody or its antigen-binding fragment comprises at least one immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H The CDR1 has the amino acid sequence SEQ ID NO: 1, the CDR2 has the amino acid sequence SEQ ID NO: 2, and the CDR3 has the amino acid sequence SEQ ID NO: 3. In one embodiment, the IL-18 antibody or its antigen-binding fragment comprises at least one immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H The CDR1 has the amino acid sequence SEQ ID NO: 4, the CDR2 has the amino acid sequence SEQ ID NO: 5, and the CDR3 has the amino acid sequence SEQ ID NO: 6.

[0176] In one embodiment, the IL-18 antibody or its antigen-binding fragment comprises at least one immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L The CDR1 has the amino acid sequence SEQ ID NO: 11, the CDR2 has the amino acid sequence SEQ ID NO: 12, and the CDR3 has the amino acid sequence SEQ ID NO: 13. In one embodiment, the IL-18 antibody or its antigen-binding fragment comprises at least one immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L The CDR1 has the amino acid sequence SEQ ID NO: 14, the CDR2 has the amino acid sequence SEQ ID NO: 15, and the CDR3 has the amino acid sequence SEQ ID NO: 16.

[0177] In one embodiment, the IL-18 antibody or its antigen-binding fragment comprises at least one immunoglobulin V. H domain and at least one immunoglobulin V L Domains, wherein: a) the immunoglobulin V HThe domain comprises (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 1, CDR2 has the amino acid sequence SEQ ID NO: 2, and CDR3 has the amino acid sequence SEQ ID NO: 3; or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 4, CDR2 has the amino acid sequence SEQ ID NO: 5, and CDR3 has the amino acid sequence SEQ ID NO: 6; and b) the immunoglobulin V. L The domains include (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 11, CDR2 has the amino acid sequence SEQ ID NO: 12, and CDR3 has the amino acid sequence SEQ ID NO: 13 or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 14, CDR2 has the amino acid sequence SEQ ID NO: 15, and CDR3 has the amino acid sequence SEQ ID NO: 16.

[0178] In one embodiment, the IL-18 antibody or its antigen-binding fragment comprises: a) an immunoglobulin heavy chain variable domain (V) containing the amino acid sequence listed in SEQ ID NO: 7. H b) An immunoglobulin light chain variable domain (V) containing the amino acid sequence listed in SEQ ID NO: 17. L c) Immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 7 H Domains and immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 17 L d) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3. H e) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 11, SEQ ID NO: 12 and SEQ ID NO: 13. L f) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6. HDomain; g) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO: 16. L Domain; h) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3. H The domain and immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 11, SEQ ID NO: 12 and SEQ ID NO: 13. L Domain; i) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6. H The domain and immunoglobulin V containing the hypervariable regions listed in SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:16. L The domain; j) contains the light chain of SEQ ID NO: 19; k) contains the heavy chain of SEQ ID NO: 9; or l) contains both the light chain of SEQ ID NO: 19 and the heavy chain of SEQ ID NO: 9.

[0179] In some embodiments, the IL-18 antibody or its antigen-binding fragment (e.g., mAb1) comprises the three CDRs of SEQ ID NO: 7. In other embodiments, the IL-18 antibody or its antigen-binding fragment comprises the three CDRs of SEQ ID NO: 17. In other embodiments, the IL-18 antibody or its antigen-binding fragment comprises both the three CDRs of SEQ ID NO: 7 and the three CDRs of SEQ ID NO: 17. In some embodiments, the IL-18 antibody or its antigen-binding fragment comprises the three CDRs of SEQ ID NO: 9. In other embodiments, the IL-18 antibody or its antigen-binding fragment comprises the three CDRs of SEQ ID NO: 19. In other embodiments, the IL-18 antibody or its antigen-binding fragment comprises both the three CDRs of SEQ ID NO: 9 and the three CDRs of SEQ ID NO: 19.

[0180] In one embodiment, the IL-18 antibody or its antigen-binding fragment (e.g., mAb1) is selected from human IL-18 antibodies, which at least comprise: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion of a human heavy chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 1, CDR2 has the amino acid sequence SEQ ID NO: 2, and CDR3 has the amino acid sequence SEQ ID NO: 3; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion of a human light chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 11, CDR2 has the amino acid sequence SEQ ID NO: 12, and CDR3 has the amino acid sequence SEQ ID NO: 13.

[0181] In one embodiment, the IL-18 antibody or its antigen-binding fragment (e.g., mAb1) is selected from human IL-18 antibodies, which at least comprise: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion of a human heavy chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 4, CDR2 has the amino acid sequence SEQ ID NO: 5, and CDR3 has the amino acid sequence SEQ ID NO: 6; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion of a human light chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 14, CDR2 has the amino acid sequence SEQ ID NO: 15, and CDR3 has the amino acid sequence SEQ ID NO: 16.

[0182] In one embodiment, the IL-18 antibody or its antigen-binding fragment is selected from a single-chain antibody or its antigen-binding fragment containing an antigen-binding site comprising: a) a first domain comprising hypervariable regions CDR1, CDR2, and CDR3 in sequence, wherein CDR1 has the amino acid sequence SEQ ID NO: 1, CDR2 has the amino acid sequence SEQ ID NO: 2, and CDR3 has the amino acid sequence SEQ ID NO: 3; and b) a second domain comprising hypervariable regions CDR1, CDR2, and CDR3 in sequence, wherein CDR1 has the amino acid sequence SEQ ID NO: 11, CDR2 has the amino acid sequence SEQ ID NO: 12, and CDR3 has the amino acid sequence SEQ ID NO: 13; and c) a peptide linker that binds to the N-terminus of the first domain and the C-terminus of the second domain or binds to the C-terminus of the first domain and the N-terminus of the second domain.

[0183] In one embodiment, the IL-18 antibody or its antigen-binding fragment (e.g., mAb1) is selected from single-chain antibodies or their antigen-binding fragments containing an antigen-binding site comprising: a) a first domain comprising, sequentially, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 4, CDR2 has the amino acid sequence SEQ ID NO: 5, and CDR3 has the amino acid sequence SEQ ID NO: 6; and b) a second domain comprising, sequentially, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 14, CDR2 has the amino acid sequence SEQ ID NO: 15, and CDR3 has the amino acid sequence SEQ ID NO: 16; and c) a peptide linker that binds to the N-terminus of the first domain and the C-terminus of the second domain or binds to the C-terminus of the first domain and the N-terminus of the second domain.

[0184] The V of the IL-18 antibody or its antigen-binding fragment used in the disclosed method H or V L The structural domain can have the same V as listed in SEQ ID NO: 7 and 17. H or V L The V domains are basically the same H and / or V LDomains. The human IL-18 antibody disclosed herein may comprise a heavy chain substantially identical to the heavy chain listed in SEQ ID NO: 9 and / or a light chain substantially identical to the light chain listed in SEQ ID NO: 19. The human IL-18 antibody disclosed herein may comprise: a heavy chain containing SEQ ID NO: 9 and a light chain containing SEQ ID NO: 19. The human IL-18 antibody disclosed herein may comprise: a) a heavy chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 7 and a constant portion of the human heavy chain; and b) a light chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 17 and a constant portion of the human light chain.

[0185] Other preferred IL-18 antagonists (e.g., antibodies) used in the disclosed methods, kits, and protocols are those listed below: U.S. Patent No. 9,376,489, which is incorporated herein by reference in its entirety.

[0186] 3. IL-1β antibody

[0187] The particularly preferred IL-1β antibody or its antigen-binding fragment used in the disclosed method is a human antibody.

[0188] For ease of reference, Table 2 below provides the hypervariable regions of specific IL-1β antibodies (referred to as mAb2) based on the Cabart and Josiah definitions, as well as V... L and V H The structural domains and the complete amino acid sequences of the heavy and light chains.

[0189] Table 2. Hypervariable region (CDR), variable domains (VH and VL), and full-strand amino acid sequence of mAb2. The DNA encoding the VL of mAb2 is listed in SEQ ID NO: 38. The DNA encoding the VH of mAb2 is listed in SEQ ID NO: 27.

[0190]

[0191] In one embodiment, the IL-1β antibody or its antigen-binding fragment comprises at least one immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. HThe CDR1 has the amino acid sequence SEQ ID NO: 21, the CDR2 has the amino acid sequence SEQ ID NO: 22, and the CDR3 has the amino acid sequence SEQ ID NO: 23. In one embodiment, the IL-1β antibody or its antigen-binding fragment comprises at least one immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H The CDR1 has the amino acid sequence SEQ ID NO: 24, the CDR2 has the amino acid sequence SEQ ID NO: 25, and the CDR3 has the amino acid sequence SEQ ID NO: 26.

[0192] In one embodiment, the IL-1β antibody or its antigen-binding fragment comprises at least one immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L The CDR1 has the amino acid sequence SEQ ID NO: 31, the CDR2 has the amino acid sequence SEQ ID NO: 32, and the CDR3 has the amino acid sequence SEQ ID NO: 33. In one embodiment, the IL-1β antibody or its antigen-binding fragment comprises at least one immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L The CDR1 has the amino acid sequence SEQ ID NO: 34, the CDR2 has the amino acid sequence SEQ ID NO: 35, and the CDR3 has the amino acid sequence SEQ ID NO: 36.

[0193] In one embodiment, the IL-1β antibody or its antigen-binding fragment contains at least one immunoglobulin V. H domain and at least one immunoglobulin V L Domains, wherein: a) the immunoglobulin V H The domain comprises (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 21, CDR2 has the amino acid sequence SEQ ID NO: 22, and CDR3 has the amino acid sequence SEQ ID NO: 23; or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 24, CDR2 has the amino acid sequence SEQ ID NO: 25, and CDR3 has the amino acid sequence SEQ ID NO: 26; and b) the immunoglobulin V. LThe domains include (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 31, CDR2 has the amino acid sequence SEQ ID NO: 32, and CDR3 has the amino acid sequence SEQ ID NO: 33 or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 34, CDR2 has the amino acid sequence SEQ ID NO: 35, and CDR3 has the amino acid sequence SEQ ID NO: 36.

[0194] In one embodiment, the IL-1β antibody or its antigen-binding fragment comprises: a) an immunoglobulin heavy chain variable domain (V) containing the amino acid sequence listed in SEQ ID NO: 27. H b) An immunoglobulin light chain variable domain (V) containing the amino acid sequence listed in SEQ ID NO: 37. L c) Immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 27 H Domains and immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 37 L d) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 21, SEQ ID NO: 22 and SEQ ID NO: 23. H e) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 31, SEQ ID NO: 32 and SEQ ID NO: 33. L f) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 24, SEQ ID NO: 25 and SEQ ID NO: 26. H g) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 34, SEQ ID NO: 35 and SEQ ID NO: 36. L Domain; h) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 21, SEQ ID NO: 22 and SEQ ID NO: 23. H The domain and immunoglobulin V containing the hypervariable regions listed in SEQ ID NO:31, SEQ ID NO:32 and SEQ ID NO:33. LDomain; i) Immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 24, SEQ ID NO: 25 and SEQ ID NO: 26. H The domain and immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 34, SEQ ID NO: 35 and SEQ ID NO: 36. L The domain; j) contains the light chain of SEQ ID NO: 37; k) contains the heavy chain of SEQ ID NO: 29; or l) contains both the light chain of SEQ ID NO: 39 and the heavy chain of SEQ ID NO: 29.

[0195] In some embodiments, the IL-1β antibody or its antigen-binding fragment (e.g., mAb2) comprises the three CDRs of SEQ ID NO: 37. In other embodiments, the IL-1β antibody or its antigen-binding fragment comprises the three CDRs of SEQ ID NO: 27. In other embodiments, the IL-1β antibody or its antigen-binding fragment comprises both the three CDRs of SEQ ID NO: 37 and SEQ ID NO: 27. In some embodiments, the IL-1β antibody or its antigen-binding fragment comprises the three CDRs of SEQ ID NO: 39. In other embodiments, the IL-1β antibody or its antigen-binding fragment comprises the three CDRs of SEQ ID NO: 29. In other embodiments, the IL-1β antibody or its antigen-binding fragment comprises both the three CDRs of SEQ ID NO: 39 and SEQ ID NO: 29.

[0196] In one embodiment, the IL-1β antibody or its antigen-binding fragment (e.g., mAb2) is selected from human IL-1β antibodies, which at least comprise: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion of a human heavy chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 21, CDR2 has the amino acid sequence SEQ ID NO: 22, and CDR3 has the amino acid sequence SEQ ID NO: 23; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion of a human light chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 31, CDR2 has the amino acid sequence SEQ ID NO: 32, and CDR3 has the amino acid sequence SEQ ID NO: 33.

[0197] In one embodiment, the IL-1β antibody or its antigen-binding fragment (e.g., mAb2) is selected from human IL-1β antibodies, which at least comprise: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion of a human heavy chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 24, CDR2 has the amino acid sequence SEQ ID NO: 25, and CDR3 has the amino acid sequence SEQ ID NO: 26; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion of a human light chain or a fragment thereof, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 34, CDR2 has the amino acid sequence SEQ ID NO: 35, and CDR3 has the amino acid sequence SEQ ID NO: 36.

[0198] In one embodiment, the IL-1β antibody or its antigen-binding fragment is selected from a single-chain antibody or its antigen-binding fragment containing an antigen-binding site comprising: a) a first domain comprising, sequentially, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 21, CDR2 has the amino acid sequence SEQ ID NO: 22, and CDR3 has the amino acid sequence SEQ ID NO: 23; and b) a second domain comprising, sequentially, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 31, CDR2 has the amino acid sequence SEQ ID NO: 32, and CDR3 has the amino acid sequence SEQ ID NO: 33; and c) a peptide linker that binds to the N-terminus of the first domain and the C-terminus of the second domain or binds to the C-terminus of the first domain and the N-terminus of the second domain.

[0199] In one embodiment, the IL-1β antibody or its antigen-binding fragment (e.g., mAb2) is selected from single-chain antibodies or their antigen-binding fragments containing an antigen-binding site comprising: a) a first domain comprising, sequentially, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 24, CDR2 has the amino acid sequence SEQ ID NO: 25, and CDR3 has the amino acid sequence SEQ ID NO: 26; and b) a second domain comprising, sequentially, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 34, CDR2 has the amino acid sequence SEQ ID NO: 35, and CDR3 has the amino acid sequence SEQ ID NO: 36; and c) a peptide linker that binds to the N-terminus of the first domain and the C-terminus of the second domain or binds to the C-terminus of the first domain and the N-terminus of the second domain.

[0200] The V of the IL-1β antibody or its antigen-binding fragment used in the disclosed method H or V L The structural domain can have the same V as listed in SEQ ID NO: 27 and 37. H or V L The V domains are basically the same H and / or V L Domains. The human IL-1β antibody disclosed herein may comprise a heavy chain substantially identical to the heavy chain listed in SEQ ID NO: 29 and / or a light chain substantially identical to the light chain listed in SEQ ID NO: 39. The human IL-1β antibody disclosed herein may comprise: a heavy chain containing SEQ ID NO: 29 and a light chain containing SEQ ID NO: 39. The human IL-1β antibody disclosed herein may comprise: a) a heavy chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 27 and a constant portion of the human heavy chain; and b) a light chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 37 and a constant portion of the human light chain.

[0201] Other preferred IL-1β antagonists (e.g., antibodies) used in the disclosed methods, kits, and protocols are those listed below: U.S. Patent Nos. 7,446,175, 7,993,878, or 8,273,350, which are incorporated herein by reference in their entirety.

[0202] 4. Fc modifier

[0203] In addition to modifications performed within the frame or CDR region, or as an alternative to modifications performed within the frame or CDR region, the antibodies of the present invention can be engineered to include modifications within the Fc region, typically to alter one or more functional properties of the antibody, such as serum half-life, complement binding, Fc receptor binding, and / or antigen-dependent cytotoxicity. Furthermore, the antibodies of the present invention can be chemically modified (e.g., one or more chemical moieties can be attached to the antibody) or modified to alter their glycosylation, thereby again altering one or more functional properties of the antibody. Each of these examples is described in more detail below. The residue numbers in the Fc region are those of Edelman et al., PNAS, May 1969, 63(1):78-85, according to the EU numbering scheme.

[0204] In one embodiment, the hinge region of CH1 is modified such that the number of cysteine ​​residues in the hinge region is altered, for example, increased or decreased. This method is further described in U.S. Patent No. 5,677,425 to Bodmer et al. Changing the number of cysteine ​​residues in the CH1 hinge region can, for example, promote the assembly of light and heavy chains or increase or decrease antibody stability.

[0205] In another embodiment, the Fc hinge region of the antibody is mutated to shorten the antibody's biological half-life. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc hinge fragment, resulting in impaired staphylococcal protein A (SpA) binding relative to the native Fc hinge domain SpA binding. This method is further described in detail in U.S. Patent No. 6,165,745 to Ward et al.

[0206] In another embodiment, the antibody is modified to increase its biological half-life. Various methods can be employed. For example, one or more of the following mutations can be introduced: T252L, T254S, T256F as described by Ward in U.S. Patent No. 6,277,375. Alternatively, to increase the biological half-life, the antibody can be modified within the CH1 or CL region to contain a salvage receptor-binding epitope with two loops of the CH2 domain derived from the Fc region of IgG, as described in U.S. Patent Nos. 5,869,046 and 6,121,022 to Presta et al.

[0207] In other embodiments, the effector function of the antibody is altered by replacing at least one amino acid residue in the Fc region with a different amino acid residue. For example, one or more amino acids can be replaced with different amino acid residues to give the antibody an altered affinity for the effector ligand while retaining the antigen-binding ability of the parent antibody. The effector ligand with altered affinity can be, for example, an Fc receptor or the C1 component of complement. This method is described in further detail in U.S. Patents 5,624,821 and 5,648,260 to Winter et al.

[0208] In another embodiment, one or more amino acids selected from amino acid residues may be substituted with different amino acid residues, such that the antibody exhibits altered C1q binding and / or reduced or eliminated complement-dependent cytotoxicity (CDC). This method is further described in detail in U.S. Patent No. 6,194,551 to Idusogie et al.

[0209] In another embodiment, one or more amino acid residues are altered to change the antibody's ability to fix complement. This method is further described in PCT Publication WO 94 / 29351 by Bodmer et al.

[0210] In yet another embodiment, the Fc region is modified to increase the antibody-mediated antibody-dependent cytotoxicity (ADCC) and / or the antibody affinity for the Fcγ receptor is increased by modifying one or more amino acids. This method is further described by Presta in PCT Publication WO 00 / 42072. Furthermore, binding sites for FcγR1, FcγRII, FcγRIII, and FcRn on human IgG1 have been located and variants with modified binding have been described (see Shields, RL et al., (2001) J. Biol Chem [Journal of Biochemistry] 276:6591-6604).

[0211] In some embodiments, the Fc domain of the IgG1 isotype is used. In some specific embodiments, mutant variants of the IgG1 Fc fragment, such as silent IgG1 Fc, are used, which reduce or eliminate the ability of the fusion peptide to mediate antibody-dependent cytotoxicity (ADCC) and / or bind to the Fcγ receptor. Examples of IgG1 isotype silencing mutants, such as those described by Hezareh et al., J. Virol [Journal of Virology] (2001); 75(24):12161-8, involve the substitution of leucine residues at amino acid positions 234 and 235 with alanine residues.

[0212] In some embodiments, the Fc domain is a mutant that prevents glycosylation at position 297 of the Fc domain. For example, the Fc domain contains an amino acid substitution of an asparagine residue at position 297. An example of such amino acid substitution is replacing N297 with glycine or alanine.

[0213] Silent effector functions can be acquired through mutations in the Fc region of antibodies, and have been described in the art: LALA and N297A (Strohl, W., 2009, Curr. Opin. Biotechnol. [Current Biotech Viewpoint] Vol. 20(6):685-691); and D265A (Baudino et al., 2008, J. lmmunol. [Journal of Immunology] 181:6664-69; Strohl, W., ibid.); and DAPA (D265A and P329A) (Shields RL., J Biol Chem. [Journal of Biochemistry]. 2001; 276(9):6591-604; US Patent Publication US 2015 / 0320880). Examples of Fc-silencing lgG1 antibodies include the so-called LALA mutant, which contains L234A and L235A mutations in the lgG1 Fc amino acid sequence. Another example of a silent IgG1 antibody contains the D265A mutation. Another example of a silent IgG1 antibody is the so-called DAPA mutant, which contains the D265A and P329A mutations in the IgG1 Fc amino acid sequence. Another silent IgG1 antibody contains the N297A mutation, which produces a glycosylated / non-glycosylated antibody. Additional Fc mutations used to provide silencing effector function are described in PCT Publication No. WO 2014 / 145806 (e.g., in Figure 7 of WO 2014 / 145806), which is incorporated herein by reference in its entirety. An example of a silent IgG1 antibody from WO 2014 / 145806 contains the E233P, L234V, L235A, and S267K mutations and the deletion of G236 (G236del). Another example of a silent IgG1 antibody from WO 2014 / 145806 contains E233P, L234V, and L235A mutations, as well as G236 deletion (G236del). Another example of a silent IgG1 antibody from WO 2014 / 145806 contains an S267K mutation.

[0214] In another embodiment, the glycosylation of the antibody is modified. For example, an antibody without glycosylation can be prepared (i.e., the antibody lacks glycosylation). Glycosylation can be altered to, for example, increase the antibody's affinity for the antigen. Such glycosylation modification can be accomplished by, for example, altering one or more glycosylation sites within the antibody sequence. For example, one or more amino acid substitutions can be performed, resulting in the elimination of one or more variable region framework glycosylation sites, thereby eliminating the glycosylation at that site. This glycosylation-free process can increase the antibody's affinity for the antigen. This method is described in more detail in U.S. Patents 5,714,350 and 6,350,861 to Co et al.

[0215] Alternatively or concurrently, antibodies with altered glycosylation patterns can be prepared, such as hypofucosylated antibodies with reduced amounts of fucosylated residues or antibodies with increased bipartite GlcNac structures. Such altered glycosylation patterns have been shown to increase the ADCC capacity of antibodies. This glycosylation modification can be accomplished, for example, by expressing antibodies in host cells with altered glycosylation mechanisms. Cells with altered glycosylation mechanisms have been described in the art and can be used as host cells in which the recombinant antibodies of the present invention are expressed, thereby producing antibodies with altered glycosylation. For example, EP 1,176,195 by Hang et al. describes a cell line with a dysfunctional FUT8 gene encoding a fucosylated enzyme, such that antibodies expressed in such cell lines exhibit hypofucosylation. Therefore, in one embodiment, the antibodies of the present invention are produced by recombinant expression in cell lines exhibiting a fucosylation pattern, such as mammalian cell lines with defective expression of the FUT8 gene encoding a fucosylated enzyme. Presta described the variant CHO cell line Lecl3 in PCT Publication WO03 / 035835, which had a reduced ability to attach fucose to Asn(297) linked sugars and also resulted in hypofucosylation of antibodies expressed in the host cells (see also Shields, RL et al., 2002 J. Biol. Chem. 277:26733-26740). Umana et al.'s PCT disclosure WO99 / 54342 describes cell lines engineered to express glycoprotein-modifying glycosyltransferases (e.g., β(1,4)-N-acetylglucosamine transferase III (GnTIII)), such that antibodies expressed in the engineered cell lines exhibit increased bipartite GlcNac structures, which lead to increased ADCC activity of the antibodies (see also Umana et al., 1999 Nat. Biotech. [Nature Biotechnology] 17:176-180). Alternatively, the antibodies of the present invention can be produced in yeast or filamentous fungi engineered for a mammalian-like glycosylation pattern and capable of producing antibodies lacking fucose as a glycosylation pattern (see, for example, EP 1297172 B1).

