Therapeutics for ocular immunoinflammatory diseases

Abstract

Methods and compositions for reducing the activity or number of memory Th17 cells in ocular tissue, and for treatment and/or prevention of ocular immunoinflammatory diseases are described.

Claims

1. A method for treating a subject afflicted with Dry Eye Disease (DED), comprising locally administering to an eye of the subject a composition consisting of a neutralizing anti-IL-15R antibody or an antibody that binds to IL-15, or a neutralizing anti-IL-7R antibody or an antibody that binds to IL-7, and an ophthalmically acceptable vehicle.

2. The method of claim 1, wherein treating the subject afflicted with the ocular immunoinflammatory disorder comprises inhibiting the survival or proliferation of a Th17 cell in an eye tissue.

3. The method of claim 1, wherein said subject suffers from a sandy or gritty feeling as if something is in the eye, eye dryness, heavy eyelids, stinging, itching, burning, irritation, pain, redness, inflammation, discharge, inability to cry when emotionally stressed, eye fatigue, blurred vision, or excessive watering, and wherein said method inhibits or reduces the severity of the sandy or gritty feeling as if something is in the eye, eye dryness, heavy eyelids, stinging, itching, burning, irritation, pain, redness, inflammation, discharge, inability to cry when emotionally stressed, eye fatigue, blurred vision, or excessive watering.

4. The method of claim 1, wherein: (a) the composition is in the form of a solid, an ointment, a gel, a liquid, an aerosol, a mist, a polymer, a contact lens, a film, an emulsion, or a suspension; (b) said composition is administered topically; (c) said method does not comprise systemic administration to non-ocular tissue of the composition; or (d) the composition is administered by the subject.

5. The method of claim 1, wherein the number of memory Th17 cells in the eye of the subject is reduced after the composition is administered.

6. The method of claim 1, wherein a symptom of the ocular immunoinflammatory disorder is reduced (a) within 5, 15, 30, or 60 minutes after administering the composition begins; or (b) within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 days after administering the composition begins.

7. The method of claim 1, wherein the composition is administered to the eye of the subject (a) less than 1, 2, 3, 4, 5, or 6 times per day; (b) about 1, 2, 3, 4, 5, 6, or 7 times per week; or (c) once daily.

8. The method of claim 1, wherein: (a) the compound that inhibits IL-7 signal transduction comprises an antibody that binds to IL-7; or (b) the compound that inhibits IL-15 signal transduction comprises an antibody that binds to IL-15.

9. The method of claim 1, wherein the composition consists of a neutralizing anti-IL-7R antibody or an antibody that binds to IL-7, and an ophthalmically acceptable excipient.

Description

DESCRIPTION OF THE DRAWINGS

(1) FIGS. 1A-1C are a graphs showing increased expression of IL-7 and IL-15 as well as their respective receptors on memory Th17 cells in chronic DED mice. (FIG. 1A) Increased expression of IL-7 and IL-15 in both draining lymph nodes (DLN) and conjunctiva was observed in chronic DED (DED D28). mRNA expression levels have been normalized to naive mice (NL). (FIG. 1B) Constitutively high protein expression of IL-7 and IL-15 in DLN, and increased both proteins in conjunctiva were observed in chronic DED. (FIG. 1C) Up-regulation of both IL-7 and IL-15 receptors (IL-7R and IL-15R) on memory Th17 cells (mTh17) was observed in chronic DED, as compared to effector Th17 cells (eTh17). MFI: mean fluorescein intensity. *, p<0.05.

(2) FIG. 2 is a graph showing that blockade of IL-7 and IL-15 receptors inhibits memory Th17 cells in chronic DED. Total DLN explant cultured in the presence of anti-IL-7R and anti-IL-15R antibodies showed a >70% reduction in the frequency of memory Th17 cells compared to that cultured with isotype IgG.

(3) FIG. 3 is a graph showing that blockade of IL-7 and IL15 inhibits memory Th17 cells in chronic DED. Total DLN explant cultured in the presence of IL-7, IL-15, or combined IL-7 and IL-15 preserved significantly more memory Th17 cells than neutralization of each of them, respectively. *, p<0.05.

(4) FIGS. 4A-4C are cartoons showing non-limiting examples of IL-7 and IL-15 inhibition. (FIG. 4A) In the absence of an inhibitor, IL-7 and IL-15 can bind their respective receptors (IL-7R and IL-15R, respectively), which leads to signal transduction. (FIG. 4B) In various embodiments, an inhibitor that binds IL-7 or IL-15 blocks receptor binding. (FIG. 4C) In certain embodiments, an inhibitor that binds IL-7R or IL-15R blocks receptor binding.

(5) FIGS. 5A-5B are graphs showing that in vivo topical treatment with anti-IL-7 or anti-IL-15 antibody significantly decreased disease severity through depleting memory Th17 cells in chronic DED mice. (FIG. SA) DED was induced in mice and then these mice were treated with topical anti-IL-7, anti-IL-15, or isotype IgG (control group) eye drops for 14 days. Each of anti-IL-7 and anti-IL-15 treatment significantly decreased disease severity. *, p<0.05 at D3, D10, and D14 as compared with control group. (FIG. 5B) At the end of the treatment, ocular surface memory Th17 cells were almost completely depleted by either anti-IL-7 or anti-IL-15 treatment.

(6) FIG. 6 is a graph showing that topical anti-IL-7 and anti-IL-15 antibody treatments have a long-term therapeutic effect. Chronic DED mice previously treated with topical anti-IL-7, anti-IL-15, or isotype IgG were re-challenged by desiccating environmental stress for 8 days without any further Ab treatment. Both anti-IL-7 and anti-IL-15 antibody pre-treated groups showed significantly reduced disease progression and severity as compared to isotype IgG. Δ epithelial disease score=disease score after re-challenge−disease score before re-challenge *, p<0.05.

DETAILED DESCRIPTION

(7) The subject matter described herein provides novel methods for the treatment of memory Th17 cell-mediated ocular disorders including DED, autoimmune uveitis, and ocular GVHD, comprising ocular delivery (e.g. topical, subconjunctival, or intravitreal administration) of an IL-7/IL-7R and/or an IL-15/IL-15R signaling inhibitor into the eye in combination with a pharmaceutically acceptable carrier. This is a novel and fundamentally different approach to the treatment of ocular immunoinflammatory diseases (e.g., DED, autoimmune uveitis, and ocular GVHD), compared to current nonspecific approaches. Current approaches for treating ocular immunoinflammatory diseases such as DED are reliant on the use of nonspecific anti-inflammatory agents, such as corticosteroids, that are fraught with side-effects.

(8) Without wishing to be bound by any scientific theory, the persistent ocular surface inflammation in DED is mediated by memory T helper 17 (Th17) cells and immunological memory has a central role in maintaining chronic ocular disease (Mucosal Immunol 2014; 7:38-45). Very little is known about the factors that regulate long-term maintenance of memory Th17 cells. Aspects of the present subject matter relate to the surprising discovery that inhibitors of the IL-7 and IL-15 pathways are useful for treating ocular immunoinflammatory diseases such as DED.

(9) Non-limiting examples of IL-7/IL-7R or IL-15/IL-15R signaling inhibitors include agents that prevent or reduce IL-7R or IL-15R mediated signal transduction. For example, IL-7R mediated signal transduction may be prevented or reduced by inhibiting the binding of IL-7 to IL-7R. Likewise, IL-15R mediated signal transduction may be reduced or prevented by inhibiting the binding of IL-15 to IL-15R. Such agents may comprise, but are not restricted to, the following: neutralizing anti-IL-7R antibodies and fragments thereof; anti-IL-15R antibodies and fragments thereof; small molecular inhibitors of IL-7R or IL-15R; proteins such as anti-IL-7 and anti-IL-15 fusion proteins (such as a soluble version or fragment of IL-7R or IL-15R); anti-IL-7 antibodies and fragments thereof; anti-IL-15 antibodies and fragments thereof; and aptamers (e.g., DNA or RNA aptamers) that bind to IL-7, IL-7R, IL-15, or IL-15R. In various embodiments, combinations of one, two, three, four or more of these agents are administered.

