WO2022245816A1 - Treatment of autoimmune inflammatory diseases with tnfrsf25-binding agents - Google Patents

Treatment of autoimmune inflammatory diseases with tnfrsf25-binding agents Download PDF

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Publication number
WO2022245816A1
WO2022245816A1 PCT/US2022/029622 US2022029622W WO2022245816A1 WO 2022245816 A1 WO2022245816 A1 WO 2022245816A1 US 2022029622 W US2022029622 W US 2022029622W WO 2022245816 A1 WO2022245816 A1 WO 2022245816A1
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Prior art keywords
patient
tnfrsf25
treatment
seq
antigen binding
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PCT/US2022/029622
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French (fr)
Inventor
Matthew M. SEAVEY
Rahul R. JASUJA
Matthew Clark
Eric P. Dixon
Vikas TAHILIANI
Charles KROGER
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Pelican Therapeutics, Inc.
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Publication of WO2022245816A1 publication Critical patent/WO2022245816A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2878—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/564—Immunoassay; Biospecific binding assay; Materials therefor for pre-existing immune complex or autoimmune disease, i.e. systemic lupus erythematosus, rheumatoid arthritis, multiple sclerosis, rheumatoid factors or complement components C1-C9
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/55—Medicinal preparations containing antigens or antibodies characterised by the host/recipient, e.g. newborn with maternal antibodies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/70—Immunoglobulins specific features characterized by effect upon binding to a cell or to an antigen
    • C07K2317/75—Agonist effect on antigen

Definitions

  • the disclosure is directed to treatment and prevention of autoimmune-related diseases and disorders using TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibodies.
  • TNF Receptor Superfamily Member 25 TNFRSF25
  • MS multiple sclerosis
  • myelin layer covers and protects neurons resulting in motor function impairment ⁇ e.g. coordination, balance, speech and vision
  • motor function impairment ⁇ e.g. coordination, balance, speech and vision
  • MS disease modifying therapies
  • DMTs disease modifying therapies
  • Many of the current pharmacological agents for MS are limited by incomplete efficacy, side effects and medical risks. These treatments have been shown to modestly reduce neurological relapses of the disease and, in some instances, incompletely slow the progression of neurological disability. Accordingly, there remains a pressing need for improved MS treatments.
  • the present disclosure relates to a method for treating or preventing an autoimmune inflammatory disease, comprising administering an effective amount of TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibody or antigen binding fragment thereof to a patient in need thereof.
  • the autoimmune inflammatory disease is a demyelinating disease.
  • the inflammation is in the central nervous system (CNS).
  • the demyelinating disease is multiple sclerosis (MS).
  • the demyelinating disease is clinically isolated syndrome (CIS).
  • the method prevents or slows the progression of CIS to MS.
  • the demyelinating disease is radiologically isolated syndrome (RIS).
  • the method prevents or slows the progression of RIS to MS.
  • the patient is likely to develop MS.
  • the patient has CIS.
  • the patient has a monofocal episode of CIS.
  • the patient has optic neuritis. In embodiments, the patient has a multifocal episode of CIS. In embodiments, the patient has optic neuritis and numbness or tingling in the legs.
  • the patient has CIS and one or more magnetic resonance imaging (MRI)- detected brain lesions.
  • MRI magnetic resonance imaging
  • the treatment occurs before or early in disease progression. In embodiments, the treatment occurs before the onset of relapsing-remitting MS (RRMS). In embodiments, the treatment occurs before the onset of secondary progressive MS (SPMS). In embodiments, the treatment occurs before the onset of primary progressive MS (PPMS).
  • RRMS relapsing-remitting MS
  • SPMS secondary progressive MS
  • PPMS primary progressive MS
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof is used to treat or prevent an autoimmune inflammatory disease selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti-MAG peripheral neuropathy, Charcot-Marie-Tooth disease, hereditary neuropathy with liability to pressure palsy, peripheral neuropathy, optic neuropathy, and progressive inflammatory neuropathy.
  • an autoimmune inflammatory disease selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating poly
  • the treatment expands and/or selectively activates a population of Tregs in the patient.
  • the treatment does not substantially expand and/or selectively activate a population of cytotoxic T cells (Tc cells) and/or helper T cells (Th cells) in the patient.
  • Tc cells cytotoxic T cells
  • Th cells helper T cells
  • the treatment does not substantially expand and/or selectively activate a population of Th17 cells in the patient.
  • the treatment increases the ratio of Tregs to Tc and/or Th cells in the patient.
  • the treatment increases the ratio of Tregs to Th17 cells in the patient.
  • the treatment reduces spinal cord inflammation, as compared to before treatment. In embodiments, the treatment reduces axon demyelination, as compared to before treatment.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises: a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 sequences, wherein the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO: 1), the heavy chain CDR2 sequence is YISSASGUSYADAVRG (SEQ ID NO: 2); and the heavy chain CDR3 sequence is DPAYTGLYALDF (SEQ ID NO: 3) or DPPYSGLYALDF (SEQ ID NO: 4); and a light chain variable region comprising light chain CDR1, CDR2, and CDR3 sequences, wherein the light chain CDR1 sequence is TLSSELSWYTIV (SEQ ID NO: 5), the light chain CDR2 sequence is LKSDGSHSKGD (SEQ ID NO: 6), and the light chain CDR3 sequence is CGAGYTLAGQY G WV (SEQ ID NO: 7).
  • the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO:
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises variable region framework (FW) sequences juxtaposed between the CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4), wherein the variable region FW sequences in the heavy chain variable region are heavy chain variable region FW sequences, and wherein the variable region FW sequences in the light chain variable region are light chain variable region FW sequences.
  • the variable region FW sequences are human.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises human heavy chain and light chain constant regions.
  • the constant regions are selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4.
  • the constant regions are lgG1.
  • the constant regions are lgG4.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTI SRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85 to about 99% identity thereto.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence
  • QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85 to about 99% identity thereto.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises 4C12, or a humanized version, or fragment thereof.
  • the treatment occurs in combination with one or more disease modifying therapies. In embodiments, the treatment occurs in combination with one or more agents of Table 1.
  • the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining an amount of ICOS protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of ICOS relative to an untreated and/or undiseased patient.
  • the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining an amount of CD103 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of CD103 relative to an untreated and/or undiseased patient.
  • the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining an amount of Tim3 and/or LAG3 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of Tim3 and/or LAG3 relative to an untreated and/or undiseased patient.
  • FIG. 1 is an image showing an overview of PTX-35 treatment during an autoimmunity disease. A treatment timeline is also shown in FIG. 1.
  • FIG. 2 is an image showing the study design.
  • FIG. 3 is a graph showing the body weight of the mice before disease induction. mPTX-35 expanded Tregs prior to model start. The measurements in FIG. 3 show body weight (g) on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice. G4 mPTX-35 was administered on both day -7 and day 7.
  • FIG. 4 is a graph showing the body weight of the mice during disease induction.
  • mPTX-35 expanded conventional T cells (“Tconv”) at model start.
  • the measurements in FIG. 4 show body weight (g) on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice.
  • G5 mPTX-35 was administered on both day 0 and day 14.
  • FIG. 5 is a graph showing the body weight of the mice after disease induction.
  • mPTX-35 expanded both Tregs and Tconv during model disease.
  • the measurements in FIG. 5 show body weight (g) on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • G6 mPTX-35 was administered on both day 7 and day 21.
  • FIG. 6 is a chart showing the clinical assessment of EAE with respect to scores and clinical signs.
  • FIG. 7 is a graph showing the clinical score of the mice before disease induction.
  • the measurements in FIG. 7 show the average clinical score on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice.
  • G4 mPTX-35 was administered on both day -7 and day 7.
  • FIG. 8 is a graph showing the clinical score of the mice during disease induction.
  • the measurements in FIG. 8 show the average clinical score on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice.
  • G5 mPTX-35 was administered on both day 0 and day 14.
  • FIG. 9 is a graph showing the clinical score of the mice after disease induction.
  • the measurements in FIG. 9 show the average clinical score on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • G6 mPTX-35 was administered on both day 7 and day 21.
  • FIG. 10 is a graph showing the disease incidence in the mice before disease induction.
  • the measurements in FIG. 10 show the percent incidence on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice.
  • G4 mPTX-35 was administered on both day -7 and day 7.
  • FIG. 11 is a graph showing the disease incidence in the mice during disease induction.
  • the measurements in FIG. 11 show the percent incidence on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice.
  • G5 mPTX-35 was administered on both day 0 and day 14.
  • FIG. 12 is a graph showing the disease incidence in the mice after disease induction.
  • the measurements in FIG. 12 show the percent incidence on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • G6 mPTX-35 was administered on both day 7 and day 21.
  • FIG. 13 is a graph showing the clinical score area under the curve (“AUC”) measurements for the G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • AUC clinical score area under the curve
  • FIG. 14 is a graph showing percent survival (x-axis) and days elapsed (y-axis) before disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice.
  • FIG. 15 is a graph showing percent survival (x-axis) and days elapsed (y-axis) during disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice.
  • FIG. 16 is a graph showing percent survival (x-axis) and days elapsed (y-axis) after disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • FIG. 17 is a chart showing the evaluation of disease progression as it relates to the spinal cord.
  • FIG. 18 is a graph showing the histology inflammation score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-353 mg/kg), G5 (mPTX-353 mg/kg), and G6 (mPTX-353 mg/kg) mice.
  • PTX-35 mediated Treg expansion reduced central nervous system pathologies.
  • FIG. 19 is a graph showing the histology lesion score for G2 (vehicle), G3 (Dex 1 mg/kg),
  • FIG. 20 is a graph showing the histology demyelination score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • FIG.21 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G1 normal mice (left panel), and G2 vehicle mice (right panel).
  • HE hematoxylin and eosin
  • FIG.22 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G3 Dex treated mice (left panel), and G4 mPTX-35 treated mice (right panel).
  • HE hematoxylin and eosin
  • FIG.23 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G5 mPTX-35 treated mice (left panel), and G6 mPTX-35 treated mice (right panel).
  • HE hematoxylin and eosin
  • FIG. 24 shows the gating strategy for the Flow cytometry data.
  • FIG. 25 is a graph showing the percentage of regulatory T cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • FIG. 26 is a graph showing the percentage of Th17 cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-353 mg/kg), G5 (mPTX- 35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • FIG. 27A - FIG. 27B show experimental design of PTX-35 treatment of EAE mouse model disease progression.
  • FIG. 27A is an image showing the study design of C57BL/6 mice immunized with MOG 35-55 and treated with PTX-35 at day 4 and 8, and day 12 and 16.
  • FIG. 27B is a graph showing the paralytic score of MOG 35-55 immunized mice treated with PTX-35 at various days post immunization compared to control mice.
  • FIG. 28A - FIG. 28B show experimental data demonstrating the weight changes of MOG
  • FIG. 28A is a graph showing the weight change in grams of MOG 35-55 immunized mice treated with PTX- 35 at various days post immunization.
  • FIG. 28B is a graph showing change in the percent of initial body weight of MOG 35-55 immunized mice treated with PTX-35 at various days post immunization.
  • FIG. 29 shows images of spinal cords from B6 mice immunized with MOG 35-55 taken at
  • FIG. 30 is a graph showing inflammatory scoring of B6 mice immunized with MOG 35-55 take at Day 30 post immunization compared to non-immunized mice and mice immunized with isotype control antibodies. Significance relative to isotype (* ⁇ 0.05, ** ⁇ 0.005, *** ⁇ 0.0005).
  • FIG. 31 is a flow cytometry histogram showing percentage distribution of FoxP3+ regulatory
  • MOG 35-55 immunized mice treated with PBS MOG 35-55 immunized mice treated with isotype control antibodies
  • MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8 MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 32 is a graph of experimental data showing the percentage of FoxP3+ regulatory T cells and conventional CD4+T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (* ⁇ 0.05, ** ⁇ 0.005, *** ⁇ 0.0005).
  • FIG. 33 is a flow cytometry histogram showing percentage of ICOS+T cells in splenic CD4+
  • MOG 35-55 immunized mice treated with PBS MOG 35- 55 immunized mice treated with isotype control antibodies
  • MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8 MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 34 is a graph of experimental data showing percentage of ICOS+ T cells in splenic
  • CD4+FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (* ⁇ 0.05, ** ⁇ 0.005, *** ⁇ 0.0005).
  • FIG. 35 is a flow cytometry histogram showing percentage of CD103+ T cells in splenic
  • CD4+ FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 36 is a graph of experimental data showing percentage of CD103+ T cells in splenic
  • CD4+FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (* ⁇ 0.05, ** ⁇ 0.005, *** ⁇ 0.0005).
  • FIG. 37 is a flow cytometry histogram showing percentage distribution of Tim3+LAG3+ T cells in splenic CD8+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 38 is a graph of experimental data showing the percentage of Tim3+LAG3+ T cells in splenic CD8+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (* ⁇ 0.05, ** ⁇ 0.005, *** ⁇ 0.0005).
  • FIG. 39 is a graph of experimental data showing percentage of CD44+ T cells in splenic
  • CD4+FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 40 is a graph of experimental data showing percentage of TIGIT+ T cells in splenic
  • CD4+FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 41 is a graph of experimental data showing percentage of CTLA4+ T cells in splenic
  • CD4+FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 42 is a graph of experimental data showing percentage of Ki67+ T cells in splenic
  • CD4+FoxP3+ regulatory T cells of non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 44 is a graph of experimental data showing MFI of CD25 in splenic CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 45 is a graph of experimental data showing the percentage of PD-1 + T cells in splenic
  • CD8+T cells in non-immunized mice MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 46 is a graph of experimental data showing the increase in percentage of PD-1+T cells in a total population of T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
  • FIG. 47 is an image showing the study design of SJL/J mice immunized with PLP 139-151 and treated with PTX-35 at day -4 and 0, day 4 and 8, day 12 and 16, or day 16 and 20.
  • FIG. 48 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
  • FIG. 49 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
  • FIG. 50 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
  • PTX-35 at day 12 and day 16 post immunization compared to control mice.
  • FIG. 51 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with PTX-35 at day 16 and day 20 post immunization compared to control mice.
  • FIG. 52 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
  • PTX-35 at day 4, day 8, day 16, and day 20 post immunization compared to control mice.
  • the present disclosure relates to compositions and methods with TNFRSF25 agonistic antibodies, including their use in the treatment of various autoimmune and/or neurodegenerative diseases.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a TNFRSF25 agonistic antibody or antigen binding fragment, such as PTX-35, which is described in PCT/US2017/036817 (WO2017214547), which is incorporated herein by reference in its entirety.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof can comprise (i) a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 sequences, wherein the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO: 1), the heavy chain CDR2 sequence is YISSASGUSYADAVRG (SEQ ID NO: 2); and the heavy chain CDR3 sequence is DPAYTGLYALDF (SEQ ID NO: 3) or DPPYSGLYALDF (SEQ ID NO: 4); and (ii) a light chain variable region comprising light chain CDR1, CDR2, and CDR3 sequences, wherein the light chain CDR1 sequence is TLSSELSWYTIV (SEQ ID NO: 5), the light chain CDR2 sequence is LKSDGSHSKGD (SEQ ID NO: 6), and the light chain CDR3 sequence is CGAGYTLAGQYGW
  • heavy chain CDR1 , CDR2, and CDR3 sequences and/or light chain CDR1 , CDR2, and CDR3 sequences each include at least one, or at least 2, or at least 3, or at least 4, or at least 5 mutations such as amino acid substitutions.
  • heavy chain CDR1 , CDR2, and CDR3 sequences and/or light chain CDR1 , CDR2, and CDR3 sequences each include more than 5 mutations.
  • an TNFRSF25 agonistic antibody includes a set of six CDRs that include no more than two, or no more than three, or no more than four, or no more than five, or no more than six total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 1, 2, 3 or 4, 5, 6, and 7.
  • an TNFRSF25 agonistic antibody includes a set of six CDRs that include at least one, or at least two, or at least three, or at least four, or at least five total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 1 , 2, 3 or 4, 5, 6, and 7.
  • an TNFRSF25 agonistic antibody includes a set of six CDRs that include one, or two, or three, or four, or five, or more than five total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEC ID NOS: 1, 2, 3 or 4, 5, 6, and 7.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises variable region framework (FW) sequences juxtaposed between the CDRs according to the formula (FW1 )-(CDR1 )-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4), wherein the variable region FW sequences in the heavy chain variable region are heavy chain variable region FW sequences, and wherein the variable region FW sequences in the light chain variable region are light chain variable region FW sequences.
  • the variable region FW sequences are human.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises human heavy chain and light chain constant regions.
  • the constant regions can be selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4. For instance, In embodiments, the constant regions are lgG1. In embodiments, the constant regions are lgG4.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence
  • QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85% to about 99% identity thereto.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence
  • EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTI SRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91 %, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99% identity thereto.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91 %, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99% identity thereto.
  • a heavy chain variable region has an amino acid sequence of SEQ ID NO: 1
  • a heavy chain variable region polypeptide can contain 24 or less (e.g., 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, ten, nine, eight, seven, six, five, four, three, two, or one) amino acid substitution as compared to SEQ ID NO: 8, or an antigen binding fragment thereof.
  • a light chain variable region has an amino acid sequence of SEQ ID NO: 1
  • a light chain variable region polypeptide can contain 23 or less (e.g., 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, ten, nine, eight, seven, six, five, four, three, two, or one) amino acid substitutions as compared to SEQ ID NO: 9, or an antigen binding fragment thereof.
  • TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises is the 4C12 antibody or antigen binding fragment thereof. In embodiments TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises is a humanized 4C12.
  • an TNFRSF25 agonistic antibody or antigen binding fragment thereof is
  • an TNFRSF25 agonistic antibody comprises (i) a heavy chain variable region sequence comprising the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEAS GFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTISRDNAKNSLFLQMNNLKSEDTAMYY CARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 10) or the amino acid sequence of SEQ ID NO: 10 with no more than 12 total amino acid substitutions (e.g., no more than 12, or no more than 11, or no more than ten, or no more than nine, or no more than eight, or no more than seven, or no more than six, or no more than five, or no more than four, or no more than three, or no more than two total
  • SEQ ID NO: 11 or the amino acid sequence of SEQ ID NO: 11 with no more than 11 total amino acid substitutions (e.g., no more than 11, or no more than ten, or no more than nine, or no more than eight, or no more than seven, or no more than six, or no more than five, or no more than four, or no more than three, or no more than two total amino acid substitutions, or no more than one total amino acid substitution).
  • An amino acid substitution refers to the replacement of one amino acid residue with another amino acid in a peptide sequence.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEAS
  • GFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTISRDNAKNSLFLQMNNLKSEDTAMYY CARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 10) or an amino acid sequence of at least about 85 to about 99% (e.g., an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91%, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99%) identity thereto.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 11), or an amino acid sequence of at least about 85 to about 99% (e.g., an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91 %, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99%) identity thereto.
  • an TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises an isolated heavy chain variable region polypeptide that binds specifically to TNFRSF25, where the polypeptide includes heavy chain CDR1, CDR2, and CDR3 sequences, where the CDR1 sequence is GFTFSNHDLN (SEQ ID NO: 12), the CDR2 sequence is YISSASGLISYADAVRG (SEQ ID NO: 13); and (c) the CDR3 sequence is DPPYSGLYALDF (SEQ ID NO: 14).
  • the isolated heavy chain variable region polypeptide can further include variable region heavy chain framework (FW) sequences juxtaposed between the heavy chain CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4).
  • the heavy chain framework sequences can be human.
  • the isolated heavy chain variable region polypeptide can be combination with a light chain variable region polypeptide.
  • the light chain variable region polypeptide comprises light chain CDR1, CDR2, and CDR3 sequences, wherein the CDR1 sequence is TLSSELSSYTIV (SEQ ID NO: 15), the CDR2 sequence is LKSDGSHSKGD (SEQ ID NO: 16), and the CDR3 sequence is GAGYTLAGQYGWV (SEQ ID NO: 17).
  • the TNFRSF25 agonistic antibody can be a humanized monoclonal antibody that specifically binds to TNFRSF25.
  • an TNFRSF25 agonistic antibody includes a set of six CDRs that include no more than two, or no more than three, or no more than four, or no more than five, or no more than six total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 12, 13, 14, 15, 16, and 17.
  • an TNFRSF25 agonistic antibody includes a set of six CDRs that include at least one, or at least two, or at least three, or at least four, or at least five, or at least 6 total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 12, 13, 14, 15, 16, and 17.
  • an TNFRSF25 agonistic antibody includes a set of six CDRs that include one, or two, or three, or four, or five, or more than five total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 12, 13, 14, 15, 16, and 17.
  • the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain comprising an amino acid of SEQ ID NO: 18, 19, 20, or 21, and a light chain comprising an amino acid of SEQ ID NO: 22 or 23, as follows:
  • an TNFRSF25 agonistic antibody or antigen binding fragment thereof is any of the antibodies or antibody fragments (or combinations thereof) described in PCT/US2015/061082 (WO2016081455).
  • variable region light chain framework (FW) sequences can be juxtaposed between the light chain CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)- (CDR3)-(FW4).
  • the light chain framework sequences can be human.
  • the antibody or antigen binding fragment can further include a human constant region (e.g., a constant region selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4), or a murine constant region (e.g., a constant region selected from the group consisting of murine lgG1, lgG2A, lgG2B, and lgG3).
  • a human constant region e.g., a constant region selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4
  • a murine constant region e.g., a constant region selected from the group consisting of murine lgG1, lgG2A, lgG2B, and lgG3
  • an TNFRSF25 agonistic antibody or antigen binding fragment thereof is as described in PCT/US2010/044218 (WQ2011017303), which is incorporated herein by reference in its entirety.
  • Embodiments of the present disclosure make use of TNFRSF25 agonistic antibodies or antigen binding fragment thereof.
  • the antibody is an antibody (e.g., human, hamster, feline, mouse, cartilaginous fish, or camelid antibodies), and any derivative or conjugate thereof, that specifically binds to TNFRSF25.
  • Non-limiting examples of antibodies include monoclonal antibodies, polyclonal antibodies, humanized antibodies, multi-specific antibodies (e.g., bi-specific antibodies), single-chain antibodies (e.g., single-domain antibodies, camelid antibodies, and cartilaginous fish antibodies), chimeric antibodies, feline antibodies, and felinized antibodies.
  • Monoclonal antibodies are homogeneous populations of antibodies to a particular epitope of an antigen.
  • Polyclonal antibodies are heterogeneous populations of antibody molecules that are contained in the sera of the immunized animals.
  • An isolated polypeptide can yield a single major band on a non-reducing polyacrylamide gel.
  • An isolated polypeptide can be at least about 75% pure ⁇ e.g., at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% pure).
  • Isolated polypeptides can be obtained by, for example, extraction from a natural source, by chemical synthesis, or by recombinant production in a host cell or transgenic plant, and can be purified using, for example, affinity chromatography, immunoprecipitation, size exclusion chromatography, and ion exchange chromatography. The extent of purification can be measured using any appropriate method, including, without limitation, column chromatography, polyacrylamide gel electrophoresis, or high- performance liquid chromatography.
  • an antigen binding fragment that specifically binds to TNFRSF25 is provided.
  • Such antigen binding fragment in embodiments, is any portion of a full-length antibody that contains at least one variable domain (e.g., a variable domain of a mammalian (e.g., feline, human, hamster, or mouse) heavy or light chain immunoglobulin, a camelid variable antigen binding domain (VHH), or a cartilaginous fish immunoglobulin new antigen receptor (Ig-NAR) domain) that is capable of specifically binding to an antigen.