[0216] Another modification of the antibodies discussed herein is polyethylene glycol (PEG) modification. PEGylation of antibodies can, for example, increase their biological (e.g., serum) half-life. To PEGylate an antibody, the antibody or a fragment thereof is typically reacted with polyethylene glycol (PEG) (such as a reactive ester or aldehyde derivative of PEG) under conditions where one or more PEG groups are attached to the antibody or antibody fragment. PEGylation can be performed by acylation or alkylation with a reactive PEG molecule (or a similar reactive water-soluble polymer). As used herein, the term "polyethylene glycol" is intended to cover any form of PEG that has been used to derive other proteins, such as mono(C1-C10)alkoxy- or aryloxy-PEG or PEG-maleimide. In some embodiments, the antibody to be PEGylated is a glycosylated antibody. Methods for PEGylating proteins are known in the art and can be applied to the antibodies of this invention. See, for example, Nishimura et al. EP 0 154 316 and Ishikawa et al. EP 0 401 384.

[0217] Another modification of the antibody contemplated by this invention is at least a conjugate or protein fusion of the antigen-binding region of the antibody of this invention with a serum protein (such as human serum albumin or a fragment thereof) to increase the half-life of the resulting molecule. Such a method is described, for example, in Ballance et al., EP 0322094.

[0218] Another modification of the antibody considered in this invention is one or more modifications to increase the formation of heterodimeric bispecific antibodies. Various methods available in the art can be used to enhance the dimerization of the two heavy chain domains of bispecific antibodies such as bbmAb, as disclosed, for example, in: EP 1870459A1; U.S. Patent No. 5,582,996; U.S. Patent No. 5,731,168; U.S. Patent No. 5,910,573; U.S. Patent No. 5,932,448; U.S. Patent No. 6,833,441; U.S. Patent No. 7,183,076; U.S. Patent Application Publication No. 2006204493 A1; and PCT Publication No. WO 2009 / 089004 A1, the contents of which are incorporated herein by reference in their entirety.

[0219] For example, the use of a pestle and mortar to produce bispecific antibodies was disclosed in PCT Publication No. WO 1996 / 027011, Ridgway et al. (1996), and Merchant et al. (1998).

[0220] When implementing some of the treatments or uses disclosed herein, a therapeutically effective dose of a bispecific antibody (e.g., bbmAb) targeting both IL-1β and IL-18 must be administered to the subject in need. It should be understood that regimen changes may be applicable to certain patients. Therefore, administration (e.g., bbmAb) may be more frequent, such as once daily, once every two weeks, or once weekly.

[0221] Some patients may benefit from a loading regimen (e.g., daily administration for several days [e.g., 1 to 4 days, such as administration on day 0, day 1, day 2, and / or day 3]) followed by a maintenance regimen (e.g., starting in week 3 or 4, where bbbmAb may be administered weekly, every two weeks, or every four weeks for several weeks). In some embodiments, administration of a bispecific antibody (e.g., bbbmAb) that simultaneously targets both IL-1β and IL-18 may last for 1, 2, 3, 4, 5, 6, or 7 days. In some embodiments, the administration of a bispecific antibody (e.g., bbmAb) that simultaneously targets both IL-1β and IL-18 may last for 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, or longer.

[0222] It should be understood that, based on the severity of the disease, dose escalation may be appropriate for certain patients (e.g., patients who do not respond adequately to treatment with bbmAb). Therefore, the dose (intravenous (iv)) may be greater than about 10 mg / kg, for example, about 11 mg / kg, 12 mg / kg, 15 mg / kg, 20 mg / kg, 25 mg / kg, 30 mg / kg, 35 mg / kg, etc. Furthermore, the subcutaneous (sc) dose (loading or maintenance dose) may be greater than about 50 mg to about 900 mg sc, for example, about 75 mg, about 100 mg, about 125 mg, about 175 mg, about 200 mg, about 250 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 600 mg, etc.

[0223] It should also be understood that dose reduction may also be applicable to certain patients, such as those who have experienced adverse events or adverse responses to treatment with bbmAb. Therefore, the dose may be lower than about 10 mg / kg, for example, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 8 mg / kg, or about 9 mg / kg. In some embodiments, the bbmAB1 dose may be adjusted as determined by a physician.

[0224] In some embodiments, the bbmAB1 antibody may be administered to the patient in a single dose of 10 mg / kg via IV delivery, wherein the dose may be adjusted to a higher or lower dose as determined by a physician if necessary, for example, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 8 mg / kg or about 9 mg / kg or, for example, about 11 mg / kg, 12 mg / kg, 15 mg / kg, 20 mg / kg, 25 mg / kg, 30 mg / kg, 35 mg / kg, etc.

[0225] In some embodiments, the bbmAB1 antibody may be administered to the patient at an initial dose of 10 mg / kg via intravenous delivery, and the dose may be adjusted to a higher or lower dose as determined by a physician if necessary.

[0226] In a specific embodiment, 10 mg / kg of bbmAB1 was administered on day 1.

[0227] In a specific embodiment, 10 mg / kg bbmAB1 was administered on day 1 (D1) and on days 2 (D2), 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, and / or 14.

[0228] In another specific embodiment, 10 mg / kg of bbmAB1 was administered intravenously on day 1.

[0229] Example 1:

[0230] The production of bbmAb has been described in detail in Examples 1 to 5 of patent application WO / 2018 / 229612. Example 1 of WO / 2018 / 229612 includes (1) vector construction, (2) host cell line and transfection, (3) cell selection and sorting, (4) cell expansion, (5) clonal stability, (6) manufacturing, (7) analytical characterization and purity assessment, and (8) analytical results, which are incorporated herein by reference in their entirety.

[0231] bbmAb is a bispecific IgG1 antibody with a LALA silencing mutation that binds to two distinct targets, IL-1β and IL-18. This antibody binds to two different antigen-binding arms (Fab fragments), with the Fab for IL-1β based on mAb2 and containing a κ light chain (Vk6). The Fab for IL-18 is based on mAb1 and consists of a λ light chain (Vλ1). To drive heterodimerization of the Fc domain during expression, a "groove" with large amino acid (aa) side chains (S354C and T366W) and a "mortar" with small aa side chains (Y349C, T366S, L368A, Y407V) are introduced into the mAb2 heavy chain.

[0232] For ease of reference, Table 3 below provides the high-variable region of bbmAb based on the Kabat and Josiah definitions, as well as V. L and V H The structural domains and the complete amino acid sequences of the heavy and light chains.

[0233] Table 3. Hypervariable regions (CDR), variable domains (VH and VL), and full-strand amino acid sequences of bbmAb. The DNA encoding the first VL is listed in SEQ ID NO: 102, and the DNA encoding the second VL is listed in SEQ ID NO: 70. The DNA encoding the first VH is listed in SEQ ID NO: 86, and the DNA encoding the second VH is listed in SEQ ID NO: 54.

[0234]

[0235] In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H1The CDR1 has the amino acid sequence SEQ ID NO:76, the CDR2 has the amino acid sequence SEQ ID NO:77, and the CDR3 has the amino acid sequence SEQ ID NO:78. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed method for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H1 The CDR1 has the amino acid sequence SEQ ID NO: 79, the CDR2 has the amino acid sequence SEQ ID NO: 80, and the CDR3 has the amino acid sequence SEQ ID NO: 81. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H1 The CDR1 has the amino acid sequence SEQ ID NO: 82, the CDR2 has the amino acid sequence SEQ ID NO: 83, and the CDR3 has the amino acid sequence SEQ ID NO: 84.

[0236] In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H2 The CDR1 has the amino acid sequence SEQ ID NO:44, the CDR2 has the amino acid sequence SEQ ID NO:45, and the CDR3 has the amino acid sequence SEQ ID NO:46. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed method for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3.H2 The CDR1 has the amino acid sequence SEQ ID NO: 47, the CDR2 has the amino acid sequence SEQ ID NO: 48, and the CDR3 has the amino acid sequence SEQ ID NO: 49. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin heavy chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. H2 The CDR1 has the amino acid sequence SEQ ID NO: 50, the CDR2 has the amino acid sequence SEQ ID NO: 51, and the CDR3 has the amino acid sequence SEQ ID NO: 52.

[0237] In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L1 The CDR1 has the amino acid sequence SEQ ID NO:92, the CDR2 has the amino acid sequence SEQ ID NO:93, and the CDR3 has the amino acid sequence SEQ ID NO:94. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L1The CDR1 has the amino acid sequence SEQ ID NO: 95, the CDR2 has the amino acid sequence SEQ ID NO: 96, and the CDR3 has the amino acid sequence SEQ ID NO: 97. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L1 The CDR1 has the amino acid sequence SEQ ID NO: 98, the CDR2 has the amino acid sequence SEQ ID NO: 99, and the CDR3 has the amino acid sequence SEQ ID NO: 100.

[0238] In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L2 The CDR1 has the amino acid sequence SEQ ID NO:60, the CDR2 has the amino acid sequence SEQ ID NO:61, and the CDR3 has the amino acid sequence SEQ ID NO:62. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3. L2 The CDR1 has the amino acid sequence SEQ ID NO: 63, the CDR2 has the amino acid sequence SEQ ID NO: 64, and the CDR3 has the amino acid sequence SEQ ID NO: 65. In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin light chain variable domain (V) containing hypervariable regions CDR1, CDR2, and CDR3.L2 The CDR1 has the amino acid sequence SEQ ID NO: 66, the CDR2 has the amino acid sequence SEQ ID NO: 67, and the CDR3 has the amino acid sequence SEQ ID NO: 68.

[0239] In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a first immunoglobulin V. H1 Domains and first immunoglobulin V L1 Domains, wherein: a) the first immunoglobulin V H1 The domain comprises (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 79, CDR2 has the amino acid sequence SEQ ID NO: 80, and CDR3 has the amino acid sequence SEQ ID NO: 81; or iii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 82, CDR2 has the amino acid sequence SEQ ID NO: 83, and CDR3 has the amino acid sequence SEQ ID NO: 84 and b) the first immunoglobulin V. L1 The domains include (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 93, and CDR3 has the amino acid sequence SEQ ID NO: 94; or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 95, CDR2 has the amino acid sequence SEQ ID NO: 96, and CDR3 has the amino acid sequence SEQ ID NO: 97; or iii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 98, CDR2 has the amino acid sequence SEQ ID NO: 99, and CDR3 has the amino acid sequence SEQ ID NO: 100.

[0240] In one embodiment, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises a second immunoglobulin V. H2 Domain and second immunoglobulin V L2 Domains, wherein: a) the second immunoglobulin V H2 The domain comprises (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 47, CDR2 has the amino acid sequence SEQ ID NO: 48, and CDR3 has the amino acid sequence SEQ ID NO: 49; or iii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 50, CDR2 has the amino acid sequence SEQ ID NO: 51, and CDR3 has the amino acid sequence SEQ ID NO: 52 and b) the second immunoglobulin V. L2 The domains include (e.g., sequentially): i) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62; or ii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 63, CDR2 has the amino acid sequence SEQ ID NO: 64, and CDR3 has the amino acid sequence SEQ ID NO: 65; or iii) hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 66, CDR2 has the amino acid sequence SEQ ID NO: 67, and CDR3 has the amino acid sequence SEQ ID NO: 68.

[0241] In one embodiment, the IL-18 / IL-1β bispecific antibody for use in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) for use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises: a) a first immunoglobulin heavy chain variable domain (VH1) comprising the amino acid sequence listed in SEQ ID NO:85; b) a first immunoglobulin light chain variable domain (VH1) comprising the amino acid sequence listed in SEQ ID NO:101. L1 c) First immunoglobulin V containing the amino acid sequence listed in SEQ ID NO:85 H1 The domain and the first immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 101 L1 d) First immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 76, SEQ ID NO: 77 and SEQ ID NO: 78. H1 e) First immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 92, SEQ ID NO: 93 and SEQ ID NO: 94. L1 f) First immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 79, SEQ ID NO: 80 and SEQ ID NO: 81. H1 Domain; g) First immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 95, SEQ ID NO: 96 and SEQ ID NO: 97. L1 Domain; h) First immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 76, SEQ ID NO: 77 and SEQ ID NO: 78. H1 The domain and the first immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 92, SEQ ID NO: 93 and SEQ ID NO: 94. L1 Domain; i) First immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 79, SEQ ID NO: 80 and SEQ ID NO: 81. H1 The domain and the first immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 95, SEQ ID NO: 96 and SEQ ID NO: 97. L1The domain; j) contains the first light chain of SEQ ID NO: 103; k) contains the first heavy chain of SEQ ID NO: 87; or l) contains the first light chain of SEQ ID NO: 103 and the first heavy chain of SEQ ID NO: 87.

[0242] In one embodiment, the IL-18 / IL-1β bispecific antibody for use in (i) the treatment or prevention of the disclosed cytokine release syndrome or cytokine storm syndrome, or (ii) for use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises: a) a second immunoglobulin heavy chain variable domain (VH2) comprising the amino acid sequence listed in SEQ ID NO:53; b) a second immunoglobulin light chain variable domain (VH2) comprising the amino acid sequence listed in SEQ ID NO:69. L2 c) A second immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 53. H2 The domain and a second immunoglobulin V containing the amino acid sequence listed in SEQ ID NO: 69 L2 d) Second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 44, SEQ ID NO: 45 and SEQ ID NO: 46. H2 e) A second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 60, SEQ ID NO: 61 and SEQ ID NO: 62. L2 f) Second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 47, SEQ ID NO: 48 and SEQ ID NO: 49. H2 g) Second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 63, SEQ ID NO: 64 and SEQ ID NO: 65. L2 Domain; h) Second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 44, SEQ ID NO: 45 and SEQ ID NO: 46. H2 The domain and the second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 60, SEQ ID NO: 61 and SEQ ID NO: 62. L2 Domain; i) Second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 47, SEQ ID NO: 48 and SEQ ID NO: 49. H2The domain and the second immunoglobulin V containing the hypervariable regions listed in SEQ ID NO: 63, SEQ ID NO: 64 and SEQ ID NO: 65. L2 The domain; j) contains the second light chain of SEQ ID NO: 81; k) contains the second heavy chain of SEQ ID NO: 55; or l) contains the second light chain of SEQ ID NO: 81 and the second heavy chain of SEQ ID NO: 55.

[0243] In some embodiments, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of cytokine release syndrome or cytokine storm syndrome disclosed herein, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises the three CDRs of SEQ ID NO: 53. In other embodiments, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of cytokine release syndrome or cytokine storm syndrome disclosed herein, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises the three CDRs of SEQ ID NO: 69. In other embodiments, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of cytokine release syndrome or cytokine storm syndrome disclosed herein, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises the three CDRs of SEQ ID NO: 53 and the three CDRs of SEQ ID NO: 69. In some embodiments, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of cytokine release syndrome or cytokine storm syndrome disclosed, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises the three CDRs of SEQ ID NO: 85. In other embodiments, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of cytokine release syndrome or cytokine storm syndrome disclosed, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises the three CDRs of SEQ ID NO: 101. In other embodiments, the IL-18 / IL-1β bispecific antibody used in (i) the treatment or prevention of cytokine release syndrome or cytokine storm syndrome disclosed, or (ii) the use in the disclosed methods for treating or preventing cytokine release syndrome or cytokine storm syndrome, comprises the three CDRs of SEQ ID NO: 85 and the three CDRs of SEQ ID NO: 101.

[0244] In some embodiments, the IL-18 / IL-1β bispecific antibody used in the treatment of autoinflammatory or disease-based by XIAP deficiency or CDC42 mutation disclosed in (i) comprises the three CDRs of SEQ ID NO: 85. In other embodiments, the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based by XIAP deficiency or CDC42 mutation disclosed in (i) comprises the three CDRs of SEQ ID NO: 101. In other embodiments, the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based by XIAP deficiency or CDC42 mutation disclosed in (i) comprises the three CDRs of SEQ ID NO: 85 and the three CDRs of SEQ ID NO: 101. In some embodiments, the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based by XIAP deficiency or CDC42 mutation disclosed in (i) comprises the three CDRs of SEQ ID NO: 53. In other embodiments, the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation disclosed in (i) comprises three CDRs of SEQ ID NO: 69. In other embodiments, the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation disclosed in (i) comprises three CDRs of SEQ ID NO: 53 and three CDRs of SEQ ID NO: 69. In one embodiment, the L-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation disclosed in (i) comprises three CDRs of SEQ ID NO: 85, three CDRs of SEQ ID NO: 101, three CDRs of SEQ ID NO: 53, and three CDRs of SEQ ID NO: 69.

[0245] In one embodiment, the first portion of the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation disclosed in (i) is selected from a human IL-18 antibody, which comprises at least: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human heavy chain, the variable domain comprising, in turn, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human light chain, the variable domain comprising, in turn, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 98, and CDR3 has the amino acid sequence SEQ ID NO: 98; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human light chain, the variable domain comprising, in turn, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 98, and CDR3 has the amino acid sequence SEQ ID NO: 98; 93, and the CDR3 has the amino acid sequence SEQ ID NO: 94. Furthermore, the second part of the IL-18 / IL-1β bispecific antibody is selected from human IL-1β antibodies, which at least comprise: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human heavy chain, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human light chain, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62.

[0246] In one embodiment, the first portion of the IL-18 / IL-1β bispecific antibody used in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation disclosed in (i) is selected from a human IL-18 antibody, which comprises at least: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human heavy chain, the variable domain comprising, in turn, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human light chain, the variable domain comprising, in turn, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 98, and CDR3 has the amino acid sequence SEQ ID NO: 98; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human light chain, the variable domain comprising, in turn, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 98, and CDR3 has the amino acid sequence SEQ ID NO: 98; 93, and the CDR3 has the amino acid sequence SEQ ID NO: 94. Furthermore, the second part of the IL-18 / IL-1β bispecific antibody is selected from human IL-1β antibodies, which at least comprise: a) an immunoglobulin heavy chain or a fragment thereof, the immunoglobulin heavy chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human heavy chain, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3; wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; and b) an immunoglobulin light chain or a fragment thereof, the immunoglobulin light chain or the fragment thereof comprising a variable domain and a constant portion or a fragment thereof of a human light chain, the variable domain comprising, in sequence, hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62.

[0247] The first V of the IL-18 / IL-1β bispecific antibody used in the disclosed method H1 or V L1 The structural domain can have the same V as listed in SEQ ID NO: 85 and 101. H or V L The first V is essentially the same as the structure domain. H1 and / or the first V L1Domains. The IL-18 / IL-1β bispecific antibody disclosed herein for use in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation as disclosed in (i) may comprise a first heavy chain substantially identical to the heavy chain listed in SEQ ID NO: 87 and / or a first light chain substantially identical to the light chain listed in SEQ ID NO: 103. The IL-18 / IL-1β bispecific antibody disclosed herein for use in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation as disclosed in (i) may comprise a first heavy chain containing SEQ ID NO: 87 and a first light chain containing SEQ ID NO: 103. As disclosed herein, the IL-18 / IL-1β bispecific antibody for use in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation as disclosed may comprise: a) a first heavy chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 85 and a constant portion of a human heavy chain having heterodimerization modifications; and b) a first light chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 101 and a constant portion of a human light chain. The constant portion of the human heavy chain may be IgG1. In one embodiment, IgG1 is human IgG1 without effector mutations. In one embodiment, the human heavy chain IgG1 comprises a silent mutation N297A, D265A, or a combination of L234A and L235A. In a particular embodiment, according to SEQ ID NO: 87, the human heavy chain IgG1 comprises a silent mutation that is a combination of L234A and L235A.

[0248] The second V of the IL-18 / IL-1β bispecific antibody used in the disclosed method H2 or V L2 The structural domain can have the same V as listed in SEQ ID NO: 53 and 69. H or V L The second V, whose structural domain is essentially the same H2 and / or the first V L2Domains. The IL-18 / IL-1β bispecific antibody disclosed herein for use in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation as disclosed in (i) may comprise a second heavy chain substantially identical to the heavy chain listed in SEQ ID NO: 55 and / or a second light chain substantially identical to the light chain listed in SEQ ID NO: 71. The IL-18 / IL-1β bispecific antibody disclosed herein for use in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation as disclosed in (i) may comprise a second heavy chain containing SEQ ID NO: 53 and a second light chain containing SEQ ID NO: 69. As disclosed herein, the IL-18 / IL-1β bispecific antibody for use in the treatment or prevention of autoinflammatory or disease-based on XIAP deficiency or CDC42 mutation as disclosed may comprise: a) a second heavy chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 53 and a constant portion of a human heavy chain having heterodimerization modifications (which are complementary to the heterodimerization of the first heavy chain); and b) a second light chain comprising a variable domain having an amino acid sequence substantially identical to that shown in SEQ ID NO: 69 and a constant portion of a human light chain. The constant portion of the human heavy chain may be IgG1. In one embodiment, IgG1 is human IgG1 without effector mutations. In one embodiment, the human heavy chain IgG1 comprises a silent mutation N297A, D265A, or a combination of L234A and L235A. In a particular embodiment, according to SEQ ID NO: 55, the human heavy chain IgG1 comprises a silent mutation that is a combination of L234A and L235A.

[0249] Other preferred IL-18 antagonists (e.g., antibodies) used as the first part of bispecific antibodies in the disclosed methods, kits, and protocols are those listed below: U.S. Patent No. 9,376,489, which is incorporated herein by reference in its entirety.

[0250] Other preferred IL-1β antagonists (e.g., antibodies) used as the second part of the bispecific antibody in the disclosed method are those listed below: U.S. Patent Nos. 7,446,175 or 7,993,878 or 8,273,350, which are incorporated herein by reference in their entirety.

[0251] Example 2: In vitro activity of bbmAb

[0252] The binding activity of bbmAb was tested in various cell assays.

[0253] (1) Materials and Methods

[0254] (a) Used for solution equilibrium titration (SET) determination

[0255] The following materials were used:

[0256] Biotinylated recombinant human IL-18 (BTP25828)

[0257] Recombinant cynomolgus monkey IL-1β (Novartis)

[0258] SULFO-TAG-labeled anti-human IgG antibody (Meso Scale Discovery, MSD) #R32AJ-5). Goat anti-human Fab-specific antibody conjugated with MSD SULFO-TAG NHS ester (Jackson Immuno Research, #109-005-097, MSD #R91AN-1), BSA (Sigma, #A-9647).