(10) Non-limiting examples of inhibitors useful in various embodiments of the present subject matter include: Human IL-7 monoclonal antibody (Clone #MAB207, R&D Systems (Minneapolis, Minn., USA)); Human IL-7 polyclonal antibody (Clone #AF-207, R&D Systems); Human IL-7R alpha (CD127) antibody (Clone #MAB306, R&D Systems); Anti-human CD127 (Clone #eBioRDRS, eBiosceience (San Diego, Calif., USA)); Soluble human IL-7R alpha (CD127)-Fc chimera (Cat #306-IR, R&D Systems); Human IL-15 monoclonal antibody (Clone #MAB247, R&D Systems); Human IL-15 monoclonal antibody (Clone #MAB647, R&D Systems); Human IL-15R alpha antibody (Clone #AF247, R&D Systems); Anti-human IL-15 (Clone #ct2nu, eBiosceience); Anti-human CD215 (IL-15R alpha) (Clone #eBioJM7A4, eBiosceience); and Soluble human IL-15R alpha-Fc chimera (Cat #147-IR, R&D Systems).

(11) In various embodiments, the inhibitor is an extracellular portion of an IL-7R or IL-15R protein subunit. In some embodiments, the extracellular portion is fused to an antibody or a fragment thereof. For example, the extracellular portion may be fused to the Fc domain of an antibody (e.g., human IgG.sub.1). The fusion may be direct or via, e.g., a polypeptide linker such as IIEGRMD (SEQ ID NO: 37) or IEGRDMD (SEQ ID NO: 38).

(12) A non-limiting example of a soluble human IL-7R alpha (CD127)-Fc chimera is available from R&D Systems (Minneapolis, Minn., USA) (Cat #306-IR).

(13) Source: Mouse myeloma cell line, NS0-derived

(14) TABLE-US-00001 Human IL-7 R alpha IIEGRMD Human IgG.sub.1 (Glu21-Lys261) (Pro100-Lys330) Accession # P16871.3 N-terminus C-terminus

(15) UniProt Accession #: P16871.3 (SEQ ID NO: 10)

(16) N-terminal Sequence Analysis: Glu21

(17) Predicted Molecular Mass: (monomer) kDa

(18) SDS-PAGE: 80-90 kDa, reducing conditions

(19) A non-limiting example of a soluble human IL-15R alpha-Fc chimera is available from R&D Systems (Minneapolis, Minn., USA) (Cat #147-IR).

(20) Source: Spodoptera frugiperda, Sf21 baculovirus)-derived

(21) TABLE-US-00002 Human IL-15 R alpha IEGRDMD Human IgG.sub.1 6-His tag (Ile31-Thr172) (Pro100-Lys330) Accession # EAW86418 N-terminus C-terminus

(22) GenBank Accession #: EAW86418 (SEQ ID NO: 35)

(23) N-terminal Sequence Analysis: Ile31

(24) Predicted Molecular Mass: 42.6 (monomer) kDa

(25) SDS-PAGE: 60-70 kDa, reducing conditions

(26) Without wishing to be bound by any scientific theory, methods and compositions herein relieve one or more symptoms of a chronic ocular inflammatory disease (e.g., DED) by suppressing the memory Th17 response in chronic ocular immunoinflammatory diseases.

(27) Ocular Immunoinflammatory Diseases

(28) DED is one of the most frequently encountered ocular morbidities and may also be referred to as Dry Eye Syndrome. Twenty-five percent of patients who visit ophthalmic clinics report symptoms of dry eye, making it a growing public health problem and one of the most common conditions seen by eye care practitioners. DED is a highly prevalent condition, estimated to affect 10-20% of the adult population (Ocul Surf 2007; 5:75-92). The disease is seen with increased prevalence in patients with autoimmune diseases, which affect approximately 8% of the population. At present, non-specific anti-inflammatory therapies are the mainstay of treatment for moderate to severe DED, along with topical cyclosporine (the only FDA approved drug for the treatment of DED).

(29) DED is a predominant ocular immunoinflammatory disease. However, other diseases may be treated using methods and compositions provided herein. Exemplary contemplated ocular surface inflammatory diseases include, but are not limited to, penetrating keratoplasty (corneal transplantation), corneal neovascularization, allergy, conjunctivitis, and microbial keratitis. Contemplated disorders can be caused by autoimmune mechanisms, bone marrow transplant, surgery (general eye surgery, corneal transplantation, refractive surgery, LASIK), allergy, infection, trauma, injury, drug use, tear film abnormalities, contact lens use, neovascularization, tumor formation or growth, exposure to airborne or liquid irritants, hormonal variation, deprivation of essential fatty acids, and genetic predisposition.

(30) In various embodiments, a subject who is afflicted with or suffering from an ocular immunoinflammatory disease may be treated using a method or composition of the invention. In certain embodiments, a subject who is “afflicted with” is “suffering from” or is “in need” of treatment for a disease may be a subject who has been affirmatively diagnosed to have that disease. In some embodiments, a subject who is in need of preventative or prophylactic treatment for a disease is a subject who is at risk of developing that disease. In some embodiments, the subject has not been diagnosed with the disease, but has one or more symptoms of the disease.

(31) As used herein, a “symptom” associated with a disease includes any clinical or laboratory manifestation associated with the disease, and is not limited to what the subject can feel or observe. Non-limiting examples of DED symptoms include pain (such as stinging or burning of the eye); ulcers or scars on the cornea; decrease tolerance for dry environments; reduced vision; a sandy or gritty feeling as if something is in the eye; episodes of excess tears following very dry eye periods; a stringy discharge from the eye; redness of the eye; episodes of blurred vision; heavy eyelids; inability to cry when emotionally stressed; uncomfortable contact lenses: decreased tolerance of reading, working on the computer, or any activity that requires sustained visual attention; and eye fatigue. Non-limiting examples of autoimmune uveitis symptoms include redness of the eye; blurred vision; photophobia; sensitivity to light; irregular pupil; eye pain; floaters, which are dark spots that appear to float in the visual field; headaches; dilated ciliary vessels; presence of cells and flare in the anterior chamber; keratic precipitates (“KP”) on the posterior surface of the cornea; a hypopyon; pigment deposits on the lens; a festooned pupil on dilation of pupil; busacca nodules (inflammatory nodules located on the surface of the iris); and synechia Photopsia or seeing flashing lights. Non-limiting examples of ocular GV-HD symptoms include blurry vision; foreign body sensation; burning sensation; severe light sensitivity; chronic conjunctivitis (pink eye); dry eyes; and eye pain. In embodiments, the method described herein may include identifying a subject having one or more of these symptoms.

(32) “Treating” (or treatment of) a disease or condition includes ameliorating at least one symptom of the particular disease or condition, even if the underlying pathophysiology is not affected. The efficacy of the treatment can be evaluated, e.g., as compared to a standard, e.g., improvement in the value or quality of a parameter (e.g., self-reported pain level, tolerance for dry environments, numbers of ulcers or scars on the cornea, and vision quality) as compared to the value or quality of the parameter prior to treatment. As another example, the efficacy of treatment can be evaluated, e.g., as compared to a standard, e.g., slowing progression of the disease as compared to a usual time course for the disease in a cohort that has not been treated or compared to historical data on disease progression. Treating a disease also includes slowing its progress; and/or relieving the disease, e.g., causing regression of the disease. In some embodiments, the progressive worsening (e.g., the increasing intensity) of a symptom is slowed, reduced, or halted.