  • a variable domain of a mammalian (e.g., feline, human, hamster, or mouse) heavy or light chain immunoglobulin, a camelid variable antigen binding domain (VHH), or a cartilaginous fish immunoglobulin new antigen receptor (Ig-NAR) domain that is capable of specifically binding to an antigen.
  • Non-limiting examples of antibody fragments include Fab, Fab’, F(ab’)2, and Fv fragments, diabodies, linear antibodies, and multi-specific antibodies formed from antibody fragments. Additional antibody fragments containing at least one camelid VHH domain or at least one cartilaginous fish Ig-NAR domain include mini-bodies, micro-antibodies, subnano-antibodies, and nano-antibodies, and any of the other forms of antibodies described, for example, in U.S. Publication No. 2010/0092470.
  • An antibody can be of the IgA-, IgD-, IgE, IgG- or IgM-type, including IgG- or IgM-types such as, without limitation, lgG1-, lgG2-, lgG3-, lgG4-, lgM1- and lgM2-types.
  • the antibody is of the lgG1-, lgG2- or lgG4- type.
  • antibodies as provided herein can be fully human or humanized antibodies.
  • the human antibody is an antibody that is encoded by a nucleic acid (e.g., a rearranged human immunoglobulin heavy or light chain locus) present in the genome of a human.
  • a human antibody can be produced in a human cell culture (e.g., feline hybridoma cells).
  • a human antibody can be produced in a non-human cell (e.g., a mouse or hamster cell line).
  • a human antibody can be produced in a bacterial or yeast cell.
  • Human antibodies can avoid certain problems associated with xenogeneic antibodies, such as antibodies that possess murine or rat variable and/or constant regions. For example, because the effector portion is human, it can interact better with other parts of the human immune system, e.g., to destroy target cells more efficiently by complement-dependent cytotoxicity or antibody-dependent cellular cytotoxicity. In addition, the human immune system should not recognize the antibody as foreign. Further, half-life in human circulation will be similar to naturally occurring human antibodies, allowing smaller and less frequent doses to be given. Methods for preparing human antibodies are known in the art.
  • the antibody is a humanized antibody, e.g., an antibody that contains minimal sequence derived from non-human ⁇ e.g., mouse, hamster, rat, rabbit, or goat) immunoglobulin.
  • Humanized antibodies generally are chimeric or mutant monoclonal antibodies from mouse, rat, hamster, rabbit or other species, bearing human constant and/or variable region domains or specific changes.
  • humanized antibodies are human antibodies (recipient antibody) in which hypervariable region (HVR) residues of the recipient antibody are replaced by HVR residues from a non-human species (donor) antibody, such as a mouse, rat, rabbit, or goat antibody having the desired specificity, affinity, and capacity.
  • HVR hypervariable region
  • Fv framework residues of the human immunoglobulin can be replaced by corresponding non-human residues.
  • humanized antibodies can contain residues that are not found in the recipient antibody or in the donor antibody. Such modifications can be made to refine antibody performance, for example.
  • a humanized antibody can contain substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops (CDRs) correspond to those of a non-human immunoglobulin, while all or substantially all of the framework regions are those of a human immunoglobulin sequence.
  • a humanized antibody also can contain at least a portion of an immunoglobulin constant (Fc) region, typically that of a human immunoglobulin.
  • a humanized antibody or antigen binding fragment as provided herein can have reduced or minimal effector function (e.g., as compared to corresponding non-humanized antibody), such that it does not stimulate effector cell action to the same extent that a corresponding non-humanized antibody would.
  • DNA sequences encoding antigen binding portions or CDRs of murine monoclonal antibodies can be grafted by molecular means into DNA sequences encoding frameworks of human antibody heavy and light chains (Jones et al., Nature 321:522, 1986; and Riechmann et al., Nature 332:323, 1988). Expressed recombinant products are called “reshaped” or humanized antibodies, and contain the framework of a human antibody light or heavy chain and antigen recognition portions, CDRs, of a murine monoclonal antibody.
  • the antibody is a single-chain antibody, e.g. a single polypeptide that contains at least one variable binding domain (e.g., a variable domain of a mammalian heavy or light chain immunoglobulin, a camelid VHH, or a cartilaginous fish (e.g., shark) Ig-NAR domain) that is capable of specifically binding to an antigen.
  • variable binding domain e.g., a variable domain of a mammalian heavy or light chain immunoglobulin, a camelid VHH, or a cartilaginous fish (e.g., shark) Ig-NAR domain
  • single-chain antibodies include single-domain antibodies.
  • the antibody is a single-domain antibody, e.g. a polypeptide that contains one camelid VHH or at least one cartilaginous fish Ig-NAR domain that is capable of specifically binding to an antigen.
  • single-domain antibodies are described, for example, in U.S. Publication No. 2010/0092470.
  • the antibody specifically binds to a particular antigen, e.g., TNFRSF25, when it binds to that antigen in a sample, and does not recognize and bind, or recognizes and binds to a lesser extent, other molecules in the sample.
  • an antibody or an antigen binding fragment thereof can selectively bind to an epitope with an affinity (Kd) equal to or less than, for example, about 1 x 10 6 M ⁇ e.g., equal to or less than about 1 x 10 9 M, equal to or less than about 1 x 10 10 M, equal to or less than about 1 x 10 11 M, or equal to or less than about 1 x 10 12 M) in phosphate buffered saline.
  • Kd affinity
  • an antibody or antigen binding fragment to specifically bind a protein epitope can be determined using any of the methods known in the art or those methods described herein (e.g., by Biacore/Surface Plasmon Resonance). This can include, for example, binding to TNFRSF25 on live cells as a method to stimulate caspase activation in live transformed cells, binding to an immobilized target substrate including human TNFRSF25 fusion proteins as detected using an ELISA method, binding to TNFRSF25 on live cells as detected by flow cytometry, or binding to an immobilized substrate by surface plasmon resonance (including ProteOn).
  • Antibodies having specific binding affinity for TNFRSF25 can be produced using standard methods.
  • a TNFRSF25 polypeptide can be recombinantly produced, purified from a biological sample ⁇ e.g., a heterologous expression system), or chemically synthesized, and used to immunize host animals, including rabbits, chickens, mice, guinea pigs, or rats.
  • Various adjuvants that can be used to increase the immunological response depend on the host species and include Freund’s adjuvant (complete and incomplete), mineral gels such as aluminum hydroxide, surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanin and dinitrophenol.
  • Monoclonal antibodies can be prepared using aTNFRSF25 polypeptide and standard hybridoma technology. In particular, monoclonal antibodies can be obtained by any technique that provides for the production of antibody molecules by continuous cell lines in culture such as described by Kohler et al. ⁇ Nature 256:495, 1975), the human B-cell hybridoma technique of Kosbor et al.
  • Such antibodies can be of any immunoglobulin class including IgG, IgM, IgE, IgA, IgD, and any subclass thereof.
  • the hybridoma producing the monoclonal antibodies can be cultivated in vitro and in vivo.
  • amino acid substitutions can be made by selecting conservative substitutions that do not differ significantly in their effect on maintaining (a) the structure of the peptide backbone in the area of the substitution, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain.
  • residues can be divided into groups based on side-chain properties: (1) hydrophobic amino acids (norleucine, methionine, alanine, valine, leucine, and isoleucine); (2) neutral hydrophilic amino acids (cysteine, serine, and threonine); (3) acidic amino acids (aspartic acid and glutamic acid); (4) basic amino acids (asparagine, glutamine, histidine, lysine, and arginine); (5) amino acids that influence chain orientation (glycine and proline); and (6) aromatic amino acids (tryptophan, tyrosine, and phenylalanine). Substitutions made within these groups can be considered conservative substitutions.
  • Non-limiting examples of conservative substitutions include, without limitation, substitution of valine for alanine, lysine for arginine, glutamine for asparagine, glutamic acid for aspartic acid, serine for cysteine, asparagine for glutamine, aspartic acid for glutamic acid, proline for glycine, arginine for histidine, leucine for isoleucine, isoleucine for leucine, arginine for lysine, leucine for methionine, leucine for phenylalanine, glycine for proline, threonine for serine, serine for threonine, tyrosine for tryptophan, phenylalanine for tyrosine, and/or leucine for valine.
  • an amino acid substitution can be nonconservative, such that a member of one of the amino acid classes described above is exchanged for a member of another class.
  • compositions for the present methods of treating which include an antibody or antigen binding fragment, as described herein, in combination with a pharmaceutically acceptable carrier.
  • a “pharmaceutically acceptable carrier” (also referred to as an “excipient” or a “carrier”) is a pharmaceutically acceptable solvent, suspending agent, stabilizing agent, or any other pharmacologically inert vehicle for delivering one or more therapeutic compounds to a subject ⁇ e.g., a mammal, such as a human, non-human primate, dog, cat, sheep, pig, horse, cow, mouse, rat, or rabbit), which is nontoxic to the cell or subject being exposed thereto at the dosages and concentrations employed.
  • Pharmaceutically acceptable carriers can be liquid or solid, and can be selected with the planned manner of administration in mind so as to provide for the desired bulk, consistency, and other pertinent transport and chemical properties, when combined with one or more of therapeutic compounds and any other components of a given pharmaceutical composition.
  • Typical pharmaceutically acceptable carriers that do not deleteriously react with amino acids include, by way of example and not limitation: water, saline solution, binding agents ⁇ e.g., polyvinylpyrrolidone or hydroxypropyl methylcell ulose), fillers ⁇ e.g., lactose and other sugars, gelatin, or calcium sulfate), lubricants ⁇ e.g., starch, polyethylene glycol, or sodium acetate), disintegrates ⁇ e.g., starch or sodium starch glycolate), and wetting agents ⁇ e.g., sodium lauryl sulfate).
  • Pharmaceutically acceptable carriers also include aqueous pH buffered solutions or liposomes (small vesicles composed of various types of lipids, phospholipids and/or surfactants which are useful for delivery of a drug to a mammal).
  • Further examples of pharmaceutically acceptable carriers include buffers such as phosphate, citrate, and other organic acids, antioxidants such as ascorbic acid, low molecular weight (less than about 10 residues) polypeptides, proteins such as serum albumin, gelatin, or immunoglobulins, hydrophilic polymers such as polyvinylpyrrolidone, amino acids such as glycine, glutamine, asparagine, arginine or lysine, monosaccharides, disaccharides, and other carbohydrates including glucose, mannose or dextrins, chelating agents such as EDTA, sugar alcohols such as mannitol or sorbitol, salt-forming counterions such as sodium, and/or nonionic surfactants such as TWEENTM,
  • compositions can be formulated by mixing one or more active agents with one or more physiologically acceptable carriers, diluents, and/or adjuvants, and optionally other agents that are usually incorporated into formulations to provide improved transfer, delivery, tolerance, and the like.
  • a pharmaceutical composition can be formulated, e.g., in lyophilized formulations, aqueous solutions, dispersions, or solid preparations, such as tablets, dragees or capsules.
  • a multitude of appropriate formulations can be found in the formulary known to all pharmaceutical chemists: Remington’s Pharmaceutical Sciences (18 th ed, Mack Publishing Company, Easton, PA (1990)), particularly Chapter 87 by Block, Lawrence, therein.
  • formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid (cationic or anionic) containing vesicles (such as LIPOFECTINTM), DNA conjugates, anhydrous absorption pastes, oil-in-water and water-in-oil emulsions, emulsions carbowax (polyethylene glycols of various molecular weights), semi-solid gels, and semi-solid mixtures containing carbowax. Any of the foregoing mixtures may be appropriate in treatments and therapies as described herein, provided that the active agent in the formulation is not inactivated by the formulation and the formulation is physiologically compatible and tolerable with the route of administration.
  • compositions include, without limitation, solutions, emulsions, aqueous suspensions, and liposome-containing formulations. These compositions can be generated from a variety of components that include, for example, preformed liquids, self-emulsifying solids and self-emulsifying semisolids.
  • Emulsions are often biphasic systems comprising of two immiscible liquid phases intimately mixed and dispersed with each other; in general, emulsions are either of the water-in-oil (w/o) or oil-in-water (o/w) variety. Emulsion formulations have been widely used for oral delivery of therapeutics due to their ease of formulation and efficacy of solubilization, absorption, and bioavailability.
  • compositions and formulations can contain sterile aqueous solutions, which also can contain buffers, diluents and other suitable additives ⁇ e.g., penetration enhancers, carrier compounds and other pharmaceutically acceptable carriers).
  • Compositions additionally can contain other adjunct components conventionally found in pharmaceutical compositions.
  • the compositions also can include compatible, pharmaceutically active materials such as, for example, antipruritics, astringents, local anesthetics or antiinflammatory agents, or additional materials useful in physically formulating various dosage forms of the compositions provided herein, such as dyes, flavoring agents, preservatives, antioxidants, opacifiers, thickening agents and stabilizers.
  • compositions can be mixed with auxiliary agents, e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings, and aromatic substances.
  • auxiliary agents e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings, and aromatic substances.
  • auxiliary agents e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings, and aromatic substances.
  • a composition containing an antibody or antigen binding fragment as used herein can be in the form of a solution or powder with or without a diluent to make an injectable suspension.
  • the composition may contain additional ingredients including, without limitation, pharmaceutically acceptable vehicles, such as saline, water, lactic acid, mannitol, or combinations thereof, for example.
  • Administration can be, for example, parenteral ⁇ e.g., by subcutaneous, intrathecal, intraventricular, intramuscular, or intraperitoneal injection, or by intravenous drip). Administration can be rapid (e.g., by injection) or can occur over a period of time (e.g., by slow infusion or administration of slow release formulations). In embodiments, administration can be topical (e.g., transdermal, sublingual, ophthalmic, or intranasal), pulmonary (e.g., by inhalation or insufflation of powders or aerosols), or oral. In addition, a composition containing an antibody or antigen binding fragment as described herein can be administered prior to, after, or in lieu of surgical resection of a tumor.
  • a composition containing an TNFRSF25 agonistic antibody or antigen binding fragment thereof can be administered to a mammal in any appropriate amount, at any appropriate frequency, and for any appropriate duration effective to achieve a desired outcome.
  • an TNFRSF25 agonistic antibody or antigen binding fragment thereof can be administered to a subject in an amount effective to stimulate proliferation of T cells in vitro or in vivo (e.g., human, murine, hamster, or macaque T cells, including CD8 + T cells and/or CD4 + FoxP3 + regulatory T cells), to stimulate apoptosis of tumor cells that express TNFRSF25, to reduce tumor size, or to increase progression-free survival of a cancer patient.
  • an TNFRSF25 agonistic antibody or antigen binding fragment thereof can be administered at a dosage of about 0.1 mg/kg to about 10 mg/kg (e.g., about 0.1 mg/kg to about 1 mg/kg, about 1 mg/kg to about 5 mg/kg, or about 5 mg/kg to about 10 mg/kg), and can be administered once every one to three weeks (e.g., every week, every 10 days, every two weeks, or every three weeks).
  • a composition containing TNFRSF25 agonistic antibody or antigen binding fragment thereof as described herein can be administered to a subject in an amount effective to increase proliferation of T cells (e.g., by at least about 10 percent, about 20 percent, about 25 percent, about 50 percent, about 60 percent, about 70 percent, about 75 percent, about 80 percent, about 90 percent, about 100 percent, or more than 100 percent), as compared to the “baseline” level of T cell proliferation in the subject prior to administration of the composition, or as compared to the level of T cell proliferation in a control subject or population of subjects to whom the composition was not administered.
  • the T cells can be, for example, CD4 + FoxP3 + T cells, regulatory T cells.
  • Any suitable method can be used to determine whether or not the level of T cell proliferation is increased in the subject.
  • Such methods can include, without limitation, flow cytometry analysis of antigen specific T cells (e.g., flow cytometry analysis of the proportion of antigen specific CD4+FoxP3+ T cells as a fraction of the total CD4 + T cell pool), analysis of cell proliferation markers (e.g., expression of Ki67) in CD4 + T cells, increased counts of CD4 + T cells, or increased proportions of individual TCR sequences of a particular clone of CD4 + T cells.
  • flow cytometry analysis of antigen specific T cells e.g., flow cytometry analysis of the proportion of antigen specific CD4+FoxP3+ T cells as a fraction of the total CD4 + T cell pool
  • analysis of cell proliferation markers e.g., expression of Ki67
  • the present TNFRSF25-specific agent is used to treat or prevent various autoimmune and/or neurodegenerative diseases.
  • the neurodegenerative disease is selected from MS (including without limitation the subtypes described herein), Alzheimer's disease (including, without limitation, early-onset Alzheimer's, late-onset Alzheimer’s, and familial Alzheimer’s disease (FAD), Parkinson’s disease and parkinsonism (including, without limitation, Idiopathic Parkinson's disease, vascular parkinsonism, drug-induced parkinsonism, dementia with Lewy bodies, inherited Parkinson's, juvenile Parkinson's), Huntington's disease, amyotrophic lateral sclerosis (ALS, including, without limitation, sporadic ALS, familial ALS, Western Pacific ALS, juvenile ALS, Hiramaya Disease).
  • the autoimmune disease is selected from Addison's disease, rheumatoid arthritis, type 1 diabetes, vasculitis, alopecia areata, lupus, polymyalgia rheumatica, ankylosing spondylitis, celiac disease, Sjogren's syndrome, and temporal arteritis, Graves' disease, Hashimoto's thyroiditis, Dermatomyositis, Myasthenia gravis, and Pernicious anemia.
  • the autoimmune inflammatory disease is a demyelinating disease.
  • the inflammation is in the central nervous system (CNS).
  • the demyelinating disease is multiple sclerosis (MS).
  • the demyelinating disease is clinically isolated syndrome (CIS).
  • the method prevents or slows the progression of CIS to MS.
  • the demyelinating disease is radiologically isolated syndrome (RIS).
  • the method prevents or slows the progression of RIS to MS.
  • the patient is likely to develop MS.
  • the present disclosure is used to treat or prevent MS.
  • TNFRSF25 agonistic antibodies, or fragments thereof as described herein are used to eliminate and reduce multiple MS symptoms.
  • Illustrative symptoms associated with multiple sclerosis include: optic neuritis, diplopia, nystagmus, ocular dysmetria, internuclear ophthalmoplegia, movement and sound phosphenes, afferent pupillary defect, paresis, monoparesis, paraparesis, hemiparesis, quadraparesis, plegia, paraplegia, hemiplegia, tetraplegia, quadraplegia, spasticity, dysarthria, muscle atrophy, spasms, cramps, hypotonia, clonus, myoclonus, myokymia, restless leg syndrome, footdrop, dysfunctional reflexes, paresthesia
  • the present TNFRSF25-specific agent is used to treat or prevent clinically isolated syndrome (CIS).
  • a clinically isolated syndrome is a single monosymptomatic attack compatible with MS, such as optic neuritis, brain stem symptoms, and partial myelitis.
  • Patients with CIS that experience a second clinical attack are generally considered to have clinically definite multiple sclerosis (CDMS). Over 80 percent of patients with a CIS and MRI lesions go on to develop MS, while approximately 20 percent have a self-limited process.
  • Patients who experience a single clinical attack consistent with MS may have at least one lesion consistent with multiple sclerosis prior to the development of clinically definite multiple sclerosis.
  • the present TNFRSF25-specific agent is used to treat CIS so it does not develop into MS, including, for example RRMS.
  • the patient has CIS.
  • the patient has a monofocal episode of CIS.
  • the patient has optic neuritis. In embodiments, the patient has a multifocal episode of CIS. In embodiments, the patient has optic neuritis and numbness or tingling in the legs.
  • the present TNFRSF25-specific agent is used to treat or prevent radiologically isolated syndrome (RIS).
  • RIS radiologically isolated syndrome
  • incidental imaging findings suggest inflammatory demyelination in the absence of clinical signs or symptoms.
  • the present TNFRSF25-specific agent is used to treat MS so it does not develop into MS, including, for example RRMS.
  • the present TNFRSF25-specific agent is used to treat benign multiple sclerosis; relapsing-remitting multiple sclerosis (RRMS); secondary progressive multiple sclerosis (SPMS); progressive relapsing multiple sclerosis (PRMS); and primary progressive multiple sclerosis (PPMS).
  • RRMS relapsing-remitting multiple sclerosis
  • SPMS secondary progressive multiple sclerosis
  • PRMS progressive relapsing multiple sclerosis
  • PPMS primary progressive multiple sclerosis
  • the treatment occurs early in disease progression. In embodiments, the treatment occurs before the onset of relapsing-remitting MS (RRMS). In embodiments, the treatment occurs before the onset of secondary progressive MS (SPMS). In embodiments, the treatment occurs before the onset of primary progressive MS (PPMS).
  • RRMS relapsing-remitting MS
  • SPMS secondary progressive MS
  • PPMS primary progressive MS
  • Benign multiple sclerosis is a retrospective diagnosis which is characterized by 1-2 exacerbations with complete recovery, no lasting disability and no disease progression for 10-15 years after the initial onset. Benign multiple sclerosis may, however, progress into other forms of multiple sclerosis.
  • the present TNFRSF25-specific agent is used to treat benign multiple sclerosis so it does not develop into MS.
  • Patients suffering from RRMS experience sporadic exacerbations or relapses, as well as periods of remission. Lesions and evidence of axonal loss may or may not be visible on MRI for patients with RRMS.
  • the present TNFRSF25-specific agent is used to treat RRMS.
  • RRMS includes patients with RRMS; patients with SPMS and superimposed relapses; and patients with CIS who show lesion dissemination on subsequent MRI scans according to McDonald's criteria.
  • RRMS includes patients with RRMS; patients with SPMS and superimposed relapses; and patients with CIS who show lesion dissemination on subsequent MRI scans according to McDonald's criteria.
  • a clinical relapse which may also be used herein as “relapse,” “confirmed relapse,” or “clinically defined relapse,” is the appearance of one or more new neurological abnormalities or the reappearance of one or more previously observed neurological abnormalities. This change in clinical state must last at least 48 hours and be immediately preceded by a relatively stable or improving neurological state of at least 30 days.
  • an event is counted as a relapse when the subject's symptoms are accompanied by observed objective neurological changes, consistent with an increase of at least 1.00 in the EDSS score or one grade in the score of two or more of the seven FS or two grades in the score of one of FS as compared to the previous evaluation.
  • SPMS may evolve from RRMS. Patients afflicted with SPMS have relapses, a diminishing degree of recovery during remissions, less frequent remissions and more pronounced neurological deficits than RRMS patients. Enlarged ventricles, which are markers for atrophy of the corpus callosum, midline center and spinal cord, are visible on MRI of patients with SPMS. In embodiments, the present TNFRSF25- specific agent is used to treat RRMS so it does not develop into SPMS.
  • PPMS is characterized by a steady progression of increasing neurological deficits without distinct attacks or remissions. Cerebral lesions, diffuse spinal cord damage and evidence of axonal loss are evident on the MRI of patients with PPMS. PPMS has periods of acute exacerbations while proceeding along a course of increasing neurological deficits without remissions. Lesions are evident on MRI of patients suffering from PRMS.
  • the present TNFRSF25-specific agent is used to treat RRMS and/or SPMS so it does not develop into PPMS.
  • the treatment reduces spinal cord inflammation, as compared to before treatment. In embodiments, the treatment reduces axon demyelination, as compared to before treatment.
  • the present TNFRSF25-specific agents are used in a method of treatment of relapsing forms of MS.
  • the present TNFRSF25-specific agents are used in a method of treatment of relapsing forms of MS to slow the accumulation of physical disability and/or reduce the frequency of clinical exacerbations, and, optionally, for patients who have experienced a first clinical episode and have MRI features consistent with MS.
  • the present TNFRSF25-specific agents are used in a method of treatment of worsening relapsing-remitting MS, progressive-relapsing MS or secondary- progressive MS to reduce neurologic disability and/or the frequency of clinical exacerbations.