[0259] MSD Read Buffer T (MSD#R92TC-1) with Surfactant

[0260] Phosphate-buffered saline (PBS) 10x (Teknova #P0195) Tris-buffered saline pH 7.5 (TBS) 10x (Teknova #T1680) Tween-20 (Fluka #93773)

[0261] Polypropylene microtiter plate (MTP) (Greiner, Inc. #781280)

[0262] 384-well plate, standard (MSD#L21XA)

[0263] (b) Used for cell assays and SET assays

[0264] mAb2, as described in the IL-1β antibody section.

[0265] mAb1, as described in the IL-18 antibody section.

[0266] As described in Example 1, bbmAb.

[0267] Recombinant human IL-18 (BTP 25829) (#B001-5) purchased from MBL Int. Corp.

[0268] Recombinant chordate IL-1β (Novartis, Inc.)

[0269] Recombinant velvet monkey IL-18 (Novartis Inc.)

[0270] Recombinant human IL-12 (#573008) was purchased from Biolegend KG-1 cell line (ATCC#CCL-246).

[0271] Normal human skin fibroblasts (#CC-2509) were purchased from Lonza.

[0272] Marmoset skin fibroblasts (#42637F(510))

[0273] HEK-Blue™ IL-18 / IL-1β cells (#hkb-il18) were purchased from InvivoGen.

[0274] PBMCs were isolated from the erythrocyte sedimentation rate (ESR) layer (obtained from Blustspendezentrum Bern).

[0275] The blood of the marmoset was obtained from Silabe, Niederhausbergen.

[0276] IL-6 ELISA: Human (Bocci, #430503); Marmoset (U-CyTech Biosciences, CT974-5)

[0277] IFNγ ELISA: Human (BD555142) and marmoset (U-CyTech Biosciences #CT340A)

[0278] The QUANTI-Blue™ assay (#rep-qb1) for detecting SEAP was purchased from InvivoGen.

[0279] Cell culture medium: RPMI 1640 (Invitrogen #31870), supplemented with 10% fetal bovine serum (Invitrogen #10108-157), 1% L-glutamine (Invitrogen #25030-03), 1% penicillin / streptomycin (Invitrogen #15140-148), 10 µM 2-mercaptoethanol (Gibco #31350-010), and 5 mM Hepes (Gibco #15630-080).

[0280] 96-well plates with round bottoms for tissue culture (Costar #3799)

[0281] Flat-bottomed 96-well plate for tissue culture (Costar #3596)

[0282] Ficoll-Pacque™ Plus (GE Healthcare LifeSciences #17-1440-02) PBS 1 X, calcium and magnesium free (Gibco #14190094)

[0283] Leucosep tubes with porous barrier, 50 ml, polypropylene (Greiner bio-one #227290).

[0284] Falcon 15 ml polypropylene tapered tube (BD Company #352096)

[0285] Falcon 50 ml polypropylene tapered tube (BD Company #352070)

[0286] (c) Affinity measurement via SET

[0287] SET Single Target Binding Assay

[0288] Twenty-two consecutive 1.6n dilutions of the antigen (maximum concentrations: huIL-18, 5 nM; marIL-18, 10 nM; huIL-1β, 0.5 nM; marIL-1β, 0.5 nM) were prepared in sample buffer (PBS containing 0.5% bovine serum albumin (BSA) and 0.02% Tween-20) with constant concentrations of antibody added (10 pM for IL-18 and 1 pM for IL-1β). Each antigen-antibody mixture was partitioned twice into 384-well polypropylene microtiter plates (MTP) at a volume of 60 µl / well. Sample buffer served as a negative control, and samples containing only the antibody served as a positive control (no maximum electrochemiluminescence signal of the antigen, B...). max Seal the plate and incubate it overnight (o / n, at least 16 hours) on a shaker at room temperature (RT).

[0289] IL-18 readings: Streptavidin-coated 384-well MSD arrays MTP were coated with 30 µl / well biotinylated huIL-18 (0.1 µg / ml, PBS) and incubated on a shaker at room temperature for 1 h.

[0290] IL-1β readings: The standard 384-well MSD array MTP was coated with 30 µl / well of huIL-1 (3 µg / ml, PBS) diluted in PBS (as a capture agent) and incubated overnight at 4°C.

[0291] At room temperature (RT), the plate was blocked with 50 µl / well of blocking buffer (PBS containing 5% BSA) for 1 hour (h). After washing (TBST, TBS containing 0.05% Tween 20), a balanced antigen-antibody mixture of 30 µl / well was transferred from the polypropylene MTP to the coated MSD plate and incubated at room temperature for 20 min. After another washing step, 30 µl of sulfonyl-tagged anti-IgG detection antibody (0.5 µg / ml) diluted in sample buffer was added to each well and incubated on a shaker at room temperature for 30 min. The MSD plate was washed, and 35 µl / well of MSD read buffer was added and incubated at room temperature for 5 min. Electrochemiluminescence (ECL) signals were generated and measured using an MSD Sector Imager 6000.

[0292] SET Simultaneous Target Binding Measurement

[0293] Except for assay A, the SET assay is performed as described above: the equilibration process (antibody / antigen mixture) is performed in the presence of an excess of one target (500 pM of IL18 or IL-1β) while simultaneously assessing the K of the other target. D .

[0294] Assay B: The equilibration process (antibody / antigen mixture) involves simultaneously performing serially diluted versions of the two targets in a single mixture (constant antibody concentration of 10 pM, maximum antigen concentration as described above). The concentration of free antibody in the same mixture on IL18 and IL-1β-coated plates is then analyzed as described above.

[0295] SET data was exported to the MS Excel add-in software Xlfit. The average ECL signal was calculated from repeated measurements in each assay. Baseline adjustment was performed by subtracting the lowest value from all data points, and titration curves were generated against the corresponding antigen concentrations. D The value is determined by fitting the graph as follows:

[0296] 1:2 binding model of monospecific Abs

[0297]

[0298] A 1:1 binding model of bispecific Abs in the mortar and pestle structure.

[0299]

[0300] in

[0301] y: Subtract the blank ECL signal

[0302] B maxThe maximum ECL signal when the antigen concentration is zero.

[0303] [IgG]: The concentration of the applied antibody

[0304] [Fab]: Total Fab concentration of the application

[0305] K D Dissociation equilibrium constant

[0306] x: The concentration of the applied antigen

[0307] (d) Cell culture

[0308] KG-1 cells were grown in RPMI 1640 supplemented with 10% fetal bovine serum, 1% L-glutamine, and 1% penicillin / streptomycin at a density of 2 x 10⁻⁶ cells / mL. 5 Up to 1 x 10 6 live cells / mL.

[0309] Normal human fibroblasts and marmoset fibroblasts were grown in an FBM (Clonetics, CC-3131) containing bFGF (1 ng / ml, CC-4065), insulin (5 μg / ml, CC-4021), and 2% FCS (CC-4101). Fibroblast basal medium (Lonza # CC-3131) was used as the starvation medium.

[0310] HEK-Blue™ IL-18 / IL-1β cells were grown in growth medium (DMEM, 4.5 g / L glucose, 10% (v / v) fetal bovine serum, 50 U / ml penicillin, 50 mg / ml streptomycin, 100 mg / ml Normocin™, 2 mM L-glutamine, supplemented with 30 μg / ml blastomycin, 200 μg / ml HygroGold™ and 100 μg / ml Zeocin™).

[0311] Human peripheral blood mononuclear cells (PBMCs) were freshly isolated from the erythrocyte sedimentation rate (ESR) amber layer using LeucoSep tubes, according to the manufacturer's instructions. Briefly, 13 ml of Ficoll-Paque was pre-packed into 14 ml LeucoSep tubes by centrifugation at 1,000 × g for 30 s. An equal volume of heparinized whole blood sample was diluted with PBS, and 25 ml of the diluted blood was added to the LeucoSep tube. The cell separation tubes were centrifuged continuously at 800 × g for 15 min at room temperature. The cell suspension layer was collected, and the cells were washed twice in PBS (at 640 × g and 470 × g for 10 min, respectively, two consecutive washes) and resuspended in culture medium, then counted.

[0312] Marmoset blood was collected in heparinized tubes and filtered using a 70 µm cell filter (BD Biosciences #352350).

[0313] (e) IL-1β neutralization assay

[0314] The assay of IL-1β-induced IL-6 production in fibroblasts was performed essentially as described in (Gram 2000), with only minor modifications. Briefly, fibroblasts were seeded at a density of 5 x 10³ cells / well (in 100 µl) in 96-well flat-bottom tissue culture plates. The next day, cells were starved in starvation medium for 5 h before the addition of the recombinant IL-1β / compound solution mixture (IL-1β concentrations shown in the table). The IL-1β / compound solution mixture was prepared in advance by incubating the recombinant IL-1β with the compound at a concentration range at 37°C for 30 min. After incubation at 37°C, the cell supernatant was collected, and the amount of IL-6 released was determined by ELISA. The assay of IL-1β-induced IL-6 production in PBMCs was performed according to the following. PBMCs were seeded at a density of 3 x 10³ cells / well (in 100 µl). 5 Cells / well (100 µl) were seeded in 96-well tissue culture plates and incubated with a recombinant IL-1β / compound solution mixture at 37°C for 24 h (IL-1β concentrations shown in the table). The IL-1β / compound solution mixture was prepared in advance by incubating recombinant IL-1β with compounds at concentration ranges at 37°C for 30 min. Cell supernatants were collected 24 h after stimulation, and the amount of IL-6 released was determined by ELISA.

[0315] (f) IL-18 neutralization assay

[0316] The determination is basically performed according to the following: A density of 3 x 10⁻⁶ is used. 5 KG-1 cells (pre-starved for 1 h in PBS + 1% FCS) or PBMCs were seeded into round-bottom 96-well cell culture plates and incubated with a solution mixture of recombinant IL-18 / IL-12 and the compound at concentration ranges (IL-18 / IL-12 concentrations shown in the table). After incubation at 37°C for 24 h, the supernatant was collected and the amount of IFNγ released was determined by ELISA. For assays using marmoset blood, 85 µl of blood / well was used.

[0317] (g) HEK-Blue™ cells neutralize dual IL1β / IL-18

[0318] The assay was performed essentially as described in the manufacturer's processing procedure. In short, HEK-Blue™ cells were placed at 4 x 10⁻⁶...4 The cells were seeded at a density of 1 / 2 well into 96-well cell culture plates and incubated with a solution mixture of recombinant IL-1β and IL-18 (to generate a 1:1 SEAP signal) and compounds in a range of concentrations. After incubation at 37°C for 24 h, the supernatant was collected and the amount of SEAP released was determined using the QUANTI-Blue™ method according to the manufacturer’s instructions.

[0319] Export all data to Excel software and calculate IC50 values ​​by plotting dose-response curves against the logistic curve fitting function using Excel / XLfit4 or GraphPad Prism software.

[0320] (2) Results

[0321] (a) Affinity to recombinant human and marmoset IL1β and IL-18

[0322] The binding affinity of bbmAb to recombinant IL-1β and IL-18 proteins in humans and marmosets was measured by solution equilibrium titration (SET), and the generated K... D The value of K binding to mAb2 for IL-1β and mAb1 for IL-18 D Values ​​were compared. Binding affinity was compared in single-target binding assays, and bbmAb showed similar mean KD for both human and marmoset IL-18 compared to mAb1 (Table 4). For human IL-1β binding, bbmAb (2.6 pM) had a slightly higher mean KD value compared to mAb2 (0.6 pM), but remained within the same low pM range. Subsequent measurements in simultaneous dual-target binding assays (Table 5) confirmed that the binding KD value of bbmAb for IL-1β was similar to that of mAb2, both using preclinical and clinical-grade materials. Therefore, bbmAb has binding affinity for targets in both humans and marmosets, similar to mAb2 and mAb1, respectively.

[0323] Table 4. Affinity to recombinant human (hu) and marmoset (mar) IL-1β and IL-18 as measured by SET (single target binding assay)

[0324]

[0325] In addition to the results of binding to a single target, it is also applied to the evaluation of binding to K DSimultaneous dual-target binding affinity of bbmAb was studied by applying either an excess of one target relative to another (Assay A) or by using a mixture of two targets in serial dilutions (Assay B) (Table 5). Simultaneous IL-1β / IL-18 affinity assays showed no significant difference between Assay A (excess of one antigen) and Assay B (a mixture of two antigens in serial dilutions), demonstrating that simultaneous binding of two targets does not affect the binding of the other target. Furthermore, the Ka obtained from the simultaneous dual-binding assays… D K value obtained from standard determination D The values ​​are similar (Table 4; in the absence of a second antigen), demonstrating that bbmAb can independently bind to both antigens. Therefore, bbmAb binds to both human IL-1β and IL-18 simultaneously and independently, and completely cross-reacts with the corresponding malathion proteins.

[0326] Table 5. Affinity to recombinant human (hu) and marmoset (mar) IL-1β and IL-18 as measured by SET (simultaneous target binding assay)

[0327]

[0328] (b) Neutralizing activity of bbmAb in human and marmoset cell assays

[0329] The neutralizing activity of bbmAb against two cytokines (IL1β and IL-18) was evaluated (mAb2mAb1). Additionally, the potency of bbmAb in neutralizing IL-1β and IL-18 in marmosets was assessed using a marmoset assay system (see section d).

[0330] (c) Neutralization of IL-1β and IL-18, both individually and simultaneously, in human cells

[0331] The neutralizing activity of bbmAb against IL-1β was assessed by inhibiting recombinant IL-1β-induced IL-6 production in human skin fibroblasts (IL-1β administered at 6 pM) and human PBMCs (IL-1β administered at 60 pM). The neutralizing activity of bbmAb against IL-18 was measured by inhibiting recombinant IL-18-induced IFN-γ production in KG-1 cells and human PBMCs (both cell types activated with 3 nM recombinant human IL-18 and 1 ng / ml recombinant human IL-12). The inhibitory potency of bbmAb against IL-1β and IL-18 was always compared to that of mAb2 or mAb1, respectively. Depending on the assay, the mean IC50 value of bbmAb was in the sub-nM or single-digit nM range, while being up to 2 to 4 times higher directly than mAb2 (for IL-1β) and mAb1 (for IL-18), respectively (Tables 6 and 7). The monovalent form of bbmAb, compared to the bivalent form of mAb2 / mAb1, but possibly due to underlying KiH mutations, is the reason for this subtle difference in bbmAb efficacy.

[0332] Table 6. Mean IC50 values ​​of IL-1β in bbmAb compared to mAb2 in human skin fibroblasts and human PBMCs. Inhibition of IL-6 production in human skin fibroblasts or PBMCs stimulated with recombinant human IL-1β (6 pM for skin fibroblasts and 60 pM for PBMCs). Figures shown are mean ± SEM (n = 3 PBMCs and n = 6 human skin fibroblasts).

[0333]

[0334] Table 7. Mean IC50 values ​​of IL-18 in bbmAb compared to mAb1 in KG-1 cells and human PBMCs. Inhibition of IFNγ production in KG-1 cells or PBMCs stimulated with recombinant human IL-18 (3 nM) and human IL-12 (1 ng / ml). Figures shown are mean ± SEM (n = 3 KG-1 cells and n = 4 PBMCs).

[0335]

[0336] bbmAb can simultaneously neutralize the biological activity of both IL-1β and IL-18, as demonstrated by the use of HEK Blue™ reporter cells (which produce SEAP in response to a 1+1 stimulation of recombinant IL-1β and IL-18) (Table 8). Similar inhibition of SEAP can only be achieved in this assay system by combination of mAb2 and mAb1, and not by using a single antibody.

[0337] Table 8. Mean IC50 values ​​for simultaneous neutralization of IL-1β and IL-18 based on SEAP reporter activity in HEK Blue™ cells. Showing mean ± SEM values ​​from n = 5 experiments.

[0338]

[0339] (d) Neutralizing activity of bbmAb against marmoset IL-1β and IL-18 in marmoset cell assays

[0340] To demonstrate the inhibitory activity of bbmAb1 in marmosets, similar in vitro assays were performed using marmoset cells and human cells, but with stimulation using recombinant marmoset IL-1β and IL-18. When assessing the inhibitory effect on IL-6 production induced by recombinant marmoset IL-1β in marmoset skin fibroblasts, bbmAb1 exhibited sub-nM potency, with an IC50 value 2 to 3 times higher than mAb2 (Table 9). Human skin fibroblasts stimulated with marmoset IL-1β produced an inhibitory spectrum similar to that produced with human IL-6.

[0341] Table 9. bbmAb1 inhibits the production of IL-6 induced by recombinant marmoset IL-1β in marmosets and human fibroblasts. Inhibition of IL-6 production in marmoset or human skin fibroblasts stimulated by recombinant marmoset IL-1β (18 pM). Results from three separate experiments (A, B, and C) are shown.

[0342]

[0343] The single- to double-digit nM IC50 values ​​of bbmAb1 confirmed its neutralizing activity against marmoset IL-18 tested in an IFNγ production assay using marmoset blood cells (Tables 4-7). When human IFNγ production was measured, bbmAb1 produced a similar inhibitory spectrum in human PBMCs stimulated with marmoset IL-18.

[0344] Therefore, in functional assays using marmoset response cells, bbmAb1 showed complete cross-reactivity with marmoset IL-1β and marmoset IL-18.

[0345] Table 10. Mean IC50 values ​​of inhibition of IFNγ production induced by recombinant marmoset IL-18 in whole blood or human PBMCs of marmosets. Inhibition of IFNγ production in whole blood of marmosets (n=3 per compound / condition) or human PBMCs (n=6) stimulated with recombinant marmoset IL-18 (concentration shown) and human IL-12 (10 ng / ml). Figures shown are mean ± SEM.

[0346]

[0347] It has been demonstrated that bbmAb1 (a KiH-type IL-1β / IL-18 bispecific mAb) retains high affinity binding to both individual targets, IL-1β and IL-18, and cytokine neutralizing potency in a variety of different cell assays compared to the original mAbs (mAb2 and mAb1). The dual IL-1β and IL-18 neutralizing properties of bbmAb1 have been demonstrated not only against human cytokines / cells but also against corresponding marmoset cytokines / cells, thus facilitating appropriate toxicology studies. The up to 2-4 times higher IC50 values ​​produced in some cell assays targeting IL-1β and IL-18 neutralization may be a result of monovalent binding of bbmAb1 rather than divalent binding of mAb2 and mAb1 respectively. However, the dual neutralization of cytokines by bbmAb1 may lead to additive or synergistic inhibitory activity in vivo, which may not be fully reflected in our in vitro cell systems.

[0348] Example 3: Effects of combined stimulation and blockade of IL-1β and IL-18 in PBMCs

[0349] Inflammasome activation-dependent cleavage of effector cytokines IL-1β and IL-18 induces secondary pro-inflammatory mediators that promote immune cell recruitment / activation not only systemically but also at sites of inflammation. In two distinct lethal mouse models of systemic inflammation (a) an LPS injection model and (b) FCAS mice (with a missense mutation activating NLRP3), the simultaneous absence / inhibition of both IL-1β and IL-18 was more protective against lethality than the absence / inhibition of IL-1β or IL-18 alone, demonstrating an additive or synergistic mechanism of immune activation (Brydges 2013, van den Berghe 2014). bbmAb1 is a bispecific mAb reactive to human / marmoset IL-1β / IL-18, lacking rodent cross-reactivity, and therefore could not be tested in mouse models. Therefore, we used LPS / IL-12 to simulate inflammasome-dependent pathway activation in vitro to stimulate human PBMCs to reveal the additive or synergistic inhibitory effects of the bbmAb1-neutralized combination of IL-1β / IL-18, and performed unbiased gene expression analysis using microarrays. As a complementary activity, we also compared gene expression profiles from PBMCs of different donors stimulated with a combination of recombinant IL-1β and recombinant IL-18 or with individual cytokines.

[0350] (3) Materials and Methods

[0351] (a) Cell culture and ELISA

[0352] RPMI 1640 (Ingenieur #31870 or Gibco #61870-010) is supplemented with 10% fetal bovine serum (Ingenieur #10108-157), 1% L-glutamine (Ingenieur #25030-03), 1% penicillin / streptomycin (Ingenieur #15140-148), 10 µM 2-mercaptoethanol (Gibco #31350-010), and 5 mM Hepes (Gibco #15630-080).

[0353] Recombinant human IL-1β was purchased from Sinocare Biotech Co., Ltd. (#10139-HNAE-5).

[0354] Recombinant human IL-18 was purchased from MBL (#B001-5).

[0355] The recombinant human IL-12 was purchased from Boch Corporation (#573008).

[0356] IFNγ ELISA: MAX Standard Kit, Boch Company, #430103 or BD OptEIA IFNγ ELISA Kit, BD#555142

[0357] IL-6 ELISA: MAX Standard Kit, Boch Company, #430503

[0358] IL-26 ELISA: Cloud Clone Corp. #SEB695Hu

[0359] mAb2, as described in the IL-1β antibody section.

[0360] mAb1, as described in the IL-18 antibody section.

[0361] As shown in Example 1, bbmAb1.

[0362] LPS is derived from Salmonella enteritidis serotype 6, Sigma-Aldrich #L7770

[0363] PBMCs were isolated from the erythrocyte sedimentation rate (ESR) layer (obtained from Blustspendezentrum Bern).

[0364] 96-well plates with round bottom for tissue culture (Costar #3799) 96-well plates with flat bottom for tissue culture (Costar #3596) Ficoll-Pacque™ Plus (GE Healthcare Life Sciences #17-1440-02) PBS 1X, calcium and magnesium free (Gibco #14190094)

[0365] Falcon 15 ml polypropylene tapered tube (BD Company #352096) Falcon 50 ml polypropylene tapered tube (BD Company #352070)

[0366] Leucosep™ tubes with porous barrier, 50 ml, Greiner bio-one #227290

[0367] Cell filter 70 µM, BD Biosciences #352350

[0368] Trypan Blue, Sigma Corporation #T8154

[0369] RNA isolation, quantity and quality measurement, and qPCR:

[0370] Nuclease-free water, Ambion Corporation #AM9938

[0371] Rnase Zap, Ambion Company #AM9780

[0372] 1.5 ml Eppendorf tubes, sterile, free of ribonuclease and deoxyribonuclease.

