CN110167593A

CN110167593A - For treating or preventing method, composition and the dosage regimen of interferon-γ correlation indication

Info

Publication number: CN110167593A
Application number: CN201780080221.6A
Authority: CN
Inventors: C.德敏; W.费尔林; F.德贝内德蒂
Original assignee: Novimmune SA
Current assignee: Swedish Orphan Biovitrum AB
Priority date: 2016-10-24
Filing date: 2017-10-24
Publication date: 2019-08-23
Also published as: BR112019008156A2; CA3041434A1; WO2018078442A3; JP2019532970A; AU2017352033A1; SG11201903659WA; IL266174A; EP3528845A2; IL266174B1; KR20190071785A; JP2022187025A; MX2019004766A; WO2018078442A2; EA201990930A1

Abstract

The disclosure relates generally to the breaking-outs or development for treating, preventing and/or postpone to increase relevant symptom to IFN γ level, or mitigate method, composition and the dosage regimen that relevant symptom is increased to IFN γ level.

Description

For treating or preventing method, composition and the administration of interferon-γ correlation indication Scheme

Related application

This application claims U.S. Provisional Application No. 62/411783 benefit and priority submitted on October 24th, 2016, Content with it by reference to being fully incorporated herein.

Invention field

The disclosure relates generally to for treating, preventing and/or postponing and interferon gamma (IFN γ, IFN-gamma) Level increase relevant symptom such as Hemophagocytic lymphohistocysis disease (HLH), Hemorrhagic fever, CAR-T cell therapy, Graft failure, graft rejection, graft versus host disease(GVH disease) (GvHD) and/or diseases associated with inflammation relevant to graft rejection breaking-out or Development, or mitigate the method and composition of the symptom.Present disclosure also relates to the survivals of the biomaterial for extending transplanting Method and composition.

Background of invention

Human interferon gamma (IFN γ, IFN-gamma) be the lymph that is generated by the T lymphocyte and natural killer cells that activate because Son.It shows antiproliferative and immunoregulatory activity, and with the heterodimer on most of primary cells of immune system by Body IFN γ-R is combined, and triggering leads to the chain of events of inflammation.The immunoregulatory activity of known IFN γ is in many clinical diseases There is beneficial effect in disease.However, there is the clinical setting that many wherein known IFN γ activity have illeffects.For example, from Body immunological diseases are related to the high-level IFN γ in the blood and illing tissue of autoimmune patient.IFN γ activity also with This morbid state such as cachexia is related with septic shock.

IFN γ is related with many obstacles, and is developing anti-IFN γ drug as therapeutic agent.

Summary of the invention

In different aspect, the present invention provides more variable dose therapeutic schemes, for treating disease relevant to the raising of IFN-γ level Disease, obstruction and illness.

In one aspect, the present invention provides a kind of combination by intravenous administration subject the first dosage and the second dosage The antibody of interferon gamma (IFN γ) is used to treat the side of human primary's Hemophagocytic lymphohistocysis disease (HLH) Method.Subject is Adult human subjects or pediatric subject.First dosage is the weight and second of 1.0 or 3.0 mg/kg subjects Dosage is the weight of 3.0,6.0 or 10.0 mg/kg subjects.Optionally, the body of 1.0 mg/kg subject of third dosage is given Weight.

On the other hand, it is dry to provide a kind of combination by the first dosage of intravenous administration subject and the second dosage by the present invention The antibody for disturbing plain γ (IFN γ) is used to treat the secondary Hemophagocytic lymphohistocysis of mankind pediatric subject The method of disease (HLH).First dosage be 6.0 mg/kg subjects weight and the second dosage be 3.0 mg/kg subjects body Weight.Optionally, the weight of 6.0 mg/kg subject of third dosage is given.

On the other hand, the present invention provides a kind of combination by intravenous administration subject the first dosage and the second dosage The secondary Hemophagocytic lymphocyte that the antibody of interferon gamma (IFN γ) is used to treat adult humans subject increases The method of more diseases (HLH).First dosage be 3.0 mg/kg or 6.0 mg/kg subjects weight and the second dosage be no more than The weight of 10 mg/kg subjects.For example, the second dosage is 1.0,3.0,6.0 or 10.0 mg/kg.Optionally, it gives and is less than The third dosage of the weight of 10.0 mg/kg subjects.Such as third dosage is the body of 1.0,3.0 or 6.0 mg/kg subjects Weight.

Still on the other hand, the present invention provides a kind of by the first dosage of intravenous administration subject and the second dosage In conjunction with interferon gamma (IFN γ) antibody be used for treat human experimenter illness method.Subject be Adult human subjects or Pediatric subject.Illness is graft rejection such as solid organ transplantation obstacle or marrow acute transplantation rejection.Illness is graft Anti- host disease, paraneoplastic cerebellar degeneration, Hemorrhagic fever, sarcoidosis or adult onset still disease.Alternatively, receiving CART cell therapy Give this method to subject later.First dosage is between the weight of 1.0-10 mg/kg subject and the second dosage is in 1.0- Between the weight of 10 mg/kg subjects.For example, the second dosage is 1.0,3.0,6.0 or 10.0 mg/kg.Preferably, second dose Amount is higher or lower than the first dosage.Optionally, the third dosage between the weight of 1.0-10 mg/kg subject is given.Example Such as, the first, second or third dosage is the weight of 1.0,3.0,6.0 or 10.0 mg/kg subjects.

Antibody in conjunction with interferon gamma (IFN γ) includes: the amino acid sequence containing SYAMS (SEQ ID NO:1) Variable heavy chain complementary determining region 1 (VH CDR1), the amino acid sequence for containing AISGSGGSTYYADSVKG (SEQ ID NO:2) The variable heavy chain complementary determining region 2 (VH CDR2) of column and the amino acid for containing DGSSGWYVPHWFDP (SEQ ID NO:3) The variable heavy chain complementary determining region 3 (VH CDR3) of sequence；Amino acid containing TRSSGSIASNYVQ (SEQ ID NO:4) The variable light complementary determining region 1 (VL CDR1) of sequence, the amino acid sequence containing EDNQRPS (SEQ ID NO:5) Variable light complementary determining region 2 (VL CDR2) and containing QSYDGSNRWM (SEQ ID NO:6) amino acid sequence can Become complementary determining region of light chain 3 (VL CDR3).Such as antibody includes the weight chain variable ammonia of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of base acid sequence and SEQ ID NO:48.

The dosage of antibody is given in 1 hour, 6 hours or 12 hours.

It gives the second dosage within every 3 days after first dosage, continues the first treatment phase.In addition, terminating it in the first treatment phase After give the second dosage, continue the second treatment phase.Second treatment phase is for example twice a week.

Antibody dosage is given as single injection.

Antibody is given as monotherapy or conjoint therapy.

Optionally, method of the invention further comprises giving dexamethasone immediately before giving antibody.Dexamethasone With at least 10 mg/m²Or at least 5 mg/m²Dosage give.

Subject had not received the treatment to HLH previously.

In different aspect, method further comprises administering to subject's at least second of drug.Second drug be therapeutic agent, Anti-inflammatory agent and/or immunosuppressor.

The invention also includes the pharmaceutical formulation of injectable, every mL includes: full people's anti-interferon gamma of 5 mg or 25 mg (IFN γ) monoclonal antibody, 1.55 mg L-Histidines, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg chlorine Change sodium (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate, wherein pH is between 5.8-6.2.

On the other hand, the present invention provides a kind of unit dose vial, contain 20 ml be suitable for injection full people it is anti-dry Plain γ (IFN γ) monoclonal antibody solution is disturbed, wherein the concentration of antibody is 5 mg/ml or 25 mg/ml, and the pH of solution exists Between 5.8-6.2.Antibody is dissolved in solution, so that solution clarification, colourless and do not precipitate.

On the other hand, the present invention provides a kind of unit dose vial, contains 10 ml or 20 ml and is suitable for the complete of injection People's anti-interferon gamma (IFN γ) monoclonal antibody solution, wherein the concentration of antibody is 25 mg/ml, and the pH of solution exists Between 5.8-6.2.Antibody is dissolved in solution, so that solution clarification, colourless and do not precipitate.

Still on the other hand, the present invention provides a kind of unit dose vial, contains 2 ml or 10 ml are suitable for injecting Full people's anti-interferon gamma (IFN γ) monoclonal antibody solution, wherein the concentration of antibody is 5 mg/ml, and the pH of solution exists Between 5.8-6.2.Antibody is dissolved in solution, so that solution clarification, colourless and do not precipitate.

Any the above or embodiment can be with any other aspect or combination of embodiment.

Unless otherwise defined, otherwise all technical and scientific terms used herein have with it is of the art general The logical normally understood identical meanings of technical staff.Although similar or equivalent method and material can be used for those described herein In practice of the invention, but suitable method and material is described below.All disclosures for being mentioned above, patent application, patent and Other bibliography with it by reference to all clearly being combined.In case of conflict, it is with this specification (including definition) It is quasi-.In addition, material described herein, method and embodiment are only illustrative, it is restrictive without being intended that.

Other feature and advantage of the invention will be apparent from described below and claim and be included therein.

Brief description

Fig. 1 is to be depicted in the ongoing 2 phase Primary Study of primary HLH patient, be administered before change of serum C XCL9 level with it is defeated The chart of correlation after note NI-0501 antibody between 24 hours total IFN γ levels.

Fig. 2 is to be depicted in the ongoing 2 phase Primary Study of primary HLH patient, change of serum C XCL9 level and infusion The chart of correlation after NI-0501 antibody before total IFN γ administration in 24 hours between level.

Fig. 3 A and 3B are a series of charts, and it is thin to be depicted in the macrophage secondary to whole body type juvenile idiopathic arthritis (sJIA) The horizontal correlation between IFN γ level of change of serum C XCL9 in born of the same parents' activation syndrome (MAS) patient and activity sJIA patient.

Fig. 4 A-1,4A-2,4B-1,4B-2,4C-1,4C-2,4D-1 and 4D-2 are a series of charts, are depicted in activity IFN γ and change of serum C XCL9 is horizontal and the correlation between clinical parameter in sJIA and MAS patient secondary to sJIA.

Fig. 5 is the chart describing IFN γ and being fully neutralized, such as the IFN γ induction type chemotactic factor (CF) institute of undetectable level Show.

Fig. 6 is the improved chart of (2 weeks and treatment end) HLH disease activity during describing NI-0501 treatment: blood is small Plate counts > 100 x109/L, neutrophil count > 1x109/L, and fibrinogen > 1.5 g/L and ferritin are reduced at least 25% patient's percentage.

Fig. 7 A and 7B are a series of charts, describe 24 hours total IFN γ water after preceding CXCL9 and NI-0501 infusion is administered Correlation between flat.The example of illustration as shown in Figure 7 B individual IFN γ and CXCL9 feature during describing NI-0501 treatment.

Fig. 8 A, 8B, 8C and 8D are a series of charts, do not have the activity of MAS during being depicted in activity MAS and in sampling Property sJIA (Act sJIA) during can get IFN γ and CXCL9, CXCL10 and CXCL11 in the individual patients of paired samples Serum levels.Significance (p) is obtained using the Wilcoxon rank tests of paired samples.

Fig. 9 A and 9B are a series of charts, are depicted in the patient that 3 MAS breaking-out is showed during its sJIA process Middle leucocyte (WBC) and blood platelet (PLT) count variation (Fig. 9 A) and IFN γ, CXCL9, CXCL10 with ferritin levels With the variation (Fig. 9 B) of the serum levels of CXCL11.

Figure 10 A, 10B, 10C, 10D, 10E, 10F, 10G, 10H, 10I and 10J are a series of charts, trouble when being depicted in sampling There is no the IFN γ of the activity sJIA patient (black triangle) of MAS when having patient's (red circle) and the sampling of activity MAS With the horizontal correlation with ferritin levels, neutrophil leucocyte and platelet count and with LDH and ALT level of CXCL9.Each phase The Spearman related coefficient (Rs) and significance (p) of closing property are as shown in table 3.

Figure 11 A, 11B, 11C, 11D, 11E and 11F be a series of charts, describe MAS in IFN γ and CXCL9 and The relationship that CXCL10 is generated.Scheme A: sampling when the patient with MAS in IFN γ level and CXCL9 and CXCL10 level phase Guan Xing.The Spearman related coefficient (Rs) and significance (p) of each correlation are as shown in table 3.

Figure 12 is the diagram of the screening of the research presented in embodiment 7, treatment and follow-up part.

Figure 13 A and 13B are that NI-0501 gives in two patients of body temperature > 37.5 DEG C when being depicted in NI-0501 treatment beginning The chart of influence to body temperature.

Figure 14 is a series of charts and table, describes NI-0501 and gives the influence to patient's neutrophil count.

Figure 15 is a series of charts and table, describes NI-0501 and gives the influence counted to Platelet.

Figure 16 is a series of charts and table, describes NI-0501 and gives the influence to patient's ferritin serum levels.

Figure 17 is a series of charts and table, describes NI-0501 and gives the influence being gradually reduced to patient's glucocorticoid.

Figure 18 is to describe giving for NI-0510 IFN γ is kept to neutralize the chart until when HSCT.HLH treats NI-0501 Reaction also continue to transplanting.

Figure 19 is the diagram of the screening of the research presented in embodiment 8, treatment and follow-up part.

Detailed description of the invention

Composition provided herein and method use full human IgG1's anti-interferon gamma (IFN γ) monoclonal antibody (mAb), herein Referred to as NI-0501, in conjunction with and neutralize IFN γ.The solubility and receptor (IFN γ R1) combining form knot of NI-0501 and IFN γ It closes.Since NI-0501 is human IgG1, retain the feature of this Immunoglobulin Isotype, including participate in Fc γ receptor and knot Close the ability of complement.IFN γ is one of most effective in immune system and pleiotropic cytokines.For virus and Intracellular bacterial The congenital and adaptive immunity of infection is most important.After in conjunction with its receptor, IFN γ, which plays, generates a variety of physiology and thin The effect of born of the same parents' reaction.Past 20 years numerous studies are by IFN γ (ginseng associated with the pathogenesis of diseases associated with inflammation and maintenance See such as Billiau A. " Interferon-gamma:biology and role in pathogenesis. " Adv. Immunol. 1996; 62:61-130; Schoenborn JR, Wilson CB. “Regulation of interferon-gamma during innate and adaptive immune responses.” Adv. Immunol. 2007; 96:41-101;With Zhang SY, Boisson-Dupuis S, Chapgier A et al. " Inborn errors of interferon (IFN)-mediated immunity in humans: insights into the respective roles of IFN-alpha/beta, IFN-gamma, and IFN-lambda in host defense." Immunol. Rev. 2008; 226:29-40).IFN γ is main as a part that congenital immunity reacts It is generated by natural kill (NK) and natural killer T (NKT) cell, and by CD4 Th1 once there is antigen specific immune It is generated with cd8 cell toxic T lymphocyte (CTL) effector T cell.

Composition provided herein and method can be used for treating disease relevant to IFN γ raising, obstacle and illness.It is suitble to Include such as Hemophagocytic lymph group in disease, obstacle and the illness treated or prevented using the compositions and methods of the invention Knit cytosis (HLH), graft versus host disease(GVH disease), paraneoplastic cerebellar degeneration, Hemorrhagic fever, sarcoidosis, adult onset still disease, Graft failure, graft rejection and/or diseases associated with inflammation relevant to graft rejection.Graft rejection includes solid organ transplantation barrier Hinder, marrow acute transplantation rejection.In addition, the composition and method can also be used for treating or mitigating the secondary work of CAR-T cell therapy With.

HLH is a kind of syndrome, it is characterized in that NK and CD8+ T cell is badly damaged or is not present cytotoxicity function, Middle immune system significantly activates.HLH includes primary (heredity/familial) HLH and secondary HLH, and the two clinically claims For immune system dysfunction, lead to serious cytokinemia, detrimental consequences are generated to various tissues and organ (Henter JI, Elinder G, Soder O et al. “Hypercytokinemia in familial hemophagocytic lymphohistiocytosis." Blood 1991; 78:2918-2922).HLH classification is such as following Shown in table 9.HLH is betided with both adult and pediatric patients.

9. HLH of table classification

Primary HLH is heterogeneous autosomal recessive hereditary diseases.Primary HLH is mainly seen in infancy and infancy, and There is 1/50000 live-born infant illness rate (Henter JI, Elinder G, Soder O, Ost A. Incidence in estimation Europe in Sweden and clinical features of familial hemophagocytic lymphohistiocytosis. Acta Paediatr. Scand. 1991; 80:428-435).If do not treated, the disease Always fatal, median survival interval less than 2 months (the Janka GE. Familial after paresthesia epilepsy of disease hemophagocytic lymphohistiocytosis. Eur. J. Pediatr. 1983; 140:221-230;With Aricò M, Janka G, Fischer A, Henter JI, Blanche S, Elinder G, Martinetti M, Rusca MP Hemophagocytic lymphohistiocytosis Report of 122 children from the International Registry. FHL Study Group of the Histiocyte Society. Leukemia. 1996 Feb; 10(2):197-203)。

The cytotoxicity function being damaged existing for HLH leads to cytokinemia and erythrophagoytosis.These are transferred Lead to all classical symptoms (Dhote R, Simon J, Papo T et al. Reactive hemophagocytic of HLH syndrome in adult systemic disease: report of twenty-six cases and literature review. Arthritis Rheum. 2003; 49:633-639; Risdall RJ, McKenna RW, Nesbit ME et al. Virus-associated hemophagocytic syndrome: a benign histiocytic proliferation distinct from malignant histiocytosis. Cancer 1979; 44:993- 1002;With Risdall RJ, Brunning RD, Hernandez JI, Gordon DH. Bacteria-associated hemophagocytic syndrome. Cancer 1984; 54:2968-2972).The classical symptom of HLH includes for example lasting Fever, hepatomegaly, haemocyte reduction, high-speed rail proteinemia, hypertriglyceridemia, hypofibrinogenemia, is bitten at splenomegaly Red blood cell effect, cytokinemia and/or lymphocyte infiltration, hypoplastic bone marrow, meninx infiltration.

Raised cell factor includes: IFN γ, interleukin-6 (IL-6), IL-10, neoplasm necrosis in HLH patient The factor (TNF) α, IL-8, macrophage colony stimulating factor (MCSF) and granulocyte-macrophage colony stimutaing factor (GM- CSF)。

During HLH can also betide infection, rheumatic or tumor disease process, and in this case it is referred to as Secondary HLH.Secondary HLH show with primary identic S&S, and may be equally serious.It is directed at present The treatment of secondary HLH aims to solve the problem that the inducement of potential disease.For as infecting HLH caused by such as leishmaniasis, situation is true It is real such.It should be noted that the presence of certain infection especially virus infection (such as due to infection caused by CMV or EBV) The triggering factors of the primary form performance of usually HLH.In the animal model of primary HLH, lymphatic choroid plexus brain Film scorching viral (LCMV) infection be evidence required for the disease develops also support the observation result (Jordan MB, Hildeman D, Kappler J, Marrack P. An animal model of hemophagocytic lymphohistiocytosis (HLH): CD8+ T cells and interferon gamma are essential for the disorder. Blood 2004; 104:735-743; Pachlopnik SJ, Ho CH, Chretien F et al. Neutralization of IFNgamma defeats haemophagocytosis in LCMV- infected perforin- and Rab27a-deficient mice. EMBO Mol. Med. 2009; 1:112-124; Kögl T, Müller J, Jessen B et al. Hemophagocytic lymphohistiocytosis in syntaxin-11- deficient mice: T- cell exhaustion limits fatal disease. Blood. 2013; 121: 604-613;With Sepulveda FE, Debeume F, Menasch é G et al. Distinct severity of HLH in both human and murine mutants with complete loss of cytotoxic effector PRF1, RAB27A, and STX11 Blood. 2013;121:595-603)。

When HLH occurs during tumor disease, especially hematologic malignancies, the severity of usual patient condition It needs to treat HLH immediately before specificity solves potential disease.

With rheumatic disease (such as whole body type juvenile idiopathic arthritis (sJIA) and systemic loupus erythematosus (SLE)) there are the S&S of HLH in patient, Macrophage Activation Syndrome is usually known as by rheumatologist It (MAS), and can be prior to the appearance of rheumatic disease itself.The NK of most of MAS patients and perforin functional test are impaired, and And a considerable amount of patients show polymorphism or heterozygous mutant in PRF1 and UNC13D.Although its to be a kind of extremely serious and The illness of threat to life, but can usually subside when starting sufficiently treatment, wherein in most cases including corticosteroid And cyclosporin.However, the disease is likely difficult to control and be contemplated that using support in about 15% patient for MAS occur Moor glycosides (Minoia F, Davi S, Horne AC et al. Clinical Features, Treatment, and Outcome of Macrophage Activation Syndrome Complicating Systemic Juvenile Idiopathic Arthritis: A Multinational, Multicenter Study of 362 Patients. Arthritis & Rheumatism 2014; 66: 3160-3169)。

Although primary HLH thinks mainly childhood illness, HLH is a kind of illness that can be found in adult, and And have become increasingly aware of this may than it is considered that generation it is more frequent.In most of adult patients, the disease is in malignant tumour Occur during (mainly non-Hodgkin lymphoma), infection, auto-inflammatory or autoimmune disease and iatrogenic immune deficiency.

It there is no the drug ratified for treating HLH at present.However, the expert in the field has been that treatment HLH patient formulates Guideline (Henter JI, Horne AC, Arico ' M, Egeler RM, Filipovich AH, Imashuku S Ladisch S, McClain K, Webb D, Winiarski J, and Janka Diagnostic and Therapeutic Guidelines for Hemophagocytic Lymphohistiocytosis Blood Cancer 2007; 48:124-13.1; Henter JI, Samuelsson-Horne A, Arico M et al. Treatment of hemophagocytic lymphohistiocytosis with HLH-94 immunochemotherapy and bone marrow transplantation. Blood 2002; 100:2367-2373;With Jordan MB, Allen CE, Weitzman S, Filipovich AH, McClain KL. How I treat hemophagocytic lymphohistiocytosis. Blood 2011; 118:4041-4052)。

The treatment of primary HLH patient includes the following steps (Henter et al., Blood Cancer at present 2007): (i) is with corticosteroid and immunosuppressive drug (such as Etoposide, CsA, alemtuzumab, anti-thymocyte ball egg It is white) combination 8 weeks inductive treatments；(ii) maintenance therapy is until transplanting；(iii) is to all genetic defectes with identification Patient transplant, and eventually in the HLH case very serious without the relevant mutation of disease.

The main target of inductive treatment is the inflammatory process for inhibiting the threat to life of the feature as HLH, so that needing Those of it patient is transplanted (Horne A, Janka G, Maarten ER et al. Haematopoietic stem cell transplantation in haemophagocytic lymphohistiocytosis. Br. J. Haematol. 2005; 129:622-630).Transplanting is unique curative therapy (Henter to HLH relevant to high penetrance gene mutation et al., Blood 2002)。

Although with this guideline, but the general mortality rate of primary HLH is still about 40-50% (Henter et al., Blood 2002; Trottestam H, Horne A, Arico M et al. Chemoimmunotherapy for hemophagocytic lymphohistiocytosis: long-term results of the HLH-94 treatment protocol. Blood 2011;118:4577-4584)。

It is needed during induction using drug relevant to serious short-term and long-term safety problem, this further promotes Through the very high death rate.Composition provided herein and method are developed as targeted therapy, it is ensured that curative effect and toxicity is smaller.

In the past few years, the key effect in terms of the development about IFN γ in HLH has produced more and more Evidence (Henter JI, Elinder G, Soder O et al. Hypercytokinemia in familial hemophagocytic lymphohistiocytosis. Blood 1991; 78:2918-2922; Jordan MB, Hildeman D, Kappler J, Marrack P. An animal model of hemophagocytic lymphohistiocytosis (HLH): CD8+ T cells and interferon gamma are essential for the disorder. Blood 2004; 104:735-743; Pachlopnik SJ, Ho CH, Chretien F et al. Neutralization of IFN gamma defeats haemophagocytosis in LCMV-infected perforin- and Rab27a-deficient mice. EMBO Mol. Med. 2009; 1:112-124; Behrens EM, Canna SW, Slade K et al. Repeated TLR9 stimulation results in macrophage activation syndrome-like disease in mice. J. Clin. Invest 2011; 121:2264- 2277; Xu XJ, Tang YM, Song H, MD, Yang SL, Xu WQ, Zhao N, Shi SW, Shen HP, Mao JQ, Zhang LY, and Pan BH, Diagnostic Accuracy of a Specific Cytokine Pattern in Hemophagocytic Lymphohistiocytosis in Children J Pediatr 2011;With Risma K, Jordan MB. Hemophagocytic lymphohistiocytosis: updates and evolving concepts. Curr. Opin. Pediatr. 2012; 24:9-15)。

The gene mutation of the feature of primary form as HLH will affect the protein for participating in same process, final to damage Evil cytotoxic activity.Perforin is mutated in HLH patient's first identified.

Perforin knockout (KO) mouse is considered the correlation model of human diseases.In fact, these mouse once infected with LCMV, will develop human diseases all diagnosis and many clinical and laboratory specific characteristic, and if without controlling Treating them will be dead.For these reasons, perforin KO mice has been used for the Pathological Physiology of research HLH.What they occurred The IFN γ that HLH sample pathology are generated dependent on CD8+ T cell and response antigenic stimulus.

The result shows that not only clinical and laboratory is different when giving in anti-IFN γ antibody with the IFN γ of high circulation level Often restore, and survival rate significantly improves.On the contrary, the ablation of any other cell factor is on survival without influencing (Jordan et al., Blood 2004; Pachlopnik et al., EMBO Mol. Med. 2009)。

Two kinds of secondary HLH models are had studied under the background of NI-0501 development plan.In a kind of model, repetition is given Giving the chronic severe overstimulation that CpG (TLR9 is caused to stimulate) has been used for simulation healthy mice (has cytotoxicity approach Normal science of heredity) as the HLH model secondary to infection.Although these mouse are not necessarily dead, there is the typical case of HLH in them Clinic and Laboratory Characteristic.When IFN γ is neutralized, clinic and laboratory spy that anti-IFN γ antibody restores the disease are given Sign.It is interesting that having confirmed that giving anti-IFN γ antibody also causes in relevant target tissue such as liver and spleen in the model The complete neutralization (manuscript in preparation) of IFN γ effect.

In order to study the physiopathology of the secondary HLH occurred under the background of rheumatic disease, expression Gao Shui is used The IL-6 transgenic mice of flat IL-6 produces animal model, and similar to the patient with sJIA, there is a situation where sJIA is most Often rheumatic disease relevant to the ondary forms of HLH.When being triggered with Toll-like receptor (TLR) ligand, these mouse are died of Many features (Strippolli R, Carvallo F, the Scianaro R et al. Amplification of human diseases of the response to Toll-like receptor ligands by prolonged exposure to interleukin-6 in mice: Implication for the pathogenesis of macrophage activation syndrome. Arthritis & Rheumatism 2012; 64: 1680-1688).In these mouse In, when IFN γ with give in anti-IFN γ antibody and when, survival significantly improves and laboratory parameters restore (Prencipe G Et al, the manuscript in preparation).

It is horizontal to further strengthen the circulation IFN γ that importance of the IFN γ in HLH is primary HLH patient middle and high concentration (Henter et al., Blood 1991; Xu et al., J Pedatr 2011).A series of from HLH diagnosis to treatment In 71 patients being monitored with follow-up, the IFN γ level of all patients is above the upper limit of normal value (17.3 pg/mL), And especially 53.5% patient levels are higher than 1000 pg/mL.Also it is reported that IFN γ horizontal early stage and rapid increase, and After effectively treatment HLH normal value can be down to from > 5000 pg/mL in 48 hours.

Recently, in the observational study of the patient of the ondary forms with HLH, with secondary to infection The patient of HLH and patient with the HLH occurred under the background of sJIA both show high-caliber IFN γ.It is known by 3 kinds of Chemokine CXCL9, CXCL10 and the horizontal also significant of CXCL11 of IFN γ induction increase.It is worth noting that, discovery The level and the laboratory parameters such as ferritin, platelet count of disease severity of IFN γ and 3 kinds of IFN γ chemotactic factor (CF)s It is significant related (Bracaglia et al., contribution are submitted) to transaminase.

Since the l&H cytokinemia of activation and organ infiltration are to cause all HLH diseases The reason of shape and horizontal dependent on CD8+ T cell overacfivity and high IFN γ, therefore the neutralization composition of IFN γ is reasonably controlled Treatment method.In fact, currently without the drug of CD8+ T cell is specifically targeted, and target the single thin of IFN γ downstream Intracellular cytokine is not necessarily feasible.

