Methods of treating eye disorders

Abstract

Provided herein are methods of treating an eye disorder, including, for example, diabetic macular edema (DME) or macular edema secondary to inflammation (MESI) or wet age-related macular degeneration (wAMD), by administering a fusion protein and / or a conjugate thereof to a subject in need thereof.

Description

embodiments, the method includes administering no more than 6 loading doses of the fusion protein conjugate.

[0005] Also provided is a method of treating an eye disorder, comprising: identifying a subject having an eye disorder; and administering at least one loading dose of a fusion protein conjugate to the subject, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non- native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non- native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 4 or 8 weeks after a final loading dose.

[0006] Provided herein is a method of treating diabetic macular edema (DME), comprising: identifying a subject having DME; and administering 3 or 4 intravitreal loading doses of a fusion protein conjugate to the subject at Q4W, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody comprising a heavy chain and a light chain; and a VEGF Trap fused to the heavy chain, wherein the heavy chain fused to the VEGF Trap comprises an amino acid sequence as set forth in SEQ ID NO: 170 (with or without the C-terminal lysine), and the light chain comprises an amino acid sequence as set forth in SEQ ID NO: 169, wherein the fusion protein conjugate comprises the following structure:O O X PC n2 PC PC = P N(CH ) O O O CH O PC O X n5 PC ,wherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L, the polymer is bonded to the heavy chain of the anti-IL-6 antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains, PC is O CH3, where the curvy line indicates the point of attachment to the rest ofa) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%, wherein each dose comprises about 1 mg to about 6 mg of the fusion protein conjugate (by weight of the fusion protein) per eye, whereby the subject retains a therapeutic result of administering the 3 or 4 intravitreal loading doses of the fusion proteinconjugate for at least 4, 8, 12, 16, 20, 24 or more weeks after a final loading dose (e.g., after the third or fourth loading dose).

[0007] Also provided is a method of treating an eye disorder, comprising: identifying a subject having an eye disorder; and administering 3 or 4 intravitreal loading doses of a fusion protein conjugate to the subject at Q4W, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain fused to the VEGF Trap comprises an amino acid sequence as set forth in SEQ ID NO: 170 (with or without the C- terminal lysine), and the light chain comprises an amino acid sequence as set forth in SEQ ID NO: 169, wherein the fusion protein conjugate comprises the following structure: ,wherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L, the polymer is bonded to the heavy chain of the anti-IL-6 antibody through thesulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains, PC is , where the curvy line indicates the point of attachment to the rest ofa) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%, wherein each dose comprises about 1 mg to about 6 mg of the fusion protein conjugate (by weight of the fusion protein) per eye, whereby the subject retains a therapeutic result of administering the 3 or 4 intravitreal doses of the fusion protein conjugate for at least 4, 8, 12, 16, 20, 24, or more weeks after a final loading dose.

[0008] Provided herein is a method of treating a subject for an inflammatory condition, comprising: administering an intravitreal or intravenous dose of a fusion protein conjugate to a subject in need thereof; and administering one or more subsequent intravitreal or intravenous doses of the fusion protein conjugate no more frequently than Q2W, wherein the fusion protein conjugate comprises: a fusion protein comprising a VEGF Trap fused to a heavy chain of an anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody, wherein each of the intravitreal dose and the one or more subsequent intravitreal doses comprises about 1-6 mg of the fusion protein conjugate (by weight of the fusion protein), or wherein each of the intravenous dose and the one or more subsequent intravenous doses comprises about 1-6 mg / kg body weight of the fusion protein conjugate (by weight of the fusion protein).

[0009] Also provided is a method of treating a subject for an inflammatory condition, comprising: administering an intravitreal or intravenous dose of a fusion protein conjugate to a subject in need thereof; and administering one or more subsequent intravitreal or intravenous doses of the fusion protein conjugate no more frequently than Q2W, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain fused to the VEGF Trap comprises an amino acid sequence as set forth in SEQ ID NO: 170 (with or without the C-terminal lysine), and the light chain comprises an amino acid sequenceas set forth in SEQ ID NO: 169, wherein the fusion protein conjugate comprises the following structure: O O X PC n2 PC PC = P N(CH ) O O O CH O PC O X n5 PC ,wherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L, the polymer is bonded to the heavy chain of the anti-IL-6 antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains, PC is O CH3, wherein the curvy line indicates the point of attachment to the restof the polymer, where –X is: a) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%, wherein each of the intravitreal dose and the one or moresubsequent intravitreal doses comprises about 1-6 mg of the fusion protein conjugate (by weight of the fusion protein), or wherein each of the intravenous dose and the one or more subsequent intravenous doses comprises about 1-6 mg / kg body weight of the fusion protein conjugate (by weight of the fusion protein).

[0010] Also provided is a method of treating an ocular inflammatory condition, comprising: administering to a subject in need of treating an ocular inflammatory condition one or more loading doses (optionally 3-5 loading doses) of a fusion protein and / or a conjugate thereof no more frequently than Q4W, the fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of the dual anti-IL-6 antibody / VEGF Trap therapy for at least 4 or 8 weeks after a final loading dose.

[0011] Provided herein is a method of treating diabetic macular edema (DME), comprising: identifying a subject with DME; administering to the subject a first dose of a fusion protein and / or a conjugate thereof, the fusion protein comprising: an anti-IL-6 antibody comprising: a heavy chain comprising the amino acid sequence of SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a light chain comprising the amino acid of SEQ ID NO: 169 or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a VEGF Trap fused to the heavy chain; administering a second dose of the fusion protein to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein to the subject about 4 weeks after the third dose; and administering a fifth dose of the fusion protein to the subject about 4 weeks after the fourth dose, wherein the first, second, third, fourth, and fifth dose each comprises about 2.5, 5, or 10 mg of the fusion protein.

[0012] Provided herein is a method of treating macular edema secondary to inflammation (MESI), comprising: identifying a subject with MESI; administering to the subject a first dose of a fusion protein and / or a conjugate thereof, the fusion protein comprising: an anti-IL-6 antibody comprising: a heavy chain comprising the amino acid sequence of SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a light chain comprising the aminoacid of SEQ ID NO: 169 or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a VEGF Trap fused to the heavy chain; administering a second dose of the fusion protein to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein to the subject about 4 weeks after the second dose; and administering a fourth dose of the fusion protein to the subject about 4 weeks after the third dose, wherein the first, second, third, and fourth, dose each comprises about 2.5, 5, or 10 mg of the fusion protein.

[0013] Also provided is a method of treating uveitic macular edema (UME), comprising: identifying a subject with UME; administering to the subject a first dose of a fusion protein and / or a conjugate thereof, the fusion protein comprising: an anti-IL-6 antibody comprising: a heavy chain comprising the amino acid sequence of SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a light chain comprising the amino acid of SEQ ID NO: 169 or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody; administering a second dose of the fusion protein to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein to the subject about 4 weeks after the third dose, wherein the first, second, third, and fourth dose each comprises about 2.5, 5, or 10 mg of the fusion protein, optionally wherein the method comprises administering an individualized dose of the fusion protein to the subject after the fourth dose and no more frequently than Q4W, wherein each individualized dose is administered upon determining that there is a decline in the subject’s eye health. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] FIGs. 1A-1C are flow diagrams depicting some non-limiting embodiments of methods of the present disclosure.

[0015] FIG.2 is a collection of graphs showing changes in visual acuity and retinal thickness over time in patients treated with aflibercept.

[0016] FIGs. 3A and 3B are a collection of schematic diagrams showing the role of IL-6 and VEGF pathways in ocular function and physiology.

[0017] FIG. 4 is a non-limiting, schematic representation of the bispecific fusion protein portion of KSI-501 (also denoted “KSI-101”) bound to VEGF and IL-6 molecules.

[0018] FIG.5 is a collection of non-limiting, schematic diagrams showing a fusion protein conjugate molecule as a combination of the fusion protein and a biopolymer.

[0019] FIG. 6 is a collection of images showing non-limiting examples of a KSI- 501 molecule, with or without VEGF bound thereto, detected by negative stain electron microscopy.

[0020] FIG. 7 is a diagram comparing various features of aflibercept, vamikibart, KSI-501 according to some non-limiting embodiments of the present disclosure.

[0021] FIG. 8 is a collection of fluorescence microscopy images and analyzed images thereof of retinal pigment epithelial (RPE) cells and primary human retinal microvascular endothelial cells (HRMVEC) stained for ZO1 (tight junction protein), VE- cadherin, and nuclei.

[0022] FIG. 9 is a collection of fluorescence microscopy images and analyzed images thereof of: RPE cells stained for ZO1 and nuclei; and HRMVECs stained for ZO1, actin, and nuclei.

[0023] FIG. 10 is a schematic diagram of a non-limiting study design for a Phase 1, multiple ascending dose study of KSI-501 in patients with diabetic macular edema (DME).

[0024] FIG. 11 depicts the demographics, general characteristics and baseline ocular characteristics of DME patients enrolled in the Phase 1 study, according to some non- limiting embodiments of the present disclosure.

[0025] FIGs.12A-12C are a collection of graphs depicting changes in visual acuity (mean change in BCVA in ETDRS letters) and retinal thickness (mean change in microns) over time in patients treated with KSI-501, according to some non-limiting embodiments of the present disclosure.

[0026] FIG. 13 depicts a non-limiting amino acid sequence of IL-6.

[0027] FIG. 14A shows OG1786.

[0028] FIG. 14B shows OG1801.

[0029] FIG. 14C shows OG1802.

[0030] FIG. 15 depicts some non-limiting embodiments of the heavy and light chain variable regions of an IL-6-Ab. Embodiments of CDRs are shown in boxed regions. These sequences can also be employed in an IL-6 Ab-VEGF Trap fusion construct.

[0031] FIG. 16 depicts some non-limiting embodiments of an IL-6-VEGF Trap fusion protein. The VEGF Trap domains are positioned either at the N-terminus immediately preceding the variable domain (left) or positioned between the Fab region and the hinge region of the antibody (right).

[0032] FIG. 17 depicts a non-limiting embodiment of a conjugate construct of VEGFR-Anti-IL6, which is a fusion of Anti-VEGF (VEGFR1 / 2) and Anti-IL-6 antibody conjugated with a phosphorylcholine-based polymer.

[0033] FIG. 18 depicts the sequences of VEGF_trap_variant_1, VEGF_trap_variant_2, and VEGF_trap_variant_3.

[0034] FIG. 19 illustrates non-limiting embodiments of Anti-IL-6 heavy chain variable region sequences. CDRs are underlined.

[0035] FIG. 20 illustrates various embodiments of VEGF trap sequences. Sections that vary between the sequences are in bold and underlined.

[0036] FIG.21 illustrates some non-limiting embodiments of linker (GS) sequence embodiments. It can be present as a double repeat Gly-Gly-Gly-Gly-Ser linker (GS).

[0037] FIG.22 illustrates some non-limiting embodiments of heavy chain sequence for Anti-IL-6 antibody molecules. CDRs are underlined.

[0038] FIG.23 illustrates some non-limiting embodiments of light chain sequences for Anti-IL-6 antibody molecules. CDRs are underlined.

[0039] FIG. 24 illustrates some non-limiting embodiments of heavy chain sequences for Anti-IL-6 antibody molecules.

[0040] FIGs. 25A-25B illustrate some non-limiting embodiments of combinations of CDRs of FIGs. 22-24.

[0041] FIG.26 illustrates some non-limiting embodiments of VEGFR-Fc sequence variants. Section that varies between the sequences are in bold and underlined.

[0042] FIG. 27 depicts the amino acid sequences of some non-limiting embodiments of the VEGFR-AntiIL6 fusion proteins. The CDRs (as defined by Kabat) are underlined. The greyed sections indicate the VEGFR constructs. The bolded text indicatesthe linker section. Mutations L234A, L235A, G237A and L443C (EU numbering) are double underlined. Each of these sections can be exchanged for other corresponding sections provided herein (e.g., alternative linkers or CDRs, etc.).

[0043] FIG.28 is a flow chart depicting a non-limiting embodiment of a method of treating a subject with wet age-related macular degeneration.

[0044] FIG.29 is a flow chart depicting a non-limiting embodiment of a method of treating a subject with wet age-related macular degeneration.

[0045] FIG.30 is a flow chart depicting a non-limiting embodiment of a method of treating a subject with wet age-related macular degeneration.

[0046] FIG.31 is a flow chart depicting a non-limiting embodiment of a method of treating a subject with wet age-related macular degeneration.

[0047] FIG.32 is a flow chart depicting a non-limiting embodiment of a method of treating a subject with eye disease.

[0048] FIG. 33 is a schematic diagram of a protocol of a dosing regimen for KSI- 501, according to some non-limiting embodiments of the present disclosure.

[0049] FIG. 34 is a non-limiting, schematic representation of KSI-101.

[0050] FIG. 35 is a schematic diagram of protocols of dosing regimens for KSI- 101 in diabetic macular edema (DME) and macular edema secondary to inflammation (MESI), according to some non-limiting embodiments of the present disclosure.

[0051] FIG. 36 is a schematic diagram of a protocol of a dosing regimen for KSI- 101 in macular edema secondary to inflammation (MESI), which may include uveitic macular edema (UME), post-surgical macular edema, and macular edema associated with inflammatory choroidal neovascularization, according to some non-limiting embodiments of the present disclosure.

[0052] FIGs. 37A-37C depict some embodiments of the treatment response of diabetic macular edema (DME) patients to KSI-101 at three different dose levels, 2.5 mg (FIG. 37A), 5 mg (FIG. 37B), and 10mg (FIG. 37C), depicting changes in visual acuity (mean change in BCVA in ETDRS letters) and central subfield thickness (CST) with the mean change in microns.

[0053] FIGs. 38A-38C depict some embodiments of the treatment response of macular edema secondary to inflammation (MESI) patients to KSI-101 at three different doselevels, 2.5 mg (FIG.38A), 5 mg (FIG.38B), and 10mg (FIG.38C), depicting changes in visual acuity (mean change in BCVA in ETDRS letters) and central subfield thickness with the mean change in microns. DETAILED DESCRIPTION

[0054] Current anti-VEGF monotherapies, such as aflibercept therapy, typically result in substantial patient-to-patient variability (FIG. 2). Individual patient variability underlies the mean best-corrected visual acuity (BCVA; FIG. 2, upper panel) and optical coherence tomography (OCT; FIG.2, lower panel) curves for patients treated with anti-VEGF monotherapy, suggesting a need for additional mechanisms of action to reduce variability.

[0055] IL-6 plays an important role in the pathophysiology of retinal vascular and hyperpermeability disorders (FIG. 3A). IL-6 is a pro-inflammatory cytokine and immune growth factor implicated in the pathophysiology of multiple retinal diseases and is associated with poor anti-VEGF treatment response. Specifically, IL-6 is known: to be associated with higher incidence of proliferative diabetic retinopathy (DR); to be associated with disease progression in age-related macular degeneration (AMD), DR and retinal vein occlusion (RVO); to be implicated in anti-VEGF treatment resistance; to upregulate VEGF; and to stimulate defective angiogenesis independent of VEGF.

[0056] KSI-501 includes a first-in-class bispecific protein (which is the same bioactive as KSI-101) that inhibits two powerful pathophysiologic mechanisms in retinal disease – IL-6 and VEGF (FIG. 3B and FIG 34). The bispecific protein of KSI-501 (which is the same bioactive as KSI-101) features a unique design that enables highly efficient binding to both IL-6 and VEGF / PlGF (FIG. 4). KSI-501ABC (“Antibody Biopolymer Conjugate”) shares the same bioactive as KSI-101, and in addition includes a biopolymer conjugated to the bispecific protein, has an increased molecular size, and in turn an extended ocular half-life (FIG. 5). KSI-501ABC includes a Trap - Antibody ABC that blocks VEGF / P1GF PlGF and IL-6.

[0057] Dual inhibition of IL-6 and VEGF can provide opportunities for clinical use of KSI-501 or KSI-101 across a range of retina disease indications. Preclinical and clinical data support the role of IL-6 as a key inflammatory modulator in retinal vascular diseases and seems to be related to the potential response to VEGF inhibition alone. In patients withmacular edema secondary to inflammation (prior or concurrent), the underlying inflammatory component of the pathophysiological process is not addressed by inhibiting VEGF alone.

[0058] Provided herein are methods of treating an eye disorder using a bispecific fusion protein conjugate (e.g., KSI-501) that binds to VEGF / PlGF and IL-6 and include a phosphorylcholine-containing polymer that extends the ocular half-life when the fusion protein conjugate is administered to a subject. The fusion protein conjugate may retain therapeutic efficacy after administration for a longer time period compared to an fusion protein without the phosphorylcholine-containing polymer. Thus, the methods of the present disclosure may provide for a course of treatment for an eye disorder that includes fewer doses (e.g., less frequent administration) of the fusion protein conjugate than conventional therapies for the eye disorder, to achieve a therapeutic effect. The present methods may encourage better patient compliance with the treatment course especially when the eye disorder treatment involves intravitreal administration of the therapeutic agent.

[0059] The present disclosure provides a method of treating an eye disorder, comprising: identifying a subject having an eye disorder; and administering at least one loading dose of a fusion protein conjugate to the subject, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 8 weeks after a final loading dose.

[0060] Also provided is a method of treating diabetic macular edema (DME), comprising: identifying a subject having DME; and administering no more than 5 loading doses of a fusion protein conjugate, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti- IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the no more than 5 loading doses of the fusion protein conjugate for at least 8 weeks after a final loading dose.

[0061] Provided herein are methods of treating eye disorders by administering a bispecific protein (e.g., KSI-101) that targets IL-6 mediated inflammation and edema, and VEGF-mediated vascular permeability. In some embodiments, the eye disorder includes an ocular inflammatory condition. In some embodiments, the method includes administering a fusion protein (e.g., an unconjugated fusion protein) to the subject. Non-limiting examples of eye disorders that can be treated by the present methods include macular edema secondary to inflammation (MESI), macular edema associated with inflammation, uveitic macular edema (UME), post-surgical macular edema, macular edema associated with inflammatory choroidal neovascularization, pediatric UME, non-infectious uveitis, macular edema, macular edema following branch retinal vein occlusion (BRVO) or central retinal vein occlusion (CRVO), wet age-related macular degeneration (AMD), diabetic retinopathy (DR), retinopathy of prematurity (ROP), idiopathic macular edema and / or diabetic macular edema (DME). TERMS

[0062] A “loading dose” has its ordinary and customary meaning as understood by a person of ordinary skill in the art, in view of the present disclosure. A loading dose may refer to an amount of a therapeutic agent administered to a subject, either before a therapeutic effect of the agent is observed in the subject, or before a desired level of therapeutic effect of the agent is achieved in the subject. A loading dose is typically administered at the beginning of a course of treatment with the therapy. In some embodiments, the loading dose is administered more frequently or at shorter intervals compared to later doses that are for maintenance of a therapeutic result. The time period during which a subject receives one or more loading doses may be referred to as a loading phase. In some embodiments, a subject is not monitored for disease progression or status (e.g., not assessed for visual acuity, retinal thickness, etc.) during the loading phase. In some embodiments, a therapeutic result (as disclosed herein) of the fusion protein conjugate therapy (e.g., KSI-501 therapy) has not reached a desired or threshold level during the loading phase. In some embodiments, a therapeutic result (as disclosed herein) of the anti-VEGF antibody conjugate therapy (e.g., KSI- 501 therapy) is observed at least at the end of the loading phase. The loading dose may be one of a series of loading doses administered to the subject, e.g., during the loading phase. A “final loading dose” may refer to the last loading dose in a series of loading doses administered tothe subject, at and / or after which a desired level of therapeutic effect of the agent is achieved. Thus, where the subject is given one loading dose, the final loading dose is the first loading dose. Where the subject is given two loading doses, the final loading dose is the second loading dose. Likewise, where the subject is given three loading doses, the final loading dose is the third loading dose, and so on. A dose of the therapeutic agent administered to a subject after the loading phase may be referred to as a subsequent dose, maintenance dose, an individualized dose, or a retreatment dose. The subsequent dose may be a maintenance dose or a retreatment dose or an individualized dose. “Maintenance dose,” “retreatment dose” and “individualized dose” are used herein interchangeably and customary meaning to one of ordinary skill in the art in view of the present disclosure. In some embodiments provided herein, the loading doses can be adequate without as frequent need for, or any need for, subsequent retreatment or maintenance doses. In some embodiments, a series of loading doses is administered to a subject at a higher frequency than a series of maintenance (or retreatment or individualized) doses administered to the subject. In some embodiments, the loading dose(s) given may be sufficient to keep disease activity under control in the subject, without requiring a subsequent dose (e.g., a maintenance or retreatment dose).

[0063] “Individualized dose” and “maintenance dose” are used interchangeably herein, and each has its ordinary and customary meaning as understood by a person of ordinary skill in the art, in view of the present disclosure. An individualized or maintenance dose may refer to one or more doses administered based on assessments of individual eye health and vision after the loading doses have been administered. In some embodiments, the timing of administering the individualized or maintenance doses for one subject is different from the timing of administering the individualized or maintenance doses for another subject, e.g., when the two subjects differ in their response to the loading doses and / or the individualized or maintenance doses. In some embodiments, criteria for determining whether individualized or maintenance doses should be administered includes but is not limited to a change in retinal thickness, the presence of intraretinal fluid (IRF), the presence of subretinal fluid (SRF), and / or the presence of new or clinically worsening macular hemorrhage due to disease activity (e.g., wAMD activity), as determined by the investigator.

[0064] The term "administered" as used herein has its plain and ordinary meaning as understood by one of ordinary skill in the art in light of the specification, and refers toproviding a dose of a fusion protein conjugate (e.g., KSI-501) to a subject such that the dose comes into contact with the region of the eye in need of therapeutic treatment. In some embodiments, the dose is provided by intravitreal injection.

[0065] A “dosing schedule” is a clinical regimen for administration of a therapeutic agent. Examples of dosing schedules include administration of a therapeutic agent every day, every week, every 4 weeks, or as needed due to monitoring of patient symptoms and symptomology. Dosing schedules can vary due to the severity of the underlying disorder or disease state.

[0066] A “dose frequency” is a clinical regimen for administration of a therapeutic agent. The dose frequency may be every day, every week, every 4 weeks, or as needed due to monitoring of patient symptoms and symptomology. Dose frequencies can vary due to the severity of the underlying disorder or disease state.

[0067] A “Ophthalmic Exam” is a comprehensive series of tests performed by a physician to assess vision and eye health of a patient. Commonly, Ophthalmic exams include patient histories, refraction tests to check vision, optical coherence tomography, and other assessments to aid a physician in rendering a professional opinion regarding a patient’s eye health. During an Ophthalmic Exam, a physician may use imaging techniques including but not limited to Fundus Photograph, SD-OCT, OCT-A, and Fluorescein Angiography (FA) to characterize anatomical features of the eye. A physician may further characterize whether anatomical structures within the eye are exudative or not, where structures that appear to “leak” bodily fluids would be “wet,” while non-exudative structures would be “dry”. On imaging an eye, the physician may also characterize finding excess intra and / or subretinal fluid and / or subretinal hyperreflective material (SHRM) affecting the central subfield of the eye as “wet”, while the lack of excess intra and / or subretinal fluid affecting the central subfield of the eye as “dry.”

[0068] The term "eye health" as used herein has its plain and ordinary meaning as understood by one of ordinary skill in the art in light of the specification and refers to the condition of a subject’s eye as determined using one or more criteria, e.g., measures of visual acuity and / or anatomical measures. In some embodiments, evaluating eye health includes evaluating a subject for ocular pain, discomfort or vision. In some embodiments, a change in eye health includes an increase or decrease in a subject’s ocular pain, discomfort or vision. Insome embodiments, eye health is determined by evaluating a subject’s eye. A person trained in evaluating eye health and / or administering treatment may perform the evaluation. In some embodiments, a subject’s eye health may worsen without the subject experiencing and / or reporting any symptoms, including pain and / or a loss of vision. In some embodiments, eye health is determined by examining a subject’s eye. In some embodiments, the criteria used for determining whether a subject has a change in eye health are subjective. In some embodiments, one or more measurements are made to determine whether a subject has had a change in eye health, and the value is compared to a standard and / or to a measurement made of the subject’s eye during a previous evaluation. In some embodiments, intraretinal fluid (IRF) is measured. In some embodiments, subretinal fluid (SRF) is measured. In some embodiments, retinal thickness is measured. In some embodiments, optical coherence tomography (OCT) is used to evaluate a subject’s eye health. In some embodiments, optical coherence tomography central subfield thickness (OCT CST) is used to evaluate a subject’s eye health. In some embodiments, the subject’s eye health is not evaluated before each loading dose is administered.

[0069] In some embodiments, a subject’s eye health is determined by evaluating an image and / or a scan of the subject’s eye. In some embodiments, the image or scan is compared to a standard image and / or a previous image and / or scan of the subject’s eye. In some embodiments, eye images are captured using fundus photography. In some embodiments, artificial intelligence is used to evaluate the images and / or scans of the subject’s eye. In some embodiments, a subject’s eye health is determined through an indirect test, for example by testing a subject’s blood, urine, spinal fluid, nasal fluid, or eye fluid including retinal fluid or tears. In some embodiments, a subject is determined to have a change in eye health if a change in macular hemorrhage due to wet AMD activity is observed. In some embodiments, the criteria for determining whether a subject has had a change in eye health includes asking the subject questions about their vision or how their eye feels. In some embodiment, the subject is asked about symptoms they may be having that are related to other regions of their head or neck, for example nasal passages, sinuses or ear canals. In some embodiments, a vision test is administered to the subject to determine whether the subject has had a change in eye health. In some embodiments, the eye test used involves a response to changes in light.

[0070] A “neovascular disease” is a disease state characterized by altered, dysregulated or unregulated angiogenesis. Examples of neovascular diseases includeneoplastic transformation (e.g. cancer) and ocular neovascular disorders including diabetic retinopathy (DR), age-related macular degeneration (AMD, e.g., wet AMD), and retinal vein occlusion (RVO). Examples of neovascular disorders or disease include neoplastic transformation (e.g. cancer) and ocular neovascular disorders including diabetic retinopathy and age-related macular degeneration. An “ocular neovascular” disorder is characterized by altered, dysregulated or unregulated angiogenesis in the eye of a patient. Such disorders include optic disc neovascularization, iris neovascularization, retinal neovascularization, choroidal neovascularization, corneal neovascularization, vitreal neovascularization, glaucoma, pannus, pterygium, macular edema, diabetic retinopathy, diabetic macular edema, vascular retinopathy, retinal degeneration, uveitis, inflammatory diseases of the retina, and proliferative vitreoretinopathy.

[0071] As used herein, “Q4W”, “Q8W” and the like refer to a dosing schedule, and have the ordinary and customary meaning to one of ordinary skill in the art in view of the present disclosure. The number may indicate the number of the unit of time specified by the subsequent letter. “W” indicates a unit of a week; “M” specifies an interval of a month. Thus, Q4W refers to a dosing interval of 4 weeks, which also includes a dosing interval of one month (QM); Q8W refers to a dosing interval of 8 weeks, which also includes a dosing interval of two months (Q2M); and so on. As used herein, specification of a dosing schedule does not necessarily imply a number of doses beyond two, unless indicated otherwise. In some embodiments, a dosing schedule refers to the dosing schedule for subsequent doses (including the interval between the last loading dose, and the first subsequent dose). A reference to a dosing schedule being “longer” or “shorter” (e.g., “Q12W or longer”) refers to the time interval between doses being longer than that specified (e.g., a dosing interval of 12 weeks or longer).

[0072] As used herein, “non-native cysteine” denotes a cysteine that is not present in the corresponding position in a naturally occurring counterpart of the protein, or portion of the protein. The cysteine can be introduced into the protein via recombinant DNA technology. In the context of an antibody, including an antibody portion of a fusion protein, a Fc region can include a cysteine that is not present in the corresponding position of a naturally occurring counterpart Fc region, e.g., in an IgG1 Fc region.

[0073] As used herein, unless designated otherwise, the term “IL-6” or “IL6” refers to human IL-6. In some embodiments, IL-6 has the amino acid sequence of SEQ ID NO: 1,shown in FIG. 13. In some embodiments, other forms of IL-6 are contemplated, and will be designated by specific reference to the other organisms, e.g., murine, canine, feline, equine, and bovine. One exemplary human IL-6 is found as UniProt Accession Number P05231.

[0074] The following terms, unless otherwise indicated, shall be understood to have the following meanings: the term “isolated molecule” as referring to a molecule (where the molecule is, for example, a polypeptide, a polynucleotide, or an antibody) that by virtue of its origin or source of derivation (1) is not associated with naturally associated components that accompany it in its native state, (2) is substantially free of other molecules from the same source, e.g., species, cell from which it is expressed, library, etc., (3) is expressed by a cell from a different species, or (4) does not occur in nature. Thus, a molecule that is chemically synthesized, or expressed in a cellular system different from the system from which it naturally originates, will be “isolated” from its naturally associated components. A molecule also may be rendered substantially free of naturally associated components by isolation, using purification techniques well known in the art. Molecule purity or homogeneity may be assayed by a number of means well known in the art. For example, the purity of a polypeptide sample may be assayed using polyacrylamide gel electrophoresis and staining of the gel to visualize the polypeptide using techniques well known in the art. For certain purposes, higher resolution may be provided by using HPLC or other means well known in the art for purification.

[0075] The term “peptide,” “polypeptide,” or “protein,” as used interchangeably herein, generally refers to a polymer of at least two amino acid residues joined by peptide bond(s). This term does not connote a specific length of polymer, nor is it intended to imply or distinguish whether the peptide is produced using recombinant techniques, chemical or enzymatic synthesis, or is naturally occurring. The terms apply to naturally occurring amino acid polymers as well as amino acid polymers comprising at least one modified amino acid. In some cases, the polymer can be interrupted by non-amino acids. The terms include amino acid chains of any length, including full length proteins, and proteins with or without secondary and / or tertiary structure (e.g., domains). The terms also encompass an amino acid polymer that has been modified, for example, by disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, oxidation, and any other manipulation such as conjugation with a labeling component. The terms “amino acid” and “amino acids,” as used herein, generally refer to natural and non-natural amino acids, including, but not limited to, modified aminoacids and amino acid analogues. Modified amino acids can include natural amino acids and non-natural amino acids, which have been chemically modified to include a group or a chemical moiety not naturally present on the amino acid. Amino acid analogues can refer to amino acid derivatives. The term “amino acid” includes both D-amino acids and L-amino acids.

[0076] Anti-IL-6 antibodies, fusion proteins, or other biologics described herein are typically provided in isolated form. This means that an antibody is typically at least 50% w / w pure of interfering proteins and other contaminants arising from its production or purification but does not exclude the possibility that the antibody or fusion protein is combined with a pharmaceutically acceptable excipient intended to facilitate its use. Sometimes antibodies are at least 60, 70, 80, 90, 95 or 99% w / w pure of interfering proteins and contaminants from production or purification. Often a fusion protein (or fusion protein conjugate) is the predominant macromolecular species remaining after its purification.

[0077] An “antibody” is an immunoglobulin molecule capable of specific binding to a target, such as a carbohydrate, polynucleotide, lipid, polypeptide, etc., through at least one antigen recognition site, located in the variable region of the immunoglobulin molecule. As used herein, the term encompasses not only intact polyclonal or monoclonal antibodies, but also, unless otherwise specified, any antigen binding portion thereof that competes with the intact antibody for specific binding, fusion proteins comprising an antigen binding portion, and any other modified configuration of the immunoglobulin molecule that comprises an antigen recognition site. Antigen binding portions include, for example, Fab, Fab’, F(ab’)2, Fd, Fv, domain antibodies (dAbs, e.g., shark and camelid antibodies), fragments including complementarity determining regions (CDRs), single chain variable fragment antibodies (scFv), maxibodies, minibodies, intrabodies, diabodies, triabodies, tetrabodies, v-NAR and bis-scFv, and polypeptides that contain at least a portion of an immunoglobulin that is sufficient to confer specific antigen binding to the polypeptide. An antibody includes an antibody of any class, such as IgG, IgA, or IgM (or sub-class thereof), and the antibody need not be of any particular class. Depending on the antibody amino acid sequence of the constant region of its heavy chains, immunoglobulins can be assigned to different classes. There are five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, and several of these may be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1and IgA2.The heavy-chain constant regions that correspond to the different classes of immunoglobulins are called alpha, delta, epsilon, gamma, and mu, respectively. The subunit structures and three- dimensional configurations of different classes of immunoglobulins are well known.