(33) “Preventing” (or prevention of) a disease includes stopping a disease from occurring in a subject, who may be predisposed to the disease but has not yet been diagnosed as having it. Preventing a disease also includes delaying the onset of the disease. The efficacy of the prevention can be evaluated, e.g., as compared to a standard, e.g., delaying onset of the disease as compared to a usual time of onset for the disease in a cohort that has not been treated or compared to historical data on disease onset.

(34) As used herein and depending on the context in which it is used, “therapeutically effective amount” refers to an amount which is effective in reducing, eliminating, treating, preventing or controlling a symptom of a disorder or condition. The term “controlling” is intended to refer to all processes wherein there may be a slowing, interrupting, arresting, or stopping of the progression of the diseases and conditions described herein, but does not necessarily indicate a total elimination of all disease and condition symptoms, and is intended to include prophylactic treatment.

(35) Aspects of the present subject matter relate to administering an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor to a subject at risk of developing an ocular immunoinflammatory disease.

(36) Dry Eye Disease (DED)

(37) DED and related diseases can be caused by autoimmune and environmental conditions as well as any activity that decreases the rate of blinking. Alternatively, DED and related diseases are caused by decreased tear production or a change in tear composition that results in inadequate lubrication of the eye. Contact lens use, eye surgery, and eye injury can induce DED. Finally, DED often occurs as a consequence of aging and hormonal changes.

(38) DED is a multifactorial disorder of the tears and ocular surface that results in symptoms of discomfort, visual disturbance, and tear film instability, with potential damage to the ocular surface. It is accompanied by increased osmolarity of the tear film and inflammation of the ocular surface (Lemp Mass. Report of the National Eye Institute/Industry Workshop on clinical trials in dry eyes. CLAO J 1995; 21:221-2). For a more detailed definition, see The definition and classification of dry eye disease: report of the Definition and Classification Subcommittee of the International Dry Eye WorkShop. Ocular Surface. 2007 April; 5(2):75-92, the entire content of which is incorporated herein by reference. The method of therapy inhibits or reduces the severity of at least one of these signs or symptoms.

(39) Synonyms and related disorders of DED include, but are not limited to, keratoconjunctivitis sicca (KCS), Sjögren syndrome (SS), Sjögren syndrome associated keratoconjunctivitis sicca, non-Sjögren syndrome associated keratoconjunctivitis sicca, keratitis sicca, sicca syndrome, xerophthalmia, tear film disorder, decreased tear production, aqueous tear deficiency (ATD), meibomian gland dysfunction (MGD), and evaporative loss. In some embodiments, a subject is identified as suffering from DED or a related disorder by detecting a sign or symptom selected from the group consisting of dry, scratchy, stingy, itchy, burning or pressured sensations, irritation, pain, redness, inflammation, discharge, and excessive eye watering. In various embodiments, a subject is identified as suffering from DED or a related disorder if their tear composition is insufficient for proper eye tissue lubrication. The method of therapy for DED inhibits or reduces the severity of at least one of sign or symptom of DED.

(40) Subjects at risk of developing DED include subjects who are taking antihistamines, nasal decongestants, tranquilizers, certain blood pressure medicines, Parkinson's medications, birth control pills and/or anti-depressants; subjects with a skin disease on or around the eyelids can result in dry eye; subjects suffering from a disease of the glands in the eyelids (such as meibomian gland dysfunction); subjects who are pregnant; female subjects who are on hormone replacement therapy (such as estrogen and/or progesterone); subjects who have had the refractive surgery known as LASIK; subjects who have suffered from a chemical or thermal burn on the membrane lining the eyelids and covering the eye; subjects afflicted with allergies; subjects afflicted with an immune system disorder (such as Sjögren's syndrome, lupus, or rheumatoid arthritis); subjects who have had an eye infection; subjects who have had ocular exposure to an irritant such as a chemical fume or tobacco smoke; and subjects with exposure keratitis.

(41) Autoimmune Uveitis

(42) The Th17 response plays a major pathogenic role is autoimmune uveitis. See, for example, Yoshimura et al., (2009) Rheumatology (Oxford). 2009; 48:347-354, the entire content of which is incorporated herein by reference. Uveitis is swelling and irritation of the uvea, the middle layer of the eye. The uvea provides most of the blood supply to the retina. Uveitis can be caused by autoimmune disorders, including rheumatoid arthritis or ankylosing spondylitis. The most common form of uveitis is anterior uveitis. This involves inflammation in the front part of the eye. It is often called iritis because it usually only affects the iris, the colored part of the eye. The inflammation may be linked with autoimmune diseases. The disorder may affect only one eye. It is most common in young and middle-aged people. Posterior uveitis affects the back part of the uvea. It involves primarily the choroid, which is a layer of blood vessels and connective tissue in the middle part of the eye. This type of uveitis is called choroiditis. If the retina is also involved, it is called chorioretinitis. Subjects with an autoimmune disease may develop this condition. Another form of uveitis is pars planitis. This inflammation affects the narrowed area (pars plana) between the colored part of the eye (iris) and the choroid. Pars planitis most often occurs in young men. It is generally not associated with any other disease. However, it may be linked to Crohn's disease and multiple sclerosis.

(43) As used herein “autoimmune uveitis” refers to uveitis that is caused or associated with an autoimmune disorder.

(44) Ocular Graft Versus Host Disease (GVHD)

(45) Increased IL-17 levels in tears of patients with ocular GVHD has been observed. See, for example, Kang et al., (2011) J Korean Med Sci, 26: 938-944, the entire content of which is incorporated herein by reference. Ocular Graft Versus Host Disease (GVHD) occurs in patients who have undergone allogenic hematological stem cell transplantation. It can occur in patients who have acute or chronic GVHD, though it is more common in patients with the chronic form. Approximately 40-90% of patients with chronic GVHD will develop ocular symptoms. Ocular manifestations can include moderate to severe keratoconjuncitvitis sicca, bilateral marginal keratitis, anterior uveitis, corneal ulceration or neovascularization.

(46) Memory Th17 Cells

(47) T lymphocytes are circulating small white blood cells that play a central role in cell-mediated immunity. T helper cells (Th) are one subgroup of T lymphocytes expressing a surface marker protein Cluster of Differentiation 4 (CD4). Memory Th17 cells are a recently-identified population of T helper cells that produce interleukin-17 (IL-17). Memory Th17 cells may be referred to herein as “Th17 cells.”

(48) Human IL-7. IL-15, IL-7R and IL-15R

(49) Amino acid sequences of IL-7 are accessible in public databases by the UniProt database accession number P13232 and are set forth herein as SEQ ID NO: 1 (corresponding to isoform 1), SEQ ID NO: 2 (corresponding to isoform 2), and SEQ ID NO: 3 (corresponding to isoform 3), and SEQ ID NO: 36 (corresponding to isoform 4). mRNA sequences which encode amino acid sequences of IL-7 are accessible in public databases by National Center for Biotechnology Information (NCBI) accession number NM_000880 the sequence of which is set forth herein as SEQ ID NO: 4 (corresponding to isoform 1); by NCBI accession number NM_001199886, the sequence of which is set forth herein as SEQ ID NO: 5 (corresponding to isoform 2); by NCBI accession number NM_001199887, the sequence of which is set forth herein as SEQ ID NO: 6 (corresponding to isoform 3); and by NCBI accession number NM_001199888, the sequence of which is set forth herein as SEQ ID NO: 7 (corresponding to isoform 4). The UniProt database entry for accession number P13232 is hereby incorporated herein by reference in its entirety.