Therefore, based on the animal model from primary and secondary HLH and to from primary and secondary The data for the observation that the patient of both HLH carries out, it was demonstrated that the key effect that IFN γ plays in the pathogenesis of the disease, IFN Targeted therapy of the neutralization of γ for exploitation for HLH provides strong theoretical basis, must be effective and without toxicity Or limited toxicity.

The disclosure also provides the composition and method that can be used for identifying or improve the patients with obstacle, wherein suffering from Person has individually or combines with one or more other interferon gamma (IFN γ) associated biomarkers raised CXCL9 is horizontal.Particularly, the disclosure provides the composition and method for detecting CXCL9 level, and CXCL9 is Hemophagocytic IFN γ in lymphohistocysis's disease (HLH), secondary HLH (secondary HLH) and/or Macrophage Activation Syndrome (MAS) The biomarker of generation.

A large amount of evidences of animal model show IFN γ in primary Hemophagocytic lymphohistocysis disease (HLH) In there are crucial pathogenic effects.In the mankind with HLH it has also been found that high-caliber IFN γ.Previously it has been reported that in activity MAS patient view is to high-caliber IFN γ and the relevant chemotactic factor (CF) (CXCL9, CXCL10 and CXCL11) of 3 kinds of IFN γs, MAS The secondary HLH occurred under the background of whole body type juvenile idiopathic arthritis (sJIA) for one kind form (see, for example, Bracaglia C., Caiello I, De Graaf K., et al. Pediatric Rheumatology 2014,12 (Suppl 1):O3).The circumstantial evidence of mouse shows that IFN γ is mainly generated in peripheral tissues, and haemoconcentration may be opposite It is lower.

Term Macrophage Activation Syndrome (MAS) refers to the potential mortality of the severe of chronic inflammation rheumatic disease simultaneously Send out disease.Under its background for generally betiding whole body type juvenile idiopathic arthritis (sJIA), wherein the patient of 10-20% is in disease Occurs the syndrome during sick process.It can also be in systemic loupus erythematosus, Kawasaki disease and other autoimmunities and itself inflammation Occur in disease property disease, although less common.In sJIA, MAS generally occurs during the disease activity phase, is included in disease hair When making.Infectious triggering factors can be identified in a high proportion of patient.The characteristic feature of MAS include fever, splenomegaly, bleeding with And it may cause the sign of the liver of multiple organ failure, central nervous system and renal involvement.Laboratory abnormalities include leucocyte, Blood platelet and decreased hemoglobin, high transaminase mass formed by blood stasis, ferritin dramatically increase, the evidence of the Intravascular activation of blood coagulation system (Ravelli, A., et al., Macrophage activation syndrome as part of systemic juvenile idiopathic arthritis: diagnosis, genetics, pathophysiology and treatment. Genes Immun. 13(4): p. 289-98).MAS leads to significant morbidity and mortality, account for due to Dead relevant portion caused by sJIA (Minoia, F., et al.,Clinical features, treatment, and outcome of macrophage activation syndrome complicating systemic juvenile idiopathic arthritis: a multinational, multicenter study of 362 patients. Arthritis Rheumatol, 2014. 66(11): p. 3160-9; Hashkes, P.J., et al.,Mortality outcomes in pediatric rheumatology in the US. Arthritis Rheum, 2010. 62(2): p. 599-608).Disease incidence mechanism is more fully understood, it is later determined that new therapy target and targeting are treated The possibility of method develops, and may result in the treatment of MAS and significantly improving for result.

MAS shares the most of Clinical symptoms and laboratory abnormalities of Hemophagocytic lymphohistocysis disease (HLH), And its be classified as really at present secondary or reactive HLH (sec-HLH) (Jordan, M.B., et al.,How I treat hemophagocytic lymphohistiocytosis. Blood, 2011. 118(15): p. 4041-52)。 The mutation that the primary form (p-HLH) of HLH is participated in the gene of the protein of granule exocytosis by coding causes, including PRF1, UNC13D, STXBP2, STX11, RAB27A and XIAP typically result in the deficient cell poison of CD8+ lymphocyte and NK cell Activity.According to current classification, in the case where no identifiable genetic cause and/or familial inheritance, HLH be defined as after Hair property is reactive.Sec-HLH can be in the case where not having verifiable triggering factors, or in infection, pernicious swollen Occur under tumor or the background of rheumatic disease, the latter is commonly referred to as MAS.The hereditary basis for MAS occur just gradually is elucidated with, many Research shows that the identical Disease-causing gene of heterozygosity or p-HLH that MAS (and usually sec-HLH) and low penetrance make a variation Mutation correlation (Kaufman, K.M., et al.,Whole-exome sequencing reveals overlap between macrophage activation syndrome in systemic juvenile idiopathic arthritis and familial hemophagocytic lymphohistiocytosis. Arthritis Rheumatol, 2014. 66(12): p. 3486-95; Vastert, S.J., et al., Mutations in the perforin gene can be linked to macrophage activation syndrome in patients with systemic onset juvenile idiopathic arthritis. Rheumatology (Oxford), 2010. 49(3): p. 441-9.; Zhang, K., et al., Macrophage activation syndrome in patients with systemic juvenile idiopathic arthritis is associated with MUNC13-4 polymorphisms. Arthritis Rheum, 2008. 58(9): p. 2892-6;And Zhang, M., et al., Genetic defects in cytolysis in macrophage activation syndrome. Curr Rheumatol Rep, 2014. 16(9): p. 439;With Bracaglia C, Sieni E, Da Ros M, et al. Mutations of familial hemophagocytic lymphohistiocytosis (FHL) related genes and abnormalities of cytotoxicity function tests in patients with macrophage activation syndrome (MAS) occurring in systemic juvenile idiopathic arthritis (sJIA). Pediatric Rheumatology 2014, 12(Suppl 1):P53).Heredity back between p-HLH and MAS These similitudes of scape further support shared pathogenic mechanism.

Such hypothesis: antigen presenting cell is supported to the research of patient and p-HLH mouse model with p-HLH (APC) the defects of-CD8+ T cell crosstalk type cytotoxic activity and the abnormal deficiency silencing and T cell for leading to immune response Activation is abnormal.This leads to the generation of T lymphocyte and macrophage uncontrolled immune activation and proinflammatory cytokine, causes Organ damage.In the p-HLH animal model that perforin and Rab27 deficient mice carry out research shows that CD8+ T by activating The key effect for the interferon gamma (IFN γ) that cell generates.In Perforin knockout type mouse, the neutralization of IFN γ leads to other causes The survival of dead syndrome, reverse biochemistry and hematological abnormality (Jordan, M.B., et al.,An animal model of hemophagocytic lymphohistiocytosis (HLH): CD8+ T cells and interferon gamma are essential for the disorder. Blood, 2004. 104(3): p. 735-43; Pachlopnik Schmid, J., et al., Neutralization of IFNgamma defeats haemophagocytosis in LCMV-infected perforin- and Rab27a-deficient mice. EMBO Mol Med, 2009. 1(2): p. 112-24).In Rab27 deficient mice, wherein the disease not will lead to death, and the neutralization of IFN γ leads to peripheral organ The involvement of (including central nervous system) significantly improves (Pachlopnik 2009).It is diagnosed according to 2004 diagnosis guide of HLH HLH patient it has also been found that the IFN γ of high circulation it is horizontal (My, L.T., et al.,Comprehensive analyses and characterization of haemophagocytic lymphohistiocytosis in Vietnamese children. Br J Haematol, 2010. 148(2): p.301-10; Takada, H., et al.,Increased serum levels of interferon-gamma-inducible protein 10 and monokine induced by gamma interferon in patients with haemophagocytic lymphohistiocytosis. Clin Exp Immunol, 2003. 133(3): p. 448-53; Tang, Y., et al., Early diagnostic and prognostic significance of a specific Th1/Th2 cytokine pattern in children with haemophagocytic syndrome. Br J Haematol, 2008. 143(1): p. 84-91; Xu, X.J., et al., Diagnostic accuracy of a specific cytokine pattern in hemophagocytic lymphohistiocytosis in children. J Pediatr, 2012. 160 (6): p. 984-90 e1.), and therefore it is not necessarily based upon the presence of gene mutation.It should refer to Out, these researchs include patient's (ibid) without verifiable genetic cause of significant (although variable) ratio.

Research provided herein be intended to assess the serum levels of IFN γ and 3 kinds of IFN γ associated chemokines with itself with And the correlation between the laboratory parameters of the disease activity of activity MAS patient, with aborning life in Searching I FN γ body Object marker.In particular, the cyclical level of IFN γ, CXCL9, CXCL10, CXCL11 and IL-6 are measured in sJIA patient, wherein About 37% (20 in 54) patient suffers from MAS in sampling.The relationship of cyclical level Yu disease activity parameter is also had evaluated, Such as the correlation of IFN γ level and CXCL9, CXCL10 and CXCL11 level.In some embodiments, biomarker is Total IFN γ is horizontal, can be used as pharmacodynamic biological marker.

As demonstrated herein, compared with the activity sJIA for not having MAS in sampling, IFN γ in activity MAS With the significant raising of level of 3 kinds of IFN γ associated chemokines CXCL9, CXCL10 and CXCL11.In activity MAS, disease is serious The laboratory parameters of degree such as ferritin, neutrophil leucocyte, blood platelet, alanine aminotransferase and lactic dehydrogenase with IFN γ is significant related to CXCL9, and related to CXCL10 and CXCL11 in lesser degree, and discovery does not have with IL-6 level Correlation.In the activity sJIA patient of not MAS, there is no significant correlation between laboratory parameters and cytokine levels. In activity MAS, IFN γ level and the level of CXCL9 are significant related, related to the level of CXCL10 in lesser degree, and And it is unrelated with the level of CXCL11.

The laboratory parameters of high-caliber IFN γ and CXCL9 and disease severity present in activity MAS patient Significant correlation.IFN γ and CXCL9 are closely related in activity MAS patient.Due to having shown that CXCL9 is only induced by IFN γ Without interferon-induced (see, for example, Groom J.R. and Luster A.D. Immunol Cell Biol by other 2011, Feb;89 (2): 207-15), result of study disclosed herein shows that CXCL9 is the biology mark that IFN γ generates in MAS Will object.

It is provided herein research it is also shown that with the concurrent sJIA of MAS patient in, IFN γ and chemotactic factor (CF) (C-X-C Motif) horizontal the increasing of ligand 9 (CXCL9), CXL10 and CXCL11 (the known 3 kinds of chemotactic factor (CF)s induced by IFN γ), but It is not increased then in the activity sJIA patient of not MAS.In addition, in these patients, IFN γ, CXCL9, CXCL10 and The level of CXCL11 is related to the laboratory parameters of disease severity.

The disclosure, which is provided, to be treated, is prevented using the anti-IFN γ antibody of neutrality or its antigen-binding fragment and/or postpone to move Plant failure, graft rejection and/or diseases associated with inflammation relevant to the graft rejection such as acute shifting of solid organ transplantation obstacle, marrow It plants the breaking-out repelled or development or mitigates the composition and method of relative symptom.The disclosure, which provides, to be received or just It is anti-using neutrality in a series of subject for receiving the graft comprising biomaterial or grafts comprising biomaterial IFN γ antibody or its antigen-binding fragment are treated, are inhibited, the development of the anti-host disease of Delayed grafting object (GvHD) or improve it The composition and method of symptom.The disclosure, which provides, uses the anti-IFN γ antibody of neutrality or its antigen-binding fragment, to extend transplanting Biomaterial survival composition and method.The disclosure, which provides, uses the anti-IFN γ antibody of neutrality or its antigen binding fragment Paraneoplastic cerebellar degeneration, Hemorrhagic fever, sarcoidosis, adult onset still disease or CART-T cell are treated, prevent and/or postponed to section The breaking-out or progress of therapy, or improve the composition and method of relative symptom.

Composition provided herein and method can be used for transplanting any biomaterial, including such as cell, tissue, marrow And/or organ, it as non-limiting examples include heart, kidney, pancreas, liver and/or enteron aisle.In some embodiments, Biomaterial to be transplanted is allogeneic biomaterial.In some embodiments, the biomaterial of transplanting is marrow.One In a little embodiments, the biomaterial of transplanting is candidate stem cell group.In some embodiments, biomaterial to be transplanted is Or it is originated from one or more liver cells.

In composition provided herein and method, NI-0501 is given to the subject for needing it, for treating, preventing And/or Delayed grafting failure, the breaking-out or development or mitigation of graft rejection and/or diseases associated with inflammation relevant to graft rejection Relative symptom.In some embodiments, graft rejection is herein also referred to as graft failure, is acute.In some realities It applies in scheme, graft rejection is hyperacute.

The anti-IFN γ antibody of neutrality of the invention include such as following table 1 A shown in complementary determining region of heavy chain (CDR), Light chain CDR and combinations thereof shown in table 1B.Comprising such as by Chothia et al. 1989, E.A. Kabat et al., The amino acid of the complementary determining region (CDR) of 1991 definition highlights in following underscore and italic text.(referring to Chothia, C, et al., Nature 342:877-883 (1989); Kabat, EA, et al., Sequences of Protein of immunological interest, Fifth Edition, US Department of Health and Human Services, US Government Printing Office (1991))。

Table 1A. is self-bonded and neutralizes the VH CDR sequence of the antibody cloning of IFN γ

Table 1B. is self-bonded and neutralizes the VL CDR sequence of the antibody cloning of IFN γ

Exemplary antibodies of the invention include such as anti-IFN γ antibody described in PCT Publication WO 2006/109191, in Hold hereby by reference to all being combined with it.

Exemplary antibodies of the invention include the antibody for being for example herein referred as NI-0501, in conjunction with people's IFN γ.NI-0501 The heavy chain of antibody, light chain, Weight variable (VH) chain and (VL) chain-ordering that can lighten are as follows, wherein CDR sequence in following VH and It is underlined in VL amino acid sequence:

NI-0501 heavy chain nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTAT ATTACTGTGCGAAAGA TGGTAGCAGTGGCTGGTACGTACCACACTGGTTCGACCCCTGGGGCCAGGGAAC CCTGGTCACCGTCTCCTCAGC CTCCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGC ACCTCTGGGGGCACAGCGGCCCTGGG CTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGT GGAACTCAGGCGCCCTGACCAGCGGCGTGCA CACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTC CCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTT GGGCACCCAGACCTACATCTGCAACGTGAATCAC AAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCCAA ATCTTGTGACAAAACTCACACATGCCCAC CGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTT CCCCCCAAAACCCAAGGACACCCT CATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCA CGAAGACCCTGAGGTCAAG TTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGA GGAGCAGTACAACA GCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGA GTACAAGTG CAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCC CCGA GAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCC TGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTA CAAGA CCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTATAGCAAGCTCACCGTGGACAAG AGCAGGTGGC AGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGC AGAAGAGCCTCTCCC TGTCTCCGGGTAAATAG (SEQ ID NO: 43)

NI-0501 heavy chain amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGSSGWYVPHWFDPWGQGTLVTVSSASTKGPSVFPLAPSSKS TSGGTA ALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH KPSNTKVDKRV EPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVK FNWYVDGVEVHNAKTK PREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPR EPQVYTLPPSREEMTKNQVSL TCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKS LSLSPGK (SEQ ID NO: 44)

0501 light chain nucleic acid sequence of NI:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACTC G CAGCAGTGGCAGCATTGCCAGCAACTATGTGCAGTGGTACCAACAGCGCCCGGGCAGTTCCCCCACCAC TGTCAT CTATGAGGATAACCAGAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAATTCTGC CTCCCTCACCATCTCTGGGCTGAAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATGGCAGCAATCG TTGGATGTTCGGCGGAGGGACCAAGCTGACCGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCC GCCCTCCTCTGAGGAGCTTCAAGCCAACAAGGCCACACTGGTGTGTCTC ATAAGTGACTTCTACCCGGGAGCCGT GACAGTGGCCTGGAAGGCAGATAGCAGCCCCGTCAAGGCGGGAG TGGAGACCACCACACCCTCCAAACAAAGCAA CAACAAGTACGCGGCCAGCAGCTACCTGAGCCTGACGCC TGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCCA GGTCACGCATGAAGGGAGCACCGTGGAGAAGACA GTGGCCCCTACAGAATGTTCATAG (SEQ ID NO: 45)

0501 light-chain amino acid sequence of NI:

NFMLTQPHSVSESPGKTVTISCTRSSGSIASNYVQWYQQRPGSSPTTVIYEDNQRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYDGSNRWMFGGGTKLTVLGQPKAAPSVTLFPPSSEELQANKATLVCL ISDFYP GAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKT VAPTECS (SEQ ID NO: 46)

NI-0501 heavy chain variable amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGSSGWYVPHWFDPWGQGTLVTVSS (SEQ ID NO: 47)

0501 chain variable region amino acid sequence of NI:

NFMLTQPHSVSESPGKTVTISCTRSSGSIASNYVQWYQQRPGSSPTTVIYEDNQRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYDGSNRWMFGGGTKLTVL (SEQ ID NO: 48)

Suitable anti-IFN γ antibody includes antibody described in United States Patent (USP) 7700098, hereby by reference to all being tied with it It closes.Several illustrative antibody include wherein be known as ARC1.2R3P2_A6 (" A6 "), ARC1.2R3P2_B4 (" B4 "), ARC1.2R3P2_B9 (“B9”)、ARC1.2R3P2_C9 (“C9”)、ARC1.2R3P2_C10 (“C10”)、ARC1.2R3P2_ D3 (“D3”)、ARC1.2R3P2_D6 (“D6”)、ARC1.2R3P2_D8(“D8”)、ARC1.2R3P2_E1(“E1”)、 ARC1.2R3P2_F8(“F8”)、ARC1.2R3P2_F9 (“F9”)、ARC1.2R3P2_G7 (“G7”)、ARC1.2R3P2_G9 The antibody of (" G9 ") and ARC1.2R3P2_G10 (" G10 ").

The sequence of these antibody is as follows.

A6 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTAT ATTACTGTGCGAAAGA TGGTAGCAGTGGCTGGTACGTACCACACTGGTTCGACCCCTGGGGCCGGGGCAC CCTGGTCACCGTCTCGAGT (SEQ ID NO: 49)

A6 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGSSGWYVPHWFDPWGRGTLVTVSS (SEQ ID NO: 50)

A6 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACTC G CAGCAGTGGCAGCATTGTCAGCAACTATGTGCAGTGGTACCAACAGCGCCCGGGCAGTGCCCCCACCAC TGTCAT CTATGAGGATAACCGGAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAATACTGC CTCCCTCACCATCTCTGGGCTGGAGGCTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATGGCAGCAATCG TTGGATGTTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 51)

A6 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIVSNYVQWYQQRPGSAPTTVIYEDNRRPSGVPDRFSGSIDSS S NTASLTISGLEAEDEADYYCQSYDGSNRWMFGGGTKLTVLG (SEQ ID NO: 52)

B4 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTAT ATTACTGTGCGAAAGA TCATAGCAGTGGCTGGTACGTAATCTCCGGTATGGACGTCTGGGGCCGAGGGAC AATGGTCACCGTCTCGAGT (SEQ ID NO: 53)

B4 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDHSSGWYVISGMDVWGRGTMVTVSS (SEQ ID NO: 54)

B4 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACCC G CAGCAGTGGCAGCATTGCCAGCAACTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTTCCCCCACCAC TGTGAT CTCTGAGGATAACCAAAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCGTCGACAGCTCC TCCAACTCTGC CTCCCTCACCATTTCTGGACTGAGGACTGAGGACGAGGCTGACTATTACTGTCAGTCTA ATGATTCCGACAATGT GGTTTTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 55)

B4 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIASNYVQWYQQRPGSSPTTVISEDNQRPSGVPDRFSGSVDSS S NSASLTISGLRTEDEADYYCQSNDSDNVVFGGGTKLTVLG (SEQ ID NO: 56)

B9 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TCCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAGGA CCTAACAGTGGGTGGTCCCTGGTACTACTTTGACTACTGGGGCCAAGGAACCCT GGTCACCGTCTCGAGT (SEQ ID NO: 57)

B9 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNPKNTLYLQMNSLRAEDTAVYYCAKDLTVGGPWYYFDYWGQGTLVTVSS (SEQ ID NO: 58)

B9 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACCC G CAGCAGTGGCAGCATTGTCAGCAACTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTGCCCCCACCAC TGTGAT CTTTGACGATGACCAAAGACCCTCTGGGGTCCCTGGTCGGTTCTCTGGCTCCCTCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGGCTGCAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATAGCAGCAATGT GGTATTCGGCGGGGGGACCAAGGTCACCGTCCTAGGT (SEQ ID NO: 59)

B9 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIVSNYVQWYQQRPGSAPTTVIFDDDQRPSGVPGRFSGSLDSS S NSASLTISGLQTEDEADYYCQSYDSSNVVFGGGTKVTVLG (SEQ ID NO: 60)

C9 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAAGA TGGATGGAACGCGCTGGGATGGCTTGAATCCTGGGGCCGGGGCACCCTGGTCAC CGTCTCGAGT (SEQ ID NO: 61)

C9 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGWNALGWLESWGRGTLVTVSS (SEQ ID NO: 62)

C9 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAGGACGATAACCATCTCCTGCACCC G CAGTGGTGGCAGCATTGGCAGCTACTATGTGCAGTGGTACCAGCAGCGCCCGGGCACTGCCCCCACCAC TGTGAT CTATGACGATAAAAAAAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTATTGTCAGTCTT ATGATAGCAACAATCT TGTGGTTTTCGGCGGAGGGACCAAGGTCACCGTCCTAGGT (SEQ ID NO: 63)

C9 VL amino acid sequence:

NFMLTQPHSVSESPGRTITISCTRSGGSIGSYYVQWYQQRPGTAPTTVIYDDKKRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYDSNNLVVFGGGTKVTVLG (SEQ ID NO: 64)

C10 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTAT ATTACTGTGCGAAAGA TGGTAGCAGTGGCTGGTACGTACCACACTGGTTCGACCCCTGGGGCAGGGGGAC AATGGTCACCGTCTCGAGT (SEQ ID NO: 65)

C10 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGSSGWYVPHWFDPWGRGTMVTVSS (SEQ ID NO: 66)

C10 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACCC G CAGCAGTGGCACCATTGCCAGCAACTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTTCCCCCACCAC TGTGAT CTATGAGGATAACCAAAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATAACAGCAATCA TTGGGTGTTCGGCGGAGGGACCAAGGTCACCGTCCTAGGT (SEQ ID NO: 67)

C10 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGTIASNYVQWYQQRPGSSPTTVIYEDNQRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYDNSNHWVFGGGTKVTVLG (SEQ ID NO: 68)

D3 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCAGGGGGGTCCCTGAAACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCAATGCCATGAGTTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AACTCTTACTGGTAGTGGTGGTACCGCATACTACGCAGACTCCGTGGAGGGCCGGTTCAGCATC TCCAGAGACAA TTCCAAGAACACACTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTAT ATTACTGTGCGAAGGG CACGGAACTCGTGGGAGGAGGACTTGACAACTGGGGCCAAGGCACCCTGGTCAC CGTCTCGAGT (SEQ ID NO: 69)

D3 VH amino acid sequence:

EVQLLESGGGLVQPGGSLKLSCAASGFTFSSNAMSWVRQAPGKGLEWVSTLTGSGGTAYYADSVEGRFSI S RDNSKNTLYLQMNSLRAEDTAVYYCAKGTELVGGGLDNWGQGTLVTVSS (SEQ ID NO: 70)

D3 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTCTGTCGGAGTCTCCGGGGAAGACGGTGACGATCTCCTGCACCG G CAGCGGAGGCAGCATTGCCACCAACTATGTGCAGTGGTATCAGCAGCGCCCGGGCAGTGCCCCCACCAC TGTGAT CCATGAGGATAACCAAAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACGGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGCAGCCTGAGGACGAGGCTGATTACTACTGTCAGTCTT ATGATAGTGACAATCA TCATGTGGTATTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 71)

D3 VL amino acid sequence:

NFMLTQPHSLSESPGKTVTISCTGSGGSIATNYVQWYQQRPGSAPTTVIHEDNQRPSGVPDRFSGSIDGS S NSASLTISGLQPEDEADYYCQSYDSDNHHVVFGGGTKLTVLG (SEQ ID NO: 72)

D6 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAAGA TGGATGGAACGCGCTGGGATGGCTTGAATCCTGGGGCAAGGGGACAATGGTCAC CGTCTCGAGT (SEQ ID NO: 73)

D6 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGWNALGWLESWGKGTMVTVSS (SEQ ID NO: 74)

D6 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACCG G CAGCAGTGGCAGCATTGCCAGCAACTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTGCCCCCACCAC TGTGAT CTATGAGGATAACCAAAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATAGCAGCAATCA AGAGGTGGTATTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 75)

D6 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTGSSGSIASNYVQWYQQRPGSAPTTVIYEDNQRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYDSSNQEVVFGGGTKLTVLG (SEQ ID NO: 76)

D8 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTAT ATTACTGTGCGAAAGA TGGTAGCAGTGGCTGGTACGTACCACACTGGTTCGACCCCTGGGGCCAGGGAAC CCTGGTCACCGTCTCGAGT (SEQ ID NO: 77)

D8 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGSSGWYVPHWFDPWGQGTLVTVSS (SEQ ID NO: 78)

D8 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTTACCATCTCCTGCACCC G CAGCAGTGGCAGCATTGTCAGCAACTATGTACAGTGGTACCAGCAGCGCCCGGGCAGTTCCCCCACCAC TGTGAT CTATGAGGATAACCAAAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATAGCAACAATTT TTGGGTGTTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 79)

D8 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIVSNYVQWYQQRPGSSPTTVIYEDNQRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYDSNNFWVFGGGTKLTVLG (SEQ ID NO: 80)

E1 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGTGAAAAG GTCCTTTGATAGTGGTGGGTCCTTTGAGTACTGGGGCCAGGGGACAATGGTCAC CGTCTCGAGT (SEQ ID NO: 81)

E1 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCVKRSFDSGGSFEYWGQGTMVTVSS (SEQ ID NO: 82)

E1 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTCACCATCTCCTGCACCC G CAGCAGTGGCTACATTGCCAGCTCCTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTTCCCCCACCAC TGTAAT CTTTGAGGATGACCGGAGACCCTCTGGGGTCCCTGATCGGTTCTCTGGCTCCATCGACGGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAGGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGATGACACCACTCC CTGGGTGTTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 83)

E1 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGYIASSYVQWYQQRPGSSPTTVIFEDDRRPSGVPDRFSGSIDGS S NSASLTISGLRTEDEADYYCQSYDDTTPWVFGGGTKLTVLG (SEQ ID NO: 84)

F8 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAGAGT CGGCAGCTGGTACCTGGAAGATTTTGATATCTGGGGCCGGGGGACAATGGTCAC CGTCTCGAGT (SEQ ID NO: 85)

F8 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCARVGSWYLEDFDIWGRGTMVTVSS (SEQ ID NO: 86)

F8 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTTACCATCTCCTGCACCC G CAGCAGTGGCAGCATTGCCAGCAACTATGTTCACTGGTATCAGCAGCGCCCGGGCAGTTCACCCACCAC TGTGAT CTATGAGGATAACCGAAGACCCTCTGGGGTCCCTGCTCGGTTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGGAGACTGACGACGAGGCTGACTACTACTGTCAGTCTT CTGATACCACCTATCA TGGAGGTGTGGTATTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 87)

F8 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIASNYVHWYQQRPGSSPTTVIYEDNRRPSGVPARFSGSIDSS S NSASLTISGLETDDEADYYCQSSDTTYHGGVVFGGGTKLTVLG (SEQ ID NO: 88)

F9 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAAGG CGGTAACTACGGTGATTACTTCGACTACTTTGACTACTGGGGCAGAGGGACAAT GGTCACCGTCTCGAGT (SEQ ID NO: 89)

F9 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKGGNYGDYFDYFDYWGRGTMVTVSS (SEQ ID NO: 90)

F9 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCACCC G CAGCAGTGGCAGCATTGCCAGCAATTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTGCCCCCACCAT TGTGAT CTATGAAGATAACCAAAGACCCTCTGGGGTCCCTCATCGGTTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT ATGAGGGGTTCGGCGG AGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 91)

F9 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIASNYVQWYQQRPGSAPTIVIYEDNQRPSGVPHRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYEGFGGGTKLTVLG (SEQ ID NO: 92)

G7 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACTATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAAGA TGGATGGAACGCGCTGGGATGGCTTGAATCCTGGGGCCAGGGGACAATGGTCAC CGTCTCGAGT (SEQ ID NO: 93)

G7 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGWNALGWLESWGQGTMVTVSS (SEQ ID NO: 94)

G7 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACGCTGTGTCGGAGTCTCCGGGGAAGACGGTGACCATTTCCTGCACCG G CAGAAATGGCAACATTGCCAGCAACTATGTGCAGTGGTACCAGCAGCGCCCGGACAGTGCCCCCACCCT TATAAT CTTTGAAGATACCCAAAGACCCTCTGGGGTCCCTACTCGGCTCTCAGGCTCCATCGACACCTCC TCCAATTCTGC CTCCCTCATCATCTCTTCATTGAGGACTGAGGACGAGGCTGATTACTACTGTCAATCTT CTGATTCCAACAGGGT GCTGTTCGGCGGAGGGACCAAGGTCACCGTCCTAGGT (SEQ ID NO: 95)

G7 VL amino acid sequence:

NFMLTQPHAVSESPGKTVTISCTGRNGNIASNYVQWYQQRPDSAPTLIIFEDTQRPSGVPTRLSGSIDTS S NSASLIISSLRTEDEADYYCQSSDSNRVLFGGGTKVTVLG (SEQ ID NO: 96)

G9 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAAGA TTTTTGGGTTATTACGAGTGGGAATGACTACTGGGGGCGGGGGACCACGGTCAC CGTCTCGAGT (SEQ ID NO: 97)

G9 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDFWVITSGNDYWGRGTTVTVSS (SEQ ID NO: 98)

G9 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTGACCATCTCCTGCACCC G CAGCAGTGGCAGCATTGCTAGCAATTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTTCCCCCACCAC TGTGAT CTTTGAAGATAACCGAAGACCCTCTGGGGTCCCTGATCGGTTTTCTGGCTCCATCGACACCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTACTGTCAGTCTT TTGATAGCACCAATCT TGTGGTGTTCGGCGGAGGGACCAAGCTGACCGTCCTAGGT (SEQ ID NO: 99)

G9 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCTRSSGSIASNYVQWYQQRPGSSPTTVIFEDNRRPSGVPDRFSGSIDTS S NSASLTISGLKTEDEADYYCQSFDSTNLVVFGGGTKLTVLG (SEQ ID NO: 100)

G10 VH nucleic acid sequence:

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAG C CTCTGGATTCACCTTTAGCAGCTATGCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTG GGTCTC AGCTATTAGTGGTAGTGGTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCGGTTCACCATC TCCAGAGACAA TTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTGT ATTACTGTGCGAAAGA TGGATGGAACGCGCTGGGATGGCTTGAATCCTGGGGGAAGGGGACCACGGTCAC CGTCTCGAGT (SEQ ID NO: 101)

G10 VH amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTI S RDNSKNTLYLQMNSLRAEDTAVYYCAKDGWNALGWLESWGKGTTVTVSS (SEQ ID NO: 102)

G10 VL nucleic acid sequence:

AATTTTATGCTGACTCAGCCCCACTCTGTGTCGGAGTCTCCGGGGAAGACGGTAACCATCTCCTGCGCCG G CAGCAGTGGCAGCATTGCCAGCAACTATGTGCAGTGGTACCAGCAGCGCCCGGGCAGTGCCCCCACCGC TGTGAT CTATGAGGATAACCAAAGACCCTCTGGGGTCCCTGATCGATTCTCTGGCTCCATCGACAGCTCC TCCAACTCTGC CTCCCTCACCATCTCTGGACTGAAGACTGAGGACGAGGCTGACTACTACTGTCAATCTT ACTCTTACAACAATCA GGTCGTGTTCGGCGGAGGGACCAAGGTCACCGTCCTAGGT (SEQ ID NO: 103)

G10 VL amino acid sequence:

NFMLTQPHSVSESPGKTVTISCAGSSGSIASNYVQWYQQRPGSAPTAVIYEDNQRPSGVPDRFSGSIDSS S NSASLTISGLKTEDEADYYCQSYSYNNQVVFGGGTKVTVLG (SEQ ID NO: 104)

In some embodiments, IFN γ antibody is with the formatting of IgG isotype.In some embodiments, IFN γ antibody with IgG1 isotype formats.