[0078] The term “polynucleotide,” “oligonucleotide,” or “nucleic acid,” as used interchangeably herein, generally refers to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof, either in single-, double-, or multi- stranded form. A polynucleotide can be exogenous or endogenous to a cell. A polynucleotide can exist in a cell-free environment. A polynucleotide can be a gene or fragment thereof. A polynucleotide can be DNA. A polynucleotide can be RNA. A polynucleotide can have any three dimensional structure, and can perform any function, known or unknown. A polynucleotide can comprise one or more analogs (e.g., altered backbone, sugar, or nucleobase).

[0079] A “variable region” of an antibody refers to the variable region of the antibody light chain or the variable region of the antibody heavy chain, either alone or in combination. As known in the art, the variable regions of the heavy and light chains each consist of four framework regions (FRs) connected by three complementarity determining regions (CDRs) also known as hypervariable regions, and contribute to the formation of the antigen binding site of antibodies. If variants of a subject variable region are desired, particularly with substitution in amino acid residues outside of a CDR region (i.e., in the framework region), appropriate amino acid substitution, preferably, conservative amino acid substitution, can be identified by comparing the subject variable region to the variable regions of other antibodies which contain CDR1 and CDR2 sequences in the same canonical class as the subject variable region (Chothia and Lesk, J Mol Biol 196(4): 901-917, 1987).

[0080] In certain embodiments, definitive delineation of a CDR and identification of residues comprising the binding site of an antibody is accomplished by solving the structure of the antibody and / or solving the structure of the antibody-ligand complex. In certain embodiments, that can be accomplished by any of a variety of techniques known to those skilled in the art, such as X-ray crystallography. In certain embodiments, various methods of analysis can be employed to identify or approximate the CDR regions. In certain embodiments, various methods of analysis can be employed to identify or approximate the CDR regions. Examples of such methods include, but are not limited to, the Kabat definition, the Chothiadefinition, the IMGT approach (Lefranc et al., 2003) Dev Comp Immunol. 27:55-77), computational programs such as Paratome (Kunik et al., 2012, Nucl Acids Res. W521-4), the AbM definition, and the conformational definition.

[0081] The Kabat definition is a standard for numbering the residues in an antibody and is typically used to identify CDR regions. See, e.g., Johnson & Wu, 2000, Nucleic Acids Res., 28: 214-8. The Chothia definition is similar to the Kabat definition, but the Chothia definition takes into account positions of certain structural loop regions. See, e.g., Chothia et al., 1986, J. Mol. Biol., 196: 901-17; Chothia et al., 1989, Nature, 342: 877-83. The AbM definition uses an integrated suite of computer programs produced by Oxford Molecular Group that model antibody structure. See, e.g., Martin et al., 1989, Proc Natl Acad Sci (USA), 86:9268-9272; “AbM™, A Computer Program for Modeling Variable Regions of Antibodies,” Oxford, UK; Oxford Molecular, Ltd. The AbM definition models the tertiary structure of an antibody from primary sequence using a combination of knowledge databases and ab initio methods, such as those described by Samudrala et al., 1999, “Ab Initio Protein Structure Prediction Using a Combined Hierarchical Approach,” in PROTEINS, Structure, Function and Genetics Suppl., 3:194-198. The contact definition is based on an analysis of the available complex crystal structures. See, e.g., MacCallum et al., 1996, J. Mol. Biol., 5:732-45. In another approach, referred to herein as the “conformational definition” of CDRs, the positions of the CDRs may be identified as the residues that make enthalpic contributions to antigen binding. See, e.g., Makabe et al., 2008, Journal of Biological Chemistry, 283:1156-1166. Still other CDR boundary definitions may not strictly follow one of the above approaches, but will nonetheless overlap with at least a portion of the Kabat CDRs, although they may be shortened or lengthened in light of prediction or experimental findings that particular residues or groups of residues do not significantly impact antigen binding. As used herein, a CDR may refer to CDRs defined by any approach known in the art, including combinations of approaches. The methods used herein may utilize CDRs defined according to any of these approaches. For any given embodiment containing more than one CDR, the CDRs may be defined in accordance with any of Kabat, Chothia, extended, IMGT, Paratome, AbM, and / or conformational definitions, or a combination of any of the foregoing.

[0082] As known in the art, a “constant region” of an antibody refers to the constant region of the antibody light chain or the constant region of the antibody heavy chain, either alone or in combination.

[0083] As used herein, “monoclonal antibody” refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally-occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. 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 may be made by the hybridoma method first described by Kohler and Milstein, 1975, Nature 256:495, or may be made by recombinant DNA methods such as described in U.S. Pat. No.4,816,567. The monoclonal antibodies may also be isolated from phage libraries generated using the techniques described in McCafferty et al., 1990, Nature 348:552-554, for example. As used herein, “humanized” antibody refers to forms of non-human (e.g. murine) antibodies that are chimeric immunoglobulins, immunoglobulin chains, or fragments thereof (such as Fv, Fab, Fab’, F(ab’)2 or other antigen-binding subsequences of antibodies) that contain minimal sequence derived from non-human immunoglobulin. Preferably, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a CDR of the recipient are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat, or rabbit having the desired specificity, affinity, and capacity. The humanized antibody may comprise residues that are found neither in the recipient antibody nor in the imported CDR or framework sequences, but are included to further refine and optimize antibody performance.

[0084] A “human antibody” is one which possesses an amino acid sequence which corresponds to that of an antibody produced by a human and / or has been made using any of the techniques for making human antibodies as disclosed herein. This definition of a human antibody specifically excludes a humanized antibody comprising non-human antigen binding residues.

[0085] The term “chimeric antibody” is intended to refer to antibodies in which the variable region sequences are derived from one species and the constant region sequences are derived from another species, such as an antibody in which the variable region sequences are derived from a mouse antibody and the constant region sequences are derived from a human antibody. The term “epitope” refers to that portion of a molecule capable of being recognized by and bound by an antibody at one or more of the antibody’s antigen-binding regions. Epitopes often consist of a surface grouping of molecules such as amino acids or sugar side chains and have specific three-dimensional structural characteristics as well as specific charge characteristics. In some embodiments, the epitope can be a protein epitope. Protein epitopes can be linear or conformational. In a linear epitope, all of the points of interaction between the protein and the interacting molecule (such as an antibody) occur linearly along the primary amino acid sequence of the protein. A “nonlinear epitope” or “conformational epitope” comprises noncontiguous polypeptides (or amino acids) within the antigenic protein to which an antibody specific to the epitope binds. The term “antigenic epitope” as used herein, is defined as a portion of an antigen to which an antibody can specifically bind as determined by any method well known in the art, for example, by conventional immunoassays. Once a desired epitope on an antigen is determined, it is possible to generate antibodies to that epitope, e.g., using the techniques described in the present specification. Alternatively, during the discovery process, the generation and characterization of antibodies may elucidate information about desirable epitopes. From this information, it is then possible to competitively screen antibodies for binding to the same epitope. An approach to achieve this is to conduct competition and cross-competition studies to find antibodies that compete or cross-compete with one another for binding to IL-6, e.g., the antibodies compete for binding to the antigen.

[0086] The term “compete,” as used herein with regard to an antibody, means that a first antibody, or an antigen-binding portion thereof, binds to an epitope in a manner sufficiently similar to the binding of a second antibody, or an antigen-binding portion thereof, such that the result of binding of the first antibody with its cognate epitope is detectably decreased in the presence of the second antibody compared to the binding of the first antibody in the absence of the second antibody. The alternative, where the binding of the second antibody to its epitope is also detectably decreased in the presence of the first antibody, can, but need not be the case. That is, a first antibody can inhibit the binding of a second antibodyto its epitope without that second antibody inhibiting the binding of the first antibody to its respective epitope. However, where each antibody detectably inhibits the binding of the other antibody with its cognate epitope or ligand, whether to the same, greater, or lesser extent, the antibodies are said to “cross-compete” with each other for binding of their respective epitope(s). Both competing and cross-competing antibodies are provided herein. Regardless of the mechanism by which such competition or cross-competition occurs (e.g., steric hindrance, conformational change, or binding to a common epitope, or portion thereof), the skilled artisan would appreciate, based upon the teachings provided herein, that such competing and / or cross- competing antibodies are encompassed and can be useful for the methods disclosed herein.

[0087] As used herein, an antibody “interacts with” IL-6 when the equilibrium dissociation constant is equal to or less than 20 nM, preferably less than about 6 nM, more preferably less than about 1 nM, most preferably less than about 0.75 nM. In some embodiments, the affinity of the antibody is between 400 and 800 pM, e.g., 450-700, or 500- 600 pM.

[0088] An IL-6 antagonist antibody encompasses antibodies that block, antagonize, suppress or reduce (to any degree including significantly) a IL-6 biological activity such as binding to IL-6R, IL-6 / IL-6R complex binding to gp130, phosphorylation and activation of Stat3, cell proliferation, and stimulation of IL-6 mediated inflammatory or pro-angiogenic pathways. For purpose of the present disclosure, it will be explicitly understood that the term “IL-6 antagonist antibody” encompasses all the previously identified terms, titles, and functional states and characteristics whereby the IL-6 itself, an IL-6 biological activity, or the consequences of the biological activity, are substantially nullified, decreased, or neutralized in any meaningful degree. In some embodiments, an IL-6 antagonist antibody binds IL-6. Examples of IL-6 antagonist antibodies are provided herein.

[0089] An antibody that “preferentially binds” or “specifically binds” (used interchangeably herein) to an epitope is a term well understood in the art, and methods to determine such specific or preferential binding are also well known in the art. A molecule is said to exhibit “specific binding” or “preferential binding” if it reacts or associates more frequently, and / or more rapidly, and / or with greater duration and / or with greater affinity with a particular cell or substance than it does with alternative cells or substances. An antibody “specifically binds” or “preferentially binds” to a target if it binds with greater affinity, and / oravidity, and / or more readily, and / or with greater duration than it binds to other substances. For example, an antibody that specifically or preferentially binds to an IL-6 epitope is an antibody that binds this epitope with greater affinity, and / or avidity, and / or more readily, and / or with greater duration than it binds to other IL-6 epitopes or non-IL-6 epitopes. It is also understood by reading this definition that, for example, an antibody (or moiety or epitope) that specifically or preferentially binds to a first target may or may not specifically or preferentially bind to a second target. As such, “specific binding” or “preferential binding” does not necessarily require (although it can include) exclusive binding. Generally, but not necessarily, reference to binding means preferential binding.

[0090] As used herein, “substantially pure” refers to material which is at least 50% pure (i.e., free from contaminants), more preferably, at least 90% pure, more preferably, at least 95% pure, yet more preferably, at least 98% pure, and most preferably, at least 99% pure.

[0091] A “host cell” includes an individual cell or cell culture that can be or has been a recipient for vector(s) for incorporation of polynucleotide inserts. Host cells include progeny of a single host cell, and the progeny may not necessarily be completely identical (in morphology or in genomic DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation. A host cell includes cells transfected in vivo with a polynucleotide(s) provided herein.

[0092] As known in the art, the term “Fc region” is used to define a C-terminal region of an immunoglobulin heavy chain. The “Fc region” may be a native sequence Fc region or a variant Fc region. Although the boundaries of the Fc region of an immunoglobulin heavy chain might vary, the human IgG heavy chain Fc region is usually defined to stretch from an amino acid residue at position Cys226, or from Pro230, to the carboxyl-terminus thereof. The numbering of the residues in the Fc region is that of the EU index as in Kabat. Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991. The Fc region of an immunoglobulin generally comprises two constant domains, CH2 and CH3. As is known in the art, an Fc region can be present in dimer or monomeric form.

[0093] As used in the art, “Fc receptor” and “FcR” describe a receptor that binds to the Fc region of an antibody. The preferred FcR is a native sequence human FcR. Moreover, a preferred FcR is one which binds an IgG antibody (a gamma receptor) and includes receptorsof the FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternatively spliced forms of these receptors. FcγRII receptors include FcγRIIA (an “activating receptor”) and FcγRIIB (an “inhibiting receptor”), which have similar amino acid sequences that differ primarily in the cytoplasmic domains thereof. FcRs are reviewed in Ravetch and Kinet, 1991, Ann. Rev. Immunol., 9:457-92; Capel et al., 1994, Immunomethods, 4:25-34; and de Haas et al., 1995, J. Lab. Clin. Med., 126:330-41. “FcR” also includes the neonatal receptor, FcRn, which is responsible for the transfer of maternal IgGs to the fetus (Guyer et al., 1976, J. Immunol., 117:587; and Kim et al., 1994, J. Immunol., 24:249).

[0094] A “functional Fc region” possesses at least one effector function of a native sequence Fc region. Exemplary “effector functions” include C1q binding; complement dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity; phagocytosis; down-regulation of cell surface receptors (e.g. B cell receptor), etc. Such effector functions generally require the Fc region to be combined with a binding domain (e.g. an antibody variable domain) and can be assessed using various assays known in the art for evaluating such antibody effector functions.

[0095] “VEGF” or “vascular endothelial growth factor,” and “PlGF” have their ordinary and customary meaning as understood by one of ordinary skill in the art in view of the present disclosure, and denotes a vascular endothelial growth factor that affects angiogenesis or an angiogenic process. In particular, the term VEGF denotes any member of the class of growth factors that (i) bind to a VEGF receptor such as VEGFR-1 (Flt-1), VEGFR- 2 (KDR / Flk-1), or VEGFR-3 (FLT-4); (ii) activates a tyrosine kinase activity associated with the VEGF receptor; and (iii) thereby affects angiogenesis or an angiogenic process. The VEGF family of factors is made up of five related glycoproteins: VEGF-A (also known as VPE), -B, -C, -D and PlGF (placental growth factor). Any reference to VEGF herein can include one or more of the VEGF family of factors.

[0096] A “native sequence Fc region” comprises an amino acid sequence identical to the amino acid sequence of an Fc region found in nature. A “variant Fc region” comprises an amino acid sequence which differs from that of a native sequence Fc region by virtue of at least one amino acid modification, yet retains at least one effector function of the native sequence Fc region. Preferably, the variant Fc region has at least one amino acid substitution compared to a native sequence Fc region or to the Fc region of a parent polypeptide, e.g. fromabout one to about ten amino acid substitutions, and preferably, from about one to about five amino acid substitutions in a native sequence Fc region or in the Fc region of the parent polypeptide. The variant Fc region herein will preferably possess at least about 80% sequence identity with a native sequence Fc region and / or with an Fc region of a parent polypeptide, and most preferably, at least about 90% sequence identity therewith, more preferably, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99% sequence identity therewith.

[0097] As used herein, “treatment” is an approach for obtaining beneficial or desired clinical results.

[0098] As used herein, “IL-6 and / or VEGF related disorders” include, for example, ocular disorders and systemic disorders. The ocular disorders include, without limitation, ophthalmic inflammatory diseases, non-proliferative diabetic retinopathy, proliferative diabetic retinopathy, diabetic macular edema, prevention of diabetic macular edema, prevention of proliferative diabetic retinopathy, wet age-related macular degeneration, prevention of wet age-related macular degeneration, dry age-related macular degeneration, venous, arterial or other blockage of the ocular and or retinal blood vessels with or without retinal edema, anterior and posterior uveitis, uveitic macular edema, and intraocular tumors. In some embodiments, the ocular disorders include macular edema secondary to inflammation (MESI). In some embodiments, MESI includes one or more of uveitic macular edema (UME), post-surgical macular edema, and macular edema associated with inflammatory choroidal neovascularization. IL-6 related disorders also include disorders where there is an elevated level of IL-6 activity due to IL-6 interacting with IL-6R or soluble IL-6R (sIL-6R). In some embodiments, any one or more of the fusion proteins provided herein and / or any one or more of the conjugates provided herein can be used for treatment or prevention of any one or more of the IL-6 and / or VEGF related disorders. In some embodiments, the disorders include systemic diseases that affect the eye such as Grave’s disease or neuromyelitis optica, or systemic diseases that do not affect the eye such as multiple sclerosis, rheumatoid arthritis. In some embodiments, the disorders include cytokine release syndrome following CAR-T or similar immune-oncology therapeutics.

[0099] Additionally, anti-IL6 molecules abrogate the induction of IL-6 expression observed following treatment with anti-PD-1 / PD-L1 molecules (Tsukamoto et al, Cancer Res;201878(17); 5011–22). It has also been shown that VEGF signaling blockade can improve anti-PD-L1 treatment (Allen et al, Sci Transl Med 2017 April 12: 9(385)). Thus, these dual inhibitors can be used in combination with PD-1 / PDL-1 modulators and / or other immune checkpoint inhibitors, to synergistically treat cancer. Other disorders can include cerebral edema in glioblastoma where anti-IL6 therapy may show additional benefits to anti-VEGF treatments. Other disorders include those with solid tumors.

[0100] As used herein, “ameliorating” means a lessening or improvement of one or more symptoms as compared to not administering an IL-6 antibody, IL-6 antibody-VEGF Trap fusion, conjugate of IL-6 antibody, and / or conjugate of IL-6 antibody-VEGF Trap fusion. “Ameliorating” also includes shortening or reduction in duration of a symptom.

[0101] As used herein, “VEGF Trap” or similar term denotes the VEGF binding domains (VEGFR1 domain 2, VEGFR2 domain 3). This fragment allows for the protein to work as a VEGF trap, preventing VEGF from binding to cellularly expressed VEGF receptors. An example of this sequence can be found in Table 10. In some embodiments, the VEGF Trap only includes VEGFR1 domain 2, VEGFR2 domain 3. Various embodiments of Trap proteins are known in the art and can be found, for example in U.S. Pub. No.20150376271, the entirety of which, with respect to various VEGF Trap embodiments (which are VEGFR proteins or fragments thereof) and fusions thereof, is incorporated herein by reference. In some embodiments, the term “VEGF Trap” or similar term refers to a full length extracellular region or any portion thereof, or combination of portions from different VEGF receptors that can antagonize signaling between at least one VEGF and VEGFR.

[0102] As used herein, “IL-6 antibody-VEGF Trap fusion”, “IL-6 antibody-VEGF Trap”, “Ab IL-6-VEGF Trap”, “AntiIL-6-VEGF Trap”, “VEGFR-AntiIL6”, “VEGFR-AntiIL- 6”, “VEGF Trap-anti-IL6 Antibody Fusion (TAF)”, “VEGF Trap-IL6”, “VEGFR IL-6”, “IL6- VEGFR” or similar term or inverse terms (e.g. “VEGF Trap-IL-6 Ab,” “VEGF Trap-IL-6 antibody fusion,” etc.) denote the fusion between the IL-6 antibody and the VEGF Trap, and may be denoted as a “fusion protein” or “fusion construct.” Embodiments are depicted in FIG. 16. When used generically, the order of the two terms can be swapped. When used specifically, the order of the two terms denotes the relative position of the components in the construct. the term “Ab-Trap”, “IL-6 Ab-VEGF Trap” “Ab IL-6 VEGF Trap” or “Ab IL-6- Trap” or “antiIL-6 VEGF Trap”, “Trap-Ab”, AntiIL-6-VEGFR, AntiIL6-VEGFR or othersimilar term or inverse terms (e.g. “VEGF Trap-IL-6 Ab,” “VEGF Trap-IL-6 antibody fusion,” etc.) denotes the arrangement of the Ab fused to the relevant domains of a VEGF binding protein so as to provide a VEGF trap. As noted above, this section of the VEGF binding protein is one that prevents VEGF from binding to VEGF receptors. As described herein, the arrangement (ordering) of the Trap and antibody sections can be varied. Thus, unless denoted otherwise explicitly or by context, the phrases used herein regarding Ab-Trap (or Il-6 / VEGF Trap, etc.) fusions, denote all disclosed embodiments for the positioning of the antibody and the Trap. Thus, unless explained otherwise, the phrase Ab-Trap (or Il-6 / VEGF Trap, etc.), denotes the left embodiment in FIG. 16, and the right embodiment in FIG. 16, and both embodiments in FIG. 16. Thus, the general language is denoted as disclosing all three options for convenience. If the orientation is specifically denoted, it can be denoted, for example, by stating that the “arrangement” can be one of: Trap-Ab, Trap IL-6 Ab, VEGF Trap Ab IL-6, VEGF Trap Ab IL6. Similarly, it will be appreciated that the context of some of the present Examples specific orientations or arrangements of the molecules, which are denoted by the context of the Example. Both arrangements (in the alternative and combined) are explicitly contemplated for all discussions of fusion proteins provided herein. In addition, due to the ordering, it is appreciated that the phrase IL-6 Ab, when used in the context of the fusion protein, includes both the option where the antibody is contiguous, FIG.16, left-hand side, and where the TRAP is positioned “within” the Ab (FIG. 16 right-hand side). Again, the term “Ab” or “antibody”, when used in the fusion protein context (or other similar term), encompasses all three options (left-hand side of FIG. 16, right-hand side of FIG. 16, and both options), unless otherwise noted. In some embodiments, the VEGF Trap is fused to IL-6 in one of the following manners: to an N-terminal end of a heavy chain comprising IL-6 VH; or between a hinge region and after a CH1 domain of a heavy chain comprising IL-6 VH. There is no difference between the designations of Ab, antibody, “anti’” or other similar term when used in a name to designate and antibody or fragment thereof. There is no difference between the designations of “Il-6” or “IL6” or “IL-6”. As used herein, when referencing a fusion construct with IL-6, the terms “VEGF”, “VEGFR”, “VEGF Trap”, “VEGFR Trap” are used interchangeably. The terms can have different meanings when used separately from the IL-6 fusion arrangement, which will depend upon the context of the term in question.

[0103] As used herein, the term “biopolymer” denotes that a polymer has been linked to the protein of interest. The term can also be described as the “conjugated” form of the protein. This can be done for all of the proteins described herein. Thus, IL-6 Ab biopolymers and IL-6 antibody-VEGF Trap biopolymers are contemplated for all such IL-6 Ab and IL-6 antibody-VEGF Trap provided herein. In addition, VEGF Trap biopolymers are also provided.

[0104] As used herein, “antagonistic antibody” denotes an antibody that blocks one or more function or activity of the molecule that the antibody binds to.

[0105] As used herein, an “effective dosage” or “effective amount” of drug, compound, or pharmaceutical composition is an amount sufficient to effect any one or more beneficial or desired results. In more specific aspects, an effective amount prevents, alleviates or ameliorates symptoms of disease, and / or prolongs the survival of the subject being treated. For prophylactic use, beneficial or desired results include eliminating or reducing the risk, lessening the severity, or delaying the outset of the disease, including biochemical, histological and / or behavioral symptoms of the disease, its complications and intermediate pathological phenotypes presenting during development of the disease. For therapeutic use, beneficial or desired results include clinical results such as reducing one or more symptoms of a disease such as, for example, diabetic macular edema (DME), decreasing the dose of other medications required to treat the disease, enhancing the effect of another medication, and / or delaying the progression of DME in patients. An effective dosage can be administered in one or more administrations. For purposes of this invention, an effective dosage of drug, compound, or pharmaceutical composition is an amount sufficient to accomplish prophylactic or therapeutic treatment either directly or indirectly. As is understood in the clinical context, an effective dosage of a drug, compound, or pharmaceutical composition may or may not be achieved in conjunction with another drug, compound, or pharmaceutical composition. Thus, an “effective dosage” may be considered in the context of administering one or more therapeutic agents, and a single agent may be considered to be given in an effective amount if, in conjunction with one or more other agents, a desirable result may be or is achieved.

[0106] Anti-IL-6 antibodies are administered in an effective regimen meaning a dosage, route of administration and frequency of administration that delays the onset, reduces the severity, inhibits further deterioration, and / or ameliorates at least one sign or symptom ofa disorder. If a patient is already suffering from a disorder, the regimen can be referred to as a therapeutically effective regimen. If the patient is at elevated risk of the disorder relative to the general population but is not yet experiencing symptoms, the regimen can be referred to as a prophylactically effective regimen. In some instances, therapeutic or prophylactic efficacy can be observed in an individual patient relative to historical controls or past experience in the same patient. In other instances, therapeutic or prophylactic efficacy can be demonstrated in a preclinical or clinical trial in a population of treated patients relative to a control population of untreated patients.

[0107] Anti-IL-6-VEGF Traps are administered in an effective regimen meaning a dosage, route of administration and frequency of administration that delays the onset, reduces the severity, inhibits further deterioration, and / or ameliorates at least one sign or symptom of a disorder. If a patient is already suffering from a disorder, the regimen can be referred to as a therapeutically effective regimen. If the patient is at elevated risk of the disorder relative to the general population but is not yet experiencing symptoms, the regimen can be referred to as a prophylactically effective regimen. In some instances, therapeutic or prophylactic efficacy can be observed in an individual patient relative to historical controls or past experience in the same patient. In other instances, therapeutic or prophylactic efficacy can be demonstrated in a preclinical or clinical trial in a population of treated patients relative to a control population of untreated patients.

[0108] The “biological half-life” of a substance is a pharmacokinetic parameter which specifies the time required for one half of the substance to be removed from an organism following introduction of the substance into the organism.

[0109] The term “preventing” or “prevent” refers to (a) keeping a disorder from occurring (b) delaying the onset of a disorder or onset of symptoms of a disorder, or (c) slowing the progression of an existing condition. Unless denoted otherwise, “preventing” does not require the absolute prohibition of the event from occurring.

[0110] An “individual” or a “subject” is a mammal or bird, more preferably, a human. Mammals also include, but are not limited to, farm animals (e.g., cows, pigs, horses, chickens, etc.), sport animals, pets, primates, horses, dogs, cats, mice and rats.

[0111] As used herein, “vector” means a construct, which is capable of delivering, and, preferably, expressing, one or more gene(s) or sequence(s) of interest in a host cell.Examples of vectors include, but are not limited to, viral vectors, naked DNA or RNA expression vectors, plasmid, cosmid or phage vectors, DNA or RNA expression vectors associated with cationic condensing agents, DNA or RNA expression vectors encapsulated in liposomes, and certain eukaryotic cells, such as producer cells.

[0112] As used herein, “expression control sequence” means a nucleic acid sequence that directs transcription of a nucleic acid. An expression control sequence can be a promoter, such as a constitutive or an inducible promoter, or an enhancer. The expression control sequence is operably linked to the nucleic acid sequence to be transcribed.

[0113] As used herein, “pharmaceutically acceptable carrier” or “pharmaceutical acceptable excipient” includes any material which, when combined with an active ingredient, allows the ingredient to retain biological activity and is non-reactive with the subject’s immune system. Examples include, but are not limited to, any of the standard pharmaceutical carriers such as a phosphate buffered saline solution, water, emulsions such as oil / water emulsion, various types of wetting agents, detergents such as polysorbate 20 to prevent aggregation, and sugars such as sucrose as cryoprotectant. Preferred diluents for aerosol or parenteral administration are phosphate buffered saline (PBS) or normal (0.9%) saline. Compositions comprising such carriers are formulated by well-known conventional methods (see, for example, Remington’s Pharmaceutical Sciences, 18th edition, A. Gennaro, ed., Mack Publishing Co., Easton, PA, 1990; and Remington, The Science and Practice of Pharmacy 20th Ed. Mack Publishing, 2000).

[0114] The term “kon”, as used herein, refers to the rate constant for association of an antibody (or bioconjugate) to an antigen. Specifically, the rate constants (kon and koff) and equilibrium dissociation constants are measured using full-length antibodies and / or Fab antibody fragments (i.e. univalent) and IL-6.

[0115] The term “koff “, as used herein, refers to the rate constant for dissociation of an antibody (or bioconjugate) from the antibody / antigen complex.

[0116] The term “KD”, as used herein, refers to the equilibrium dissociation constant of an antibody-antigen (or bioconjugate-antigen) interaction.

[0117] Reference to “about” a value or parameter herein includes (and describes) embodiments that are directed to that value or parameter per se. For example, descriptionreferring to “about X” includes description of “X.” Numeric ranges are inclusive of the numbers defining the range.

[0118] The term “patient” includes human and other subjects (including mammals) that receive either prophylactic or therapeutic treatment.

[0119] A “neovascular disorder” or “vascular disorder” has its ordinary and customary meaning as understood by one of ordinary skill in the art, in view of the present disclosure, and denotes a disorder or disease state characterized by altered, dysregulated or unregulated angiogenesis. Examples of vascular / neovascular disorders include neoplastic transformation (e.g. cancer) and ocular vascular / neovascular disorders including diabetic retinopathy, age-related macular degeneration, and retinal vein occlusion.

[0120] An “ocular vascular / neovascular” disorder is a disorder characterized by altered, dysregulated or unregulated angiogenesis in the eye of a patient. Such disorders include retinal vein occlusion, optic disc neovascularization, iris neovascularization, retinal neovascularization, choroidal neovascularization, corneal neovascularization, vitreal neovascularization, glaucoma, pannus, pterygium, macular edema, diabetic retinopathy, diabetic macular edema, vascular retinopathy, retinal degeneration, uveitis, inflammatory diseases of the retina, and proliferative vitreoretinopathy.

[0121] Percentage sequence identities are determined with antibody sequences maximally aligned by the Kabat numbering convention for a variable region or EU numbering for a constant region. After alignment, if a subject antibody region (e.g., the entire mature variable region of a heavy or light chain) is being compared with the same region of a reference antibody, the percentage sequence identity between the subject and reference antibody regions is the number of positions occupied by the same amino acid in both the subject and reference antibody region divided by the total number of aligned positions of the two regions, with gaps not counted, multiplied by 100 to convert to percentage. Sequence identities of other sequences can be determined by aligning sequences using algorithms, such as BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package Release 7.0, Genetics Computer Group, 575 Science Dr., Madison, WI, using default gap parameters, or by inspection, and the best alignment (i.e., resulting in the highest percentage of sequence similarity over a comparison window). Percentage of sequence identity is calculated by comparing two optimally aligned sequences over a window of comparison, determining the number of positions at which theidentical residues occur in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison (i.e., the window size), and multiplying the result by 100 to yield the percentage of sequence identity.

[0122] The term “antibody-dependent cellular cytotoxicity”, or ADCC, is a mechanism for inducing cell death that depends upon the interaction of antibody-coated target cells (i.e., cells with bound antibody) with immune cells possessing lytic activity (also referred to as effector cells). Such effector cells include natural killer cells, monocytes / macrophages and neutrophils. ADCC is triggered by interactions between the Fc region of an antibody bound to a cell and Fcγ receptors, particularly FcγRI and FcγRIII, on immune effector cells such as neutrophils, macrophages and natural killer cells. The target cell is eliminated by phagocytosis or lysis, depending on the type of mediating effector cell. Death of the antibody- coated target cell occurs as a result of effector cell activity.