  • the present TNFRSF25-specific agents reduce the frequency and/or severity of relapses.
  • the present TNFRSF25-specific agents are used in a method of treatment of relapsing forms of MS in patients who have had an inadequate response to (or are refractory to) one, or two, or three, or four, or five, or six, or seven, or eight, or nine, or ten or more DMTs.
  • the subject's symptoms may be assessed quantitatively, such as by EDSS, or decrease in the frequency of relapses, or increase in the time to sustained progression, or improvement in the magnetic resonance imaging (MRI) behavior in frequent, serial MRI studies and compare the patient's status measurement before and after treatment.
  • the patient status will have improved ⁇ e.g., the EDSS measurement number or frequency of relapses will have decreased, or the time to sustained progression will have increased, or the MRI scans will show less pathology).
  • the patient can be evaluated, e.g., before, during or after receiving the present TNFRSF25-specific agent, e.g., for indicia of responsiveness.
  • Various clinical or other indicia of effectiveness of treatment e.g., EDSS score; MRI scan; relapse number, rate, or severity; multiple sclerosis functional composite (MSFC); multiple sclerosis quality of life inventory (MSQLI); Paced Serial Addition Test (PASAT); symbol digit modalities test (SDMT); 25-foot walk test; 9-hole peg test; low contrast visual acuity; Modified Fatigue Impact Scale; expanded disability status score (EDSS); multiple sclerosis functional composite (MSFC); Beck Depression Inventory; and 7/24 Spatial Recall Test can be used and, in embodiments, the present TNFRSF25-specific agent causes an improvement in one or more of these measures.
  • the present TNFRSF25-specific agents cause a disease improvement as assessed by assessment of MacDonald dissemination in space and time.
  • lesion imaging such as, by way of illustration, Barkhof-Tintore MR imaging criteria, may be used, including at least one gadoliniumenhancing lesion or 9 T2 hyperintense lesions; at least one infratentorial lesion; at least one juxtacortical lesion; at least about three periventricular lesions; and a spinal cord lesion.
  • MRI can also be used; for example, if an MRI scan of the brain performed at >3 months after an initial clinical event demonstrates a new gadolinium-enhancing lesion, this may indicate a new CNS inflammatory event, because the duration of gadolinium enhancement in MS is usually less than 6 weeks. If there are no gadolinium-enhancing lesions but a new T2 lesion (presuming an MRI at the time of the initial event), a repeat MR imaging scan after another 3 months may be needed with demonstration of a new T2 lesion or gadolinium-enhancing lesion.
  • disease effects are assessed using any of the measures described in Lavery, et al. Multiple Sclerosis International, Vol 2014 (2014), Article ID 262350, the entire contents of which are hereby incorporated by reference.
  • the present TNFRSF25-specific agent results in one or more of: (a) prevention of worsening in disability defined as deterioration by 1.0 point on EDSS, (b) increase in time to relapse, (c) reduction or stabilization of number and/or volume of gadolinium enhancing lesions, (d) decreased annualized relapse rate, (e) increased relapse duration and severity by NRS score, (f) decrease in disease activity as measured by MRI (annual rate of new or enlarging lesions), (g) lower average number of relapses at 1 year, or 2 years, (h) sustained disease progression as measured by the EDSS at 3 months, (i) prevention of conversion to CDMS, (j) no or few new or enhancing T2 lesions, (k) minimal change in hyperintense T2 lesion volume, (I) increased time to McDonald defined MS, (m) prevention of progression of disability as measured by sustained worsening of EDSS at 12 weeks, (n) reduction in time to re
  • the present TNFRSF25-specific agent is administered and is effective to result in a decreased rate of relapse ⁇ e.g., at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80% or greater reduction in rate of relapse) compared to the rate of relapse before administration ⁇ e.g., compared to the rate of relapse following administration for 12 months or for less than 12 months, e.g., about 10, or about 8, or about 4, or about 2 or less months) of treatment, or before commencement of treatment, when measured between 3-24 months ⁇ e.g., between 6-18 months, e.g., 12 months) after a previous relapse.
  • a decreased rate of relapse e.g., at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80% or greater reduction in rate of relapse
  • the rate of relapse before administration e.g., compared to the rate of relapse following administration for 12 months or for less than 12 months, e.g., about 10,
  • the present TNFRSF25-specific agent is administered and is effective to result in a prevention of an increase in EDSS score from a pre-treatment state.
  • the Kurtzke Expanded Disability Status Scale (EDSS) is a method of quantifying disability in multiple sclerosis. The EDSS replaced the previous Disability Status Scales which used to bunch people with MS in the lower brackets. The EDSS quantifies disability in eight Functional Systems (FS) and allows neurologists to assign a Functional System Score (FSS) in each of these.
  • the Functional Systems are: pyramidal, cerebellar, brainstem, sensory, bowel and bladder, visual & cerebral.
  • the present TNFRSF25-specific agent is administered and is effective to result in a decreased EDSS score ⁇ e.g., a decrease of 1 , 1.5, 2, 2.5, 3 points or more, e.g., over at least three months, six months, one year, or longer) compared to the EDSS score following administration of the present TNFRSF25-specific agent ⁇ e.g. for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before the commencement of treatment).
  • the present TNFRSF25-specific agent is administered and is effective to result in a decreased number of new lesions overall or of any one type ⁇ e.g., at least 10%, 20%, 30%, 40% decrease), compared to the number of new lesions following administration of the present TNFRSF25- specific agent for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
  • the present TNFRSF25-specific agent is administered and is effective to result in a decreased number of lesions overall or of any one type ⁇ e.g., at least 10%, 20%, 30%, 40% decrease), compared to the number of lesions following administration of the present TNFRSF25-specific agent for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
  • the present TNFRSF25-specific agent is administered and is effective to result in a reduced rate of appearance of new lesions overall or of any one type ⁇ e.g., at least 10%, 20%, 30%, 40% reduced rate), compared to the rate of appearance of new lesions following administration for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
  • the present TNFRSF25-specific agent is administered and is effective to result in a decreased increase in lesion area overall or of any one type ⁇ e.g., at least 10%, 20%, 30%, 40% decreased increase), compared to an increase in lesion area following administration for 12 months or less than 12 months, e.g., less than 10 months, 8 months, 4 months, or 2 months, or before commencement of treatment.
  • the human has an age in a range of from about 1 to about 18 months old, from about 18 to about 36 months old, from about 1 to about 5 years old, from about 5 to about 10 years old, from about 10 to about 15 years old, from about 15 to about 20 years old, from about 20 to about 25 years old, from about 25 to about 30 years old, from about 30 to about 35 years old, from about 35 to about 40 years old, from about 40 to about 45 years old, from about 45 to about 50 years old, from about 50 to about 55 years old, from about 55 to about 60 years old, from about 60 to about 65 years old, from about 65 to about 70 years old, from about 70 to about 75 years old, from about 75 to about 80 years old, from about 80 to about 85 years old, from about 85 to about 90 years old, from about 90 to about 95 years old or from about 95 to about 100 years old. In embodiments, the human has an age of more than 30 years old.
  • the present TNFRSF25-specific agents are used to treat or prevent an autoimmune inflammatory disease selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti-MAG peripheral neuropathy, Charcot-Marie-Tooth disease, hereditary neuropathy with liability to pressure palsy, peripheral neuropathy, optic neuropathy, and progressive inflammatory neuropathy.
  • an autoimmune inflammatory disease selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti-
  • the present TNFRSF25-specific agents are used to prevent or reduce symptoms of an autoimmune inflammatory disease, such as blurred double vision (diplopia), ataxia, clonus, dysarthria, fatigue, clumsiness, hand paralysis, hemiparesis, genital anesthesia, incoordination, paresthesia, ocular paralysis (cranial nerve palsy), impaired muscle coordination, weakness (muscle), loss of sensation, impaired vision, neurological symptoms, unsteady gait, spastic paraparesis, incontinence, hearing problems, and speech problems.
  • an autoimmune inflammatory disease such as blurred double vision (diplopia), ataxia, clonus, dysarthria, fatigue, clumsiness, hand paralysis, hemiparesis, genital anesthesia, incoordination, paresthesia, ocular paralysis (cranial nerve palsy), impaired muscle coordination, weakness (muscle), loss of sensation, impaired vision, neurological symptoms, unsteady gait, spastic parapare
  • the treatment expands and/or selectively activates a population of Tregs in the patient.
  • the treatment does not substantially expand and/or selectively activate a population of cytotoxic T cells (Tc cells) and/or helper T cells (Th cells) in the patient.
  • Tc cells cytotoxic T cells
  • Th cells helper T cells
  • the treatment does not substantially expand and/or selectively activate a population of Th17 cells in the patient.
  • the treatment increases the ratio of Tregs to Tc and/or Th cells in the patient.
  • the treatment increases the ratio of Tregs to Th17 cells in the patient.
  • the treatment increases the ratio of Tregs to conventional T cells in the patient compared to an untreated patient.
  • the treatment increases the population of Tregs expressing ICOS compared to an untreated patient. In embodiments, the treatment increases the population of Tregs expressing CD103 compared to an untreated patient. In embodiments, the treatment increases the population of Tc cells expressing Tim3 and/or LAG3 compared to an untreated patient.
  • the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining the amount of ICOS in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to a patient in need of an increase of ICOS.
  • the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining the amount of CD103 in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to a patient in need of an increase of CD103.
  • the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining the amount of Tim3 and/or LAG3 in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to a patient in need of an increase of Tim3 and/or LAG3.
  • a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment comprising, determining an amount of ICOS protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of ICOS relative to an untreated and/or undiseased patient.
  • a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment comprising, determining an amount of CD103 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of CD103 relative to an untreated and/or undiseased patient.
  • a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment comprising, determining an amount of Tim3 and/or LAG3 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of Tim3 and/or LAG3 relative to an untreated and/or undiseased patient.
  • a sample is biological fluid, such as whole blood or whole blood components including red blood cells, white blood cells, platelets, serum and plasma, ascites, vitreous fluid, lymph fluid, synovial fluid, follicular fluid, seminal fluid, amniotic fluid, milk, saliva, sputum, tears, perspiration, mucus, cerebrospinal fluid, and urine.
  • the sample is a body sample from any animal.
  • the sample is from a mammal.
  • the sample is from a human subject.
  • an effective amount of the TNFRSF25-specific agents can be any amount that has a desired effect (e.g., treating).
  • dosages can vary depending on the relative potency of individual treatments (e.g., TNFRSF25-specific agents), and can generally be estimated based on ECso found to be effective in in vitro and in vivo animal models. In embodiments, dosage is from 0.01 g to 100 g per kg of body weight.
  • an effective amount of the TNFRSF25-specific agents can be from about 0.1 mg/kg to about 50 mg/kg (e.g., about 0.4 mg/kg, about 2 mg/kg, about 5 mg/kg, about 10 mg/kg, about 20 mg/kg, about 30 mg/kg, about 40 mg/kg, or about 50 mg/kg), or any range there between, such as about 0.1 mg/kg to about 10 mg/kg, about 0.4 mg/kg to about 20 mg/kg, about 2 mg/kg to about 30 mg/kg, or about 5 mg/kg to about 40 mg/kg.
  • the amount of TNFRSF25-specific agents is increased by, for example, two fold.
  • the subject is monitored for both responsiveness to the treatment and toxicity symptoms, and adjustments made accordingly.
  • the effective amount remains constant or is adjusted as a sliding scale or variable dose depending on the subject’s response to treatment.
  • the frequency of administration, duration of treatment, use of multiple treatment agents, route of administration, and severity of the cancer may require an increase or decrease in the actual effective amount administered.
  • the frequency of administration of the TNFRSF25-specific agents is any frequency that, for example, can be used to treat or prevent an autoimmune inflammatory disease.
  • the frequency of administration is once or more daily, biweekly, weekly, monthly, or even less.
  • the frequency of administration remains constant or is variable during the duration of treatment.
  • a course of treatment includes rest periods.
  • the TNFRSF25- specific agents are administered over a two-week period followed by a two-week rest period, and such a regimen is repeated multiple times.
  • various factors can influence the frequency of administration used for a particular application.
  • an effective duration for administering the TNFRSF25-specific agents is any duration that is needed to effectively treat or prevent an autoimmune inflammatory disease.
  • an effective duration can vary from several days to several weeks, months, or years.
  • an effective duration is for as long as an individual subject is alive.
  • multiple factors influence the actual effective duration used for a particular treatment.
  • an effective duration varies with the frequency of administration, effective amount, use of multiple treatment agents, and route of administration.
  • the TNFRSF25-specific agents described herein will typically contain an effective amount of the TNFRSF25-specific agents for achieving the desired effect.
  • therapeutically effective amount and “effective amount,” used interchangeably, applied to a dose or amount refers to a quantity of a composition, compound or pharmaceutical formulation that is sufficient to result in a desired activity upon administration to an animal in need thereof.
  • the effective amount of the combination may or may not include amounts of each ingredient that would have been effective if administered individually.
  • the dosage of the therapeutic formulation will vary, depending upon the nature of the disease or condition, the patient’s medical history, the frequency of administration, the manner of administration, the clearance of the agent from the host, and the like.
  • the initial dose may be larger, followed by smaller maintenance doses.
  • the dose may be administered, e.g., weekly, biweekly, daily, semi-weekly, etc., to maintain an effective dosage level.
  • the present TNFRSF25-specific agent is administered and is effective to result in a reduced incidence or symptom of optic neuritis (e.g ., improved vision), compared to the incidence or symptom of optic neuritis following administration for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
  • a reduced incidence or symptom of optic neuritis e.g ., improved vision
  • the present TNFRSF25-specific agent is administered and is effective to result in a reduced incidence or symptom of optic neuritis (e.g ., improved vision), compared to the incidence or symptom of optic neuritis following administration for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
  • the methods described herein are intended for use with any subject that may experience the benefits of these methods.
  • “subjects,” “patients,” and “individuals” include humans as well as non-human subjects, particularly domesticated animals.
  • the subject and/or animal is a mammal, e.g., a human, mouse, rat, guinea pig, dog, cat, horse, cow, pig, rabbit, sheep, or non-human primate, such as a monkey, chimpanzee, or baboon.
  • the subject and/or animal is a non-mammal, such, for example, a zebrafish.
  • the subject and/or animal may comprise fluorescently-tagged cells (with e.g. GFP).
  • the subject and/or animal is a transgenic animal comprising a fluorescent cell.
  • methods of the disclosure are useful in treatment a human subject.
  • the human is a pediatric human.
  • the human is an adult human.
  • the human is a geriatric human.
  • the human may be referred to as a patient.
  • the human is a female.
  • the present TNFRSF25-specific agents is used in a method of treating multiple sclerosis in combination with one or more of the DMTs described herein ⁇ e.g. the agents of Table 1).
  • the present disclosure provides an improved therapeutic effect as compared to use of one or more of the DMTs described herein (e.g. the agents of Table 1) without the one or more TNFRSF25- specific agents.
  • the combination of the one or more TNFRSF25-specific agents and the one or more DMTs produce synergistic therapeutic effects.
  • the improved therapeutic benefit of adding an TNFRSF25-specific agent to treatment with a DMT is manifested in a further reduction in one or more of relapse rate, e.g. annualized relapse rate (ARR) relative risk reduction over 2 years (%); disability progression e.g. disability progression relative risk reduction (%); reduction in lesions, e.g. MRI lesions - reduction in Gd-enhancing (%) or MRI lesions - reduction in T2-weighted new or enlarging lesions (%).
  • the present TNFRSF25-specific agents provide an improvement in therapeutic response as compared to the results of Table 2 below ⁇ e.g. about 10%, or about 20%, or about 30%, or about 40%, or about 50%, or about 60%, or about 70%, or about 80%, or about 90%, or about 100% improvement over the values of Table 2).
  • the present TNFRSF25-specific agents reduce deleterious effects or dosing features of one or more MS-treating agents described herein.
  • the present TNFRSF25- specific agents may reduce the dose and/or frequency of administration of any of the DMTs, mitigate inconvenient dosing and therefore increase adherence, reduce side effects, allow for access to further therapies, and bridge the therapeutic benefits while changing therapies, among others.
  • MS disease progression may be most intensive, and most damaging, at the earliest stages of disease progression. Accordingly, counter to many reimbursement policies and physician practice in light of, for example, costs and side effect mitigation, it may be most beneficial for a patient’s long term disease status to begin treatment with the most intensive DMTs, for instance, so-called second-line therapies.
  • a patient is treated with a regimen of the present TNFRSF25-specific agents in combination with a second-line therapy. Such a combination is used to reduce the side effect profile of one or more second-line therapies. In embodiments, the combination is used to reduce dose of frequency of administration of one or more second-line therapies.
  • the doses of Table 1 may be reduced by about 75%, about 50%, or about 40%, or about 30%, or about 25% in the context of the combination and the/or the frequency of dosing may be decreased to be half as often, or a third as often or may be reduced from, for example, daily to every other day or weekly, every other day to weekly or bi-weekly, weekly to biweekly or monthly, etc.
  • the present TNFRSF25-specific agents increase patient adherence by allowing for more convenient treatment regimens.
  • some DMTs have a suggested lifetime dose limitation e.g. for mitoxantrone, the lifetime cumulative dose should be strictly limited to 140 mg/m 2 , or 2 to 3 years of therapy (Fow Clin Thera 28(4): 461).
  • supplementation with the present TNFRSF25-specific agents preserves patient’s access to mitoxantrone by allowing for lower or less frequent dosing with this DMT.
  • the patient is a naive patient, who has not received treatment with one or more DMTs, and the present TNFRSF25-specific agents are used to buffer the side effects of a second-line therapy. Accordingly, the naive patient is able to benefit from the long-term benefits of a second-line therapy at disease outset.
  • the present TNFRSF25-specific therapy is used as an entry therapy that precedes the use of a second-line therapy.
  • the present TNFRSF25-specific therapy may be administered for an initial treatment period of about 3 months to stabilize disease and then the patient may be transitioned to a maintenance therapy of a second line agent.
  • the present TNFRSF25-specific agents find use in patients that have received, and perhaps failed one or more DMT.
  • the present TNFRSF25-specific agents increase the therapeutic effect in patients that have received, and perhaps failed one or more DMT and may allow these patients to respond like naive patients.
  • the patient has received or is receiving treatment with one or more DMTs and is not responding well.
  • the patient may be refractory or poorly responsive to one or more DMTs.
  • the patient is refractory, or poorly responsive to one or more of agents of Table 1.
  • the patient is refractory, or poorly responsive to one or more of teriflunomide (AUBAGIO (GENZYME)); interferon beta-1 a (AVONEX (BIOGEN IDEC); interferon beta-1 b (BETASERON (BAYER HEALTHCARE PHARMACEUTICALS, INC.); glatiramer acetate (COPAXONE (TEVA NEUROSCIENCE); interferon beta-1 b (EXTAVIA (NOVARTIS PHARMACEUTICALS CORP.); fingolimod (GILENYA (NOVARTIS PHARMACEUTICALS CORP.); alemtuzumab (LEMTRADA (GENZYME); mitoxantrone (NOVANTRONE (EMD SERONO); pegylated interferon beta-1 a (PLEGRIDY (BIOGEN I DEC); interferon beta-1 a (REBIF (EMD SERONO, INC.); dimethyl fumarate (BG-12) (AUBAG
  • the present TNFRSF25-specific agents result in a therapeutic benefit of one or more DMTs in the patient and therefore reduce or eliminate the nonresponsiveness to the DMT. For instance, this may spare the patient therapy with one or more DMTs at a higher dosing or frequency.
  • the present TNFRSF25-specific agents may increase a patient’s response such as to avoid the need to change the patient’s therapeutic regiment from a first line therapy to a second line therapy.
  • First line therapies or “platform therapies,” e.g. glatiramer acetate and interferons
  • platforms therapies e.g. glatiramer acetate and interferons
  • first line therapies are considered to be only mildly effective.
  • the present TNFRSF25-specific agents increase the therapeutic efficacy of the first line therapy and reduce the need to alter a patient’s treatment to a second line therapy ⁇ e.g. oral agents (e.g.
  • the present TNFRSF25-specific agents reduce the need for escalation of therapy.
  • the present TNFRSF25-specific agents reduce or eliminate one or more side effects and therefore allow patients access to second line therapies.
  • oral agents are often considered more palatable for MS patients as a second line therapy as compared to the monoclonal antibodies, which often require a medical site visit for infusion administration (and indeed, in some contexts, the oral agents are considered to be first line therapies).
  • the present TNFRSF25-specific agents prevent the need to escalate therapy from an oral agent to a monoclonal antibody.
  • the present TNFRSF25-specific agents are used in combination to allow a patient access to a more efficacious second line therapy, e.g.
  • natalizumab reduces the relapse rate more than first-line agents; however, due to issues of adverse effects is a second-line agent reserved for those who do not respond to other treatments or with severe disease.
  • patients may be given clinical access to natalizumab as their baseline therapy and therefore enjoy natalizumab’s better efficacy relative to first line agents.
  • mitoxantrone whose use is limited by severe adverse effects, is a third-line option for those who do not respond to other medications.
  • patients may be given clinical access to mitoxantrone.
  • the present TNFRSF25-specific agents find use to supplement a DMT with which a patient is comfortable and therefore avoid a need to switch therapies to improve therapeutic effect.
  • fingolimod therapy has been reported to result in a higher probability of no evidence of disease activity than dimethyl fumarate and teriflunomide therapy (Nixon, et al. Adv Thera. 2014; 31 (11): 1134-54).
  • the present TNFRSF25-specific agents supplement dimethyl fumarate and/or teriflunomide therapy to bridge the gap to the clinical effect observed with fingolimod therapy.
  • the present TNFRSF25-specific agents are used to provide the optimal treatment regimen for the patient, in light of therapeutic costs and benefits.
  • one approach is an induction treatment model, whereby a therapy with strong efficacy but strong safety concerns would be given first, followed by a maintenance therapy.
  • An example of such a model might include initial treatment with alemtuzumab, followed by interferons ⁇ e.g. IFN-b), glatiramer acetate, or BG-12.
  • the present TNFRSF25-specific agents are used to prevent the need to switch therapies for maintenance.
  • the present TNFRSF25-specific agents are used to as maintenance therapy to one or more DMTs, including second line therapies.
  • the present TNFRSF25-specific agents are used to as first therapy in an induction, followed by another DMT as a maintenance therapy - such as, for example, a first line therapy.
  • the present TNFRSF25-specific therapy may be administered for an initial treatment period of about 3 months to stabilize disease and then the patient may be transitioned to a maintenance therapy of a first line agent.
  • the present TNFRSF25-specific therapy is used to bridge the gap between the initiation of treatment with one or more DMTs and the onset of clinical effect. For example, in some patients glatiramer acetate slowly begins to have an effect in a patient and the present TNFRSF25-specific therapy provides the necessary therapy during this period. Further, some have suggested a transition period between different DMTs of about 8 to about 12 weeks (by way of non-limiting example, form Tysabri (natalizumab) to GILENYA (fingolimod) to maintain disease control and avoid potentially harmful additive effects on immune surveillance. In embodiments, the present TNFRSF25-specific therapy is used to bridge this gap.
  • the present TNFRSF25-specific agents are used to prevent or reduce one or more side effects of a DMT, including without limitation any agent of Table 1.
  • the present TNFRSF25-specific agents may be used in a regimen that allows dose sparing for one or more DMTs and therefore results in fewer side effects.
  • the present TNFRSF25-specific agents may reduce one or more side effects of AUBAGIO or related agents, which may include hair thinning, diarrhea, flu, nausea, abnormal liver tests and unusual numbness or tingling in the hands or feet (paresthesias), levels of white blood cells, which can increase the risk of infections; increase in blood pressure; and severe liver damage.