(50) Amino acid sequences of interleukin-7 receptor-α (CD127) are accessible in public databases by the UniProt database accession number P16871 and are set forth herein as SEQ ID NO: 8 (corresponding to isoform 1), SEQ ID NO: 9 (corresponding to isoform 3), SEQ ID NO: 10 (corresponding to isoform 4), and SEQ ID NO: 11 (corresponding to isoform 2). An mRNA sequence which encodes an amino acid sequence of interleukin-7 receptor-α (CD127) is accessible in public databases by NCBI accession number NM_002185, the sequence of which is set forth herein as SEQ ID NO: 12 (all isoforms). The UniProt database entry for accession number P16871 is hereby incorporated herein by reference in its entirety.

(51) Amino acid sequences of common-T chain receptor (CD132) are accessible in public databases by the UniProt database accession number P31785 and are set forth herein as SEQ ID NO: 13 (corresponding to isoform 1) and SEQ ID NO: 14 (corresponding to isoform 2). An mRNA which encodes a common-γ chain receptor (CD132) amino acid sequence is accessible in public databases by NCBI accession number NM_000206, the sequence of which is set forth herein as SEQ ID NO: 15 (corresponding to isoform 1). The UniProt database entry for accession number P31785 is hereby incorporated herein by reference in its entirety.

(52) Amino acid sequences of IL-15 are accessible in public databases by the UniProt database accession number P40933 and are set forth herein as SEQ ID NO: 16 (corresponding to isoform IL15-S48AA) and SEQ ID NO: 17 (corresponding to isoform IL15-S21AA). mRNA sequences which encode amino acid sequences of IL-15 are accessible in public databases by NCBI accession number NM_000585, the sequence of which is set forth herein as SEQ ID NO: 18 (corresponding to isoform IL15-S48AA); and by NCBI accession number NM_172175, the sequence of which is set forth herein as SEQ ID NO: 19 (corresponding to isoform IL15-S21AA). The UniProt database entry for accession number P40933 is hereby incorporated herein by reference in its entirety.

(53) An amino acid sequence of IL-2/IL-15 receptor beta chain (CD122) is accessible in public databases by the UniProt database accession number P14784 and is set forth herein as SEQ ID NO: 20. An mRNA sequence which encodes an amino acid sequence of IL-2/IL-15 receptor beta chain (CD122) is accessible in public databases by NCBI accession number NM_000878, the sequence of which is set forth herein as SEQ ID NO: 21. The UniProt database entry for accession number P14784 is hereby incorporated herein by reference in its entirety.

(54) Amino acid sequences of IL-15R alpha are accessible in public databases by the UniProt database accession number Q13261 and are set forth herein as SEQ ID NO: 22 (corresponding to isoform 1), SEQ ID NO: 23 (corresponding to isoform 2), SEQ ID NO: 24 (corresponding to isoform 3), SEQ ID NO: 25 (corresponding to isoform 4), SEQ ID NO: 26 (corresponding to isoform 5), SEQ ID NO: 27 (corresponding to isoform 6), SEQ ID NO: 28 (corresponding to isoform 7), SEQ ID NO: 29 (corresponding to isoform 8), and SEQ ID NO: 30 (corresponding to isoform 9). mRNA sequences which encode amino acid sequences of IL-15R alpha are accessible in public databases by NCBI accession number NM_002189, the sequence of which is set forth herein as SEQ ID NO: 31 (corresponding to isoform 1); by NCBI accession number NM_172200, the sequence of which is set forth herein as SEQ ID NO: 32 (corresponding to isoform 2); by NCBI accession number NM_001243539, the sequence of which is set forth herein as SEQ ID NO: 33 (corresponding to isoform 9); and by NCBI accession number NM_001256765, the sequence of which is set forth herein as SEQ ID NO: 34 (corresponding to isoform 4). The UniProt database entry for accession number Q13261 is hereby incorporated herein by reference in its entirety.

(55) With respect to isoform 1 of interleukin-7 receptor-α (CD127), the signal peptide is predicted to include amino acid positions 1-20, the extracellular domain is predicted to include amino acids 21-239, the transmembrane domain is predicted to include 240-264, and the cytoplasmic domain is predicted to include amino acids 265-459, where the amino acid positions are numbered as in SEQ ID NO: 8.

(56) With respect to isoform 1 of common-γ chain receptor (CD132), the signal peptide is predicted to include amino acid positions 1-22, the extracellular domain is predicted to include amino acids 23-262, the transmembrane domain is predicted to include 263-283, and the cytoplasmic domain is predicted to include amino acids 284-369, where the amino acid positions are numbered as in SEQ ID NO: 13.

(57) With respect to IL-2/IL-15 receptor beta chain (CD122), the signal peptide is predicted to include amino acid positions 1-26, the extracellular domain is predicted to include amino acids 27-240, the transmembrane domain is predicted to include 241-265, and the cytoplasmic domain is predicted to include amino acids 266-551, where the amino acid positions are numbered as in SEQ ID NO: 20.

(58) With respect to isoform 1 of IL-15R alpha, the signal peptide is predicted to include amino acid positions 1-30, the extracellular domain is predicted to include amino acids 31-205, the transmembrane domain is predicted to include 206-228, and the cytoplasmic domain is predicted to include amino acids 229-267, where the amino acid positions are numbered as in SEQ ID NO: 22.

(59) Extracellular domains of proteins within the IL-7 and IL-15 receptor complexes are known. For example, an extracellular domain of interleukin-7 receptor-α (CD127) may include amino acids 1-239 or 21-239 of SEQ ID NO: 8; an extracellular domain of common-γ chain receptor (CD132) may include amino acids 1-262 or 23-262 of SEQ ID NO: 13; an extracellular domain of IL-15R alpha may include amino acids 1-205 or 31-205 of SEQ ID NO: 22; and an extracellular domain of IL-2/IL-15 receptor beta chain (CD122) may include amino acids 1-240 or 27-240 of SEQ ID NO: 20. In certain embodiments, an inhibitor of IL-7 transduction comprises a soluble compound comprising an extracellular domain of a protein within interleukin-7 receptor-α (CD127) or common-γ chain receptor (CD132). In some embodiments, an inhibitor of IL-15 transduction comprises a soluble compound comprising an extracellular domain of IL-15R alpha, IL-2/IL-15 receptor beta chain (CD122), or common-γ chain receptor (CD132). In various embodiments, an extracellular domain is part of a chimeric or fusion protein comprising the extracellular domain and a polypeptide such as an antibody Fc domain or a portion thereof.

(60) Antibodies

(61) The term “antibody” is used in the broadest sense and specifically covers monoclonal antibodies (including full length monoclonal antibodies), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), monovalent antibodies, multivalent antibodies, and antibody fragments so long as they exhibit the desired biological activity (e.g., Fab and/or single-armed antibodies).

(62) An “antibody fragment” refers to a molecule other than an intact antibody that comprises a portion of an intact antibody that binds the antigen to which the intact antibody binds. Examples of antibody fragments include but are not limited to Fv, Fab, Fab′, Fab′-SH, F (ab′).sub.2; diabodies; linear antibodies; single-chain antibody molecules (e.g., scFv); and multispecific antibodies formed from antibody fragments.

(63) The terms “full length antibody,” “intact antibody,” and “whole antibody” are used herein interchangeably to refer to an antibody having a structure substantially similar to a native antibody structure or having heavy chains that contain an Fc region.