[0123] A humanized antibody is a genetically engineered antibody in which the CDRs from a non-human “donor” antibody are grafted into a human “acceptor” antibody sequence (see, e.g., Queen, US 5,530,101 and 5,585,089; Winter, US 5,225,539, Carter, US 6,407,213, Adair, US 5,859,205 6,881,557, Foote, US 6,881,557). The acceptor antibody sequences can be, for example, a mature human antibody sequence, a composite of such sequences, a consensus sequence of human antibody sequences, or a germline region sequence. Thus, a humanized antibody is an antibody having some or all CDRs entirely or substantially from a donor antibody and variable region framework sequences and constant regions, if present, entirely or substantially from human antibody sequences. Similarly, a humanized heavy chain has at least one, two and usually all three CDRs entirely or substantially from a donor antibody heavy chain, and a heavy chain variable region framework sequence and heavy chain constant region, if present, substantially from human heavy chain variable region framework and constant region sequences. Similarly, a humanized light chain has at least one, two and usually all three CDRs entirely or substantially from a donor antibody light chain, and a light chain variable region framework sequence and light chain constant region, if present, substantially from human light chain variable region framework and constant region sequences. Other than nanobodies and dAbs, a humanized antibody comprises a humanized heavy chain and a humanized light chain. A CDR in a humanized antibody is substantiallyfrom a corresponding CDR in a non-human antibody when at least 85%, 90%, 95% or 100% of corresponding residues (as defined by Kabat) are identical between the respective CDRs. The variable region framework sequences of an antibody chain or the constant region of an antibody chain are substantially from a human variable region framework sequence or human constant region respectively when at least 85, 90, 95 or 100% of corresponding residues defined by Kabat are identical.

[0124] Although humanized antibodies often incorporate all six CDRs (preferably as defined by Kabat) from a mouse antibody, they can also be made with less than all CDRs (e.g., at least 3, 4, or 5 CDRs from a mouse antibody) (e.g., Pascalis et al, J. Immunol. 169:3076, 2002; Vajdos et al., Journal of Molecular Biology, 320: 415-428, 2002; Iwahashi et al., Mol. Immunol. 36: 1079-1091, 1999; Tamura et al, Journal of Immunology, 164: 1432- 1441, 2000).

[0125] A chimeric antibody is an antibody in which the mature variable regions of light and heavy chains of a non-human antibody (e.g., a mouse) are combined with human light and heavy chain constant regions. Such antibodies substantially or entirely retain the binding specificity of the mouse antibody and are about two-thirds human sequence.

[0126] A veneered antibody is a type of humanized antibody that retains some and usually all of the CDRs and some of the non-human variable region framework residues of a non-human antibody but replaces other variable region framework residues that may contribute to B- or T-cell epitopes, for example exposed residues (Padlan, Mol. Immunol. 28:489, 1991) with residues from the corresponding positions of a human antibody sequence. The result is an antibody in which the CDRs are entirely or substantially from a non-human antibody and the variable region frameworks of the non-human antibody are made more human-like by the substitutions. A human antibody can be isolated from a human, or otherwise result from expression of human immunoglobulin genes (e.g., in a transgenic mouse, in vitro or by phage display). Methods for producing human antibodies include the trioma method of Oestberg et al., Hybridoma 2:361-367 (1983); Oestberg, U.S. Patent No. 4,634,664; and Engleman et al., US Patent 4,634,666, use of transgenic mice including human immunoglobulin genes (see, e.g., Lonberg et al., W093 / 12227 (1993); US 5,877,397, US 5,874,299, US 5,814,318, US 5,789,650, US 5,770,429, US 5,661,016, US 5,633,425, US 5,625,126, US 5,569,825, US 5,545,806, Nature 148, 1547-1553 (1994), Nature Biotechnology 14, 826 (1996),Kucherlapati, WO 91 / 10741 (1991) and phage display methods (see, e.g., Dower et al., WO 91 / 17271 and McCafferty et al., WO 92 / 01047, US 5,877,218, US 5,871,907, US 5,858,657, US 5,837,242, US 5,733,743 and US 5,565,332.

[0127] A “polymer” is a molecule composed of many repeating subunits. The subunits, also sometimes referred to as “monomers” can be the same or different. There are both natural and synthetic polymers. DNA, protein and complex carbohydrates are examples of natural polymers. Poly-styrene and poly-acrylamide are examples of synthetic polymers. A polymer composed of repeating units of a single monomer is called a homopolymer. A polymer composed of two or more monomers is called a copolymer or sometimes a heteropolymer. A copolymer in which certain monomer types are clustered together are sometimes called block copolymers. Polymers can be linear or branched. When the polymer is branched, polymer chains having a common origin are sometimes referred to as a polymer arm(s).

[0128] An “initiator” is a compound capable of serving as a substrate on which one or more polymerizations can take place using monomers or comonomers as described herein. The polymerization can be a conventional free radical polymerization or preferably a controlled / ”living” radical polymerization, such as Atom Transfer Radical Polymerization (ATRP), Reversible Addition-Fragmentation-Termination (RAFT) polymerization or nitroxide mediated polymerization (NMP). The polymerization can be a “pseudo” controlled polymerization, such as degenerative transfer. Initiators suitable for ATRP contain one or more labile bonds which can be homolytically cleaved to form an initiator fragment, I, being a radical capable of initiating a radical polymerization, and a radical scavenger, I’, which reacts with the radical of the growing polymer chain to reversibly terminate the polymerization. The radical scavenger I’ is typically a halogen, but can also be an organic moiety, such as a nitrile. Also provided herein, the initiator can contain one or more 2-bromoisobutyrate groups as sites for polymerization via ATRP.

[0129] A “chemical linker” refers to a chemical moiety that links two groups together, such as a half-life extending moiety and a protein. The linker can be cleavable or non-cleavable. Cleavable linkers can be hydrolysable, enzymatically cleavable, pH sensitive, photolabile, or disulfide linkers, among others. Other linkers include homobifunctional and heterobifunctional linkers. A “linking group” is a functional group capable of forming acovalent linkage consisting of one or more bonds to a bioactive agent. Non-limiting examples include those illustrated in Table 1 of WO2013059137 (incorporated by reference).

[0130] The term “reactive group” refers to a group that is capable of reacting with another chemical group to form a covalent bond, i.e. is covalently reactive under suitable reaction conditions, and generally represents a point of attachment for another substance. The reactive group is a moiety, such as maleimide or succinimidyl ester, is capable of chemically reacting with a functional group on a different moiety to form a covalent linkage. Reactive groups generally include nucleophiles, electrophiles and photoactivatable groups.

[0131] As used herein, “phosphorylcholine,” also denoted as “PC,” refers to the following: O *O P ON+(CH3)3where * The phosphorylcholine is azwitterionic group and includes salts (such as inner salts), and protonated and deprotonated forms thereof.

[0132] As used herein, “phosphorylcholine-based polymer” is a polymer that contains phosphorylcholine. “Zwitterion containing polymer” refers to a polymer that contains a zwitterion.

[0133] Poly(acryloyloxyethyl phosphorylcholine) containing polymer refers to a polymer containing 2-(acryloyloxy)ethyl-2-(trimethylammonium)ethyl phosphate as monomer.

[0134] Poly(methacryloyloxyethyl phosphorylcholine) containing polymer refers to a polymer containing 2-(methacryloyloxy)ethyl-2-(trimethylammonium)ethyl phosphate as monomer.

[0135] As used herein, “molecular weight” in the context of the polymer can be expressed as either a number average molecular weight, or a weight average molecular weight or a peak molecular weight. Unless otherwise indicated, all references to molecular weight herein refer to the peak molecular weight. These molecular weight determinations, number average (Mn), weight average (Mw) and peak (Mp), can be measured using size exclusion chromatography or other liquid chromatography techniques. Other methods for measuring molecular weight values can also be used, such as the use of end-group analysis or themeasurement of colligative properties (e.g., freezing-point depression, boiling-point elevation, or osmotic pressure) to determine number average molecular weight, or the use of light scattering techniques, ultracentrifugation or viscometry to determine weight average molecular weight. Also provided herein, the molecular weight can be measured by SEC-MALS (size exclusion chromatography – multi angle light scattering). The polymeric reagents provided herein are typically polydisperse (i.e., number average molecular weight and weight average molecular weight of the polymers are not equal). The Poly Dispersity Index (PDI) provides a measure for the dispersity of polymers in a mixture. PDI is given by the formula Mw / Mn. In this regard a homogenous protein will have a PDI of 1.0 (Mn is the same as Mw). Typically, the PDI for polymers will be above 1.0. Polymers provided herein preferably have relatively low polydispersity (PDI) values of, for example, less than about 1.5, as judged, for example, by SEC-MALS. In other embodiments, the polydispersities (PDI) are more preferably in the range of about 1.4 to about 1.2, still more preferably less than about 1.15, and still more preferably less than about 1.10, yet still more preferably less than about 1.05, and most preferably less than about 1.03.

[0136] As used herein, “protected,” “protected form,” “protecting group” and “protective group” refer to the presence of a group (i.e., the protecting group) that prevents or blocks reaction of a particular chemically reactive functional group in a molecule under certain reaction conditions. Protecting groups vary depending upon the type of chemically reactive group being protected as well as the reaction conditions to be employed and the presence of additional reactive or protecting groups in the molecule, if any. Suitable protecting groups include those such as found in the treatise by Greene et al., “Protective Groups In Organic Synthesis,” 3rdEdition, John Wiley and Sons, Inc., New York, 1999.

[0137] As used herein, “alkyl” refers to a straight or branched, saturated, aliphatic radical having the number of carbon atoms indicated. For example, C1-C6 alkyl includes, but is not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, hexyl, etc. Other alkyl groups include, but are not limited to heptyl, octyl, nonyl, decyl, etc. Alkyl can include any number of carbons, such as 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8, 1-9, 1-10, 2-3, 2-4, 2-5, 2-6, 3-4, 3-5, 3-6, 4-5, 4-6 and 5-6 carbons.

[0138] The term “lower” referred to above and hereinafter in connection with organic radicals or compounds respectively defines a compound or radical which can bebranched or unbranched with up to and including 7, preferably up to and including 4 and (as unbranched) one or two carbon atoms.

[0139] As used herein, “alkylene” refers to an alkyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the alkylene can be linked to the same atom or different atoms of the alkylene. For instance, a straight chain alkylene can be the bivalent radical of -(CH2)n, where n is 1, 2, 3, 4, 5 or 6. Alkylene groups include, but are not limited to, methylene, ethylene, propylene, isopropylene, butylene, isobutylene, sec-butylene, pentylene and hexylene.

[0140] Substituents for the alkyl, alkenyl, alkylene, heteroalkyl, heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl radicals can be one or more of a variety of groups selected from, but not limited to: -OR’, =O, =NR’, =N-OR’, -NR’R”, -SR’, -halogen, -SiR’R”R”‘, -OC(O)R’, -C(O)R’, -CO2R’, -CONR’R”, -O C(O)NR’R”, -NR”C(O)R’, -NR’-C(O)NR”R”‘, -NR”C(O)2R’, -NR-C(NR’R”R’”)=NR”“, -N R-C(NR’R”)=NR’”, -S(O)R’, -S(O)2R’, -S(O)2NR’R”, -NRSO2R’, -CN and –NO2 in a number ranging from 1 to (2m’+1), where m’ is the total number of carbon atoms in such radical. Each of R’, R”, R”‘ and R”“ independently refers to hydrogen, substituted or unsubstituted heteroalkyl, substituted or unsubstituted aryl, e.g., aryl substituted with 1-3 halogens, substituted or unsubstituted alkyl, alkoxy or thioalkoxy groups, or arylalkyl groups. When R’ and R” are attached to the same nitrogen atom, they can be combined with the nitrogen atom to form a 5-, 6-, or 7-membered ring. For example, -NR’R” is meant to include, but not be limited to, 1-pyrrolidinyl and 4-morpholinyl.

[0141] As used herein, “alkoxy” refers to alkyl group attached to an oxygen atom and forms radical –O-R, wherein R is alkyl. Alkoxy groups include, for example, methoxy, ethoxy, propoxy, iso-propoxy, butoxy, 2-butoxy, iso-butoxy, sec-butoxy, tert-butoxy, pentoxy, hexoxy, etc. The alkoxy groups can be further substituted with a variety of substituents described herein. For example, the alkoxy groups can be substituted with halogens to form a “halo-alkoxy” group.

[0142] As used herein, “carboxyalkyl” means an alkyl group (as defined herein) substituted with a carboxy group. The term “carboxycycloalkyl” means an cycloalkyl group (as defined herein) substituted with a carboxy group. The term alkoxyalkyl means an alkylgroup (as defined herein) substituted with an alkoxy group. The term “carboxy” employed herein refers to carboxylic acids and their esters.

[0143] As used herein, “haloalkyl” refers to alkyl as defined above where some or all of the hydrogen atoms are substituted with halogen atoms. Halogen (halo) preferably represents chloro or fluoro, but may also be bromo or iodo. For example, haloalkyl includes trifluoromethyl, fluoromethyl, 1,2,3,4,5-pentafluoro-phenyl, etc. The term “perfluoro” defines a compound or radical which has all available hydrogens that are replaced with fluorine. For example, perfluorophenyl refers to 1,2,3,4,5-pentafluorophenyl, perfluoromethyl refers to 1,1,1-trifluoromethyl, and perfluoromethoxy refers to 1,1,1-trifluoromethoxy. Haloalkyl can also be referred to as halo-substitute alkyl, such as fluoro-substituted alkyl.

[0144] As used herein, “cytokine” in the context provided herein is a member of a group of protein signaling molecules that may participate in cell-cell communication in immune and inflammatory responses. Cytokines are typically small, water-soluble glycoproteins that have a mass of about 8-35 kDa.

[0145] As used herein, “cycloalkyl” refers to a saturated mono- or multi- cyclic aliphatic ring system that contains from about 3 to 12, from 3 to 10, from 3 to 7, or from 3 to 6 carbon atoms. When cycloalkyl group is composed of two or more rings, the rings may be joined together with a fused ring or a spiro ring structure. When cycloalkyl group is composed of three or more rings, the rings may also join together forming a bridged ring structure. Monocyclic rings include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cyclooctyl. Bicyclic and polycyclic rings include, for example, bicyclo[1.1.1]pentane, bicyclco[2.1.1]heptane, norbornane, decahydronaphthalene and adamantane. For example, C3-8 cycloalkyl includes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl, and norbornane.

[0146] As used herein, “endocyclic” refers to an atom or group of atoms which comprise part of a cyclic ring structure.

[0147] As used herein, “exocyclic” refers to an atom or group of atoms which are attached but do not define the cyclic ring structure.

[0148] As used herein, “cyclic alkyl ether” refers to a 4 or 5 member cyclic alkyl group having 3 or 4 endocyclic carbon atoms and 1 endocyclic oxygen or sulfur atom (e.g., oxetane, thietane, tetrahydrofuran, tetrahydrothiophene); or a 6 to 7 member cyclic alkyl grouphaving 1 or 2 endocyclic oxygen or sulfur atoms (e.g., tetrahydropyran, 1,3-dioxane, 1,4-dioxane, tetrahydrothiopyran, 1,3-dithiane, 1,4-dithiane, 1,4-oxathiane).

[0149] As used herein, “alkenyl” refers to either a straight chain or branched hydrocarbon of 2 to 6 carbon atoms, having at least one double bond. Examples of alkenyl groups include, but are not limited to, vinyl, propenyl, isopropenyl, 1-butenyl, 2-butenyl, isobutenyl, butadienyl, 1-pentenyl, 2-pentenyl, isopentenyl, 1,3-pentadienyl, 1,4-pentadienyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 1,3-hexadienyl, 1,4-hexadienyl, 1,5-hexadienyl, 2,4-hexadienyl, or 1,3,5-hexatrienyl. Alkenyl groups can also have from 2 to 3, 2 to 4, 2 to 5, 3 to 4, 3 to 5, 3 to 6, 4 to 5, 4 to 6 and 5 to 6 carbons.

[0150] As used herein, “alkenylene” refers to an alkenyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the alkenylene can be linked to the same atom or different atoms of the alkenylene. Alkenylene groups include, but are not limited to, ethenylene, propenylene, isopropenylene, butenylene, isobutenylene, sec-butenylene, pentenylene and hexenylene.

[0151] As used herein, “alkynyl” refers to either a straight chain or branched hydrocarbon of 2 to 6 carbon atoms, having at least one triple bond. Examples of alkynyl groups include, but are not limited to, acetylenyl, propynyl, 1-butynyl, 2-butynyl, isobutynyl, sec-butynyl, butadiynyl, 1-pentynyl, 2-pentynyl, isopentynyl, 1,3-pentadiynyl, 1,4-pentadiynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 1,3-hexadiynyl, 1,4-hexadiynyl, 1,5-hexadiynyl, 2,4-hexadiynyl, or 1,3,5-hexatriynyl. Alkynyl groups can also have from 2 to 3, 2 to 4, 2 to 5, 3 to 4, 3 to 5, 3 to 6, 4 to 5, 4 to 6 and 5 to 6 carbons.

[0152] As used herein, “alkynylene” refers to an alkynyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the alkynylene can be linked to the same atom or different atoms of the alkynylene. Alkynylene groups include, but are not limited to, ethynylene, propynylene, butynylene, sec-butynylene, pentynylene and hexynylene.

[0153] As used herein, “cycloalkylene” refers to a cycloalkyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the cycloalkylene can be linked to the same atom or different atoms of the cycloalkylene. Cycloalkylene groups include, but are not limited to, cyclopropylene, cyclobutylene, cyclopentylene, cyclohexylene, and cyclooctylene.

[0154] As used herein, “heterocycloalkyl” refers to a ring system having from 3 ring members to about 20 ring members and from 1 to about 5 heteroatoms such as N, O and S. Additional heteroatoms can also be useful, including, but not limited to, B, Al, Si and P. The heteroatoms can also be oxidized, such as, but not limited to, -S(O)- and -S(O)2-. For example, heterocycle includes, but is not limited to, tetrahydrofuranyl, tetrahydrothiophenyl, morpholino, pyrrolidinyl, pyrrolinyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperazinyl, piperidinyl, indolinyl, quinuclidinyl and 1,4-dioxa-8-aza-spiro[4.5]dec-8-yl.

[0155] As used herein, “heterocycloalkylene” refers to a heterocyclalkyl group, as defined above, linking at least two other groups. The two moieties linked to the heterocycloalkylene can be linked to the same atom or different atoms of the heterocycloalkylene.

[0156] As used herein, “aryl” refers to a monocyclic or multicyclic (e.g., fused bicyclic, tricyclic or greater) aromatic ring assembly containing 6 to 16 carbon atoms. For example, aryl may be phenyl, benzyl or naphthyl, preferably phenyl. Aryl groups can be mono-, di- or tri-substituted by one, two or three radicals selected from alkyl, alkoxy, aryl, hydroxy, halogen, cyano, amino, amino-alkyl, trifluoromethyl, alkylenedioxy and oxy-C2-C3-alkylene; all of which are optionally further substituted, for instance as hereinbefore defined; or 1- or 2-naphthyl; or 1- or 2-phenanthrenyl. Alkylenedioxy is a divalent substitute attached to two adjacent carbon atoms of phenyl, e.g. methylenedioxy or ethylenedioxy. Oxy-C2-C3-alkylene is also a divalent substituent attached to two adjacent carbon atoms of phenyl, e.g. oxyethylene or oxypropylene. An example for oxy- C2-C3-alkylene-phenyl is 2,3-dihydrobenzofuran-5-yl.

[0157] Preferred as aryl is naphthyl, phenyl or phenyl mono- or disubstituted by alkoxy, phenyl, halogen, alkyl or trifluoromethyl, especially phenyl or phenyl-mono- or disubstituted by alkoxy, halogen or trifluoromethyl, and in particular phenyl.

[0158] Examples of substituted phenyl groups as R are, e.g. 4-chlorophen-1-yl, 3,4-dichlorophen-1-yl, 4-methoxyphen-1-yl, 4-methylphen-1-yl, 4-aminomethylphen-1-yl, 4-methoxyethylaminomethylphen-1-yl, 4-hydroxyethylaminomethylphen-1-yl, 4-hydroxyethyl-(methyl)-aminomethylphen-1-yl, 3-aminomethylphen-1-yl, 4-N-acetylaminomethylphen-1-yl, 4-aminophen-1-yl, 3-aminophen-1-yl, 2-aminophen-1-yl, 4-phenyl-phen-1-yl, 4-(imidazol-1-yl)-phenyl, 4-(imidazol-1-ylmethyl)-phen-1-yl,4-(morpholin-1-yl)-phen-1-yl, 4-(morpholin-1-ylmethyl)-phen-1-yl, 4-(2-methoxyethylaminomethyl)-phen-1-yl and 4-(pyrrolidin-1-ylmethyl)-phen-1-yl, 4-(thiophenyl)-phen-1-yl, 4-(3-thiophenyl)-phen-1-yl, 4-(4-methylpiperazin-1-yl)-phen-1-yl, and 4-(piperidinyl)-phenyl and 4-(pyridinyl)-phenyl optionally substituted in the heterocyclic ring.

[0159] As used herein, “arylene” refers to an aryl group, as defined above, linking at least two other groups. The two moieties linked to the arylene are linked to different atoms of the arylene. Arylene groups include, but are not limited to, phenylene.

[0160] As used herein, “arylene-oxy” refers to an arylene group, as defined above, where one of the moieties linked to the arylene is linked through an oxygen atom. Arylene-oxy groups include, but are not limited to, phenylene-oxy.

[0161] Similarly, substituents for the aryl and heteroaryl groups are varied and are selected from: -halogen, -OR’, -OC(O)R’, -NR’R”, -SR’, -R’, -CN, -NO2, -CO2R’, -CONR’R”, -C(O) R’, -OC(O)NR’R”, -NR”C(O)R’, -NR”C(O)2R’, -NR’-C(O)NR”R”‘, -NH-C(NH2)=NH, -NR ’C(NH2)=NH, -NH-C(NH2)=NR’, -S(O)R’, -S(O)2R’, -S(O)2NR’R”, -N3, -CH(Ph)2, perfluoro(C1-C4)alkoxy, and perfluoro(C1-C4)alkyl, in a number ranging from zero to the total number of open valences on the aromatic ring system; and where R’, R” and R”‘ are independently selected from hydrogen, (C1-C8)alkyl and heteroalkyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl)-(C1-C4)alkyl, and (unsubstituted aryl)oxy-(C1-C4)alkyl.

[0162] Two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced with a substituent of the formula -T-C(O)-(CH2)q-U-, wherein T and U are independently -NH-, -O-, -CH2- or a single bond, and q is an integer of from 0 to 2. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced with a substituent of the formula -A-(CH2)r-B-, wherein A and B are independently -CH2-, -O-, -NH-, -S-, -S(O)-, -S(O)2-, -S(O)2NR’- or a single bond, and r is an integer of from 1 to 3. One of the single bonds of the new ring so formed may optionally be replaced with a double bond. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced with a substituent of the formula -(CH2)s-X-(CH2)t-, where s and t are independently integers of from 0 to 3, and Xis -O-, -NR’-, -S-, -S(O)-, -S(O)2-, or -S(O)2NR’-. The substituent R’ in -NR’- and -S(O)2NR’- is selected from hydrogen or unsubstituted (C1-C6)alkyl.

[0163] As used herein, “heteroaryl” refers to a monocyclic or fused bicyclic or tricyclic aromatic ring assembly containing 5 to 16 ring atoms, where from 1 to 4 of the ring atoms are a heteroatom each N, O or S. For example, heteroaryl includes pyridyl, indolyl, indazolyl, quinoxalinyl, quinolinyl, isoquinolinyl, benzothienyl, benzofuranyl, furanyl, pyrrolyl, thiazolyl, benzothiazolyl, oxazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl, imidazolyl, thienyl, or any other radicals substituted, especially mono- or di-substituted, by e.g. alkyl, nitro or halogen. Pyridyl represents 2-, 3- or 4-pyridyl, advantageously 2- or 3-pyridyl. Thienyl represents 2- or 3-thienyl. Quinolinyl represents preferably 2-, 3- or 4-quinolinyl. Isoquinolinyl represents preferably 1-, 3- or 4-isoquinolinyl. Benzopyranyl, benzothiopyranyl represents preferably 3-benzopyranyl or 3-benzothiopyranyl, respectively. Thiazolyl represents preferably 2- or 4-thiazolyl, and most preferred, 4-thiazolyl. Triazolyl is preferably 1-, 2- or 5-(1,2,4-triazolyl). Tetrazolyl is preferably 5-tetrazolyl.

[0164] Preferably, heteroaryl is pyridyl, indolyl, quinolinyl, pyrrolyl, thiazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl, imidazolyl, thienyl, furanyl, benzothiazolyl, benzofuranyl, isoquinolinyl, benzothienyl, oxazolyl, indazolyl, or any of the radicals substituted, especially mono- or di-substituted.

[0165] The term “heteroalkyl” refers to an alkyl group having from 1 to 3 heteroatoms such as N, O and S. Additional heteroatoms can also be useful, including, but not limited to, B, Al, Si and P. The heteroatoms can also be oxidized, such as, but not limited to, -S(O)- and -S(O)2-. For example, heteroalkyl can include ethers, thioethers, alkyl-amines and alkyl-thiols.

[0166] The term “heteroalkylene” refers to a heteroalkyl group, as defined above, linking at least two other groups. The two moieties linked to the heteroalkylene can be linked to the same atom or different atoms of the heteroalkylene.

[0167] As used herein, “electrophile” refers to an ion or atom or collection of atoms, which may be ionic, having an electrophilic center, i.e., a center that is electron seeking, capable of reacting with a nucleophile. An electrophile (or electrophilic reagent) is a reagent that forms a bond to its reaction partner (the nucleophile) by accepting both bonding electrons from that reaction partner.

[0168] As used herein, “nucleophile” refers to an ion or atom or collection of atoms, which may be ionic, having a nucleophilic center, i.e., a center that is seeking an electrophilic center or capable of reacting with an electrophile. A nucleophile (or nucleophilic reagent) is a reagent that forms a bond to its reaction partner (the electrophile) by donating both bonding electrons. A “nucleophilic group” refers to a nucleophile after it has reacted with a reactive group. Non limiting examples include amino, hydroxyl, alkoxy, haloalkoxy and the like.

[0169] As used herein, “maleimido” refers to a pyrrole-2,5-dione-1-yl group having the structure: O which upon reaction with a forms an -S-maleimido group havingthe structurewhere “•” indicates the point ofgroup and “ “indicates the pointof the sulfur atom the thiol to the remainder of thesulfhydryl bearing group.

[0170] For the purpose of this disclosure, “naturally occurring amino acids” found in proteins and polypeptides are L-alanine, L-arginine, L-asparagine, L-aspartic acid, L-cysteine, L-glutamine, L-glutamic acid, L-glycine, L-histidine, L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylalanine, L-proline, L-serine, L-threonine, L-tryptophan, L-tyrosine, and or L-valine. “Non-naturally occurring amino acids” found in proteins are any amino acid other than those recited as naturally occurring amino acids. Non-naturally occurring amino acids include, without limitation, the D isomers of the naturally occurring amino acids, and mixtures of D and L isomers of the naturally occurring amino acids. Other amino acids, such as 4-hydroxyproline, desmosine, isodesmosine, 5-hydroxylysine, epsilon-N-methyllysine, 3-methylhistidine, although found in naturally occurring proteins, are considered to be non-naturally occurring amino acids found in proteins for the purpose of thisdisclosure as they are generally introduced by means other than ribosomal translation of mRNA.

[0171] As used herein, “linear” in reference to the geometry, architecture or overall structure of a polymer, refers to polymer having a single polymer arm.

[0172] As used herein, “branched,” in reference to the geometry, architecture or overall structure of a polymer, refers to a polymer having 2 or more polymer “arms” extending from a core structure contained within an initiator. The initiator may be employed in an atom transfer radical polymerization (ATRP) reaction. A branched polymer may possess 2 polymer chains (arms), 3 polymer arms, 4 polymer arms, 5 polymer arms, 6 polymer arms, 7 polymer arms, 8 polymer arms, 9 polymer arms or more. Each polymer arm extends from a polymer initiation site. Each polymer initiation site is capable of being a site for the growth of a polymer chain by the addition of monomers. For example and not by way of limitation, using ATRP, the site of polymer initiation on an initiator is typically an organic halide undergoing a reversible redox process catalyzed by a transition metal compound such as cuprous halide. Preferably, the halide is a bromine.

[0173] As used herein, “pharmaceutically acceptable excipient” refer to an excipient that can be included in the compositions provided herein and that causes no significant adverse toxicological effect on the patient and is approved or approvable by the FDA for therapeutic use, particularly in humans. Non-limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, lactated Ringer’s, normal sucrose, normal glucose and the like.

[0174] As used herein, “OG1786” is a 9-arm initiator used for polymer synthesis with the structure shown in FIG. 14A, which depicts that salt form of OG1786 with trifluororacetic acid. OG1786 may also be used as provided herein as other salts or as the free base.

[0175] As used herein, “OG1801” is an approximately (+ / - 15%) 750 kDa polymer (either by Mn or Mp) made using OG1786 as an initiator for ATRP synthesis using the monomer HEMA-PC. The structure of OG1801 is shown in FIG. 14B.

[0176] As used herein, “OG1802” is OG1801 with a maleimide functionality added, and it has the structure shown in FIG. 14C, wherein each of n1, n2, n3, n4, n5, n6, n7, n8and n9is an integer (positive) (from 0 up to about 3000) such that the total molecular weightof the polymer is (Mw) 750,000 ± 15% Daltons. When the term OG1802 is used to modify a protein term (such as VEGF trap or anti-IL-6 antibody), it designates that the protein is the conjugate protein.

[0177] As used herein “unconjugated protein” and “free protein” are used interchangeably to denote the protein that is not conjugated to a polymer (e.g., not conjugated to a phosphorylcholine-containing polymer).

[0178] As used herein, “molar amount” denotes a measure of the molar quantity of a molecule. In some embodiments, molar amount is a molar concentration (e.g., M, mM, μM, nM, etc.). In some embodiments, molar amount is expressed in units of moles (e.g., moles, millimoles, micromoles, etc.).

[0179] As used herein, CDR positions follow their order of appearance in variable domain when described. For example, heavy chain positions can be described as S35H or G66D. As used herein, Fc positions follow EU numbering when described or when noted to be in EU numbering. For example, mutations of antibodies can be described as L234A or L235A, which would be according to EU numbering. Positions may also be defined according to specified positions within a specific SEQ ID NO or sequence provided herein.

[0180] Multi-angle light scattering (MALS) is a technique of analyzing macromolecules where the laser light impinges on the molecule, the oscillating electric field of the light induces an oscillating dipole within it. This oscillating dipole will re-radiate light and can be measured using a MALS detector such as Wyatt miniDawn TREOS. The intensity of the radiated light depends on the magnitude of the dipole induced in the macromolecule which in turn is proportional to the polarizability of the macromolecule, the larger the induced dipole, and hence, the greater the intensity of the scattered light. Therefore, in order to analyze the scattering from a solution of such macromolecules, one should know their polarizability relative to the surrounding medium (e.g., the solvent). This may be determined from a measurement of the change, Δn, of the solution’s refractive index n with the molecular concentration change, Δc, by measuring the dn / dc (=Δn / Δc) value using a Wyatt Optilab T- rEX differential refractometer. Two molar weight parameters that MALS determination employ are number average molecular weight (Mn) and weight average molecular weight (Mw) where the polydispersity index (PDI) equals Mw divided by Mn. SEC also allowsanother average molecular weight determination of the peak molecular weight Mp which is defined as the molecular weight of the highest peak at the SEC.

[0181] The PDI is used as a measure of the broadness of a molecular weight distribution of a polymer and bioconjugate which is derived from conjugation of a discrete protein to a polydisperse biopolymer (e.g., OG1802). For a protein sample, its polydispersity is close to 1.0 due to the fact that it is a product of translation where every protein molecule in a solution is expected to have almost the same length and molar mass. In contrast, due to the polydisperse nature of the biopolymer where the various length of polymer chains are synthesized during the polymerization process, it is very important to determine the PDI of the sample as one of its quality attribute for narrow distribution of molecular weight.

[0182] Size exclusion chromatography (SEC) is a chromatography technique in which molecules in solution are separated by their size. Typically an aqueous solution is applied to transport the sample through the column which is packed with resins of various pore sizes. The resin is expected to be inert to the analyte when passing through the column and the analytes separate from each other based on their unique size and the pore size characteristics of the selected column.