In some embodiments, people and/or machin IFN γ are combined to IFN γ antibody specificity of the invention, wherein Antibody is incorporated into and NI-0501 antibody, A6 antibody, B4 antibody, B9 antibody, C9 antibody, C10 antibody, D3 antibody, D6 antibody, D8 Antibody, E1 antibody, F8 antibody, F9 antibody, G7 antibody, G9 antibody and/or the identical epitope of G10 antibody.

Treatment method

Composition and method can be used for treating with interferon gamma (IFN γ) expression and/or active relevant various obstacles (including Any one of IFN γ expression and/or activity are abnormal).The composition and method of the disclosure can be used for treating Hemophagocytic Lymphohistocysis's disease (HLH).HLH is a kind of rare, serious and threat to life pathologic immune activation disease, Feature is the clinical sign and symptom (fever, splenomegaly, haemocyte reduction, coagulopathy) of extreme inflammation, leads to abnormal exempt from occur The pathology that epidemic disease mediates finally can lead to multiple organ failure and death (Henter JI, Elinder by tissue damage G, Söder O, Hansson M, et al: Hypercytokinemia in familial hemophagocytic lymphohistiocytosis. Blood 1991, 78:2918-2922).HLH includes primary (heredity/familial) HLH and secondary HLH.

Primary HLH is heterogeneous autosomal recessive hereditary diseases, is mainly seen in infancy and infancy, and estimate Europe There is 1/50000 live-born infant illness rate (Janka GE:Familial hemophagocytic in continent lymphohistiocytosis. Eur. J. Pediatr. 1983, 140:221-230).If do not treated, the disease is total It is fatal, median survival interval less than 2 months (Filipovich AH:Hemophagocytic after paresthesia epilepsy lymphohistiocytosis (HLH) and related disorders. Hematology Am Soc Hematol Educ Program 2009:127-131)。

Genetic defect in primary HHL influences to participate in the cytotoxicity of NK cell and/or cytotoxic lymphocyte The gene of approach, cytotoxicity approach are that protein, the cell of the macrophage of elimination activation, coding for perforin synthesis are molten It solves particle maturation, granule exocytosis and discharges (Filipovich, A., K. required for granule exocytosis effect or function McClain, and A. Grom. 2010. Histiocytic disorders: recent insights into pathophysiology and practical guidelines. Biol. Blood Marrow Transplant. 16 (1 Suppl): S82-S89).In the primary HLH patient of about 20-40%, become the cytotoxicity function of the feature of HLH syndrome It can be damaged as since the gene (PRF1) of coding perforin is mutated, PRF1 is a kind of cytolytic proteins of cytotoxicity particle, Its cytolytic key regulator 10 mediated for T cell and natural killer cells.In about 10% patient, which is Caused by the mutation of UNC13D gene, gene coding participates in perforin and discharges the protein into target cell.In addition, some exempt from Epidemic disease deficit syndrome, such as 2 type of Griscelli syndrome (GS-2) and Chediak-Higashi syndrome (CHS) are often in Reveal HLH (Janka GE, Lehmberg K:Hemophagocytic lymphohistiocytosis: pathogenesis and treatment. Hematology Am Soc Hematol Educ Program 2013, 2013:605-611)。

The ondary forms of HLH can occur in infection, during autoimmunity/rheumatic disease process or with malignant tumour phase It closes.Ondary forms show S&S identical with primary HLH, and may be equally serious.

The composition and method of the disclosure can be used for treating secondary HLH.The composition and method of the disclosure can be used for controlling It treats Macrophage Activation Syndrome (MAS).

MAS is a kind of serious, rheumatic disease complication of possible threat to life, thin by T lymphocyte and macrophage The overactivity of born of the same parents and expanding causes.These immunocytes it is uncontrolled expand cause apparent cytokinemia and High inflammatory conditions relevant to fever, haemocyte reduction, hepatosplenomegaly, hepatosis, disturbance of blood coagulation and high-speed rail proteinemia, And multiple organ failure and death (Schulert GS, Grom AA:Pathogenesis of may be developed into macrophage activation syndrome and potential for cytokine- directed therapies. Annu. Rev. Med. 2015, 66:145-159).[00161] due to it strong clinical with HLH and Pathology similitude, MAS are classified as the secondary or acquired form of HLH.In fact, it has been confirmed that, most of MAS suffer from recently The NK of person and perforin functional test are impaired, and a considerable amount of MAS patients PRF1 and UNC13D show polymorphism or Heterozygous mutant (Zhang M, Behrens EM, Atkinson TP, Shakoory B, et al:Genetic defects in cytolysis in macrophage activation syndrome. Curr Rheumatol Rep 2014, 16: 439)。

MAS is most commonly in sJIA patient, and rarer in systemic loupus erythematosus (SLE) patient, but in vasculitis spy It is not also to be described in patients with Kawasaki disease, although more rare.There is apparent MAS (Sawhney in the sJIA patient of about 7-17% S, Woo P, Murray KJ: Macrophage activation syndrome: a potentially fatal complication of rheumatic disorders. Arch. Dis. Child. 2001, 85:421-426; Moradinejad MH, Ziaee V: The incidence of macrophage activation syndrome in Children with rheumatic disorders. Minerva Pediatr. 2011,63:459-466), some cards According to show in the patient with activity systemic disease of up to one third visible subclinical MAS (Behrens EM, Beukelman T, Paessler M, Cron RQ: Occult macrophage activation syndrome in patients with systemic juvenile idiopathic arthritis. J. Rheumatol. 2007, 34: 1133-1138)。

Since MAS may be fatal, diagnosis and immediate treatment intervention are essential for suitably treating disease in time.Institute The MAS death rate of report reaches 20-30%, and is still main source (the Grom AA, Horne of the paediatrics rheumatism death rate A, De Benedetti F: Macrophage activation syndrome in the era of biologic therapy. Nat Rev Rheumatol. 2016 Mar 24. doi:10.1038/nrrheum.2015.179)。

Different set of standard is had proposed for diagnosing the MAS of sJIA patient.Sometimes recommend (to lose mainly for the primary of HLH Pass) the HLH-2004 diagnosis guide of Informal development (Henter J, Horne A, Aric ó M, Egeler RM, et al: HLH-2004: Diagnostic and therapeutic guidelines for hemophagocytic lymphohistiocytosis. Pediatr Blood Cancer 2007, 48:124-131).However, they there are some Limitation, and sJIA patient may be suitable for.For example, the standard such as haemocyte lower than threshold value needed for HLH-2004 subtracts Few and hypofibrinogenemia only just becomes obviously in the later period of MAS, because these patients usually will increase leucocyte and blood A part (the Schulert GS, Grom of platelet number and raising serum fibrinogen level as sJIA inflammatory reaction AA: Pathogenesis of macrophage activation syndrome and potential for cytokine- directed therapies. Annu. Rev. Med. 2015, 66:145-159).The MAS of significant proportion Erythrophagoytosis (Minoia F, Dav ì S, Horne A, Demirkaya E, et may be not present when presenting in patient al: Clinical features, treatment, and outcome of macrophage activation syndrome complicating systemic juvenile idiopathic arthritis: a multinational, multicenter study of 362 patients. Arthritis & rheumatology (Hoboken, N.J.) 2014, 66:3160-3169).In addition, making under the background of MAS without routine evaluations erythrophage With, NK cell activity and sCD25.

Alternative approach is to be applied to the concurrent sJIA of MAS based on tentative diagnosis guide (PDG), for by analyzing one group of MAS What patient obtained compared with one group of patient with sJIA1 burst.

Recently, its differentiation of the HLH-2004 diagnosis guide and tentative diagnosis guide for sJIA correlation MAS is compared The systemic infection of sJIA/MAS and sJIA (in the case where no MAS) and a large amount of patients ability (Dav ì S, Minoia F, Pistorio A, Horne A, et al: Performance of current guidelines for diagnosis of macrophage activation syndrome complicating systemic juvenile idiopathic arthritis. Arthritis & rheumatology (Hoboken, N.J.) 2014, 66:2871- 2880).Although due to its retrospective property, having some limitations property, the research seem to indicate that the preliminary guide of MAS sensitive Reach optimum balance between degree and specificity, and there is best uniform with the diagnosis for the treatment of physician.HLH-2004 covers standard Sensitivity < 30%.However, also it is reported that reach the Proportion of patients variation of each single standard of PDG very greatly, and some face Bed feature (such as CNS dysfunction and bleeding) may come out in the Late manifestations of MAS, so that it is low in the sensitivity of early stage MAS (Lehmberg K, Pink I, Eulenburg C, Beutel K, et al: Differentiating macrophage activation syndrome in systemic juvenile idiopathic arthritis from other forms of hemophagocytic lymphohistiocytosis. The Journal of pediatrics 2013, 162:1245-1251)。

Recently, it develops and demonstrates diagnostic score (HScore) in retrospective group of 312 patients, wherein 162 quilts Be determined as with reactive hemophagocytic syndrome (Fardet L, Galicier L, Lambotte O, Marzac C, Aumont C, Chahwan D, Coppo P, Hejblum G: Development and validation of the HScore, a score for the diagnosis of reactive hemophagocytic syndrome. Arthritis & rheumatology (Hoboken, N.J.) 2014, 66:2613-2620)。

9 variables (3 clinical [i.e. known potential immunosupress, high temperature, organ enlargement], 5 are remained with HScore [i.e. triglycerides, ferritin, serum glutamic oxalacetic transaminase, fibrinogen level and haemocyte are reduced] of a biology and 1 Cytological [i.e. to the erythrophagoytosis feature of bone marrow aspiration liquid]), and suffer from hemophagocytic syndrome a possibility that < In the range of 1% (HScore≤90) to > 99% (HScore >=250).

Before the MAS diagnostic criteria based on verifying reaches final common recognition, the clinical diagnosis of expert physician is still by MAS The key of the challenge distinguished with the illness (such as burst or pyemia sample syndrome of SJIA) for showing overlapping feature.

It there is no the drug ratified for treating MAS at present.In general, high dose glucocorticoid is the first-line treatment of MAS.It is right In the patient for not having reaction to glucocorticoid, it has been suggested that cyclosporin A (CsA) as other treatment (St é phan JL, Koné-Paut I, Galambrun C, Mouy R, Bader-Meunier B, Prieur AM: Reactive haemophagocytic syndrome in children with inflammatory disorders. A retrospective study of 24 patients. Rheumatology (Oxford, England) 2001, 40: 1285-1292)。

As a part developed for treating the HLH-94 therapeutic scheme of pHLH, lost in high dose glucocorticoid treatment The patient lost also considers to give Etoposide.However, the genotoxic potential of the drug is still main problem.Other current lines HLH treatment includes dexamethasone.However, treatment such as Etoposide and/or dexamethasone are bone marrow suppression and/or widely exempt from Epidemic disease inhibits.Currently without the standard care of two wires HLH treatment, and treat the immune suppression that such as alemtuzumab/ATG has extreme Property processed, and think that the survival of these treatments is excessively poor.

Biological products inhibit the effectiveness of IL-1, IL-6R or TNF α approach still unclear in MAS treatment.Although it is reported that It is effective in sporadic cases to inhibit the biological products of these approach, but also has been reported that in the environment of these treatments occur MAS's Patient (Stern A, Riley R, Buckley L:Worsening of macrophage activation syndrome in a patient with adult onset Still's disease after initiation of etanercept therapy. J Clin Rheumatol 2001, 7:252–256; Ramanan AV, Schneider R: Macrophage activation syndrome following initiation of etanercept in a child with systemic onset juvenile rheumatoid arthritis. J. Rheumatol. 2003, 30: 401-403; De Benedetti F, Brunner HI, Ruperto N, Kenwright A, et al: Randomized trial of tocilizumab in systemic juvenile idiopathic arthritis. N. Engl. J. Med. 2012, 367:2385–2395; Ruperto N, Brunner HI, Quartier P, Constantin T, et al: Two randomized trials of canakinumab in systemic Juvenile idiopathic arthritis. N. Engl. J. Med. 2012,367:2396-2406) and to this Responseless patient is treated, shows the complete protection for inhibiting IL-1, IL-6R or TNF α that cannot provide for MAS development, Effectively treatment terminal phase syndrome cannot be provided.

One retrospective multicenter study of large size investigated in total the clinic of the MAS/sJIA of 362 patients, laboratory and Histopathological Characteristics and at present treatment and effect (Minoia F, Dav ì S, Horne A, Demirkaya E, et al: Clinical features, treatment, and outcome of macrophage activation syndrome complicating systemic juvenile idiopathic arthritis: a multinational, multicenter study of 362 patients. Arthritis & rheumatology (Hoboken, N.J.) 2014, 66:3160-3169).In the patient of about half, the background in activity sJIA occurs for MAS During lower or sJIA happens suddenly, wherein 30% occurs in seizure of disease.The patient of one third identify infectious triggering because Element.In 24 patients for reporting infection type, EBV is the most common pathogen (25%).Think in 11 patients's (3.8%) MAS is related to treatment side effect: wherein 8 lifes for being related to targeting IL-6 (N=4), IL-1 (N=3) or TNF α (N=1) approach Object drug.Nearly all patient gives glucocorticoid.Cyclosporin, bio-pharmaceutical and Etoposide give 61% respectively, 15% and 12% patient.

Therefore, effective therapeutic scheme of identification of M AS represents unsatisfied high medical demand field.SJIA more than 50% and MAS patient does not react individual systemic glucocorticoid, or may need relevant to significant disease incidence in high agent Amount is lower to treat for a long time.When patient does not have reaction to glucocorticoid, (for example CsA or pool is not relied on about other treatment Glycosides) validity good evidence-based data.The process of MAS may become rapidly irreversible, so as to cause fatal result.

It is current statistics indicate that, the death rate of the relevant MAS of sJIA is 8%, and the patient of about one third needs ICU to be admitted to hospital. The result of study of the key effect about IFN γ in terms of the disease incidence mechanism shows that IFN γ blocking may represent one recently The new therapy target of kind.

The composition (including NI-0501 composition) and method of the disclosure are better than the existing treatment of primary and secondary HLH Method.

MAS and HLH is characterized by the relevant cell factor storm of lasting activated immune cell and proinflammatory cytokine (the excessive generation of IFN γ, TNF α, IL-1 and IL-6) (Henter JI, Elinder G, S der O, Hansson M, et al: Hypercytokinemia in familial hemophagocytic lymphohistiocytosis. Blood 1991, 78:2918-2922; Imashuku S, Hibi S, Fujiwara F, Todo S: Hyper-interleukin (IL)-6-naemia in haemophagocytic lymphohistiocytosis. Br. J. Haematol. 1996, 93:803-807; Xu X, Tang Y, Song H, Yang S, et al: Diagnostic accuracy of a specific cytokine pattern in hemophagocytic lymphohistiocytosis in children. J. Pediatr. 2012, 160:984-90.e1; Put K, Avau A, Brisse E, Mitera T, et al: Cytokines in systemic juvenile idiopathic arthritis and haemophagocytic lymphohistiocytosis: tipping the balance between interleukin-18 and interferon-γ. Rheumatology (Oxford) 2015).In the past few years, evidence has been had accumulated to support Is there is HLH (Jordan MB, Hildeman D, Kappler J, Marrack P:An animal model in IFN γ of hemophagocytic lymphohistiocytosis (HLH): CD8+ T cells and interferon gamma are essential for the disorder. Blood 2004, 104:735-743; Pachlopnik Schmid J, Ho C, Chrétien F, Lefebvre JM, et al: Neutralization of IFNgamma defeats haemophagocytosis in LCMV-infected perforin- and Rab27a-deficient mice. EMBO Mol Med 2009, 1:112-124; Zoller EE, Lykens JE, Terrell CE, Aliberti J, et al: Hemophagocytosis causes a consumptive anemia of Inflammation. J. Exp. Med. 2011,208:1203-1214) and MAS (Behrens EM, Canna SW, Slade K, Rao S, et al: Repeated TLR9 stimulation results in macrophage activation syndrome-like disease in mice. J. Clin.Invest. 2011, 121:2264- 2277) key effect in terms of the two.

For primary HLH, perforin knockout mice is considered correlation model because these mouse once infected with LCMV, just will appear human diseases all diagnosis and many clinical and laboratory specific characteristic.The HLH sample disease that they occur Dependent on CD8+ T cell and response antigenic stimulus generate IFN γ (Imashuku S, Hibi S, Fujiwara F, Todo S: Hyper-interleukin (IL)-6-naemia in haemophagocytic lymphohistiocytosis. Br. J. Haematol. 1996, 93:803-807).The result shows that when giving anti-IFN In gamma antibodies and when the high circulation level of IFN γ, not only clinical and laboratory abnormalities are restored, but also survival rate also significantly improves. On the contrary, the ablation of many other cell factors on survival without influence (Imashuku S, Hibi S, Fujiwara F, Todo S: Hyper-interleukin (IL)-6-naemia in haemophagocytic lymphohistiocytosis. Br. J. Haematol. 1996, 93:803-807; Xu X, Tang Y, Song H, Yang S, et al: Diagnostic accuracy of a specific cytokine pattern in hemophagocytic lymphohistiocytosis in children. J. Pediatr. 2012, 160:984- 90.e1).Further strengthening importance of the IFN γ in HLH is the circulation IFN γ level in the high concentration of these Finding cases (Henter JI, Elinder G, Söder O, Hansson M, et al: Hypercytokinemia in familial hemophagocytic lymphohistiocytosis. Blood 1991, 78:2918-2922; Xu X, Tang Y, Song H, Yang S, et al: Diagnostic accuracy of a specific cytokine pattern in hemophagocytic lymphohistiocytosis in children. J. Pediatr. 2012, 160:984-90.e1).A series of from 71 patients that HLH diagnosis is monitored to treatment and follow-up, all patients' IFN γ level is above the upper limit (17.3 pg/mL) of normal value, and especially 53.5% patient levels are higher than 1000 pg/ mL.Also it is reported that IFN γ horizontal early stage and rapid increase, and can be from > 5000 pg/ in 48 hours after effectively treatment HLH ML is down to normal value.

Two kinds of secondary HLH animal models are had studied, under the background of NI-0501 development plan to illustrate IFN γ Potential pathogenic effects.Firstly, it is high thin that the activated TLR9 triggering of CpG is given in repetition in the mouse model of the HLH of simulated infection driving Intracellular cytokine mass formed by blood stasis, cause HLH33 clinic (such as weight loss, splenomegaly) and laboratory (such as haemocyte reduce, high-speed rail egg White mass formed by blood stasis) feature.When by giving in anti-IFN γ antibody with IFN γ, the clinic and Laboratory Characteristic of the disease are restored. It is also complete that the neutralization of IFN γ, which is shown in relevant target tissue such as liver and spleen,.It is interesting that anti-IFN γ antibody Give disclose IFN γ amount it is 500-2000 times higher than the IFN γ measured in blood, may preferably reflect the IFN in tissue γ is generated.In both blood and liver after TLR9 stimulation, two kinds of IFN γ induction type chemotactic factor (CF)s (CXCL9 and CXCL10) Up-regulation, and significant correlation is observed between the serum levels of IFN γ and CXCL9 and CXCL10 serum-concentration.IFNγ The blood serum induced CXCL9 and CXCL10 of neutralization and its mRNA level in-site in liver significantly reduce (Buatois V, Chatel L, Cons L, Lory S, et al: IFNγ drives disease in the TLR9-mediated secondary HLH in mice: rationale for a new therapeutic target in secondary HLH, in preparation)。

Secondly, the animal model of the IL-6 transgenic mice of expression high level IL-6 is had studied, because it simulates sJIA The illness of patient, sJIA are most normal rheumatic disease relevant to the ondary forms of HLH.When with Toll-like receptor (TLR) ligand When triggering, observe that lethal increases, inflammatory cytokine generation increases and the overactivity of inflammatory signals pathway.This Outside, these mouse show that blood platelet and neutrophil count decline, sCD25, ferritin and LDH level increase, and are similar to Many features (Strippoli R, Carvello F, the Scianaro R, De generally presented in MAS patient Pasquale L, et al: Amplification of the response to Toll-like receptor ligands by prolonged exposure to interleukin-6 in mice: implication for the pathogenesis of macrophage activation syndrome. Arthritis Rheum. 2012, 64: 1680-1688).In these mouse, when IFN γ with give in anti-IFN γ antibody and when, survival significantly improves and laboratory Parameter reconstruction (Prencipe G et al, the manuscript in preparation).

Have collected similar evidence in an observational study recently, the research with secondary to infection or it is unknown come Source (pHLH excluded by normal cell cytotoxic activity, in known without result in the mutation of pHLH and there is no family history) HLH ondary forms or the MAS occurred under the background of sJIA patient implement.

In 14 patients with secondary HLH (wherein 7 latent infections are identifiable), in activity terminal During phase disease and disease reaction period analysis blood serum sample.Compared with disease reaction, the water of IFN γ, CXCL9 and CXCL10 Put down that significantly higher (IFN γ: 34.7 relative to < 3.5 pg/ml in active stage;CXCL9:33598 is relative to 745 pg/ml; CXCL10:4420 is relative to 132 pg/ml;Intermediate value).Horizontal significant related (p=0.0018) of IFN γ level and CXCL9, And in lesser degree to horizontal related (p=0.014) of CXCL10.The water of IFN γ and chemotactic factor (CF) (especially CXCL9) It is flat significant related to the parameter of disease severity such as neutrophil leucocyte and platelet count, ferritin and ALT, further prop up Hold pathogenic effects and chemotactic factor (CF) potential use as the associated biomarkers of the disease of the IFN γ in secondary HLH (Buatois V, Chatel L, Cons L, Lory S, et al:IFN γ drives disease in the on the way TLR9-mediated secondary HLH in mice: rationale for a new therapeutic target in secondary HLH)。

Similar result of study is had shown that in the patient that MAS occurs for sJIA patient.IFN is measured in 54 sJIA patients The serum-concentration of γ, IFN γ induction type chemotactic factor (CF) (CXCL9, CXCL10, CXCL11) and IL-6, wherein 20 suffer from MAS. In terminal phase MAS patient and sampling with activity sJIA but being on close level without those of MAS patient IL-6.On the contrary, It is significant higher in MAS to recycle IFN γ and Chemokines Levels, especially for CXCL9, median level is than not MAS's (13392 relative to 837 pg/mL by 15 times of activity sJIA patient Gao Yue; p=0.005).

It is worth noting that, only in MAS patient, in CXCL9 horizontal and generally abnormal parameter such as ferritin (p= 0.041), neutrophil leucocyte (p=0.010) and blood platelet (p=0.022) counting, ALT (p=0.044) and LDH (p=0.013) Between be confirmed the existence of significant correlation.IFN γ level it is also related to the laboratory parameters of disease severity (except LDH, It is not up to statistical significance).

In short, these data are the targeted therapy and its in a clinical setting for neutralizing IFN γ as secondary HLH and MAS Research strong theoretical basis is provided.

The composition (including NI-0501 composition) and method of the disclosure are better than the existing therapy of sJIA.For example, the disclosure Composition (including NI-0501 composition) and method can be used for treating the MAS/sHLH of sJIA patient, main purpose is real Existing MAS reaction.

Have benefited from the patients of NI-0501 treatment and for assessing NI-0501 in MAS/sHLH for identifying The theoretical basis of curative effect is based on many factors.Firstly, the preclinical data obtained in animal model relevant to MAS in sJIA Show that IFN γ neutralizes the change for significantly improving survival and restoring laboratory parameters.Secondly, in the observed number of MAS/sHLH patient According to showing that there are high-caliber IFN γs, and importantly, the Chemokine CXCL9 of IFN γ induction, CXCL10 and The horizontal of CXCL11 extremely increases.Third, in MAS/sHLH patient, the concentration and disease parameters such as iron egg of IFN γ and CXCL9 White, platelet count is significant related to transaminase.Next, it is reported that observing that pHLH patient's is good in research previous Tolerance characteristics and no relevant safety problem, wherein the equal tolerance of all infusions given is good, it was demonstrated that volunteer in health The observation that person carries out is not infected caused by the benefited pathogen of IFN γ by neutralizing as known, and suffered from some pHLH The infection that person occurs does not think related with NI-0501 treatment, but with its immune state, disease duration and previous or companion It is related with treating.5th, the preliminary data of previous clinical research shows there is Beneficial Effect to disease parameters, for the treatment of Have apparent action in one day: after giving NI-0501 for the first time, the typical clinical signs and symptom of HLH start to improve rapidly (a few houres internal heat generation, splenomegaly/hepatomegaly in several days)；It can assess in patient with 18 in cut-off, be treated with NI-0501 So that 10 patients are transferred to HSCT.Next, the evidence from PK modeling and analogy method shows that NI-0501's is predictable Pharmacokinetic Characteristics, and realize and maintain the neutralization of IFN γ.Finally, if NI-0501 fails fully to control disease Disease can start conventional therapy (such as CsA) without cleaning the phase immediately.