  • the present TNFRSF25-specific agents may reduce one or more side effects of AVONEX or related agents which include flu-like symptoms following injection, depression, mild anemia, liver abnormalities, allergic reactions, and heart problems.
  • the present TNFRSF25-specific agents may reduce one or more side effects of BETASERON or related agents which include flu-like symptoms following injection, injection site reactions, allergic reactions, depression, liver abnormalities, and low white blood cell counts.
  • the present TNFRSF25-specific agents may reduce one or more side effects of COPAXONE or related agents which include injection site reactions, vasodilation (dilation of blood vessels); chest pain; a reaction immediately after injection, which includes anxiety, chest pain, palpitations, shortness of breath, and flushing.
  • the present TNFRSF25-specific agents may reduce one or more side effects of EXTAVIA or related agents which include flu-like symptoms following injection, injection site reactions, allergic reactions, depression, liver abnormalities, and low white blood cell counts.
  • the present TNFRSF25-specific agents may reduce one or more side effects of GILENYA or related agents which include headache, flu, diarrhea, back pain, liver enzyme elevations, cough, slowed heart rate following first dose, infections, and swelling in the eye.
  • the present TNFRSF25-specific agents may reduce one or more side effects of LEMTRADA or related agents which include rash, headache, fever, nasal congestion, nausea, urinary tract infection, fatigue, insomnia, upper respiratory tract infection, hives, itching, thyroid gland disorders, fungal Infection, pain in joints, extremities and back, diarrhea, vomiting, flushing, and infusion reactions (including nausea, hives, itching, insomnia, chills, flushing, fatigue, shortness of breath, changes in the sense of taste, indigestion, dizziness, pain).
  • LEMTRADA LEMTRADA or related agents which include rash, headache, fever, nasal congestion, nausea, urinary tract infection, fatigue, insomnia, upper respiratory tract infection, hives, itching, thyroid gland disorders, fungal Infection, pain in joints, extremities and back, diarrhea, vomiting, flushing, and infusion reactions (including nausea, hives, itching, insomnia, chills, flushing, fatigue, shortness of breath, changes in the sense of taste, indigestion, dizziness, pain).
  • the present TNFRSF25-specific agents may reduce one or more side effects of NOVANTRONE or related agents which include blue-green urine 24 hours after administration; infections, bone marrow suppression (fatigue, bruising, low blood cell counts), nausea, hair thinning, bladder infections, mouth sores, and serious liver and heart damage.
  • the present TNFRSF25-specific agents may reduce one or more side effects of PLEGRIDY or related agents which include flu-like symptoms following injection, injection site reactions, depression, mild anemia, liver abnormalities, allergic reactions, and heart problems.
  • the present TNFRSF25-specific agents may reduce one or more side effects of REBIF or related agents which include flu-like symptoms following injection, injection site reactions, liver abnormalities, depression, allergic reactions, and low red or white blood cell counts.
  • the present TNFRSF25-specific agents may reduce one or more side effects of TECFIDERA or related agents which include flushing (sensation of heat or itching and a blush on the skin), gastrointestinal issues (nausea, diarrhea, abdominal pain), rash, protein in the urine, elevated liver enzymes; and reduction in blood lymphocyte (white blood cell) counts.
  • the present TNFRSF25-specific agents may reduce one or more side effects of TYSABRI or related agents which include headache, fatigue, urinary tract infections, depression, respiratory tract infections, joint pain, upset stomach, abdominal discomfort, diarrhea, vaginitis, pain in the arms or legs, rash, allergic or hypersensitivity reactions within two hours of infusion (dizziness, fever, rash, itching, nausea, flushing, low blood pressure, difficulty breathing, chest pain).
  • the present TNFRSF25-specific agents are used to prevent or reduce progressive multifocal leukoencephalopathy (PML). In embodiments, the present TNFRSF25-specific agents are used to prevent or reduce progressive multifocal leukoencephalopathy (PML) caused by treatment with TECFIDERA or related agents.
  • PML progressive multifocal leukoencephalopathy
  • a method for treating or preventing an autoimmune inflammatory disease comprises administering an effective amount of TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibody or antigen binding fragment thereof (e.g., PTX-35, SEQ ID NOs: 8 and 9, or a murine version thereof “mPTX-35”) to a patient in need thereof.
  • TNF Receptor Superfamily Member 25 TNFRSF25
  • PTX-35 e.g., SEQ ID NOs: 8 and 9, or a murine version thereof “mPTX-35”
  • a classical in vivo experimental model based on chronic experimental autoimmune encephalomyelitis (“EAE”) in mice was utilized to test whether PTX-35 is effective to treat or prevent an autoimmune disease.
  • EAE chronic experimental autoimmune encephalomyelitis
  • the following experiments show how mPTX-35 expanded both Tregs and Tconv during model disease.
  • Data is shown in FIG. 3 of the body weight of the mice before disease induction.
  • mPTX-35 was administered on both day -7 and day 7, and mPTX-35 expanded Tregs prior to model start. Data is shown in FIG.
  • FIG. 6 is a chart showing the clinical assessment of EAE with respect to scores and clinical signs.
  • FIG.7 is a graph showing the clinical score of the mice when given mPTX-35 before disease induction
  • FIG. 10 is a graph showing the disease incidence in the mice before disease induction.
  • FIG. 14 is a graph showing percent survival (x-axis) and days elapsed (y-axis) before disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice.
  • mPTX- 35 was administered on both day -7 and day 7, and mPTX-35 expand Tregs prior to the start of the experimental model.
  • FIG. 8 is a graph showing the clinical score of the mice when given mPTX-35 during disease induction
  • FIG. 11 is a graph showing the disease incidence in the mice during disease induction
  • FIG. 15 is a graph showing percent survival (x-axis) and days elapsed (y-axis) during disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice.
  • mPTX- 35 was administered on both day 0 and day 14, and mPTX-35 expanded conventional T cells (“Tconv”) at the start of the experimental model.
  • FIG. 9 is a graph showing the clinical score of the mice when given mPTX-35 after disease induction
  • FIG. 12 is a graph showing the disease incidence in the mice after disease induction
  • FIG. 16 is a graph showing percent survival (x-axis) and days elapsed (y-axis) after disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • mPTX- 35 was administered on both day 7 and day 21, and mPTX-35 expanded both Tregs and Tconv during the model disease.
  • FIG. 13 is a graph showing the clinical score area under the curve (“AUC”) measurements for the G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • AUC clinical score area under the curve
  • FIG. 17 is a chart showing the evaluation of disease progression as it relates to the spinal cord.
  • FIG. 18 is a graph showing the histology inflammation score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • PTX-35 mediated Treg expansion reduced central nervous system pathologies
  • these experiments show how expansion of Tregs before disease induction or after initiation can control outcome.
  • FIG. 19 is a graph showing the histology lesion score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • PTX-35 mediated Treg expansion reduced central nervous system pathologies. These experiments show expansion of Tregs before disease induction or after initiation can control outcome.
  • FIG. 20 is a graph showing the histology demyelination score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • G2 vehicle
  • G3 Dex 1 mg/kg
  • G4 mPTX-35 3 mg/kg
  • G5 mPTX-35 3 mg/kg
  • G6 mPTX-35 3 mg/kg mice.
  • FIG. 21 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G1 normal mice (left panel), and G2 vehicle mice (right panel). These experiments show PTX-35 mediated Treg expansion reduced central nervous system pathologies.
  • HE hematoxylin and eosin
  • FIG. 22 and FIG. 23 Representative HE and fast blue staining are shown in FIG. 22 and FIG. 23 for G3 Dex treated mice, G4 mPTX-35 treated mice, G5 mPTX-35 treated mice, and G6 mPTX-35 treated mice.
  • FIG. 24 shows the gating strategy for the Flow cytometry data.
  • FIG. 25 is a graph showing the percentage of regulatory T cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • FIG. 24 shows the gating strategy for the Flow cytometry data.
  • FIG. 25 is a graph showing the percentage of regulatory T cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehi
  • 26 is a graph showing the percentage of Th17 cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
  • Example 2 MOG 35-55 Immunized Mice Treated with mPTX-35 at Various Days Show Reduced or Minimal Disease Severity
  • PTX-35 treatment on Days 12&16 showed the greatest protective capacity from EAE.
  • Analysis of splenocytes from PTX-35 treated mice revealed an increase FoxP3 + Treg.
  • PTX-35 treated T cells displayed increased expression of ICOS and CD103.
  • CD8 + T cells showed signs of exhaustion with increased Tim3 and LAG3 after PTX-35 treatment.
  • MOG 35-55 immunized C57BL/6 were treated with PBS, Isotype control antibody (10 mg/kg; clone MOPC- 21, InVivoMab), PTX-35 (10 mg/kg) at days 4 and 8 post immunization, or PTX-35 (10 mg/kg) at days 12 and 16 post immunization (FIG. 27A).
  • Splenocytes were isolated and stained for flow cytometry. Spinal cords were kept for histology.
  • mice were euthanized at day 30 post immunization for immune profiling of splenocytes by flow cytometry and demyelination of the spinal cord by histology. Images of spinal cords from B6 mice immunized with MOG 35-55 taken at Day 30 post immunization (FIG.29). Luxol Fast Blue staining specifically stained the myelin sheath of nerve cells highlighting the white matter of the spinal cord. Hematoxylin and eosin (H&E) staining highlighted lymphocytic infiltration of the spinal cord (indicated by arrows) (FIG. 29).
  • H&E Hematoxylin and eosin
  • PTX-35 treatment on days 12 and 16 showed the greatest protective capacity from EAE.
  • Analysis of splenocytes from PTX-35 treated mice revealed an increase of FoxP3+ Treg.
  • PTX-35 treated T cells displayed increased expression of ICOS and CD103.
  • CD8+ T cells showed signs of exhaustion with increased Tim3 and LAG3 after PTX-35 treatment.
  • FIG. 31 shows percentage distribution of Foxp3+ regulatory T cells and conventional CD4+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with mPTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with mPTX-35 at day 12 and day 16.
  • B6 mice immunized with MOG 35-55 were sacrificed 30 days post immunization.
  • Splenic CD4+ T cells were analyzed and results showed immunized mice treated with PTX- 35 at day 4 and 8 and immunized mice at day 12 and 16 had significantly higher percentages of activated FoxP3+T cells compared to isotype mice (FIG. 32).
  • the splenic CD4+ FoxP3+ regulatory T cells were also examined for expression levels of immunosuppressive markers including ICOS+ T (FIG. 33 - FIG. 34), CD103+ T cells (FIG. 35 - FIG. 36). These results showed that immunized mice treated with PTX-35 at days 4 and 8 and immunized mice treated with PTX-35 at days 12 and 16 had significantly higher percentages of regulatory T cells expressing these markers.
  • the splenic CD4+ FoxP3+ regulatory T cells were also examined for expression levels of other immunosuppressive markers including CD44 (FIG. 39), TIGIT (FIG. 40), CTLA4 (FIG. 41), and Ki67 (FIG. 42). These results showed that immunized mice treated with PTX-35 at days 12 and 16 had slightly higher expression levels of CD44, TIGIT, CTLA4, and Ki67 compared to mice treated with isotype control antibodies.
  • the splenic CD8+ T cells were also examined for percentage of PD-1 + T cells, which did not change significantly between non-immunized and immunized mice samples (FIG.45). Percentage of PD- 1+T cells increased after treatment of PTX-35 between non-immunized and immunized mice samples (FIG. 46).
  • Results showed FoxP3 + Treg are expanded after PTX-35 administrations during autoimmunity and reduces EAE severity.
  • ICOS and CD103 were significantly upregulated markers on FoxP3 + Treg. Other markers showed a trend towards increasing but were variable in expression.
  • CD8 + T cells significantly upregulated LAG3 and TIM3 after PTX-35 therapy.
  • An increased frequency of FoxP3+ Treg suggests enhanced suppression of pathogenic T cells after PTX-35 therapy.
  • ICOS expression by FoxP3+ Treg is known to be important for regulatory T cell function in autoimmune settings.
  • increased expression of CD103 on FoxP3+Treg suggests a greater capacity for tissue residency from PTX-35 treated FoxP3+Treg.
  • PTX-35 treatment also enhanced dysfunctional CD8+T cell markers during EAE.
  • CD8+ T cells showed increased expression of the inhibitory molecules Tim3 and Lag3 implying PTX-35 treated CD8+T cells may be functionally exhausted which could attenuate autoimmunity.
  • PTX-35 treatments also showed prevention or reversal of primary progressive EAE.
  • Example 3 PLP 139-151 Immunized SJUL Mice Showed Relapsing Remitting EAE Prevention or Reversal via PTX-35
  • mice were immunized with PLP 139-151 and treated with PTX-35 at day -4 and 0, day 4 and 8, day 12 and 16, day 16 and 20, or day 4, 8, 16, and 20 post immunization (FIG. 47).
  • Results show disease progression in mice treated with PTX-35 showed signs of improved compared to untreated mice (FIG. 48-52).
  • mice treated with PTX-35 at day 4 and day 8 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 10 until day 43 (FIG. 49).
  • Mice treated with PTX-35 at day 12 and day 16 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 13-15 and day 24-43 (FIG. 50).
  • Mice treated with PTX-35 at day 16 and day 20 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 13 until day 43 (FIG. 51).
  • Mice treated with PTX-35 at day 4 and 8, and day 12 and 16 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 13-15 and day 20-43 (FIG. 52).
  • Results showed that PTX-35 treatment at various days prevented or reversed relapsing remitting EAE disease progression compared to mice treated with isotype control antibodies.

Abstract

The present disclosure relates to, in part, compositions and methods for the treatment of various autoimmune and/or neurodegenerative diseases, including, for example, multiple sclerosis, e.g. with TNFRSF25 agonistic antibodies or antigen binding fragments thereof.

Description

TREATMENT OF AUTOIMMUNE INFLAMMATORY DISEASES WITH TNFRSF25-BINDING AGENTS
FIELD
[0001] The disclosure is directed to treatment and prevention of autoimmune-related diseases and disorders using TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibodies.
CROSS-REFERENCE TO RELATED APPLICATIONS
[0002] This application claims priority to and the benefit of U.S. Provisional Patent Application No.
63/189,774, filed on May 18, 2021, and U.S. Provisional Patent Application No. 63/323,252 filed on March 24, 2022, the entire content of which is hereby incorporated herein by reference in its entireties.
SEQUENCE LISTING
[0003] The application contains a Sequence Listing which has been submitted in ASCII format via
EFS-Web and is hereby incorporated by reference in its entirety. Said ASCII copy, created on May 18, 2021, is named PEL-017PC_ST25.txt and is 16,384 bytes in size.
BACKGROUND
[0004] Neurodegenerative diseases are presently incurable and debilitating conditions that result in progressive degeneration and/or death of nerve cells. Of these, multiple sclerosis (MS) is a chronic, progressive neurodegenerative disorder that results from an attack on the central nervous system (brain, spinal cord and optical nerves) by the body’s own immune system, causing inflammation and damage to the myelin layer that covers and protects neurons resulting in motor function impairment {e.g. coordination, balance, speech and vision), irreversible neurological disability and paralysis. It is estimated that more than 2.3 million people in the world suffer from MS, and 400,000 people in the U.S. alone have the disease.
[0005] Current pharmacological treatments for MS, including disease modifying therapies (DMTs), either modify or suppress the body's immune system. Many of the current pharmacological agents for MS are limited by incomplete efficacy, side effects and medical risks. These treatments have been shown to modestly reduce neurological relapses of the disease and, in some instances, incompletely slow the progression of neurological disability. Accordingly, there remains a pressing need for improved MS treatments.
SUMMARY
[0006] Accordingly, in various aspects, the present disclosure relates to a method for treating or preventing an autoimmune inflammatory disease, comprising administering an effective amount of TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibody or antigen binding fragment thereof to a patient in need thereof. In embodiments, the autoimmune inflammatory disease is a demyelinating disease. For example, In embodiments the inflammation is in the central nervous system (CNS). In embodiments, the demyelinating disease is multiple sclerosis (MS). In embodiments, the demyelinating disease is clinically isolated syndrome (CIS). In embodiments, the method prevents or slows the progression of CIS to MS. In embodiments, the demyelinating disease is radiologically isolated syndrome (RIS). In embodiments, the method prevents or slows the progression of RIS to MS. In embodiments, the patient is likely to develop MS. In embodiments, the patient has CIS. In embodiments, the patient has a monofocal episode of CIS.
[0007] In embodiments, the patient has optic neuritis. In embodiments, the patient has a multifocal episode of CIS. In embodiments, the patient has optic neuritis and numbness or tingling in the legs.
[0008] In embodiments, the patient has CIS and one or more magnetic resonance imaging (MRI)- detected brain lesions.
[0009] In embodiments, the treatment occurs before or early in disease progression. In embodiments, the treatment occurs before the onset of relapsing-remitting MS (RRMS). In embodiments, the treatment occurs before the onset of secondary progressive MS (SPMS). In embodiments, the treatment occurs before the onset of primary progressive MS (PPMS).
[0010] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof is used to treat or prevent an autoimmune inflammatory disease selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti-MAG peripheral neuropathy, Charcot-Marie-Tooth disease, hereditary neuropathy with liability to pressure palsy, peripheral neuropathy, optic neuropathy, and progressive inflammatory neuropathy.
[0011] In embodiments, the treatment expands and/or selectively activates a population of Tregs in the patient. In embodiments, the treatment does not substantially expand and/or selectively activate a population of cytotoxic T cells (Tc cells) and/or helper T cells (Th cells) in the patient. In embodiments, the treatment does not substantially expand and/or selectively activate a population of Th17 cells in the patient. In embodiments, the treatment increases the ratio of Tregs to Tc and/or Th cells in the patient. In embodiments, the treatment increases the ratio of Tregs to Th17 cells in the patient.
[0012] In embodiments, the treatment reduces spinal cord inflammation, as compared to before treatment. In embodiments, the treatment reduces axon demyelination, as compared to before treatment.
[0013] In various aspects, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises: a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 sequences, wherein the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO: 1), the heavy chain CDR2 sequence is YISSASGUSYADAVRG (SEQ ID NO: 2); and the heavy chain CDR3 sequence is DPAYTGLYALDF (SEQ ID NO: 3) or DPPYSGLYALDF (SEQ ID NO: 4); and a light chain variable region comprising light chain CDR1, CDR2, and CDR3 sequences, wherein the light chain CDR1 sequence is TLSSELSWYTIV (SEQ ID NO: 5), the light chain CDR2 sequence is LKSDGSHSKGD (SEQ ID NO: 6), and the light chain CDR3 sequence is CGAGYTLAGQY G WV (SEQ ID NO: 7).
[0014] In embodiments, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises variable region framework (FW) sequences juxtaposed between the CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4), wherein the variable region FW sequences in the heavy chain variable region are heavy chain variable region FW sequences, and wherein the variable region FW sequences in the light chain variable region are light chain variable region FW sequences. In embodiments, wherein the variable region FW sequences are human.
[0015] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises human heavy chain and light chain constant regions. In embodiments, the constant regions are selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4. In embodiments, the constant regions are lgG1. In embodiments, the constant regions are lgG4.
[0016] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTI SRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85 to about 99% identity thereto. In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence
QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85 to about 99% identity thereto.
[0017] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises 4C12, or a humanized version, or fragment thereof.
[0018] In embodiments, the treatment occurs in combination with one or more disease modifying therapies. In embodiments, the treatment occurs in combination with one or more agents of Table 1.
[0019] In embodiments, the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining an amount of ICOS protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of ICOS relative to an untreated and/or undiseased patient.
[0020] In embodiments, the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining an amount of CD103 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of CD103 relative to an untreated and/or undiseased patient.
[0021] In embodiments, the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining an amount of Tim3 and/or LAG3 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of Tim3 and/or LAG3 relative to an untreated and/or undiseased patient.
BRIEF DESCRIPTION OF THE FIGURES
[0022] FIG. 1 is an image showing an overview of PTX-35 treatment during an autoimmunity disease. A treatment timeline is also shown in FIG. 1.
[0023] FIG. 2 is an image showing the study design. [0024] FIG. 3 is a graph showing the body weight of the mice before disease induction. mPTX-35 expanded Tregs prior to model start. The measurements in FIG. 3 show body weight (g) on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice. G4 mPTX-35 was administered on both day -7 and day 7.
[0025] FIG. 4 is a graph showing the body weight of the mice during disease induction. mPTX-35 expanded conventional T cells (“Tconv”) at model start. The measurements in FIG. 4 show body weight (g) on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice. G5 mPTX-35 was administered on both day 0 and day 14.
[0026] FIG. 5 is a graph showing the body weight of the mice after disease induction. mPTX-35 expanded both Tregs and Tconv during model disease. The measurements in FIG. 5 show body weight (g) on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. G6 mPTX-35 was administered on both day 7 and day 21.
[0027] FIG. 6 is a chart showing the clinical assessment of EAE with respect to scores and clinical signs.
[0028] FIG. 7 is a graph showing the clinical score of the mice before disease induction. The measurements in FIG. 7 show the average clinical score on the x-axis and the number of days after immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice. G4 mPTX-35 was administered on both day -7 and day 7.
[0029] FIG. 8 is a graph showing the clinical score of the mice during disease induction. The measurements in FIG. 8 show the average clinical score on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice. G5 mPTX-35 was administered on both day 0 and day 14.
[0030] FIG. 9 is a graph showing the clinical score of the mice after disease induction. The measurements in FIG. 9 show the average clinical score on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. G6 mPTX-35 was administered on both day 7 and day 21.
[0031] FIG. 10 is a graph showing the disease incidence in the mice before disease induction. The measurements in FIG. 10 show the percent incidence on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice. G4 mPTX-35 was administered on both day -7 and day 7.
[0032] FIG. 11 is a graph showing the disease incidence in the mice during disease induction. The measurements in FIG. 11 show the percent incidence on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice. G5 mPTX-35 was administered on both day 0 and day 14.
[0033] FIG. 12 is a graph showing the disease incidence in the mice after disease induction. The measurements in FIG. 12 show the percent incidence on the x-axis and the number of days after first immunization on the y-axis for G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. G6 mPTX-35 was administered on both day 7 and day 21.
[0034] FIG. 13 is a graph showing the clinical score area under the curve (“AUC”) measurements for the G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
[0035] FIG. 14 is a graph showing percent survival (x-axis) and days elapsed (y-axis) before disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice.
[0036] FIG. 15 is a graph showing percent survival (x-axis) and days elapsed (y-axis) during disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice.
[0037] FIG. 16 is a graph showing percent survival (x-axis) and days elapsed (y-axis) after disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
[0038] FIG. 17 is a chart showing the evaluation of disease progression as it relates to the spinal cord.
[0039] FIG. 18 is a graph showing the histology inflammation score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-353 mg/kg), G5 (mPTX-353 mg/kg), and G6 (mPTX-353 mg/kg) mice. PTX-35 mediated Treg expansion reduced central nervous system pathologies.
[0040] FIG. 19 is a graph showing the histology lesion score for G2 (vehicle), G3 (Dex 1 mg/kg),
G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. PTX-35 mediated Treg expansion reduced central nervous system pathologies. [0041] FIG. 20 is a graph showing the histology demyelination score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
[0042] FIG.21 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G1 normal mice (left panel), and G2 vehicle mice (right panel).
[0043] FIG.22 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G3 Dex treated mice (left panel), and G4 mPTX-35 treated mice (right panel).
[0044] FIG.23 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G5 mPTX-35 treated mice (left panel), and G6 mPTX-35 treated mice (right panel).