[0183] Coupling the SEC with MALS or SEC / MALS provides accurate distribution of molar mass and size (root mean square radius) as opposed to relying on a set of SEC calibration standards. This type of arrangement has many advantages over traditional column calibration methods. Since the light scattering and concentration are measured for each eluting fraction, the molar mass and size can be determined independently of the elution position. This is particularly relevant for species with non-globular shaped macromolecules such as the biopolymers (OG1802) or bioconjugates; such species typically do not elute in a manner that might be described by a set of column calibration standards.

[0184] In some embodiments, a SEC / MALS analysis includes a Waters HPLC system with Alliance 2695 solvent delivery module and Waters 2996 Photodiole Array Detector equipped with a Shodex SEC-HPLC column (7.8x300mm). This is connected online with a Wyatt miniDawn TREOS and Wyatt Optilab T-rEX differential refractometer. The Empower software from Waters can be used to control the Waters HPLC system and the ASTRA V 6.1.7.16 software from Wyatt can be used to acquire the MALS data from the Wyatt miniDawn TREOS, dn / dc data from the T-rEX detector and the mass recovery data using theA280 absorbance signal from the Waters 2996 Photodiole Array detector. SEC can be carried out at 1ml / min in 1xPBS pH 7.4, upon sample injection, the MALS and RI signals can be analyzed by the ASTRA software for determination of absolute molar mass (Mp, Mw, Mn) and polydisperse index (PDI). In addition, the calculation also involves the input dn / dc values for polymer and protein as 0.142 and 0.183, respectively. For bioconjugates dn / dc value, the dn / dc is calculated based on the weighted MW of the polymer and the protein to be about 0.148 using the formula below: Conjugate dn / dc = 0.142 x [ MWpolymer / (MWpolymer+MWprotein)]+ 0.183 x [MWprotein / (MWpolymer+MWprotein)] where MWpolymer for OG1802 measured by SEC-MALS is about 800 kDa and the MWprotein for anti-IL-6 measured by SEC-MALS is about 145 kDa, the expected total molecular weight of the bioconjugate measured by SEC-MALS is about 1000 kDa. The MWprotein for the antiIL-6 VEGF Trap or VEGF Trap-antiIL-6 is about 192 kDa, and the expected total molecular weight of the bioconjugate is 1000-1100 kDa.

[0185] As used herein “KSI-501” or “KSI-501ABC” is a bioconjugate of a recombinant, fusion protein that includes an anti-IL-6 antibody fused to a VEGF Trap, and that is covalently conjugated to a branched high molecular weight phosphorylcholine based biopolymer. In some embodiments, KSI-501 is supplied as a preservative free, sterile, aqueous solution in a single-use glass vial. FIG. 27 displays the amino acid sequence of the fusion protein portion of KSI-501 (with or without the C-terminal lysine in the heavy chain of the anti-IL-6 antibody portion). KSI-501 is a dual anti-vascular endothelial growth factor (VEGF) / anti-IL-6 biopharmaceutical with an extended ocular half-life. KSI-501 is a bioconjugate of two intermediates: (1) a recombinant, fusion protein intermediate, a fusion between an anti-IL-6 antibody and a VEGF Trap, and (2) OG1802 biopolymer intermediate, a phosphorylcholine biopolymer. “KSI-501P” denotes a recombinant, fusion protein that includes an anti-IL-6 antibody fused to a VEGF Trap (without the phosphorylcholine based biopolymer).

[0186] The term “percent composition” refers to the percent amount (in mass or concentration units) of a component present in a composition. Percent composition is calculated by determining the amount of a component in mass units (e.g., µg) or inconcentration units (e.g., mg / mL), dividing that amount by the total amount of all components in the composition in the corresponding unit, and multiplying by 100. For compositions and formulations of a conjugate and an unconjugated protein described herein, the amount of the unconjugated protein can be divided by the total amount of the protein component in the solution (excluding the contribution from the polymer component of the conjugate to the mass of the conjugate) to obtain a percent composition.

[0187] As used herein, “% total molar amount” denotes the proportion (in percent) of the amount (in moles or a molar concentration) of one component of a composition relative to the amount(s) (in moles or a molar concentration) of one or more other component of the composition, that together make up the whole (100%). It is understood that percent composition and % total molar amount can be converted between each other where the molecular weight of all of the relevant components is known.

[0188] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art in view of the present disclosure. In case of conflict, the present specification, including definitions, will control. Throughout this specification and claims, the word “comprise,” or variations such as “comprises” or “comprising” will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers. Unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular. Any example(s) following the term “e.g.” or “for example” is not meant to be exhaustive or limiting.

[0189] The phrase “a” or “an” entity refers to one or more of that entity; for example, a compound refers to one or more compounds or at least one compound. As such, the terms “a” (or “an”), “one or more”, and “at least one” can be used interchangeably herein.

[0190] As used herein, “about” means variation one might see in measurements taken among different instruments, samples, and sample preparations. METHODS

[0191] Provided herein are methods for treating an eye disorder (e.g., DME) with a protein fusion conjugate (e.g., KSI-501) that provides within the same molecule, binding domains to VEGF and IL-6, and thereby targets the VEGF / PlGF pathway and IL-6 pathwaysin a subject in need of treatment for the eye disorder, and can further provide for a course of treatment for the eye disorder that includes fewer doses (e.g., less frequent administration) of the protein fusion conjugate than conventional therapies, to achieve a therapeutic effect. With reference to FIG. 1A, a method of treating diabetic macular edema (DME) 100a can include, at block 110a, identifying a subject having DME. Then the method can include, at block 120a, administering no more than 5 loading doses of a fusion protein conjugate, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the loading doses (e.g., the no more than 5 loading doses) of the fusion protein conjugate for at least 4 or at least 8 weeks after a final loading dose. In some embodiments, the method includes administering one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8 or more) loading doses of the fusion protein conjugate. In some embodiments, the subject retains a therapeutic result of administering the loading doses (e.g., no more than 5 loading doses) of the fusion protein conjugate for at least 4 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 20 weeks, at least 24 weeks, at least 28 weeks, or longer, or for a length of time in a range defined by any two of the preceding values (e.g., 4-28 weeks, 8-28 weeks, 10-24 weeks, 12-20 weeks, 8-16 weeks, etc.) after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the subject retains a therapeutic result of administering the loading doses (e.g., no more than 5 loading doses) of the fusion protein conjugate for at least 12 weeks after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the subject retains a therapeutic result of administering the loading doses (no more than 5 loading doses) of the fusion protein conjugate for at least 16 weeks after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the subject retains a therapeutic result of administering the loading doses (no more than 5 loading doses) of the fusion protein conjugate for up to 16 weeks after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the fusion protein and / or fusion protein conjugate comprises any one or more of the constructs in the accompanying figures, including proteins comprising SEQ ID NOs: 169 and 170 (with or without the terminal lysine). Also provided is a method oftreating diabetic macular edema (DME) that includes identifying a subject having DME; and administering one or more loading doses of a fusion protein conjugate, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody.

[0192] With reference to FIG. 1B, a method of treating an eye disorder 100b is provided. The method can include, at block 110b, identifying a subject having an eye disorder. At block 120b, the method can include administering at least one loading dose of a fusion protein conjugate to the subject, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti- IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 4 or at least 8 weeks after a final loading dose. In some embodiments, the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 4 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 20 weeks, at least 24 weeks, at least 28 weeks, or longer, or for a length of time in a range defined by any two of the preceding values (e.g., 4-28 weeks, 8-28 weeks, 10-24 weeks, 12-20 weeks, 8-16 weeks, etc.) after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 12 weeks after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 16 weeks after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for up to 16 weeks after a final loading dose (e.g., after the third or fourth loading dose). In some embodiments, the fusion protein and / or fusion protein conjugate comprises any one or more of the constructs in the accompanying figures, including proteins comprising SEQ ID NOs: 169 and 170. Alsoprovided is a method of treating an eye disorder, comprising: identifying a subject having an eye disorder; and administering at least one loading dose of a fusion protein conjugate to the subject, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti- IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody.

[0193] The method can include treating any suitable eye disorder. In some embodiments, the eye disorder includes, without limitation, diabetic macular edema (DME), wet age-related macular degeneration (wAMD), retinal vein occlusion (RVO), diabetic retinopathy (DR), macular edema secondary to inflammation (MESI), and / or uveitis. In some embodiments, the eye disorder is DME. In some embodiments, the disorder is selected from the group consisting of wet or neovascular macular degeneration, wet or neovascular age- related macular degeneration, dry age-related macular degeneration, venous, arterial or other blockage of the ocular and or retinal blood vessels with or without retinal edema, anterior and posterior uveitis, uveitic macular edema, diabetic retinopathy, proliferative diabetic retinopathy, diabetic macular edema, non-proliferative diabetic macular edema, and intraocular tumors. In some embodiments, the fusion protein and / or fusion protein conjugate comprises any one or more of the constructs in the accompanying figures, including proteins comprising SEQ ID NOs: 169 and 170.

[0194] In some embodiments, the eye disorder is a VEGF therapy-resistant eye disorder. In some embodiments, the eye disorder is resistant to a VEGF Trap therapy, e.g., an aflibercept therapy. In some embodiments, the eye disorder is resistant to an anti-VEGF antibody therapy, e.g., a ranibizumab or bevacizumab therapy. In some embodiments, the eye disorder is resistant to the VEGF therapy when a subject receiving a standard-of-care VEGF therapy exhibits a therapeutic response that is, is about, or is at most 50%, 40%, 30%, 20%, 10%, 5%, 2%, 1% or less, or a percentage in a range defined by any two of the preceding values (e.g., 1-50%, 2-40%, 5-50%, 1-30%, etc.) of an average therapeutic response for the VEGF therapy.

[0195] In some embodiments, the subject has DME. In some embodiments, the subject has type 2 diabetes. In some embodiments, the subject having DME has type 2diabetes. In some embodiments, the subject having DME has type 1 diabetes. In some embodiments, the subject has a hemoglobin A1c (HbA1c) level of about 6.5-12 %. In some embodiments, the subject has a HbA1c level of or of about 6.5, 6.6, 6.7, 6.8, 6.9, 7, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8, 8.1, 8.2, 8.4, 8.4, 8.5, 8.6, 8.7, 8.8, 9, 9.5, 10, 10.5, 11, 11.5, or 12%, or a percentage in a range defined by any two of the preceding values (e.g., about 6.5- 12%, about 6.5-10%, about 6.5-9%, etc.). In some embodiments, the subject has a hemoglobin A1c (HbA1c) level of at most 12%.

[0196] In some embodiments, the subject has a baseline central subfield retinal thickness (CST) in an affected eye (e.g., as measured by optical coherence tomography (OCT)), before being treated with the fusion protein conjugate, of about 200 microns or more, about 250 microns or more, about 275 microns or more, about 300 microns or more, about 325 microns or more, about 350 microns or more, about 375 microns or more, about 400 microns or more, about 425 microns or more, about 450 microns or more, about 475 microns or more, about 500 microns or more, about 525 microns or more, about 550 microns or more, about 575 microns or more, about 600 microns or more, about 625 microns or more, about 650 microns or more, about 675 microns or more, about 700 microns or more, about 725 microns or more, about 750 microns or more, about 775 microns or more, about 800 microns or more, about 825 microns or more, about 850 microns or more, about 875 microns or more, about 900 microns or more, or a thickness in a range defined by any two of the preceding values (e.g., about 200- 900 microns, about 250-800 microns, about 300-600 microns, including about 300-500 microns, etc.). In some embodiments, the subject has, before being treated with the fusion protein conjugate, a CST of at least about 320 microns.

[0197] In some embodiments, a subject with the eye disorder (e.g., DME) has, before being treated with the fusion protein conjugate, a mean best-corrected visual acuity (BCVA), in Early treatment diabetic retinopathy study (ETDRS) letters, of about 80 or less, about 75 or less, about 70 or less, about 68 or less, about 66 or less, about 64 or less, about 62 or less, about 60 or less, about 58 or less, about 56 or less, about 54 or less, about 52 or less, about 50 or less, about 48 or less, about 46 or less, about 44 or less, about 42 or less, about 40 or less, about 38 or less, about 36 or less, about 34 or less, about 32 or less, about 30 or less, about 28 or less, about 26 or less, about 25 or less, or a ETDRS letters value within a range defined by any two of the preceding values (e.g., 25-80 ETDRS letters, 25-70 ETDRS letters,30-70 ETDRS letters, etc.). In some embodiments, a subject with the eye disorder (e.g., DME) has, before being treated with the fusion protein conjugate, a BCVA of about 25-70 ETDRS letters. In some embodiments, the subject’s BCVA is represented using the corresponding value in the Snellen chart. In some embodiments, a subject with the eye disorder (e.g., DME) has, before being treated with the fusion protein conjugate, a BCVA of about 20 / 40 – 20 / 320 Snellen.

[0198] The method can include any suitable number of loading doses of the fusion protein conjugate. In some embodiments, the number of loading doses is or is at most 1, 2, 3, 4, or 5. In some embodiments, the number of loading doses is in a range of 1-5, 2-5, 3-5, 4-5, 1-4, 2-4, 3-4, 1-3, 2-3, or 1-2. In some embodiments, the number of loading doses is 3. In some embodiments, the number of loading doses is 4.

[0199] The loading doses can be administered under any suitable dosing schedule. In some embodiments, the loading doses are administered no more frequently than once every 2 weeks (Q2W), once every 4 weeks (Q4W), once every 8 weeks (Q8W), once every 12 weeks (Q12W), once a month (QM), once every two months (Q2M), once every three months (Q3M), once every four months (Q4M), once every 6 months (Q6M), or less frequently, or at a dosing schedule in a range defined by any two of the preceding values (e.g., administered once every 2 weeks to 6 months, every 2-12 weeks, every 4-12 weeks, every 1-6 months, etc.).

[0200] After receiving the last dose (e.g., the final loading dose, or any dosing that occurred last), the subject may retain a therapeutic result of administering the fusion protein conjugate for a sustained period of time without the subject receiving a subsequent dose (e.g., a maintenance dose) of the fusion protein conjugate. A therapeutic result of administering the fusion protein conjugate may include an improvement in one or more of visual acuity or retinal health (e.g., retinal thickness, extent of retinal perfusion, etc.) at or around the time of the final loading dose compared to before or at the time of the first loading dose. Any suitable therapeutic result of administering the fusion protein may be used according to methods of the present disclosure. Suitable measures for determining therapeutic results include, e.g., visual acuity, retinal thickness, perfusion in at least one eye, diabetic retinopathy severity score (DRSS), disease activity of the eye disorder, or any combination thereof. In some embodiments, wAMD, DME, RVO, or DR disease activity includes one or more of increased intraretinal fluid, increased subretinal fluid, new intraretinal hemorrhage, or new exudates.

[0201] In some embodiments, the therapeutic result includes an improvement, or at least a slowed decline, in visual acuity. Visual acuity may be monitored using any suitable method. In some embodiments, the visual acuity is measured by best corrected visual acuity (BCVA) using e.g., ETDRS letters or Snellen chart, etc. In some embodiments, the therapeutic result includes an improvement in BCVA measured by ETDRS letters of 3 or more, e.g., 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, 12 or more, 15 or more, 18 or more, 20 or more, 22, or more, including 25 or more, or by a ETDRS letters value within a range defined by any two of the preceding values, compared to pre-treatment (e.g., 3- 25 ETDRS letters, 5-15 ETDRS letters, 5-10 ETDRS letters, 3-15 ETDRS letters, etc.). In some embodiments, the therapeutic result includes an improvement in BCVA measured by ETDRS letters of about 5 letters or more. In some embodiments, the therapeutic result includes an improvement in BCVA measured by ETDRS letters of about 10 letters or more. In some embodiments, the therapeutic result may include a reduction in the rate of deterioration of visual acuity as measured by BCVA ETDRS letters by at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage reduction in the rate of deterioration in a range defined by any two of the preceding values (e.g., about 10-100%, about 15-90%, about 25-75%, etc.), over pre-treatment. In some embodiments, the therapeutic result includes an improvement in BCVA measured by ETDRS of, or at least, or of about 1, 2, 3, or 4 lines, or a number in a range defined by any two of the preceding values (e.g., about 1-4 lines, about 2-4 lines, about 1-3 lines, about 2-3 lines, etc.). In some embodiments, the therapeutic result includes an improvement in BCVA measured by ETDRS of at least 2 lines. In some embodiments, the therapeutic result includes an improvement in BCVA measured by ETDRS of about 2-3 lines.

[0202] In some embodiments, the therapeutic result includes a reduction, or at least a slowed increase, in retinal thickness (e.g., central subfield thickness (CST)). The retinal thickness may be measured using any suitable method, including, but not limited to, optical coherence tomography (OCT) or OCT-A. In some embodiments, the therapeutic result includes a reduction in central subfield thickness (CST) of about 10 µm or more, e.g., about 15 µm or more, about 20 µm or more, about 25 µm or more, about 30 µm or more, about 40 µm or more, about 50 µm or more, about 75 µm or more, about 100 µm or more, about 125 µm or more, about 150 µm or more, about 175 µm or more, about 200 µm or more, about 225µm or more, about 250 µm or more, about 275 µm or more, about 300 µm or more, about 325 µm or more, about 350 µm or more, about 375 µm or more, about 400 µm or more, or a reduction within a range defined by any two of the preceding values, compared to pre-treatment (e.g., about 10-400 µm, about 15-300 µm, about 20-200 µm, about 10-150 µm, etc.). In some embodiments, the therapeutic result includes a reduction in CST of about 20 µm or more. In some embodiments, the therapeutic result includes a reduction in CST of about 50 µm or more. In some embodiments, the therapeutic result includes a reduction in CST of about 100 µm or more. In some embodiments, the therapeutic result may include a reduction in the rate of increase in retinal thickness by at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage reduction in the rate of increase in retinal thickness in a range defined by any two of the preceding values (e.g., about 10-100%, about 15-90%, about 25-75%, etc.), over pre-treatment.

[0203] In some embodiments, the therapeutic result includes improved perfusion, or at least a reduction in the rate of expansion of non-perfusion, of the retina. Perfusion may be monitored using any suitable method. Suitable methods include, without limitation, OCT- angiography (OCT-A), fluorescein angiogram or ultrawide-field fluorescein angiogram. The degree of perfusion, or non-perfusion, may be measured using any suitable measure. In some embodiments, non-perfusion area or area of capillary non-perfusion is measured. In some embodiments, an ischemic index is calculated by dividing the non-perfusion area by the total retinal area. In some embodiments, the presence or absence of retinal non-perfusion in retinal quadrants on the angiogram is measured. In some embodiments, the therapeutic result may include a reduction in the area of non-perfusion of at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage in a range defined by any two of the preceding values (e.g., about 10-100%, about 15-90%, about 25- 75%, etc.), over pre-treatment. In some embodiments, the therapeutic result may include a reduction in the rate of progressive non-perfusion of at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage in a range defined by any two of the preceding values (e.g., about 10-100%, about 15-90%, about 25-75%, etc.), over pre-treatment.

[0204] In some embodiments, the therapeutic result includes improved, or prevented worsening of, diabetic retinopathy severity score (DRSS). In some embodiments,the therapeutic result may include an improved DRSS of 2 steps or more, or 3 steps or more compared to pre-treatment. In some embodiments, the therapeutic result may include preventing worsening of DRSS by 2 steps or more, or 3 steps or more compared to pre- treatment.

[0205] The therapeutic result is retained if the level of visual acuity or retinal health (e.g., retinal thickness, degree of non-perfusion, etc.) does not worsen by more than a predetermined amount compared to the improved level. In some embodiments, the therapeutic result is retained if the level of visual acuity or retinal health does not revert by 30% or more, e.g., 50% or more, 75% or more, 90% or more, including 100% or more to the pretreatment level of visual acuity or retinal health after the last dose (e.g., final loading dose). In some embodiments, the therapeutic result is retained if the rate of change of visual acuity or retinal health does not revert by 30% or more, e.g., 50% or more, 75% or more, 90% or more, including 100% or more to the pretreatment level of the rate of change of visual acuity or retinal health after the last dose (e.g., final loading dose).

[0206] In some embodiments, the method includes administering one or more subsequent doses (e.g., maintenance or retreatment dose) of the fusion protein conjugate to the subject after a final loading dose. Without limitation, the subsequent dose(s) may be administered to the subject no more frequently than once every 8 weeks, e.g., every 10 weeks, every 12 weeks, every 14 weeks, every 16 weeks, every 18 weeks, every 20 weeks, every 24 weeks, every 28 weeks, every 32 weeks, every 40 weeks, including every 52 weeks. In some embodiments, no subsequent dose is administered to the subject within or within about 4, 6, 8, 10, 12, 14, 16, 20, 24, 28, 32 weeks, or longer, or a length of time in a range defined by any two of the preceding values (e.g., 4-32 weeks, 6-28 weeks, 8-24 weeks, 8-16 weeks, etc.) of a final loading dose. In some embodiments, no subsequent dose is administered to the subject within about 8 weeks of a final loading dose. In some embodiments, no subsequent dose is administered to the subject within about 12 weeks of a final loading dose. In some embodiments, no subsequent dose is administered to the subject within about 16 weeks of a final loading dose. In some embodiments, no subsequent dose of the fusion protein conjugate is administered to the subject after a final loading dose

[0207] In some embodiments, the subsequent dose may be administered based on a predetermined schedule (e.g., a schedule determined before the subject is administered anyof the one or more of the loading doses). In some embodiments, the subsequent dose may be administered based on a predetermined schedule based on the severity of the eye disorder (e.g., DME), the subject’s previous response, or lack thereof, to other therapies for the eye disorder (e.g., DME), or any other clinically relevant factors associated with the subject. In some embodiments, the subsequent dose may be administered based on the outcome of one or more assessment tests for ocular health and / or function carried out on the subject during the course of treatment with the fusion protein conjugate. In some embodiments, the subsequent dose may be administered based on the outcome of one or more assessment tests carried out on the subject every 4 or more weeks, e.g., every 6 or more weeks, every 8 or more weeks, every 10 or more weeks, every 12 or more weeks, every 16 or more weeks, every 20 or more weeks, every 24 or more weeks, every 28 or more weeks, every 32 or more weeks, every 36 or more weeks, including every 40 or more weeks. In some embodiments, the subsequent dose may be administered if one or more assessment tests indicates a diminishment of the therapeutic result of administering the fusion protein conjugate that is greater than a predetermined threshold.

[0208] Any suitable amount of the fusion protein conjugate (and / or an unconjugated fusion protein) can be administered to the subject in a loading dose, individualized dose, maintenance dose, or a subsequent dose. As used herein, the amount of a fusion protein conjugate in a dose or composition is denoted by the amount or weight of the protein component (e.g., the fusion protein portion excluding the contribution of the polymer to the total mass) of the fusion protein conjugate. In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes or includes about, without limitation, 1, 1.1, 1.2, 1.25, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3, 3.1, 3.2, 3.3., 3.4, 3.5, 3.6, 3.7, 3.75, 3.8, 3.9, 4, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6 mg, or more, or an amount in a range defined by any two of the preceding values (e.g., about 1-6 mg, about 1.3-5.3 mg, about 1.25- 5.3 mg, about 1.3-5 mg, about 1.25-5 mg, about 2.6-5.3 mg, about 1.5-4.5 mg, about 2-4 mg, etc.) of the fusion protein conjugate (by weight of the fusion protein) per eye. In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes between about 1.3 mg and about 5 mg of the fusion protein conjugate (by weight of the fusion protein) per eye. In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes between about 1.25 mg and about 5mg of the fusion protein conjugate (by weight of the fusion protein) per eye. In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes about 1.3 mg, about 2.8 mg, about 3.9 mg, or about 5.3 mg of the fusion protein conjugate (by weight of the fusion protein) per eye. In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes about 1.3 mg, about 2.5 mg, about 3.8 mg, or about 5 mg of the fusion protein conjugate (by weight of the fusion protein) per eye. In some embodiments, the fusion protein conjugate and the unconjugated fusion protein are present in the dose (e.g., loading dose and / or subsequent dose) in therapeutically effective amounts. In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes about 3.5 mg of the fusion protein conjugate (by weight of the fusion protein portion) and about 1.5 mg of the unconjugated fusion protein (as described herein). In some embodiments, a dose (e.g., loading dose and / or subsequent dose) of the fusion protein conjugate includes 2-5.5 mg of the fusion protein conjugate (by weight of the fusion protein portion) and 1-2 mg of the unconjugated fusion protein (as described herein).

[0209] In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of, or of about 25, 30, 35, 40, 45, 47.5, 50, 52.5, 55, 57.5, 60, 65 mg / ml, or a concentration in a range defined by any two of the preceding values (e.g., about 30-65 mg / ml, about 40-60 mg / ml, about 50-55 mg / ml, etc.) (by weight of fusion protein). In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of 25 to 55 mg / ml (by weight of the fusion protein). In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of between about 30 to about 65 mg / ml (by weight of the fusion protein). In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of about 52.5 mg / ml (by weight of fusion protein). In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of about 50 mg / ml (by weight of fusion protein). In some embodiments, the dose (e.g., loading dose and / or subsequent dose) includes the fusion protein conjugate, and the concentration of the fusion protein (with or without a polymer conjugated thereto) in the dose is, or is about 25, 30, 35, 40, 45, 47.5, 50, 52.5, 55, 57.5, 60, 65 mg / ml, or a concentration in a range defined byany two of the preceding values (e.g., about 25-65 mg / ml, about 30-65 mg / ml, about 40-60 mg / ml, about 50-55 mg / ml, etc.). In some embodiments, the dose (e.g., loading dose and / or subsequent dose) includes the fusion protein conjugate, and the concentration of the fusion protein (with or without a polymer conjugated thereto) in the dose is between about 30 and about 65 mg / ml. In some embodiments, the dose (e.g., loading dose and / or subsequent dose) includes the fusion protein conjugate, and the concentration of the fusion protein (with or without a polymer conjugated thereto) in the dose is about 52.5 mg / ml. In some embodiments, the dose (e.g., loading dose and / or subsequent dose) includes the fusion protein conjugate, and the concentration of the fusion protein (with or without a polymer conjugated thereto) in the dose is about 50 mg / ml. In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of, of about, or of at least 35 mg / ml. In some embodiments, the fusion protein conjugate and the unconjugated fusion protein are present in the dose (e.g., loading dose and / or subsequent dose) in therapeutically effective concentrations. In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of, of about, or of at least 35 mg / ml, where the dose also includes unconjugated fusion protein (as described herein) at a concentration of, of about, or of at least 15 mg / ml. In some embodiments, the fusion protein conjugate is present in the dose (e.g., loading dose and / or subsequent dose) at a concentration of 25 to 55 mg / ml (by weight of the fusion protein), where the dose also includes unconjugated fusion protein (as described herein) at a concentration of 10-20 mg / ml.

[0210] In some embodiments, the same amount of the fusion protein conjugate is in each dose (e.g., each loading dose and / or each subsequent dose) administered to the subject. In some embodiments, the same amount of the fusion protein conjugate is in each loading dose. In some embodiments, the same amount of the fusion protein conjugate is in each subsequent dose.

[0211] In some embodiments, the dose (e.g., loading dose and / or subsequent dose) includes a mixture of the fusion protein conjugate and an unconjugated fusion protein, where the total amount of the fusion protein (e.g., sum of the amount of the protein component of the fusion protein conjugate and the amount of the unconjugated fusion protein) in the dose is any of the amount or in a range as described above. In some embodiments, substantially all (e.g., 85%, 90%, or 95% or more) of the fusion protein is present in the dose (e.g., loading doseand / or subsequent dose) as the fusion protein conjugate. In any of the methods herein, in some embodiments, the fusion protein conjugate can be replaced with the unconjugated fusion protein. In some embodiments, instead of the fusion protein conjugate, a method of the present disclosure includes administering the fusion protein (e.g., without a polymer conjugated thereto).

[0212] The dose (e.g., loading dose or subsequent dose) of the fusion protein conjugate can be administered using any suitable route of administration. In some embodiments, the method includes oral, intravitreal, intravenous, subcutaneous, intramuscular, intraosseous, intranasal, topical, intraperitoneal, and intralesional administration of the dose (e.g., loading dose or subsequent dose) of the fusion protein conjugate. In some embodiments, suitable parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration or routes among others. In some embodiments, the method includes intravitreal administration of the dose (e.g., loading dose or subsequent dose) of the fusion protein conjugate to the subject. The dose can be prepared in any suitable pharmaceutical composition or therapeutic formulation, as disclosed herein, for the desired route of administration.

[0213] Also provided is a method of treating diabetic macular edema (DME), comprising identifying a subject having DME; and administering 3 or 4 intravitreal doses of a fusion protein conjugate at Q4W, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody comprising a heavy chain and a light chain; and a VEGF Trap fused to the heavy chain, wherein the heavy chain fused to the VEGF Trap comprises an amino acid sequence as set forth in SEQ ID NO: 170 (with or without the C- terminal lysine), and the light chain an amino acid sequence as set forth in SEQ ID NO: 169, wherein the fusion protein conjugate comprises the following structure:O O X PC n2 PC PC = P N(CH ) O O O CH O PC O X n5 PCwherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to the heavy chain of the anti-IL-6 antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains, PC is O CH3, wherein the curvy line indicates the point of attachment to the restis: a) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I, d) –SCN, or e) –NCS; and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%, wherein each dose comprises about 1 mg to about 6 mg of the fusion protein per eye, whereby the subject retains a therapeutic result of administering thethree doses of the fusion protein conjugate for at least 16 weeks after the third dose. In some embodiments, –X is –Br.

[0214] Also provided is a method of treating an eye disorder that includes: identifying a subject having an eye disorder; and administering 3 or 4 intravitreal loading doses of a fusion protein conjugate to the subject at Q4W, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain fused to the VEGF Trap comprises an amino acid sequence as set forth in SEQ ID NO: 170 (with or without the C- terminal lysine), and the light chain comprises an amino acid sequence as set forth in SEQ ID NO: 169, wherein the fusion protein conjugate comprises the following structure: O O X PC n2 PC PC = P N(CH ) O O O CH O PC O X n5 PCwherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L, the polymer is bonded to the heavy chain of the anti-IL-6 antibody through thesulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains, PC is , where the curvy line indicates the point of attachment to the rest ofa) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%, wherein each dose comprises about 1 mg to about 6 mg of the fusion protein per eye, whereby the subject retains a therapeutic result of administering the 3 or 4 intravitreal doses of the fusion protein conjugate for at least 16 weeks after a final loading dose. In some embodiments, –X is –Br.

[0215] In any method of treating an eye disorder (e.g., DME, wAMD, etc.) provided herein, in some embodiments, the method includes administering to the subject a therapeutic formulation or pharmaceutical composition containing the dose (e.g., loading dose, subsequent dose) of the fusion protein conjugate as provided herein. The pharmaceutical composition or therapeutic formulation can include any suitable components, as disclosed herein. In some embodiments, the therapeutic formulation or pharmaceutical composition includes a mixture of the conjugated and unconjugated form of the fusion protein that includes an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine, as described herein (e.g., mixture of KSI-501ABC and KSI-501P). In some embodiments, the therapeutic formulation or pharmaceutical composition includes: an unconjugated fusion protein comprising the fusion protein with or without the non-native cysteine in the Fc region, wherein the unconjugated fusion protein is not covalently attached to a phosphorylcholine-containing polymer; and the fusion protein conjugate, wherein the unconjugated fusion protein is present in the formulation in a range of 10-60% of a total molar amount of the fusion protein conjugate and the unconjugated fusion protein, wherein the total molar amount is the sum of the molar amount of the fusion protein conjugate and the molar amount of the unconjugated fusion protein. In some embodiments, the unconjugated fusion protein is present in the formulation at 15-45% of the total molar amount of the fusion protein conjugate and the unconjugated fusion protein. In some embodiments, the unconjugated fusion protein is present in theformulation at 20-40% of the total molar amount of the fusion protein conjugate and the unconjugated fusion protein. In some embodiments, the unconjugated fusion protein is present in the formulation at about 30% of the total molar amount of the fusion protein conjugate and the unconjugated fusion protein.