In short, preclinical and clinical evidence is based on, to the neutrality IFN γ in the MAS/sHLH secondary to rheumatic disease There are very strong theoretical basis, and the preliminary data of pHLH patient shows that NI-0501 has advantageous benefit feature of risk, and shows Write the normalization for improving HLH feature.

Therefore, NI-0501 represents a kind of for treating the wound of the rheumatic disease complication of this serious threat to life New and effective treatment method may limit the side effect of glucocorticoid treatment in long-term high dose.

Give anti-IFN γ antibody

It should be recognized that treatment entity of the invention give by with suitable carrier, excipient and incorporation preparation in other Substance is given together, to provide improved transhipment, transmitting, tolerance etc..It can be found in the formulary known to all pharmacy men Many suitable preparations: Remington ' s Pharmaceutical Sciences (15th ed., Mack Publishing Company, Easton, PA (1975)), the 87th chapter of especially wherein Blaug, Seymour.These preparations include for example Pulvis, paste, ointment, gelling agent, wax, oil, lipid, containing vesica lipid (cation or anion) (such as Lipofectin), DNA conjugate, it is anhydrous absorb cream, oil-in-water and water-in-oil emulsion, carbowax emulsion (various molecular weight it is poly- Ethylene glycol), semi-solid gel and the semi-solid mixtures containing carbowax.Any aforementioned mixture is applicable to treatment of the invention And therapy, condition are that the active constituent in preparation is not inactivated by preparation, and preparation is physiologically compatible and for giving Approach is tolerable.Other information relevant to preparation known to pharmacy man, excipient and carrier also can be found in Baldrick P. ‘‘Pharmaceutical excipient development: the need for preclinical guidance.” Regul. Toxicol Pharmacol. 32(2):210-8 (2000), Wang W. “Lyophilization and development of solid protein pharmaceuticals.” Int. J. Pharm. 203(1-2):1-60 (2000), Charman WN “Lipids, lipophilic drugs, and oral drug delivery- some emerging concepts.” J Pharm Sci. 89(8):967-78 (2000), Powell et al. “Compendium of excipients for parenteral formulations” PDA J Pharm Sci Technol. 52:238-311 (1998) and quotation therein.

Combine the validity for determining treatment with any known method for diagnosing or treating specific immune associated disorders. The mitigation of one or more symptoms of immune associated disorders shows that the antibody assigns clinical benefit.

Antibody (including polyclonal, monoclonal, humanization and human antibody) of the invention can be used as therapeutic agent.This treatment Agent is commonly used in subject or prevention disease relevant to the expression of given target or activation exception or pathology.It will resist Body preparation preferably gives the antibody preparation with high specific and high-affinity to its target antigen to subject, and usually Since it has effect in conjunction with target.Giving antibody can eliminate or inhibit or the signal transduction function of disturb target.It gives Antibody can be eliminated or inhibit or the combination of disturb target and its endogenic ligand naturally combined.

Amount needed for the antibody of the present invention of therapeutically effective amount is usually directed to realization therapeutic purposes.As described above, this can be anti- Binding interactions between body and its target antigen, in some cases, meeting disturb target work.The amount for needing to give this It is outer depending on antibody to the binding affinity of its specific antigen, and additionally depend on that the antibody given gives from it other by The rate exhausted in the free volume of examination person.As non-limiting examples, the treatment of antibody or antibody fragment of the invention is effective The Typical ranges of administration can be about the mg/kg weight of 0.1 mg/kg weight-about 50.Preferred dosage includes 1,3,6,10 mg/kg Weight.Common administration frequency can for example once a day to twice a week in the range of.Treat sustainable 2,3,4,5,6,7, 8,9,10,11,12 or more week.

Antibody or its segment of the invention can be given in the form of Pharmaceutical composition to treat various diseases and obstacle.Such as In Remington:The Science And Practice Of Pharmacy 19th ed. (Alfonso R. Gennaro, et al., editors) Mack Pub. Co., Easton, Pa.: 1995; Drug Absorption Enhancement: Concepts, Possibilities, Limitations, And Trends, Harwood Academic Publishers, Langhorne, Pa., 1994;With Peptide And Protein Drug Delivery (Advances In Parenteral Sciences, Vol. 4), 1991, M. Dekker, NewYork In provide the guideline for preparing principle involved in this composition and Consideration and component selection.

Preparation is such as necessary for the specific indication treated anti-also containing more than one reactive compound IFN γ antagonist, it is therefore preferable to there is complementary activity and the reactive compound that those of do not have an adverse effect each other.Or it is or another Outside, composition may include the drug for enhancing its function, such as cytotoxic agent, cell factor, chemotherapeutant or growth inhibitor. This molecule suitably exists effectively to measure combination to expected purpose.

In one embodiment, reactive compound (such as anti-IFN γ antagonist) is given with combination treatment, i.e., with one kind Or a variety of it can be used for treating pathological condition or the other pharmaceutical composition of obstacle is given.In this context, term " combination " means Drug substantially simultaneously (simultaneously or sequentially) is given.If sequentially given, when starting to give second of compound, two kinds of chemical combination The first in object preferably can still be detected in therapentic part with effective concentration.

For example, combination treatment may include the therapeutic agent co-formulation other with one or more of and/or give jointly The anti-IFN γ antibody of one or more neutralities of the invention: for example one or more cells as described in more detail below because Son and growth factor receptor inhibitors, immunosuppressor, anti-inflammatory agent, metabolic poison, enzyme inhibitor and/or cytotoxic agent or cell are raw Long inhibitor.The therapeutic agent to be administered of relatively low-dose is advantageously used in this combination treatment, thus avoid with it is various single The relevant possible toxicity of therapy or complication.

In some embodiments, drug in addition is immunosuppressor.In some embodiments, immunosuppressor is Cyclosporin A (CsA).In some embodiments, subject has received CsA before giving NI-0501.In some realities It applies in scheme, drug in addition includes at least Etoposide.In some embodiments, subject is before giving NI-0501 Etoposide is received.

In some embodiments, drug in addition is intrathecal methotrexate (MTX) and/or glucocorticoid.In some embodiment party In case, subject has received intrathecal methotrexate (MTX) and/or glucocorticoid before giving NI-0501.

In some embodiments, drug in addition is IV immunoglobulin (IVIG).In some embodiments, IVIG It is given in the subject that immunoglobulin deficiency is recorded as replacement therapy.Subject is recorded immunoglobulin and lacks wherein In weary some embodiments, with the dosage of 0.5 g/kg, every 4 weeks or IVIG is given more frequently to maintain enough IgG water It is flat.

In some embodiments, one or more other drugs are analgesia therapy, blood transfusion product, electrolyte and grape Sugared infusion, antibiotic, antimycotic and antiviral therapy and/or general supportive treatment.

Using antibody fragment, preferably press down with the minimum of the binding structural domain of target protein specific binding Film-making section and/or interference or the minimum of antagonism IFN γ signal transduction inhibit segment.For example, the variable region sequences based on antibody, The peptide molecule for retaining the ability for combining target protein sequence can be designed.This peptide can chemical synthesis and/or pass through recombinant DNA technology produce It is raw.(see, for example, Marasco et al., Proc. Natl. Acad. Sci. USA, 90:7889-7893 (1993)).Preparation is also containing more than one for the necessary reactive compound of specific indication treated, it is therefore preferable to have Those of there is complementary activity and do not have an adverse effect each other reactive compound.Alternatively or in addition, composition may include enhancing it The drug of function, such as cytotoxic agent, cell factor, chemotherapeutant or growth inhibitor.This molecule is suitably to pre- Phase purpose is effectively measured combination and is existed.

Dosage regimen

The present invention further provides for treating, preventing and/or postponing the raised breaking-out of IFN γ or development or mitigate and its phase The dosage regimen of the symptom of pass.Dosage regimen is more variable dose schemes.Dosage is the weight in 1.0-10 mg/kg subject Between combination interferon gamma (IFN γ) antibody.Dosage is given at least the first and second dosage.Second dosage is lower than or height In the first dosage.

For treating more agent variables of primary Hemophagocytic lymphohistocysis disease (HLH) in human experimenter Amount scheme includes the anti-IFN-g monoclonal antibody of induction or the first dosage and treatment or the second dosage.Subject is adult tested Person or pediatric subject.

First dosage is 1.0 or 3.0 mg/kg weight.Second dosage is 3.0,6.0 or 10.0 mg/kg weight.First dose Amount is the antibody of 1.0 mg/kg weight and the second dosage is 3.0,6.0 or 10.0 mg/kg weight.Alternatively, the first dosage is 3.0 The antibody of mg/kg weight and the second dosage are 6.0 or 10.0 mg/kg weight.

First dosage is given once as single dose.Or first dosage be administered more than once in the inductive treatment phase.

Second dosage gives one or more treatment phases.Second dosage gives the first treatment phase.During the first treatment phase, Give the second dosage within every 3 days after first dosage.First treatment phase is for about 1,2,3,4,5,6 or more weeks.Preferably first Treatment phase is 2 weeks.Optionally, second the second treatment phase of dosage is given after the first treatment phase terminates.In the second treatment phase phase Between, the second dosage is given weekly twice.Second treatment phase was for about 1-20 weeks, 2-20 weeks, 3-20 weeks, 4-20 weeks, 5-20 weeks, 6- 20 weeks, 1-10 weeks, 2-10 weeks, 3-10 weeks, 4-10 weeks, 5-10 weeks, 6-10 weeks.Second treatment phase be 1,2,3,4,5,6,7,8,9, 10,11,12 or more week.

In some embodiments, the second dosage increases or decreases during the process of first or second treatment phase.Increase Or the dosage of reduction is known as third dosage.Third dosage can be 1.0,3.0 or 6.0 mg/kg weight.In the first and second treatments Third dosage is given in the remaining time of phase.Alternatively, third dosage gives third treatment phase.In some embodiments, third Dosage is known as maintenance dose and third treatment phase is known as the maintenance phase.

It can for treating the more of secondary Hemophagocytic lymphohistocysis disease (HLH) in mankind pediatric subject Become the anti-IFN-g monoclonal antibody that dosage includes induction or the first dosage and treatment or the second dosage.

First dosage is 6.0 mg/kg weight.Second dosage is 3.0 mg/kg weight.10.0 mg/kg weight.

In some embodiments, the second dosage increases or decreases during the process of first or second treatment phase.Increase Or the dosage of reduction is known as third dosage.Preferably, third dosage is 6.0 mg/kg weight.In the first and second treatment phases Third dosage is given in remaining time.Alternatively, third dosage gives third treatment phase.In some embodiments, third dosage Referred to as maintenance dose and third treatment phase is known as the maintenance phase.

For can in secondary the more of Hemophagocytic lymphohistocysis disease (HLH) of adult humans subject Become the anti-IFN-g monoclonal antibody that dosage includes induction or the first dosage and treatment or the second dosage.

First dosage is 3.0 or 6.0 mg/kg weight.Preferably, the first dosage is 6.0 mg/kg weight.Second dosage For 6.0 or 10.0 mg/kg weight.Preferably, the second dosage is 10.0 mg/kg weight.First dosage is 6.0 mg/kg weight Antibody and the second dosage be 10.0 mg/kg weight.First dosage is given once as single dose.Or first dosage exist The inductive treatment phase is administered more than once.

More variable dose schemes for treating illness in human experimenter include induction or the first dosage and treatment or the The anti-IFN-g monoclonal antibody of two dosage.Subject is Adult human subjects or pediatric subject.The raising of illness and IFNg level It is related.Illness is graft rejection such as solid organ transplantation obstacle or marrow acute transplantation rejection, graft versus host disease(GVH disease), pair swell Tumor cerebellar degeneration, Hemorrhagic fever, sarcoidosis or adult onset still disease.In other embodiments, receiving CAR-T cell therapy Give dosage regimen to subject later.

First dosage is between 1.0-10 mg/kg weight.For example, the first dosage is 1.0,3.0,6.0 or 10 mg/kg The antibody of weight.Second dosage is higher or lower than the first dosage.Second dosage is between 1.0-10 mg/kg weight.For example, the Two dosage are the antibody of 1.0,3.0,6.0 or 10 mg/kg weight.First dosage is 1.0 mg/kg weight and the second dosage is 3.0,6.0 or 10.0 mg/kg weight.Alternatively, the first dosage be the antibody of 3.0 mg/kg weight and the second dosage be 6.0 or 10.0 mg/kg weight.First dosage is the antibody of 6.0 mg/kg weight and the second dosage is 10.0 mg/kg weight.

In some embodiments, the second dosage increases or decreases during the process of first or second treatment phase.Increase Or the dosage of reduction is known as third dosage.Third dosage can be 1.0,3.0,6.0 or 10.0 mg/kg weight.First and second Third dosage is given in the remaining time for the treatment of phase.Alternatively, third dosage gives third treatment phase.In some embodiments, Third dosage is known as maintenance dose and third treatment phase is known as the maintenance phase.

In more variable dose schemes of the invention, can the health status based on such as subject adjust second dose every now and then First and second periods of amount.For example, the time between administration can be adjusted to ± 1,2,3 or 4 day.Preferably ± 2 days.

In more variable dose schemes of the invention, dosage was given with 1,2,3,4,5,6,7,8,9,10,11 or 12 hour. Preferably, dosage was given with 1 hour.For infusion dosage period be based on many factors known in the art, such as the age, Weight, physical condition or the adverse reaction to therapeutic agent.Period for infusion dosage is typically that subject is resistant to most Fast infusion rates (not causing adverse reaction).

Dosage is given as single injection.Antibody is as the monotherapy for treated disease or illness or combines treatment Method is given.For example, giving subject second of drug.Second of drug is such as anti-inflammatory agent and/or immunosuppressor.

Optionally, subject gives dexamethasone immediately before antibody administration.Dexamethasone is at least 10 mg/m²'s Dosage is given.Alternatively, dexamethasone is at least 5 mg/m²Dosage give.

In some embodiments, subject had not carried out the treatment of HLH previously.

Pharmaceutical composition

It is antibody of the invention or soluble chimeric polypeptide (herein also referred to as " reactive compound ") and its derivative, segment, similar Object and homologue can mix in the Pharmaceutical composition being suitable for administering to.This composition generally includes that antibody or soluble chimeric are more Peptide and pharmaceutically acceptable carrier.Terms used herein " pharmaceutically acceptable carrier " intention includes giving phase with medicinal Hold any and all solvents, decentralized medium, coating, antibacterium and antifungal agent, etc. blend absorption delaying agent etc..It is suitable to carry Body is described in Remington ' the s Pharmaceutical Sciences (the canonical reference text of this field) of latest edition In, by reference to being incorporated herein in.The preferred embodiment of this carrier or diluent includes but is not limited to water, salt water, woods Grignard solution, dextrose solution and 5% human serum albumins.Liposome and non-aqueous vehicles such as fixing oil can also be used.This What a little media and reagent were well known in the art for the purposes of active medicinal matter.Unless any conventional media or reagent and activity Compound is incompatible, otherwise considers to use them in the composition.The reactive compound of supplement can also mix in composition.

Pharmaceutical composition of the invention be configured to its expected to give approach compatible.The example for giving approach includes non-enteric Road, for example, intravenously, intradermal, subcutaneous, oral (such as sucking), percutaneous (i.e. local), transmucosal and rectal administration.For non-enteric Road, intradermal or subcutaneous application solution or suspension may include following components: sterile diluent such as water for injection, salt is water-soluble Liquid, fixing oil, polyethylene glycol, glycerol, propylene glycol or other synthetics；Antibacterial agent such as benzyl alcohol or methylparaben； Antioxidant such as ascorbic acid or sodium hydrogensulfite；Chelating agent such as ethylenediamine tetra-acetic acid (EDTA)；Buffer such as acetic acid Salt, citrate or phosphate, and reagent such as sodium chloride or dextrose for adjusting tension.PH usable acid or alkali are such as Hydrochloric acid or sodium hydroxide are adjusted.Parenteral formulation can be encapsulated in ampoule, disposable syringe or by glass or plastics system At multiple dose vials in.

Preferably giving approach is for example to be injected intravenously by injection.Intravenous injection can be quick or slow infusion.Example Such as, infusion is more than about 1,2,3,4,5,6,7,8,9,10,11,12 hour periods.Period for infusion is based on this field Known many factors, such as age, weight, physical condition or the adverse reaction to therapeutic agent.Period one for infusion As for subject tolerance most fast infusion rates (not causing adverse reaction).

Being suitable for the Pharmaceutical composition that uses of injection includes aseptic aqueous solution (when water solubility) or dispersion liquid and for facing When preparation sterile injectable solution or dispersion liquid aseptic powdery.For intravenous administration, suitable carrier include physiological saline, Bacteriostatic water, Cremophor EL (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS) (PBS).It is in love in institute Under condition, composition is necessary for sterile and should be fluid on there is the degree that can be easy to injectivity.It must prepared Stablize under condition of storage, and the contamination of microorganism such as bacterium and fungi must be protecteded from.Carrier can be molten Agent or decentralized medium contain such as water, ethyl alcohol, polyalcohol (such as glycerol, propylene glycol and liquid macrogol) and its close Suitable mixture.Such as by using coating such as lecithin, by keeping required granularity in the case where dispersing agent and leading to It crosses using surfactant, mobility appropriate can be kept.Pass through various antibacteriums and antifungal agent such as parabens, neoprene Alcohol, phenol, ascorbic acid, thimerosal etc. are, it can be achieved that prevent the effect of microorganism.In many cases it is preferred to be to combine It include isotonic agent, such as carbohydrate, polyalcohol such as mannitol, sorbierite, sodium chloride in object.By in the composition comprising delay The substance of absorption, such as aluminum monostearate and gelatin are, it can be achieved that the extension of Injectable composition absorbs.

Sterile injectable solution can be by mixing the desired amount of reactive compound in solvent appropriate, and the latter is as needed It is prepared containing one of ingredient listed above or combination, subsequent filtration sterilization.In general, dispersion liquid is by by active ingredient It is prepared in object incorporation sterile carrier, sterile carrier contains basic dispersion medium and needed for those listed above Other compositions.In the case where being used to prepare the aseptic powdery of sterile injectable solution, preparation method be vacuum drying and it is cold Dry, the powder of generation active constituent is lyophilized, in addition any in addition desired ingredient from its previous sterilefiltered solutions.

Preferred injectable Pharmaceutical composition includes excipient, for example L-Histidine, L-Histidine mono-hydrochloric salts one are hydrated Object, sodium chloride (NaCl), polyoxyethylene sorbitan monoleate.

The pH of injectable Pharmaceutical composition is between about 5.8-6.2.The preferably pH of injectable Pharmaceutical composition is pH 6.0。

In some embodiments, anti-IFN γ antibody such as NI-0501 is prepared (every ml) as follows: 5 mg NI-051, 1.55 mg L-Histidines, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg sodium chloride (NaCl) and 0.05 Mg polyoxyethylene sorbitan monoleate, wherein pH is 6.0 ± 0.2.

In some respects, Pharmaceutical composition is with unit dose packaging.Unit dose packaging is in container.Container be glass or Plastic containers.Container is syringe, bottle, infusion bottle, ampoule or cartridge.Container can accommodate the volume of 1-25 ml.For example, Container can accommodate the volume of 2,5,10 or 20 ml.

The unit dose of full people's anti-interferon gamma (IFN γ) monoclonal antibody (such as NI-501) 5-25 mg/ml it Between.Preferably unit dose is 5 mg/ml or 25 mg/ml.Antibody is in solution (such as water for injection), and every ml contains: 1.55 Mg L-Histidine, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg sodium chloride (NaCl) and the poly- mountain 0.05 mg Pear ester 80.

In some embodiments, the present invention provides a kind of unit-dose container, and having 20 ml concentration is 5 mg/ml Or 25 mg/ml antibody full people's anti-interferon gamma (IFN γ) monoclonal antibody solution, wherein the pH of solution 5.8-6.2 it Between.It is dissolved in antibody in solution, so that solution is clarified, it is colourless and do not precipitate.Preferably, antibody (such as is injected in solution With water) in, every ml contains: 1.55 mg L-Histidines, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg chlorine Change sodium (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate.

In other embodiments, the present invention provides a kind of unit-dose container, has 10 ml or 20 ml concentration are Full people's anti-interferon gamma (IFN γ) monoclonal antibody solution of 25 mg/ml antibody, wherein the pH of solution is between 5.8-6.2. It is dissolved in antibody in solution, so that solution is clarified, it is colourless and do not precipitate.Preferably, antibody is in solution (such as injection Water) in, every ml contains: 1.55 mg L-Histidines, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg chlorinations Sodium (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate.

In other embodiments, the present invention provides a kind of unit-dose container, has 2 ml or 10 ml concentration are 5 Full people's anti-interferon gamma (IFN γ) monoclonal antibody solution of mg/ml antibody, wherein the pH of solution is between 5.8-6.2.Make Antibody is dissolved in solution, so that solution is clarified, it is colourless and do not precipitate.Preferably, antibody is at solution (such as water for injection) In, every ml contains: 1.55 mg L-Histidines, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg sodium chloride (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate.

Orally administered composition generally comprises inert diluent or edible carrier.They can be encapsulated in gelatine capsule or suppress At tablet.For the purpose that oral medication is given, reactive compound and excipient can be blended and with tablet, pastille or capsule Form use.Orally administered composition can also be used fluid carrier preparation to be used as collutory, wherein the compound warp in fluid carrier Mouth application, rinse simultaneously spue or swallow.It may include one of the adhesive and/or adjunct materials of pharmaceutically compatible as composition Point.The compound containing any following component or similarity such as tablet, pill, capsule, pastille: adhesive such as crystallite Cellulose, bassora gum or gelatin；Excipient such as starch or lactose；Disintegrating agent such as alginic acid, Primogel or cornstarch； Lubricant such as magnesium stearate or Sterotes；Glidant such as colloidal silicon dioxide；Sweetener such as sucrose or saccharin；Or it rectifys Taste agent such as peppermint, gaultherolin or orange taste corrigent.

For administration by inhalation, compound is transmitted from pressurizing vessel or distributor in the form of aerosol spray, is held Device or distributor contain suitable stock solution (such as gas such as carbon dioxide) or atomizer.

Whole body, which is given, to be carried out by transmucosal or transcutaneous modalities.For transmucosal or for percutaneous administration of making in the formulation With the bleeding agent for being suitable for barrier to be infiltrated.This bleeding agent is usually known in the art, and including for example for passing through Detergent, bile salt and the fusidic acid derivatives that mucous membrane is given.

Transmucosal, which is given, to be completed by using nasal spray or suppository.In order to for percutaneous administration of reactive compound is matched Ointment commonly known in the art, ointment, gelling agent or cream is made.

The suppository that compound can be used for rectum transmitting (such as contains conventional suppository bases such as cocoa butter and other sweet Grease) or enema,retention form preparation.

In one embodiment, reactive compound is prepared together with carrier, and carrier will protect compound from from internal It quickly eliminates, such as controlled release preparation, including implantation material and microencapsulated delivery systems.Biodegradable biocompatibility can be used Polymer, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoester and polylactic acid.It is used to prepare The method of this preparation is obvious for those skilled in the art.Material can also be from Alza Corporation and Nova Pharmaceuticals, Inc. are commercially available.Liposomal suspensions are (comprising anti-with the monoclonal for viral antigen The liposome of body targeting infection cell) it also is used as pharmaceutically acceptable carrier.These can be according to those skilled in the art The method preparation known, such as described in U.S. Patent No. 4522811.

Be particularly advantageous to prepare with dosage unit form orally or parenterally composition in order to giving and dose uniformity Property.Dosage unit form used herein refers to the physical discrete list being suitable as the single dose of subject to be treated Position, each unit contain the activation for combining and being computed the predetermined amount for generating desired therapeutic effect with required pharmaceutical carrier Close object.The specification of dosage unit form of the invention for individual treatment is by the specific characteristic of reactive compound and to be achieved Particular treatment acts on and mixes limitation intrinsic in the technology of this reactive compound and determines and depend directly on factors above.

Pharmaceutical composition can be included in container, packaging or distributor with being described together with book.

The detection of CXCL9 and other biological marker

Use the level of any one of various standard detection technologies detection CXCL9 and other biological marker.Detection reagent It can be used in test sample giving the presence of object (or its protein fragments).In some embodiments, detection reagent contains Detectable marker.In some embodiments, detection reagent is antibody (or its segment) or probe.In some embodiments In, reagent or probe are labeled.About probe or antibody, term " labeled " intention includes by making detectable substance and visiting Needle or antibody coupling (i.e. physical connection) come direct label probe or antibody and the reagent by directly marking with another kind is anti- It should come indirect labelling probe or antibody.The example of indirect labelling includes detecting primary antibody using the secondary antibody of fluorescent marker and using biotin End mark is carried out to DNA probe, enable it is used the Streptavidin of fluorescent marker to detect.

Term " biological sample " intention includes out of, subject separates tissue, cell and biofluid and subject's body Existing tissue, cell and fluid.It therefore, include a part or group of blood and blood in the use of term " biological sample " Point, including serum, blood plasma or lymph.Body fluid can be the fluid that separates from anywhere in out of subject body, it is therefore preferable to periphery Position, including but not limited to for example blood, blood plasma, serum, synovial fluid, urine, sputum, spinal fluid, cerebrospinal fluid, liquor pleurae, Respiratory tract, the fluid of enteron aisle and urogenital tract, saliva, fluid, ascites, tumour cystic fluid, amniotic fluid and its group in tract It closes.Biological sample further includes the experiment separate section of all aforesaid fluids.Biological sample further includes containing homogeneous solid material Solution or mixture, such as excrement, tissue and biopsy samples.Detection method of the invention can be used for detecting life in vitro and in vivo Analyte mRNA, albumen or genomic DNA in object sample.For example, the ex vivo technique for testing and analyzing object mRNA includes Northern hybridization and in situ hybridization.Ex vivo technique for testing and analyzing object albumen includes enzyme linked immunosorbent assay (ELISA) (ELISA), western blot, immunoprecipitation and immunofluorescence.Ex vivo technique for testing and analyzing object genomic DNA includes Southern hybridization.For carry out the program descriptions of immunoassays in for example " ELISA:Theory and Practice: Methods in Molecular Biology”, Vol. 42, J.R. Crowther (Ed.) Human Press, Totowa, NJ, 1995; “Immunoassay”, E. Diamandis and T. Christopoulus, Academic Press, Inc., San Diego, CA, 1996;" Practice and Theory of Enzyme Immunoassays", P. Tijssen, Elsevier Science Publishers, Amsterdam, 1985.In addition, Vivo techniques for testing and analyzing object albumen include the analysis resistant object protein antibodies that label is introduced into subject's body.For example, Radioactive mark's substance markers antibody can be used, radioactive mark's object can be detected it by standard imaging techniques and be deposited subject is intracorporal And position.

Definition

Unless otherwise defined, the scientific and technical terms being otherwise used in conjunction with the invention should have those skilled in the art Normally understood meaning.Further, unless the context requires otherwise, otherwise singular references should include plural number and plural term It should include odd number.In general, in conjunction with the techniques described herein, cell and tissue culture, molecular biology and protein and widow-or more Nucleotide chemistry and the nomenclature that uses of hybridization those of are well known in the art and commonly use.Standard technique is for recombinant DNA, few core Thuja acid synthesis and tissue cultures and conversion (such as electroporation, liposome transfection).Enzymatic reaction and purification technique are according to manufacturer Specification or usually the realize or progress as described herein such as this field.Aforementioned techniques and program generally according to it is well known that Conventional method and as this specification quote and discuss in the whole text various general and referring more particularly to described in document into Row.See, for example, Sambrooket al. Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989)).In conjunction with this The name that analytical chemistry described in text, synthetic organic chemistry and medicine and pharmaceutical chemistry and its laboratory procedure and technology use Method those of is well known in the art and commonly uses.Standard technique is used for chemical synthesis, chemical analysis, medicine preparation, preparation and transmitting And the treatment of patient.

Terms used herein " dosage " refer to the amount for giving the anti-IFN γ antibody of subject.

Term " more variable doses " include be given various dose of the subject to carry out therapeutic treatment anti-IFN γ it is anti- Body." more variable dose schemes " or " more variable dose therapies " description is given not based on the different time points in entire therapeutic process The treatment schedule of the anti-IFN γ antibody of same amount.In one embodiment, the present invention was described comprising induction period and treatment phase More variable dose treatment methods, wherein it is anti-with the dosage more higher or lower than treatment phase to give anti-FN γ during induction period Body.In another embodiment, the present invention describes more variable dose treatment methods comprising induction period, the first treatment phase With the second treatment phase, wherein giving anti-IFN γ antibody during induction period with the dosage more higher or lower than treatment phase.First During treatment phase can the dosage more higher or lower than the second treatment phase give anti-IFN γ antibody.Preferably, dosage is controlled first The treatment phase is identical with the second treatment phase.