(64) An “Fv” fragment is an antibody fragment which contains a complete antigen recognition and binding site. This region consists of a dimer of one heavy and one light chain variable domain in tight association, which can be covalent in nature, for example in scFv. It is in this configuration that the three hypervariable regions (HVRs) of each variable domain interact to define an antigen binding site on the surface of the VH-VL dimer. Collectively, the six HVRs or a subset thereof confer antigen binding specificity to the antibody. However, even a single variable domain (or half of an Fv comprising only three HVRs specific for an antigen) has the ability to recognize and bind antigen, although usually at a lower affinity than the entire binding site.

(65) A “Fab” fragment contains a variable and constant domain of the light chain and a variable domain and the first constant domain (CHI) of the heavy chain. F(ab′) 2 antibody fragments comprise a pair of Fab fragments which are generally covalently linked near their carboxy termini by hinge cysteines between them. Other chemical couplings of antibody fragments are also known in the art.

(66) “Single-chain Fv” or “scFv” antibody fragments comprise the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain. Generally the Fv polypeptide further comprises a polypeptide linker between the VH and L domains, which enables the scFv to form the desired structure for antigen binding. For a review of scFv, see Pluckthun in The Pharmacology of Monoclonal Antibodies, Vol 113, Rosenburg and Moore eds. Springer-Verlag, New York, pp. 269-31S (1994).

(67) The term “diabodies” refers to small antibody fragments with two antigen-binding sites, which fragments comprise a heavy chain variable domain (VH) connected to a light chain variable domain (VL) in the same polypeptide chain (VH and VL). By using a linker that is too short to allow pairing between the two domains on the same chain, the domains are forced to pair with the complementary domains of another chain and create two antigen-binding sites. Diabodies are described more fully in, for example, BP 404,097; WO 93/11161; and Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993).

(68) The expression “linear antibodies” refers to the antibodies described in Zapata et al., Protein Eng., 8 (10): 1057-1062 (1995). Briefly, these antibodies comprise a pair of tandem Fd segments (V.sub.H-C.sub.H1-V.sub.H-C.sub.H1) which, together with complementary light chain polypeptides, form a pair of antigen binding regions. Linear antibodies can be bispecific or monospecific.

(69) The term “monoclonal antibody” as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and/or bind the same epitope, except for possible variant antibodies, e.g., containing naturally occurring mutations or arising during production of a monoclonal antibody preparation, such variants generally being present in minor amounts. In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. Thus, the modifier “monoclonal” indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies, and is not to be construed as requiring production of the antibody by any particular method. For example, the monoclonal antibodies to be used may be made by a variety of techniques, including but not limited to the hybridoma method (see, e.g., Kohler and Milstein, Nature 256:495-97 (1975); Kozbor et al., Immunol. Today 4:72 (1983); Cole et al., in Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc. 77-96 (1985)), recombinant DNA methods (see, e.g., U.S. Pat. No. 4,816,567), phage-display methods (see, e.g., Clackson et al., Nature 352:624-28 (1991) and Marks et al., J. Mol. Biol. 222 (3):581-97 (1991)), and methods utilizing transgenic animals containing all or part of the human immunoglobulin loci, such methods and other exemplary methods for making monoclonal antibodies being described herein.

(70) The term “chimeric” antibody refers to an antibody in which a portion of the heavy and/or light chain is derived from a particular source or species, while the remainder of the heavy and/or light chain is derived from a different source or species.

(71) A “humanized” antibody refers to a chimeric antibody comprising amino acid residues from non-human HVRs and amino acid residues from human FRs. In certain embodiments, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the HVRs (e.g., CDRs) correspond to those of a non-human antibody, and all or substantially all of the FRs correspond to those of a human antibody. A humanized antibody optionally may comprise at least a portion of an antibody constant region derived from a human antibody. A “humanized form” of an antibody, e.g., a non-human antibody, refers to an antibody that has undergone humanization.

(72) A “human antibody” is one which possesses an amino acid sequence which corresponds to that of an antibody produced by a human or a human cell or derived from a non-human source that utilizes human antibody repertoires or other human antibody-encoding sequences. This definition of a human antibody specifically excludes a humanized antibody comprising non-human antigen-binding residues.

(73) Antisense Oligonucleotides

(74) Antisense oligonucleotides are nucleotide sequences which are complementary to a specific DNA or RNA sequence. Once introduced into a cell, the complementary nucleotides combine with natural sequences produced by the cell to form complexes and block either transcription or translation. Preferably, an antisense oligonucleotide is at least 11 nucleotides in length, but can be at least 12, 15, 20, 25, 30, 35, 40, 45, or 50 or more nucleotides long. Longer sequences also can be used. Antisense oligonucleotide molecules can be directly administered or provided in a DNA construct and introduced into a cell to decrease the level of IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha gene products in the cell.

(75) In some embodiments, antisense oligonucleotides can be deoxyribonucleotides, ribonucleotides, or a combination of both. In various embodiments, oligonucleotides mag be modified to increase the half-life of the oligonucleotide in vivo. Oligonucleotides can be synthesized manually or by an automated synthesizer, by covalently linking the 5′ end of one nucleotide with the 3′ end of another nucleotide with non-phosphodiester internucleotide linkages such alkylphosphonates, phosphorothioates, phosphorodithioates, alkylphosphonothioates, alkylphosphonates, phosphoramidates, phosphate esters, carbamates, acetamidate, carboxymethyl esters, carbonates, and phosphate triesters.

(76) Modifications of gene expression can be obtained by designing antisense oligonucleotides which will form duplexes to the control, 5′, or regulatory regions of the gene. Oligonucleotides derived from the transcription initiation site, e.g., between positions −10 and +10 from the start site, are used in various embodiments. Similarly, inhibition can be achieved using “triple helix” base-pairing methodology. Triple helix pairing is useful because it causes inhibition of the ability of the double helix to open sufficiently for the binding of polymerases, transcription factors, or chaperons. Therapeutic advances using triplex DNA have been described in the literature (Nicholls et al., 1993, J Immunol Meth 165:81-91). An antisense oligonucleotide also can be designed to block translation of mRNA by preventing the transcript from binding to ribosomes.

(77) Precise complementarity is not required for successful complex formation between an antisense oligonucleotide and the complementary sequence of an IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha polynucleotide. Antisense oligonucleotides which comprise, for example, 2, 3, 4, or 5 or more stretches of contiguous nucleotides which are precisely complementary to an IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha polynucleotide, each separated by a stretch of contiguous nucleotides which are not complementary to adjacent IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha nucleotides, can provide sufficient targeting specificity for IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha mRNA, respectively. Preferably, each stretch of complementary contiguous nucleotides is at least 4, 5, 6, 7, or 8 or more nucleotides in length. Noncomplementary intervening sequences are preferably 1, 2, 3, or 4 nucleotides in length. One skilled in the art can easily use the calculated melting point of an antisense-sense pair to determine the degree of mismatching which will be tolerated between a particular antisense oligonucleotide and a particular IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha polynucleotide sequence. Antisense oligonucleotides can be modified without affecting their ability to hybridize to an IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha polynucleotide. These modifications can be internal or at one or both ends of the antisense molecule. For example, internucleoside phosphate linkages can be modified by adding cholesteryl or diamine moieties with varying numbers of carbon residues between the amino groups and terminal ribose. Modified bases and/or sugars, such as arabinose instead of ribose, or a 3′, 5′-substituted oligonucleotide in which the 3′ hydroxyl group or the 5′ phosphate group are substituted, also can be employed in a modified antisense oligonucleotide. These modified oligonucleotides can be prepared by methods well known in the art. In some embodiments, the antisense oligonucleotide is a phosphorodiamidate morpholino oligomer (also referred to herein as a “PMO” or a “morpholino”).