[0216] Some non-limiting embodiments disclosed herein (with reference to Figure 28) are a method of treating wet age-related macular degeneration (wet AMD) (2800), the method comprising identifying a subject with wet AMD (2801). The method can further include administering to the subject a first dose of a fusion protein conjugate (e.g., KSI-501) comprising a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody (2802). The method can also include administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose (2803). The method can include administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose (2804). The method can also include administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose (2805). Further, the method can include administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose (2806). The method can also include administering an individualized dose of the fusion protein conjugate to the subject at least 4 weeks (+ / - 7 days) after the fourth dose or about every 8 weeks thereafter, wherein each individualized dose is administered upon determining that there is a decline in the subject’s eye health, wherein the subsequent dose and the individualized dose are collectively administered no more frequently than about once every four weeks (or no more frequently than Q4W or QM) (2807). The first, second, third and fourth doses can each be a loading dose. In some embodiments, the subject’s eye health is not determined after administering the first dose and before administering the second, third, and / or fourth doses. In some embodiments, the method includes administering an individualized dose of the fusion protein conjugate to the subject upon determining that the subject has a decline in eye health. In some embodiment, the method includes determining that the subject has a decline in eye health, and administering the individualized dose of the fusion protein conjugate to the subjectbased on the determined decline in eye health. The decline in eye health can be based on any suitable anatomical and / or visual acuity-based measure of eye health, as described herein. In some embodiments, administering the dose (e.g., first, second, third, fourth dose) is performed no sooner than the designated point in time (e.g., no sooner than four weeks after the last dose). In some embodiments, administering the dose (e.g., first, second, third, fourth dose) is performed no sooner than four weeks after the last dose. In some embodiments, the dosing is ±1, 2, 3, 4, 5, 6, or 7 days from the designated point in time. In some embodiments, the first, second, third, and fourth doses are each a loading dose. In some embodiments, administering the first, second, third, and fourth doses does not depend on the eye health of the subject. In some embodiments, the subject’s eye health is not evaluated to determine whether to administer the first, second, third, and fourth doses. In some embodiments, the subject’s eye health is not evaluated until after the fourth dose. In some embodiments, the subject’s eye health is evaluated only after the fourth dose.

[0217] In some embodiments, any one or more of the dosing intervals between consecutive doses among the first, second, third and fourth doses (e.g., between the first and second doses, between the second and third doses, between the third and fourth doses) is 3, 4, or 5 weeks. In some embodiments, any one or more of the dosing intervals between consecutive doses (e.g., between the first and second doses, between the second and third doses, between the third and fourth doses) is in the range of 3-4 weeks, or 4-5 weeks. In some embodiments, any one or more of the dosing intervals between consecutive doses (e.g., between the first and second doses, between the second and third doses, between the third and fourth doses) is in the range of 3-5 weeks, e.g., 21-35 days, 22-34 days, 23-33 days, 24-32 days, 25-31 days, 26-30 days, or 27-29 days. In some embodiments, any one or more of the dosing intervals between consecutive doses (e.g., between the first and second doses, between the second and third doses, between the third and fourth doses) is 4 weeks. In some embodiments, any one or more of the dosing intervals between consecutive doses (e.g., between the first and second doses, between the second and third doses, between the third and fourth doses) is or is about one month. In some embodiments, the method includes administering the individualized dose of the fusion protein conjugate (e.g., KSI-501) to the subject at least one month (+ / - 7 days) after the fourth dose, wherein thereafter the individualized dose is administered no more frequently than about once every 8 weeks (or nomore frequently than Q8W or Q2M). In some embodiments, the method includes administering the individualized dose of the fusion protein conjugate to the subject at least one month (+ / - 7 days) after the fourth dose or about every 8 weeks thereafter, wherein each individualized dose is administered upon determining that there is a decline in the subject’s eye health, wherein the individualized dose is administered no more frequently than about once every 8 weeks (or no more frequently than Q8W or Q2M). In some embodiments, the subsequent doses are administered at, at about, or no more frequently than Q8W, Q12W, Q16W, Q20W, or Q24W. In some embodiments, the subsequent dose is not administered to the subject until there is a decline in the subject’s eye health (e.g., until after administering the first individualized dose after the final loading dose).

[0218] Some non-limiting embodiments disclosed herein (with reference to Figure 29) are a method of treating wet AMD (2900), the method comprising identifying a subject with wet AMD (2901). The method can further include administering to the subject a first dose of a fusion protein conjugate (e.g., KSI-501) comprising a fusion protein comprising: an anti- IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody (2902). The method can also include administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose (2903). The method can also include administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose (2904). The method can also include administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose (2905). Further, the method can include administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose (2906). The first, second, third and fourth doses can each be a loading dose. In some embodiments, the subject’s eye health is not determined after administering the first dose and before administering the second, third, and / or fourth doses. In some embodiments, the subject’s eye health is not determined until after the fourth dose. In some embodiments, the subject’s eye health is evaluated only after the fourth dose. In some embodiments, the method includes evaluating the subject’s eye health after the fourth dose to determine if the subject has a decline in eye health; and administering an individualized doseof the fusion protein conjugate to the subject at least 4 weeks (+ / - 7 days) after the fourth dose or about every 8 weeks thereafter, wherein each individualized dose is administered upon determining that there is a decline in the subject’s eye health. In some embodiments, the subsequent dose and the individualized dose are collectively administered no more frequently than about once every four weeks (or no more frequently than Q4W or QM). In some embodiments, administering the dose is no sooner than the designated point in time. In some embodiments, the dosing is ±1, 2, 3, 4, 5, 6, or 7 days from the designated point in time. The decline in eye health can be based on any suitable anatomical and / or visual acuity-based measure of eye health, as described herein.

[0219] Some embodiments disclosed herein (with reference to Figure 30) are a method of treating wet AMD (3000), the method comprising identifying a subject with wet AMD (3001). The method can further include administering to the subject a first dose of a fusion protein conjugate comprising a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody (3002). The method can also include administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose (3003). The method can also include administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose (3004). The method can also include administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose (3005). The first, second, third and fourth doses can each be a loading dose. In some embodiments, the subject’s eye health is not determined after administering the first dose and before administering the second, third, and / or fourth doses. In some embodiments, the subject’s eye health is not determined until after the fourth dose. In some embodiments, the subject’s eye health is evaluated only after the fourth dose. The method can also include administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose (3006). The method can also include evaluating the subject’s eye health at least 4 weeks (+ / - 7 days) after the fourth dose and subsequently no more frequently than about once every eight weeks (or no more frequently than Q8W or Q2M) (3007). Further, the method can include administering an individualized dose to the subjectbased on the subject’s evaluated eye health (3007). If the subject has a decline in eye health upon the evaluating, the method can include administering an individualized dose of the fusion protein conjugate to the subject (3008). In some embodiments, the dosing is no sooner than the designated point in time. In some embodiments, the dosing is ±1, 2, 3, 4, 5, 6, or 7 days from the designated point in time.

[0220] Some non-limiting embodiments disclosed herein (with reference to Figure 31) are a method of treating eye disease (3100), the method comprising identifying a subject with a neovascular eye disease (3101). The method can further include administering to the subject a first dose of a fusion protein conjugate (e.g., KSI-501) comprising a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody (3102). The method can also include administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose (3103). The method can also include administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose (3104). The method can also include administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose (3105). The first, second, third and fourth doses can each be a loading dose. In some embodiments, the subject’s eye health is not determined after administering the first dose and before administering the second, third, and / or fourth doses. In some embodiments, the subject’s eye health is not determined until after the fourth dose. In some embodiments, the subject’s eye health is evaluated only after the fourth dose. Further, the method can include administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose (3106). The method can also include administering an individualized dose of the fusion protein conjugate to the subject at least 4 weeks (+ / - 7 days) after the fourth dose or about every 8 weeks thereafter, wherein each individualized dose is administered upon determining a decline in the subject’s eye health, wherein the subsequent dose and the individualized dose are collectively administered no more frequently than about once every four weeks (or no more frequently than Q4W or QM) (3107). The decline in eye health can be based on any suitable anatomical and / or visual acuity-based measure of eye health, as described herein.

[0221] The methods provided herein can treat any suitable neovascular eye disease. In some embodiments, the neovascular eye disease is diabetic retinopathy (DR), age-related macular degeneration (AMD, e.g., wet AMD), diabetic macular edema (DME), or retinal vein occlusion (RVO). In some embodiments, the eye disease is wet AMD. In some embodiments, the eye disease is diabetic macular edema (DME).

[0222] In some embodiments, the individualized dose is administered 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 days after the fourth dose. In some embodiments, the dosing is no sooner than the designated point in time. In some embodiments, the dosing is ±1, 2, 3, 4, 5, 6, or 7 days. The individualized dose(s) can be administered at any suitable dosing interval, e.g., upon determining that the subject has a decline in eye health based on evaluating the subject’s eye health.

[0223] Any suitable number of individualized doses can be administered to the subject under the recited dosing regimen. In some embodiments, the method includes administering, or administering at least, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or more individualized doses to the subject, or optionally a number of individualized doses that is in a range defined by any two of the preceding values (e.g., 1- 24, 2-12, 3-21, 4-10, 2-6, etc.). In some embodiments, no individualize dose is administered to the subject.

[0224] In some embodiments, the method achieves a therapeutic effect of the dual IL-6 / VEGF therapy at least by the last loading dose. In some embodiments, the method achieves a therapeutic effect of the dual IL-6 / VEGF therapy at least by Week 16, 20, 24, 28, 32, 36, 40, 44, or 48 from the first dose (e.g., first loading dose). In some embodiments, the therapeutic effect of the dual IL-6 / VEGF therapy includes an improvement in visual acuity. In some embodiments, the therapeutic effect of the dual IL-6 / VEGF therapy that is an improvement in visual acuity includes a BCVA increase by ETDRS letters of 3 letters or more, e.g., 4 letters or more, 5 letters or more, 6 letters or more, 7 letters or more, 8 letters or more, 9 letters or more, 10 letters or more, 12 letters or more, 15 letters or more, 18 letters or more, 20 letters or more, 22 letters, or more, including 25 letters or more compared to pre-treatment, or optionally an increase by a number within a range defined by any two of the preceding values (e.g., 3-25 letters, 4-20 letters, 5-15 letters, 3-18 letters, 4-10 letters, etc.) compared to pre-treatment. In some embodiments, the therapeutic effect of the dual IL-6 / VEGF therapyincludes a BCVA increase by ETDRS letters of, of about, or of at least 5, 10, or 15 letters. In some embodiments, the therapeutic effect of the dual IL-6 / VEGF therapy includes an improvement in retinal anatomical features. In some embodiments, the therapeutic effect of the dual IL-6 / VEGF therapy that is an improvement in retinal anatomical features includes a reduction, or at least a slowed increase, in retinal thickness (e.g., central subfield thickness). The retinal thickness may be measured using any suitable method, including, but not limited to, optical coherence tomography (OCT) or OCT-A. In some embodiments, the therapeutic result may include a reduction in retinal thickness of about 25 µm or more, e.g., about 30 µm or more, about 40 µm or more, about 50 µm or more, about 75 µm or more, about 100 µm or more, about 125 µm or more, about 150 µm or more, about 175 µm or more, about 200 µm or more, about 225 µm or more, about 250 µm or more, about 275 µm or more, about 300 µm or more, about 325 µm or more, about 350 µm or more, about 375 µm or more, about 400 µm or more compared to pre-treatment, or optionally a reduction within a range defined by any two of the preceding values (e.g., 25-400 µm, 30-300 µm, 40-250 µm, 30-200 µm, 40-375 µm, etc.) compared to pre-treatment.

[0225] In some embodiments, the method includes evaluating the subject’s eye health after the fourth dose (e.g., after the last loading dose). In some embodiments, the method includes evaluating the subject’s eye health after the fourth dose (or after the last loading dose) to determine if the subject has a decline in eye health. In some embodiments, the method includes evaluating the subject’s eye health no more frequently than about once every four weeks (or no more frequently than Q4W or QM). In some embodiments, the method includes evaluating the subject’s eye health no more frequently than once every 3-5 weeks. In some embodiments, the method includes evaluating the subject’s eye health about once every four weeks. In some embodiments, the method includes evaluating the subject’s eye health once every 4, 5, 6, 7, 8, 9, 10, 11, 12 weeks or less frequently, optionally evaluating the subject’s eye health at a frequency of once per a number of weeks in a range defined by any two of the preceding values (e.g., once per 4-12 weeks, once per 4-8 weeks, once per 4-6 weeks, etc.). In some embodiments, the method includes evaluating the subject’s eye health after administering an individualized dose. In some embodiments, the method includes evaluating the subject’s eye health after each administration of an individualized dose.

[0226] In some embodiments, the decline in eye health is based on the presence of intraretinal fluid (IRF), the presence of subretinal fluid (SRF), and / or new or worsening macular hemorrhage due to wet AMD activity. In some embodiments, the decline in eye health is based on the presence of intraretinal fluid (IRF). In some embodiments, the decline in eye health is based on the presence of subretinal fluid (SRF). In some embodiments, the decline in eye health is based on new or worsening macular hemorrhage due to wet AMD activity. In some embodiments, the decline in eye health is based on the presence of intraretinal fluid (IRF) and subretinal fluid (SRF).

[0227] In some embodiments, the decline in eye health comprises an increase in retina thickness. In some embodiments, the retina thickness is based on an optical coherence tomography central subfield thickness (OCT CST). In some embodiments, the decline in eye health comprises an increase in OCT CST compared to a previous measurement of OCT CST. In some embodiments, the increase in OCT CST is compared to a prior lowest measurement. In some embodiments, the subject’s eye health is evaluated by performing optical coherence tomography (OCT) and / or optical coherence tomography angiography (OCT-A).

[0228] In some embodiments, the method of treating a subject’s eye disease comprises evaluating the subject’s eye health and determining that the subject has new or worsening macular hemorrhage due to wet AMD activity. In some embodiments, the new or worsening macular hemorrhage due to wet AMD activity is determined based on a fundus photograph. In some embodiments, the evaluating the subject’s eye health includes taking a fundus photograph of the subject.

[0229] In some embodiments, the individualized dose is administered no more frequently than once every four weeks (+ / - 7 days). In some embodiments the individualized dose is administered 1, 2, 3, 4, 5, 6, or 7 days before or after the at least 4-weeks after the fourth dose.

[0230] Some non-limiting embodiments disclosed herein (with reference to Figure 32) are a method of treating a subject with wet AMD (3200), the method comprising administering to a subject in need thereof 4 loading doses of a fusion protein conjugate (e.g., KSI-501) comprising a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymercovalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody (3201). The method can also include administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after a last loading dose (3202). The method can further include determining, at least 4 weeks (+ / - 7 days) after a last loading dose, the presence of intraretinal fluid (IRF) and / or subretinal fluid (SRF) in an eye of the subject, and / or if macular hemorrhage due to wAMD activity has clinically worsened (3203). Further, if there is IRF or SRF in the eye, or the subject has clinically worsened macular hemorrhage due to wAMD activity, the method can include administering a maintenance dose of the fusion protein conjugate (3204).

[0231] In some embodiments, the decline in eye health is based on the presence of intraretinal fluid (IRF), the presence of subretinal fluid (SRF), and / or new or worsening macular hemorrhage due to wet AMD activity. In some embodiments, the method includes evaluating an IRF volume and / or SRF volume in an optical coherence tomography (OCT) scan of the subject to determine that the subject has the presence of intraretinal fluid (IRF) and / or subretinal fluid (SRF), respectively. In some embodiments, determining the presence of IRF and / or SRF in the subject’s eye comprises performing optical coherence tomography (OCT). In some embodiments, the subject is determined to have the presence of intraretinal fluid (IRF) and / or subretinal fluid (SRF) when the amount of fluid in the subject’s eye above predetermined normal levels (for example, around 5-20 nanoliters for IRF at 3mm, or 10-50 nanoliters for SRF at 3mm, or IRF exceeding 5-10 nL at 1mm, or SRF exceeding 10-25 nL at 1mm), e.g., as determined by OCT. The OCT scan can be analyzed using any suitable option. In some embodiments, the OCT scan or image is analyzed by automated image analysis.

[0232] In some embodiments, subjects in need of retreatment for wAMD can be identified by specific criteria indicating a decline in eye health. In some embodiments, subjects in need of retreatment for wAMD can be identified by an IRF exceeding 5-20 nL or SRF exceeding 10-50 nL at 3mm. In some embodiments, subjects in need of retreatment for wAMD can be identified by an IRF exceeding 5-10 nL or SRF exceeding 10-25 nL at 1mm, or CST- RPE / CST-BM increase above 25-100 μm. In some embodiments, subjects in need of retreatment for wAMD can be identified by a CST-RPE / CST-BM increase above 25-100 μm. In some embodiments, subjects in need of retreatment for wAMD can be identified by an IRFexceeding 5-20 nL or SRF exceeding 10-50 nL at 3mm or IRF exceeding 5-10 nL or SRF exceeding 10-25 nL at 1mm. In some embodiments, subjects in need of retreatment for wAMD can be identified by new onset of retinal hemorrhage from CNV. In some embodiments, criteria are assessed after the fourth dose of a fusion protein conjugate (e.g., KSI-501), and the criteria are compared to the prior lowest measurement. In some embodiments, the subject is determined to have the presence of intraretinal fluid (IRF) when the amount of fluid in an OCT IRF volume is above a predetermined normal level, optionally wherein the predetermined normal level is 5-20 nanoliters over a 3 mm diameter circle, or 5-10 nanoliters over a 1 mm diameter circle. In some embodiments, the subject is determined to have the presence of subretinal fluid (SRF) when the amount of fluid in an OCT SRF volume is above a predetermined normal level, optionally wherein the predetermined normal level is 10-50 nanoliters over a 3 mm diameter circle, or 10-25 nanoliters over a 1 mm diameter circle.

[0233] In some embodiments, a vision assessment, as measured by Best Corrected Visual Acuity (BCVA), increases by at least a threshold after administering at least the fourth dose relative to a baseline, wherein the threshold is 1-5 Early Treatment Diabetic Retinopathy Study (ETDRS) letters. In some embodiments, the threshold is 1, 2, 3, 4, or 5 ETDRS letters. In some embodiments, a vision assessment, as measured by Best Corrected Visual Acuity (BCVA), increases by at least 1 ETDRS letters after administering at least the fourth dose relative to a baseline. In some embodiments, a vision assessment, as measured by Best Corrected Visual Acuity (BCVA), increases by at least 4.5 ETDRS letters after administering at least the fourth dose relative to a baseline.

[0234] In some embodiments, the decline in eye health comprises an increase in central subfield thickness (CST) as measured by optical coherence tomography (OCT) of at least 25-50 microns after the fourth dose, compared to the prior lowest measurement. In some embodiments, the decline in eye health includes an increase in CST of, of about, or of at least 15, 20, 25, 30, 35, 40, 45, 50, 55, or 60 microns, or more, or optionally an increase in CST in a range defined by any two of the preceding values, for example, 15-60, 25-45, 25-40, 25-35, 25-30, 30-35, 30-40, 30-45, 30-50, 35-40, 35-50, 40-50, or 45-40 microns, compared to the prior lowest measurement.

[0235] In some embodiments, each dose (e.g., a first, second, third, fourth dose, or a loading dose, or an individualized dose, or a maintenance dose) of the fusion proteinconjugate comprises 1-6 mg of protein of the fusion protein conjugate. In some embodiments, the fusion protein conjugate comprises 1, 2, 3, 4, 5, or 6 mg of protein of the fusion protein conjugate, or a range defined by any two of the preceding values, for example, 1-5 mg, 1-4 mg, 1-3 mg, 1-2 mg, 2-5 mg, 2-4 mg, 2-3 mg, 3-4 mg, 3-5, or 4-5 mg. In some embodiments, each dose comprises about 5 mg of protein of the fusion protein conjugate. In some embodiments, each loading dose of the fusion protein conjugate includes the same or about the same amount (e.g., 5 mg) of the fusion protein conjugate (e.g., as measured based on the amount of protein). In some embodiments, each individualized or maintenance dose of the fusion protein conjugate includes the same or about the same amount (e.g., 5 mg) of the fusion protein conjugate (e.g., as measured based on the amount of protein). In some embodiments, each subsequent dose of the fusion protein conjugate includes the same or about the same amount (e.g., 5 mg) of the fusion protein conjugate (e.g., as measured based on the amount of protein).

[0236] In some embodiments, the fusion protein conjugate comprises a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti- IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody.

[0237] In some embodiments, the fusion protein conjugate comprises a fusion protein comprising an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti- IL-6 antibody conjugate, wherein the heavy chain comprises an Fc region comprising a non- native cysteine; and a phosphorylcholine containing polymer, wherein the polymer is covalently bonded to the fusion protein conjugate at a non-native cysteine outside a variable region of the fusion protein conjugate, wherein the fusion protein conjugate comprises a light chain and heavy chain, said heavy chain comprising an Fc region, wherein the cysteine is in the Fc region of the heavy chain, wherein the fusion protein conjugate has the following structure:, nd each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to the anti-IL-6 antibody through a sulfhydryl at C443 according to EU numbering, which bond is O CH3CH3depicted on one of the heavy chains above; PC , where the curvyline indicates the point of attachment to the rest n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%. In some embodiments, the sequence of the heavy chain comprises SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto, and wherein the sequence of the light chain comprises SEQ ID NO: 169, or a sequence at least 90% identical thereto.

[0238] After receiving the last dose (e.g., the final loading dose, or any dosing that occurred last), the subject may retain a therapeutic result of the fusion protein conjugatetherapy for a sustained period of time without the subject receiving a maintenance or individualized dose of the antibody conjugate. A therapeutic result of the fusion protein conjugate therapy may include an improvement in one or more of visual acuity, eye health, or retinal health (e.g., retinal thickness, extent of retinal perfusion, etc.) at or around the time of the final loading dose compared to before or at the time of the first loading dose. Any suitable therapeutic result of dual IL-6 / VEGF therapy may be used according to methods of the present disclosure. Suitable measures for determining therapeutic results include, e.g., visual acuity, retinal thickness, perfusion in at least one eye, diabetic retinopathy severity score (DRSS), disease activity of the eye disorder, the absence or a decrease in intraretinal fluid (IRF), subretinal fluid (SRF), or any combination thereof.

[0239] In some embodiments, the therapeutic result includes an improvement, or at least a slowed decline, in visual acuity. Visual acuity may be monitored using any suitable method. In some embodiments, the visual acuity is measured by best corrected visual acuity (BCVA) using e.g., ETDRS letters or Snellen chart, etc. In some embodiments, the therapeutic result may include an improvement in BCVA measured by ETDRS letters of 1 letter or more, e.g., 2 letters or more, 3 letters or more, 4 letters or more, 5 letters or more, 6 letters or more, 7 letters or more, 8 letters or more, 9 letters or more, 10 letters or more, 12 letters or more, 15 letters or more, 18 letters or more, 20 letters or more, 22 letters, or more, including 25 letters or more, or by a number within a range defined by any two of the preceding values, compared to pre-treatment. In some embodiments, the therapeutic result may include a reduction in the rate of deterioration of BCVA by at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage in a range defined by any two of the preceding values, over pre-treatment.

[0240] In some embodiments, the therapeutic result includes a reduction, or at least a slowed increase, in retinal thickness (e.g., central subfield thickness). The retinal thickness may be measured using any suitable method, including, but not limited to, optical coherence tomography (OCT) or OCT-A. In some embodiments, the therapeutic result may include a reduction in retinal thickness of about 25 µm or more, e.g., about 30 µm or more, about 40 µm or more, about 50 µm or more, about 75 µm or more, about 100 µm or more, about 125 µm or more, about 150 µm or more, about 175 µm or more, about 200 µm or more, about 225 µm or more, about 250 µm or more, about 275 µm or more, about 300 µm or more, about 325 µm ormore, about 350 µm or more, about 375 µm or more, about 400 µm or more, or a reduction within a range defined by any two of the preceding values, compared to pre-treatment. In some embodiments, the therapeutic result may include a reduction in the rate of increase in retinal thickness by at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage in a range defined by any two of the preceding values, over pre-treatment.

[0241] In some embodiments, the therapeutic result includes improved perfusion, or at least a reduction in the rate of expansion of non-perfusion, of the retina. Perfusion may be monitored using any suitable method. Suitable methods include, without limitation, OCT- angiography (OCT-A), fluorescein angiogram or ultrawide-field fluorescein angiogram. The degree of perfusion, or non-perfusion, may be measured using any suitable measure. In some embodiments, non-perfusion area or area of capillary non-perfusion is measured. In some embodiments, an ischemic index is calculated by dividing the non-perfusion area by the total retinal area. In some embodiments, the presence or absence of retinal non-perfusion in retinal quadrants on the angiogram is measured. In some embodiments, the therapeutic result may include a reduction in the area of non-perfusion of at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage in a range defined by any two of the preceding values, over pre-treatment. In some embodiments, the therapeutic result may include a reduction in the rate of progressive non-perfusion of at least 10%, e.g., at least 15%, at least 25%, at least 50%, at least 75%, at least 90%, including about 100%, or any percentage in a range defined by any two of the preceding values, over pre- treatment.

[0242] In some embodiments, the therapeutic result includes improved, or prevented worsening of, diabetic retinopathy severity score (DRSS). In some embodiments, the therapeutic result may include an improved DRSS of 2 steps or more, or 3 steps or more compared to pre-treatment. In some embodiments, the therapeutic result may include preventing worsening of DRSS by 2 steps or more, or 3 steps or more compared to pre- treatment.

[0243] The therapeutic result is retained if the level of visual acuity or retinal health (e.g., retinal thickness, presence or absence of IRF or SRF, etc.) does not worsen by more than a predetermined amount compared to the improved level. In some embodiments, thetherapeutic result is retained if the level of visual acuity or retinal health does not revert by 10% or more, e.g., 20% or more, 30% or more, 40% or more, 50% or more, 75% or more, 90% or more, including 100% or more to the pretreatment level of visual acuity or retinal health after the last dose (e.g., final loading dose). In some embodiments, the therapeutic result is retained if the rate of change of visual acuity or retinal health does not revert by 10% or more, e.g., 20% or more, 30% or more, 40% or more, 50% or more, 75% or more, 90% or more, including 100% or more to the pretreatment level of the rate of change of visual acuity or retinal health after the last dose (e.g., final loading dose).

[0244] In some embodiments, the therapeutic result includes an improvement in visual acuity. In some embodiments the therapeutic result may be retained if BCVA does not fall by 1 letter or more, 2 letters or more, 3 letters or more, 4 letters or more, 5 letters or more, 6 letters or more, 7 letters or more, 8 letters or more, 9 letters or more, or 10 letters or more from the BCVA score at the time of the final loading dose (e.g., at Week 12 after three monthly loading doses). In some embodiments the therapeutic result may be retained if BCVA does not fall by 1 letter or more, 2 letters or more, 3 letters or more, 4 letters or more, 5 letters or more, 6 letters or more, 7 letters or more, 8 letters or more, 9 letters or more, or 10 letters or more from the BCVA score measured at the last assessment (e.g., 4 weeks ago). In some embodiments the therapeutic result may be retained if BCVA does not fall by 1 letter or more, 2 letters or more, 3 letters or more, 4 letters or more, 5 letters or more, 6 letters or more, 7 letters or more, 8 letters or more, 9 letters or more, or 10 letters or more from the best measured BCVA score, or the average of the 2 best measured BCVA scores, of the subject.

[0245] In some embodiments, the therapeutic result includes a reduction in retinal thickness (e.g., central subfield thickness). In some embodiments, the therapeutic result may be retained if retinal thickness (e.g., central subfield thickness) does not increase by 25 µm or more, 30 µm or more, 40 µm or more, 50 µm or more, 75 µm or more, 100 µm or more, 125 µm or more, or 150 µm or more from the retinal thickness at the time of the last dose (e.g., final loading dose) (e.g., at Week 12 after three monthly loading doses). In some embodiments, the therapeutic result may be retained if retinal thickness (e.g., central subfield thickness) does not increase by 25 µm or more, 30 µm or more, 40 µm or more, 50 µm or more, 75 µm or more, 100 µm or more, 125 µm or more, or 150 µm or more from the retinal thickness measured at the last assessment (e.g., 4 weeks ago). In some embodiments, a retainedtherapeutic result includes retinal thickness that is not greater than 150 µm, 125 µm, 100 µm, 75 µm, 50 µm, 40 µm, or 30 µm, compared to the lowest measured retinal thickness of the subject.

[0246] In some embodiments, the therapeutic result includes improved perfusion of the retina. In patients with DR and DME, the retina can have an area of non-perfusion, or absence of blood flow. In some embodiments, non-perfusion is visualized on angiograms. In some embodiments, a therapeutic result of the fusion protein conjugate, e.g., KSI-501, administration, according to methods of the present disclosure, includes regression of non- perfusion, or re-perfusion of the retina. In some embodiments, the therapeutic result may be retained if the area of non-perfusion is reduced by 10% or more, e.g., 15% or more, 25% or more, 50% or more, 75% or more, 90% or more, or about 100% relative to the area of non- perfusion at the time of the last dose (e.g., final loading dose). In some embodiments, the therapeutic result may be retained if the area of non-perfusion is not increased by 10% or more, e.g., 15% or more, 25% or more, 50% or more, 75% or more, 90% or more, or about 100% relative to the area of non-perfusion at the time of the last dose (e.g., final loading dose). In some embodiments, the therapeutic result may be retained if the area of non-perfusion is not increased by 10% or more, e.g., 15% or more, 25% or more, 50% or more, 75% or more, 90% or more, or about 100% relative to the area of non-perfusion measured at the last assessment (e.g., 4 weeks ago). In some embodiments, the therapeutic result may be retained if the area of non-perfusion is not increased by 10% or more, e.g., 15% or more, 25% or more, 50% or more, 75% or more, 90% or more, or about 100% relative to the smallest area of non-perfusion measured in the subject.

[0247] In some embodiments, the method includes administering at least four loading doses of the fusion protein conjugate (e.g., KSI-501) to the subject, and administering one or more maintenance or individualized doses of the fusion protein conjugate, e.g., KSI- 501, after the final loading dose. The maintenance or individualized doses may be administered to the subject no more frequently than once every 4 weeks, e.g., every 6 weeks, every 8 weeks, every 10 weeks, every 12 weeks, every 14 weeks, every 16 weeks, every 18 weeks, every 20 weeks, or every 24 weeks. Whether to administer a maintenance or individualized dose of the fusion protein conjugate may be determined based on the presence of intraretinal fluid (IRF), subretinal fluid (SRF) and or new or clinically worsening macularhemorrhage due to wet AMD activity. In some embodiments, patients who do not have the presence of intraretinal fluid (IRF), subretinal fluid (SRF) and or new or clinically worsening macular hemorrhage due to wet AMD activity may receive treatment if there is an increase in OCT CST compared to prior lowest measurement. In some embodiments, the presence of new or clinically worsening macular hemorrhage due to wAMD activity is documented with a fundus photograph.