Terms used herein " induction period " or " load phase " refer to including giving subject's IFN γ antibody to reach threshold value The treatment of horizontal a period of time.During induction period, the IFN γ of at least one inductive dose is given with wherein IFN γ The subject of harmful obstacle.

Terms used herein " threshold level " refer to the treatment effective level of IFN γ antibody in subject.By controlling At least one inductive dose is given during the induction period for the treatment of to reach threshold level.Any amount of inductive dose can be given to reach To the threshold level of IFN γ.Once reaching threshold level begins to treatment phase.

Term " inductive dose " or " load dosage " used interchangeably herein refers to the first dosage of anti-IFN γ antibody, It is more greater or lesser compared with maintenance or therapeutic dose.Inductive dose can be single dose or one group of dosage.Inductive dose Commonly used in making intracorporal drug reach steady-state quantity, and it can be used for being rapidly achieved maintenance levels of drugs.Subsequent inductive dose it After give smaller or biggish anti-IFN γ antibody dosage, i.e. therapeutic dose.Inductive dose is given during the induction period of therapy.

Terms used herein " treatment phase " or " maintaining the phase " refer to including giving the anti-IFN γ antibody of subject with retention period The treatment of a period of time of the therapeutic effect of prestige.Therefore treatment phase is opened after induction period once reaching threshold level Begin.There may be more than one treatment phases, that is, shorten or the extended treatment phase is to keep certain threshold value or realize desired clinic Reaction.Treatment phase preferably gives the substance in every 3 days after inductive dose (such as predose).Alternatively, the substance is every Week is given once or twice.

Term " therapeutic dose " or " maintenance dose " are that subject takes to keep or continue the anti-of desired therapeutic effect The amount of IFN γ antibody.Therapeutic dose is given after inductive dose.Therapeutic dose can be single dose or one group of dosage.It controls Dosage is treated to give during the treatment phase of therapy.Therapeutic dose be less than or greater than inductive dose, but and when continuously give when that This is equal.When there are more than one treatment phase, it is also possible to which there are more than one therapeutic doses.

" dosage " or " dosage regimen " includes the therapeutic scheme of the dosage determined based on one group.

Terms used herein " administration " refer to give substance (such as anti-IFN γ antibody) with realize therapeutic purposes (such as Treat IFN γ associated disorders).

Terms used herein " double week dosage regimens ", " double week administrations " and " double weeks are given ", which refer to, gives subject's substance (such as anti-IFN γ antibody) is to realize the time courses of therapeutic purposes (such as treatment IFN γ associated disorders).Double week dosage regimens Intention includes weekly administration scheme.

Term " combination " such as in phrase " combination of the first drug and second of drug " includes giving first jointly Kind drug and second of drug, such as can dissolve or be mixed in identical pharmaceutically acceptable carrier, or successively give The first drug and second of drug are given, or successively gives second of drug and the first drug.Therefore, the present invention includes group The method closed therapeutic treatment and combine Pharmaceutical composition.

Term " adjoint " such as in phrase " adjoint therapeutic treatment " is included in the case where there are second of drugs Under give a kind of drug.Adjoint therapeutic treatment method includes the medicine for wherein giving first, second, third kind or other jointly The method of object.Adjoint therapeutic treatment method further include wherein the first or other drug there are second or other The method given in the case where drug, wherein second or other of drug can for example be given in advance.Adjoint therapeutic treatment Method can gradually be implemented by different participants.For example, a participant can give the first drug of subject and second place ginseng Second of drug of subject can be given with person, and giving step can be at the same time or almost simultaneously or real to be separated by certain time It applies, as long as the first drug (and other drug) gives it there are second of drug (and other drug) Afterwards.Participant and subject can be same entity (such as people).

Terms used herein " combination treatment ", which refer to, gives two or more therapeutic substances, for example, anti-IFN γ antibody and Another drug, such as DMARD or NSAID.Other drugs can while giving anti-IFN γ antibody, before or after give.

Embodiment

In composition provided herein and method, by NI-0501 be configured to be used for be transfused without bacterium concentrate (every mL).One In a little embodiments, NI-0501 is formulated as follows: 5 mg NI-051,1.55 mg L-Histidines, 3.14 mg L-Histidine lists Hydrochloride monohydrate, 7.31 mg sodium chloride (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate, wherein pH is between 5.8-6.2.? In some embodiments, NI-0501 is formulated as follows: 5 mg NI-051,1.55 mg L-Histidines, 3.14 mg L-Histidines Mono-hydrochloric salts monohydrate, 7.31 mg sodium chloride (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate, wherein pH is 6.0.

In composition provided herein and method, NI-0501 is given to the subject for needing it, for treating, preventing And/or the breaking-out or development or the relative symptom of mitigation of delay HLH.In some embodiments, by with 1 mg/kg Predose be transfused through the IV of 1 hour period, NI-0501 is given to the subject for needing it.In certain patients, Such as with under-weight and/or very those of young patients, IV is transfused sustainable 1 hour or more, for example, at least 90 points Clock, at least 2 hours or at least 3 hours or longer time.

In some embodiments, by initial IV infusion after, with the predose of 1 mg/kg through 1 hour when Between section other at least once IV infusion, NI-0501 is given to the subject for needing it.In some embodiments, at least The dosage of primary other IV infusion is higher than the predose of 1 mg/kg.In some embodiments, other IV at least once Infusion dosage is 3 mg/kg.In some embodiments, at least once other IV infusion at least 3 after initial IV infusion It is given.In some embodiments, other IV infusion is given in the time selected from the following at least once: being transfused in initial IV 3 days later, 6 days after initial IV infusion, 9 days after initial IV infusion, 12 days and initial defeated after initial infusion 15 days after note.In some embodiments, at least once other IV infusion 3 days after initial IV infusion, in initial IV It is given within 9 days, 12 days and 15 days after initial infusion after initial infusion 6 days after infusion, after initial IV infusion.

In some embodiments, by initial IV infusion after, with the predose of 1 mg/kg through 1 hour when Between section at least a series of other IV infusions, NI-0501 is given to the subject for needing it, wherein the other IV of the series Infusion includes at least a series of infusions of IV twice a week.In some embodiments, a series of at least IV twice a week infusion with The dosage of predose higher than 1 mg/kg is given.In some embodiments, at least a series of infusions of IV twice a week are with 3 The dosage of mg/kg is given.In some embodiments, at least once other IV infusion at least 3 weeks after initial IV infusion It gives.In some embodiments, other IV infusion is given in the time selected from the following at least once: being transfused it in initial IV 3 weeks afterwards, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, 6 weeks after initial infusion, initial infusion it 7 weeks and 8 weeks after initial infusion afterwards.In some embodiments, other IV is transfused and is transfused it in initial IV at least once 3 weeks afterwards, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, 6 weeks after initial infusion, initial infusion it It gives within 7 weeks and 8 weeks after initial infusion afterwards.

In some embodiments, it is transfused by the IV other at least twice after initial IV infusion by NI-0501 Give the subject for needing it.In some embodiments, the dosage of IV other at least twice infusion is first higher than 1 mg/kg Beginning dosage.In some embodiments, IV other for the first time infusion and second of other IV infusion are given with identical dosage It gives.In some embodiments, IV other for the first time infusion and second of other IV infusion are with the phase higher than predose It is given with dosage.In some embodiments, in first and second other IV infusions at least once with the agent of 3 mg/kg Amount is given.In some embodiments, IV other for the first time infusion is given at least 3 days after initial IV infusion.Some In embodiment, IV infusion other for the first time is given in the time selected from the following: 3 days after initial IV infusion, initial 6 days after IV infusion, 9 days after initial IV infusion, 12 days and 15 days after initial infusion after initial infusion.One In a little embodiments, IV infusion other for the first time 3 days after initial IV infusion, 6 days after initial IV infusion, initial It is given within 9 days, 12 days and 15 days after initial infusion after initial infusion after IV infusion.In some embodiments, Secondary other IV infusion is given in the time selected from the following: 3 weeks after initial IV infusion, 4 weeks after initial IV is transfused, 5 weeks after initial IV infusion, 6 weeks after initial infusion, 7 weeks and 8 weeks after initial infusion after initial infusion.? In some embodiments, second of other IV infusion 3 weeks after initial IV infusion, 4 weeks after initial IV is transfused, first Beginning IV infusion after 5 weeks, 6 weeks after initial infusion, give within 7 weeks and 8 weeks after initial infusion after initial infusion.? In some embodiments, for the first time other IV infusion 3 days after initial IV infusion, 6 days after initial IV infusion, first It is given within 9 days, 12 days and 15 days after initial infusion after initial infusion after beginning IV infusion and IV that second other is defeated Note initial IV infusion after 3 weeks, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, 6 after initial infusion It gives for 7 weeks and 8 weeks after initial infusion after initial infusion in week.

In some embodiments, IV other for the first time infusion and second of other IV infusion are given with different dosage It gives.In some embodiments, IV other for the first time infusion and second of other IV infusion are given with different dosage, In second other IV infusion dosage be higher than IV infusion other for the first time.In some embodiments, other for the first time IV infusion and second of other IV infusion are given with different dosage, wherein second of other IV infusion dosage is higher than first Secondary other IV infusion, and wherein first and second other IV infusion dosages are both higher than predose.In some realities It applies in scheme, being given at least once with the dosage of 3 mg/kg in first and second other IV infusions.In some embodiment party In case, IV infusion other for the first time is given with the dosage of 3 mg/kg to be transfused with second of other IV with the dosage of 6 mg/kg It gives.In some embodiments, IV other for the first time infusion is given at least 3 days after initial IV infusion.In some realities It applies in scheme, IV infusion other for the first time is given in the time selected from the following: 3 days after initial IV infusion, in initial IV 6 days after infusion, 9 days after initial IV infusion, 12 days and 15 days after initial infusion after initial infusion.Some In embodiment, for the first time other IV infusion 3 days after initial IV infusion, 6 days after initial IV infusion, in initial IV 9 days after infusion, give within 12 days and 15 days after initial infusion after initial infusion.In some embodiments, second Secondary other IV infusion is given in the time selected from the following: 3 weeks after initial IV infusion, 4 weeks after initial IV is transfused, 5 weeks after initial IV infusion, 6 weeks after initial infusion, 7 weeks and 8 weeks after initial infusion after initial infusion.One In a little embodiments, second of other IV infusion 3 weeks after initial IV infusion, 4 weeks after initial IV is transfused, initial IV infusion after 5 weeks, 6 weeks after initial infusion, give within 7 weeks and 8 weeks after initial infusion after initial infusion.One In a little embodiments, IV infusion other for the first time 3 days after initial IV infusion, 6 days after initial IV infusion, initial The IV infusion that IV gives for 9 days, 12 days and 15 days after initial infusion after initial infusion after being transfused and second other Initial IV infusion after 3 weeks, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, 6 after initial infusion It gives for 7 weeks and 8 weeks after initial infusion after initial infusion in week.

In some embodiments, IV other for the first time infusion includes the infusion of IV twice a week of at least First Series, It include the infusion of IV twice a week of at least second series with second of other IV infusion.In some embodiments, the first system The infusion of IV twice a week of column and the infusion of IV twice a week of second series are given with the dosage of the predose higher than 1 mg/kg It gives.In some embodiments, the infusion of IV twice a week of First Series is given with the dosage of 3 mg/kg and second series IV infusion is given twice a week with the dosage of 6 mg/kg.In some embodiments, the other IV of First Series is transfused first It is given at least 3 days after beginning IV infusion.In some embodiments, the other IV of First Series is transfused when selected from the following Between give: initial IV infusion after 3 days, initial IV infusion after 6 days, initial IV infusion after 9 days, in initial infusion 12 days later and 15 days after initial infusion.In some embodiments, the other IV of First Series is transfused in initial IV 3 days after infusion, 6 days after initial IV infusion, 9 days after initial IV infusion, 12 days and first after initial infusion Begin to give within 15 days after infusion.In some embodiments, the other IV infusion of second series is given in the time selected from the following Give: initial IV infusion after 3 weeks, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, after initial infusion 6 weeks, 7 weeks and 8 weeks after initial infusion after initial infusion.In some embodiments, the other IV of second series Infusion initial IV infusion after 3 weeks, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, initial infusion it It gives within 6 weeks afterwards, 7 weeks and 8 weeks after initial infusion after initial infusion.In some embodiments, First Series is another Outer IV infusion 3 days after initial IV infusion, 6 days after initial IV infusion, 9 days after initial IV infusion, initial It 12 days and is given within 15 days after initial infusion and the other IV infusion of second series is after initial IV infusion after infusion 3 weeks, initial IV infusion after 4 weeks, initial IV be transfused after 5 weeks, 6 weeks after initial infusion, 7 after initial infusion Week and give within 8 weeks after initial infusion.

In some embodiments, it is transfused within every 3 days after predose, until most 15 days after predose. In some embodiments, it is transfused within every 3 days after predose, until most 15 days after predose, then first Start to be transfused weekly twice at least 15 days after beginning dosage.In some embodiments, any time after predose Point, infusion dosage increase to 3 mg/kg.In some embodiments, after being at least transfused twice with 3 mg/kg, NI-0501 Dosage increase to 6 mg/kg, at most infusion 4 times.

In composition provided herein and method, NI-0501 is given to the subject for needing it, for treating, preventing And/or the breaking-out or development or the relative symptom of mitigation of delay HLH.In some embodiments, by with 1 mg/kg Predose be transfused through the IV of 1 hour period, NI-0501 is given to the subject for needing it.In some embodiments In, it is transfused within every 3 days after predose, until most 15 days after predose.In some embodiments, initial It is transfused within every 3 days after dosage, until then starting at least 15 days after predose every most 15 days after predose Week is transfused twice.In some embodiments, any time point after predose, infusion dosage increase to 3 mg/kg. In some embodiments, after being at least transfused twice with 3 mg/kg, the dosage of NI-0501 increases to 6 mg/kg, at most Infusion 4 times.

In composition provided herein and method, NI-0501 is given to the subject for needing it, for treating, preventing And/or the breaking-out or development or the relative symptom of mitigation of delay HLH.In some embodiments, being transfused by IV will NI-0501 gives the subject for needing it, and dosage is greater than 6 mg/kg.In some embodiments, by after predose IV infusion NI-0501 given need its subject, the second dosage is greater than 6 mg/kg.In some embodiments, second Dosage is at least 10 mg/kg.In some embodiments, the second dosage is 10 mg/kg.In some embodiments, second Dosage is 10 mg/kg, daily to repeat.In some embodiments, the second dosage is 10 mg/kg, is repeated daily for 1 week. In some embodiments, the second dosage is 10 mg/kg, is repeated daily for 2 weeks.In some embodiments, the second dosage For 10 mg/kg, it is repeated more than daily 2 weeks.

In some embodiments, NI-0501 is given and needs its subject, for treat, prevent and/or postpone after The breaking-out or development of hair property HLH mitigate relative symptom.In some embodiments, NI-0501 is given and is needed Its subject, for treating, preventing and/or postponing the breaking-out or development or mitigation of secondary HLH under the background of sJIA Relative symptom.In some embodiments, using NI-0501 as the predose of 6 mg/kg give need it by Examination person.In some embodiments, continue NI-0501 treatment with subsequent NI-0501 dosage.In some embodiments In, continue NI-0501 treatment with the NI-0501 dosage of subsequent every 3 days 3 mg/kg, continues (to be up at least 4 weeks SD27)。

In some embodiments, after realizing desired clinical effectiveness, reduce, stop or shorten NI-0501 treatment. In some embodiments, the evidence (i.e. MAS reaction) based on complete clinical response shortens NI-0501 treatment.

In some embodiments, after 4 weeks, continue NI-0501 treatment and be up to other 4 weeks (being up to SD56), It carries out maintaining as needed until realizing MAS reaction.In some embodiments, after 4 weeks, continue NI-0501 treatment Up to other 4 weeks (being up to SD56), carry out maintaining as needed until realization MAS reaction, can be reduced dosage to 1 mg/kg And it is primary to giving weekly to extend infusion interval.

In composition provided herein and method, NI-0501 is given to the subject for needing it, for treating, preventing And/or the breaking-out or development or the relative symptom of mitigation of delay HLH, wherein subject has given the back of dexamethasone Scape.In some embodiments, subject is the patient's (previously not carrying out treatment to HLH) for not receiving treatment, and Dexamethasone is at least 10 mg/m²Dosage give.In some embodiments, subject receives NI-0501 as two wires HLH treatment, and dexamethasone is with 10 mg/m²-5 mg/m²Dosage in range is given.In some embodiments, subject Receive NI-0501 to treat as two wires HLH, and dexamethasone is at least 5 mg/m²Dosage give.In some embodiments In, subject receives NI-0501 and treats as two wires HLH, and dexamethasone is to be less than 5 mg/m²Dosage give.

In some embodiments, NI-0501 before treatment and/or during and/or after with it is one or more following Other pharmaceutical composition is given, such as (as non-limiting examples) therapeutic agent, anti-inflammatory agent and/or immunosuppressor.Some In embodiment, second of drug is the drug for becoming known for treating HLH.In some embodiments, drug in addition is at least Including Etoposide.In some embodiments, NI-0501 and other drug are configured to single therapy composition, and same When give NI-0501 and other drug.Alternatively, NI-0501 and other drug are separated from each other, such as every kind of drug is configured to Individual therapeutic combination, and NI-0501 and other drug give simultaneously or NI-0501 and other drug are being controlled Different time during treatment scheme is given.For example, NI-0501 gives before giving other drug, NI-0501 is after giving It gives after other drug or NI-0501 and other drug is given in an alternating fashion.As described herein, NI-0501 It is given with other drug with single dose or with multi-dose.

In some embodiments, NI-0501 and other drug are given simultaneously.For example, NI-0501 and other drug It can be prepared using single composition or be given as two or more individual compositions.In some embodiments, NI-0501 It is sequentially given with other drug or different time during therapeutic scheme of NI-0501 and other drug is given.

In some embodiments, drug in addition is immunosuppressor.In some embodiments, immunosuppressor is Cyclosporin A (CsA).In some embodiments, subject has received CsA before giving NI-0501.In some implementations In scheme, drug in addition includes at least Etoposide.In some embodiments, subject before giving NI-0501 Receive Etoposide.

In some embodiments, drug in addition is IV immunoglobulin (IVIG).In some embodiments, IVIG It is given in the subject that immunoglobulin deficiency is recorded as replacement therapy.Subject is recorded immunoglobulin and lacks wherein In weary some embodiments, with the dosage of 0.5 g/kg, every 4 weeks or IVIG is given more frequently to keep enough IgG horizontal.

Composition provided herein and method can be used for treating, prevent and/or Delayed grafting failure, graft rejection and/or The breaking-out or development of diseases associated with inflammation relevant to graft rejection mitigate relative symptom.Composition provided herein It can be used for receiving or receiving graft or a series of transplanting comprising biomaterial comprising biomaterial with method The subject of object, inhibition, the development of the anti-host disease of Delayed grafting object (GvHD) mitigate its symptom.Provided herein group Conjunction object and method can be used for extending the survival of the biomaterial of transplanting.

Composition provided herein and method can be used for transplanting any biomaterial, including such as cell, tissue, marrow And/or at least part of organ or organ, it as non-limiting examples include heart, kidney, pancreas, liver and/or enteron aisle. In some embodiments, biomaterial to be transplanted is allogeneic biomaterial.In some embodiments, the life of transplanting Object material is marrow.In some embodiments, the biomaterial of transplanting is candidate stem cell group.In some embodiments, Biomaterial to be transplanted is or from one or more liver cell.

In composition provided herein and method, by NI-0501 be configured to be used for be transfused without bacterium concentrate (every mL). In some embodiments, NI-0501 is formulated as follows: 5 mg NI-051,1.55 mg L-Histidines, 3.14 mg L- group ammonia Sour mono-hydrochloric salts monohydrate, 7.31 mg sodium chloride (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate, wherein pH 5.8-6.2 it Between.In some embodiments, NI-0501 is formulated as follows: 5 mg NI-051,1.55 mg L-Histidines, 3.14 mg L- Histidine mono-hydrochloric salts monohydrate, 7.31 mg sodium chloride (NaCl) and 0.05 mg polyoxyethylene sorbitan monoleate, wherein pH is 6.0.

In some embodiments, graft rejection is chronic.In some embodiments, by with the first of 1 mg/kg Beginning dosage is transfused through the IV of 1 hour period, and NI-0501 is given to the subject for needing it.In certain patients, example Such as there are under-weight and/or very those of young patients, IV is transfused sustainable 1 hour or more, for example, at least 90 minutes, At least 2 hours or at least 3 hours or longer time.

The disclosure also provides the composition and method that can be used for identifying or improve the patients with obstacle, wherein suffering from Person has individually or combines with one or more other interferon gamma (IFN γ) associated biomarkers raised CXCL9 is horizontal.Particularly, the disclosure provides the composition and method for detecting CXCL9 level, and CXCL9 is to suffer from or suspect The biomarker that IFN γ generates in patient with Hemophagocytic lymphohistocysis disease (HLH).Particularly, originally The open composition and method provided for detecting CXCL9 level, CXCL9 are to suffer from or suspect with secondary Hemophagocytic The biomarker that IFN γ generates in the patient of lymphohistocysis's disease (HLH).In some embodiments, composition It is used to detect CXCL9 level with method, CXCL9 is to suffer from or suspect in the patient with Macrophage Activation Syndrome (MAS) The biomarker that IFN γ generates.In some embodiments, composition and method are horizontal for detecting CXCL9, and CXCL9 is In the biology for suffering from or suspecting IFN γ generation in the case where autoimmune disease or diseases associated with inflammation in the patient with MAS Marker.In some embodiments, composition and method be for detecting CXCL9 level, and CXCL9 is to suffer from suffering from or suspect The biomarker that IFN γ generates in the case where systemic autoimmune disease or diseases associated with inflammation in the patient of MAS.One In a little embodiments, composition and method are used to detect CXCL9 level, and CXCL9 is to suffer from or suspecting in the patient with MAS The biomarker that IFN γ generates in the case where whole body type juvenile idiopathic arthritis (sJIA).In some embodiments, Composition and method are used to detect CXCL9 level, and CXCL9 is to suffer from or suspecting in the patient with MAS in systemic erythema The biomarker that IFN γ generates in the case where lupus (SLE).

Be accredited as with the horizontal raised patient of CXCL9 be confirmed as with drug (such as antibody or other based on polypeptide Therapeutic agent, the therapeutic agent based on peptide, micromolecular inhibitor, therapeutic agent and its derivative based on nucleic acid) treatment suitable candidate One or more bioactivity of person, the interfering effects of drug or antagonism IFN γ such as IFN γ signal transduction, and neutralize IFN γ At least one bioactivity.

Suffering from or suspecting some patients with obstacle, fluid and other biological sample contain individually or with other IFN The raised CXCL9 of γ associated biomarkers (such as CXCL10 and/or CXCL11) combination is horizontal.

CXCL9 and these other biological markers are the indicator that internal IFN γ generates.Therefore, using interference, inhibition, Reduce or antagonism IFN γ signal transduction anti-IFN γ antagonist, such as the anti-IFN γ antibody of neutrality or other be based on polypeptide Therapeutic agent, the therapeutic agent based on peptide, micromolecular inhibitor, therapeutic agent and its derivative based on nucleic acid, block or inhibit IFN γ activity.Therefore, by give show CXCL9 and/or other biological marker the raised patient of expression it is anti- IFN γ antagonist, such as the anti-IFN γ antibody of neutrality or other therapeutic agents based on polypeptide, the therapeutic agent based on peptide, small molecule Inhibitor, therapeutic agent and its derivative based on nucleic acid, composition and method can be used for treating, delay-dependent is expressed in IFN γ And/or activity abnormal (such as raising), proinflammatory cytokine generate it is abnormal and/or combination thereof, be driven by it, it is associated therewith or by The development of its obstacle influenced improves its symptom.By detect individually or with one or more IFN γ associated ligands or The level of the CXCL9 of other biological marker combination, being accredited as may be suitble to the anti-IFN of anti-IFN γ antagonist such as neutrality Patient of the gamma antibodies than the candidate of anti-IFN γ Antybody therapy as those described herein.In some embodiments, individually or The horizontal raised patient of CXCL9 that do not have combined with other IFN γ associated biomarkers still can be used anti-IFN γ antagonist to control Treat, including any anti-IFN γ antibody of neutrality as described herein or other therapeutic agents based on polypeptide, the therapeutic agent based on peptide, Micromolecular inhibitor, therapeutic agent and its derivative based on nucleic acid.

It is increased with CXCL9 level that is independent or being combined with one or more other IFN γ associated biomarkers Patient be confirmed as being treated with one or more anti-IFN γ antagonists (such as the anti-IFN γ antibody of neutrality as described herein) Appropriate candidates.Phrase " expression raising " used herein, which refers to come from, to be suffered from or suspects with primary or secondary In the sample of the patient of HLH or HLH associated disorders or from another control sample, be higher than individually or with it is one or more The expression of the baseline expression level of the CXCL9 of other biomarker combinations.In some embodiments, CXCL9 and/ Or the expression of other biological marker increases as significant horizontal raising.

The level of detect independent or the CXCL9 combined with other one or more IFN γ associated biomarkers, It can be used for improving or being layered patients.In some embodiments, the level detected be used for it is determined that give to Determine the dosage of the anti-IFN γ antagonist of patient.In some embodiments, the level detected is for dividing patients Class or layering.For example, horizontal based on the CXCL9 detected, patient can be classified as having " severe " or advanced MAS, Huo Zhexiang Instead, not serious or rudimentary MAS.

Sample is such as blood or blood constituent, such as serum, blood plasma.In some embodiments, sample is another kind Body fluid, such as (as non-limiting examples) urine, synovial fluid, bronchovesicular liquid, cerebrospinal fluid, BAL fluid (BAL) and/or saliva.In some embodiments, biological sample CSF.In some embodiments, biological sample be from The CSF of HLH patient.

Other than the level of detection IFN γ and/or other IFN γ associated biomarkers, controlled with anti-IFN γ antagonist The sensitivity and specificity of biomarker can also can be improved in the appropriate patient for the treatment of by assessing, for identifying or improving patient Any one of many other biology and clinical parameter of crowd are identified.Alternatively, these other biology and facing Bed parameter can be used alone as being accredited as the patient's of the appropriate candidates with anti-IFN γ antagonist or other suitable therapy treatments Means.As non-limiting examples, these biology and clinical parameter include following any one: ferritin levels, neutral grain Cell count, platelet count, alanine aminotransferase level and/or lactate dehydrogenase levels.

The obstacle useful to the compositions and methods of the invention includes wherein IFN γ expression and/or activity is abnormal (such as rises It is high) any obstacle, especially HLH, including secondary HLH, MAS and/or sJIA.

As non-limiting examples, method and composition provided herein is suitable for diagnosing and/or treating obstacle such as former Hair property and/or secondary HLH obstacle.As non-limiting examples, suitable autoimmunity and/or diseases associated with inflammation include with IFN γ activity and/or the relevant primary of abnormal expression and/or secondary HLH obstacle.