[0373] RLT buffer, Qiagen #1015762

[0374] Rneasy Mini Reagent Kit, Qiagen #74104

[0375] Deoxyribonuclease kit without ribonuclease, Qiagen #79254

[0376] Agilent RNA 6000 Nano Kit, Agilent Technologies #5067-1511

[0377] Chip trigger station, Agilent Technologies #5065-4401

[0378] IKA Vortex Mixer

[0379] RNaseZAP®, Ambion Corporation #9780

[0380] Agilent Technologies 2100 Bioanalyzer

[0381] High-capacity cDNA reverse transcription kit, Applied Biosystems, #PN4374966

[0382] Nase-free, thin-walled, capped 0.2 ml PCR tubes, Ambion, #AM12225

[0383] MicroAmp Optical 384-well reaction plate, Applied Biosystems, #4309849

[0384] TaqMan GenEx premixed solutions, Applied Biosystems, #4369514

[0385] PCR primers (Applied Biosystems)

[0386]

[0387] PBMC Preparation: PBMCs were isolated from the ESR (erythrocyte sedimentation rate) amber layer by Ficoll-Paque gradient centrifugation in Leucosep tubes, following the manufacturer's instructions. Briefly, 15 mL of Histopaque was placed in a 50 mL Leucosep™ tube and centrifuged at 1300 rpm for 30 seconds at room temperature. 30 mL of the diluted suspension of the ESR amber layer was added to the top of the Histopaque solution using a pipette, and the tube was centrifuged continuously at 1000 g for 15 min at room temperature. The plasma (approximately 20 mL) was discarded, and the interfacial loop (= human PBMCs) was collected and transferred to a 50 mL Falcon tube. The tube was filled with 50 mL of sterile PBS and centrifuged once at 1200 rpm for 5 min at room temperature. This centrifugation was repeated twice. The supernatant was gently discarded, and the cells were resuspended in 50 mL PBS containing 2% FCS and 2 mM EDTA. The cell suspension was filtered using a 70 µm cell filter, and the cells were counted using trypan blue staining (500 µL trypan blue + 200 µL cells + 300 µL PBS).

[0388] LPS / IL-12 Stimulation of PBMCs: Cytokine production in the supernatant was prepared according to the following method. 250,000 cells / well (final volume 100 μl) were dispensed into 96-well round-bottom plates. LPS was applied at concentrations ranging from 0.3 μg / ml to 3000 μg / ml, along with 10 ng / ml of recombinant IL-12. The supernatant was harvested after 24 h at 37°C and 10% CO2.

[0389] RNA extraction from cell pellets is performed according to the following procedure. Take 3 x 10 6 Cells / well were fractionated into 24-well plates with a final volume of 1000 μl. Cells were harvested after incubation at 37°C and 10% CO2 for 24 h using 3 μg / ml LPS along with 10 ng / ml recombinant IL-12.

[0390] Stimulation of PBMCs with recombinant cytokines: 7 x 10⁶ cells per well of a 12-well plate. 6 One PBMC was used in 1.5 ml of final complete RPMI medium. Recombinant cytokines were added at the following final concentrations: 10 ng / ml recombinant IL-1β, 3 nM recombinant IL-18, and 1 ng / ml recombinant IL-12. Both supernatant and cells were collected after 4 h and 24 h at 37°C and 10% CO2.

[0391] RNA isolation, quantity and quality assessment: Cell precipitation was performed by lysing the precipitate in 350 µl of Qiager RTL buffer containing 2% β-mercaptoethanol and freezing at -20°C or -80°C until all study samples were collected. RNA isolation was performed using Qiager's standard protocol. Briefly, 350 µl of 70% ethanol was added to all samples, which were then transferred to an RNeasy column and centrifuged at 8000 g for 15 s. After discarding the runoff, 350 μl of buffer RW1 was added, and the column was centrifuged at 8000 g for 15 s to wash the column membrane. A deoxyribonuclease I incubation mixture was prepared according to the manufacturer's instructions and added to the RNeasy column, which was incubated at room temperature for 15 min. After washing with 350 μl and 500 μl of buffer RW1, the RNeasy column was transferred to a new 2 ml collection tube and centrifuged at full speed for 1 min. RNA was finally collected by adding 35 μl of ribonuclease-free water directly to the centrifuge column membrane and centrifuging at 8000 g for 1 min to elute RNA. The amount of RNA was measured using a Nanodrop ND-1000, and the RNA was stored at -20°C. RIN measurements were performed according to the manufacturer's instructions for RNA quality assessment. In short, 1 µl of RNA or a gradient was pipetted into an Agilent RNA 6000 Nano chip and measured using an Agilent 2100 Bioanalyzer.

[0392] Cytokine gene expression was analyzed by qPCR:

[0393] Perform this method according to the manufacturer's instructions. In short, reverse transcribe 400 ng of RNA using the High-Capacity cDNA Reverse Transcription Kit according to the instructions. Dilute the cDNA solution 1 / 10 with RNA / DNA-free water, then transfer 1 µl of cDNA to a 384-well plate and mix with 1 µl of 20X TaqMan® gene expression assay target FAM gene, 10 µl of 2xTaqMan® gene expression premix, and 10 µl of RNA / DNA-free water. Load the plate onto an Applied Biosystems ViiA™ 7 Real-Time PCR System and use the following instrument settings:

[0394]

[0395] The housekeeping genes used in this study were HPRT1 and RLP27. The relative expression levels of the target genes were calculated using the following formula:

[0396] 1) Ct [reference] = (Ct [HPRT1] + Ct [RLP27]) / 2

[0397] 2) dCt [reference] = 40 - Ct [reference]

[0398] 3) dCt [target] = Ct [target] - Ct [reference]

[0399] 4) ddCt = dCt [reference] - dCt [target]

[0400] 5) Relative target gene expression = 2^ddCt

[0401] Microarrays were performed according to the following methods. Samples were processed by CiToxLAB France on an Affymetrix HG_U133_Plus2 microarray. They were RMA normalized and analyzed in GeneSpring 11.5.1 (Agilent Technologies, Santa Clara, CA). Pathway analyses were performed using Ingenuity Pathway Analysis (IPA) and Nextbio (Illumina). The two datasets were processed independently.

[0402] Initially, the data underwent standard quality control (QC) by CiToxLAB, with internal QC performed using an R script (MA_AffyQC.R) in the Rstudio suite and GeneSpring (PCA, hybridization control). Subsequently, the data were filtered to eliminate unreliable expression levels: entities (probe sets) were retained where at least 100% of the samples had values ​​above the 20th percentile in any one experimental condition.

[0403] Differentially expressed genes (DEGs) were identified using the "filter on volcano plot" feature in GeneSpring. For the filtered genes (expressed between the 20.0 and 100.0 percentiles), probe groups with a corrected p-value below 0.05 and a fold change greater than 2.0 were considered differentially expressed using an unpaired T-test. Where possible, i.e., in the LPS (NUID-0000-0202-4150) study, Benjamini-Hochberg multiplex testing was used for correction.

[0404] For cytokine stimulation experiments, synergistic effects are calculated using the following formula: Signal A + B / (Signal A + Signal B - Control) ≥ 1.5

[0405] Each feature (or DEG list) is used to calculate the p-value in the Fisher exact test case, which represents the statistical significance of the overlap between the observed feature and the "disease gene list" (lesions versus non-lesions) in the public dataset. To do this, the list is uploaded to the Enomina Basic Spatial Association Engine (formerly Nextbio) and compared using meta-analysis and disease-specific keyword searches.

[0406] Export all data to Excel and calculate IC by plotting dose-response curves against the logistic curve fitting function using Excel / XLfit4 or GraphPad Prism software. 50 Values ​​were determined using GraphPad Prism software. Differences between treatment groups were analyzed using one-way ANOVA followed by Dunnett multiple comparisons, and results were considered statistically significant when p < 0.05.

[0407] (4) Results

[0408] (a) bbmAb1 is highly effective in inhibiting LPS / IL-12-induced IFNγ production in whole blood.

[0409] Exposure of human whole blood to LPS supplemented with 10 ng / ml IL-12 resulted in an IFNγ response, which was largely (but not entirely) dependent on “natural” IL-18 produced by blood cells. The addition of IL-12 may enhance the LPS-induced IFNγ response by upregulating the IL-18 receptor on responder cells.

[0410] Under the experimental conditions used, IL-18 neutralization with mAb1 resulted only incomplete inhibition of IFNγ production, while IL-1β blockade (with mAb2) had only a small effect on the IFNγ response. Interestingly, compared to single cytokine neutralization, the combination of bbmAb1 or mAb2 and mAb1 resulted in a more profound and complete inhibition of IFNγ production through combined inhibition of IL-1β and IL-18.

[0411] In our cellular assays, no other tested cytokines (IL-2, -4, -6, -8, -10, -13, and TNFα) were inhibited by the combined neutralization and additive effect of IL-1β and IL-18 (data not shown). Considering the monovalent form of the bispecific molecules, the potency of bbmAb1 is in the same range as the combination of mAb2 and mAb1.

[0412] (b) In LPS / IL-12 activated human PBMCs, IFNγ was cumulatively inhibited by bbmAb1 (i.e., combined IL-1β / IL-18 inhibition) compared to single IL-1β or IL-18 inhibition.

[0413] Unbiased transcriptomic assessment was required to reveal additional additive effects (besides IFNγ) using bbmAb1 through combined IL-1β / IL-18 inhibition. Since whole blood is not the optimal material for transcriptomic analysis, we adapted the LPS / IL-12 stimulation assay conditions (as described in the Materials and Methods section above) to human PBMC samples. Using PBMCs from a total of nine donors, we were able to confirm that bbmAb1 cumulatively inhibited IFNγ protein secretion into the PBMC supernatant. IFNγ production was suppressed at approximately 10-fold lower concentrations than in whole blood assays compared to the individual mAbs used. Importantly, a similar pattern of inhibition was observed at the IFNγ mRNA level, confirming the suitability of the samples for gene expression analysis based on unbiased microarrays. The data showed that bbmAb1, mAb2, and mAb1 (each at 10 nM) inhibited LPS (0.3 µg / ml) / IL-12-induced IFNγ protein production and IFNγ gene expression in human PBMCs.

[0414] The Affymetrix microarrays were performed using n = 5 individual donors from PBMCs sampled from the LPS / IL-12 stimulation assays described in the Materials and Methods section above. Unfortunately, overall assessment of gene expression profiles confirmed strong LPS / IL-12 stimulation, with PCA showing clustering of compounds per donor rather than within stimulated or unstimulated groups. However, comparisons of LPS / IL-12-stimulated samples with stimulated samples plus bbmAb1 for differentially expressed genes revealed a list of genes downregulated by the combined IL-1β / IL-18 blockade using bbmAb1 (Table 11). In addition to the strong downregulation of the IFNγ gene (which reaffirmed our microarray data), the IL-26 gene was another cytokine gene that was cumulatively suppressed by bbmAb1 compared to single IL-1β inhibition (via mAb2) or IL-18 inhibition (via mAb1). Gene expression levels of IFNγ and IL-26 derived from microarray data were observed, as well as the inhibitory effects of bbmAb1, mAb2, and mAb1 (each 10 nM) on PBMCs stimulated with LPS (0.3 μg / ml) / IL-12 at 24 h.

[0415] Table 11. Differentially expressed genes (genes downregulated only between bbmAb1 and the control group in LPS / IL-12 stimulated samples). FC = fold change.

[0416]

[0417] (c)IL-26 is another pro-inflammatory cytokine that is cumulatively inhibited by bbbAb1 in LPS / IL-12 stimulated PBMCs.

[0418] To further confirm that bbmAb1 most effectively inhibits LPS / IL-12-driven IL-26 gene expression and protein production via combined IL-1β / IL-18 blockade, the study was expanded to a total of n = 9 PBMC donors, with IL-26 gene expression investigated by qPCR and IL-26 protein production investigated by ELISA. The results largely confirmed the inhibition of IL-26 gene expression obtained via the microarray method. Interestingly, the addition of mAb resulted in only a partial reduction in IL-26 protein levels in the supernatant at 24 h. The reason for this difference is unclear, but it may be related to the kinetic differences between IL-26 gene expression and protein production, as well as the difference in IL-26 consumption compared to IFNγ. However, bbmAb1 showed an advantage in reducing IL-26 protein levels in PBMC supernatant compared to mAb2 and mAb1. The results showed that bbmAb1, mAb2 and mAb1 (10 nM each) inhibited LPS (0.3 μg / ml) / IL-12-induced IL-26 gene expression (by qPCR) and IL-26 protein levels in human PBMCs.

[0419] (d) Disease-related IL1β / IL18 signaling characteristics

[0420] Previously established PBMC culture conditions (where recombinant IL-1β stimulation leads to IL-6 production or recombinant IL-18 / IL-12 stimulation leads to IFNγ production) were combined to reveal additive or synergistic downstream target genes or signatures (data not shown). PBMCs from n = 4 donors were sampled at two different time points (6 h and 24 h) for Affymetrix microarray evaluation to perform unbiased assessment of gene expression profiles. Genes synergistically upregulated at 6 h and 24 h under combined IL-1β and IL-18 stimulation were revealed (data not shown). Adding IL-12 to the IL-1β / IL-18 combination significantly enhanced the synergistic effect of a range of upregulated genes. Signal transduction signatures produced by single or combined IL-1β / IL-18 pathway stimulation (genes upregulated only) were used to inquire about datasets spanning several autoimmune diseases. For example, a correlation was observed with a common sarcoidosis dataset. The p-values ​​(calculated using Fisher's exact test) show significant correlations with several public studies comparing healthy tissues and diseased tissues from sarcoidosis patients. Tissues included skin, as well as lungs, lacrimal glands, and the anterior orbit. Among all datasets, the combination of IL1β / IL18 signaling showed the best correlation with the disease, followed by IL-1β and IL-18 alone. Differential upregulated genes for IL-1β / IL-18 in PBMCs compared to the five sarcoidosis tissue "disease-to-health" DEGs (x-axis). The p-values ​​(y-axis) represent the statistical significance of the overlap between the observed features and the "disease gene list." Black bars represent skin from sarcoidosis lesions relative to skin from healthy patients. Light gray bars represent skin from sarcoidosis lesions relative to non-lesion skin. White bars represent lacrimal glands from sarcoidosis patients relative to lacrimal glands from healthy individuals. Dark gray bars represent anterior orbital tissue from sarcoidosis patients relative to anterior orbital tissue from healthy individuals. Striped bars represent lung samples from progressive fibrosis and sarcoidosis, as opposed to lung samples from nodular self-limiting sarcoidosis.

[0421] (e) Conclusion

[0422] LPS and recombinant IL-12 were used to mimic pathogen-associated molecular pattern (PAMP)-dependent NLRP3 inflammasome activation during the first 24 hours of in vitro culture. It was demonstrated that the cumulative effect of LPS / IL-12 stimulation in PBMCs was reduced / inhibited by using bbmAb, IL-1β, and IL-18. The synergistic effect of IL-12 and IL-18 to induce IFNγ production in T, B, NK cells, macrophages, and dendritic cells has been previously described (as reviewed by Nakanishi, 2001), but it can now be demonstrated under the experimental conditions used that IL-1β has an additional stimulatory effect on IFNγ. Therefore, co-incubation of PBMCs with LPS / IL-12 effectively drove the production of “native” IL-1β and IL-18 (both of which contribute to a strong IFNγ response). Using unbiased microarray transcriptomics, additional genes were identified that were neutralized and cumulatively downregulated by a combination of IL-1β / IL-18, relative to single IL-1β or IL-18 blockade. These included IL-26, a member of the IL-20 cytokine subfamily (IL-19, IL-20, IL-22, IL-24, and IL-26), which is conserved in most vertebrate species but absent in most rodent strains (including mice and rats) (Donnelly 2010). It transmits signals via a heterodimeric receptor complex composed of the IL-20R1 and IL-10R2 chains. The IL-26 receptor is primarily expressed on non-hematopoietic cell types, particularly epithelial cells. Elevated IL-26 levels have been reported in serum and, especially, in the synovial fluid of RA patients, suggesting that IL-26 may function as a factor promoting Th17 cell growth and differentiation. Unfortunately, the strong effect of LPS / IL-12 stimulation of PBMC samples hindered the discovery of additional genes / pathways induced by combined blockade of IL-1β and IL-18. However, IFNγ and IL-26, as well as IL-22, were also found to some extent in genes co-regulated by combined stimulation of IL-1β and IL-18 in PBMCs, confirming that these two factors are downstream effectors of this activation pathway. Therefore, the IL-20 subfamily of cytokines (including IL-26 and IL-22) appears to be strongly dependent on simultaneous signaling from IL-1β and IL-18. These comparisons, taking into full account the selectivity of individual signaling signatures and the potential efficacy of blockade, help to demonstrate the activity of various pathways in a variety of inflammatory diseases.

[0423] Example 4: Therapeutic Use

[0424] A three-phase, multicenter study using a randomized, double-blind, placebo-controlled design was conducted to evaluate the clinical efficacy, safety, and tolerability of bbmAb in autoinflammatory or disease-associated conditions based on XIAP deficiency or CDC42 mutation.

[0425] Summary of Plans

[0426]

[0427] 1. Introduction

[0428] 1.1. Background

[0429] X-linked apoptosis inhibitor (XIAP) deficiency

[0430] X-linked inhibitor of apoptosis (XIAP) deficiency is a rare, inherited immunodeficiency that occurs almost exclusively in young men (Rigaud et al. 2006). XIAP exhibits pleiotropic functions in cell survival, innate immunity, and inflammation. In particular, in addition to its role in regulating caspase activity, XIAP is considered an essential regulator of NLRP3 inflammasome activation (Miyazawa and Wado 2022). Key features of XIAP deficiency include inflammatory bowel disease, typically presenting as abdominal pain and diarrhea, relapsing fever, splenomegaly, and hemophagocytic lymphohistiocytosis (HLH). The latter manifestation is usually triggered by infection with Epstein-Barr virus (EBV). Once diagnosed, the initial goal of treatment is to suppress inflammation, typically using corticosteroids and biologics. Inflammatory bowel disease can be treated with standard immunosuppressive drugs, although response rates are generally low. EBV infection can be treated with specific antiviral therapies, and in some cases, immunoglobulin therapy. Currently, the only potentially curative treatment for XIAP deficiency is hematopoietic stem cell transplantation (HSCT). However, due to the characteristics of the underlying disease, a low-intensity conditioning regimen is required, and HSCT results are generally worse than normal for patients of this age. Although life expectancy in XIAP patients has increased over the past decade, the condition still presents significant morbidity and mortality (Mudde et al. 2021).

[0431] CDC42 mutation

[0432] Cell division control protein 42 (CDC42) belongs to the Rho family of small monomeric GTPases (Heasman et al. 2008). It influences multiple cellular processes, including cell division and migration, and regulates the nervous, immune, and hematopoietic systems (Melendez et al. 2011). Pathogenic mutations in CDC42 can have a variety of effects on protein function, leading to increasingly prominent and diverse disease phenotypes centered on neurodevelopment, hematopoiesis, and immune responses (Takenouchi et al. 2015, Asiri et al. 2021, Coppola et al. 2022). In particular, aberrant palmitoylation of CDC42R186C has been shown to trap CDC42 within the Golgi apparatus, which may induce excessive activation of the heat protein inflammasome and significant increases in IL-1β and IL-18 (Coppola et al. 2022). In four pediatric patients, a neonatal missense mutation at the C-terminus (p.R186C) has been shown to affect CDC42 localization and cause a specific set of neonatal episodic cytopenia, autoinflammation, and recurrent HLH (Lam et al. 2019). The presence of chronically elevated serum IL-18 (even after a clinical response to anti-IL-1 therapy) suggests that IL-18 is a promising therapeutic target for CDC42 C-terminal disease (Lam et al. 2019).

[0433] Table 4. Clinical phenotypes, cytokine profiles, and responses to investigational treatments for XIAP and CDC42.

[0434]

[0435] summary

[0436] bbmAb is a heterodimeric Fc, monovalent bispecific IgG1 monoclonal antibody (mAb) composed of Novartis' clinical-stage anti-IL-1β and anti-IL-18 mAbs in single-molecule form. By simultaneously targeting and neutralizing both inflammasome effector cytokines IL-1β and IL-18, bbmAb has the potential for excellent clinical efficacy in autoinflammatory diseases in which inflammasome overactivation and both IL-1β and IL-18 directly promote the pathophysiology of the disease, such as: hypogammaglobulinemia, cytopenia, inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, hemophagocytic lymphohistiocytosis, neonatal episodic cytopenia, syndromic thrombocytopenia, megaloblastic thrombocytopenia, hematopoietic insufficiency, autoinflammation, syndromic immunodeficiency, or recurrent hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome, or AIFEC.

[0437] Non-clinical data

[0438] Nonclinical pharmacology

[0439] In most cell assays, bbmAb binds to both IL-1β and IL-18 with single to double-digit pM affinity, resulting in sub-nM inhibition of cytokine signaling. Although bbmAb has monovalent binding to IL-1β and IL-18, its in vitro neutralization potency for human IL-1β and IL-18 is similar to that of bivalent mAb1 and mAb2, and its inhibitory activity against primary marmoset cells is similar.

[0440] Clinical data

[0441] Clinical human pharmacokinetics

[0442] Preliminary pharmacokinetic data from the FIH trial in healthy volunteers (HV) were consistent with predictions based on marmoset data and modeling (i.e., typically belonging to human IgG1 immunoglobulin). Peak serum concentrations of bbmAb were observed shortly after the completion of the intravenous infusion. The median Tmax was approximately 3 hours from the start of the infusion (lasting 120 minutes). Cmax and AUCinf increased with increasing dose in a slightly overproportional manner. The mean terminal elimination half-life (T1 / 2) ranged from approximately 21 days (at 0.1 mg / kg) to approximately 28 days (at 30 mg / kg). The volume of distribution was low, with a mean Vz of approximately 5.2 L to approximately 6.4 L. Additionally, bbmAb was administered subcutaneously at doses of 100 mg, 300 mg, and 600 mg. The mean Cmax of bbmAb ranged from 7.1 to 54.7 μg / mL approximately 7 days after administration. Based on AUCinf, at 10 mg / kg IV, Japanese subjects had approximately 1.16 times higher exposure than non-Japanese subjects, while at 600 mg SC, the exposure was 1.25 times higher compared to non-Japanese subjects. For the Japanese cohort, the bioavailability of the 600 mg SC dose was approximately 65% ​​compared to the 10 mg / kg IV dose. For the non-Japanese cohort, the bioavailability of the 100 mg SC, 300 mg SC, and 600 mg SC doses was approximately 45%, 69%, and 65%, respectively, compared to the 10 mg / kg IV dose.

[0443] 2. Targets and endpoints related to XIAP deficiency or CDC42 mutation

[0444] Table 5 Objectives and Relevant Endpoints

[0445]

[0446] Main estimators

[0447] The main clinical issues of concern regarding autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation are:

[0448] In patients with autoinflammatory or disease-related conditions based on XIAP deficiency or CDC42 mutation, who achieved a full clinical response after approximately 28 weeks of bbmAb treatment despite discontinuing glucocorticoids, what is the effect of continuing bbmAb treatment on disease onset within 24 weeks?

[0449] The main estimators include the following components:

[0450] 1. Population: Patients with autoinflammatory or disease-associated with XIAP deficiency or CDC42 mutation who have achieved a complete response after approximately 28 weeks of bbmAb treatment and have discontinued cyclosporine and glucocorticoids or are receiving a maintenance / replacement dose of glucocorticoids (< 0.2 mg / kg / day).

[0451] 2. Endpoint: The occurrence of disease onset within 24 weeks.

[0452] 3. Purpose treatment: Randomized study treatment (bbmAb or placebo study treatment).

[0453] 4. Management of Concurrent Events: The primary analysis will employ the treatment policy strategy; therefore, treatment discontinuation due to any reason other than disease onset will be ignored. Patients who prematurely discontinued treatment in Phase 2 (not due to disease onset) will be analyzed in the same manner as patients who continued treatment as planned.