[0045] FIG. 24 shows the gating strategy for the Flow cytometry data.
[0046] FIG. 25 is a graph showing the percentage of regulatory T cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
[0047] FIG. 26 is a graph showing the percentage of Th17 cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-353 mg/kg), G5 (mPTX- 35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
[0048] FIG. 27A - FIG. 27B show experimental design of PTX-35 treatment of EAE mouse model disease progression. FIG. 27A is an image showing the study design of C57BL/6 mice immunized with MOG 35-55 and treated with PTX-35 at day 4 and 8, and day 12 and 16. FIG. 27B is a graph showing the paralytic score of MOG 35-55 immunized mice treated with PTX-35 at various days post immunization compared to control mice.
[0049] FIG. 28A - FIG. 28B show experimental data demonstrating the weight changes of MOG
35-55 immunized mice treated with PTX-35 at various days post immunization compared to control mice. FIG. 28A is a graph showing the weight change in grams of MOG 35-55 immunized mice treated with PTX- 35 at various days post immunization. FIG. 28B is a graph showing change in the percent of initial body weight of MOG 35-55 immunized mice treated with PTX-35 at various days post immunization.
[0050] FIG. 29 shows images of spinal cords from B6 mice immunized with MOG 35-55 taken at
Day 30 post immunization compared to non-immunized mice and mice immunized with isotype control antibodies. [0051] FIG. 30 is a graph showing inflammatory scoring of B6 mice immunized with MOG 35-55 take at Day 30 post immunization compared to non-immunized mice and mice immunized with isotype control antibodies. Significance relative to isotype (*<0.05, **<0.005, ***<0.0005).
[0052] FIG. 31 is a flow cytometry histogram showing percentage distribution of FoxP3+ regulatory
T cells and conventional CD4+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0053] FIG. 32 is a graph of experimental data showing the percentage of FoxP3+ regulatory T cells and conventional CD4+T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (*<0.05, **<0.005, ***<0.0005).
[0054] FIG. 33 is a flow cytometry histogram showing percentage of ICOS+T cells in splenic CD4+
FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35- 55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0055] FIG. 34 is a graph of experimental data showing percentage of ICOS+ T cells in splenic
CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (*<0.05, **<0.005, ***<0.0005).
[0056] FIG. 35 is a flow cytometry histogram showing percentage of CD103+ T cells in splenic
CD4+ FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0057] FIG. 36 is a graph of experimental data showing percentage of CD103+ T cells in splenic
CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (*<0.05, **<0.005, ***<0.0005).
[0058] FIG. 37 is a flow cytometry histogram showing percentage distribution of Tim3+LAG3+ T cells in splenic CD8+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0059] FIG. 38 is a graph of experimental data showing the percentage of Tim3+LAG3+ T cells in splenic CD8+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16. Significance relative to isotype (*<0.05, **<0.005, ***<0.0005).
[0060] FIG. 39 is a graph of experimental data showing percentage of CD44+ T cells in splenic
CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0061] FIG. 40 is a graph of experimental data showing percentage of TIGIT+ T cells in splenic
CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0062] FIG. 41 is a graph of experimental data showing percentage of CTLA4+ T cells in splenic
CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0063] FIG. 42 is a graph of experimental data showing percentage of Ki67+ T cells in splenic
CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0064] FIG. 43 is a graph of experimental data showing MFI of GITR in splenic CD4-H=oxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0065] FIG. 44 is a graph of experimental data showing MFI of CD25 in splenic CD4+FoxP3+ regulatory T cells of non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0066] FIG. 45 is a graph of experimental data showing the percentage of PD-1 + T cells in splenic
CD8+T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0067] FIG. 46 is a graph of experimental data showing the increase in percentage of PD-1+T cells in a total population of T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with PTX- 35 at day 4 and day 8, and MOG 35-55 immunized mice treated with PTX-35 at day 12 and day 16.
[0068] FIG. 47 is an image showing the study design of SJL/J mice immunized with PLP 139-151 and treated with PTX-35 at day -4 and 0, day 4 and 8, day 12 and 16, or day 16 and 20.
[0069] FIG. 48 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
PTX-35 at day -4 and day 0 post immunization compared to control mice.
[0070] FIG. 49 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
PTX-35 at day 4 and day 8 post immunization compared to control mice.
[0071] FIG. 50 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
PTX-35 at day 12 and day 16 post immunization compared to control mice.
[0072] FIG. 51 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with PTX-35 at day 16 and day 20 post immunization compared to control mice.
[0073] FIG. 52 is a graph showing the paralytic score of PLP 139-151 immunized mice treated with
PTX-35 at day 4, day 8, day 16, and day 20 post immunization compared to control mice.
DETAILED DESCRIPTION
[0074] In one aspect, the present disclosure relates to compositions and methods with TNFRSF25 agonistic antibodies, including their use in the treatment of various autoimmune and/or neurodegenerative diseases.
TNFRSF25 Agonistic Antibodies
[0075] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a TNFRSF25 agonistic antibody or antigen binding fragment, such as PTX-35, which is described in PCT/US2017/036817 (WO2017214547), which is incorporated herein by reference in its entirety.
[0076] In any of the methods of treatment or prevention of a disease or condition in accordance with the present disclosure, the TNFRSF25 agonistic antibody or antigen binding fragment thereof can comprise (i) a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 sequences, wherein the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO: 1), the heavy chain CDR2 sequence is YISSASGUSYADAVRG (SEQ ID NO: 2); and the heavy chain CDR3 sequence is DPAYTGLYALDF (SEQ ID NO: 3) or DPPYSGLYALDF (SEQ ID NO: 4); and (ii) a light chain variable region comprising light chain CDR1, CDR2, and CDR3 sequences, wherein the light chain CDR1 sequence is TLSSELSWYTIV (SEQ ID NO: 5), the light chain CDR2 sequence is LKSDGSHSKGD (SEQ ID NO: 6), and the light chain CDR3 sequence is CGAGYTLAGQYGWV (SEQ ID NO: 7). In embodiments, heavy chain CDR1 , CDR2, and CDR3 sequences and/or light chain CDR1 , CDR2, and CDR3 sequences each include at least one, or at least 2, or at least 3, or at least 4, or at least 5 mutations such as amino acid substitutions. In embodiments, heavy chain CDR1 , CDR2, and CDR3 sequences and/or light chain CDR1 , CDR2, and CDR3 sequences each include more than 5 mutations.
[0077] In embodiments, an TNFRSF25 agonistic antibody includes a set of six CDRs that include no more than two, or no more than three, or no more than four, or no more than five, or no more than six total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 1, 2, 3 or 4, 5, 6, and 7. In embodiments, an TNFRSF25 agonistic antibody includes a set of six CDRs that include at least one, or at least two, or at least three, or at least four, or at least five total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 1 , 2, 3 or 4, 5, 6, and 7. In embodiments, an TNFRSF25 agonistic antibody includes a set of six CDRs that include one, or two, or three, or four, or five, or more than five total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEC ID NOS: 1, 2, 3 or 4, 5, 6, and 7.
[0078] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises variable region framework (FW) sequences juxtaposed between the CDRs according to the formula (FW1 )-(CDR1 )-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4), wherein the variable region FW sequences in the heavy chain variable region are heavy chain variable region FW sequences, and wherein the variable region FW sequences in the light chain variable region are light chain variable region FW sequences. In embodiments, the variable region FW sequences are human. In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises human heavy chain and light chain constant regions. The constant regions can be selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4. For instance, In embodiments, the constant regions are lgG1. In embodiments, the constant regions are lgG4.
[0079] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence
EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTI SRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85% to about 99% identity thereto. In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence
QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85% to about 99% identity thereto.
[0080] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence
EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTI SRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91 %, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99% identity thereto.
[0081] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91 %, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99% identity thereto.
[0082] In embodiments, a heavy chain variable region has an amino acid sequence of SEQ ID
NO: 8, or an antigen binding fragment thereof, but with one to 24 sequence modifications, as well as polypeptides having at least about 80% (e.g., about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99%) amino acid sequence identity to SEQ ID NO: 8, or an antigen binding fragment thereof. In embodiments, a heavy chain variable region polypeptide can contain 24 or less (e.g., 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, ten, nine, eight, seven, six, five, four, three, two, or one) amino acid substitution as compared to SEQ ID NO: 8, or an antigen binding fragment thereof.
[0083] In some embodiment, a light chain variable region has an amino acid sequence of SEQ ID
NO: 9, or an antigen binding fragment thereof, but with one to 23 sequence modifications, as well as polypeptides having at least about 80% (e.g., about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99%) amino acid sequence identity to SEQ ID NO: 9, or an antigen binding fragment thereof. In embodiments, a light chain variable region polypeptide can contain 23 or less (e.g., 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, ten, nine, eight, seven, six, five, four, three, two, or one) amino acid substitutions as compared to SEQ ID NO: 9, or an antigen binding fragment thereof.
[0084] In embodiments TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises is the 4C12 antibody or antigen binding fragment thereof. In embodiments TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises is a humanized 4C12.
[0085] In embodiments, an TNFRSF25 agonistic antibody or antigen binding fragment thereof is
PTX-25 antibody or antigen binding fragment thereof. The PTX-25 antibody is described, for example, in PCT/US2015/061082 (WO2016081455), which is incorporated herein by reference in its entirety. In embodiments, an TNFRSF25 agonistic antibody comprises (i) a heavy chain variable region sequence comprising the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEAS GFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTISRDNAKNSLFLQMNNLKSEDTAMYY CARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 10) or the amino acid sequence of SEQ ID NO: 10 with no more than 12 total amino acid substitutions (e.g., no more than 12, or no more than 11, or no more than ten, or no more than nine, or no more than eight, or no more than seven, or no more than six, or no more than five, or no more than four, or no more than three, or no more than two total amino acid substitutions); and (ii) a light chain variable region sequence comprising the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSEL
SSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSSGAHRYLSISNVQSEDDATYFCGAGYTL AGQYGWVFGSGTKVTVL (SEQ ID NO: 11) or the amino acid sequence of SEQ ID NO: 11 with no more than 11 total amino acid substitutions (e.g., no more than 11, or no more than ten, or no more than nine, or no more than eight, or no more than seven, or no more than six, or no more than five, or no more than four, or no more than three, or no more than two total amino acid substitutions, or no more than one total amino acid substitution). An amino acid substitution refers to the replacement of one amino acid residue with another amino acid in a peptide sequence.
[0086] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEAS
GFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAVRGRFTISRDNAKNSLFLQMNNLKSEDTAMYY CARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 10) or an amino acid sequence of at least about 85 to about 99% (e.g., an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91%, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99%) identity thereto.
[0087] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDRFSGSSS GAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 11), or an amino acid sequence of at least about 85 to about 99% (e.g., an amino acid sequence of at least about 85%, or at least about 86%, or at least about 87%, or at least about 88%, or at least about 89%, or at least about 90%, or at least about 91 %, or at least about 92%, or at least about 93%, or at least about 94%, or at least about 95%, or at least about 96%, or at least about 97%, or at least about 98%, or at least about 99%) identity thereto.
[0088] In embodiments, an TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises an isolated heavy chain variable region polypeptide that binds specifically to TNFRSF25, where the polypeptide includes heavy chain CDR1, CDR2, and CDR3 sequences, where the CDR1 sequence is GFTFSNHDLN (SEQ ID NO: 12), the CDR2 sequence is YISSASGLISYADAVRG (SEQ ID NO: 13); and (c) the CDR3 sequence is DPPYSGLYALDF (SEQ ID NO: 14). The isolated heavy chain variable region polypeptide can further include variable region heavy chain framework (FW) sequences juxtaposed between the heavy chain CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4). The heavy chain framework sequences can be human. In embodiments, the isolated heavy chain variable region polypeptide can be combination with a light chain variable region polypeptide. In embodiments, the light chain variable region polypeptide comprises light chain CDR1, CDR2, and CDR3 sequences, wherein the CDR1 sequence is TLSSELSSYTIV (SEQ ID NO: 15), the CDR2 sequence is LKSDGSHSKGD (SEQ ID NO: 16), and the CDR3 sequence is GAGYTLAGQYGWV (SEQ ID NO: 17). The TNFRSF25 agonistic antibody can be a humanized monoclonal antibody that specifically binds to TNFRSF25.
[0089] In embodiments, an TNFRSF25 agonistic antibody includes a set of six CDRs that include no more than two, or no more than three, or no more than four, or no more than five, or no more than six total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 12, 13, 14, 15, 16, and 17. In embodiments, an TNFRSF25 agonistic antibody includes a set of six CDRs that include at least one, or at least two, or at least three, or at least four, or at least five, or at least 6 total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 12, 13, 14, 15, 16, and 17. In embodiments, an TNFRSF25 agonistic antibody includes a set of six CDRs that include one, or two, or three, or four, or five, or more than five total amino acid substitutions in the set of six CDRs having the amino acid sequences set forth in SEQ ID NOS: 12, 13, 14, 15, 16, and 17.
[0090] In embodiments, the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain comprising an amino acid of SEQ ID NO: 18, 19, 20, or 21, and a light chain comprising an amino acid of SEQ ID NO: 22 or 23, as follows:
[0091] EVQLVESGGGLVQPGGSLRLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLIS
YADAVRGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 18),
[0092] EVQLVESGGGLVQPGGSLRLSCAASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLIS
YADAVRGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 19),
[0093] EVQLVESGGGLVQPGGSLRLSCEASGFTFSNHDLNWVRQAPGKGLEWVSYISSASGLIS
YADAVRGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 20),
[0094] EVQLVESGGGLVQPGGSLRLSCAASGFTFSNHDLNWVRQAPGKGLEWVSYISSASGLIS
YADAVRGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 21),
[0095] QPVLTQSSSASASLGSSVKLTCTLSSELSSYTIVWHQQQPGKAPRYLMYLKSDGSHSKG
DGVPDRFSGSSSGADRYLTISNLQSEDEADYYCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 22),
[0096] QLVLTQSPSASASLGASVKLTCTLSSELSSYTIVWHQQQPEKGPRYLMYLKSDGSHSKG
DGIPDRFSGSSSGAERYLTISSLQSEDEADYYCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 23).
[0097] In embodiments, an TNFRSF25 agonistic antibody or antigen binding fragment thereof is any of the antibodies or antibody fragments (or combinations thereof) described in PCT/US2015/061082 (WO2016081455). In embodiments, variable region light chain framework (FW) sequences can be juxtaposed between the light chain CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)- (CDR3)-(FW4). The light chain framework sequences can be human. The antibody or antigen binding fragment can further include a human constant region (e.g., a constant region selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4), or a murine constant region (e.g., a constant region selected from the group consisting of murine lgG1, lgG2A, lgG2B, and lgG3).
[0098] In embodiments, an TNFRSF25 agonistic antibody or antigen binding fragment thereof is as described in PCT/US2010/044218 (WQ2011017303), which is incorporated herein by reference in its entirety.
[0099] Embodiments of the present disclosure make use of TNFRSF25 agonistic antibodies or antigen binding fragment thereof. In embodiments, the antibody is an antibody (e.g., human, hamster, feline, mouse, cartilaginous fish, or camelid antibodies), and any derivative or conjugate thereof, that specifically binds to TNFRSF25. Non-limiting examples of antibodies include monoclonal antibodies, polyclonal antibodies, humanized antibodies, multi-specific antibodies (e.g., bi-specific antibodies), single-chain antibodies (e.g., single-domain antibodies, camelid antibodies, and cartilaginous fish antibodies), chimeric antibodies, feline antibodies, and felinized antibodies. Monoclonal antibodies are homogeneous populations of antibodies to a particular epitope of an antigen. Polyclonal antibodies are heterogeneous populations of antibody molecules that are contained in the sera of the immunized animals.
[00100] An isolated polypeptide can yield a single major band on a non-reducing polyacrylamide gel. An isolated polypeptide can be at least about 75% pure {e.g., at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% pure). Isolated polypeptides can be obtained by, for example, extraction from a natural source, by chemical synthesis, or by recombinant production in a host cell or transgenic plant, and can be purified using, for example, affinity chromatography, immunoprecipitation, size exclusion chromatography, and ion exchange chromatography. The extent of purification can be measured using any appropriate method, including, without limitation, column chromatography, polyacrylamide gel electrophoresis, or high- performance liquid chromatography.
[00101] In embodiments, an antigen binding fragment that specifically binds to TNFRSF25 is provided. Such antigen binding fragment, in embodiments, is any portion of a full-length antibody that contains at least one variable domain (e.g., a variable domain of a mammalian (e.g., feline, human, hamster, or mouse) heavy or light chain immunoglobulin, a camelid variable antigen binding domain (VHH), or a cartilaginous fish immunoglobulin new antigen receptor (Ig-NAR) domain) that is capable of specifically binding to an antigen. Non-limiting examples of antibody fragments include Fab, Fab’, F(ab’)2, and Fv fragments, diabodies, linear antibodies, and multi-specific antibodies formed from antibody fragments. Additional antibody fragments containing at least one camelid VHH domain or at least one cartilaginous fish Ig-NAR domain include mini-bodies, micro-antibodies, subnano-antibodies, and nano-antibodies, and any of the other forms of antibodies described, for example, in U.S. Publication No. 2010/0092470.
[00102] An antibody can be of the IgA-, IgD-, IgE, IgG- or IgM-type, including IgG- or IgM-types such as, without limitation, lgG1-, lgG2-, lgG3-, lgG4-, lgM1- and lgM2-types. For example, in some cases, the antibody is of the lgG1-, lgG2- or lgG4- type.
[00103] In embodiments, antibodies as provided herein can be fully human or humanized antibodies. In embodiments, the human antibody is an antibody that is encoded by a nucleic acid (e.g., a rearranged human immunoglobulin heavy or light chain locus) present in the genome of a human. In embodiments, a human antibody can be produced in a human cell culture (e.g., feline hybridoma cells). In embodiments, a human antibody can be produced in a non-human cell (e.g., a mouse or hamster cell line). In embodiments, a human antibody can be produced in a bacterial or yeast cell.
[00104] Human antibodies can avoid certain problems associated with xenogeneic antibodies, such as antibodies that possess murine or rat variable and/or constant regions. For example, because the effector portion is human, it can interact better with other parts of the human immune system, e.g., to destroy target cells more efficiently by complement-dependent cytotoxicity or antibody-dependent cellular cytotoxicity. In addition, the human immune system should not recognize the antibody as foreign. Further, half-life in human circulation will be similar to naturally occurring human antibodies, allowing smaller and less frequent doses to be given. Methods for preparing human antibodies are known in the art.
[00105] In embodiments, the antibody is a humanized antibody, e.g., an antibody that contains minimal sequence derived from non-human {e.g., mouse, hamster, rat, rabbit, or goat) immunoglobulin. Humanized antibodies generally are chimeric or mutant monoclonal antibodies from mouse, rat, hamster, rabbit or other species, bearing human constant and/or variable region domains or specific changes. In nonlimiting examples, humanized antibodies are human antibodies (recipient antibody) in which hypervariable region (HVR) residues of the recipient antibody are replaced by HVR residues from a non-human species (donor) antibody, such as a mouse, rat, rabbit, or goat antibody having the desired specificity, affinity, and capacity. In embodiments, Fv framework residues of the human immunoglobulin can be replaced by corresponding non-human residues. In embodiments, humanized antibodies can contain residues that are not found in the recipient antibody or in the donor antibody. Such modifications can be made to refine antibody performance, for example.
[00106] In embodiments, a humanized antibody can contain substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops (CDRs) correspond to those of a non-human immunoglobulin, while all or substantially all of the framework regions are those of a human immunoglobulin sequence. A humanized antibody also can contain at least a portion of an immunoglobulin constant (Fc) region, typically that of a human immunoglobulin.
[00107] In embodiments, a humanized antibody or antigen binding fragment as provided herein can have reduced or minimal effector function (e.g., as compared to corresponding non-humanized antibody), such that it does not stimulate effector cell action to the same extent that a corresponding non-humanized antibody would.
[00108] Techniques for generating humanized antibodies are well known to those of skill in the art. In embodiments, controlled rearrangement of antibody domains joined through protein disulfide bonds to form new, artificial protein molecules or “chimeric” antibodies can be utilized (Konieczny et al., Haematologia (Budap.) 14:95, 1981). Recombinant DNA technology can be used to construct gene fusions between DNA sequences encoding mouse antibody variable light and heavy chain domains and human antibody light and heavy chain constant domains (Morrison et al, Proc Natl Acad Sci USA 81 :6851 , 1984). For example, DNA sequences encoding antigen binding portions or CDRs of murine monoclonal antibodies can be grafted by molecular means into DNA sequences encoding frameworks of human antibody heavy and light chains (Jones et al., Nature 321:522, 1986; and Riechmann et al., Nature 332:323, 1988). Expressed recombinant products are called “reshaped” or humanized antibodies, and contain the framework of a human antibody light or heavy chain and antigen recognition portions, CDRs, of a murine monoclonal antibody.
[00109] Other methods for designing heavy and light chains and for producing humanized antibodies are described in, for example, U.S. Patent Nos. 5,530,101; 5,565,332; 5,585,089; 5,639,641; 5,693,761; 5,693,762; and 5,733,743. Yet additional methods for humanizing antibodies are described in U.S. Patent Nos. 4,816,567; 4,935,496; 5,502,167; 5,558,864; 5,693,493; 5,698,417; 5,705,154; 5,750,078; and 5,770,403, for example.
[00110] In embodiments, the antibody is a single-chain antibody, e.g. a single polypeptide that contains at least one variable binding domain (e.g., a variable domain of a mammalian heavy or light chain immunoglobulin, a camelid VHH, or a cartilaginous fish (e.g., shark) Ig-NAR domain) that is capable of specifically binding to an antigen. Non-limiting examples of single-chain antibodies include single-domain antibodies.
[00111] In embodiments, the antibody is a single-domain antibody, e.g. a polypeptide that contains one camelid VHH or at least one cartilaginous fish Ig-NAR domain that is capable of specifically binding to an antigen. Non-limiting examples of single-domain antibodies are described, for example, in U.S. Publication No. 2010/0092470.
[00112] In embodiments, the antibody specifically binds to a particular antigen, e.g., TNFRSF25, when it binds to that antigen in a sample, and does not recognize and bind, or recognizes and binds to a lesser extent, other molecules in the sample. In embodiments, an antibody or an antigen binding fragment thereof can selectively bind to an epitope with an affinity (Kd) equal to or less than, for example, about 1 x 106M {e.g., equal to or less than about 1 x 109 M, equal to or less than about 1 x 1010 M, equal to or less than about 1 x 1011 M, or equal to or less than about 1 x 1012M) in phosphate buffered saline. The ability of an antibody or antigen binding fragment to specifically bind a protein epitope can be determined using any of the methods known in the art or those methods described herein (e.g., by Biacore/Surface Plasmon Resonance). This can include, for example, binding to TNFRSF25 on live cells as a method to stimulate caspase activation in live transformed cells, binding to an immobilized target substrate including human TNFRSF25 fusion proteins as detected using an ELISA method, binding to TNFRSF25 on live cells as detected by flow cytometry, or binding to an immobilized substrate by surface plasmon resonance (including ProteOn).