(78) Ribozymes

(79) Ribozymes are RNA molecules with catalytic activity (Uhlmann et al., 1987, Tetrahedron. Lett. 215, 3539-3542). Ribozymes can be used to inhibit gene function by cleaving an RNA sequence, as is known in the art. The mechanism of ribozyme action involves sequence-specific hybridization of the ribozyme molecule to complementary target RNA, followed by endonucleolytic cleavage. Examples include engineered hammerhead motif ribozyme molecules that can specifically and efficiently catalyze endonucleolytic cleavage of specific nucleotide sequences. The coding sequence of a polynucleotide can be used to generate ribozymes which will specifically bind to rnRNA transcribed from the polynucleotide. Methods of designing and constructing ribozymes which can cleave other RNA molecules in trans in a highly sequence specific manner have been developed and described in the art. For example, the cleavage activity of ribozymes can be targeted to specific RNAs by engineering a discrete “hybridization” region into the ribozyme. The hybridization region contains a sequence complementary to the target RNA and thus specifically hybridizes with the target RNA.

(80) Specific ribozyme cleavage sites within an RNA target can be identified by scanning the target molecule for ribozyme cleavage sites which include the following sequences: GUA, GUU, and GUC. Once identified, short RNA sequences of between 15 and 20 ribonucleotides corresponding to the region of the target RNA containing the cleavage site can be evaluated for secondary structural features which may render the target inoperable. Suitability of candidate RNA targets also can be evaluated by testing accessibility to hybridization with complementary oligonucleotides using ribonuclease protection assays. The nucleotide sequences shown in nucleotide sequences described herein (e.g., SEQ ID NOs 4-7, 12, 15, 18, 19, 21, and 31-34) and their complements provide sources of suitable hybridization region sequences. Longer complementary sequences can be used to increase the affinity of the hybridization sequence for the target. The hybridizing and cleavage regions of the ribozyme can be integrally related such that upon hybridizing to the target RNA through the complementary regions, the catalytic region of the ribozyme can cleave the target.

(81) Ribozymes can be introduced into cells as part of a DNA construct. Mechanical methods, such as microinjection, liposome-mediated transfection, electroporation, or calcium phosphate precipitation, can be used to introduce a ribozyme-containing DNA construct into cells in which it is desired to decrease an IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha expression. Alternatively, if it is desired that the cells stably retain the DNA construct, the construct can be supplied on a plasmid and maintained as a separate element or integrated into the genome of the cells, as is known in the art. A ribozyme-encoding DNA construct can include transcriptional regulatory elements, such as a promoter element, an enhancer or VAS element, and a transcriptional terminator signal, for controlling transcription of ribozymes in the cells (U.S. Pat. No. 5,641,673). Ribozymes also can be engineered to provide an additional level of regulation, so that destruction of mRNA occurs only when both a ribozyme and a target gene are induced in the cells.

(82) RNA Interference

(83) As used herein, “RNA interference” compound refers to a compound capable of inducing RNA interference or “RNAi” of an IL-7, IL-15, CD132, CD127, CD122, or IL-15R alpha expression, depending on the context. RNAi involves mRNA degradation, but many of the biochemical mechanisms underlying this interference are unknown. The use of RNAi has been described in Fire et al., 1998, Carthew et al., 2001, and Elbashir et al., 2001, the contents of which are incorporated herein by reference.

(84) Isolated RNA molecules can mediate RNAi. That is, the isolated RNA molecules of the present invention mediate degradation or block expression of mRNA that is the transcriptional product of the gene, which is also referred to as a target gene. For convenience, such mRNA may also be referred to herein as mRNA to be degraded. The terms RNA, RNA molecule (s), RNA segment(s) and RNA fragment(s) may be used interchangeably to refer to RNA that mediates RNA interference. These terms include double-stranded RNA, small interfering RNA (siRNA), hairpin RNA, single-stranded RNA, isolated RNA (partially purified RNA, essentially pure RNA, synthetic RNA, recombinantly produced RNA), as well as altered RNA that differs from naturally occurring RNA by the addition, deletion, substitution and/or alteration of one or more nucleotides. Such alterations can include addition of non-nucleotide material, such as to the end(s) of the RNA or internally (at one or more nucleotides of the RNA). Nucleotides in the RNA molecules of the present invention can also comprise nonstandard nucleotides, including non-naturally occurring nucleotides or deoxyribonucleotides. Collectively, all such altered RNAi molecules are referred to as analogs or analogs of naturally-occurring RNA. RNA of the present invention need only be sufficiently similar to natural RNA that it has the ability to mediate RNAi.

(85) As used herein the phrase “mediate RNAi” refers to and indicates the ability to distinguish which mRNA molecules are to be afflicted with the RNAi machinery or process. RNA that mediates RNAi interacts with the RNAi machinery such that it directs the machinery to degrade particular mRNAs or to otherwise reduce the expression of the target protein. In one embodiment, the present invention relates to RNA molecules that direct cleavage of specific mRNA to which their sequence corresponds. It is not necessary that there be perfect correspondence of the sequences, but the correspondence must be sufficient to enable the RNA to direct RNAi inhibition by cleavage or blocking expression of the target mRNA.

(86) As noted above, the RNA molecules of the present invention in general comprise an RNA portion and some additional portion, for example a deoxyribonucleotide portion. The total number of nucleotides in the RNA molecule is suitably less than in order to be effective mediators of RNAi. In preferred RNA molecules, the number of nucleotides is 16 to 29, more preferably 18 to 23, and most preferably 21-23.

(87) Aptamers

(88) As used herein, the terms “aptamer(s)” or “aptamer sequence(s)” are meant to refer to single stranded nucleic acids (RNA or DNA) whose distinct nucleotide sequence determines the folding of the molecule into a unique three dimensional structure. Aptamers comprising 15 to 120 nucleotides can be selected in vitro from a randomized pool of oligonucleotides (1014-1015 molecules). Aptamers that bind to pre-selected targets including proteins and peptides with high affinity and specificity can be designed and/or selected using methods known in the art. See, e.g., Cox, J. C.; Ellington, A. D. (2001) Bioorganic & Medicinal Chemistry 9 (10): 2525-2531; Cox, J. C.; Hayhurst, A.; Hesselberth, J.; Bayer, T. S.; Georgiou, G.; Ellington, A. D. (2002) Nucleic Acids Research 30 (20): e108; and Neves, M. A. D.; O. Reinstein; M. Saad; P. E. Johnson (2010) Biophys Chem 153 (1): 9-16, the entire content of each of which is incorporated herein by reference.

(89) Pharmaceutical Formulations and Delivery to the Eye

(90) Dosages, formulations, dosage volumes, regimens, and methods for antagonizing the IL-7/IL-7R and IL-15/IL-17R pathways can vary. Thus, minimum and maximum effective dosages vary depending on the method of administration.

(91) In various embodiments of the invention, a composition comprising an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor may be administered only once or multiple times. For example, an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor may be administered using a method disclosed herein at least about once, twice, three times, four times, five times, six times, or seven times per day week, month, or year. In some embodiments, a composition comprising an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor is administered once per month. In certain embodiments, the composition is administered once per month via intravitreal injection. In various embodiments, such as embodiments involving eye drops, a composition is self-administered.

(92) Preferred formulations are in the form of a solid, a paste, an ointment, a gel, a liquid, an aerosol, a mist, a polymer, a contact lens, a film, an emulsion, or a suspension. The formulations are administered topically, e.g., the composition is delivered and directly contacts the eye. The composition is present at a concentration of 0.01-50% (weight/volume). For example, the inhibitory composition is present at concentrations of 1% (weight/volume), 10% (weight/volume), 20% (weight/volume), 25% (weight/volume), 30% (weight/volume), 40% (weight/volume), 50% (weight/volume), or any percentage point in between. The method does not involve systemic administration or planned substantial dissemination of the composition to non-ocular tissue.