[0454] 5. Overview: Differences in the proportion of patients experiencing disease onset between treatment groups.

[0455] 3. Research Design

[0456] like Figure 1 This is a three-phase study, with Phase 1 being an open-label, single-arm aggressive treatment regimen, Phase 2 being a randomized, placebo-controlled, double-blind design, and Phase 3 being an open-label, long-term safety follow-up. Patients who meet the criteria for XIAP deficiency or CDC42 mutation and are currently receiving bbmAb will be eligible to directly enter Phase 3 for open-label long-term safety follow-up.

[0457] Patients will be assigned to relevant groups:

[0458] • Group 1: Phase 1 of the study will include patients diagnosed with XIAP deficiency or CDC42 mutation, so that patients can be randomly assigned to Phase 2 of the study.

[0459] • Group 2: Additional patients diagnosed with XIAP deficiency or CDC42 mutation using bbmAb after completing screening / baseline can be directly enrolled in Phase 3 of the study. The total study duration from screening to study end (EoS) is expected to be between 3 and 4 years.

[0460] Figure 1 The overall research design is outlined.

[0461] This three-phase study includes:

[0462] filter:

[0463] The screening phase lasts approximately 30 days to confirm compliance with study inclusion and exclusion criteria. The screening phase also allows Group 1 patients to safely discontinue or stabilize their permitted medication dosage during Phase 1. For Group 2 patients, enrollment in Phase 3 can proceed directly after screening and baseline completion. If necessary, assessments can be performed within a few days for the patient's best interests or travel arrangements. If results are available, laboratory tests performed several days prior to screening as part of routine patient care can be used to avoid collecting additional blood samples from the patient.

[0464] The screening window may be extended in the following circumstances (if informed consent has been obtained):

[0465] • For group 1, sufficient time is required to demonstrate the presence of active disease after discontinuation of current treatment as outlined in the inclusion and exclusion criteria.

[0466] • For patients with undocumented molecular diagnoses of NLRC4, XIAP deficiency, or CDC42 mutation (Group 1), all other screening assessments (except for informed consent) should be performed after the molecular diagnosis is available and within the screening window, in order to make the molecular diagnosis available.

[0467] Baseline:

[0468] Eligible patients will be admitted (if not already hospitalized) and assessed at the baseline visit; this may be done on day -1 or in combination with day 1 before administration.

[0469] To mitigate the potential for SARS-CoV-2 infection in patients, guidelines and requirements provided by local regulatory agencies or local research centers will be followed (e.g., patients may be screened for SARS-CoV-2 by PCR or similar approved methods before any overnight stay at a research / hospital research center, in accordance with local research center-specific SOPs).

[0470] All baseline safety assessment results must be obtained prior to drug administration. If results are available, laboratory tests performed several days prior to baseline as part of routine patient care can be used to avoid collecting additional blood samples from the patient.

[0471] Phase 1, Open-Label Treatment Phase:

[0472] Only applicable to Group 1.

[0473] Phase 1 is an open-label treatment phase designed to identify responders to bbmAb treatment and allow these patients to gradually reduce their glucocorticoid doses and / or discontinue cyclosporine treatment. Phase 1 is divided into three sub-parts (Phase 1a, 1b, and 1c).

[0474] Eligible patients will proceed to Phase 1 and receive their first dose of bbmAb (10 mg / kg) via intravenous infusion on Day 1 of Phase 1a. Due to the nature of the disease, patients may remain hospitalized for the duration of Phase 1; however, this is not mandatory, and investigators should determine when a patient is ready for discharge based on their condition. During Phase 1, patients will undergo efficacy, pharmacokinetic (PK), and disease progression (PD) assessments as outlined in the assessment schedule (Table 14).

[0475] Phase 1a

[0476] Phase 1a lasts for 4 weeks, with bbmAb administered every 2 weeks.

[0477] For patients currently receiving stable doses of glucocorticoids and / or cyclosporine, these doses will remain stable throughout phase 1a. Gradual reduction of glucocorticoids or cyclosporine is not permitted.

[0478] On day 29 (week 4), response assessment will be conducted using PGA, CRP, and ferritin. Patients who achieve at least a partial response will proceed to phase 1b of the study. Patients who do not achieve a partial response during phase 1a will be withdrawn from the study.

[0479] If a patient discontinues treatment during phase 1a, the patient should return approximately one month after discontinuation to complete the phase 1 end assessment (i.e., week 28 of phase 1c in the assessment schedule) as a treatment end visit.

[0480] Phase 1b (gradual reduction of glucocorticoids and cessation of cyclosporine use)

[0481] Phase 1b lasts a maximum of 20 weeks, with bbmAb administered every 2 weeks. Patients will proceed to Phase 1b after successfully completing Phase 1a.

[0482] At the start of the randomization withdrawal phase (Phase 2) prior to randomization, patients receiving a stable dose of glucocorticoids will be gradually reduced to the lowest possible dose for 4 weeks. Similarly, any patient receiving a stable dose of glucocorticoids will have their dose reduced to achieve cyclosporine discontinuation for 4 weeks prior to randomization at the start of the randomization withdrawal phase (Phase 2). Guidelines for gradual reduction of glucocorticoids, discontinuation of cyclosporine, and patient eligibility for Phase 1c and Phase 2 are given in Section 6.2.1.1.

[0483] For patients who were not hospitalized during Phase 1b, the research center will call the patient / parent / caregiver weekly to monitor their response during the gradual reduction of glucocorticoids.

[0484] Patients who meet the eligibility criteria for entering Phase 1c before week 24 may enter Phase 1c earlier, but must complete the assessments listed for week 24 as detailed in the assessment schedule before entering Phase 1c.

[0485] For patients who are not receiving glucocorticoids and cyclosporine at the start of the study (phase 1a) (i.e., no gradual reduction of glucocorticoids or discontinuation of cyclosporine is required in phase 1b), upon completion of phase 1a, they will proceed to phase 1b and complete only the assessments and treatments listed for weeks 22 and 24 of phase 1b, before proceeding to phase 1c. This will ensure that all patients receive at least 12 weeks of bbmAb treatment during phase 1 of the study.

[0486] Patients who cannot reduce their glucocorticoid dose or discontinue cyclosporine treatment by week 24 may discontinue the study. If a patient discontinues during phase 1b, they should return approximately one month after discontinuation to complete the phase 1 end assessment (i.e., week 28 of phase 1c in the assessment schedule) as part of the treatment end visit. Patients who have achieved a partial response (with or without gradual reduction of glucocorticoids, but with cyclosporine discontinuation) as assessed at the phase 1 end visit may be directly enrolled in phase 3 open-label treatment at the discretion of the investigator and their families.

[0487] Phase 1c

[0488] Phase 1c lasts for 4 weeks, during which bbmAb is administered once every 2 weeks.

[0489] After successfully completing phase 1b, the patient will enter phase 1c at the next scheduled bbmAb dose.

[0490] Any patient continuing glucocorticoid therapy must maintain a stable dose throughout the entire duration of phase 1c. Gradual reduction of glucocorticoids is not permitted in phase 1c, nor is cyclosporine therapy.

[0491] The goal of Phase 1c is to ensure that all patients who have discontinued cyclosporine therapy and / or gradually reduced and maintained low doses of glucocorticoids are clinically stable for at least 4 weeks before entering Phase 2.

[0492] At the end of Phase 1c (day 197, week 28), response assessments will be conducted using PGA, CRP, and ferritin, and patients with a complete response will be randomized to Phase 2 of the study. Patients who do not meet the criteria for a complete response but achieve a partial response (with or without gradual reduction of glucocorticoids, but with discontinuation of cyclosporine) may be directly enrolled in Phase 3 open-label treatment at the discretion of the investigator and their families.

[0493] If a patient discontinues treatment during phase 1c, the patient should return approximately one month after discontinuation to complete the phase 1 end assessment (i.e., week 28 of phase 1c in the assessment schedule) as a treatment end visit.

[0494] Phase 2, Random Exit Phase:

[0495] Applicable to Group 1

[0496] Phase 2 consists of a 24-week placebo-controlled, double-blind, randomized withdrawal phase, primarily evaluating the efficacy of bbmAb compared to placebo. At the start of Phase 2, bbmAb responders (those who achieved a complete response to treatment at the end of open-label Phase 1 treatment) will be randomized in a 1:1 ratio to either bbmAb treatment (i.e., continued use at 10 mg / kg) or placebo.

[0497] The first planned blinded dosing after randomization in Phase 2 will be two weeks after the last dose of Phase 1c, and will continue every two weeks until disease onset or 24 weeks have elapsed in Phase 2. Patients may remain hospitalized for the duration of Phase 2; however, this is not mandatory, and investigators should determine when a patient can be discharged based on their condition.

[0498] If the patient meets the criteria for onset in Phase 2, an unplanned visit should be conducted as detailed in the assessment schedule, blinded treatment should be discontinued, and the patient should be transferred to open-label bbbAb therapy to continue treatment for the remainder of Phase 2.

[0499] If a patient discontinues treatment during Phase 2, the patient should return approximately one month after discontinuation to complete the Phase 2 end assessment (i.e., week 24 of Phase 2 in the assessment schedule) as a treatment end visit.

[0500] Phase 3, long-term safety, open-label treatment:

[0501] Phase 3 consists of a 3-year long-term safety period, with treatment using open-label bbmAb (10 mg / kg).

[0502] The first scheduled dose for Phase 3 will be administered 2 weeks after the last dose of Phase 1 or Phase 2, and will continue at intervals of at least 2 weeks. For Group 2, scheduled dosing will begin after screening and baseline visits have been completed and all group-specific eligibility criteria have been met.

[0503] Phase 3 dosing will continue approximately every 2 weeks, with protocol evaluation featuring reduced visit rates, as outlined in the evaluation schedule. Any Phase 3 visits may be conducted at sponsor-approved alternative research centers (e.g., satellite centers that may be more convenient for the patient) only if the principal investigator, who will maintain investigator responsibilities, agrees and only if permitted by local regulations.

[0504] At the investigator’s discretion (and provided that the patient remains clinically stable without evidence of increased disease activity), dosing in phase 3 may be administered approximately every 14 days, with the window specified in Table 14 to allow for flexibility based on individual patient response.

[0505] For patients receiving maintenance doses of glucocorticoids, further gradual reduction of glucocorticoids is recommended, with complete discontinuation if possible. All patients who have not maintained a minimum partial response may discontinue treatment unless their loss of response is considered a result of the gradual reduction in glucocorticoids. Patients in group 2 who have transitioned to the study will continue in phase 3, provided their treating physician determines there is clinical benefit.

[0506] If a patient discontinues treatment during Phase 3, the patient should return approximately one month after discontinuation to complete the Phase 3 end assessment (i.e., Phase 3, week 152 in the assessment schedule) as a treatment end visit.

[0507] Screening and baseline visits will be used to confirm eligibility for study inclusion and exclusion criteria and to perform baseline clinical observation and biological sampling. Patients eligible for enrollment in this study may have been treated with anakinase, canagencinumab, imarumab, and / or investigational IL-18 / IL-1 / IFN-γ combination or blockade therapy and may be screened. Enrollment will take approximately 30 days after screening, and bbmAb treatment will be administered once criteria for active disease are met. This induction period for previously treated patients shortens the clearance time compared to the classic design, but this approach is considered reasonable because subjects may require urgent enrollment to bbmAb after failure of other treatment options.

[0508] Phase 1 is an open-label, aggressive treatment phase designed to identify patients with monogenic IL-18-driven autoinflammatory diseases (including XIAP deficiency and CDC42 mutations) who respond to bbbab treatment, and then allow patients receiving glucocorticoids and / or cyclosporine to gradually reduce / discontinue these therapies. In Phase 1a, patients are initially treated with two doses of bbbab to ensure a response by day 29, particularly achieving control of MAS and resolution of intestinal clinical manifestations (enterocolitis) simultaneously over several weeks. In Phase 1b, patients with a clear bbbab response to cyclosporine and glucocorticoids will discontinue cyclosporine, and glucocorticoids will be gradually reduced or discontinued to a maintenance / replacement dose over a period of up to 20 weeks to avoid long-term morbidity associated with both treatments in pediatric populations. Phase 1c ensures that patients discontinue glucocorticoids or receive a maintenance / replacement dose and are clinically stable for at least 4 weeks before assessing response to bbbab treatment at the end of Phase 1.

[0509] At the start of the randomization withdrawal phase (Phase 2), patients who fully responded to bbbab and had discontinued glucocorticoids or received maintenance / replacement doses were randomly assigned in a double-blind, 1:1 ratio to receive either bbbab or a matched placebo. This design allows patients who have withdrawn from bbbab treatment (placebo patients) to immediately restart bbbab treatment upon reaching the study endpoint (disease flare-up), thus addressing clinical issues and patient preference for placebo allocation by minimizing the time patients spend receiving potentially ineffective treatment. All patients who experience a flare-up at any time during Phase 2 will have their blinded treatment discontinued and will be transferred to open-label bbbab treatment, allowing the investigator and their families to decide whether to continue treatment for the remainder of Phase 2. A 1:1 randomization ratio was chosen to maximize the statistical power of the primary analysis while minimizing the overall sample size, given the rarity of the condition. Blinding was reasonable to prevent intentional or unintentional bias in the study design and its implementation. The duration of the randomized exit phase of the study was based on clinical trial experience with episodes in similar pediatric populations of autoinflammatory diseases treated with canagencinumab (e.g., CAPS and SJIA) and modeling from free IL-18 (where elevated levels were expected to lead to episodes in patients with monogenic IL-18-driven autoinflammatory diseases, including XIAP deficiency and CDC42 mutations, in the absence of effective therapies).

[0510] Long-term safety of patients who have responded to bbbab, open-label treatment (Phase 3) will allow them to continue bbbab treatment and provide long-term safety data.

[0511] Basic principles of dosage / regimen

[0512] Intravenous administration of bbmAb has been evaluated in healthy volunteers (up to 30 mg / kg IV) and COVID-19 patients without any drug-related SAEs (10 mg / kg IV) in a single-dose escalation FIH study; the pharmacokinetic (PK) of bbmAb in humans was as expected for typical IgG1 antibody targeting soluble ligand cytokines. In a pre-planned PK analysis that enabled subcutaneous administration in the FIH study, bbmAb showed dose-proportional increases in exposure, matching the predicted human PK. Peak serum concentrations of bbmAb were observed shortly after IV infusion. The median Tmax was approximately 0.146 days, or approximately 3.5 hours, from the start of infusion. Cmax and AUC0-inf increased with dose in a dose-proportional manner. bbmAb concentrations decreased exponentially, with a mean terminal elimination half-life (T1 / 2) ranging from 21.1 to 286.3 days. The volume of distribution was low, with a mean Vz ranging from 0.06662 to 0.083 mL / kg. In addition, when bbmAb was administered subcutaneously at a dose of 100 mg, a Cmax of approximately 7 µg / mL was observed approximately 9 days after administration. Bioavailability was estimated at 70% (over the duration) by comparing AUCinf divided by 100 mg sc with AUCinf divided by 1 mg / kg iv. Cmax and AUCinf increased with increasing dose in a slightly overproportional manner. The mean terminal elimination half-life (T1 / 2) ranged from approximately 21 days (at 0.1 mg / kg) to approximately 28 days (at 30 mg / kg). The volume of distribution was low, with a mean Vz of approximately 5.2 L to approximately 6.4 L. Furthermore, when bbmAb was administered subcutaneously at doses of 100 mg, 300 mg, and 600 mg, the mean Cmax of bbmAb ranged from 7.1 to 54.7 μg / mL approximately 7 days after administration. Based on AUCinf, at 10 mg / kg IV, Japanese subjects had approximately 1.16 times higher exposure than non-Japanese subjects, while at 600 mg SC, the exposure was 1.25 times higher compared to non-Japanese subjects. For the Japanese group, the bioavailability of the 600 mg SC dose was approximately 65% ​​compared to the 10 mg / kg IV dose. For the non-Japanese group, the bioavailability of the 100 mg SC, 300 mg SC, and 600 mg SC doses was approximately 45%, 69%, and 65%, respectively, compared to the 10 mg / kg IV dose. (Figure 5)

[0513] All subjects in all groups (including those treated with bbbmAb and those receiving placebo) tested negative (i.e., treated with anti-bbmAb antibodies), except for the following: one subject from group A4 (3 mg / kg IV) on day 29, one subject from group A5 (10 mg / kg IV) on day 29, one subject from group A6 (30 mg / kg IV) on day 197, and one subject from group D1 (10 mg / kg IV, Japanese) on day 197 (these subjects were therefore all considered to have transient ADA responses). On day 15, one subject from group A5 (10 mg / kg IV) tested positive. All subjects from group A6 (both those treated with bbbmAb and those receiving placebo) tested negative on day 253.

[0514] 4. Basic Principles

[0515] Basic principles of dosage / regimen

[0516] Patients with monogenic IL-18-driven autoinflammatory diseases, including XIAP deficiency or CDC42 mutations, exhibit significantly and chronically elevated serum free IL-18. The kinetics of free IL-18 potentially limit the efficacy of bispecific antibodies and thus guide dosing principles in these patients. Under normal physiological conditions, almost all circulating IL-18 is biologically inactive when bound to its binding protein (IL-18BP); however, in severe inflammatory conditions, IL-18 levels exceed available IL-18BP, resulting in a higher proportion of free / biologically active IL-18 that drives pathology. Modeling using measurements of total IL-18, IL-18BP, and free IL-18 from pediatric patients with NLRC4-GOF mutations (Weiss et al. 2018), a dose of 10 mg / kg iv bbmAb was confirmed to achieve rapid and sustained reductions in free IL-18 in both pediatric and adult populations in those with NLRC4-GOF, XIAP deficiency, or CDC42 mutations.

[0517] The model used to predict the kinetics of serum anti-IL-18 / IL-1β bispecific antibodies and their targets consisted of a general competitive binding model describing the kinetics of free and total IL-18 in the IL-18 group (Yan et al. 2012) and a previously published canaginumumab model (Chakraborty et al. 2012) (for the IL-1β group, the parameters were applied to bbmAb). The model was adjusted using baseline values ​​(Weiss et al. 2018) and internal measurements of free IL-18, total IL-18, and IL-18BP in the serum of patients with several autoimmune diseases (including NLRC4-GOF), and the assumption was that neonatal patients weighed 3 kg. Based on simulations of the effect of bbmAb on free IL-18 during intravenous administration, complete control (neutralization) of free IL-18 was expected to last approximately 14 days at a dose of 10 mg / kg, while also neutralizing IL-1β to completely control inflammatory syndromes, allowing for the reversal of clinical features in these patients (such as gastrointestinal pathology and MAS or HLH) during treatment.

[0518] The effects of simulating bbmAb 10 mg / kg on patients with elevated free IL-18 and IL-1β were demonstrated to be both immediate and durable. The therapeutic range of 2 mg / kg q2d dose of rhIL-18BP (tadalafil α) (Tak et al. 2006) is shown in gray shading (Figure 5), which has been reported to be clinically effective in NLRC4-GOF infants (Canna et al. 2017, Moghaddas et al. 2018) and is currently being evaluated in a phase 3 study (NCT03113760). Compared with this, a 10 mg / kg q2W dose of bbmAb is expected to be equivalent to neutralizing free IL-18 at 2 weeks and then completely inhibiting free / bioactive IL-18 to undetectable levels similar to those in healthy individuals over subsequent weeks.

[0519] The rationale for the bbmAb dose of 10 mg / kg IV q2w in this study is further demonstrated by the following:

[0520] • This dose is expected to result in rapid and simultaneous neutralization of free IL-18 and IL-1β, thereby rapidly inducing a clinical response in patients with already measured elevated levels of IL-1β and IL-18 and in pediatric patients enrolled in this study.

[0521] • A single 10 mg / kg IV dose was administered to healthy volunteers in FIH to establish the safety of this dose for treating patients with gain-of-function mutations (e.g., NLRC4 mutations) that lead to overexpression of IL-1β and IL-18 (Romberg et al. 2014). This dose has provided combined IL-18 and IL-1β inhibition for several months in healthy volunteers and is currently being administered to COVID-19 patients without safety issues.

[0522] • IL-18BP treatment modeling in pediatric patients with NLRC4-GOF was used to estimate the clinically effective bbmAb dose, demonstrating that patients with IL-18-driven diseases (including XIAP deficiency and CDC42 mutations) should achieve a sustained clinical response at 10 mg / kg IV q2w, with neutralization of free IL-18 lasting approximately 14 days—lower doses and longer dosing intervals may not achieve a complete response and patients are at risk of relapse due to inadequate treatment. In the open-label Phase 3, disease-stable patients could have dosing intervals up to 28 days, with investigators closely monitoring their clinical status.

[0523] • Based on clinical experience with canageninumab in treating pediatric patients with severe CAPS, these patients (especially those under 2 years of age) may require higher doses (greater than 8 mg / kg of canageninumab) to achieve a complete clinical response and may require more frequent dose titrations compared to adults.

[0524] • Allometric scaling of the bbmAb pop-PK parameters from 70 kg adults to 3 kg newborns predicted higher clearance per body weight in newborns and young infants. Therefore, the expected lower exposure per dose justifies higher doses in the pediatric population to maximize clinical response. No significant accumulation was expected in this population after multiple dosings at the proposed dose of 10 mg / kg IV q2w. Similarly, increased body weight-based clearance (L / day / kg) and decreased canaginumab exposure were observed in younger pediatric patients (Zhung et al. 2019).

[0525] • The total systemic exposure in the expected study was significantly lower than that achieved in the 26-week marmoset study, with the predicted AUC exposure ratio to non-clinical NOEL exposure being 14.4 times for 3kg newborns, and the Cmax being 14.4 times.

[0526] • In the 26-week toxicology study of NHP, no adverse events were observed (no level of action was observed) at 100 mg / kg twice a week.

[0527] In summary, based on available data, a dose of 10 mg / kg q2w to q4w has been defined as potentially effective while having minimal risk in both pediatric and adult populations, depending on the disease state.

[0528] 5. Research Group

[0529] The study population included approximately 18 patients in total:

[0530] • Group 1: Patients with monogenic IL-18-driven autoinflammatory diseases (including XIAP deficiency or CDC42 mutation)

[0531] • Group 2: Patients diagnosed with XIAP deficiency or CDC42 mutation directly enter Phase 3.

[0532] Inclusion criteria

[0533] For all patients: Patients eligible for inclusion in this study must meet all of the following criteria:

[0534] 1. Male and female patients weighing at least 3 kg at the time of screening.

[0535] 2. Written informed consent from the child patient's parents / legal guardians and the child patient's consent (subject to local requirements) must be obtained before conducting any study-specific evaluations. For adult patients, written informed consent must be signed by the patient who is able to give consent, or, if the patient is unable to give consent, by their legal / authorized representative (if permitted by local requirements).

[0536] Group 1 specific inclusion criteria:

[0537] 3. Patients with a single-gene IL-18-driven autoinflammatory disease who have a genetic diagnosis of XIAP deficiency or CDC42 mutation (this analysis may be performed as part of a screening procedure if not already available).