[00113] Antibodies having specific binding affinity for TNFRSF25 can be produced using standard methods. For example, a TNFRSF25 polypeptide can be recombinantly produced, purified from a biological sample {e.g., a heterologous expression system), or chemically synthesized, and used to immunize host animals, including rabbits, chickens, mice, guinea pigs, or rats. Various adjuvants that can be used to increase the immunological response depend on the host species and include Freund’s adjuvant (complete and incomplete), mineral gels such as aluminum hydroxide, surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanin and dinitrophenol. Monoclonal antibodies can be prepared using aTNFRSF25 polypeptide and standard hybridoma technology. In particular, monoclonal antibodies can be obtained by any technique that provides for the production of antibody molecules by continuous cell lines in culture such as described by Kohler et al. {Nature 256:495, 1975), the human B-cell hybridoma technique of Kosbor et al. ( Immunology Today, 4:72, 1983) or Cote et al. {Proc. Natl. Acad. Sci. USA, 80:2026, 1983), and the EBV-hybridoma technique described by Cole et al. (Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp. 77-96, 1983). Such antibodies can be of any immunoglobulin class including IgG, IgM, IgE, IgA, IgD, and any subclass thereof. The hybridoma producing the monoclonal antibodies can be cultivated in vitro and in vivo. [00114] In embodiments, amino acid substitutions can be made by selecting conservative substitutions that do not differ significantly in their effect on maintaining (a) the structure of the peptide backbone in the area of the substitution, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. For example, naturally occurring residues can be divided into groups based on side-chain properties: (1) hydrophobic amino acids (norleucine, methionine, alanine, valine, leucine, and isoleucine); (2) neutral hydrophilic amino acids (cysteine, serine, and threonine); (3) acidic amino acids (aspartic acid and glutamic acid); (4) basic amino acids (asparagine, glutamine, histidine, lysine, and arginine); (5) amino acids that influence chain orientation (glycine and proline); and (6) aromatic amino acids (tryptophan, tyrosine, and phenylalanine). Substitutions made within these groups can be considered conservative substitutions. Non-limiting examples of conservative substitutions include, without limitation, substitution of valine for alanine, lysine for arginine, glutamine for asparagine, glutamic acid for aspartic acid, serine for cysteine, asparagine for glutamine, aspartic acid for glutamic acid, proline for glycine, arginine for histidine, leucine for isoleucine, isoleucine for leucine, arginine for lysine, leucine for methionine, leucine for phenylalanine, glycine for proline, threonine for serine, serine for threonine, tyrosine for tryptophan, phenylalanine for tyrosine, and/or leucine for valine. In embodiments, an amino acid substitution can be nonconservative, such that a member of one of the amino acid classes described above is exchanged for a member of another class.
Pharmaceutical Compositions
[00115] In addition, the disclosure also provides pharmaceutical compositions for the present methods of treating, which include an antibody or antigen binding fragment, as described herein, in combination with a pharmaceutically acceptable carrier. A “pharmaceutically acceptable carrier” (also referred to as an “excipient” or a “carrier”) is a pharmaceutically acceptable solvent, suspending agent, stabilizing agent, or any other pharmacologically inert vehicle for delivering one or more therapeutic compounds to a subject {e.g., a mammal, such as a human, non-human primate, dog, cat, sheep, pig, horse, cow, mouse, rat, or rabbit), which is nontoxic to the cell or subject being exposed thereto at the dosages and concentrations employed. Pharmaceutically acceptable carriers can be liquid or solid, and can be selected with the planned manner of administration in mind so as to provide for the desired bulk, consistency, and other pertinent transport and chemical properties, when combined with one or more of therapeutic compounds and any other components of a given pharmaceutical composition. Typical pharmaceutically acceptable carriers that do not deleteriously react with amino acids include, by way of example and not limitation: water, saline solution, binding agents {e.g., polyvinylpyrrolidone or hydroxypropyl methylcell ulose), fillers {e.g., lactose and other sugars, gelatin, or calcium sulfate), lubricants {e.g., starch, polyethylene glycol, or sodium acetate), disintegrates {e.g., starch or sodium starch glycolate), and wetting agents {e.g., sodium lauryl sulfate). Pharmaceutically acceptable carriers also include aqueous pH buffered solutions or liposomes (small vesicles composed of various types of lipids, phospholipids and/or surfactants which are useful for delivery of a drug to a mammal). Further examples of pharmaceutically acceptable carriers include buffers such as phosphate, citrate, and other organic acids, antioxidants such as ascorbic acid, low molecular weight (less than about 10 residues) polypeptides, proteins such as serum albumin, gelatin, or immunoglobulins, hydrophilic polymers such as polyvinylpyrrolidone, amino acids such as glycine, glutamine, asparagine, arginine or lysine, monosaccharides, disaccharides, and other carbohydrates including glucose, mannose or dextrins, chelating agents such as EDTA, sugar alcohols such as mannitol or sorbitol, salt-forming counterions such as sodium, and/or nonionic surfactants such as TWEEN™, polyethylene glycol (PEG), and PLURONICS™.
[00116] Pharmaceutical compositions can be formulated by mixing one or more active agents with one or more physiologically acceptable carriers, diluents, and/or adjuvants, and optionally other agents that are usually incorporated into formulations to provide improved transfer, delivery, tolerance, and the like. A pharmaceutical composition can be formulated, e.g., in lyophilized formulations, aqueous solutions, dispersions, or solid preparations, such as tablets, dragees or capsules. A multitude of appropriate formulations can be found in the formulary known to all pharmaceutical chemists: Remington’s Pharmaceutical Sciences (18th ed, Mack Publishing Company, Easton, PA (1990)), particularly Chapter 87 by Block, Lawrence, therein. These formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid (cationic or anionic) containing vesicles (such as LIPOFECTIN™), DNA conjugates, anhydrous absorption pastes, oil-in-water and water-in-oil emulsions, emulsions carbowax (polyethylene glycols of various molecular weights), semi-solid gels, and semi-solid mixtures containing carbowax. Any of the foregoing mixtures may be appropriate in treatments and therapies as described herein, provided that the active agent in the formulation is not inactivated by the formulation and the formulation is physiologically compatible and tolerable with the route of administration. See, also, Baldrick, R egul Toxicol Pharmacol 32:210-218, 2000; Wang, Int JPharm 203:1-60, 2000; Charman, JPharm Sci 89:967-978, 2000; and Powell et al. PDA J Pharm Sci Techno 152:238-311 , 1998), and the citations therein for additional information related to formulations, excipients and carriers well known to pharmaceutical chemists. [00117] Pharmaceutical compositions include, without limitation, solutions, emulsions, aqueous suspensions, and liposome-containing formulations. These compositions can be generated from a variety of components that include, for example, preformed liquids, self-emulsifying solids and self-emulsifying semisolids. Emulsions are often biphasic systems comprising of two immiscible liquid phases intimately mixed and dispersed with each other; in general, emulsions are either of the water-in-oil (w/o) or oil-in-water (o/w) variety. Emulsion formulations have been widely used for oral delivery of therapeutics due to their ease of formulation and efficacy of solubilization, absorption, and bioavailability.
[00118] Compositions and formulations can contain sterile aqueous solutions, which also can contain buffers, diluents and other suitable additives {e.g., penetration enhancers, carrier compounds and other pharmaceutically acceptable carriers). Compositions additionally can contain other adjunct components conventionally found in pharmaceutical compositions. Thus, the compositions also can include compatible, pharmaceutically active materials such as, for example, antipruritics, astringents, local anesthetics or antiinflammatory agents, or additional materials useful in physically formulating various dosage forms of the compositions provided herein, such as dyes, flavoring agents, preservatives, antioxidants, opacifiers, thickening agents and stabilizers. Furthermore, the composition can be mixed with auxiliary agents, e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings, and aromatic substances. When added, however, such materials should not unduly interfere with the biological activities of the polypeptide components within the compositions provided herein. The formulations can be sterilized if desired.
[00119] In embodiments, a composition containing an antibody or antigen binding fragment as used herein can be in the form of a solution or powder with or without a diluent to make an injectable suspension. The composition may contain additional ingredients including, without limitation, pharmaceutically acceptable vehicles, such as saline, water, lactic acid, mannitol, or combinations thereof, for example.
[00120] Any appropriate method can be used to administer an antibody or antigen binding fragment as described herein to a mammal. Administration can be, for example, parenteral {e.g., by subcutaneous, intrathecal, intraventricular, intramuscular, or intraperitoneal injection, or by intravenous drip). Administration can be rapid (e.g., by injection) or can occur over a period of time (e.g., by slow infusion or administration of slow release formulations). In embodiments, administration can be topical (e.g., transdermal, sublingual, ophthalmic, or intranasal), pulmonary (e.g., by inhalation or insufflation of powders or aerosols), or oral. In addition, a composition containing an antibody or antigen binding fragment as described herein can be administered prior to, after, or in lieu of surgical resection of a tumor.
[00121] A composition containing an TNFRSF25 agonistic antibody or antigen binding fragment thereof can be administered to a mammal in any appropriate amount, at any appropriate frequency, and for any appropriate duration effective to achieve a desired outcome. For example, an TNFRSF25 agonistic antibody or antigen binding fragment thereof can be administered to a subject in an amount effective to stimulate proliferation of T cells in vitro or in vivo (e.g., human, murine, hamster, or macaque T cells, including CD8+ T cells and/or CD4+FoxP3+ regulatory T cells), to stimulate apoptosis of tumor cells that express TNFRSF25, to reduce tumor size, or to increase progression-free survival of a cancer patient. In embodiments, an TNFRSF25 agonistic antibody or antigen binding fragment thereof can be administered at a dosage of about 0.1 mg/kg to about 10 mg/kg (e.g., about 0.1 mg/kg to about 1 mg/kg, about 1 mg/kg to about 5 mg/kg, or about 5 mg/kg to about 10 mg/kg), and can be administered once every one to three weeks (e.g., every week, every 10 days, every two weeks, or every three weeks).
[00122] In some cases, a composition containing TNFRSF25 agonistic antibody or antigen binding fragment thereof as described herein can be administered to a subject in an amount effective to increase proliferation of T cells (e.g., by at least about 10 percent, about 20 percent, about 25 percent, about 50 percent, about 60 percent, about 70 percent, about 75 percent, about 80 percent, about 90 percent, about 100 percent, or more than 100 percent), as compared to the “baseline” level of T cell proliferation in the subject prior to administration of the composition, or as compared to the level of T cell proliferation in a control subject or population of subjects to whom the composition was not administered. The T cells can be, for example, CD4+FoxP3+ T cells, regulatory T cells. Any suitable method can be used to determine whether or not the level of T cell proliferation is increased in the subject. Such methods can include, without limitation, flow cytometry analysis of antigen specific T cells (e.g., flow cytometry analysis of the proportion of antigen specific CD4+FoxP3+ T cells as a fraction of the total CD4+ T cell pool), analysis of cell proliferation markers (e.g., expression of Ki67) in CD4+ T cells, increased counts of CD4+ T cells, or increased proportions of individual TCR sequences of a particular clone of CD4+ T cells.
Methods of Treatment
[00123] In embodiments, the present TNFRSF25-specific agent is used to treat or prevent various autoimmune and/or neurodegenerative diseases. In embodiments, the neurodegenerative disease is selected from MS (including without limitation the subtypes described herein), Alzheimer's disease (including, without limitation, early-onset Alzheimer's, late-onset Alzheimer’s, and familial Alzheimer’s disease (FAD), Parkinson’s disease and parkinsonism (including, without limitation, Idiopathic Parkinson's disease, vascular parkinsonism, drug-induced parkinsonism, dementia with Lewy bodies, inherited Parkinson's, juvenile Parkinson's), Huntington's disease, amyotrophic lateral sclerosis (ALS, including, without limitation, sporadic ALS, familial ALS, Western Pacific ALS, juvenile ALS, Hiramaya Disease). In embodiments, the autoimmune disease is selected from Addison's disease, rheumatoid arthritis, type 1 diabetes, vasculitis, alopecia areata, lupus, polymyalgia rheumatica, ankylosing spondylitis, celiac disease, Sjogren's syndrome, and temporal arteritis, Graves' disease, Hashimoto's thyroiditis, Dermatomyositis, Myasthenia gravis, and Pernicious anemia.
[00124] In embodiments, the autoimmune inflammatory disease is a demyelinating disease. For example, In embodiments the inflammation is in the central nervous system (CNS). In embodiments, the demyelinating disease is multiple sclerosis (MS). In embodiments, the demyelinating disease is clinically isolated syndrome (CIS). In embodiments, the method prevents or slows the progression of CIS to MS. In embodiments, the demyelinating disease is radiologically isolated syndrome (RIS). In embodiments, the method prevents or slows the progression of RIS to MS. In embodiments, the patient is likely to develop MS.
[00125] In embodiments, the present disclosure is used to treat or prevent MS. In embodiments, TNFRSF25 agonistic antibodies, or fragments thereof as described herein are used to eliminate and reduce multiple MS symptoms. Illustrative symptoms associated with multiple sclerosis, which can be prevented or treated with the compositions and methods described herein, include: optic neuritis, diplopia, nystagmus, ocular dysmetria, internuclear ophthalmoplegia, movement and sound phosphenes, afferent pupillary defect, paresis, monoparesis, paraparesis, hemiparesis, quadraparesis, plegia, paraplegia, hemiplegia, tetraplegia, quadraplegia, spasticity, dysarthria, muscle atrophy, spasms, cramps, hypotonia, clonus, myoclonus, myokymia, restless leg syndrome, footdrop, dysfunctional reflexes, paresthesia, anesthesia, neuralgia, neuropathic and neurogenic pain, Lhermitte’s sign, proprioceptive dysfunction, trigeminal neuralgia, ataxia, intention tremor, dysmetria, vestibular ataxia, vertigo, speech ataxia, dystonia, dysdiadochokinesia, frequent micturation, bladder spasticity, flaccid bladder, detrusor-sphincter dyssynergia, erectile dysfunction, anorgasmy, frigidity, constipation, fecal urgency, fecal incontinence, depression, cognitive dysfunction, dementia, mood swings, emotional lability, euphoria, bipolar syndrome, anxiety, aphasia, dysphasia, fatigue, Uhthoffs symptom, gastroesophageal reflux, and sleeping disorders. Mitigation or amelioration or one more of these symptoms in a subject can be achieved by the one or more TNFRSF25 agonistic antibodies, or fragments thereof and one or more additional agents as described herein.
[00126] In embodiments, the present TNFRSF25-specific agent is used to treat or prevent clinically isolated syndrome (CIS). A clinically isolated syndrome (CIS) is a single monosymptomatic attack compatible with MS, such as optic neuritis, brain stem symptoms, and partial myelitis. Patients with CIS that experience a second clinical attack are generally considered to have clinically definite multiple sclerosis (CDMS). Over 80 percent of patients with a CIS and MRI lesions go on to develop MS, while approximately 20 percent have a self-limited process. Patients who experience a single clinical attack consistent with MS may have at least one lesion consistent with multiple sclerosis prior to the development of clinically definite multiple sclerosis. In embodiments, the present TNFRSF25-specific agent is used to treat CIS so it does not develop into MS, including, for example RRMS. In embodiments, the patient has CIS. In embodiments, the patient has a monofocal episode of CIS.
[00127] In embodiments, the patient has optic neuritis. In embodiments, the patient has a multifocal episode of CIS. In embodiments, the patient has optic neuritis and numbness or tingling in the legs.
[00128] In embodiments, the present TNFRSF25-specific agent is used to treat or prevent radiologically isolated syndrome (RIS). In RIS incidental imaging findings suggest inflammatory demyelination in the absence of clinical signs or symptoms. In embodiments, the present TNFRSF25-specific agent is used to treat MS so it does not develop into MS, including, for example RRMS.
[00129] In embodiments, the present TNFRSF25-specific agent is used to treat benign multiple sclerosis; relapsing-remitting multiple sclerosis (RRMS); secondary progressive multiple sclerosis (SPMS); progressive relapsing multiple sclerosis (PRMS); and primary progressive multiple sclerosis (PPMS).
[00130] In embodiments, the treatment occurs early in disease progression. In embodiments, the treatment occurs before the onset of relapsing-remitting MS (RRMS). In embodiments, the treatment occurs before the onset of secondary progressive MS (SPMS). In embodiments, the treatment occurs before the onset of primary progressive MS (PPMS).
[00131] Benign multiple sclerosis is a retrospective diagnosis which is characterized by 1-2 exacerbations with complete recovery, no lasting disability and no disease progression for 10-15 years after the initial onset. Benign multiple sclerosis may, however, progress into other forms of multiple sclerosis. In embodiments, the present TNFRSF25-specific agent is used to treat benign multiple sclerosis so it does not develop into MS. [00132] Patients suffering from RRMS experience sporadic exacerbations or relapses, as well as periods of remission. Lesions and evidence of axonal loss may or may not be visible on MRI for patients with RRMS. In embodiments, the present TNFRSF25-specific agent is used to treat RRMS. In embodiments, RRMS includes patients with RRMS; patients with SPMS and superimposed relapses; and patients with CIS who show lesion dissemination on subsequent MRI scans according to McDonald's criteria. In embodiments, RRMS includes patients with RRMS; patients with SPMS and superimposed relapses; and patients with CIS who show lesion dissemination on subsequent MRI scans according to McDonald's criteria. A clinical relapse, which may also be used herein as “relapse,” “confirmed relapse,” or “clinically defined relapse,” is the appearance of one or more new neurological abnormalities or the reappearance of one or more previously observed neurological abnormalities. This change in clinical state must last at least 48 hours and be immediately preceded by a relatively stable or improving neurological state of at least 30 days. In embodiments, an event is counted as a relapse when the subject's symptoms are accompanied by observed objective neurological changes, consistent with an increase of at least 1.00 in the EDSS score or one grade in the score of two or more of the seven FS or two grades in the score of one of FS as compared to the previous evaluation.
[00133] SPMS may evolve from RRMS. Patients afflicted with SPMS have relapses, a diminishing degree of recovery during remissions, less frequent remissions and more pronounced neurological deficits than RRMS patients. Enlarged ventricles, which are markers for atrophy of the corpus callosum, midline center and spinal cord, are visible on MRI of patients with SPMS. In embodiments, the present TNFRSF25- specific agent is used to treat RRMS so it does not develop into SPMS.
[00134] PPMS is characterized by a steady progression of increasing neurological deficits without distinct attacks or remissions. Cerebral lesions, diffuse spinal cord damage and evidence of axonal loss are evident on the MRI of patients with PPMS. PPMS has periods of acute exacerbations while proceeding along a course of increasing neurological deficits without remissions. Lesions are evident on MRI of patients suffering from PRMS. In embodiments, the present TNFRSF25-specific agent is used to treat RRMS and/or SPMS so it does not develop into PPMS.
[00135] In embodiments, the treatment reduces spinal cord inflammation, as compared to before treatment. In embodiments, the treatment reduces axon demyelination, as compared to before treatment.
[00136] In embodiments, the present TNFRSF25-specific agents are used in a method of treatment of relapsing forms of MS. In embodiments, the present TNFRSF25-specific agents are used in a method of treatment of relapsing forms of MS to slow the accumulation of physical disability and/or reduce the frequency of clinical exacerbations, and, optionally, for patients who have experienced a first clinical episode and have MRI features consistent with MS. In embodiments, the present TNFRSF25-specific agents are used in a method of treatment of worsening relapsing-remitting MS, progressive-relapsing MS or secondary- progressive MS to reduce neurologic disability and/or the frequency of clinical exacerbations. In embodiments, the present TNFRSF25-specific agents reduce the frequency and/or severity of relapses.
[00137] In embodiments, the present TNFRSF25-specific agents are used in a method of treatment of relapsing forms of MS in patients who have had an inadequate response to (or are refractory to) one, or two, or three, or four, or five, or six, or seven, or eight, or nine, or ten or more DMTs.
[00138] In embodiments, the subject's symptoms may be assessed quantitatively, such as by EDSS, or decrease in the frequency of relapses, or increase in the time to sustained progression, or improvement in the magnetic resonance imaging (MRI) behavior in frequent, serial MRI studies and compare the patient's status measurement before and after treatment. In a successful treatment, the patient status will have improved {e.g., the EDSS measurement number or frequency of relapses will have decreased, or the time to sustained progression will have increased, or the MRI scans will show less pathology).
[00139] In embodiments, the patient can be evaluated, e.g., before, during or after receiving the present TNFRSF25-specific agent, e.g., for indicia of responsiveness. Various clinical or other indicia of effectiveness of treatment, e.g., EDSS score; MRI scan; relapse number, rate, or severity; multiple sclerosis functional composite (MSFC); multiple sclerosis quality of life inventory (MSQLI); Paced Serial Addition Test (PASAT); symbol digit modalities test (SDMT); 25-foot walk test; 9-hole peg test; low contrast visual acuity; Modified Fatigue Impact Scale; expanded disability status score (EDSS); multiple sclerosis functional composite (MSFC); Beck Depression Inventory; and 7/24 Spatial Recall Test can be used and, in embodiments, the present TNFRSF25-specific agent causes an improvement in one or more of these measures. Further, the patient can be monitored at various times during a regimen. In embodiments, the present TNFRSF25-specific agents cause a disease improvement as assessed by assessment of MacDonald dissemination in space and time. For example, for dissemination in space, lesion imaging, such as, by way of illustration, Barkhof-Tintore MR imaging criteria, may be used, including at least one gadoliniumenhancing lesion or 9 T2 hyperintense lesions; at least one infratentorial lesion; at least one juxtacortical lesion; at least about three periventricular lesions; and a spinal cord lesion. For dissemination in time, MRI can also be used; for example, if an MRI scan of the brain performed at >3 months after an initial clinical event demonstrates a new gadolinium-enhancing lesion, this may indicate a new CNS inflammatory event, because the duration of gadolinium enhancement in MS is usually less than 6 weeks. If there are no gadolinium-enhancing lesions but a new T2 lesion (presuming an MRI at the time of the initial event), a repeat MR imaging scan after another 3 months may be needed with demonstration of a new T2 lesion or gadolinium-enhancing lesion.
[00140] In embodiments, disease effects are assessed using any of the measures described in Lavery, et al. Multiple Sclerosis International, Vol 2014 (2014), Article ID 262350, the entire contents of which are hereby incorporated by reference.
[00141] In embodiments, the present TNFRSF25-specific agent results in one or more of: (a) prevention of worsening in disability defined as deterioration by 1.0 point on EDSS, (b) increase in time to relapse, (c) reduction or stabilization of number and/or volume of gadolinium enhancing lesions, (d) decreased annualized relapse rate, (e) increased relapse duration and severity by NRS score, (f) decrease in disease activity as measured by MRI (annual rate of new or enlarging lesions), (g) lower average number of relapses at 1 year, or 2 years, (h) sustained disease progression as measured by the EDSS at 3 months, (i) prevention of conversion to CDMS, (j) no or few new or enhancing T2 lesions, (k) minimal change in hyperintense T2 lesion volume, (I) increased time to McDonald defined MS, (m) prevention of progression of disability as measured by sustained worsening of EDSS at 12 weeks, (n) reduction in time to relapse at 96 weeks, and (o) reduction or stabilization of brain atrophy {e.g. percentage change from baseline).
[00142] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a decreased rate of relapse {e.g., at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80% or greater reduction in rate of relapse) compared to the rate of relapse before administration {e.g., compared to the rate of relapse following administration for 12 months or for less than 12 months, e.g., about 10, or about 8, or about 4, or about 2 or less months) of treatment, or before commencement of treatment, when measured between 3-24 months {e.g., between 6-18 months, e.g., 12 months) after a previous relapse.
[00143] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a prevention of an increase in EDSS score from a pre-treatment state. The Kurtzke Expanded Disability Status Scale (EDSS) is a method of quantifying disability in multiple sclerosis. The EDSS replaced the previous Disability Status Scales which used to bunch people with MS in the lower brackets. The EDSS quantifies disability in eight Functional Systems (FS) and allows neurologists to assign a Functional System Score (FSS) in each of these. The Functional Systems are: pyramidal, cerebellar, brainstem, sensory, bowel and bladder, visual & cerebral.