[0248] In some embodiments, the maintenance or individualized dose may be administered based on a predetermined schedule (e.g., a schedule determined before the subject is administered any of the one or more of the loading doses). In some embodiments, the maintenance or individualized dose may be administered based on a predetermined schedule based on the severity of the eye disorder, the subject’s previous response, or lack thereof, to other therapies for the eye disorder, or any other clinically relevant factors associated with the subject. In some embodiments, the maintenance or individualized dose may be administered based on the outcome of one or more assessment tests for ocular health and / or function carried out on the subject during the course of treatment with the fusion protein conjugate. In some embodiments, the maintenance or individualized dose may be administered based on the outcome of one or more assessment tests carried out on the subject every 4 or more weeks, e.g., every 6 or more weeks, every 8 or more weeks, every 10 or more weeks, every 12 or more weeks, every 16 or more weeks, every 20 or more weeks, every 24 or more weeks, every 28 or more weeks, every 32 or more weeks, every 36 or more weeks, including every 40 or more weeks. In some embodiments, the maintenance or individualized dose may be administered if one or more assessment tests indicates a diminishment of the therapeutic result of the fusion protein conjugate therapy that is greater than a predetermined threshold. In some embodiments, the maintenance or individualized dose may be administered if one or more assessment tests does not indicates a diminishment of the therapeutic result of the fusion protein conjugate therapy or the diminishment is not greater than a predetermined threshold

[0249] As used herein, “Q4W”, “QM” and the like each refers to a dosing schedule, and have the ordinary and customary meaning to one of ordinary skill in the art in view of the present disclosure. The number may indicate the number of the unit of time specified by the subsequent letter. “W” indicates a unit of a week; “M” specifies an interval of a month. Thus, Q4W refers to a dosing interval of 4 weeks, which also includes a dosing interval of one month;Q8W refers to a dosing interval of 8 weeks, which also includes a dosing interval of two months; and so on. QM refers to a dosing interval of one month. Q4W and QM are used interchangeably herein. As used herein, specification of a dosing schedule does not necessarily imply a number of doses beyond two, unless indicated otherwise. In some embodiments, a dosing schedule refers to the dosing schedule for maintenance doses (including the interval between the last loading dose, and the first maintenance dose). A reference to a dosing schedule being “longer” or “shorter” (e.g., “Q12W or longer”) refers to the time interval between doses being longer than that specified (e.g., a dosing interval of 12 weeks or longer).

[0250] In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-01) administered to the subject in the first year of treatment is 10 times or less, 9 times or less, 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. As used herein, “first year of treatment” and similar reference to the first “X” years of treatment denotes the time period measured from the first dose (e.g., first loading dose) of the fusion protein conjugate administered to the subject under the particular treatment regimen or schedule. In some embodiments, the time period is measured from the first dose (e.g., first loading dose) of the fusion protein conjugate administered to a treatment naïve subject (e.g., a subject who has not received an dual IL-6 / VEGF therapy before being administered the fusion protein conjugate of the present disclosure). In some embodiments, the time period is measured from the first dose (e.g., first loading dose) of the fusion protein conjugate administered to a subject who has previously been treated with an dual IL-6 / VEGF therapy (e.g., an dual IL-6 / VEGF therapy that does not include the fusion protein conjugate of the present disclosure) but has not responded to the dual IL-6 / VEGF therapy. In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject in the first two years of treatment is 10 times or less, 9 times or less, 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject in the first three years of treatment is 10 times or less, 9 times or less, 8 times or less,7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. In any method of the present disclosure, in some embodiments, at least 4 doses (e.g., loading doses and any maintenance or individualized doses) of the fusion protein conjugate is administered to the subject in the first year of treatment.

[0251] In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject in a one-year period for treatment is 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject in a two-year period for treatment is 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject in a three-year period for treatment is 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy.

[0252] In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject with wAMD in the first year of treatment is 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject with wAMD in the first two years of treatment is 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject with wAMD in the first three years of treatment is 8 times or less, 7 times or less, 6 times orless, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy.

[0253] In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with wAMD in a one-year period for treatment is 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with wAMD in a two-year period for treatment is 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with wAMD in a three-year period for treatment is 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy.

[0254] In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in the first year of treatment is 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in the first two years of treatment is 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of injections (including loading and maintenance doses) of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in the first three years of treatment is 8 times or less, 7 times or less, 6 times or less, 5 times or less, 4 times or less, 3 times or less, 2 times or less, or once, in order to retain the therapeutic result of the fusion protein conjugate therapy.

[0255] In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in a one-year period during treatment is 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments,the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in a two-year period during treatment is 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in a three-year period during treatment is 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy.

[0256] In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in a one-year period during treatment is 5 times or less, 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in a two-year period during treatment is 5 times or less, 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy. In some embodiments, the total number of maintenance doses of the fusion protein conjugate (e.g., KSI-501) administered to the subject with DME in a three-year period during treatment is 5 times or less, 4 times or less, 3 times or less, 2 times or less, once or less, or zero, in order to retain the therapeutic result of the fusion protein conjugate therapy.

[0257] In some embodiments, a first maintenance or individualized dose is administered at a first time after the last loading dose, and a second maintenance or individualized dose is administered at a second period of time after the first maintenance or individualized dose, where no other dose is administered between the last loading dose and the first maintenance or individualized dose, or between the first maintenance or individualized dose and the second maintenance or individualized dose. The second period of time between the first and second subsequent doses may be the same or different from the first period of time between the last loading dose and the first maintenance or individualized dose. In some embodiments, the first time period is 8 weeks or more, e.g., 10 weeks or more, 12 weeks or more, 14 weeks or more, 16 weeks or more, 18 weeks or more, 20 weeks or more, or 24 weeks or more. In some embodiments, the second period of time is longer than the first period of time by 0 weeks or more, e.g., by 4 weeks or more, by 6 weeks or more, by 8 weeks or more,by 10 weeks or more, by 12 weeks or more, by 16 weeks or more, by 20 weeks or more. The timing for administering the second maintenance or individualized dose may depend on the outcome of one or more assessments for ocular health and / or function of the subject.

[0258] Any suitable amount of the fusion protein conjugate (e.g., KSI-501), may be administered to the subject in loading, subsequent, individualized or maintenance dose. In some embodiments, the loading, subsequent, individualized or maintenance dose includes about 1 mg or more, e.g., about 1.25 mg or more, about 1.5 mg or more, about 1.75 mg or more, about 2 mg or more, about 2.5 mg or more, about 3 mg or more, about 3.5 mg or more, about 4 mg or more, about 4.5 mg of more, including about 5 mg or more (by weight of the fusion protein portion) of the fusion protein conjugate. In some embodiments, the loading, subsequent, individualized or maintenance dose includes from about 1 mg to about 10 mg, e.g., about 1 mg to about 7.5 mg, about 1.25 mg to about 5 mg, including about 2 mg to about 5 mg (by weight of the fusion protein portion) of the fusion protein conjugate.

[0259] As the therapeutic result of the dual IL-6 / VEGF therapy is retained for a sustained period of time after the last dose (e.g., final loading dose), the subject may not need to receive a dose of the fusion protein conjugate, while the therapeutic effect lasts. In some embodiments, no further administration of the fusion protein conjugate (e.g., KSI-501), is made to the subject within 4 weeks or more, e.g., within 6 weeks or more, within 8 weeks or more, within 10 weeks or more, within 12 weeks or more, within 14 weeks or more, within 16 weeks or more, within 20 weeks or more, within 24 weeks or more, within 28 weeks or more, within 32 weeks or more, within 36 weeks or more, within 40 weeks or more, within 44 weeks or more, within 48 weeks or more, including within 52 weeks or more, after the last dose (e.g., final loading dose).

[0260] In some embodiments, the route of administration is via an intravitreal injection. In some embodiments, a fusion protein conjugate (e.g., KSI-501) can be administered every 3 - 4 months, after a loading dose completion, or even less frequently.

[0261] Also provided herein is a method of treating wet age-related macular degeneration (wet AMD) that includes: identifying a subject with wet AMD; to the subject a first dose of a fusion protein conjugate comprising a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti- IL-6 antibody; administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose; administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose; and administering an individualized dose of the fusion protein conjugate to the subject at least 4 weeks (+ / - 7 days) after the fourth dose or about every 8 weeks thereafter, wherein each individualized dose is administered upon determining that there is a decline in the subject’s eye health, wherein the subsequent dose and the individualized dose are collectively administered no more frequently than about once every four weeks (or no more frequently than Q4W or QM), wherein the first, second, third, fourth doses, the subsequent dose and the individualized dose each comprises about 5 mg of protein of the fusion protein conjugate, wherein the sequence of the heavy chain comprises SEQ ID NO: 170 (with or without the C-terminal lysine), and wherein the sequence of the light chain comprises SEQ ID NO: 169, wherein the antibody conjugate has the following structure:, ch light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to the anti-IL- 6 antibody through a sulfhydryl at C443 according to EU numbering, which bond is depicted O CH3CH3on one of the heavy chains above; PC , where the curvy lineindicates the point of attachment to the n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%.

[0262] Also provided is a method of treating wet age-related macular degeneration (wet AMD) that includes: identifying a subject with wet AMD; to the subject a first dose of a fusion protein conjugate comprising a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containingpolymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody; administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose; administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose, wherein the first, second, third, fourth doses, the subsequent dose and the individualized dose each comprises about 5 mg of protein of the fusion protein conjugate, wherein the sequence of the heavy chain comprises SEQ ID NO: 170 (with or without the C-terminal lysine), and wherein the sequence of the light chain comprises SEQ ID NO: 169, wherein the antibody conjugate has the following structure: ,letter H, and each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to the anti-IL-6 antibody through a sulfhydryl at C443 according to EU numbering,which bond is depicted on one of the heavy chains above; PC i , where the curvy line indicates the point of attachment to the rest , n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%. Remote monitoring of treatment progress

[0264] In some embodiments, determining a subject’s eye health after administration of a fusion protein conjugate according to embodiments described herein includes the subject obtaining an image of the subject’s own retina using a suitable imaging device. In some embodiments, a device is provided herein, comprising an imaging device for remote monitoring of treatment progress in a subject wherein the subject has been, is, or will be treated with any of the compositions or methods described herein. In some embodiments, the device can comprise wearable goggles and / or a compact desktop model. In some embodiments, the device can assess retinal fluid volume and / or retinal thickness using traditional and / or machine learning algorithms. In some embodiments, the device can use scanning laser ophthalmoscopy (SLO) and / or optical coherence tomography (OCT) imaging modalities to capture images. In some embodiments, the device can be controlled remotely (i.e., by a physician or other healthcare professional). In some embodiments, the device can provide real-time image viewing and evaluation by a physician or other healthcare professional. In some embodiments, the device can detect potential bacterial infections. In some embodiments, the device can be used to detect bacterial infections within the first 24 hours following treatment with any of the compositions or methods described herein. In some embodiments, the device is used daily by a subject to capture images. In some embodiments, the device uploads the captured images to a cloud system for storage and / or analysis. In some embodiments, the device transmits the images and / or processed data to a physician or other healthcare professional. In some embodiments, the device provides diagnostic feedback to the subject regarding the uploaded images. In some embodiments, the physician or other healthcare professional can analyze the images and / or processed data and provide arecommendation regarding further course of treatment. In some embodiments, the subject has wet age-related macular degeneration (AMD).

[0265] In some embodiments, systems for remote monitoring of treatment progress in a subject, wherein the subject has been, is, or will be treated with any of the compositions or methods described herein are provided herein, comprising, a memory; at least one processor; and at least one non-transitory computer readable medium containing instructions that, when executed by the at least one processor, cause the at least one processor to perform operations comprising: capturing images for assessment of retinal fluid volume and retinal thickness, uploading the captured images to a cloud system, processing the images with traditional and / or machine learning algorithms to generate processed data, sending the processed data to a physician or other health care professional, diagnosing a subject based upon the processed data, providing the diagnosis directly to the subject, detecting a bacterial infection based upon the processed data, or any or all of the steps recited herein. In some embodiments, the subject has wet age-related macular degeneration (AMD).

[0266] In some embodiments, methods for remote monitoring of treatment progress in a subject, wherein the subject has been, is, or will be treated with any of the compositions or methods described herein are provided herein. In some embodiments, the methods comprise capturing images for assessment of retinal fluid volume and retinal thickness. In some embodiments, the methods comprise uploading the captured images to a cloud system. In some embodiments, the methods comprise processing the images with traditional and / or machine learning algorithms to generate processed data. In some embodiments, the methods comprise sending the processed data to a physician or other health care professional. In some embodiments, the methods comprise diagnosing a subject based upon the processed data. In some embodiments, the methods comprise providing the diagnosis directly to the subject. In some embodiments, the methods comprise retreatment of the subject with any of the compositions or methods described herein. In some embodiments, the methods comprise detection of a bacterial infection based upon the processed data. In some embodiments, capturing comprises the use of a device comprising wearable goggles or compact desktop model.FUSION PROTEINS AND CONJUGATES THEREOF

[0267] Provided herein are fusion proteins and conjugates thereof that find use in methods of the present disclosure. The fusion protein conjugate can include any suitable combination of the anti-IL-6 antibody, VEGF Trap and the phosphorylcholine-containing polymer, as provided herein. Suitable, non-limiting fusion proteins and conjugate thereof, and suitable, non-limiting methods of making the same, are provided in U.S. Pub. No. 2019 / 0270806, the entirety of which is incorporated herein by reference. In some embodiments, the fusion protein conjugate is KSI-501ABC. In some embodiments, the fusion protein is KSI-101. Fusion Proteins, Anti-IL6 Antibodies, VEGF Traps

[0268] Provided herein are anti-IL-6 antibodies that block, suppress or reduce (including significantly reduces) IL-6 biological activity, including downstream events mediated by IL-6. In some embodiments, the IL-6 antagonist antibody will have one or more of the CDR sequences provided herein.

[0269] In some embodiments, an isolated antagonist antibody specifically binds to IL-6. In some embodiments, the isolated antagonist antibody specifically binds to IL-6 having the amino acid sequence of SEQ ID NO:1. In some embodiments, the antibody preferably reacts with IL-6 in a manner that inhibits IL-6 signaling function. In some embodiments, the IL-6 antagonist antibody specifically binds primate IL-6. In some embodiments, the IL-6 antagonist antibody specifically binds human IL-6.

[0270] Also provided are antibodies that encompass monoclonal antibodies, polyclonal antibodies, antibody fragments (e.g., Fab, Fab’, F(ab’)2, Fv, Fc, etc.), chimeric antibodies, bispecific antibodies, heteroconjugate antibodies, single chain (ScFv), mutants thereof, fusion proteins comprising an antibody portion (e.g., a domain antibody), humanized antibodies, and any other modified configuration of the immunoglobulin molecule that comprises an antigen recognition site of the required specificity, including glycosylation variants of antibodies, amino acid sequence variants of antibodies, and covalently modified antibodies. The antibodies may be murine, rat, human, or any other origin (including chimeric or humanized antibodies). In some embodiments, the IL-6 antagonist antibody is a monoclonal antibody. In some embodiments, the antibody is a human or humanized antibody.

[0271] In some embodiments, the antibody comprises a heavy chain amino acid variable region as shown in Tables 1, 2, 6, 7, 8 and / or 9 or FIG. 15. In some embodiments, an isolated antagonist antibody comprises a heavy chain variable region (VH) comprising a VH complementarity determining region one (CDR1), VH CDR2, and VH CDR3 of the VH having an amino acid sequence of that shown in Tables 1, 2, 6, 7, 8 and / or 9, and a light chain variable region (VL) comprising a VL CDR1, VL CDR2, and VL CDR3 of the VL having an amino acid sequence of that shown in the table.

[0272] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided. The antibody comprises a heavy chain constant domain comprising one or more mutations to reduce effector function. In some embodiments, the one or more mutations reduce effector functions of the antibody related to the complement cascade, for example, a reduced activation of the complement cascade. In some embodiments, the reduction in effector function is at least about 50%.

[0273] In some embodiments, an isolated antagonist antibody that specifically binds to IL-6 comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the antibody comprises the mutations L234A, L235A, and G237A (based on EU numbering) is provided. In some embodiments, the isolated antagonist antibody comprises the mutations L234A, L235A, and G237A. In some embodiments, the isolated antagonist antibody with mutations has minimized binding to FC gamma receptors or C1q. In some embodiments, the isolated antagonist antibody with mutations L234A, L235A, and G237A has minimized binding to FC gamma receptors or C1q. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the mutation(s) is located at one or more of the following amino acid positions (EU numbering): E233, L234, L235, G236, G237, A327, A330, and P331. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the mutation (s) is selected from the group consisting of E233P, L234V, L234A, L235A, G237A, A327G, A330S, and P331S.

[0274] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided. The heavy chain constant domain further comprises a cysteine residue introduced by recombinant DNA technology (or a non-native cysteine). In some embodiments, the cysteine residue is selected from the group consisting of Q347C and L443C (EU numbering). In some embodiments, the cysteine residue is L443C (EU numbering).

[0275] In some embodiments, the antibody comprises all three of the following mutations (EU numbering) L234A, L235A, and G237A, and the antibody comprises L443C (EU numbering). In some embodiments, the antibody is a human IgG1, and a heavy chain constant domain of the antibody comprises one or more mutations that reduce an immune- mediated effector function.

[0276] In some embodiments, an isolated antagonist antibody is provided that binds an epitope on human IL-6 that is the same as or overlaps with the epitope recognized by an antibody comprising the amino acid sequences in any one or more of Tables: 1 and 2, and / or 6-9. In some embodiments, an IL-6 antibody with a cys and that is linked through that cysteine to a polymer is provided (as shown in Formula 17 or 17A herein).

[0277] In some embodiments, an isolated antagonist antibody that binds to IL-6 is provided. In some embodiments, the isolated antagonist antibody that binds to IL-6 comprises a heavy chain comprising the amino acid sequence shown in Tables 1, 2, 6, 7, 8 and / or 9, with or without the C-terminal lysine and a light chain comprising the amino acid sequence shown in Tables 1, 2, 6, 7, 8 and / or 9.

[0278] In some embodiments, an isolated antagonist antibody that binds to IL-6 is provided. The antibody comprises a VH comprising the amino acid sequence shown in Tables 1, 2, 6, 7, 8 and / or 9, or a sequence that is at least 90% identical thereto, having amino acid substitutions in residues that are not within a CDR. In some embodiments, the antibody comprises one or more of: HCDR1: in FIG. 15, HCDR2: in FIG. 15, HCDR3: in FIG. 15; LCDR1: in FIG. 15, LCDR2: in FIG. 15, LCDR3: in FIG. 15, for example, 1, 2, 3, 4, 5, or all 6 CDRs.

[0279] In some embodiments, an antibody that binds to IL-6 is provided, wherein the antibody comprises a CDRH1 that is the CDRH1 in Tables 1, 2, 6, 7, 8 and / or 9, a CDRH2 that is the CDRH2 in Tables 1, 2, 6, 7, 8 and / or 9, a CDRH3 that is the CDRH3 in Tables 1, 2, 6, 7, 8 and / or 9, a CDRL1 that is the CDRL1 in Tables 1, 2, 6, 7, 8 and / or 9, a CDRL2 that is the CDRL2 in Tables 1, 2, 6, 7, 8 and / or 9, a CDRL3 that is the CDRL3 in Tables 1, 2, 6, 7, 8 and / or 9, at least one of the following mutations: L234A, L235A, and G237A based on EU numbering, and at least one of the following mutations:Q347C or L443C based on EU numbering.

[0280] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided. The heavy chain variable region of the antibody comprises three complementarity determining regions (CDRs) comprising the amino acid sequences shown in Table 1. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the light chain variable region of the antibody comprises three complementarity determining regions (CDRs) comprising the amino acid sequences shown in Table 2. In some embodiments, the antibody is one that contains one or more of the identified sequences in FIG.15, e.g., one or more of the CDRs (including 2, 3, 4, 5 or 6 of the boxed CDRs) and / or the entire heavy and light chain variable regions.

[0281] In some embodiments, an isolated antagonist anti-IL-6 antibody comprises a heavy chain variable region (VH) that comprises three CDRs comprising the amino acid sequences shown in Table 1, and the light chain variable region (VL) of the antibody comprises three CDRs comprising the amino acid sequences shown in Table 2.

[0282] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided. The VH comprises the amino acid sequences shown in Table 1 and the light chain variable region of the antibody comprises three CDRs comprising the amino acid sequences shown in Table 2.

[0283] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody comprises a VL comprising the amino acid sequence shown in Table 2, or a variant thereof with one amino acid substitution in amino acids that are not within a CDR. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody comprises a VH comprising the amino acid sequence shown in Table 1, or a variant thereof with several amino acid substitutions in amino acids that are not within a CDR.

[0284] In some embodiments, an isolated antagonist antibody is provided, wherein the antibody comprises a heavy chain comprising the amino acid sequence shown in Table 1, with a C-terminal lysine, and a light chain comprising the amino acid sequence shown in Table 2. In some embodiments, an isolated antagonist antibody is provided, wherein the antibody comprises a heavy chain comprising the amino acid sequence shown in Table 1, without a C- terminal lysine, and a light chain comprising the amino acid sequence shown in Table 2.

[0285] The IL-6 antagonist antibodies may be made by any method known in the art. General techniques for production of human and mouse antibodies are known in the art and / or are described herein.

[0286] IL-6 antagonist antibodies can be identified or characterized using methods known in the art, whereby reduction, amelioration, or neutralization of IL-6 biological activity is detected and / or measured. In some embodiments, an IL-6 antagonist antibody is identified by incubating a candidate antibody with IL-6 and monitoring binding to IL-6R or IL-6 / IL-6R binding to gp130 and / or attendant reduction or neutralization of a biological activity of IL-6. The binding assay may be performed with, e.g., purified IL-6 polypeptide(s), or with cells naturally expressing (e.g., various strains), or transfected to express, IL-6 polypeptide(s). In one embodiment, the binding assay is a competitive binding assay, where the ability of a candidate antibody to compete with a known IL-6 antagonist antibody for IL-6 binding is evaluated. The assay may be performed in various formats, including the ELISA format.

[0287] Following initial identification, the activity of a candidate IL-6 antagonist antibody can be further confirmed and refined by bioassays, known to test the targeted biological activities. In some embodiments, an in vitro cell assay is used to further characterize a candidate IL-6 antagonist antibody.

[0288] IL-6 antagonist antibodies may be characterized using methods well known in the art. For example, one method is to identify the epitope to which it binds, or “epitope mapping.” There are many methods known in the art for mapping and characterizing the location of epitopes on proteins, including solving the crystal structure of an antibody-antigen complex, competition assays, gene fragment expression assays, and synthetic peptide-based assays, as described, for example, in Chapter 11 of Harlow and Lane, Using Antibodies, a Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 1999. In an additional example, epitope mapping can be used to determine the sequence to which an IL-6 antagonist antibody binds. IL-6 antagonist antibody Epitope mapping is commercially available from various sources, for example, Pepscan Systems (Edelhertweg 15, 8219 PH Lelystad, The Netherlands). The epitope can be a linear epitope, i.e., contained in a single stretch of amino acids, or a conformational epitope formed by a three-dimensional interaction of amino acids that may not necessarily be contained in a single stretch. Peptides of varying lengths (e.g., at least 4-6 amino acids long) can be isolated or synthesized (e.g.,recombinantly) and used for binding assays with an IL-6 antagonist antibody. In another example, the epitope to which the IL-6 antagonist antibody binds can be determined in a systematic screening by using overlapping peptides derived from the IL-6 sequence and determining binding by the IL-6 antagonist antibody. According to the gene fragment expression assays, the open reading frame encoding IL-6 is fragmented either randomly or by specific genetic constructions and the reactivity of the expressed fragments of IL-6 with the antibody to be tested is determined. The gene fragments may, for example, be produced by PCR and then transcribed and translated into protein in vitro, in the presence of radioactive amino acids. The binding of the antibody to the radioactively labeled IL-6 fragments is then determined by immunoprecipitation and gel electrophoresis. Certain epitopes can also be identified by using large libraries of random peptide sequences displayed on the surface of phage particles (phage libraries) or yeast (yeast display). Alternatively, a defined library of overlapping peptide fragments can be tested for binding to the test antibody in simple binding assays. In an additional example, mutagenesis of an antigen, domain swapping experiments and alanine scanning mutagenesis can be performed to identify residues required, sufficient, and / or necessary for epitope binding. For example, alanine scanning mutagenesis experiments can be performed using a mutant IL-6 in which various residues of the IL-6 polypeptide have been replaced with alanine. By assessing binding of the antibody to the mutant IL-6, the importance of the particular IL-6 residues to antibody binding can be assessed.

[0289] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody binds human IL-6 with an affinity of between about 0.01 pM to about 10 nM. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody binds human IL-6 with an affinity of between about 0.1 pM to about 2 nM. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody binds human IL-6 with an affinity of about 0.01, 0.05, 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95100, 200, 300, 400, 500, 600, 700, 800, 900, 1000 pM.

[0290] The binding affinity (KD) of an IL-6 antagonist antibody to IL-6 can be about 0.001 to about 200 nM. In some embodiments, the binding affinity is any of about 200 nM, about 100 nM, about 50 nM, about 10 nM, about 1 nM, about 500 pM, about 100 pM, about 60 pM, about 50 pM, about 20 pM, about 15 pM, about 10 pM, about 5 pM, about 2 pM,or about 1 pM. In some embodiments, the binding affinity is less than any of about 250 nM, about 200 nM, about 100 nM, about 50 nM, about 10 nM, about 1 nM, about 500 pM, about 100 pM, about 50 pM, about 20 pM, about 10 pM, about 5 pM, about 2 pM, about 1 pM, about 0.5 pM, about 0.1 pM, about 0.05 pM, about 0.01 pM, about 0.005 pM, or about 0.001 pM.

[0291] In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody binds human IL-6 with a koff that is at least 5.0E-03 at 37 degrees. In some embodiments the koff is 5E-04. In some embodiments, an isolated antagonist anti-IL-6 antibody is provided, wherein the antibody binds human IL-6 with a koff that is better than 5.0E-04 at 37 degrees.

[0292] In some embodiments, binding affinity can be defined in terms of one or more of association constant (ka), dissociation constant (kd), and analyte concentration that achieves half-maximum binding capacity (KD). In some embodiments, ka can range from about 0.50E+05 to about 5.00E+08. In some embodiments, kd can range from about 0.50E-06 to about 5.00E-03. In some embodiments, KDcan range from about 0.50E-12 to about 0.50E- 07.

[0293] In some embodiments, a pharmaceutical composition comprising any of the fusion proteins disclosed herein is provided. In some embodiments, a pharmaceutical composition comprising any of the fusion protein conjugates disclosed herein is provided. In some embodiments, a pharmaceutical composition comprising any of the fusion proteins disclosed herein and any of the conjugates disclosed herein is provided. In some embodiments, the pharmaceutical composition comprises one or more pharmaceutically acceptable carriers. In some embodiments, the pharmaceutical composition is a liquid. In some embodiments, the pharmaceutical composition has an endotoxin level less than about 0.2 EU / ml. In some embodiments, the pharmaceutical composition is a liquid and has an endotoxin level less than about 0.2 EU / ml. In some embodiments, the pharmaceutical composition is a liquid and has an endotoxin level less than about 2.0, 1, 0.5, 0.2 EU / ml. In some embodiments, for example in intravitreal injection, the endotoxin limit is 0.01-0.02 EU / injection / eye.

[0294] Some non-limiting embodiments provide any of the following, or compositions (including pharmaceutical compositions) comprising an antibody having a partial light chain sequence and a partial heavy chain sequence as found in Tables 1 and 2, orvariants thereof. In Tables 1 and 2, the underlined sequences are some embodiments of CDR sequences as provided herein. TABLE 1 Table 1. Anti-IL-6 heavy chain variable region sequences. CDRs are underlined. ID Sequence I EVQLVESGGGLVQPGGSLRLSCAASGFTFSPFAISWVRQAPGKGLEWVAKISPGGSWTYYSDTVTDRFTFSL FS FS FS FS FS FS FS FSTable 2. Anti-IL-6 light chain variable region sequences. CDRs are underlined. ID Sequence LT LT LT

[0295] In some embodiments, the antibody does not have one or more (or any) of the following CDRs, Tables 3, 4, and / or 5. TABLE 3 SequencetCDR1 SASISVSYMY h n(SEQ ID NO: 96)giiah CDR2 DMSNLAS(SEQ ID NO: 97) LTABLE 4 Sequence CDR1 PFAMS (SEQ ID NO: 244)TABLE 5 Sequence

[0296] In some embodiments, a composition as disclosed herein comprises an antibody having a partial or complete light chain sequence and a partial or complete heavy chain sequence from any of the options provided in Tables 1, 2, 6, 7, 8 and / or 9, or variants thereof. In some embodiments, the antibody (or binding fragment thereof) can include any one or more of the CDRs provided in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody (or binding fragment thereof) can include any three or more of the CDRs provided in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody (or binding fragment thereof) can include any all six of the CDRs provided in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the heavy and / or light chain can be any one or more of the other antibodyconstructs provided herein, including, for example, those provided in FIGs. 15, 19, and / or 22- 25B and Tables 1, 3, 4, 5, 6, 7, 8 and / or 9.

[0297] In some embodiments, a composition as disclosed herein comprises an antibody having a partial or complete light chain CDR sequence and a partial or complete heavy chain CDR sequence from any of the options provided in Tables 1, 2, 6, 7, 8 and / or 9.

[0298] In some embodiments, CDR portions of IL-6 antagonist antibodies are also provided. Determination of CDR regions is well within the skill of the art. It is understood that in some embodiments, CDRs can be a combination of the IMGT and Paratome CDRs (also termed “combined CDRs” or “extended CDRs”). Determination of CDRs is well within the skill of the art. In some embodiments, the CDRs are the IMGT CDRs. In other embodiments, the CDRs are the Paratome CDRs. In other embodiments, the CDRs are the extended, AbM, conformational, Kabat, or Chothia CDRs. In embodiments with more than one CDR, the CDRs may be any of IMGT, Paratome, extended, Kabat, Chothia, AbM, conformational CDRs, or combinations thereof. In some embodiments, other CDR definitions may also be used. In some embodiments, only residues that are in common between 2, 3, 4, 5, 6, or 7 of the above definitions are used (resulting in a shorter sequence). In some embodiments, any residue in any of 2, 3, 4, 5, 6, or 7 of the above definitions can be used (resulting in a longer sequence).

[0299] In some embodiments, an IL-6 antagonist antibody comprises three CDRs of any one of the heavy chain variable regions shown in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody comprises three CDRs of any one of the light chain variable regions shown in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody comprises three CDRs of any one of the heavy chain variable regions shown in Table 1, and three CDRs of any one of the light chain variable regions shown in Table 2. In some embodiments, the CDRs are one or more of those designated in Tables 6 and / or 7, or 8 and / or 9 below: Table 6 ANTI IL-6 HEAVY CHAIN CDR SEQUENCES. ID CDR1 CDR2 CDR3 D DID CDR1 CDR2 CDR3 IIa GFTFSPFAMH (SEQ ID NO: VAKISPGGSWTYYSDTVTD ARQAWGYYALDI (SEQ ID D ID D ID ID IDTable 7. ANTI IL-6 HEAVY CHAIN CDR SEQUENCES. Kabat CDR1 CDR2 CDR3 PFAMH SEQ ID NO 172) KISPGGSWTYYSDTVTD QAWGYYALDI SEQ ID NO. O. O. O. O. O. O.Table 8. ANTI IL-6 LIGHT CHAIN CDR SEQUENCES. ID CDR1 CDR2 CDR3 :ID CDR1 CDR2 CDR3 IV SASISVSYLY (SEQ ID NO: 108) LLIYDASSLAS (SEQ ID NO: QQWSGYPYT (SEQ ID NO: :Table 9 ANTI IL-6 LIGHT CHAIN CDR SEQUENCES. CDR1 CDR2 CDR3 SASISVSYLY SEQ ID NO. DDSSLAS SEQ ID NO.200) QQWSGYPYT SEQ ID NO. . .[ ] n some em o men s, e an o y use or n ng o - can e one a includes one or more of the sequences in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody used for binding to IL-6 can be one that includes three or more of the sequences in Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody used for binding to IL-6 can be one that includes six of the sequences in any one of Tables 1, 2, 6, 7, 8 and / or 9. In some embodiments, the antibody that binds to IL-6 can be one that competes for binding with an antibody that includes 6 of the specified CDRs in any one of Tables 1, 2, 6, 7, 8 and / or 9.