Once patient is accredited as with combining individually or with one or more IFN γ associated biomarkers CXCL9 level increases, then it is then with anti-IFN γ antagonist for treating.For example, anti-IFN γ antagonist is anti-for the anti-IFN γ of neutrality Body or its immunocompetence (such as antigen binding) segment.The suitable anti-IFN γ antibody of neutrality includes as described herein any anti- IFN γ antibody.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include to contain and SYAMS (SEQ ID NO:1 amino acid sequence) at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence can be changed Complementary determining region of heavy chain 1 (VH CDR1) contains the amino acid sequence with AISGSGGSTYYADSVKG (SEQ ID NO:2) The variable heavy chain complementary determining region 2 of at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence (VH CDR2) and containing with DGSSGWYVPHWFDP (SEQ ID NO:3) amino acid sequence at least 90%, 92%, 95%, 96%, the variable heavy chain complementary determining region 3 (VH CDR3) of 97%, 98%, 99% or more identical amino acid sequence, containing with The amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more of TRSSGSIASNYVQ (SEQ ID NO:4) The variable light complementary determining region 1 (VL CDR1) of identical amino acid sequence contains and EDNQRPS (SEQ ID NO:5) The identical amino acid sequence of amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more variable light it is mutual Mend determine area 2 (VL CDR2) and containing with QSYDGSNRWM (SEQ ID NO:6) amino acid sequence at least 90%, 92%, 95%, the variable light complementary determining region 3 (VL CDR3) of 96%, 97%, 98%, 99% or more identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include containing SYAMS (SEQ ID NO: 1) VH CDR1 of amino acid sequence, containing AISGSGGSTYYADSVKG (SEQ ID NO:2) amino acid sequence VH The area CDR2 and the area VH CDR3 of the amino acid sequence containing DGSSGWYVPHWFDP (SEQ ID NO:3) are contained The variable light complementary determining region 1 (VL CDR1) of the amino acid sequence of TRSSGSIASNYVQ (SEQ ID NO:4) contains The VL CDR2 of the amino acid sequence of EDNQRPS (SEQ ID NO:5) and the ammonia for containing QSYDGSNRWM (SEQ ID NO:6) The VL CDR3 of base acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include to contain VH CDR1 sequence, VH The combined heavy chain of CDR2 sequence and VH CDR3 sequence, wherein group is combined into table 1A 3 kinds of heavy CDR sequences (VH shown in uniline CDR1, VH CDR2, VH CDR3) combination.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include to contain VL CDR1 sequence, VL The combined light chain of CDR2 sequence and VL CDR3 sequence, wherein group is combined into table 1B 3 kinds of CDR sequence (VL shown in uniline CDR1, VL CDR2, VL CDR3) combination.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include to contain VH CDR1 sequence, VH (wherein group is combined into table 1A 3 kinds of heavy CDR sequences (VH shown in uniline to the combined heavy chain of CDR2 sequence and VH CDR3 sequence CDR1, VH CDR2, VH CDR3) combination) and combination containing VL CDR1 sequence, VL CDR2 sequence and VL CDR3 sequence Light chain (wherein group is combined into table 1B the group of 3 kinds of CDR sequences (VL CDR1, VL CDR2, VL CDR3) shown in uniline It closes).

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino with SEQ ID NO:47 The heavy chain variable amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% of acid sequence or more identical amino acid sequence Column.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino with SEQ ID NO:48 The light chain variable amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% of acid sequence or more identical amino acid sequence Column.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino with SEQ ID NO:47 The heavy chain variable amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% of acid sequence or more identical amino acid sequence Column and with the light chain variable amino acid sequence at least 90% of the amino acid sequence of SEQ ID NO:48,92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino acid of SEQ ID NO:47 The heavy chain variable amino acid sequence of sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino acid of SEQ ID NO:48 The light chain variable amino acid sequence of sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino acid of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of the heavy chain variable amino acid sequence and SEQ ID NO:48 of sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino with SEQ ID NO:44 The heavy chain amino acid sequence of acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino with SEQ ID NO:46 The light-chain amino acid sequence of acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino with SEQ ID NO:44 The heavy chain amino acid sequence of acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence and With the light-chain amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% of the amino acid sequence of SEQ ID NO:46 or more Mostly identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino acid of SEQ ID NO:44 The heavy chain amino acid sequence of sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino acid of SEQ ID NO:46 The light-chain amino acid sequence of sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include the amino acid of SEQ ID NO:44 The light-chain amino acid sequence of the amino acid sequence of the heavy chain amino acid sequence and SEQ ID NO:46 of sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include and are selected from SEQ ID NO:50, 54,58,62,66,70,74,78,82,86,90,94,98 and 102 heavy chain variable amino acid sequence is at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include and are selected from SEQ ID NOs:52, 56,60,64,68,72,76,80,84,88,92,96,100 and 104 light chain variable amino acid sequence is at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include and are selected from SEQ ID NO:50, 54,58,62,66,70,74,78,82,86,90,94,98 and 102 heavy chain variable amino acid sequence is at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence and be selected from SEQ ID NOs:52,56, 60,64,68,72,76,80,84,88,92,96,100 and 104 light chain variable amino acid sequence at least 90%, 92%, 95%, 96%, 97%, 98%, 99% or more identical amino acid sequence.In some embodiments, anti-IFN γ antibody or Its immunoreactive fragments includes to be selected from SEQ ID NO:50,54,58,62,66,70,74,78,82,86,90, 94,98 and 102 heavy chain variable amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include to be selected from SEQ ID NOs:52, 56,60,64,68,72,76,80,84,88,92,96,100 and 104 light chain variable amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments include to be selected from SEQ ID NO:50, It 54,58,62,66,70,74,78,82,86,90,94,98 and 102 heavy chain variable amino acid sequence and is selected from The light chain of SEQ ID NOs:52,56,60,64,68,72,76,80,84,88,92,96,100 and 104 can Become amino acid sequence.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments are given with therapeutically effective amount.Treatment is effective Amount needed for the antibody of the present invention of amount is usually directed to realization therapeutic purposes.The therapeutic purposes can be between antibody and its target antigen Binding interactions, in some cases, meeting disturb target work.As non-limiting examples, antibody of the invention or The Typical ranges that the treatment of antibody fragment is effectively administered can be about the mg/kg weight of 0.1 mg/kg weight-about 50.Common administration Frequency can for example twice daily to once a week in the range of.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments are in about 2 mg/kg of about 0.5 mg/kg- In the range of, such as in the range of about 0.5 mg/kg- about 1.5 mg/kg and/or about 0.5 mg/kg- about 1.0 mg/kg Predose (i.e. load dosage) is given.In some embodiments, anti-IFN γ antibody or its immunoreactive fragments are with about 1.0 The predose of mg/kg is given.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments are successively with initial loading dose and one Or multiple maintenance doses are given.In some embodiments, one or more maintenance doses are and the basic phase of initial loading dose As dosage.In some embodiments, one or more maintenance doses are the dosage less than initial loading dose.In some realities It applies in scheme, one or more maintenance doses are the dosage greater than initial loading dose.

In some embodiments, one or more maintenance doses contain at least two or multiple dosage, wherein each dimension Holding dosage is identical dosage.In some embodiments, two or more maintenance doses are substantially similar to initial loading dose. In some embodiments, two or more maintenance doses are greater than initial loading dose.In some embodiments, two or more A maintenance dose is less than initial loading dose.

In some embodiments, one or more maintenance doses contain at least two or multiple dosage, wherein each dimension Holding dosage not is identical dosage.In some embodiments, two or more maintenance doses are given with increased dosage.One In a little embodiments, two or more maintenance doses are given with the dosage of reduction.

In some embodiments, one or more maintenance doses contain at least two or multiple dosage, wherein each dimension Dosage is held to give with periodic time interval.In some embodiments, two or more dosage are with increased time interval It gives.In some embodiments, two or more dosage are given with the time interval of reduction.

In some embodiments, anti-IFN γ antibody or its immunoreactive fragments are in about 2 mg/kg of about 0.5 mg/kg- In the range of, such as in the range of about 0.5 mg/kg- about 1.5 mg/kg and/or about 0.5 mg/kg- about 1.0 mg/kg Initial loading dose is given, and is then at least one, such as two or more, 3 or multiple, 4 or multiple or 5 or more A maintenance dose.In some embodiments, anti-IFN γ antibody or its immunoreactive fragments are added with the initial of about 1.0 mg/kg Carrier amount is given, and is then at least one, such as two or more, 3 or multiple, 4 or multiple or 5 or multiple maintenances Dosage.

Pharmaceutical composition of the invention may include anti-IFN γ antibody and carrier of the invention.These Pharmaceutical compositions can wrap It is contained in kit such as diagnostic kit.

The present invention also provides the kits for implementing any method provided herein.For example, in some embodiments, Kit include individually or the CXCL9 specific detection agents that are combined with one or more IFN γ associated biomarkers and Method for detecting detection reagent.

The present invention will further describe in the examples below, these embodiments do not limit this hair described in claim Bright range.

Embodiment

The biomarker that 1. CXCL9 level of embodiment is generated as IFN γ in Macrophage Activation Syndrome (MAS)

Research presented herein be intended to assess the serum levels of IFN γ and 3 kinds of IFN γ associated chemokines with itself and with Correlation between the laboratory parameters of the disease activity of activity MAS patient, with aborning potential life in Searching I FN γ body Object marker.

IFN γ, CXCL9, CXCL10, CXCL11 and IL- are measured using Luminex multiplexing measuring method in sJIA patient 6 cyclical level, 20 patients in 54 suffer from MAS in sampling.It also has evaluated these cyclical levels and disease activity is joined The correlation of several relationship and IFN γ level and CXCL9, CXCL10 and CXCL11 level.

Compared with the activity sJIA for not having MAS in sampling, IFN γ and 3 kinds of IFN γ correlations become in activity MAS Change the significant raising (all p value < 0.005) of level of the factor (CXCL9, CXCL10 and CXCL11).In activity MAS, disease is tight Weight degree laboratory parameters (ferritin, neutrophil leucocyte, blood platelet, alanine aminotransferase and lactic dehydrogenase) with IFN γ is significant related to CXCL9, and related to CXCL10 and CXCL11 in lesser degree, and discovery does not have with IL-6 level Correlation.In the activity sJIA patient of not MAS, there is no significant correlation between laboratory parameters and cytokine levels, As shown in the following Table 7.In activity MAS, significant related (r=0.69 of level of IFN γ level and CXCL9; r2=0.47; p =0.001), (r=0.53 related to the level of CXCL10 in lesser degree; r2=0.28;P=0.015), and and CXCL11 Unrelated (the r=- 0.04 of level; p=0.886).

The laboratory parameters of the disease activity of 7. MAS patient of table and activity sJIA patient and IFN γ, CXCL9, The correlation of CXCL10, CXCL11 and IL-6.

N=neutrophil count；PLT=platelet count；ALT=alanine aminotransferase；¹=median (IQR)；r* = Spearman r

The laboratory parameters of the high-caliber IFN γ present in activity MAS patient and CXCL9 and disease severity are significant It is related.It is closely related in activity MAS patient IFN γ and CXCL9.It is only induced by IFN γ without by it since CXCL9 has been displayed He is interferon-induced (see, for example, Groom J.R. and Luster A.D. Immunol Cell Biol 2011, Feb; 89 (2): 207-15), these results of study show that CXCL9 is the biomarker that IFN γ generates in MAS.

The CXCL9 and IFN γ water of 2. primary Hemophagocytic lymphohistocysis disease (HLH) patient of embodiment Flat correlation

Research presented herein is received from the primary HLH patient for giving NI-0501 antibody and in sympathizing with use The ongoing 2 phase Primary Study of the patient of NI-0501 antibody.

As shown in Figure 1, sympathizing in the sample obtained using patient from 6 primary HLH patients and 3, pass through respectively Luminex and Meso Scale Discovery (MSD) technology measures the serum levels of CXCL9 and IFN γ.In CXCL9 and Correlation research is carried out between total IFN γ concentration.Count and examine using Spearman obtaining p value.

As shown in Fig. 2, sympathizing in the sample obtained using patient from 6 primary HLH patients and 3, pass through respectively Luminex and MSD technology measures serum levels before the administration of change of serum C XCL9 and IFN γ.CXCL9 and total IFN γ concentration it Between carry out correlation research.Count and examine using Spearman obtaining p value.

The CXCL9 and IFN γ water of 3. secondary Hemophagocytic lymphohistocysis disease (HLH) patient of embodiment Flat correlation

Research presented herein is received from the secondary HLH patient for giving NI-0501 antibody and in sympathizing with use The observational study of the patient of NI-0501 antibody.

Particularly, these are with whole body type juvenile idiopathic arthritis (sJIA) and macrophage activation synthesis occur Levy the patient of (a kind of form of MAS, secondary HLH).For these patients, in CXCL9 or IFN γ and disease parameters such as iron There is also correlations between albumen, platelet count (PLT), neutrophil count (Neu) and alanine aminotransferase (ALT) Property.

As shown in figs.3 a and 3b, using Luminex technology by multiplex measuring method measure sample CXCL9 and The serum levels of IFN γ, sample from 19 with secondary to sJIA MAS patient and 24 sampling when suffer from activity sJIA Patient obtain.Correlation research is carried out between CXCL9 and total IFN γ concentration.Count and is examined using Spearman Obtain p value.

As shown in Fig. 4 A-1,4A-2,4B-1,4B-2,4C-1,4C-2,4D-1 and 4D-2, passed through using Luminex technology Multiplexing measuring method is measured with the MAS patient secondary to sJIA and the patient with activity sJIA in sampling The serum levels of CXCL9 and IFN γ.IFN γ or CXCL9 level and ferritin, platelet count, neutrophil count or Correlation research is carried out between ALT (alanine aminotransferase).Count and examine using Spearman obtaining p value.

The CXCL9 and IFN γ of 4. severe Hemophagocytic lymphohistocysis disease (HLH) patient of embodiment is horizontal Correlation

Research presented herein is from the patient for receiving NI-0501 antibody in sympathizing with use.Patient shows NLRC4 phase The symptom of related disorders and severe Hemophagocytic lymphohistocysis disease (HLH).The mutation of NLRC4 gene has been reported recently Lead to recurrent Macrophage Activation Syndrome and increases the generation of IL-18 (its known induction IFN γ).

Patient in this research has the feature that 20 ages in days are fallen ill, with fever, fash, significant hepatosplenomegaly, whole blood Leukopenia, hypofibrinogenemia, hypertriglyceridemia, significant ferritin and sCD25 increase.It is followed by multiple organ Failure needs ICU to be admitted to hospital.HLH is diagnosed based on 6 in 8 HLH-2004 standards.Cause primary HLH gene (PRF1, UNC13D, STXBP2, STX11, RAB27A, XIAP) and functional test (perforin expression, threshing and cytotoxicity) be yin Property.High dose i.v. glucocorticoid and i.v. cyclosporin A gradually improve ordinary circumstance and laboratory abnormalities.It is (white by infecting Color candida albicans (Candida Albicans) and Klebsiella Pneumoniae septicemia (Klebsiella Pneumoniae sepsis)) The HLH of triggering is reactivated, and ordinary circumstance deteriorates rapidly, and new ICU is needed to be admitted to hospital.Due to immune impairment by There are Active infections by examination person, do not consider to be carried out with Etoposide and/or ATG treatment.

Record the high serum of the serum levels of measurable IFN γ and the Chemokine CXCL9 of IFN γ induction and CXCL10 Horizontal and IL-18 serum levels significantly increase (table 8).

When table 8. starts NI-0501 treatment and the IFN γ during being treated with NI-0501, IFN γ associated chemokine and The level of IL-18

Start to be used with the sympathy of NI-0501 under the background of dexamethasone (13.6 mg/m2) and i.v. injection cyclosporin A Treatment.According to pharmacokinetics, NI-0501 is given within every 3 days, and is then given within every 7 days primary.It is anti-that infusion is not observed It answers.NI0501 tolerance is good.HLH Clinical symptoms and laboratory abnormalities gradually improve.Activity persistent infection is quickly removed. After treatment 5 months, patient is kept in order.Patient continues to receive oral cyclosporin A (6 mg/kg) and prednisone (0.3 mg/kg is equal to 0.9 m/m²Dexamethasone).All HLH parameters normalization.

Subject still has unaccountable inflammatory episode,NLRC4Analysis shows that go out from the beginning missense mutation (T337N).Serum IL-18 raising is recorded, it was confirmed thatNLRC4The correlation of mutation.Pass through the high level compound with NI-0501 IFN γ confirms that the height of IFN γ generates.IFN γ is fully neutralized, such as undetectable IFN γ induction type chemotactic factor (CF) water Shown in flat (Fig. 5 and table 8).The cyclical level of IL-18 persistently increases.

Therefore, the study show that, with severe intractable HLH (due toNLRC4Mutation) patient hindered with NI-0501 Disconnected IFN γ tolerance is good, without safety problem, enables control over all HLH features, so that glucocorticoid is rapidly gradually Decrement, and it is related to the recession of activity persistent infection.

Embodiment 5. treats the targeted approach of Hemophagocytic lymphohistocysis disease (HLH) with NI-0501

The 2 phase Primary Studies of research presented herein from the children with primary HLH.Primary HLH (pHLH) is one The rare immunomodulatory disorders of kind, are fatal always if not treating.It is activated by pathologic immune and is driven, and causes to occur Fever, splenomegaly, haemocyte reduces and coagulopathy, this may cause multiple organ failure and death.It is based on anti-come anti-IFN γ of using by oneself The primary of body treatment and the mouse model of secondary HLH (sHLH) and the data of the observational study in HLH patient, IFN γ Height generate and be considered that the key factor of the disease occurs in driving.Immunochemotherapy (scheme for being mainly based upon Etoposide), It is at present control HLH and the unique pharmacology for making treatment patients' property allogeneic hematopoietic stem cell transplantation (allo- HSCT) Method.Although attempting to further strengthen therapeutic scheme recently, the death rate and disease incidence are still very high, and partly cause is and drug Relevant toxicity causes.

As described above, NI-0501 be a kind of complete anti-IFN γ mAb of people's high-affinity, in conjunction with and neutralize people's IFN γ, be Control HLH provides method that is a kind of new and having targeting.

Method: the 2 phases research an of open label has been carried out in US and European, has made a definite diagnosis or doubting to assess NI-0501 Like the safety and curative effect of the children with pHLH.NI-0501 was given with the predose of 1 mg/kg every 3 days once, initial Background dexamethasone 5-10 mg/m²Under, instruct possible dosage to increase according to the PK data and/or clinical response of every patient. Duration for the treatment of is in 4-8 weeks range.It has rated the ability for turning to allo-HSCT, correlation HLH disease parameters and deposits for 8 weeks It is living.

Study population: having recruited and amounted to 13 patients: 8F/5M, and the median age 1.0 years old (range was at -13 years old 2.5 months). It is receiving conventional therapy and is reactivating, after not obtaining satisfied reaction or not tolerating to treatment, 12 patients are received NI-0501 is as second line treatment.One patient receives NI-0501 as first-line treatment.9 patients carry known HLH heredity Defect (3 FHL2,2 FHL3,2 GS-2,1 XLP1,1 XLP2).Most patients are in the severe end of HLH spectrum, general Under damage, the significant toxicity treated from previous HLH is carried.Patient's ferritin 12/13 increases and in 8 patients SCD25 is increased, and haemocyte is presented in 10 patients and reduces, 8 presentation splenomegaly, 9 presentation hypofibrinogenemias and height Triglyceride.Hepatic disorder and CNS involvement is presented in 7 and 3 patients respectively.

As a result: in general, NI-0501 treatment significantly improves the parameter (Fig. 6) of HLH disease activity, and 13 trouble There are 9 to realize satisfied reaction in person.6 patients have continued HSCT.Two patient care plans with good HLH control Continue HSCT after the suitable donor of determination.A patient (it realizes disease control with a line NI-0501), in view of There is no pathogenic HLH gene mutation, not yet plan HSCT.

There are 11 survivals at 8 weeks in 13 patients.Subside in 2 appreciable patient CNS S&Ss.In NI- During first 4 weeks of 0501 treatment, 50% patient can reduce by 50% or more dexamethasone dosage.

A kind of evaluation of biomarker (especially CXCL9, known chemotactic factor (CF) accurately induced by IFN γ), not only Make it possible to confirm that complete IFN γ neutralizes, and seemingly diagnose HLH relevant to the generation of IFN γ new parameter (Fig. 7 A and 7B)。

NI-0501 tolerance is good, and does not find safety problem.It does not report in known IFN γ and advantageous infection, And there is no infection in the patient for not receiving previous chemotherapy.7 patients report at least an example SAE, are commented by DMC Valence is unrelated to give with NI-0501.Accident (such as the marrow of " missing the target " effect for being attributed to NI-0501 is not observed Toxicity, hemodynamic Effects).

Conclusion: NI-0501 targeting neutralizes IFN γ and provides a kind of method of innovation and potential low toxicity for HLH treatment.It should Research the result shows that, do not obtaining satisfied reaction to conventional therapy or showing the primary HLH patient not tolerated to it, NI-0501 is a kind of safely and effectively therapeutic choice.In addition, being treated with NI-0501 relevant to the scheme based on Etoposide Any typical short-term or long term toxicity is unrelated.It is in progress as the first-line treatment of pHLH patient to evaluate NI-0501, it is contemplated that can Realize similar significant clinical benefit.

The cyclical level of 6. interferon gamma of embodiment and interferon-induced chemotactic factor (CF), which increases to become, suffers from macrophage The feature of the patient of activation syndrome concurrent system JIA

Interferon gamma (IFN γ) is that the crucial of the mouse model of primary Hemophagocytic lymphohistocysis disease (HLH) is situated between Matter.In view of the similitude between primary and secondary HLH (sec-HLH) (including Macrophage Activation Syndrome (MAS)), Analyze whole body type juvenile idiopathic arthritis (sJIA) and the IFN γ level and its bioactivity of MAS patient.

In research provided herein, Luminex multiplexing measuring method is for evaluating sec-HLH patient (n=11) and its In 20 IL-1 β of the sJIA patients (n=54) with MAS in sampling, IL-6, IFN γ and IFN induction and/or IFN phase Close the serum levels of Chemokine CXCL9, CXCL10 and CXCL11.Wherein by stimulating TLR4 to induce MAS feature with LPS IL-6 transgene mouse model has rated expression (CXCL9 and CXCL10 in liver and spleen of the chemotactic factor (CF) of IFN γ induction MRNA level in-site) and its correlation with serum ferritin level.

If shown in further detail below, the cyclical level of IFN γ and the chemotactic factor (CF) of IFN induction is significantly risen during MAS Height, herein also referred to as activity MAS and sec-HLH.Compared with the activity sJIA patient of not MAS, the IFN of MAS patient The level of the chemotactic factor (CF) of γ and IFN induction is significant higher.In later group, IFN γ and IFN γ induction chemotactic factor (CF) with face Bed inactivity sJIA patient is suitable.During MAS, become the laboratory abnormalities of the feature of the syndrome, including ferritin and Alanine transaminase level and neutrophil leucocyte and platelet count are significant related to the level of IFN γ and CXCL9.In MAS In mouse model, the serum levels of ferritin are significant related to the mRNA level in-site of the CXCL9 in liver and spleen.

Therefore, presented below studies have shown that high-caliber IFN γ and IFN induction chemotactic factor (CF) and its (especially CXCL9) show that IFN γ plays a crucial role in MAS with the correlation of the severity of the laboratory abnormalities of MAS.Interferon gamma Increasing with the cyclical level of interferon-induced chemotactic factor (CF) becomes Macrophage Activation Syndrome concurrent system JIA patient's Feature.

Materials and methods: patient and sample.In 3 paediatrics rheumatism center (the Ospedale Pediatrico Bambino Ges ù in Rome, the Istituto Giannina Gaslini in Genoa and Cincinnati children's hospital Medical centre (the Cincinnati Children ' s Hospital Medical Centre)), from being with or without MAS's SJIA patient collects peripheral blood sample.To 54 sJIA patient's (age of onset for meeting ILAR whole body type arthritis classification standard 7.9 years old, interquartile-range IQR 4.6-13.6 years old, women 48%) studied (Petty, R.E., et al.,International League of Associations for Rheumatology classification of juvenile idiopathic arthritis: second revision, Edmonton, 2001. J Rheumatol, 2004. 31(2): p. 390-2).For 20 SJIA patients, sample is collected during activity terminal phase MAS breaking-out, by 3 The treating physician of each in a center diagnoses.Posterior analysis shows that having 17 (85%) to meet in this 20 breaking-outs newly mentions MAS classification standard (Minoia F, Dav ì S, Bovis F, et al. Development of new out classification criteria for macrophage activation syndrome complicating systemic juvenile idiopathic arthritis. Pediatric Rheumatology 2014, 12(Suppl 1):O1).28 activity SJIA patients without MAS evidence can get sample.According to Wallace standard, clinical inactive Property disease during, can be obtained from 35 sJIA patients's (being with or without MAS in its history of disease) 35 samples (Wallace, C.A., et al., Preliminary criteria for clinical remission for select categories of juvenile idiopathic arthritis. J Rheumatol, 2004. 31(11): p. 2290-4)。

Increase due to having been displayed in sec-HLH patient's (eliminating rheumatic disease) IFN γ, also from seeing Ospedale Pediatrico Bambino Ges ù 11 sec-HLH patients (age of onset 8.6 years old, interquartile-range IQR 4.1- 12.9 years old, women 36%) sample is collected, and it is used as positive control.All sec-HLH patients meet 2004-HLH diagnosis guide (Henter, J.I., et al., HLH-2004: Diagnostic and therapeutic guidelines for hemophagocytic lymphohistiocytosis. Pediatr Blood Cancer, 2007. 48(2): p. 124-31): 6 patients meet 5 standards and 5 patients meet 4 standards.It should be noted that there is no and be with U/ml The sCD25 of unit is horizontal, because not testing routinely in the mechanism for recruiting these patients.Based on no family history, known Cause not have in the gene of HLH pathogenic mutation and there are normal function research (including NK activity, perforin expression and CD107 Threshing), eliminate the diagnosis of primary HLH.All 11 sec-HLH patients respectively contribute to one during active disease The sample of acquisition.

Clinic and Laboratory Characteristic when all patients are about diagnosis and sampling are collected by the researcher at each center In the network data base of concentration.In the 20 MAS patients sampled during active disease, 6 sampling when do not receive it is any Treatment, and remaining 14 patient has received one of MAS specific treatment, including glucocorticoid pulse, cyclosporin A, Anakinra or cyclophosphamide.There are 6 not yet to connect in patient in 11 active diseases with sec-HLH in sampling By specific treatment, and remaining 5 patient has received at least one of above-mentioned treatment.Ospedale Pediatrico The Ethics Committee of Bambino Ges ù has approved this research.Have collected the written consent form of all participants.

Cell factor quantifies: by Luminex multiplex pearl technology analyze IL-6, IL-1 β, IFN γ, The level of CXCL9, CXCL10 and CXCL11.Reagent is purchased from Millipore and all reagents with Milliplex^®MAP reagent Box provides.Reagent is prepared according to the scheme of manufacturer.25 holes μ l/ are added in duplicate in 96 orifice plate of Milliplex MAP Standard items, blank and quality examination sample, then be added 25 μ l serum matrix.25 μ l measurement is added into each sample well 25 μ l samples are then added in buffer.Two parts of sample or triplicate addition, the volume available depending on sample.? Luminex 200^®The plate is measured in system (Luminex Corp.).Use 3.1 (Luminex of x PONENT software version Corp. initial data) is obtained, and analyzes number using Milliplex Analyst software version 3.5.5.0 (Millipore) According to.Then with the customized macros (NI-Sc-ESM-MAC-012-v01 and Sc-ESM-MAC-013-v01) analyzed for Luminex into One step analyzes the initial data obtained with Milliplex Analyst software.

Zoopery.The generation and phenotype of IL-6 transgenic mice is described previously, and is lured by giving TLR ligand The MAS sample syndrome led feature (Strippoli, R., et al.,Amplification of the response to Toll-like receptor ligands by prolonged exposure to interleukin-6 in mice: implication for the pathogenesis of macrophage activation syndrome. Arthritis Rheum, 2012. 64(5): p. 1680-8).Mouse is maintained under conditions of no-special pathogen and according to national policy It is handled.Research approach is ratified through the local ethics committee.It is all to test the mouse progress for being based on 10-14 week old.With list Mouse (LPS, e. coli serotype 055:B5 are given in the lipopolysaccharides peritonaeum of 5 μ g/g weight of dosage; Sigma- Aldrich).Mouse is put to death after 30 hours.It is mentioned using Trizol (Life technologies) from spleen and liver organization Take total serum IgE.CDNA is obtained using Superscript Vilo kit (Invitrogen).Use TaqMan Universal PCR Master Mix (Applied Biosystems) and mouseCxcl9WithCxcl10 Determination of gene expression (Applied Biosystems real-time PCR measurement) is carried out.Use mouseHprt (Applied Biosystems) normalized gene expresses number According to.Tables of data is shown to employ 2^-ΔctThe arbitrary unit (AU) of method measurement.According to the manufacturer's instructions, it is arrived using commercially available ELISA kit (ALPCO Diagnostics) measure serum ferritin concentration.

Statistical analysis.It is for statistical analysis using 5 software of GraphPad Prism.Continuous variable (quantitative demographics number According to clinical and laboratory data) it is expressed as median and interquartile-range IQR (IQR), and examined and carried out using Mann-Whitney U Compare.Wilcoxon signed rank test is for comparing two pairs of group, and it is not assumed that the different distribution of pre-post difference follows Gaussian Profile.

Spearman rank correlation is for evaluating and the relationship of laboratory parameters.Think with statistically significant p value < 0.05 Property.