[00144] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a decreased EDSS score {e.g., a decrease of 1 , 1.5, 2, 2.5, 3 points or more, e.g., over at least three months, six months, one year, or longer) compared to the EDSS score following administration of the present TNFRSF25-specific agent {e.g. for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before the commencement of treatment).
[00145] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a decreased number of new lesions overall or of any one type {e.g., at least 10%, 20%, 30%, 40% decrease), compared to the number of new lesions following administration of the present TNFRSF25- specific agent for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
[00146] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a decreased number of lesions overall or of any one type {e.g., at least 10%, 20%, 30%, 40% decrease), compared to the number of lesions following administration of the present TNFRSF25-specific agent for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
[00147] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a reduced rate of appearance of new lesions overall or of any one type {e.g., at least 10%, 20%, 30%, 40% reduced rate), compared to the rate of appearance of new lesions following administration for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
[00148] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a decreased increase in lesion area overall or of any one type {e.g., at least 10%, 20%, 30%, 40% decreased increase), compared to an increase in lesion area following administration for 12 months or less than 12 months, e.g., less than 10 months, 8 months, 4 months, or 2 months, or before commencement of treatment. [00149] In certain embodiments, the human has an age in a range of from about 1 to about 18 months old, from about 18 to about 36 months old, from about 1 to about 5 years old, from about 5 to about 10 years old, from about 10 to about 15 years old, from about 15 to about 20 years old, from about 20 to about 25 years old, from about 25 to about 30 years old, from about 30 to about 35 years old, from about 35 to about 40 years old, from about 40 to about 45 years old, from about 45 to about 50 years old, from about 50 to about 55 years old, from about 55 to about 60 years old, from about 60 to about 65 years old, from about 65 to about 70 years old, from about 70 to about 75 years old, from about 75 to about 80 years old, from about 80 to about 85 years old, from about 85 to about 90 years old, from about 90 to about 95 years old or from about 95 to about 100 years old. In embodiments, the human has an age of more than 30 years old.
[00150] In embodiments, the present TNFRSF25-specific agents are used to treat or prevent an autoimmune inflammatory disease selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti-MAG peripheral neuropathy, Charcot-Marie-Tooth disease, hereditary neuropathy with liability to pressure palsy, peripheral neuropathy, optic neuropathy, and progressive inflammatory neuropathy.
[00151] In embodiments, the present TNFRSF25-specific agents are used to prevent or reduce symptoms of an autoimmune inflammatory disease, such as blurred double vision (diplopia), ataxia, clonus, dysarthria, fatigue, clumsiness, hand paralysis, hemiparesis, genital anesthesia, incoordination, paresthesia, ocular paralysis (cranial nerve palsy), impaired muscle coordination, weakness (muscle), loss of sensation, impaired vision, neurological symptoms, unsteady gait, spastic paraparesis, incontinence, hearing problems, and speech problems.
[00152] In embodiments, the treatment expands and/or selectively activates a population of Tregs in the patient. In embodiments, the treatment does not substantially expand and/or selectively activate a population of cytotoxic T cells (Tc cells) and/or helper T cells (Th cells) in the patient. In embodiments, the treatment does not substantially expand and/or selectively activate a population of Th17 cells in the patient. In embodiments, the treatment increases the ratio of Tregs to Tc and/or Th cells in the patient. In embodiments, the treatment increases the ratio of Tregs to Th17 cells in the patient. [00153] In embodiments, the treatment increases the ratio of Tregs to conventional T cells in the patient compared to an untreated patient. In embodiments, the treatment increases the population of Tregs expressing ICOS compared to an untreated patient. In embodiments, the treatment increases the population of Tregs expressing CD103 compared to an untreated patient. In embodiments, the treatment increases the population of Tc cells expressing Tim3 and/or LAG3 compared to an untreated patient.
[00154] In embodiments, the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining the amount of ICOS in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to a patient in need of an increase of ICOS.
[00155] In embodiments, the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining the amount of CD103 in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to a patient in need of an increase of CD103.
[00156] In embodiments, the present disclosure provides a method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising determining the amount of Tim3 and/or LAG3 in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to a patient in need of an increase of Tim3 and/or LAG3.
[00157] A method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising, determining an amount of ICOS protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of ICOS relative to an untreated and/or undiseased patient.
[00158] A method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising, determining an amount of CD103 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of CD103 relative to an untreated and/or undiseased patient.
[00159] A method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising, determining an amount of Tim3 and/or LAG3 protein, nucleic acid, or activity in a sample from the patient, and administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of Tim3 and/or LAG3 relative to an untreated and/or undiseased patient.
[00160] In embodiments, a sample is biological fluid, such as whole blood or whole blood components including red blood cells, white blood cells, platelets, serum and plasma, ascites, vitreous fluid, lymph fluid, synovial fluid, follicular fluid, seminal fluid, amniotic fluid, milk, saliva, sputum, tears, perspiration, mucus, cerebrospinal fluid, and urine. In embodiments, the sample is a body sample from any animal. In embodiments, the sample is from a mammal. In embodiments, the sample is from a human subject.
[00161] In embodiments, an effective amount of the TNFRSF25-specific agents can be any amount that has a desired effect (e.g., treating). In embodiments, dosages can vary depending on the relative potency of individual treatments (e.g., TNFRSF25-specific agents), and can generally be estimated based on ECso found to be effective in in vitro and in vivo animal models. In embodiments, dosage is from 0.01 g to 100 g per kg of body weight. In embodiments, an effective amount of the TNFRSF25-specific agents can be from about 0.1 mg/kg to about 50 mg/kg (e.g., about 0.4 mg/kg, about 2 mg/kg, about 5 mg/kg, about 10 mg/kg, about 20 mg/kg, about 30 mg/kg, about 40 mg/kg, or about 50 mg/kg), or any range there between, such as about 0.1 mg/kg to about 10 mg/kg, about 0.4 mg/kg to about 20 mg/kg, about 2 mg/kg to about 30 mg/kg, or about 5 mg/kg to about 40 mg/kg.
[00162] In embodiments, if a particular subject fails to respond to a particular amount of TNFRSF25- specific agents, then the amount of TNFRSF25-specific agents is increased by, for example, two fold. In embodiments, after receiving this higher concentration, the subject is monitored for both responsiveness to the treatment and toxicity symptoms, and adjustments made accordingly. In embodiments, the effective amount remains constant or is adjusted as a sliding scale or variable dose depending on the subject’s response to treatment. Various factors can influence the actual effective amount used for a particular application. In embodiments, the frequency of administration, duration of treatment, use of multiple treatment agents, route of administration, and severity of the cancer may require an increase or decrease in the actual effective amount administered.
[00163] In embodiments, the frequency of administration of the TNFRSF25-specific agents is any frequency that, for example, can be used to treat or prevent an autoimmune inflammatory disease. In embodiments, the frequency of administration is once or more daily, biweekly, weekly, monthly, or even less. In embodiments, the frequency of administration remains constant or is variable during the duration of treatment. In embodiments, a course of treatment includes rest periods. In embodiments, the TNFRSF25- specific agents are administered over a two-week period followed by a two-week rest period, and such a regimen is repeated multiple times. In embodiments, as with the effective amount, various factors can influence the frequency of administration used for a particular application.
[00164] In embodiments, an effective duration for administering the TNFRSF25-specific agents is any duration that is needed to effectively treat or prevent an autoimmune inflammatory disease. Thus, an effective duration can vary from several days to several weeks, months, or years. In embodiments, an effective duration is for as long as an individual subject is alive. In embodiments, multiple factors influence the actual effective duration used for a particular treatment. In embodiments, an effective duration varies with the frequency of administration, effective amount, use of multiple treatment agents, and route of administration.
[00165] In embodiments, the TNFRSF25-specific agents described herein will typically contain an effective amount of the TNFRSF25-specific agents for achieving the desired effect. As used herein the terms “therapeutically effective amount” and “effective amount,” used interchangeably, applied to a dose or amount refers to a quantity of a composition, compound or pharmaceutical formulation that is sufficient to result in a desired activity upon administration to an animal in need thereof. When a combination of active ingredients is administered, the effective amount of the combination may or may not include amounts of each ingredient that would have been effective if administered individually. The dosage of the therapeutic formulation will vary, depending upon the nature of the disease or condition, the patient’s medical history, the frequency of administration, the manner of administration, the clearance of the agent from the host, and the like. The initial dose may be larger, followed by smaller maintenance doses. The dose may be administered, e.g., weekly, biweekly, daily, semi-weekly, etc., to maintain an effective dosage level.
[00166] In embodiments, the present TNFRSF25-specific agent is administered and is effective to result in a reduced incidence or symptom of optic neuritis ( e.g ., improved vision), compared to the incidence or symptom of optic neuritis following administration for 12 months or for less than 12 months, e.g., less than 10, 8, 4 or less months, or before commencement of treatment.
[00167] In embodiments, the methods described herein are intended for use with any subject that may experience the benefits of these methods. Thus, “subjects,” “patients,” and “individuals” (used interchangeably) include humans as well as non-human subjects, particularly domesticated animals.
[00168] In embodiments, the subject and/or animal is a mammal, e.g., a human, mouse, rat, guinea pig, dog, cat, horse, cow, pig, rabbit, sheep, or non-human primate, such as a monkey, chimpanzee, or baboon. In other embodiments, the subject and/or animal is a non-mammal, such, for example, a zebrafish. In embodiments, the subject and/or animal may comprise fluorescently-tagged cells (with e.g. GFP). In embodiments, the subject and/or animal is a transgenic animal comprising a fluorescent cell.
[00169] In embodiments, methods of the disclosure are useful in treatment a human subject. In embodiments, the human is a pediatric human. In other embodiments, the human is an adult human. In other embodiments, the human is a geriatric human. In other embodiments, the human may be referred to as a patient. In embodiments, the human is a female.
Additional Agents and Combination Therapy
[00170] In embodiments, the present TNFRSF25-specific agents is used in a method of treating multiple sclerosis in combination with one or more of the DMTs described herein {e.g. the agents of Table 1). In embodiments, the present disclosure provides an improved therapeutic effect as compared to use of one or more of the DMTs described herein (e.g. the agents of Table 1) without the one or more TNFRSF25- specific agents. In an embodiment, the combination of the one or more TNFRSF25-specific agents and the one or more DMTs produce synergistic therapeutic effects.
Table 1 - Illustrative Disease Modifying Therapies (DMTS)
Figure imgf000037_0001
Figure imgf000038_0001
Figure imgf000039_0001
Figure imgf000040_0001
Figure imgf000041_0001
Figure imgf000042_0001
[00171] In embodiments, the improved therapeutic benefit of adding an TNFRSF25-specific agent to treatment with a DMT, such as one or more of those of Table 1, is manifested in a further reduction in one or more of relapse rate, e.g. annualized relapse rate (ARR) relative risk reduction over 2 years (%); disability progression e.g. disability progression relative risk reduction (%); reduction in lesions, e.g. MRI lesions - reduction in Gd-enhancing (%) or MRI lesions - reduction in T2-weighted new or enlarging lesions (%). In embodiments, the present TNFRSF25-specific agents provide an improvement in therapeutic response as compared to the results of Table 2 below {e.g. about 10%, or about 20%, or about 30%, or about 40%, or about 50%, or about 60%, or about 70%, or about 80%, or about 90%, or about 100% improvement over the values of Table 2).
Table 2: Illustrative Therapeutic Benefits of DMTs
Figure imgf000042_0002
Figure imgf000043_0001
[00172] In embodiments, the present TNFRSF25-specific agents reduce deleterious effects or dosing features of one or more MS-treating agents described herein. For example, the present TNFRSF25- specific agents may reduce the dose and/or frequency of administration of any of the DMTs, mitigate inconvenient dosing and therefore increase adherence, reduce side effects, allow for access to further therapies, and bridge the therapeutic benefits while changing therapies, among others.
[00173] MS disease progression may be most intensive, and most damaging, at the earliest stages of disease progression. Accordingly, counter to many reimbursement policies and physician practice in light of, for example, costs and side effect mitigation, it may be most beneficial for a patient’s long term disease status to begin treatment with the most intensive DMTs, for instance, so-called second-line therapies. In embodiments, a patient is treated with a regimen of the present TNFRSF25-specific agents in combination with a second-line therapy. Such a combination is used to reduce the side effect profile of one or more second-line therapies. In embodiments, the combination is used to reduce dose of frequency of administration of one or more second-line therapies. For example, the doses of Table 1 may be reduced by about 75%, about 50%, or about 40%, or about 30%, or about 25% in the context of the combination and the/or the frequency of dosing may be decreased to be half as often, or a third as often or may be reduced from, for example, daily to every other day or weekly, every other day to weekly or bi-weekly, weekly to biweekly or monthly, etc. Accordingly, In embodiments, the present TNFRSF25-specific agents increase patient adherence by allowing for more convenient treatment regimens. Further, some DMTs have a suggested lifetime dose limitation e.g. for mitoxantrone, the lifetime cumulative dose should be strictly limited to 140 mg/m2, or 2 to 3 years of therapy (Fow Clin Thera 28(4): 461). In embodiments, supplementation with the present TNFRSF25-specific agents preserves patient’s access to mitoxantrone by allowing for lower or less frequent dosing with this DMT.
[00174] In embodiments, the patient is a naive patient, who has not received treatment with one or more DMTs, and the present TNFRSF25-specific agents are used to buffer the side effects of a second-line therapy. Accordingly, the naive patient is able to benefit from the long-term benefits of a second-line therapy at disease outset. In embodiments, the present TNFRSF25-specific therapy is used as an entry therapy that precedes the use of a second-line therapy. For example, the present TNFRSF25-specific therapy may be administered for an initial treatment period of about 3 months to stabilize disease and then the patient may be transitioned to a maintenance therapy of a second line agent.
[00175] It is generally believed that naive patients are more likely to respond to therapy as compared to patients that have received, and perhaps failed one or more DMT. In embodiments, the present TNFRSF25-specific agents find use in patients that have received, and perhaps failed one or more DMT. For example, In embodiments, the present TNFRSF25-specific agents increase the therapeutic effect in patients that have received, and perhaps failed one or more DMT and may allow these patients to respond like naive patients.
[00176] In embodiments, the patient has received or is receiving treatment with one or more DMTs and is not responding well. For example, the patient may be refractory or poorly responsive to one or more DMTs. In embodiments, the patient is refractory, or poorly responsive to one or more of agents of Table 1. In embodiments, the patient is refractory, or poorly responsive to one or more of teriflunomide (AUBAGIO (GENZYME)); interferon beta-1 a (AVONEX (BIOGEN IDEC); interferon beta-1 b (BETASERON (BAYER HEALTHCARE PHARMACEUTICALS, INC.); glatiramer acetate (COPAXONE (TEVA NEUROSCIENCE); interferon beta-1 b (EXTAVIA (NOVARTIS PHARMACEUTICALS CORP.); fingolimod (GILENYA (NOVARTIS PHARMACEUTICALS CORP.); alemtuzumab (LEMTRADA (GENZYME); mitoxantrone (NOVANTRONE (EMD SERONO); pegylated interferon beta-1 a (PLEGRIDY (BIOGEN I DEC); interferon beta-1 a (REBIF (EMD SERONO, INC.); dimethyl fumarate (BG-12) (TECFIDERA (BIOGEN IDEC); and natalizumab (TYSABRI (BIOGEN IDEC). In embodiments, the present TNFRSF25-specific agents result in a therapeutic benefit of one or more DMTs in the patient and therefore reduce or eliminate the nonresponsiveness to the DMT. For instance, this may spare the patient therapy with one or more DMTs at a higher dosing or frequency.
[00177] Similarly, In embodiments, the present TNFRSF25-specific agents may increase a patient’s response such as to avoid the need to change the patient’s therapeutic regiment from a first line therapy to a second line therapy. First line therapies (or “platform therapies,” e.g. glatiramer acetate and interferons) are often the entry choice of therapeutic for MS patients and are commonly thoughts of as having lower side effects than second line therapies and are often the first reimbursed therapy for an MS patient. However, first line therapies are considered to be only mildly effective. In embodiments, the present TNFRSF25-specific agents increase the therapeutic efficacy of the first line therapy and reduce the need to alter a patient’s treatment to a second line therapy {e.g. oral agents (e.g. teriflunomide, fingolimod, dimethyl fumarate) and monoclonal antibodies (e.g. natalizumab, alemtuzumab)) which are believed to carry greater side effects. Accordingly, In embodiments, the present TNFRSF25-specific agents reduce the need for escalation of therapy.
[00178] However, as described herein, In embodiments, the present TNFRSF25-specific agents reduce or eliminate one or more side effects and therefore allow patients access to second line therapies. For instance, oral agents are often considered more palatable for MS patients as a second line therapy as compared to the monoclonal antibodies, which often require a medical site visit for infusion administration (and indeed, in some contexts, the oral agents are considered to be first line therapies). Accordingly, In embodiments, the present TNFRSF25-specific agents prevent the need to escalate therapy from an oral agent to a monoclonal antibody. In embodiments, the present TNFRSF25-specific agents are used in combination to allow a patient access to a more efficacious second line therapy, e.g. by allowing for dose and/or frequency reduction and/or by mitigating side effects. For instance, natalizumab reduces the relapse rate more than first-line agents; however, due to issues of adverse effects is a second-line agent reserved for those who do not respond to other treatments or with severe disease. In combination with the present TNFRSF25-specific agents, In embodiments, patients may be given clinical access to natalizumab as their baseline therapy and therefore enjoy natalizumab’s better efficacy relative to first line agents. Further, mitoxantrone, whose use is limited by severe adverse effects, is a third-line option for those who do not respond to other medications. In combination with the present TNFRSF25-specific agents, In embodiments, patients may be given clinical access to mitoxantrone.
[00179] Further, in embodiments, the present TNFRSF25-specific agents find use to supplement a DMT with which a patient is comfortable and therefore avoid a need to switch therapies to improve therapeutic effect. For example, within the oral DMTs, fingolimod therapy has been reported to result in a higher probability of no evidence of disease activity than dimethyl fumarate and teriflunomide therapy (Nixon, et al. Adv Thera. 2014; 31 (11): 1134-54). In embodiments, the present TNFRSF25-specific agents supplement dimethyl fumarate and/or teriflunomide therapy to bridge the gap to the clinical effect observed with fingolimod therapy. Accordingly, In embodiments, the present TNFRSF25-specific agents are used to provide the optimal treatment regimen for the patient, in light of therapeutic costs and benefits.
[00180] In patients with more aggressive disease, one approach is an induction treatment model, whereby a therapy with strong efficacy but strong safety concerns would be given first, followed by a maintenance therapy. An example of such a model might include initial treatment with alemtuzumab, followed by interferons {e.g. IFN-b), glatiramer acetate, or BG-12. In embodiments, the present TNFRSF25-specific agents are used to prevent the need to switch therapies for maintenance. In embodiments, the present TNFRSF25-specific agents are used to as maintenance therapy to one or more DMTs, including second line therapies.
[00181] In embodiments, the present TNFRSF25-specific agents are used to as first therapy in an induction, followed by another DMT as a maintenance therapy - such as, for example, a first line therapy.
[00182] In embodiments, the present TNFRSF25-specific therapy may be administered for an initial treatment period of about 3 months to stabilize disease and then the patient may be transitioned to a maintenance therapy of a first line agent.
[00183] In embodiments, the present TNFRSF25-specific therapy is used to bridge the gap between the initiation of treatment with one or more DMTs and the onset of clinical effect. For example, in some patients glatiramer acetate slowly begins to have an effect in a patient and the present TNFRSF25-specific therapy provides the necessary therapy during this period. Further, some have suggested a transition period between different DMTs of about 8 to about 12 weeks (by way of non-limiting example, form Tysabri (natalizumab) to GILENYA (fingolimod) to maintain disease control and avoid potentially harmful additive effects on immune surveillance. In embodiments, the present TNFRSF25-specific therapy is used to bridge this gap.
[00184] In embodiments, the present TNFRSF25-specific agents are used to prevent or reduce one or more side effects of a DMT, including without limitation any agent of Table 1. For example, the present TNFRSF25-specific agents may be used in a regimen that allows dose sparing for one or more DMTs and therefore results in fewer side effects. For example, In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of AUBAGIO or related agents, which may include hair thinning, diarrhea, flu, nausea, abnormal liver tests and unusual numbness or tingling in the hands or feet (paresthesias), levels of white blood cells, which can increase the risk of infections; increase in blood pressure; and severe liver damage. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of AVONEX or related agents which include flu-like symptoms following injection, depression, mild anemia, liver abnormalities, allergic reactions, and heart problems. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of BETASERON or related agents which include flu-like symptoms following injection, injection site reactions, allergic reactions, depression, liver abnormalities, and low white blood cell counts. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of COPAXONE or related agents which include injection site reactions, vasodilation (dilation of blood vessels); chest pain; a reaction immediately after injection, which includes anxiety, chest pain, palpitations, shortness of breath, and flushing. In embodiments, the present TNFRSF25- specific agents may reduce one or more side effects of EXTAVIA or related agents which include flu-like symptoms following injection, injection site reactions, allergic reactions, depression, liver abnormalities, and low white blood cell counts. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of GILENYA or related agents which include headache, flu, diarrhea, back pain, liver enzyme elevations, cough, slowed heart rate following first dose, infections, and swelling in the eye. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of LEMTRADA or related agents which include rash, headache, fever, nasal congestion, nausea, urinary tract infection, fatigue, insomnia, upper respiratory tract infection, hives, itching, thyroid gland disorders, fungal Infection, pain in joints, extremities and back, diarrhea, vomiting, flushing, and infusion reactions (including nausea, hives, itching, insomnia, chills, flushing, fatigue, shortness of breath, changes in the sense of taste, indigestion, dizziness, pain). In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of NOVANTRONE or related agents which include blue-green urine 24 hours after administration; infections, bone marrow suppression (fatigue, bruising, low blood cell counts), nausea, hair thinning, bladder infections, mouth sores, and serious liver and heart damage. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of PLEGRIDY or related agents which include flu-like symptoms following injection, injection site reactions, depression, mild anemia, liver abnormalities, allergic reactions, and heart problems. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of REBIF or related agents which include flu-like symptoms following injection, injection site reactions, liver abnormalities, depression, allergic reactions, and low red or white blood cell counts. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of TECFIDERA or related agents which include flushing (sensation of heat or itching and a blush on the skin), gastrointestinal issues (nausea, diarrhea, abdominal pain), rash, protein in the urine, elevated liver enzymes; and reduction in blood lymphocyte (white blood cell) counts. In embodiments, the present TNFRSF25-specific agents may reduce one or more side effects of TYSABRI or related agents which include headache, fatigue, urinary tract infections, depression, respiratory tract infections, joint pain, upset stomach, abdominal discomfort, diarrhea, vaginitis, pain in the arms or legs, rash, allergic or hypersensitivity reactions within two hours of infusion (dizziness, fever, rash, itching, nausea, flushing, low blood pressure, difficulty breathing, chest pain).
[00185] In embodiments, the present TNFRSF25-specific agents are used to prevent or reduce progressive multifocal leukoencephalopathy (PML). In embodiments, the present TNFRSF25-specific agents are used to prevent or reduce progressive multifocal leukoencephalopathy (PML) caused by treatment with TECFIDERA or related agents.
[00186] Any aspect or embodiment described herein can be combined with any other aspect or embodiment as disclosed herein.
[00187] In order that the disclosure disclosed herein may be more efficiently understood, examples are provided below. It should be understood that these examples are for illustrative purposes only and are not to be construed as limiting the invention in any manner.
EXAMPLES Example 1: Expansion of Tregs using an TNFRSF25 Agonistic Antibody
[00188] For the experiments of this example, a method for treating or preventing an autoimmune inflammatory disease is described, and the method comprises administering an effective amount of TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibody or antigen binding fragment thereof (e.g., PTX-35, SEQ ID NOs: 8 and 9, or a murine version thereof “mPTX-35”) to a patient in need thereof.