(93) Optionally, the composition further contains a pharmaceutically-acceptable carrier. Exemplary pharmaceutical carriers include, but are not limited to, compounds selected from the group consisting of a physiological acceptable salt, poloxamer analogs with carbopol, carbopol/hydroxypropyl methyl cellulose (HPMC), carbopol-methyl cellulose, a mucolytic agent, carboxymethylcellulose (CMC), hyaluronic acid, cyclodextrin, and petroleum. In one embodiment, the mucolytic agent is N-acetyl cysteine.

(94) For the treatment of an ocular immunoinflammatory disease, an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor (e.g., a pharmaceutical composition comprising an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor) may be administered locally, e.g., as a topical eye drop, peri-ocular injection (e.g., sub-tenon), intraocular injection, intravitreal injection, retrobulbar injection, intraretinal injection, subconjunctival injection, or using iontophoresis, or peri-ocular devices which can actively or passively deliver drug.

(95) Pharmaceutical formulations adapted for topical administration may be formulated as aqueous solutions, ointments, creams, suspensions, lotions, powders, solutions, pastes, gels, sprays, aerosols, liposomes, microcapsules, microspheres, or oils.

(96) Pharmaceutical formulations adapted for topical administrations to the eye include eye drops wherein an IL-7, IL-7R, IL-15, and/or IL-15R inhibitor is dissolved or suspended in a suitable carrier, especially an aqueous solvent. Formulations to be administered to the eye will have ophthalmically compatible pH and osmolality. The term “ophthalmically acceptable vehicle” means a pharmaceutical composition having physical properties (e.g., pH and/or osmolality) that are physiologically compatible with ophthalmic tissues.

(97) In some embodiments, an ophthalmic composition of the present invention is formulated as sterile aqueous solutions having an osmolality of from about 200 to about 400 milliosmoles/kilogram water (“mOsm/kg”) and a physiologically compatible pH. The osmolality of the solutions may be adjusted by means of conventional agents, such as inorganic salts (e.g., NaCl), organic salts (e.g., sodium citrate), polyhydric alcohols (e.g., propylene glycol or sorbitol) or combinations thereof.

(98) In various embodiments, the ophthalmic formulations of the present invention may be in the form of liquid, solid or semisolid dosage form. The ophthalmic formulations of the present invention may comprise, depending on the final dosage form, suitable ophthalmically acceptable excipients. In some embodiments, the ophthalmic formulations are formulated to maintain a physiologically tolerable pH range. In certain embodiments, the pH range of the ophthalmic formulation is in the range of from about 5 to about 9. In some embodiments, pH range of the ophthalmic formulation is in the range of from about 6 to about 8, or is about 6.5, about 7, or about 7.5.

(99) In some embodiments, the composition is in the form of an aqueous solution, such as one that can be presented in the form of eye drops. By means of a suitable dispenser, a desired dosage of the active agent can be metered by administration of a known number of drops into the eye, such as by one, two, three, four, or five drops.

(100) One or more ophthalmically acceptable pH adjusting agents and/or buffering agents can be included in a composition of the invention, including acids such as acetic, boric, citric, lactic, phosphoric, and hydrochloric acids; bases such as sodium hydroxide, sodium phosphate, sodium borate, sodium citrate, sodium acetate, and sodium lactate; and buffers such as citrate/dextrose, sodium bicarbonate, and ammonium chloride. Such acids, bases, and buffers can be included in an amount required to maintain pH of the composition in an ophthalmically acceptable range. One or more ophthalmically acceptable salts can be included in the composition in an amount sufficient to bring osmolality of the composition into an ophthalmically acceptable range. Such salts include those having sodium, potassium, or ammonium cations and chloride, citrate, ascorbate, borate, phosphate, bicarbonate, sulfate, thiosulfate, or bisulfite anions.

(101) Pharmaceutical compositions for ocular delivery also include in situ gellable aqueous composition. Such a composition comprises a gelling agent in a concentration effective to promote gelling upon contact with the eye or with lacrimal fluid. Suitable gelling agents include but are not limited to thermosetting polymers. The term “in situ gellable” as used herein includes not only liquids of low viscosity that form gels upon contact with the eye or with lacrimal fluid, but also includes more viscous liquids such as semi-fluid and thixotropic gels that exhibit substantially increased viscosity or gel stiffness upon administration to the eye. See, for example, Ludwig, Adv. Drug Deliv. Rev. 3; 57:1595-639 (2005), the entire content of which is incorporated herein by reference.

(102) Drug Delivery by Contact Lens

(103) The invention comprises a contact lens and a composition that inhibits an activity of an inflammatory interleukin-1 cytokine. For example, the composition is incorporated into or coated onto the lens. The composition is chemically bound or physically entrapped by the contact lens polymer. Alternatively, a color additive is chemically bound or physically entrapped by the polymer composition that is released at the same rate as the therapeutic drug composition, such that changes in the intensity of the color additive indicate changes in the amount or dose of therapeutic drug composition remaining bound or entrapped within the polymer. Alternatively, or in addition, an ultraviolet (UV) absorber is chemically bound or physically entrapped within the contact lens polymer. The contact lens is either hydrophobic or hydrophilic.

(104) Exemplary materials used to fabricate a hydrophobic lens with means to deliver the compositions of the invention include, but are not limited to, amefocon A, amsilfocon A, aquilafocon A, arfocon A, cabufocon A, cabufocon B, carbosilfocon A, crilfocon A, crilfocon B, dimefocon A, enflufocon A, enflofocon B, erifocon A, flurofocon A, flusilfocon A, flusilfocon B, flusilfocon C, flusilfocon D, flusilfocon E, hexafocon A, hofocon A, hybufocon A, itabisfluorofocon A, itafluorofocon A, itafocon A, itafocon B, kolfocon A, kolfocon B, kolfocon C, kolfocon D, lotifocon A, lotifocon B, lotifocon C, melafocon A, migafocon A, nefocon A, nefocon B, nefocon C, onsifocon A, oprifocon A, oxyfluflocon A, paflufocon B, paflufocon C, paflufocon D, paflufocon E, paflufocon F, pasifocon A, pasifocon B, pasifocon C, pasifocon D, pasifocon E, pemufocon A, porofocon A, porofocon B, roflufocon A, roflufocon B, roflufocon C, roflufocon D, roflufocon E, rosilfocon A, satafocon A, siflufocon A, silafocon A, sterafocon A, sulfocon A, sulfocon B, telafocon A, tisilfocon A, tolofocon A, trifocon A, unifocon A, vinafocon A, and wilofocon A.

(105) Exemplary materials used to fabricate a hydrophilic lens with means to deliver the compositions of the invention include, but are not limited to, abafilcon A, acofilcon A, acofilcon B, acquafilcon A, alofilcon A, alphafilcon A, amfilcon A, astifilcon A, atlafilcon A, balafilcon A, bisfilcon A, bufilcon A, comfilcon A, crofilcon A, cyclofilcon A, darfilcon A, deltafilcon A, deltafilcon B, dimefilcon A, droxfilcon A, elastofilcon A, epsilfilcon A, esterifilcon A, etafilcon A, focofilcon A, galyfilcon A, genfilcon A, govafilcon A, hefilcon A, hefilcon B, hefilcon C, hilafilcon A, hilafilcon B, hioxifilcon A, hioxifilcon B, hioxifilcon C, hydrofilcon A, lenefilcon A, licryfilcon A, licryfilcon B, lidofilcon A, lidofilcon B, lotrafilcon A, lotrafilcon B, mafilcon A, mesafilcon A, methafilcon B, mipafilcon A, nelfilcon A, netrafilcon A, ocufilcon A, ocufilcon B, C, ocufilcon D, ocufilcon E, ofilcon A, omafilcon A, oxyfilcon A, pentafilcon A, perfilcon A, pevafilcon A, phemfilcon A, polymacon, senofilcon A, silafilcon A, siloxyfilcon A, surfilcon A, tefilcon A, tetrafilcon A, trilfilcon A, vifilcon A, vifilcon B, and xylofilcon A.