[0538] •3a. Patients with XIAP deficiency or CDC42 mutation who have previously undergone bone marrow transplantation may be permitted if they show evidence of primary or secondary graft failure or failure to achieve phenotypic correction, and have evidence of XIAP / CDC42-related disease relapse or clinically significant mixed chimerism.

[0539] 4. Clinical history and studies consistent with autoinflammation and infantile enterocolitis (XIAP or CDC42), including elevated IL-18 levels (this analysis may be performed as part of the screening procedure if not already available).

[0540] •4a. XIAP patients only: Patients must have a persistent disease or be resistant to escalating therapy.

[0541] 5. At the time of initial treatment (day 1 of phase 1), there is evidence of active disease as assessed by the following criteria:

[0542] a. PGA of disease activity > very mild

[0543] and

[0544] b. Ferritin > 600 ng / ml

[0545] or

[0546] c. CRP level > 20 mg / L.

[0547] Group 2 specific inclusion criteria:

[0548] Patients with a genetic diagnosis of XIAP deficiency or CDC42 mutation, elevated IL-18 and / or IL-1β, who are being treated with bbmAb in Novartis’s Managed Access Program (MAP), have demonstrated treatment failure while receiving other immunomodulatory therapies, and, according to their treating physician, have no available alternative treatment options.

[0549] Exclusion criteria

[0550] Patients meeting any of the following criteria are not eligible for inclusion in this study:

[0551] 1. History of hypersensitivity to any investigational drug, or to drugs of a similar chemical classification, or to any excipient.

[0552] 2. Researchers determine that the signs and symptoms indicate a clinically significant active bacterial, fungal, or viral infection. If appropriate treatment has been initiated and no signs of infection progression are present at screening, the infection is considered under control. Infection progression is defined as hemodynamic instability attributable to sepsis, new symptoms, worsening physical signs, or radiological findings attributable to infection. Persistent fever without other signs or symptoms is not considered a progressive infection.

[0553] 3. COVID-19 Specificity: If in accordance with health and governmental guidelines, it is strongly recommended to complete a COVID-19 PCR test or similar approved method within one week prior to the first dose. If testing is performed, a negative result is required before enrollment in the study. Additional testing may be determined at the discretion of the investigation physician. For pediatric patients, COVID-19 testing should be performed via nasal or throat swabs or other approved methods. If testing is not performed, the investigator must document in the source documentation their discussions with the patient / parent / caregiver regarding testing and the rationale for not performing testing. This requirement may be disregarded if the country where the research center is located declares the pandemic over, and will be reinstated if the pandemic resurges.

[0554] 4. Any condition or major medical problem that researchers believe would put patients at an unacceptable risk (in cases of uncertainty, this can be discussed on a case-by-case basis with Novartis).

[0555] 5. Prior to bbmAb treatment, the patient has received prior treatment with anti-rejection and / or immunomodulatory drugs within the past 28 days or within 5 half-lives of an immunomodulatory therapeutic antibody (or as listed in the prohibited treatment section of 6.2.2, whichever is longer).

[0556] The exception is:

[0557] •6.2.2 For drugs listed in the Prohibited Drugs section, the washout period outlined therein should be followed.

[0558] • Prior to bbmAb treatment, a stable dose of glucocorticoids ≤ 2.0 mg / kg / day (maximum 60 mg / day for patients over 30 kg) or a stable dose of oral prednisone (or equivalent, regardless of route of administration or dosing schedule) should have been maintained for at least 24 hours.

[0559] • Prior to bbmAb treatment, a stable cyclosporine dose of < 5 mg / kg / day should be maintained for at least 3 days.

[0560] • Prior to treatment with bbmAb, anakinase, canagencinumab, imarumumab, and / or investigational IL-18 / IL-1 / IFN-γ binding or blocking therapy must be discontinued (see section 6.2.2). Once the evidence criteria for active disease are met, patients may receive bbmAb treatment (according to inclusion 5).

[0561] 5. Participation in any other investigational trial within 4 weeks or longer prior to administration (if required by local regulations), except for treatment with anakinase, canagencinumab, imarumumab, and / or investigational IL-18 / IL-18BP / IL-1 / IFN-□ binding or blocking therapy. Participation in Novartis' MAP using bbmAb is not excluded.

[0562] 6. A positive HIV test result at screening (ELISA and Western blot). Sufficient prior testing evidence within 3 months.

[0563] 7. A positive hepatitis B surface antigen (HBsAg) or hepatitis C test result at the time of screening. Sufficient prior testing evidence within 3 months is required.

[0564] 8. Tuberculosis infection as defined by a positive TB test is present at the time of screening. Sufficient prior testing evidence within 3 months is required.

[0565] 9. Administer the live vaccine within one month before treatment with bbmAb, during the trial, and for up to three months after the last dose.

[0566] 10. History of any malignant tumor of an organ system (including post-transplant lymphoproliferative disorders) that has been treated or not treated within the past 5 years (excluding adequately treated local basal cell carcinoma of the skin or cervical cancer in situ).

[0567] 11. Pregnant or breastfeeding women, where pregnancy is defined as the state from the time a woman conceives until the termination of pregnancy, as confirmed by a positive hCG laboratory test.

[0568] 12. Female patients who are of reproductive potential (or Tanner stage 2 or above) and who are sexually active or likely to be pregnant must be informed of the potential teratogenic risks of bbbab and agree to use highly effective contraception to prevent pregnancy while receiving bbbab therapy.

[0569] • Highly effective contraception (abstinence, oral, injectable, or implantable hormonal contraception, or intrauterine device (IUD) or intrauterine system (IUS) or other forms of hormonal contraception with equivalent efficacy (failure rate < 1%), such as hormonal vaginal rings or transdermal hormonal contraception) must be used during the study period and for 5 months after discontinuation of bbmAb treatment, during which time IL-18 and IL-1β are not expected to be neutralized by bbmAb. The decision on contraceptive method should be reviewed at least every 3 months to assess individual needs and compatibility with the chosen method.

[0570] Please note that only highly effective contraceptive methods approved in accordance with local regulations should be used.

[0571] • Women are considered postmenopausal if they have experienced 12 months of spontaneous amenorrhea and have appropriate clinical features (e.g., a history of age-appropriate vasomotor symptoms). Women who are postmenopausal or who have undergone surgical bilateral oophorectomy (with or without hysterectomy), total hysterectomy, or bilateral tubal ligation at least six weeks prior are considered to have no reproductive potential. In cases of oophorectomy alone, a woman is considered to have no reproductive potential only if her reproductive status has been confirmed by subsequent hormone level assessments.

[0572] 13. Patients weighing > 160 kg at the time of screening.

[0573] 14. For patients with CDC42 mutations: Takenouchi-Kosaki syndrome—a CDC42 mutation associated with a variety of syndromes characterized by variable developmental delay, cardiac, cerebral, and hematological abnormalities.

[0574] 6. Treatment

[0575] 6.1. Research on treatment

[0576] Details regarding the storage and administration requirements of investigational therapeutics, as well as patient numbering, prescription / dispensing, and the instructions for use of investigational therapeutics, are outlined in the pharmacy manual.

[0577] 6.1.1. Investigational drugs and control drugs

[0578] bbmAb is still under development, and the investigational drug and control drug will be administered in the same manner as in the Phase 1 and Phase 2 studies. The investigational drug bbmAb and the matched placebo will be manufactured by Novartis and provided as an open-label, bulk drug to pharmacists at unblinded research centers. Unblinded pharmacists or authorized designated personnel are required to dispense the investigational drug. The drug will be administered by researchers at clinical centers via intravenous infusion over approximately 120 minutes, following the prescribed study procedures.

[0579] During an epidemic or pandemic (such as the COVID-19 pandemic) that restricts or prevents research visits at research centers, research center staff may be arranged to visit patients at their homes so that research treatment can continue in accordance with the protocol, as permitted by local regulations.

[0580] Table 13 Investigational Drugs

[0581]

[0582] 6.1.2. Other research treatments

[0583] This trial does not include treatments other than the investigational drug. The research center will provide supportive care in addition to the investigational treatment.

[0584] 6.1.3. Treatment group (arm / group)

[0585] Phase 1 – Open-label treatment phase (Group 1)

[0586] The patient will be administered bbmAb 10 mg / kg q2w iv on day 1.

[0587] Phase 2 – Random Exit Phase (Group 1)

[0588] Responseing patients will be randomly assigned to one of the following treatment groups at a 1:1 ratio at the end of Phase 1:

[0589] •bbmAb 10 mg / kg IV q2w

[0590] • Matched placebo IV q2w

[0591] Phase 3 – Long-term security of open-label systems (all groups)

[0592] Patients will begin receiving bbmAb 10 mg / kg q2w IV infusion on day 1 of phase 3 (week 0). The frequency of administration can be adjusted within the allowable window listed in Table 14 and at the investigator's discretion.

[0593] 6.1.4. Post-test visit

[0594] Provided there is evidence of clinical benefit to patients, Novartis will provide bbmAb to patients who complete the study, or provide bbmAb up to: [details to be provided].

[0595] • Researchers stopped treatment.

[0596] • Products or alternative treatments are available commercially.

[0597] 6.2. One or more other treatments

[0598] 6.2.1. Companion Therapy

[0599] All key medications, procedures, and significant nonpharmacological therapies (including physical therapy and blood transfusions) administered to the patient within six months prior to enrollment in the study for the treatment of the disease under investigation (if available) should be documented in the appropriate case report form.

[0600] All medications, procedures, and significant nonpharmacological therapies (including physical therapy and blood transfusions) administered after patient enrollment in the study must be documented in the appropriate case report form.

[0601] Each concomitant medication must be evaluated individually for all exclusion criteria / contraindicated drugs. If in doubt, the investigator should contact Novartis's medical monitor before enrolling a patient or allowing the initiation of new drug treatment. If a patient is already enrolled, contact Novartis to determine whether the patient should continue participating in the study.

[0602] During the study and prior to screening, patients may receive gastric protectants, folic acid, acetaminophen, NSAIDs, analgesics, antibiotics, vasopressors, and nutritional supplements (such as vitamins, fluid supplements, enteral nutrition, total parenteral nutrition) and other medications / treatments, which form part of the supportive care for the disease under investigation at the center where the patient is participating in the study (as determined by medical judgment).

[0603] 6.2.1.1. Permitted concomitant therapies requiring caution and / or action

[0604] Phase 1 – Gradual reduction of glucocorticoids

[0605] This only applies to group 1.

[0606] During the 4-week treatment period of Phase 1a, to stabilize patients receiving bbmAb, patients receiving a stable dose of glucocorticoids were permitted to enter the study and receive an initial dose of bbmAb (10 mg / kg) intravenously. The glucocorticoid dose should be maintained throughout the entire duration of Phase 1a, up to at least day 29.

[0607] After day 29, at the start of the randomization exit phase (Phase 2), researchers should gradually reduce the glucocorticoid dose to progressively decrease the dosage, with the aim of stopping glucocorticoids (based on medical judgment) or reaching ≤ [missing value] before randomization. A stable maintenance dose of 0.2 mg / kg / day of glucocorticoids (prednisone or equivalent) was maintained for 4 weeks.

[0608] • If the patient achieves at least a partial response, steroids can be gradually reduced starting after day 29 (section 8.3.5).

[0609] Table 16 Guidelines for Gradual Reduction of Glucocorticoids

[0610]

[0611] • If the following situations occur, the reduction can be further gradually reduced:

[0612] 1. Between research visits during the weekly research center calls, based on responses to the telephone script questionnaire, patients / parents did not report any loss of response.

[0613] 2. During the visit to the research center, the patient maintained at least a partial response to bbmAb.

[0614] • The gradual decrease in glucocorticoids will continue until one of the following occurs first:

[0615] 1. The patient has achieved the goal of discontinuing glucocorticoids and is no longer using steroids.

[0616] 2. The longest duration of gradual reduction in glucocorticoid levels in patients has reached 20 weeks.

[0617] 3. The patient's three attempts to gradually reduce glucocorticoid levels all failed.

[0618] • All patients who fail to maintain at least a partial response to bbmAb may discontinue the study unless the loss of response is considered to be a result of the gradual reduction of glucocorticoids.

[0619] 1. If a patient loses response to bbbmAb while gradually reducing glucocorticoids, the glucocorticoid dose should be increased to the previous level and the patient should remain in phase 1b; the increased steroid dose should be maintained for at least 2 weeks. If a patient remains unresponsive for more than 2 weeks after the initial event after increasing to the previous steroid dose, the study may be discontinued.

[0620] 2. For patients who lose response during the gradual reduction period, gradual reduction of steroids should only be attempted if the patient has received a stable dose of steroids for at least 2 weeks and has at least a partial response.

[0621] A patient is eligible to proceed directly to Phase 2 of the study if they meet one of the following criteria:

[0622] 1. Any patient who has reached the point of discontinuing glucocorticoids and has not used steroids for 4 weeks in phase 1c.

[0623] 2. Patients who, during phase 1c, consistently maintain a stable maintenance dose of glucocorticoids ≤ 0.2 mg / kg / day (for patients weighing over 30 kg) or glucocorticoids (prednisone or equivalent) ≤ 0.4 mg / kg / day (for pediatric patients who may require a higher replacement dose) for 4 weeks.

[0624] If a patient discontinues treatment during Phase 1, the Phase 1 end assessment (i.e., week 28 of Phase 1c in the assessment schedule) should be completed as the end-of-treatment visit. Patients who have achieved a partial response (with or without gradual reduction of glucocorticoids, but with discontinuation of cyclosporine) as assessed at the Phase 1 end-of-treatment visit may be directly enrolled in Phase 3 open-label treatment at the discretion of the investigator and their family.

[0625] Phase 2 – Glucocorticoids (random exit phase)

[0626] During Phase 2, patients who achieved a stable maintenance dose of glucocorticoids (prednisone or equivalent) < 0.2 mg / kg / day (for patients weighing over 30 kg) or < 0.4 mg / kg / day (for pediatric patients) in Phase 1c should maintain that dose, and gradual reduction of glucocorticoids is not permitted in Phase 2. Rescue medication - glucocorticoids

[0627] Based on medical judgment and local guidelines, patients experiencing an attack (see Section 8.3.5) are permitted to receive an increased maintenance dose of glucocorticoids or intermittent glucocorticoid therapy as a rescue medication for a limited period of time.

[0628] Phase 1 – Cyclosporine Cessation

[0629] During the 4-week treatment period of Phase 1a, patients receiving a stable dose of cyclosporine may receive bbmAb treatment to stabilize their condition. Investigators should maintain a stable cyclosporine dose throughout the entire duration of Phase 1a, up to at least day 29.

[0630] After day 29, researchers should gradually reduce the dose of cyclosporine (as determined by medical judgment) with the aim of stopping cyclosporine use for 4 weeks before randomization at the start of the randomization exit phase (Phase 2).

[0631] • If the patient achieves at least a partial response, cyclosporine reduction can begin (section 8.3.5).

[0632] • Cyclosporine dosage reduction will continue until one of the following occurs first:

[0633] • The patient reached the point where cyclosporine treatment could be discontinued.

[0634] • The longest period of time a patient has been able to stop taking cyclosporine has reached 20 weeks.

[0635] • The patient's three attempts to stop cyclosporine treatment all failed.

[0636] If patients have stopped taking cyclosporine for 4 weeks, they are eligible to proceed directly to Phase 2 of the study.

[0637] All patients who fail to maintain at least a partial response to bbmAb and discontinue cyclosporine may be discontinued from the study. Patients who have achieved a partial response (with or without gradual reduction of glucocorticoids, but with discontinuation of cyclosporine) as assessed at the end of the Phase 1 visit may be directly enrolled in Phase 3 open-label treatment at the discretion of the investigator and their families.

[0638] If a patient discontinues treatment during Phase 1, the Phase 1 end assessment (i.e., week 28 of Phase 1c in the assessment schedule) should be completed as the end-of-treatment visit. Patients who have achieved a partial response (with or without gradual reduction of glucocorticoids, but with discontinuation of cyclosporine) as assessed at the Phase 1 end-of-treatment visit may be directly enrolled in Phase 3 open-label treatment at the discretion of the investigator and their family.

[0639] contraception

[0640] Oral, injectable, or implantable hormonal contraceptive methods are permitted when receiving bbmAb.

[0641] 6.2.2. Prohibited Drugs

[0642] The following treatments are not permitted before Day 1 of Phase 1 (see below for the time interval before Day 1) and throughout the study period:

[0643] Enalapril within one week prior to day 1

[0644] Adalimumab within 4 weeks prior to Day 1

[0645] Infliximab within 2 weeks prior to Day 1

[0646] Tocilizumab within 3 weeks prior to day 1

[0647] Vido-group monoclonal antibodies within 4 weeks prior to Day 1

[0648] IV immunoglobulin (IV Ig) was administered for 4 weeks prior to Day 1, except in cases of replacement therapy for patients diagnosed with hypogammaglobulinemia. For guidelines on discontinuing immunoglobulin therapy, please refer to section 6.2.1.1.

[0649] Any other investigational or non-investigational immunomodulatory antibody that was present within the past 30 days or 5 half-lives (whichever is longer) prior to day 1.

[0650] Leflunomide within 4 weeks prior to Day 1

[0651] Thalidomide within 4 weeks prior to Day 1

[0652] 6-Mercaptopurine, azathioprine, cyclophosphamide, or chlorambucil within 12 weeks prior to day 1

[0653] Tacrolimus within 4 weeks prior to day 1

[0654] Colchicine, dapsone, and mycophenolate mofetil within 4 weeks prior to day 1

[0655] Ruxolitinib and other JAK inhibitors within 4 weeks prior to day 1

[0656] Any other investigational or non-investigational anti-rejection and immunomodulatory drugs administered within the past 28 days prior to day 1.

[0657] Anti-thymocyte globulin and alenzumab (Campath) within 12 weeks prior to Day 1

[0658] Depending on the patient's clinical condition, patients receiving glucocorticoid therapy may continue as needed. Before bbmAb treatment, the glucocorticoid dose should be stable for at least 24 hours (see section 6.2.1.1 for the gradual reduction of glucocorticoids during Phase 1).

[0659] Patients receiving cyclosporine treatment may continue as needed, depending on their clinical condition. The cyclosporine dose should be stable for at least 3 days prior to bbmAb treatment (see section 6.2.1.1 for discontinuation of cyclosporine during Phase 1).

[0660] Patients receiving anakinase, canagencinumab, imarumumab, and / or investigational IL-18 / IL-1 / IFN-γ combination or blockade therapy need to discontinue this treatment. Patients may receive bbmAb treatment once the evidence criteria for active disease are met (section 5.1, inclusion criteria). This introductory period shortens the classic clearance period to a medically meaningful time and avoids unnecessary suffering for patients in cases where the protocol-predefined clearance period is too long for the individual patient.

[0661] No live vaccine was administered within 4 months prior to Day 1 of Phase 1, during the trial, and within 3 months after the last dose. At the investigator's discretion and in accordance with local guidelines, approved (including conditional marketing authorization from the HA) killer vaccines, inactivated vaccines, peptide vaccines, DNA vaccines, and RNA vaccines may be permitted.

[0662] Patients entering the study from MAP using bbbmAb will be required to maintain MAP's requirements regarding permitted contraindications and medications.

[0663] Novartis' qualified medical personnel are available to advise investigators on trial-related medical issues regarding concomitant therapies and contraindicated medications.

[0664] 6.2.3. Rescue Drugs

[0665] Increased maintenance doses of glucocorticoids or intermittent steroid therapy may be used as rescue therapy. Information regarding the administration of glucocorticoids to study patients is provided in Section 6.2.1.1, which describes the use and gradual reduction of glucocorticoids during this study.

[0666] The use of rescue medications must be documented on the accompanying medications page of the CRF.

[0667] Patients who do not improve after treatment, do not meet the partial response criteria on day 29 of phase 1, or whose seizures are not due to the gradual reduction of glucocorticoids during phase 1c may discontinue the study and be treated according to medical judgment and local practice. Patients who have achieved at least a partial response and discontinued cyclosporine may be eligible to enter phase 3, where they will receive bbmAb.

[0668] Prescription and instructions for receiving research treatment

[0669] Table 6 Dosage and Treatment Schedule

[0670] (a) Table 7 Dosage and Treatment Schedule

[0671]

[0672] Note: The total infusion time using an IV bag or pump syringe is approximately 60 to 120 minutes.

[0673] 8. Visit schedule and assessment

[0674] The assessment schedule (Table 14) lists all assessments conducted during the assessment process. All data obtained from these assessments must be supported by patient source documentation.

[0675] All patient visits / assessments should be conducted in accordance with the assessment schedule outlined in Table 14 or as close as possible to the designated date / time.

[0676] Missed or rescheduled visits should not result in automatic termination. Patients who prematurely discontinue the study for any reason should be scheduled for a visit as soon as possible, at which time all assessments listed for the final visit should be performed. At the final visit, all assigned study products should be discontinued, and adverse events and concomitant medications should be documented on the CRF.

[0677] In Phase 3, to maintain q2w dosing between planned study visits as outlined in Table 14, home dosing visits may be conducted by a mobile nurse, depending on local regulations and capabilities. At each dosing visit, weight will be measured and any adverse events (AEs) evaluated (by the investigator or an appropriately designated member of the research team based on information obtained from the mobile nurse). Weight from previous dosing days can be used for dose calculations at home dosing visits.

[0678] If an epidemic or pandemic (e.g., the COVID-19 pandemic) restricts or prevents in-person research visits at research centers, alternative methods for providing ongoing care may be implemented. During a pandemic, depending on local regulations and capabilities, phone calls, virtual contacts with patients (e.g., telemedicine), or visits by research center staff to patients' homes may be alternatives to in-person research visits at research centers until it is safe for patients to visit the research center again.

[0679]

[0680] 8.1. Filtering

[0681] filter

[0682] If a patient fails the initial screening, rescreening is permitted; however, each case must be discussed and agreed upon on a sponsor-led case-by-case basis. Patients undergoing rescreening must re-sign an informed consent form and complete a rescreening CRF.

[0683] If the safety laboratory assessment at screening exceeds the range specified in the exclusion criteria, it may be repeated once before treatment. If the repeated value remains outside the specified range, the patient must be excluded from the study.

[0684] 8.1.1. Qualification Screening

[0685] 8.1.1.1. Hepatitis screening, HIV screening

[0686] When appropriate, patients may be screened for hepatitis B surface antigen (HBsAg) and, following standard local practice, for hepatitis B core antigen (HBcAg). Hepatitis C screening will be based on HCV antibodies, and if positive, the level of HCV RNA should be determined. If available, negative test results from the previous 3 months may be used.

[0687] HIV serological positivity will be evaluated, and if positive, confirmed using a second technique available at the laboratory center, such as Western blotting. In the event of a positive confirmatory test, the investigator will provide appropriate counseling as available. The investigator is responsible for notifying state and federal agencies as required by law. If available, negative test results from the previous three months may be used.