[00189] In this example, a classical in vivo experimental model based on chronic experimental autoimmune encephalomyelitis (“EAE”) in mice was utilized to test whether PTX-35 is effective to treat or prevent an autoimmune disease. The following experiments show how mPTX-35 expanded both Tregs and Tconv during model disease. An experimental overview of PTX-35 treatment during an autoimmunity disease, as well as a timeline, is shown in FIG. 1, and features of the study design are shown in FIG.2. Data is shown in FIG. 3 of the body weight of the mice before disease induction. In these experiments, mPTX-35 was administered on both day -7 and day 7, and mPTX-35 expanded Tregs prior to model start. Data is shown in FIG. 4 of the body weight of the mice during disease induction. In these experiments, mPTX-35 was administered on both day 0 and day 14, and mPTX-35 expanded conventional T cells (“Tconv”) at model start. Data is shown in FIG. 5 of the body weight of the mice after disease induction. In these experiments, mPTX-35 was administered on both day 7 and day 21, and mPTX-35 expanded both Tregs and Tconv during model disease. Collectively, the data from the experiments in FIG. 3, FIG. 4, and FIG. 5 demonstrate how intervention and treatment timing matters due to the Treg/Tconv Expansion Potential of PTX-35. Specifically, mPTX-35 treatment before disease induction is best.
[00190] The following experiments show how intervention and treatment timing matters due to the Treg/Tconv expansion potential of PTX-35.
[00191] FIG. 6 is a chart showing the clinical assessment of EAE with respect to scores and clinical signs. FIG.7 is a graph showing the clinical score of the mice when given mPTX-35 before disease induction, and FIG. 10 is a graph showing the disease incidence in the mice before disease induction. FIG. 14 is a graph showing percent survival (x-axis) and days elapsed (y-axis) before disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G4 (mPTX-35 3 mg/kg) mice. In these experiments, mPTX- 35 was administered on both day -7 and day 7, and mPTX-35 expand Tregs prior to the start of the experimental model.
[00192] FIG. 8 is a graph showing the clinical score of the mice when given mPTX-35 during disease induction, and FIG. 11 is a graph showing the disease incidence in the mice during disease induction. FIG. 15 is a graph showing percent survival (x-axis) and days elapsed (y-axis) during disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G5 (mPTX-35 3 mg/kg) mice. In these experiments, mPTX- 35 was administered on both day 0 and day 14, and mPTX-35 expanded conventional T cells (“Tconv”) at the start of the experimental model.
[00193] FIG. 9 is a graph showing the clinical score of the mice when given mPTX-35 after disease induction, and FIG. 12 is a graph showing the disease incidence in the mice after disease induction. FIG. 16 is a graph showing percent survival (x-axis) and days elapsed (y-axis) after disease induction for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. In these experiments, mPTX- 35 was administered on both day 7 and day 21, and mPTX-35 expanded both Tregs and Tconv during the model disease.
[00194] FIG. 13 is a graph showing the clinical score area under the curve (“AUC”) measurements for the G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. This graph underscores how intervention and treatment timing matters due to the Treg/Tconv expansion potential of PTX-35. Specifically, mPTX-35 treatment before disease induction is best.
[00195] The following experiments show how expansion of Tregs before disease induction or after initiation can control outcome. FIG. 17 is a chart showing the evaluation of disease progression as it relates to the spinal cord. FIG. 18 is a graph showing the histology inflammation score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. In these experiments, PTX-35 mediated Treg expansion reduced central nervous system pathologies, and these experiments show how expansion of Tregs before disease induction or after initiation can control outcome.
[00196] FIG. 19 is a graph showing the histology lesion score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. PTX-35 mediated Treg expansion reduced central nervous system pathologies. These experiments show expansion of Tregs before disease induction or after initiation can control outcome.
[00197] FIG. 20 is a graph showing the histology demyelination score for G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. These experiments show PTX-35 mediated Treg expansion reduced central nervous system pathologies. Expansion of Tregs before disease induction or after initiation can control outcome. A reduced spinal cord inflammation and axon demyelination was observed with PTX-35 treatment. FIG. 21 shows representative images for hematoxylin and eosin (“HE”) and fast blue staining for G1 normal mice (left panel), and G2 vehicle mice (right panel). These experiments show PTX-35 mediated Treg expansion reduced central nervous system pathologies.
[00198] Representative HE and fast blue staining are shown in FIG. 22 and FIG. 23 for G3 Dex treated mice, G4 mPTX-35 treated mice, G5 mPTX-35 treated mice, and G6 mPTX-35 treated mice. FIG. 24 shows the gating strategy for the Flow cytometry data. FIG. 25 is a graph showing the percentage of regulatory T cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice. FIG. 26 is a graph showing the percentage of Th17 cells in the spleen (left panel), and the blood (right panel), for the G1 (normal), G2 (vehicle), G3 (Dex 1 mg/kg), G4 (mPTX-35 3 mg/kg), G5 (mPTX-35 3 mg/kg), and G6 (mPTX-35 3 mg/kg) mice.
[00199] Collectively, inter alia, the experiments of this example demonstrate that expansion of T regs cells prior to disease, or early in disease, gives a beneficial result in autoimmune inflammatory disease.
Example 2: MOG 35-55 Immunized Mice Treated with mPTX-35 at Various Days Show Reduced or Minimal Disease Severity
[00200] PTX-35 treatment on Days 12&16 showed the greatest protective capacity from EAE. Analysis of splenocytes from PTX-35 treated mice revealed an increase FoxP3+Treg. PTX-35 treated T cells displayed increased expression of ICOS and CD103. CD8+T cells showed signs of exhaustion with increased Tim3 and LAG3 after PTX-35 treatment.
[00201] 12-week-old female C57BL/6 mice were left unimmunized or were immunized with 50 pL of
200 pg MOG 35-55 in CFA between the shoulder blades and at the base of the tail. At 2hrs and 24hrs following immunization, intraperitoneal injection with 100 pL of pertussis toxin at 200 ng/pL was administered. MOG 35-55 immunized C57BL/6 were treated with PBS, Isotype control antibody (10 mg/kg; clone MOPC- 21, InVivoMab), PTX-35 (10 mg/kg) at days 4 and 8 post immunization, or PTX-35 (10 mg/kg) at days 12 and 16 post immunization (FIG. 27A). Splenocytes were isolated and stained for flow cytometry. Spinal cords were kept for histology. N=5 mice per group (FIG. 27B). Mice were monitored for paralysis by daily observation beginning at 5 days post MOG 35-55 immunization. Level of paralysis was characterized on a scale of 1 to 5 (1=limp tail, 2=one hind limb paralysis, 3= two limb paralysis, 4= three of more limb paralysis/weakness, 5=moribund). Additionally, immunized mice were monitored every 2-3 days for loss in body weight.
[00202] The results demonstrate that beginning PTX-35 treatment at an early stage of EAE disease progression slightly reduced disease severity but the overall weight loss was comparable to the isotype treated mice. Beginning PTX-35 treatment at the late stage of EAE disease progression demonstrate minimal disease severity, as noted for example by the paralytic score and low degree of weight loss. Weight change in grams of MOG 35-55 immunized mice treated with mPTX-35 at various days post immunization showed less decrease in weight compared to mice treated with isotype control antibodies (FIG. 28A). The percent of initial body weight of MOG 35-55 immunized mice treated with mPTX-35 at day 12 and 16 post immunization was similar to percent of initial body weight of non-immunized mice (FIG. 28B). Mice were euthanized at day 30 post immunization for immune profiling of splenocytes by flow cytometry and demyelination of the spinal cord by histology. Images of spinal cords from B6 mice immunized with MOG 35-55 taken at Day 30 post immunization (FIG.29). Luxol Fast Blue staining specifically stained the myelin sheath of nerve cells highlighting the white matter of the spinal cord. Hematoxylin and eosin (H&E) staining highlighted lymphocytic infiltration of the spinal cord (indicated by arrows) (FIG. 29). Levels of inflammation were scored based on a scale of 0 to 4 (0= no infiltration, 1= infiltration of the perivascular regions, 2=infiltration of less than 1/3 of the white matter, 3=infiltration of more than 1/3 of the white matter, 4= infiltration of the entire white matter) (FIG. 30). B6 mice immunized with MOG 35-55 treated with PTX-35 at day 12 and day 16 had significantly less inflammation compared to isotype mice (FIG. 30).
[00203] PTX-35 treatment on days 12 and 16 showed the greatest protective capacity from EAE. Analysis of splenocytes from PTX-35 treated mice revealed an increase of FoxP3+ Treg. PTX-35 treated T cells displayed increased expression of ICOS and CD103. CD8+ T cells showed signs of exhaustion with increased Tim3 and LAG3 after PTX-35 treatment. FIG. 31 shows percentage distribution of Foxp3+ regulatory T cells and conventional CD4+ T cells in non-immunized mice, MOG 35-55 immunized mice treated with PBS, MOG 35-55 immunized mice treated with isotype control antibodies, MOG 35-55 immunized mice treated with mPTX-35 at day 4 and day 8, and MOG 35-55 immunized mice treated with mPTX-35 at day 12 and day 16. B6 mice immunized with MOG 35-55 were sacrificed 30 days post immunization. Splenic CD4+ T cells were analyzed and results showed immunized mice treated with PTX- 35 at day 4 and 8 and immunized mice at day 12 and 16 had significantly higher percentages of activated FoxP3+T cells compared to isotype mice (FIG. 32). [00204] The splenic CD4+ FoxP3+ regulatory T cells were also examined for expression levels of immunosuppressive markers including ICOS+ T (FIG. 33 - FIG. 34), CD103+ T cells (FIG. 35 - FIG. 36). These results showed that immunized mice treated with PTX-35 at days 4 and 8 and immunized mice treated with PTX-35 at days 12 and 16 had significantly higher percentages of regulatory T cells expressing these markers. Mice treated with PTX-35 at day 12 and 16 had a significantly higher percentage T cells expressing ICOS compared to mice treated with isotype control antibodies (FIG. 34). Mice treated with PTX-35 at day 12 and 16 had a significantly higher percentage T cells expressing CD103 compared to mice treated with isotype control antibodies (FIG. 36).
[00205] The splenic CD8+ T cells were also examined for Tim3 and LAG3 (FIG. 37 - FIG. 38). Results showed immunized mice at day 12 and 16 had significantly higher percentages of Tim3+LAG3+ CD8+T cells compared to mice treated with isotype control antibodies. (FIG. 38). In all figures, significance is relative to isotype control antibodies with asterisk in graphs meaning *<0.05, **<0.005, ***<0.0005.
[00206] The splenic CD4+ FoxP3+ regulatory T cells were also examined for expression levels of other immunosuppressive markers including CD44 (FIG. 39), TIGIT (FIG. 40), CTLA4 (FIG. 41), and Ki67 (FIG. 42). These results showed that immunized mice treated with PTX-35 at days 12 and 16 had slightly higher expression levels of CD44, TIGIT, CTLA4, and Ki67 compared to mice treated with isotype control antibodies.
[00207] The splenic CD4+ FoxP3+ regulatory T cells were also examined for median fluorescence intensity (MFI) of GITR (FIG. 43) and CD25 (FIG. 44). The results showed that immunized mice treated with PTX-35 at days 12 and 16 had slightly higher amounts of GITR compared to mice treated with isotype control antibodies. CD25 levels did not change significantly between non-immunized and immunized mice samples.
[00208] The splenic CD8+ T cells were also examined for percentage of PD-1 + T cells, which did not change significantly between non-immunized and immunized mice samples (FIG.45). Percentage of PD- 1+T cells increased after treatment of PTX-35 between non-immunized and immunized mice samples (FIG. 46).
[00209] Results showed FoxP3+ Treg are expanded after PTX-35 administrations during autoimmunity and reduces EAE severity. ICOS and CD103 were significantly upregulated markers on FoxP3+ Treg. Other markers showed a trend towards increasing but were variable in expression. CD8+ T cells significantly upregulated LAG3 and TIM3 after PTX-35 therapy. An increased frequency of FoxP3+ Treg suggests enhanced suppression of pathogenic T cells after PTX-35 therapy. ICOS expression by FoxP3+ Treg is known to be important for regulatory T cell function in autoimmune settings. Furthermore, increased expression of CD103 on FoxP3+Treg suggests a greater capacity for tissue residency from PTX-35 treated FoxP3+Treg. PTX-35 treatment also enhanced dysfunctional CD8+T cell markers during EAE. Interestingly, CD8+ T cells showed increased expression of the inhibitory molecules Tim3 and Lag3 implying PTX-35 treated CD8+T cells may be functionally exhausted which could attenuate autoimmunity. PTX-35 treatments also showed prevention or reversal of primary progressive EAE.
Example 3: PLP 139-151 Immunized SJUL Mice Showed Relapsing Remitting EAE Prevention or Reversal via PTX-35
[00210] SJL/J mice were immunized with PLP 139-151 and treated with PTX-35 at day -4 and 0, day 4 and 8, day 12 and 16, day 16 and 20, or day 4, 8, 16, and 20 post immunization (FIG. 47). Results show disease progression in mice treated with PTX-35 showed signs of improved compared to untreated mice (FIG. 48-52). Mice treated with PTX-354 days prior to immunization and at day 0 had lower paralytic scores compared to immunized mice treated with isotype control antibodies at day 10-15 and day 27-41 (FIG. 48). Mice treated with PTX-35 at day 4 and day 8 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 10 until day 43 (FIG. 49). Mice treated with PTX-35 at day 12 and day 16 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 13-15 and day 24-43 (FIG. 50). Mice treated with PTX-35 at day 16 and day 20 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 13 until day 43 (FIG. 51). Mice treated with PTX-35 at day 4 and 8, and day 12 and 16 had lower paralytic scores compared to immunized mice treated with isotype control antibodies starting at day 13-15 and day 20-43 (FIG. 52).
[00211] Results showed that PTX-35 treatment at various days prevented or reversed relapsing remitting EAE disease progression compared to mice treated with isotype control antibodies.
EQUIVALENTS
[00212] While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essential features hereinbefore set forth and as follows in the scope of the appended claims.
[00213] Those skilled in the art will recognize, or be able to ascertain, using no more than routine experimentation, numerous equivalents to the specific embodiments described specifically herein. Such equivalents are intended to be encompassed in the scope of the following claims.
INCORPORATION BY REFERENCE
[00214] All patents and publications referenced herein are hereby incorporated by reference in their entireties.
[00215] As used herein, all headings are simply for organization and are not intended to limit the disclosure in any manner. The content of any individual section may be equally applicable to all sections.

Claims

CLAIMS What is claimed is:
1. A method for treating or preventing an autoimmune inflammatory disease, comprising administering an effective amount of TNF Receptor Superfamily Member 25 (TNFRSF25) agonistic antibody or antigen binding fragment thereof to a patient in need thereof.
2. The method of claim 1 , wherein the autoimmune inflammatory disease is a demyelinating disease.
3. The method of claim 1 or claim 2, wherein the inflammation is in the central nervous system (CNS).
4. The method of claim 2 or claim 3, wherein the demyelinating disease is multiple sclerosis (MS).
5. The method of claim 2, wherein the demyelinating disease is clinically isolated syndrome (CIS).
6. The method of claim 5, wherein the method prevents or slows the progression of CIS to MS.
7. The method of claim 2, wherein the demyelinating disease is radiologically isolated syndrome (RIS).
8. The method of claim 7, wherein the method prevents or slows the progression of RIS to MS.
9. The method of any one of claims 1 to 8, wherein the patient is likely to develop MS.
10. The method of any one of claims 1 to 9, wherein the patient has CIS.
11. The method of claim 10, wherein the patient has a monofocal episode of CIS.
12. The method of any one of claims 1 to 11 , wherein the patient has optic neuritis.
13. The method of claim 12, wherein the patient has a multifocal episode of CIS.
14. The method of claim 13, wherein the patient has optic neuritis and numbness or tingling in the legs.
15. The method of any one of claims 1 to 14, wherein the patient has CIS and one or more magnetic resonance imaging (MRI)-detected brain lesions.
16. The method of any one of claims 1 to 15, wherein the treatment occurs early in disease progression.
17. The method of any one of claims 1 to 16, wherein the treatment occurs before the onset of relapsing- remitting MS (RRMS).
18. The method of any one of claims 1 to 17, wherein the treatment occurs before the onset of secondary progressive MS (SPMS).
19. The method of any one of claims 1 to 18, wherein the treatment occurs before the onset of primary progressive MS (PPMS).
20. The method of any one of claims 1 to 19, wherein the treatment expands and/or selectively activates a population of regulatory T cells (Tregs) in the patient.
21. The method of any one of claims 1 to 19, wherein the treatment does not substantially expand and/or selectively activate a population of cytotoxic T cells (Tc cells) and/or helper T cells (Th cells) in the patient.
22. The method of any one of claims 1 to 19, wherein the treatment does not substantially expand and/or selectively activate a population of Th17 cells in the patient.
23. The method of any one of claims 1 to 20, wherein the treatment increases the ratio of Tregs to Tc and/or Th cells in the patient compared to an untreated patient.
24. The method of any one of claims 1 to 20, wherein the treatment increases the ratio of Tregs to Th17 cells in the patient compared to an untreated patient.
25. The method of any one of claims 1 to 20, wherein the treatment increases the ratio of Tregs to conventional T cells (Tconv) in the patient compared to an untreated patient.
26. The method of any one of claims 1 to 20, wherein the treatment increases the population of Tregs expressing ICOS compared to an untreated patient.
27. The method of any one of claims 1 to 20, wherein the treatment increases the population of Tregs expressing CD103. compared to an untreated patient.
28. The method of any one of claims 1 to 21, wherein the treatment increases the population of Tc cells expressing Tim3 and/or LAG3 compared to an untreated patient.
29. The method of any one of claims 1 to 28, wherein the treatment reduces spinal cord inflammation, as compared to before treatment.
30. The method of any one of claims 1 to 28, wherein the treatment reduces axon demyelination, as compared to before treatment.
31. The method of any one of the above claims, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises:
(i) a heavy chain variable region comprising heavy chain CDR1 , CDR2, and CDR3 sequences, wherein the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO: 1), the heavy chain CDR2 sequence is YISSASGUSYADAVRG (SEQ ID NO: 2); and the heavy chain CDR3 sequence is DPAYTGLYALDF (SEQ ID NO: 3) or DPPYSGLYALDF (SEQ ID NO: 4); and
(ii) a light chain variable region comprising light chain CDR1, CDR2, and CDR3 sequences, wherein the light chain CDR1 sequence is TLSSELSWYTIV (SEQ ID NO: 5), the light chain CDR2 sequence is LKSDGSFISKGD (SEQ ID NO: 6), and the light chain CDR3 sequence is CGAGYTLAGQY GWV (SEQ ID NO: 7).
32. The method of claim 31, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises variable region framework (FW) sequences juxtaposed between the CDRs according to the formula (FW1)-(CDR1)-(FW2)-(CDR2)-(FW3)-(CDR3)-(FW4), wherein the variable region FW sequences in the heavy chain variable region are heavy chain variable region FW sequences, and wherein the variable region FW sequences in the light chain variable region are light chain variable region FW sequences.
33. The method of claim 32, wherein the variable region FW sequences are human.
34. The method of any one of claims 31 to 33, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof further comprises human heavy chain and light chain constant regions.
35. The method of any one of claims 31 to 34, wherein the constant regions are selected from the group consisting of human lgG1, lgG2, lgG3, and lgG4.
36. The method of claim 35, wherein the constant regions are lgG1.
37. The method of claim 35, wherein the constant regions are lgG4.
38. The method of any one of claims 31 to 37, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAV RGRFTISRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85 to about 99% identity thereto.
39. The method of any one of claims 31 to 38, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDR FSGSSSGAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85 to about 99% identity thereto.
40. The method of any one of claims 31 to 39, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises 4C12, or a humanized version, or fragment thereof.
41. The method of any one of claims 1 to 40, wherein the treatment occurs in combination with one or more disease modifying therapies.
42. The method of any one of claims 1 to 41, wherein the treatment occurs in combination with one or more agents of Table 1.
43. A method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising:
(i) determining an amount of ICOS protein, nucleic acid, or activity in a sample from the patient, and
(ii) administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of ICOS relative to an untreated and/or undiseased patient.
44. A method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising:
(i) determining the amount of CD103 protein, nucleic acid, or activity in a sample from the patient, and
(ii) administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of CD103 relative to an untreated and/or undiseased patient.
45. A method of selecting a patient with an autoimmune inflammatory disease for treatment with TNFRSF25 agonistic antibody or antigen binding fragment, comprising:
(i) determining the amount of Tim3 and/or LAG3 protein, nucleic acid, or activity in a sample from the patient, and
(ii) administering an effective amount of the TNFRSF25 agonistic antibody or antigen binding fragment to the patient, if the patient presents with a reduced amount of Tim3 and/or LAG3 relative to an untreated and/or undiseased patient.
46. The method of any one of claims 1 to 45, wherein the autoimmune inflammatory disease is a demyelinating disease.
47. The method of any one of claims 1 to 46, wherein the inflammation is in the central nervous system (CNS).
48. The method of any one of claims 1 to 47, wherein the demyelinating disease is selected from multiple sclerosis (MS), clinically isolated syndrome (CIS), and radiologically isolated syndrome (RIS).
49. The method of any one of claims 1 to 48, wherein the autoimmune inflammatory disease is selected from neuromyelitis optica, Devic’s disease, idiopathic inflammatory demyelinating diseases, central nervous system neuropathies, central pontine myelinolysis, myelopathies, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti-MAG peripheral neuropathy, Charcot-Marie-Tooth disease, hereditary neuropathy with liability to pressure palsy, peripheral neuropathy, optic neuropathy, and progressive inflammatory neuropathy.
50. The method of any one of the above claims, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises:
(i) a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 sequences, wherein the heavy chain CDR1 sequence is GFTFSNFIDLN (SEQ ID NO: 1), the heavy chain CDR2 sequence is Yl SSASGLI SYADAVRG (SEQ ID NO: 2); and the heavy chain CDR3 sequence is DPAYTGLYALDF (SEQ ID NO: 3) or DPPYSGLYALDF (SEQ ID NO: 4); and
(ii) a light chain variable region comprising light chain CDR1 , CDR2, and CDR3 sequences, wherein the light chain CDR1 sequence is TLSSELSWYTIV (SEQ ID NO: 5), the light chain CDR2 sequence is LKSDGSFISKGD (SEQ ID NO: 6), and the light chain CDR3 sequence is CGAGYTLAGQY G WV (SEQ ID NO: 7).
51. The method of claim 50, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a heavy chain variable region having the amino acid sequence EVQLVESGGGLSQPGNSLQLSCEASGFTFSNHDLNWVRQAPGKGLEWVAYISSASGLISYADAV RGRFTISRDNAKNSLFLQMNNLKSEDTAMYYCARDPPYSGLYALDFWGQGTQVTVSS (SEQ ID NO: 8), or an amino acid sequence of at least about 85 to about 99% identity thereto.
52. The method of claims 50 or 51, wherein the TNFRSF25 agonistic antibody or antigen binding fragment thereof comprises a light chain variable region having the amino acid sequence QPVLTQSPSASASLSGSVKLTCTLSSELSSYTIVWYQQRPDKAPKYVMYLKSDGSHSKGDGIPDR FSGSSSGAHRYLSISNVQSEDDATYFCGAGYTLAGQYGWVFGSGTKVTVL (SEQ ID NO: 9), or an amino acid sequence of at least about 85 to about 99% identity thereto.
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