General Definitions

(106) Unless specifically defined otherwise, all technical and scientific terms used herein shall be taken to have the same meaning as commonly understood by one of ordinary skill in the art (e.g., in cell culture, molecular genetics, and biochemistry).

(107) As used herein, the term “about” in the context of a numerical value or range means ±10% of the numerical value or range recited or claimed, unless the context requires a more limited range.

(108) In the descriptions above and in the claims, phrases such as “at least one of” or “one or more of” may occur followed by a conjunctive list of elements or features. The term “and/or” may also occur in a list of two or more elements or features. Unless otherwise implicitly or explicitly contradicted by the context in which it is used, such a phrase is intended to mean any of the listed elements or features individually or any of the recited elements or features in combination with any of the other recited elements or features. For example, the phrases “at least one of A and B;” “one or more of A and B;” and “A and/or B” are each intended to mean “A alone, B alone, or A and B together.” A similar interpretation is also intended for lists including three or more items. For example, the phrases “at least one of A, B, and C;” “one or more of A, B, and C;” and “A, B, and/or C” are each intended to mean “A alone, B alone, C alone, A and B together, A and C together, B and C together, or A and B and C together.” In addition, use of the term “based on,” above and in the claims is intended to mean, “based at least in part on,” such that an unrecited feature or element is also permissible

(109) It is understood that where a parameter range is provided, all integers within that range, and tenths thereof, are also provided by the invention. For example, “0.2-5 mg” is a disclosure of 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg etc. up to and including 5.0 mg.

(110) The transitional term “comprising,” which is synonymous with “including,” “containing,” or “characterized by,” is inclusive or open-ended and does not exclude additional, unrecited elements or method steps. By contrast, the transitional phrase “consisting of” excludes any element, step, or ingredient not specified in the claim. The transitional phrase “consisting essentially of” limits the scope of a claim to the specified materials or steps “and those that do not materially affect the basic and novel characteristic(s)” of the claimed invention.

(111) Examples are provided below to facilitate a more complete understanding of the invention. The following examples illustrate the exemplary modes of making and practicing the invention. However, the scope of the invention is not limited to specific embodiments disclosed in these Examples, which are for purposes of illustration only, since alternative methods can be utilized to obtain similar results.

Example 1: Effects of IL-7/IL-7R and IL-15/IL-15R Signaling Inhibition on the Maintenance of DED Memory Th17 Cells

(112) A goal of this study was to show that diminishing or eliminating memory Th17 cell-mediated ocular surface inflammation in DED can be achieved by interfering with the survival of memory Th17 cells through the topical blockade of IL-7/IL-7R and IL-15/IL-15R signaling. Experiments were performed as described below.

(113) Chronic DED develops when mice are exposed to desiccating stress using a controlled environment chamber for 14 days and then housed in a standard environment for additional 14 days. There is an increase in the expression levels of IL-7 and IL-15 in both ocular surface and draining lymph nodes (DLNs) (FIG. 1A). In addition, memory Th17 cells from chronic DED also exhibit a significant up-regulation of both IL-7R and IL-15R (FIG. 1B).

(114) Next examined were the effects of IL-7/IL-7R and IL-15/IL-15R signaling blockade on the maintenance of DED memory Th17 cells using an in vitro culture system. Total DLNs were isolated from chronic DED and cultured for 72 hours. Initially, antibodies targeting the cytokine receptors were tested. After adding anti-IL-7R and anti-IL-15R antibodies to the explant culture, the frequency of recovered memory Th17 cells was reduced by >70% (FIG. 2). Thereafter, effects of modifying the environmental cytokine milieu on the maintenance of memory Th17 cells were investigated. Specifically, the DLN explant cultures were supplemented with the following various factors: IL-7, IL-15, combination of IL-7 and IL-15, anti-IL-7 antibody, anti-IL-15 antibody, combination of anti-IL-7 and anti-IL-15 antibodies, or control IgG. The presence of IL-7, IL-15, or combination of IL-7 and IL-15 led to significantly higher frequencies of recovered memory Th17 cells than the presence of anti-IL-7, anti-IL-15, or combination of anti-IL-7 and anti-IL-15 antibodies (p<0.05) (FIG. 3).

(115) These findings demonstrate that both IL-7 and IL-15 maintain memory Th17 cells in DED. As described herein, reducing IL-7 and/or IL-15 signaling is a useful and specific therapeutic approach for treating DED.

Example 2: Topical Treatment with IL-7 and IL-15 Inhibitors Reduces DED Severity

(116) In vivo topical treatment with anti-IL-7 or anti-IL-15 antibodies significantly decreased disease severity and depleted memory Th17 cells in chronic DED mice. DED was induced in mice and the mice were treated with topical anti-IL-7, anti-IL-15, or isotype IgG (control group) eye drops for 14 days. Both anti-IL-7 and anti-IL-15 treatment significantly decreased disease severity as compared with control group (FIG. 5A). At the end of the treatment, ocular surface memory Th17 cells were almost completely depleted by each of anti-IL-7 and anti-IL-15 treatment (FIG. 5B).

(117) Mouse IL-7 polyclonal antibody was used as the anti-IL-7 antibody in this study (Clone #AB-407, R&D Systems, Minneapolis, Minn., USA) at a dose of 10 μg three times per day (1 mg/ml). This antibody was provided in lyophilized form by the manufacture (R&D Systems) and was reconstituted in sterile phosphate buffered saline (PBS).

(118) Anti-mouse IL-15 antibody was used as the anti-IL-15 antibody in this study (Clone #AIO.3, eBioscience, San Diego, Calif., USA) at a dose of 10 μg three times per day (1 mg/ml). This antibody was provided in azide-free aqueous buffer by the manufacture (eBioscience).

(119) Isotype IgG is Polyclonal goat IgG (Catalog #ab37373, Abcam, Cambridge, Mass., USA). It was administered at a dose of 10 μg three times per day (1 mg/ml). This antibody was provided in borate buffered saline by the manufacture (Abcam).

(120) Corneal fluorescein staining was used as a clinical evaluation tool for DED severity. Fluorescein (Sigma-Aldrich, St. Louis, Mo., USA; 1 μl 2.5%) was applied into the lateral conjunctival sac of the mice, and after 3 minutes, corneas were examined with a slit lamp biomicroscope under cobalt blue light. Punctate staining was recorded in a masked manner with the standard National Eye Institute grading system of 0-3 for each of the five areas of the cornea-central, superior, inferior, nasal, and temporal (CLAO J. 1995; 21:221-32).

Other Embodiments

(121) While the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

(122) The patent and scientific literature referred to herein establishes the knowledge that is available to those with skill in the art. All United States patents and published or unpublished United States patent applications cited herein are incorporated by reference. All published foreign patents and patent applications cited herein are hereby incorporated by reference. Genbank and NCBI submissions indicated by accession number cited herein are hereby incorporated by reference. All other published references, documents, manuscripts and scientific literature cited herein are hereby incorporated by reference.

(123) While this invention has been particularly shown and described with references to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the invention encompassed by the appended claims.