[0688] 8.1.2. Tuberculosis (TB) test

[0689] To assess a patient's TB status, TB testing can be performed at screening using one of the following methods, in accordance with local regulations / guidelines:

[0690] •QuantiFERON ® -TB Measurement

[0691] • Chest X-ray

[0692] If available, negative test results from the previous 3 months can be used.

[0693] If necessary, any significant findings will be documented in the relevant medical history / current medical condition section of the eCRF.

[0694] 8.1.3. Information collected when filtering fails

[0695] Patients who have signed informed consent / agreement forms (or whose parents / legal guardians) are subsequently found to be ineligible will be considered screening failures. The reason for screening failure should be entered in the applicable case report form (treatment form). Patients who fail screening must also complete the visit information, demographic information, informed consent form, inclusion / exclusion, disease-specific history (e.g., XIAP deficiency or CDC42 mutation), and treatment pages. No additional data will be entered into the clinical database for patients who fail screening unless the patient experiences a serious adverse event during the screening period. Investigators will follow up on adverse events that are not classified as SAEs and will only record them in the patient's source data.

[0696] Patients (or their parents / legal guardians) who have signed informed consent / agreement forms will be considered early terminaters if the patient is deemed eligible but fails to begin treatment for any reason. The reason for early termination should be documented in the appropriate treatment case report form (treatment page). If a patient (or their parent / legal guardian) voluntarily withdraws from the study during the screening period, visit information, demographic information, informed consent form, NLRC4-GOF medical history, inclusion / exclusion page, withdrawal of informed consent form, and treatment must be completed.

[0697] 8.2. Patient Demographics / Other Baseline Characteristics

[0698] Demographic information

[0699] For the collection of demographic and baseline characteristics, nationally specific regulations consistent with the CRF should be considered. Patient race and ethnicity should be collected and analyzed to identify variations in safety or efficacy due to these factors, and to assess the diversity of the study population as required by health authorities.

[0700] Patient Demographics: Year of birth (age), sex, race, primary ethnicity (if permitted), and relevant medical history / current medical condition (up to the date of informed consent) will be recorded in the eCRF. Where possible, diagnoses should be recorded instead of symptoms. Vaccination status should be recorded as part of the medical history / current medical condition collection.

[0701] Disease-specific history and diagnosis (e.g., NLRC4-GOF, XIAP deficiency, or CDC42 mutation)

[0702] A detailed medical history of the patient's condition should be documented in the eCRF to show how the condition was managed and diagnosed. Details (including the assessment date) should include:

[0703] Symptoms

[0704] Molecular diagnostics for XIAP deficiency or CDC42 mutation (if not yet available, can be performed by the research center at screening time according to locally approved diagnostic procedures).

[0705] IL-18 assay levels (if not yet available, can be performed by the research center at screening according to locally approved diagnostic procedures).

[0706] Treatment intervention and outcome / response

[0707] Time and frequency of attacks

[0708] Clinically significant laboratory values, such as cytokines, CRP, and ferritin.

[0709] Nutritional support

[0710] Hospitalization

[0711] Family history

[0712] Any other research

[0713] Any other clinically relevant information deemed clinically relevant to support a broader understanding of the disease or diagnosis.

[0714] 8.3. Efficacy

[0715] The efficacy assessment will be conducted at the time points defined in the assessment schedule (Table 14).

[0716] If an epidemic or pandemic (e.g., the COVID-19 pandemic) restricts or prevents research visits at research centers, alternative approaches to providing ongoing care and collecting efficacy assessments can be implemented.

[0717] 8.3.1. Physician Overall Assessment (PGA) of Disease Activity

[0718] The time points for assessing physician overall assessment (PGA) of the disease will be outlined in the assessment schedule (Table 14) (Appendix 4).

[0719] PGA will be performed before CRP results are available from the local laboratory to prevent evaluation bias. An investigator is encouraged to evaluate the same patient throughout the study to ensure consistency between assessments.

[0720] The physician's overall assessment will be based on a 5-point scale:

[0721] • 0 = Lack of (no) disease-related clinical signs and symptoms

[0722] • 1 = Very mild disease-related signs and symptoms

[0723] • 2 = Mild disease-related signs and symptoms

[0724] • 3 = Moderate disease-related signs and symptoms

[0725] • 4 = Signs and symptoms related to severe illness

[0726] 8.3.2. Physician's assessment of the severity of disease signs and symptoms

[0727] The physician severity assessment of key disease-specific signs and symptoms will be conducted at the time points outlined in the assessment schedule (Table 14) (Appendix 5). A different assessment will be used for each disease diagnosis, and the appropriate questionnaire should be completed based on the diagnosis.

[0728] Researchers are encouraged to assess the same patient throughout the study to ensure consistency of assessments. The following signs and symptoms will be assessed:

[0729] • The following signs and symptoms will be assessed in patients with XIAP deficiency:

[0730] • Gastrointestinal diseases

[0731] •fever

[0732] • Decreased blood cell count

[0733] • Skin diseases

[0734] •Infect

[0735] •splenomegaly

[0736] •CNS affected

[0737] • The following signs and symptoms will be assessed in patients with CDC42 mutations:

[0738] • Gastrointestinal diseases

[0739] •fever

[0740] • Skin diseases

[0741] •splenomegaly

[0742] •CNS affected

[0743] • The physician severity assessment of key disease-specific signs and symptoms will be based on a 5-point scale:

[0744] •0 = None

[0745] •1 = extremely slight

[0746] •2 = Mild

[0747] •3 = Moderate

[0748] •4 = Severe

[0749] 8.3.3. Inflammatory markers

[0750] CRP and ferritin will be measured by a local laboratory as shown in Table 14. Where possible, these analyses should be included as part of routine laboratory safety monitoring to avoid the need for additional sample collection.

[0751] 8.3.4. Patient / Parent Overall Assessment of Disease Activity (PPGA)

[0752] Patient assessments of disease activity (PPGA) will be collected on the paper CRF for transcription into the electronic CRF (Appendix 6).

[0753] The PPGA should be completed before any clinical assessment at any given visit. Depending on the patient's age, according to local guidelines, this document needs to be completed by the patient or parent / caregiver at the time points outlined in Table 14. Parent / caregiver assistance is possible where possible. However, for consistency, the assessment should be performed by the same assessor (the same patient or parent / caregiver) throughout the study period.

[0754] This will guide patients or parents / caregivers to complete the PPGA.

[0755] Researchers or research center staff should not give verbal or nonverbal prompts that could affect PPGA responses. Researchers or research center staff are only permitted to check the completeness of the documents.

[0756] PPGA is based on a 5-component table:

[0757] • 0 = Lack of (no) disease-related clinical signs and symptoms

[0758] •1 = Very mild disease-related signs and symptoms

[0759] •2 = Mild disease-related signs and symptoms

[0760] •3 = Moderate disease-related signs and symptoms

[0761] •4 = Signs and symptoms related to severe illness

[0762] 8.3.5. Criteria for Response to Treatment

[0763] The response to treatment will be collected via PGA (part 8.3.1) and inflammatory markers (part 8.3.3).

[0764] Phase 1

[0765] Complete response: A patient is considered to have a complete response if the following conditions are met (to be assessed on the same day):

[0766] • The physician's overall assessment of disease activity is very mild or better.

[0767] • Ferritin and / or CRP decreased by 60% or more from baseline or ferritin (< 400 ng / mL) and / or CRP (< 10 mg / L) normalized.

[0768] Partial Response Criteria: A patient is considered to have an incomplete (partial) response if the following conditions are met (to be assessed on the same day):

[0769] • The physician's overall assessment of disease activity showed a step improvement from baseline and

[0770] • Ferritin or CRP decreased by 30% or more from baseline.

[0771] Phase 2

[0772] Seizure criteria: A patient is considered to have a seizure if the following conditions are met (to be assessed on the same day):

[0773] • Overall physician assessment of disease activity > Very mild and

[0774] • Patients whose ferritin and / or CRP levels have increased by 60% or more from the level at the start of stage 2 or whose levels have normalized have ferritin elevation > 2500 ng / mL and / or CRP elevation > 20 mg / L.

[0775] When obtaining standard inflammatory markers for treatment response, researchers should, based on medical judgment, rule out alternative common causes of changes in CRP or ferritin in the pediatric study population (e.g., childhood infection, iron supplementation, blood transfusion).

[0776] Table 8 Clinical Laboratory Safety Assessment (Local)

[0777]

[0778] 9. Efficacy and / or one or more pharmacodynamic endpoints

[0779] This section will use FAS from each research phase for analysis.

[0780] The following secondary efficacy endpoints will be analyzed:

[0781] Responses at the end of day 29, phase 1, and phase 2 (for group 1)

[0782] Serological remission at the end of day 29, phase 1, and phase 2 (for group 1)

[0783] Phase 1 glucocorticoid therapy (for group 1)

[0784] The time of first onset in phase 2 (for group 1)

[0785] Physician severity assessment of disease signs and symptoms (all groups)

[0786] Overall assessment of disease activity in patients / parents (all groups)

[0787] Overall physician assessment of disease activity (all groups)

[0788] The proportion of patients responding to bbmAb treatment on day 29 and thereafter until the end of Phase 1 will be calculated. The definition of a complete responder to bbmAb treatment is given in section 8.3.6 of the protocol. The proportion of patients responding to both bbmAb and placebo will also be evaluated at the end of Phase 2.

[0789] Inflammatory markers (CRP and ferritin) will be summarized through treatment and visits.

[0790] The proportion of patients achieving serological remission will be evaluated at day 29, at the end of phase 1, and at the end of phase 2.

[0791] The proportion of patients who achieved gradual reduction in glucocorticoids and cyclosporine by day 29 until the end of Phase 1 will be calculated. Definitions of gradual reduction in glucocorticoids and cyclosporine are given in section 6.2.1.1 of the protocol.

[0792] attack

[0793] As given in section 8.3.6 of the protocol, the onset criteria are used to assess the onset of disease activity through a physician’s overall assessment, ferritin and / or CRP.

[0794] The time of first onset will be summarized according to the treatment phase 2. Kaplan-Meier diagrams with separate lines for each treatment group will be provided for graphical representation.

[0795] Other efficacy endpoints

[0796] The report will provide summaries of absolute values ​​and changes from baseline for physician severity assessments of disease signs and symptoms, patient / parent overall assessments of disease activity, and physician overall assessments of disease activity, based on treatment and visits. A frequency table of each symptom assessed by physicians and patients / parents will be presented based on visits. The frequency distribution of severity scores (none, very mild, mild, moderate, severe) will be calculated based on treatment and visits.

[0797] 10. References

[0798] 1.Anon (2004) The Fourth Report on the Diagnosis, Evalutation, andTreatment of High Blood Pressure in Children and Adolescents. Pediatrics; 114Suppl 2:555-576.

[0799] 2.Anon (2017) The Hospital for Sick Children Research Ethics BoardBlood Sampling Guidelines (internet) Available at: http: / / www.sickkids.ca / Research / Research-Ethics / forms-guidelines-templates / index.html (Accessed on06-Oct-2020).

[0800] 3.Asiri A, Alwadaani D, et al (2021) Pancytopenia, RecurrentInfection, Poor Wound Healing, Heterotopia of the Brain Probably Associatedwith A Candidate Novel de Novo CDC42 Gene Defect: Expanding the Molecular andPhenotypic Spectrum. Genes. 2021; 12(2):294.

[0801] 4.Baracaglia C, Gatto A, Pardeo M, et al (2015) Anti interferon-gamma(IFNγ) monoclonal antibody treatment in a patient carrying an NLRC4 mutationand severe hemophagocytic lymphohistiocytosis. Pediatric Rheumatology; 13(Suppl 1):O68.

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[0804] 7.Booshehri LM and Hoffman HM (2019) CAPS and NLRP3. J Clin Immunol;39:277-286.

[0805] 8.Canna SW, Almeida de Jesus A, Gouni S, et al (2014) An activatingNLRC4 inflammasome mutation causes autoinflammation with recurrent macrophageactivation syndrome. Nat Genet; 10:1140-1146.

[0806] 9.Canna SW, Girard C, Malle L, et al (2017) Life-threatening NLRC4-associated hyperinflammation successfully treated with IL-18 inhibition. JAllergy Clin Immunol; 139:1698-1701.

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[0809] 12.Coppola S, Insalaco A, Zara E, et al (2022) Mutations at the C-terminus of CDC42 cause distinct hematopoietic and autoinflammatorydisorders, Journal of Allergy and Clinical Immunology, Volume 150, Issue 1,Pages 223-228, ISSN 0091-6749, https: / / doi.org / 10.1016 / j.jaci.2022.01.024.

[0810] 13.FDA (2007) Guidance for Industry. Drug-Induced Liver Injury: Pre-marketing Clinical Evaluation (Draft Guidance) (internet) Available from:https: / / evs.nci.nih.gov / ftp1 / CTCAE / Related / UCM072278.pdf (Accessed 08-Oct-2020).

[0811] 14.Feld LG and Corey H (2007) Hypertension in Childhood. Pediatric inReview; 28:283-98.

[0812] 15.Fleming S, Thompson M, Stevens R, et al (2011) Normal ranges ofheart rate and respiratory rate in children from birth to 18 years of age: asystematic review of observational studies. Lancet; 19; 377(9770):1011-8.

[0813] 16.Geerlinks AV Dvorak AM. et al (2022) XIAP Deficiency TreatmentConsortium. A Case of XIAP Deficiency Successfully Managed with TadekinigAlfa (rhIL-18BP). J Clin Immunol 42, 901-903. https: / / doi.org / 10.1007 / s10875-022-01236-2.

[0814] 17.Girard C, Rech J, Brown M, et al (2016) Elevated serum levels offree interleukin-18 in adult-onset Still’s disease. Rheumatology; 55:2237-2247.

[0815] 18.Heasman S, Ridley A. (2008) Mammalian Rho GTPases: new insightsinto their functions from in vivo studies. Nat Rev Mol Cell Biol 9, 690-701.https: / / doi.org / 10.1038 / nrm2476.

[0816] 19.Kofoed EM and Vance RE (2011) Innate immune recognition ofbacterial ligands by NAIPs determines inflammasome specificity. Nature; 477:592-595.

[0817] 20.Koike T, Harigai M, Mimura T et al (2014) Guidelines forPrevention of Immunosuppressive Therapy or Chemotherapy-induced Hepatitis BVirus Infection (Amendment). Japan College of Rheumatology.

[0818] 21.Kou R, and Shuei L (2009) Bradypnea, Encylopedia of MolecularMechanisms of Disease p241-243.

[0819] 22.Lam MT, Coppola S, Krumbach OHF et al (2019) A novel disorderinvolving dyshematopoiesis, inflammation, and HLH due to aberrant CDC42function. J Exp Med 2; 216 (12): 2778-2799. doi: https: / / doi.org / 10.1084 / jem.20190147 .

[0820] 23.Liang J, Alfano DN, Squires JE, et al (2017) Autoinflammation WithHemophagocytic Lymphohistiocytosis, Hepatosplenomegaly, Fetal ThromboticVasculopathy, and Congenital Anemia and Ascites. Pediatr Dev Pathol; 20(6):498-505.

[0821] 24.Melendez J, Grogg M, Zheng Y (2011) Signaling Role of Cdc42 inRegulating Mammalian Physiology, Journal of Biological Chemistry, Volume 286,Issue 4.

[0822] 25.Miao EA, Mao DP, Yudkovsky N, et al (2010) Innate immune detectionof the type III secretion apparatus through the NLRC4 inflammasome. Proc NatlAcad Sci USA; 107:3076-3080.

[0823] 26.Miyazawa H and Wada T (2022) Immune-mediated inflammatory diseaseswith chronic excess of serum interleukin-18. Front. Immunol. 13:930141. doi:10.3389 / fimmu.2022.930141

[0824] 27.Moghaddas F, Zeng P, Zhang Y, et al (2018) Autoinflammatorymutation in NLRC4 reveals a leucine-rich repeat (LRR)-LRR oligomerizationinterface. J Allergy Clin Immunol; 142:1956-1967 e1956.

[0825] 28.Mudde A, Booth C, Marsh R (2021) Evolution of our understanding ofXIAP deficiency. Front Pediatr; Jun 17;9;660520.

[0826] 29.Perez EE, Orange JS, Bonilla F, et al (2017) Update on the use ofimmunoglobulin in human disease: A review of evidence. J Allergy ClinImmunol; Mar;139(3S):S1-S46.

[0827] 30.Rigaud S, Fondanèche MC, Lambert N, et al (2006) XIAP deficiencyin humans causes an X-linked lymphoproliferative syndrome. Nature; 444(7115):110-4.

[0828] 31.Romberg N, Al Moussawi K, Nelson-Williams C, et al (2014) Mutationof NLRC4 causes a syndrome of enterocolitis and autoinflammation. NatureGenetics; 46(10):1135-1139.

[0829] 32.Romberg N, Vogel TP and Canna SW (2017) NLRC4 inflammasomopathies.Curr Opin Allergy Clin Immunol; 17:398-404.

[0830] 33.Tak PP, Bacchi M and Bertolino M (2006) Pharmacokinetics of IL-18binding protein in healthy volunteers and subjects with rheumatoid arthritisor plaque psoriasis. Eur J Drug Metab Pharmacokinet; 31:109-116.

[0831] 34.Takenouchi, T, Kosaki, R, Niizuma, T, et al (2015)Macrothrombocytopenia and Developmental Delay with a de novo CDC42 Mutation:Yet Another Locus for Thrombocytopenia and Developmental Delay. Am J MedGenet Part A 167A: 2822- 2825.

[0832] 35.Wada T, Kanegane H, Ohta K, et al (2014) Sustained elevation ofserum interleukin-18 and its association with hemophagocyticlymphohistiocytosis in XIAP deficiency. Cytokine. 2014;65(1):74-8.

[0833] 36.Weiss ES, Girard-Guyonvarc’h C, Holzinger D, et al (2018)Interleukin-18 diagnostically distinguishes and pathogenically promotes humanand murine macrophage activation syndrome. Blood; 131(13):1442-1455.

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[0836] 39. Zhao Y, Yang J, Shi J et al (2011) The NLRC4 inflammasomereceptors for bacterial flagellin and type III secretion apparatus. Nature;477: 596-600.

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[0838] Appendix 1: Clinically Obvious Laboratory Values ​​and Vital Signs

[0839] Researchers will inform Novartis of any obvious laboratory or vital sign abnormalities as defined below. Novartis will then determine whether further consultation with one or more researchers is appropriate.

[0840] Newly occurring significant laboratory abnormalities in pediatric patients (<16 years of age):

[0841] • Albumin: < LLN

[0842] •AST, ALT, and ALT or AST Increases >3 x-, 5 x-, 10 x-, and 20 x ULN

[0843] • Any increase in bilirubin; bilirubin increases to >1.5 x ULN and >2 x ULN.

[0844] • Any ALP elevation > 1.5 x ULN

[0845] • Elevated ALT and / or AST (> 3 x ULN) accompanied by elevated bilirubin (> 1.5 x ULN, > 2 x ULN)

[0846] • Gamma-glutamyl transferase (GGT): > 3 x ULN

[0847] • Creatinine (serum): ≥ 1.5 x ULN

[0848] Potassium: ≥ 5.5 mmol / L, or ≤ 3.5 mmol / L

[0849] • Magnesium: ≥ 1.2 mmol / L, or ≤ 0.7 mmol / L

[0850] • Sodium: ≥ 150 mmol / L, or ≤ 130 mmol / L

[0851] • Hemoglobin: A decrease of ≥ 2 g / dL or < 8.5 g / dL from baseline.

[0852] • Platelet count: < Lower limit of normal (LLN)

[0853] • White blood cell count: ≤ 0.8 x LLN or ≥ 1.2 x ULN

[0854] • Neutrophils: ≤ 0.9 x LLN or ≥ 1.2 x ULN

[0855] • Eosinophils: ≥ 1.1 x ULN

[0856] • Lymphocytes: < LLN or ≥ 1.1 x ULN

[0857] • Urine protein test strip: Positive (trace amount, ≥ +)

[0858] Significant abnormalities in vital signs in pediatric patients (<16 years old):

[0859] • Systolic / diastolic blood pressure 1 :

[0860] • Tall: ≥ 95th percentile of age and height group

[0861] • Low: ≤ 5th percentile of age and height group

[0862] • Oral temperature (°C)

[0863] • Height: ≥ 38.4°C

[0864] • Low: ≤ 35.0°C

[0865] • Pulse (bpm): Refer to Table 17-1

[0866] Table 9 Abnormal pulse rates (bpm) in children 2

[0867]

[0868] •weight:

[0869] •High: Compared to baseline 3 Increase BMI percentile categories for ages ≥ 2 4

[0870] • Low: Compared to baseline 3 BMI percentile reduction ≥ 2 years 4

[0871] • Respiratory rate: Refer to Table 17-2

[0872] Table 10 Abnormal respiratory rates (breaths / minute) in children 2, 5

[0873]

[0874] 1 The blood pressure percentile for each blood BP record was calculated using the methods described in Appendix B of The Fourth Report on the Diagnosis, Evaluation and Treatment of High Blood Pressure in Children and Adolescents (Anon 2004).

[0875] 2 Fleming S et al. 2011.

[0876] 3 Baseline age, BMI, and weight status categories are: underweight (below the 5th percentile), healthy weight (5th percentile to below the 85th percentile), overweight (85th percentile to below the 95th percentile), and obese (equal to or above the 95th percentile).

[0877] 4Obtain age BMI percentile categories (P3, P5, P10, P25, P50, P75, P85, P90, P95, P97) from WHO growth charts (www.who.int / childgrowth / en / ) (WHO 2020);

[0878] Note: For patients under 2 years old, growth charts are based on supine length, not height.

[0879] 5 Kou R and Shuei L 2009.

[0880] Notice: Only baseline values ​​will be marked as obvious anomalies.

[0881] Newly occurring significant laboratory abnormalities in adult patients (≥ 16 years of age):

[0882] • Albumin: < LLN

[0883] •AST, ALT, and ALT or AST Increase ULN by 3 x-, 5 x-, 10 x-, and 20 x.

[0884] • Any increase in bilirubin; bilirubin increases to >1.5 x ULN and >2 x ULN.

[0885] • Any ALP elevation > 1.5 x ULN

[0886] • Elevated AST and / or ALT (> 3 x ULN) accompanied by elevated bilirubin (> 1.5 x ULN, > 2 x ULN)

[0887] • Gamma-glutamyl transferase (GGT): > 3 x ULN

[0888] • Creatinine (serum): ≥ 1.5 x ULN

[0889] • Creatinine clearance rate: (Cockroft-Gault formula) : ≥ 25% reduction from baseline

[0890] Potassium: ≥ 5.5 mmol / L or ≤ 3.0 mmol / L

[0891] • Magnesium: ≥ 1.5 mmol / L, or ≤ 0.5 mmol / L

[0892] • Sodium: ≥ 150 mmol / L, or ≤ 130 mmol / L

[0893] • Calcium: ≥ 1.2 x ULN or < Lower limit of normal (LLN)

[0894] • Hemoglobin: A decrease of ≥ 2 g / dL or < 10.0 g / dL from baseline.