[0301] In some embodiments, the antibody can be linked or fused to a VEGF Trap sequence. In some embodiments, this Trap sequence can be as shown in Table 10. In some embodiments, the sequence is at least 80% identical to that shown in Table 10, e.g., at least 80, 85, 90, 95, 96, 97, 98, or 99% identical to that shown in Table 10. In some embodiments, any of the VEGF Trap molecules in U.S. Pub. No.20150376271 can be employed herein. In some embodiments, the VEGF Trap sequence is fused to an anti-IL-6 antibody in one of the following manners: to an N-terminal end of a heavy chain comprising IL-6 antibody VH (FIG. 16 left), or between a hinge region and after a CH1 domain of a heavy chain comprising IL-6 antibody VH (FIG. 16 right). Unless designated otherwise, both options in the alternative and together are contemplated for the embodiments provided herein wherein any Ab-Trap fusion is discussed. In some embodiments, the term “Trap” refers to a full length extracellular region or any portion thereof, or combination of portions from different VEGF receptors that can antagonize signaling between at least one VEGF and VEGFR. Preferably, the extracellular trapsegment includes at least one domain from one of VEGFR-1, -2 or -3, and more preferably at least two contiguous domains, such as D2 and D3. Optionally, an extracellular domain includes at least one domain from at least two different VEGFRs. A preferred extracellular domain comprises or consists essentially of D2 of VEGFR-1 and D3 of VEGFR-2. TABLE 10. VEGFR1, Domain 2 and VEGFR2, Domain 3 Fusion sequence SDTGRPFVEMYSEIPEIIHMTEGRELVIPCRVTSPNITVTLKKFPLDTLIPDGKRIIWDSRKGFIISNATYKEIGLLTCEA TVNGHLYKTNYLTHRQTNTIIDVVLSPSHGIELSVGEKLVLNCTARTELNVGIDFNWEYPSSKHQHKKLVNRDLKT QSGSEMKKFLSTLTIDGVTRSDQGLYTCAASSGLMTKKNSTFVRVHEK (SEQ ID NO: 114)

[0302] In some embodiments, the IL-6 Ab VEGF Trap construct can have any of the sequences provided in TABLE 11. In some embodiments, the construct can be at least identical to the sequences in Table 11, e.g., 80, 85, 90, 95, 96, 97, 98, or 99% identical, or higher. In some embodiments, the fusion protein can be in line with those percentages, with the exception that the antibody IL-6 domain does not contain one or more of the CDRs in Tables 3, 4, and / or 5. In some embodiments, the fusion protein is one that contains one or more of the identified sequences in FIG. 15, e.g., one of more of the CDRs (including 2, 3, 4, 5 or 6 of the boxed CDRs) and / or the entire heavy and light chain variable regions, along with a VEGF Trap sequence (e.g., Table 10). In some embodiments, the sequences can be directly fused to one another. In some embodiments, one or more flexible linking sequences or sections can be used. The linking sequence can be positioned between the Ab sequence and the VEGF Trap sequence. These sequences can be 5 to 30 amino acids in length. In some embodiments, the linking sequence can include G and S in a ratio of about 4:1. In some embodiments, the linker includes the following sequence: GGGGSGGGGS (SEQ ID NO: 115). In some embodiments, any flexible linker can be employed. In some embodiments, the Fc portion of the Il-6 Ab is IgG1. TABLE 11 Heavy and light chain sequences for dual inhibitor molecules. CDRs are underlined in the heavy and light chains, VEGF trap sequence is bolded in black, Gly-Ser linker is italicized. ID Heavy chain Light chain Q TI VID Heavy chain Light chain LSPSHGIELSVGEKLVLNCTARTELNVGIDFNWE AAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKV TL Q TI V V L Q TI V V L Q TIID Heavy chain Light chain KEIGLLTCEATVNGHLYKTNYLTHRQTNTIIDVV SSLQPEDFATYYCQQWSGYPYTFGQGTKVEIKRTV V L Q TI V V L Q TI V V LID Heavy chain Light chain G SDTGRPFVEMYSEIPEIIHMTEGRELVIPCRVTSP DIQLTQSPSSLSASVGDRVTITCSASISVSYLYWYQQ TI V V L Q TI V V L Q TI V V LID Heavy chain Light chain KSRWQQGNVFSCSVMHEALHNHYTQKSLSCSPGK Q TI V V L Q TI V V L Q TI V V LID Heavy chain Light chain VEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVD Q TI V V L Q TI V V L Q TI V V LID Heavy chain Light chain QPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDI Q TI V V L Q TI V V L Q TI V V LID Heavy chain Light chain TVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGd herein, DNA fragments encoding VH and VL regions described can first be obtained. Various modifications, e.g. mutations, deletions, and / or additions can also be introduced into the DNA sequences using standard methods known to those of skill in the art. For example, mutagenesis can be carried out using standard methods, such as PCR-mediated mutagenesis, in which the mutated nucleotides are incorporated into the PCR primers such that the PCR product contains the desired mutations or site-directed mutagenesis.

[0304] Also provided are modifications to the variable regions shown herein. For example, also provided are antibodies comprising functionally equivalent variable regions and CDRs which do not significantly affect their properties as well as variants which have enhanced or decreased activity and / or affinity. For example, the amino acid sequence may be mutated to obtain an antibody with the desired binding affinity to IL-6. Modification of polypeptides is routine practice in the art and need not be described in detail herein. Examples of modified polypeptides include polypeptides with conservative substitutions of amino acid residues, one or more deletions or additions of amino acids which do not significantly deleteriously change the functional activity, or which mature (enhance) the affinity of the polypeptide for its ligand, or use of chemical analogs.

[0305] Amino acid sequence insertions include amino- and / or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intra-sequence insertions of single or multiple amino acid residues. Examples of terminal insertions include an antibody with an N-terminal methionyl residue or the antibody fused to an epitope tag. Other insertional variants of the antibody molecule include the fusion to the N- or C-terminus of the antibody of an enzyme or a polypeptide which increases the half-life of the antibody in the blood circulation.

[0306] Substitution variants have at least one amino acid residue in the antibody molecule removed and a different residue inserted in its place. The sites of greatest interest for substitutional mutagenesis include the hypervariable regions, but framework alterations arealso contemplated. Conservative substitutions are shown in Table 12 under the heading of “conservative substitutions.” If such substitutions result in a change in biological activity, then more substantial changes, denominated “exemplary substitutions” in Table 12, or as further described below in reference to amino acid classes, may be introduced and the products screened. Table 12 - Amino Acid Substitutions Original Residue Conservative Substitutions Exemplary Substitutions Ala (A) Val Val; Leu; Ile a;

[0307] Substantial modifications in the biological properties of the antibody are accomplished by selecting substitutions that differ significantly in their effect on maintaining (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a β-sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the targetsite, or (c) the bulk of the side chain. Naturally occurring residues are divided into groups based on common side-chain properties: (1) Non-polar: Norleucine, Met, Ala, Val, Leu, Ile; (2) Polar without charge: Cys, Ser, Thr, Asn, Gln; (3) Acidic (negatively charged): Asp, Glu; (4) Basic (positively charged): Lys, Arg; (5) Residues that influence chain orientation: Gly, Pro; and (6) Aromatic: Trp, Tyr, Phe, His.

[0308] Non-conservative substitutions are made by exchanging a member of one of these classes for another class.

[0309] One type of substitution, for example, that may be made is to change one or more cysteines in the antibody, which may be chemically reactive, to another residue, such as, without limitation, alanine or serine. For example, there can be a substitution of a non- canonical cysteine. The substitution can be made in a CDR or framework region of a variable domain or in the constant region of an antibody. In some embodiments, the cysteine is canonical. Any cysteine residue not involved in maintaining the proper conformation of the antibody also may be substituted, generally with serine, to improve the oxidative stability of the molecule and prevent aberrant cross-linking. Conversely, cysteine bond(s) may be added to the antibody to improve its stability, particularly where the antibody is an antibody fragment such as an Fv fragment.

[0310] The antibodies may also be modified, e.g. in the variable domains of the heavy and / or light chains, e.g., to alter a binding property of the antibody. Changes in the variable region can alter binding affinity and / or specificity. In some embodiments, no more than one to five conservative amino acid substitutions are made within a CDR domain. In other embodiments, no more than one to three conservative amino acid substitutions are made within a CDR domain. For example, a mutation may be made in one or more of the CDR regions to increase or decrease the KDof the antibody for IL-6, to increase or decrease koff, or to alter the binding specificity of the antibody. Techniques in site-directed mutagenesis are well-known in the art. See, e.g., Sambrook et al. and Ausubel et al., supra.

[0311] According to an aspect, the IgG domain of an IL-6 antagonist antibody or fusion protein can be IgG1, IgG2, IgG3 or IgG4. According to another aspect, the IgG domaincan be a composite in which a constant regions is formed from more than one of the above isotypes (e.g., CH1 region from IgG2 or IgG4, hinge, CH2 and CH3 regions from IgG1). In choosing an isotype, it is known in the art that human isotopes IgGl and IgG3 have complement-mediated cytotoxicity whereas human isotypes IgG2 and IgG4 have poor or no complement-mediated cytotoxicity. In some embodiments the IL-6 antagonist antibody isotype is IgG1.

[0312] The light chain constant region can be either human lambda or kappa. In some embodiments, the IL-6 antagonist antibody has a human kappa light chain constant region.

[0313] Human constant regions show allotypic variation and isoallotypic variation between different individuals, that is, the constant regions can differ in different individuals at one or more polymorphic positions. Isoallotypes differ from allotypes in that sera recognizing an isoallotype binds to a non-polymorphic region of one or more other isotypes. Reference to a human constant region includes a constant region with any natural allotype or any permutation of residues occupying polymorphic positions in natural allotypes or up to 3, 5 or 10 substitutions for reducing or increasing effector function as described below.

[0314] One or several amino acids at the amino or carboxy terminus of the light and / or heavy chains such as the C-terminal lysine of the heavy chain, may be missing or derivatized in a proportion or all of the molecules. In some embodiments, the fusion protein does not include a C-terminal lysine in the heavy chain.

[0315] Substitutions can be made in the constant regions to reduce or increase effector function such as complement-mediated cytotoxicity (CDC), antibody-dependent cell- mediated cytotoxicity (ADCC) (see, e.g., Winter et al., US Patent No.5,624,821; Tso et al., US Patent No. 5,834,597; and Lazar et al., Proc. Natl. Acad. Sci. USA 103:4005, 2006), or to prolong half-life in humans (see, e.g., Hinton et al., J. Biol. Chem. 279:6213, 2004).

[0316] In some embodiments, the IL-6 antagonist antibodies provided herein include one more substitutions that reduce complement mediated cytotoxicity. Reduction in complement mediated cytotoxicity can be accomplished with or without reduction in Fc receptor binding depending on the nature of the mutation(s). Antibodies with reduced complement mediated cytotoxicity but little or no reduction in Fc receptor allow a desired effect of Fc-mediated phagocytosis of iC3b without activating complement, which maycontribute to side effects. Exemplary mutations known to reduce complement-mediated cytotoxicity in human constant regions include mutations at positions 241, 264, 265, 270, 296, 297, 322, 329 and 331 by EU numbering. Mutations in positions 318, 320, and 322 have been reported to reduce complement activation in mouse antibodies. Alanine is a preferred residue to occupy these positions in a mutated constant region. Some exemplary human mutations that have been used include F241A, V264A, D265A, V296A, N297A, K322A, and P331S in human IgG3 and D270A or E, N297Q, K322A, P329A, and P331S in human IgGl (EU numbering).

[0317] Here, as elsewhere, the EU numbering scheme is used for numbering amino acids in the constant region of an antibody. When a residue in a variable region is referenced herein (unless designated otherwise) the residue numbering is according to the variable domain (or, if designated, the SEQ ID NO). Substitution at any or all of positions 234, 235, 236 and / or 237 reduce affinity for Fcγ receptors, particularly FcγRI receptor and also reduces complement binding and activation (see, e.g., US 6,624,821, WO / 2009 / 052439). An alanine substitution at positions 234, 235 and 237 reduces effector functions, particularly in the context of human IgGl. Optionally, positions 234, 236 and / or 237 in human IgG2 are substituted with alanine and position 235 with glutamine. (See, e.g., US 5,624,821) to reduce Fc receptor binding. Exemplary substitutions for increasing half-life include a Gln at position 250 and / or a Leu at position 428. Also provided are aspects where the anti-IL-6 antibody presented has a human IgG1 isotype, it is preferred that the antibody has at least one mutation in the constant region. Preferably, the mutation reduces complement fixation or activation by the constant region. Also provided are particularly preferred aspects, where the antibody has one or more mutations at positions E233, L234, L235, G236, G237, A327, A330 and P331 by EU numbering. Still more preferably, the mutations constitute one or more of the following E233P, L234V, L234A, L235A, G237A, A327G, A330S and P331S by EU numbering. In the most preferred embodiments the human IgG1 has the following mutations L234A, L235A and G237A by EU numbering. Fusion Protein Conjugates

[0318] In some embodiments, the half-life of IL-6 antagonist antibodies and / or IL- 6 Ab VEGF Traps can be extended by attachment of a “half-life extending moieties” or “half- life extending groups,” which terms are herein used interchangeably to refer to one or morechemical groups attached to one or more amino acid site chain functionalities such as -SH, - OH, -COOH, -CONH2, -NH2, or one or more N- and / or O-glycan structures and that can increase in vivo circulatory half-life of proteins / peptides when conjugated to these proteins / peptides. Examples of half-life extending moieties include polymers described herein, particularly those of zwitterionic monomers, such as HEMA-phosphorylcholine, PEG, biocompatible fatty acids and derivatives thereof, Hydroxy Alkyl Starch (HAS) e.g. Hydroxy Ethyl Starch (HES), Poly Ethylene Glycol (PEG), Poly (Glyx-Sery) (HAP), Hyaluronic acid (HA), Heparosan polymers (HEP), Fleximers, Dextran, Poly-sialic acids (PSA), Fc domains, Transferrin, 25 Albumin, Elastin like (ELP) peptides, XTEN polymers, PAS polymers, PA polymers, Albumin binding peptides, CTP peptides, FcRn binding peptides and any combination thereof.

[0319] In some embodiments, the antibody is conjugated with a phosphorylcholine containing polymer. In some embodiments, the antibody is conjugated with a poly(acryloyloxyethyl phosphorylcholine) containing polymer, such as a polymer of acrylic acid containing at least one acryloyloxyethyl phosphorylcholine monomer such as 2- methacryloyloxyethyl phosphorylcholine (i.e., 2-methacryloyl-2’-trimethylammonium ethyl phosphate).

[0320] In some embodiments, the antibody and / or antibody VEGF Trap fusion is conjugated with a water-soluble polymer, which refers to a polymer that is soluble in water. A solution of a water-soluble polymer may transmit at least about 75%, more preferably at least about 95% of light, transmitted by the same solution after filtering. On a weight basis, a water- soluble polymer or segment thereof may be at least about 35%, at least about 50%, about 70%, about 85%, about 95% or 100% (by weight of dry polymer) soluble in water.

[0321] In one embodiment, a half-life extending moiety can be conjugated to an IL-6 antagonist antibodies and / or IL-6 Ab VEGF Trap via free amino groups of the protein using N-hydroxysuccinimide (NHS) esters. Reagents targeting conjugation to amine groups can randomly react to ^-amine group of lysines, α-amine group of N-terminal amino acids, and δ-amine group of histidines.

[0322] However, also provided are IL-6 antagonist antibodies which have many amine groups available for polymer conjugation. Conjugation of polymers to free amino groups, thus, might negatively impact the ability of the antibody to bind to the epitope.

[0323] In another embodiment, a half-life extending moiety is coupled to one or more free SH groups using any appropriate thiol-reactive chemistry including, without limitation, maleimide chemistry, or the coupling of polymer hydrazides or polymer amines to carbohydrate moieties of the IL-6 antagonist antibodies and / or IL-6 Ab VEGF Traps after prior oxidation. The use of maleimide coupling is a particularly preferred embodiment. Coupling preferably occurs at cysteines naturally present or introduced via genetic engineering.

[0324] In some embodiments, polymers are covalently attached to cysteine residues introduced into IL-6 antagonist antibodies and / or IL-6 Ab VEGF Traps by site directed mutagenesis. In some embodiments, the cysteine residues in the Fc portion of the IL- 6 antagonist antibody and / or IL-6 Ab VEGF Trap can be used. In some embodiments, sites to introduce cysteine residues into an Fc region are provided in WO 2013 / 093809, US 7,521,541, WO 2008 / 020827, US 8,008,453, US 8,455,622 and US2012 / 0213705, incorporated herein by reference for all purposes. In some embodiments, cysteine mutations are Q347C and L443C referring to the human IgG heavy chain by the EU index of Kabat. In some embodiments, the cysteine added by directed mutagenesis for subsequent polymer attachment is L443C. In some embodiments, the stoichiometry of IL-6 antagonist antibody to polymer is 1:1; in other words, a conjugate consists essentially of molecules each comprising one molecule of IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap conjugated to one molecule of polymer. In some embodiments, coupling can occur at one or more lysines.

[0325] In some embodiments, a conjugate comprises an isolated antagonist antibody that specifically binds to IL-6 or IL-6 Ab VEGF Trap is conjugated to a polymer. In some embodiments, the polymer comprises a zwitterionic monomer. In some embodiments, the zwitterionic monomer, without limitations, is HEMA-phosphorylcholine, PEG, biocompatible fatty acids and derivatives thereof, Hydroxy Alkyl Starch (HAS) e.g. Hydroxy Ethyl Starch (HES), Poly Ethylene Glycol (PEG), Poly (Glyx-Sery) (HAP), Hyaluronic acid (HA), Heparosan polymers (HEP), Fleximers, Dextran, Poly-sialic acids (PSA), Fc domains, Transferrin, 25 Albumin, Elastin like (ELP) peptides, XTEN polymers, PAS polymers, PA polymers, Albumin binding peptides, CTP peptides, or FcRn binding peptides. In some embodiments, the polymer comprising a zwitterionic monomer is a half-life extending moiety.

[0326] In some embodiments, the IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap can have a half-life extending moiety attached. In some embodiments, the half-lifeextending moiety is a zwitterionic polymer but PEG or other half-life extenders discussed below can alternatively be used. In some embodiments, the zwitterionic polymer is formed of monomers having a phosphorylcholine group. In some embodiments, the monomer is 2- (acryloyloxyethyl)-2’-(trimethylammoniumethyl) phosphate. In some embodiments, the monomer is 2-(methacryloyloxyethyl)-2’-(trimethylammoniumethyl) phosphate (HEMA-PC).

[0327] In some embodiments, the polymer conjugated to the IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap has at least 2 or 3 or more arms. Some polymers have 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 arms. In some embodiments, the polymer has 3, 6 or 9 arms. In some embodiments, the polymer has 9 arms. In some embodiments, the polymer peak molecular weight is between 300,000 and 1,750,000 Da. In some embodiments, the polymer has a peak molecular weight between 500,000 and 1,000,000 Da. In some embodiments, the polymer has a peak molecular weight between 600,000 to 800,000 Da.

[0328] In some embodiments, a conjugate of antagonistic anti-IL-6 antibody and / or IL-6 Ab VEGF Trap and a polymer is provided. In some embodiments, the polymer has a peak molecular weight between 300,000 and 1,750,000 Daltons as measured by size exclusion chromatography – multi angle light scattering (hereinafter “SEC-MALS”). In some embodiments, the polymer has a peak molecular weight between 500,000 and 1,000,000 Daltons as measured by SEC-MALS. In some embodiments, the polymer has a peak molecular weight between 600,000 to 800,000 Daltons as measured by SEC-MALS.

[0329] In some embodiments, a half-life extending moiety may be conjugated to a naturally occurring cysteine residue of an IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap provided herein. In some embodiments, the half-life extending moiety is conjugated to a cysteine that is added via site directed mutagenesis. In some embodiments, the cysteine is added by using recombinant DNA technology to add the peptide SGGGC or CAA to the C- terminus of either the light or heavy chain. In some embodiments, the peptide is added to the heavy chain. In some embodiments, the cysteine residue introduced via recombinant DNA technology is selected from the group consisting of (EU numbering) Q347C and L443C.

[0330] Also provided is, a pharmaceutical composition presented having an IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap and a pharmaceutically acceptable excipient.

[0331] IL-6 antagonist antibodies and / or IL-6 Ab VEGF Traps (or other constructs provided herein) can be produced by recombinant expression including (i) the production ofrecombinant DNA by genetic engineering, (ii) introducing recombinant DNA into prokaryotic or eukaryotic cells by, for example and without limitation, transfection, electroporation or microinjection, (iii) cultivating the transformed cells, (iv) expressing anti-IL-6 antibodies and / or IL-6 Ab VEGF Traps, e.g. constitutively or on induction, and (v) isolating the anti-IL- 6 antibody and / or IL-6 Ab VEGF Trap, e.g. from the culture medium or by harvesting the transformed cells, in order to (vi) obtain purified anti-IL-6 antibody and / or IL-6 Ab VEGF Trap.

[0332] In some embodiments, a conjugate comprising an anti-IL-6 antibody and / or IL-6 Ab VEGF Trap and a polymer is provided (as well as the other constructs provided herein, such as a bioconjugate). The antibody and / or Ab-Trap is any one of the antibodies and / or Ab- Traps disclosed herein. In some embodiments, the antibody is an IgG. In some embodiments, the antibody is IgA, IgE, IgD or IgM. In some embodiments, the polymer is covalently bonded to a sulfhydryl group. In some embodiments, the polymer is covalently bonded to a sulfhydryl group from a cysteine residue. In some embodiments, the polymer is covalently bonded to a sulfhydryl group from a cysteine residue on the heavy chain. In some embodiments, the polymer is covalently bonded to a sulfhydryl group from a cysteine residue on the heavy chain of the IgG. In some embodiments, the antibody comprises a cysteine residue at position 347 or 443 (EU numbering). In some embodiments, the polymer is covalently bonded to a sulfhydryl group from a cysteine residue at position 347 or 443 (EU numbering).

[0333] IL-6 antagonist antibodies and / or IL-6 Ab VEGF Trap can be produced by expression in a suitable prokaryotic or eukaryotic host system characterized by producing a pharmacologically acceptable anti-IL-6 antibody molecule. Examples of eukaryotic cells are mammalian cells, such as CHO, COS, HEK 293, BHK, SK-Hip, and HepG2. Other suitable expression systems are prokaryotic (e.g., E. coli with pET / BL21 expression system), yeast (Saccharomyces cerevisiae and / or Pichia pastoris systems), and insect cells. In some embodiments, an isolated cell line that produces any of the antibodies and / or Ab-Traps disclosed herein is provided. In some embodiments, the isolated cell line is selected, without limitations, from one or more of CHO, k1SV, XCeed, CHOK1SV, GS-KO.

[0334] In some embodiments, an isolated nucleic acid encoding any of the antibodies and / or Ab-Traps disclosed herein is provided. In some embodiments, a recombinantexpression vector comprising the isolated nucleic acid is provided. In some embodiments, a host cell comprises the expression vector.

[0335] A wide variety of vectors can be used for the preparation of the IL-6 antagonist antibodies and / or IL-6 Ab VEGF Traps and may be selected from eukaryotic and prokaryotic expression vectors. Examples of vectors for prokaryotic expression include plasmids such as, and without limitation, preset, pet, and pad, wherein the promoters used in prokaryotic expression vectors include one or more of, and without limitation, lac, trc, trp, recA, or araBAD. Examples of vectors for eukaryotic expression include, without limitation: (i) for expression in yeast, vectors such as, and without limitation, pAO, pPIC, pYES, or pMET, using promoters such as, and without limitation, AOX1, GAP, GAL1, or AUG1; (ii) for expression in insect cells, vectors such as and without limitation, pMT, pAc5, pIB, pMIB, or pBAC, using promoters such as and without limitation PH, p10, MT, Ac5, OpIE2, gp64, or polh, and (iii) for expression in mammalian cells, vectors such as, and without limitation, pSVL, pCMV, pRc / RSV, pcDNA3, or pBPV, and vectors derived from, in one aspect, viral systems such as and without limitation vaccinia virus, adeno-associated viruses, herpes viruses, or retroviruses, using promoters such as and without limitation CMV, SV40, EF-1, UbC, RSV, ADV, BPV, and beta-actin.

[0336] In some embodiments, a method of producing an IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap is provided. In some embodiments, the method comprises culturing a cell line that recombinantly produces any of the antibodies and / or Ab-Traps disclosed herein under conditions wherein the antibody is produced and recovered. In some embodiments, a method of producing an IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap is provided. In some embodiments, the method comprises culturing a cell line comprising nucleic acid encoding an antibody and / or Ab-Trap comprising a heavy chain comprising the amino acid sequence shown in Table 1 and a light chain comprising the amino acid sequence shown in Table 2 under conditions wherein the antibody and / or IL-6 Ab VEGF Trap is produced and recovered. In some embodiments, the heavy and light chains of the antibody and / or IL-6 Ab VEGF Trap are encoded on separate vectors. In some embodiments, the heavy and light chains of the antibody and / or IL-6 Ab VEGF Trap are encoded on the same vector.

[0337] Also provided are methods for synthesizing a zwitterionic polymer-IL-6 antagonist antibody and / or IL-6 Ab VEGF Trap conjugates, the conjugate having one or morefunctional agents and one or more polymer arms wherein each of the polymer arms has one or more monomer units wherein at least one of the units has a zwitterion. For example, such a method can have the steps of: a. providing an initiator having one or more sites for monomer polymerization and a first linker having an amine group wherein the initiator is a trifluoro acetic acid salt; b. providing one or more monomers suitable for polymerization wherein at least one of the monomers is zwitterionic; c. reacting the monomers with the initiator to form one or more polymer arms each corresponding to the sites for monomer polymerization to provide an initiator-polymer conjugate having the first linker with the amine group; d. providing a second linker having at least second and third reactive groups; e. coupling one of the second and third reactive groups of the second linker to the amine group of the first linker of the initiator-polymer conjugate to provide a linker-initiator-polymer conjugate having one or more reactive groups that were not used in the coupling step; and f. coupling one or more functional agents to one or more of the unreacted reactive groups of the linker-initiator-polymer moiety to provide the polymer- functional agent conjugate.

[0338] In some embodiments, a conjugate comprising an isolated antagonist antibody and / or IL-6 Ab VEGF Trap that specifically binds to IL-6 and a phosphorylcholine- containing polymer is provided. In some embodiments, the polymer is covalently bonded to the antibody and / or Ab-Trap. In some embodiments, the polymer is non-covalently bonded to the antibody and / or Ab-Trap.

[0339] In some embodiments, a conjugate comprising an anti-IL-6 antibody and / or IL-6 Ab VEGF Trap and a phosphorylcholine containing polymer is provided. In some embodiments, the polymer is covalently bonded to the antibody outside a variable region of the antibody. In some embodiments, a conjugate comprising an anti-IL-6 antibody and / or IL- 6 Ab VEGF Trap and a phosphorylcholine containing polymer is provided. The polymer is covalently bonded to the antibody at a cysteine outside a variable region of the antibody. Insome embodiments, a conjugate comprising an anti-IL-6 antibody and / or IL-6 Ab VEGF Trap and a phosphorylcholine containing polymer is provided, wherein the polymer is covalently bonded to the antibody at a cysteine outside a variable region of the antibody wherein said cysteine has been added via recombinant DNA technology (or is a non-native cysteine). In some embodiments of the conjugate, the polymer comprises 2(methacryloyloxy)ethyl (2- (trimethylammonio)ethyl) phosphate (MPC) monomers.

[0340] Also provided is a method where the conjugation group (e.g. maleimide) is added after polymer synthesis. This is sometimes referred to as a “snap-on strategy” or “universal polymer strategy”. See, e.g., U.S. Patent Application No.14 / 916,180 (published as U.S. Patent Application Publication No. 20160199501), hereby incorporated by reference in its entirety. In some embodiments, a single initiator moiety can be used for large scale polymer synthesis. Thus, conditions can be developed for scaled up optimal polymer synthesis. Such polymers can then be adapted to various types of functional agents by “snapping-on” various types of linkers. For example, if it is desired to conjugate a larger functional agent to a polymer provided herein such as an antibody of even a Fab fragment, a longer linker sequence can be snapped on to the polymer. In contrast, smaller functional agents may call for relatively shorter linker sequences.

[0341] In some embodiments of the methods, the initiator has about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 sites for polymer initiation. In some embodiments, the initiator has about 3, about 6, or about 9 sites for polymer initiation.

[0342] Also provided herein is a second linker which has second, third, fourth, fifth, and sixth reactive groups. More preferably, a second linker has just second and third reactive groups.

[0343] Also provided herein, each polymer arm has from about 20 to about 2000 monomer units. Preferably, each arm has from about 100 to 500 monomer units or from about 500 to 1000 monomer units or from about 1000 to 1500 monomer units or from about 1500 to 2000 monomer units.

[0344] Also provided herein, the peak molecular weight of the polymer-functional agent conjugate is about 100,000 to 1,500,000 Da. Preferably, the peak molecular weight of the polymer-functional agent conjugate is about 200,000 to about 300,000 Da, about 400,000 to about 600,000 Da or about 650,000 to about 850,000 Da.

[0345] Also provided herein, the first linker is preferably alkyl, substituted alkyl, alkylene, alkoxy, carboxyalkyl, haloalkyl, cycloalkyl, cyclic alkyl ether, alkenyl, alkenylene, alkynyl, alkynylene, cycloalkylene, heterocycloalkyl, heterocycloalkylene, aryl, arylene, arylene-oxy, heteroaryl, amino, amido or any combination thereof. More preferably, the first linker has the formula:wherein m is 1 to 10. In some embodiments, the first linker has the above formula (Formula (1)) and m is 4.

[0346] In some embodiments, the initiator preferably includes a structure selected from group consisting of,CH H C 3 3 Xwherein X is selected from the group consisting of NCS, F, Cl, Br and I. More preferably, X in Formula (2), Formula (3) and / or Formula (4) is Br.

[0347] In some embodiments, the monomer is selected from the group consisting of,Formula (9) wherein R7 is H or C1-6 alkyl and t is 1 to 6.

[0348] More preferably, the monomer is selected from the group consisting of 2- (methacryloyloxyethyl)-2’-(trimethylammoniumethyl) phosphate (HEMA-PC) and 2- (acryloyloxyethyl)-2’-(trimethylammoniumethyl) phosphate.

[0349] Most preferably, the monomer is 2-(methacryloyloxyethyl)-2’- (trimethylammoniumethyl) phosphate.

[0350] The second linker moiety preferably comprises an activated ester having the structurewherein R8 is selected from the group consisting ofpolymer has 9 arms, m is 2-4, R9 iswherein p is 4 to 15. Still more preferably, m is 4 and p is 12.

[0352] In some embodiments, the radically polymerizable monomer isormu a ( ) wherein R1 is H or C1-6 alkyl, R2, R3, R4 are the same or different and are H or C1-4alkyl and X and Y are the same or different and are integers from 1-6. In some embodiments, R1, R2, R3 and R4 are each methyl and X and Y are each 2 in Formula (12).

[0353] In some embodiments, the radically polymerizable monomer isFormula (13) wherein R1 is H or C1-6alkyl, R2 and R3 are the same or different and are H or C1-4alkyl, R4 is PO4-, SO3- or CO2- and X and Y are the same or different and are integers from 1-6. Insome embodiments, R1, R2 and R3 are methyl, R4 is PO4- and X and Y are each 2 in Formula (13).