As a result: being increased in the Chemokines Levels that MAS patient's IFN γ and IFN γ induce.There is no the work of MAS when sampling When dynamic property sJIA patient is compared with the patient sampled during clinical inactivity disease, (de as expected is found Benedetti, F., et al., Correlation of serum interleukin-6 levels with joint involvement and thrombocytosis in systemic juvenile rheumatoid arthritis. Arthritis Rheum, 1991. 34 (9): p. 1158-63), the IL-6 level of activity sJIA patient, which is significantly higher than, faces Bed inactivity Disease (p < 0.01).It is most of to suffer from as reported in the research of first first few items activity SJIA - 1 β level of serum IL of person is lower than detection limit, independently of disease activity state.It is worth noting that, Clinical Activity sJIA suffers from The level of the chemotactic factor (CF) of the IFN γ and the induction of 3 kinds of IFN γs of person and clinical inactivity Disease does not have difference.

When sampling, MAS patient when sampling with not having the activity sJIA patient of MAS to be compared, IL-1 β and IL-6 Be on close level, show the level of the known two kinds of cell factors to play a crucial role in activity sJIA in MAS phase terminal phase Between not will increase.It should be noted that recycling IL-1 β level lower than fixed in most of sJIA patients with or without MAS Amount limit (i.e. 3.5 pg/ml).In contrast, there is no MAS when the circulation IFN γ level of activity MAS patient is significantly higher than sampling Activity sJIA patient.With sampling when do not have compared with the activity sJIA patient of MAS, 3 kinds of IFN of activity MAS patient γ associated chemokine CXCL9, CXCL10 and CXCL11's is horizontal also significant higher.This species diversity is particularly evident to CXCL9, Activity sJIA patient Gao Yue 15 times of the Median levels of middle MAS patient than not MAS.

In sec-HLH patient, the level of the level of IFN γ and 3 kinds of IFN γ associated chemokines is significant to be increased.IFNγ It is largely difficult to differentiate between with the level of IFN γ associated chemokine and the level of MAS patient, and difference is aobvious without statistics Work property.Incidentally, in activity MAS patient and activity sec-HLH patient, IFN γ and the induction of 3 kinds of IFN γs become Patient receiving treatment is suitable in non-patient receiving treatment and for the level of the change factor.

The level of IFN γ and CXCL9, CXCL10 and CXCL11 in individual patient MAS there are related.Fig. 8 A-8D is aobvious Show during activity MAS and during the activity sJIA of not MAS can get paired samples individual patient in IFN γ and The level of CXCL9, CXCL10 and CXCL11.It is consistent with the result obtained in crossed section analysis, it is analyzed by paired samples, The level of IFN γ and the chemotactic factor (CF) of 3 kinds of IFN γs induction is significant higher in the sample obtained during MAS.In addition, suffering from several Person can get sample before and after MAS breaking-out, and confirm that IFN γ and IFN γ lure after MAS clinical symptoms recession The Chemokines Levels led restore to normal value.For example, the 3 MAS breaking-outs of a patient experience in the research, in these hairs Blood serum sample is obtained during the disease phase for not having MAS during work and in sampling.Further confirm that IFN γ and 3 kinds of IFN γs lure The generation for the chemotactic factor (CF) led increases the relationship between activity MAS, only has found IFN γ when MAS breaks out in the patient The horizontal of chemotactic factor (CF) relevant with 3 kinds of IFN γs increases (Fig. 9 A-9B).

The level of IFN γ and IFN γ associated chemokine is related to the laboratory abnormalities of MAS.Then IFN γ and 3 is checked The correlation of the laboratory parameters of MAS when the level and sampling of the chemotactic factor (CF) of kind IFN γ induction.In the activity of not MAS The level of the chemotactic factor (CF) of sJIA patient, IFN γ and the induction of 3 kinds of IFN γs is unrelated with the laboratory parameters of MAS, but has an example It is outer: level, the r related to ALT slight level of CXCL9, CXCL10 and CXCL11²In the range of 0.17-0.25 (table 2).This The meaning of kind of correlation is unclear, it is noted, however, that either with or without MAS activity sJIA patient, ALT water It puts down in the normal range.The patient with MAS in sampling, does not have found the Laboratory Characteristic and the significant phase of IL-1 and IL-6 of MAS It closes.In contrast, the level and ferritin of the chemotactic factor (CF) of the patient with MAS in sampling, IFN γ and IFN γ induction It is horizontal, to neutrophil leucocyte and platelet count and related with the raising of LDH and ALT, it is all these generally in MAS patient Abnormal (table 2).IFN γ and CXCL9 and the correlation of laboratory abnormalities are particularly evident, and unique exception is the phase of IFN γ with LDH Guan Xing is not up to statistical significance (table 2 and Figure 10 A-10J).Equally, as described above, there is no the activity of MAS in sampling These correlations are not present in sJIA patient.A patient in the group have significant high-caliber IFN γ (336.2 pg/ml), CXCL9 (549400 pg/ml) and CXCL10 (35066 pg/ml).The patient suffers from the MAS of especially severe, and because serious Intensive care unit is moved in central nervous system involvement.The observation result further supports IFN γ and the level and disease of CXCL9 There are the hypothesis of strong correlation between sick severity.In short, these are the result shows that IFN γ and the relevant chemotactic factor (CF) of IFN γ Generation increase be activity MAS feature, it is strongly related to the severity of the laboratory abnormalities of MAS.

Do not have when patient when patient of the table 2. with the secondary HLH of activity, sampling with activity MAS, sampling IL-1 β of the activity sJIA patient of MAS and the patient with clinical inactivity sJIA, IL-6, IFN γ and 3 kinds of IFN γ phases The serum levels of the Chemokine CXCL9 of pass, CXCL10 and CXCL11.

Numerical value is shown as median (interquartile-range IQR)

* activity sJIA and clinical inactivity sJIA:p < 0.01

3. MAS patient of table and sampling when do not have the disease activity of the activity sJIA patient of MAS laboratory parameters and IFN γ, The correlation of CXCL9, CXCL10, CXCL11 and IL-6 level.

NEU=neutrophil count；PLT=platelet count；ALT=alanine aminotransferase；LDH=lactic acid is de- Hydrogen enzyme；

1=median (IQR)；r*= Spearman r.

The laboratory parameters of the disease activity of MAS patient and activity sJIA patient and IFN γ, CXCL9, CXCL10, The correlation of CXCL11 and IL-6

The horizontal correlation for the chemotactic factor (CF) that the IFN γ of MAS patient is induced with IFN γ.In order to further characterize MAS patient's Relationship between IFN γ and the chemotactic factor (CF) of 3 kinds of IFN γs induction, has evaluated IFN γ level and every kind of single Chemokines Levels Correlation.It is worth noting that, CXCL9 seems main and is specifically induced by IFN γ, and CXCL10 and CXCL11 also by I type is interferon-induced.It is consistent with this, in activity MAS patient, the cyclical level of IFN γ and CXCL9 it is significant it is related (r= 0.693; r2=0.48;P=0.001), but weaker (r=0.535 of correlation with CXCL10 level; r2=0.29; p= 0.015) (Figure 11 A-11F).It is also weaker with the correlation of CXCL11 level, and not up to statistical significance (r=0.447; r2=0.20;P=0.08) (not shown).

The chemotactic factor (CF) of IFN γ induction is related to the disease activity of MAS mouse model.It is lured to further study IFN γ The chemotactic factor (CF) led generates the correlation with MAS, has studied these chemotactic factor (CF)s at target tissue (liver and spleen) in the mouse model of MAS In expression.In the model, more by the background Imitating TLR4 agonist rouge in IL-6 transgenic mice high level IL-6 The acute infection of sugared (LPS) induces MAS clinic and Laboratory Characteristic (Strippoli et al., Arthritis Rheum 2012).This method summarises the symptom of sJIA patient's generation: infecting there are active disease may trigger MAS/HLH, really characterized by high-caliber IL-6.After being induced with LPS, deposited in the liver of IL-6 transgenic mice and spleen In the CXCL9 and CXCL10 of high mRNA level in-site.It is worth noting that, CXCL9 in the serum levels and spleen and liver of ferritin and The significant correlation of the expression of CXCL10 in liver shows the IFN γ associated upstream event in target tissue (i.e. in liver and spleen CXCL9 and CXCL10 are generated) and the such as high ferritin levels of typical downstream laboratory abnormalities between relationship.In short, MAS suffers from Person and MAS mouse model statistics indicate that the generation of IFN γ increases the expression with CXCL9 (CXCL10 in lesser degree) increases And the clear correlation of laboratory abnormalities of MAS.

The research of patient and both animal models to p-HLH are it has been confirmed that core of the IFN γ in disease incidence mechanism Effect.However, effect of the IFN γ in sec-HLH (including in the MAS under sJIA background) is still unclear.The research is finally demonstrate,proved In fact, in the patient of sJIA generation MAS, there are the chemotactic factor (CF)s that high-caliber IFN γ and IFN γ induce.In addition, IFN γ, CXCL9 It is strongly related to the laboratory parameters of MAS severity to the level of CXCL10.Should the study found that in activity sJIA patient and The serum levels of IFN γ and the relevant chemotactic factor (CF) of 3 kinds of IFN γs are suitable between clinical inactivity Disease.The result is no Determine pathogenic effects of the IFN γ in sJIA, and really consistent with many observation results of other authors.3 gene expressions are ground Study carefully and exists significantly in the peripheral blood mononuclear cells (PBMC) that do not send out when being sampled there are currently no the activity sJIA patient of MAS IFN γ induction feature (Fall, N., et al.,Gene expression profiling of peripheral blood from patients with untreated new-onset systemic juvenile idiopathic arthritis reveals molecular heterogeneity that may predict macrophage activation syndrome. Arthritis Rheum, 2007. 56(11): p. 3793-804; Ogilvie, E.M., et al., Specific gene expression profiles in systemic juvenile idiopathic arthritis. Arthritis Rheum, 2007. 56(6):p. 1954-65; Pascual, V., et al., Role of interleukin-1 (IL-1) in the pathogenesis of systemic onset juvenile idiopathic arthritis and clinical response to IL-1 blockade. J Exp Med, 2005. 201(9): p. 1479-86).After stimulation PBMC in vitro, IFN γ is generated in activity sJIA patient Cell number it is similar to control group (Lasiglie, D., et al.,Role of IL-1 beta in the development of human T(H)17 cells: lesson from NLPR3 mutated patients. PLoS One, 2011. 6(5): p. e20014).Always, the patient with both activity and inactivity SJIA is not presented Out the serum of IFN γ or synovial fluid level increase (de Jager, W., et al.,Blood and synovial fluid cytokine signatures in patients with juvenile idiopathic arthritis: a cross- sectional study. Ann Rheum Dis, 2007. 66(5): p. 589-98).Support IFN γ in the joint of sJIA It is not acted in inflammation, is nearly no detectable CXCL9 and CXCL10 in the synovial tissue of sJIA patient, and in few joint or more Can be found in the synovial tissue of joint JIA patient these high-caliber chemotactic factor (CF)s (Sikora, K.A., et al.,The limited role of interferon-gamma in systemic juvenile idiopathic arthritis cannot be explained by cellular hyporesponsiveness. Arthritis Rheum, 2012. 64 (11): p. 3799-808).Nearest mouse statistics indicate that, immune thorn is carried out to IFN γ knock-out mice with Freund's complete adjuvant Swashing can produce the inflammatory syndrome of whole body type including sJIA feature, further support effect of the IFN γ in sJIA limited (Avau, A., et al., Systemic juvenile idiopathic arthritis-like syndrome in mice following stimulation of the immune system with Freund's complete adjuvant: regulation by interferon-gamma. Arthritis Rheumatol, 2014. 66(5): p.1340-51)。

Sharp contrast is formed with this, researches show that the IFN γs and IFN γ phase when activity MAS patient samples out for this The Chemokines Levels of pass are apparently higher than the activity sJIA patient for not having MAS when sampling.For during activity MAS and not having There is the individual patient for obtaining series of samples during the activity sJIA of MAS to also demonstrate this point.Incidentally, this is studied Do not find that horizontal the dramatically increasing of IL-6 or IL-1 β of the patient sampled during MAS does not have with the laboratory parameters of MAS yet Any association, show these cell factors although seriously participate in sJIA pathogenic mechanism (De Benedetti, F., et al.,Randomized trial of tocilizumab in systemic juvenile idiopathic arthritis. N Engl J Med, 2012. 367(25): p. 2385-95; Ruperto, N., et al., Two randomized trials of canakinumab in systemic juvenile idiopathic arthritis. N Engl J Med, 2012. 367 (25): p. 2396-406), but may be not most important in terms of maintaining MAS.IFN γ and IFN γ This raised result of study of relevant Chemokines Levels is consistent with some previous observation results.The report such as Shimizu, with There is no the activity sJIA patient of MAS to compare, neopterin (three phosphorus of guanosine synthesized after being stimulated with IFN γ by human macrophage The catabolite of acid) MAS patient levels during sJIA it is higher (Shimizu, M., et al.,Distinct cytokine profiles of systemic-onset juvenile idiopathic arthritis-associated macrophage activation syndrome with particular emphasis on the role of interleukin-18 in its pathogenesis. Rheumatology (Oxford), 2010. 49(9): p. 1645-53).Recently, Put etc. reports the IFN γ of 5 primary and secondary HLH patient and CXCL10 level increases, wherein 3 during sJIA with MAS (Put, K., et al.,Cytokines in systemic juvenile idiopathic arthritis and haemophagocytic lymphohistiocytosis: tipping the balance between interleukin-18 and interferon-gamma. Rheumatology (Oxford), 2015).Consistent with these results, whens 5 samplings, does not have the IFN γ of the activity sJIA patient of MAS and CXCL10 horizontal significant It reduces (Put et al., Rheumatology 2015).

It is interesting that this is the study found that not only IFN γ and the relevant Chemokines Levels of IFN γ significantly increase, but also Its horizontal (especially level of CXCL9) is strictly related to the Laboratory Characteristic of MAS, shows associated with disease severity. Further support is associated with disease severity, the study find that one refreshing with multiple organ failure and maincenter with serious disease Need to extend the IFN γ and CXCL9 and CXCL10 water of the patient that intensive care unit is moved in generalized seizures through system involvement Head up display writes higher.

In MAS patient, in the chemotactic factor (CF) of 3 kinds of IFN γs induction, discovery CXCL9 has strongest phase with IFN γ level Guan Xing.This observation result with it is established identical of views, i.e. what the generation of CXCL9 was seemingly only induced by IFN γ specificity, And the generation of CXCL10 and CXCL11 can also it is interferon-induced by I type (Groom, J.R. and A.D. Luster,CXCR3 ligands: redundant, collaborative and antagonistic functions. Immunol Cell Biol, 2011. 89(2): p. 207-15).It is movable sensitive raw with specificity that this shows that the level of CXCL9 can be used as MAS Object marker.Really, it is simulated using the mouse model of MAS and MAS is triggered by infection sexual stimulus under the background of high IL-6 level (Strippoli et al., Arthritis Rheum 2012), the expression water of CXCL9 in liver and spleen is also found in this research It is flat significant related to the cyclical level of ferritin.For CXCL10 expression, this correlation exists only in liver, and for Spleen level is not present.This point is also supported to the result of study of MAS patient, wherein all experiments of CXCL9 level and MAS Room parameter is strictly related.In short, these the mankind and mouse observation indicate that, CXCL9 and MAS feature and IFN γ are generated It is strong related, it further supports the excessive of IFN γ and generates the hypothesis for playing principal causative in MAS.These observation results Also the continuous leaching of the identical SJIA patient obtained during the activity sJIA from not MAS and during MAS is used with Put etc. The immunohistochemistry data for fawning on biopsy generation are consistent.They are reported in the tissue obtained during MAS, pass through immuning tissue Chemical detection is not having to high-caliber CXCL10 and indoleamine 2,3-dioxygenase (the two is IFN γ inducible protein) The testing result obtained during the activity sJIA of MAS is not high (Put et al., Rheumatology 2015).

The observation result of obtainable p-HLH patient together, is supported in these of MAS and sec-HLH result and document One independently of basic reason that the increase of IFN γ and the relevant chemotactic factor (CF) of IFN γ (especially CXCL9) is HLH is only The hypothesis of feature.In this respect, enjoyably it is noted that being examined in the recurrent MAS patient for obtaining function mutation induction by NLRC4 Measure high-caliber CXCL9 (Canna, S.W., et al.,An activating NLRC4 inflammasome mutation causes autoinflammation with recurrent macrophage activation syndrome. Nat Genet, 2014. 46 (10): p. 1140-6), show even if only by inflammatory corpusculum functional disturbance Under the background of the HLH of induction, it is also possible to there may suitably be IFN γ and excessively generate.

Be unequivocally established the data (in both perforin and Rab27a knock-out mice) of p-HLH animal model IFN γ Pathogenic effects.Similarly, the latest data of the HLH model (the HLH model secondary to infection) of TLR9 induction also shows that increase IFN γ generate main function (Behrens, E.M., et al.,Repeated TLR9 stimulation results in macrophage activation syndrome-like disease in mice. J Clin Invest, 2011. 121 (6): p. 2264-77 and (Bautois et al., progress in).It is carried out recently in above-mentioned MAS mouse model other Studies have shown that causing to improve survival with anti-IFN γ Antybody therapy and restoring the clinic and Laboratory Characteristic (Prencipe et of MAS Al., in progress).In short, the result of this research and in animal these observation results be IFN γ in and controlling as MAS Treatment method provides reasonability.

The 7. primary Hemophagocytic lymphohistocysis multiple intravenous administration of disease (HLH) infant of embodiment is anti- Safety, tolerance, pharmacokinetics and the therapeutic evaluation of the anti-IFN γ monoclonal antibody of interferon gamma

Research presented herein is intended to determine the peace that the multiple vein (IV) for the anti-IFN γ antibody for being herein referred as NI-0501 is given Full property and tolerance characteristics determine that NI-0501 in the curative effect and benefit/risk feature of HLH patient, describes NI-0501 and suffers from HLH Pharmacokinetics (PK) feature of person determines suitable NI-0501 therapeutic dose scheme for HLH and evaluates the immune of NI-0501 Originality.

Preclinical study: previous research have confirmed NI-0501 to from non-human primate species (including rhesus macaque and Machin) IFN γ there is similar binding affinity and blocking activity, but to from dog, cat, pig, rabbit, rat or mouse IFN γ does not have then.The toxicology and safety research of machin show to give NI-0501 and do not miss the target toxicity, give weekly NI-0501 tolerance is good and does not need antibiotic prophylaxis, and is not observed during these previous research abnormal Histopathology or behavior discovery.

Since NI-0501 combines the ability of IFN γ free and that IFN γ R1 is combined, research has been carried out to study NI- 0501 mediates the active potentiality of ADCC and CDC there are target.It confirms shortage ADCC activity, does not observe CDC Active induction.

I phase clinical research: one is studied 20 healthy adult volunteers the safety that single dose intravenous (IV) gives NI-0501 Property, tolerance and Pharmacokinetic Characteristics 1 phase randomized double-blind placebo single dose be incremented by research.During the research, 6 Name subject receives placebo, and 3,3,4 and 4 subjects's (amounting to 14 subjects) receive 0.01,0.1,1 and 3 respectively The NI-0501 dosage of mg/kg.

The PK analysis of NI-0501 discloses the expection feature of IgG1, i.e. long half time (about 22 days), slowly remove (≤ 0.007 L/h) and volume of distribution it is low (average < 6 L).

After starting infusion of drug, 14 (70%) in 20 subjects, which are observed, amounts to 41 adverse events (AE), wherein 10 4 subjects reporteds by receiving placebo.36 (87.8%) AE have slight intensity and 5 (12.2%) there is moderate strength.Serious or threat to life AE is not reported.10 in 14 subjects for living through AE Name has 23 AE (56.1%) to be reported as related (a possibility that at least reasonable) with drug.Most of AE are single incident, And it is not observed and increases the dose-dependent trend of NI-0501.All NI-0501 infusions are without incident.

In short, the infusion tolerance of NI-0501 is good, and observed during the monitorings in 8 weeks after infusion of drug Effect does not show any serious or unexpectedly miss the target safety or Immunogenicity.

2/3 phase clinical research material and method: these researchs are carried out based on primary HLH patient.Research is divided into 3 portions Point: screening, treatment and follow-up.It summarizes as shown in figure 12.

In these researchs, suitable patient includes the patient's (herein also referred to as " line trouble for not receiving HLH treatment Person "), or conventional H LH treatment may have been received and do not obtain satisfactory reaction (such as according to treating physician) or Show the patient (herein also referred to as " two wires patient ") for the sign not tolerated to it.In conventional H LH treatment failure or show The patient for receiving NI-0501 after not tolerating to it represents crucial group of the research, to confirm NI-0501 as primary HLH Second line treatment curative effect.The patient for not receiving treatment has participated in the collection of the efficacy and saferry data under a line background.

Following patient is excluded except this research: being diagnosed as secondary HLH after being diagnosed as rheumatic or tumor disease Patient；Previously before screening before during 2 weeks with any T cell depletor (such as antithymocyte globulin (ATG), Anti- CD52 treatment) it treats or (is felt in addition to B cell EBV is recorded in the half-life period of its determination at 5 times with any other bio-pharmaceutical Rituximab in the case of dye) treatment patient；With activity mycobacteria (mycobacteria), histoplasmin Bacterium (Histoplasma capsulatum), Shigella (Shigella), salmonella (Salmonella), campylobacter (Campylobacter) and Leishmania (Leishmania) infection patient；With past tuberculosis or latency knot The patient of core medical history evidence；The patient of HIV antibody, hepatitis B surface antibody or antibody to hepatitis C positive serology；In the presence of The patient of malignant tumour；The disease of angiocarpy, lung, liver or renal function or the patient of deformity are seriously affected with another kind simultaneously； There is the patient of allergy or allergy history to any component part of research approach；Receive living or attenuation in 12 weeks before screening The patient of (including BCG) vaccine living and/or gestation or lactating female patient.

Research presented herein uses anti-interferon gamma antibodies NI-0501, is a kind of full human IgG1 for people's IFN γ Monoclonal antibody (mAb).NI-0501 provides (every mL) without bacterium concentrate as infusion, as shown in the following Table 4.

4. NI-0501 preparation of table

Ingredient	Quantity (every mL)
		NI-0501	5 mg
L-Histidine	1.55 mg
		L-Histidine mono-hydrochloric salts monohydrate	3.14 mg
Sodium chloride (NaCl)	7.31 mg
		Polyoxyethylene sorbitan monoleate	0.05 mg
pH	6.0 ± 0.2

In these researchs, NI-0501 is transfused by IV and was given with the predose of 1 mg/kg through 1 hour period.It is expected that The dosage inhibits IFN γ effect 3 days of at least 99% in patient of the baseline IFN γ concentration less than or equal to 3400 pg/mL.Infusion It carries out once within every 3 days, until the 15th day (SD15) of research (infusion #6), and twice a week later.According to during research The preassigned guided any time as the clinic of every patient and laboratory reaction (as described in following table 5), NI-0501 dosage It is possible for increasing to 3 mg/kg.After least twice is transfused 3 mg/kg, if discovery makes patient have money after reappraising The identical clinical and laboratory standard that lattice receive the NI-0501 of 3 mg/kg stands good, then the dosage of NI-0501 is with periodically Monitoring is clinical and HLH parameter in laboratory can increase to 6 mg/kg, at most infusion 4 times.Differentiation based on these parameters, NI-0501 Dosage can i) drop back to and 6 mg/kg are maintained at for other IV infusion to 3 mg/kg or ii) (or increase to 6 mg/kg More than), if PK and PD evidence shows that IFN γ generates excessively high and therefore quickly eliminates NI-0501.If met as described herein Clinical and laboratory standard, then dosage increase can occur for any time during research.

Table 5. instructs the increased clinical and laboratory standard of dosage

^aIf these standards are suitable for after SD6, the dosage of NI-0501 must increase to 3 mg/kg from 1 mg/kg.

^bIt, must be before standard revaluation with the dosage of 3 mg/kg if having increased the dosage of NI-0501 in SD3 It is transfused at least twice.

^c3 mg/kg are increased to depending on whether SD3 or SD6 has had occurred dosage.

^dAt most infusion 4 times.

Abbreviation: bsl.=baseline；ANC=Absolute Neutrophil Count；US=ultrasound

In these researchs, NI-0501 is given 8 weeks, and treatment phase is classified into 2 different times: treatment phase 1 and 2, such as Figure 12 It is shown.

After NI-0501 gives 8 weeks, the pretreating scheme of hematopoietic stem cell transplantation (HSCT) can be begun preparing.If The state of an illness and donor availability of patient allows to transplant, then can shorten expected duration for the treatment of, although 4 will not be less than Week.If suitable donor was not yet determined by the 8th week, or if with NI-0501 give it is unrelated due to need to postpone Schedule is transplanted, then NI-0501 treatment can continue in the case where long-term follow-up study, and condition has been established for patient The benefit/risk of benefit.

In these researchs, NI-0501 is given under the background of dexamethasone, can be according to the state of an illness of patient gradually Decrement.In the patient for not receiving treatment, NI-0501 will be in 10 mg/m²It is given under the background of dexamethasone.Receiving NI- 0501 patient as two wires HLH treatment, dexamethasone must be at least 5 mg/m²Dosage give, or before screening (if higher) is given with identical dosage.Patient needs to receive dexamethasone since SD-1.

According to the judgement for the treatment of physician, dexamethasone can be gradually reduced according to the state of an illness of patient.Treating physician may be selected by The scheme decrescence measured, condition are that the dexamethasone dosage of each step is no more than half and the frequency of variation is no more than weekly.

If disease progression after dexamethasone is gradually reduced, the dosage of dexamethasone can increase and keep, until root Satisfied reaction is realized according to treating physician.

According to the recommendation in HLH treatment guidelines, the previous day of patient since being treated NI-0501 terminates to receive up to research Ye Shi lung pityrosporion ovale (Pneumocystis jiroveci), fungi and shingles zoster (Herpes Zoster) virus infection it is pre- Anti- property treatment.Patient receives prophylactic treatment up to research terminates the previous day (i.e. SD-1) since being treated NI-0501. For example, for Ye Shi lung pityrosporion ovale (Pneumocystis jiroveci) prevention, patient is acceptable twice daily, connects weekly Such as 750 mg/m are given so that equal part dosage is oral within continuous 3 days²The sulfamethoxazole in/day and 150 mg/m²The trimethoprim in/day.It is right In the prevention of fungal infection, acceptable daily such as 12 mg/kg of Fluconazole of patient, daily maximum dose is 400 mg.For HZ Virus prevention, for two years old or more child patient be subjected to such as four times a day 200 mg of acyclovir, for two years old with Under children 100 mg 4 times a day.These treatments will take orally as far as possible, otherwise intravenous administration.

Patient is also subjected to any one of various concomitant therapies, such as cyclosporin A, intrathecal methotrexate (MTX) and sugared skin Matter hormone etc..If having given patient before screening, cyclosporin A (CsA) can be continued to use.CsA can be removed at any time It returns.Once NI-0501 is given, CsA will not be reintroduced back to during research process.

If patient receives intrathecal methotrexate (MTX) and glucocorticoid when NI-0501 is treated and started, will be as needed Continue the treatment.If NI-0501 treatment start before there is CNS symptom, must give for the first time NI-0501 it Before begin to use the treatment of intrathecal methotrexate (MTX) and glucocorticoid.

IV immunoglobulin (IVIG) only just allows in the case where immunoglobulin deficiency is recorded as replacement therapy. For example, if immunoglobulin deficiency, which is recorded, meets replacement therapy, IVIG can be with every 4 weeks of the dosage of 0.5 g/kg or more frequency It gives numerously, to keep enough IgG horizontal.During any infusion in first 4 weeks and NI-0501 before screening are treated Any infusion be acceptable.

Allow analgesia therapy, blood transfusion product, electrolyte and glucose infusion, antibiotic, antimycotic and antiviral therapy with And general supportive treatment.Once reaching maximum NI-0501 dosage level, allowed if HLH improvement is non-continuous or limited another Outer HLH treatment.HLH used herein improve it is non-continuous refer to not being able to maintain from baseline 3 HLH parameters improve at least 50% patient (referring to following table 6).The forfeiture of HLH improvement must be recorded in continuous measurement at least twice.HLH used herein Improve limited refer in minimum 3 HLH clinics and laboratory standard from the variation of baseline less than 50%.Etoposide should be used as separately Outer HLH treatment is given, and reacts or does not tolerate unless showing that tangible proof shows to lack drug from previous medical history.