[0895] • Platelet count: < LLN

[0896] • White blood cell count: ≤ 0.8 x LLN or ≥ 1.2 x ULN

[0897] • Neutrophils: ≤ 0.9 x LLN or ≥ 1.2 x ULN

[0898] • Eosinophils: ≥ 1.1 x ULN

[0899] • Lymphocytes: < LLN or ≥ 1.1 x ULN

[0900] • Urine protein test strips: ≥ ++

[0901] Newly occurring, selective, significant vital sign abnormalities in adult patients (≥ 16 years of age):

[0902] • Systolic / diastolic blood pressure: ≥25% decrease or ≥25% increase from baseline, or ≥140 / 90 mmHg

[0903] • Pulse: ≥ 110 bpm with a change of ≥ 15% from baseline, or < 50 bpm with a change of ≥ 15% from baseline.

[0904] Source: Draft October 2007 FDA Guidance for Industry Drug-Induced Liver Injury: Premarketing Clinical Evaluation (FDA 2007)

[0905] Cockroft-Gault Formula (Male): Creatinine clearance (mL / min) = [((140 - Age (years)) x Weight (kg)) / (Serum creatinine (μmol / L) / 88.4)(mg / dL) x 72]

[0906] Cockroft-Gault Formula (Women): Creatinine clearance (mL / min) = [((140 - Age (years)) x Weight (kg)) / (Serum creatinine (μmol / L) / 88.4)(mg / dL) x 72] x 0.85

[0907] Notice: Only baseline values ​​will be marked as obvious anomalies.

[0908] Table 11 Recommended Sizes for Blood Pressure Cuff Cuffs

[0909]

[0910] Source: Feld and Corey in review [Pediatrics Review] (2007)

[0911] Appendix 2: Allergic Reactions

[0912] An allergic reaction is highly likely to occur when any one of the following three criteria is met:

[0913] 1. Acute onset of a disease involving the skin, mucous membranes, or both (from minutes to hours) (e.g., generalized urticaria, itching or flushing, swelling of the lips, tongue, and uvula).

[0914] And it meets at least one of the following:

[0915] a. Impaired breathing (e.g., dyspnea, wheezing-bronchospasm, stridor, decreased PEF, hypoxemia)

[0916] b. Decreased blood pressure or related end-organ dysfunction symptoms (e.g., hypotonia [collapse], syncope, incontinence)

[0917] 2. The patient experiences two or more of the following conditions rapidly after exposure to a possible allergen (within minutes to hours):

[0918] a. Involvement of skin and mucous membrane tissues (e.g., generalized urticaria, itching or flushing, swelling of the lips, tongue, and uvula).

[0919] b. Impaired breathing (e.g., dyspnea, wheezing-bronchospasm, stridor, decreased PEF, hypoxemia)

[0920] c. Decreased blood pressure or related symptoms (e.g., hypotonia [collapse], syncope, incontinence)

[0921] d. Persistent gastrointestinal symptoms (e.g., abdominal cramps, vomiting)

[0922] 3. Blood pressure decreases after exposure to a known allergen (from minutes to hours):

[0923] a. Adults: systolic blood pressure below 90 mm Hg or a decrease of more than 30% from baseline.

[0924] Appendix 3: Blood Collection Guidelines of the Toronto Hospital for Sick Children Research Ethics Board (REB)

[0925] The following recommendations from the Hospital for Sick Children's Research Ethics Committee in Toronto will guide researchers on the maximum amount of blood collected during a study. Researchers should closely monitor total blood volume to ensure compliance with the limits listed in the guidelines or local restrictions set forth by the IRB / EC.

[0926] For studies involving infants, children, and adolescents, the guidelines allow for a maximum of 5% of the patient’s total blood volume to be drawn in a single or multiple sessions over an eight-week period.

[0927] Table 12 shows the variation of blood volume with age; therefore, the usable amount per kg is:

[0928]

[0929] Adapted from The Hospital for Sick Children Research Ethics Board Blood Sampling Guide (Anon 2017). sequence list

[0930] Table 13 discloses useful amino acid and nucleotide sequences for carrying out the present invention.

[0931] Table 13. Sequences according to embodiments of the present invention

[0932]

[0933] In the entirety of this application, if there are any discrepancies between the specification text (e.g., Table 13) and the sequence list, the specification text shall prevail.

Claims

1. A method for treating or preventing XIAP-based autoinflammatory or disease in a subject in need, the method comprising administering a therapeutically effective amount of a bispecific antibody to the subject, wherein the antibody comprises a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18.

2. The method of claim 1, wherein the first and second constant heavy chains of the bispecific antibody are IgG1, and wherein... a. The first constant heavy chain has a point mutation that produces a pestle structure, and the second constant heavy chain has a point mutation that produces a mortar structure, or b. The first constant heavy chain has a point mutation that produces a mortar structure, and the second constant heavy chain has a point mutation that produces a pestle structure, and optionally... c. The first and second constant heavy chains have mutations that lead to disulfide bridges.

3. The method according to claims 1-2, wherein: a. The first immunoglobulin VH1 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; or ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 79, CDR2 has the amino acid sequence SEQ ID NO: 80, and CDR3 has the amino acid sequence SEQ ID NO: 81; and d. The first immunoglobulin VL1 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 93, and CDR3 has the amino acid sequence SEQ ID NO: 94 or... ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 95, CDR2 has the amino acid sequence SEQ ID NO: 96, and CDR3 has the amino acid sequence SEQ ID NO: 97; and e. The second immunoglobulin VH2 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; or ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 47, CDR2 has the amino acid sequence SEQ ID NO: 48, and CDR3 has the amino acid sequence SEQ ID NO: 49; and f. The second immunoglobulin VL2 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62 or... ii. Hypervariable regions CDR1, CDR2 and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 63, CDR2 has the amino acid sequence SEQ ID NO: 64 and CDR3 has the amino acid sequence SEQ ID NO:

65.

4. The method according to any one of the preceding claims, wherein: a. The first immunoglobulin VH1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

85. b. The first immunoglobulin VL1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

101. c. The second immunoglobulin VH2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 53, and d. The second immunoglobulin VL2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

69.

5. The method according to any one of the preceding claims, wherein: a. The first immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

87. b. The first immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

103. c. The second immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 55, and d. The second immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

71.

6. The method according to any one of the preceding claims, wherein the subject has a loss-of-function mutation in the XIAP / BIRC4 gene encoding the XIAP protein.

7. The method according to any one of the preceding claims, wherein the subject suffers from macrophage activation syndrome (MAS).

8. The method according to any one of the preceding claims, wherein the subject suffers from autoinflammation with infantile enterocolitis (AIFEC).

9. The method according to any one of the preceding claims, wherein the subject has excessively elevated serum IL-18 and serum IL-1β levels compared to a control group of subjects who do not have autoinflammatory or disease-based on XIAP deficiency.

10. The method according to any one of the preceding claims, wherein the patient with XIAP-based autoinflammatory or disease suffers from inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, or hemophagocytic lymphohistiocytosis.

11. The method according to any one of the preceding claims, wherein the subject has a serum ferritin level of >600 ng / mL or a serum C-reactive protein (CRP) level of >20 mg / L.

12. The method according to any one of the preceding claims, wherein the XIAP-based autoinflammatory or disease is resistant to treatment with cyclosporine, anti-TNFα therapy, systemic glucocorticoids, and anti-IL-1β therapy as a single therapy or as a combination thereof.

13. A bispecific antibody, said bispecific antibody comprising a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18 for use in treating or preventing autoinflammatory or disease-based on XIAP deficiency in subjects in need.

14. The bispecific antibody of claim 13 for use in treating or preventing XIAP-based autoinflammatory or disease in subjects in need, wherein the first and second constant heavy chains of the bispecific antibody are IgG1, and wherein... a. The first constant heavy chain has a point mutation that produces a pestle structure, and the second constant heavy chain has a point mutation that produces a mortar structure, or b. The first constant heavy chain has a point mutation that produces a mortar structure, and the second constant heavy chain has a point mutation that produces a pestle structure, and optionally... c. The first and second constant heavy chains have mutations that lead to disulfide bridges.

15. The bispecific antibody according to any one of claims 13-14 for use in treating or preventing XIAP-based autoinflammatory or disease-related conditions in subjects in need, wherein: a. The first immunoglobulin VH1 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; or ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 79, CDR2 has the amino acid sequence SEQ ID NO: 80, and CDR3 has the amino acid sequence SEQ ID NO: 81; and b. The first immunoglobulin VL1 domain of the bispecific antibody contains iii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 93, and CDR3 has the amino acid sequence SEQ ID NO: 94 or... iv. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 95, CDR2 has the amino acid sequence SEQ ID NO: 96, and CDR3 has the amino acid sequence SEQ ID NO: 97; and c. The second immunoglobulin VH2 domain of the bispecific antibody contains v. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; or vi. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 47, CDR2 has the amino acid sequence SEQ ID NO: 48, and CDR3 has the amino acid sequence SEQ ID NO: 49; and d. The second immunoglobulin VL2 domain of the bispecific antibody contains vii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62 or... viii. Hypervariable regions CDR1, CDR2 and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 63, CDR2 has the amino acid sequence SEQ ID NO: 64 and CDR3 has the amino acid sequence SEQ ID NO:

65.

16. The bispecific antibody according to any one of claims 13-15 for use in treating or preventing XIAP-based autoinflammatory or disease-related conditions in subjects in need, wherein: a. The first immunoglobulin VH1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

85. b. The first immunoglobulin VL1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

101. c. The second immunoglobulin VH2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 53, and d. The second immunoglobulin VL2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

69.

17. The bispecific antibody according to any one of claims 13-16 for use in treating or preventing XIAP-based autoinflammatory or disease-related conditions in subjects in need, wherein: a. The first immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

87. b. The first immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

103. c. The second immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 55, and d. The second immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

71.

18. The bispecific antibody for use in treating or preventing XIAP-based autoinflammatory or disease in subjects in need, according to any one of claims 13-17, wherein the subject has a loss-of-function mutation in the XIAP gene.

19. The bispecific antibody according to any one of claims 13-18 for use in treating or preventing XIAP-deficient autoinflammatory or disease in subjects in need, wherein the patient with XIAP-deficient autoinflammatory or disease suffers from inflammatory bowel disease, abdominal pain and diarrhea, relapsing fever, splenomegaly, or hemophagocytic lymphohistiocytosis (HLH).

20. The bispecific antibody according to any one of claims 13-19 for use in treating or preventing autoinflammatory or disease-based on XIAP deficiency in subjects in need, wherein the subjects suffer from macrophage activation syndrome (MAS).

21. The bispecific antibody according to any one of claims 13-20 for use in treating or preventing autoinflammatory or disease-based on XIAP deficiency in subjects in need, wherein the subject suffers from autoinflammatory disease with infantile enterocolitis (AIFEC).

22. The bispecific antibody according to any one of claims 13-21 for use in treating or preventing XIAP-deficient autoinflammatory or disease in subjects in need, wherein the subjects have excessively elevated serum IL-18 and serum IL-1β levels compared to a control group of subjects who do not have XIAP-deficient autoinflammatory or disease.

23. The bispecific antibody according to any one of claims 13-22 for use in treating or preventing XIAP-based autoinflammatory or disease in subjects in need, wherein the subject has a serum ferritin level of >600 ng / mL or a serum C-reactive protein (CRP) level of >20 mg / L.

24. The method according to any one of claims 1-12, the method comprising administering the bispecific antibody to the subject at a dose of about 1 mg / kg to about 35 mg / kg.

25. The method of claim 24, wherein the method comprises administering about 10 mg / kg of the bispecific antibody to the subject.

26. The method of claim 25, wherein the bispecific antibody is administered intravenously or subcutaneously.

27. The method of claim 25 or 26, wherein the dose of the administered bispecific antibody is about 10 mg / kg intravenously, optionally wherein the bispecific antibody is administered every other week.

28. The method of claim 26, wherein the dose of the applied bispecific antibody is from about 50 mg to about 900 mg subcutaneously.

29. The bispecific antibody for use in treating or preventing XIAP-deficient autoinflammatory or disease in subjects in need, according to any one of claims 13-23, wherein the use comprises administering the bispecific antibody to the subject at a dose of 1 mg / kg to 35 mg / kg.

30. The bispecific antibody of claim 29 for use in treating or preventing XIAP-deficient autoinflammatory or disease in subjects in need, said use comprising intravenous administration of 10 mg / kg of said bispecific antibody to said subject.

31. The bispecific antibody according to any one of claims 13-23 for use in treating or preventing autoinflammatory or disease-based on XIAP deficiency in subjects in need, wherein the bispecific antibody is administered subcutaneously at a dose of about 50 mg to about 900 mg.

32. A method for treating or preventing autoinflammatory or disease based on a CDC42 mutation in a subject in need, the method comprising administering a therapeutically effective amount of a bispecific antibody to the subject, wherein the antibody comprises a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18.

33. The method of claim 32, wherein the first and second constant heavy chains of the bispecific antibody are IgG1, and wherein... a. The first constant heavy chain has a point mutation that produces a pestle structure, and the second constant heavy chain has a point mutation that produces a mortar structure, or b. The first constant heavy chain has a point mutation that produces a mortar structure, and the second constant heavy chain has a point mutation that produces a pestle structure, and optionally... c. The first and second constant heavy chains have mutations that lead to disulfide bridges.

34. The method according to claims 32-33, wherein: a. The first immunoglobulin VH1 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; or ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 79, CDR2 has the amino acid sequence SEQ ID NO: 80, and CDR3 has the amino acid sequence SEQ ID NO: 81; and b. The first immunoglobulin VL1 domain of the bispecific antibody contains iii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 93, and CDR3 has the amino acid sequence SEQ ID NO: 94 or... iv. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 95, CDR2 has the amino acid sequence SEQ ID NO: 96, and CDR3 has the amino acid sequence SEQ ID NO: 97; and c. The second immunoglobulin VH2 domain of the bispecific antibody contains v. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; or vi. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 47, CDR2 has the amino acid sequence SEQ ID NO: 48, and CDR3 has the amino acid sequence SEQ ID NO: 49; and d. The second immunoglobulin VL2 domain of the bispecific antibody contains vii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62 or... viii. Hypervariable regions CDR1, CDR2 and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 63, CDR2 has the amino acid sequence SEQ ID NO: 64 and CDR3 has the amino acid sequence SEQ ID NO:

65.

35. The method according to claims 32-34, wherein: a. The first immunoglobulin VH1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

85. b. The first immunoglobulin VL1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

101. c. The second immunoglobulin VH2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 53, and d. The second immunoglobulin VL2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

69.

36. The method according to claims 32-35, wherein: a. The first immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

87. b. The first immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

103. c. The second immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 55, and d. The second immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

71.

37. The method according to claims 32-36, wherein the subject has a mutation in the CDC42 gene that causes abnormal palmitoylation of the CDC42 protein or a missense mutation at the C-terminus of the CDC42 protein that affects the localization of the CDC42 protein.

38. The method according to claims 32-37, wherein the subject suffers from macrophage activation syndrome (MAS).

39. The method according to claims 32-37, wherein the subject suffers from autoinflammation with infantile enterocolitis (AIFEC).

40. The method of claims 32-39, wherein the subject has excessively elevated serum IL-18 and serum IL-1β levels compared to a control group of subjects who do not have autoinflammatory or disease-based on CDC42 mutations.

41. The method according to claims 32-40, wherein the patient with autoinflammatory or disease-based CDC42 mutation suffers from neonatal episodic cytopenia, autoinflammatory disease, or recurrent hemophagocytic lymphohistiocytosis (HLH).

42. The method according to claims 32-41, wherein the subject has a serum ferritin level of >600 ng / mL or a serum C-reactive protein (CRP) level of >20 mg / L.

43. The method according to any one of claims 32-42, wherein the patient with autoinflammatory or disease-based CDC42 mutation is resistant to treatment with cyclosporine, anti-TNFα therapy, systemic glucocorticoids, and anti-IL-1β therapy as monotherapy or as a combination thereof.

44. A bispecific antibody, said bispecific antibody comprising a. A first portion, wherein the first portion is an immunoglobulin having a first variable light chain (VL1) and a first variable heavy chain (VH1) and a first constant heavy chain (CH1) with heterodimerization modification, the first portion specifically binding to IL1β, and b. The second part is an immunoglobulin having a second variable light chain (VL2) and a second variable heavy chain (VH2) and a second constant heavy chain (CH2) having a heterodimerization modification complementary to the heterodimerization modification of the first constant heavy chain, the second part specifically binding to IL-18 for use in treating or preventing autoinflammatory or disease-based on CDC42 mutations in subjects in need.

45. The bispecific antibody of claim 44 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein the first and second constant heavy chains of the bispecific antibody are IgG1, and wherein... d. The first constant heavy chain has a point mutation that produces a pestle structure, and the second constant heavy chain has a point mutation that produces a mortar structure, or e. The first constant heavy chain has a point mutation that produces a mortar structure, and the second constant heavy chain has a point mutation that produces a pestle structure, and optionally... f. The first and second constant heavy chains have mutations that lead to disulfide bridges.

46. ​​The bispecific antibody according to any one of claims 44-45 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein: e. The first immunoglobulin VH1 domain of the bispecific antibody contains i. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 76, CDR2 has the amino acid sequence SEQ ID NO: 77, and CDR3 has the amino acid sequence SEQ ID NO: 78; or ii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 79, CDR2 has the amino acid sequence SEQ ID NO: 80, and CDR3 has the amino acid sequence SEQ ID NO: 81; and f. The first immunoglobulin VL1 domain of the bispecific antibody contains iii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 92, CDR2 has the amino acid sequence SEQ ID NO: 93, and CDR3 has the amino acid sequence SEQ ID NO: 94 or... iv. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 95, CDR2 has the amino acid sequence SEQ ID NO: 96, and CDR3 has the amino acid sequence SEQ ID NO: 97; and g. The second immunoglobulin VH2 domain of the bispecific antibody contains v. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 44, CDR2 has the amino acid sequence SEQ ID NO: 45, and CDR3 has the amino acid sequence SEQ ID NO: 46; or vi. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 47, CDR2 has the amino acid sequence SEQ ID NO: 48, and CDR3 has the amino acid sequence SEQ ID NO: 49; and h. The second immunoglobulin VL2 domain of the bispecific antibody contains... vii. Hypervariable regions CDR1, CDR2, and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 60, CDR2 has the amino acid sequence SEQ ID NO: 61, and CDR3 has the amino acid sequence SEQ ID NO: 62 or... viii. Hypervariable regions CDR1, CDR2 and CDR3, wherein CDR1 has the amino acid sequence SEQ ID NO: 63, CDR2 has the amino acid sequence SEQ ID NO: 64 and CDR3 has the amino acid sequence SEQ ID NO:

65.

47. The bispecific antibody according to any one of claims 44-46 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein: e. The first immunoglobulin VH1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

85. f. The first immunoglobulin VL1 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

101. g. The second immunoglobulin VH2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 53, and h. The second immunoglobulin VL2 domain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

69.

48. The bispecific antibody according to any one of claims 44-47 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein: e. The first immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

87. f. The first immunoglobulin light chain of the bispecific antibody comprises the amino acid sequence SEQ ID NO:

103. g. The second immunoglobulin heavy chain of the bispecific antibody contains the amino acid sequence SEQ ID NO: 55, and h. The second immunoglobulin light chain of the bispecific antibody contains the amino acid sequence SEQ ID NO:

71.

49. The bispecific antibody according to any one of claims 44-48 for use in treating or preventing autoinflammatory or disease-based on CDC42 mutations in a subject in need, wherein the subject has a mutation in the CDC42 gene that causes abnormal palmitoylation of the CDC42 protein or a missense mutation at the C-terminus of the CDC42 protein that affects the localization of the CDC42 protein.

50. The bispecific antibody according to any one of claims 44-49 for use in treating or preventing autoinflammatory or disease-based on CDC42 mutations in subjects in need, wherein the patient suffers from neonatal episodic cytopenia, autoinflammatory disease, or recurrent hemophagocytic lymphohistiocytosis (HLH).

51. The bispecific antibody according to any one of claims 44-50 for use in treating or preventing autoinflammatory or disease-based on CDC42 mutation in a subject in need, wherein the subject suffers from macrophage activation syndrome (MAS).

52. The bispecific antibody of claim 50 for use in treating or preventing autoinflammatory or disease-based on CDC42 mutations in subjects in need, wherein the subject suffers from autoinflammatory disease with infantile enterocolitis (AIFEC).

53. The bispecific antibody according to any one of claims 44-52 for use in treating or preventing CDC42 mutation-based autoinflammatory or disease in subjects in need, wherein the subjects have excessively elevated serum IL-18 and serum IL-1β levels compared to a control group of subjects who do not have CDC42 mutation-based autoinflammatory or disease.

54. The bispecific antibody according to any one of claims 44-53 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein the subject has a serum ferritin level of >600 ng / mL or a serum C-reactive protein (CRP) level of >20 mg / L.

55. The method according to any one of claims 33-43, the method comprising administering the bispecific antibody to the subject at a dose of about 1 mg / kg to about 35 mg / kg.

56. The method of claim 55, wherein the method comprises administering about 10 mg / kg of the bispecific antibody to the subject.

57. The method of claim 56, wherein the bispecific antibody is administered intravenously or subcutaneously.

58. The method according to claims 56-57, wherein the dose of the administered bispecific antibody is about 10 mg / kg intravenously, optionally wherein the bispecific antibody is administered every other week.

59. The method of claim 57, wherein the dose of the applied bispecific antibody is from about 50 mg to about 900 mg administered subcutaneously.

60. The bispecific antibody for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in a subject in need, according to any one of claims 44-54, wherein the use comprises administering the bispecific antibody to the subject at a dose of 1 mg / kg to 35 mg / kg.

61. The bispecific antibody of claim 60 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in a subject in need, wherein the use comprises intravenous administration of 10 mg / kg of the bispecific antibody to the subject.

62. The bispecific antibody according to claims 60-61 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein the bispecific antibody is administered subcutaneously at a dose of about 50 mg to about 900 mg.

63. The treatment method according to claims 1-12, wherein the subject has a loss-of-function mutation in the XIAP gene.

64. The treatment method according to claims 32-43, wherein the subject has a loss-of-function mutation in the CDC42 gene.

65. The bispecific antibody according to any one of claims 44-54 and 60-62 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in a subject in need, wherein the subject has a loss-of-function mutation in the CDC42 gene.

66. The bispecific antibody according to claims 15-23 and 29-31 for use in treating or preventing autoinflammatory or disease-based on XIAP deficiency in subjects in need, wherein the subjects are resistant to treatment with cyclosporine, anti-TNFα therapy, systemic glucocorticoids, and anti-IL-1β therapy as monotherapy or as a combination thereof.

67. The bispecific antibody according to claims 44-54 and 60-62 for use in treating or preventing autoinflammatory or disease-associated with CDC42 mutations in subjects in need, wherein the subjects are resistant to treatment with cyclosporine, anti-TNFα therapy, systemic glucocorticoids, and anti-IL-1β therapy as monotherapy or as a combination thereof.

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