[0354] In some embodiments, the monomer iswherein R1 is H or C1-6alkyl, R2, R3 and R4 are the same or different and are H or C1-4alkyl, R5 is PO4-, SO3- or CO2- and X and Y are the same or different and are integers from 1-6. In some embodiments, R1, R2, R3 and R4 are methyl, R5 is PO4- and X and Y are 2 in Formula (14).

[0355] When a polymer is the to be conjugated via a cysteine (or other specified residue), the polymer can be linked directly or indirectly to the residue (e.g., with an intervening initiator, and or spacer or the like).

[0356] In some embodiments, the phosphorylcholine containing polymer comprises 2-(methacryloyloxyethyl)-2’-(trimethylammonium)ethyl phosphate (MPC) monomers as set forth below:, such that the polymer comprises the following repeating units:where n is an integer from 1 to 3000 and the wavy lines indicate the points of attachment between monomer units in the polymer.

[0357] In some embodiments, the polymer has three or more arms, or is synthesized with an initiator comprising 3 or more polymer initiation sites. In some embodiments, the polymer has 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 arms, or is synthesized with an initiator comprising 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 polymer initiation sites. More preferably, the polymer has 3, 6, or 9 arms, or is synthesized with an initiator comprising 3, 6, or 9 polymer initiation sites.In some embodiments, the polymer has 9 arms, or is synthesized with an initiator comprising 9 polymer initiation sites.

[0358] In some embodiments, the polymer that is added has a molecular weight between about 300,000 and about 1,750,000 Da (e.g., SEC-MALs). In some embodiments, the polymer has a molecular weight between about 500,000 and about 1,000,000 Da. In some embodiments, the polymer has a molecular weight of between about 600,000 to about 900,000 Da. In some embodiments, the polymer has a molecular weight of between about 750,000 to about 850,000 Da. In some embodiments, the polymer has a molecular weight of between about 800,000 to about 850,000 Da. In some embodiments, the polymer has a molecular weight of between about 750,000 to about 800,000 Da.

[0359] In some embodiments, any of the antibodies and / or Ab-Traps described herein can be further conjugated to a polymer to form a bioconjugate. The molecular weight of the bioconjugate (in total, SEC-MALs) can be between about 350,000 and 2,000,000 Daltons, for example, between about 450,000 and 1,900,000 Daltons, between about 550,000 and 1,800,000 Daltons, between about 650,000 and 1,700,000 Daltons, between about 750,000 and 1,600,000 Daltons, between about 850,000 and 1,500,000 Daltons, between about 900,000 and 1,400,000 Daltons, between about 950,000 and 1,300,000 Daltons, between about 900,000 and 1,000,000 Daltons, between about 1,000,000 and 1,300,000 Daltons, between about 850,000 and 1,300,000 Daltons, between about 850,000 and 1,000,000 Daltons, and between about 1,000,000 and 1,200,000 Daltons. In some embodiments, the bioconjugate has a molecular weight between about 350,000 and 1,900,000 Daltons.

[0360] In some embodiments, the antibody and / or Ab-Trap conjugate is purified. In some embodiments, the polymer in aspect of the antibody and / or Ab-Trap conjugate is polydisperse, i.e. the polymer PDI is not 1.0. In some embodiments, the PDI is less than 1.5. In some embodiments, the PDI is less than 1.4. In some embodiments, the PDI is less than 1.3. In some embodiments the PDI is less than 1.2. In some embodiments the PDI is less than 1.1. In some embodiments, the conjugate PDI is equal to or less than 1.5.

[0361] In some embodiments, the antibody and / or Ab-Trap conjugate has an anti- IL-6 immunoglobulin G (IgG) bonded to a polymer, which polymer comprises MPC monomers, wherein the sequence of the anti-IL-6 heavy chain is in Table 1, and the sequence of the anti-IL-6 light chain is in Table 2, and wherein the antibody and / or Ab-Trap is bondedonly at C442 to the polymer. In some embodiments, the polymer has 9 arms and has a molecular weight of between about 600,000 to about 1,000,000 Da.

[0362] In some embodiments, the antibody and / or Ab-Trap conjugate has an anti- IL-6 immunoglobulin G (IgG) bonded to a polymer, which polymer comprises MPC monomers, wherein the sequence of the anti-IL-6 heavy chain comprises in Table 1, and the sequence of the anti-IL-6 light chain comprises in Table 2, and wherein the antibody is bonded only at C443 (EU numbering) to the polymer. In some embodiments, the polymer has 9 arms and has a molecular weight of between about 600,000 to about 1,000,000 Da. In some embodiments, the conjugate comprises a polymer that has 9 arms and the polymer has a molecular weight of between about 600,000 to about 900,000 Da.

[0363] In some embodiments, the fusion protein conjugate has the following structure: O O X PC n2 PC PC = P N(CH ) O O O CH O PC O X n5 PCwhere each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to the antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains; PC , where the curvy line indicates the point of attachment to the restis: a) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) – H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS; and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%. In some embodiments, –X is –Br.

[0364] In some embodiments, the fusion protein conjugate has the following structure:+ N(CH3)3PC O X n5 PCwherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H, and each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to the anti-IL- 6 antibody (and / or Ab-Trap) through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains; PC iswhere –X is: a) –OR where R is isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, and n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15% wherein the VEGF Trap is fused: to the N-terminal end of the heavy chain; orbetween a hinge region and a Fab region (after the CH1 domain) of the heavy chain. In some embodiments, n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different and are integers from 0 to 3000. In some embodiments, the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is about 1500 to about 3500 plus or minus about 10% to about 20%. In some embodiments, n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different and are integers from 0 to 500. In some embodiments, –X is –Br. In some embodiments, X is OR, where R is a sugar, an aminoalkyl, mono-substituted, poly-substituted or unsubstituted variants of the following residues: saturated C1 -C24 alkyl, unsaturated C2 -C24 alkenyl or C2 -C24 alkynyl, acyl, acyloxy, alkyloxycarbonyloxy, aryloxycarbonyloxy, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, heteroaryl, arylalkoxy carbonyl, alkoxy carbonylacyl, amino, aminocarbonyl, aminocarboyloxy, nitro, azido, phenyl, hydroxy, alkylthio, arylthio, oxysulfonyl, carboxy, cyano, and halogenated alkyl including polyhalogenated alkyl, --CO--O--R7, carbonyl --CCO- -R7, --CO--NR8R9, --(CH2)n--COOR7, --CO--(CH) n--COOR7, --(CH2) n--NR8R9, ester, alkoxycarbonyl, aryloxycarbonyl, wherein n is an integer from 1 to 6, wherein each R7, R8and R9 is separately selected from the group consisting of a hydrogen atom, halogen atom, mono- substituted, poly-substituted or unsubstituted variants of the following residues: saturated C1- C24alkyl, unsaturated C2-C24alkenyl or C2- C24alkynyl, acyl, acyloxy, alkyloxycarbonyloxy, aryloxycarbonyloxy, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, heteroaryl, arylalkoxy carbonyl, alkoxy carbonylacyl, amino, aminocarbonyl, aminocarboyloxy, nitro, azido, phenyl, hydroxy, alkylthio, arylthio, oxysulfonyl, carboxy, cyano, and halogenated alkyl including polyhalogenated alkyl, a 5-membered ring, and a 6-membered ring. In some embodiments, Formula 17 can be part of a fusion protein, which would further include some or all of the sequence of Table 10, so as to make an IL-6 Ab VEGF Trap fusion protein. In some embodiments, the fusion protein in FIG. 27 can be used within Formula 17, with the fusion protein replacing the depicted antibody.

[0365] In some embodiments, the antibody and / or Ab-Trap conjugate is present in a liquid formulation. In some embodiments, the antibody and / or Ab-Trap conjugate is combined with a pharmaceutically acceptable carrier.

[0366] In some embodiments, the isotype of the anti-IL-6 antibody (and / or IL-6 Ab-VEGF Trap) heavy chain, is IgG1 and has a CH1, hinge, CH2and CH3domains. In some embodiments the light chain isotype is kappa.

[0367] In some embodiments, the IgG1 domain of the anti-IL-6 antibody (and / or IL-6 Ab-VEGF Trap) has one or more mutations to modulate effector function, such as ADCC, ADCP, and CDC. In some embodiments, the IgG1 mutations reduce effector function. In some embodiments the amino acids to use for effector function mutations include (EU numbering) E233X, L234X, L235X, G236X, G237X, G236X, D270X, K322X, A327X, P329X, A330X, A330X, P331X, and P331X, in which X is any natural or non-natural amino acid. In some embodiments, the mutations include one or more of the following: E233P, L234V, L234A, L235A, G237A, A327G, A330S and P331S (EU numbering). In some embodiments, the anti-IL-6 antibody (and / or IL-6 Ab-VEGF Trap) heavy chain has the following mutations (EU numbering): L234A, L235A and G237A. In some embodiments, the number of effector function mutations relative to a natural human IgG1 sequence is no more than 10. In some embodiments the number of effector function mutations relative to a natural human IgG1 sequence is no more than 5, 4, 3, 2 or 1. In some embodiments, the antibody (and / or IL-6 Ab-VEGF Trap) has decreased Fc gamma binding and / or complement C1q binding, such that the antibody’s ability to result in an effector function is decreased. This can be especially advantageous for ophthalmic indications / disorders.

[0368] In some embodiments, the anti-IL-6 antibody (and / or IL-6 Ab-VEGF Trap) comprises one or more of the following amino acid mutations: L234A, L235A, G237A, and L443C (EU numbering, or 451A, 452A, and 454A, and 660C in SEQ ID NO: 170, as shown in double underlining in FIG. 27).

[0369] In some embodiments, the anti-IL-6 antibody (and / or IL-6 Ab VEGFTrap) is or is part of a human immunoglobulin G (IgG1).

[0370] In some embodiments, the IL-6 antibody (and / or IL-6 Ab-VEGF Trap) comprises a heavy chain constant domain that comprises one or more mutations that reduce an immune-mediated effector function.

[0371] In some embodiments, the anti-IL-6 heavy chain has a cysteine residue added as a mutation by recombinant DNA technology which can be used to conjugate a half- life extending moiety. In some embodiments, the mutation is Q347C (EU numbering) and / or L443C (EU numbering). In some embodiments, the mutation is L443C (EU numbering). In some embodiments, the stoichiometry of antibody to polymer is 1:1; in other words, a conjugate has one molecule of antibody conjugated to one molecule of polymer.

[0372] The half-life of the anti-IL-6 antibodies can be extended by attachment of a “half-life (“half life”) extending moieties” or “half-life (“half life”) extending groups”. Half- life extending moieties include peptides and proteins which can be expressed in frame with the biological drug of issue (or conjugated chemically depending on the situation) and various polymers which can be attached or conjugated to one or more amino acid side chain or end functionalities such as -SH, -OH, -COOH, -CONH2, -NH2, or one or more N- and / or O-glycan structures. Half-life extending moieties generally act to increase the in vivo circulatory half- life of biologic drugs.

[0373] Examples of peptide / protein half-life extending moieties include Fc fusion (Capon DJ, Chamow SM, Mordenti J, et al. Designing CD4 immunoadhesions for AIDS therapy. Nature. 1989. 337:525-31), human serum albumin (HAS) fusion (Yeh P, Landais D, Lemaitre M, et al. Design of yeast-secreted albumin derivatives for human therapy: biological and antiviral properties of a serum albumin-CD4 genetic conjugate. Proc Natl Acad Sci USA. 1992.89:1904-08 ), carboxy terminal peptide (CTP) fusion (Fares FA, Suganuma N. Nishimori K, et al. Design of a long-acting follitropin agonist by fusing the C-terminal sequence of the chorionic gonadotropin beta subunit to the follitropin beta subunit. Proc Natl Acad Sci USA. 1992. 89:4304-08), genetic fusion of non-exact repeat peptide sequence (XTEN) fusion (Schellenberger V, Wang CW, Geething NC, et al. A recombinant polypeptide extends the in vivo half-life of peptides and proteins in a tunable manner. Nat Biotechnol.2009.27:1186-90), elastin like peptide (ELPylation) (MCpherson DT, Morrow C, Minehan DS, et al. Production and purification of a recombinant elastomeric polypeptide, G(VPGVG19-VPGV, from Escheriachia coli. Biotechnol Prog. 1992. 8:347-52), human transferrin fusion (Prior CP, Lai C-H, Sadehghi H et al. Modified transferrin fusion proteins. Patent WO2004 / 020405. 2004), proline-alanine-serine (PASylation) (Skerra A, Theobald I, Schlapsky M. Biological active proteins having increased in vivo and / or vitro stability. Patent WO2008 / 155134 A1. 2008), homo-amino acid polymer (HAPylation) (Schlapschy M, Theobald I, Mack H, et al. Fusion of a recombinant antibody fragment with a homo-amino acid polymer: effects on biophysical properties and prolonged plasma half-life. Protein Eng Des Sel. 2007. 20:273-84) and gelatin like protein (GLK) fusion (Huang Y-S, Wen X-F, Zaro JL, et al. Engineering a pharmacologically superior form of granulocyte-colony-stimulating-factor by fusion with gelatin-like protein polymer. Eur J. Pharm Biopharm. 2010. 72:435-41).

[0374] Examples of polymer half-life extending moieties include polyethylene glycol (PEG), branched PEG, PolyPEG® (Warwick Effect Polymers; Coventry, UK), polysialic acid (PSA), starch, hydroxylethyl starch (HES), hydroxyalkyl starch (HAS), carbohydrate, polysaccharides, pullulane, chitosan, hyaluronic acid, chondroitin sulfate, dermatan sulfate, dextran, carboxymethyl-dextran, polyalkylene oxide (PAO), polyalkylene glycol (PAG), polypropylene glycol (PPG), polyoxazoline, polyacryloylmorpholine, polyvinyl alcohol (PVA), polycarboxylate, polyvinylpyrrolidone, polyphosphazene, polyoxazoline, polyethylene-co-maleic acid anyhydride, polystyrene-co-maleic acid anhydride, poly(1- hydroxymethyethylene hydroxymethylformal) (PHF), a zwitterionic polymer, a phosphorylcholine containing polymer and a polymer comprising MPC, Poly (Glyx-Sery), Hyaluronic acid (HA), Heparosan polymers (HEP), Fleximers, Dextran, and Poly-sialic acids (PSA).

[0375] In one embodiment a half-life extending moiety can be conjugated to an antibody via free amino groups of the protein using N-hydroxysuccinimide (NHS) esters. Reagents targeting conjugation to amine groups can randomly react to ^-amine group of lysines, α-amine group of N-terminal amino acids, and δ-amine group of histidines.

[0376] However, the anti-IL-6 antibodies (and / or IL-6 Ab-VEGF Trap) disclosed herein can have many amine groups available for polymer conjugation. Conjugation of polymers to free amino groups, thus, might negatively impact the ability of the antibody proteins to bind to IL-6 (and / or IL-6 Ab-VEGF Trap).

[0377] In some embodiments, a half-life extending moiety is coupled to one or more free SH groups using any appropriate thiol-reactive chemistry including, without limitation, maleimide chemistry, or the coupling of polymer hydrazides or polymer amines to carbohydrate moieties of the antibody after prior oxidation. In some embodiments maleimide coupling is used In some embodiments, coupling occurs at cysteines naturally present or introduced via genetic engineering.

[0378] In some embodiments, conjugates of fusion proteins and high MW polymers serving as half-life extenders are provided. In some embodiments, a conjugate comprises an antibody (and / or IL-6 Ab-VEGF Trap) that is coupled to a zwitterionic polymer wherein the polymer is formed from one or more monomer units and wherein at least one monomer unit has a zwitterionic group is provided. In some embodiments, the zwitterionicgroup is phosphorylcholine. In some embodiments, 50%, 60%, 70%, 80% or 90% of the antibody is conjugated.

[0379] In some embodiments, one of the monomer units is HEMA-PC. In some embodiments, a polymer is synthesized from a single monomer which is HEMA-PC.

[0380] In some embodiments, some antibody (and / or IL-6 Ab VEGFTrap) conjugates have 2, 3, or more polymer arms wherein the monomer is HEMA-PC. In some embodiments, the conjugates have 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 polymer arms wherein the monomer is HEMA-PC. In some embodiments, the conjugates have 3, 6 or 9 arms. In some embodiments, the conjugate has 9 arms.

[0381] In some embodiments, polymer-antibody (and / or polymer-IL-6 Ab- VEGFTrap) conjugates have a polymer portion with a molecular weight of between 100,000 and 1,500,000 Da. In some embodiments, the conjugate has a polymer portion with a molecular weight between 500,000 and 1,000,000 Da. In some embodiments, the conjugate has a polymer portion with a molecular weight between 600,000 to 800,000 Da. In some embodiments, the conjugate has a polymer portion with a molecular weight between 600,000 and 850,000 Da and has 9 arms. When a molecular weight is given for an antibody conjugated to a polymer, the molecular weight will be the addition of the molecular weight of the protein, including any carbohydrate moieties associated therewith, and the molecular weight of the polymer.

[0382] In some embodiments, an anti-IL-6 antibody (and / or IL-6 Ab VEGF Trap) has a HEMA-PC polymer which has a molecular weight measured by Mw of between about 100 kDa and 1650 kDa is provided. In some embodiments, the molecular weight of the polymer as measured by Mw is between about 500 kDa and 1000 kDa. In some embodiments, the molecular weight of the polymer as measured by Mw is between about 600 kDa to about 900 kDa. In some embodiments, the polymer molecular weight as measured by Mw is 750 or 800 kD...

Claims

WHAT IS CLAIMED IS:

1. A method of treating diabetic macular edema (DME), comprising: identifying a subject having DME; and administering one or more loading doses of a fusion protein conjugate, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non- native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non- native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the loading doses of the fusion protein conjugate for at least 4 or 8 weeks after a final loading dose, optionally wherein the one or more loading doses comprise no more than 5 loading doses.

2. A method of treating an eye disorder, comprising: identifying a subject having an eye disorder; and administering at least one loading dose of a fusion protein conjugate to the subject, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non- native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non- native cysteine in the Fc region of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of administering the at least one loading dose of the fusion protein conjugate for at least 8 weeks after a final loading dose.

3. The method of claim 2, wherein the eye disorder comprises diabetic macular edema (DME), wet age-related macular degeneration (wAMD), retinal vein occlusion (RVO), diabetic retinopathy (DR), macular edema secondary to inflammation (MESI), and / or uveitis.

4. The method of claim 2, wherein the eye disorder is a VEGF therapy-resistant eye disorder.

5. The method of claim 2, comprising administering 2-5 loading doses of the fusion protein conjugate, optionally wherein the method comprises administering the loading doses no more frequently than Q4W.

6. The method of claim 5, wherein the loading dose comprises from 1 mg to 6 mg of the fusion protein conjugate (by weight of the fusion protein) per eye.

7. The method of claim 5, wherein the loading dose comprises from 25 to 65 mg / ml of the fusion protein conjugate (by weight of the fusion protein).

8. The method of claim 1, wherein the therapeutic result comprises one or more of improved visual acuity, reduced retinal thickness, improved retinal perfusion, improved diabetic retinopathy severity score (DRSS), or reduced disease activity, compared to a pre- treatment level.

9. The method of claim 1, comprising administering one or more subsequent doses of the fusion protein conjugate to the subject after a final loading dose, optionally wherein no subsequent dose is administered to the subject within about 8 weeks of a final loading dose.

10. The method of claim 1, wherein the VEGF Trap is positioned either: at an N-terminal end of a heavy chain variable region of the anti-IL-6 antibody; or between a hinge region and after a CH1 domain of a heavy chain comprising the heavy chain variable region of the anti-IL-6 antibody.

11. The method of claim 1, wherein the fusion protein comprises: a first polypeptide comprising an amino acid sequence at least 80% identical to SEQ ID NO: 170; and a second polypeptide comprising an amino acid sequence at least 80% identical to SEQ ID NO:

169.

12. The method of claim 1, wherein the polymer has a molecular weight between 300,000 and 1,750,000 Daltons.

13. The method of claim 10, wherein the fusion protein conjugate comprises the following structure:, wherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to a heavy chain of the anti-IL-6 antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains; O CH3PC i , where the curvy line indicates the point ofattachment to the rest of the polymer, where –X is: a) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, andn1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%.

14. The method of claim 1, wherein the administering comprises intravitreal administration.

15. A method of treating diabetic macular edema (DME), comprising: identifying a subject having DME; and administering 3 or 4 intravitreal loading doses of a fusion protein conjugate to the subject at Q4W, wherein the fusion protein conjugate comprises: a fusion protein comprising: an anti-IL-6 antibody comprising a heavy chain and a light chain; and a VEGF Trap fused to the heavy chain, wherein the heavy chain fused to the VEGF Trap comprises an amino acid sequence as set forth in SEQ ID NO: 170 (with or without the C-terminal lysine), and the light chain comprises an amino acid sequence as set forth in SEQ ID NO: 169, wherein the fusion protein conjugate comprises the following structure:, wherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H and is fused at the N-terminal end to the VEGF Trap, and each light chain of the anti-IL-6 antibody is denoted by the letter L, the polymer is bonded to the heavy chain of the anti-IL-6 antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains, O CH3PC i , where the curvy line indicates the point ofattachment to the rest of the polymer, where –X is: a) –OR where R is H, methyl, ethyl, propyl, isopropyl; b) –H; or c) any halogen, including –Br, –Cl, or –I; d) –SCN; or e) –NCS, andn1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%, wherein each dose comprises from 1 mg to 6 mg of the fusion protein conjugate (by weight of the fusion protein) per eye, whereby the subject retains a therapeutic result of administering the 3 or 4 intravitreal loading doses of the fusion protein conjugate for at least 4, 8, 12, 16, 20, 24, or more weeks after the final loading dose.

16. The method of claim 2, wherein the administering comprises administering to the subject a therapeutic formulation comprising: an unconjugated fusion protein comprising the fusion protein with or without the non-native cysteine in the Fc region, wherein the unconjugated fusion protein is not covalently attached to a phosphorylcholine-containing polymer; and the fusion protein conjugate, wherein the unconjugated fusion protein is present in the formulation in a range of 10-60% of a total molar amount of the fusion protein conjugate and the unconjugated fusion protein, wherein the total molar amount is the sum of the molar amount of the fusion protein conjugate and the molar amount of the unconjugated fusion protein.

17. A method of treating a subject for an inflammatory condition, comprising: administering an intravitreal or intravenous dose of a fusion protein conjugate to a subject in need thereof; and administering one or more subsequent intravitreal or intravenous doses of the fusion protein conjugate no more frequently than Q2W, wherein the fusion protein conjugate comprises: a fusion protein comprising a VEGF Trap fused to a heavy chain of an anti-IL- 6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody, wherein each of the intravitreal dose and the one or more subsequent intravitreal doses comprises from 1 to 6 mg of the fusion protein conjugate (by weight of the fusion protein), or wherein each of the intravenous dose and the one or more subsequent intravenous dosescomprises from 1 to 6 mg / kg body weight of the fusion protein conjugate (by weight of the fusion protein).

18. A method of treating wet age-related macular degeneration (wet AMD), the method comprising: identifying a subject with wet AMD; administering to the subject a first dose of a fusion protein conjugate comprising: a fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, wherein the heavy chain comprises an Fc region comprising a non-native cysteine; and a phosphorylcholine-containing polymer covalently attached to the non-native cysteine in the Fc region of the anti-IL-6 antibody; administering a second dose of the fusion protein conjugate to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein conjugate to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein conjugate to the subject about 4 weeks after the third dose; and administering a subsequent dose of the fusion protein conjugate to the subject no more frequently than about once every 8 weeks (or Q8W, or Q2M) after the fourth dose, optionally wherein the method comprises administering an individualized dose of the fusion protein conjugate to the subject at least 4 weeks (+ / - 7 days) after the fourth dose or about every 8 weeks thereafter, wherein each individualized dose is administered upon determining a decline in the subject’s eye health, wherein the subsequent dose and the individualized dose are collectively administered no more frequently than about once every four weeks (or no more frequently than Q4W or QM).

19. The method of claim 18, wherein the decline in eye health is based on the presence of intraretinal fluid (IRF), the presence of subretinal fluid (SRF), and / or new or worsening macular hemorrhage due to wet AMD activity, optionally wherein the method comprises evaluating an IRF volume and / or SRF volume in an optical coherence tomographyindividualized dose is administered upon determining that there is a decline in the subject’s eye health, wherein the subsequent dose and the individualized dose are collectively administered no more frequently than about once every four weeks (or no more frequently than Q4W or QM), wherein the first, second, third, fourth doses, the subsequent dose and the individualized dose each comprises about 5 mg of protein of the fusion protein conjugate, wherein the sequence of the heavy chain comprises SEQ ID NO: 170 (with or without the C-terminal lysine), and wherein the sequence of the light chain comprises SEQ ID NO: 169, wherein the antibody conjugate has the following structure: ,wherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H, and each light chain of the anti-IL-6 antibody is denoted by the letter L;the polymer is bonded to the anti-IL-6 antibody through a sulfhydryl at C443 according to EU numbering, which bond is depicted on one of the heavy chains above; PC , where the curvy line indicates the point of attachmentand n1, n2, n3, n4, n5, n6, n7, n8 and n9 are the same or different such that the sum of n1, n2, n3, n4, n5, n6, n7, n8 and n9 is 2500 plus or minus 15%.

22. The method of claim 18, wherein the administering comprises administering to the subject a therapeutic formulation comprising: an unconjugated fusion protein comprising the fusion protein with or without the non-native cysteine in the Fc region, wherein the unconjugated fusion protein is not covalently attached to a phosphorylcholine-containing polymer; and the fusion protein conjugate, wherein the unconjugated fusion protein is present in the formulation in a range of 10-60% of a total molar amount of the fusion protein conjugate and the unconjugated fusion protein, wherein the total molar amount is the sum of the molar amount of the fusion protein conjugate and the molar amount of the unconjugated fusion protein.

23. A method of treating an ocular inflammatory condition, comprising: administering to a subject in need of treating an ocular inflammatory condition one or more loading doses (optionally 3-5 loading doses) of a fusion protein and / or a conjugate thereof no more frequently than Q4W, the fusion protein comprising: an anti-IL-6 antibody; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody, whereby the subject retains a therapeutic result of the dual anti-IL-6 antibody / VEGF Trap therapy for at least 4 or 8 weeks after a final loading dose.

24. The method of claim 23, wherein the ocular inflammatory condition comprises diabetic macular edema (DME), diabetic retinopathy (DR), macular edema secondary to inflammation (MESI), uveitic macular edema (UME), post-surgical macular edema, macular edema associated with inflammatory choroidal neovascularization, pediatric UME, non- infectious uveitis, macular edema, macular edema following branch retinal vein occlusion(BRVO), wet age-related macular degeneration (AMD), retinopathy of prematurity (ROP), idiopathic edema, central retinal vein occlusion (CRVO), and / or uveitis.

25. The method of claim 23, wherein each of the loading doses comprises at least 2 mg of the fusion protein, optionally wherein each of the loading doses comprises from 2 mg to 15 mg of the fusion protein.

26. A method of treating diabetic macular edema (DME), comprising: identifying a subject with DME; administering to the subject a first dose of a fusion protein and / or a conjugate thereof, the fusion protein comprising: an anti-IL-6 antibody comprising: a heavy chain comprising the amino acid sequence of SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a light chain comprising the amino acid of SEQ ID NO: 169 or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a VEGF Trap fused to the heavy chain; administering a second dose of the fusion protein to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein to the subject about 4 weeks after the third dose; and administering a fifth dose of the fusion protein to the subject about 4 weeks after the fourth dose, wherein the first, second, third, fourth, and fifth dose each comprises about 2.5, 5, or 10 mg of the fusion protein.

27. A method of treating macular edema secondary to inflammation (MESI), comprising: identifying a subject with MESI;administering to the subject a first dose of a fusion protein and / or a conjugate thereof, the fusion protein comprising: an anti-IL-6 antibody comprising: a heavy chain comprising the amino acid sequence of SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a light chain comprising the amino acid of SEQ ID NO: 169 or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a VEGF Trap fused to the heavy chain; administering a second dose of the fusion protein to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein to the subject about 4 weeks after the second dose; and administering a fourth dose of the fusion protein to the subject about 4 weeks after the third dose, wherein the first, second, third, and fourth dose each comprises about 2.5, 5, or 10 mg of the fusion protein.

28. A method of treating MESI (e.g., with uveitic macular edema (UME)), comprising: identifying a subject with MESI (e.g., a subject with UME); administering to the subject a first dose of a fusion protein and / or a conjugate thereof, the fusion protein comprising: an anti-IL-6 antibody comprising: a heavy chain comprising the amino acid sequence of SEQ ID NO: 170 (with or without the C-terminal lysine) or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a light chain comprising the amino acid of SEQ ID NO: 169 or a sequence at least 90% identical thereto having one or more variations outside the CDR regions; and a VEGF Trap fused to a heavy chain of the anti-IL-6 antibody;administering a second dose of the fusion protein to the subject about 4 weeks after the first dose; administering a third dose of the fusion protein to the subject about 4 weeks after the second dose; administering a fourth dose of the fusion protein to the subject about 4 weeks after the third dose, wherein the first, second, third, and fourth dose each comprises about 2.5, 5, or 10 mg of the fusion protein, optionally wherein the method comprises administering an individualized dose of the fusion protein to the subject after the fourth dose and no more frequently than Q4W, wherein each individualized dose is administered upon determining that there is a decline in the subject’s eye health.

29. The method of claim 26, wherein a vision assessment as measured by Best Corrected Visual Acuity (BCVA) increases after administering at least the fourth dose relative to a baseline.

30. The method of claim 26, wherein a vision assessment as measured by Best Corrected Visual Acuity (BCVA) increases by at least 5, 10, or 15 Early Treatment Diabetic Retinopathy Study (ETDRS) letters.

31. The method of claim 23, wherein the VEGF Trap is positioned either: at an N-terminal end of a heavy chain variable region of the anti-IL-6 antibody; or between a hinge region and after a CH1 domain of a heavy chain comprising the heavy chain variable region of the anti-IL-6 antibody.

32. The method of claim 23, wherein the sequence of the heavy chain comprises a SEQ ID NO: 170 (with or without the C-terminal lysine), and wherein the sequence of the light chain comprises SEQ ID NO:

169.

33. The method of claim 23, wherein the heavy chain of the fusion protein conjugate comprises a Fc region comprising a non-native cysteine, wherein the fusion protein conjugate comprises a phosphorylcholine-containing polymer covalently attached to the non- native cysteine in the Fc region of the anti-IL-6 antibody.

34. The method of claim 23, wherein the polymer has a molecular weight between 300,000 and 1,750,000 Daltons.

35. The method of claim 23, wherein the fusion protein conjugate comprises the following structure: O O X PC n2 PC PC = P N(CH ) O O O CH O PC O X n5 PCwherein: each heavy chain of the anti-IL-6 antibody is denoted by the letter H, and each light chain of the anti-IL-6 antibody is denoted by the letter L; the polymer is bonded to a heavy chain of the antibody through the sulfhydryl of C443 (EU numbering), which bond is depicted on one of the heavy chains; O CH33PC i , where the curvy line indicates the point ofattachment to the rest of the polymer, where –X is: a) –OR where R is H, methyl, ethyl,43. The method of claim 23, wherein the administering comprises administering to the subject a therapeutic formulation comprising: an unconjugated fusion protein comprising the fusion protein with or without the non-native cysteine in the Fc region, wherein the unconjugated fusion protein is not covalently attached to a phosphorylcholine-containing polymer; and the fusion protein conjugate, wherein the unconjugated fusion protein is present in the formulation in a range of 10-60% of a total molar amount of the fusion protein conjugate and the unconjugated fusion protein, wherein the total molar amount is the sum of the molar amount of the fusion protein conjugate and the molar amount of the unconjugated fusion protein.

Images