Following therapy cannot be given with NI-0501 and use simultaneously: being generally not allowed and use Etoposide, T cell depletor Or any other bio-pharmaceutical, in addition to following situations: G-CSF, if long-term Neutrophilic granulocytopenia；Rituximab, if B cell EBV infection is recorded；It is treated with other HLH, if HLH improves non-continuous or has under maximum NI-0501 dosage level It limits (as defined herein).Etoposide should be given, unless showing that tangible proof shows to lack drug from previous medical history It reacts or does not tolerate.During entire research (follow-up period including 4 weeks), it is necessary to avoid inoculation living or attenuation (including BCG) epidemic disease Seedling.If NI-0501 concentration is maintained at treatment level after research terminates, the nonvaccinated time should be extended, until no longer It can detect the NI-0501 of measurable concentration.

Characterize the clinical sign (fever, splenomegaly, CNS symptom) and laboratory parameters (CBC, fibrinogen, iron of disease Albumen, sCD25 are horizontal) differentiation for evaluation response realization and reaction time.Primary efficacy endpoint include overall reaction rate, That is the realization that reaction or HLH improve completely or partially at treatment end (EoT), as following table 6 defines.Secondary efficacy terminal Duration including reaction time of any time during research, reaction, (any time during maintaining research was realized Reaction until EoT and after (including the data collected in any long-term follow-up study)), glucocorticoid can be reduced Patient's number of 50% or more of Baseline dose, patient's number that HSCT is able to carry out when thinking to have indication, the 8th week (or EoT) and Survival at the end of research, the serum-concentration of NI-0501 for determining NI-0501 pharmacokinetics (PK) feature, pharmacodynamics (PD) The measurement (including recycling total IFN γ and its neutralizing the level of marker (i.e. CXCL9 and CXCL10)) of effect and other biological mark The measurement of will object (such as sCD25, IL-10).

The definition that table 6. reacts

Security parameters wait collect and evaluate include the disease incidence of adverse events (AE) (serious and not serious), severity, Causality and as a result, pay special attention to infection, laboratory parameters such as whole blood count (CBC) differentiation, pay close attention to red Cell (hemoglobin), neutrophil leucocyte and blood platelet, liver inspection, kidney function test and blood coagulation are exited because of security reason Patient's number and other parameters, such as the circulating antibody of NI-0501 level (if any) to determine immunogenicity (ADA)。

Assessment Primary Endpoint (overall reaction rate) is examined using the horizontal exact binomial in unilateral side 0.025.It is the time of reaction, anti- The duration and time-to-live answered are indicated using Kaplan-Meier curve, if it is available, calculating median.Calculate it is each this 95% confidence interval of the median of a little terminals.By based on binary outcome other terminal (including by glucocorticoid reduce 50% Or more patient's number) and be able to carry out patient's number of HSCT) conversion is proportional and calculates relevant 95% confidence interval.It is only right The statistical significance of Primary Endpoint acquisition p value.Every other terminal is accordingly to be regarded as supporting Primary Endpoint, and therefore will not show end The formal hierarchical structure of point.

It is transfused after NI-0501 in a few hours for the first time, patient, which gives NI-0501, leads to rapid normalization of generating heat.Figure 13 A The influence to body temperature is transfused with 13B two patient NI-0501 for being depicted in body temperature > 37.5 DEG C when NI-0501 treatment starts.Figure 14 be a series of charts and table, describes NI-0501 and gives the influence to patient's neutrophil count.Figure 15 is a series of figures Table and table describe NI-0501 and give the influence counted to Platelet.Figure 16 is a series of charts and table, describes NI- 0501 gives the influence to patient's ferritin serum levels.Figure 17 is a series of charts and table, describes NI-0501 and gives to trouble The influence that person's glucocorticoid is gradually reduced.Figure 18 is to describe giving for NI-0510 IFN γ is kept to neutralize the figure until when HSCT Table.HLH also continues to transplanting the reaction that NI-0501 is treated.After NI-0501 gives, also have evaluated any CNS of patient by It is tired.Baseline CNS involvement and the general introduction of state when treatment end (EOT) are as shown in the following Table 11.

Table 11. is involved to the reaction-CNS that NI-0501 is treated

Annotation: patient receives IT treatment, in addition to 4# patient does not carry out routine administration LP

* it treats and is carrying out

^STreatment was since 2 weeks

^ΛEOT control is not carried out

10 experienced in the patient of hematopoietic stem cell transplantation (HSCT), and all patients are implanted into, after 1 patient is due to HSCT The Mixed chimerism of D+145 and need CD34 stem cell enhance.Second transplant failure occurs for 1 patient, and subsequent HLH is reactivated. Due to acute respiratory failure and bacterium infection, patient D+68 after HSCT is dead.Another patient D+47 after HSCT is dead (septic shock in the case of serious GvHD).Other 3 patients report slight GvHD, and have subsided/subsided.

10 experience transplanting patient in 8 measure HSCT when NI-0501 neutrality serum-concentration, this reflection It is horizontal in the CXCL9 (a kind of chemotactic factor (CF) accurately induced by IFN γ) lower than quantitative limit.Therefore, these are statistics indicate that NI- 0501 can avoid the short-term or long term toxicity to the scheme report based on Etoposide.This, which can be translated into, reduces allo-HSCT The risk of related complication.

These statistics indicate that NI-0501 treatment improve and/or the HLH that can subside relevant clinical and laboratory abnormalities, including CNS S&S.Presence and type and/or infectious triggering factors to the reaction of NI-0501 independent of pathogenic mutation Presence and type.NI-0501 tolerance is good.Do not occur so far safety problem (such as without bone marrow toxicity, without extensive Immunosupress).It is not observed known as in IFN γ and infection caused by the pathogen that promotes.NI-0501 is to IFN γ A kind of method innovated and have targeting can be provided for the treatment of HLH by neutralizing.

Is there is Macrophage Activation Syndrome/secondary HLH (MAS/sHLH) whole body type childhood spy in embodiment 8. (a kind of anti-interferon gamma (anti-IFN γ) monoclonal is anti-by hair property arthritis (sJIA) patient's middle or short term intravenous administration NI-0501 Body) safety, tolerance, pharmacokinetics and curative effect

Research provided herein is intended to confirm that NI-0501 for treating the efficacy and saferry of MAS/sHLH in sJIA patient, is ground Study carefully and is divided into two parts: (i) Primary Study, to assess the PK feature and administration strategy of NI-0501, and preliminary assessment NI- 0501 the patients benefit/risk；(ii) crucial research, to confirm the efficacy and saferry of NI-0501 (research continued after the dosage regimen of confirmation NI-0501 and positive benefit/risk feature).The general introduction of the researching and designing is such as Shown in Figure 19.

The main purpose of Primary Study are as follows: (i) is that the sJIA patient with MAS/sHLH determines that suitable NI-0501 is controlled Treat dosage；(ii) benefit/risk feature of the evaluation NI-0501 in the sJIA patient with MAS/sHLH；(iii) is retouched NI-0501 is stated in pharmacokinetics (PK) feature of the sJIA patient with MAS/sHLH.The main purpose of key research are as follows: (i) determine NI-0501 in the curative effect of the sJIA patient with MAS/sHLH；(ii) it assesses and suffers from the sJIA with MAS/sHLH The short-term vein of person (i.v.) gives safety and the tolerance characteristics of NI-0501；(iii) confirmation NI-0501 is with MAS/ The positive benefit/risk feature of the sJIA patient of sHLH；(iv) exploratory evaluation Chemokine CXCL9 and CXCL10 conduct MAS/sHLH diagnostic biomarkers and predictive factor as the reaction treated to NI-0501；(v) evaluation NI-0501 exists Immunogenicity in sJIA patient with MAS/sHLH.

Study population includes that hyporeactive sJIA with MAS/sHLH is shown to high dose glucocorticoid treatment Patient.The standard of being included in includes the following contents: (i) gender: male and female；(ii) age: when sJIA is diagnosed < 16 years old；(iii) In the case where there is at least 2 or less laboratories and clinical criteria, activity is diagnosed as by treatment rheumatologist's confirmation MAS/sHLH:(a) laboratory standard: platelet count≤262 x10⁹/ L, WBC count≤4.0 x10⁹/ L, AST level > 59 U/L and/or fibrinogen level≤2.5 g/L；(b) clinical criteria: hepatomegaly, bleeding performance and/or CNS function Obstacle；(iv) according to local treatment standard, patient is (including (but unlimited to high dose i.v. glucocorticoid treatment at least 3 days In) 30 mg/kg mPDN pulse for three days on end) show underaction；(v) high dose i.v. glucocorticoid is not lower than 2 Mg/Kg/ days mPDN equivalents, daily 2 individually dosed, highests 60 mg/ days.If conditions of patients and/or laboratory parameters are fast Speed deteriorate, can after starting high dose i.v. glucocorticoid less than 3 days in included.(vi) patient agrees to (or through law The representative of authorization is agreed to)；(vii) receives contraceptives when patient was in after puberty.

Exclusion criteria includes: the HLH that (i) is diagnosed as after primary HLH that is doubtful or making a definite diagnosis or tumor disease；(ii) The following treatment of patient in the half-life period of its determination at 5 times: anakinra, Torr pearl monoclonal antibody, Kang Na monoclonal antibody, tnf inhibitor, benefit Appropriate former times monoclonal antibody or any other bio-pharmaceutical；(iii) activity mycobacteria (mycobacteria) (typical and atypia), Histoplasma capsulatum (Histoplasma capsulatum), Shigella (Shigella), salmonella (Salmonella), campylobacter (Campylobacter) and Leishmania (Leishmania) infection；(iv) latency Evidence lungy；(v) HIV antibody positive serum；(vi) there are malignant tumours；(vii) patient suffers from another kind simultaneously The disease or deformity of angiocarpy, lung, CNS, liver or renal function are seriously affected, opinion according to the study may be significantly affected to controlling A possibility that evaluation of the reaction for the treatment of and/or NI-0501 safety；(viii) had to any component part of research approach Quick or allergy history；(ix) receive BCG vaccine in 12 weeks before screening；(x) receive it in 6 weeks before screening He lives or attenuated live vaccine；And/or (xi) gestation or lactating female patient.

Dosage regimen gives frequency and duration for the treatment of: in these researchs, NI-0501 is for shown in embodiment 7 Preparation in.In part 1, by being transfused in SD0 through 1 hour period, NI- is given with the predose of 6 mg/kg 0501.NI-0501 is (being up to SD27) for 4 weeks with the every 3 days one time continual cure of the dosage of 3 mg/kg.NI-0501 treatment It can shorten afterwards realizing complete clinical response (i.e. MAS reaction).After 4 weeks, NI-0501 treatment can continue to grow as needed It was used as maintenance therapy up to other 4 weeks (being up to SD56), until realizing MAS reaction, it is possible to reduce dosage to 1 mg/kg simultaneously It is primary to giving weekly to extend infusion interval.If PK feature show unexpected TMDD (therefore issue especially height generate The signal of IFN γ), the dosage of NI-0501 can increase to 10 mg/kg according to clinical and PK advice on evidence.Only it is somebody's turn to do in careful evaluation Just ratify dosage increase after the benefit/risk feature of individual patient.

It is suitable in dosage regimen that confirmation proposes and after confirming the positive benefit/risk of NI-0501 in part 2, it should Research will continue.If desired, minor modifications can be carried out to dosage regimen based on the evidence obtained in part 1.

Background therapy and drug combination: NI-0501 is at least 2 mg/kg methylprednisolone (mPDN) equivalents at most 60 It gives under the background of mg/ days (in the patient of 30 kg or more), can be gradually reduced according to conditions of patients during treatment.Suffer from Person preferably starts to receive herpes zoster infection in the previous day (and under any circumstance before starting NI-0501 treatment) Prophylactic treatment, until serum N I-0501 level no longer can be detected.If opened at least 3 days before starting NI-0501 treatment Begin, then can continue to use cyclosporin A (CsA).CsA dosage is allowed to adjust to keep treatment level.According to sentencing for researcher Disconnected, CsA can be recalled at any time during research.It is had begun once NI-0501 gives, CsA cannot be reintroduced back to.If patient Receive intrathecal methotrexate (MTX) and glucocorticoid when NI-0501 is treated and started, then can continue the treatment as needed.Entire Must avoid inoculation living during research or attenuation (including BCG) vaccine, and anyway, until serum N I-0501 level not It can be detected again.Allow analgesia therapy, blood transfusion product, electrolyte and glucose infusion, antibiotic, antimycotic and antiviral therapy And general supportive treatment.

Sample size: in part 1, at least 5 appreciable patients will be recruited.In part 2, continuing to study Afterwards, at least 10 appreciable patients will be recruited, amounts to 15 appreciable patients to obtain.In view of rare orphan's property of disease Matter and the treatment for lacking any approval, it is suitable that 15 sample size not yet formally proves.Nevertheless, based on the assumption that at least 50% Patient to individual systemic glucocorticoid underaction, i.e., 50% based on the patient of glucocorticoid to treating beginning Realization MAS reaction in 8th week afterwards, this research carry out the ability for having 70% to detect from 50% to 77% using 5% one-sided significance level Improvement.

Studying duration and research terminates definition: the research duration of every patient is 8 weeks (plus up to 1 week Screening period).It is medical that the end of research is defined as last patient last time.All patients for receiving at least one NI-0501 It will be required that entering NI-0501-05 research carries out long term follow-up.

It studies terminal: in the part 1 (preliminary) of the research, evaluating the following contents to confirm the administration of the patients Scheme: the benefit/risk feature of (i) NI-0501；(ii) the PK feature of NI-0501；(iii) by IFN γ induction known to The level of chemotactic factor (CF) (such as CXCL9, CXCL10, CXCL11)；(iv) at the 2nd, 4,6 and 8 week after NI-0501 starts, The reduction of different characteristic haemocyte, the differentiation of hepatosis and coagulopathy of MAS；(v) dosage and hold that NI-0501 is treated The continuous time.In the part 2 (key) of the research, effectiveness study terminal is as follows: (a) primary efficacy endpoint: arriving NI-0501 Treat the patient's number for realizing MAS reaction after starting on the 8th week；(b) secondary efficacy terminal: the time of MAS reaction, initial anti- Time for answering can gradually be reduced to MAS and it occurs according to the evaluation of researcher, any time glucocorticoid during research Before patient's number of identical (or lower) dosage for giving；Realize the time, depositing at the end of research that glucocorticoid is gradually reduced Patient's number that is living and withdrawing from the study due to lacking curative effect.In the part 2 (key) of the research, safety research terminal is such as Under: (a) disease incidence of AE (serious and not serious), severity, causality and as a result, pay special attention to infect；Laboratory The parameter especially differentiation of CBC (paying close attention to hemoglobin, neutrophil leucocyte and blood platelet), LFT and coagulation parameters；Due to peace The level (if any) of patient's number and the circulating antibody for NI-0501 that full reason is withdrawn from the study is to determine immunogenicity (ADA)。

Pharmacokinetics and pharmacodynamics pass through following assessment: the PK feature of NI-0501；The free IFN γ of circulation before administration Level and total IFN γ (the free IFN γ+in conjunction with NI-0501) after NI-0501 starts；It is known to be lured by IFN γ The level for the chemotactic factor (CF) (such as CXCL9, CXCL10, CXCL11) led；Chemokines Levels (CXCL9, CXCL10) and free Correlation between NI-0501, free IFN γ (before administration) and the level of total IFN γ；Chemotactic factor (CF) and total IFN γ level with The correlation of the laboratory parameters of MAS severity such as ferritin, platelet count, LFT (exploratory analysis)；And other The level of potential disease biomarker (such as sCD25, IL-10, IL-6, IL-18, TNF α, neopterin).

Other embodiments

Although its detailed description has been combined to describe the present invention, foregoing description intention illustrates and nots limit the scope of the present invention, The scope of the present invention is defined by the appended claims.Other aspects, advantage and modification are in the model of following following claims In enclosing.

Claims

1. one kind in human experimenter in need for treating primary Hemophagocytic lymphohistocysis disease (HLH) more variable dose methods comprising intravenous administration subject:

A) combination that the first dosage of the weight of the 1.0 or 3.0 mg/kg subjects of the subject is given in 12 hours is dry Disturb the antibody of plain γ (IFN γ)；With

B) antibody of the second dosage of the weight of 3.0, the 6.0 or 10.0 mg/kg subjects in 12 hours,

Wherein the antibody includes: the variable heavy chain complementary determining region of the amino acid sequence containing SYAMS (SEQ ID NO:1) The variable heavy chain complementation of 1 (VH CDR1), the amino acid sequence for containing AISGSGGSTYYADSVKG (SEQ ID NO:2) are determined The variable heavy chain for determining area 2 (VH CDR2) and the amino acid sequence containing DGSSGWYVPHWFDP (SEQ ID NO:3) is complementary It determines area 3 (VH CDR3)；The variable light of amino acid sequence containing TRSSGSIASNYVQ (SEQ ID NO:4) is complementary Determine the variable light complementary determining region 2 of area 1 (VL CDR1), the amino acid sequence for containing EDNQRPS (SEQ ID NO:5) 3 (the VL of variable light complementary determining region of (VL CDR2) and the amino acid sequence containing QSYDGSNRWM (SEQ ID NO:6) CDR3)。

2. the method for claim 1 wherein the antibody includes the weight chain variable amino of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of acid sequence and SEQ ID NO:48.

3. the method for claim 1 wherein the antibody of the dosage to give in 6 hours.

4. method for claim 3, wherein the antibody of the dosage is given in 1 hour.

5. continuing the first treatment phase the method for claim 1 wherein every 3 days give the second dosage after the first dosage.

6. method for claim 5, wherein after the first dosage complete the first treatment phase 1,2,3,4,5,6,7,8 week it Afterwards, the second dosage is given, the second treatment phase is continued.

7. method for claim 6, wherein second treatment phase is twice a week.

8. method of claim 1, the method further includes give given in 12 hours 1.0 mg/kg of subject by The antibody of the third dosage of the weight of examination person.

9. the method for claim 1 wherein the antibody dosages to give as single injection.

10. the method for claim 1 wherein the antibody to give as monotherapy or conjoint therapy.

11. the method for claim 1 wherein the subject is Adult human subjects or pediatric subject.

12. one kind in mankind pediatric subject in need for treating secondary Hemophagocytic lymphohistocysis More variable dose methods of disease (HLH) comprising intravenous administration subject:

A) the combination interferon of the first dosage of the weight of the 6.0 mg/kg subjects of the subject is given in 12 hours The antibody of γ (IFN γ)；With

B) antibody of the second dosage of the weight of the 3.0 mg/kg subjects in 12 hours,

13. the method for claim 12, wherein the antibody of the dosage is given in 6 hours.

14. the method for claim 13, wherein the antibody of the dosage is given in 1 hour.

15. the method for claim 12 continues the first treatment phase wherein every 3 days after the first dosage give the second dosage.

16. the method for claim 15 continues the second treatment phase wherein giving the second dosage after completing the first treatment phase.

17. the method for claim 16, wherein second treatment phase is twice a week.

18. the method for claim 12, the method further includes given after second dosage in 12 hours to The antibody of the other dosage of the weight of the 6.0 mg/kg subjects given.

19. the method for claim 12, wherein the antibody dosage is given as single injection.

20. the method for claim 12, wherein the antibody is given as monotherapy or conjoint therapy.

21. one kind in adult humans subject in need for treating secondary Hemophagocytic lymphohistocysis More variable dose methods of disease (HLH) comprising intravenous administration subject:

A) knot of the first dosage of 3 mg/kg of the subject or the weight of 6.0 mg/kg subjects is given in 12 hours Close the antibody of interferon gamma (IFN γ)；With

B) antibody of the second dosage of the weight no more than 10.0 mg/kg subjects in 12 hours,

22. the method for claim 21, wherein the antibody of the dosage is given in 6 hours.

23. the method for claim 22, wherein the antibody of the dosage is given in 1 hour.

24. the method for claim 22 continues the first treatment phase wherein every 3 days after the first dosage give the second dosage.

25. the method for claim 24 continues the second treatment phase wherein giving the second dosage after completing the first treatment phase.

26. the method for claim 25, wherein second treatment phase is twice a week.

27. the method for claim 21, the method further includes giving to give in 12 hours after the second dosage 1.0, the antibody of the other dosage of the weight of 3.0 or 6.0 mg/kg subjects.

28. the method for claim 21, wherein the antibody dosage is given as single injection.

29. the method for claim 21, wherein the antibody is given as monotherapy or conjoint therapy.

30. a kind of for treating more variable dose methods of illness in human experimenter in need comprising intravenously give Give subject:

A) antibody of the combination interferon gamma (IFN γ) of the first dosage of the subject is given in 12 hours；With

B) antibody of the second dosage of the weight of the subject in 12 hours,

31. the method for claim 30, wherein the antibody includes the weight chain variable ammonia of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of base acid sequence and SEQ ID NO:48.

32. the method for claim 30, wherein the antibody of the dosage is given in 6 hours.

33. the method for claim 32, wherein the antibody of the dosage is given in 1 hour.

34. the method for claim 30 continues the first treatment phase wherein every 3 days after the first dosage give the second dosage.

35. the method for claim 34 continues the second treatment phase wherein giving the second dosage after completing the first treatment phase.

36. the method for claim 35, wherein second treatment phase is twice a week.

37. the method for claim 30, wherein first dosage is between the weight of 1.0-10 mg/kg subject.

38. the method for claim 30, wherein the dosage is between the weight of 1.0-10 mg/kg subject, wherein described Two dosage are below or above first dosage.

39. the method for claim 30, the method further includes give given in 12 hours the subject The antibody of third dosage between the weight of 1.0-10 mg/kg subject.

40. the method for claim 30, wherein the antibody dosage is given as single injection.

41. the method for claim 30, wherein the antibody is given as monotherapy or conjoint therapy.

42. the method for claim 30, wherein the subject is Adult human subjects or pediatric subject.

43. the method for claim 30, wherein the illness is graft rejection.

44. the method for claim 43, wherein the graft rejection is solid organ transplantation obstacle, marrow acute transplantation rejection.

45. the method for claim 30, wherein the illness be graft versus host disease(GVH disease), paraneoplastic cerebellar degeneration, Hemorrhagic fever, Sarcoidosis, adult onset still disease.

46. the method for claim 30, wherein the method gives subject after receiving CART cell therapy.

47. the method for any one of preceding claims, wherein the subject gives immediately before giving the antibody Dexamethasone.

48. the method for claim 30, wherein the dexamethasone is at least 10 mg/m²Dosage give.

49. the method for claim 30, wherein the subject is at least 5 mg/m²Dosage give dexamethasone.

50. the method for any one of preceding claims, wherein the subject had not received the treatment to HLH previously.

51. the method for any one of preceding claims, wherein the method further includes giving the subject at least Second of drug.

52. the method for claim 523, wherein second of drug is therapeutic agent, anti-inflammatory agent and/or immunosuppressor.

53. a kind of pharmaceutical formulation of injectable, every mL includes:

A) full people's anti-interferon gamma (IFN γ) monoclonal antibody of 5 mg or 25 mg；With

B) 1.55 mg L-Histidines, 3.14 mg L-Histidine mono-hydrochloric salts monohydrates, 7.31 mg sodium chloride (NaCl) With 0.05 mg polyoxyethylene sorbitan monoleate, wherein pH is between 5.8-6.2.

54. the preparation of claim 53, wherein the antibody includes: the amino acid sequence containing SYAMS (SEQ ID NO:1) Variable heavy chain complementary determining region 1 (VH CDR1), contain AISGSGGSTYYADSVKG (SEQ ID NO:2) amino acid The variable heavy chain complementary determining region 2 (VH CDR2) of sequence and the amino for containing DGSSGWYVPHWFDP (SEQ ID NO:3) The variable heavy chain complementary determining region 3 (VH CDR3) of acid sequence；Amino containing TRSSGSIASNYVQ (SEQ ID NO:4) The variable light complementary determining region 1 (VL CDR1) of acid sequence, the amino acid sequence for containing EDNQRPS (SEQ ID NO:5) Variable light complementary determining region 2 (VL CDR2) and amino acid sequence containing QSYDGSNRWM (SEQ ID NO:6) Variable light complementary determining region 3 (VL CDR3).

55. the preparation of claim 54, wherein the antibody includes the weight chain variable ammonia of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of base acid sequence and SEQ ID NO:48.

56. a kind of unit dose vial, full people's anti-interferon gamma (IFN γ) monoclonal for being suitable for injection it includes 20 ml is anti- Liquid solution, wherein the concentration of antibody is 5 mg/ml or 25 mg/ml, and the pH of the solution is between 5.8-6.2.

57. the unit dose vial of claim 56, wherein the antibody is dissolved in the solution, so that the solution is clear Clearly, colourless and do not precipitate.

58. the unit dose vial of claim 56, wherein the antibody includes: the ammonia containing SYAMS (SEQ ID NO:1) The variable heavy chain complementary determining region 1 (VH CDR1) of base acid sequence contains AISGSGGSTYYADSVKG (SEQ ID NO:2) Amino acid sequence variable heavy chain complementary determining region 2 (VH CDR2) and containing DGSSGWYVPHWFDP (SEQ ID NO: 3) the variable heavy chain complementary determining region 3 (VH CDR3) of amino acid sequence；Containing TRSSGSIASNYVQ (SEQ ID NO: 4) the variable light complementary determining region 1 (VL CDR1) of amino acid sequence, the ammonia for containing EDNQRPS (SEQ ID NO:5) The variable light complementary determining region 2 (VL CDR2) of base acid sequence and the amino for containing QSYDGSNRWM (SEQ ID NO:6) The variable light complementary determining region 3 (VL CDR3) of acid sequence.

59. the unit dose vial of claim 58, wherein the antibody includes the weight of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of chain variable amino acid sequence and SEQ ID NO:48.

60. a kind of unit dose vial, it includes full people's anti-interferon gammas (IFN γ) that 10 ml or 20 ml are suitable for injection Monoclonal antibody solution, wherein the concentration of antibody is 25 mg/ml, and the pH of the solution is between 5.8-6.2.

61. the unit dose vial of claim 60, wherein the antibody is dissolved in the solution, so that the solution is clear Clearly, colourless and do not precipitate.

62. the unit dose vial of claim 60, wherein the antibody includes: the ammonia containing SYAMS (SEQ ID NO:1) The variable heavy chain complementary determining region 1 (VH CDR1) of base acid sequence contains AISGSGGSTYYADSVKG (SEQ ID NO:2) Amino acid sequence variable heavy chain complementary determining region 2 (VH CDR2) and containing DGSSGWYVPHWFDP (SEQ ID NO: 3) the variable heavy chain complementary determining region 3 (VH CDR3) of amino acid sequence；Containing TRSSGSIASNYVQ (SEQ ID NO: 4) the variable light complementary determining region 1 (VL CDR1) of amino acid sequence, the ammonia for containing EDNQRPS (SEQ ID NO:5) The variable light complementary determining region 2 (VL CDR2) of base acid sequence and the amino for containing QSYDGSNRWM (SEQ ID NO:6) The variable light complementary determining region 3 (VL CDR3) of acid sequence.

63. the unit dose vial of claim 62, wherein the antibody includes the weight of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of chain variable amino acid sequence and SEQ ID NO:48.

64. a kind of unit dose vial, the full people's anti-interferon gamma (IFN γ) for being suitable for injection it includes 2 ml or 20 ml is single Clonal antibody solution, wherein the concentration of antibody is 5 mg/ml, and the pH of the solution is between 5.8-6.2.

65. the unit dose vial of claim 64, wherein the antibody is dissolved in the solution, so that the solution is clear Clearly, colourless and do not precipitate.

66. the unit dose vial of claim 64, wherein the antibody includes the ammonia containing SYAMS (SEQ ID NO:1) The variable heavy chain complementary determining region 1 (VH CDR1) of base acid sequence contains AISGSGGSTYYADSVKG (SEQ ID NO:2) Amino acid sequence variable heavy chain complementary determining region 2 (VH CDR2) and containing DGSSGWYVPHWFDP (SEQ ID NO: 3) the variable heavy chain complementary determining region 3 (VH CDR3) of amino acid sequence；Containing TRSSGSIASNYVQ (SEQ ID NO: 4) the variable light complementary determining region 1 (VL CDR1) of amino acid sequence, the ammonia for containing EDNQRPS (SEQ ID NO:5) The variable light complementary determining region 2 (VL CDR2) of base acid sequence and the amino for containing QSYDGSNRWM (SEQ ID NO:6) The variable light complementary determining region 3 (VL CDR3) of acid sequence.

67. the unit dose vial of claim 66, wherein the antibody includes the weight of the amino acid sequence of SEQ ID NO:47 The light chain variable amino acid sequence of the amino acid sequence of chain variable amino acid sequence and SEQ ID NO:48.